UNIVERSIDAD COMPLUTENSE DE MADRID 
FACULTAD DE VETERINARIA 

 
 

 
 
 

TESIS DOCTORAL 
 

Dinámicas Evolutivas de los Cassettes de Resistencia en 
Integrones. Profundizando en el Modelo de Integrón 

 
Evolutionary Dynamics of Integron Resistance Cassettes. A 

Deeper Look into the Integron Model 
 
 

MEMORIA PARA OPTAR AL GRADO DE DOCTOR 
 

PRESENTADA POR 
 

Alberto Hipólito Carrillo de Albornoz 
 

Director 
 

José Antonio Escudero García-Calderón 
 
 

Madrid 
 
 

© Alberto Hipólito Carrillo de Albornoz, 2024 



 

 
  

Dinámicas Evolutivas de los Cassettes de Resistencia en 
Integrones. Profundizando en el Modelo de Integrón.

UNIVERSIDAD COMPLUTENSE DE MADRID

FACULTAD DE VETERINARIA

Evolutionary Dynamics of Integron Resistance Cassettes.
A Deeper Look into the Integron Model.

MEMORIA PARA OPTAR AL GRADO DE DOCTOR

PRESENTADA POR
Alberto Hipólito Carrillo de Albornoz

DIRECTOR
José Antonio Escudero García-Calderón

TESIS DOCTORAL



 

 
  



 

 

UNIVERSIDAD COMPLUTENSE DE MADRID 
FACULTAD DE VETERINARIA 

 
 
 
 
 
 
 
 
 

 
 
 

TESIS DOCTORAL 
 
 
Dinámicas Evolutivas de los Cassettes de Resistencia en Integrones. 

Profundizando en el Modelo de Integrón. 
 

Evolutionary Dynamics of Integron Resistance Cassettes.  
A Deeper Look into the Integron Model. 

 

MEMORIA PARA OPTAR AL GRADO DE DOCTOR 
 

PRESENTADA POR 
 

Alberto Hipólito Carrillo de Albornoz 
 

DIRECTOR 
 

José Antonio Escudero García-Calderón 
 
 
 



 

  



 

 

UNIVERSIDAD COMPLUTENSE DE MADRID 
FACULTAD DE VETERINARIA 

 
 
 
 
 
 
 
 
 

 
 
 

TESIS DOCTORAL 
 
 
Dinámicas Evolutivas de los Cassettes de Resistencia en Integrones. 

Profundizando en el Modelo de Integrón. 
 

Evolutionary Dynamics of Integron Resistance Cassettes.  
A Deeper Look into the Integron Model. 

 

MEMORIA PARA OPTAR AL GRADO DE DOCTOR 
 

PRESENTADA POR 
 

Alberto Hipólito Carrillo de Albornoz 
 

DIRECTOR 
 

José Antonio Escudero García-Calderón 
 
 

Programa de Doctorado de Bioquímica, Biología Molecular y Biomedicina 



 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 



 

 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 “Imagination is more  
important than knowledge” 

 
Albert Einstein 

 



 

 

 
 
 
 
 
 



 

 

 
 
 
 
 
 
 
 
 



Acknowledgements 

 9 

Acknowledgements 
 

Ubuntu is a Bantu term that can be translated as “I am because we are”, and I cannot think 
of a better word to start writing this thesis. Looking back to the beginning of this journey, I can 
remember countless moments when this word acquired great relevance, being essential to thank 
every person who has helped and guided me along this trip. 

 
En primer lugar querría dar las gracias a mi familia, especialmente a mis padres. Sin ellos 

este camino hubiera sido imposible de recorrer. Todo lo que soy os lo debo a vosotros. Gracias 
por animarme a perseguir mis metas y alumbrar el camino cuando se hacía más oscuro. Mil 
gracias al resto de mi familia, que tanto se ha preocupado por mí en estos años, en especial a 
mis abuelos, siempre seréis un referente para mí. 

 
In my case, the decision to dedicate my professional career to research and teaching 

science (a tough but gratifying task) is also inspired by the examples and dedication of several 
teachers and researchers who have transmitted their curiosity and vocation over the years. In 
these lines, I would also like to thank Manuel and Toñi for guiding me on my first steps into the 
scientific world at school; and Cristina, Nacho, and Chema for their passionate dedication to 
teaching at the university. I am especially grateful to Rocío; thanks for taking care of me inside 
and outside the lab, and showing me how a great mentor can become an irreplaceable friend. 

 
All work begins with an idea in the human intellect, and this thesis was born in Jose's mind, 

my thesis supervisor. Thank you for the trust placed in me to join your lab and turn your ideas into 
scientific discoveries. During these years, I had the opportunity and luck to develop myself as a 
scientist and as a person following your example. Thank you for every piece of knowledge and 
advice.  

 
Alongside José, I have been lucky enough to share fortunes and misfortunes with amazing 

colleagues: Lucía, for your support and care both personally and professionally; Paula, for being 
my partner in crime for any crazy and funny idea; Filipa, for sharing all these years of smiles and 
tears; Nico and André, for every discussion and laugh over a beer; Laura and Amalia, for your 
energy and push in this last part of this thesis; good luck in your PhDs; Carlota, for your trust and 
friendship; Cabesa, for making a regular day simply better. And last but not least, Ester, thanks 
for being the heart of this lab and making a place for me in your own one. 

 
I am also very grateful to Prof. Dr. Bärbel Stecher and her lab members for hosting me in 

Munich for three months and giving me the opportunity to elevate the relevance of this study to 
another level. I would also like to thank every great scientist, professor, and doctor that I met 
during this PhD for their advice and guidance, especially Bruno, Nacho, and Álvaro, together with 
their teams (Bosco, Emilia, Irene, Manu, José, Carlos, Mario, Javi; Miguel, the Javis; Jero, Richi, 
Javi, Alfonso,…). Discussing science with such great teams and people is one of the best 
experiences of this journey. 

 



Acknowledgements 
 

 10 

I would like to express my gratitude to all the financial agencies that have supported my 
research, including Universidad Complutense de Madrid, EMBO, and FEMS. 

 
On another note, I want to thank my friends—you have always been there when I needed 

you, and these years would certainly have been more difficult and sad without you. Christina, 
Quique, Pablo, Cris, Bea, Nuria, Luis, Miguel, Patri, Alex, Manuela... I could name a long list of 
people who have supported me during these 5 years. Some of you have been around for the last 
few years, others for 10 or 20 years, but I know that there is much more to come than what we 
have already experienced together. 

 
Finally, I would also like to thank the Music. Even if science is something that helps you 

living, music is one of the things that make life worth living. Listening and playing to music is my 
second activity in which I spend more time right after researching and it has been crucial to stand 
and enjoy these last years. You will find some of the songs that have accompanied me during this 
PhD among these pages. 

 
Five years is enough time to find amazing people but also to lose contact with some of 

them. I feel I have also to thank every person that has taken an important role along the way, even 
if it is not present at the end of the journey. Thank you all for every moment, every memory, and 
every lesson learned.   

 
 
 
 
 
 

                                                                                A.H.C.A. 
 

 
 
 
 
 



 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

TABLE OF  
CONTENTS 

 Scan me on Spotify 



 

 

  
 
 



Table of contents 

 13 

Table of Contents 

ACKNOWLEDGEMENTS 9 

TABLE OF CONTENTS 13 
List of Figures 16 
List of Tables 18 
List of Abbreviations 19 

RESUMEN 23 

SUMMARY 25 

1 INTRODUCTION 29 
1.1 Antimicrobial Resistance. 29 

1.1.1 A brief history of antibiotics. 29 
1.1.2 The antibiotic era: discovery of penicillin and antibiotic development. 29 
1.1.3 The rise of antimicrobial resistance. 30 
1.1.4 Mechanisms of antimicrobial resistance. 32 
1.1.5 Transmission of antimicrobial resistance: horizontal gene transfer (HGT). 32 
1.1.6 Mobile Genetic Elements (MGEs). 33 

1.2 Integrons. 35 
1.2.1 Structure and function of integrons. 35 
1.2.2 Origin of integrons: chromosomal integrons. 37 
1.2.3 Mobile integrons: prevalence and clinical relevance. 37 
1.2.4 Recombination system. 39 
1.2.5 Recombination reactions. 41 
1.2.6 Integron cassettes. 43 

2 OBJECTIVES 47 

3 MATERIALS AND METHODS 51 
3.1 Bacterial Strains, Plasmids, and Culture Conditions. 51 
3.2 Generation of the pMBA collection and derivatives. 51 
3.3 Antimicrobial resistance characterization by agar diffusion test. 52 
3.4 Minimal inhibitory concentration (MIC) determination. 52 



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 14 

3.5 Quantitation of GFP expression by flow cytometry. 53 
3.6 Quantification of gfp translation by Western blot. 53 
3.7 Quantification of gfp transcription by RT-qPCR. 53 
3.8 In vitro competition assays by flow cytometry. 54 
3.9 OMM12 stocks preparation. 55 
3.10 In vitro competition assays by phenotypic selection. 56 
3.11 In vivo competition assays. 57 
3.12 gDNA extraction from faecal pellets, caecal samples and in vitro competitions. 57 
3.13 Quantification of OMM12 microbiota members and E. coli abundance by qPCR. 58 
3.14 RNA-Seq analysis. 58 
3.15 Motility quantitation by swarming assays. 58 
3.16 Recombination assays. 59 
3.17 Calculation of the probability of folding attC sites. 60 
3.18 Growth curves. 60 
3.19 Inducibility of aacA5. 60 

4 RESULTS 65 
4.1 Evolutionary dynamics of integron resistance cassettes. 69 

4.1.1 Generation of the pMBA collection. 71 
4.1.2 Antimicrobial resistance characterisation. 73 

4.1.2.1 Aminoglycosides resistance cassettes. 73 
4.1.2.2 Beta-lactam resistance cassettes. 76 
4.1.2.3 Antifolate resistance cassettes. 78 
4.1.2.4 Fosfomycin resistance cassettes. 79 
4.1.2.5 Chloramphenicol resistance cassettes. 80 
4.1.2.6 Rifampicin resistance cassettes. 80 
4.1.2.7 Erythromycin resistance cassettes. 80 
4.1.2.8 Quaternary ammonium compounds ARCs (qac, smr). 81 
4.1.2.9 Influence of the genetic context in ARC activity. 82 

4.1.3 Polar effects characterisation. 84 
4.1.3.1 ARC identity affects the expression of downstream ICs. 84 
4.1.3.2 ARC identity affects the transcription of downstream ICs. 85 
4.1.3.3 attC sites are involved in the transcriptional regulation of the array. 86 

4.1.4 Fitness cost characterisation. 89 
4.1.4.1 ARCs can provide a fitness gain in the absence of antibiotics in vitro. 89 
4.1.4.2 Anaerobiosis is a crucial condition affecting the fitness dynamics of some ARCs. 92 



Table of contents 

 15 

4.1.4.3 ARCs can provide a fitness gain in the absence of antibiotics in vivo. 97 
4.1.4.4 ereA2 reduces E. coli motility. 98 

4.1.5 ARC recombination frequency characterisation. 102 
4.1.5.1 Generation of the pMBA recombination collection. 102 
4.1.5.2 Recombination frequency quantitation. 104 
4.1.5.3 Correlation between recombination frequencies and in silico analysis. 106 

4.2 A deeper look into the integron model. 111 
4.2.1 The expression of AgR genes in ICs  is not controlled by riboswitches. 113 

4.2.1.1 Expression of aacA5 is not induced by Ags in its native genetic context. 115 
4.2.1.2 Generation of the pMBA 5´aaX collection. 117 
4.2.1.3 AgR cassettes are not repressed in the absence of antibiotics. 117 
4.2.1.4 Aminoglycosides do not induce the expression of AgR cassettes. 119 
4.2.1.5 Higher concentrations of Ags neither induce resistance genes. 121 
4.2.1.6 Induction is higher in the presence of unrelated antibiotics. 122 
4.2.1.7 Increased expression is due to pleiotropic effects of antibiotics. 123 

5 DISCUSSION 127 

6 CONCLUSIONS 137 

7 BIBLIOGRAPHY 141 

8 SUPPLEMENTARY MATERIAL 155 
 
 
 
 
 
 



Table of contents 

 16 

List of Figures 
 
Figure 1. Timeline showing the discovery and first report of clinical resistance. (31)                                     
Figure 2. Horizontal gene transfer. (33)                                                                                                         
Figure 3. Schematic representation of the mobilisation of ICs in integrons. (36)                                           
Figure 4. Mobile integron prevalence and classification. (38) 
Figure 5. Sequence of a double stranded (ds) attI1 site. (39) 
Figure 6. Sequence and structure of an attC site. (40) 
Figure 7. Recombination reaction between attC bs and attI sites. (42) 
Figure 8. Recombination reaction between two attC bs sites. (42) 
Figure 9. IC occurrence and distribution in MIs. (43) 
Figure 10. Schematic representation of the cloning region in pMBA derivatives. (51) 
Figure 11. Flow diagram of the experimental process. (55) 
Figure 12. Occurrence of each ARC within its gene family across databases. (69) 
Figure 13. Generation of the pMBA collection. (71) 
Figure 14. MICs of aminoglycoside resistance cassettes. (75) 
Figure 15. MICs of b-lactam resistance cassettes. (77) 
Figure 16. Antifolates resistance characterisation. (79) 
Figure 17. MICs of ARCs. (81) 
Figure 18. MIC characterisation of qac and smr ARCs. (82) 
Figure 19. Comparison of the MIC of ARCs in different genetic contexts. (83) 
Figure 20. ARC identity affects the expression of downstream ICs. (84) 
Figure 21. GFP expression of a subset of pMBAdfr strains. (85) 
Figure 22. GFP fluorescence and protein yields in a subset of pMBAdfr strains. (85) 
Figure 23. GFP fluorescence, protein yields, and GFP mRNA levels in a subset of pMBAdfr strains. (86)      
Figure 24. Compilation of attC site parameters in pMBAdfr strains. (87) 
Figure 25. GFP expression comparison between strains with modified and deleted attC sites. (87) 
Figure 26. GFP fluorescence comparison between strains with and without PcS promoter. (88) 
Figure 27. Growth comparison between different pMBA derivative strains. (89) 
Figure 28. Correlation between fitness cost and GFP fluorescence. (90) 
Figure 29. Effect of each ARC in the fitness of the host strain. (91) 
Figure 30. OMM12 consortium phylogenetic tree. (92) 

Figure 31. Pairwise competitions between pMBA and pMBAARCs  strains. (93) 

Figure 32. Relative abundance of each OMM12 member in batch culture competitions. (94) 
Figure 33. Effect of each ARC in the fitness of the host strain in anaerobic conditions. (95) 
Figure 34. Comparison between ARCs-fitness effect in aerobiosis and anaerobiosis. (96) 
Figure 35. ARCs can provide a fitness gain in the absence of antibiotics in vivo. (97) 
Figure 36. Volcano plot showing differential gene expression of pMBAereA2 in comparison to pMBA. (99)   



Table of contents 

 17 

Figure 37. ereA2 reduces E. coli motility. (100) 
Figure 38. MIC characterisation of pMBA derived strains against erythromycin. (100) 
Figure 39. ereA2 expression reduces E. coli motility. (101) 
Figure 40. Schematic representation of a recombination assays. (102) 
Figure 41. attC site recombination. (103) 
Figure 42. Recombination frequency of each ARC. (104) 
Figure 43. Analysis of the average recombination capacity of each ARC family. (104) 
Figure 44. Structures of bottom strands of attC sites. (105) 
Figure 45. Comparative analysis of the recombination frequencies of ARCs. (106) 
Figure 46. Correlation between recombination frequencies and in silico analysis. (107) 
Figure 47. The AG sensing riboswitch described in an integron cassette is counterintuitive. (114) 
Figure 48. Mobile integrons contain cassettes encoding varied functions. (115) 
Figure 49. The expression of aacA5 is not induced by aminoglycosides in its native genetic context. (116)    
Figure 50. Aminoglycoside resistance cassettes are not repressed in the absence of antibiotics. (118) 
Figure 51.  Aminoglycosides do not induce the expression of aminoglycoside resistance cassettes. (120) 
Figure 52.  High concentrations of aminoglycosides do not induce AgR cassettes. (121) 
Figure 53. Induction is higher in the presence of unrelated antibiotics. (122) 
Figure 54.  Increased expression is due to pleiotropic effects of antibiotics. (123) 
Figure 55.  Distribution of ARCs by antibiotic class among different bacterial genus. (128)   
Figure 56.  Correlation between ARC recombination rate and GC content of the gene encoded. (132) 

 
Supplementary Figure S1. Resistance characterisation of aa ARCs by agar diffusion test. (243) 
Supplementary Figure S2. Resistance characterisation of bla ARCs by agar diffusion test. (244) 
Supplementary Figure S3. pMBAARCs fluorescence do not change in stationary growth phase. (245) 
Supplementary Figure S4. Growth curves of all pMBA derivatives. (246) 
Supplementary Figure S5. Relative abundance of each OMM12 member in vivo competitions. (249) 
Supplementary Figure S6. Volcano plots showing DE genes of pMBA and pMBA derivatives. (250) 
Supplementary Figure S7. Non-corrected recombination frequency of each ARC. (251) 
Supplementary Figure S8. Cassette expression correlates with the presence of SD-like sequences. (252) 
Supplementary Figure S9. Antibiotic concentrations used do not affect growth. (253) 
Supplementary Figure S10. Alternative 5´UTRs are not induced by aminoglycosides. (257) 
Supplementary Figure S11. Results from statistical models applied to induction assays. (258) 
Supplementary Figure S12. Hierarchical clustering tree of all attC sites present in ARCs. (259)  



Table of contents 

 18 

List of Tables 
 
Supplementary Table S1. Strains and plasmids used in this study. (155) 
Supplementary Table S2. Primers and probes used in this study. (164) 
Supplementary Table S3. Antibiotic discs used in this study. (167) 
Supplementary Table S4. Antimicrobial compounds used in this study. (168) 
Supplementary Table S5. Antimicrobial resistance cassettes (ARCs) sequences. (169) 
Supplementary Table S6. Transcriptomic analysis of pMBA containing E. coli MG1655. (188) 
Supplementary Table S7. Transcriptomic analysis of pMBA derivatives. (190) 
Supplementary Table S8. Sequences of all integron cassette 5’UTRs used in this work. (241)   
 

 

  



Table of contents 

 19 

List of Abbreviations  
 
Ab: antibiotic. 
Ag: aminoglycoside. 

AgR gene: aminoglycoside resistance gene. 
aHJ: atypical Holliday junction. 

AMEs: aminoglycoside modifying enzymes. 

ARCs: antimicrobial resistance cassettes. 

ARGs: antimicrobial resistance genes. 
BC: before Christ. 

bp: base pairs. 

bs: bottom strand. 

BSA: bovine serum albumin. 

BZK: benzalkonium chloride. 

ºC: Celsius degrees. 

Cat. Mut.: catalytic mutant. 

cDNA: complementary DNA. 
CFUs: colony forming units. 

CHX: chlorhexidine. 

CLSI: Clinical and Laboratory Standards Institute. 

CmR: chloramphenicol resistance. 

cpm: cycles per minute. 

CTAB: hexadecyltrimethylammonium bromide. 

DDs: disubstituted deoxystreptamines. 

ddH2O: double distilled water. 
DE: differentially expressed. 

DG: Gibbs free energy. 

DNA: deoxyribonucleic acid. 

DR: direct repeats. 

ds: double strand. 
E. coli: Escherichia coli. 

EHB: extra helical bases. 

ESBL: extended spectrum beta-lactamase. 

EUCAST: European Society of Clinical 

Microbiology and Infectious Diseases. 

g: gram. 

gcu: gene cassette of unknown function. 

gDNA: genomic DNA. 

GFP: green fluorescent protein.  

HGT: horizontal gene transfer. 

HJ: Holliday junction. 

IC: integron cassette. 

ICE: integrative conjugative element. 

IME: integrative mobile element 

IQR: interquartile range. 
IR: induction ratio. 

IRs: inverted repeats. 

IS: insertion sequence. 

K: Kelvin degrees. 

kb: kilobases. 

kcal: kilocalories. 

KO: knockout. 

L: litre. 
LB: lysogeny broth. 

MGE: mobile genetic element. 

mg: milligram. 

µg: microgram. 

MH: Müller Hinton. 
MI: mobile integron. 

MIC: minimal inhibitory concentration. 

MICs: mobile insertion cassettes. 

MITE: miniature inverted repeats transposable 

element. 

mL: millilitre. 

µL: microlitre. 

µM: micromolar. 

mm: millimetre. 

mRNA: messenger RNA. 

nm: nanometres. 
OD600: optical density at 600 nm. 

OMM12: oligo-mouse microbiota 12. 

ORF: open reading frame. 

PBPs: penicillin binding proteins. 

PCR: polymerase chain reaction. 



Table of contents 

 20 

PICI: phage-inducible chromosomal islands. 

QACs: quaternary ammonium compounds. 

 

qPCR: quantitative PCR. 

RNA: ribonucleic acid. 

rpm: revolutions per minute. 

rRNA: ribosomal RNA. 

RT-qPCR: reverse transcription qPCR. 
SCI: sedentary chromosomal integron. 

SD: Shine Dalgarno. 

SEM: standard error of the mean.  

SI: super integron. 

spp.: several species. 

ss: single strand. 

TA: toxin-antitoxin. 

TBS: tris buffered saline. 
tIS: transporter IS. 

Tn: transposon. 

ts: top strand. 

UCS: uncoupled central spacer. 

VCRs: Vibrio cholerae repeats. 

VTS: variable terminal structure. 

WB: western blot. 

WHO: World Health Organisation. 
5´UTR: 5´untranslated region.  

 

 

 

 

 

 
 



 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

RESUMEN 
SUMMARY  

 
 

 
 
 
 
 

Scan me on Spotify 



 

 

 



Resumen / Summary 

 23 

Resumen  
 

La resistencia a antimicrobianos es una de las principales amenazas para la salud global 
del siglo XXI. El uso indiscriminado de antibióticos, entre otros factores, ha provocado un 
incremento exponencial de bacterias resistentes a antimicrobianos en las últimas décadas. Se 
estima que cada año mueren entre 1.5 y 5 millones de personas debido a infecciones causadas 
por bacterias multirresistentes. Estos datos ponen de manifiesto la necesidad de tomar medidas 
urgentes para combatir esta amenaza, empezando por entender las fuerzas que rigen el éxito de 
las bacterias multirresistentes. 
 

La excepcional capacidad de las bacterias para evolucionar y adaptarse a compuestos 
antimicrobianos está principalmente promovida por la transferencia horizontal de genes. Gran 
cantidad de genes de resistencia se transfieren de unas bacterias a otras mediante elementos 
genéticos móviles y plataformas como los plásmidos, transposones, e integrones.  

 
Los integrones son plataformas genéticas capaces de capturar y almacenar nuevos genes 

embebidos en pequeños elementos genéticos móviles llamados cassettes. Los integrones 
poseen una estructura conservada, compuesta por una plataforma estable; donde se encuentran 
codificados el gen de la integrasa, los promotores Pc y Pint, y el sitio attI de integración; y por una 
colección variable de cassettes. La colección de cassettes se expresa desde el promotor Pc 
siguiendo un gradiente de expresión que mantiene silenciados los cassettes más alejados del 
promotor. Ante una situación de estrés, la integrasa se expresa, permitiendo la captación 
cassettes exógenos, así como la escisión y reordenamiento de los cassettes. Este 
reordenamiento permite la expresión de cassettes distales recolocándolos en primera posición 
de la colección. Por ello, los integrones actúan como memorias bacterianas de muy bajo coste 
capaces de proporcionar adaptación bajo demanda a su hospedador.    

 
Los integrones móviles desempeñan un papel crucial en el aumento y propagación de la 

resistencias a antimicrobianos, siendo capaces de capturar y expresar más de 170 genes de 
resistencia antimicrobiana frente a la mayoría de antibióticos de relevancia clínica. Los integrones 
son plataformas de gran éxito, frecuentemente presentes en aislados clínicos de bacterias Gram 
negativas multirresistentes. Por el contrario, la prevalencia de los cassetes de resistencia en 
integrones varía ampliamente, incluso entre genes que confieren la misma resistencia a 
antimicrobianos. Este fenómeno sugiere que existen otros rasgos específicos que gobiernan el 
éxito diferencial de los cassettes de resistencia distintos al fenotipo que confieren.  

 
Esta tesis doctoral tiene como objetivo caracterizar las fuerzas que rigen el éxito evolutivo 

de los cassettes de resistencia en integrones, así como profundizar en el modelo actual de 
integrón y su regulación. Para ello, hemos cuantificado los niveles de resistencia conferidos por 
estos cassettes así como el coste asociado a su expresión. Adicionalmente, y debido a su 
presencia en esta plataforma, hemos caracterizado el posible efecto de un cassette en la 
expresión de la colección posterior, así como la movilidad de cada uno de ellos. 

 



Resumen / Summary 
 

 24 

A pesar de la gran relevancia clínica de esta plataforma, el conocimiento acerca de los 
genes de resistencia que porta se encuentra disperso en la literatura, impidiendo un análisis 
comparativo de los mismos. Para estudiar de manera comparable las fuerzas que rigen el éxito 
de estos cassettes hemos generado la colección pMBA, compuesta por 136 cassettes de 
resistencia clonados en un entorno genético que mimetiza el natural. Abarcando el 76% de los 
cassettes de resistencia descritos en integrones móviles, es la mayor colección de cassettes de 
resistencia en integrones en la actualidad. 

 
La caracterización del perfil de resistencia de cada cassette ha permitido confirmar la ya 

conocida especificidad por familias de genes, a la vez que pone de manifiesto grandes diferencias 
en los niveles de resistencia conferidos por genes filogenéticamente muy cercanos. Asimismo, 
hemos detectado genes que no confieren resistencia alguna, lo cual pone en duda el papel de 
las familias de genes qac y smr como genes de resistencia a desinfectantes.  

 
Por su parte, la cuantificación del coste asociado a cada cassette ha revelado que ciertas 

familias, como los genes de resistencia a beta-lactámicos, poseen un gran coste en E. coli. Por 
el contrario, genes como aacA7 han demostrado ser beneficiosos para el fitness bacteriano en 
ausencia de selección, tanto in vitro como in vivo, sugiriendo un posible rol metabólico secundario 
de los mismos. Adicionalmente, competiciones bacterianas realizadas tanto en aerobiosis como 
en anaerobiosis, han demostrado que los cassettes de resistencia pueden modificar 
drásticamente su efecto en el fitness bacteriano dependiendo de las condiciones oxigénicas del 
entorno, como en el caso de ereA2. Este hecho tiene una gran relevancia a la hora de entender 
la ecología y su prevalencia de los cassettes en entornos clínicos y ambientales. 

 
Además de caracterizar la resistencia que confieren los cassettes y el coste que implican 

para su hospedador, hemos caracterizado la influencia de un cassettes en la colección posterior 
y su capacidad de recombinación en la plataforma. Por un lado, observamos que la identidad de 
un cassette puede modular la expresión de la colección posterior de manera transcripcional, 
añadiendo un nuevo nivel de complejidad al modelo de integrón. Este hecho posee importantes 
implicaciones en posibles fenómenos de co-selección ocurridos en infecciones tratadas con 
varios antibióticos. Por otro lado, las frecuencias de recombinación obtenidas para cada cassettes 
son muy variables, existiendo diferencias de hasta un millón de veces entre las tasas de 
recombinación de aacA52 y fosN. Si bien no hemos podido relacionar parámetros in silico como 
DG y pfold de los sitios attC de los cassettes y su tasa de recombinación en nuestros 
experimentos, encontramos una correlación entre los cassettes de resistencia a aminoglucósidos 
y altas tasas de recombinación. 

 
Adicionalmente, durante el transcurso de este estudio hemos podido demostrar que los 

genes de resistencia a aminoglucósidos presentes en integrones no están regulados por 
riboswitches, tema de controversia en el campo hasta ahora.  

 
Si bien todavía estamos lejos de comprender en su totalidad el éxito diferencial de cada 

cassette de resistencia, esta tesis doctoral ofrece una visión general y cuantificada de las fuerzas 
que rigen el éxito evolutivo de los cassettes de resistencia en integrones.  



Resumen / Summary 

 25 

Summary  
 

Antimicrobial resistance is one of the major global health concerns of the 21st century. The 
indiscriminate use of antibiotics, among other factors, has led to an exponential increase in 
antimicrobial-resistant bacteria in recent decades. It is estimated that every year between 1.5 and 
5 million people die due to infections caused by multidrug-resistant bacteria. These data highlight 
the need to take urgent measures to tackle this threat, with a priority on understanding the forces 
that govern the success of multidrug-resistant bacteria. 

 
The exceptional ability of bacteria to evolve and adapt to antimicrobial compounds is 

mainly promoted by horizontal gene transfer. A large number of resistance genes are transferred 
from one bacterium to another through mobile genetic elements and platforms such as plasmids, 
transposons, and integrons. 

 
Integrons are genetic platforms capable of capturing and storing new genes embedded in 

small mobile genetic elements called cassettes. Integrons have a conserved structure, composed 
of a stable platform encoding the integrase gene, the Pc and Pint promoters, and the attI 
integration site; and a variable array of cassettes. The cassettes’ array is expressed from the Pc 
promoter following an expression gradient that maintains the cassettes furthest from the promoter 
silenced. Under stress conditions, the integrase is expressed, allowing to capture exogenous 
cassettes, as well as the excision and rearrangement of the cassettes present in the array. This 
rearrangement allows the expression of distal cassettes by relocating them in the first position of 
the array, via recombination between the attI site placed in the platform and the attC site of the 
cassette. Therefore, integrons act as low-cost bacterial memories capable of providing adaptation 
on-demand to their hosts. 

 
Mobile integrons play a crucial role in the increase and spread of antimicrobial resistance, 

capable of capturing and expressing over 170 antimicrobial resistance genes against most 
clinically relevant antibiotics. Integrons are highly successful platforms, often found in clinical 
isolates of multidrug-resistant Gram-negative bacteria. Conversely, the prevalence of 
antimicrobial resistance cassettes varies widely, even among genes conferring the same 
antimicrobial resistance. This phenomenon suggests that there are other specific features 
governing the differential success of resistance cassettes other than the phenotype they confer. 

 
 This PhD thesis aims to characterize the forces governing the evolutionary success of 

resistance cassettes in integrons, as well as to delve into the current model of integron and its 
regulation. To achieve this, we have quantitate the resistance levels conferred by these cassettes, 
as well as the cost associated to their expression. Furthermore, we have explored the potential 
impact of a cassette on the expression of the subsequent collection and assessed their mobility, 
given their presence in this specific platform. 

 
  
 



Resumen / Summary 
 

 26 

Despite the significant clinical relevance of this platform, our knowledge about their 
resistance genes is scattered in the literature, hindering a comparative analysis. To study the 
forces governing the success of these cassettes in a comparable manner, we have generated the 
pMBA collection, composed of 136 resistance cassettes cloned in a genetic environment that 
mimics their natural context. This collection encompasses 76% of the resistance cassettes 
described in mobile integrons, being largest collection of resistance cassettes in integrons to date. 

 
The characterisation of the resistance profile of each cassette has confirmed the already 

known specificity by gene families, while revealing significant differences in the resistance levels 
conferred by closely related genes. Additionally, we have identified genes that do not confer any 
resistance, challenging the role of qac and smr gene families as disinfectant resistance genes. 

 
The quantification of the cost entailed by each cassette has revealed that certain families, 

such as beta-lactam resistance genes, pose a significant cost in E. coli. Conversely, certain genes, 
such as aacA7, have proven to be beneficial for bacterial fitness in the absence of selective 
pressure, both in vitro and in vivo. This suggests potential secondary metabolic roles of 
antimicrobial resistance cassettes in integrons. Furthermore, bacterial competitions conducted 
both in aerobic and anaerobic conditions have shown that resistance cassettes can drastically 
modify their effect on bacterial fitness depending on the oxygen availability in the environment, as 
observed in the case of ereA2. This fact is highly relevant for understanding the ecology of 
cassettes and their prevalence in clinical and environmental settings. 

 
In addition to the characterisation of the resistance conferred by the cassettes and the cost 

they impose on their host, we have investigated the influence of a cassette on the subsequent 
collection and its recombination capacity in the platform. On one hand, we observe that the identity 
of a cassette can modulate the expression of the collection transcriptionally, adding a new level 
of complexity to the integron model. This has important implications for potential co-selection 
phenomena occurring in infections treated with various antibiotics. On the other hand, 
recombination frequencies obtained for each cassette are highly diverse, with differences of up to 
a million times between the recombination rates of aacA52 and fosN. Although we have not been 
able to correlate in silico parameters such as DG and pfold of the attC sites of the cassettes with 
their recombination rate in our experiments, we have found a correlation between cassettes 
conferring resistance to aminoglycosides and high recombination rates. 

 
Additionally, during the course of this study, we have demonstrated that the 

aminoglycoside resistance genes present in integrons are not regulated by riboswitches, a topic 
of controversy in the field until now. 

 
Although we are still far from a complete understanding of the differential success of each 

resistance cassette, this PhD thesis provides a comprehensive and quantified view of the forces 
governing the evolutionary success of resistance cassettes in integrons. 
 
 
 
 



 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

INTRODUCTION  
 
 

  Scan me on Spotify 



 

 

 
 



   Introduction 
 

29 

1 Introduction 
 
1.1 Antimicrobial Resistance. 

 
1.1.1 A brief history of antibiotics. 
 

The word “antibiotic” was first introduced in our vocabulary by Selman Waksman in 1942 
to describe “substances produced by microorganisms that provoke an antagonistic effect in other 
microorganisms growth when present in high dilution”1. This initial and rather simplistic definition 
of antibiotic laid the foundations for further descriptions of these molecules up to the present day. 
Currently, antibiotics and antimicrobials can be defined as natural, synthetic, or semi-synthetic 
substances that, at low concentrations, inhibit or kill susceptible microorganisms2.  

 
Although antibiotics were defined during the 20th century, the use of antibiotic-producing 

microorganisms to treat and prevent diseases dates back thousands of years. The oldest 
documented report of these medical practices is the Eber´s papyrus from 1550 BC. In this 
document, the use of mouldy bread and medicinal soil is advised as therapeutic remedies3. 
Nevertheless, the first use of antibiotics that we can attributed to the modern concept of 
chemotherapy is generally credited to Paul Ehrlich in 1907 for the development of the synthetic 
antibiotic Salvarsan, which exhibited activity against Treponema pallidum, the causative agent of 
syphilis4. Ehrlich´s studies inspired colleagues to discover other synthetic antibiotics such as the 
sulfonamides, firstly described by Gerhard Domagk and commercialized under the name of 
Prontosil in 19355. Therefore, sulfonamides are accredited as the first broad spectrum 
antimicrobials used in the clinic since the decade of 1930 to date.  

 
1.1.2 The antibiotic era: discovery of penicillin and antibiotic development. 
 

While the development of synthetic antibiotics was improving the clinical practice, the 
discovery of the first natural antibiotic by Alexander Fleming in 1928 marked a breaking point in 
medicine and human wellbeing. His initial observations about the inhibitory, bacteriolytic, and 
bacteriostatic properties of Penicillium ushered the beginning of the antibiotic era6. Penicillin was 
finally produced and distributed on a large scale in 1945 due to the effective purification protocol 
developed by Howard Florey and Ernest Chain in 19407.  

 
From 1940 to 1970 there was an exponential increase in the discovery of new molecules, 

acknowledged as the golden age of antibiotics. During this period, past studies in bacterial 
pathogens and antibiosis performed by renown scientists such as Louis Pasteur or Robert Koch 
served as a starting point for the systematic search of natural antibiotics8. The study of potential 
antimicrobial producers in soil samples by Selman Waksman and Albert Schatz resulted in the 
identification of Streptomyces genus as one of the most prolific producers of natural antimicrobials, 
being streptomycin the first discovered compound produced by this genus9.  

 
 



Introduction  

30 

Waksman´s findings inspired the pharmaceutical industry and triggered the discovery of 
the vast majority of antibiotics currently in use. All along this golden age, around 20 classes of 
antibiotics were isolated from bacterial and fungal species10. During this period, the use of 
antibiotics became a general practice in medicine. This fact resulted in an increase in life 
expectancy and a great reduction in human mortality derived from infectious diseases. Altogether, 
we can consider antibiotic discovery and development a milestone in human history and one of 
the most important improvements in modern medicine. In addition to the extensive research and 
discovery of new natural antibiotics, the structural and mechanistic description of these new 
molecules enabled the production of semi-synthetic antimicrobials. A well know example of these 
studies is the determination of the b-lactam structure of penicillin by Dorothy Hodgkin in 194511. 

 
From 1980 onwards, the discovery of new antibiotics slowed down. Finding new classes 

of molecules exerting antibiotic activity became exceptional, giving way to the medical chemistry 
age. During this period of time (1980-1990), antibiotic research was focused on the development 
of semi-synthetic antibiotics from natural molecules discovered in the golden era. These semi-
synthetic antibiotics permitted antibiotic usage in lower doses, expanding its action spectrum and 
avoiding incipient resistances against these molecules12 .   

 
Since 1990, antibiotic discovery and development decreased radically. The amount of new 

antibiotics as well as the investment on their development diminished until our days, with no new 
antibiotic classes commercially developed since 1980s13. Nevertheless, the emergence of new 
technologies provides alternative antibiotic screening approaches based on metagenomics and 
combinatorial biosynthesis14. Unfortunately, these metagenomics and combinatorial biosynthesis 
screenings have not been successful so far.  

 
On the contrary, the prevalence of antibiotic resistant bacteria increased unstoppably. The 

massive use of antibiotics together with the outstanding evolvability of bacteria triggered the end 
of the antibiotic era giving rise to the contemporary antimicrobial resistance situation. 

 
1.1.3 The rise of antimicrobial resistance. 

 
Antimicrobial resistance is defined as the ability of a bacterium to survive and grow when 

it is exposed to the hazardous action of a drug against it. Antimicrobial resistance can be displayed 
due to inherent properties of the bacterium (intrinsic), induced by specific signals or conditions 
transiently (adaptive), or achieved by genetic mutations or incorporation of exogenous genetic 
material via horizontal gene transfer (HGT) (acquired)15. 
 

Although antibiotic resistance is ancient, with resistance genes found in DNA from 30000 
year old sediments16, the selective pressure applied by the excessive use of antibiotics in the last 
century has caused a great increase in antimicrobial resistant bacteria. Fleming himself advised 
about the risks of misuse and abuse of these molecules, suggesting that subinhibitory 
concentrations of these molecules could lead to the selection of resistant bacteria. 



   Introduction 
 

31 

As expected, antibiotic resistant bacteria have been discovered all over the world after few 
years of antibiotic use; starting at 1936 when the first sulfonamide-resistant isolate was described, 
and continuing to the present day, when we encounter resistant bacteria against every single 
antibiotic used in the clinical practice (Figure 1). Nowadays we can find multirresistant bacteria 
against several antibiotic in a number of clinical isolates. 

  
Figure 1.Timeline showing the discovery (blue pill) and first report of clinical resistance (red 
bacterium).10,17. Modified from 18. 
 

Antibiotic resistant bacteria are able to survive under antimicrobial-selective conditions, but 
this ability also entails a theorical cost in the absence of any selective pressure, leading to a 
reduction in bacterial fitness. Initial hypotheses suggested that a decrease in antibiotic usage 
could lead to an enriched susceptible bacterial population by outcompeting the resistant ones19. 
Nevertheless, events such as compensatory evolution and genetic co-selection20, and platforms 
such as integrons (see below) alleviate the cost produced by the acquired resistance. The fact 
that bacteria can alleviate the cost derived from acquiring antibiotic resistance, together with the 
possibility of sharing DNA by HGT facilitates the emergence of multidrug resistant bacteria. 
Multidrug resistance was first described in Japan among Shigella spp. isolates during 1950´s 
decade21. These isolates were resistant to several antimicrobials such as streptomycin, 
tetracycline, chloramphenicol, and sulfonamides. Subsequent research revealed that these 
resistance determinants were carried in a conjugative plasmid, which also contained an integron22. 

 
The emergence of multirresistant bacteria is considered by the World Health Organisation 

(WHO) as one of the major global health concerns of the 21st century23. Actual estimations reveal 
that every year between 1.5 and 5 million people pass away from antimicrobial resistant bacterial 
infections24. This datum, together with the economic cost derived from antimicrobial resistant 
infections, has alerted numerous international organizations about the irreversible consequences 
of a post-antibiotic era where antibiotics become ineffective. 

1930 1940 1950 1960 1970 1980 1990 2000 2010 2020

Sulphonamides

Penicillin

Tetracycline

Erythromycin

Vancomycin

Methicillin

Gentamicin

Cefotaxime

Ceftazidime

Imipenem

Ciprofloxacin

Daptomycin

Ceftazidime - Avibactam



Introduction  

32 

1.1.4 Mechanisms of antimicrobial resistance. 
 

As mentioned before, bacteria are capable of avoiding the activity of antibiotics by 
acquiring resistance to these compounds via mutation or HGT. The mechanisms underlying 
antibiotic resistance are generally antibiotic modification, target alteration, or reduction of the 
antibiotic accumulation15. 

 
Antibiotic modification consists of the alteration of the antimicrobial structure (via 

hydrolysis or addition of radicals) to inactivate the molecule. This is the most common mechanism 
of resistance against b-lactams such in Gram-negative bacteria. b-lactamases (encoded in bla 
genes) hydrolyse the b-lactam ring (the main structure of these compounds), rendering the 
antimicrobial harmless25. Also, the action of aminoglycoside modifying enzymes (AMEs) causes 
antibiotic resistance via acetylation, adenylation, or phosphorylation of aminoglycosides (mediated 
by aac, aad, and aph genes respectively)26. Enzymatic modification of other antibiotics such as 
chloramphenicol and rifampicin (via catB and arr genes respectively) are common mechanisms of 
resistance against these compounds 27,28. 

 
Antibiotic target alteration such as point genetic mutations allow the target to escape 

from the antibiotic activity. A common case of acquired resistance by genetic mutations is the 
fluoroquinolone resistance mediated by the chromosomal mutations in gyrase and topoisomerase 
IV genes (gyrA and parC) 29. In addition, antibiotic targets can suffer non-genetic modification to 
escape from antibiotic activity. An example of post-translational modification leading to antibiotic 
resistance is the methylation of the 16S ribosomal RNA by armA, which protects against 
aminoglycosides30. Furthermore, the overproduction or acquisition of resistance homologous 
genes can bypass the antimicrobial action. Common examples are sulfonamides and trimethoprim 
resistance, generally mediated by the acquisition of dfr and sul genes that avoid the antimicrobial 
action31. Moreover, antibiotic evasion can be achieved by antibiotic target protection. Quinolone 
resistance proteins (qnr) act as DNA analogues that bind to gyrase and topoisomerase IV lowering 
the available targets, rendering bacteria resistant to quinolones29.  
 

Efflux pumps are the main drivers of antibiotic accumulation reduction. These energy-
dependent membrane complexes are able to export compounds out of the cell. Multidrug efflux 
mechanisms are responsible for the intrinsic resistance of some bacterial species against  multiple 
drugs. In addition, several substrate-specific efflux pumps have been described to confer 
resistance against chloramphenicol, quaternary ammonium compounds (QACs), macrolides and 
tetracyclines32. 

 
1.1.5 Transmission of antimicrobial resistance: horizontal gene transfer (HGT). 
 

Horizontal gene transfer (HGT) is a key process in the development of resistant bacteria. 
HGT is defined as the ability of a cell to acquire new genetic material from another organism. This 
mechanism allows bacteria to evolve quickly under stress conditions, such as the presence of 
antimicrobials, acquiring antimicrobial resistance genes (ARGs)33,34. 



   Introduction 
 

33 

Horizontal acquisition of exogenous DNA has to be facilitated by three main mechanisms: 
natural transformation, conjugation, and transduction35 (Figure 2A). Nevertheless alternative HGT 
ways mediated by vesicles or nanotubes have also been described 36,37. 
 

Natural transformation consists of the active uptake of free DNA present in the 
environment by a cell38. This mechanism relies on the natural competence of the recepient cell to 
capture, recombine and express exogenous DNA. Genera such as Vibrio, Acinetobacter and 
Streptococcus are examples of bacteria that are able to acquire exogenous genes by natural 
transformation39. Besides, HGT mediated by conjugation needs a type IV secretion system that 
produces a pilus (cell to cell junction) to perform DNA transference from the donor cell to the 
recipient one. This mechanism is the main driver of antimicrobial resistance dissemination 
mediated by plasmids among clinical settings40. Likewise, transduction is mediated by a 
bacteriophage that transmits DNA from one cell to another. This process occurs due to the 
packaging of bacterial DNA during encapsulation of the viral genetic material prior to cell lysis and 
viral dissemination41,42. We can differentiate between three main transduction mechanisms; 
generalized43, specialized44 and lateral45 transduction. 

 
Figure 2. Horizontal gene transfer A) Graphic representation of the three main mechanisms of HGT: 
transformation, conjugation, and transduction. B) Example of the modular and hierarchical composition of 
mobile genetic elements (inter and intracellular). The model shows a plasmid that encodes a transposon 
which also encodes an integron that carries an ARG. 
 
1.1.6 Mobile Genetic Elements (MGEs). 

 
Except for natural transformation, HGT requires genetic elements such as plasmids or 

prophages that mediate gene transmission. These genetic carriers are denominated mobile 
genetic elements (MGEs). MGEs play a key role in the evolution of multirresistant bacteria due to 
intra and intercellular sharing of ARGs. We can differentiate between intercellular MGEs such as 
conjugative plasmids, integrative conjugative elements (ICEs), and prophages; and intracellular 
MGEs generally encoded in intercellular MGEs as non-conjugative transposons, insertion 
sequences and integrons (Figure 2B)46,47. 

Transduction

Conjugation
Transformation

Plasmid

Transposon

Integron

ARG

A B



Introduction  

34 

• Intercellular MGEs. 
 
Among all intercellular MGEs, conjugative elements (plasmids and ICEs), prophages and 

phage-plasmids (elements that are both phages and plasmids48) are the main vectors of ARGs 
that participate in multirresistances dissemination. Nevertheless, other MGEs, such as integrative 
mobilizable elements (IMEs), and phage satellites also contribute to HGT49.  

 
Plasmids can be defined as circular self-replicative extrachromosomal genetic elements. 

They usually present a modular anatomy. Conjugative plasmids also carry the essential machinery 
for conjugation, enabling their transfer to adjacent cells. Other conjugative elements, such as 
ICEs, resemble plasmids in their conjugative machinery and modular structure. However, ICEs 
encode their own enzymes to catalyse their insertion into host chromosomes, making them neither 
self-replicative nor extrachromosomal50. 

 
These intercellular MGEs can also carry platforms and genetic elements that are not able 

to move between cells per se but contribute to ARGs spread by collecting and transposing genes 
between genomic locations. Insertion sequences (ISs), transposons (Tns), and integrons are the 
main vectors of this intracellular HGT47.  

 
• Intracellular MGEs. 
 

Insertion sequences (ISs) are classically defined as short DNA sequences encoding 
exclusively the genetic information for their transposition and insertion in genetic locations 
surrounded by inverted repeats (IRs). Nevertheless, IS derived structures lacking the transposase 
gene or/and encoding accessory genes have also being reported. Examples of this IS derived 
structures are the miniature inverted repeats transposable elements (MITEs), the transporter IS 
(tIS) and mobile insertion cassettes (MICs)51. Moreover, two close ISs can mobilise genes 
encoded between them forming a composite transposon52. Transposons are generally associated 
with spread and dissemination of virulence, infection, and antimicrobial resistance genes. As an 
example, Tn3 transposon family is known to play a major role in antimicrobial resistance via 
mobilisation of several resistance determinants53.  

 
Furthermore, transposons and conjugative elements can carry additional structures known 

as integrons, which facilitate the mobilization, storage, and rearrangement of various resistance 
determinants. In fact, the discovery of multirresistant Shigella spp. isolates in Japan around 1960, 
which was regarded as the initial description of a strain resistant to multiple antimicrobial agents, 
has been conclusively associated with the existence of an integron encoding several ARGs22. Due 
to the relevance of this platform in antimicrobial resistance dissemination54, a detailed and 
extensive description of it is summarized in the following sections. Integrons and their role in 
antimicrobial resistance conform the main object of this thesis. 
 
 
 



   Introduction 
 

35 

1.2 Integrons. 
 
1.2.1 Structure and function of integrons. 
 

Integrons are genetic structures able to capture, accumulate and reorganise genes 
embedded in small mobile elements described as integron cassettes (ICs). The plasticity of this 
platform provides a plethora of functions to its host, allowing bacteria to evolve quickly and survive 
in challenging environmental conditions55. All integrons share the same structure with two well-
defined elements: a stable platform and a variable cassette array (Figure 3A).  

 
The platform contains the necessary elements for the acquisition and expression of ICs: 

the gene encoding the integrase (intI) and its promoter (Pint), the integron recombination site (attI 
site), and the cassette array promoter (Pc). The integrase belongs to the tyrosine-recombinase 
family and mediates the excision and reshuffling of ICs present in the array, and the insertion of 
external ICs in the primary integration site (attI), generally located upstream of the intI gene. 
Meanwhile, the Pc promoter, found within the intI gene or between the intI gene and the attI site, 
promotes the transcription of the cassettes stockpiled in the array in the opposite orientation to 
that of the integrase 56,57. 

 
Integrase expression is regulated by the SOS response; which can be triggered by several 

challenges such as starvation, HGT, DNA damage and the presence of antibiotics58. SOS 
response is mainly controlled by the transcriptional repressor LexA, which regulates intI gene 
repression by its binding to a conserved LexA-binding motif located at the Pint  region. Under 
stress conditions, RecA mediates LexA cleavage allowing the expression of intI, letting ICs 
recombination59. 

 
Integron cassettes are defined as non-replicative circular elements that usually encode a 

promoterless gene and a recombination site called attC site 60. attC sites are imperfect palindromic 
sequences of a variable lengths (ranging from 55 to 143 base pairs, according to IntegrAll 
database61) downstream the cassette coding region and play a central role in cassette 
recombination. Every acquired IC is inserted into the platform in the attI site, generating an array 
of cassettes via consecutive IC integrations. These ICs are transcribed from the Pc  promoter and 
therefore, only cassettes placed close to it will be expressed, remaining the distal ones silent62. 
Nevertheless, some ICs contain their own promoters affecting the polar expression of the array of 
cassettes. Examples of self-expressed ICs are toxin-antitoxin (TA) systems63, and some 
antimicrobial resistance cassettes (ARCs) such as cmlA, qacE, and ereA64–66 
 

Summarizing, under stress conditions the integrase is expressed modifying the array via 
excision and integration of ICs. The integrase can mediate the recombination of exogenous ICs 
in the first position of the array to modulate their expression and survive the external stressor. In 
consequence, the identity and location of ICs present in the array can vary depending on the 
circumstances, postulating the integron as an adaptive low-cost bacterial memory56.  

 
 



Introduction  

36 

Figure 3. Schematic representation of the mobilisation of ICs in integrons. A) Integrons are composed 
by a stable platform and a variable array of ICs (antimicrobial resistance cassettes (ARCs), gene cassettes 
of unknown function (gcu), etc). The platform contains the integrase gene, the Pc and Pint promoters, and 
the integron insertion site (attI). Under stress conditions, the integrase is expressed and it is able to excise 
and reshuffle IC from the platform, as well as to capture foreign ICs. The insertion of these IC in the first 
position of the array occurs through a reaction between the attI site and the cassette recombination site 
(attC). ICs are expressed from a Pc promoter following a gradient of expression from the first to the last IC 
of the array. B) Integrons are able to capture, exchange, and reshuffle integron cassettes (ICs) between 
mobile genetic elements and chromosomal integrons. Moreover, several mobile integrons are related to 
transposons that mobilize these platforms between plasmids and genomes. Integrons can be mobilized 
between cells through plasmid conjugation or transduction by bacteriophages67. 



   Introduction 
 

37 

1.2.2 Origin of integrons: chromosomal integrons. 
 

Integrons are ancient chromosomal structures present in 17% of the bacterial species 
sequenced genomes68, commonly found across multiple bacterial phyla. Furthermore, some 
studies even suggest their presence in archaeal genomes69. Comparative analyses of all integron 
integrases show a significative sequence homology grouping together within the tyrosine-
recombinase family. Moreover, there is a correlation between bacterial speciation and intI gene 
divergence, being integrases organised in genus-specific clades70. This fact suggests a main 
vertical transference of these platforms through punctual HGT events between bacteria sharing 
the same environments, dating the potential origin of sedentary chromosomal integrons (SCIs) 
hundreds of millions years ago.  

 
We can distinguish three major bacterial groups in relation to their chromosomal 

integrases: soil and freshwater proteobacteria, marine g-proteobacteria, and the inverted integrase 
group57. This last group is composed by bacteria of several taxonomic groups as Cyanobacteria, 
Spirochaetes or Planctomycetes, sharing a non-canonical integron structure with the integrase 
gene oriented towards the cassette array71.  

 
Among chromosomal integrons it is worth to highlight a subset denoted as superintegrons 

(SI). Superintegrons carry more than 20 ICs in their arrays with a high rate of identity in their 
cassette recombination sites (attC sites). SIs are especially prevalent in Vibrio species´ genomes; 
being Vibrio cholerae SI the paradigm of these structures72. Vibrio cholerae SI is located in 
chromosome two of this bacterium, covering the 3% of the entire bacterial genome. It contains 
179 ICs distributed between intergenic repeated sequences, that correspond to attC sites, initially 
known as Vibrio cholerae repeats (VCRs). SI ICs functions are in general not known with few 
exceptions like a chloramphenicol resistance gene (catB9) and 19 TA systems among others73. 
 
1.2.3 Mobile integrons: prevalence and clinical relevance. 
 

Sedentary chromosomal integrons (SCI) are not mobile by themselves, lacking the 
necessary machinery for their own integration and excision as a platform. Nevertheless, their 
association with transposable elements and HGT has mobilised integrons to a broader set of 
clinical bacteria via mobile integrons (MI) (Figure 3B)71. MIs can mobilize ARCs in arrays up to 11 
ICs, playing a central role in antimicrobial dissemination. 

 
Currently, we can distinguish five canonical classes of MIs based on the sequence of their 

integrases (intI1-5). In addition, each class is usually linked to a certain MGE; being integron 
classes 1 and 3 commonly associated with Tn402 transposon74,75, class 2 unequivocally linked to 
Tn7 derivates76, and classes 4 and 5 generally found in Vibrio spp. elements conferring resistance 
to trimethoprim and sulfamethoxazole77,78.  

 
 
 
 



Introduction  

38 

Class 1 integrons are by far the most prevalent MIs across databases. Approximately 96% 
of the classified MIs recorded in the IntegrAll61 database are identified as belonging to this 
particular class (Figure 4A). Class 1 MIs carry a vast pool of ARCs against 12 different antibiotic 
families and antiseptics. Due to their adaptive potential, class 1 integrons can be found virtually in 
all species of Gram-negative pathogens79,80. It is estimated that humans and animals shed to the 
environment 1023 copies of class 1 integrons per day81. As a result of integron propagation, 
genomes of human and animal pathogens are constantly connected with those of environmental 
bacteria, highlighting the importance of evaluating the antimicrobial resistance issue from a One 
Health perspective.  

 
It is estimated that 50% multirresistant uropathogenic E. coli harbour a class 1 integron, 

being highly prevalent in clinical isolates82. In this line, recent metagenomic analyses have 
demonstrated the high prevalence (42%) of integrons carrying ARCs in Klebsiella pneumoniae 
genomes, a closely surveyed pathogen by the WHO due to its high virulence and 
multirresistance83. 

 
 
Figure 4. Mobile integron prevalence and classification. A) Prevalence of each MI class across IntegrAll 
database based on the identity of the integrase detected within the integron (intI1-5). B) Occurrence of each 
Pc variant and Pc P2 combinations across class 1 MIs. Data collected from 84. 
 

Among class 1 MIs, we can find several Pc variants classified according to their strength 
and sequence homology with the s70 promoter (Figure 4B). PcS (strong) variant leads to the 
highest transcription rate in contrast with the PcW (weak) variant, which corresponds to the lowest 
rates, about 25-fold weaker than PcS. In between PcS and PcW we can find other hybrid promoter 
variants such as PcH1 and PcH2, as well as PcSS (super strong) being the strength of this variant 
still inconsistent between publications84. In addition, class 1 integrons can harbour a second P2 
promoter placed in the attI region that increases the strength of every Pc variant84–86. 

 
Summarizing, around 13 promoter variants and configurations have been already reported. 

These promoters are present in databases with different abundance, being  PcW (41.7%) the most 
prevalent, followed by PcH1 (28%), and PcS (24.3%) variants. (Figure 4B). Interestingly, because 
Pc is encoded within intI1, Pc polymorphism affects integrase sequence modifying its activity in 
an inverse manner, the weaker the promoter, the stronger the excision activity of the integrase84. 
 

A B

0.09%  Others

0.01%   intI5

0.12%  intI4

0.39%  intI3

3.72%  intI2

95.67%  intI1

intI1 96%

24.00%  PcS
0.30%  PcS P2
4.40%  PcH2
26.80%  PcH1
1.20%  PcH1 P2 
0.30%  PcSS
33.60%  PcW
8.10%  PcW P2
1.30%  Others

PcS
24%

PcH1
26,8%

PcW
33,6% intI1



   Introduction 
 

39 

1.2.4 Recombination system. 
 

As mentioned above, integrons are able to incorporate and excise several ICs via integrase 
mediated recombination. This unique recombination system requires the three main elements to 
accomplish its function: the integron attachment site (attI site), the cassette attachment site (attC 
site), and the integrase to catalyse the recombination reaction.  

 
• The attI site. 

 
The attI site is located upstream the intI gene and it is recognised by the integrase as a 

double stranded (ds) substrate where ICs are integrated. attI sites vary between integrons in line 
with integrase sequence diversification87. Nevertheless, integrases are able to recombine ICs into 
non-cognate attI sites, albeit less efficiently. As an example, intI1 can integrate ICs into attI2 and 
attI3 sites with recombination efficiencies 100 times lower than into attI1 site88.  

 
Despite sequence differences, attI sites are essentially composed by two integrase binding 

sites named L and R that constitute the core site (Figure 5). The recombination point is located in 
the R box, specifically in the 5´ GTT 3´ triplet, where the integrase is going to perform a cleavage 
in the complementary or bottom strand (bs) between the C and the A base pairs (bp). On the 
contrary, the L box and the central region between boxes varies widely across attI sites.  

 
Regarding the attI1 site, two imperfect direct repeats (DR1 and DR2) located upstream of 

the core site have been identified as additional binding domains for the integrase. The presence 
of these DRs is not essential for the recombination process, suggesting that their role is to attract 
two more integrase monomers in close proximity to the core site89–91. 

 
 
 
 
 
 
 
Figure 5. Sequence of a double stranded (ds) attI1 site. L and R boxes and DRs are indicated with grey 
arrows. GTT triplet is coloured in purple and cleavage point is indicated with a purple arrow. Integrase 
binding domains are marked with pink shadows. Modified from 92. 
 
• The attC site. 

 
attC sites conform the 3´end of ICs and are responsible for ICs mobility. The structure of 

these recombination sites is more complex than the one described for attI sites. attC site 
exceptional characteristics make integron recombination a unique process, differentiating it from 
the canonical tyrosine(Y)-recombinase performance56. 

 
 

5´ TTTGATGTTATGGAGCAGCAACGATGTTACGCAGCAGGGCAGTCGCCCTAAAACAAAGTTAGTGGATCC 3´
3´ AAACTACAATACCTCGTCGTTGCTACAATGCGTCGTCCCGTCAGCGGGATTTTGTTTCAATCACCTAGG 5´

R boxL boxDR1DR2

“core site”



Introduction  

40 

attC sites present in MIs can widely differ in sequence and length, from 55 to 143 bp. 
Nevertheless, all of them conserve the same palindromic organisation with two regions of inverted 
homology named R’’-L’’ and R’-L’ separated by a variable spacer (Figure 6)91. These palindromic 
sequences allow attC sites to fold, generating hairpin-like secondary structures that mimic a 
canonical recombination core site with L and R complete boxes93. Due to the formation of these 
hairpins, attC sites are recognized by the integrase as single-stranded (ss) substrates, in contrast 
to the attI site which is identified as a double stranded (ds) sequence. It is worth to mention that 
the bottom strand (bs) of every attC site is around 1000 times more recombinogenic than the top 
strand (ts)94. 

 
Figure 6. Sequence and structure of an attC site. A) Schematic representation of a double strand (ds) 
attC recombination site designating L’, L’’, R’ and R’’ boxes,  as well as the variable terminal structure (VTS). 
B) Sequence of aadA7 attC site specifying top and bottom strands (ts and bs). C) Proposed secondary 
structure of aadA7 attC site indicating the uncoupled central spacer (UCS), the extrahelical bases (EHB) 
and the variable terminal structure (VTS). L’, L’’, R’, R’’ boxes are indicated with grey arrows/lines while the 
GTT triplet and the cleavage point are coloured in purple. EHB are also pointed in orange. Integrase binding 
domains are marked with pink shadows. Modified from 92. 
 

Comparative studies between attC sites show a clear sequence conservation in two 
triplets, 5´ AAC 3´ and 5´ GTT 3´, located in R’’ and R’ regions, being the cleavage point located 
between C and A bps as in attI sites. In addition, it is possible to extend the conserved region up 
to 7 nucleotides conforming the inverse core and the core (Figure 6A, 6B). These conserved 
regions consist on the complementary sequences 5´ RYYYAAC 3´and 5´ GTTYYYR 3´ where R 
stands for a purine and Y represents a pyrimidine56.  

 
 

A

B

5´ ATGTCTAACAATTCATTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAATTCAAGCGTTAGACAT 3´attC aadA7 ts
3´ TACAGATTGTTAAGTAAGTTCGGCTGCGGCGAAGCGCCGCGCCGAATTAAGTTCGCAATCTGTA 5´attC aadA7 bs

R’R” L” L’

5´ --RYYYAAC--------------- VTS ----------------GTTRRRY 3´
3´ --YRRRTTG--------------- 20 to 104 bp ----------------CAAYYYR 5´

R’R” L” L’

attC site ds

C

“core”“inverse core”

C  GA
5´---GTCTAACG  TT ATTAAGCCGCGCCGC
3´---CAGATTGT  AA  TAATTCGGCGCGGCG

T  G   G T

G

A
A attC aadA7 bs folded

R box L box

UCS VTSEHB



   Introduction 
 

41 

attC site folding adds an extra layer of information and regulation to the recombination 
process (Figure 6C). The presence of single bases in the R’’-L’’ region with no complementarity 
in the R’ - L’ region give rise to extrahelical bases (EHBs) in the folded structure. EHBs have an 
important role determining the recombinogenic strand (bs), stabilising the synaptic complex and 
avoiding a second cut of the integrase in the L box. Other structures that take part in the correct 
attC site folding are the variable terminal structure (VTS), located at the end of the stem; and the 
uncouple central spacer (UCS), which is the region between R’’-R’ and L’’-L’  boxes, also 
participating in the stabilization of the synaptic complex95–98. 
 
• The integrase. 
 

Integron integrases are members of the Y-recombinases family, with Xer recombinases as 
their closest relatives99,100. Consequently, they share some characteristics such as the same active 
site residues RKHRHY101. In contrast, integrases possess an extra domain named I2 domain that 
facilitates their unique recombination activity. I2 domain consists in 19 amino acids forming a a-
helix structure which is not present in any other Y-recombinases102. This domain plays a crucial 
role stabilising the synaptic complex and accommodating the EHBs present in attC sites97. 
 

Notably, integrases are able to recognise different substrates in terms of sequence and 
structure; attI sites are dsDNA sequences while attC sites are ssDNA hairpins with little sequence 
homology. This dual-recognition implies a fined-tuned regulation of the synaptic complex and the 
interactions between monomers during recombination103. 
 
 
1.2.5 Recombination reactions. 
 

Canonical recombination mediated by Y-recombinases follows a very conserved process. 
Initially, four monomers of the protein are assembled to both DNA substrates forming the synaptic 
complex. The process continues with a first strand exchange between DNA molecules, forming a 
transient structure called Holliday Junction (HJ), which is finally resolved through a second strand 
exchange101.  

 
Nevertheless, integrases perform a different recombination process by reason of the 

structural peculiarities of the attI and attC sites. Three different integrase-mediated recombination 
reactions can occur between attI and attC sites: the attC x attI reaction, which leads to IC insertion; 
the attC x attC, responsible for IC excision from the array; and the attI x attI reaction, potentially 
implicated in the rearrangement of ICs arrays located in different integrons, although 1000 times 
less efficient than the previous ones56. 

 
 As a result of their participation in shuffling ICs, attC x attI and attC x attC reactions will 

be described in detail below. 
 
 
 



Introduction  

42 

• attC x attI: IC insertion 
 

The insertion of new ICs in the array occurs through attCbs x attI recombination (Figure 7). 
This reaction starts as a canonical recombination up to the formation of the HJ. attC sites are 
recognised as ssDNA substrates so the resolution of this atypical HJ (aHJ) by  the classical second 
strand exchange would generate a linear DNA product and therefore, an abortive reaction. attC 
site EHBs impede this second cut of the integrase, resolving the aHJ by replication. Consequently, 
IC insertion is a semiconservative process producing the initial attI-containing substrate and the 
insertion of the IC97,104. As mentioned before, the cleavage previous to the strand transfer happens 
between the C and AA nucleotides in both substrates. This cut implies the formation of “chimeric” 
attI and attC sites where the last 6 bp of R boxes belong to the other substrate.  

Figure 7. Recombination reaction between attC bs and attI sites. An atypical Holliday Junction (aHJ) is 
formed after the first strand exchange between attI site (coloured in red) and the bottom strand of the attC 
site (coloured in green). The aHJ is resolved by replication (pointed blue arrows) producing the IC insertion 
and the initial attI-containing substrate. Integrase monomers are represented in purple. Modified from 56. 
 
• attC x attC: IC excision 

 
The recombination reaction between two attC sites leads to the excision of a IC (Figure 8). 

Both attC sites have to be folded simultaneously to allow the first strand exchange performed by 
the integrase, generating  an aHJ, as happens in the attI x attC reaction. In this case, a second 
cut of the integrase would lead to an attC site exchange between ICs so the aHJ is also resolved 
by replication60,105. This semiconservative mechanism generates as products the excised IC, the 
original substrate and the substrate lacking the IC excised. 

Figure 8. Recombination reaction between two attC bs sites. The integrase recognise the bottom strand 
of two folded attC sites (coloured in orange and green) and cleaves provoking the strand transfer and aHJ 
formation. The aHJ is resolved by replication (pointed blue arrows) producing the excision of the IC, as well 
as releasing the original substrate and the substrate lacking the IC excised. Integrase monomers are 
represented in purple. Modified from 56. 

attC-containing 
substrate

ss attC bs 

cleavage

OH

OH

strand
transfer

aHJ
formation

replication products

attC1/2 attC2/3

attC1/2 attC2/3

attC1/3

substrate

IC2 excised

attC2

IC2

ds attI

ss attC bs

GC

attI- containing
substrate

cleavage

O
H

O
H

strand
transfer

aHJ
formation

replication products

ds attI

attI/attC at
tC
/a
ttI

substrate

IC
inserted

IC



   Introduction 
 

43 

1.2.6 Integron cassettes. 
 

Integrons carry a pool of more than a thousand different ICs71 recovered from virtually 
every environmental source and condition106. These ICs generally contain a promoterless open 
reading frame (ORF) followed by the attC site needed for its recombination in the platform. The 
size of integron ICs varies depending on the contained ORF generally falling within the range of 
500 to 1500 bp.  

 
Functions encoded in most ICs are still cryptic and vary depending on their presence in 

CIs or MIs. CIs harbour a diverse pool of ICs of which only the function of 25% of them is known. 
This 25% of ICs encode genes with roles in DNA modification, virulence, TA systems, and phage-
related functions71. In contrast, gene cassettes of unknown function (gcus) represent just the 11% 
of the total ICs present in MIs, being the remaining 89% involved in antimicrobial resistance 
(Figure 9A).  
 
 

 
Figure 9. IC occurrence and distribution in MIs. A) Occurrence of ICs of unknown function (gcu) and 
antimicrobial resistance cassettes (ARCs) in class 1 MIs (data collected from IntegrAll database). B) 
Distribution of ARCs found in MIs in the IntegrAll database. (QACs: quaternary ammonium compounds; Cm: 
chloramphenicol). Modified from 107. 
 

The fact that most MI ICs are involved in antimicrobial resistance is probably a 
consequence of the selective pressure exerted within the clinical settings. To date, the IntegrAll 
database records approximately 200 distinct ARC alleles within MIs. However, when a 95% 
threshold for nucleotide sequence identity is implemented, the count of unique ARCs decreases 
to 177 cassettes107. These ARCs confer resistance against 13 different antibiotic families (Figure 
9B), including the most clinically relevant compounds like aminoglycosides, beta-lactams, 
sulphonamides, macrolides, phenicols, fosfomycin and quaternary ammonium compounds 
(QACs)107,108. Interestingly, no ARCs conferring resistance against tetracyclines have being found 
in MIs. 
 

 
 

A B 

intI1 ARCs

gcu

11%

89%



 
 

 
  
 

 

 
 

 



 

 
 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

OBJECTIVES 
 
 

 
 
 
  

Scan me on Spotify 



 
 

 
  
 

 

 



Objectives 

 47 

2 Objectives 
 
Antibiotic resistance stands as one of the most significant health challenges of the 21st 

century. The development of bacterial resistance is facilitated by the spread of resistance factors 
through horizontal gene transfer (HGT), which facilitates the genetic connection between 
unrelated bacterial genomes. Several genetic platforms such as plasmids or transposons can 
mobilize genes. During the rise of multidrug resistance, integrons carrying multiple antimicrobial 
resistance genes were found associated to both structures, circulating among clinical isolates. 
Mobile integrons (MIs) carry around 200 antimicrobial resistance cassettes (ARCs) against most 
antibiotic families and are hence major drivers of multidrug resistance. These genetic platforms 
are successful in the clinical setting for the adaptive value of their cassettes. Yet, marked 
differences in the abundance of individual cassettes suggest that each one has distinct 
evolutionary dynamics. 

 
Thus, the main objectives of this PhD Thesis are to characterize and quantitate the 

forces driving the evolutionary success of each ARC in integrons (namely: resistance levels, 
fitness cost, polar effects, and cassette recombination frequency), as well as to widen the 
understanding of the integron platform mechanisms. To accomplish the main objective of this PhD 
Thesis, a deeper look into the integron model is needed as well as the implementation of  the next 
specific tasks: 

 
1. Generate a comparative and comprehensive collection of strains bearing specific 

ARCs to characterize them on their native environment. 
 

2. Determine the resistance levels conferred by each ARC. 
 

3. Evaluate the influence of each ARC in the expression of downstream ARCs in the 
array, also known as polar effects. 

 
4. Assess the fitness cost imposed by individual ARCs to the host. 

 
5. Measure the capacity of single ARCs to be integrated or excised from the integron. 

 
6. Revisit the literate to give light to controversial topics related with integron 

regulation. 
 
 



 
 

 
  
 

 

  



 

 
 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

MATERIALS  
AND METHODS 

 
 

 
 
 
  

Scan me on Spotify 



 
 

 
  
 

 

 



Materials and Methods 

 51 

 

3 Materials and Methods  
 

3.1 Bacterial Strains, Plasmids, and Culture Conditions. 
 

Escherichia coli MG1655 and DH5a were used as recipients for all plasmid constructions 
in this study, unless otherwise stated (Supplementary Table S1). Bacterial strains were cultured 
at 37ºC in Müller Hinton (MH; Oxoid, UK) or lysogeny broth (LB; Oxoid, UK)  liquid media and 
MH/LB agar (1.5%) solid media (BD, France). Zeocin (Invivogen, USA) was added at 100 µg/mL 
to maintain pMBA plasmid collection in E. coli. Liquid cultures were incubated in an Infors Multitron 
shaker at 200 rpm (Infors HT, Switzerland). 
 
3.2 Generation of the pMBA collection and derivatives. 
 

Most plasmids used in this study (Supplementary Table S1) derive from pMBA107; a vector 
that mimics the genetic environment of a class I integron with a cassette of interest in the first 
position of the array. Antimicrobial Resistance Cassettes (ARCs) were synthesized in vitro (IDT, 
US) adding homology regions with pMBA vector in both 5´and 3´ends. To generate pMBA 
collection we cloned each ARC in the first position of the array (attI site) using Gibson Assembly 
(Figure 10)109. 

 
Briefly, pMBA vector was linearized and amplified by Polymerase Chain Reaction (PCR) 

using Int R bb and GFP F bb primers while each ARC was amplified by PCR using gBlock F and 
gBlock R primers (Supplementary Table S2). Amplifications were performed using a PCR ProFlex 
Thermocycler (Applied Biosystems, US) and 2x Phusion Green HSII High-Fidelity PCR MasterMix 
(Thermo Scientific, US) following manufacturer´s instructions. PCR thermocycling conditions 
contained an initial stage of 30 seconds at 98°C followed by 30 cycles of amplification (10 seconds 
at 98°C, 15 seconds at 55°C, and 15 seconds/kb. amplified at 72ºC), with a final stage of 10 min 
at 72ºC. PCR products were run by agarose (Biotools, Spain) gel (1%) electrophoresis and 
visualized using a GelDoc apparatus (BioRad). Gibson assembly reaction was performed in a final 
volume of 4 µl containing 1 µl of the linearised vector, 1 µl of the insert (each AR cassette), and 2 
mL of the 2X Gibson Assembly Buffer (5X ISOBuffer, 10.000, 147 u/mL T5 exonuclease, 2,000 
u/mL Phusion polymerase, 40,000 u/mL Taq ligase, ddH2O). The mix was incubated for 30 
minutes at 50 ºC before being transformed into E. coli MG1655 competent cells.  
 

Figure 10. Schematic representation of the cloning region in pMBA derivatives. Each ARC was cloned 
in its native environment in the first position of the array (integron attachment site or attI site) and followed 
by a GFP gene. The primers used both for backbone and ARCs amplifications are represented in orange. 



Materials and Methods 

 52 

Competent cells used in this study were either chemo-competent or electro-competent. 
Overnight culture of the strain of interest was diluted 1/100 in 100 mL of fresh MH broth and 
incubated at 37ºC with shaking until reaching optical density (OD600) 0.6 for chemo-competent 
cells and OD600 1 for electro-competent cells. Cells were then placed in ice and washed twice 
(centrifuge 6000 rpm, 10 min, 4ºC and resuspended again) using 10 mL of CaCl (0,1 M) or 10% 
glycerol (Sigma, US) autoclaved ddH20 for acquiring chemo or electro-competence respectively. 
Cells were finally resuspended in 5 mL of 15% glycerol CaCl (0,1 M) (chemo) or 300 µl of 10% 
glycerol autoclaved ddH20 (electro), before aliquoting and freezing at -80ºC. Chemo-competent 
cells were transformed by thermal shock via 45 seconds incubation at 42ºC, while an Eporator 
(Eppendorf, Germany) was needed to transform electro-competent cells (2 mm electroporation 
cuvettes (Molecular Bioproducts, US), 2500V).  After transformation, cells were resuspended in 1 
mL of fresh MH broth and incubated at 37ºC with shaking for an hour prior to plating in MH agar 
supplemented with selective antibiotics. 
 

Primers used to generate pMBA derivatives and mutants are listed in Supplementary Table 
S2. All construction sequences were verified after transformation by Sanger sequencing 
(Macrogen, South Korea) of the region of interest (from Pc promoter to the end of gfp gene) using 
the following primers: Int F, GFP R and GFP 2.0 R (Supplementary Table S2).  
 
3.3 Antimicrobial resistance characterization by agar diffusion test. 
 

We assessed the antimicrobial resistance of each strain in pMBA collection using disc 
diffusion tests in MH-agar broth. Overnight cultures of pMBA-derived strains were adjusted to a 
0.5 in the McFarland scale using filtered saline solution (NaCl 0.9%). The resulting solutions were 
diluted 1/200 and plated on MH agar plates by inundation using approximately 3 mL of solution 
for 3 minutes. After disposing any excess of inoculum and allowing the plates to dry, antibiotic 
discs (Supplementary Table S3) (Oxoid, BioRad) were deposited on top of the agar. Plates were 
incubated overnight at 37ºC. The resulting inhibitory halos around the antibiotic discs were 
measured to characterise the antibiotic susceptibility of each strain.  
 
3.4 Minimal inhibitory concentration (MIC) determination. 

 
MIC determination, defined as the minimal concentration at which bacterial growth was 

inhibited , was performed according to guidelines set by the Clinical and Laboratory Standards 
Institute (CLSI)110. To summarize, 105 colony-forming units (CFUs) were inoculated in 200 µl of 
fresh MH with doubling dilutions of each selected antibiotic. The 96 well-plates (Nunc, US) were 
then incubated overnight at 37ºC in static conditions. We considered the MIC as the media of at 
least three biological replicates for each antibiotic and strain. Antimicrobials tested in MIC 
determination are listed in  Supplementary Table S4. 
 
 
 
 
 



Materials and Methods 

 53 

3.5 Quantitation of GFP expression by flow cytometry. 
 

To determine Green Fluorescent Protein (GFP) expression, three independent colonies of 
each pMBA derived strain were inoculated in MH/LB zeocin and incubated at 37ºC overnight. 
Cultures were then diluted 1/400 in filtered saline solution to measure fluorescence using a 
Cytoflex-S flow cytometer (Beckman Coulter) for stationary phase measurements. For exponential 
phase measurements, overnight cultures were diluted 1/100 in MH supplemented with zeocin and 
incubated at 37ºC with shaking for 2 hours prior to measure fluorescence diluting samples 1/20 in 
saline solution. The 488 nm laser was used to detect gfp expression through 525/40 nm (FITC) 
band pass filter. 20,000 events were recorded per sample. Data were analysed using CytExpert 
software (v.2.4; Beckman Coulter). 
 
3.6 Quantification of gfp translation by Western blot. 
 

Specific strains were selected for relative GFP protein quantitation through Western blot. 
Western blot protocol was followed as described in Hipólito et al, 2022. Briefly, strains were 
incubated in MH at 37ºC overnight, one mL of each culture was collected, pelleted and 
resuspended in 50 µl of Laemli buffer (BioRad, US) with 20% b-mercaptoethanol (BioRad).  The 
solutions were boiled, and 10 µl of the supernatants were separated by electrophoresis (30 min 
200 V) using 4–12% gradient polyacrylamide precast gels (Invitrogen, USA). Proteins were then 
transferred to nitrocellulose membrane using iBlot2 system (Invitrogen). Membranes were blocked 
using a solution containing 2% BSA and 0.2% Tween-20 TBS  (BSA TBS-T) for 1 hour, followed 
by a 2-hour incubation with primary antibodies diluted in fresh 2% BSA TBS-T (mouse a-GFP 
1/50,000  (Invitrogen) and mouse a-DnaK 1/6.000 (Enzo Life Sciences, US)). Next, the 
membranes were washed thrice and incubated for 1 hour with the secondary antibody HRP-
labelled goat anti-mouse IgG (Invitrogen) diluted 1/10,000 in fresh 2% BSA TBS-T. Finally, the 
membranes were developed using SuperSignal West Pico Plus Chemiluminescent Substrate and 
ChemiDoc XRS+ imaging system Data were analysed with ImageLab software (v.6.0.1, BioRad) 
to quantitate band intensities. 

 
3.7 Quantification of gfp transcription by RT-qPCR. 
 

Reverse transcription qPCR was performed to quantitate gfp differential transcription in 
each pMBA derivative. Three biological replicates of each strain were diluted 1/100 in MH broth 
supplemented with zeocin and grown at 37°C with shaking until reaching exponential phase 
(OD600 = 0.6-0.8). Two mL of each culture were pelleted and resuspended in 50 µl of lysozyme 3 
mg/mL (Thermo Scientific) prior to freezing at -20ºC. RNA extraction was performed using the 
RNeasy Mini Kit (QIAGEN), following the manufacturer’s protocol. Residual DNA was removed 
using the TURBO DNA-freeTM Kit (Invitrogen). RNA concentration and integrity were measured 
using a BioSpectrometer (Eppendorf). For cDNA synthesis, 1 µg of RNA was used for reverse 
transcription using the QuantiTect Reverse Transcription Kit (Qiagen, Germany) according to 
manufacturer’s instructions.  
 



Materials and Methods 

 54 

cDNA was diluted 1/2000 and 1µL of this dilution was used for quantitative PCR (qPCR). 
qPCR was performed in QuantStudio 3 Real-Time PCR system (Applied Biosystems) with the 
Sybr Green Fast MasterMix Kit (Applied Biosystems) according to manufacturer’s instructions. 
Primers used in the mix are listed in Supplementary Table S2. qPCR reactions were previously 
optimised, with reaction efficiencies within the range of 90-110%. Each PCR reaction contained 
400 μM of each primer, Sybr Green Fast MasterMix 2x, and 1 μL of template cDNA. PCR 
thermocycling conditions contained an initial 10 min at 95 °C step, followed by 40 cycles of 15 s 
at 95ºC and 1 min at 60ºC. Fluorescence for each cycle was recorded after the step at 60 °C. 
Design and Analysis software version 2.3.3 (Applied biosystems) automatically determined 
quantification cycle (Cq) and baseline. Negative controls included both a reaction containing water 
instead of template and a reverse transcriptase-free reaction. For data analysis, the relative 
abundance of gfp transcripts was normalized to that of the housekeeping genes rssA and rpoA. 
 
3.8 In vitro competition assays by flow cytometry. 
 

To assess the cost of each ARC, we performed in vitro competition assays following the 
protocol published in DeLaFuente et al, 2020111 (Figure 11). In short, we competed strains 
containing different ARCs and a strain harbouring the empty vector pMBA with the GFP knockout 
(GFP KO). This allowed us to differentiate both population using flow cytometry. Measuring the 
initial and final proportions of each population along the experiment permitted us to determine the 
fitness cost associated to each ARC.  

 
At least 6 biological replicates of each competition were performed per condition. 

Precultures were incubated overnight in MH broth without any selective pressure at 37ºC with 
shaking. 1:1 mix of both competitor strains was performed and diluted 1/400 in filtered saline 
solution to determine the initial distribution of each population. Measurements were performed 
using a Cytoflex-S flow cytometer (Beckman Coulter, US), as described above. The mixture was 
also diluted 1/400 in fresh MH broth and incubated at 37ºC with shaking for 22 hours. After 22 h 
of incubation, final proportions were quantitated as described previously. The fitness of each 
pMBA derivative strain relative to the parental strain (pMBA KO) was calculated using the following 
formula: 

 
 
 
 
In the formula, wpMBA ARC corresponds with the relative fitness of each pMBA derived strain 

(pMBA ARC) in comparison with the strain harbouring the empty vector with the GFP knockout 
(pMBA GFO KO). Ni and Nf refer to the number of cells at the beginning and the end of each 
competitor along the experiment.  

𝑤!"#$	$&' = 	
ln	(𝑁(,	!"#$	$&' 𝑁*,	!"#$	$&'⁄ )

ln	(𝑁(,	!"#$	+,-	./ 𝑁*,	!"#$	+,-	./⁄ )
 



Materials and Methods 

 55 

 
Figure 11. Flow diagram of the experimental process. Distinguishable bacterial cultures were mixed in 
1:1 ratio prior to measure their initial populations by flow cytometry. The mixture was diluted and incubated 
for 22 h at 37 ºC with shaking. Lastly, final populations were measured by flow cytometry again. 
 

When competitions were performed under anaerobic conditions, 96-well plates were 
incubated inside hermetic recipients where anaerobe container system sachets (BD, US) were 
deposited to generate the anaerobic atmosphere. The fact that GFP needs oxygen to fluoresce, 
forced us to expose cells to oxygen 2h with shaking when diluted 1/400 in filtered saline solution 
before measurements.  
 
3.9 OMM12 stocks preparation. 
 

The Oligo Mouse Microbiota (OMM12) is a synthetic bacterial community comprising twelve 
different bacterial species (Enterococcus faecalis KB1 (DSM32036), Bifidobacterium animalis YL2 
(DSM26074), Acutalibacter muris KB18 (DSM26090), Muribaculum intestinale YL27 (DSM28989), 
Flavonifractor plautii YL31 (DSM26117), Enterocloster clostridioformis YL32 (DSM26114), 
Akkermansia muciniphila YL44 (DSM26127), Turicimonas muris YL45 (DSM26109), Clostridium 
innocuum I46 (DSM26113), Bacteroides caecimuris I48 (DSM26085), Limosilactobacillus reuteri 
I49 (DSM32035) and Blautia coccoides YL58 (DSM26115)). The OMM12 are able to colonize the 
murine intestinal system and serve as a model of bacterial mouse gut consortia as its members 
represent the 5 mayor eubacterial phyla present in the gut112.  

 
OMM12 stocks used in competitions assays in vitro and in vivo were prepared from 

individual monoculture inocula. Monocultures were grown from frozen glycerol stocks under strict 
anaerobic conditions at 37ºC in 10 mL culture flasks (flask T25, Sarstedt, Germany) filled with 
anaerobic medium (18 g/L  glucose free brain-heart infusion (Oxoid), 15 g/L glucose free trypticase 
soy broth (USBiological), 5 g/L yeast extract, 2.5 g/L K2HPO4, 1 mg/L haemin, 0.5 mg/L 
menadione, 3% heat-inactivated fetal calf serum, 0.25 g/L cysteine-HCl‧H2O). Monocultures were 
diluted 1/10 in fresh anaerobic medium and incubated again for 24 hours. Prior to OMM12 
generation, monocultures were checked for contaminations performing Gram stainings.  

Mix 1:1

Ti

Incubate 22 h

Tf

22 h

Growth rates

22 h

Growth rates

pMBA ARCpMBA GFP KO



Materials and Methods 

 56 

After checking for contaminations, monocultures were diluted to an OD600 of 0,1 in fresh 
AMM medium and mixed in equal proportions. Final OMM12 stock was then aliquoted and frozen 
at -80ºC in anaerobic vials with glycerol (final concentration 10%). A 500 µL aliquot was collected 
to quantify the relative abundance of each member of the OMM12 consortia in the final mixture via 
qPCR. 
 
3.10 In vitro competition assays by phenotypic selection. 
 

Bacterial fitness in the presence of each ARC was also assessed by phenotypic selection 
in different environments and conditions. In these experiments we competed pMBA derived strains 
without GFP (pMBA ARC DGFP) against a strain harbouring the empty vector without the GFP 
(pMBA DGFP). At least three independent biological replicates were performed per competition 
and condition. We performed three independent competition experiments selecting by phenotype: 
in the presence of oxygen, in the absence of oxygen, and in the presence of OMM12

 bacterial 
consortia under anaerobic conditions. 

 
Aerobic competitions were performed at 37 ºC with shaking. Precultures of pMBA ARC 

DGFP and pMBA DGFP were grown overnight in MH broth. Then, 106 CFUs of each culture were 
mixed in 5 mL of MH broth without selective pressure and incubated overnight. A total of 2x106 
CFU were transferred to  5 mL of fresh MH broth every 24 hours for 5 days. Samples were taken 
every day, diluted 100.000 times and plated in MH agar plates. The proportion of resistant colonies 
was inferred by replica plating of 50 colonies on MH agar plates with specific selective pressure 
depending the ARC encoded in the vector (Supplementary Table S5). 

 
Anaerobic competitions were performed under a specific gas atmosphere (7% H2, 10% 

CO2, 83% N2). Precultures were grown overnight in 24-well plates (Nunc) and mixed adding 0.1 
mL of each culture (OD600 = 0.1) to a final volume of 1 mL of fresh MH broth. After 24h incubation 
each mixture was diluted 1/10 in fresh MH every 24 hours for 5 days. Plating was performed as 
stated in the previous paragraph.  

 
Anaerobic competitions in the presence of OMM12 consortia were performed following the 

previous protocol with an exception: initial proportions in the mixture were 10:1, adding 100 µL of 
OMM12 stock and 10 µl of each competitor to 900 µL of fresh M10 broth (18 g/L  glucose free 
brain-heart infusion (Oxoid), 15 g/L glucose free trypticase soy broth (USBiological, US), 5 g/L 
yeast extract, 2.5 g/L K2HPO4, 1 mg/L haemin, 0.5 mg/L menadione, 3% heat-inactivated fetal 
calf serum, 0.25 g/L cysteine-HCl‧H2O, 0.025% mucin, 0.5 g/L of fucose, xylose, arabinose, 
rhamnose and lyxose, 2 g/L of xylan from beechwood (Roth, Germany)). Moreover; 500 µL of day 
5 mixed cultures were collected to quantitate the relative abundance of each OMM12 member. 
 
 
 
 
 



Materials and Methods 

 57 

3.11 In vivo competition assays. 
 

In vivo competitions were performed using C57B1/6 mice stably colonized with OMM12 

defined bacterial consortia. Mice were housed under germfree conditions in IsoCage P systems 
(Tecniplast, Italy). Mice were supplied with autoclaved ddH2O and Mouse-breeding complete food 
for mice (Ssniff, Germany) ad libitum. All mice were males between 10-15 weeks. They were 
randomly assigned to experimental groups and kept in groups of 2-4 individuals per cage during 
the experiment. Animal health was scored twice a day along the experiment. 

 
Previous to bacterial inoculation, strains were cultured in MH broth for 12 hours at 37ºC on 

a wheel rotor and mixed 1:1 (pMBA DGFP : pMBA ARC DGFP). Mice were inoculated with mixing 
cultures by gavage (100 µL on the fur and 50 µL orally). Faecal samples were taken every 24 
hours for the first 4 days and at the end of the experiment at day 7 for plating and qPCR analysis. 
All mice were sacrificed by cervical dislocation at day 7 before collecting their caecal content.  

 
E. coli loads in faeces and caecal contents were determined by plating on MacConkey 

agar (Oxoid) supplemented with vancomycin (7,5 µg/mL), or with vancomycin and zeocin to 
determine E. coli pMBA loads. The proportion of resistant colonies was inferred by replica plating 
of 50 colonies per sample on MacConkey agar plates with specific selective pressure depending 
the ARC encoded in the vector (Supplementary Table S5). 

 
3.12 gDNA extraction from faecal pellets, caecal samples and in vitro competitions. 

 
Genomic DNA (gDNA) was extracted from in vitro and in vivo competitions where OMM12 

community was present. The protocol used was stablished based on Turnbaugh et at., 2009 and 
Ubeda et al., 2012 as references113,114.  
 

Faecal, caecal or in vitro competition samples were thawed and resuspended in 500 µL of 
extraction buffer (200 mM Tris (pH 8.0), 200 mM NaCl, 20 mM EDTA), 210 mL of 20% SDS, 500 
µl phenol:chloroform:isoamylalcohol (25:24:1, pH 7.9, Roth) and 500 µL of 0.1 mm-diameter 
zirconia/silica beads (BioSpec Products, Bartlesville, OK, Roth). Bacteria were lysed using a bead 
beater (TissueLyser LT, Qiagen) at max speed (50 s-1) for 4 min and centrifuged 5 min at full 
speed (14,000 x g). Supernatants were collected without taking phenol from the lower phase and 
transferred to a new tube containing 500 µL of phenol:chloroform:isoamylalcohol. Samples were 
mixed and centrifuged again, collecting the upper phase of the supernatant. This phase was then 
mixed by inversion with 1 mL of 96 % ethanol supplemented with 50 µl Sodium Acetate 3M to 
precipitate gDNA. After 30 min centrifugation (14,000 x g), at 4 °C; pellets were washed with 500 
µl 70% ice-cold ethanol and centrifuged again 15 min (14,000 x g), at 4 °C. gDNA pellets were air 
dried and resuspended in 100 µL of TE buffer. Finally gDNA samples were purified using a 
NucleoSpin gDNA clean-up kit (Macherey-Nagel, Germany), following manufacturer´s 
instructions. 

 
 
 



Materials and Methods 

 58 

3.13 Quantification of OMM12 microbiota members and E. coli abundance by qPCR. 
 

qPCR of bacterial 16S rRNA genes was performed following Brugiroux et al, 2016 
protocol112 to evaluate relative abundance of OMM12 members and E. coli along in vitro and in vivo 
competition assays. Primers and probes used to this purposed are listed in Supplementary Table 
S2. qPCR reactions were previously optimised, with reaction efficiencies within the range of 90-
110%. qPCR reactions with gDNA templates extracted from in vitro competitions, faeces, or caecal 
content were run in duplicate in a LightCycler96 Thermocycler (Roche, Switzerland). Each PCR 
reaction contained 30 μM of each primer, 25 μM of each probe, FastStart Essential DNA Probes 
MasterMix (Roche) and 5 ng template gDNA. PCR thermocycling conditions contained an initial 
10 min at 95 °C step, followed by 45 cycles of 15 s at 95ºC and 1 min at 60ºC. Fluorescence for 
each cycle was recorded after the step at 60 °C. LightCycler96 software version 1.1 (Roche) 
automatically determined quantification cycle (Cq) and baseline. 
 
3.14 RNA-Seq analysis. 
 

Total RNA was extracted from E.coli MG1655, pMBA and pMBA derivatives to assess 
differences in their transcriptomes as stated before. The rRNA depletion and subsequent RNA-
Seq library and sequencing was carried out by Oxford Genomics Centre using Illumina´s 
NovaSeq6000 sequencing platform. Transcriptomic data processing consisted in a first trimming 
step using Trim galore v0.6.6 (https://github.com/FelixKrueger/TrimGalore). In this step, a 
trimming quality threshold of 20 was used and adapters and reads shorter than 50 bp were 
removed. Next, trimmed reads were mapped to E. coli MG1655 reference genome (Accession 
number: NC_000913.3) using BWA-MEM v0.7.17115. FeatureCounts from the Rsubread v2.10.2 
package was then used to obtained read counts116. Differential expression analysis was performed 
from raw count data using DESeq2, version 1.36.0117 using as threshold parameters log2fold 
change > 1 and padj < 0.05. 
 
3.15 Motility quantitation by swarming assays. 

 
To determine motility in pMBA derived strains we performed swarming assays. 

Independent colonies were cultured in MH medium at 37ºC overnight. Swarming media (TSB 10 
g/L, NaCl 5 g/L, Agar 2.5 g/L) was prepared and poured into squared Petri dishes plates (Deltalab, 
Spain), letting them dry for 25 min before inoculation. Three microlitres of the overnight cultures 
were inoculated inside swarming media and incubated at 37ºC for 12 hours. Anaerobic swarming 
assays were performed following the same protocol with some modifications. Agar plates were 
placed inside hermetic recipients where anaerobe container system sachets (BD) were deposited 
to generate the anaerobic atmosphere. Then, these recipients were incubated at 37ºC for 24 
hours. At least 6 biological replicates were used for each strain and condition. Motility was then 
quantitated by measuring the halo diameter around the inoculation point. Images were taken using  
a GelDoc apparatus using epilight (BioRad). 
 
 
 

https://github.com/FelixKrueger/TrimGalore


Materials and Methods 

 59 

3.16 Recombination assays. 
 

We performed recombination assays to compare attI x attC recombination rates depending 
on the ARC recombined. The purpose of this test is to introduce a recombination substrate (attI 
site in our experiments) by conjugation into a recipient bacteria that express IntI1 integrase and 
already possesses the other recombination substrate (each attC site in our experiments). The attI 
site delivered by conjugation is encoded in a suicide vector from the R6K-based pSW family118, 
while the recipient strain harbours a p3938 plasmid that encodes an IntI1 integrase under the 
control of a PBAD promoter, and each pMBA derived plasmids encoding the recombinogenic attC 
sites. E. coli b2163 (dapA-, pir+) was used as donor strain as it can sustain pSW replication by the 
expression of the pir gene, although it requires DAP to grow. On the other hand, E. coli DH5a was 
selected as recipient strain. Donor and recipient strains used in recombination assays are 
described in Supplementary Table S1. 

 
We mimicked the cassette insertion process following the protocol stated in Biskri et al, 

2005; with some modifications119. Overnight cultures of donor and recipient strains were diluted 
1/100 in 200 µL of fresh MH broth supplemented with arabinose 20% and DAP 0,3 mM, and 
incubated in 96-well plates at 37ºC with shaking for 3h reaching an OD600 of 0,6. Strains were 
mixed 1:4 (40 µL donor : 160 µL recipient)  and pelleted by centrifugation (3 min. 4.400 rpm). 160 
µL of supernatant was removed and the mixture was then resuspended in the 40 
remaining microlitres. 10 µL of the concentrated mixtures were plated in 96-well plates coated with 
100 µL of LB agar supplemented with DAP and arabinose and incubated overnight at 37ºC. 
Finally, each well was resuspended in 120 µL of fresh MH and serial dilutions were plated by spots 
in different selective markers to determine total recipients and transconjugants. In case of low 
recombination frequencies, 100 µL of the direct resuspension were also plated independently. The 
frequency of recombination in the recipient bacteria was determined by counting the number of 
recipients expressing the pSW resistance marker (Chloramphenicol) over the total number of 
recipients in the absence of DAP, since the replication of pSW depends on the presence of the P 
protein. In this case, the recombination rate was calculated as the ratio of transconjugants 
resistant to chloramphenicol (Cm+, Zeo+, Carb+) to the total number of recipients (Zeo+, Carb+). 
The presence of the attI-attC cointegrate was checked by PCR with specific primers (Swbeg and 
CheckConjGFP2.0) (Supplementary Table S2) randomly selecting six clones per recombination. 

 
Three biological replicates and two technical replicates were performed per recipient strain. 

A negative control of recombination was also performed in the absence of integrase to stablish 
background recombination. 
 
 
 
 
 
 
 
 



Materials and Methods 

 60 

3.17 Calculation of the probability of folding attC sites. 
 

The probability of folding the recombinogenic structure of each attC site was calculated 
using the online RNA webserver (http://rna.tbi.univie.ac.at/cgi-bin/RNAWebSuite/RNAfold.cgi). 
First, we submitted the attC sequence for folding using “Matthews model, 2004” as DNA folding 
parameter. Then, the same sequence was submitted for folding denoting R and L boxes as 
constrains in the sequence. The free energies of both thermodynamic ensembles were then used 
to calculate the probability of folding the recombinogenic structure using the following formula120: 

 
 
 In the formula, p is the probability of forming the recombinogenic structure, DGu and DGc 

are the free energy values of both the unconstrained and constrained sequence foldings 
respectively, R is the gas constant (R=1.987x10-3 kcal/mol/K), and T is the temperature of folding 
measured in Kelvin (310K). 

 
3.18 Growth curves. 
 

Three independent colonies were resuspended in MH zeocin and incubated overnight at 
37ºC with shaking. Cultures were then diluted 1:200 in fresh MH zeocin media and 200 µl were 
inoculated in 96-well plates. Plates were incubated for 22 h and OD600 was measured every 10 
minutes using an incubated Synergy HTX plate reader. Prior to measuring, the plates were 
subjected to shaking at 280 cycles per minute. Growth curves under anaerobic conditions were 
performed using an incubated GenTech Epoch2 plate reader under strict anaerobic conditions 
(gas atmosphere 7% H2, 10% CO2, 83% N2). 
 
3.19 Inducibility of aacA5. 
 

In an effort to recreate Jia et al.’s experiments121, we studied the inducibility of aacA5 using 
disc diffusion tests and induction in broth. 
 

Disc diffusion tests: Overnight cultures of pMBA, pMBA5´-aacA5 and pBGT were adjusted to 
a 0.5 in the Mc-Farland scale using saline solution. These solutions were further diluted 1/200 and 
seeded on MH agar plates by inundation (∼3 ml) for 3 minutes. After discarding the remainder of 
the inoculum and letting the plates dry, discs containing kanamycin (30 µg), sisomycin (30 µg) or 
arabinose (2 µg) were placed on top of the agar. Kanamycin disks were obtained from Oxoid 
(Supplementary Table S3) while sisomicin and arabinose discs were prepared in house using 
commercial 6-mm sterile discs (Oxoid) and powder reagents (Supplementary Table S3). Plates 
were incubated overnight at 37ºC. Pictures were taken in a GelDoc apparatus using epilight for 
growth and UV transillumination for GFP fluorescence (BioRad). Fluorescence pictures were 
taken using constant exposition values across all plates to avoid misinterpretations derived from 
exposition differences. Fluorescence profiles across the diameter of the plate were obtained using 
ImageJ plot profile tool (https://imagej.nih.gov/ij/). 

 
 

𝑝 = 	𝑒[(∆+3!4∆+")/&7] 

http://rna.tbi.univie.ac.at/cgi-bin/RNAWebSuite/RNAfold.cgi
https://imagej.nih.gov/ij/


Materials and Methods 

 61 

Induction in broth: three independent colonies of pMBA and pMBA5´-aacA5 were inoculated 
in LB and incubated at 37ºC overnight. Cultures were then diluted 1/50 in fresh LB containing a 
range of inhibitory and subinhibitory concentrations of kanamycin (0.125 - 16 µg/mL) and 2 × 200 
µl of each were placed in 96-well plates and incubated at 37ºC without shaking. Growth and GFP 
expression were followed along 24 h using a Biotek Synergy HTX plate reader and BioStack plate 
feeder (Agilent, USA). Growth (OD600) and fluorescence intensity at 488 nm were measured every 
15 min with prior shaking at 567 cpm for 10 seconds. Growth and fluorescence curves are the 
result of the mean of the two technical and three biological replicates (six points) of each strain. 
In an interpretation of the experiments performed by Jia et al. we also took a sample of these 
cultures after 1 h of incubation, diluted 1/20 in filtered saline solution and measured fluorescence 
using a Cytoflex-S flow cytometer (Beckman Coulter). 





 

 
 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

RESULTS 
 

 
 

Scan me on Spotify 



 
 

 
  



Results 

 65 

 

4 Results 
 

The results section of the present PhD thesis is structured into two main parts, each of 
them includes a series of studies. The first part of this section, named “Evolutionary dynamics 
of integron resistance cassettes”, develops the results obtained from the characterisation and 
quantitation of the evolutionary forces that drive ARC success. Likewise, this part is subdivided in 
5 chapters that enclose the generation of the strains collection, the antimicrobial resistance 
profiling of all ARCs, the study of ARC-dependent polar effects occurring in integron arrays, the 
measurement of the fitness cost entailed by each ARC, and the quantitation of the differential 
mobility of each ARC. The second part of the results section, named “A deeper look into the 
integron model”, seeks to clarify aspects of the integron regulation that remain controversial in 
the field. This part deals with the counterintuitive presence of an aminoglycoside-sensing 
riboswitch that regulates the expression of ARCs in integrons. 

 
It is worth mentioning that some of the results shown in this work have been already 

published in prestigious and  peer-reviewed journals. The antimicrobial resistance characterisation  
of each ARC, enclosed in the first part of the results,  has been publish in npj  Antimicrobials and 
Resistance journal  under the title “Profile and Resistance levels of 136 Integron resistance 
Genes”122. In addition, the second part of the results have been successfully published in Nucleic 
Acid Research journal titled “The expression of aminoglycoside resistance genes in integron 
cassettes is not controlled by riboswitches”107. 

 
 





 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 

PART 1 

 
Evolutionary dynamics of integron 

resistance cassettes 
 
 
 
 
 
 
 
 





Results: Evolutionary dynamics of integron resistance cassettes 

 69 

4.1 Evolutionary dynamics of integron resistance cassettes. 
 

Mobile integrons are present in a plethora of Gram-negative bacterial species carrying 
around 200 different antimicrobial resistant cassettes (ARCs). These ARCs provide resistance 
against 13 different antimicrobial families and are widespread across environmental and clinical 
settings81,82. Despite its general and boundless role in the rise of antimicrobial resistances, 
individual ARCs have very different prevalence across databases. ARCs displaying the same 
activity and closely related are found in very different frequencies across datasets (Figure 12).  
 

 
Figure 12. Occurrence of each ARC within its gene family across databases. Graph showing ARC 
occurrence in IntegrAll database in the three main ARC families: aminoglycoside resistance cassettes (aa), 
beta-lactam resistance cassettes (bla), and trimethoprim resistance cassettes (dfr). ARCs that comprise half 
of the datasets for each ARC family are specified by their names and occurrences in the graph. Data 
collected from the 10 genera with more than 100 class 1 MI sequences described (Acinetobacter, 
Aeromonas, Citrobacter, Enterobacter, Escherichia, Klebsiella, Proteus, Pseudomonas, Salmonella, and 
Vibrio).  
 

This fact indicates that antibiotic selection is not the only parameter that governs the fate 
of ARCs, so there should be other factors linked to the identity of the ARC contributing to its 
success. We believe that this factors are the antimicrobial activity and the fitness cost entailed by 
each ARC, as any other antimicrobial resistance gene. In addition, due to its presence within the 
integron platform, the potential polar effects in downstream cassettes as well as its differential 
capacity to recombine should be taken into account as elements driving ARCs success. In order 
to study these four evolutionary forces, we have generated a collection of strains bearing specific 
ARCs in a vector (pMBA) which mimics its natural environment, an integron. 

0

50

100

aa

aadA2
(33,8%)

aadA5
(15,7%)

aadB
(11,4%)

A
R

C
s 

oc
ur

re
nc

e 
(%

)

bla

blaVIM-2
(19,2%)

blaOXA-1
(15,7%)

blaVIM-1
(13,5%)

blaOXA-2
(10,8%)

dfr

dfrA12
(32,5%)

dfrA17
(24,6%)





Results: Evolutionary dynamics of integron resistance cassettes 

 71 

4.1.1 Generation of the pMBA collection. 
 

First, we retrieve all ARCs from the IntegrAll database, the main curated repository of 
integron sequences. Applying a 95% nucleotidic sequence cut-off, we identified 177 different 
ARCs in class 1 mobile integrons107, as mentioned in the introduction of this work (Supplementary 
Table S5). We synthetised each ARC and cloned them in pMBA, a vector designed ad hoc for this 
study that mimics the natural environment of a class 1 integron allowing us to characterise each 
ARC on its native background (Figure 13A). pMBA is a non-conjugative small size plasmid based 
on a p15a origin of replication (9 copies per cell123) including a zeocin resistance marker, ensuring 
a universal selection that does not interfere with any specific ARC cloned. pMBA, recreates the 
genetic environment of a cassette in the first position of a class 1 mobile integron, cloning each 
ARC in the integration site (attI1) simulating an integration event mediated by an integrase. These 
ARCs are positioned close to a strong Pc promoter (PcS)124 that is encoded within the integrase 
gene. We choose a PcS promoter since it would allow us to identify even subtle differences in 
ARC performance, although it is not the most common described in class 1 MI. It is worth 
mentioning that the intI1 gene present in the vector is truncated to avoid the undesired 
recombination events and deleterious effects of the integrase activity125. Downstream the integron 
there is a GFP gene acting as a second cassette in the array. This GFP will help us to determine 
the effect of each ARC in the expression of the following cassette in the array. Furthermore, it will 
be useful for fitness cost quantitation by in vitro competition assays using flow cytometry111.    

Figure 13. Generation of the pMBA collection. A) Diagram of pMBA derived vectors encoding ARCs. 
pMBA vector is built from a p15a origin of replication, adding to it a zeocin resistance marker and an 
integron-like platform. This integron is composed by a truncated integrase (DintI1) encoding native PcS and 
Pint promoters and the array. Each ARC is cloned in the integration site (attI1) in the first position of the 
array, followed by a GFP placed as second cassette of the array after the ARC attC site. B)  Histogram 
showing the number of ARCs cloned or not against each antimicrobial family (aa: aminoglycoside resistant 
gene; dfr: dihydrofolate reductase; bla: beta-lactamase; fos: phosphomycin resistance gene; qac: 
quaternary ammonium compounds resistance gene;  smr: small multidrug resistance gene; cat and cml: 
chloramphenicol resistance genes; arr: rifampicin resistance gene; sul: sulphonamide resistance gene; 
qnrVC: quinolone resistance gene; lnu: lincosamide resistance gene; ere: erythromycin resistance gene). 
Modified from 122 

A B 



Results: Evolutionary dynamics of integron resistance cassettes 

  72 

Unless otherwise specified, all ARCs were introduced into Escherichia coli MG1655 and 
their sequences were verified by Sanger. The selection of this particular bacterium is justified by 
its significant role as an opportunistic pathogen in both community and hospital settings, along 
with its tendency to  harbour and acquire antibiotic resistance genes. Its relevance it is also 
confirmed as it is part of the ESKAPEE group, bacterial pathogens highly virulent and multidrug 
resistant. 

 
 Among the 177 ARCs, we were able to establish 136 of them in pMBA with the correct 

sequence (Figure 13B). In an attempt to clone the remaining 41, we used E. coli MG1655 high 
competent cells with a transformation efficiency approximated of 108 CFU/mL µg DNA. 
Nevertheless, any colony sequenced of these 41 ARCs showed mutations in essential locations 
of pMBA derivatives such as the promoter, the ARC open reading frame (ORF), and the attI and 
attC sites. The 136 ARCs we managed to clone account for 83% of all ARCs ever identified in E. 
coli and 77% of all ARCs documented within class 1 integrons to date. In summary, the 
combination of pMBA's native environment, the diverse range of cloned ARCs, and the substantial 
size of this collection nominate pMBA collection as the most extensive compilation of integron 
ARCs to date. Consequently, it emerges as a unique tool for understanding the factors that 
contribute to the success of each individual ARC. 
  



Results: Evolutionary dynamics of integron resistance cassettes 

 73 

4.1.2 Antimicrobial resistance characterisation. 
 
As mentioned in the introduction, the emergence of multirresistant bacteria is considered 

nowadays one of the main global health threads. Integrons are able to express and mobilise more 
than a hundred ARCs against most of antibiotics and among the main human pathogens. 
Integrons prevalence and ubiquity rule them as one of the main drivers of antimicrobial resistance 
in Gram-negative bacteria. Nevertheless, a comparative analysis of the resistance levels 
conferred by each ARC is a complex task as data is dispersed between several studies performed 
in different bacterial species, plasmid structures and integron classes containing disparate Pc 
promoters. Moreover, the antimicrobial activity of some ARCs has not been tested experimentally, 
being identified as ARCs just by sequence similarities. Using pMBA collection, we have being able 
to achieve a comprehensive and comparable characterisation of the resistance profiles of each 
ARC. 
 

We determine the resistance provided by each ARC by measuring the minimum inhibitory 
concentration (MIC) of every strain in pMBA collection for a variety of relevant antimicrobials. In 
the case of antibiotic families composed by several members with very different features 
(structure, class or generation), as aminoglycosides and beta-lactams, we obtained a preliminary 
resistance profile using disc diffusion assays (Supplementary Figures S1 and S2) to select for the 
most relevant molecules to study. This section is divided by ARC families and therefore, 
antimicrobials tested; summarizing the antimicrobial mechanism of action and the different 
resistance mechanisms described to bypass it prior to the MIC quantitation for each ARC and 
antibiotic.  

 
Data is presented as column graphs showing both, resistance levels (in µg/mL) and the 

resistance fold change relative to the empty vector pMBA, representing the mean of three 
independent replicates. EUCAST clinical breakpoints are also shown as red-dotted lines when 
available. Gene order in the graph is the result of a hierarchical clustering tree of the proteins 
encoded in ARCs revealing important phenotypic differences in closely related genes. 

 
4.1.2.1 Aminoglycosides resistance cassettes. 

 
Aminoglycosides are included in the WHO’s list of critically important antimicrobials for 

their efficacy in a wide range of bacterial infections, particularly those caused by Gram-negative 
bacteria126. They alter protein synthesis by tightly attaching to the A-site on the 16 S ribosomal 
RNA of the 30 S ribosome subunit127. Resistance to these antibiotics is commonly conferred by 
numerous antibiotic modifying enzymes (AMEs) that transfer acetyl-, phosphoryl- or 
nucleotidyl/adenyl groups to the antibiotic molecule, lowering the affinity for its target26,128. 
Depending on their substrate specificity, these enzymes have different resistance profiles against 
the broad variety of molecules in the family.  Another group of resistance enzymes are 16S-RNA 
methylases, that modify the target instead of the antibiotic. These enzymes confer extremely high 
levels of resistance against most aminoglycosides129. 

 



Results: Evolutionary dynamics of integron resistance cassettes 

  74 

Aminoglycoside resistance is the largest group of ARCs in our collection, with 54 different 
genes (Supplementary Table S5). Integrons encode numerous AMEs including acetyltransferases 
(aacA (31 genes), aacC (8) and sat (1)), nucleotidyl/adenyltransferases (aadA (14) and aadB (1)), 
and phosphotransferases (aph (2)), but do not encode, to date, RNA methylases. In this work we 
use the nomenclature in Partridge et al, 2009108 and the IntegrAll database, but another 
nomenclature is also used in the field128. We provide the equivalence between nomenclatures 
when possible, and we include the exact sequence used here for disambiguation purposes 
(Supplementary Table S5). 

 
Given the large number of genes and molecules in this family, we first measured resistance 

by disc diffusion against seven structurally different aminoglycosides. These included 4,6-
disubstituted deoxystreptamines (4,6-DDs) like kanamycin, gentamicin, tobramycin and amikacin; 
the 4,5-DD neomycin; the 4-monosubstituted deoxystreptamine apramycin -of use in Veterinary 
medicine-; and the non deoxystreptamine streptomycin (Supplementary Figure S1). We then 
determined the MIC of kanamycin, tobramycin, amikacin, gentamicin, streptomycin, and 
apramycin for all ARCs (Figure 14). 

 
As a general overview, we can see the known trends in resistance where aacAs confer 

resistance against kanamycin and tobramycin; aacCs against gentamicin; aadAs against 
streptomycin; aadB against gentamicin; aph genes confer resistance to kanamycin and sat2 did 
not confer resistance against any of the antibiotics tested (Figure 14). In general, phenotypic 
differences seem to correlate with protein families. Yet, superposing a dendrogram of sequence 
relationships reveals known incoherencies in the branching of genes. This highlights that striking 
phenotypic differences can be found among closely related genes, justifying the need for a study 
like this. 

 
Most aacA genes confer increased resistance against kanamycin (Figure 14A) and 

tobramycin (Figure 14B) often with >64-fold increases. Surprisingly, aacA2, aacA16 and aacA45 
provide very low resistance (2- to 8-fold increases) despite being closely related to genes with 
high resistance levels. In fact, aacA16 does not clearly reach the clinical breakpoint to tobramycin 
in our setup. Among the aacAs, amikacin- and gentamicin- resistance is clearly ARC-dependent 
(Figure 14C,D); and aacAX does not confer resistance to any antibiotic.  On their side, aacCs 
confer clinically relevant resistance to gentamicin (MIC ³ 2 µg/mL), with up to 128-fold increase; 
(Figure 14D). Notably, aacC5 and aacC13, that branch together, also confer clinical resistance 
against tobramycin (Figure 14B) 

 
Among the adenyltransferases, aadAs present great specificity against streptomycin, 

(Figure 14E) conferring 16- to 128-fold increases in resistance. Interestingly, aadA4 and aadA10 
do not confer increased resistance to streptomycin -or any other antibiotic. It is possible that this 
is specific of the alleles chosen here, and that other alleles of these genes do confer resistance. 
The fact that certain alleles might not represent a threat highlights the need for functional and 
detailed studies like this one. aadB shows a profile more related to aacAs, not showing activity 
against streptomycin but instead conferring resistance to kanamycin, tobramycin and gentamicin 
(Figure 14A,B,D). 



Results: Evolutionary dynamics of integron resistance cassettes 

 75 

 
Figure 14. MICs of aminoglycoside resistance cassettes. Antimicrobial resistance to kanamycin (A), 
tobramycin (B), amikacin (C), gentamicin (D), streptomycin (E), and apramycin (F) is shown as MIC (µg/mL) 
in the right axis, and resistance fold increase compared to pMBA in the left axis. The MIC is the mean of 
three biological replicates (black dots) for each strain. A red dotted line represents the clinical breakpoint 
(EUCAST) for E. coli against this antimicrobial. A hierarchical clustering tree showing protein sequence 
similarity is shown under the graphs. Figure from 122. 

Kanamycin

sa
t2

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15

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hA
16

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1

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4

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1
2
4
8

16
32
64

128
256
512

1024

1
2
4
8
16
32
64
128
256
512
1024

Gentamicin

sa
t2

aa
cA
X

ap
hA
15

ap
hA
16

aa
cA
49

aa
cA
43

aa
cA
64

aa
cA
28

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56

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5

aa
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aa
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Results: Evolutionary dynamics of integron resistance cassettes 

  76 

Within the pMBA collection, aphA15 and aphA16 are the only phosphotransferase 
encoding cassettes (aphs). Both genes confer clinical resistance against kanamycin (Figure 14A), 
but at different levels: aphA16 provides 8-fold the resistance of aphA15 (128-fold vs. 16-fold 
increases respectively). 
 

sat2 is described as an acetyl-transferase conferring resistance against streptothricin130. 
This compound has very little clinical relevance due to its toxicity131, and has not been tested here. 
Our data do not corroborate this phenotype, but at least rule out its role in resistance against other 
more recent or clinically relevant aminoglycosides (Figure 14). Regarding apramycin  resistance 
(Figure 14F), an antibiotic of use in Veterinary medicine, it is known that aac(3)-IV is the only 
resistance gene, and it is not found in integrons. Our results generally corroborate this, yet the 4-
fold increase in resistance provided by aacA47 and aacA61 is a call for caution. 
 
4.1.2.2 Beta-lactam resistance cassettes. 
 

Beta-lactam antibiotics are likely the class of antimicrobials with the highest use to treat 
infectious diseases132, with some of its members classified as last resource antibiotics by the 
WHO. These molecules affect cell wall synthesis by binding to specific proteins called penicillin-
binding proteins (PBPs). PBPs are transpeptidases involved in the crosslinking of 
peptidoglycan133. Beta-lactams can be classified into penicillins, cephalosporins, carbapenems, 
and monocyclic beta-lactams132. These antibiotics are used to treat a plethora of infections in many 
clinical situations. 

 
In our study, we successfully cloned and characterized a total of 19 different integron 

cassettes, encoding most beta-lactamase classes (Supplementary Table S5). Attending to 
Ambler´s classification134, these included 11 class D enzymes, commonly referred to as 
oxacillinases (blaOXA); 5 class B enzymes known as metallo-beta-lactamases (blaVIM and blaIMP 

variants) and 3 cassettes encoding class A enzymes, specifically blaGES, blaBEL and blaPBL. 
Applying Bush-Medeiros-Jacoby135 classification, we can find 14 enzymes enclosed in group II 
and 5 group III beta-lactamases in pMBA collection. Given the variety of antibiotics and 
generations within the family, we performed preliminary diffusion antibiograms to select 
representative antibiotics from different classes (Supplementary Table S3) (Supplementary Figure 
S2). We chose a member of the most clinically-relevant beta-lactam classes for MIC 
determination, namely amoxicillin (an aminopenicillin), cefaclor and ceftazidime (1st and 3rd 
generation cephalosporins), ertapenem (a carbapenem), and aztreonam (a monobactam) (Figure 
15). 
 

Every beta-lactamase encoding ARC confers high resistance to amoxicillin in a clinically 
relevant manner (MIC ≥ 8 µg/mL) (Figure 15) and only blaOXA-1, blaOXA-9, blaPBL-1, and blaBEL-1 were 
inhibited by clavulanic acid (Supplementary Figure S2).  



Results: Evolutionary dynamics of integron resistance cassettes 

 77 

 
Figure 15. MICs of b-lactam resistance cassettes. Antimicrobial resistance to amoxicillin (A), cefaclor 
(B), ceftazidime (C), ertapenem (D), and aztreonam (E) is shown as MIC (µg/mL) in the right axis, and 
resistance fold increase compared to the empty pMBA control in the left axis. The MIC is the mean of three 
biological replicates (black dots) for each strain. A red dotted line represents the clinical breakpoint 
(EUCAST) for E. coli against this antimicrobial. A hierarchical clustering tree showing protein sequence 
similarity is shown under the graphs. Figure from 122. 
 

OXA-2

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Results: Evolutionary dynamics of integron resistance cassettes 

  78 

Oxacillinases (blaOXA genes) are the most abundant group of beta-lactamases in our 
collection. This is a broad family of proteins with different resistance profiles, ranging from 
penicillinases to carbapenemases. Despite not having a clinical breakpoint for cefaclor, fold 
increases in resistance were high in general, except for blaOXA-1; while only blaOXA-2 and, blaOXA-46 

showed an extended spectrum beta-lactamase (ESBL) profile, conferring resistance against 3rd 
generation cephalosporins. Importantly, although many oxacillinases in our collection have 
carbapenemase activity, none of them reached clinical resistance against ertapenem, and only 
blaOXA-9,  blaOXA-10, and blaOXA-129 conferred resistance against aztreonam.  
 

blaVIM genes are well known carbapenemases found in integrons. Together with blaIMPs in 
pMBA collection they confer the expected resistance against cephalosporins and carbapenems, 
but not monobactams. blaBEL-1 and blaGES-1 are ESBLs with similar resistance profiles. They were 
described for the first time in Pseudomonas aeruginosa136 and Klebsiella pneumoniae137 and they 
both confer resistance to 3rd generation cephalosporins and monobactams, although at different 
levels. Also, blaBEL-1 and blaGES-1 are inhibited by clavulanate. blaPBL1 is also an ESBL, that reaches 
clinically relevant resistance to cephalosporins in our conditions, but differs from the previous ones 
in that it does not confer resistance to monobactams. 
 
4.1.2.3 Antifolate resistance cassettes. 
 

Antifolates such as sulphonamides and trimethoprim, inhibit the synthesis of 
tetrahydrofolate at different stages and are commonly used synergistically to treat urinary, 
respiratory, and gastrointestinal infections. Sulphonamides inhibit the dihydropteroate synthase 
(DHPS) (FolP), while trimethoprim inhibits the dihydrofolate reductase (DHFR) (FolA) enzymes 
(Figure 14)138 

 
Resistance genes encode homologs of these enzymes with reduced affinity to the drugs139. 

They generally confer extremely high resistance levels (beyond solubility of the antibiotic). Only 
exceptionally, some alleles can provide intermediate levels of resistance if expression is low 
enough. In integrons, several dfr genes conferring resistance to trimethoprim have been reported, 
while resistance to sulphonamides is conferred by a hybrid cassette in which qacE is truncated 
and fused to sul1. This cassette is immobile because it has lost its recombination site, and it is 
hence often found at the end of arrays, what led to the misconception of it being a conserved 3’ 
region of class 1 integrons. 
 
Our collection contains 27 dfrA/B ARCs and 1 qacEDsul1 (Supplementary Table S5). As expected, 
all genes conferred extreme levels of resistance with MICs of 1024 µg/mL or more (Figure 14 
B,C). Incidentally qacE∆sul1 had to be tested in DH5a because of the high intrinsic resistance 
levels of MG1655 to sulfamethoxazole. 
 
 
 



Results: Evolutionary dynamics of integron resistance cassettes 

 79 

 
Figure 16. Antifolates resistance characterisation. A) Folate biosynthesis pathway. FolP/DHPS 
condenses P-aminobenzoic acid (PABA) and 6-hydroxymethyl-7,8- dihydropterin pyrophosphate (DHPP) 
into 7,8 dihydropteroate (DHP). FolP is inhibited by sulfonamides (PABA analogs). DHP is then converted 
to 7,8 dihydrofolate (DHF) by the action of FolC/DHFS; which is again modified by the action of FolA/DHFR 
into 5,6,7,8-tetrahydrofolate (THF). The action of FolA/DHFR can be inhibited by the drug trimethoprim. 
Modified from 31 B and C) MIC of qacE∆sul1 and dfr cassettes against antifolate antibiotics. 
Antimicrobial resistance to sulfamethoxazole (SMX) (B) and trimethoprim (C) is shown as MIC (µg/mL) in 
the right axis, and resistance fold increase compared to the empty pMBA control in the left axis. The MIC is 
the mean of three biological replicates (black dots) for each strain. A red dotted line represents the clinical 
breakpoint (EUCAST) for E. coli against this antimicrobial. A hierarchical clustering tree showing protein 
sequence similarity is shown under the graph for trimethoprim. Figure from 122. 
 
4.1.2.4 Fosfomycin resistance cassettes. 
 

Fosfomycin is used to treat urinary tract infections, and skin and soft tissue infections140. It 
inhibits MurA, interfering with peptidoglycan biosynthesis at a step prior to that of beta-lactams. 
Resistance often arises through mutations in active transporters that prevent the entrance of the 
antibiotic; the acquisition of plasmid encoded enzymes that degrade the antibiotic, or the 
modification of MurA141. Integrons encode Fos enzymes that inactivate fosfomycin either through 
hydrolysation (fosL and fosM genes), addition of glutathione (fosC2, fosF, fosG, fosK) or addition 
of bacillithiol (fosE, fosH, and fosI)142–144. Here we have determined the MIC to fosfomycin of 10 
different fos ARCs (Supplementary Table S1) (Figure 17A). Most alleles confer strong increases 
in resistance that are clinically relevant (MIC ³ 32 µg/mL), with the exception of fosM, that only 
increases 8-fold the MIC of the strain, and fails to reach the clinical breakpoint. 

 

B C

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Results: Evolutionary dynamics of integron resistance cassettes 

  80 

4.1.2.5 Chloramphenicol resistance cassettes. 
 

Chloramphenicol is a broad-spectrum antibiotic, that is now usually reserved for serious 
infections that have not responded to other antibiotics, due to the risk of serious side effects145. 
This antibiotic inhibits protein synthesis by binding to the 50 S subunit of the bacterial ribosome. 
Chloramphenicol resistance can be mediated by the modification of the chemical structure of this 
compound -either by acetylation (CatB)27 or phosphorylation-, as well as via efflux pumps such as 
CmlA64. A number of catB and cmlA ARCs are present in mobile integrons108. In the pMBA 
collection we could only clone catB alleles (Supplementary Table S5) that conferred high 
resistance levels against chloramphenicol, that in all cases reached the clinical breakpoint (MIC ³ 
8) (Figure 17B). 
 
4.1.2.6 Rifampicin resistance cassettes. 
 

Rifampicin acts inhibiting bacterial RNA polymerase (RNAP)146. Resistance can be 
mediated by mutation in the target (the b-subunit of the RNA polymerase encoded in the rpoB 
gene), or by enzymatic modification of the antibiotic; an example of these enzymes are the ADP-
ribosylating transferases (encoded in arr genes)28,147 present in integrons148. We have cloned and 
characterized 8 arr variants (Supplementary Table S5) (Figure 17C). Resistance levels vary 
significantly across alleles, with two genes providing >64-fold increases in resistance, while arr7 
increasing resistance only 2-fold. 
 
4.1.2.7 Erythromycin resistance cassettes. 
 

Erythromycin is commonly used to treat skin and respiratory tract infections caused by 
Gram positive bacteria. This macrolide inhibits bacterial protein synthesis by binding to the 50 S 
subunit of the bacterial ribosome. Erythromycin resistance can be due to ribosome methylation 
(erm genes)149, erythromycin esterification (ereA genes)66, and antibiotic transportation (mef 
genes)150. Enterobacteria are not generally susceptible to erythromycin, but can be treated in 
some cases with azithromycin, a broader spectrum macrolide with several therapeutical uses151.  

 
We have cloned two ereA genes in our collection (ereA2 and ereA3) and characterized the 

resistance against both antibiotics (Figure 17D). These two alleles increased more than 16-fold 
the resistance of E. coli to erythromycin, reaching MICs > 1024 µg/mL. In contrast, with only two-
fold increases in resistance to azithromycin, neither gene is likely to confer clinical resistance to 
this antibiotic, for which EUCAST provides a loose estimation of the threshold above 16 µg/mL. 
 
 
 
 
 



Results: Evolutionary dynamics of integron resistance cassettes 

 81 

 
Figure 17. MICs of ARCs. Antimicrobial resistance to fosfomycin (A), chloramphenicol (B), rifampicin (C), 
erythromycin and azithromycin (D) is shown as MIC (µg/mL) in the right axis, and resistance fold increase 
compared to the empty pMBA control in the left axis. The MIC is the mean of three biological replicates 
(black dots) for each strain. A red dotted line represents the clinical breakpoint (EUCAST) for E. coli against 
this antimicrobial. A hierarchical clustering tree showing protein sequence similarity is shown under the 
graphs. Figure from 122. 
 
4.1.2.8 Quaternary ammonium compounds ARCs (qac, smr). 
 

Quaternary ammonium compounds (QACs) and antiseptics are commonly used in 
healthcare settings, food processing facilities, and households to disinfect surfaces and products. 
QACs antiseptic potential is based on its ability to interact with and disrupt the cell membranes of 
bacteria, viruses, and fungi152. Chlorhexidine (CHX), benzalkonium chloride (BZK), 
hexadecyltrimethylammonium bromide (CTAB), and sodium hypochlorite (NaOCl) are examples 
of commonly used antiseptics. QAC resistance genes (qacs) and small multidrug resistance (smr) 
ARCs are generally considered to provide resistance against antiseptics via efflux pumps153. 
Studies  characterising resistance mediated by qac and smr ARCs show, at best, 2 to 4-fold 
increases154,155.  

 
 
 
 

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5

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1

2

4

8

16

32

64

128

256

4

8

16

32

64

128

256

512

1024

fo
ld

 in
cr

ea
se

Chloramphenicol 

M
IC

 (µg/m
L)

ar
r2

ar
r8
b

ar
r6

ar
r7

ar
r5

1

2

4

8

16

32

64

16

32

64

128

256

512

1024

fo
ld

 in
cr

ea
se

Rifampicin

M
IC

 (µg/m
L)

> >

ere
A2

ere
A3

1

2

4

8

16

32

64

128

256

4

8

16

32

64

128

256

512

1024

fo
ld

 in
cr

ea
se

Azithromycin

M
IC

 (µg/m
L)

A B

C D

fo
sE fo
sI

fo
sL

fo
sH

fo
sM fo
sF

fo
sN

fo
sK

fo
sC
2

fo
sG

1

2

4

8

16

32

64

128

256

512

2

4

8

16

32

64

128

256

512

1024

fo
ld

 in
cr

ea
se

Fosfomycin

M
IC

 (µg/m
L)

>> >>>

ere
A2

ere
A3

1

2

4

8

16

64

128

256

512

1024

fo
ld

 in
cr

ea
se

Erythromicyn 

M
IC

  (µg/m
L

> >



Results: Evolutionary dynamics of integron resistance cassettes 

  82 

Here we tested the resistance conferred by 8 qac and 3 smr ARCs (Supplementary Table 
S5) against CHX, BZK, CTAB, and NaOCl and found no increase in MIC against any compound 
(Figure 18). Only against CHX we found single replicates growing at 2-fold higher concentrations, 
a variability accepted in the field as not significant, and not changing the final MIC. In our hands, 
none of these genes confer resistance against disinfectants, a relevant finding with strong 
implications for co-selection phenomena. 

 
Figure 18. MIC characterisation of qac and smr ARCs. Antimicrobial resistance to antiseptics 
chlorhexidine CHX, benzalkonium chloride (BZK), hexadecyltrimethylammonium bromide (CTAB), and 
sodium hypochlorite (NaOCl). MIC (µg/mL) is shown in the right axis, and resistance fold increase compared 
to pMBA in the left axis. The MIC is the mean of three biological replicates (black dots) for each strain. A 
hierarchical clustering tree showing protein sequence similarity is shown under the graphs. Figure from 122. 
 
4.1.2.9 Influence of the genetic context in ARC activity. 

 
One of the main aims of this thesis is to provide a comparative view of ARCs. As mentioned 

before, cassettes can be found in a variety of genetic backgrounds that influence their expression 
levels. Therefore it is impossible to provide a single (universal) measure of resistance for ARCs. 
Consequently, MIC values provided here might not be directly translatable to the clinical setting. 
Indeed, some peculiarities of the integron platform need to be considered to better interpret these 
results. Our genetic setup mimics a class 1 integron with a strong variant of the Pc promoter, and 
is located on a plasmid with approximately 9 copies per cell123. This setting was chosen to 
maximize the phenotype of each gene.  

 
 



Results: Evolutionary dynamics of integron resistance cassettes 

 83 

 
Yet integrons are normally borne on large conjugative plasmids with low copy number, and 

they can encode a variety of Pc promoters with different strengths, or even the combination of two 
promoters84. It is therefore probable that phenotypes in clinical settings are not as strong as the 
ones reported here, but the impact of genetic context is probably different among resistance 
mechanisms.  

 
Typically, antifolate genes generally have an almost digital behaviour with all genes 

conferring extremely high resistance, while genes encoding modifying enzymes are known to 
display a more linear behaviour with copy number156. To exemplify the influence of genetic context, 
we have investigated both types of resistance mechanisms in a low number/low expression 
genetic context. We have determined the MIC of dfrA5 and aadB borne in the low copy plasmid 
R388157 and located in first position of an integron array with a truncated integrase, as in pMBA 
vector, but under the control of a weak Pc promoter (PcW, 26 to 30 times weaker than PcS) (Figure 
19). As expected, trimethoprim resistance is not conditioned by the genetic context while 
aminoglycoside resistance conferred by aadB is clearly lower when expressed from a PcW 
promoter 57 in a low copy plasmid. 

Figure 19. Comparison of the MIC of ARCs in different genetic contexts. Antimicrobial resistance of 
dfrA5 (A), and aadB (B), in R388 is shown as resistance fold increase compared to MG1655 without the 
plasmid. The MIC is the mean of three biological replicates (black dots) for each strain. MIC values for pMBA 
derivatives are taken from figures 14 and 16 and plotted side by side for comparison. Figure from 122.  

R38
8

pM
BA

1
2
4
8

16
32
64

128
256
512

1024
2048

fo
ld

 in
cr

ea
se

dfrA5
> >

R38
8

pM
BA

R38
8

pM
BA

 R
38

8
pM

BA

1

2

4

8

16

32

64

128

256

512

fo
ld

 in
cr

ea
se

aadB

Kanamycin

Tobramycin

Gentamicin

Trimethoprim

A B



Results: Evolutionary dynamics of integron resistance cassettes 

  84 

4.1.3  Polar effects characterisation. 
 
Integron cassettes (ICs) are generally expressed from a Pc promoter located at the 5´ 

integron conserved segment86. Due to this operon-like regulation of IC expression, ICs placed at 
the 3´end of the integron array tend to be less expressed or even silent. It is known that some ICs 
can encode promoters modulating the expression of the following cassettes64–66; nevertheless the 
potential effect of each IC, on particular ARCs, in the expression of the following ones has not 
being assessed in detailed yet. 

 
4.1.3.1 ARC identity affects the expression of downstream ICs. 
 

We measured GFP expression in all strains of the pMBA collection to analyse the effect of 
each ARCs on the following ones. gfp acts as the second cassette of the array in our model so 
any fluctuation in its expression/fluorescence in comparison with the empty vector (pMBA) should 
be caused by the identity of the previous cassette. Results show huge differences in GFP 
expression among pMBA derived strains in an ARC-dependent manner (Figure 20).  
 

Some ARCs such as  catB5, arr8b, dfrA31, smr1, dfrB4, and aacA43 enhance the 
fluorescence of the downstream gfp up to 2 fold in comparison with the empty vector, with no 
known promoters associated with them. In contrast, most ARCs tend to repress GFP fluorescence 
being blaOXA-20 the highest repressor with a 52-fold GFP repression level. Taking into account GFP 
expression levels of the entire pMBA collection, we find a 100-fold variation in expression levels 
between strains with the highest and lowest GFP expression. Data was also collected when strains 
where in stationary phase with a high correlation rate between GFP fluorescence in both phases 
(r = 0.895) (Supplementary Figure S3). A possible explanation for this range of GFP fluorescence 
within pMBA collection could be size of each ARC cloned. We perform a correlation between 
fluorescence and ARC size to demonstrate that (r2= 0.03), ARC identity rules the expression of 
the following ICs of the array independently of their size (Figure 20 inset) .  

Figure 20. ARC identity affects the expression of downstream ICs. GFP fluorescence normalized to 
pMBA levels of each pMBA derived strain in exponential phase. Data is expressed as the mean of three 
independent replicates, error bars represent the standard error of the mean (SEM). GFP expression and 
ARC size do not correlate (inset: r2= 0.03). 
 

bla
OX
A-
20

aa
cA
54

aa
cA
35

qa
cE
Δs
ul1

aa
cA
27arr

7

dfr
A1
2

aa
cA
38

aa
cA
59

bla
IM
P-
31

bla
OX
A-
1

bla
OX
A-
10

bla
BE
L-1

dfr
A2
1

bla
IM
P-
2

dfr
A2
2

aa
cA
45

aa
cC
11

aa
cA
48
qa
cL

dfr
A3
4
dfr
B7
fos
M

ap
hA
15

aa
cA
31
aa
dA
5

bla
OX
A-
46

bla
OX
A-
11
8

aa
cA
3

aa
dA
16

aa
cA
61
aa
dA
4
arr
6
arr
5

bla
OX
A-
2

bla
PB
L-1

bla
OX
A-
21
qa
cK

aa
cA
64

dfr
A1
4

aa
dA
34sa

t2
aa
cA
7

aa
cA
29
dfr
B6
aa
dA
6

bla
OX
A-
5

bla
OX
A-
19
8

aa
cC
3

aa
cA
51
dfr
B5
aa
cC
1
dfr
B1
qa
cH
aa
cC
4

bla
OX
A-
12
9
qa
cF
dfr
A2
5
fos
L
qa
cE
aa
cA
2

ca
tB
10

aa
cA
50

aa
cA
X
ere
A3

bla
VI
M-
7

aa
dA
1
aa
cC
2

dfr
A3
0

aa
dA
29
qa
cGfos

G
aa
dA
7

aa
dA
24

dfr
A2
9
aa
cC
6

aa
cA
42
ca
tB
6

aa
cA
34

aa
dA
11
dfr
B9

dfr
A3
5

aa
cA
47

aa
cA
49

dfr
A1
6
sm
r2
qa
cM
aa
cA
4
fos
H

aa
cA
17
aa
cA
8
dfr
A5

aa
cA
16
dfr
A6

aa
dA
10

aa
cA
52 fos

I

bla
GE
S-
1

bla
OX
A-
9
dfr
A7
dfr
B3
aa
dA
2

dfr
A1
7
dfr
A1

dfr
A2
7
ere
A2
ca
tB
3
ca
tB
2
dfr
B2

aa
cA
28

aa
cA
30
dfr
B8

aa
cC
13
aa
cC
5

aa
dA
28fos

E

bla
VI
M-
1

ap
hA
16
fos
N
dfr
A1
5
sm
r3
fos
F
arr
2

bla
VI
M-
2
fos
K
fos
C2

aa
cA
37

aa
dA
13
aa
dB

aa
cA
56
ca
tB
5
arr
8b

dfr
A3
1
sm
r1
dfr
B4

aa
cA
43

0.01

0.1

1

10

ARCs

G
FP

 fl
uo

re
sc

en
ce

 (n
or

m
. p

M
B

A
)

0.0 0.5 1.0 1.5 2.0 2.5
0

500

1000

1500

2000

GFP expression (norm. pMBA)

A
R

C
 s

iz
e 

(b
p.

)

Correlation GFP expression vs. ARC lenght (bp.)

r2 = 0.03371 



Results: Evolutionary dynamics of integron resistance cassettes 

 85 

4.1.3.2 ARC identity affects the transcription of downstream ICs. 
 
There are two main possible explanations for this phenomenon; ARCs can regulate the 

array expression in a translational and/or transcriptional level. To decipher how deeply ARCs can 
regulate ICs expression we select a subset of ten ARCs for further studies (Figure 21). We 
selected ten different strains that harbour members of the dfr family, covering a significant range 
of GFP expression (20-fold). These strains contain ARCs with varying lengths, attC sites, and 
molecular features. To this purpose, we quantitated the amount of GFP protein and RNA in these 
strains via Western blot and qPCR respectively.  

Figure 21. GFP expression of a subset of pMBAdfr strains. GFP fluorescence normalized to pMBA levels 
of each pMBA derived strain. Data is expressed as the mean of three independent replicates, error bars 
represent the standard error of the mean (SEM). 
 

As expected, GFP expression and protein yields follow the same trend validating our flow 
cytometry results via Western blot (Figure 22). In this experiment, we quantitated DnaK protein 
levels as an internal control to normalize absolute GFP protein levels.  

 
Figure 22. GFP fluorescence and protein yields in a subset of pMBAdfr strains. GFP protein yields 
were measured by Western blot and normalize to DnaK as an internal control and to the empty vector pMBA 
levels for easier comparison with GFP fluorescence. Data is expressed as the mean of four independent 
replicates, error bars represent the standard error of the mean (SEM). 
 

dfr
A2
1

dfr
A2
2

dfr
A3
4

dfr
B5

dfr
A1
6

dfr
A5

dfr
B2

dfr
B8

dfr
A1
5

dfr
B4

0.01

0.1

1

10

ARCs

G
FP

 (n
or

m
.  

pM
BA

)

Fluorescence

Protein

DnaK
70 kDa

GFP
27 kDa

MG    21     22        34       5       15       16 pMBA MG        5        8         2        4     pMBA

- dfrA                              + - dfrB                       +

dfr
A2
1

dfr
A2
2

dfr
A3
4

dfr
B5

dfr
A1
6

dfr
A5

dfr
B2

dfr
B8

dfr
A1
5

dfr
B4

0.01

0.1

1

10

ARCs

G
FP

 fl
uo

re
sc

en
ce

 (n
or

m
.  

pM
BA

) 



Results: Evolutionary dynamics of integron resistance cassettes 

  86 

This approach helps us determine whether GFP fluorescence levels are influenced by each 
ARC or are a result of a general change in the bacterial proteome. We used E. coli MG1655 as a 
negative control for GFP and the empty vector pMBA to normalize our results, thus acting as a 
positive control. As it happens with GFP fluorescence, dfrB4 contains the higher yields of GFP 
and dfrA21 presents the lower GFP levels. We can see mild discrepancies between GFP 
fluorescence and protein amount in some pMBA derived strains as in dfrA34. This is likely due to 
technical reasons such as Western blot analysis and sensitivity. 

 
We also quantitated the amount of gfp mRNA in each strain of this subset by RT-qPCR. 

We used two different housekeeping genes (rssA158 and rpoA159) to normalise gfp mRNA levels 
(Figure 23A). Focusing on GFP RNA levels, strains with the lower GFP fluorescence intensity 
(dfrA21 and dfrA22 containing strains) also display the lower gfp mRNA levels, in agreement with 
protein yields (Figure 23B). In addition, dfrA15 and dfrB4 containing strains, display the higher 
levels of gfp mRNA, correlating with GFP fluorescence and protein amount. Strains containing 
mid-range repression ARCs show variable levels of GFP mRNA within the same range but not 
following an exact trend. This fact could be explained by technical reasons such as batch effect 
or low number of replicates; and/or by biological reasons like the participation of posttranscriptional 
regulation of the GFP transcript in certain strains. This data clearly shows that the identity of the 
first IC of the array affect the transcription of the downstream cassettes. 

 

Figure 23. GFP fluorescence, protein yields, and GFP mRNA levels in a subset of pMBAdfr strains A) 
GFP mRNA levels were measured by qPCR and normalize to two housekeeping genes (rssA and rpoA) as 
an internal controls and to the empty vector pMBA GFP mRNA levels. B) GFP quantitation via flow 
cytometry, western blot and qPCR for comparison purposes. GFP mRNA levels are expressed as the mean 
of two independent replicates, error bars represent the standard error of the mean (SEM). 
 
4.1.3.3 attC sites are involved in the transcriptional regulation of the array.  
 

Translational and transcriptional regulation of ICs in integron arrays has been studied by 
several authors using different approaches. Some studies postulate that attC sites could be the 
main regulators of these processes due to their hairpin secondary structure and variable 
sequence. Collis and Hall suggested that attC sites could act as transcriptional terminators62, while 
Jacquier et al. described attC sites as translational regulators of downstream ICs160.  

 
 

A B 

dfr
A2
1

dfr
A2
2

dfr
A3
4

dfr
B5

dfr
A1
6

dfr
A5

dfr
B2

dfr
B8

dfr
A1
5

dfr
B4

0.01

0.1

1

10

ARCs

G
FP

 (n
or

m
.  

pM
BA

)

Fluorescence

Protein

mRNA

0.1 1 10

dfrA21

dfrA22

dfrA34

dfrB5

dfrA16

dfrA5

dfrB2

dfrB8

dfrA15

dfrB4

 GFP mRNA (norm. pMBA)

AR
C

s

rssA

rpoA



Results: Evolutionary dynamics of integron resistance cassettes 

 87 

We first studied in silico the possible implications of the attC sites on the expression of ICs. 
The main factors that could affect downstream expression of ICs could be attC size  (Figure 24A) 
and the energy needed to release the secondary structure of the attC site, in other words the 
folding stability of each attC site (Figure 24B). While the strain containing dfrB4 is associated with 
the highest GFP expression levels within this subset and has the shortest attC site with the lowest 
energy required for unfolding it, other pMBA derived strains with similar attC site lengths and free 
energy levels significantly inhibit the expression of downstream ICs, such as dfrB5.In conclusion, 
neither attC site size nor folding explain entirely the ARC-dependent expression of downstream 
GFP. 

Figure 24. Compilation of attC site parameters in pMBAdfr strains. A) attC site size (base pairs) of each 
dfr containing strain. B) attC site folding energy of each dfr containing strains, measured as free energy (DG 
(kcal/mol)). To calculate DG, we used the online RNAfold webserver from Vienna University web page. 

 
After the in silico analysis we decided to perform some experiments to elucidate the impact 

of particular attC sites in the expression of the downstream array. To do so, we followed the double 
approach of modifying or deleting attC sites in our pMBA derived strains (Figure 25). We modified 
dfrB4 ARC changing its attC site for the ones belonging to the strains displaying the lowest GFP 
fluorescence levels within the dfr family: pMBAdfrA12 and pMBAdfrA21. Additionally, we deleted the 
attC sites of these  strains to test whether the GFP located afterwards changed its expression.  

Figure 25. GFP expression comparison between strains with modified and deleted attC sites. dfrB4 
attC site was modified replacing it by dfrA12 and dfrA21 attC sites. In addition, dfrA21 and dfrA12 attC sites 
were deleted. GFP fluorescence was measured by flow cytometry and normalised to empty vector pMBA 
levels. GFP fluorescence levels are represented as the mean of three independent replicates, error bars 
represent the standard error of the mean (SEM). Unpaired t-test with Welch´s correction were used to 
assess statistical significance among strains. These differences are marked as  * (p < 0.05), ** (p < 0.01).  
 

0 20 40 60

dfrA21

dfrA22

dfrA34

dfrB5

dfrA16

dfrA5

dfrB2

dfrB8

dfrA15

dfrB4

ΔG (kcal/mol)

AR
C

s

attC site folding

0 50 10
0

dfrA21

dfrA22

dfrA34

dfrB5

dfrA16

dfrA5

dfrB2

dfrB8

dfrA15

dfrB4

 size (bp.)

AR
C

s

attC site size A B 

dfr
B4

dfr
B4 a

ttC
 df

rA
12

 

dfr
B4 a

ttC
 df

rA
21

 

dfr
A21

dfr
A21

 Δa
ttC

dfr
A12

dfr
A12

 Δa
ttC

0.01

0.1

1

10

Strains

G
FP

 fl
uo

re
sc

en
ce

 (n
or

m
.  

pM
BA

)

*
*

***

 



Results: Evolutionary dynamics of integron resistance cassettes 

  88 

These results show that changing dfrB4 attC site for other potentially repressing attC sites 
lead to a statistically significant yet minor reduction in the expression of the downstream GFP 
(using an unpaired t-test with Welch´s correction). In agreement with these results, the complete 
deletion of the attC sites of dfrA21 and dfrA12 increases the expression of the GFP located 
afterwards. Although there is a repressive effect of dfrA21 and dfrA12 attC sites in the expression 
of the following ICs of the array, this effect does not entirely explain GFP repression reduction in 
comparison with the empty vector. This fact suggests the participation of other factors in the array 
transcription and translation. 

 
Contrarily, the presence of additional promoters within ARCs can upregulate the 

expression of the following ICs. Several examples of ARCs encoding secondary promoters have 
been described in the literature such as cmlA and ereA1 ARCs64,66. Here, we want to check for 
secondary promoters within ARCs that lead to a high expression of the following array such as 
dfrA15 and dfrB4. To do so we deleted the primary PcS promoter in the integron platform in this 
pMBA derived strains and measure their fluorescence intensity, as well as in pMBAereA3 strain as 
a potential positive control due to its sequence similarity with ereA1(Figure 26).  

Figure 26. GFP fluorescence comparison between strains with and without PcS promoter. pMBA 
strains were modified by deleting the main promoter PcS from the integron platform. GFP fluorescence was 
measured by flow cytometry and normalised to empty vector pMBA levels. GFP fluorescence levels are 
represented as the mean of three independent replicates, error bars represent the standard error of the 
mean (SEM). A red dotted line represents GFP expression level (norm. to pMBA) of DPcS pMBA strain. 
Unpaired t-test with Welch´s correction were used to assess statistical significance among strains. These 
differences are marked as n.s. (non-significative), * (p < 0.05), ** (p < 0.01), *** (p < 0.001).  
 

The deletion of the main PcS promoter in the empty vector pMBA leads to a GFP 
fluorescence reduction of approximately 800 fold. When comparing GFP fluorescence between 
promoter and promoterless strains we can see a dramatic decrease of GFP expression in 
pMBAdfrA15 and pMBAdfrB4 promoterless strains (DPcS). DPcS dfrB4 containing strain exerts similar 
GFP fluorescence levels to the ones presented by DPcS pMBA, coherent with the absence of any 
secondary promoter. Nevertheless, DPcS dfrA15 and ereA3 containing strain shows significantly 
higher GFP fluorescence levels, suggesting the potential existence of a secondary weak promoter 
within the ARCs. Nevertheless, the existence of secondary promoters within ICs do not explain 
every case where IC array expression is enhanced as it happens in pMBAdfrB4 strain, which shows 
higher GFP expression than pMBA strain. This fact suggests other mechanisms not yet described.  

ΔP
cS

 pM
BA

dfr
A15

ΔP
cS

 df
rA

15
dfr

B4

ΔP
cS

 df
rB

4
ere

A3

ΔP
cS

 er
eA

3
0.001

0.01

0.1

1

10

Strains

G
FP

 fl
uo

re
sc

en
ce

 (n
or

m
.  

pM
BA

) 

***

n.s.

**



Results: Evolutionary dynamics of integron resistance cassettes 

 89 

4.1.4 Fitness cost characterisation. 
 
Bacterial fitness is a complex parameter that encompasses various elements defining a 

microorganism's ability to thrive in a competitive environment. It is widely recognized that the 
acquisition of genes conferring resistance to antimicrobials is linked to alterations in bacterial 
metabolism, which may result in a reduction in overall fitness20,161. In the absence of any selective 
pressure, carrying antimicrobial resistance genes (ARGs) imposes a fitness burden on the 
resistant bacterium. However, when antibiotics are present, the resistant strain can survive and 
outcompete the susceptible one162. This equilibrium between fitness cost and adaptability is crucial 
for bacterial evolvability and survival. 

 
Integrons are low-cost structures entailing a general reduction in the host fitness ranging 

between 1.3 and 4%163. This cost is mitigated by a tight regulation of the integrase activity. It is 
also known that the fitness of a certain integron correlates with the strength of the Pc promoter 
and the IC content; the higher number of ICs in the array, the costlier the integron163. However, it 
is noteworthy that ICs in the array generally follow a polar expression from the Pc promoter, which 
ameliorates the cost of long arrays, with distal ICs slightly expressed or even silent. Few studies 
have assessed the cost of each ARC164 and none of them have been performed in the native 
context of an integron with a representative amount of ARCs. Here we aim to characterize the 
fitness cost associated with each ARC of pMBA collection under different and relevant 
environmental and clinical conditions. 

 
 

4.1.4.1 ARCs can provide a fitness gain in the absence of antibiotics in vitro. 
 

ARC fitness cost can be estimated from exponential growth rates of resistant (ARC 
encoded) and sensitive bacteria in monocultures. When we compared growth curves of each 
pMBA derived strain we can observe ARC-dependent differences in bacterial growth if the cost of 
this ARCs is high (Figure 27) (Supplementary Figure S4). However, we conducted pairwise 
competitions of isogenic resistance and sensitive strains due to the precision of the technique, 
which can detect differences in fitness up to 1%20.    

Figure 27. Growth comparison between different pMBA derivative strains. Growth (OD600 nm) was 
measured along 24h. Growth curves are represented as the mean of three independent replicates, the 
standard error of the mean (SEM) is represented as a shadow in lighter colour. 

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
pt

ic
al

 d
en

si
ty

 (O
D

60
0 

nm
.)

pMBA

pMBAblaVIM-2

pMBAfosN



Results: Evolutionary dynamics of integron resistance cassettes 

  90 

In vitro pairwise competitions were performed in the absence of any selective pressure by 
flow cytometry111 (see Materials and Methods). We first quantitated the cost of the pMBA vector 
in our genetic background (E. coli vs. E coli bearing pMBA), resulting in a 22% of cost. Then, we 
competed each pMBA derivative against a pMBA GFP KO mutant strain allowing us to differentiate 
both bacterial populations and measure the fitness cost of each ARC by flow cytometry.  

 
 We also have to consider and correct for the differential cost imposed by the GFP in each 

pMBAARC derivative. As shown in the previous chapter, the identity of each ARC modulates the 
expression of the GFP afterwards, resulting in different GFP expression levels. To determine the 
fitness cost associated to each GFP expression level we selected a subset of 17 different strains 
from pMBA collection that yielded different GFP fluorescence intensities to generate their GFP 
KO mutants. We quantitated GFP cost of each of this strains by pairwise competition against their 
respective GFP KO mutants (Figure 28).  

Figure 28. Correlation between fitness cost and GFP fluorescence. A) Graph showing the fitness cost 
of 17 different strains from pMBA collection (named beside each point) and its correlation with GFP 
fluorescence levels. Simple linear regression is represented as a dotted line along the 95% confidence 
interval (CI)  B) Simple linear regression forced to go through y = 1. Data is represented as the mean of 
three independent replicates. 
 

GFP fluorescence and fitness cost were directly proportional following a simple linear 
regression (r2= 0.82) (Figure 28A). In principle, Y-intercept should be 1, as the absence GFP 
fluorescence should not cause any fitness modulation. To maintain this premise, we adjusted the 
linear fit to pass through y = 1, obtaining an equation ( y = 7.64*10-7x + 1) that allowed us to correct 
the cost associated with ARC-dependent GFP fluorescence for each pMBA derivative (Figure 
28B). We then performed every in vitro competition (pMBA GFP KO vs pMBAARC) by flow 
cytometry. After data collection we corrected the fitness effect of each ARC by subtracting the cost 
relative to the GFP expression using the equation described in Figure 28B. Finally, we could 
estimate the fitness effect of each ARC in our experimental conditions: aerobic, in vitro pairwise 
competitions. (Figure 29A). 

0 100000 200000 300000 400000
0.9

1.0

1.1

1.2

1.3

1.4

GFP fluorescence (a.u.)

Fi
tn

es
s 

co
st

Fitness cost vs. GFP expression

aacA59

aacA8

aadB

aacC2

aacA28

aacA52
aacA30

aadA24

aadA13

aacC13

dfrA35

dfrB4

dfrA22.2 
dfrA16

dfrA14

dfrA34

pMBA

y = 1.1*10-6 x + 0.9343

r2 =  0.82

0 100000 200000 300000 400000
0.9

1.0

1.1

1.2

1.3

1.4

GFP fluorescence (a.u.)

Fi
tn

es
s 

co
st

Fitness cost vs. GFP expression

y = 7.64*10-7 x + 1

A B 



Results: Evolutionary dynamics of integron resistance cassettes 

 91 

Figure 29. Effect of each ARC in the fitness of the host strain. A) Graph showing the fitness effect of 
each ARC. This graph shows data already corrected taken into account ARC-dependent GFP fluorescence 
intensity. ARCs conferring a gain in fitness are represented with green bars while ARCs entailing a cost are 
coloured in red. Data is represented as the mean of at least six independent replicates, error bars represent 
the standard error of the mean (SEM). B) Histograms showing the fitness distribution of ARCs from the main 
gene families: dfrs, AgRs genes (aa, aph and sat) and blas. Grey lines represent gaussian fits to the data. 
Gain in fitness is represented in green, no fitness effect in yellow and fitness cost in red. Dark red represents 
a fitness cost >30%. 
 

Data collected from pairwise competitions show that approximately the 60% of ARCs 
(82/136) are costly in the absence of any selective pressure. It is worth mentioning that 9,5% of 
all ARCs (13/136) entail more than 30% cost, making it impossible to exactly quantify their fitness 
effect, even leading to the loss of the plasmid during the competition assays. Unexpectedly, 19% 
of all ARCs (26/136) are able to confer more than a 5% gain in fitness to their hosts reaching up 
the 38% of fitness gain in the case of ereA2.  

 
Focusing on the distribution of fitness effects in ARCs of the same family (Figure 29B), we 

can see Gaussian distributions for dfrs and aminoglycoside-resistance (AgR) genes, with 
members of both families conferring fitness cost and gain. dfrs distribution shows fitness levels 
close to 1, with all members showing a fitness effect of +/- 10% with the exception of dfrB4 and 
dfrA31 which are costlier (15%). AgR genes cause a higher modulation of bacterial fitness with 
some members providing a 18% of gain such as aphA16, and others such as aacA56, aacA43 
and aacA17 imposing a substantial cost (>30%) that impedes their precise quantitation in our 
experimental setting. Besides, bla genes impose a great cost with 31% of them entailing >30% 
cost, while only 4 of them (blaOXA-10, blaBEL-1, blaOXA-1 and blaOXA-198) confer a mild growth advantage 
to their hosts in the absence of any selective pressure. The fact that bla genes tend to be costly 
could be related to both the mechanism of action of their encoded enzymes and their location 
within the periplasm, which requires a potentially costly signal peptide for translocation162,165. 

0.7 0.8 0.9 1.0 1.1 1.2 1.3

2

4

6

AR
C

s 
(n

)

dfrs 

0.7 0.8 0.9 1.0 1.1 1.2 1.3

Fitness

aa, aph, sat 

0.7 0.8 0.9 1.0 1.1 1.2 1.3

bla 

A 

B 

qa
cH

aa
cA

56

aa
cA

43
qa

cK

bla
OXA-5

bla
OXA-9

bla
OXA-21

aa
cA

17

bla
OXA-46

bla
OXA-20arr

2
sm

r1
arr

8b

bla
VIM

-1

bla
VIM

-2

bla
IM

P-31

bla
OXA-2

bla
OXA-11

8
aa

dB

bla
OXA-12

9
dfr

B4

aa
cA

30

dfr
A31

qa
cE
Δs

ul1

aa
cA

X
qa

cE
aa

cC
4

aa
cA

49

aa
cA

28
qa

cM

aa
cA

16fos
F
aa

cC
3
qa

cG

aa
cA

64fos
K
ca

tB
2
dfr

A7

aa
dA

24

bla
VIM

-7
dfr

B2
ca

tB
3
fos

N

aa
dA

13

bla
IM

P-2

aa
cC

13
aa

cC
6
ca

tB
6
ca

tB
5
fos

G
dfr

A12

aa
cA

51sa
t2
dfr

B5
sm

r2
dfr

B9
sm

r3
dfr

B8

aa
cA

29
dfr

A5

dfr
A35

aa
dA

28

aa
dA

29
dfr

B3
fos

H
fos

C2

aa
cA

35
aa

cC
5

aa
dA

16
dfr

B1

dfr
A17

dfr
A30

dfr
A14fos

E
arr

5
aa

dA
1

aa
cA

37

dfr
A15

bla
PBL-1

aa
cA

52

ca
tB

10
aa

cC
1

aa
dA

10

dfr
A27

qa
cF

aa
dA

34 arr
6

aa
cA

54
dfr

A1
aa

dA
2

aa
cA

38

dfr
A16

bla
GES-1

aa
cA

50 arr
7
aa

dA
4

dfr
A29fos

L

aa
cC

11
dfr

B6

bla
OXA-19

8
qa

cL
aa

dA
7

dfr
A22

 

aa
dA

6
dfr

B7
dfr

A6

aa
cA

48
fos

M

aa
cA

61

dfr
A34

dfr
A25

aa
cA

8

bla
OXA-1

bla
BEL-1

ap
hA

15

aa
cA

34

aa
cA

31
aa

cA
2

dfr
A21 fos

I

aa
cA

4

bla
OXA-10

aa
cA

27
aa

cA
3

aa
cA

59
aa

cA
7

aa
cA

45
aa

cC
2
aa

dA
5
ere

A3

aa
cA

47

aa
cA

42

aa
dA

11

ap
hA

16
ere

A2
0.6

0.7

0.8

0.9

1.0

1.1

1.2

1.3

1.4

ARCs

Fi
tn

es
s 

<



Results: Evolutionary dynamics of integron resistance cassettes 

  92 

4.1.4.2 Anaerobiosis is a crucial condition affecting the fitness dynamics of some ARCs. 
 

The observation that many ARCs provide a growth advantage to their host in the absence 
of selective pressure raised questions about the clinical relevance of our results. Specifically, 
explaining the fitness advantage of ereA2 (38% gain) in our experimental conditions proved 
challenging. Our bacterial host, E. coli MG1655, is intrinsically resistant to high levels of 
erythromycin (MIC = 64 µg/mL), so the acquisition of an erythromycin resistance gene should not 
confer any adaptative advantage to the bacterium. Moreover, this ARC is mostly found truncated 
or interrupted in E. coli isolates described in databases. The disparities observed between our 
data and the clinical situation documented in databases prompted us to bring our data closer to 
clinical scenarios. The presence of microbial communities as well as the lack of oxygen are 
definitory elements of life in the gut, therefore we decided to quantitate the fitness effect of ARCs 
in these conditions: anaerobiosis and within a microbial community. 

 
We modified the experimental procedure by conducting pairwise competitions based on 

phenotype selection over a period of four days (as described in the Materials and Methods). This 
allowed us to track the resistant bacterial population's dynamics throughout the competition. To 
achieve this, we generated DGFP mutants of a subset of pMBA derivatives, eliminating the need 
for GFP fitness correction in pairwise competitions between pMBADGFP and pMBAARCDGFP. The 
microbial community selected for batch culture competitions was the controlled Oligo-Mouse-
Microbiota (OMM12)112. The OMM12 consortium comprises twelve different bacterial species 
representing the five major eubacterial phyla found in the murine gastrointestinal tract (Figure 30). 
This consortium has been widely utilized to study microbial interactions and enteric infections and 
has demonstrated stability both in vivo and in vitro 112,166,167.  

 
Figure 30. OMM12 consortium phylogenetic tree. Phylogenetic relation between the twelve strains that 
composed the OMM12 consortium based on its 16S sequence. OMM12 includes the five mayor phyla present 
in the murine gastrointestinal track: Bacteroidetes (orange), Verrucomicrobia (purple), Actinobacteria (blue) 
and Proteobacteria (red) and Firmicutes (green). Modified from 166 



Results: Evolutionary dynamics of integron resistance cassettes 

 93 

Pairwise competitions were then performed under aerobic and anaerobic conditions, and 
in batch cultures in the presence of the OMM12 consortium (Figure 31). First, dfr ARCs exhibit 
consistent fitness trends in both the presence and absence of oxygen. Specifically, dfrA21 
promotes bacterial growth, while dfrA31 has a detrimental effect. In contrast, the rest of ARCs 
tested modify their fitness effect in an oxygen-dependent manner. aacA7, which provides a gain 
in fitness (8.4%) in our flow cytometry competition assays, seems to be detrimental in our long 
term competitions. In the same line, blaOXA-10, which entails an initial fitness gain in both 
competition assays, after four days confers a fitness cost to its host. Both, aacA7 and blaOXA-10  
confer slight fitness gain in anaerobic conditions, or at least they do not impose any cost. The 
opposite phenomenon happens with ereA2 which is beneficial for aerobic bacterial growth but 
entails a huge cost in anaerobiosis (>40% cost after 4 days). These data suggest that the presence 
or the absence of oxygen can modulate the fitness effect of ARCs in integrons. 

Figure 31. Pairwise competitions between pMBA and pMBAARCs  strains. In vitro competitions of DGFP 
strains (denominated as pMBA and pMBAARC in the figure) were perform (from left to right) in aerobic, 
anaerobic and anaerobic adding OMM12 microbiota conditions. Along four days pMBA (grey line) strain was 
co-cultured with (from top to bottom) pMBAdfrA21 (orange), pMBAdfrA31 (brown), pMBAaacA7 (purple), 
pMBAblaOXA-10 (blue), or pMBAereA2 (green) strains to assess ARCs fitness effect. Each time point represents 
the mean of three independent replicates, error bars represent the standard error of the mean (SEM). 

0

10

20

30

40

50

60

70

80

90

100

df
rA
21

 (%
)

Aerobiosis

0

10

20

30

40

50

60

70

80

90

100

df
rA
31

 (%
)

0

10

20

30

40

50

60

70

80

90

100

aa
cA
7 

(%
)

0

10

20

30

40

50

60

70

80

90

100

bl
a O

XA
-1

0 
(%

)

1 2 3 4
0

10

20

30

40

50

60

70

80

90

100

Time (days)

er
eA
2 

(%
)

0

10

20

30

40

50

60

70

80

90

100
Anaerobiosis

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4
0

10

20

30

40

50

60

70

80

90

100

Time (days)

0

10

20

30

40

50

60

70

80

90

100
Anaerobiosis + OMM12

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

60

70

80

90

100

1 2 3 4
0

10

20

30

40

50

60

70

80

90

100

Tiime (days)

pMBAereA2pMBAblaOXA-10pMBAaacA7pMBAdfrA21 pMBAdfrA31 pMBA



Results: Evolutionary dynamics of integron resistance cassettes 

  94 

 
However, conducting these competitions within a controlled microbiota, OMM12, does not 

appear to significantly alter the fitness effects of ARCs. ARCs follow a similar trend to competitions 
performed in anaerobic conditions. It is worth mentioning that ARCs that impose a fitness cost in 
anaerobic conditions are even more costly in the presence of OMM12 consortium. Bacteria 
encoding dfrA31 or ereA2 almost disappeared from the mixture after 4 days competitions due to 
the high cost of these ARCs in this environmental conditions.  

 
We also checked whether ARCs could modulate OMM12 consortium composition (Figure 

32). To do so, we quantitated by qPCR the relative abundance of each member of the consortium 
in the beginning and in the end of each competition. We do not appreciate relevant differences in 
OMM12 composition depending on the ARC encoded in the competed strain. Differences between 
stock and final OMM12 community are usually related to substrate depletion in the media during 
the experiment166. We can conclude that E. coli can colonize on top of the OMM12 community 
without destabilizing the microbiota independently of the ARC encoded.  

 
Figure 32. Relative abundance of each OMM12 member in batch culture competitions. Relative 
abundances of every member of OMM12 consortia before (stock) and after each replicate of the batch culture 
competition of pMBAARC DGFP (dfrA21, dfrA31, aacA7, blaOXA-10 and ereA2) vs. pMBA DGFP. OMM12 
members are represented in different colours following the figure legend pattern. 
 

In light of these results, we decided to measure the fitness effect of all ARCs from the 
original pMBA collection in anaerobic conditions via pairwise competitions by flow cytometry 
(Figure 33). This data will enable us to compare the fitness effects of ARCs based on the presence 
of oxygen in the environment. In this scenario, we can see clear differences in the fitness effect 
derived from each ARC in comparison with the results collected from aerobic pairwise competition 
assays (Figure 33A).   

0

50

100

 OMM12 Community batch culture

re
la

tiv
e 

ab
un

da
nc

e 
(%

 o
f t

ot
al

 g
en

e 
co

pi
es

)

YL2

I46

I49
YL31

YL58

YL32
KB1
YL45
YL27
I48
YL44

aacA7 blaOXA-10 ereA2dfrA21 dfrA31

KB18

Stock



Results: Evolutionary dynamics of integron resistance cassettes 

 95 

If we arrange all ARCs in the graph by their costs (Figure 33B) we can observe that the 
vast majority of ARCs (81% vs 60% in aerobiosis) entail fitness cost, with 43 ARCs (32% of all 
ARCs) conferring more than 5% cost, and 10 of them entailing more than a 30% of fitness 
detriment. In addition, only arr7 confers more than 5% benefit to the bacterial growth (26 ARCs 
that have this beneficial effect in aerobiosis). In general, anaerobic conditions reduced the 
potential fitness gain provided by ARCs. The three main ARC families (dfrs, AgR genes and blas) 
tend to be costly (Figure 33C). bla genes follow a clear cost trend, with milder costs than the ones 
displayed in aerobiosis (Figure 29). In this line, dfrs tend to be detrimental with a mean family cost 
of 3% in the absence of oxygen. However, AgR genes generally had a mixed effect on bacterial 
fitness, with variations of up to 5%. Some of its members encode huge cost (aacA17, aacA49 and 
aacA58) being the exception to AgR gene family trend, which is centred in 1.  

 

Figure 33. Effect of each ARC in the fitness of the host strain in anaerobic conditions. A) Graph 
showing the fitness effect of each ARC in anaerobiosis. This graph shows data already corrected taken into 
account ARC-dependent GFP fluorescence intensity. ARCs conferring a gain in fitness are represented with 
green bars while ARCs entailing a cost are coloured in red. Data is represented as the mean of at least six 
independent replicates, error bars represent the standard error of the mean (SEM). B) Graph showing the 
ARC fitness effect in anaerobiosis organized by their cost. C) Histograms showing anaerobic fitness 
distribution of ARCs from the main families: dfrs, AgR genes (aa, aph and sat) and blas. Grey lines represent 
gaussian fits to the data. Gain in fitness is represented in green, no fitness effect in yellow and fitness cost 
in red. Dark red represents a fitness cost >30%. 

0.7 0.8 0.9 1.0 1.1 1.2 1.3

2

4

6

8

10

AR
C

s 
(n

)

dfrs 

0.7 0.8 0.9 1.0 1.1 1.2 1.3

Fitness

aa, aph, sat 

0.7 0.8 0.9 1.0 1.1 1.2 1.3

bla 

C 

A 

B 
qa

cH

aa
cA

56

aa
cA

43
qa

cK

bla
OXA-5

bla
OXA-9

bla
OXA-21

aa
cA

17

bla
OXA-46

bla
OXA-20arr

2
sm

r1
arr

8b

bla
VIM

-1

bla
VIM

-2

bla
IM

P-31

bla
OXA-2

bla
OXA-11

8
aa

dB

bla
OXA-12

9
dfr

B4

aa
cA

30

dfr
A31

qa
cE
Δs

ul1

aa
cA

X
qa

cE
aa

cC
4

aa
cA

49

aa
cA

28
qa

cM

aa
cA

16
fos

 F
aa

cC
3
qa

cG

aa
cA

64
fos

 K
ca

tB
2
dfr

A7

aa
dA

24

bla
VIM

-7
dfr

B2
ca

tB
3
fos

 N

aa
dA

13

bla
IM

P-2

aa
cC

13
aa

cC
6
ca

tB
6
ca

tB
5
fos

 G
dfr

A12

aa
cA

51sa
t2
dfr

B5
sm

r2
dfr

B9
sm

r3
dfr

B8

aa
cA

29
dfr

A5

dfr
A35

aa
dA

28

aa
dA

29
dfr

B3
fos

 H
fos

 C
2

aa
cA

35
aa

cC
5

aa
dA

16
dfr

B1

dfr
A17

dfr
A30

dfr
A14

fos
 E arr

5
aa

dA
1

aa
cA

37

dfr
A15

bla
PBL-1

aa
cA

52

ca
tB

10
aa

cC
1

aa
dA

10

dfr
A27

qa
cF

aa
dA

34 arr
6

aa
cA

54
dfr

A1
aa

dA
2

aa
cA

38

dfr
A16

bla
GES-1

aa
cA

50 arr
7
aa

dA
4

dfr
A29

fos
 L

aa
cC

11
dfr

B6

bla
OXA-19

8
qa

cL
aa

dA
7

dfr
A22

aa
dA

6
dfr

B7
dfr

A6

aa
cA

48
fos

 M

aa
cA

61

dfr
A34

dfr
A25

aa
cA

8

bla
OXA-1

bla
BEL-1

ap
hA

15

aa
cA

34

aa
cA

31
aa

cA
2

dfr
A21 fos

 I

aa
cA

4

bla
OXA-10

aa
cA

27
aa

cA
3

aa
cA

59
aa

cA
7

aa
cA

45
aa

cC
2
aa

dA
5
ere

A3

aa
cA

47

aa
cA

42

aa
dA

11

ap
hA

16
ere

A2

0.6

0.7

0.8

0.9

1.0

1.1

ARCs

Fi
tn

es
s 

< < < < < << < <

arr
2
arr

8b
sm

r1

aa
cA

17

aa
cA

49

aa
cA

56

bla
OXA-20

bla
OXA-21

bla
PBL-1

bla
OXA-46

ap
hA

16

bla
IM

P-31

bla
OXA-2

bla
OXA-12

9

ca
tB

2
dfr

A7
ere

A2

aa
cA

16

aa
cA

43
dfr

B4
dfr

A6

bla
VIM

-2

bla
OXA-1

dfr
A31

aa
cA

30
aa

cA
8

bla
OXA-5

aa
cA

28
ere

A3

aa
cC

13

aa
cA

64

aa
cA

42
dfr

A1

bla
VIM

-1

dfr
A12

ca
tB

5

bla
IM

P-2
qa

cK

qa
cE
Δs

ul1
fos

 C
2

dfr
A27

dfr
A17

aa
cA

7
fos

 K
ca

tB
6
dfr

A5

aa
cA

X
aa

cC
5
sm

r3

aa
dA

24

dfr
A16

dfr
B8

dfr
B3

aa
dB
dfr

A15
fos

 E
fos

 N
fos

 F
fos

 H
dfr

A35

bla
OXA-9

aa
cA

27

bla
OXA-11

8

ca
tB

3
sm

r2

aa
cA

61

aa
dA

13
qa

cF

bla
GES-1

bla
BEL-1

fos
 G
aa

cC
3
dfr

B1
dfr

B9

aa
cA

47
qa

cG
dfr

B6
qa

cH

aa
cA

34

aa
cA

52

bla
VIM

-7

dfr
A14

dfr
A25

dfr
A30

ca
tB

10

aa
cA

35 fos
 I
qa

cM

aa
cC

11
aa

dA
2

aa
cA

29

aa
cA

51

dfr
A34

aa
cA

31

aa
cA

50
aa

cC
2
aa

cC
1
qa

cL
fos

 L

aa
dA

28 arr
5
dfr

B7

dfr
A29

bla
OXA-19

8
arr

6

aa
dA

11
dfr

B5
aa

cC
6
dfr

B2
aa

cC
4
sa

t2
fos

 M

aa
cA

45

aa
cA

38
aa

cA
4

aa
dA

29
qa

cE

aa
dA

10

aa
cA

59

aa
cA

48
aa

dA
4

ap
hA

15

aa
cA

37
aa

dA
1
aa

cA
3

bla
OXA-10

aa
dA

34

aa
cA

54
aa

dA
5

aa
dA

16
aa

cA
2

dfr
A21

dfr
A22

aa
dA

6
aa

dA
7
arr

7

0.6

0.7

0.8

0.9

1.0

1.1

ARCs

Fi
tn

es
s 

<



Results: Evolutionary dynamics of integron resistance cassettes 

  96 

 
 After quantifying the fitness effects of each ARC in both aerobic and anaerobic conditions, 

we can conduct a comprehensive analysis of how the presence of ARCs impacts bacterial fitness 
(Figure 34). In general, ARCs tend to display a weaker fitness effect (cost or gain) in anaerobiosis. 
In aerobic conditions, the fitness effects exhibit an interquartile range (IQR) of 12%, compared 
with the 5% in anaerobiosis. Moreover, the difference between minimum and maximum fitness 
values is way higher in aerobiosis (70%) than in anaerobiosis (30%).  

 
Figure 34. Comparison between ARCs-fitness effect in aerobiosis and anaerobiosis. Violin plots 
showing the fitness effect of all ARCs in the presence (dark blue) and in the absence (light blue) of oxygen. 
Median and quartiles are represented as dashed lines. ARCs measured in aerobiosis and anaerobiosis are 
linked with dotted lines, highlighting ereA2 (red) and qacE (green) ARCs. Each data point is the mean of at 
least six independent replicates. 
 

We can conduct a detailed analysis of the data, examining specific ARCs and their 
responses to oxygen-dependent changes. Most of ARCs are influenced by the lack of oxygen by 
slightly reducing either their cost or gain; nevertheless, the growth of some pMBAARC strains can 
be drastically affected by the absence of oxygen changing the sign of the ARC fitness effect. ARCs 
such as ereA2 (Figure 34 red line) and aphA16 shift their aerobic fitness gain (38% and 18% 
respectively) into a cost (13% and 25% respectively) when oxygen is removed from the 
environment. This drastic modulation also occurs in the opposite sense, with qacE being 
detrimental in aerobic environments (13% cost) and changing its fitness effect to slightly beneficial 
(1% gain) (Figure 34 green line). In contrast, the fitness effect of other ARCs seem to be oxygen-
independent, as it happens with aacA54 and aacA16 which entails 2% benefit and 11% cost 
respectively in both conditions. Summing up, ARCs fitness effect is generally conditioned by the 
presence or the absence of oxygen in the environment. Consequently, specific ARCs can 
modulate the fitness of the bacterial host in opposite directions (from cost to gain and vice versa) 
depending on the oxygenic conditions.  

aero anaero
0.6

0.7

0.8

0.9

1.0

1.1

1.2

1.3

1.4

fit
ne
ss

qacE

ereA2



Results: Evolutionary dynamics of integron resistance cassettes 

 97 

4.1.4.3 ARCs can provide a fitness gain in the absence of antibiotics in vivo. 
 

Next, we aimed to validate our in vitro experiments by replicating pairwise competitions 
(pMBADGFP vs. pMBAARCDGFP) in vivo using mice that were stably colonised by the OMM12 
bacterial consortium. The use of mouse models to perform pairwise bacterial competitions 
increases the environmental complexity, adding the immune system of the host as a new factor 
involved in the competition. Thus, we believe this is the best approach to approximate our 
experimental setup to the clinical reality. For this purpose, we selected three different pMBA 
derivatives to assess the fitness effect of their ARCs: ereA2, aacA7 and blaOXA-10. These three 
ARCs belong to different families, target different antibiotics and exhibit different fitness effect 
under the tested conditions. This diversity between ARCs allows us to draw general conclusions 
from this experimental data. 

 
We performed in vivo bacterial competitions (Figure 35) inoculating the 50:50 mixed culture 

at day 0 and collecting faecal samples from mice every day until day 4. At day 7 we performed the 
final sampling and mice were sacrificed to collect caecal samples from mice intestine. Samples 
were used to quantitate pMBA loads in faeces, relative populations of pMBAARC and potential 
fluctuations in the OMM12 consortium along the competition. 

Figure 35. ARCs can provide a fitness gain in the absence of antibiotics in vivo. A) pMBA loads 
quantitated as CFU/g faeces or GFU/mL caecal content at day seven (7C). B) Fitness effect of each ARC 
during 7 days in vivo competition assays. Each competition is represented with a different colour which 
stands for the ARC fitness effect assessed: ereA2 in green, aacA7 in purple and blaOXA-10 in blue. 
Experimental groups were composed of at least 5 individuals represented in the graphs as data points. 

A 

B 

0 1 2 3 4 7 7C 0 1 2 3 4 7 7C 0 1 2 3 4 7 7C
104

105

106

107

108

109

1010

Time (days)

C
FU

/g
 o

r C
FU

/m
L

E. coli::pMBA loads 

0 1 2 3 4 7 7C 0 1 2 3 4 7 7C 0 1 2 3 4 7 7C
0

25

50

75

100

Time (days)

pM
BA

AR
C

 (%
)

in vivo competitions

ereA2 aacA7 blaOXA-10



Results: Evolutionary dynamics of integron resistance cassettes 

  98 

pMBA loads (Figure 35A) in faeces and caecal content are generally stable at around 107 
CFUs/g or CFUs/mL along the experiment in aacA7 and blaOXA-10 groups. Nevertheless, pMBA 
mice infected with pMBA-pMBAereA2 mixture show a lower colonisation capacity during the initial 4 
days of the experiment (106 mean CFUs/g faeces). In contrast, by day 7, their levels reach those 
observed in the other groups.  

 
When examining each of the three groups, we can observe two contrasting fitness effects 

associated with the encoded ARCs (Figure 35B). pMBAereA2 population is nearly depleted within 
24 hours  of competition, in line with our anaerobiosis results, underscoring the substantial cost 
associated with this ARC in vivo. In contrast, aacA7 and blaOXA-10 clearly provide a growth 
advantage to their bacterial hosts in the mouse gut. After four days of competition pMBAaacA7 and 
pMBAblaOXA-10 populations almost replace bacteria carrying the empty vector, with a prevalence of 
98% in mouse faeces. By the end of the competition, the percentages of pMBAaacA7 and 
pMBAblaOXA-10 in both faeces and caecal content have decreased to 83%. This shift in the 
composition of the mixture might be explained as a result of adaptive mutations in the pMBA 
plasmid, which enhance the fitness of the strain containing the empty vector and stabilize its 
population. Attending to OMM12 composition, no relevant differences between groups are found 
in faecal and caecal samples in day 0 and day 7 (Supplementary Figure S5).  

 
In conclusion, while some ARCs impose a significant cost to their bacterial host in vivo, 

others like aacA7 and blaOXA-10 clearly confer a growth advantage in the absence of any selective 
pressure within the mouse gut.  
 
4.1.4.4 ereA2 reduces E. coli motility. 
 

In light of ARCs fitness results, we wanted to study in detail the molecular processes that  
lead to ARC bacterial fitness modulation. To do so, we performed transcriptomic analysis via RNA-
Seq of certain strains harbouring ARCs grown in aerobic conditions. We selected ereA2 and ereA3 
as they confer a gain in fitness in aerobic conditions, conferring large cost in the absence of 
oxygen. Similarly, we chose dfrA21 and dfrA31 for their distinct fitness effects in various 
environments, entailing fitness gains and costs, respectively. To date, transcriptomic data needs 
a further analysis, being the following preliminary results. We considered transcriptional changes 
significant if they exceeded a 2-fold change and had a significance level of padj < 0.05. 

 
We first exanimated if pMBA plasmid presence modify in E.coli MG1655 transcriptomic 

profile. We found few transcriptomic changes between MG1655 and pMBA strains, potentially due 
to the plasmid presence (Supplementary Figure S6A, Supplementary table S6). Genes up or 
downregulated when pMBA plasmid is transformed exceptionally surpass 4-fold change after 
analysing the raw data using DESeq2 method117, which corrects data dispersion to obtain better 
estimations of transcription fold change. When comparing pMBAereA3 or pMBAdfrA31 strain 

transcription against their parental strain pMBA we can see clear differences with several genes 
up and downregulated in both cases (Supplementary Figure S6B,C, Supplementary table S7). In 
contrast, dfrA21 does not seem to modify extensively the host transcriptome with few genes up or 
downregulated. (Supplementary Figure S6D, Supplementary table S7). 



Results: Evolutionary dynamics of integron resistance cassettes 

 99 

 
pMBAereA2 transcriptome has been analysed further (Figure 36). Following DESeq2 

analysis, we can conclude that several of the genes that are upregulated in pMBAereA2 belong to 
the s32 regulon. This transcriptional factor (encoded in the rpoH gene) controls the heat-shock 
response and therefore, the expression of most genes involved in it168–170 (Figure 36, indicated by 
purple dots). Our transcriptomic analysis revealed a slight increase in rpoH gene transcription 
(1.62-fold increase). This increase is further amplified in downstream genes associated with the 
heat-shock response, including ibpA/B, groL, dnaK/J, lon, hsIU, etc. For instance, ibpA is 
expressed 30 times more in pMBAereA2 than in pMBA. This massive modulation of heat-shock 
response it is not observed in ereA3, dfrA31 or dfrA21 pMBA derivatives (Supplementary Table 
S7), and needs to be exanimated in more detail.  

 
 Among all the genes that are downregulated in pMBAereA2, we wanted to highlight flg and 

fli operons (Figure 36, indicated by yellow dots), as these genes do not show this massive 
downregulation in any other pMBA derivative in the RNA-seq analysis (Supplementary Table S7). 
These genes belong to FlhDC-dependent operons which are crucial in the E. coli flagellar 
regulatory network171. In our analysis these genes show significantly lower transcriptional levels 
in pMBAereA2 strain, being flgB 20 times less transcribed than in pMBA. In this case, the master 
flagellar regulator flhDC shows the same transcription levels in both strains.  

Figure 36. Volcano plot showing differential gene expression of pMBAereA2 in comparison to pMBA.  
Gene expression was measured by RNA-seq both in pMBAereA2 and pMBA. Dotted-lines represent 
significance and fold change thresholds (padj <0.05 and fold change >2 respectively). Genes with a similar 
transcription in both strains are coloured in blue, while upregulated and downregulated genes are 
represented in green and red respectively. Genes involved in flagellar regulatory network (flg, fli) are named 
and coloured in yellow. A variety of genes involved in heat-shock response are also named close to their 
dots and coloured in purple. Data is represented as the mean of three independent replicates.  
 

-6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6

0

50

100

150

200

log2 (FoldChange)

-lo
g 10

(p
ad

j)

pMBAereA2 vs pMBA

flgB flgD

flgC

flgF
flgE

fliF
fliI
fliA

flgG
fliN

flgH

fliG

fliH

rpoH

ybeZ

lon

cspB

hsIU

groL

dnaJ

hsIV

ycjF

groS

dnaK

clpB

ibpB

ibpA



Results: Evolutionary dynamics of integron resistance cassettes 

  100 

The downregulation of genes related to flagellar proteins in pMBAereA2 led us to consider a 
potential motility defect associated to ereA2 expression or activity. To investigate this, we 
quantitated bacterial motility of pMBA and pMBA derivatives ereA2 and ereA3 after 12 hours via 
swarming assays. Results reveal a significant reduction of bacterial motility in pMBAereA2 compared 
to the parental strain, with some replicates showing complete immobility. pMBAereA3 does not 
exhibit a significant motility defect. (Figure 37). These results correlate with the transcriptomic 
downregulation of flagella-related genes observed in pMBAereA2 but not in pMBAereA3. 

 

 
Figure 37. ereA2 reduces E. coli motility. A) Bacterial motility is measured after 12 hours. The graph 
shows the mean of three independent experiments with 3 biological replicates each, error bars correspond 
to the standard error of the mean (SEM). Unpaired t-test with Welch´s correction were used to assess 
statistical significance among strains. These differences are marked as * (p < 0.05). B) Representative 
images showing bacterial motility of each strain after 12 hours. 

 
Next, we investigated whether the motility defect associated with ereA2 is due to the 

protein expression or its enzymatic activity. To address this, we generated a subset of mutants, 
including ereA2 Dmet, in which we deleted the initial methionine of the ARC, and ereA2 cat. mut. 
(H50P E47A), a catalytic mutant of ereA2 with mutations in its active site172. In parallel we 
generated the same mutants for ereA3 as a control, where flagellar genes are not downregulated. 
We measured resistance levels against erythromycin for each strain to confirm whether the protein 
is not expressed or it has lost its enzymatic activity (Figure 38). Results show a decrease in 
resistance in both Dmet and cat. mut. Strains, reaching the parental levels and  confirming the 
loss of expression or activity of ereA2 and ereA3 ARCs. 

Figure 38. MIC characterisation of pMBA derived strains against erythromycin. The MIC is the mean 
of three biological replicates (black dots) for each strain. The parental strain pMBA is shown in green while 
ereA2 mutants are coloured in blue and ereA3 mutants in a red to yellow palette.  

pM
BA 

ere
A2 

ere
A2 c

at.
 m

ut.

ere
A2 Δ

met

ere
A3 

ere
A3 c

at.
 m

ut.

ere
A3 Δ

met
16

32

64

128

256

512

1024

pMBA derived strains

M
IC

 (µ
g/

m
L 

er
ith

ro
m

yc
in

)

> >

pMBA ereA2 ereA3 
0

5

10

15

20

pMBA derived strains

di
am

et
er

 (m
m

.)

*
pMBA ereA2 ereA3

A B 



Results: Evolutionary dynamics of integron resistance cassettes 

 101 

Finally, we quantitated the bacterial motility of pMBAereA mutants (Figure 39). The motility 
reduction observed in pMBAereA2 is maintained when ereA2 is not functional (ereA2 cat. mut.) but 
returns to motility levels similar to pMBA when ereA2 is not translated (ereA2 Dmet). ereA3 and 
its mutants (ereA3 cat. mut. and ereA3 Dmet) exhibit similar motility rates than the parental strain 
pMBA. These data suggest that the motility defect observed in ereA2 encoding strains is due to 
the protein expression itself rather than its enzymatic activity. 

 

 

 
Figure 39. ereA2 expression reduces E. coli motility. A) Bacterial motility is measured after 12 hours. 
The graph shows the mean of three independent experiments with 3 biological replicates each, error bars 
correspond to the standard error of the mean (SEM). The parental strain pMBA is shown in green while 
ereA2 mutants are coloured in blue and ereA3 mutants in a red to yellow palette. Unpaired t-test with 
Welch´s correction were used to assess statistical significance among strains. These differences are 
marked as * (p < 0.05). B) Representative images showing bacterial motility of each strain after 12 hours. 
 

In light of these results we can assume that the expression of ereA2 reduces E. coli motility 
through the transcriptional downregulation of FlhDC-dependent operons, which are involved in the 
formation and regulation of the flagellar complex. Whether this reduction in motility is a 
moonlighting activity of ereA2 or simply arises from nonspecific interactions with other proteins 
remains to be studied. In any case, the observation that E. coli strains harbouring ereA2 exhibit a 
motility defect has significant implications for their bacterial ecology and proliferation, as it may 
interfere with their ability to search for nutrients in a competitive environment. 
  

pM
BA 

ere
A2 

ere
A2 c

at.
 m

ut.

ere
A2 Δ

met

ere
A3 

ere
A3 c

at.
 m

ut.

ere
A3 Δ

met
0

10

20

30

40

pMBA derived strains

di
am

et
er

 (m
m

.) *
*

A 

B 
pMBA ereA2 ereA3ereA2 cat. mut. ereA2 Dmet ereA3 cat. mut. ereA3 Dmet



Results: Evolutionary dynamics of integron resistance cassettes 

  102 

4.1.5 ARC recombination frequency characterisation. 
 

Integrons are genetic platforms able to capture, excise, and reshuffle ICs. These processes 
require the integrase to interact with the attI site situated on the platform and/or the attC sites 
associated to each IC. It is important to note that the attI site is conserved within class 1 MIs, while 
the attC site is cassette-specific. Despite the low sequence similarity among attC sites in MIs, the 
integrase is able to recognise attC sites due to their conserved hairpin-folded structure94,173. 
Nevertheless, several studies suggest that specific features in the sequence and structure of attC 
sites have important consequences in the differential recombination frequency of each 
IC95,96,174,175. In this chapter we quantitate the recombination frequency of each ARC present in 
pMBA collection and attempt to find correlations between these values and specific features of 
attC sites. 
 
4.1.5.1 Generation of the pMBA recombination collection. 
 

Recombination assays are generally conducted by conjugating an attC site, located in a 
suicide vector from a donor strain, to a recipient strain that contains an attI site. The delivered attC 
is inserted via integrase recombination (see Materials and Methods). These assays are designed 
to provide the optimal conditions for recombination to occur. The attC site is delivered into the 
recipient bacterium as ssDNA, which facilitates its recombinogenic folding. Simultaneously, the 
attI site in the recipient strain is already structured as dsDNA. 

 
To mimic a more natural scenario, we chose to perform the conjugation in reverse. We 

conjugated an attI site, located in a suicide vector, to a recipient strain which harbours each 
pMBAARC vector from pMBA collection (Figure 40). 

 
 
 
 
 
 
 
 
 
 
 
 
 

 
Figure 40. Schematic representation of a recombination assays. Briefly, a suicide vector (pSW) 
containing an attI site is conjugated from a donor strain to a recipient strain that harbours a pMBAARC vector. 
When the integrase is expressed in the recipient strain, attI x attC recombination takes place, generating 
chloramphenicol-resistant (CmR) transconjugants . We calculated the recombination rate of each strain as 
the ratio between CmR transconjugants and total number of recipient bacteria.  

Donor Recipient 

Transconjugant 



Results: Evolutionary dynamics of integron resistance cassettes 

 103 

 In the modified recombination assay, the attI site is conjugated in the recipient cell as a 
ssDNA which needs to reach a dsDNA form to be recombinogenic. In addition, attC sites need to 
be extruded from the dsDNA integron sequence to be recognised as ssDNA recombinogenic 
sequences, as it happens naturally. In contrast, the classic recombination assay requires fewer 
substrate modification because the attC sites are delivered as ssDNA sequences in the recipient 
strain, which already contains the attI site in its recombinogenic form as dsDNA. Thus, this new 
conjugation assay will also provide information about the likelihood of each structure reaching its 
recombinogenic state. 

 
It is worth noting that each ARC in pMBA collection is located in the first position of the 

array, followed by a gfp gene which act as a second cassette of the array. To be coherent with IC 
natural integration, the 3´end of each attC site does not belong to the actual cassette downstream 
the integrase cleavage site (G/TTRRRY). We decided to standardize every R’ box along ARCs 
cloned in pMBA collection using AGGC as the RRRY sequence, as it is the most prevalent R’ box 
sequence in ARCs (logo).  
 

Every vector from pMBA collection was subcloned into the DH5a  E. coli strain, which is 
recA-, reducing its capacity for homologous recombination176. This approach helps to quantitate 
specific integrase-mediated recombination without the interference of homologous recombination. 
We also subcloned a p3839 plasmid encoding an int1 gene under the control of an inducible 
promoter which is required for recombination. We will refer to each DH5a strain harbouring 
pMBAARC and p3839 vectors as pMBAARC R strain in an effort to simplify the nomenclature in this 
chapter (Supplementary Table S1).  

 
We first validate our recombination assay procedure comparing recombination frequencies 

achieved using the classical assay, where aadA7 attC site is encoded in a donor strain, with the 
recombination rates reached when the same attC is present in the recipient strain: pMBAaadA7 R. 
(Figure 41). Recombination frequency decreases by one logarithm when the attC site is located  
in the recipient compared to when the same attC site is delivered from the donor strain. This 
phenomenon could be consistent with the extra processes needed to achieve recombination in 
our modified procedure.  

Figure 41. attC site recombination. Comparison between recombination frequencies achieved when 
aadA7 attC site is present in the donor or the recipient strain of the recombination assay. Graph represents 
the mean of at least 15 biological replicates and error bars stand for the standard error of the mean (SEM).     
 

attC donor attC recipient
10-8

10-7

10-6

10-5

10-4

10-3

10-2

10-1

100

Strains

R
. F

.



Results: Evolutionary dynamics of integron resistance cassettes 

  104 

4.1.5.2 Recombination frequency quantitation. 
 

Despite the decrease in recombination rates observed in our modified recombination 
assay, the achieved recombination frequencies are also consistent with those published in the 
bibliography177. Therefore, we quantitated recombination frequency of each ARC present our 
pMBA recombination collection (Figure 42). It is worth considering that the following recombination 
frequencies might be one logarithm lower than the actual values, as indicated by our previous 
validation experiment. The recombination rates presented here have been corrected to subtract 
non-integron-mediated recombination. To do this, we performed PCR on every transconjugant to 
identify attI-attC cointegrates. Recombination rates before this correction are shown in 
Supplementary Figure S7. 

Figure 42. Recombination frequency of each ARC. Graph showing integron-mediated recombination 
rates of all ARCs present in pMBA recombination collection. Bars represents the mean of at least 3 biological 
replicates and error bars correspond to the standard error of the mean (SEM). Recombination frequency 
and attC site length do not correlate (inset: r2 = 0.007981). 
 

Recombination rates vary drastically between ARCs with a million-fold change between 
the highest (aacA52) and the lowest (fosN) integron-mediated recombination rates. These 
differences in recombination do not seem to correlate with attC site length (Figure 42 inset). 
However, when we group ARCs by gene families, we can observe that cassettes encoding 
aminoglycoside resistance genes (AgR genes) recombine with substantially higher rates than the 
rest of families (Figure 43). The other gene families display similar average recombination 
frequencies, although dfr and fos ARCs exhibit greater data dispersion compared to bla and 
qac/smr ARCs. This findings suggest that AgR ICs may, on average, have more opportunities for 
recombination than any other ARC, potentially making them more mobile.  

Figure 43. Analysis of the average recombination capacity of each ARC family. Graph showing 
the recombination tendency of each ARC family by grouping the recombination frequencies of every 
member of each ARC family. Only ARC families with at least 10 members are represented in this graph. 

AgR bla dfr fos  qac, smr
10-8

10-6

10-4

10-2

100

ARC families

R
. F

.

dfr
A3
5
fos
K

qa
cE
Δs
ul1fos

N
fos
F
dfr
A1
6
dfr
A5

dfr
A3
0

dfr
A1
5

dfr
A1
4
aa
dA
4

dfr
A2
5

dfr
A3
4

dfr
A1
2

ap
ha
16

dfr
A2
9

bla
OXA-5

bla
IM

P-31

aa
dA
16

dfr
A2
7
qa
cK
sm
r2

bla
OXA-1

aa
dA
10
ere
A3

aa
dA
13
fos
G
qa
cG

ap
ha
15
ca
tB
6
dfr
B6

bla
OXA-2

bla
OXA-12

9

bla
OXA-21

aa
dA
5
arr
8b

aa
cC
11

bla
VIM

-7

ca
tB
2
arr
2

aa
cC
13
dfr
B3

aa
dA
11
aa
dB

bla
OXA-46

aa
dA
24
sm
r3
dfr
B5
dfr
A7
fos
C2

aa
dA
28
aa
cC
6
dfr
B7
dfr
B8
dfr
A1

bla
OXA-19

8
dfr
B1

dfr
A1
7
ca
tB
3
dfr
B2

bla
OXA-20

ca
tB
5

bla
OXA-9

bla
OXA-10

aa
cA
49

aa
cA
16
aa
dA
2

ca
tB
10

bla
PBL-1
dfr
B4
ere
A2fos

L

aa
dA
34
qa
cL
fos
M
qa
cE

bla
OXA-11

8

bla
VIM

-1

dfr
A3
1
aa
cC
2
fos
H

bla
VIM

-2 fos
I
dfr
B9
qa
cH
qa
cM
aa
dA
1
arr
6
arr
5

aa
cA
30

aa
cA
42arr

7

dfr
A2
2
sa
t2

bla
IM

P-2
qa
cF

bla
BEL-1sm

r1
dfr
A6
aa
cC
5

aa
cA
38

dfr
A2
1

aa
cA
43
aa
cC
3
fos
E
aa
cC
1

aa
cA
27
aa
cA
3

aa
cA
54

bla
GES-1

aa
cA
28

aa
dA
29

aa
cA
17

aa
cA
56

aa
cA
47
aa
dA
6

aa
cA
48
aa
dA
7

aa
cA
35
aa
cA
4
aa
cC
4
aa
cA
8

aa
cA
59
aa
cA
2

aa
cA
31

aa
cA
34

aa
cA
64

aa
cA
45

aa
cA
29

aa
cA
37

aa
cA
51

aa
cA
X

aa
cA
50

aa
cA
61
aa
cA
7

aa
cA
52

10-8

10-7

10-6

10-5

10-4

10-3

10-2

10-1

100

ARCs

R
. F

. 10-8 10-7 10-6 10-5 10-4 10-3 10-2 10-1
0

50

100

150

R. F.

at
tC

 s
iz

e 
(b

p)

Correlation R.F. vs. attC site lenght (b.p)

r2 = 0.007981



Results: Evolutionary dynamics of integron resistance cassettes 

 105 

Remarkably, some ARCs exhibit very low or none integrase-mediated recombination in 
our experimental setup (Figure 42). Specifically, dfrA35, fosK, and qacEDsul1 do not show any 
integrase-mediated recombination, while fosN and fosF display extremely low recombination 
rates, with values lower than 10-7. It is known that qacEDsul1 do not contain any attC site which 
aligns with the lack of integrase-mediated recombination. However, a detailed analysis is required 
to understand this phenotype in the rest of ARCs. Certain anomalies in the attC site structure of 
these ARCs may interfere with proper integrase-mediated recombination (Figure 44).  

Figure 44. Structures of bottom strands of attC sites. fosK (A), fosN (B), fosF (C), and dfrA35 (D) attC 
site structures predicted by Geneious DNA fold plugin using DNA Matthews 2004 energy model. Colour 
code represents base-pair pairing probability (red: high, green: mild, blue: low). R and L boxes are described 
in de sequence when possible as well as the integrase binding sites represented as purple ellipses. 
 

The canonical attC site structure of fosK (Figure 44A) differs from an archetypical attC site 
as it appears to lack any L box with EHB protruding from the hairpin. Additionally, as mentioned 
earlier, the 3´end of every ARC attC site in our pMBA collection is generic, potentially leading to 
mismatches in the R box of attC sites. In the case of fosK, this modification disrupts the hairpin 
structure, completely destabilizing R box formation. These two factors could be the main drivers 
of the non-recombinogenic phenotype of fosK in our assays. 

attC - fosK 

attC logo - fosK 

attC – dfrA35 

attC logo – dfrA35 

attC - fosN 

attC logo - fosN 

attC - fosF 

attC logo - fosF 

R box 

R box 

R box 

R box 

R box 

R box 

L box 

L box 

L box 

L box 

A 

C 

B 

D 



Results: Evolutionary dynamics of integron resistance cassettes 

  106 

 Similarly, the attC site structure of fosN (Figure 44B) is altered when the logo is added to 
its 3´end, disrupting the R box and reducing substantially its recombination capacity. The attC site 
structure of fosF (Figure 44C) is not deeply modified in pMBAfosK but it seems to lack of a canonical 
L box, which could explain its low recombination frequency.  

 
Conversely; dfrA35, which exhibits a non-recombinogenic phenotype in our setup, contains 

a well-structured attC site (Figure 44D) with a minor variation in the R box This minor variation 
should not lead to such a dramatic decrease in recombination capacity, as similar variations are 
naturally found in integron arrays during IC excision. 
 

After confirming that certain ARCs might have reduced their recombination rates due to 
3´end modifications, we aimed to analyse if this phenomenon could significantly bias our results. 
If the presence of mismatches in the  R box of attC sites were always linked to a reduction in 
recombination frequency, we should find all of these ARCs in the lower quartile of our 
recombination rates distribution. However, we can find attC sites with the same R box mismatches 
both in the 10% of ARCs with the highest and lowest recombination rates (Figure 45). Some 
cassettes differ from their canonical attC site 3´end sequence in 1,2 or even 3 base pairs resulting 
in recombination rates that are evenly distributed between the highest (10-2) and the lowest (10-7) 
rates, validating our experimental setup and highlighting the biological relevance of this result.  

 

 
Figure 45. Comparative analysis of the recombination frequencies of ARCs. Graph showing by pairs 
recombination rates of ARCs with the same number of mismatches in their attC site 3´ends. R boxes of 
each ARC are described above each pair of ARCs, being mismatches with the logo coloured in red. Data 
collected from figure 42. 
 
4.1.5.3 Correlation between recombination frequencies and in silico analysis. 
 

ARC recombination frequency is known to depend on certain features of the attC site, such 
as EHB position, free energy of the folded structure, positional entropy, probability of folding in a 
recombinogenic hairpin, GC content, etc178. Here, we aim to evaluate if there is a direct numerical 
correlation between the recombination frequencies obtained in vitro and some of these 
parameters measured in silico.  

aacA52 aadA4 a aacA64 dfrA12 a aacA37 dfrA5 a aacA45 dfrA16
10-8

10-7

10-6

10-5

10-4

10-3

10-2

10-1

100

ARCs

 5´GTTAGGC3´
 3´CAATCCG5´

R
.F

.

 5´GTTAACC3´
 3´CAATCCG5´

 5´GTTAAAA3´
 3´CAATCCG5´

 5´GTTAGCC3´
 3´CAATCCG5´

 5´GTTAACT3´ 
 3´CAATCCG5´

   



Results: Evolutionary dynamics of integron resistance cassettes 

 107 

We believe that the free energy of the structure and its probability of folding generating a 
recombinogenic substrate (pfold) could potentially be the main parameters governing 
recombination. These parameters are related to the stability of the attC site hairpin and its 
recombinogenic conformation. The Gibbs free energy (∆G) indicates whether the folding of a 
ssDNA molecule into a particular secondary structure is energetically favourable or not, with lower 
(negative) values suggesting that the molecule is more stable in its folded state. Additionally, pfold 
calculation considers ∆G of both the unconstrained and constrained (recombinogenic) potential 
structures of the attC site to quantitate the probability of achieving the recombinogenic 
conformation of the attC site. 

 
For this analysis, we selected a subset of 34 ARCs, all of which have attC site 3´ ends 

matching the logo. This selection aims to minimize the introduction of additional parameters that 
could potentially affect the recombination capacity of ARCs and increase the background noise of 
our results (Figure 46A). Subsequently, we quantitated DG of the potential secondary structures 
of their attC sites as well as the probability of folding into recombinogenic structures (pfold) We 
then compared these values with their respective recombination frequencies (Figure 46B,C). 

Figure 46. Correlation between recombination frequencies and in silico analysis. A) Graph showing 
recombination frequencies of ARCs that have attC site 3´ends that match with the logo. B) Correlation 
between recombination frequency and DG of the unconstrained attC site sequence C) Correlation between 
recombination frequency and pfold of the attC site sequence. Recombination rates collected from figure 41. 
 

Recombination frequencies of ARCs achieved in our in vitro experiments do not exhibit 
correlations with either the ∆G or pfold values of their attC sites calculated in silico (r2 DG = 
0.002644 and r2 pfold = 0.001408) (Figure 45B,C). Thus, more detailed studies of attC site 
structure are needed to establish a numerical correlation between in vitro recombination 
frequencies and potential attC site features measured in silico. 

aa
dA
4
aa
dA
5
arr
8b
ca
tB
2

aa
cC
13
aa
dB
dfr
B7
dfr
B2

bla
OXA-20

bla
PBL-1fos

M
dfr
B9 arr

6
arr
5

aa
cA
30

aa
cA
42 arr

7
sa
t2
aa
cC
5

aa
cA
38
aa
cC
3

aa
cA
27
aa
cA
3

aa
cA
54

aa
cA
35
aa
cA
4
aa
cA
8

aa
cA
59
aa
cA
2

aa
cA
31

aa
cA
51

aa
cA
61
aa
cA
7

aa
cA
52

10-8

10-6

10-4

10-2

100

ARCs

R
. F

.

0 25 50 75 100
10-8

10-6

10-4

10-2

100

pfold (%)

R
. F

.

r2 = 0.001408

-60 -50 -40 -30 -20 -10
10-8

10-6

10-4

10-2

100

ΔG (kcal/mol)

R
. F

.

r2 = 0.002644

A 

C B 





 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 

PART 2 

 
A deeper look  

into the integron model 
 
 
 
 
 
 





Results: A deeper look into the integron model 

 111 

4.2 A deeper look into the integron model. 
 

The integron model describes this platform as an operon-like structure. ICs present in the 
integron array are generally promoterless and are transcribed from the Pc promoter located in the 
integron platform. However, the literature describes several exceptions to this model, such as 
cassettes containing promoters64–66, the presence of secondary structures that regulate IC 
translation121, or the existence of small ORFs acting as control elements for translational initiation 
in class 1 integrons179,180. These regulators of IC expression directly effect to the modulation of the 
potential cost and resistance attributed to ICs, making  their study essential for understanding the 
success of each IC.  

 
While the existence of cassette containing promoters and small ORFs affecting IC 

translation is generally accepted in the field, the presence of a common mechanism that regulates 
the translation of AgR ICs via a riboswitch remains controversial181. In this section, we provide 
experimental evidences addressing the validity of this puzzling and counterintuitive integron 
regulatory mechanism. 

 





Results: A deeper look into the integron model 
 

 113 

4.2.1 The expression of AgR genes in ICs  is not controlled by riboswitches. 
 

The expression of antibiotic resistance (AR) genes can influence the phenotype and the 
fitness of the bacterial host, and is a fundamental factor driving their fate in the clinical setting161. 
Inducible resistance mechanisms can alleviate the cost of resistance in the absence of antibiotics 
while preserving a high resistance phenotype in its presence182–184. Several mechanisms are 
known to induce the expression of AR genes in the presence of antibiotics, like ribosome 
stalling185,186, ribosome-mediated transcriptional attenuation187,188, and the sensing of the antibiotic 
molecules189 or their effects190,191. Because the cost of resistance is the main driver of its 
reversibility at the community level192, a clear understanding of the transcriptional and translational 
mechanisms regulating the expression of resistance genes is key in the fight against AR. 

 
Aminoglycosides (Ag) are critically important antibiotics against which the most common 

resistance mechanism is enzymatic modification. Many such modifying enzymes are plasmid-
borne and circulate among clinically relevant species. To fight against the emergence and spread 
of Ag resistant bacteria it is necessary to understand the expression of these determinants. 
Notably, the expression of AgR genes has recently been claimed to be under the control of a 
riboswitch121. The authors observed a strong level of conservation of the 5´ leader RNA of several 
AgR genes and suspected a common regulation mechanism. They investigated experimentally 
the (6´)-N-acetyltransferase encoded in aacA5 (GenBank L06163) and found a potential 
secondary structure in the 5´ untranslated region (5´UTR) of its mRNA that involved a large part 
of the conserved region (Figure 47). Through a series of experiments combining bioinformatic, 
genetic and biochemical techniques, they concluded that 4,6 disubstituted deoxystreptamine (4,6-
DDs) aminoglycosides bind to this structure and destabilize it, releasing the trapped Shine-
Dalgarno (SD) sequence and allowing for the translation of the gene. This structure would hence 
act as a riboswitch. Given the sequence conservation they observed among AgR genes, in follow 
up works from the same group, they investigated the regulation of other AgR genes using a similar 
methodology and concluded that riboswitches are also present in other acetyltransferases193 and 
adenyltransferases194. º 

 
A puzzling aspect of these riboswitch-controlled genes is that they are encoded in integron 

cassettes, and that the conserved region encoding a large part of the riboswitch is actually the 
integration site (attI) of a class 1 integron, where genes of unrelated functions are inserted (Figure 
46). The variety of functions encoded in integron cassettes makes the presence of such a 
riboswitch counterintuitive, since the expression of unrelated genes would be contingent on the 
presence of aminoglycosides. Also, in the initial report on these putative riboswitches, the 
induction levels observed for aacA5 were arguably very low (1.5–2.5-fold) and it has been 
suggested that these effects might be the result of pleiotropic effects of antibiotics on protein 
production, rather than the consequence of a specific induction mechanism181. Yet, data 
supporting this claim was not provided and is still nowadays unavailable. Hence the presence of 
Ag riboswitches in integron AgR cassettes remains today counterintuitive and controversial, and 
experimental proof from independent laboratories is necessary. 
 



Results: A deeper look into the integron model 
 

  114 

 
 
Figure 47. The Ag-sensing riboswitch described in an integron cassette is counterintuitive. Diagram 
of the riboswitch described in Jia et al.121,  upstream the aacA5 integron cassette. A large part of the structure 
is composed of the attI1 integration site of class 1 integrons (blue shade) including the direct repeats (DR) 
1 and 2 (partially) and the L box. This region is the sequence found by the authors to be conserved upstream 
aminoglycoside resistance genes. Cassettes start at the crossover point between the G and the T of the 
conserved 5′-GTT-3′ triplet (dotted box). It is of note that most of the R box (5′-GTTRRRY-3′) in attI sites is 
provided by cassettes in first position, but are a functional part of the site. Modified from 107. 
 

Two strong argument against the biological meaning of an Ag-sensing riboswitch are IC 
mobility and variety. This mobility goes way beyond the reshuffling of positions within the same 
platform and deserves a deeper look into it. A wealth of data in the field put forward that mobile 
integrons recruit their cassettes from sedentary chromosomal integrons in environmental bacteria. 
It is remarkable that cassettes can be exchanged between different integrons. The variety of 
functions found in cassettes also argues against the biological rationale behind an Ag-sensing 
riboswitch that is composed mostly (47/75 bp) of the attI1 site of a class 1 integron, since the 
expression of unrelated genes would be contingent on the presence of aminoglycosides. As 
mentioned previously, we have found 177 different ARCs against most antibiotic families.  

 
From this ARC pool, AgR genes are found in the first position of the array in only 35% of 

class 1 integrons of the database, which correlates well with their relative abundance (36%: 
64/177) and suggests there is no particular bias on the function encoded in first position. Indeed, 
genes conferring resistance against other antibiotics are also found frequently in first position 
(beta-lactams: 25%; trimethoprim: 20%; other families: 8.7%) together with cassettes not related 
to resistance or even cassettes of unknown function (10%) (Figure 48A). Hence, we could not find 
a bias towards the first position for cassettes that could potentially use the attI1 site to form a 
riboswitch. 

 



Results: A deeper look into the integron model 
 

 115 

4.2.1.1 Expression of aacA5 is not induced by Ags in its native genetic context.  
To test the behaviour of aacA5 in the presence of aminoglycosides, we exchanged the 

5´UTR of the gfp in pMBA vector by the 5´UTR of aacA5, obtaining pMBA5´-aacA5 (Figure 48B). This 
is the native genetic context of cassettes in class 1 integrons and is a more biologically relevant 
genetic environment than the cloning vector used in Jia et al.’s work in which expression is 
controlled by an inducible Plac, preceded by the lacI repressor gene.  

 
Figure 48. Mobile integrons contain cassettes encoding varied functions. A) Distribution of functions 
encoded in cassettes in first position of class 1 integrons in IntegrAll. B) Diagram of pMBA derived vector 
used to analyse cassette expression by fusing 5′UTRs to a GFP encoding gene. Not to scale. Modified 
from107. 
 

We subjected this construction to induction with kanamycin and sisomicin (members of the 
4,6-DDs family) in diffusion antibiograms, in an effort to recreate Jia et al.’s experiments in which 
they used a beta-galactosidase reporter instead (Figure 49A). As controls, we exposed a strain 
containing pMBA to both antibiotics, and a strain containing the pBGT plasmid, in which the gfp 
gene is under the control of a PBAD promoter156, to an arabinose-containing disc to induce its 
expression. Results with pBGT validated the assay and provided a plot with a clear inducibility 
profile. pMBA5′-aacA5  did not show increased levels of fluorescence at the border of the inhibition 
halos, as would have been expected from a riboswitch-mediated induction of expression, and 
instead had a profile similar to that of the pMBA control.  
 

We have also recreated Jia et al.’s induction experiments in broth, by growing strains 
carrying pMBA and pMBA5′-aacA5 in doubling concentrations of kanamycin ranging from 
subinhibitory to inhibitory and measuring fluorescence after 1 h through flow cytometry (Figure 
49B). We did not observe an increase in fluorescence at any concentration. Instead, fluorescence 
started decreasing as antibiotic concentrations approached the MIC of our strain (1 µg/ml). It is 
worth noting that the bacterial inoculum used in these experiments was higher than the one used 
to determine the MIC, which explains the visible growth at concentrations above the MIC (a well-
known phenomenon named inoculum effect195). 

A B 



Results: A deeper look into the integron model 
 

  116 

To test whether a longer time period was necessary to observe an induction, we analysed 
growth and expression levels in all concentrations of kanamycin for 24 h (Figure 49C). Our data 
revealed an inhibitory effect in growth at all concentrations that was overlooked in Jia et al.’s 
experiments and could be relevant since it seems to affect bacterial physiology. We took it into 
account by normalizing fluorescence to optical density. 

 
 Data showed that the expression of gfp was not induced in the presence of kanamycin at 

any concentration tested. Instead, fluorescence (both raw, and normalized by the OD) was lower 
in the presence of the antibiotic than in its absence in agreement with previous results. We 
observed a similar behaviour for the empty pMBA control, suggesting that the decrease in 
fluorescence/OD600 is a consequence of the general inhibition of protein synthesis caused by 
kanamycin rather than a specific effect on the 5′UTR of aacA5.  
 
 

 
 
Figure 49. The expression of aacA5 is not induced by aminoglycosides in its native genetic context. 
A) Diffusion antibiograms of pMBA5′-aacA5 and pMBA in the presence of sisomicin and kanamycin, and pBGT 
confronted to an arabinose disc (positive control). B) GFP fluorescence of pMBA and pMBA5′-aacA5 in broth 
experiments after 1 hour of induction with kanamycin. Statistically significant differences compared to the 
no-antibiotic condition are marked with * (p < 0.05), **(p < 0.01) and ***(p < 0.001). C) pMBA5′-aacA5 and 
pMBA growth (OD600), fluorescence (arbitrary units) and cassette expression (measured as 
fluorescence/OD600) along 24 h. Figure from 107. 
 

Altogether, our data do not support a riboswitch-controlled model of expression for the 
ARC aacA5 cassette. Yet it is possible that other AgR genes in integrons possess such control 
mechanisms. 
 
 
 
 

A 

B 

C 



Results: A deeper look into the integron model 
 

 117 

4.2.1.2 Generation of the pMBA 5´aaX collection. 
 

The expression of aacA5 is, in our hands, not induced by kanamycin or sisomicin. 
Nevertheless, we sought to analyse the inducibility of all known AgR cassettes. As we did for 
pMBA5′-aacA5, we generated a collection of 64 pMBA derivatives that harbour the 5´UTRs of each 
AgR gene cassette present in class 1 integrons (Figure 50A, Supplementary Table S8) between 
the attI site and the gfp start codon in pMBA. 

 
These UTRs were defined as the sequence starting at the TT of the 5′-GTT-3′ triplet (where 

the recombination crossover takes place in integrons) to the start codon of the gene. We used 
gene and attC site annotations from IntegrAll, but in the case of cassettes with potentially dubious 
annotations, alternative (alt) UTRs were also tested (see below). The 5′UTRs selected here varied 
considerably in size from 3 to 86 bp (Figure 50B). Eleven out of 13 aadA cassettes in our collection 
had the same 5′ UTR, but were cloned and treated independently as biological replicates (mauve 
dots in Figure 50A and onwards). This was also the case for the 5′UTRs of aacC2 and aacC6 
(brown dots in Figure 50A). This provides a control of technical and biological variability in our 
assays.  

 
Once this collection of 5′ UTRs from AgR cassettes was established and verified, we 

included as additional controls: the pMBA vector; a plasmid-less MG1655 strain that does not 
contain a gfp gene (MG); three constructions with the 5′UTRs of integron cassettes that confer 
resistance to unrelated antibiotics; such as trimethoprim (dfrA5), fosfomycin (fosG), and beta-
lactams (blaOXA-9); and a thermometer riboswitch that responds to temperature, not antibiotics196. 

 
 

4.2.1.3 AgR cassettes are not repressed in the absence of antibiotics. 
 

To characterize the expression of AgR cassettes in resting conditions, we measured the 
fluorescence of all 5′ UTR-gfp fusions in the absence of antibiotics (Figure 50C). Fluorescence 
levels varied 170-fold between the least (pMBA5′-aacA38 ) and the most expressed (pMBA5′-aacA42) 
strains, and the rest of pMBA5´ strains were evenly distributed throughout this scale.  

 
To put these levels of expression in context, we have included as a landmark of expression 

a PBAD promoter in repressed, unrepressed and induced conditions (yellow dots in Figure 50C). 
This promoter is known to have a broad dynamic range with good levels of induction and 
repression. pMBA5´ derivatives with the highest expression levels were similar to the induced PBAD. 
Instead, the least expressed strain was 10-fold higher than the repressed PBAD or the no-GFP 
control (MG1655 strain). pMBA5´ strains derived from cassettes conferring resistance against 
other antibiotics did not group together in a qualitatively distinct expression range, but instead 
were found among the least and the most expressed strains.  

 
 
 
 



Results: A deeper look into the integron model 
 

  118 

Thus, the differences in expression levels among pMBA5´ derivatives do not support a 
common repressing mechanism. Contrarily, expression levels correlated well (r2 = 0.69) with the 
presence of a Shine-Dalgarno sequence in the vicinity of the start codon (Supplementary Figure 
S8). The net expression levels of most pMBA5´ derivatives are incompatible with a biologically 
relevant repressed status, as one would expect for an uninduced riboswitch. 
 

 
Figure 50. Aminoglycoside resistance cassettes are not repressed in the absence of antibiotics. A) 
Tree showing the phylogenetic relationship of the 64 AgR proteins found in integron cassettes. The 
enzymatic activity is marked with coloured lines. Proteins encoded in cassettes with identical 5′ UTRs are 
marked with mauve and brown dots, and pMBA5′-aacA5 is marked with a red dot. B) Size distribution of 5′ 
UTRs within AgR cassettes. C) Expression of AgR cassettes in resting conditions measured as the 
fluorescence of 5 ́UTR-gfp fusions. Expression levels of the PBAD promoter are depicted as landmarks of 
expression (yellow dots). Controls include the MG1655 strain without the gfp gene (MG: black dot), pMBA 
(green dot), cassettes conferring resistance to unrelated antibiotics (light blue dots) and the thermometer 
riboswitch (dark blue dot). As in panel A, constructions with identical 5′ UTRs are marked with mauve and 
brown dots, and pMBA5′-aacA5 with a red dot. Modified from 107. 
 
 

A B 

C 



Results: A deeper look into the integron model 
 

 119 

4.2.1.4 Aminoglycosides do not induce the expression of AgR cassettes. 
 

As mentioned previously, aminoglycosides are a large family of antibiotics that can be 
classified in three subclasses attending to their chemical structure: 4,5- and 4,6-disubstituted 
deoxystreptamines (-DD), and the non-deoxystreptamines. Most clinically relevant molecules like 
kanamycin, sisomicin, gentamicin or amikacin are 4,6-DDs, while neomycin and paromomycin are 
4,5-DDs, and streptomycin is a non-DD. Induction of aacA5 was shown to occur in the presence 
of 4,6- but not 4,5-DDs. Therefore, we will use ‘inducing’ Ags to refer to 4,6-DDs and ‘non- 
inducing’ for 4,5-DDs. 

 
We aimed to better verify the inducibility of all AgR cassettes by aminoglycosides by 

broadening the molecules tested to include the inducers gentamicin and amikacin. All antibiotics 
were used at 1/2 of the MIC to provide conditions in which concentrations are maximal before 
strongly inhibiting bacterial growth (0.5 µg/mL kanamycin, 0.06 µg/mL sisomicin, 0.5 µg/mL 
amikacin, 0.125 µg/mL gentamicin). As a control, we included the non- inducing aminoglycoside 
neomycin (1/2 MIC = 4 µg/mL). Correct growth was verified for all conditions (Supplementary 
Figure S9). 

 
We calculated the induction ratio (IR) of all cassettes as the ratio between the fluorescence 

in the presence over the absence of the antibiotic, so that non-inducible genes should have an IR 
around 1, while inducible genes should have an IR > 1. As an example, the IR of the PBAD 
promoter in Figure 50C is 170. IRs of all cassettes showed distributions with a median close to 1 
in the presence of inducer aminoglycosides (0.99 for kanamycin, 1.05 for sisomicin, 1 for amikacin 
and 1.07 for gentamicin) (Figure 51A). The non-inducer neomycin showed the highest median of 
induction (1.077), although very similar to that of gentamicin. Maximal and minimal IR values 
observed were around 1.3 and 0.8 respectively for all Ags except for gentamicin that showed a 
higher dispersion (1.56–0.63). We also tested alternative UTRs for cassettes with potentially 
dubious annotations, and showed a similar non-inducible profile (Supplementary Figure S10).  

 
The small increase observed for maximal IR values argues against a biologically relevant 

induction for any gene. Instead, the distribution of IR values suggests that variations are likely due 
to noise, whether biological or technical. To address this, we examined more closely the 11 
constructions with identical 5´UTRs (from aad ICs) that were treated as independent biological 
replicates. Their IR values showed differences that comprised a large part of the variation in the 
whole dataset (Figure 51A, mauve dots). Taking kanamycin as an example, the IR values of 76% 
(49/64) AgR cassettes fell between the highest and lowest IR of these identical 5´UTRs. If we 
include the controls that are not related to AgR, this figure reached 81% for kanamycin and 
sisomicin, 72% for gentamicin, 87% for amikacin and 84% for neomycin.  

 
This strongly suggests that the variability found among genes is non-specific, and that it 

most likely reflects biological or technical noise, and/or the pleiotropic effect of the antibiotic.  
 
 
 



Results: A deeper look into the integron model 
 

  120 

We performed a statistical analysis of our data comparing fluorescence levels through 
linear regression. We did not find significant differences in the presence of an aminoglycoside 
compared with the baseline readings in the absence of antibiotic (p > 0.3). We then applied a 
linear mixed model accounting for genes and experimental batches as random effects, and 
observed significant changes in the fluorescence readings in the presence of gentamicin and 
amikacin, but not kanamycin, sisomicin nor neomycin (Supplementary Figure S11). These effects 
were extremely mild, with a mean increase in expression of 5% for gentamicin, and a mean 
decrease of 7.5% for amikacin.  

 
Figure 51.  Aminoglycosides do not induce the expression of aminoglycoside resistance cassettes. 
A) Violin plots showing the induction ratios of all cassettes in the presence of four inducing aminoglycosides, 
and the non-inducing neomycin. Colour code of the dots as in Figure 49. Median and quartiles are 
represented by dashed lines. Linear regression analysis did not find significant differences (p > 0.3) in 
fluorescence readings in the absence versus the presence of Ags. Each data point represents the mean of 
at least three measurements. B) Induction ratio measured through Western blot of the 10 pMBA5´ derivatives 
with highest and lowest IRs in kanamycin measured by flow cytometry (orange and violet boxes in panel A). 
GFP abundance was normalized to DnaK. Cytometry and Western blot results do not correlate (inset: linear 
regression r2 = 0.02). Figure from 107 

A 

B 



Results: A deeper look into the integron model 
 

 121 

We have also quantitated GFP protein in the presence and the absence of kanamycin 
using Western blot (Figure 51B) as we previously did for polar effects characterisation. We have 
analyzed ten strains showing the highest IR values with kanamycin, and have compared them to 
the ten pMBA5´ derivatives with the lowest IRs (orange and violet boxes in the kanamycin violin 
plot in Figure 51A). Cytometry and WB data clearly show a lack of correlation (r2 = 0.02) (inset in 
Figure 51B) suggesting that genes with high IRs in cytometry do not represent a distinct subset of 
cassettes with a specific induction mechanism.  

 
Altogether, the lack of coherent effects among inducer molecules and of biological 

significance of the induction ratios, rules out the presence of a common regulatory mechanism 
capable of sensing aminoglycosides and inducing gene expression. 
 
4.2.1.5 Higher concentrations of Ags neither induce resistance genes. 
 

The concentrations of antibiotics used for in vivo experiments in the first report of the 
putative riboswitch121 were close or above the minimal inhibitory concentration of a susceptible E. 
coli K-12 derivative such as their (and our) strain. Indeed, the highest induction level they observe 
for kanamycin is at 2.4 µg/ml (5 µM), in a strain with a MIC around 1 µg/ml. Therefore, we decided 
to address whether riboswitches could be acting at concentrations above the MIC, despite not 
conforming to a biologically sound rationale. To do so, we introduced the AgR gene armA in a 
subset of our strains. ArmA is a 16S rRNA methylase that provides high levels of resistance 
against 4,6-DDs, but does not modify the antibiotic. Crucially, armA is not encoded in an integron 
cassette197. We repeated induction experiments of cassettes in the presence of 256-fold higher 
concentrations of kanamycin and confirmed that they remain uninduced (Figure 52). 

Figure 52.  High concentrations of aminoglycosides do not induce AgR cassettes. A) Induction ratio 
of a subset of AgR cassettes in the presence of 128 µg/ml of kanamycin (blue dots). As a reference, the IR 
values at 0.5 µg/ml of kanamycin from Figure 50 are also represented (green dots). B) Induction ratio 
(calculated as the ratio of fluorescence/OD600 with kanamycin over fluorescence/OD600 without kanamycin) 
of these strains along their growth curve (18 h). Colour code is as in A). Green: pMBA; red: pMBA5′-aacA5; 
blue: control strains; mauve: strains with identical 5′UTRs; grey: rest of strains. 

A 

B 



Results: A deeper look into the integron model 
 

  122 

4.2.1.6 Induction is higher in the presence of unrelated antibiotics. 
 

Our experiments at 1/2 the MIC show that aminoglycosides can have a very mild and non-
specific influence in the measures of IRs. To provide a broader context to our data we have tested 
the effect of unrelated antibiotics in the induction of all 5′UTRs (Figure 53). We chose molecules 
with different modes of action, such as tetracycline (1/2 MIC = 0.125 µg/mL); a protein synthesis 
inhibitor, like aminoglycosides, but with a distinct mode of action; ciprofloxacin (1/2 MIC = 0.008 
µg/mL), a topoisomerase inhibitor; and trimethoprim (1/2 MIC = 0.06 µg/mL), that inhibits the 
biosynthesis of folate.  

 
With these antibiotics, the medians of IR were all higher than with aminoglycosides 

(tetracycline: 1.08; ciprofloxacin: 1.26; trimethoprim: 1.19), and estimated differences in our linear 
mixed model due to the antibiotic were larger than those associated with the addition of 
aminoglycosides (Supplementary Figure S11). No construction showed an IR > 2 in any antibiotic, 
and >93% of the IR values of AgR cassettes fell within those of control strains, pointing to a non-
specific effect of these molecules on the expression of AgR cassettes. To address if there is any 
cassette with a mild but consistent inducible behaviour across our experiments, we analysed 
independently the IRs of each 5´UTR in the presence of inducing aminoglycosides (4,6-DDs) and 
non-inducing antibiotics (4,5-DDs and unrelated antibiotics) (Supplementary Figure S11). Our 
data clearly shows that 4,6-DDs do not induce the expression of AgR cassettes to biologically 
relevant levels in any case. The stronger effects of other antibiotics are also evident in this analysis 
since 83% (53/64) of the genes had higher median IRs in the presence of non-inducing molecules 
compared to 4,6-DDs, and statistically significant differences were observed in 10 of them. 

 
Figure 53. Induction is higher in the presence of unrelated antibiotics. Violin plots showing the 
induction ratio of all genes in the presence of non-aminoglycoside antibiotics. Colour code as in Figure 50. 
Median and quartiles are represented by dashed lines. Statistically significant differences compared to the 
no-antibiotic condition are marked with **(p < 0.01).  Each data point represents the mean of at least three 
measurements.  
 
 

0.5

1.0

1.5

2.0

In
du

ct
io

n 
ra

tio

TetracyclineCiprofloxacin

PROTEIN SYNTHESISDNA TOPOLOGY FOLATE SYNTHESISAffected
Process

Trimethoprim MH

**



Results: A deeper look into the integron model 
 

 123 

4.2.1.7 Increased expression is due to pleiotropic effects of antibiotics. 
 

Other authors as Roth and Breaker suggested that the mild induction observed by Murchie 
and collaborators could be due to pleiotropic effects of antibiotics181. Conveniently, we can test 
this experimentally because the presence of trimethoprim can be decoupled from its antibiotic 
effect, by measuring IRs in Lysogeny Broth (LB), instead of Müller Hinton (MH). In contrast to MH, 
LB contains traces of thymine that render the folate route redundant so that trimethoprim is not 
lethal in LB. We analyzed the induction of all cassettes in the presence of trimethoprim in LB and 
compared them to our previous results in MH (Figure 54). The observed mean of IR values 
decreased from 1.17 to 0.98 in the absence of antibiotic and this effect was statistically significant 
using both linear regression and a random linear mixed model (p < 0.0001). This confirms that 
pleiotropic effects of antibiotics can alter the expression of genes 1.5-fold range in a non-specific 
manner, which is approximately the range observed by Jia, et al. 

 
Figure 54.  Increased expression is due to pleiotropic effects of antibiotics. Induction ratio of all 
cassettes in the presence of trimethoprim in LB compared to MH (data from Figure 52). Colour code as in 
Figure 49. Median and quartiles are represented by dashed lines. Statistically significant differences 
compared to the no-antibiotic condition are marked with ****(p < 0.0001).  Each data point represents the 
mean of at least three measurements.  

Altogether, our results rule out the existence, in integron AgR ICs, of a regulatory 
mechanism that induces gene expression in the presence of aminoglycosides. It therefore 
challenges the validity of the Ag-sensing riboswitch.  





 

  

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

DISCUSSION 

 
 

 

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Discussion 
 

 127 

5 Discussion 
 

Antimicrobial resistance is a major threat for global health, causing between 1.5 and 5 
million deaths per year24. Evolution towards resistance is mainly driven by the dissemination of 
resistance determinants through horizontal gene transfer facilitated by various agents, including 
phages, plasmids, transposons, and mobile integrons. Notably, integrons are key elements in 
multidrug resistance among Gram negatives, harbouring more than 170 resistance determinants 
against 12 antibiotic families122. This, together with their high prevalence in clinical isolates make 
integrons crucial genetic elements for the study and tackling of antimicrobial resistance. 

 
While the success of mobile integrons among clinical and environmental isolates is 

undeniable, the prevalence of each IC in databases varies significantly, even among ICs 
conferring the same phenotype. We believe that various evolutionary forces, such as antimicrobial 
resistance, fitness cost, polar effects, and mobility, as well as the factors that modulate these 
forces, play a significant role in driving the success of each IC. In the literature, we can find studies 
that focus on characterizing each of these forces separately. However, the data is scattered and 
reported in different genetic backgrounds and conditions, which makes impossible to conduct 
comparable analysis. 

 
The establishment of the pMBA collection facilitates a comprehensive and comparable 

study of all features potentially associated with the success of each ARC in its biologically-sound 
genetic environment. It is worth noting that this collection represents the most extensive 
compilation of ARCs to date, encompassing 136 ARCs (Figure 13). However, as mentioned in the 
Results section, this number accounts for 77% of all ARCs found in mobile integrons, which 
constitutes a limitation of our study.  

 
More than half of the non-cloned ARCs (22 out of 41) belong to the bla ARC family. In our 

experiments, most cloned members of this family impose a significant fitness cost to their host, 
which may explain our limitation in cloning blas in E. coli. For instance, when we examine the 
distribution of ARCs among bacterial species, blas constitute a small percentage of all ARCs in E. 
coli (4.1%), in contrast to other species like Pseudomonas spp. (34.4%). An analysis of the 
distribution by resistance mechanism reveals a similar pattern, with nearly 50% of bla ARCs found 
in Pseudomonas spp., while only 7% are present in E. coli (Figure 55). These data highlight the 
potential existence of species-specific barriers to horizontal gene transfer mediated by 
integrons122, phenomenon that would be worth assessing in the future. 

 
In addition, the high expression levels achieved in the pMBA vector, a consequence of its 

copy number and the PcS promoter, could hinder the cloning of costly ARCs, such as blaVEB-1, 
which has been successfully cloned in R388 under a PcW promoter198 but not in pMBA. However, 
we consider that our genetic context, in which ARCs are strongly expressed, offers more benefits 
than drawbacks, as it allows us to distinguish subtle phenotypic differences between the studied 
ARCs. 
 
 



Discussion 
 

  128 

 
 
Figure 55.  Distribution of ARCs by antibiotic class among different bacterial genus.  Each column 
represents the whole set of reports of ARCs against a given antimicrobial family available in IntegrAll (each 
ARC can be reported more than once). Colours represent the bacterial genus in which it was found.  
 

Characterizing the resistance profile of each ARC not only provides essential information 
for understanding their success but also serves as a valuable guide in clinical contexts. By 
coupling MICs to hierarchical clustering trees, we observe that profiles do not follow strictly the 
identity signal between proteins, highlighting that genes annotated automatically might not behave 
as predicted. Indeed, some ARCs do not follow the resistance pattern of the family, like aadB, that 
shows a resistance profile closer to that of acetylases than to other adenylases (Figure 14). In 
other cases, closely related genes such as blaOXA-1 and blaOXA-9 (Figure 15) show different 
resistance profiles against cephalosporins and aztreonam, similarly to what we see for aadA4 and 
aadA5 in which the latter confers high streptomycin resistance, while the former (the closest 
related protein) does not (Figure 15). 

 
 In more extreme cases, the annotation of a gene as a resistance gene might be mistaken, 

as for aadA10 or aacAX (Figure 14); or at least it is unlikely that these genes are clinically relevant, 
like for aacA16 and fosM (Figures 14 and 17). The most striking case revealed by our results is 
that of complete families of genes that do not confer resistance against the compounds described 
in the literature in E. coli. Such is the case of qac, and smr, that did not rise the MIC against any 
quaternary ammonium compound tested (Figure 18), although we cannot rule out a stronger 
impact of these determinants in different conditions, such as in biofilms155, or species.  
 

Beta
-la

cta
ms

Amino
gly

co
sid

es

Chlo
ram

ph
en

ico
l

Tri
meth

op
rim

Fos
fom

yc
in

Small
 dr

ug
s

Rifa
mpic

in

Mac
rol

ide
s

Quin
olo

ne
s0

50

100

Antibiotic class

%
 

Distribution of ARCs among genus

Acinetobacter (n=415)

Citrobacter (n=200)

Enterobacter (n=336)

Aeromonas (n=261)

Klebsiella (n=1259)

Escherichia (n=1512)

Pseudomonas (n=1055)

Proteus (n=187)

Salmonella (n=647)

Vibrio (n=101)

n=1960 n=5624 n=857 n=2668 n=19 n=401 n=413 n=149 n=37



Discussion 
 

 129 

In this context, the fact that some ARCs do not appear to confer resistance in our 
experiments does not necessarily imply that these genes would behave similarly in other genetic 
backgrounds. As an example, it is also noteworthy that R388 carrying aadB in first position confers 
significantly higher resistance levels to gentamicin in P. aeruginosa (64-fold)198, compared to E. 
coli (4-fold) (Figure 19B). Therefore, it becomes essential to expand this resistance 
characterization to other clinically relevant bacteria from the ESKAPEE family, as it is crucial to 
effectively combat antimicrobial resistance. 
 

On a different note, an interesting aspect of this resistance profiling is its potential impact 
in biotechnology. Integron ARCs have been broadly used as resistance markers, and sometimes 
the availability of different markers is a limiting factor when delivering strains with multiple genetic 
modifications. Our results provide researchers with a useful guide with options that are 
counterintuitive for many, like the possibility of using erythromycin resistance genes in E. coli or 
the possibility of selecting 5 consecutive markers within a single family of antibiotics: for example 
aacC11 and aadA5 can be independently selected with gentamycin and streptomycin. Then, 
respecting the order, aphA16 can be selected with kanamycin; aacA34 with tobramycin; and 
aacA49 with amikacin (Figure 14). 

 
Together with the resistance conferred, the cost entailed by an antimicrobial resistance 

gene is one of the main drivers of its success. We have quantitated the fitness effect of each ARC 
under different conditions such as aerobic, anaerobic and in the presence of a controlled 
microbiota. It is worth noting, that to bring our data closer to the clinical reality we also performed 
in vivo competition experiments to complement and reinforce data acquired in vitro.  

 
The cost associated with each ARC varies in our experiments depending on the 

environmental conditions and is partially linked to the encoded resistance mechanism. 
Competition assays show a strong tendency toward cost associated with bla ARCs, independently 
of the experimental conditions. This, together with their low prevalence in E. coli isolates described 
in the IntegrAll database (Figure 55) and our cloning limitations linked to this family of ARCs, 
supports the hypothesis of species-specific barriers to horizontal gene transfer in integrons. 

 
We could apply the same hypothesis to explain the differential prevalence of individual 

ARCs, expecting that the costlier ARCs may be rare in databases, while beneficial ARCs should 
be reported more often. This hypothesis holds in some cases, with aadA5 being the most reported 
ARC in class 1 MIs in E. coli and also conferring a growth benefit to E. coli in our experiments (9% 
and 2% in aerobiosis and anaerobiosis, respectively) (Figures 29 and 33). However, this 
approximation could be biased due to the lack of metadata for the samples submitted to 
databases. Several factors, such as the sample origin (clinical or environmental), the MGE that 
carries the integron, and, in clinical samples, specific antibiotic exposure, play a crucial role in the 
selection of specific arrays or ARCs. Therefore, the fitness effect of each ARC in databases cannot 
be directly correlated with its prevalence and should be considered only as a possible explanation 
for its success. 
 



Discussion 
 

  130 

One of the most remarkable results of this thesis is the fact that some ARCs, in certain 
conditions, confer a gain in fitness to their hosts in the absence of selective pressure both in vivo 
and in vitro. This finding suggests that certain ARCs play an accessory role, along with their 
function as resistant determinants, enhancing bacterial growth. These beneficial ARCs might play 
a key role in the spread and stabilization of plasmid-mediated resistance in antibiotic-free 
conditions, breaking the general fitness-resistance trade-off199 without needing compensatory 
mutations to alleviate the “cost” derived from ARCs200,201. 

 
It is known that several of the so-called ARGs may not act as resistance determinants in 

their native host context, displaying their function in bacterial metabolism202. In this line of thought, 
we suggest that ARCs conferring a gain in fitness in the absence of selective pressure could be 
playing a dual role, conferring resistance to antimicrobials and participating in bacterial 
metabolism. As an example, we have discovered that ereA2, in aerobiosis, modulates the 
transcriptome of E. coli repressing motility-related genes (Figure 36). This transcriptomic 
modulation could be related to the gain in fitness conferred by this ARC in aerobic competitions, 
where saving energy destinated to motility in shaking conditions could lead to a growth benefit, 
while in anaerobiosis and in vivo might be detrimental causing great fitness cost to the bacterium.  
 

This dual role of certain ARCs could explain the significant prevalence of ARCs that confer 
low resistance levels or even fail to confer resistance, such as qac and smr ICs. Nevertheless, 
further studies are needed to confirm this hypothesis and unveil these hidden accessory functions. 

 
Another observation worth considering is the influence of oxygen in fitness effects of most 

ARCs (Figure 34). This phenomenon could lead to strikingly different fitness effects due to 
environmental changes or in clinical settings, which could be key in the ecology and maintenance 
of these ARCs, even if they are costly to the bacterium under certain conditions. Using ereA2 as 
an example again, this ARC is beneficial under aerobic conditions but detrimental in anaerobiosis 
and in vivo competition assays. This duality in fitness effect could explain why ereA2 is rarely 
found in databases, where clinical isolates are overrepresented compared to environmental 
samples, or when found, it is often truncated. 

 
Nevertheless, all these hypotheses are only valid for cassettes located in the first position 

of the array. ARCs located further from the dedicated Pc promoter are less expressed and, 
therefore, entail a lower cost. Thus, the operon-like structure of the integron and the modulation 
of ARCs expression by distal ICs within the platform make direct correlations between prevalence 
and fitness effect a simplistic approximation to a complex phenomenon. Integrons can harbour 
more than one ARC, forming IC arrays. We have demonstrated that the identity of an ARC can 
modulate the expression of the following ICs in the array at a transcriptional level, adding an extra 
layer of complexity to the canonical operon-like integron model. This phenomenon has been 
demonstrated for a subset of ARCs from the dfr family (Figure 23). However, further experiments 
with a broader subset of ARCs from different families would be necessary to generalize this 
regulation.  

 



Discussion 
 

 131 

This finding has important clinical implications, impacting in potential co-selection 
phenomena during antibiotic treatment when bacteria are exposed to different antimicrobials and 
antiseptics. In this scenario, it is crucial to understand that ARCs in distal positions of the integron 
array can be expressed and exert their functions as antimicrobial resistance genes, depending on 
the identity of previous ICs in the array. These arrays can be selected during antibiotic treatments, 
fixing a resistant bacterial population both in patients and clinical environments, despite their 
higher cost in the absence of selection. Hence, ARCs present in mobile integrons could be defined 
by the resistance conferred, the fitness cost entailed, and their capacity to modulate the 
expression of the cassettes in the array. Nevertheless, one of the main features of ICs is their 
capacity to reshuffle within the platform under stress conditions and their mobility between 
integrons via excision and recombination. The differential recombination frequency of each IC 
should also be taken into account when exploring the forces that drive the success of each ARC. 

 
The difference in the recombination frequency between ARCs is classically attributed to 

attC site structural features and sequence95,96,178. In the classic recombination assay, a donor 
strain delivers an attC site as ssDNA into the recipient strain to recombine with an attI site 
structured as dsDNA. Therefore, the folding of each attC site into a recombinogenic structure, as 
well as its sequence, are key for successful recombination. Conversely, our modified 
recombination assay adds an extra parameter to the process, as the attC site (present in the 
recipient strain as dsDNA) needs to be extruded from a dsDNA sequence to form a 
recombinogenic structure as a ssDNA hairpin, as it would happen naturally in excision reactions. 

 
 This extra processing of the recombination substrates might explain why recombination 

rates observed in our experiments tend to be significantly lower than the ones published in the 
literature174. In addition, the wide range of recombination rates observed in our assays (from 10-2 
to 10-8) (Figure 42) could be due to the differential ability of each attC to acquire its recombinogenic 
conformation from a dsDNA, which is not tested when the attC site is delivered following the classic 
recombination assay. 

 
This modification in the recombination assay might also explain why in silico parameters 

such as DG and pfold do not directly correlate with the recombination frequencies measured in 
vitro following this modified protocol (Figure 46). In our experiments, attC sites are initially dsDNA 
sequences before their extrusion, while in silico parameters are calculated considering the attC a 
ssDNA sequence. Therefore, in silico calculation does not take into account this process. To 
correlate both in silico and in vitro results we should find a way to quantitate the energy needed 
for a specific attC site to be extruded from dsDNA to a ssDNA conformation before structuring as 
a ssDNA hairpin. 
 

It is remarkable that AgR cassettes show higher recombination rates on average than the 
rest of ARC families (Figure 43). This observation might correlate with any distinctive feature of 
the attC site such as a potential similarity in sequence. It could be also possible that the co-
evolution of the attC site´s sequence with the antimicrobial resistance gene encoded might lead 
to this observation.  

 



Discussion 
 

  132 

Mobile integrons are believed to appear from mobilised cassettes of sedentary 
chromosomal integrons, where attC site sequences are very conserved within the same 
integron.70,203. Thus, ARCs arising from close origins/environments might contain attC sites with 
similar sequences. A hierarchical clustering tree of the nucleotidic sequences of all attC sites in 
the pMBA collection (Supplementary Figure S12) shows great differences in sequence between 
ARCs, particularly in the AgR genes family, ruling out the idea of a common origin of AgR genes 
with high recombination rates. 

 
We also wonder if any specific feature of the antimicrobial resistance gene encoded in the 

cassette might influence attC site structuration and hence, the recombination frequency of the 
cassette. It is demonstrated that high GC-content near the apical loop of attC sites plays a crucial 
role in attC folding and recombination204. In this line, gene GC-content might also have an 
influence in this process as a proxy of the bonding strength of the dsDNA in the vicinity of the attC 
site structure, which needs to get extruded and form a ssDNA hairpin. If we correlate 
recombination frequency with GC content of the encoded gene we find a significant but mild 
positive correlation (r = 0.3130) (Figure 56A).  

 

Figure 56.  Correlation between ARC recombination rate and GC content of the gene encoded.  
Graphs showing the general correlation of all ARCs (A) or specific ARC families (B). Spearman´s rank 
correlation coefficient (r) and p-values are indicated. 
 

This significance is conserved in the AgR family but disappears in the rest of ARC families 
(Figure 56B), potentially due to the smaller sample size of these families. However, a higher 
correlation is observed in bla and fos ARC families (r > 0.4), suggesting a stronger effect of the 
GC content on the recombination frequency. While there is a correlation between gene GC content 
and recombination frequency, this correlation is mild, and further experiments are needed to affirm 
that the gene GC content influences the recombination frequency of the ARC.  

 
 
 
 
 

10-8 10-7 10-6 10-5 10-4 10-3 10-2 10-1
30

40

50

60

R. F.

%
G

C

ρ = 0.3130
p-value = 0.0004

10-8 10-7 10-6 10-5 10-4 10-3 10-2 10-1
30

40

50

60

R. F.

%
G

C

AgR genes

ρ = 0.3007
p-value = 0.0231

10-8 10-7 10-6 10-5 10-4 10-3 10-2 10-1
30

40

50

60

R. F.

%
G

C

dfrs 

ρ = 0.2931
p-value =  0.1379

10-8 10-7 10-6 10-5 10-4 10-3 10-2 10-1
30

40

50

60

R. F.

%
G

C

bla

ρ = 0.4363
p-value = 0.0618

 

10-8 10-7 10-6 10-5 10-4 10-3 10-2 10-1
30

40

50

60

R. F.

%
G

C

fos

ρ = 0.4667
p-value = 0.1786 

10-8 10-7 10-6 10-5 10-4 10-3 10-2 10-1
30

40

50

60

R. F.

%
G

C
qacs, smr

ρ = 0.2121
p-value =  0.5603

A B 



Discussion 
 

 133 

Apart from the already mentioned general evolutionary forces ruling the fate of ARCs, 
certain specific regulatory elements have being described to participate in ARC function and 
success121,179,180. In the second part of this PhD thesis, we have also provided experimental 
evidences against the existence of one of these regulatory elements: a controversial Ag-sensing 
riboswitch present in the integron platform121,193,194. This study emphasizes the importance of 
revisiting the existing literature, challenging it, and building a solid base to continue expanding  our 
knowledge. 

 
In summary, this PhD thesis offers a detailed quantitation of the forces that we believe 

govern the success of ARCs: antimicrobial resistance, fitness cost, polar effects, and mobility. To 
achieve this, we have generated the most extensive collection of ARCs in their native environment 
to date, the pMBA collection122. Although we are still far from a complete understanding of the 
success of each ARC, this work contributes to an holistic approach in explaining the ecology of 
ARCs and presents novel and promising observations regarding the evolutionary dynamics that 
determine their fate. 
 
 
 
 
 
 
 
 





 

  

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

CONCLUSIONS 

 
 

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Conclusions 

 137 
 

6 Conclusions  
 

I. Closely related antimicrobial resistance cassettes (ARCs) confer drastically different 
resistance levels even finding ARCs not conferring resistance, suggesting possible 
mistaken annotations of them as resistance determinants. 
 
 

II. Integron cassette (IC) identity modulates downstream expression of the array at a 
transcriptional level.  
 
 

III. ARC fitness effect is variable being some of them beneficial for bacterial growth in the 
absence of any selective pressure both in vitro and in vivo. 
 
 

IV. Anaerobiosis is a crucial condition affecting the fitness dynamics of ARCs. 
 
 

V. ARCs such as ereA2 can modulate bacterial success independently of their antimicrobial 
resistance function via transcription regulation. 
 
 

VI. ARC recombination frequencies vary widely across cassettes tested, finding a positive 
correlation between aminoglycoside resistance cassettes and high recombination rates.  
 
 

VII. The expression of aminoglycoside resistance genes in integron cassettes is not controlled 
by riboswitches. 

 
 





 

  

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

BIBLIOGRAPHY 

 
 

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Bibliography 

 141 
 

7 Bibliography  
 
1. Waksman, S. A. What is an antibiotic or an antibiotic substance? Mycologia 39, 565–569 

(1947). 
2. Singleton, P., & Sainsbury, D. Dictionary of microbiology and molecular biology (3rd ed. 

rev). Wiley. (2006). doi:10.1002/9780470056981 
3. Haas, L. F. Papyrus of Ebers and Smith. J. Neurol. Neurosurg. Psychiatry 67, 578 LP – 578 

(1999). 
4. Gensini, G. F., Conti, A. A. & Lippi, D. The contributions of Paul Ehrlich to infectious 

disease. J. Infect. 54, 221–224 (2007). 
5. Otten, H. Domagk and the development of the sulphonamides. J. Antimicrob. Chemother. 

17, 689–690 (1986). 
6. Fleming, A. On the antibacterial action of cultures of a penicillium, with special reference to 

their use in the isolation of B. influenzae. The British Journal of Experimental Pathology 10, 
226–236 (1929). 

7. Chain, E. et al. THE CLASSIC: penicillin as a chemotherapeutic agent. 1940. Clin. Orthop. 
Relat. Res. 439, 23–26 (2005). 

8. Brunel, J. Antibiosis from Pasteur to Fleming*. J. Hist. Med. Allied Sci. VI, 287–301 (1951). 
9. Waksman, S. A., Schatz, A. & Reynolds, D. M. Production of antibiotic substances by 

actinomycetes. Ann. N. Y. Acad. Sci. 1213, 112–124 (2010). 
10. Hutchings, M., Truman, A. & Wilkinson, B. Antibiotics: past, present and future. Curr. Opin. 

Microbiol. 51, 72–80 (2019). 
11. Hodgkin, D. C. The X-ray analysis of the structure of penicillin. Adv. Sci. 6, 85–89 (1949). 
12. Brown, E. D. & Wright, G. D. Antibacterial drug discovery in the resistance era. Nature 529, 

336–343 (2016). 
13. Durand, G. A., Raoult, D. & Dubourg, G. Antibiotic discovery: history, methods and 

perspectives. Int. J. Antimicrob. Agents 53, 371–382 (2019). 
14. Kealey, C., Creaven, C. A., Murphy, C. D. & Brady, C. B. New approaches to antibiotic 

discovery. Biotechnol. Lett. 39, 805–817 (2017). 
15. Christaki, E., Marcou, M. & Tofarides, A. Antimicrobial Resistance in Bacteria: Mechanisms, 

Evolution, and Persistence. J. Mol. Evol. 88, 26–40 (2020). 
16. Dcosta, V. M. et al. Antibiotic resistance is ancient. Nature 477, 457–461 (2011). 
17. Stennett, H. L., Back, C. R. & Race, P. R. Derivation of a Precise and Consistent Timeline 

for Antibiotic Development. Antibiotics 11, (2022). 
18. Torres, P. B. Inducible and acquired antibiotic resistance in stenotrophomonas maltophilia. 

(Universidad Autónoma de Madrid Faculty of Science, 2019). 
19. Levin, B. R. et al. The population genetics of antibiotic resistance. Clin. Infect. Dis. 24, 

(1997). 
20. Andersson, D. I. & Levin, B. R. The biological cost of antibiotic resistance. Curr. Opin. 

Microbiol. 2, 489–493 (1999). 



Bibliography 

  142 

21. Mitshuhashi, S., Harada, K., Hashimoto, H. & Egawa, R. On the drug-resistance of enteric 
bacteria. 4. Drug-resistance of Shigella prevalent in Japan. Jpn. J. Exp. Med. 31, 47–52 
(1961). 

22. Partridge, S. R. & Hall, R. M. Complex multiple antibiotic and mercury resistance region 
derived from the r-det of NR1 (R100). Antimicrob. Agents Chemother. 48, 4250–4255 
(2004). 

23. O´Neill, J. Antimicrobial Resistance: Tackling a crisis for the health and wealth of nations 
The Review on Antimicrobial Resistance Chaired by J O’Neill, and supported by the 
Wellcome Trust and the UK Government. (2014). doi:10.1038/510015a 

24. Murray, C. J. et al. Global burden of bacterial antimicrobial resistance in 2019: a systematic 
analysis. Lancet 399, 629–655 (2022). 

25. Abraham, E. P. & Chain, E. An enzyme from bacteria able to destroy penicillin. 1940. Rev. 
Infect. Dis. 10, 677–678 (1988). 

26. Shaw, K. J., Rather, P. N., Hare, R. S. & Miller, G. H. Molecular genetics of aminoglycoside 
resistance genes and familial relationships of the aminoglycoside-modifying enzymes. 
Microbiol. Rev. 57, 138–163 (1993). 

27. Shaw, W. V. The enzymatic acetylation of chloramphenicol by extracts of R factor-resistant  
Escherichia coli. J. Biol. Chem. 242, 687–693 (1967). 

28. Quan, S. et al. ADP-ribosylation as an intermediate step in inactivation of rifampin by a  
mycobacterial gene. Antimicrob. Agents Chemother. 43, 181–184 (1999). 

29. Aldred, K. J., Kerns, R. J. & Osheroff, N. Mechanism of quinolone action and resistance. 
Biochemistry 53, 1565–1574 (2014). 

30. González-Zorn, B. et al. armA and aminoglycoside resistance in Escherichia coli. Emerg. 
Infect. Dis. 11, 954–956 (2005). 

31. Kordus, S. L. & Baughn, A. D. Revitalizing antifolates through understanding mechanisms 
that govern susceptibility and resistance. Medchemcomm 10, 880–895 (2019). 

32. Poole, K. Efflux-mediated antimicrobial resistance. Journal of Antimicrobial Chemotherapy 
56, (2005). 

33. Sørensen, S. J., Bailey, M., Hansen, L. H., Kroer, N. & Wuertz, S. Studying plasmid 
horizontal transfer in situ: a critical review. Nat. Rev. Microbiol. 3, 700–710 (2005). 

34. Charpentier, X., Polard, P. & Claverys, J. P. Induction of competence for genetic 
transformation by antibiotics: Convergent evolution of stress responses in distant bacterial 
species lacking SOS? Curr. Opin. Microbiol. 15, 570–576 (2012). 

35. Daubin, V. & Szöllősi, G. J. Horizontal Gene Transfer and the History of Life. Cold Spring 
Harb. Perspect. Biol. 8, (2016). 

36. Ficht, T. A. Bacterial exchange via nanotubes : lessons learned from the history of 
molecular biology. 2, 1–4 (2011). 

37. Fischer, S. et al. Indication of Horizontal DNA Gene Transfer by Extracellular Vesicles. 
PLoS One 11, 1–22 (2016). 

38. Redfield, R. J., Schragt, M. R. & Dean, A. M. The Evolution of Bacterial Transformation: 
Sex With Poor Relations. Genetics 146, 27–38 (1997). 

 



Bibliography 

 143 
 

39. Johnston, C., Martin, B., Fichant, G., Polard, P. & Claverys, J.-P. Bacterial transformation: 
distribution, shared mechanisms and divergent control. Nat. Rev. Microbiol. 12, 181–196 
(2014). 

40. Thomas, C. M., Nielsen, K. M. & N, S. P. Mechanisms of, and barriers to, horizontal gene 
transfer between bacteria. Nat. Rev. Microbiol. 3, 711–721 (2005). 

41. Zinder, N. D. Bacterial transduction. 23–49 
42. Haaber, J., Penadés, J. R. & Ingmer, H. Transfer of Antibiotic Resistance in Staphylococcus 

aureus. Trends Microbiol. 25, 893–905 (2017). 
43. Lennox, E. S. Transduction of linked genetic characters of the host by bacteriophage P1. 

Virology 1, 190–206 (1955). 
44. Morse, M. L., Lederberg, E. M. & Lederberg, J. Transduction in Escherichia coli K-12. 

Genetics 41, 142–56 (1956). 
45. Chen, J. et al. Genome hypermobility by lateral transduction. Science (80-. ). 362, 207–212 

(2018). 
46. Frost, L. S., Leplae, R., Summers, A. O., Toussaint, A. & Edmonton, A. Mobile genetic 

elements : the agents of open source evolution. 3, 722–732 (2005). 
47. Partridge, S. R., Kwong, S. M., Firth, N. & Jensen, S. O. Mobile genetic elements associated 

with antimicrobial resistance. Clin. Microbiol. Rev. 31, 1–61 (2018). 
48. Ravin, N. V, Svarchevsky, A. N. & Dehò, G. The anti-immunity system of phage-plasmid 

N15: identification of the  antirepressor gene and its control by a small processed RNA. 
Mol. Microbiol. 34, 980–994 (1999). 

49. Horne, T., Orr, V. T. & Hall, J. P. How do interactions between mobile genetic elements 
affect horizontal gene transfer? Curr. Opin. Microbiol. 73, 102282 (2023). 

50. Hall, J. P. J., Brockhurst, M. A. & Harrison, E. Sampling the mobile gene pool: Innovation 
via horizontal gene transfer in bacteria. Philos. Trans. R. Soc. B Biol. Sci. 372, 1–10 (2017). 

51. Siguier, P., Gourbeyre, E., Varani, A., Ton-Hoang, B. & Chandler, M. Everyman’s guide to 
bacterial insertion sequences. Microbiol. Spectr. 3, (2015). 

52. Bennett, P. Genome plasticity. Methods in Molecular Biology (Humana Press, 2004). 
53. Nicolas, E. et al. The Tn3-family of replicative transposons. Mob. DNA III 693–726 (2015). 

doi:10.1128/9781555819217.ch32 
54. Macesic, N. et al. Genomic dissection of endemic carbapenem resistance reveals  metallo-

beta-lactamase dissemination through clonal, plasmid and integron transfer. Nat. Commun. 
14, 4764 (2023). 

55. Stokes, H. W. & Hall, R. M. A novel family of potentially mobile DNA elements encoding 
site-specific gene-integration functions: integrons. Mol. Microbiol. 3, 1669–1683 (1989). 

56. Escudero JA, Loot C, Nivina A, M. D. The integron: Adaption on demand. Microbiol. Spectr. 
25–32 (2014). doi:10.1128/microbiolspec 

57. Mazel, D. Integrons: agents of bacterial evolution. Nat. Rev. Microbiol. 4, 608–620 (2006). 
58. Fonseca, É. L. & Vicente, A. C. Integron Functionality and Genome Innovation: An Update 

on the Subtle and Smart Strategy of Integrase and Gene Cassette Expression Regulation. 
Microorganisms 10, (2022). 



Bibliography 

  144 

59. Guerin, É. et al. The SOS Response Controls Integron Recombination. Science (80). 324, 
1034 (2009). 

60. Collis, C. M. & Hall, R. M. Gene cassettes from the insert region of integrons are excised 
as covalently closed circles. Mol. Microbiol. 6, 2875–2885 (1992). 

61. Moura, A. et al. INTEGRALL: A database and search engine for integrons, integrases and 
gene cassettes. Bioinformatics (2009). doi:10.1093/bioinformatics/btp105 

62. Collis, C. M. & Hall, R. M. Expression of antibiotic resistance genes in the integrated 
cassettes of integrons. Antimicrob. Agents Chemother. 39, 155–162 (1995). 

63. Szekeres, S., Dauti, M., Wilde, C., Mazel, D. & Rowe-Magnus, D. A. Chromosomal toxin-
antitoxin loci can diminish large-scale genome reductions in  the absence of selection. Mol. 
Microbiol. 63, 1588–1605 (2007). 

64. Bissonnette, L., Champetier, S., Buisson, J. P. & Roy, P. H. Characterization of the 
nonenzymatic chloramphenicol resistance (cmlA) gene of  the In4 integron of Tn1696: 
similarity of the product to transmembrane transport proteins. J. Bacteriol. 173, 4493–4502 
(1991). 

65. da Fonseca, É. L. & Vicente, A. C. P. Functional characterization of a Cassette-specific 
promoter in the class 1  integron-associated qnrVC1 gene. Antimicrob. Agents Chemother. 
56, 3392–3394 (2012). 

66. Biskri, L. & Mazel, D. Erythromycin esterase gene ere(A) is located in a functional gene 
cassette in an  unusual class 2 integron. Antimicrob. Agents Chemother. 47, 3326–3331 
(2003). 

67. Qi, Q., Rajabal, V., Ghaly, T. M., Tetu, S. G. & Gillings, M. R. Identification of integrons and 
gene cassette-associated recombination sites in  bacteriophage genomes. Front. Microbiol. 
14, 1091391 (2023). 

68. Cury, J., Jové, T., Touchon, M., Néron, B. & Rocha, E. P. Identification and analysis of 
integrons and cassette arrays in bacterial  genomes. Nucleic Acids Res. 44, 4539–4550 
(2016). 

69. Ghaly, T. M. et al. Discovery of integrons in Archaea: Platforms for cross-domain gene 
transfer. Sci. Adv. 8, 1–11 (2022). 

70. Rowe-Magnus, D. A. et al. The evolutionary history of chromosomal super-integrons 
provides an ancestry for multiresistant integrons. Proc. Natl. Acad. Sci. U. S. A. 98, 652–
657 (2001). 

71. Boucher, Y., Labbate, M., Koenig, J. E. & Stokes, H. W. Integrons: mobilizable platforms 
that promote genetic diversity in bacteria. Trends Microbiol. 15, 301–309 (2007). 

72. Mazel, D., Dychinco, B., Webb, V. A. & Davies, J. A distinctive class of integron in the Vibrio 
cholerae genome. Science 280, 605–608 (1998). 

73. Escudero, J. A., Mazel, D. & Escudero, J. A. Genomic Plasticity of Vibrio cholerae. Int. 
Microbiol. 20, 138–148 (2017). 

74. Arakawa, Y. et al. A novel integron-like element carrying the metallo-beta-lactamase gene 
blaIMP. Antimicrob. Agents Chemother. 39, 1612–1615 (1995). 

75. Collis, C. M., Kim, M.-J., Partridge, S. R., Stokes, H. W. & Hall, R. M. Characterization of 
the class 3 integron and the site-specific recombination  system it determines. J. Bacteriol. 
184, 3017–3026 (2002). 



Bibliography 

 145 
 

76. Sundström, L., Roy, P. H. & Sköld, O. Site-specific insertion of three structural gene 
cassettes in transposon Tn7. J. Bacteriol. 173, 3025–3028 (1991). 

77. Hochhut, B. et al. Molecular analysis of antibiotic resistance gene clusters in vibrio cholerae 
O139  and O1 SXT constins. Antimicrob. Agents Chemother. 45, 2991–3000 (2001). 

78. Sørum, H., Roberts, M. C. & Crosa, J. H. Identification and cloning of a tetracycline 
resistance gene from the fish  pathogen Vibrio salmonicida. Antimicrob. Agents Chemother. 
36, 611–615 (1992). 

79. Ghaly, T. M., Chow, L., Asher, A. J., Waldron, L. S. & Gillings, M. R. Evolution of class 1 
integrons: Mobilization and dispersal via food-borne bacteria. PLoS One 12, 1–11 (2017). 

80. Martinez-Freijo, P. et al. Class I integrons in Gram-negative isolates from different 
European hospitals and  association with decreased susceptibility to multiple antibiotic 
compounds. J. Antimicrob. Chemother. 42, 689–696 (1998). 

81. Zhu, Y.-G. et al. Microbial mass movements. Science 357, 1099–1100 (2017). 
82. Halaji, M. et al. The Global Prevalence of Class 1 Integron and Associated Antibiotic 

Resistance in  Escherichia coli from Patients with Urinary Tract Infections, a Systematic 
Review and Meta-Analysis. Microb. Drug Resist. 26, 1208–1218 (2020). 

83. Néron, B. et al. IntegronFinder 2.0: Identification and Analysis of Integrons across Bacteria, 
with a Focus on Antibiotic Resistance in Klebsiella. Microorganisms 10, (2022). 

84. Jové, T., Da Re, S., Denis, F., Mazel, D. & Ploy, M. C. Inverse correlation between promoter 
strength and excision activity in class 1 integrons. PLoS Genet. 6, (2010). 

85. Papagiannitsis, C. C., Tzouvelekis, L. S. & Miriagou, V. Relative strengths of the class 1 
integron promoter hybrid 2 and the combinations of strong and hybrid 1 with an active P2 
promoter. Antimicrob. Agents Chemother. 53, 277–280 (2009). 

86. Lévesque, C., Brassard, S., Lapointe, J. & Roy, P. H. Diversity and relative strength of 
tandem promoters for the antibiotic-resistance genes of several integron. Gene 142, 49–54 
(1994). 

87. Rowe-Magnus, D. A. et al. The evolutionary history of chromosomal super-integrons 
provides an ancestry for multiresistant integrons. Proc. Natl. Acad. Sci. U. S. A. 98, 652–
657 (2001). 

88. Collis, C. M., Kim, M.-J., Stokes, H. W. & Hall, R. M. Integron-encoded IntI integrases 
preferentially recognize the adjacent cognate  attI site in recombination with a 59-be site. 
Mol. Microbiol. 46, 1415–1427 (2002). 

89. Gravel, A., Fournier, B. & Roy, P. H. DNA complexes obtained with the integron integrase 
IntI1 at the attI1 site. Nucleic Acids Res. 26, 4347–4355 (1998). 

90. Collis, C. M., Kim, M. J., Stokes, H. W. & Hall, R. M. Binding of the purified integron DNA 
integrase Intl1 to integron- and cassette-associated recombination sites. Mol. Microbiol. 29, 
477–490 (1998). 

91. Stokes, H. W., O’Gorman, D. B., Recchia, G. D., Parsekhian, M. & Hall, R. M. Structure and 
function of 59-base element recombination sites associated with  mobile gene cassettes. 
Mol. Microbiol. 26, 731–745 (1997). 

92. Cambray, G., Guerout, A. M. & Mazel, D. Integrons. Annu. Rev. Genet. 44, 141–166 (2010). 
 



Bibliography 

  146 

93. Hall, R. M., Brookes, D. E. & Stokes, H. W. Site-specific insertion of genes into integrons: 
role of the 59-base element and  determination of the recombination cross-over point. Mol. 
Microbiol. 5, 1941–1959 (1991). 

94. Francia, M. V, Zabala, J. C., de la Cruz, F. & García Lobo, J. M. The IntI1 integron integrase 
preferentially binds single-stranded DNA of the attC site. J. Bacteriol. 181, 6844–6849 
(1999). 

95. Bouvier, M., Demarre, G. & Mazel, D. Integron cassette insertion: A recombination process 
involving a folded single strand substrate. EMBO J. 24, 4356–4367 (2005). 

96. Bouvier, M., Ducos-Galand, M., Loot, C., Bikard, D. & Mazel, D. Structural features of 
single-stranded integron cassette attC sites and their role in strand selection. PLoS Genet. 
5, e1000632 (2009). 

97. MacDonald, D., Demarre, G., Bouvier, M., Mazel, D. & Gopaul, D. N. Structural basis for 
broad DNA-specificity in integron recombination. Nature 440, 1157–1162 (2006). 

98. Loot, C., Bikard, D., Rachlin, A. & Mazel, D. Cellular pathways controlling integron cassette 
site folding. EMBO J. 29, 2623–2634 (2010). 

99. Nunes-Düby, S. E., Kwon, H. J., Tirumalai, R. S., Ellenberger, T. & Landy, A. Similarities 
and differences among 105 members of the Int family of site-specific recombinases. Nucleic 
Acids Res. 26, 391–406 (1998). 

100. Boyd, E. F., Almagro-Moreno, S. & Parent, M. A. Genomic islands are dynamic, ancient 
integrative elements in bacterial evolution. Trends Microbiol. 17, 47–53 (2009). 

101. Grindley, N. D. F., Whiteson, K. L. & Rice, P. A. Mechanisms of site-specific recombination. 
Annu. Rev. Biochem. 75, 567–605 (2006). 

102. Messier, N. & Roy, P. H. Integron integrases possess a unique additional domain necessary 
for activity. J. Bacteriol. 183, 6699–6706 (2001). 

103. Demarre, G., Frumerie, C., Gopaul, D. N. & Mazel, D. Identification of key structural 
determinants of the IntI1 integron integrase that influence attC × attI1 recombination 
efficiency. Nucleic Acids Res. 35, 6475–6489 (2007). 

104. Loot, C., Ducos-Galand, M., Escudero, J. A., Bouvier, M. & Mazel, D. Replicative resolution 
of integron cassette insertion. Nucleic Acids Res. 40, 8361–8370 (2012). 

105. Collis, C. M. & Hall, R. M. Site-specific deletion and rearrangement of integron insert genes 
catalyzed by  the integron DNA integrase. J. Bacteriol. 174, 1574–1585 (1992). 

106. Elsaied, H. et al. Marine integrons containing novel integrase genes, attachment sites, attI, 
and  associated gene cassettes in polluted sediments from Suez and Tokyo Bays. ISME J. 
5, 1162–1177 (2011). 

107. Hipólito, A. et al. The expression of aminoglycoside resistance genes in integron cassettes 
is not controlled by riboswitches. Nucleic Acids Res. 50, 8566–8579 (2022). 

108. Partridge, S. R., Tsafnat, G., Coiera, E. & Iredell, J. R. Gene cassettes and cassette arrays 
in mobile resistance integrons: Review article. FEMS Microbiol. Rev. 33, 757–784 (2009). 

109. Gibson, D. G. et al. Enzymatic assembly of DNA molecules up to several hundred 
kilobases. Nat. Methods 6, (2009). 

110. Wiegand, I., Hilpert, K. & Hancock, R. E. W. Agar and broth dilution methods to determine 
the minimal inhibitory concentration (MIC) of antimicrobial substances. Nat. Protoc. 3, 163–
175 (2008). 



Bibliography 

 147 
 

111. DelaFuente, J., Rodriguez-Beltran, J. & San Millan, A. Methods to Study Fitness and 
Compensatory Adaptation in Plasmid-Carrying  Bacteria. Methods Mol. Biol. 2075, 371–
382 (2020). 

112. Brugiroux, S. et al. Genome-guided design of a defined mouse microbiota that confers 
colonization resistance against Salmonella enterica serovar Typhimurium. Nat. Microbiol. 
2, 16215 (2016). 

113. Ubeda, C. et al. Familial transmission rather than defective innate immunity shapes the 
distinct  intestinal microbiota of TLR-deficient mice. J. Exp. Med. 209, 1445–1456 (2012). 

114. Turnbaugh, P. J. et al. A core gut microbiome in obese and lean twins. Nature 457, 480–
484 (2009). 

115. Li, H. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. 
(2013). doi:10.48550/arxiv.1303.3997. 

116. Liao, Y., Smyth, G. K. & Shi, W. featureCounts: an efficient general purpose program for 
assigning sequence reads  to genomic features. Bioinformatics 30, 923–930 (2014). 

117. Love, M. I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for 
RNA-seq data with DESeq2. Genome Biol. 15, 1–21 (2014). 

118. Demarre, G. et al. A new family of mobilizable suicide plasmids based on broad host range 
R388  plasmid (IncW) and RP4 plasmid (IncPalpha) conjugative machineries and their 
cognate Escherichia coli host strains. Res. Microbiol. 156, 245–255 (2005). 

119. Biskri, L., Bouvier, M., Guérout, A. M., Boisnard, S. & Mazel, D. Comparative study of class 
1 integron and Vibrio cholerae superintegron integrase activities. J. Bacteriol. 187, 1740–
1750 (2005). 

120. Ding, Y. & Lawrence, C. E. A statistical sampling algorithm for RNA secondary structure 
prediction. Nucleic Acids Res. 31, 7280–7301 (2003). 

121. Jia, X. et al. Riboswitch control of aminoglycoside antibiotic resistance. Cell 152, 68–81 
(2013). 

122. Hipólito, A., García-Pastor, L., Vergara, E., Jové, T. & Escudero, J. A. Profile and resistance 
levels of 136 integron resistance genes. npj Antimicrob. Resist. 1, 13 (2023). 

123. Shao, B. et al. Single-cell measurement of plasmid copy number and promoter activity. Nat. 
Commun. 12, (2021). 

124. Vinué, L., Jové, T., Torres, C. & Ploy, M. C. Diversity of class 1 integron gene cassette Pc 
promoter variants in clinical Escherichia coli strains and description of a new P2 promoter 
variant. Int. J. Antimicrob. Agents (2011). doi:10.1016/j.ijantimicag.2011.07.007 

125. Cambray, G. et al. Prevalence of SOS-mediated control of integron integrase expression 
as an  adaptive trait of chromosomal and mobile integrons. Mob. DNA 2, 6 (2011). 

126. Hernando-Amado, S., Coque, T. M., Baquero, F. & Martínez, J. L. Defining and combating 
antibiotic resistance from One Health and Global Health perspectives. Nat. Microbiol. 4, 
1432–1442 (2019). 

127. Kotra, L. P., Haddad, J. & Mobashery, S. Aminoglycosides: perspectives on mechanisms 
of action and resistance and  strategies to counter resistance. Antimicrob. Agents 
Chemother. 44, 3249–3256 (2000). 

128. Tolmasky, M. S. R. and M. E. Aminoglycoside Modifying Enzymes. Drug Resist Updat. 13, 
151–171 (2010). 



Bibliography 

  148 

129. Doi, Y. & Arakawa, Y. 16S ribosomal RNA methylation: Emerging resistance mechanism 
against aminoglycosides. Clin. Infect. Dis. 45, 88–94 (2007). 

130. Tietze, E. & Brevet, J. Nucleotide sequence of the streptothricin-acetyl-transferase gene 
sat-2. Nucleic Acids Res. 18, 1283 (1990). 

131. Webb, H. E., Angulo, F. J., Granier, S. A., Scott, H. M. & Loneragan, G. H. Illustrative 
examples of probable transfer of resistance determinants from food  animals to humans: 
Streptothricins, glycopeptides, and colistin. F1000Research 6, 1805 (2017). 

132. Bush, K. & Jacoby, G. A. Updated functional classification of β-lactamases. Antimicrob. 
Agents Chemother. 54, 969–976 (2010). 

133. Georgopapadakou, N. H. & Liu, F. Y. Penicillin-binding proteins in bacteria. Antimicrob. 
Agents Chemother. 18, 148–157 (1980). 

134. Ambler, R. P. The structure of beta-lactamases. Philos. Trans. R. Soc. London. Ser. B, Biol.  
Sci. 289, 321–331 (1980). 

135. Bush, K., Jacoby, G. A. & Medeiros, A. A. A functional classification scheme for β-
lactamases and its correlation with molecular structure. Antimicrob. Agents Chemother. 39, 
1211–1233 (1995). 

136. Poirel, L., Brinas, L., Verlinde, A., Ide, L. & Nordmann, P. BEL-1, a novel clavulanic acid-
inhibited extended-spectrum beta-lactamase, and  the class 1 integron In120 in 
Pseudomonas aeruginosa. Antimicrob. Agents Chemother. 49, 3743–3748 (2005). 

137. Poirel, L., Le Thomas, I., Naas, T., Karim, A. & Nordmann, P. Biochemical sequence 
analyses of GES-1, a novel class A extended-spectrum  beta-lactamase, and the class 1 
integron In52 from Klebsiella pneumoniae. Antimicrob. Agents Chemother. 44, 622–632 
(2000). 

138. Green, J. M. & Matthews, R. G. Folate Biosynthesis, Reduction, and Polyglutamylation and 
the Interconversion of Folate Derivatives. EcoSal Plus 2, (2007). 

139. Thomson, C. J. Trimethoprim and Brodimoprim Resistance of Gram-Positive and Gram-
Negative Bacteria. J. Chemother. 5, 458–464 (1993). 

140. Kahan, F. M., Kahan, J. S., Cassidy, P. J. & Kropp, H. The mechanism of action of 
fosfomycin (phosphonomycin). Ann. N. Y. Acad. Sci. 235, 364–386 (1974). 

141. Falagas, M. E., Athanasaki, F., Voulgaris, G. L., Triarides, N. A. & Vardakas, K. Z. 
Resistance to fosfomycin: Mechanisms, Frequency and Clinical Consequences. Int. J. 
Antimicrob. Agents 53, 22–28 (2019). 

142. Zurfluh, K., Treier, A., Schmitt, K. & Stephan, R. Mobile fosfomycin resistance genes in 
Enterobacteriaceae—An increasing threat. Microbiologyopen 9, 1–13 (2020). 

143. Castañeda-García, A., Blázquez, J. & Rodríguez-Rojas, A. Molecular Mechanisms and 
Clinical Impact of Acquired and Intrinsic Fosfomycin Resistance. Antibiotics 2, 217–236 
(2013). 

144. Kieffer, N., Poirel, L., Descombes, M.-C. & Nordmann, P. Characterization of FosL1, a 
Plasmid-Encoded Fosfomycin Resistance Protein Identified in Escherichia coli. Antimicrob. 
Agents Chemother. 64, e02042-19 (2020). 

145. Schwarz, S., Kehrenberg, C., Doublet, B. & Cloeckaert, A. Molecular basis of bacterial 
resistance to chloramphenicol and florfenicol. FEMS Microbiol. Rev. 28, 519–542 (2004). 

 



Bibliography 

 149 
 

146. Campbell, E. A. et al. Structural mechanism for rifampicin inhibition of bacterial rna 
polymerase. Cell 104, 901–912 (2001). 

147. Tribuddharat, C. & Fennewald, M. Integron-mediated rifampin resistance in Pseudomonas 
aeruginosa. Antimicrob. Agents Chemother. 43, 960–962 (1999). 

148. Goldstein, B. P. Resistance to rifampicin: A review. J. Antibiot. (Tokyo). 67, 625–630 (2014). 
149. Weisblum, B. Erythromycin resistance by ribosome modification. Antimicrob. Agents 

Chemother. 39, 577–585 (1995). 
150. Pozzi, G., Iannelli, F., Oggioni, M. R., Santagati, M. & Stefani, S. Genetic elements carrying 

macrolide efflux genes in streptococci. Curr. Drug Targets. Infect. Disord. 4, 203–206 
(2004). 

151. Firth, A. & Prathapan, P. Azithromycin: The First Broad-spectrum Therapeutic. Eur. J. Med. 
Chem. 207, 112739 (2020). 

152. Buffet-Bataillon, S., Tattevin, P., Bonnaure-Mallet, M. & Jolivet-Gougeon, A. Emergence of 
resistance to antibacterial agents: the role of quaternary ammonium compounds—a critical 
review. Int. J. Antimicrob. Agents 39, 381–389 (2012). 

153. Paulsen, I. T. et al. The 3’ conserved segment of integrons contains a gene associated with 
multidrug  resistance to antiseptics and disinfectants. Antimicrob. Agents Chemother. 37, 
761–768 (1993). 

154. Kazama, H., Hamashima, H., Sasatsu, M. & Arai, T. Characterization of the antiseptic-
resistance gene qacEΔ1 isolated from clinical and environmental isolates of Vibrio 
parahaemolyticus and Vibrio cholerae non-O1. FEMS Microbiol. Lett. 174, 379–384 (1999). 

155. Slipski, C. J., Jamieson-Datzkiw, T. R., Zhanel, G. G. & Bay, D. C. Characterization of 
proteobacterial plasmid integron-encoded qac efflux pump sequence diversity and 
quaternary ammonium compound antiseptic selection in Escherichia coli grown 
planktonically and as biofilms. Antimicrob. Agents Chemother. 65, (2021). 

156. San Millan, A., Escudero, J. A., Gifford, D. R., Mazel, D. & MacLean, R. C. Multicopy 
plasmids potentiate the evolution of antibiotic resistance in bacteria. Nat. Ecol. Evol. 1, 0010 
(2016). 

157. Souque, C., Escudero, J. A. & MacLean, R. C. Integron activity accelerates the evolution 
of antibiotic resistance. Elife 10, 1–47 (2021). 

158. Danielewicz, N. et al. In-Depth Characterization of a Re-Engineered Cholera Toxin 
Manufacturing Process Using Growth-Decoupled Production in Escherichia coli. Toxins 
(Basel). 14, 1–23 (2022). 

159. Rocha, D. J. P., Santos, C. S. & Pacheco, L. G. C. Bacterial reference genes for gene 
expression studies by RT-qPCR: survey and analysis. Antonie van Leeuwenhoek, Int. J. 
Gen. Mol. Microbiol. 108, 685–693 (2015). 

160. Jacquier, H., Zaoui, C., Sanson-Le Pors, M. J., Mazel, D. & Berçot, B. Translation regulation 
of integrons gene cassette expression by the attC sites. Mol. Microbiol. 72, 1475–1486 
(2009). 

161. Andersson, D. I. & Hughes, D. Antibiotic resistance and its cost: is it possible to reverse 
resistance? Nat. Rev. Microbiol. 8, 260–271 (2010). 

162. Rajer, F. & Sandegren, L. The Role of Antibiotic Resistance Genes in the Fitness Cost of 
Multiresistance Plasmids. MBio 13, 1–14 (2022). 



Bibliography 

  150 

163. Lacotte, Y., Ploy, M. C. & Raherison, S. Class 1 integrons are low-cost structures in 
Escherichia coli. ISME J. 11, 1535–1544 (2017). 

164. Porse, A., Schou, T. S., Munck, C., Ellabaan, M. M. H. & Sommer, M. O. A. Biochemical 
mechanisms determine the functional compatibility of heterologous genes. Nat. Commun. 
9, 1–11 (2018). 

165. Kadonaga, J. T. et al. The role of the β-lactamase signal sequence in the secretion of 
proteins by Escherichia coli. J. Biol. Chem. 259, 2149–2154 (1984). 

166. Weiss, A. S. et al. In vitro interaction network of a synthetic gut bacterial community. ISME 
J. 16, 1095–1109 (2022). 

167. Eberl, C. et al. E. coli enhance colonization resistance against Salmonella Typhimurium by 
competing for galactitol, a context-dependent limiting carbon source. Cell Host Microbe 29, 
1680-1692.e7 (2021). 

168. Nonaka, G., Blankschien, M., Herman, C., Gross, C. A. & Rhodius, V. A. Regulon and 
promoter analysis of the E. coli heat-shock factor, sigma32, reveals a multifaceted cellular 
response to heat stress. Genes Dev. 20, 1776–1789 (2006). 

169. Zhao, K., Liu, M. & Burgess, R. R. The global transcriptional response of Escherichia coli 
to induced σ32 protein involves σ32 regulon activation followed by inactivation and 
degradation of σ32 in vivo. J. Biol. Chem. 280, 17758–17768 (2005). 

170. Guisbert, E., Yura, T., Rhodius, V. A. & Gross, C. A.  Convergence of Molecular, Modeling, 
and Systems Approaches for an Understanding of the Escherichia coli Heat Shock 
Response . Microbiol. Mol. Biol. Rev. 72, 545–554 (2008). 

171. Fitzgerald, D. M., Bonocora, R. P. & Wade, J. T. Comprehensive Mapping of the 
Escherichia coli Flagellar Regulatory Network. PLoS Genet. 10, (2014). 

172. Zieliński, M., Park, J., Sleno, B. & Berghuis, A. M. Structural and functional insights into 
esterase-mediated macrolide resistance. Nat. Commun. 12, 1–9 (2021). 

173. Johansson, C., Kamali-Moghaddam, M. & Sundström, L. Integron integrase binds to bulged 
hairpin DNA. Nucleic Acids Res. 32, 4033–4043 (2004). 

174. Nivina, A., Escudero, J. A., Vit, C., Mazel, D. & Loot, C. Efficiency of integron cassette 
insertion in correct orientation is ensured by the interplay of the three unpaired features of 
attC recombination sites. Nucleic Acids Res. 44, 7792–7803 (2016). 

175. Mukhortava, A. et al. Structural heterogeneity of attC integron recombination sites revealed 
by optical tweezers. Nucleic Acids Res. (2018). doi:10.1093/nar/gky1258 

176. Chen, Z., Yang, H. & Pavletich, N. P. Mechanism of homologous recombination from the 
RecA-ssDNA/dsDNA structures. Nature 453, 489–494 (2008). 

177. Bouvier, M., Demarre, G. & Mazel, D. Integron cassette insertion: a recombination process 
involving a folded single  strand substrate. EMBO J. 24, 4356–4367 (2005). 

178. Nivina, A. et al. Structure-specific DNA recombination sites: Design, validation, and 
machine learning-based refinement. Sci. Adv. 6, (2020). 

179. Hanau-Berçot, B., Podglajen, I., Casin, I. & Collatz, E. An intrinsic control element for 
translational initiation in class 1 integrons. Mol. Microbiol. 44, 119–130 (2002). 

 
 



Bibliography 

 151 
 

180. Papagiannitsis, C. C., Tzouvelekis, L. S., Tzelepi, E. & Miriagou, V. attI1-Located Small 
Open Reading Frames ORF-17 and ORF-11 in a Class 1 Integron  Affect Expression of a 
Gene Cassette Possessing a Canonical Shine-Dalgarno Sequence. Antimicrob. Agents 
Chemother. 61, (2017). 

181. Roth, A. & Breaker, R. R. Integron attI1 sites, not riboswitches, associate with antibiotic 
resistance genes. Cell 153, 1417 (2013). 

182. Foucault, M.-L., Depardieu, F., Courvalin, P. & Grillot-Courvalin, C. Inducible expression 
eliminates the fitness cost of vancomycin resistance in  enterococci. Proc. Natl. Acad. Sci. 
U. S. A. 107, 16964–16969 (2010). 

183. Foucault, M.-L., Courvalin, P. & Grillot-Courvalin, C. Fitness cost of VanA-type vancomycin 
resistance in methicillin-resistant  Staphylococcus aureus. Antimicrob. Agents Chemother. 
53, 2354–2359 (2009). 

184. Morosini, M. I., Ayala, J. A., Baquero, F., Martínez, J. L. & Blázquez, J. Biological cost of 
AmpC production for Salmonella enterica serotype Typhimurium. Antimicrob. Agents 
Chemother. 44, 3137–3143 (2000). 

185. Alexieva, Z., Duvall, E. J., Ambulos, N. P. J., Kim, U. J. & Lovett, P. S. Chloramphenicol 
induction of cat-86 requires ribosome stalling at a specific site  in the leader. Proc. Natl. 
Acad. Sci. U. S. A. 85, 3057–3061 (1988). 

186. Weisblum, B., Siddhikol, C., Lai, C. J. & Demohn, V. Erythromycin-inducible resistance in 
Staphylococcus aureus: requirements for  induction. J. Bacteriol. 106, 835–847 (1971). 

187. Reilman, E., Mars, R. A. T., van Dijl, J. M. & Denham, E. L. The multidrug ABC transporter 
BmrC/BmrD of Bacillus subtilis is regulated via a ribosome-mediated transcriptional 
attenuation mechanism. Nucleic Acids Res. 42, 11393–11407 (2014). 

188. Dar, D. et al. Term-seq reveals abundant ribo-regulation of antibiotics resistance in bacteria. 
Science 352, aad9822 (2016). 

189. Evers, S. & Courvalin, P. Regulation of VanB-type vancomycin resistance gene expression 
by the VanS(B)-VanR(B) two-component regulatory system in Enterococcus faecalis V583. 
J. Bacteriol. 178, 1302–1309 (1996). 

190. Normark, S. Beta-Lactamase induction in gram-negative bacteria is intimately linked to 
peptidoglycan recycling. Microb. Drug Resist. 1, 111–114 (1995). 

191. Da Re, S. et al. The SOS response promotes qnrB quinolone-resistance determinant 
expression. EMBO Rep. 10, 929–933 (2009). 

192. Levin, B. R. Minimizing potential resistance: a population dynamics view. Clin. Infect. Dis.  
an Off. Publ. Infect. Dis.  Soc. Am. 33 Suppl 3, S161-9 (2001). 

193. Wang, S. et al. Integron Derived Aminoglycoside-Sensing Riboswitches Control 
Aminoglycoside Acetyltransferase Resistance Gene Expression. Antimicrob. Agents 
Chemother. (2019). doi:10.1128/AAC.00236-19 

194. Zhang, J. et al. Aminoglycoside riboswitch control of the expression of integron associated  
aminoglycoside resistance adenyltransferases. Virulence 11, 1432–1442 (2020). 

195. Tan, C. et al. The inoculum effect and band-pass bacterial response to periodic antibiotic  
treatment. Mol. Syst. Biol. 8, 617 (2012). 

 
 



Bibliography 

  152 

196. Weber, G. G., Kortmann, J., Narberhaus, F. & Klose, K. E. RNA thermometer controls 
temperature-dependent virulence factor expression in Vibrio cholerae. Proc. Natl. Acad. 
Sci. U. S. A. 111, 14241–14246 (2014). 

197. González-Zorn, B. et al. Genetic basis for dissemination of armA. J. Antimicrob. Chemother. 
56, 583–585 (2005). 

198. Souque, C., Escudero, J. A. & MacLean, R. C. Integron activity accelerates the evolution 
of antibiotic resistance. Elife 10, (2021). 

199. Basra, P. et al. Fitness Tradeoffs of Antibiotic Resistance in Extraintestinal Pathogenic  
Escherichia coli. Genome Biol. Evol. 10, 667–679 (2018). 

200. Brockhurst, M. A. & Harrison, E. Ecological and evolutionary solutions to the plasmid 
paradox. Trends Microbiol. 30, 534–543 (2022). 

201. Loftie-Eaton, W. et al. Compensatory mutations improve general permissiveness to 
antibiotic resistance  plasmids. Nat. Ecol. Evol. 1, 1354–1363 (2017). 

202. Dantas, G. & Sommer, M. O. A. Context matters - the complex interplay between resistome 
genotypes and resistance phenotypes. Curr. Opin. Microbiol. 15, 577–582 (2012). 

203. Ghaly, T. M., Tetu, S. G. & Gillings, M. R. Predicting the taxonomic and environmental 
sources of integron gene cassettes using structural and sequence homology of attC sites. 
Commun. Biol. 4, (2021). 

204. Grieb, M. S. et al. Dynamic stepwise opening of integron attC DNA hairpins by SSB 
prevents toxicity and ensures functionality. Nucleic Acids Res. 45, 10555–10563 (2017). 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
  
 



 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

ANNEX 
Supplementary Material 

 

 
 
 
 

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Supplementary Material 

 155 

8 Supplementary Material  
 
Supplementary Table S1. Strains and plasmids used in this study. 
All strains constructed for this work were built in E. coli backgrounds otherwise stated. Strains corresponding 
to the OMM12 are numbered following DSM (German collection of microorganisms and cell cultures) 
classification. 
 

Strain Genetic 
background Plasmid ARC cloned Source 

A072 MG1655 - - Lab collection 
A093 DH5a - - Lab collection 
A249 MG1655 pMBA empty vector - This study 
C381 DH5a pMBA empty vector - This study 
A223 MG1655 pMBAdfrA1 dfrA1 This study 
A224 MG1655 pMBAdfrA5 dfrA5 This study 
A225 MG1655 pMBAdfrA6 dfrA6 This study 
A226 MG1655 pMBAdfrA7 dfrA7 This study 
A241 MG1655 pMBAdfrA12 dfrA12 This study 
A227 MG1655 pMBAdfrA14 dfrA14 This study 
A228 MG1655 pMBAdfrA15 dfrA15 This study 
A229 MG1655 pMBAdfrA16 dfrA16 This study 
A230 MG1655 pMBAdfrA17 dfrA17 This study 
A231 MG1655 pMBAdfrA21 dfrA21 This study 
A500 MG1655 pMBAdfrA22 dfrA22 This study 
A244 MG1655 pMBAdfrA22.2 dfrA22.2 This study 
A232 MG1655 pMBAdfrA25 dfrA25 This study 
A233 MG1655 pMBAdfrA27 dfrA27 This study 
A613 MG1655 pMBAdfrA29 dfrA29 This study 
A248 MG1655 pMBAdfrA30 dfrA30 This study 
A234 MG1655 pMBAdfrA31 dfrA31 This study 
A235 MG1655 pMBAdfrA34 dfrA34 This study 
A242 MG1655 pMBAdfrA35 dfrA35 This study 
A236 MG1655 pMBAdfrB1 dfrB1 This study 
A245 MG1655 pMBAdfrB2 dfrB2 This study 
A246 MG1655 pMBAdfrB3 dfrB3 This study 
A237 MG1655 pMBAdfrB4 dfrB4 This study 
A238 MG1655 pMBAdfrB5 dfrB5 This study 
A239 MG1655 pMBAdfrB6 dfrB6 This study 
A240 MG1655 pMBAdfrB7 dfrB7 This study 
A247 MG1655 pMBAdfrB8 dfrB8 This study 
A243 MG1655 pMBAdfrB9 dfrB9 This study 
A390 MG1655 pMBAblaBEL-1 blaBEL-1 This study 
A380 MG1655 pMBAblaGES-1 blaGES-1 This study 
A387 MG1655 pMBAblaIMP-2 blaIMP-2 This study 
A383 MG1655 pMBAblaIMP-31 blaIMP-31 This study 
A441 MG1655 pMBAblaOXA-1 blaOXA-1 This study 
A373 MG1655 pMBAblaOXA-2 blaOXA-2 This study 
A381 MG1655 pMBAblaOXA-5 blaOXA-5 This study 
A384 MG1655 pMBA blaOXA-9 blaOXA-9 This study 
A374 MG1655 pMBAblaOXA-10 blaOXA-10 This study 
A385 MG1655 pMBAblaOXA-20 blaOXA-20 This study 



Supplementary Material 

  156 

A382 MG1655 pMBAblaOXA-21 blaOXA-21 This study 
A395 MG1655 pMBAblaOXA-46 blaOXA-46 This study 
A375 MG1655 pMBAblaOXA-118 blaOXA-118 This study 
A391 MG1655 pMBAblaOXA-129 blaOXA-129 This study 
A376 MG1655 pMBAblaOXA-198 blaOXA-198 This study 
A803 MG1655 pMBAblaPBL-1 blaPBL-1 This study 
A388 MG1655 pMBAblaVIM-1 blaVIM-1 This study 
A396 MG1655 pMBAblaVIM-2 blaVIM-2 This study 
A389 MG1655 pMBAblaVIM-7 blaVIM-7 This study 
A327 MG1655 pMBAaacA2 aacA2 This study 
A266 MG1655 pMBAaacA3 aacA3 This study 
A328 MG1655 pMBAaacA4 aacA4 This study 
A267 MG1655 pMBAaacA7 aacA7 This study 
A268 MG1655 pMBAaacA8 aacA8 This study 
A263 MG1655 pMBAaacA16 aacA16 This study 
A543 MG1655 pMBAaacA17 aacA17 This study 
A264 MG1655 pMBAaacA27 aacA27 This study 
A256 MG1655 pMBAaacA28 aacA28 This study 
A257 MG1655 pMBAaacA29 aacA29 This study 
A270 MG1655 pMBAaacA30 aacA30 This study 
B657 MG1655 pMBAaacA31 aacA31 This study 
A272 MG1655 pMBAaacA34 aacA34 This study 
A329 MG1655 pMBAaacA35 aacA35 This study 
A258 MG1655 pMBAaacA37 aacA37 This study 
A603 MG1655 pMBAaacA38 aacA38 This study 
A273 MG1655 pMBAaacA42 aacA42 This study 
C117 MG1655 pMBAaacA43 aacA43 This study 
A265 MG1655 pMBAaacA45 aacA45 This study 
A274 MG1655 pMBAaacA47 aacA47 This study 
A259 MG1655 pMBAaacA48 aacA48 This study 
B656 MG1655 pMBAaacA49 aacA49 This study 
A260 MG1655 pMBAaacA50 aacA50 This study 
A261 MG1655 pMBAaacA51 aacA51 This study 
A326 MG1655 pMBAaacA52 aacA52 This study 
B655 MG1655 pMBAaacA54 aacA54 This study 
A262 MG1655 pMBAaacA56 aacA56 This study 
A303 MG1655 pMBAaacA59 aacA59 This study 
A302 MG1655 pMBAaacA61 aacA61 This study 
A566 MG1655 pMBAaacA64 aacA64 This study 
A331 MG1655 pMBAaacAX aacAX This study 
A308 MG1655 pMBAaacC1 aacC1 This study 
A304 MG1655 pMBAaacC2 aacC2 This study 
B625 MG1655 pMBAaacC3 aacC3 This study 
A320 MG1655 pMBAaacC4 aacC4 This study 
A305 MG1655 pMBAaacC5 aacC5 This study 
A306 MG1655 pMBAaacC6 aacC6 This study 
A332 MG1655 pMBAaacC11 aacC11 This study 
A309 MG1655 pMBAaacC13 aacC13 This study 
A311 MG1655 pMBAaadA1 aadA1 This study 
A333 MG1655 pMBAaadA2 aadA2 This study 
A319 MG1655 pMBAaadA4 aadA4 This study 
A313 MG1655 pMBAaadA5 aadA5 This study 
A321 MG1655 pMBAaadA6 aadA6 This study 
A397 MG1655 pMBAaadA7 aadA7 This study 



Supplementary Material 

 157 

A315 MG1655 pMBAaadA10 aadA10 This study 
A312 MG1655 pMBAaadA11 aadA11 This study 
A322 MG1655 pMBAaadA13 aadA13 This study 
A316 MG1655 pMBAaadA16 aadA16 This study 
A318 MG1655 pMBAaadA24 aadA24 This study 
A604 MG1655 pMBAaadA28 aadA28 This study 
A314 MG1655 pMBAaadA29 aadA29 This study 
A398 MG1655 pMBAaadA34 aadA34 This study 
A292 MG1655 pMBAaadB aadB This study 
A422 MG1655 pMBAaphA15 aphA15 This study 
C116 MG1655 pMBAaphA16 aphA16 This study 
A293 MG1655 pMBAsat2 sat2 This study 
A368 MG1655 pMBAarr2 arr2 This study 
A334 MG1655 pMBAarr5 arr5 This study 
A363 MG1655 pMBAarr6 arr6 This study 
A440 MG1655 pMBAarr7 arr7 This study 
A364 MG1655 pMBAarr8b arr8b This study 
A335 MG1655 pMBAcatB2 catB2 This study 
A336 MG1655 pMBAcatB3 catB3 This study 
A365 MG1655 pMBAcatB5 catB5 This study 
A625 MG1655 pMBAcatB6 catB6 This study 
A393 MG1655 pMBAcatB10 catB10 This study 
A338 MG1655 pMBAereA2 ereA2 This study 
A423 MG1655 pMBAereA3 ereA3 This study 
A657 MG1655 pMBAfosC2 fosC2 This study 
A339 MG1655 pMBAfosE fosE This study 
A624 MG1655 pMBAfosF fosF This study 
A354 MG1655 pMBAfosG fosG This study 
A355 MG1655 pMBAfosH fosH This study 
A356 MG1655 pMBAfosI fosI This study 
A626 MG1655 pMBAfosK fosK This study 
A621 MG1655 pMBAfosL fosL This study 
A622 MG1655 pMBAfosM fosM This study 
A627 MG1655 pMBAfosN fosN This study 
A337 MG1655 pMBAsmr1 smr1 This study 
A359 MG1655 pMBAsmr2 smr2 This study 
A366 MG1655 pMBAsmr3 smr3 This study 
A367 MG1655 pMBAqacE qacE This study 
B091 MG1655 pMBAqacEDsul1 qacEDsul1 This study 
C382 DH5a pMBAqacEDsul1 qacEDsul1 This study 
A323 MG1655 pMBAqacF qacF This study 
A357 MG1655 pMBAqacG qacG This study 
A324 MG1655 pMBAqacH qacH This study 
A623 MG1655 pMBAqacK qacK This study 
A325 MG1655 pMBAqacL qacL This study 
A358 MG1655 pMBAqacM qacM This study 
A086 MG1655 R388dfrA5-aadB-blaVEB-1 dfrA5-aadB-blaVEB-1 Souque et al, 2021 
A078 MG1655 R388aadB-dfrA5-blaVEB-1 aadB-dfrA5-blaVEB-1 Souque et al, 2021 
A487 MG1655 pMBAdfrB4-attCdfrA12 dfrB4-attCdfrA12 This study 
A471 MG1655 pMBAdfrB4-attCdfrA21 dfrB4-attCdfrA21 This study 
A483 MG1655 pMBAdfrA21 DattC dfrA21 DattC This study 
A482 MG1655 pMBAdfrA12 DattC dfrA12 DattC This study 
A480 MG1655 DPcS pMBA - This study 



Supplementary Material 

  158 

A497 MG1655 DPcS pMBAdfrA15 dfrA15 This study 
A485 MG1655 DPcS pMBAdfrB4 dfrB4 This study 
C457 MG1655 DPcS pMBAereA3 ereA3 This study 
A696 MG1655 pMBA DmetGFP KO - This study 
A744 MG1655 pMBAaacA59 DmetGFP KO aacA59 This study 
A745 MG1655 pMBAaacA8 DmetGFP KO aacA8 This study 
A746 MG1655 pMBAaadB DmetGFP KO aadB This study 
A747 MG1655 pMBAaacC2 DmetGFP KO aacC2 This study 
A748 MG1655 pMBAaacA28 DmetGFP KO aacA28 This study 
A749 MG1655 pMBAaacA52 DmetGFP KO aacA52 This study 
A750 MG1655 pMBAaacA30 DmetGFP KO aacA30 This study 
A751 MG1655 pMBAaadA24 DmetGFP KO aadA24 This study 
A752 MG1655 pMBAaadA13 DmetGFP KO aadA13 This study 
A753 MG1655 pMBAaacC13 DmetGFP KO aacC13 This study 
A754 MG1655 pMBAdfrA35 DmetGFP KO dfrA35 This study 
A755 MG1655 pMBAdfrB4 DmetGFP KO dfrB4 This study 
A756 MG1655 pMBAdfrA22.2 DmetGFP KO dfrA22.2 This study 
A757 MG1655 pMBAdfrA16 DmetGFP KO dfrA16 This study 
A758 MG1655 pMBAdfrA14 DmetGFP KO dfrA14 This study 
A759 MG1655 pMBAdfrA34 DmetGFP KO dfrA34 This study 
B878 MG1655 pMBA DGFP - This study 
B880 MG1655 pMBAereA2 DGFP ereA2 This study 
B881 MG1655 pMBAdfrA31 DGFP dfrA31 This study 
B882 MG1655 pMBAdfrA21 DGFP dfrA21 This study 
B883 MG1655 pMBAaacA7 DGFP aacA7 This study 
B886 MG1655 pMBAblaOXA-10 DGFP blaOXA-10 This study 
26074 YL2 - - Brugiroux et al, 2016 
28989 YL27 - - Brugiroux et al, 2016 
26085 I48 - - Brugiroux et al, 2016 
26109 YL45 - - Brugiroux et al, 2016 
26127 YL44 - - Brugiroux et al, 2016 
32036 KB1 - - Brugiroux et al, 2016 
32035 I49 - - Brugiroux et al, 2016 
26114 YL32 - - Brugiroux et al, 2016 
26115 YL58 - - Brugiroux et al, 2016 
26117 YL31 - - Brugiroux et al, 2016 
26090 KB18 - - Brugiroux et al, 2016 
26113 I46 - - Brugiroux et al, 2016 
B966 MG1655 pMBADmet ereA2 Dmet ereA2 This study 
B965 MG1655 pMBA cat. mut. ereA2 cat. mut. ereA2 This study 
C271 MG1655 pMBADmet ereA3 Dmet ereA3 This study 
B951 MG1655 pMBA cat. mut. ereA3 cat. mut. ereA3 This study 
A123 b2163 pSW23T::attCaadA7 - Demarre et al, 2005 
A785 Top10 pSU38D::attI / p3938 - Lab collection 
A122 b2163 pSW23T::attI - Demarre et al, 2005 
B963 DH5a p3938 - This study 
B964 DH5a pMBAaadA7 / p3938 aadA7 This study 
B965 DH5a pMBAaadA7 / p3938 Dint1 aadA7 This study 
B967 DH5a pMBAaacA2 / p3938 aacA2 This study 
B968 DH5a pMBAaacA3 / p3938 aacA3 This study 
B969 DH5a pMBAaacA4 / p3938 aacA4 This study 



Supplementary Material 

 159 

B970 DH5a pMBAaacA7 / p3938 aacA7 This study 
B971 DH5a pMBAaacA8 / p3938 aacA8 This study 
B972 DH5a pMBAaacA16 / p3938 aacA16 This study 
B973 DH5a pMBAaacA17 / p3938 aacA17 This study 
B974 DH5a pMBAaacA27 / p3938 aacA27 This study 
B975 DH5a pMBAaacA28 / p3938 aacA28 This study 
B976 DH5a pMBAaacA29 / p3938 aacA29 This study 
B977 DH5a pMBAaacA30 / p3938 aacA30 This study 
B978 DH5a pMBAaacA31 / p3938 aacA31 This study 
B979 DH5a pMBAaacA34 / p3938 aacA34 This study 
B980 DH5a pMBAaacA35 / p3938 aacA35 This study 
B981 DH5a pMBAaacA37 / p3938 aacA37 This study 
B982 DH5a pMBAaacA38 / p3938 aacA38 This study 
B983 DH5a pMBAaacA42 / p3938 aacA42 This study 
B984 DH5a pMBAaacA43 / p3938 aacA43 This study 
B985 DH5a pMBAaacA45 / p3938 aacA45 This study 
B986 DH5a pMBAaacA47 / p3938 aacA47 This study 
B987 DH5a pMBAaacA48 / p3938 aacA48 This study 
B988 DH5a pMBAaacA49 / p3938 aacA49 This study 
B989 DH5a pMBAaacA50 / p3938 aacA50 This study 
B990 DH5a pMBAaacA51 / p3938 aacA51 This study 
B991 DH5a pMBAaacA52 / p3938 aacA52 This study 
B992 DH5a pMBAaacA54 / p3938 aacA54 This study 
B993 DH5a pMBAarr8b / p3938 arr8b This study 
B994 DH5a pMBAaacA56 / p3938 aacA56 This study 
B995 DH5a pMBAaacA59 / p3938 aacA59 This study 
B996 DH5a pMBAaacA61 / p3938 aacA61 This study 
B997 DH5a pMBAaacA64 / p3938 aacA64 This study 
B998 DH5a pMBAaacAX / p3938 aacAX This study 
B999 DH5a pMBAaacC1 / p3938 aacC1 This study 
C001 DH5a pMBAaacC2 / p3938 aacC2 This study 
C002 DH5a pMBAaacC3 / p3938 aacC3 This study 
C003 DH5a pMBAaacC4 / p3938 aacC4 This study 
C004 DH5a pMBAaacC5 / p3938 aacC5 This study 
C005 DH5a pMBAaacC6 / p3938 aacC6 This study 
C006 DH5a pMBAaacC11 / p3938 aacC11 This study 
C007 DH5a pMBAaacC13 / p3938 aacC13 This study 
C009 DH5a pMBAaadA1 / p3938 aadA1 This study 
C010 DH5a pMBAaadA2 / p3938 aadA2 This study 
C011 DH5a pMBAaadA4 / p3938 aadA4 This study 
C012 DH5a pMBAaadA5 / p3938 aadA5 This study 
C013 DH5a pMBAaadA6 / p3938 aadA6 This study 
C015 DH5a pMBAaadA10 / p3938 aadA10 This study 
C016 DH5a pMBAaadA11 / p3938 aadA11 This study 
C017 DH5a pMBAaadA13 / p3938 aadA13 This study 
C018 DH5a pMBAaadA16 / p3938 aadA16 This study 
C019 DH5a pMBAaadA24 / p3938 aadA24 This study 
C020 DH5a pMBAaadA28 / p3938 aadA28 This study 
C021 DH5a pMBAaadA29 / p3938 aadA29 This study 
C022 DH5a pMBAaadA34 / p3938 aadA34 This study 
C023 DH5a pMBAaadB / p3938 aadB This study 



Supplementary Material 

  160 

C024 DH5a pMBAaphA15 / p3938 aphA15 This study 
C025 DH5a pMBAaphA16 / p3938 aphA16 This study 
C026 DH5a pMBAsat2 / p3938 sat2 This study 
C027 DH5a pMBAdfrA1 / p3938 dfrA1 This study 
C028 DH5a pMBAdfrA5 / p3938 dfrA5 This study 
C029 DH5a pMBAdfrA6 / p3938 dfrA6 This study 
C030 DH5a pMBAdfrA7 / p3938 dfrA7 This study 
C031 DH5a pMBAdfrA12 / p3938 dfrA12 This study 
C032 DH5a pMBAdfrA14 / p3938 dfrA14 This study 
C033 DH5a pMBAdfrA15 / p3938 dfrA15 This study 
C034 DH5a pMBAdfrA16 / p3938 dfrA16 This study 
C035 DH5a pMBAdfrA17 / p3938 dfrA17 This study 
C036 DH5a pMBAdfrA21 / p3938 dfrA21 This study 
C037 DH5a pMBAdfrA22 / p3938 dfrA22 This study 
C038 DH5a pMBAdfrA25 / p3938 dfrA25 This study 
C039 DH5a pMBAdfrA27 / p3938 dfrA27 This study 
C040 DH5a pMBAdfrA29 / p3938 dfrA29 This study 
C041 DH5a pMBAdfrA30 / p3938 dfrA30 This study 
C042 DH5a pMBAdfrA31 / p3938 dfrA31 This study 
C043 DH5a pMBAdfrA34 / p3938 dfrA34 This study 
C044 DH5a pMBAdfrA35 / p3938 dfrA35 This study 
C045 DH5a pMBAdfrB1 / p3938 dfrB1 This study 
C046 DH5a pMBAdfrB2 / p3938 dfrB2 This study 
C047 DH5a pMBAdfrB3 / p3938 dfrB3 This study 
C048 DH5a pMBAdfrB4 / p3938 dfrB4 This study 
C049 DH5a pMBAdfrB5 / p3938 dfrB5 This study 
C050 DH5a pMBAdfrB6 / p3938 dfrB6 This study 
C051 DH5a pMBAdfrB7 / p3938 dfrB7 This study 
C052 DH5a pMBAdfrB8 / p3938 dfrB8 This study 
C053 DH5a pMBAdfrB9 / p3938 dfrB9 This study 
C054 DH5a pMBAblaBEL-1 / p3938 blaBEL-1 This study 
C055 DH5a pMBAblaGES-1 / p3938 blaGES-1 This study 
C056 DH5a pMBAblaIMP-2 / p3938 blaIMP-2 This study 
C057 DH5a pMBAblaIMP-31 / p3938 blaIMP-31 This study 
C058 DH5a pMBAblaOXA-1 / p3938 blaOXA-1 This study 
C059 DH5a pMBAblaOXA-2 / p3938 blaOXA-2 This study 
C060 DH5a pMBAblaOXA-5 / p3938 blaOXA-5 This study 
C061 DH5a pMBAblaOXA-9 / p3938 blaOXA-9 This study 
C062 DH5a pMBAblaOXA-10 / p3938 blaOXA-10 This study 
C063 DH5a pMBAblaOXA-20 / p3938 blaOXA-20 This study 
C064 DH5a pMBAblaOXA-21 / p3938 blaOXA-21 This study 
C065 DH5a pMBAblaOXA-46 / p3938 blaOXA-46 This study 
C066 DH5a pMBAblaOXA-118 / p3938 blaOXA-118 This study 
C067 DH5a pMBAblaOXA-129 / p3938 blaOXA-129 This study 
C068 DH5a pMBAblaOXA-198 / p3938 blaOXA-198 This study 
C069 DH5a pMBAblaPBL-1 / p3938 blaPBL-1 This study 
C070 DH5a pMBAblaVIM-1 / p3938 blaVIM-1 This study 
C071 DH5a pMBAblaVIM-2 / p3938 blaVIM-2 This study 
C072 DH5a pMBAblaVIM-7 / p3938 blaVIM-7 This study 
C073 DH5a pMBAarr2 / p3938 arr2 This study 
C074 DH5a pMBAarr5 / p3938 arr5 This study 



Supplementary Material 

 161 

C075 DH5a pMBAarr6 / p3938 arr6 This study 
C076 DH5a pMBAarr7 / p3938 arr7 This study 
C077 DH5a pMBAcatB2 / p3938 catB2 This study 
C078 DH5a pMBAcatB3 / p3938 catB3 This study 
C079 DH5a pMBAcatB5 / p3938 catB5 This study 
C080 DH5a pMBAcatB6 / p3938 catB6 This study 
C081 DH5a pMBAcatB10 / p3938 catB10 This study 
C082 DH5a pMBAereA2 / p3938 ereA2 This study 
C083 DH5a pMBAereA3 / p3938 ereA3 This study 
C084 DH5a pMBAsmr1 / p3938 smr1 This study 
C085 DH5a pMBAsmr2 / p3938 smr2 This study 
C086 DH5a pMBAsmr3 / p3938 smr3 This study 
C087 DH5a pMBAfosC2 / p3938 fosC2 This study 
C088 DH5a pMBAfosE / p3938 fosE This study 
C089 DH5a pMBAfosF / p3938 fosF This study 
C090 DH5a pMBAfosG / p3938 fosG This study 
C091 DH5a pMBAfosH / p3938 fosH This study 
C092 DH5a pMBAfosI / p3938 fosI This study 
C093 DH5a pMBAfosK / p3938 fosK This study 
C094 DH5a pMBAfosL / p3938 fosL This study 
C095 DH5a pMBAfosM / p3938 fosM This study 
C096 DH5a pMBAfosN / p3938 fosN This study 
C097 DH5a pMBAqacE / p3938 qacE This study 
C098 DH5a pMBAqacF / p3938 qacF This study 
C099 DH5a pMBAqacG / p3938 qacG This study 
C100 DH5a pMBAqacH / p3938 qacH This study 
C101 DH5a pMBAqacK / p3938 qacK This study 
C102 DH5a pMBAqacL/ p3938 qacL This study 
C103 DH5a pMBAqacM / p3938 qacM This study 
C104 DH5a pMBAqacEDsul1 / p3938 qacEDsul1 This study 
A798 MG1655 pMBA5´- aacA1 - This study 
A439 MG1655 pMBA5´- aacA2 - This study 
A551 MG1655 pMBA5´- aacA3 - This study 
A438 MG1655 pMBA5´- aacA4 - This study 
A615 MG1655 pMBA5´- aacA5 - This study 
A424 MG1655 pMBA5´- aacA7 - This study 
A560 MG1655 pMBA5´- aacA8 - This study 
A605 MG1655 pMBA5´- aacA16 - This study 
A610 MG1655 pMBA5´- aacA17 - This study 
A425 MG1655 pMBA5´- aacA27 - This study 
A426 MG1655 pMBA5´- aacA28 - This study 
A612 MG1655 pMBA5´- aacA29 - This study 
A427 MG1655 pMBA5´- aacA30 - This study 
A428 MG1655 pMBA5´- aacA31 - This study 
A802 MG1655 pMBA5´- aacA32 - This study 
A429 MG1655 pMBA5´- aacA34 - This study 
A430 MG1655 pMBA5´- aacA35 - This study 
A437 MG1655 pMBA5´- aacA37 - This study 
A567 MG1655 pMBA5´- aacA38 - This study 
A565 MG1655 pMBA5´- aacA39 - This study 



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  162 

A564 MG1655 pMBA5´- aacA40 - This study 
A502 MG1655 pMBA5´- aacA42 - This study 
A507 MG1655 pMBA5´- aacA43 - This study 
A801 MG1655 pMBA5´- aacA44 - This study 
A535 MG1655 pMBA5´- aacA45 - This study 
A808 MG1655 pMBA5´- aacA46 - This study 
A508 MG1655 pMBA5´- aacA47 - This study 
A509 MG1655 pMBA5´- aacA48 - This study 
A510 MG1655 pMBA5´- aacA49 - This study 
A534 MG1655 pMBA5´- aacA50 - This study 
A511 MG1655 pMBA5´- aacA51 - This study 
A540 MG1655 pMBA5´- aacA52 - This study 
A361 MG1655 pMBA5´- aacA54 - This study 
A512 MG1655 pMBA5´- aacA56 - This study 
A513 MG1655 pMBA5´- aacA59 - This study 
A473 MG1655 pMBA5´- aacA61 - This study 
A800 MG1655 pMBA5´- aacA64 - This study 
A549 MG1655 pMBA5´- aacAX - This study 
A514 MG1655 pMBA5´- aacC1 - This study 
A542 MG1655 pMBA5´- aacC2 - This study 
A537 MG1655 pMBA5´- aacC3 - This study 
A799 MG1655 pMBA5´- aacC4 - This study 
A541 MG1655 pMBA5´- aacC5 - This study 
A533 MG1655 pMBA5´- aacC6 - This study 
A539 MG1655 pMBA5´- aacC11 - This study 
A606 MG1655 pMBA5´- aacC13 - This study 
A531 MG1655 pMBA5´- aadA1 - This study 
A607 MG1655 pMBA5´- aadA2 - This study 
A550 MG1655 pMBA5´- aadA4 - This study 
A530 MG1655 pMBA5´- aadA5 - This study 
A553 MG1655 pMBA5´- aadA6 - This study 
A609 MG1655 pMBA5´- aadA7 - This study 
A796 MG1655 pMBA5´- aadA9 - This study 
A608 MG1655 pMBA5´- aadA10 - This study 
A554 MG1655 pMBA5´- aadA11 - This study 
A558 MG1655 pMBA5´- aadA13 - This study 
A555 MG1655 pMBA5´- aadA16 - This study 
A557 MG1655 pMBA5´- aadA24 - This study 
A797 MG1655 pMBA5´- aadA28 - This study 
A556 MG1655 pMBA5´- aadA29 - This study 
A559 MG1655 pMBA5´- aadA34 - This study 
A532 MG1655 pMBA5´- aadB - This study 
A538 MG1655 pMBA5´- aphA15 - This study 
A552 MG1655 pMBA5´- aphA16 - This study 
A563 MG1655 pMBA5´- dfrA5 - This study 
A561 MG1655 pMBA5´- fosG - This study 
A562 MG1655 pMBA5´- blaOXA-9 - This study 
A611 MG1655 pMBATm - This study 
A601 MG1655 pBGT - San Millán et al, 2016 
B536 MG1655 pMBA5´- aacA43alt - This study 



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 163 

B537 MG1655 pMBA5´- aacA47alt - This study 
B538 MG1655 pMBA5´- aacAXalt - This study 
B539 MG1655 pMBA5´- aadA9alt - This study 
B540 MG1655 pMBA5´- aacA4alt - This study 
A617 MG1655 pMBA5´- aacA1alt - This study 
B628 MG1655 pMBA5´- aacC5alt - This study 
B771 A605 pBAD::armA - This study 
B772 A606 pBAD::armA - This study 
B773 A607 pBAD::armA - This study 
B774 A608 pBAD::armA - This study 
B775 A609 pBAD::armA - This study 
B776 A610 pBAD::armA - This study 
B777 A611 pBAD::armA - This study 
B778 A612 pBAD::armA - This study 
B779 A615 pBAD::armA - This study 
B780 A796 pBAD::armA - This study 
B781 A797 pBAD::armA - This study 
B782 A799 pBAD::armA - This study 
B783 A801 pBAD::armA - This study 
B784 A802 pBAD::armA - This study 

 
  



Supplementary Material 

  164 

Supplementary Table S2. Primers and probes used in this study. 
 

Primer/Probe Sequence (5´à 3´) Description 

int R bb CTTTGTTTTAGGGCGACTGC pMBA linearisation gfp F bb TTAGGCGTCGACGCTGCA 
gBlock F GCAGTCGCCCTAAAACAAAG ARC amplification gBlock R GCTGCAGCGTCGACGCCTAA 

int F AACGCAATTACAGAAATGCCTCGACTTCGC 
pMBA sequencing gfp R AAAGCGCTGTCTAGACTATT 

gfp 2.0 R CTCGATTCTATTAACAAGGG 
gfp RT F GTTGGCCATGGAACAGGTAG 

gfp qPCR gfp RT R AGTGGAGAGGGTGAAGGTGA 
rpoA RT F TTGAGCAGGATTTCCAGGAT 

rpoA qPCR rpoA RT R CCGAGGTTGAGATTGATGGT 
rssA RT F CAATTGAACATCCTGCAACG 

rssA qPCR rssA RT R AGAGGTTGGTCGCATATTGG 
DPcS F ACGAACCCAGGTAATGCAAGTAGCGTATGC 

pMBA PcS deletion  
DPcS R ACTTGCATTACCTGGGTTCGTGCCTTCATC 

DattC gfp F GTCGACGCTGCAGCGATT pMBA attC deletion 
dfrA21 DattC R AATCGCTGCAGCGTCGACTTAGCCGTTACGACGCGCAT pMBAdfrA21 attC deletion 
dfrA12 DattC R GCAATCGCTGCAGCGTCGACTTAGCCGTTTCGACGCGCAT pMBAdfrA12 attC deletion 
B4 cassette R CTTGTTAGCCTGTGGACGGTGCCGCATGAT 

pMBAdfrB4 attC change 
(to dfrA21 or dfrA12) bb A21B4 F ACCGTCCACAGGCTAACAAGTCCGTCAACG 

bb dfrA12 F ATCATGCGGCACCGTCCACAGGCTAACCATTCCGTCAACG 
Dmet gfp F GTAAATATAGAGTAAATGAGAAGAACTTTT 

pMBA gfp KO and Dmet 
Dmet gfp R ATTTACTCTATATTTACCTCCTTTATATTG 

Dgfp F TGCAATATAAAATTGTCGGCACGTAAGAGG 
pMBA gfp deletion 

Dgfp R GCCGACAATTTTATATTGCAATCGCTGCAG 
IsoI46 Exonucl.2 F CGGATCGTAAAGCTCTGTTGTAAG 

I46 qPCR 
(Brugiroux et al, 2016) IsoI46 Exonucl.3 R GCTACCGTCACTCCCATAGCA 

Probe3 IsoI46 FAM-AAGAACGGCTCATAGAGG-BHQ1 
IsoI49 Exonucl. F GCACTGGCTCAACAGATTGATG 

I49 qPCR 
(Brugiroux et al, 2016) IsoI49 Exonucl. R CCGCCACTCACTGGTGATC 

Probe IsoI49 HEX-CTTGCACCTGATTGACGA-BHQ1 
YL58 Exonucl. F GAAGAGCAAGTCTGATGTGAAAGG 

YL58 qPCR 
(Brugiroux et al, 2016) YL58 Exonucl. R CGGCACTCTAGAAAAACAGTTTCC 

Probe YL58 FAM-TAACCCCAGGACTGCAT-BHQ1 
YL27 Exonucl.2 F TCAAGTCAGCGGTAAAAATTCG 

YL27 qPCR 
(Brugiroux et al, 2016) YL27 Exonucl.2 R CCCACTCAAGAACATCAGTTTCAA 

Probe2 YL27 HEX-CAACCCCGTCGTGCC-BHQ1 
YL31 Exonucl.2 F AGGCGGGATTGCAAGTCA 

YL31 qPCR 
(Brugiroux et al, 2016) YL31 Exonucl.3 R CCAGCACTCAAGAACTACAGTTTCA 

Probe2 YL31 FAM-CAACCTCCAGCCTGC-BHQ1 
YL32 Exonucl.2 F AATACCGCATAAGCGCACAGT 

YL32 qPCR 
(Brugiroux et al, 2016) YL32 Exonucl.2 R CCATCTCACACCACCAAAGTTTT 

Probe2 YL32 HEX-CGCATGGCAGTGTGT-BHQ1 
KB1 Exonucl. F CTTCTTTCCTCCCGAGTGCTT 

KB1 qPCR 
(Brugiroux et al, 2016) KB1 Exonucl. R CCCCTCTGATGGGTAGGTTACC 

Probe KB1 FAM-CACTCAATTGGAAAGAGGAG-BHQ1 
YL2 Exonucl. F GGGTGAGTAATGCGTGACCAA 

YL2 qPCR 
(Brugiroux et al, 2016) YL2 Exonucl. R CGGAGCATCCGGTATTACCA 

Probe2 YL2 HEX-CGGAATAGCTCCTGGAAA-BHQ1 



Supplementary Material 

 165 

KB18 Exonucl.2 F TGGCAAGTCAGTAGTGAAATCCA 
KB18 qPCR 

(Brugiroux et al, 2016) KB18 Exonucl.2 R TCACTCAAGCTCGACAGTTTCAA 
Probe2 KB18 FAM-CTTAACCCATGAACTGC-BHQ1 

YL44 Exonucl. F CGGGATAGCCCTGGGAAA 
YL44 qPCR 

(Brugiroux et al, 2016) YL44 Exonucl. R GCGCATTGCTGCTTTAATCTTT 
Probe YL44 HEX-TGGGATTAATACCGCATAGTA-BHQ1 

YL45 Exonucl. F AGACGGCCTTCGGGTTGTA 
YL45 qPCR 

(Brugiroux et al, 2016) YL45 Exonucl. R CGTCATCGTCTATCGGTATTATCAA 
Probe YL44 FAM-ACCACTTTTGTAGAGAACGA-BHQ1 

IsoI48 Exonucl. F GGCAGCATGGGAGTTTGCT 
I48 qPCR 

(Brugiroux et al, 2016) IsoI48 Exonucl. R TTATCGGCAGGTTGGATACGT 
Probe IsoI48 HEX-CAAACTTCCGATGGCGAC-BHQ1 

E. coli Exonucl. F GGACCTTCGGGCCTCTTG 
E. coli qPCR 

(Brugiroux et al, 2016) E. coli Exonucl. R CCTTTACCCCACCTACTAGCTAATCC 
Probe E. coli FAM-ATCGGATGTGCCCAGAT-BHQ1 
Dmet ereA2 F AAAAATAGTAGTGGAGAACGACCAGAACAC 

ereA2 Dmet in pMBA 
Dmet ereA2 R TCCACTACTATTTTTTATCCTTTTGCGTTTATTGCT 

Dmet ereA3 F AAAAATAGTAGTGGAGAACTACCAGAACACTT 
ereA3 Dmet in pMBA 

Dmet ereA3 R CTCCACTACTATTTTTTAGCCTTTGCGCTCAT 
mut ereA2 F2 GCGGGTGCTCCCTTTGTCGCGGAGTTC 

ereA2 cat. mut. mut ereA2 R2 GGGAGCACCCGCGCCAATGCCGACAAT 
mut ereA3 F2 GCGGGCGCTCCCTTTGTCGCGGAGTTT 

ereA3 cat. mut. mut ereA3 R2 GGGAGCGCCCGCGCCAAGGCCGAC 
check conj p2714 F GGAATTCGGCTTGTTATGAC Integrase-dependent 

recombination checking check conj gfp2.0 R CTCGATTCTATTAACAAGGG 
ribo GFP F ATGAGTAAAGGAGAAGAACTTTT pMBA5´UTR generation 

aacA1 GFP R AGTTCTTCTCCTTTACTCATTAGCGGCGTCGCCCTAAC pMBA5´UTR generation 
aacA2 GFP R GTTCTTCTCCTTTACTCATGACGCCTAACTTTGTTTTAGG pMBA5´UTR generation 
aacA3 GFP R AAGTTCTTCTCCTTTACTCATGAAATGGTCGCTCTGTGCT pMBA5´UTR generation 
aacA4 GFP R AGTTCTTCTCCTTTACTCATTGTGACGGAATCGTTGCTG pMBA5´UTR generation 
aacA5 GFP R AAGTTCTTCTCCTTTACTCATGCTGAAGTGTCTCCGTGCT pMBA5´UTR generation 
aacA7 GFP R AAAAGTTCTTCTCCTTTACTCATTGGTGCCTAACTTTG pMBA5´UTR generation 
aacA8 GFP R AAAAGTTCTTCTCCTTTACTCATGAGGACGGAGTTTGTGC pMBA5´UTR generation 
aacA16 GFP R AAAAGTTCTTCTCCTTTACTCATATAGGTAATGCTAGAAC pMBA5´UTR generation 
aacA17 GFP R AAAAGTTCTTCTCCTTTACTCATATAGGTAATACTAGAAC pMBA5´UTR generation 
aacA27 GFP R AAAAGTTCTTCTCCTTTACTCATGAGATGTTGATTCCGTG pMBA5´UTR generation 
aacA28 GFP R AAAAGTTCTTCTCCTTTACTCATAAAACCTCTTCGCGCGC pMBA5´UTR generation 
aacA29 GFP R AAAAGTTCTTCTCCTTTACTCATAGCCGTCTAACTTTG pMBA5´UTR generation 
aacA30 GFP R AAAAGTTCTTCTCCTTTACTCATTTAAGGTCTCCGAGTAG pMBA5´UTR generation 
aacA31 GFP R AAAAGTTCTTCTCCTTTACTCATGAGAACGGTCTTTGTGC pMBA5´UTR generation 
aacA32 GFP R AAAAGTTCTTCTCCTTTACTCATGGGACGGTCCTTTTGTG pMBA5´UTR generation 
aacA34 GFP R AAAAGTTCTTCTCCTTTACTCATAGCCTGGGCCTTCTAAC pMBA5´UTR generation 
aacA35 GFP R AAAAGTTCTTCTCCTTTACTCATAAGAAGGTGGCCCTGTG pMBA5´UTR generation 
aacA37GFP R AAAAGTTCTTCTCCTTTACTCATAGCCGCGGTTAACTTTG pMBA5´UTR generation 
aacA38 GFP R AAAGTTCTTCTCCTTTACTCATGGTGTGACGGTGGTTATG pMBA5´UTR generation 
aacA39 GFP R AAAAGTTCTTCTCCTTTACTCATAATACCTCTTTGCGCG pMBA5´UTR generation 
aacA40 GFP R AAGTTCTTCTCCTTTACTCATGCGGAGTCTGGACTGTGCT pMBA5´UTR generation 
aacA42 GFP R AAAAGTTCTTCTCCTTTACTCATTCAAAGTCTCCGAGTAG pMBA5´UTR generation 
aacA43 GFP R AAAAGTTCTTCTCCTTTACTCATTTGTCCTCGGCGCGAA pMBA5´UTR generation 
aacA44 GFP R AAAAGTTCTTCTCCTTTACTCATTTAATCACCAATTGTGCTCAGTC pMBA5´UTR generation 
aacA45 GFP R AAAAGTTCTTCTCCTTTACTCATTGGCTGAGCCTTTTAAC pMBA5´UTR generation 
aacA46 GFP R AAAAGTTCTTCTCCTTTACTCATAAGATCCTCTTTCTTCACGC pMBA5´UTR generation 
aacA47 GFP R AAAAGTTCTTCTCCTTTACTCATAACTTTGTTTTAGGGCG pMBA5´UTR generation 



Supplementary Material 

  166 

aacA48 GFP R AAAAGTTCTTCTCCTTTACTCATGCTCGGTGCAGGTTTAG pMBA5´UTR generation 
aacA49 GFP R AAAAGTTCTTCTCCTTTACTCATTGAAACTCTCCTCTGCG pMBA5´UTR generation 
aacA50 GFP R AAAGTTCTTCTCCTTTACTCATGAGAATTGTCTTTGTGC pMBA5´UTR generation 
aacA51 GFP R AAAAGTTCTTCTCCTTTACTCATACTGGGACGGTTCCTTG pMBA5´UTR generation 
aacA52 GFP R AAAAGTTCTTCTCCTTTACTCATCTTGTGCTGCCTAACTT pMBA5´UTR generation 
aacA54 GFP R AAAGTTCTTCTCCTTTACTCATAAGGCGTTGATTTTGTGC pMBA5´UTR generation 
aacA56 GFP R AAAAGTTCTTCTCCTTTACTCATTCTTACTCCTGCGCGAA pMBA5´UTR generation 
aacA59 GFP R AAAAGTTCTTCTCCTTTACTCATGGGATGCGGCTTCTGT pMBA5´UTR generation 
aacA61 GFP R AAAAGTTCTTCTCCTTTACTCATAAGCGGTGGCCCTGTGC pMBA5´UTR generation 
aacA64 GFP R AAAAGTTCTTCTCCTTTACTCATTTTATTCCTAGCGCCGA pMBA5´UTR generation 
aacAX GFP R AAAAGTTCTTCTCCTTTACTCATTTGGTAGATGGCTTCTC pMBA5´UTR generation 

aacC1C4 GFPR AAAAGTTCTTCTCCTTTACTCATACTTGAGCCACCTAACT pMBA5´UTR generation 
aacC2 GFP R AAAAGTTCTTCTCCTTTACTCATTGAGCCACCTAACTTTG pMBA5´UTR generation 
aacC3 GFP R AAAAGTTCTTCTCCTTTACTCATCTTAGCTGCTGCTGCCT pMBA5´UTR generation 
aacC5 GFP R AAAAGTTCTTCTCCTTTACTCATCGCAACATCGTTTCCAG pMBA5´UTR generation 
aacC6 GFP R AAAAGTTCTTCTCCTTTACTCATTGAGCCACCTAACTTTG pMBA5´UTR generation 
aacC11 GFP R AAAAGTTCTTCTCCTTTACTCATACGTGAGCCACCTAAC pMBA5´UTR generation 
aacC13 GFP R AAAAGTTCTTCTCCTTTACTCATTCATCGGCTCCTAATGC pMBA5´UTR generation 
aadA1 GFP R TTCTTCTCCTTTACTCATGATGTTTAACTTTGTTTTAGGGC pMBA5´UTR generation 
aadA4 GFP R AAAAGTTCTTCTCCTTTACTCATGAAGATGCCTAACTTTG pMBA5´UTR generation 
aadA5 GFP R AAAAGTTCTTCTCCTTTACTCATGATGCCTAACTTTG pMBA5´UTR generation 
aadA9 GFP R AAGTTCTTCTCCTTTACTCATGTCTAACTTTGTTTTAGGGCG pMBA5´UTR generation 
aadB GFP R AAAAGTTCTTCTCCTTTACTCATGCGGCCTAACTTTGTTT pMBA5´UTR generation 

aphA15 GFP R AAAAGTTCTTCTCCTTTACTCATAGCGGTCTAACTTTGTT pMBA5´UTR generation 
aphA16 GFP R AAAAGTTCTTCTCCTTTACTCATTTTCTCTTTCCTGCGCG pMBA5´UTR generation 
dfrA5 GFP R GTTCTTCTCCTTTACTCATTAATGATACTTTCACAATTTTGGTTCC pMBA5´UTR generation 
fosG GFP R AGTTCTTCTC CTTTACTCAT TCCTCGGAGCACAAATCTAC pMBA5´UTR generation 

blaOXA-9 GFP R AGTTCTTCTCCTTTACTCATTGCTCCGCTGTGCGCTTAAT pMBA5´UTR generation 
Tm F AGTCGCCCTAAAACAAAGTTTCGATTTTAAGATAACTTTACGTG pMBA5´UTR generation 
Tm R AAAAGTTCTTCTCCTTTACTCATTGCGTTCTACTCTGAAG pMBA5´UTR generation 

aacA1 alt R CTCTTTGTATTCGCGCCGCAATAGCGGCGTCGCCC pMBA5´UTR generation 
aacA43 alt R GTTCTTCTCCTTTACTCATCAATTGTCCTCGGCGC pMBA5´UTR generation 
aacA4 alt R TCTTCTCCTTTACTCATGATGCTGTACTTTGTGATGC pMBA5´UTR generation 
aadA9 alt R TCTCCTTTACTCATCATGTCTAACTTTGTTTTAGGGC pMBA5´UTR generation 
aacA47alt R GTTCTTCTCCTTTACTCATAGGTATGGTGGTTCTGTG pMBA5´UTR generation 
aacAX alt R TTCTTCTCCTTTACTCATTACCAAGCAATACAATTGGTAG pMBA5´UTR generation 
aacC5 alt R TTCTCCTTTACTCATTCGTCTGCTCCTGATGCC pMBA5´UTR generation 

aadA2 6 7 10 11 
13 16 24 28 

29 34 GFP R 
CTAAAACAAAGTTAGACATCATGAGTAAAGGAGAAGAACT pMBA5´UTR generation 

  



Supplementary Material 

 167 

Supplementary Table S3. Antibiotic discs used in this study. 
 

Reference Antibiotic (Ab.) Ab. content (mg) 

CT0026B Kanamycin (K) 30 
CT1897B Streptomycin (S) 300 
CT0024B Gentamicin (CN) 10 
CT0056B Tobramycin (TOB) 10 
CT0107B Amikacin (AK) 30 
CT0545B Apramycin (APR) 15 
CT0033B Neomycin (N) 30 
CT0003B Ampicillin (AMP) 10 
CT0623B Ciprofloxacin (CIP) 1 
CT0152B Penicillin G (P) 1 iu 

68042 Amoxycillin (AMO) 20 
66178 Amoxycillin Clavulanic Acid (AMC) 20/10 

CT0149B Cefaclor (CEC) 30 
CT0166B Cefotaxime (CTX) 30 
CT0119B Cefoxitin (FOX) 30 
CT0412B Ceftazidime (CAZ) 30 
CT0455B Imipenem (IPM) 10 
CT0774B Meropenem (MEM) 10 
CT1761B Ertapenem (ETP) 10 
CT0264B Aztreonam (ATM) 30 
CT0052B Trimethoprim-Sulfamethoxazole 1:19 (SXT) 25 
CT0998B Blank dics (-) - 

  



Supplementary Material 

  168 

Supplementary Table S4. Antimicrobial compounds used in this study. 
 

Reference Trader Antimicrobial 
T7883-5G  Sigma Aldrich Trimethoprim 
K4000-5G  Sigma Aldrich Kanamycin sulfate 
S6501-25G  Sigma Aldrich Streptomycin sulfate salt 
G1914-5G  Sigma Aldrich Gentamicin sulfate 

PHR1079-1G  Sigma Aldrich Tobramycin 
A1774-1G  Sigma Aldrich Amikacin 
A2024-1G  Sigma Aldrich Apramycin sulfate salt 
A8523-5G  Sigma Aldrich Amoxicillin 
C6895-1G  Sigma Aldrich Cefaclor 

CDS020667-50MG  Sigma Aldrich Ceftazidime 
901967-H  MSD INVANZ (Ertapenem) 

PZ0038-25MG  Sigma Aldrich Aztreonam 
P5396-1G  Sigma Aldrich Fosfomycin disodium salt 
E5389-1G  Sigma Aldrich Erythromycin 
R3501-1G Sigma Aldrich Rifampicin 
C0378-25G  Sigma Aldrich Chloramphenicol 
B6295-100G  Sigma Aldrich Benzalkonium chloride 
282227-1G  Sigma Aldrich Chlorhexidine 

H6269-100G  Sigma Aldrich CTAB (Hexadecyltrimethylammonium bromide) 
S7507-10G Sigma Aldrich Sulfamethoxazole 
Ant-Zn-1G InvivoGen Zeocin 
0242.3-1G Roth Vancomycin hydrochloride 
S7796-1G Sigma Aldrich Sisomicin sulfate salt 
N6386-5G Sigma Aldrich Neomycin trisulfate salt hidrate 
17850-56-F Sigma Aldrich Ciprofloxacin 
T3383-25G Sigma Aldrich Tetracycline hydrochloride 

  



Supplementary Material 

 169 

Supplementary Table S5. Antimicrobial resistance cassettes (ARCs) sequences. 
In silico predicted main ORFs of each antimicrobial gene are shown in blue. ARCs described in E. coli in 
the data bases are underlined. 
 

ARC Sequence (5´ à 3´) 

dfrA1 

TTAACCTCTGAGGAAGAATTGTGAAACTATCACTAATGGTAGCTATATCGAAGAATGGAGTTATCGGGAATGGCCCTGATATTCCATG
GAGTGCCAAAGGTGAACAGCTCCTGTTTAAAGCTATTACCTATAACCAATGGCTGTTGGTTGGACGCAAGACTTTTGAATCAATGGG
AGCATTACCCAACCGAAAGTATGCGGTCGTAACACGTTCAAGTTTTACATCTGACAATGAGAACGTATTGATCTTTCCATCAATTAAA
GATGCTTTAACCAACCTAAAGAAAATAACGGATCATGTCATTGTTTCAGGTGGTGGGGAGATATACAAAAGCCTGATCGATCAAGTA
GATACACTACATATATCTACAATAGACATCGAGCCGGAAGGTGATGTTTACTTTCCTGAAATCCCCAGCAATTTTAGGCCAGTTTTTA
CCCAAGACTTCGCCTCTAACATAAATTATAGTTACCAAATCTGGCAAAAGGGTTAACAAGTGGCAGCAACGGATTCGCAAACCTGTC
ACGCCTTTTGTGCCAAAAGCCGCGCCAGGTTTGCGATCCGCTGTGCCAGGCG 

dfrA5 

TTAACCCGGAACCAAAATTGTGAAAGTATCATTAATGGCTGCAAAAGCGAAAAACGGAGTGATTGGTTGCGGTCCACACATACCCTG
GTCCGCGAAAGGAGAGCAGCTACTCTTTAAAGCCTTGACGTACAACCAGTGGCTTTTGGTGGGCCGCAAGACGTTCGAATCTATGG
GAGCACTCCCTAATAGGAAATACGCGGTCGTTACTCGCTCAGCCTGGACGGCCGATAATGACAACGTAATAGTATTCCCGTCGATC
GAAGAGGCCATGTACGGGCTGGCTGAACTCACCGATCACGTTATAGTGTCTGGTGGCGGGGAGATTTACAGAGAAACATTGCCCAT
GGCCTCTACGCTCCATATATCGACGATTGATATTGAGCCGGAAGGAGATGTTTTCTTTCCGAATATTCCCAATACCTTCGAAGTTGTT
TTTGAGCAACACTTTAGCTCAAACATTAACTATTGCTATCAAATTTGGCAAAAGGGTTAACAAAGCTATGCAATTGACGGTAAAAAGCT
TCGTTCGCTTCGCTTGCTACGCTTCTTACCGCAATTGATAACGGCG 

dfrA6 

TTAGCCCTCAGGAGGAAAAATGAAAATATCTCTTATGGCAGCTGTTTCCGAGAATGGAGTAATTGGCTCTGGATTGGATATACCTTG
GCATGTACAAGGCGAGCAGCTCCTATTCAAAGCCATGACTTACAATCAATGGCTTCTAGTTGGTCGTAAAACCTTCGACTCAATGGG
TAAACTTCCGAATAGAAAATATGCAGTGGTTACTCGTTCTAAAATTATCTCGAATGACCCTGATGTTGTGTATTTCGCAAGTGTTGAAT
CGGCATTAGCTTACCTAAACAATGCGACAGCACATATCTTTGTTTCTGGTGGTGGTGAAATATATAAAGCTTTAATCGATCAAGCAGA
TGTTATCCATCTTTCAGTGATTCACAAGCATATCTCTGGCGATGTGTTTTTTCCTCCAGTTCCACAGGGCTTCAAGCAAACATTTGAG
CAAAGTTTCAGTTCAAATATTGATTACACGTACCAAATTTGGGCAAAGGGCTAACAATCTGTTTAAGAGTGATTCGCAACGCGTGGAA
TTTTTACTATGCGTTGCGTTTAGTGTTTAAGGTGGTATGCGGAGGCTTCGGTATTGCGTTGCTCACACCTTAACAGGGCG 

dfrA7 

TTAGCCATTACGGGGGTTGAATTGAAAATTTCATTGATTTCTGCAACGTCAGAAAATGGCGTAATCGGTAATGGCCCTGATATCCCAT
GGTCAGCAAAAGGTGAGCAGTTACTCTTTAAAGCGCTCACATATAATCAGTGGCTCCTTGTTGGAAGGAAAACATTTGACTCTATGG
GTGTTCTTCCAAATCGAAAATATGCAGTAGTGTCGAGGAAAGGAATTTCAAGCTCAAATGAAAATGTATTAGTCTTTCCTTCAATAGAA
ATCGCTTTGCAAGAACTATCGAAAATTACAGATCATTTATATGTCTCTGGTGGCGGTCAAATCTACAATAGTCTTATTGAAAAAGCAGA
TATAATTCATTTGTCTACTGTTCACGTTGAGGTTGAAGGTGATATCAATTTTCCTAAAATTCCAGAGAATTTCAATTTGGTTTTTGAGCA
GTTTTTTTTGTCTAATATAAATTACACATATCAGATTTGGAAAAAAGGCTAACAAGTCGTTCCAGCACCAGTCGCTGCGCTCCTTGGA
CAGTTTTTTAAGTCGCGGTTTTATGGTTTTGCTGCGCAAAAGTATTCCATAAAACCACAACTTAAAAACCGCCGCTGAACTCGGCG 

dfrA12 

TTAGCCATATGAACTCGGAATCAGTACGCATTTATCTCGTTGCTGCGATGGGAGCCAATCGGGTTATTGGCAATGGTCCTAATATCC
CCTGGAAAATTCCGGGTGAGCAGAAGATTTTTCGCAGACTCACTGAGGGAAAAGTCGTTGTCATGGGGCGAAAGACCTTTGAGTCT
ATCGGCAAGCCTCTACCGAACCGTCACACATTGGTAATCTCACGCCAAGCTAACTACCGCGCCACTGGCTGCGTAGTTGTTTCAAC
GCTGTCGCACGCTATCGCTTTGGCATCCGAACTCGGCAATGAACTCTACGTCGCGGGCGGAGCTGAGATATACACTCTGGCACTAC
CTCACGCCCACGGCGTGTTTCTATCTGAGGTACATCAAACCTTCGAGGGTGACGCCTTCTTCCCAATGCTCAACGAAACAGAATTCG
AGCTTGTCTCAACCGAAACCATTCAAGCTGTAATTCCGTACACCCACTCCGTTTATGCGCGTCGAAACGGCTAACCATTCCGTCAAC
GGGACGCCAAAATGCTGCGCATTTTGGTTCCCTCCGCTGCGCTCCGGCTCTCGTTACGTCCAACG 

dfrA14 

TTAACCCAGGATGAGAACCTTGAAAGTATCATTGATAGCTGCGAAAGCGAAAAACGGCGTGATTGGTTGCGGTCCAGACATACCGT
GGTCCGCGAAAGGGGAGCAGCTACTTTTTAAAGCATTGACCTACAATCAGTGTCTTCTGGTGGGTCGCAAGACGTTTGAATCTATGG
GCGCACTCCCCAATAGGAAATACGCGGTCGTTACCCGCTCAGGTTGGACATCAAATGATGACAATGTAGTTGTATTTCAGTCAATCG
AAGAGGCCATGGACAGGCTAGCTGAATTCACCGGTCACGTTATAGTGTCTGGTGGCGGAGAAATTTACCGAGAAACATTACCCATG
GCCTCTACGCTCCACTTATCGACGATCGACATCGAGCCAGAGGGGGATGTTTTCTTCCCGAGTATTCCAAATACCTTCGAAGTTGTT
TTTGAGCAACACTTTACTTCAAACATTAACTATTGCTATCAAATTTGGAAAAAGGGTTAACAAAGCTATGCAATCGACGGCAAAAAGCT
TCGTTCGCTTCGCGCACTACGCTTTTTCCGCGATTGATAGCGACG 

dfrA15 

TTAACCCCTAAGGAAGTATCGTGAAACTATCACTAATGGCAGCAATTTCGAAGAATGGAGTTATCGGAAATGGCCCAGATATTCCAT
GGAGTGCCAAAGGGGAACAATTACTCTTCAAAGCGATTACCTATAATCAGTGGCTTTTGGTAGGCCGAAAGACTTTCGAGTCAATGG
GGGCTTTACCCAACCGAAAATATGCCGTTGTAACTCGTTCAAGCTTCACTTCCAGTGATGAGAATGTATTGGTATTTCCATCTATCGA
TGAAGCGCTAAATCATCTGAAGACGATAACGGATCATGTGATTGTGTCTGGTGGTGGTGAAATATACAAAAGCCTGATCGATAAAGT
TGATACTTTACATATTTCAACAATCGACATTGAGCCAGAAGGTGATGTCTATTTTCCAGAAATCCCCAGTAGTTTTAGGCCAGTTTTTA
GCCAAGACTTCGTGTCTAACATAAATTATAGTTACCAAATCTGGCAAAAGGGTTAACAAGTGGCAGCAACTGACCGCCAAAAGTGTC
ACTTGTTTTGCCAAAAAGCCGGCAAAACAAGCGCCAATTTTGTCGGCAGCTGTGCCAGGCG 

dfrA16 

TTAACTCGAGGGAGAAATCGTGAAGTTATCACTAATGGCTGCCAAGTCGAAGAACGGTATTATCGGTAATGGACCAGATATTCCATG
GAGCGCCAAAGGCGAGCAACTTCTATTTAAGGCAATTACATATAATCAATGGCTTTTAGTTGGACGCAAAACTTTTGAGTCAATGGGC
GCTCTCCCAAATCGAAAGTATGCAGTTGTAACTCGCTCTAATTTTTCTACGAATGATGAGGGTGTAATGGTTTTCTCCTCAATTCAGG
ATGCCTTAATAAATTTAGAGGAAATCACGGATCATGTTATCGTTTCTGGTGGTGGTGAAATATACAAAAGCTTGATTTCCAAAGTAGAT
ACTTTGCATATTTCAACAGTCGACATCGAGCGAGATGGAGACATAGTTTTTCCTGAAATCCCAGATACATTCAAGTTGGTATTTGAGC
AAGATTTCGAGTCTAACATTAACTATTGTTATCAAATCTGGCAAAAGAGTTAACAAGCGCCTGCAATCTGACCTCCGGTTACTGTCAC
CTTTTTTTGCGGTGGAGCTGCAAAAAAGGCGCCATTAACCTCCGGCAGTTGAGGCGGGCG 

dfrA17 

TTAGCCATTAAGGGAGTTAAATTGAAAATATCATTGATTTCTGCAGTGTCAGAAAATGGCGTAATCGGTAGTGGTCCTGATATCCCGT
GGTCAGTAAAAGGTGAGCAACTACTCTTTAAAGCGCTCACATATAATCAATGGCTCCTTGTCGGAAGAAAAACATTTGACTCTATGGG
TGTTCTTCCAAATCGCAAATATGCAGTAGTGTCAAAGAACGGAATTTCAAGCTCAAATGAAAACGTCCTAGTTTTTCCTTCAATAGAAA
ATGCTTTGAAAGAGCTATCAAAAGTTACAGATCATGTATATGTCTCTGGCGGGGGTCAAATCTATAATAGCCTTATTGAAAAAGCAGA
TATAATTCATTTGTCTACTGTTCACGTTGAAGTCGAAGGTGATATCAAATTCCCTATAATGCCTGAGAATTTCAATTTGGTTTTTGAAC
AGTTTTTTATGTCTAATATAAATTATACATACCAGATTTGGAAAAAAGGCTAACAATGCGTTGCAGCACCAGTCGCTTCGCTCCTTGG
ACAGCTTTTAAGTCGCGTCTTTGTGGTTTTGCTGCGCAAAAGTATTCCACAAAGCCGCAACTTAAAAGCTGCCGCTGAACTTAACG 

dfrA21 

TTAGCCGTATGAACCCGGAATCGGTCCGCATTTATCTGGTCGCTGCCATGGGTGCCAATCGGGTTATTGGCAATGGTCCCGATATC
CCCTGGAAAATCCCAGGTGAGCAGAAGATTTTTCGCAGGCTCACCGAGAGCAAAGTGGTCGTTATGGGCCGCAAGACATTTGAGTC
CATAGGCAAGCCCTTACCAAACCGCCACACAGTGGTGCTCTCGCGCCAAGCTCGTTATAGCGCTCCTGGTTGTGCAGTTGTTTCAA
CGCTGTCACAGGCTATCGCCATCGCAGCCGAACACGGCAAAGAACTCTACGTAGCCGGCGGAGCCGAGGTATATGCGCTGGCGCT
ACCGCATGCCAACGGCGTCTTTCTATCTGAGGTACATCAAACCTTTGAGGGTGACGCCTTCTTCCCAGTGCTTAACGCAGCAGAATT
CGAGGTTGTCTCATCCGAAACCATTCAAGGCACAATCACGTACACGCACTCCGTCTATGCGCGTCGTAACGGCTAACAAGTCCGTC
AACGGGACACCCAAATGCTGCGCATTTGGGTTCCCTTGGCTGCGCCTCGGCGCCCGTTACGTCCAACG 

dfrA22 
TTAGCCGTATGAACCCGGAATTGGTCCGCATTTATCTGGTCGCTGCCATGGGTGCCAATCGGGTTATTGGCAATGGCCCCGATATT
CCCTGGAAAATCCCGGGTGAGCAAAAGATCTTTCGCAGGCTCACCGAGGGCAAAGTGGTCGTTATGGGCCGCAAGACGTTTGAGT
CCATAGGCAAGCCCTTACCAAACCGCCGCACAGTGGTGCTCTCGCGCCAAGCCAGTTATAGCGCTGCTGGTTGTGCAGTTGTTTCA
ACGCTGTCGCAGGCTATTGCCATCGCAGCCGAACACGGCAAAGAGCTCTACGTGGCCGGCGGAGCCGAGGTATATGCACTGGCAC
TACCTCGTGCCGACGGCGTCTTTCTATATGAGGTACATCAAACCTTCGAGGGTGACGCCTTCTTCCCTGTGCTCGACGAAGCAGAAT



Supplementary Material 

  170 

TCGAGGTTGTCTCAGCCGAAACCGTTCAAGCCACAATCACGTACACGCACTCCGTCTATGCACGTCGTAACGGCTAACAAGTCCGT
CAACGGGACACCCAAATGCTGCGCATTTGGGGTCCCTCGGCTGCGCCTCGGTGCCCGTTACGTCCAACG 

dfrA25 

TTAACCCAGGACGAGTACCTTGAAAGTATCATTGATGGCTGCAAGAGCGAAAAATGGCGTAATCGGTTGCGGTCCTGACATTCCTTG
GTCTGCCAAAGGGGAACAGCTTCTTTTCAAAGCACTGACCTATAACCAATGGCTTTTGGTAGGGCGCAAAACATTTGAGTCTATGGG
GCCGCTGCCCAATAGGAAATACGCGGTTGTTACCCGCTCAAACTGGACAGCGGCTAATGAAAACGTAGTGGTTTTCCCGTCGATTG
ACGAAGCGATGGGTAGATTAGGCGAGATCACTGACCATGTCATCGTCGCCGGTGGTGGAGAAATCTACCATGAAACGATACCCATG
GCCTCTACTCTGCATGTGTCGACAATCGACGTTGAGCCAGAGGGAGACGTTTTCTTTCCGAACATTCCTGGGAAGTTTGATGTCGTT
TTTGAGCAACAATTTACATCAAACATTAACTATTGCTATCAAATCTGGCAAAAGGGTTAACAAAGCTATGCAATTGACGGCAAAAAGC
TTCGTTCGCTTCACTCACTACGCAATTTGCCGCACTTGATAGCGGCG 

dfrA27 

TTAACCCAAAGGAGTATCGTGAAAATATCACTAATGGCTGCAAAAGCAAGAAATGGGGTTATTGGCTGCGGCTCGGATATCCCGTGG
AACGCTAAAGGTGAGCAGCTGCTTTTTAAAGCAATAACTTACAATCAATGGCTCTTAGTCGGCCGTAAAACATTTGAGGCAATGGGG
GCTCTCCCAAATAGAAAGTATGCAGTTGTCAGCCGCTCAGGATCGGTAGCTACTAACGATGATGTGGTTGTGTTTCCATCTATAGAA
GCAGCAATGAGGGAGCTAAAGACTCTTACGAACCATGTTGTTGTTTCTGGTGGTGGAGAGATCTACAAGAGTCTGATCGCCCATGC
CGACACGCTACATATCTCGACAATAGATTCCGAGCCAGAGGGCAATGTTTTCTTTCCGGAAATCCCCAAAGAGTTCAATGTGGTGTT
CGAGCAGGAATTTCATTCAAATATAAATTATCGCTATCAAATCTGGCAAAGGGGTTAACCATCCAAGCCATCGGACACATTTTGCTTC
GCTGCGCTCAAAACGCAAAATGTGCCGCTGCTTAGCGGCG 

dfrA29 

TTAACCATTACGGGAATTTAATTGAAAATTTCTTTAATCGCAGCTCAGTCAGAAAACGGTGTTATTGGTAATGGCCCAGATATTCCATG
GTCAGCAAAAGGGGAGCAGTTACTTTTCAAAGCGCTAACATATAATCAGTGGCTTCTTGTCGGAAGAAAAACATTTGAGTCAATGGG
TATTCTTCCTAATCGAAAGTATGCTGTTCTTTCAAAAAATGGAATTTCACACCTTCCTGAAAACGTACTAGTTTTTTCGTCTATAGAAAA
TGCATTATATGAACTGGCTAAGGTAACAGACCATTTATATATTTCTGGCGGTGGTCAAATATATAATAGTCTTATTGAAAGTGCTGATA
CCATCCACTTATCTATCATCCACAAAGAGGTAGAAGGTGAAGTAAGGTTTCCCAAAATACCTCCTAATTACAAGTTGGTATTTGAGCA
ATATTATTCTTCAAATATTAATTACACTTATCAAATTTGGCAAAAAGGTTAACAAGTCGCTCAAGCACCAGTCGCTTCGCTCCTTGGAC
AGTTTTTAAGTCGCAGTTTTGTGGTTTTGCTGCGCAAAAAGTTTCCACAAAACCACAACTTAAAAACTGCCGCTTAGCTCGGCG 

dfrA30 

TTAACCCGGGACCAAAATTGTGAAAGTATCATTAATGGCTGCAAGAGCGAGAAACGGAGTGATCGGTTGCGGTCCACACATACCCT
GGTCCGCGAAAGGAGAGCAGCTACTCTTTAAAGCCCTGACGTACAACCAGTGGCTTTTGGTTGGCCGCAAGACGTTCGAATCAATG
GGGGCGCTCCCCAACAGGAAATACGCGGTCGTTACTCGCTCAGCCTGGACGGCCAATAATGACAACGTAGTAGTATTCCCGTCGAT
CGAAGAGGCCATGGGCGGTCTAGCTAAACTCAACGGTCACGTTATAGTGTCTGGTGGCGGGGAGATTTACAGAGAAACGTTGCCCA
TGGCCTCTACGCTCCATGTATCGACGATCGACATTGAGCCAGAAGGGGATGTTTTCTTCCCGAATATTCCCAACTTCTTCGAAGTTG
TTTTTGAGCAACATTTTAGTTCAAACATTAACTATTGCTATCAAATTTGGAAAAAGGGTTAACAAAGCTATGCAATTGACGGCAAAAAG
CTTCGTTCGCCGCGCTCACTACGCTTTTTACCGCAATTGATAGCGGCG 

dfrA31 

TTAGCCCTCAGGAGGAAAAATGAAAATATCCATTATGGCAGCAGTTTCTGAGAATGGAGTAATTGGCTCTGGATTGGATATACCTTG
GCATGTACAAGGTGAGCAGCTCCTGTTCAAAGCTATGACTTACAATCATTGGCTTTTAGTCGGTCGTAAAACTTTCGACTCAATGGGT
AAACTTCCCAATAGGAAATATGCTGTGGTTACTCGCTCAGAAATGGTCTCGAATGATCCAGATGTTATTTATTTCACCAGCATTGAAT
CGGCATTATCTTACTTAGACAATACGACAACACATGTCTTTGTTTCTGGTGGTGGTGAAATTTACAAAGCATTAATCGAACAAGCAGA
TGTTATCCATCTTTCAGTGATTCATAAGCACATCTCTGGCGACGTGTTTTTCCCTTCAGTTCCACAGAGTTTCAAACAAACATTTGAGC
AAAGTTTCAGTTCAAATATTGATTACACGTACCAAATTTGGGCAAAGGGCTAACAAACTGTTTAAGAGGGATTCACAACGCGTGGCAT
TTTTACTATGCGTTGGTTTTAGTGGTTAAGGCGGTATGCGGAAGCTTCGGTATTGCGTTGCTCACCCCTTAACAGGGCG 

dfrA32 

TTAGCCATATCGGGAGTTAAATTGAAAATTTCATTGATTTCTGCAGTGTCAGAAAATGGCGTAATCGGTAGTGGTCCTGATATTCCGT
GGTCAGCAAAAGGTGAGCAGCTAATCTTTAAGGCGCTCACATACAATCAGTGGCTTCTTGTTGGAAGGAAAACATTTGACTCTATGG
GAGTTCTTCCAAATCGCAAATATGCAGTAGTGTCAAAGAATGGAATTTCAGGGTCAAATGAAAACGTCTTGGTTTTTCCTTCAATAGA
AAATGCTTTGCAAGAACTATCTAAAATTACAGATCATGTATATATTTCGGGTGGGGGGCAAATCTATGAAAGCCTTATTGAAAAAGCA
GATATAATTCATCTATCTACTATTCATGTTGAGGTTGAAGGTGATATTAAATTCCCTATATTACCTGAAGGTTTCAACTTGGTTTTTGAA
CAGTTTTTTGTGTCTAATATAAATTATACATATCAAATTTGGAAAAAAGGCTAACAAGTCGTTGCAGCACCAGTCGCTCCGCTCCTTG
GACAGTTTTTAAGTTGTGGTTTTATGGTTTTGCTGCGCAAAAATATTCCATAAAACCACAACTTAAAAACTGCCGCTGAACTCGGCG 

dfrA34 

TTAACCCAGGACCAAAACCTTGAAAGTATCATTAATGGCTGCAAGAGCGAAAAACGGAGTGATCGGTTGTGGTCCAGACATACCCTG
GTCCGCGAAAGGAGAGCAGCTACTTTTCAAAGCCTTGACGTACAATCAGTGGCTTTTGGTGGGCCGCAAGACGTTCGAATCTATCG
GGGCGCTCCCTAATAGGAAGTACGCGGTCGTTACCCGCTCAGACTGGACAATGGATCGTGACGATGTAGTAGTATTCCGGTCAATC
GAAGAGGCGATGGACGGGCTAGCTAAACACACCAGTCACGTTATAGTGTCTGGTGGTGGGGAGATTTACAGAGAAACCTTGCCGAT
AGCCTCTACCCTCCACGTATCGACAATCGACATTGAGCCAGAGGGAGACGTTTTCTTCCCGAATATTCCGAATACCTTCGACGTTGT
TTTTGAGCAACACTTTACTTCAAATATTAATTACTGCTATCAAATTTGGCAAAAAGGGTTAACAAAGCTATGCAATTGACGGCAAAAAG
CTTCGTTCGCTTCACTCACTACGCCTTTTGCCGCAATTGATAGCGGCG 
 

dfrA35 

TTAACTAGGAGAAATCGTTAAACTATCACTAATGGCAGCAATATCGAAGAATGGAGTCATCGGAAATGGCCCAGATATTCCATGGAG
TGCCAAAGGGGAACAATTACTGTTTAAGGCGATTACCTATAATCAATGGCTTTTGGTAGGTCGAAAGACTTTCGAATCTATGGGAGCT
TTACCCAATCGAAAGTATGCCGTTGTAACTCGTTCAAGCTTCACATCAAATGAAGAGAATGTATTGGTATTTCCATCTATCGAAGATG
CGCTAAACCAATTAGAGAAAATAACGGATCATGTGATTGTGTCTGGTGGTGGTGAAATATACAAAAGCCTAATATCTAAAGTAGATAC
ATTACATATCTCTACCATTGATATTGAGCCTGAAGGTGATGTTGTTTTTCCTAAAATTCCAGATTCATTTAAGTTAGTATTTAGCCAGG
AATTCGAGTCTAACATTAATTATTGTTACCAAATCTGGCAGATGGGTTAACAAGGGCCTGCAATCTGACCTCCGGCCACTGTCACCTT
TTGTGCAAAAAGCCGCACAAAAGGCGCCATTGGCCTCCGGCAGTTGAGGCGGGCG 

dfrB1 
TTGGGCGTGACCAATGCAGCGTGTCGTCGGGCCACATAGAACACCTAGAAGTTCACAAGAAAGGTCGGAAATGGAACGAAGTAGCA
ATGAAGTCAGTAATCCAGTTGCTGGCAATTTTGTATTCCCATCGAACGCCACGTTTGGTATGGGAGATCGCGTGCGCAAGAAATCCG
GCGCCGCCTGGCAAGGTCAGATTGTCGGGTGGTACTGCACAAATTTGACCCCCGAAGGCTACGCCGTCGAGTCTGAGGCTCACCC
AGGCTCAGTACAGATTTATCCTGTTGCGGCGCTTGAACGCATCAACTGAGTTCAAGGTTGAAGTGCGCCGCTCAACGGTCAAACCG
GGCATCACCGACTACAGCCCAACTTGTCGCTCCAGCGGACGGCTTCGCCGCCCGCTGAGCTAATTCG 

dfrB2 
TTAGGCAGCCGTTGTGCTGGTGCTTTCTGATAGTTGTTGTGGGGTAGGCAGTCAGAGTTCGATTTGCTTGTCGCCATAATAGATTCA
CAAGAAGGATTCGACATGGGTCAAAGTAGCGATGAAGCCAACGCTCCCGTTGCAGGGCAGTTTGCGCTTCCCCTGAGTGCCACCTT
TGGCTTAGGGGATCGCGTACGCAAGAAATCTGGTGCCGCTTGGCAGGGTCAAGTCGTCGGTTGGTATTGCACAAAACTCACTCCTG
AAGGCTATGCGGTCGAGTCCGAATCCCACCCAGGCTCAGTGCAAATTTATCCTGTGGCTGCACTTGAACGTGTGGCCTAACAATTC
GCTCCAGCGGACGGCTTCGCCGCCCGCTGAGCTTCATCG 

dfrB3 
TTGGGCTTCACCGACGCAGCGTTGTCGGGCTAGCAAGCATCTCAAGAAATTCACAAGAAAGGTCAGATATGGACCAACACAACAAT
GGAGTCAGTACTCTAGTTGCTGGCCAGTTTGCGCTCCCATCGCACGCCACGTTTGGCCTGGGAGATCGCGTGCGCAAGAAATCTG
GCGCCGCTTGGCAGGGTCAAGTTGTCGGGTGGTACTGCACAAAACTGACCCCTGAAGGCTATGCCGTCGAGTCCGAGTCTCACCC
CGGTTCAGTACAGATTTACCCTGTGGCTGCGCTTGAACGCGTGGCCTGAATTCACGGTTGAGTGCACCGCTCAGCGCTCAAGCCGT
GCAGTACCGCCCACAGCCCAACTGGTCGCTCCAGCGGACGGCTTCGCCGCCCGCTGAGCTAGAGCG 

dfrB4 
TTGGGCTTCACCAGAGTATCAAGTTGCCGAGCTGGCGATAACCTAAAACATTAAAAAGAGGTTTGAAATGAATGAAGGAAAAAATGA
GGTCAGTACTTCAGCTGCTGGCCGGTTCGCATTCCCATCAAACGCCACGTTTGCCTTGGGGGATCGCGTACGCAAGAAGTCTGGC
GCTGCTTGGCAGGGGCGCATTGTCGGGTGGTACTGCACAACACTTACCCCTGAAGGCTACGCCGTCGAGTCCGAATCTCACCCAG
GCTCAGTCCAGATTTATCCCATGACTGCGCTTGAACGGGTGGCCTGAAGTCACGGTTGAAATGCACCGCCTGACGGCCAAATCATG
CGGCACCGTCCACAGCCCAACTAGTCGCTCCAGCGGATGGCTTCGCCGCCCGCTGAGCTAATTCG 

dfrB5 
TTGGGCGTGACGATTGCAGCGTGTTGTCGGGCTACGCAGCAACCCTAGAAATTCAAAAGAAGGGTCATAAATGGACCAAGGCAGAA
GTGAAGTCAGTAATCCAGTTGCTGGCCAGTTTGCGTTCCCTTCAAACGCCGCGTTCGGAATGGGAGATCGCGTGCGCAAGAAATCT
GGCGCCGCTTGGCAAGGCCAGATTGTCGGGTGGTACTGCACAAAATTGACCCCTGAAGGGTACGCTGTCGAGTCTGAGGCTCACC
CTGGCTCGGTACAGATTTATCCTGTTGCGGCACTGGAACGCATCAACTGAGTTCAAGATTGAAGTGCTCCACTCAACGGCCATACC
GTGCATCACCGCCCACGGCCCAACTTGTCGCTCCAGCGGACGGCTTCGCCGCCCACTGAGCTAATTCG 

dfrB6 TTGGGCGTGACCACAGCTGCGTGTTGTCCGGCTACATAGCACCCCTAAAATTCACAAGAAAGGTCAGAAATGGACCAAGGTAGCAA
TGAAGTCATTAATCCAGTCGCTGGCCAGTTTGCGTCCCCATCGAACGCCACGTTTGGTATGGGAGATCGCGTGCGCAAGAAATCTG



Supplementary Material 

 171 

GCGCCGCCTGGCAAGGTCAGATTGTCGGGTGGTACAGCACAAAGTTGACCCCTGAAGGCTACGCTGTCGAGTCTGAGGCTCACCC
TGGCTCGGTGCAGATTTATCCTGTTGCCGCGCTTGAACGCGTCAACTGAGTTCAAGGTTGAAGTGCGCCACTCAAGGGCCAAACCG
TGCATCACCGCCAAAGGCCCAACTTGTCGCTCCAGCGGACGGCTTCGCCGCCCGCTGAGCTAATTCG 

dfrB7 

TTAGGCCCCGCAGGGGAGAGTCGGCGCTTTGAACTTCTTAGCCTGCGCAGAAAGTTGTAGTTCCCACGCCTAGCGCCGCAGTTCCT
GGCAGCTCACCGCGGATGCAACCAACGCTCACCCAGTCAAATGGTAGATATGGATCACCAGATCCCCTCAAGGCATCCACAAGAAA
GGTTAGGTATGGATCAAAGTAGTAAGGAAGTCAGTTCTCCAGCTACTGACCAGTTTGCGCTCCCATTCCGCGCCACGTTTGGCCTG
GGAGATCGCGTACGCAAGAAATCTGGCGCCGCTTGGCAGGGTCAAGTTGTCGGCTGGTACAGCACAAAACTAACCCCAGAAGGCT
ATGCCGTCGAGTCCGAGTCTCATCCGGGCTCTGTACAAATCTATCCTGTTGCCGCGCTTGAACGCGTGGCCTAACCCTTCCATCGA
GAGGACGTCACAAGGGCTACGCCCTTGCGCCGCCTCTCATGTCAAACG 

dfrB8 
TTGGGCATGACCACCGCAGCGTGTTGTCGGGCGATATTGCACCCCTAGAAATTCACAAGAGAGGTCAGAAATGGACCAAGGTAGCA
ATGAAGTCGGTAATCCAGTTGCGGGCCAGTTTTCGTTCCCATCGAACGCCGCGTTTAGTATGGGAGATCGCGTGCGCAAGAAATCG
GGCGCCGCTTGGCAAGGTCAGATTGTCGGGTGGTACTGCACAAAGTTGACCCCTGAAGGCTACGCTGTCGAGTCTGAGGCTCACC
CTGGCTCGGTACAGATTTATCCTGTTGCGGCGCTTGAACGCATCAACGGAGTTCAAGGTTGAAGTGCGCCACTCAACGGCCAAACC
GTGCATCACCGCCCACGGCCCAACCTGTCGCTCCAGCGGACGGCTTCGCCGTCCGCTGAGCTAATTCG 

dfrB9 

TTAGGCCCCGCAATGCGAACCTCTCGTGGTGCCAGACGATTTCGCGCATCCCGTCAGGAATTGAACAGTCGCCGGTGCCTGAATTC
AATCTCCCTACGACATTCAACGAGGCTGCCTGCCCACCACGCAGACCCCAGTGGATCATGGGCGAAAGACTTCAAGGTCATCGACC
AATCACGTAGTAAATGGACCAGTCGAAGGTCGCAAGACGATCACAAGAAAGGTTATGTATGAATCAAAGTAGCAATTGCATCAGCAC
TCCAGTTGTTGGACAGTTTGCGCTGCCATTTCAACCCACGTTCGGCCTGGGAGATCGCGTACGCAAGAAGTCTGGCGCCGCTTGGC
AAGGTAAAGTTGTCGGCTGGTACTGCACAAAATTAACCCCTGAAGGCTACGCGGTCGAGTCCGAAGCTCATCCAGGCTCAGTGCAG
ATTTATCCTGTGGCTGCGCTTGAACGCGTGGCCTAACCCCTCAGTCGAGGCGACAAATTGCAGCAAGCTGCAATTTGCGCCTCACC
TCGAACG 

blaBEL-1 

TTAGACGTAAGCCTATAATCTCTAGTATTTTGAAAATCGAGATCAAGATGAAACTGCTGCTCTACCCGTTATTGCTGTTCCTTGTCATT
CCAGCCTTTGCCCAGGCGGACTTTGAACATGCCATTTCAGATCTTGAGGCGCACAATCAAGCCAAGATCGGAGTGGCCCTAGTTAG
TGAAAATGGCAACCTGATTCAAGGGTATCGTGCGAATGAAAGGTTCGCGATGTGCTCAACTTTCAAGTTGCCGTTGGCCGCTCTTGT
TCTGAGTCGCATTGACGCTGGGGAAGAGAATCCTGAGCGCAAGCTTCATTACGATTCCGCGTTCCTTGAAGAGTACGCCCCAGCCG
CAAAACGGTATGTGGCAACTGGATATATGACTGTAACTGAGGCAATTCAATCCGCCCTCCAACTCAGCGACAATGCCGCAGCTAACC
TGCTGTTAAAAGAGGTTGGCGGCCCACCTTTATTGACAAAGTATTTCCGTAGCCTGGGTGATAAAGTAAGTCGCCTTGATCGTATTG
AACCGACTTTGAACACCAATACGCCCGGCGATGAAAGAGATACAACAACGCCCATGTCCATGGCACAGACTGTGTCAAAGCTGATTT
TTGGAGACACGTTGACATATAAATCCAAGGGGCAGCTAAGGCGATTACTCATCGGCAATCAGACCGGGGACAAAACCATTCGAGCT
GGCTTGCCTGATTCATGGGTAACGGGTGACAAGACAGGCTCGTGTGCGAATGGCGGCCGTAACGATGTGGCGTTTTTTATAACCAC
TGCCGGAAAAAAATATGTTCTTTCTGTATATACCAATGCACCTGAATTGCAAGGCGAGGAAAGGGCGTTATTAATTGCTTCTGTAGCA
AAGTTAGCACGTCAATATGTTGTTCACTGAATCCGCATACGTCTAACAATTCGCCTAAGCCGGACGCTCGCATGGCTCGCGCCGCTT
GGCT 

blaBIM-1 

TTAGAAAGGAATAAAAATGAGAATATCGTTAGCTTTATGCTTGCTTAGTATTTTAAATTTTGCTGTTGCAGAAGAGGTATTGCCAGAAA
TAAAAGTTGAAAAGCTAGAAGAAGGAATCTATCTTCACACATCTTATCTAGAGTATCAGGGAAATATTTATGAAAAACATGGGTTGGT
AGTTATTGATGATCGTAAGGCATATATAATTGATACGCCAGTTTTGGCTATAGATACTGAGCGGCTAGTAAAGTGGTTTGAAGAGCGT
AACTTTACTATAGGGGCTAGTTTCTCAACACATTTTCATAGTGACAGTTCCGGCGGCATAGAATGGCTAAATAAAAATTCTATTCCTAC
ATATGCATCTGAATTAACAAATGAACTTCTAAAAAAAGACGGCAAGGCGCAAGCTAAAAACTCTTTTAATGCAGTTAGTTTTTGGCTA
GTTAAAAACAAAATTGAAGTTTTTTATCCGGGGCCAGGGCATACACAGGATAACGAGGTCGTGTGGATACCTAGTAAGAAAATATTAT
TCGGTGGTTGTTTTGTTAAGCCAGACGGTCTTGGGTATTTGGGTGACGCAAATTTAGAAGCTTGGCCAAATTCTGCTAGAAAGTTAAT
GTCTAAATATAGCAACGCAAAACTGGTTATTCCAAGCCATAGTGAAATAGGAAATGCATCACTCTTGAAGCGTACATGGGAGCAGGC
TGTTAAAGGGCTAAATGACAGTAAAAAAACATCACAGCTCAGCAAATAAATTTCTAACAAGTCGCTCAAGCACCAGTCGCTACGCTCC
TTGGACAGTTTTTAAGTCGCAGTTTTGTGGTTTTGCTGCGCAAAAGTATTCCACAAAACCACAACTTAAAAACTGCCGCTTAGCTCGG
CG 

blaCARB-1 

TTAGCCATATTATGGAGCCTCATGCTTTTATATAAAATGTGTGACAATCAAAATTATGGGGTTACTTACATGAAGTTTTTATTGGCATTT
TCGCTTTTAATACCATCCGTGGTTTTTGCAAGTAGTTCAAAGTTTCAGCAAGTTGAACAAGACGTTAAGGCAATTGAAGTTTCTCTTTC
TGCTCGTATAGGTGTTTCCGTTCTTGATACTCAAAATGGAGAATATTGGGATTACAATGGCAATCAGCGCTTCCCGTTAACAAGTACT
TTTAAAACAATAGCTTGCGCTAAATTACTATATGATGCTGAGCAAGGAAAAGTTAATCCCAATAGTACAGTCGAGATTAAGAAAGCAG
ATCTTGTGACCTATTCCCCTGTAATAGAAAAGCAAGTAGGGCAGGCAATCACACTCGATGATGCGTGCTTCGCAACTATGACTACAA
GTGATAATACTGCGGCAAATATCATCCTAAGTGCTGTAGGTGGCCCCAAAGGCGTTACTGATTTTTTAAGACAAATTGGGGACAAAG
AGACTCGTCTAGACCGTATTGAGCCTGATTTAAATGAAGGTAAGCTCGGTGATTTGAGGGATACGACAACTCCTAAGGCAATAGCCA
GTACTTTGAATAAATTTTTATTTGGTTCCGCGCTATCTGAAATGAACCAGAAAAAATTAGAGTCTTGGATGGTGAACAATCAAGTCACT
GGTAATTTACTACGTTCAGTATTGCCGGCGGGATGGAACATTGCGGATCGCTCAGGTGCTGGCGGATTTGGTGCTCGGAGTATTAC
AGCAGTTGTGTGGAGTGAGCATCAAGCCCCAATTATTGTGAGCATCTATCTAGCTCAAACACAGGCTTCAATGGAAGAGCGAAATGA
TGCGATTGTTAAAATTGGTCATTCAATTTTTGACGTTTATACATCACAGTCGCGCTGATAAGGCTAACAAGGCCATCAAGTTGACGGC
TTTTCCGTCGCTTGTTTTGTGGTTTAACGCTACGCTACCACAAAACAATCAACTCCAAAGCCGCAACTTATGGCGGCG 

blaCARB-4 

TTAGCCTTATTTGAATACTTCACACTCTGATATAAGATGTAAGCCAATCAAAATGATGAGGTTATTACATGAAGCTTTTACTGGTATTTT
CGCTTTTAATACCGTCTATGGTGTTTGCAAATAGTTCAAAGTTTCAACAGGTTGAACAAGATGCTAAGGTAATTGAAGCATCTCTTTCT
GCGCATATAGGGATTTCTGTTCTTGATACTCAAACTGGAGAGTATTGGGATTACAATGGCAATCAGCGTTTTCCTTTGACAAGTACTT
TTAAAACAATAGCTTGTGCTAAATTATTATATGATGCTGAGCAAGGGGAAATAAACCCTAAGAGTACAATTGAGATCAAAAAAGCAGA
TCTTGTGACCTATTCTCCCGTAATAGAAAAGCAAGTAGGACAAGCAATAACGCTCGATGATGCGTGTTTTGCAACTATGACGACAAG
TGATAATGCAGCAGCAAATATCATCCTAAATGCCCTAGGAGGTCCTGAAAGCGTGACGGATTTTCTAAGACAAATCGGAGATAAAGA
AACCCGTCTAGACCGTATTGAACCTGAATTAAATGAAGGCAAGCTTGGTGATTTGAGGGATACGACAACTCCTAATGCAATAGTGAA
TACTTTAAATGAATTATTATTTGGTTCCACATTGTCTCAAGATGGCCAGAAAAAATTAGAGTATTGGATGGTGAATAATCAAGTCACTG
GTAATTTATTGCGGTCAGTATTGCCAGAGGGATGGAATATTGCGGATCGTTCAGGTGCTGGCGGATTTGGTGCTCGGAGTATTACA
GCCGTTGTTTGGAGTGAAGCTCAATCCCCAATCATAGTTAGTATCTATCTAGCGCAAACAGAGGCTTCAATAGCAGATCGAAATGAT
GCAATTGTTAAAATTGGTCGTTCAATTTTTGAAGTTTATTCATCACAATCGCGTTGATAAGGCTAACAAAGCATTTAAGAGGGACAGC
CAACGCGTGGCACTTTTATTATGCGTTGGTTTTTGTGGTTACGGTGTTATGCGGTAAGTTGGTAGTAGCGTTGGCTGCCCCTTAATG
CGGCG 

blaDIM-1 

TTAGAGGGAAAAATCGAATGAGAACACATTTTACAGCGTTATTACTTCTATTCAGCTTGTCTTCGCTTGCTAACGACGAGGTACCTGA
GCTAAGAATCGAGAAAGTAAAAGAGAACATCTTTTTGCACACATCATACAGTCGTGTGAATGGGTTTGGTTTGGTCAGTTCAAACGG
CCTTGTTGTCATAGATAAGGGTAATGCTTTCATTGTTGATACACCTTGGTCAGACCGAGATACAGAAACGCTCGTACATTGGATTCGT
AAAAATGGTTATGAGCTACTGGGGAGTGTTTCTACTCATTGGCATGAGGATAGAACCGCAGGAATTAAATGGCTTAATGACCAATCA
ATTTCTACGTATGCCACGACTTCAACCAACCATCTCTTGAAAGAAAATAAAAAAGAGCCAGCGAAATACACCTTGAAAGGAAATGAGT
CCACATTGGTTGACGGCCTTATCGAAGTATTTTATCCAGGAGGTGGTCATACAATAGACAACGTAGTGGTGTGGTTGCCAAAGTCGA
AAATCTTATTTGGCGGCTGTTTTGTGCGTAGCCTTGATTCCGAGGGGTTAGGCTACACTGGTGAAGCCCATATTGATCAATGGTCCC
GATCAGCTCAGAATGCTCTGTCTAGGTACTCAGAAGCCCAGATAGTAATTCCTGGCCATGGGAAAATCGGGGATATAGCGCTGTTAA
AACACACCAAAAGTCTGGCTGAGACAGCCTCTAACAAATCAATCCAGCCGAACGCTAACGCGTCGGCTGATTGAGGCG 

blaGES-1 

TTAGACGGGCGTACAAAGATAATTTCCATCTCAAGGGATCACCATGCGCTTCATTCACGCACTATTACTGGCAGGGATCGCTCACTC
TGCATATGCGTCGGAAAAATTAACCTTCAAGACCGATCTTGAGAAGCTAGAGCGCGAAAAAGCAGCTCAGATCGGTGTTGCGATCG
TCGATCCCCAAGGAGAGATCGTCGCGGGCCACCGAATGGCGCAGCGTTTTGCAATGTGCTCAACGTTCAAGTTTCCGCTAGCCGC
GCTGGTCTTTGAAAGAATTGACTCAGGCACCGAGCGGGGGGATCGAAAACTTTCATATGGGCCGGACATGATCGTCGAATGGTCTC
CTGCCACGGAGCGGTTTCTAGCATCGGGACACATGACGGTTCTCGAGGCAGCGCAAGCTGCGGTGCAGCTTAGCGACAATGGGGC
TACTAACCTCTTACTGAGAGAAATTGGCGGACCTGCTGCAATGACGCAGTATTTTCGTAAAATTGGCGACTCTGTGAGTCGGCTAGA
CCGGAAAGAGCCGGAGATGGGCGACAACACACCTGGCGACCTCAGAGATACAACTACGCCTATTGCTATGGCACGTACTGTGGCT
AAAGTCCTCTATGGCGGCGCACTGACGTCCACCTCGACCCACACCATTGAGAGGTGGCTGATCGGAAACCAAACGGGAGACGCGA



Supplementary Material 

  172 

CACTACGAGCGGGTTTTCCTAAAGATTGGGTTGTTGGAGAGAAAACTGGTACCTGCGCCAACGGGGGCCGGAACGACATTGGTTTT
TTTAAAGCCCAGGAGAGAGATTACGCTGTAGCGGTGTATACAACGGCCCCGAAACTATCGGCCGTAGAACGTGACGAATTAGTTGC
CTCTGTCGGTCAAGTTATTACACAACTCATCCTGAGCACGGACAAATAGTTGACGCCCGTCTAACAATTCGTTCAAGCCGACGTTGC
TTCGTGGCGGCGCTTGCGTGCTACGCTAAGCTTCGCACGCCGCTTGCCACTGCGCACCGCGGCTTAACTCAGGCG 

blaGIM-1 

TTAGAAGGATGATTTCCAAATGAAAAATGTATTAGTGTTTTTAATATTACTTGTAGCGTTGCCAGCTTTAGCTCAGGGTCATAAACCGC
TAGAAGTTATAAAAATTGAAGATGGAGTATATCTTCATACCTCCTTTAAGAATATTGAAGGCTATGGGTTAGTTGATTCGAATGGGTTG
GTAGTTCTGGATAATAATCAAGCCTATATTATCGACACACCTTGGTCTGAAGAAGACACGAAGTTGTTATTATCCTGGGCGACTGACA
GGGGATACCAGGTTATGGCTAGCATCTCAACTCATTCTCATGAAGATCGCACTGCTGGTATCAAGTTGCTAAATTCAAAGTCAATTCC
TACATACACATCAGAGTTAACTAAAAAGCTTCTTGCCCGTGAAGGAAAGCCGGTTCCTACCCACTACTTTAAAGACGACGAATTCACA
CTGGGAAATGGGCTTATAGAGCTCTACTATCCAGGTGCTGGGCATACAGAGGATAATATTGTTGCTTGGTTACCCAAAAGCAAAATA
CTATTTGGTGGCTGCCTCGTGAGGAGTCATGAGTGGGAAGGCTTAGGTTACGTAGGCGACGCCTCAATTAGCTCTTGGGCTGACTC
AATTAAAAATATTGTATCGAAAAAATATCCCATTCAAATGGTCGTTCCGGGGCATGGCAAAGTTGGAAGTTCAGATATATTAGATCAC
ACCATTGATCTTGCTGAATCAGCTTCTAACAAATTAATGCAACCGACCGCTGAAGCGTCGGCTGATTAAGGCG 

blaIMP-1 

TTAGAAAAGGAAAAGTATGAGCAAGTTATCTGTATTCTTTATATTTTTGTTTTGCAGCATTGCTACCGCAGCAGAGTCTTTGCCAGATT
TAAAAATTGAAAAGCTTGATGAAGGCGTTTATGTTCATACTTCGTTTGAAGAAGTTAACGGGTGGGGCGTTGTTCCTAAACATGGTTT
GGTGGTTCTTGTAAATGCTGAGGCTTACCTAATTGACACTCCATTTACGGCTAAAGATACTGAAAAGTTAGTCACTTGGTTTGTGGAG
CGTGGCTATAAAATAAAAGGCAGCATTTCCTCTCATTTTCATAGCGACAGCACGGGCGGAATAGAGTGGCTTAATTCTCGATCTATC
CCCACGTATGCATCTGAATTAACAAATGAACTGCTTAAAAAAGACGGTAAGGTTCAAGCCACAAATTCATTTAGCGGAGTTAACTATT
GGCTAGTTAAAAATAAAATTGAAGTTTTTTATCCAGGCCCGGGACACACTCCAGATAACGTAGTGGTTTGGTTGCCTGAAAGGAAAAT
ATTATTCGGTGGTTGTTTTATTAAACCGTACGGTTTAGGCAATTTGGGTGACGCAAATATAGAAGCTTGGCCAAAGTCCGCCAAATTA
TTAAAGTCCAAATATGGTAAGGCAAAACTGGTTGTTCCAAGTCACAGTGAAGTTGGAGACGCATCACTCTTGAAACTTACATTAGAGC
AGGCGGTTAAAGGGTTAAACGAAAGTAAAAAACCATCAAAACCAAGCAACTAAATTTCTAACAAGTCGTTGCAGCACGCCACTACGT
GGCTGGACAGTTTGTAAGTTGCGCTTTTGTGGTTTGCTTCGCAAAGTATTCCACAACGCGCAACTTACAAACTGCCGCTGAACTTAG
CG 

blaIMP-2 

TTAGAAAAGGGCAAGTATGAAGAAATTATTTGTTTTATGTGTATGCTTCCTTTGTAGCATTACTGCCGCGGGAGCGCGTTTGCCTGAT
TTAAAAATCGAGAAGCTTGAAGAAGGTGTTTATGTTCATACATCGTTCGAAGAAGTTAACGGTTGGGGTGTTGTTTCTAAACACGGTT
TGGTGGTTCTTGTAAACACTGACGCCTATCTGATTGACACTCCATTTACTGCTACAGATACTGAAAAGTTAGTCAATTGGTTTGTGGA
GCGCGGCTATAAAATCAAAGGCACTATTTCCTCACATTTCCATAGCGACAGCACAGGGGGAATAGAGTGGCTTAATTCTCAATCTAT
TCCCACGTATGCATCTGAATTAACAAATGAACTTCTTAAAAAAGACGGTAAGGTGCAAGCTAAAAACTCATTTAGCGGAGTTAGTTAT
TGGCTAGTTAAAAATAAAATTGAAGTTTTTTATCCCGGCCCGGGGCACACTCAAGATAACGTAGTGGTTTGGTTACCTGAAAAGAAAA
TTTTATTCGGTGGTTGTTTTGTTAAACCGGACGGTCTTGGTAATTTGGGTGACGCAAATTTAGAAGCTTGGCCAAAGTCCGCCAAAAT
ATTAATGTCTAAATATGTTAAAGCAAAACTGGTTGTTTCAAGTCATAGTGAAATTGGGGACGCATCACTCTTGAAACGTACATGGGAA
CAGGCTGTTAAAGGGCTAAATGAAAGTAAAAAACCATCACAGCCAAGTAACTAAATTTCTAACAAGGCGGTGAAACTCATTCCGCTTC
GCTTCACTGGACGCCGCAAGCGGCGCCGTTTACCTTCGCG 

blaIMP-4 

TTAGAAAAGGGAAAGTATGAGCAAGTTATCTGTATTCTTTATATTTTTGTTTTGTAGCATTGCTACCGCAGCAGAGCCTTTGCCAGATT
TAAAAATTGAAAAACTTGATGAAGGCGTTTATGTTCATACTTCGTTTGAAGAAGTTAACGGGTGGGGCGTTGTTCCTAAACATGGTTT
GGTTGTTCTTGTAGATGCTGAAGCTTATCTAATTGACACTCCATTTACGGCTAAAGATACTGAAAAGTTAGTCACTTGGTTTGTGGAA
CGTGGCTATAAAATAAAAGGCAGTATTTCCTCTCATTTTCATAGTGACAGCACGGGCGGAATAGAGTGGCTTAATTCTCAATCCATCC
CCACGTATGCGTCTGAATTAACTAATGAGCTGCTTAAAAAAGACGGTAAGGTTCAAGCTAAAAATTCATTTGGCGGGGTTAACTATTG
GCTAGTTAAAAATAAAATTGAAGTTTTTTATCCAGGCCCAGGACACACTCCAGATAACCTAGTAGTTTGGCTGCCTGAAAGGAAAATA
TTATTCGGTGGTTGTTTTATTAAACCGTACGGTCTAGGTAATTTGGGTGACGCAAATTTAGAAGCTTGGCCAAAGTCCGCTAAATTAT
TAATATCCAAATATGGTAAGGCAAAACTGGTTGTTCCAAGTCACAGTGAAGCTGGAGACGCATCACTCTTGAAACTTACATTAGAGCA
GGCGGTTAAAGGGTTAAACGAAAGTAAAAAACCATCAAAACTAAGCAACTAAATTTCTAACAAGTCGTTGCAGCATCGTGCGCTGCG
CGCACTGGACAGTTTTTAAGTCGCGGTTTTATGGTTTTGCTTCGCAAAAATATTCCATAAAACCACAACTTAAAAACTGCCGCTGAAC
TCGGCG 

blaIMP-5 

TTAGAAAAGGGAAAGTATGAGCAAGTTATTTGTATTCTTTATGTTTTTGTTTTGTAGCATTACTGCCGCAGCAGAGTCTTTGCCAGATT
TAAAAATTGAGAAGCTTGACGAAGGCGTTTATGTTCATACTTCGTTTGAAGAAGTTAACGGTTGGGGTGTTGTTCCTAAACACGGCTT
GGTGGTTCTTGTAAATACTGAGGCCTATCTGATTGACACTCCATTTACGGCTAAAGATACTGAAAAGTTAGTCACTTGGTTTGTGGAA
CGCGGCTATAAAATAAAAGGCAGTATTTCCTCTCATTTTCATAGCGACAGCACGGGCGGAATAGAGTGGCTTAATTCTCAATCTATCC
CCACGTATGCATCTGAATTAACAAATGAACTTCTTAAAAAAGACGGTAAAGTACAAGCTAAAAATTCATTTAGCGGAGCTAGCTATTG
GCTAGTTAAGAAAAAGATTGAAGTTTTTTATCCTGGTCCAGGGCACACTCCAGATAACGTAGTGGTTTGGCTACCTGAAAATAGAGTT
TTGTTCGGTGGTTGTTTTGTTAAACCGTACGGTCTAGGTAATTTGGGTGACGCAAATGTAGAAGCTTGGCCAAAGTCCGCCAAATTA
TTAATGTCCAAATATGGTAAGGCAAAACTGGTAGTTCCAAGTCACAGTGAAGTTGGAGACGCATCACTCTTGAAACGTACGTTAGAA
CAGGCGGTTAAAGGGTTAAACGAAAGTAAAAAACCATCAAAACCAAGTAACTAAATTTCTAACAAGTCGTTGCAGCACCGTTCCGCTT
CGCTGCACTGGACAGTTTTTAAGTTGCAGTTTTATGGTTTGCTGCGCAAATATTTCCATAAAACCACAACTTAAAAACTGCCGCTGAA
CTCGGCG 

blaIMP-9 

TTAGAAAAAGGGAAAGTATGAGCAAGTTATTTGTATTCTTTATGTTTTTGTTTTGTAGCATTACTGCCGCAGGAGAGTCTTTGCCAGAT
TTAAAAATTGAGAAGCTTGACGAAGGCGTTTATGTTCATACTTCGTTTGAAGAAGTTAACGGTTGGGGTGTTATTCCTAAACACGGCT
TGGTGGTTCTTGTAAATACTGATGCCTATCTGATAGACACTCCATTTACTGCTAAAGATACTGAAAATTTAGTTAATTGGTTTGTTGAG
CGCGGCTATAGAATAAAAGGCAGTATTTCCTCACATTTCCATAGCGACAGCACGGGTGGAATAGAGTGGCTTAATTCTCAATCTATC
CCCACGTATGCATCTGAATTAACAAATGAACTTCTTAAAAAAGACGGTAAGGTACAAGCTAAATATTCATTTAGCGGAGTTAGCTATT
GGCTAGTTAAGAAAAAGATTGAAGTTTTTTATCCTGGTCCAGGGCACGCTCCAGATAACGTAGTGGTTTGGCTGCCTGAAAATAGAG
TTTTGTTCGGTGGTTGTTTTGTTAAACCCTACGGTCTAGGTAATTTGGGTGACGCAAATTTAGAAGCTTGGCCAAAATCCGCCAAATT
ATTAATGTCAAAATATAGTAAGGCAAAACTGGTTGTACCAAGTCATAGTGACATAGGAGATTCGTCGCTCTTGAAGCTTACATGGGAG
CAGACGGTAAAAGGATTCAATGAAAGCAAAAAAAGTACCACTGCACATTAACCAAATTTCTAACAAGTCGCTCAAGCATCGCACCTTC
GGTGCTGGACAGTTTTTAAGTCGCGCTTTTGTGGTTTTGCTACGCAAAAGGTTTCCACAAAATCACAACTTAAAAACTGCCGCTTAGC
TCGGCG 

blaIMP-11 

TTAGAAAGGAGTAAGTATGAAAAAACTATTTGTTTTATGTATATTTTTGTTTTGTAGCATTACTGCCGCAGGAGCGTCTTTGCCTGATT
TAAAAATTGAGAAGCTTGAAGAGGGTGTTTATGTTCATACATCGTTTGAAGAAGTTAACGGCTGGGGTGTTGTTTCTAAACACGGTTT
GGTGGTTCTTGTAAATACTGACGCCTATCTGATTGACACTCCATTTACTGCTAAAGATACTGAAAAGTTAGTCAATTGGTTTGTGGAG
CGCGGCTATAAAATCAAAGGCAGTATTTCCTCACATTTCCATAGCGACAGCACGGGTGGAATAGAGTGGCTTAATTCTCAATCTATTC
CCACGTATGCATCTGTATTAACAAATGAACTTCTCAAAAAAGACGGTAAGGTGCAAGCTAAAAACTCATTTAGCGGAGTTAGCTATTG
GCTAGTTAAAAATAAAATTGAAGTTTTTTATCCAGGCCCAGGGCACACTCAAGATAACGTAGTGGTTTGGCTACCTAAAAATAAAATC
TTATTTGGTGGTTGTTTTGTTAAACCATATGGTCTTGGTAATCTAGATGACGCAAATGTTGAAGCATGGCCACATTCGGCTGAAAAAT
TAATATCTAAGTATGGTAATGCAAAACTGGTTGTTCCAAGCCATAGTGACATAGGAGATGCGTCGCTCTTGAAGCTTACGTGGGAAC
AGGCGGTAAAAGGGCTTAATGAAAGCAAAAAAAGTAACACTGTTCATTAACCAAATTTCTAACAAGTCGCTGAAGCATCGCACCTTC
GGTGCTGGACAGTTTTTAAGTCGCGCTTTTGTGGTTTTGCTACGCAAAAGGTTTCCACAAAATCACAACTTAAAAACTGCCGCTTAGC
TCGGCG 

blaIMP-12 

TTAGAAAAGGGAAAGTATGAAGAAATTATTTGTTTTATGCATTTTTTTGTTTTTAAGTATTACTGCCTCAGGTGAGGTTTTGCCTGATTT
GAAAATTGAGAAGCTTGAAGAGGGTGTTTATCTTCATACATCTTTTGAAGAGGTTAGCGGTTGGGGTGTTGTTACTAAACATGGTTTG
GTAGTTCTTGTAAATAATGACGCCTATCTAATTGACACTCCATTTACAAATAAAGATACTGAAAAATTAGTTGCTTGGTTTGTAGGGCG
CGGCTTTACAATAAAGGGAAGTGTTTCCTCACATTTTCATAGCGACAGTACGGGTGGAATAGAGTGGCTTAATTCTCAATCTATTCCC
ACGTATGCATCTGAGTTAACAAATGAACTTCTGAAAAAGAACGGTAAGGTGCAAGCTACAAATTCATTTAGCGGGGTTAGTTATTGGC
TAGTTAAAAATAAAATTGAAATTTTTTATCCCGGCCCAGGACATACTCAAGATAACGTAGTGGTTTGGCTACCTGAAAACAAAATTTTA
TTCGGTGGTTGTTTTGTTAAACCGGACGGTCTTGGTAATTTGGATGACGCAAATTTAAAAGCTTGGCCAAAGTCCGCAAAAATATTAA
TGTCTAAATATGGTAAAGCAAAGTTAGTTGTTTCAGGTCATAGTGAAATTGGGAACGCATCACTCTTGAAACTTACTTGGGAGCAGGC



Supplementary Material 

 173 

TGTTAAAGGGCTAAAAGAAAGTAAAAAACCATTACTGCCAAGTAACTAATTTTCTAACAAGTCGTTGCAGCACCGTTCCTTCGCTCCA
CTGGACAGTTTTTAAGTCGCGGTTTTATGGTTTGTGCGCAAAAGTATTTCATAAAACCACAACTTAAAAACTGCCGCTGAACTCGGCG 

blaIMP-13 

TTAGAAAAGGAAAGGTATGAAGAAATTATTTGTTTTATGTGTATGCTTCTTTTGTAGCATTACTGCCGCAGGAGCGGCTTTACCTGATT
TAAAAATCGAGAAGCTTGAAGAAGGTGTTTTTGTTCATACATCGTTCGAAGAGGTTAACGGTTGGGGGGTTGTTACTAAACACGGTTT
AGTGGTGCTTGTAAACACAGACGCCTATCTAATTGACACTCCATTTACTGCTACAGACACTGAAAAATTAGTCAATTGGTTTGTGGAG
CGCGGCTATGAAATCAAAGGCACTATTTCATCACATTTCCATAGCGACAGCACAGGAGGAATAGAGTGGCTTAATTCTCAATCTATTC
CCACGTATGCATCTGAATTAACAAATGAACTTTTGAAAAAATCCGGTAAGGTACAAGCTAAATATTCATTTAGCGAAGTTAGCTATTGG
CTAGTTAAAAATAAAATTGAAGTTTTCTACCCTGGCCCAGGTCACACTCAAGATAACCTAGTGGTTTGGTTGCCTGAAAGTAAAATTTT
ATTCGGTGGTTGCTTTATTAAACCTCACGGTCTTGGCAATTTAGGTGACGCAAATTTAGAAGCTTGGCCAAAGTCCGCCAAAATATTA
ATGTCTAAATATGGCAAAGCAAAGCTTGTTGTTTCAAGTCATAGTGAAAAAGGGGACGCATCACTAATGAAACGTACATGGGAACAA
GCCCTTAAAGGGCTTAAAGAAAGTAAAAAAACATCATCACCAAGTAACTAAATTTCTAACAAGTCGTTGCAGCACCGCGCACTTCGTG
CGCTGGACAGTTTTTAAGTCGCGGCTTTATGGTTTTGCTGCGCAAAAGTATTCCATAAAACCACAACTTAAAAACTGCCGCTGAACTC
GGCG 

blaIMP-14 

TTAGAAAAGGATAAGTATGAAAAAATTATTTGTTTTATGTGTATTCTTCTTCTGCAACATTGCAGTTGCAGAAGAATCTTTGCCTGATTT
AAAAATTGAGAAGCTTGAAGAAGGCGTTTATGTTCATACTTCGTTTGAAGAAGTTAAAGGTTGGAGTGTGGTCACTAAACACGGTTTG
GTGGTTCTTGTGAAAAATGACGCCTATCTGATTGATACTCCAATTACTGCTAAAGATACTGAAAAATTAGTCAATTGGTTTGTTGAGC
GGGGCTATAAAATCAAAGGCAGTATTTCAACACATTTCCATGGTGACAGTACGGCTGGAATAGAGTGGCTTAATTCTCAATCTATCCC
CACATATGCTTCTGAATTAACAAATGAACTTCTTAAAAAAGACAATAAGGTACAAGCTAAACACTCTTTTAATGGGGTTAGTTATTCAC
TAATTAAAAACAAAATTGAAGTTTTTTATCCAGGCCCAGGGCACACTCAAGATAACGTAGTGGTTTGGTTACCTGAAAAGAAAATTTTA
TTCGGTGGTTGCTTTGTTAAACCGGACGGTCTTGGCTATTTGGGGGACGCAAATTTAGAAGCTTGGCCAAAGTCCGCTAAAATATTA
ATGTCTAAATATGGTAAAGCAAAACTAGTTGTGTCGAGTCATAGTGATATTGGAGATGTATCACTCTTGAAACGTACATGGGAGCAGG
CTGTTAAAGGGCTGAATGAAAGTAAAAAATCATCACAGCCAAGCGACTAAATTTCTAACAAGGCGCTTCAGCACCGCGCACTTCGTG
CGCTCGACAGTTCGTAAGCCGCTTTTTTGTGGTTTTGCTACGCAAAAGGTTTCCACAAAAAATCAACTTACAAACTGCGGCTGAGCTT
AACG 

blaIMP-15 

TTAGAAAAGGTAAAGTATGAACAAGTTATCTGTATTCTTTATGTTTATGTTTTGTAGCATTACTGCCGCAGGAGAGTCTTTGCCAGATT
TAAAAATTGAGAAGCTTGACGAAGGTGTTTATGTTCATACTTCGTTTGAAGAAGTTAACGGTTGGGGTGTTGTTCCTAAACACGGCTT
GGTGGTTCTTGTAAATACTGAGGCCTATCTGATTGACACTCCATTTACGGCAAAAGATACTGAAAAGTTAGTCACTTGGTTTGTGGAG
CGCGGCTATAAAATAAAAGGCAGTATTTCCTCTCATTTTCATAGCGACAGCACGGGCGGAATAGAGTGGCTTAATTCTCAATCTATCC
CCACGTATGCATCTGAATTAACAAATGAACTTCTTAAAAAAGACGGTAAGGTACAAGCTAAAAATTCATTTAGCGGAGGTAGCTATTG
GCTAGTTAATAATAAGATTGAAGTTTTTTATCCTGGTCCAGGGCACACTCCAGATAACGTAGTGGTTTGGCTACCTGAAAATAGAGTT
TTGTTCGGTGGTTGTTTTGTTAAACCGTACGGTCTTGGTAATTTGGGTGACGCAAATTTAGAAGCTTGGCCAAAGTCCGCCAAAATAT
TAATGTCTAAATATGGTAAAGCAAAGTTGGTTGTTTCAAGTCATAGTGAAACTGGGAACGCATCACTCTTGAAACTTACTTGGGAGCA
GGCTGTTAAAGGGCTAAAAGAAAGTAAAAAACCATCACTGCCAAGTAACTAATTTTCTAACAAGTCGTTGCAGCACCGCTCCGGCAC
TTCGTGCCTGCGCTGGACAGTTTTTAAGTCGCAGTTTTATGGTTTTGCTGCGCAAAAGTATTCCATAAAACCACAACTTAAAAACTGC
CGCTGAACTCAGCG 

blaIMP-16 

TTAGAAAAGGGCGAGTATGAAAAAATTATTTGTTTTATGTATCTTTTTGTTTTGTAGCATTACTGCCGCAGGAGAGTCTTTGCCTGATT
TAAAAATTGAGAAGCTTGAAGACGGTGTTTATGTTCATACATCGTTTGAAGAAGTTAACGGTTGGGGTGTTGTTACTAAACACGGTTT
GGTGTTTCTTGTAAACACAGACGCCTATCTGATTGACACTCCATTTGCTGCTAAAGACACTGAAAAGTTAGTAAATTGGTTTGTGGAG
CGCGGTTATAAAATAAAAGGCAGTATTTCCTCACATTTTCATAGCGACAGCTCGGGTGGAATAGAATGGCTTAACTCTCAATCTATTC
CCACGTATGCATCTGAATTAACAAACGAACTTCTTAAAAAGAACGGTAAGGTGCAAGCTAAAAACTCATTTAGCGGAGTTAGTTATTG
GCTACTTAAAAATAAAATTGAAATTTTTTATCCGGGCCCTGGGCACACTCAAGATAACGTAGTGGTTTGGTTGCCTGAAAAGAAAATT
TTATTTGGTGGGTGTTTTGTTAAACCGTACGGTCTTGGAAATCTCGATGATGCAAATGTTGAAGCGTGGCCACATTCTGCTGAAATAT
TAATGTCTAGGTATGGTAATGCAAAACTGGTTGTTCCAAGCCATAGTGACGTCGGAGATGCGTCGCTCTTGAAGCTTACATGGGAGC
AGGCTGTTAAAGGGCTAAAAGAAAGTAAAAAACCATCACAGCCAAGTAACTAATTTTCTAACAAGTCGCTCAAGCATCGCGCACTTC
GTGCGCTGGACAGTTTTTAAGTCGCAGTTTTGTGGTTTTGCTGCGCAAAAGTATTCCACAAAACTACAACTTAAAAACTGCCGCTTAG
CTCGGCG 

blaIMP-18 

TTAGAAAAGGGTAAGTATGAAAAAATTATTTGTTTTATGTGTATTCTTCCTTTGCAACATTGCTGCTGCAGATGATTCTTTGCCTGATTT
AAAAATTGAGAAGCTTGAAAAAGGCGTTTATGTTCATACTTCGTTTGAAGAAGTTAAAGGTTGGGGTGTAGTCACAAAACACGGTTTA
GTGGTTCTTGTAAAGAATGATGCTTATCTGATAGATACTCCAATTACCGCTAAAGATACTGAAAAATTAGTTAATTGGTTTATTGAGCA
CGGCTATAGAATCAAAGGCAGTATTTCCACACATTTCCATGGCGACAGTACGGCTGGAATAGAGTGGCTTAATTCTCAATCTATCTC
CACGTATGCCTCTGAATTAACAAATGAACTTCTAAAAAAAGACAATAAGGTGCAAGCTACAAATTCTTTTAGTGGAGTTAGTTATTCAC
TTATCAAAAACAAAATTGAAGTTTTCTATCCAGGTCCAGGACACACTCAAGATAACGTAGTGGTTTGGTTACCTGAAAAGAAAATTTTA
TTCGGTGGTTGCTTTGTTAAACCGGACGGTCTTGGAAATTTAGGGGATGCAAATTTAGAAGCTTGGCCAAAGTCCGCTAAAATATTAA
TGTCTAAATATGGTAAAGCAAAACTGGTTGTTTCAAGTCATAGTGAAATTGGAAACGCATCACTCTTGCAGCGCACATGGGAGCAGG
CTGTTAAAGGGTTAAATGAAAGTAAAAAACCGTTACAGCCAAGTAGCTAAATTTCTAACAAGTCGTTGCAGCATCGTTCCGCTTTGCT
CCACTGGACGGGTTTTAAGTCGCATTTTTTGTGGCTCGCTGGCGCTCACTTTATCACAAAAATACAACTTAAAACCCGCCGCTGAAC
TCGGCG 

blaIMP-22 

TTAGAAAAGGGGAAAGTATGAAGAAATTATTTGTTTTATGTGTGTTTTTGTTTTGTAGCATTACTGCCGCAGGAGAGTCTTTGCCCGA
TTTAAAAATTGAAAAGCTTGAAGAAGGTGTTTATGTTCATACATCGTTTGAAGAAGTTAATGGTTGGGGCGTTGTTTCTAAACACGGTT
TGGTTATTCTTGTGAATACTGACGCCTATCTGATTGACACTCCATTCACGGCTAAAGATACTGAAAAGTTAGTCACCTGGTTTGTGGA
GCGCGGCTATAAAATCAAAGGTAGCATTTCCTCACATTTCCATAGCGACAGCACGGGTGGAATAGAGTGGCTTAATTCTCAATCAAT
TCCCACGTATGCATCTGAATTAACAAATGACCTTCTTAAACAAAACGGTAAGGTACAAGCTAAAAACTCATTTAGCGGAGTTAGTTATT
GGTTAGTTAAAAATAAAATTGAAGTTTTCTATCCCGGCCCCGGGCACACTCAAGATAACGTAGTGGTTTGGTTGCCTGAAAAGAAAAT
TTTATTTGGTGGGTGCTTTGTTAAACCGTACGGTCTTGGAAATCTCGATGACGCAAATGTTGTAGCATGGCCACATTCTGCTGAAATA
TTAATGTCTAGGTATGGTAATGCAAAACTGGTTGTTCCAAGCCATAGTGACATCGGAGATGCGTCGCTCTTGAAGCTTACATGGGAG
CAGGCTGTTAAAGGGCTAAAAGAAAGTAAAAAACCATCAGAGCCAAGTAACTAATTTCTAACAAGTCGCTTAAGCATCAGTCGCTAC
GCTCCTTGGACAGCTTTTAAGTCGCAGTTTTGTGGTTTTGCTGCGCAAAAGTATTCCACAAAACCGCAACTTAAAAACTGCCGCTTAG
CTCGGCG 

blaIMP-31 

TTAGAAAATGGTGCGTATGAAAAAAATATTTGTGTTATTTGTATTTTTGTTTTGCAGTATTACTGCCGCCGGAGAGTCTTTGCCTGATA
TAAAAATTGAGAAACTTGACGAAGATGTTTATGTTCATACTTCTTTTGAAAAGATAACCGGCTGGGGTGTTATTACTAAACACGGCTT
GGTGGTTCTTGTAAATACTGATGCCTATATAATTGACACTCCATTTACAGCTAAAGATACTGAAAAATTAGTCCGCTGGTTTGTGGGG
CGTGGTTATAAAATCAAAGGCAGTATTTCCTCACATTTTCATAGCGATAGCGCAGGTGGAATTGAGTGGCTTAATTCTCAATCTATCC
CCACATATGCATCTAAATTAACAAATGAGCTTCTTAAAAAGAACGGTAATGCGCAAGCCGAAAACTCATTTAGTGGCGTTAGCTATTG
GCTAGTTAAACATAAAATTGAAGTTTTCTATCCAGGACCAGGGCACACTCAGGATAATGTAGTGGTTTGGTTGCCTGAAAAGAAAATT
TTATTTGGCGGTTGTTTTATTAAGCCGGACGGTCTTGGTTATTTGGGAGACGCAAATCTAGAAGCATGGCCTAAGTCCGCAGAAACA
TTAATGTCTAAGTATGGTAATGCAAAACTGGTTGTTTCGAGTCATAGTGAAATTGGGGGCGCATCACTATTGAAGCGCACTTGGGAG
CAGGCTGTTAAGGGGCTAAAAGAAAGTAAAAACCATCACAGCCCCAAATAACTAATTTTCTAACAAGTCTCTCAAGCGGGACTGCCC
CAACCGGCGCTGCTTTCAGCATAGCGCCGGTTGCGTACGTTTCGCGCTGCGCGCTCCACTCAGCCCCTTAGCTCTGCG 

blaOXA-1 

TTGGGCGAACCCGGAGCCTCATTAATTGTTAGCCGTTAAAATTAAGCCCTTTACCAAACCAATACTTATTATGAAAAACACAATACATA
TCAACTTCGCTATTTTTTTAATAATTGCAAATATTATCTACAGCAGCGCCAGTGCATCAACAGATATCTCTACTGTTGCATCTCCATTAT
TTGAAGGAACTGAAGGTTGTTTTTTACTTTACGATGCATCCACAAACGCTGAAATTGCTCAATTCAATAAAGCAAAGTGTGCAACGCA
AATGGCACCAGATTCAACTTTCAAGATCGCATTATCACTTATGGCATTTGATGCGGAAATAATAGATCAGAAAACCATATTCAAATGG
GATAAAACCCCCAAAGGAATGGAGATCTGGAACAGCAATCATACACCAAAGACGTGGATGCAATTTTCTGTTGTTTGGGTTTCGCAA
GAAATAACCCAAAAAATTGGATTAAATAAAATCAAGAATTATCTCAAAGATTTTGATTATGGAAATCAAGACTTCTCTGGAGATAAAGA
AAGAAACAACGGATTAACAGAAGCATGGCTCGAAAGTAGCTTAAAAATTTCACCAGAAGAACAAATTCAATTCCTGCGTAAAATTATT
AATCACAATCTCCCAGTTAAAAACTCAGCCATAGAAAACACCATAGAGAACATGTATCTACAAGATCTGGATAATAGTACAAAACTGT



Supplementary Material 

  174 

ATGGGAAAACTGGTGCAGGATTCACAGCAAATAGAACCTTACAAAACGGATGGTTTGAAGGGTTTATTATAAGCAAATCAGGACATA
AATATGTTTTTGTGTCCGCACTTACAGGAAACTTGGGGTCGAATTTAACATCAAGCATAAAAGCCAAGAAAAATGCGATCACCATTCT
AAACACACTAAATTTATAAAAAATCTAATGGCAAAATCGCCCAACCCTTCAATCAAGTCGGGACGGCCAAAAGCAAGCTTTTGGCTCC
CCTCGCTGGCGCTCGGCGCCCCTTATTTCAAACG 

blaOXA-2 

TTGGGCATTAAGGAAAAGTTAATGGCAATCCGAATCTTCGCGATACTTTTCTCCATTTTTTCTCTTGCCACTTTCGCGCATGCGCAAG
AAGGCACGCTAGAACGTTCTGACTGGAGGAAGTTTTTCAGCGAATTTCAAGCCAAAGGCACGATAGTTGTGGCAGACGAACGCCAA
GCGGATCGTGCCATGTTGGTTTTTGATCCTGTGCGATCGAAGAAACGCTACTCGCCTGCATCGACATTCAAGATACCTCATACACTT
TTTGCACTTGATGCAGGCGCTGTTCGTGATGAGTTCCAGATTTTTCGATGGGACGGCGTTAACAGGGGCTTTGCAGGCCACAATCA
AGACCAAGATTTGCGATCAGCAATGCGGAATTCTACTGTTTGGGTGTATGAGCTATTTGCAAAGGAAATTGGTGATGACAAAGCTCG
GCGCTATTTGAAGAAAATCGACTATGGCAACGCCGATCCTTCGACAAGTAATGGCGATTACTGGATAGAAGGCAGCCTTGCAATCTC
GGCGCAGGAGCAAATTGCATTTCTCAGGAAGCTCTATCGTAACGAGCTGCCCTTTCGGGTAGAACATCAGCGCTTGGTCAAGGATC
TCATGATTGTGGAAGCCGGTCGCAACTGGATACTGCGTGCAAAGACGGGCTGGGAAGGCCGTATGGGTTGGTGGGTAGGATGGGT
TGAGTGGCCGACTGGCTCCGTATTCTTCGCACTGAATATTGATACGCCAAACAGAATGGATGATCTTTTCAAGAGGGAGGCAATCGT
GCGGGCAATCCTTCGCTCTATTGAAGCGTTACCGCCCAACCCGGCAGTCAACTCGGACGCTGCGCGATAAAACCGCGCAGCGCCG
GTTACTTCAACG 

blaOXA-5 

TTAGCCACCAAGGTACCATGAAAACCATAGCCGCATATTTAGTTCTAGTTTTTTATGCAAGCACCGCGCTCTCAGAGTCTATTTCTGA
AAATTTGGCGTGGAATAAAGAATTTTCTAGTGAATCCGTACATGGCGTTTTTGTACTTTGTAAAAGTAGTAGCAATTCCTGTACTACAA
ATAATGCGGCACGTGCATCTACAGCCTATATTCCAGCATCAACATTCAAAATTCCTAATGCTCTAATAGGTCTTGAAACCGGCGCCAT
AAAAGATGAACGGCAGGTTTTCAAATGGGACGGCAAGCCCAGAGCCATGAAGCAATGGGAAAAAGACTTAAAGCTAAGGGGCGCTA
TACAGGTTTCTGCTGTTCCGGTATTTCAACAAATTGCCAGAGAAGTTGGCGAAATAAGAATGCAAAAATACCTTAACCTGTTTTCATA
CGGCAACGCCAATATAGGGGGAGGCATTGACAAATTCTGGCTAGAAGGTCAGCTTAGAATCTCAGCATTCAATCAAGTTAAATTTTT
AGAGTCGCTCTACCTGAATAATTTGCCAGCATCAAAAGCAAACCAACTAATAGTAAAAGAGGCAATAGTTACAGAAGCAACTCCAGA
ATATATAGTTCATTCAAAAACTGGGTATTCCGGTGTTGGCACAGAATCAAGTCCTGGTGTCGCTTGGTGGGTTGGTTGGGTAGAGAA
AGGAACTGAGGTTTACTTTTTTGCTTTTAACATGGACATAGACAATGAGAGTAAATTGCCGTCAAGAAAATCCATTTCAACGAAAATCA
TGGCAAGTGAAGGCATCATCATTGGTGGCTAACAAGGCGCTCAAGGCTCGGTTCGCCACTGGACGTCTCCAATCGGCGCATCGTTC
GCATTTTACGCGCCGCTTGTAGCCGCCCCTTAGCTTTGCG 

blaOXA-9 

TTATGCACCTATTAAGCGCACAGCGGAGCAATGAAGGATACCTTGATGAAAAAAATTTTGCTGCTGCATATGTTGGTGTTCGTTTCCG
CCACTCTCCCAATCAGTTCCGTGGCTTCTGATGAGGTTGAAACGCTTAAATGCACCATCATCGCAGACGCCATTACCGGAAATACCT
TATATGAGACCGGAGAATGTGCCCGTCGTGTGTCTCCGTGCTCGTCTTTTAAACTTCCATTGGCAATCATGGGGTTTGATAGTGGAA
TCTTGCAGTCGCCAAAATCACCTACGTGGGAATTGAAGCCGGAATACAACCCGTCTCCGAGAGATCGCACATACAAACAAGTCTATC
CGGCGCTATGGCAAAGCGACTCTGTTGTCTGGTTCTCGCAGCAATTAACAAGCCGTCTGGGAGTTGATCGGTTCACGGAATACGTA
AAGAAATTTGAGTACGGTAATCAAGATGTTTCCGGTGACTCGGGGAAGCATAACGGCTTGACCCAGTCATGGCTGATGTCGTCGCT
CACCATATCTCCCAAGGAGCAAATTCAGTTTCTTCTACGCTTTGTCGCGCATAAGCTGCCTGTATCCGAAGCGGCTTATGACATGGC
GTATGCCACAATCCCGCAGTACCAGGCAGCCGAAGGATGGGCTGTACATGGAAAAAGCGGCAGCGGCTGGCTTCGGGACAATAAC
GGCAAGATAAATGAAAGTCGTCCGCAGGGCTGGTTCGTGGGCTGGGCTGAAAAAAACGGACGGCAAGTTGTTTTCGCCCGATTGG
AAATAGGAAAGGAAAAGTCCGATATTCCCGGCGGGTCTAAAGCACGAGAGGATATTCTCGTGGAATTACCCGTGTTGATGGGTAAC
AAATGATATGTGGCGTCATCGAGAGCAGATGCATAACCCTGCGCTCGAGCGGACCTCGCGCATAAAGCCGCGCGAGTCCGCTCAC
CTTGAACG 

blaOXA-10 

TTAGCCACCAAGAAGGTGCCATGAAAACATTTGCCGCATATGTAATTATCGCGTGTCTTTCGAGTACGGCATTAGCTGGTTCAATTAC
AGAAAATACGTCTTGGAACAAAGAGTTCTCTGCCGAAGCCGTCAATGGTGTCTTCGTGCTTTGTAAAAGTAGCAGTAAATCCTGCGC
TACCAATGACTTAGCTCGTGCATCAAAGGAATATCTTCCAGCATCAACATTTAAGATCCCCAACGCAATTATCGGCCTAGAAACTGGT
GTCATAAAGAATGAGCATCAGGTTTTCAAATGGGACGGAAAGCCAAGAGCCATGAAGCAATGGGAAAGAGACTTGACCTTAAGAGG
GGCAATACAAGTTTCAGCTGTTCCCGTATTTCAACAAATCGCCAGAGAAGTTGGCGAAGTAAGAATGCAGAAATACCTTAAAAAATTT
TCCTATGGCAACCAGAATATCAGTGGTGGCATTGACAAATTCTGGTTGGAAGGCCAGCTTAGAATTTCCGCAGTTAATCAAGTGGAG
TTTCTAGAGTCTCTATATTTAAATAAATTGTCAGCATCTAAAGAAAACCAGCTAATAGTAAAAGAGGCTTTGGTAACGGAGGCGGCAC
CTGAATATCTAGTGCATTCAAAAACTGGTTTTTCTGGTGTGGGAACTGAGTCAAATCCTGGTGTCGCATGGTGGGTTGGGTGGGTTG
AGAAGGAGACAGAGGTTTACTTTTTCGCCTTTAACATGGATATAGACAACGAAAGTAAGTTGCCGCTAAGAAAATCCATTCCCACCAA
AATCATGGAAAGTGAGGGCATCATTGGTGGCTAACAAGTCGCTCAAGGTCGCTCCCTGCGGTCGCTGGACAGTCCCAGTCGGCGC
ATGCTTCGCATTTTATGCGCCGCCTGTGCCTGCCCCTTAGCTCCAACG 

blaOXA-20 

TTAGGCACCAAAGGAGGCTCCTTGATAATCCGATTTCTAGCACTGCTTTTCTCAGCTGTTGTACTTGTCTCTCTTGGTCATGCACAAG
AAAAAACGCATGAGAGCTCTAATTGGGGGAAATACTTTAGTGATTTCAACGCTAAAGGTACAATAGTTGTAGTAGATGAACGCACAAA
CGGTAATTCCACATCGGTTTATAATGAATCCCGGGCTCAGCAGCGCTATTCGCCTGCGTCCACATTCAAGATTCCGCATACCCTTTT
TGCGCTGGATGCAGGGGCGGTTCGCGATGAGTTTCATGTTTTTCGATGGGACGGCGCTAAAAGAAGCTTTGCAGGTCACAATCAAG
ACCAAAACCTACGATCGGCAATGCGCAATTCTACCGTTTGGGTCTATCAACTATTCGCAAAAGAAATAGGCGAAAACAAAGCACGAA
GCTACCTAGAAAAATTAAACTACGGCAATGCAGACCCCTCGACCAAGAGCGGTGACTACTGGATAGATGGAAATCTTGCAATTTCAG
CAAATGAACAAATTTCCATCCTAAAGAAGCTTTATCGAAATGAGCTTCCTTTTAGGGTAGAGCACCAACGCTTGGTTAAAGACTTGAT
GATTGTCGAAGCCAAACGCGATTGGATACTACGTGCCAAAACAGGCTGGGATGGTCAAATGGGTTGGTGGGTCGGTTGGGTAGAG
TGGCCTACAGGCCCAGTATTTTTTGCGTTAAATATCGACACGCCAAACAGGATGGAAGACCTTCATAAACGAGAGGCAATTGCGCGT
GCTATTCTTCAATCCGTCAATGCTTTGCCACCCAACTAGCAGCCCAAACCCCCTGTTGTGCCTAACAAGGCGCTCAAGTCGGACAGC
CCAAACCGGCGCATGCTTCGCATTATGCGCGCCGGTTCGGTACGTTGCGCGCTTCGCGCTCCACTCTGCCGCTTAGCTTGGCG 

blaOXA-21 

TTGGGCGTCAAGGAAAACTTAATGGCAATCCGAATCTTCGCAATACTTTTCTCCACTTTTGTTTTTGGCACGTTCGCGCATGCACAAG
AAGGCATGCGCGAACGTTCTGACTGGCGGAAGTTTTTCAGCGAATTTCAAGCCAAAGGCACGATAGTTGTGGCAGACGAACGCCAA
ACAGATCGTGTCATATTGGTTTTTGATCAGGTGCGGTCAGAGAAACGCTACTCGCCGGCCTCGACATTCAAGATTCCACATACACTT
TTTGCACTTGACGCAGGCGCTGCACGTGATGAGTTTCAAGTTTTCCGATGGGACGGCATCAAAAGAAGCTTTGCAGCTCACAACCAA
GACCAAGACTTGCGATCAGCAATGCGGAATTCTACTGTCTGGATTTATGAGCTATTTGCAAAAGAGATCGGTGAAGACAAGGCTCGA
CGCTATTTGAAGCAAATCGACTATGGCAACGCCGATCCTTCGACAAGTAATGGCGATTACTGGATAGATGGCAATCTTGCTATCGCG
GCACAAGAACAGATTGCATTTCTCAGGAAGCTCTATCATAACGAGTTGCCCTTTCGGGTAGAACATCAGCGCTTGGTCAAGGACCTC
ATGATTGTGGAAGCCGGTCGCAACTGGATACTGCGCGCAAAGACGGGCTGGGAAGGCCGCATGGGTTGGTGGGTAGGATGGGTT
GAGTGGCCGACTGGCCCCGTATTCTTCGCACTGAATATTGATACGCCAAACAGGATGGATGACCTTTTCAAAAGGGAGGCAATAGT
GCGGGCAATCCTTCGCTCTATCGAAGCGTTGCCGCCCAACCCGGCAGTCAACTCGGACGCAGCGCGATAAAGCCGCGCAGCGCC
GGTTACTTCTACG 

blaOXA-46 

TTGGGCATCAAAGGAAATTTAATGGCAATCCGATTCTTCACCATACTGCTATCCACCTTCTTTCTTACCTCATTCGTGTATGCGCAAG
AACATGTGGTAATCCGTTCGGACTGGAAAAAGTTCTTCAGCGACCTCCAGGCCGAAGGTGCAATCGTTATTGCAGACGAACGTCAA
GCGAAGCATACTTTATCGGTTTTTGATCAAGAGCGAGCGGCAAAGCGTTACTCGCCAGCTTCAACCTTCAAGATACCCCACACACTT
TTTGCACTTGATGCAGACGCCGTTCGTGATGAGTTCCAGGTTTTTCGATGGGACGGCGTTAACCGAAGCTTTGCAGGTCACAATCAA
GACCAAGATTTGCGATCAGCGATGCGAAATTCTACGGTTTGGGTTTATGAGCTGTTTGCAAAAGATATCGGAGAGGACAAAGCAAGA
CGTTATTTAAAGCAAATTGATTATGGCAACGTCGATCCTTCGACAATCAAGGGCGATTACTGGATAGATGGAAATCTTAAAATCTCAG
CGCACGAACAGATTTTGTTTCTCAGAAAACTCTATCGAAATCAGTTACCATTTAAGGTGGAGCACCAGCGCTTGGTGAAAGATCTCAT
GATTACGGAAGCCGGGCGCAGTTGGATACTACGCGCAAAGACCGGCTGGGAAGGCAGGTTTGGCTGGTGGGTAGGGTGGATTGA
ATGGCCAACAGGCCCCGTATTCTTTGCGCTGAATATTGATACGCCAAACAGAACGGACGATCTTTTCAAAAGAGAGGCCATCGCAC
GGGCAATCCTTCGTTCTATTGACGCATTGCCACCCAACTAACCAATCCAGCCGACGCCTTCGACGCGGCTGATTTCAAACG 

blaOXA-53 
TTGGGCATCAAGGAAAAATTAATGGCAATCCAAATCTTCGCAATACTTTTCTCCACTTTTGTTCTTGCCACTTTTGCACATGCGCAAGA
TGGCACGCTGGAACGTTCTGACTGGGGGAAATTTTTCAGCGATTTTCAGGCCAAAGGTACGATAGTTGTGGCAGACGAACGCCAAG
CGGATCATGCGATATTGGTTTTTGATCAAGCACGGTCAATGAAACGCTACTCGCCTGCGTCGACATTCAAGATTCCACATACACTTTT
TGCACTTGATGCAGGCGCCGTTCGCGATGAGTTTCAGATTTTCCGCTGGGACGGCGTCAAAAGGAGCTTTGCAGGTCACAATAAAG
ACCAAGATTTGCGATCAGCAATGCGAAATTCTACTGTCTGGGTTTATGAGCTATTTGCAAAGGAAATCGGTGATGGCAAGGCTCGAC



Supplementary Material 

 175 

GCTATTTGAAGCAAATCGGCTATGGCAACGCCGATCCTTCGACAAGTCATGGCGATTACTGGATAGAAGGCAGCCTTGCAATCTCA
GCACAGGAACAGATCGCGTTTCTCAGAAAGCTCTATCAAAACGATCTGCCCTTTAGGGTGGAACATCAGCGCTTGGTCAAGGATCT
GATGATTGTGGAAGCGGGACGCAACTGGATTCTGCGCGCGAAGACGGGCTGGGAAGGCAGCATGGGTTGGTGGGTGGGGTGGGT
TGAATGGCCAACCGGTCCCGTATTCTTTGCCTTGAATATCGATACGCCAAACAGAATGGACGATCTTTTCAAGAGGGAAGCAATAGC
GCGAGCGATACTTCTCTCTATCGAAGCGTTGCCGCCCAACCCGGCAGTCCACTCGGACGCTGCGCGATGAGGCAGCGCAGCGCC
GGTAACTTCTACG 

blaOXA-118 

TTGGGCGTCAAAGGAAACTTAATGGCAATCCGATTCCTCACCATACTGCTATCTACTTTTTTTCTTACCTCATTCGTGCATGCGCAAG
AACACGTGCTAGAGCGTTCTGACTGGAAGAAGTTCTTCAGCGACCTCCGGGCCGAAGGTGCAATCGTTATTTCAGACGAACGTCAA
GCGGAGCATGCTTTATTGGTTTTTGGTCAAGAGCGAGCAGCAAAGCGTTACTCGCCTGCTTCAACCTTCAAGCTTCCACACACACTT
TTTGCACTCGATGCAGACGCCGTTCGTGATGAGTTCCAGGTTTTTCGATGGGACGGCGTTAAACGGAGCTTTGCGGGCCATAATCA
AGACCAAGACTTGCGATCAGCGATGCGAAATTCTGCGGTCTGGGTTTATGAGCTATTTGCAAAAGAGATCGGAAAGGACAAAGCAA
GACACTATTTAAAGCAAATTGATTATGGCAACGCCGACCCTTCGACAATCAAGGGCGATTACTGGATAGATGGCAATCTTGAAATCT
CAGCGCACGAACAGATTTCGTTTCTCAGAAAACTCTATCGAAATCAGCTGCCATTTCAGGTGGAACATCAGCGCTTGGTCAAAGATC
TCATGATTACGGAAGCCGGGCGCAACTGGATACTACGCGCAAAGACCGGCTGGGAAGGCAGGTTTGGCTGGTGGGTAGGGTGGG
TGGAGTGGCCAACCGGTCCCGTATTCTTCGCGCTGAATATTGATACGCCAAACAGAACGGATGATCTTTTCAAAAGAGAGGCAATCG
CGCGGGCAATCCTTCGCTCTATCGACGCATTGCCGCCCAACTAATCAATCCAGCGGACGCCTTCGGCGCCGCTGATTTCAACG 

blaOXA-129 

TTAGCCACCAAGGTACCATGAAAACCATAGCCGCATATTTAGTTCTAGTATTTTTTGCAGGCACTGCACTTTCAGAGTCTATTTCTGA
AAATTTAGCTTGGAATAAAGAATTTTCCAGTGAATCAGTGCATGGTGTTTTTGTACTTTGTAAAAGCAGTAGTAATTCCTGTACAACAA
ATAATGCAACACGTGCATCTACGGCCTATATTCCAGCATCAACATTCAAAATTCCCAATGCTCTCATAGGCCTTGAAACCGGCGCCAT
AAAAGATGCGCGGCAGGTTTTCAAATGGGACGGCAAGCCCAGAGCCATGAAGCAATGGGAAAAAGACTTAACGCTAAGGGGCGCT
ATACAAGTTTCTGCTGTTCCGGTATTTCAACAAATTGCCAGAGACATTGGCAAAAAAAGAATGCAAAAATACCTTAACCTTTTTTCATA
TGGCAACGCCAATATAGGCGGAGGCATTGACAAATTTTGGCTAGAAGGTCAGCTTAGAATCTCAGCAGTCAATCAAGTTAAATTTTTA
GAGTCGCTTTACCTAAATAATTTGCCAGCATCTAAAGCAAACCAACTTATAGTAAAAGAGGCAATAGTTACAGAAGCAACTCCAGAAT
ATATAGTGCATTCAAAAACCGGGTATTCCGGTGTGGGCACAGAATCAAATCCTGGTGTCGCTTGGTGGGTTGGTTGGGTAGAAAAA
GGAACTGAGGTTTACTTTTTTGCATTTAACATGGACATAGACAATGAGAGTAAGTTGCCGTCAAGAAAATCCATTCCAACGAAAATCA
TGGCAAGTGAAGGTATCATCATTGGTGGCTAACAAGGCGCTCAAGGCCGTGGCTGCGCCACTGGACGTCTCCAATCGGCGCATGC
TTCGCATTTTGCGCGCCGCTTGTAGCCGCCCCTTAGCTTTGCG 

blaOXA-198 

TTATGCATAAACACATGAGTAAGCTCTTCATCGCTTTTTTAGCCTTTCTGCTGTCGGTGCCAGCAGCCGCTGAAGACCAGACACTTG
CCGAGCTCTTTGCCCAACAAGGCATTGACGGGACTATAGTGATTTCGTCGCTACACAACGGAAAGACATTTATCCACAACGATCCCC
GCGCAAAACAGAGATTCTCGACAGCATCCACGTTCAAGATACTGAACACGCTGATCTCGCTCGAAGAAAAAGCCATCTCTGGAAAAG
ACGATGTGCTGAAATGGGACGGGCATATTTACGATTTTCCAGATTGGAATCGTGACCAGACGCTAGAAAGTGCGTTCAAGGTTTCCT
GTGTCTGGTGTTATCAGGCGCTTGCACGCCAGGTCGGCGCGGAGAAGTATCGAAATTATTTACGCAAGTCAGTTTACGGAGAATTA
CGCGAGCCTTTTGAGGAAACAACATTCTGGCTTGATGGTTCACTTCAAATCAGCGCAATTGAACAAGTGAATTTCCTCAAGAAAGTTC
ATCTGCGCACTCTCCCATTCAGTGCATCGTCCTACGAAACGCTACGACAAATCATGCTTATCGAGCAAACGCCGGCTTTTACGCTGC
GGGCCAAGACAGGCTGGGCAACAAGAGTAAAGCCGCAAGTTGGCTGGTATGTGGGCCATGTCGAAACTCCAACGGATGTATGGTT
CTTTGCCACGAATATTGAAGTCCGTGACGAAAAAGACTTGCCCTTACGTCAGAAGCTAACGCGAAAAGCATTACAAGCAAAGGGGAT
CATCGAATAATGCATAACATGGCGCTCAAGCGGGACGCGGCAAAATGCCGCCGCGCCCCTTAGCTCTACG 

blaPBL-1 

TTAGGCCGCACAATCAGCACATCATGTGACCTTATGTTTTCTTGAAACAAACATATGCACGTCATATCGCGGTGTTCATAGACAATCA
TTAAAGGAAGGAGATAGGAATGGTTACAAGGCGAGATTTTTTGATTGCTTCAGCGGCAACGGCCGCTCTTCTAACTTTGCCAGCTAC
TGCAAAAGTTCCCTCGCGTCGCTTTGACGCATCAGCGAAGCTTCAGTCGCTTGAAGCGGGACAGGCGCGCCTTGGTGTTTGCTTCC
TCGATACGGTTACTGGTGAAGTCAGCGGTAACCGTATCGAGGAGCGCTTTGCAATGTGCTCGACGGTCAAGCTGGCAATAGTTGCC
GCCTGCTTGCGCGAGGCAGATAAGGCGCGCTTGAATCTCGAGGAGATACTGACCTATTCAGAGGCGGATCTTCTTCCTTGGGCGC
CAGTGACGCGCAAGAATCTCGCCAAAGGCGGTCTGAGTATTTCCGCTCTGGCGCAAGCGGCTCAAGAAATGAGCGATGGTGTTGC
CGCTAACCTTTTAATCAAACGCCTAGGCGGTCCTGCCGCCGTCACCGCGAAGTTCCGAGAAATGGGCGATTCGGTGACCCGCCTTG
ATCGTTATGAGCCAGACTTAGGGTTAGTTCTTTCAGCCGATGTTCGCGACACCACCACGCCCCTTGCTTATGCGCAACTTGTCCGTC
GAATCACGACTGGCCGTGTTTTGTCGCACGGATCGCGGGAACAACTTTTAGAGTGGATGCGAAATACAGTCACAGGCGCCAGTCGG
CTGCGAGCCGGCCTCCCCACTGAATGGCGTACTGGGAACAAGACAGGGACTGGGCGCGACGAAGGGACGACCAACAAGTGCAAT
GACGTCGCTATTACTTTTCCGCCAAGCAAGAACCCAATCATCATTGCAGCCTATTTCGACAGCGGCGAATACACAGAAAAGGTAGAG
GCGAGGCATGAGGCTGTCCTCGCTGAAGTGGGAAAGATTGCTGCCGAATGGGGCGAGAGTTAGTGTGCGAACTACACAATCACTTT
ATTTAAAAAATTAGAATCTAGGTAGGTACTTACGTCTTTTGAAGAATTTCTAATTTTTGTCATGAAAAAGGGATTAAAAAAAAGAAGCCT
AACAAATCGCTCAAGCACCGCACACTTCGTGTGCGGGACAGTTTTTAAGTCGCAGTTTTGTGGTTTTGCTGCGCAAAATTATTCCAC
AAAACCGCAACTTAAAAACTGCCGCTTAGCTCGGCG 

blaSIM-1 

TTAGAGGTAGTTAAAAATGAGAACTTTATTGATTTTATGTTTATTCGGCACTTTAAATACCGCGTTTGCGGAAGAAGCCCAGCCAGAT
TTAAAAATTGAAAAAATAGAAGAAGGGATCTATCTTCATACATCTTTTCAAGAGTACAAGGGATTCGGCATCGTTAAAAAACAAGGCTT
AGTAGTTCTTGACAATCACAAGGCATATCTCATCGACACTCCAGCTTCCGCAGGAGATACTGAAAAGCTAGTAAACTGGCTCGAAAA
AAATGATTTCACTGTCAATGGAAGCATTTCAACACATTTCCACGACGACAGTACTGCTGGGATAGAGTGGCTTAATACAAAGTCCATC
CCCACATATGCATCTAAATTGACAAATGAATTGCTAAATAAAAATGGCAAAACTCAAGCCAAGCACTCTTTTGATAAAGAGAGCTTTTG
GTTGGTCAAAAATAAAATTGAAATTTTTTATCCAGGCCCAGGACACACTCAAGATAACGAAGTTGTCTGGATACCTAATAAAAAAATC
CTATTCGGGGGCTGTTTTATAAAACCGAATGGCCTTGGCAATCTAAGTGACGCAAATTTGGAAGCTTGGCCAGGCTCCGCAAAAAAA
ATGATATCAAAATACAGTAAGGCAAAACTTGTTATCCCAAGCCACAGTGAAATCGGAGACGCATCACTATTGAAACTCACATGGGAA
CAGGCCATTAAAGGTTTAAATGAAAGCAAATCAAAACCGCCGCTCATTAATTAACCCCCCCAAGGTCATTACGAAAACTCTGAGTGC
CTCTAACAATGCGCTAAACACCGCTCGCTTCGCTCACTGGACTCGCAAAAGCTGCGCTTTTGCTCGCCTGTTAGCTTAATCG 

blaTMB-1 

TTAGAGGAATAATGGAATGCGACCATTTTTATTTTTAATAATTTTTATCAGTCATTTCGCTTTTGCCAACGAAGAAATACCCGGATTGG
AAGTTGAGGAAATTGACAACGGCGTTTTTTTGCACAAGTCATACAGCCGGGTGGAAGGCTGGGGCCTGGTAAGTTCAAACGGACTT
GTTGTCATCAGCGGCGGAAAAGCATTCATTATTGACACTCCATGGTCGGAATCAGATACAGAAAAGCTTGTAGATTGGATACGATCA
AAAAAGTATGAGCTGGCGGGAAGCATTTCTACACATTCACACGAAGACAAGACTGCCGGTATAAAATGGCTAAACGGCAAATCCATT
ACTACATATGCCTCAGCGCTGACTAATGAAATTCTAAAAAGAGAGGGTAAGGAGCAGGCAAGGAGCTCATTCAAAGGTAATGAATTT
TCGCTGATGGACGGTTTTCTAGAAGTCTATTATCCCGGAGGCGGCCATACTATTGATAACTTAGTGGTATGGATCCCTAGTTCAAAAA
TATTGTATGGCGGCTGTTTCATACGTAGCTTGGAATCCAGTGGGCTAGGTTACACTGGTGAAGCTAAAATTGATCAGTGGCCACAAT
CCGCTAGAAATACAATTTCGAAGTATCCTGAAGCTAAGATTGTGGTGCCTGGTCATGGAAAAATTGGCGATTTCGAGTTGTTAAAACA
TACCAAGGTCCTTGCAGAAAAGGCCTCTAACAAGGCCAATCACGGCGACCGCTGACGCGGCGCGTGTTGGCATAG 

blaVEB-1 

TTAGCGGTAATTTAACCAGATAGGAGTACAGACATATGAAAATCGTAAAAAGGATATTATTAGTATTGTTAAGTTTATTTTTTACAATTG
TGTATTCAAATGCTCAAACTGACAACTTAACTTTGAAAATTGAGAATGTTTTAAAGGCAAAAAATGCCAGAATAGGAGTAGCAATATTC
AACAGCAATGAGAAGGATACTTTGAAGATTAATAACGACTTCCATTTCCCGATGCAAAGCGTTATGAAATTTCCGATTGCTTTAGCCG
TTTTGTCTGAGATAGATAAAGGGAATCTTTCTTTTGAACAAAAAATAGAGATTACCCCTCAAGACCTTTTGCCTAAAACGTGGAGTCC
GATTAAAGAGGAATTCCCTAATGGAACAACTTTGACGATTGAACAAATACTAAATTATACAGTATCAGAGAGCGACAATATTGGTTGT
GATATTTTGCTAAAATTAATCGGAGGAACTGATTCTGTTCAAAAATTCTTGAATGCTAATCATTTCACTGATATTTCAATCAAAGCAAAC
GAAGAACAAATGCACAAGGATTGGAATACCCAATATCAAAATTGGGCAACCCCAACAGCGATGAACAAACTGTTAATAGATACTTATA
ATAATAAGAACCAATTACTTTCTAAAAAAAGTTATGATTTTATTTGGAAAATTATGAGAGAAACAACAACAGGAAGTAACCGATTAAAA
GGACAATTACCAAAGAATACAATTGTTGCTCATAAAACAGGGACTTCCGGAATAAATAATGGAATTGCAGCAGCCACTAATGATGTTG
GGGTAATTACTTTACCGAATGGACAATTAATTTTTATAAGCGTATTTGTTGCAGAGTCCAAAGAAACTTCGGAAATTAATGAAAAGATT
ATTTCAGACATTGCAAAAATAACGTGGAATTACTATTTGAATAAATAAAAAACTACCGCTAACACTGGCTCATAGGCAATGGCGGGTT
GAAGTGCAATTTGCAAAGTCGGTAGCCCGCCCGAGCGTTTTCTCGGTTTGACAGGAAAGGCTCACGCAAACCGCCACTGCCCATAG
CCCAAACCG 

blaVIM-1 TTATGCCGCACCCACCCCTATGGAGTCTTGATGTTAAAAGTTATTAGTAGTTTATTGGTCTACATGACCGCGTCTGTCATGGCTGTCG
CAAGTCCGTTAGCCCATTCCGGGGAGCCGAGTGGTGAGTATCCGACAGTCAACGAAATTCCGGTCGGAGAGGTCCGACTTTACCA



Supplementary Material 

  176 

GATTGCCGATGGTGTTTGGTCGCATATCGCAACGCAGTCGTTTGATGGCGCGGTCTACCCGTCCAATGGTCTCATTGTCCGTGATG
GTGATGAGTTGCTTTTGATTGATACAGCGTGGGGTGCGAAAAACACAGCGGCACTTCTCGCGGAGATTGAAAAGCAAATTGGACTT
CCCGTAACGCGTGCAGTCTCCACGCACTTTCATGACGACCGCGTCGGCGGCGTTGATGTCCTTCGGGCGGCTGGGGTGGCAACGT
ACGCATCACCGTCGACACGCCGGCTAGCCGAGGCAGAGGGGAACGAGATTCCCACGCATTCTCTAGAAGGACTCTCATCGAGCGG
GGACGCAGTGCGCTTCGGTCCAGTAGAGCTCTTCTATCCTGGTGCTGCGCATTCGACCGACAATCTGGTTGTATACGTCCCGTCAG
CGAACGTGCTATACGGTGGTTGTGCCGTTCATGAGTTGTCAAGCACGTCTGCGGGGAACGTGGCCGATGCCGATCTGGCTGAATG
GCCCACCTCCGTTGAGCGGATTCAAAAACACTACCCGGAAGCAGAGGTCGTCATTCCCGGGCACGGTCTACCGGGCGGTCTAGAC
TTGCTCCAGCACACAGCGAACGTTGTCAAAGCACACAAAAATCGCTCAGTCGCCGAGTAGCAGATGCGGCATAACAAATCGTTGGA
GCGGGACTTTTGCTACGCAGGCTGCGCCTACTCCGCAAAAGCCCCTCAACTCAGGCG 

blaVIM-2 

TTATGCCGCACTCACCCCCATGGAGTTTTGATGTTCAAACTTTTGAGTAAGTTATTGGTCTATTTGACCGCGTCTATCATGGCTATTG
CGAGTCCGCTCGCTTTTTCCGTAGATTCTAGCGGTGAGTATCCGACAGTCAGCGAAATTCCGGTCGGGGAGGTCCGGCTTTACCAG
ATTGCCGATGGTGTTTGGTCGCATATCGCAACGCAGTCGTTTGATGGCGCAGTCTACCCGTCCAATGGTCTCATTGTCCGTGATGGT
GATGAGTTGCTTTTGATTGATACAGCGTGGGGTGCGAAAAACACAGCGGCACTTCTCGCGGAGATTGAGAAGCAAATTGGACTTCC
TGTAACGCGTGCAGTCTCCACGCACTTTCATGACGACCGCGTCGGCGGCGTTGATGTCCTTCGGGCGGCTGGGGTGGCAACGTAC
GCATCACCGTCGACACGCCGGCTAGCCGAGGTAGAGGGGAACGAGATTCCCACGCACTCTCTAGAAGGACTCTCATCGAGCGGGG
ACGCAGTGCGCTTCGGTCCAGTAGAACTCTTCTATCCTGGTGCTGCGCATTCGACCGACAACTTAGTTGTGTACGTCCCGTCTGCG
AGTGTGCTCTATGGTGGTTGTGCGATTTATGAGTTGTCACGCACGTCTGCGGGGAACGTGGCCGATGCCGATCTGGCTGAATGGCC
CACCTCCATTGAGCGGATTCAACAACACTACCCGGAAGCACAGTTCGTCATTCCGGGGCACGGCCTGCCGGGCGGTCTAGACTTG
CTCAAGCACACAACGAATGTTGTAAAAGCGCACACAAATCGCTCAGTCGTTGAGTAGCAGGCAGATGCGGCATAACATGAAGTTGC
AGCCGACCATCACTCCGCTGCGCTCCGTTCTGGCGGCTGAACTTCGGCG 

blaVIM-7 

TTATCGCAGTCGGCCCCCGAGGAGTATTGATGTTTCAAATTCGCAGCTTTCTGGTTGGTATCAGTGCATTCGTCATGGCCGTACTTG
GATCAGCAGCATATTCCGCACAGCCTGGCGGTGAATATCCGACAGTAGATGACATACCGGTAGGGGAAGTTCGGCTGTACAAGATT
GGCGATGGCGTTTGGTCGCATATCGCAACTCAGAAACTCGGTGACACGGTGTACTCGTCTAATGGACTTATCGTCCGCGATGCTGA
TGAGTTGCTTCTTATTGATACAGCGTGGGGGGCGAAGAACACGGTAGCCCTTCTCGCGGAGATTGAAAAGCAAATTGGACTTCCAG
TAACGCGCTCAATTTCTACGCACTTCCATGACGATCGAGTCGGTGGAGTTGATGTCCTCCGGGCGGCTGGAGTGGCAACGTACACC
TCACCCTTGACACGCCAGCTGGCCGAAGCGGCGGGAAACGAGGTGCCTGCGCACTCTCTAAAAGCGCTCTCCTCTAGTGGAGATG
TGGTGCGCTTCGGTCCCGTAGAGGTTTTCTATCCTGGTGCTGCGCATTCGGGCGACAATCTTGTGGTATACGTGCCGGCCGTGCGC
GTACTGTTTGGTGGCTGTGCAGTTCATGAGGCGTCACGCGAATCCGCGGGTAATGTTGCCGATGCCAATTTGGCAGAATGGCCTGC
TACCATTAAACGAATTCAACAGCGGTATCCGGAAGCAGAGGTCGTCATCCCCGGCCACGGTCTACCGGGCGGTCTGGAATTGCTCC
AACACACAACTAACGTTGTCAAAACGCACAAAGTACGCCCGGTGGCCGAGTAACAAATGCGGCATAACAACGCCAAGGAGGGGGA
CGCATTTTCCACTACGCCCTTCGGGCTTCCCTCCAAAAGCGCCCCTCATGGCGGGCG 

blaVIM-13 

TTATGCCGCACCCACCCCTATGGAGTTTTGATGTTAAAAGTTATTAGTAGTTTATTGTTCTACATGACCGCCTCTCTAATGGCTGTAG
CTAGTCCGTTAGCCCATTCCGGGGAGTCGAGAGGTGAGTATCCGACAGTCAGCGAAATTCCGGTCGGAGAAGTTCGGCTGTACCA
GATTGACGATGGTGTTTGGTCGCATATCGCAACGCATACGTTTGATGGCGTGGTGTACCCGTCCAATGGTCTCATTGTCCGTGATG
GCGATGAGTTGCTTTTGATTGATACAGCTTGGGGTACGAAAAACACAGTGGCCCTTCTCGCGGAGATTGAGAAGCAAATTGGACTTC
CCGTAACGCGTTCAGTCTCCACGCACTTTCATGACGACCGCGTCGGCGGAGTTGATGCCCTTAGGGCGGCTGGAGTGGCGACGTA
CGCATCGCCCTCGACACGCCGTCTAGCCGAGGCAGAGGGGAACGAGGTTCCCACACACTCTCTAGAAGGGCTCTCATCGAGTGGG
GACGCAGTGCGTTTCGGTCCAGTAGAGCTCTTCTATCCTGGTGCTGCGCATTCGACCGACAATCTGGTTGTATACGTCCCGTCAGC
GAACGTGCTATACGGTGGTTGTGCCGTTCTGGAATTGTCACGCACATCCGCGGGAAACGTGGCCGATGCCGACCTGGCTGAATGG
CCCGGTTCCGTTGAGCGGATTCAACAACATTACCCAGAAGCAGAGGTGGTCATTCCCGGGCACGGTCTACCGGGCGGTCTAGACT
TGCTCCAGCACACAGCGAACGTTGTCAAAGCACACACAAATCGCTCAGTCGCCGAGTAGCAGATGCGGCATAACAAATCGCTGCAG
TTGACCGCCCAACCCGCTGTCGCGGCTTGGGTTCCCTCCGCGCTTCGCGCTCCGGCGGCAACTGAGCTCAGGCG 

aacA1 
AAC(6´)-Ia 

TTAGGGCGACGCCGCTATTGCGGCGCGAATACAAAGAGGAAGAGATGAATTATCAAATTGTGAATATTGCGGAATGCAGCAATTATC
AGTTAGAAGCAGCAAATATACTAACAGAAGCGTTCAATGATCTTGGTAACAATTCATGGCCAGATATGACGAGTGCAACAAAAGAAG
TAAAAGAATGTATTGAGAGTCCAAACCTTTGTTTCGGTCTGCTAATAAATAACTCCTTAGTTGGCTGGATAGGCTTAAGGCCAATGTA
CAAGGAAACCTGGGAATTGCATCCATTGGTTGTCAGACCAGATTATCAAAATAAAGGTATTGGCAAGATCCTGCTTAAGGAATTAGAA
AACAGAGCTAGAGAGCAAGGTATTATTGGAATCGCTTTAGGAACAGATGATGAATACTATAGAACAAGTCTCTCTTTAATAACTATAA
CAGAAGATAATATATTTGATTCAATAAAAAATATTAAAAATATTAATAAACATCCATATGAGTTTTATCAGAAGAATGGTTATTATATTGT
TGGAATAATTCCAAATGCCAATGGTAAAAACAAACCAGATATTTGGATGTGGAAAAGTTTAATCAAAGAGTAAAACAATGGAACAGAT
TAATAACATAGAACTTGTAGATCCATCTATCTACCCAACAGATGAAATCTTAAAAAGAATACTTGGAAAATCGTTTAGTGTATATTTAAA
ATTACTACGACTCTATGAAAATTATAGTTTAATACCAGAGTGGAAGTATTATAAAGATGGAAAAGCTTGGTTATGTAAAGTTATAAAGG
GAAAGAAAACGATTGTTTGGATGTCTGCATGGAAGAACTACATAAAAGCTACTATATATCTTCCAGAAAAACATATTAACGGTGTATTA
GTATTAGATATTCATGAGATTACAAAGAAAGCATTTATAGAGACAAATAATATTGGAAGATCAAGGCCTTGCATGTTTGAATTAAAAGA
AGAAAATATATTAGAAGATTTCATAAAGGTAATGCAGTTCAAAATGACATTAAAATGATCAACCGAAATTTGCCCTAACAAATGCTTCA
ACCTGACAATTCCTGTTGTCATGGTTTGTGCTGTCGCTTTGCTCAGCACAAACCACGCCAAGCCCTTCGGGCCGGAATTGCAGGTTA
AGCAAATG 

aacA2 
AAC(6´)-Id 

TTAGGCGTCATGATCGAAGCGTGTCACTCCGTCGAATGCCCTGGCTGGCTGCAACTTCGTTTTCTCCTCTGGCCGCAAGATAGCGC
CGACGAACACCTTGCTGAAATGGCAATCTTCGTGGCTGAGCCAAATCGGTTCGCGCAGTTCATCGCTTACGACGAAGCGAACAAAC
CACTAGGCTTCGTTGAGGCTGCGCTCCGATCTGACTACGTAAACGGAACCAATTCGTCCCCAGTAGCGTTTCTTGAAGGGGTCTAT
GTCTTACCGGAAGCACGGCGTCGCGGCATCGCTCATGCCTTGGTCGGTGCAGTCGAGATATGGGCTCGTAATCGGGCCTGTACCG
AGTTCGCATCCGATGCATCAACTGACAACCCGGAAAGCCATAGATTTCACCAGTCACTTGGGTTCAAAGAAACCGAGCGTGTCGTGT
ACTTCAGGAAGATGCTTGCGCCAGAATGACGCCTAACCCTTCCATCGAGGGGATGCCCAAAAGGCTGCGCCTTCTGTGCACCCCTC
ATGTCAAACG 

aacA3 
AAC(6´)-IIa 

TTAGGCAGCACAGAGCGACCATTTCATGTCCGCGAGCACCCCCCCCATAACTCTTCGCCTCATGACCGAGCGCGACCTGCCGATG
CTCCATGATTGGCTCAACCGGCCGCACATCGTTGAGTGGTGGGGTGGTGACGAAGAGCGACCGACTCTTGATGAAGTGCTGGAAC
ACTACCTGCCCAGAGCGATGGCGGAAGAGTCCGTAACACCGTACATCGCAATGCTGGGCGAGGAACCGATCGGCTATGCTCAGTC
GTACGTCGCGCTCGGAAGCGGTGATGGCTGGTGGGAAGATGAAACTGATCCAGGAGTGCGAGGAATAGACCAGTCTCTGGCTGAC
CCGACACAGTTGAACAAAGGCCTAGGAACAAGGCTTGTCCGCGCTCTCGTTGAACTACTGTTCTCGGACCCCACCGTGACGAAGAT
TCAGACCGACCCGACTCCGAACAACCATCGAGCCATACGCTGCTATGAGAAGGCAGGATTCGTGCGGGAGAAGATCATCACCACG
CCTGACGGGCCGGCGGTTTACATGGTTCAAACACGACAAGCCTTCGAGAGAAAGCGCGGTGTTGCCTAACAACTCATTCAAGCCGA
CGCCGCTTCGCGGCGCGGCTTAATTCAGGTG 

aacA4 
AAC(6´)-Ib 

TTAGGCATCACAAAGTACAGCATCGTGACCAACAGCAACGATTCCGTCACACTGCGCCTCATGACTGAGCATGACCTTGCGATGCT
CTATGAGTGGCTAAATCGATCTCATATCGTCGAGTGGTGGGGCGGAGAAGAAGCACGCCCGACACTTGCTGACGTACAGGAACAGT
ACTTGCCAAGCGTTTTAGCGCAAGAGTCCGTCACTCCATACATTGCAATGCTGAATGGAGAGCCGATTGGGTATGCCCAGTCGTAC
GTTGCTCTTGGAAGCGGGGACGGATGGTGGGAAGAAGAAACCGATCCAGGAGTACGCGGAATAGACCAGTTACTGGCGAATGCAT
CACAACTGGGCAAAGGCTTGGGAACCAAGCTGGTTCGAGCTCTGGTTGAGTTGCTGTTCAATGATCCCGAGGTCACCAAGATCCAA
ACGGACCCGTCGCCGAGCAACTTGCGAGCGATCCGATGCTACGAGAAAGCGGGGTTTGAGAGGCAAGGTACCGTAACCACCCCAG
ATGGTCCAGCCGTGTACATGGTTCAAACACGCCAGGCATTCGAGCGAACACGCAGTGATGCCTAACCCTTCCATCGAGGGGGACG
TCCAAGGGCTGGCGCCCTTGGCCGCCCCTCATGTCAAACG 

aacA5 
AAC(6´)-IIb 

TTAGGCAGCACGGAGACACTTCAGCATGCATCCCGGCGTTGTTACTCTGCGTCCGATGACCGAAGACGACATCGGTATGCTTCACG
AATGGTTGAATCGGCCGCACATTGTCGAATGGTGGGGTGGTGAGCGGCCCTCGCTCGAAGAGGTGAAAGAGGACTATCGGCCCAG
CGCGTTGGCCGAAGAAGGAGTGACGCCGTACATCGGTTTGCTTGACGGAACTCCATTCGCGTTCGCACAGTCGTACGTTGCGCTC
GGGTCGGGTGGTGGATGGTGGGAGGAAGAGACCGATCCTGGTGTCCGCGGAATCGATCAATCAATCGCCGATTCCGGGCTTCTCG
GAAGAGGTTACGGCACTCGGCTGGTGCAGGCGCTTGTTGATTTGCTGTTCGCCGACCCGCAGGTATCCAAGGTTCAGACGGACCC
CTCCCCGAACAACATGCGCGCGATACGCTGCTATGAGAAGGCAGGCTTCCGGAAGGTCAAGGTCGTTTCAACACCGGATGGGCCG



Supplementary Material 

 177 

GCCATGTACATGTTGCACGAGCGTCCGTTGGTGAACGGTTTGCGCAGTGCGGCCTAACTTTTCGCTCCAACGGACGCTTGACCCGT
CGGCCCGGCCGCTGCCGCAGCCGGACCGCCGCGTCAACCGCCGCTGACCTCCGGAG 

aacA7 
AAC(6´)-Il 

TTAGGCACCAATGGATAGTTCGCCGCTCGTCAGGCCTGTTGAAACTACCGATTCGGCCAGTTGGCTAAGCATGCGCTGTGAGCTGT
GGCCAGATGGCACATGTCAAGAGCACCAGTCAGAGATCGCAGAATTTCTGTCCGGAAAAGTCGCCCGGCCTGCTGCTGTCCTCATT
GCTGTAGCACCCGACGGAGAAGCACTAGGGTTTGCCGAGCTTTCGATCCGCCCGTATGCGGAGGAGTGCTACTCCGGCAACGTTG
CGTTCTTGGAGGGTTGGTACGTTGTGCCAAGTGCGCGGCGTCAGGGCGTAGGTGTAGCTCTGGTAAAAGCCGCCGAGCATTGGGC
TCGTGGTCGCGGATGCACCGAATTCGCCTCCGACACTCAACTTACCAACAGCGCAAGCACCTCGGCGCACCTGGCGGCTGGATTC
ACGGAGGTTGCTCAAGTACGCTGCTTCCGGAAACCGTTGTGAGGGGCGCCGCGTTGGTGCCTAACAATTCGTTCAAGCCGAACTTG
CTTCGTTACACCAAAGCCATGGCAGAATGAGCTTGCCATGGCTTTGGCTCCACTACGCAAGTCGGCTTAACTCAGGCG 

aacA8 

TTAGGCAGCACAAACTCCGTCCTCATGACATCCAGCATTAGTTTCGTCAAGCTGCGCCTCATGACCGAGCAAGACCTTCCGATGCTC
CATGAGTGGCTAAACCGGCCTCACATCGTTGAGTGGTGGGGCGGAGAAGAAGCACGTCCAACACTTGCTGAAGTGCAGGAGCAAT
ACCTGCCAAGCGCCTTGGCGAGAGAGTCCGTCACTCCCTACATCGCAATGCTGGATGAAGAACCGATTGGGTACGCTCAGTCGTAC
GTTGCACTCGGAAGCGGTGGCGGATGGTGGGAAGACGAAACGGATCCAGGAGTACGCGGAATTGACCAGTCCCTGGCAAATCCAT
CGCAGCTGGGAAAGGGGCTAGGAACCAAGCTTGTTCGCGCGCTCGTTGAGATGCTGTTCAAAGACGCTAAGGTAACCAATATCCAA
ACGGACCCGTCGCCGAACAACTTGCGCGCAATCCGGTGCTACGAGAAGGCGGGGTTTGTGACGCAAAGAACCATAACCACCCCAG
ACGGGCCGGCTGTGTACATGGTTCAAACACGTCAGGCGTTCGAGCAGGCGCGCAGTGCTGCCTAACCCTTCCATCGAGAGGGACG
TCCAAGAGCTATCGCTCTTGGCCGCCCCTCATGTCAAACG 

aacA16 
AAC(6´)-Ip 

TTGGGCTGATTGATTTGTTTGTTCTAGCATTACCTATCTGGAGTTTGTTTTGAACTATTCAATATGCGATATAGCTGAATCAAATGAAT
TAATCCTTGAAGCAGCAAAGATGCTTAAGAAAAGCTTTCTTGATGCTGGAAATGAATCATGGGGAGATATTAAAAATGCTATTGAAGA
AGTTGAAGAATGTATAGAACATCCAAATATATGCTTGGGAATATGTCTGGATGATAAACTGATTGGATGGACCGGATTAAGGCCGAT
GTACGATAAGACCTGGGAACTTCATCCCATGGTTATAAAAACTGAATATCAAGGCAAGGGTTTTGGGAAAGTACTACTAAGAGAACT
AGAGACGAGAGCGAAGAGTAGGGGAATTATCGGAATAGCTCTTGGAACTGATGACGAATATCAGAAAACTAGTTTGTCTATGATTGA
TATAAACGAACGAAATATCTTCGATGAAATCGGGAATATAAAGAACGTTAATAATCATCCATATGAGTTTTATAAGAAATGTGGTTATA
TGATCGTTGGAATAATCCCTAATGCTAATGGAAAAAGAAAACCAGATATATGGATGTGGCAGATATTAGCTAGGAAGAACAGCCCAA
CAATCGCTTCAACCTGACTCAGGGCGCCGTCACGATTTCTGCTAGTTATCCTGGGACGCAGAAATCGCGCCAACGCCCTTCGCAGG
TTAAGCGAAGTG 

aacA17 
AAC(6´)-Iq 

TTGGGCTGATTGATTTGTTTGTTCTAGTATTACCTATCTGGAGTTTGTTTTGGACTATTCAATATGCGATATAGCGGAATCAAATGAAT
TAATCCTTGAAGCAGCAAAAATACTTAGGAAAAGCTTTCTTGATGCTGGAAATGAATCATGGGTAGATATCAAAAAGGCTATTGAAGA
AGTTGAGGATTGTATAGAACACCCAAATCTATGCTTGGGAATATGTCTGGATGATAAACTGATTGGCTGGACCGGATTAAGGCCGAT
GTACGATAAGACCTGGGAACTTCATCCCATGGTTATAAAAACTGAGTATCAATGCAGGGGTATTGGGAAAGTCTTAATAAAAGAACTA
GAGAAGAGAGCGAAGGGTAGGGGAATTATCGGAATAGCTCTTGGAACTGATGATGAATATCAGAAAACTAGTTTGTCTATGATTGAT
ATAAACGAACGAAACATCTTCGATGAAATCGGGAACATAAAGAACGTTACTAATCATCCATATGAGTTTTATAAGAAATGTGGTTATAT
GATCGTTGGAATAATCCCTAATGCTAATGGAAAAAGAAAACCAGATATTTGGATGTGGAAAGATATTAGCTAGGAAGAACAGCCCAA
CAACCGCTTCAACCTGACTCAGGGCGCTGTCACGATTTCTGCAGGTAATTCTGGACGCAGAAATCGCGCCAACGCCCTTCGCAGGT
TAAGCGAAGTG 

aacA27 
AAC(6´)-IIc 

TTAGGCCCGCACGGAATCAACATCTCATGTCCGCCAACAATGCCGCAATAGTTCTACGAGTCATGGCCGAGAACGATCTGCCAATG
CTCCATGCTTGGCTGAACCGCCCCCACATAGTCGAGTGGTGGGGCGGCGAGGATGAACGCCCAACTCTTGACGAAGTCTTAGAAC
ACTATTCGCCCGAAGTTCTGGCAAAGCAAGCTGTAGTGCCTTACATCGCAATGCTAGATGACGAACCCATCGGCTACGCCCAATCCT
ACATCGCACTTGGAAGTGGCGATGGATGGTGGGAAGACGAAACTGATCCAGGGGTCCGCGGGATTGACCAGTCTTTGGCTAATCC
ATCACAGTTAAACAAGGGGTTGGGTACAAAGCTCGTACGCTCGCTCGTTGAACTCCTGTTTAGCGACCCGGCCGTAACGAAAATCC
AAACCGATCCATCTCCTAGCAACCATCGCGCCATTCGCTGCTACGAGAAGGCCGGGTTCGTTCAAGAAAAAAACATCCTCACACCT
GACGGCCCTGCGGTGTACATGGTCCAAACACGCCAGGCGTTCGAAAGCCTGCGCACTGTTCAAAGCTTCAAAATCAAGGGGAAGT
GGTCATGAATGGCTGCACCATCTCTACCAACTGGTATCTCGGCGTGTTCTCATCTGCCTTCGCATCTGTCGTCAGCCTAACCGGGC
GTTCAACCGGACATCAACGTGCTGCGCACGTTGATGTCGGTTAACTGGGGCG 

aacA28 
AAC(6')-Iae 

TTAGCCGGACGCTGCGCGCGAAGAGGTTTTATGAAATACAACATTGTTAATATTAAAGATTCTGAAAAGTATATAACGCAAGCTGCAG
AAATTCTATTTGATGTATTTTCACACATAAATTTCGATTCTTGGCCGTCACTCCAAAAGGCTACAGAAACTGTAATAGAATGTATTAGC
GCCGAAAACATTTGTATTGGCATTTTAATAAACGATGAATTGTGTGGTTGGGTTGGATTAAGAGAAATGTATAAAAAAACTTGGGAAC
TACATCCAATGGTTATTAAGAAAAAACATCAAAATAAGGGATTTGGTAAAATACTAATTTTTGAAACAGAAAAGAAAGCGAAAGAAAGA
AATTTAGAAGGAATTGTACTTGGAACAGACGATGAAACATTTAGAACTACATTATCAATGTCAGAATTAAATAATGAAAATATATTCCA
TGAAATTAAAAATATAAAAAATCTAAAAAATCATCCATTTGAATTTTATGAAAAATGTGGTTACAGTATTATTGGTGTGATTCCTAATGC
AAATGGGAAAAATAAACCTGATATATTAATGTGGAAAAATATAATGTAAAAAAATTTCGGCTAACAACTGCTTCAACTTGATAATTGCT
TGAGCAATTACAAGTTAAGCAAATG 

aacA29 
AAC(6´)-29 

TTAGACGGCTTTGAGCGTTTCGATCTTACCTGTGAAAGAACAAGACGCTGCCGACTGGCTAGCGCTGCGGAATCTTCTTTGGCTCG
CGGATGATCACGCCTCGGAGATTGAGCAGTACTTCTCTGGTGGACTTGAGGAGCCTGTAGAAGTGCTCATCGCACGTGATGCTACC
GGCGCGGCTGTTGGGCATGTCGAACTCTCGATAAGACATGACTTGGAAGAACTCCAAGGAATCAAGACCGGCTACATCGAAGGCCT
TTATGTGGCCCCAAGCCATCGATCAACAGACCTTGTGAAGCGTTTCTTGCGTGAGTCCGAGAAGTGGGCCCTAGAACAAGGGTGCA
GCGCATTTGCCTCAGACAGAAGTGATCGGGTCATCACGCACCGCAAGTTCGCAGGCAGCGCCGTCTAACAACTCGTTCAAGCCGAA
CCCGCTTCGCTCCGGCAACGGCGTGGCAGGTTAAGCTTGCCACGCCGCCGCCTCCACTATGCGGGTCGGCTTAACTCAGGCG 

aacA30 
AAC(6')-I30 

TTAGGCTGGCGCGCTTCGCGCGGAAGACTTTATGGCTACTCGGAGACCTTAAATGGCATATGCGTTCTGCGAAATTGGAGAATCAA
ATGAATATATTATTCAGGCAGCTAGAATCTTAACGAAATCATTCCTTGATATTGGAAATGATTCCTGGCCTGATATGAAAAGTGCCAC
CAAAGAAGTTGAAGAATGTATTGAGAAGCCAAACATATGTCTTGGAATACATGAAAACGAAAAACTACTTGGATGGATTGGTCTTAGG
CCCATGTACAAATTAACATGGGAATTACATCCCTTGGTAATAAGTACGCAATATCAGAATAAAGGTATTGGAAGACTTCTAATAAATGA
ATTGGAAAAACAAGCAAAGCAAAATGGAATAATCGGAATAGTATTGGGAACTGACGATGAATACTTTAAGACTTCATTATCAGATGTG
GATCTTTCCGGGAAAAATATACTTGATGAGATAAGGAATATTAAAAATATAAGGAATCATCCGTACGAATTCTATCAACGATGTGGTTA
TTCCATTGTCGGAGTAATACCCGATGCAAATGGCAAAAGAAAGCCAGATATTTGGATGTGGAAGAAGATTAGTGATTAGGGCAAAAA
CGACAGCCTAACAATCGCTTTCACCTCGACAACCCGGCCTGTCACGCAATCTGCTAGTTTGATCCATGAGGCAGATTGCGCGCCAG
GCTCGGGTTGCGGGTGAAGCGTATG 

aacA31 
AAC(6´)-31 

TTAGGCAGCACAAAGACCGTTCTCATGACCACTACCATTAGCTTCGTCACGCTGCGCCTCATGACCGAGCACGACCTTCCGATGCT
CCATGACTGGCTAAATCGGCCTCACATCGTTGAGTGGTGGGGCGGAGAAGAAACACGTCCAACACTTGCTGAAGTGCTGGAGCAAT
ACCTACCAAGCGCCCTGGCGAAAGAGTCCGTCACTCCCTACATCGCAATGCTGGATGAAGAACCGATTGGGTACGCTCAGTCGTAC
ATTGCACTCGGAAGCGGTGACGGATGGTGGGAAGACGAAACCGATCCAGGAGTACGCGGAATAGACCAGTCTCTGGCGAATCCAT
CGCAGCTGGGCAAGGGCTTGGGAACCAAGCTCGTTTGCGCGCTCGTTGAGATGCTGTTCAAAGACGCTGAGGTAACCAAGATCCA
AACGGACCCGTCGCCGAACAACTTACGCGCAATCCGGTGCTACGAGAAGGCGGGTTTTGTGGCGCAAAGAACCATAAACACCCCA
GATGGACCGGCCGTATACATGGTTCAAACACGTCAGGCGTTCGAGCAGGCGCGCAGTGCTGTCTAACCCTTCCATCGAGGGGGAC
GTCCAAGGGCTATCGTCCTTGGCCGCCCCTCATGTCAAACG 

aacA32 
AAC(6´)-32 

TTAGGCAGCACAAAAGGACCGTCCCATGTCCCCGAGCAAAACACCCGTTACCTTGCGCCTCATGACCGAGCGCGACCTACCGATG
CTGCATGCATGGCTGAACCGGCCGCACATTGTCGAGTGGTGGGGTGGAGAAGAAGAACGCCCGACTCTTCATGAAGTGGTCAAAC
ACTACCTGCCGAGGGTTTTGGCAGAAGAAGCCGTCACACCATACATCGCGATGTTGGGCGACGAACCCATCGGCTACGCTCAGTCA
TACGTCGCACTCGGAAGCGGTGATGGATGGTGGGAGGATGAAACCGACCCAGGCGTACGAGGGATAGACCAATTCCTGTCGAACC
ATACACAGTTGAACCAGGGCCTAGGTACAAAGCTCGTCCAGGCACTCGTTGAACTGCTGTTCTCAGATCCTACCGTGACGAAGATC
CAAACCGACCCGGCGCCAAACAACCATCGAGCGATTCGCTGCTACGAGAAAGCTGGCTTTGTTCAGCAAAACGTCATCACCACACC
AGACGGCCCAGCCGTCTACATGGTTCAAACCAGGCAGGCCTTCGAGCGTGTGCGCAGTGCTGCCTAACCCCTCGCTCAAGCGGAG
CGCCAACGGCAGGCCACCAGGCCCGGGCCGGTGGTACGCTGTACATTTTCACCGGCCCGGGCCTGGCGTCCTGCCGTTGGCGCC
CGCTTAGCTCGAACG 

aacA34 TTAGAAGGCCCAGGCTATGCAGTACTCCATTCGCTCGGTTCGTGTTTCGGATACATCTGATTGGTTACGCCTTCGCAATCTCCTGTG
GGAAGGGGATGACCACGAAACCGAGATCGCCCAGTTTTTCGCCGGAGCCCTGGCCGAGCCCAACGAAGTGCTGGTAGCCCATGAT



Supplementary Material 

  178 

GATGCGGGGGCCGTTGTTGGGCATGTCGAGTTATCCATCCGCGAGGATGTCGCAGGGCTGGAAGGCATCAGAGCGGGCTATATCG
AAGGCCTGTACATCGAGGAGGCCCATCGCTCGTCCAGCGTCGCGACGCAGTTACTACGGCACTCCGAGCAATGGGCCCAAAGTCA
GGGATGCCGGGCGTTTGCATCGGATCGAGAGGATCGCCTGATCATCCATAAGCGGTTTTCTGTGAGTCCGCTTTCTAACCCTTCATT
CCAGCGGACCGCCTTCGGCGGCCGCTGAATTTGAACG 

aacA35 

TTAGGCAGCACAGGGCCACCTTCTTATGCCCTCCCACGATCATCCTGTCACCTTGCGGCTTATGACGGAGCAAGACCTGCCTATGC
TCCATGAATGGCTGAATCGGCCGCACATAGTCGAGTGGTGGGGCGGTGAAGAGCAACGTCCGACACTGGCGGATGTACTGGAACA
CTACAGGCCCAGAATCTTGGCGCACGAGTCGGTCACTCCATACATCGCCATGCTGGGCGAAGAACCAATCGGATATGCGCAGTCGT
ACGTCGCGCTCGGAAGCGGCGATGGATGGTGGGAAGAGGAAACCGACCCAGGAGTACGAGGAATCGATCAGTCGTTGGCGAATC
CTACGCAGTTGAACATGGGCCTGGGAACAGAGCTTGTGCGAGCGCTGGTTGATCGGTTGTTCTCCGACCCAACGGTGACAAAGATC
CAGACTGATCCGGCCCCAAACAATCGCCGCGCGATCCGCTGCTACGAGAAGGCGGGCTTTGTGCAGGAGAGAGTCATCACGACAC
CCGACGGGCAGGCCGTCTACATGACCCAATCTAGGCAAGCCTACGAGCGTGCGCGCGGTGCTGCCTAACCCCTCGCTCAACCCCC
GACTCGCTACGGCAGGCGTCGTAAGCCCGGTCCGCGCCAGCCGTAACATCATCGCGTACCGGGCTTACGACACCTGCCTCCGCTC
GCGGGGTTAGCTCGAACG 

aacA37 

TTAACCGCGGCTATGAAGTACTCTTTAAATCCAATCCGTCCCGAAGAATCACACGATTGGCTGCGCCTTCGAGATCTGCTTTGGGAA
GCTGATGACCATGCGATAGAAATTGCGAGATTCTTTACTGGCGAGCTAGAGGAAACGGTTGAAGTACTGATCGCACGCGACCTCGA
AGGACGTGCGGTAGGGCACGTTGAGCTTTCCATCCGTGAAGACATAGATGGTCTGAATGGCGTCAAGACCGGATACATCGAGGGG
TTGTACGTTGATGTATTGCATCGCTCAAGCGGTCTGGTCAGGCAGTTTCTGAGAGCATCGGAACAATGGGCAAAAGATCAAGGCTG
TAGCGCATTCGCATCTGATCGGCAAGACCGAGTAATCATCCACAGGCGGTTCTCCGGTGGCTCCGCCTAACAATTCATTCAAGCCG
ACGCCGCTTCGCGGCGCGGCTTAATTCACGCG 

aacA38 

TTAGGCAGCACATAACCACCGTCACACCATGCCTACCAAAGCGATCGCTGTCACCTTACGCGCCATGACCGAGGATGATCTTCCGA
TGCTGTACGACTGGCTCAACCGACCCCACATCGTTGAATGGTGGGGAGGCGAGGGGCAGCGTCCCACCCTCTCAGACGTGGTGGA
GCACTACCGGCCACGAGTCTTGGCAGAAGAGCGCGTCACGCCGTACATCGCCATGCTTGGCAATGAACCAATCGGCTATGCGCAG
TCGTACATTGCTCTCGGGAGTGGCGAAGGTTGGTGGCAGGACGAAACTGACCCTGGAGTTCGCGGCATCGATCAATCGCTTGCCA
ATCCCCTCCAGTTGAACAAAGGTCTCGGAACGGAGCTTGTGCGATCACTCGTCGAACTGTTGTTCTTGGATCCCGAGGTAACCAAG
ATTCAGACTGATCCGGCTCCTACTAACCATCGTGCAATCAGGTGCTACGAAAAAGCAGGGTTTGTGGTGCAGAACACAATCACTACC
CCCGATGGTCCGGCCGTCTATATGGTTCAAACTCGTCACGACTTCGAGCGAGCGCGCTGTGCTGCCTAACTGTTTGGTCAAGCTGA
ACGCCAAAAGCTGCGCTTTTGGTGCCCTCACTCCGCTTCGCTCCTTGCGGCGCAGCTTACCGCGGGCG 

aacA39 
AAC(6')-Iai 

TTAGCCGGACGCTTCGCGCAAAGAGGTATTATGAAATACACTATTATTGATATTAAAGATTCAGAAACGTACATTACTCAAGCTGCAG
AAATATTATTTGATGTATTTTCAGAAATAAGCCCAGAATCATGGCCAACACTCCAAAAAGCAAAAGAAGATGTTATTGAATGTATAGAA
GGTGAAAACATTTGCATTGGCATTATAATAAATAAAGAATTAATTGGATGGATTGGATTAAGAGAAATGTATAAAAAAACATGGGAATT
ACATCCTATGGTTATCAAGAAAACACATCATAATATGGGATTTGGAAAAATACTAATTAATGAAATAGAAAAAAAAGCAAGAGAAAGAA
ATTTAGAAGGTATTGTACTTGGAACAGATGATGAAACATATAGAACTTCATTATCAATGATTGAATTAAATAATGAAAATATTTTGCAAG
AAATAAAGAATATTAGAAATTTAGAAAATCATCCTTATGAATTTTATAAAAAATGTGGATATTGTATTATTGGTGTAATTCCAAACGCAA
ATGGGAAGAATAAGCCAGATATATTAATGTGGAAAAATATTATGGAAGAAAATTGCGGCTAACAACTGCTTCAACCTGACTCACCTAT
TGGCATGGTTTATGCTTTCGCTTCGCTTGGCATAAACCATGCCAATGTACGGTTCGCAGGTTAAGCAAATG 

aacA40 

TTAGGCAGCACAGTCCAGACTCCGCATGCCCTCCACTGCCCCTGAAGTCACCCAGCGACTCATGGTCAAGCGCGACCTGGTAATG
CTTCACGAATGGCTCAATCGCCCTCACATCGTTGAGTGGTGGGGAGGCGAAGAAGCCCGCCCGACGCTGCAAGAAGTGCAATCGC
ACTACCTACCTCGCGTGTTGGCCGAAGAGGCGGTCACGCCGTATATCGCGATGCTCGGGAGTGAGCCAATTGGGTACGCGCAGTC
TTACGTTGCATTGGGAAGTGGCGATGGGTGGTGGGAGGATGAAACAGACCCGGGTGTGCGCGGAATAGATCAGTCGCTGGCCAAT
CCAGAACAACTGGGCAAGGGCCTCGGGACCAGGCTTGTTTGTACCTTGGTCAAGACACTGTTCAACGATCCATCCGTGACCAAGAT
TCAAACCGACCCGGCGCCAAACAACGTCCGCGCGATCCGCTGCTACGAGAAGGCTGGCTTCAGACAACAGAAGGTCATCACCACG
CCGGACGGCCCGGCCGTCTACATGGTTCAAACCCGCGCCTCTTTCGAGGCAGCGCGCGGTGCTGCCTAACCCCTCGCCCAAGGC
GCCGACGCGCTACGGCAAGCAGCGTAAGCCTAGTCCGCGGCCTATACCGCATCATCCCGCACTAGGCTTACGCTGCTCGCCTCCG
CGCGCAGCTTAGCTCGAACG 

aacA42 

TTAGGCTGACGCGCTTCGCGCGGAAGACTTTATGGCTACTCGGAGACTTTGAATGGCGTATGAGTTCTGCGAAATAGGTGAATCAA
ACGAATATATTATTCTGGCGGCTAGAATCTTAACGAAATCATTCCTAGATATCGGTAATAATTCCTGGCCTGACATGAAAAGTGCTAC
TAAAGAAGTTGAAGAATGCATTGAGAAGCCAAACATATGTCTTGGAATACATGAAAATGAAAAATTGCTTGGATGGATTGGCCTTAGG
CCCATGTACAAATTAACATGGGAATTACATCCCTTGGTAATAAGTACTCAATATCAGAATAAAGGTATTGGAAGACTTTTAATAAATGA
ATTAGAAAAAAAAGCAAAGCAAATTGGAATAATTGGAATAGTATTGGGAACTGACGATGAATACTTTAAAACTTCATTATCAGCTGTTG
ATCTTTACGGCGAAAATATTCTTGATGAGATAAGGACTATTAAAAACATAAAAAATCATCCGTACGAATTCTATCAAAAATGTGGGTAT
TCCATTGTCGGAGTAATACCCGATGCAAATGGAAAAAGGAAGCCAGATATTTGGATGTGGAAGAAGATAAATGATTAGGGTAAAAAT
GACAGCCTAACAATCGCTTTCACCTCGACAACCCGGCCTGTCACGCAATCTGCTAGTTTAATCCTTGTGGAAGATTGCGCGCCAGG
CTCGGGTTGCGGGTGAAGCGTATG 

aacA43 

TTAGCCAGACGCTTCGCGCCGAGGACAATTGATGATTTATAACATAATTAACATTGCTGATTCTGAAAAGAACAAGGAAGACGCTGC
ACGAATTCTATATTCTGCATTTCGCGGAAAGGGAAAAGATGCTTGGCCTACATTAGATTCAGCTCGTGAAGAAATAGCAGAGTGCAT
AGCAAGTCCTAATATTTGCTTGGGCATAACCTTAGATGATCGCTTAGTAGGGTGGGGCGGACTTCGTCCCATGTATGAAACCACATG
GGAATTGCATCCCTTAGTAATAGATCCTGATTATCAAGGTAATGGATTAGGAAGACTGCTCCTATCAAAGATTGAGAGCACTGCAACC
ACAAATAGAATAATTGGTATAATGCTTGGAACAGATGATGAGACATTGAGTACAAGTCTTTCAATGACTGATATAGATGAGTCTAATAT
TTTCCAAGAGATAAAAAATATAATTAATATAAAGAATCATCCATTTGAATTTTATAAAAAATGCGGGTACATCATTGTCGGTATAGTACC
TAACGCAAATGGGTATAGAAAACCTGACATTTGGATGTGGAAGAATCTAGAAAAGAAAAGTGGCTAACAATCGCTTCAAGCGGATAA
AACGGCGTGTCACGGTTCTTGCTTACGCAAGAACACGCGCCACACTCGTTTTACCGCTTAAGCGTATG 

aacA44 

TTAGGCAGCACAAGACCACCTGTTCATGCCCGCGAACGAAAACACCGTAACCCTACGTCTGATGACTGAGCACAATTGGTGATTAAA
TGAACATTGACAACAGAACCGTAACCCTACGTCTGATGACTGAGCACGACCTCTCGATGCTCCATGAGTGGCTGAACCGGCCACAC
ATAGTTGAATGGTGGGGCGGTGAGGACGAACGTCCAACACTGGACGAGGTATATGAACACTACCTGCCAAGAGTTCTGGCGCAGG
AATCGGTCACCCCGTACATCGCCATGCTCGGCAACGAACCGATTGGCTACGCGCAGTCGTACGTAGCGCTCGGTAGCGGTGACGG
ATGGTGGGAAGAGGAAACTGACCCAGGGGTGCGAGGAATCGATCAGTCGCTGGCCAACCCAACGAAACTGAACAAAGGCCTCGGA
ACGAAGCTTGTACGCGCGCTGGCTGATCTGCTGTTCTCAGATGCGTCAGTGACAAAGATCCAGACCGACCCGGCTCCGGGTAACC
ATAGAGCTATTCGTTGCTACGAGAAAGCGGGGTTTGAGAGGCAAGGTACCGTAACCACCCCAGATGGTCCAGCCGTGTACATGGTT
CAAACACGCCAGGCATTCGAGCGAACACGCAGTGATGCCTAACCCTTCCATCGAGGGGGACGTCCAAGGGCTGGCGCCCTTGGCC
GCCCCTCATGTCAAACG 

aacA45 

TTAAAAGGCTCAGCCAATGCTGTACCTCATCCACCCGGTCAGTAGTTCGGACTCGCTCGATTGGCTTCGCCTTCGAAACCGGCTCT
GGACTGGACACGACCACGCGGAAGAAATTGCTGAATTCTTCAACGGAAATTTGGTGGAGCTGGACGAGGTATTGATCGCACATGAC
GACACGGGCATGGCCGTAGCTCATGTTGAGTTGTCAATCCGACAAGACATTGTTGGACTGGAGGGAGTCCGAACAGGTTACATCGA
AGGGCTCTATATCGATGAATTTCATCGCTCGTCCGGCATCGCACTCCAACTCTTGCGGGCGTCCGAGCTTTGGGCTCTGGATCACG
GGTGCCAAGCATTCGCTTCTGACCGCGAAGATCGCATCATCGTTCACAAGCGGTTTCCTGGAACTCCGCCTTCTAACATTTCAGTCG
ACCGGACCGCCTGCGGCGTCCGGTCACCTTCACGTTGGGCATTAAGGAAAAGTTAATGGCAATCCGAATCTTCGCGATACTTTTCT
CCATTTTTTCTCTTGCCACTTTCGCGCATGCGCAAGAAGGCACGCTAG 

aacA46 
AAC(6')-Iaj 

TTAGGGCGACGCCGCATTCGCGGCGCGTGAAGAAAGAGGATCTTATGGAATATTCAATTATCAATATAGTAGAGCAAAACAATTATC
AGATCGATGCTGCAAGAATTCTTACAAATACTTTTCTTGATATAGGTAATAAAACTTGGCCAACTATTCAAAGCGCAATCGATGAAGTC
GAAGAGTGTATTGATCTGCCCAATATATGTATAGGTTTAATTCATAACAATCAATTAATTGGATGGGTCGGATTACGTCCGATGTATG
ATAAAACGTGGGAATTGCACCCATTAGTTGTAAGAACTGACTATCAAAGTAAGGGGATCGGTAGTGTATTACTTGCTGAAGTTGAAAA
AAGGGCAAGAGAAGTTGGAATAATTGGAATAATATTAGGAACTGATGATGAATATAACAAAACAAGTCTTTCTGAAATAACTATAGAT
GAAAATAATATATTCGATGCAATACAAAATATTAAGAATATACATAATCATCCATATGAGTTTTACCAAAAAAATGGATATATGATTGTT
GGAATAATTCCAAATGCAAATGGACTAAGAAAACCCGATATTTGGATGTGGAAAAGTCTACTCAATTGAATGGAAACAATAATAACTA
TAGATAAGCTGTAAAAAATACTTTGACGAAATAACTTCGGTTGAAGTTATTATAGAAAAGAATATTAAAAAGAAAATTGTAAAAGAAATA
ATTTTTTATCAAAAAATATCATTGATTCTTCTGGAAGAAACAGAAAAAGAAATGATTTACTTTGGTAATAATCTACTTAATACACAATAG



Supplementary Material 

 179 

TTTTATTCAAAAATAATGTTCATTAGGTTATAGGAAATTTGCCCTAACAACTGTTTCAACCTGACATTGCTATTGTCACACTTTTTGCTT
TCGCTTCGCGGCAAAAAGTGCGCCAATTGACGCAATGCAGGTTAAACAAATG 

aacA47 

TTATGCATCACAGAACCACCATACCTATGTCCGCGATCGACACCCCCGTTACCTTGCGCCTCATGACCGAGCAAGATCTTCCGATGC
TCCATGACTGGCTGAACCGACCCCACATATTCGAGTGGTGGGGCGGTGAAGAGGAACGCCCGACTCTTGATGAAGTGCTGGAACA
CTACCTGCCAAGAGTTCTGGCAGAAGAGTCCGTCACGCCGTACATCGCAATGCTGGGCGAGGAGCCGATCGGCTACGCCCAGTCT
TACGTCGCACTCGGCAGCGGTGATGGATGGTGGGAAGACGAAACTGACCCAGGAGTGAGGGGAATTGACCAGTCTCTGGCTAATC
CGACACAGTTGAGCAAGGGTCTGGGAACAAAGCTTGTCCGTGCGCTCGTTGAGCGGTTGTTCTTGGACTCCACTGTGACGAAGATC
CAAACCGACCCAACTCCGAACAACCATCGGGCGATCCGTTGCTACGAGAAGGCGGGGTTTGTACGGCAGAAGATAATCACCACGC
CTGATGGCCCAGCCGTCTACATGGTTCAAACACGTCAGTCATTCGAGAACGCGCGTAGTACTGCCTAACCAGCCAATGGAGCCGAT
GCGGTGACGTATTGCGAGGTTTTAGTTCGGTGGCCCGCGCGGCTCATCGGCAACG 

aacA48 
AAC(6')-Iag 

TTAGGTCCCACTAAACCTGCACCGAGCATGGCGAACACTCCGGTTGGAAACGTCGTGCCATGCAAGACGCCAGATCACCCTGGCT
GGCTTGAGTTGCGCCTGCAGCTGTGGCCAGATGGCTCAACCGAGGAGTTCCTTCCCGAGATGGCTGCGGCCTGCGCTGAACCCGA
CCGCTTCGGCCAGTTTTTGTTCCTGTCGCCGGGCGGCTTGGCGGAGGGCCTGGTCGAGGTGGCGCTTCGCACGGACTACGTCAAC
GGCACCGAAAGCTCGCCAGTCGCCTTTCTCGAAGGCGTCTTCGTGGTGCCAGCGAGTCGAGGCCTTGGCATCGCCAGAGCGCTG
GTGGCTGCGGCGGAAGGCTGGGCTAGAGATCGTGGCTGCACTGAGTTCGCCTCGGACGCCGAGGTCAGCAACGTTGGTAGTCAC
GCCATGCACGCCGCTCTTGGTTTCGTCGAAACTGAGCGCGTCGTGTTCTTCCGTAAGGTCGTGGCACCGTGAGACCTAACCCCTCC
ATAGAGCGGACAGCCAAAAGCTGGCCGCTCATGTCGAACG 

aacA49 

TTAGCTTGACGCTTCGCGCAGAGGAGAGTTTCAATGAATATACAAATTTTGAACCTAGCAGAATGTACCGAATTTCAAGAGAGTGCA
GCAAGAGTCCTATTAGATGGATTTAGAGAGGTTGGCAAGATTGCATGGGCTACCTATGAAGAAGCTATGGTCGAAGTACAGGAATGT
ACTGAGATTCCTAATATAGCAATATGTGCTGTCGATAATAATAAAGTCGTTGGTTGGGTTGGAATTCGTCCAATGTACGATTACGTTT
GGGAACTACATCCAATGATAGTAACCAAGAAATACCAAAAAAAAGGAATAGGGACCAAACTACTTAAGGAAATTGAACAGATTGCGA
AAGAGAAAGGTTTACTAGGACTGGCTCTTGGAACTGATGATGAAACTGATAGTACAAGTCTATCGAAATGCGATTTTTCCCGAGATAA
TATATTGATCGAGATAGCGAATATCAAATCATCTATGTCGCACCCCTATGCGTTCTATGAAAAGAACGGATACTTTATTGTAGGAGTG
ATTCCGAATGCGAATGGAAAAAGAAAACCAGACATTTGGATGTGGAAAGAACTTGAAAGCTAACATCTAGTTCAACTTGACTTATTTA
GCCGTCACGGGTTCTTGCTTTCGCAACAACCGCGGCCAGCCAAAACGCAAGTTAATATAGGCG 

aacA50 

TTAGACAGCACAAAGACAATTCTCATGATCAACAGCATTAGCCTCGTTACGCTGCGCCTCATGACCGAGCAAGACCTACCGATGCTC
CACGACTGGCTAAACCGGCCGCACATCGTTGAGTGGTGGGGTGGAGAAGAGGCACGGCCATCACTCGCTGAGGTACACGAGCAAT
ACCTACCAAGTGTTCTGGCGAAAGAGTCGGTTACTCCATACATCGCAATGCTGAATCAAGAGCCGATCGGGTACGCTCAGTCATAC
GTTGCCCTCGGAAGCGGTGACGGATGGTGGGAAGACGAAACCGATCCAGGGGTGCGTGGAATCGACCAGTTCCTCGCGAGTCCA
CTACAACTTGGAAAAGGCCTTGGAACCAAGCTCGTTCGCGCGCTGGTAGAGACGCTGTTCAAAGATCCCGAGGTAACCAAGATCCA
AACGGACCCGTCGCCGAGTAACCTACGCGCGATCCGCTGCTATGAGAAGGCGGGGTTCGTGAGACAAAAGACAGTAGCCACGCCA
GATGGGCCAGCTGTGTACATGGTTCAAACACGCCACGAGTTCGGGCGGGCGCGCAGTGCTGCCTAACCCTTCCATCGAGAGGACA
TGCCCCGGCAAGCCGGGTCATGCCTCTCATGTCAAACG 

aacA51 

TTAGGCCACAAGGAACCGTCCCAGTATGAGCCCCGGCGTTGTCACCTTGCGGAGCATGACCGAAGAGGATCTCGGTATGCTTCAC
GACTGGTTGAACCGGCCCCACATCGTCGAATGGTGGGGTGGCGAGCGTCCTTCGCTTGAGGAGGTGCAAGAGCACTATCACCCTT
GTGCCCTTGCAGAAGCCAACGTGACTCCGTACGTCGGGATGCTCGATGGGCGGCCTTTTGCATATGCACAGTCGTATGTCGCCCTG
GGATCGGGAGATGGGTGGTGGCAGGACGAAACCGATCCGGGCATACGTGGTATCGACCAGTCAATTGGCGAGTCCGCGCTTCTAG
GGCAAGGTTACGGTACGTTGCTGGTACGCGCGCTCGTTGATCTGCTCTTCGCCGACCCGCGCGTATCGAAAGTTCAGACAGATCCT
TCTCCCCGGAACTTGCGCGCCATACGATGCTATGAGAAGGCTGGCTTCCGCAGGATCAAGACCATTGAGACACCTGATGGACCAGC
GATATACATGTTGCACGAGCGCCCATAGCCAGGGAATCCGCACGGTGTGGCCTAACAGATCGCTGCAACGGACAGTTGACCGAGC
CGCTCGCGCGCTACCGCACGCGATCGTCCGCGTCAACTGCCGCTGAGCTCCGGAG 

aacA52 

TTAGGCAGCACAAGATGACCGCGAACGAAAACACCGTAACCCTGCGCCTGATGACTGAGCACGACCTCCCCATGCTCCATGAATGG
CTGAACCGGCCACACATAGTTGAATGGTGGGGCGGTGAGGAAGAACGTCCAACACTGGACGAGGTATATGACCACTACCTGCCAA
CAGTTCTGGCGCAGGAATCAGTCACCCCGTACATCGCCATGCTAGGCAACGAACCGATTGGCTACGCGCAGTCGTATGTCGCGCTT
GGCAGCGGTGACGGGTGGTGGGAAGAGGAAACCGACCCAGGCGTGCGAGGAATCGATCAGTCGCTGGCCAACCCAACGAAACTG
AACAAAGGCCTCGGGACCAAGCTTGTACGCGCGCTGGCTGATCTGCTGTTCTCAGATGCGTCAGTGACAAAGATCCAGACCGACCC
GGCTCCGGGTAACCATAGAGCTATCCGTTGCTACGAGAAGGCAGGCTTTTTGCAGGAAAAGGTCATCACCACACCTGACGGGCCG
GCTGTCTACATGGTTCAGTCAAGGCAAACATACGAGCGTGCGCGCGGTGCTGCCTAACCCCTCGTTCAAGCGGACCGCCAACGGC
GTGGCGCCTTGGCGGCCGCTTAACTCGAACG 

aacA54 

TTAGGCCGCACAAAATCAACGCCTTATGTACACCAAGAATGCCGCAATAGTTCTGCGCCTCATGACTGAGAGCGATCTGCCAATGCT
CCATGCGTGGCTGAACCGGCCCCACATAGTCGAGTGGTGGGGAGGAGAAGATAAACGCCCCACACTTGGCGAAGTTTTAGAACAT
TATTCGCCCCGAGTTCTCGCAGAGCAAGCAGTAGTACCCTACATCGCAATGCTAGATGATGAACCCATTGGCTACGCGCAGTCCTA
CACCGCCCTTGGAAGTGGCGATGGATGGTGGGAAGACGAAACTGATCCAGGGGTCCGTGGCATTGACCAGTCATTGGCCAATCCG
TCGCAGCTAAACAAGGGCCTTGGAACAACGCTCGTACGCTCACTGGTTGAACTCCTGTTCAGTGATCCGGCCGTGTCGAAGATCCA
AACTGATCCTTCTCCTAACAACCATCGCGCTATCCGCTGTTACGAAAAAGCCGGGTTCGCACAAGACAAAATCATCCTGACGCCTGA
CGGCCCTGCGGTGTATATGGTTCAAACACGCCAAGCGTTCGAAAGCCAGCGCAATGCTGCCTAAAATTGAGGTCAAGGCGGCTCG
CGTTCATGAAGAGCCTCATCATCACTCTATCCATTCGCATCTCGGCGTGTTTTCCTACGCCTTGGCATCTGTCCCTAGCCTAACTGG
GCGCTCAACTGGACATCAACGTGCTGCGCACGTTGCTGCCAGTTAACTGGGGCG 

aacA56 

TTAGCCGGACGCTTCGCGCAGGAGTAAGAATGGAATACAAAATAGTAGATATTGCACTTGATAGTAAGCTGGTGAAAGTTGCAGCAG
AGATTTTGTTTTATACTTTTTCTGAAATTAATAATGAATCTTGGCCAACAATTAACTCTGCCACTGAGGAAGTGAAAGAATGTATAGAA
GATAAGAATATTTGTATTGGAGTTTTGGTAGAAGACAAGCTGGTAGGTTGGATTGGATTACGTCCAATGTATGAAAATACTTGGGAAT
TACATCCAATGGTGGTTTTATCAAAATGGCAAGGCAAAGGATTAGGGAAAATATTAATATTTGAATTAGAAAAAAGAGCAAAAGAACA
AGGAATAAACGGAATTGTTTTAGGAACTGACGATGAGACATTTCGAACATCTTTATCTATGAAAGAATTGGATAAAAATGATCTATTTG
AAGAAATAAAAAACATTAAAAATATTAATCATCATCCGTATGAGTTTTATCAGAAATGTGGATATAAAATTATTGGAGTAATCCCAGATG
CTAATGGTAAGAATAAACCGGATATTTGGATGTGGAAGAAAATAATGTAGGAAGAACGGCTAACAACGGTTTCAACCTGACTTGCCC
TTTGTCACGGTTTGTGCATGACGCTTCGCGGCTGCGCCGCTTCGCTTCTTCATGCACAAACCGCGCCAATCCGCTTCGCGGAACGG
GCTGCGCAGGTTAAACCAATG 

aacA59 

TTAGGCAGCACAGAAGCCGCATCCCATGCCCGCGAGCACATCCGTTGTGACCCTGCGCCTGATGACCGAGCACGACCTGCCAATG
CTCCACGAATGGCTGAACCGGCCGCACATCGTCGAGTGGTGGGGCGGCGAGGAAGAGCGCCCCAGTCTTGACGAAGTCCGCGAG
CACTATCTTCCGAGAGTCTTGGCTGAAGAGGCGGTTACGCCGTACATCGCAATGCTGGACGGCGAGGCAATTGGCTACGCCCAGT
CTTACGTCGCACTGGGAAGCGGCGATGGGTGGTGGGAAGACGAAACTGATCCGGGAGTACGAGGGATTGACCAGTCGCTGGCCA
ACCCCACACAGTTGAGCCGGGGCCTCGGAACGCAGCTTGTCCGTGCGCTCGTAGAGATGCTGTTCTCCAACCCCGCCGTGACTAA
AATTCAGACCGATCCCGATCCCAAGAACCTACGTGCAATCCGGTGCTATGAGAAGGCAGGGTTTGTACAGCAGAAGGTCATCACCA
CGCCCGATGGCCCCGCCGTCTACATGGTTCAAACGCGCAACGCGTTCGAGAGTTCGCGCAGTGCTGCCTAACACCTCGTTCAACC
CCCGACCCGCTACGGCAGGCACTGTAAGCCTGGCCCGAGGTACTCCGTACATTGTCTCGGTCCAGGCTTACAGTGCCCGCCTCCG
CGGTCGGGGTTAACTCGAACG 

aacA61 

TTAGGCAGCACAGGGCCACCGCTTATGTCCTCCAGCGAATCCAAAGTCACGTTGCGCCTCATGACCGAGCACGACCTGCCAATGCT
CCACGACTGGCTAAACCGGCCTCACATCGTTGAATGGTGGGGCGGTGAGGATGAGCGCCCTACTCTCGACGAAGTACTCGAGCAC
TACCTGCCACGAGTAATGGCGGAGGAATCAGTAACTCCATACATTGCCATGCTGGGCGATGAACCGATCGGCTACGCACAGTCTTA
CGTCGCACTCGGGAGCGGTGATGGATGGTGGGAAGACGAGACCGACGCAGGGGTACGAGGAATTGACCAATCTCTAGCCAATCCA
GCGCAGTTGAGCAAGGGCCTGGGAACACTCCTGGTCCGAACGCTCGTTGAAACCTTGTTCGCGGATCCGGCCGTCACGAAGATCC
AAACCGATCCGTCTCCAAACAACTACCGCGCAATCCGCTGCTACGAGAAGGCAGGGTTTGCACAGCAGGGCGTCATCACCACGCC
TGACGGGCCGGCGGTCTACATGGTTCAAACACGGCAAGCGTTCGAGCGCGCGCGCGGTGCTGCCTAACAACTCGCTCAACCGGA
CCCGCTACGGCAGGCCGGTTAGCTCGAACG 

aacA64 TTAGCCGGACGCCTTCGGCGCTAGGAATAAAATGGAATATTCAATTGTTAATATTGGCTTAAACGATAATTATATTACCCAAGCAGCT
GTTGTTTTATATAATGTATTTAATAATCTTGAAAACCAATCATGGCCAACTATTGATTCTGCCAAAATGGAAGTAAATGAATGCTTAGAA



Supplementary Material 

  180 

AATAATAACATCTGTATTGGAATGTTGATAGATAATAAAATAGTCGGTTGGGTAGGCCTAAGGCCAATGTATGAAAAAACTTGGGAAT
TACATCCATTAGTTGTCATTGATGATTATCAAAATATGGGTGTTGGTAAAATATTAATGAATGAAATAGAAAAAAGGGCAAAAGAAAAG
GGAATTATAGGGGTTGTACTTGGAACTGATGATGAGAAATATAAAACATCGTTATCAAAAGTTGATTTAAATAATAAGAATATATTCAC
TGAAATAGAAAATATTAAAAATATTAATCATCATCCATTCGAGTTTTATCAAAAATGTGGATATTTTATTGTTGGTGTAATACCAAATGC
AAATGGAAAAAATAAGCCAGATATATGGATGTGGAAAGAAATACAATAAGAAAGTTCGGCTAACAACTGCTTCAACCTGAGTCGCCTA
TTGTCATGATTTTTGCTGCTCGCTTCGCTCGTTCACGCAAAAATCACGCCAATTTACGGCTCCCAGGTTAAGCAAATG 

aacAX 

TTATGCATACAAATCATCACCGTGATTTACTCTTACCGGAAAGCTGAAGAAACAGATAGAGAAGCCATCTACCAATTGTATTGCTTGG
TAATGCGCGGCTTCATTTCTGAAATTTGGGGTTGGGATGAACAGTGGCAAGGAAACGATTTTTCTGCTCACTTTGATACCAAAGGCA
TTACGTTGGTACACCAAAAACACGAGTTGGTTGGGTATTCCCATGTCGAGAATCGAGGGGCCAGTGTCATAAGAATGATCGTTGTTC
ATCCTCATCACCAGCGGAAGGGTATCGGGAGAAAACTGCTTGAGTCTGTTATTGCGTCTGGCAATGAGCAATCCAAGGGCATCGGA
TTGGAGGTATTCAAAATCAATGATGAGGCAAAGAAATTCTATGAAAGATTTGGCTTTAATGTTGAAGGTGAAAACCCCACCAGTTACG
TCATGGCACATGCATAACCCGGCGCTCAACCTCGCTCCCTTCGGTCGCTGGACTCCGGGCGATAAGGCCGCCCGGAGCCGGTTAG
CTCTACG 

aacC1 
AAC(3)-Ia 

TTAGGTGGCTCAAGTATGGGCATCATTCGCACATGTAGGCTCGGCCCTGACCAAGTCAAATCCATGAGGGCTGCTCTTGATCTTTTC
GGTCGTGAGTTCGGAGACGTAGCCACCTACTCCCAACATCAGCCGGACTCCGATTACCTCGGGAACTTGCTCCGTAGTAAGACATT
CATCGCGCTTGCTGCCTTCGACCAAGAAGCGGTTGTTGGCGCTCTCGCGGCTTACGTTCTGCCAAAGTTTGAGCAGGCGCGTAGTG
AGATCTATATCTATGATCTCGCAGTCTCCGGCGAGCACCGGAGGCAAGGCATTGCCACCGCGCTCATCAATCTCCTCAAGCATGAG
GCCAACGCGCTTGGTGCTTATGTGATCTACGTGCAAGCAGATTACGGTGACGATCCCGCAGTGGCTCTCTATACAAAGTTGGGCAT
ACGGGAAGAAGTGATGCACTTTGATATCGACCCAAGTACCGCCACCTAACAATTCGTTCAAGCCGAGATCGCTTCGCGGCCGCGGA
GTTGTTCGGAAAAATTGTCACAACGCCGCGGCCGCAAAGCGCTCCGGCTTAACTCAGGCG 

aacC2 
AAC(3)-Ib 

TTAGGTGGCTCAATGAGCATCATTGCAACCGTCAAGATCGGCCCTGACGAAATTTCAGCCATGAGGGCTGTGCTCGATCTCTTCGG
CAAAGAGTTTGAGGACATTCCAACCTACTCTGATCGCCAGCCGACCAATGAGTATCTTGCCAATCTTCTGCACAGCGAGACGTTCAT
CGCGCTCGCTGCTTTTGACCGCGGAACAGCAATAGGTGGGCTCGCCGCCTACGTTCTACCCAAGTTCGAGCAAGCGCGAAGCGAG
ATCTACATTTATGACTTGGCAGTCGCTTCCAGCCATCGAAGGCTAGGAGTCGCAACTGCCCTGATTAGCCACCTGAAGCGTGTGGC
GGTTGAACTTGGCGCGTATGTAATCTATGTGCAAGCAGACTACGGTGACGATCCGGCAGTCGCTCTCTACACAAAGCTTGGAGTTC
GGGAAGACGTCATGCACTTCGACATTGATCCAAGAACCGCCACCTAACAATTCGTTCAAGCCGAGACCGCTTCGCGGCTCGGCTTA
GTGCGGTAACGTGTACCACACCGCCGTGCCGCGCAGCGGTCCGGCTTAACTCAGGCG 

aacC3 
AAC(3)-Ic 

TTAGGCAGCAGCAGCTAAGATGATCTCTACTCAAACCAAGATTACCCGCCTCAACTCTCAAGACGTTGGTGTAATGCGGGCAATGCT
AGGCATGTTCGGCGAGGCTTTTGAGGACGCTGAGAACTATTGCCGCGCTCAACCAAGCGACAGTTACCTACAAGACTTACTGTGTG
GCTCTGGCTTCATCGCAATCGCTGCGTTACAGGGGCAAGAGGTCATCGGTGGGCTCGCCGCGTATGTGCTCCCAAAGTTTGAACAA
CAGCGCAAAGAAATCTATATCTACGACTTAGGCGTGCAGGGAGCCTATCGCCGACGAGGCATCGCCACAGCCTTGATCAATGAACT
CCAGCGTATCGCACATGATATTGGCGCTTATGTAATTTTTGTCCAGGCTGACTATGGGGACGATCCTGCGGTAGCGCTCTACACAAA
ACTCGGTATCCGGGAGGACGTGATGCACTTTGACATAGAACCTCAACCTGCTGCCTAACAAGTCGTTCAAGCCGACGCTGCTTCGT
GGCAGCAACTGCGTGCATACGCTACGCTAGCACGCGTTGCCGGCACTACCGCAGCGCGGCTTAACTCAAGTG 

aacC4 

TTAGGTGGCTCACGTATGGGCATCATTCGCACATGTAGGCTCGGCCCTGACCAAGTCCAATCCATGCGGGCTGCTCTTGATCTTTTC
GGTCGTGAGTTCGGAGACGTAGCCACCTACTCCCAACATCAGCCGGACTCCGATTACCTCGGGAACTTGCTCCGTAGTAGGACATT
CATCGCGCTTGCTGCCTTCGAGCAAGAAGCGGTTGTTGGCGCTCTCGCGGCTTACGTTCTGCCCAAGTTTGAGCAGGCGCGTAGT
GAGATCTATATCTATGATCTCGCAGTCTCCGGCGAGCACCGCCGGCAGGGCATTGCCACCGCGCTCATCAATCTCCTCAAGCATGA
GGCCAACGCGCTTGGTGCTTACGTGATCTACGTGCAAGCGGATTACGGTGACGATCCCGCAGTGGCTCTCTATACAAAGTTGGGCA
TACGGGAAGACGTGATGCACTTTGATATCGACCCAAGTTCCGCCACCTAACAATTCGTTCAAGCCGAGATCGCTTCGCGGCCGCGG
AGTTGTTCGGTAAATTGTCAAAACGCCGCGGCCGCAAAGCGCTCCGGCTTAACTCAGGCG 

aacC5 
AAC(3)-Id 

TTAGGCATCAGGAGCAGACGAGTGTCAGTCGAAATCATCCATCTCACTGGAAACGATGTTGCGTTGTTGCAGTCAATAAATGCCATG
TTCGGCGAGGCATTCAACGACCAAGATAGTTATGCCCGCAACAAGCCGTCATCAAGCTATCTTCAAAAACTGCTTAGCACTTCTAGT
TTTATTGCGTTGGCTGCGGTTGACGAGCAAAAAGTCATTGGCGCTATCGCCGCGTATGAGTTGCAAAAATTCGAGCAGCAAAGAAG
CGAGATTTATATCTACGATCTCGCTGTAGCGGCAACCCGCCGCAGAGAAGGCATAGCTACAGCTCTAATTAAAAAACTCAAGGCTAT
AGGCGCAGCGCGTGGAGCTTATGTGATTTACGTCCAAGCTGATAAAGGCGTAGAAGACCAACCAGCCATAGAGCTCTATAAAAAAC
TAGGAACCATCGAAGACGTATTTCATTTCGACATTGCGGTTGAGCAGAGTAAAAATCATGCCTAACAAGGCACTCCAGCCGACGGCT
TTGCCTTCGCTGCGCTACGGCAAATCCGCGGCTGAGTTTGGGCG 

aacC6 
AAC(3)-If 

TTAGGTGGCTCAATGAGCATCACTGCTACCATCAGGATTGGTCCTGATGAGATTCCGGCCATGCGGGCTGTGCTCGACCTGTTCGG
CAGAGAGTTTGAGGACATGTCCGCCTACTCGGATCGCCAGCCGACAAATGATTACTTGGCCAAGCTTCTCCGCAGCGAGACGTTCA
TCGCGCTAGCCGCATTTGACCAAGGAACAGCAATAGGTGGGCTCGCTGCCTACGTTTTGCCCAAGTTCGAGCAAGCGCGTAGTGA
GATCTACATTTATGACTTAGCAGTCGCATCCAGCTATCGACGGCAAGGAATCGCAACTTCCCTGATTAGCCACCTGAAGCGTGAGGC
AATCAAGATTGGGGCATATGTGATCTATGTGCAAGCAGACTACGGAGACGACCCCGCGGTGGCTCTCTACACCAAGCTTGGTGTTC
GGGAAGATGTCATGCACTTTGACATTGATCCAGGAACCGCCACCTAACAATTCGTTCAAGCCGAACTTGCTTCGTTACACCAGCGCC
GTGGCAGGTTAAGCTTGCCACGTCGCCGGCTACACTTCGCAAGTCGGCTTAACTCAGGCG 

aacC11 

TTAGGTGGCTCACGTATGGGCATCATTCGCACATGTAGGCTCGGTCCGGACCAAGTGCAATCCATGCGGGCTGCTCTTGATCTCTT
CGGTCGTGAGTTCGGAGACGTAGCCAGCTACTCCCAACATCAGCCGGACTCCGATTACCTCGCGAAATTGCTCCACAGCAGGACAT
TCATCGCGCTTGCCGCGTTCGACCAAGAATCGGTTGTTGGCGCTCTTGCCGCCTACGTTTTGCCAAAGTTTGAGCAAGCACGTAGC
GAGATCTACATCTATGATCTCGCAGTCTCCGGCGAGCACCGGCGGCAAGGCATAGCCACCGCGCTCATCAATCTCCTCAAGCAAGA
GGCCAATGCGCTTGGTGCTTACGTGATCTATGTGCAAGCTGACTACGGTGACGATCCCGCAGTGGCCCTCTACACAAAGTTGGGCA
TACGGGAAGACGTCATGCACTTTGATATCGACCCAAGTACCGCCACCTAACAATTCGTTCAAGCCGAACCTGCTTCGTGGCCCGGC
TAATGTGCTAGCGTGTAGCACAAAGCCGTGCCACTACGCAGGTCGGCTTAACTCAGGCG 

aacC13 

TTAGGCATTAGGAGCCGATGAATGTCAGTCAAGATTATTCAACTCACTGAAAAAGATGTTGGCTTAATGCAGTCTATAAATGCCATGT
TCGGTGAGGCATTCGATGATAAAGAGCATTATTCCAGCAACAAGCCGACCTCAAGCTATCTTAAAAAACTGCTTGGCAGTTCAAGTTT
TATTGCGTTGGCTGCGGTTCACGAGGAAAAAGTCATAGGCGCTATAGCTGCGTATGAGTTGCAAAAATTTGAGCAGCAAAGAAGCG
AAATCTACATCTACGACCTCGCTGTAGCGGCAACCCACCGCAGAGCAGGCGTAGCTACAGCACTAATTCACAAATTGAAGGCGATA
GGCGCCGAGCGCGGCGCTTATGTCATATACGTCCAAGCCGATAAAGGCGTAGAAGACCAACCGGCCATAGAGCTTTATAAAAAACT
GGGAACCATCGAAGACGTATTCCACTTCGATATTGCAGTTGAGGATAATAAAAATGCCTAACAAGGCGTTGCACCGGAAAAATTTGC
CTTCGCTTCGCTACGGCAATTTTTCAGGTGAACTTGGGCG 

aadA1 
ANT(3’’)-1 

TTAAACATCATGAGGGAAGCGGTGATCGCCGAAGTATCGACTCAACTATCAGAGGTAGTTGGCGTCATCGAGCGCCATCTCGAACC
GACGTTGCTGGCCGTACATTTGTACGGCTCCGCAGTGGATGGCGGCCTGAAGCCACACAGTGATATTGATTTGCTGGTTACGGTGA
CCGTAAGGCTTGATGAAACAACGCGGCGAGCTTTGATCAACGACCTTTTGGAAACTTCGGCTTCCCCTGGAGAGAGCGAGATTCTC
CGCGCTGTAGAAGTCACCATTGTTGTGCACGACGACATCATTCCGTGGCGTTATCCAGCTAAGCGCGAACTGCAATTTGGAGAATG
GCAGCGCAATGACATTCTTGCAGGTATCTTCGAGCCAGCCACGATCGACATTGATCTGGCTATCTTGCTGACAAAAGCAAGAGAACA
TAGCGTTGCCTTGGTAGGTCCAGCGGCGGAGGAACTCTTTGATCCGGTTCCTGAACAGGATCTATTTGAGGCGCTAAATGAAACCT
TAACGCTATGGAACTCGCCGCCCGACTGGGCTGGCGATGAGCGAAATGTAGTGCTTACGTTGTCCCGCATTTGGTACAGCGCAGTA
ACCGGCAAAATCGCGCCGAAGGATGTCGCTGCCGACTGGGCAATGGAGCGCCTGCCGGCCCAGTATCAGCCCGTCATACTTGAAG
CTAGACAGGCTTATCTTGGACAAGAAGAAGATCGCTTGGCCTCGCGCGCAGATCAGTTGGAAGAATTTGTCCACTACGTGAAAGGC
GAGATCACCAAGGTAGTCGGCAAATAATGTCTAACAATTCGTTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAACTCAAGCG 

aadA2 

TTAGACATCATGAGGGTAGCGGTGACCATCGAAATTTCGAACCAACTATCAGAGGTGCTAAGCGTCATTGAGCGCCATCTGGAATCA
ACGTTGCTGGCCGTGCATTTGTACGGCTCCGCAGTGGATGGCGGCCTGAAGCCATACAGCGATATTGATTTGTTGGTTACTGTGGC
CGTAAAGCTTGATGAAACGACGCGGCGAGCATTGCTCAATGACCTTATGGAGGCTTCGGCTTTCCCTGGCGAGAGCGAGACGCTC
CGCGCTATAGAAGTCACCCTTGTCGTGCATGACGACATCATCCCGTGGCGTTATCCGGCTAAGCGCGAGCTGCAATTTGGAGAATG
GCAGCGCAATGACATTCTTGCGGGTATCTTCGAGCCAGCCATGATCGACATTGATCTAGCTATCCTGCTTACAAAAGCAAGAGAACA
TAGCGTTGCCTTGGTAGGTCCGGCAGCGGAGGAATTCTTTGACCCGGTTCCTGAACAGGATCTATTCGAGGCGCTGAGGGAAACCT
TGAAGCTATGGAACTCGCAGCCCGACTGGGCCGGCGATGAGCGAAATGTAGTGCTTACGTTGTCCCGCATTTGGTACAGCGCAATA



Supplementary Material 

 181 

ACCGGCAAAATCGCGCCGAAGGATGTCGCTGCCGACTGGGCAATAAAACGCCTACCTGCCCAGTATCAGCCCGTCTTACTTGAAGC
TAAGCAAGCTTATCTGGGACAAAAAGAAGATCACTTGGCCTCACGCGCAGATCACTTGGAAGAATTTATTCGCTTTGTGAAAGGCGA
GATCATCAAGTCAGTTGGTAAATGATGTCTAACAATTCGTTCAAGCCGACCGCGCTACGCGCGGCGGCTTAACTCCGGCG 

aadA4 

TTAGGCATCTTCATGGGTGAATTCTTTCCTGCACAAATTTCCGAGCAGCTATCCCACGCTCGCGGGGTGATCGAGCGCCATCTAGCT
GCAACGCTGGACACAATCCACCTGTTCGGATCTGCGCTCGATGGAGGGTTGAAGCCGGACAGCAACATCGACTTGCTCGTGACCG
TCAGCGCCGCACCTAACGATTCGCTCCGGCAGGCACTAATGCTCGACCTGCTAAAAGTCTCATCACCGCCAGGCAATGGCGGACC
ATGGCGACCGCTGGAGGTGACTGTTGTCGCTCGAAGCGAAGTAGTGCCCTGGCGCTATCCGGCGCGACGTGGGCTTCAGTTCGGT
GAGTGGCTCCGCCACGACATCCTCTCCGGAACGTTCGAGCCTGCCGTTCTGGATCACGATCTTGCGATTTTGCTGACCAAGGCGAG
GCAACACAGCCTTGCACTGCTAGGTCCATCCGCAGTCACGTTCTTCGAGCCGGTGCCGAACGAGCATTTTTCCAAGGCGCTTTTCG
ACACGATTGCCCAGTGGAATTCAGAGTCGGATTGGAAGGGTGACGAGCGGAACGTCGTTCTTGCTCTTGCTCGCATTTGGTACAGT
GCTTCAACGGGTCTCATTGCTCCTAAGGACGTTGCTGCCGCATGGGTATCGGAGCGTTTGCCTGCCGAGCATCGGCCCATCATTTG
CAAGGCACGCGCGGCGTACCTGGGTAGCGAGGACGACGACCTAGCAATGCGCGTCGAAGAGACGGCTGCGTTCGTTCGATATGC
CAAAGCAACGATTGAGAGAATCTTGCGTTGAGCACGACGTGCGAAAGTGCATCGACCGGCGCCCAGGCACTTGATGCCTAACTCC
GCGTTGAAGTGGAGGCTGCGCCCGCCGCTCAACTATGCG 

aadA5 

TTAGGCATCATGGGTGAATTTTTCCCTGCACAAGTTTTCAAGCAGCTGTCCCACGCTCGCGCGGTGATCGAGCGCCATCTGGCTGC
GACACTGGACACAATCCACCTGTTCGGATCTGCGATCGATGGAGGGCTGAAGCCGGACAGCGACATAGACTTGCTCGTGACCGTC
AGCGCCGCACCTAACGATTCGCTCCGGCAGGCGCTAATGCTCGATTTGCTGAAAGTCTCATCACCGCCAGGCGATGGCGGAACAT
GGCGACCGCTGGAGCTAACTGTTGTCGCTCGAAGCGAAGTAGTGCCTTGGCGCTATCCGGCGCGGCGTGAGCTTCAGTTCGGTGA
GTGGCTCCGCCACGACATCCTTTCCGGAACGTTCGAGCCTGCCGTTCTGGATCACGATCTTGCGATTTTGCTGACCAAGGCGAGGC
AACACAGCCTTGCGCTTCTAGGCCCATCCGCAGCCACGTTTTTCGAGCCGGTGCCGAAGGAGCATTTCTCCAAGGCGCTTTTCGAC
ACTATTGCCCAGTGGAATGCAGAGTCGGATTGGAAGGGTGACGAGCGGAACGTCGTTCTTGCTCTTGCTCGCATTTGGTACAGCGC
TTCAACTGGTCTCATTGCTCCTAAGGACGTTGCTGCCGCATGGGTATCGGAGCGTTTGCCTGCCGAGCATCGGCCCCTCATCTGCA
AGGCACGCGCGGCGTACCTGGGTAGCGAGGACGACGACCTAGCAATGCGCGTCGAAGAGACGGCCGCGTTCGTTCGATATGCCA
AAGCAACGATTGAGAGAATCTTGCGTTGAGCGGCATGTGCGAAAAGTGCATCGACCCGCGCCGAGGGCATCTGATGCCTAACTCG
GCGTTCAAGCGGACGGGCTGCGCCCGCCGCTCAACTATGCG 

aadA6 

TTAGACATCATGAGTAACGCAGTACCCGCCGAGATTTCGGTACAGCTATCACTGGCTCTCAACGCCATCGAGCGTCATCTGGAATCA
ACGTTGCTGGCCGTGCATTTGTACGGCTCTGCACTGGACGGTGGCCTGAAGCCATACAGTGATATTGATTTGCTGGTTACTGTGGC
TGCACGGCTCGATGAGACTGTCCGACAAGCCCTGGTCGTAGATCTCTTGGAAATTTCTGCCTCCCCTGGCCAAAGTGAAGCTCTCC
GCGCCTTGGAAGTTACCATCGTCGTGCATGGTGATGTTGTCCCTTGGCGTTATCCGGCCAGACGGGAACTGCAATTCGGGGAGTG
GCAGCGTAAGGACATTCTTGCGGGCATCTTCGAGCCCGCCACAACCGATGTTGATCTGGCTATTCTGCTAACTAAAGTAAGGCAGC
ATAGCCTTGCATTGGCAGGTTCGGCCGCAGAGGATTTCTTTAACCCAGTTCCGGAAGGCGATCTATTCAAGGCATTGAGCGACACT
CTGAAACTATGGAATTCGCAGCCGGATTGGGAAGGCGATGAGCGGAATGTAGTGCTTACCTTGTCTCGCATTTGGTACAGCGCAGC
AACCGGCAAGATCGCACCGAAGGATATCGTTGCCAACTGGGCAATGGAGCGTCTGCCAGATCAACATAAGCCCGTACTGCTTGAAG
CCCGGCAGGCTTATCTTGGACAAGGAGAAGATTGCTTGGCCTCACGCGCGGATCAGTTGGCGGCGTTCGTTCACTTCGTGAAACAT
GAAGCCACTAAATTGCTTAGTGCCATGCCAGTGATGTCTAACAATTCATTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAATTCAG
GCG 

aadA7 

TTAGACATCATGAGTGAAAAAGTGCCCGCCGAGATTTCGGTGCAACTATCACAAGCACTCAACGTCATCGGGCGCCACTTGGAGTC
GACGTTGCTGGCCGTGCATTTGTACGGCTCCGCACTGGATGGCGGATTGAAACCGTACAGTGATATTGATTTGCTGGTGACTGTAG
CTGCACCGCTCAATGATGCCGTGCGGCAAGCCCTGCTCGTCGATCTCTTGGAGGTTTCAGCTTCCCCTGGCCAAAACAAGGCACTC
CGCGCCTTGGAAGTGACCATCGTCGTGCACAGTGACATCGTACCTTGGCGTTATCCGGCCAGGCGGGAACTGCAGTTCGGAGAGT
GGCAGCGCAAAGACATCCTTGCGGGCATCTTCGAGCCCGCCACAACCGATTCTGACTTGGCGATTCTGCTAACAAAGGCAAAGCAA
CATAGCGTCGTCTTGGCAGGTTCAGCAGCGAAGGATCTCTTCAGCTCAGTCCCAGAAAGCGATCTATTCAAGGCACTGGCCGATAC
TCTGAAGCTATGGAACTCGCCGCCAGATTGGGCGGGCGATGAGCGGAATGTAGTGCTTACTTTGTCTCGTATCTGGTACACCGCAG
CAACCGGCAAGATCGCGCCAAAGGATGTTGCTGCCACTTGGGCAATGGCACGCTTGCCAGCTCAACATCAGCCCATCCTGTTGAAT
GCCAAGCGGGCTTATCTTGGGCAAGAAGAAGATTATTTGCCCGCTCGTGCGGATCAGGTGGCGGCGCTCATTAAATTCGTGAAGTA
TGAAGCAGTTAAACTGCTTGGTGCCAGCCAATGATGTCTAACAATTCATTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAATTCAAG
CG 

aadA9 

TTAGACATGATGAGCAACTCTATACACACCGGAATCTCAAGACAGCTTTCACAGGCACGCGATGTAATTAAACGCCATTTGGCATCA
ACGCTGAAAGCCATACACTTGTATGGTTCTGCAATTGATGGTGGCCTCAAACCATATAGCGACATTGATCTGCTGGTTACCGTGGAT
GCACGCTTGGATGAAGCTACCAGACGCTCCCTGATGCTCGATTTCTTGAATATCTCGGCACCACCATGCGAAAGCTCAATACTCCG
GCCGCTAGAGGTAACTGTTGTTGCATGCAACGAAGTAGTGCCTTGGCGTTATCCGGCACGACGAGAACTGCAGTTCGGGGAGTGG
CTGCGGGAGGATATTCTTGAAGGTGTCTTCGAGCCAGCCGCCTTGGACGCCGACCTTGCAATTCTAATAACGAAAGCTAGGCAACA
CAGCATCGCTTTAGTAGGTCCAGTGGCTCAAAAAGTCTTCATGCCGGTGCCAGAGCATGACTTTCTCCAGGTGCTTTCCGATACCCT
TAAGCTGTGGAATACTCATGAGGATTGGGAAAATGAGGAGCGGAACATCGTACTCACGTTAGCTCGGATCTGGTATAGCACTGAAA
CTGGAGGAATCGTCCCCAAGGATGTGGCCGCCGAATGGGTTTTAGAGCGCTTGCCAGCTGAGCATAAGCCAATACTGGTTGAGGC
GCGGCAAGCCTATCTTGGGCTTTGCAAGGATAGTCTTGCTTTGCGTGCAGATGAGACTTCGGCGTTCATTGGCTATGCAAAGTCTG
CGGTCGCTGATTTGCTCGAAAAGCGAAAATCTCAAACTTCGCATATTTGCGATGGCGCCAAGAACGTCTAACGTCTAACTATTCATTT
AAGCCGAAGCCGCTTCGCGGCTCGGCTTAATTCAGGCG 

aadA10 

TTAGACATCATGATAAACACAGTGCCCGCCGAGATTTCGGTACAGTTATCACAGGCACTCAACGTCATCGAGCATCATCTGGGATCG
ACGTTGCTGGCCATGCATTTGTATGGCTCTGCACTCGACGGTGGCCTGAAGCCATACAGTGATATTGATTTGCTGGTTACTGTGACC
GCACGGCTCGATGAGAGTGTGCGGCAAGCTCTGTTCGTCGATCTCTTGGGGGTTTCCGTTTTCCCTGGTCAAAGCAGAGTTCTCCG
CGCCTTGGAAGTTACCATTGTCGTGCACAGTGACATCGTTCCTTGGCGCTATCCGGACAGACGGGAACTGCAATTCGGGGAGTGGC
AGCGCAAAGACATTCTTGCGGGCATCTTCGAGCCTGCGACAACCGATGTTGATCTAGCCATTCTGCTAACAAAAGCAAGGCAACATA
GCCTTGCCTTGGCCGGTTCGGCTGGGGAAGATTTCTTCAACCCAGTCCCGGAAAGCGATCTATTCAAGGCACTGGCCGACACCCTG
AAACTATGGAACTCACAGCCGGATTGGATAGGTGACGAGCGGAATGTAGTGCTTACTTTGTCTCGTATTTGGTACAGCGCAGCAACC
GGCAAGATCGCGCCGAAGGATGTTGCCGCCAACTGGGTAATGGAGCGTTTGCCAGTTCAACATCAGCCCGTGCTGCTTGAAGCCC
GGCAGGCTTATCTTGGACAAGGAGAAGATTGCTTGGCTTCGCTCACGGATCAGTTAGAGGCGTTTGTTCACTTCGTGAAGCATGAA
GCCACTAAACTGCTTGGTGCCACGCCAATGATGTCTAACAATTCATTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAATTCAGGCG 

aadA11 

TTAGACATCATGAGGGAAGCGGTGACCGCCGAAATTTCAACACAACTATCAGAGGTGTTTAGCGTCATCGAGCGCCATCTGGAGCC
GACGTTGCTTGCCGTGCATTTGTACGGCTCCGCAGTGGATGGCGGCTTGAAGCCATACAGCGATATTGATTTGCTGGTTACTGTGA
CCGCAAGGCTTAATGAAGCAACACGGCAAGCTTTGCTCAACGACCTTTTGGAGGCTTCGGCTTTCCCTGGCGAGAGCGAGACTCTC
CGCGCTATAGAAGTCACCTTTGTCGTGCACGACGACATCATCCCGTGGCGTTATCCAGCTAAGCGCGAACTGCAATTTGGAGAATG
GCAGCGTAATGACATTCTTGCGGGTATCTCCGAGCCAGCCGCGATCGACGTTGATCTGGCTATCTTGCTGACAAAAGCAAGAGAAC
ATAGCGTTGCCTTGGTAGGTCCAGCTGCGGAGGAACTCTTTGATCCCGTTCCTGAACAGGATCTATTCGAGGCACTGAATGAAACCT
TGAAGCTATGGAACTCGCAGCCCGACTGGGCCGGCGATGAGCGAAATGTAGTGCTCACGTTGTCCCGCATTTGGTACACCGAAGTA
ACCGGAAAAATCGTGCCGAAGGATGTCGCTGCCGACTGGGCAATGGAGCGCCTACCTGCCCAGCATCAGCCCGTCTTACTTGAAG
CTAGACAGGCTTATCTTGGACAAAAAGAAGATCGCTTGGCCTCGCGCGCAGATCAGTTGGAAGAATTTGTTCACTTCGTGAAAGGC
GAGATCACTAAGGTAGTCGGCAAATGATGTCTAACAATTCGTTCAAGCCGATTCCGCTTCGCGGCACGGCTTAACTCAGGCG 

aadA13 

TTAGACATCATGAGGGACTCAGTGACCGCCGAAATTTCGACGCAACTATCCAAGGTGCTTAGTGTTATCGAGCACCATCTGGAACCG
ACGTTGCTTGCCGTACATTTGTACGGCTCCGCAGTGGATGGCGGCCTGAAGCCATACAGTGATATTGATTTGCTGGTTACTGTGACC
GCAAGGCTTGATGACACAACGCGGCGAGCTTTGTTCAACGATCTTTTGGAGGTTTCGGCTTTCCCAGGCGAGAGTGAGATTCTCCG
CGCTATAGAAGTCACCATTGTCGTGCACGAAGACATTATGCCGTGGCGTTATCCAGCCAAGCGCGAACTGCAATTTGGAGAATGGC
AGCGCAATGACATTCTTGCGGGTATCTTCGAGCCAGCCACGATCGACATCGATCTGGCTATCTTGCTAACGAAAGCGAGAGAACAT
AGCGTGGCTTTGGTAGGTCCGGCGGCGGAGGAACTCTTTGATCCAGTTCCTGAACAAGATCTAATCAAGGCGCTGAATGAAACCTT
GAAGCTATGGAACTCGCAGCCCGACTGGGCCGGCGATGAGCGAAATGTAGTGCTTACGTTGTCCCGCATTTGGTACAGCGCAGCA
ACTGGTAAAATCGCGCCGAAGGATGTCGCTGCCAACTGGGCAATGGAACATCTACCTGCCCAGCATCAGTCTGTCTTGCTTGAAGC



Supplementary Material 

  182 

TAGACAGGCTTATCTTGGGCAAGAGGAAGATCGCTCGGTCTTGCGCGCAGATAAGTTGGAAGAATTTATTCACTTCATGAAAAGCGA
GATCACCAAGGTGCTCGGCAATGATGTCTAACAATGCGTTCAAGCCGATGCCGCTTCGCGGCACGACTTAACTTCGGCG 

aadA16 

TTAGACATCATGAGCAACGCAGTGCCCGCCGAGATTTCGGTACAGCTATCACAGGCACTCAACGTCATCGAGCATCATCTGGGATC
GACGTTGCTGGCCGTGCATTTGTACGGCTCTGCACTCGACGGTGGCCTGAAGCCATGCAGTGATATTGATTTGCTGGTTACTGTGA
CTGCACAGCTCGATGAGACTGTGCGGCAGGCTCTGTTCGTAGATTTCCTGGAAGTTTCCGCTTCTCCCGGCCAAAGTGAAGCTCTC
CGTGCCTTGGAAGTTACCATCGTCGTGTACGGCGATGTTGTTCCTTGGCGTTATCCAGCCAGACGGGAACTGCAATTCGGGGAGTG
GCAGCGCAAGGACATTCTTGCGGGCATCTTCGAGCCCGCGACAACCGATGTTGATCTGGCTATTCTGCTAACTAAAGCAAGGCAAC
ACAGCCTTGCCTTGGCAGGTTCGGCCGCGGAAGATTTCTTCAACTCAGTCCCGGAAAGCGATCTATTCAAAGCACTGGCCGACACC
TTGAAACTATGGAACTCACAACCGGATTGGGCAGGCGACGAGCGGAATGTAGTGCTTACTTTGTCTCGCATTTGGTACAGCGCAGC
AACCGGCAAGATCGCGCCGAAGGATGTAGCTGCCAACTGGGTAATGGAACGCCTGCCCGTCCAACATCAGCCCGTGCTGCTTGAA
GCCCAGCAGGCTTACCTTGGACAAGGGATGGATTGCTTGGCCTCACGCGCTGATCAGTTGACTGCGTTCATTTACTTTGTGAAGCA
CGAAGCCGCCAGTCTGCTCGGCTCCACGCCAATGATGTCTAACAGTTCATTCAAGCCGACGCCGCTTCGCGGCGCAGCTTAATTCA
GGCG 

aadA24 

TTAGACATCATGAGGGACGCAGTGATCGCCGAAATTTCGACACAACTGTTAGAGGTGCTTAGTGTCATTGAGCGCCATCTGGAGCC
GACGTTGCTGGCCGTGCATTTGTACGGCTCCGCAGTGAATGGCGGCCTGAAGCCATACAGCGATATTGATTTGCTGGTTACTGTGA
CTGTAAGGCTTAATGAAACAACGCGGCGAGCTTTGCTCAACGACCTTCTGGAGGTTTCGACTTTCCCCGGCGAGAGTGAGGCTCTC
CGCGCTATAGAAGTCACCATTGTCGTGCACGACGACATCATTCCGTGGCGTTATCCAGCTAAGCGAGAACTGCAATTTGGAGAATG
GCAGCGCAATGACATTCTTGCGGGTATCTTCGAGCCAGCCACGATCGACATTGATCTGGCTATCTTGCTGACAAAAGCAAGAGAAC
ATAGCGTTGCCTTGGTAGGTCCAGCGGCGGAGGAACTCTTTGATCCGGTCCCTGAACAGGATCTATTCGAGGCACTAAATGAAACC
TTGAAGCTATGGAACTCGCAGCCCGACTGGGCTGGCGATGAGCGAAATGTAGTGCTTACGTTGTCTCGTATTTGGTACAGCGCAGT
AACCGGCAAAATCGCGCCGAAGGATGTCGCTGCCGACTGGGCAATGGAGCGCCTACCGGCCCAGTATCAGCCCGTCTTGCTTGAA
GCTAGACAGGCTTATCTTGGACAAGAAGAAGATCGCTTGGCCTTGCACGCTGATCAGTTGGAAGAATTTGTTCACTACGTGAAAGGC
GAGAGCACCAAGGTAGTCGGCAAATGATGTCTAACAATTCGTTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAACTCGAGCG 

aadA28 

TTAGACATCATGAGGGACTCAGTGACCGCCGAAATTTCGACGCAACTATCCAAGGTGCTTAGTGTTATCGAGCACCATCTGGAACCG
ACGTTGCTTGCCGTACATTTGTACGGCTCCGCAGTGGATGGCGGCCTGAAGCCATACAGTGATATTGATTTGCTGGTTACTGTGACC
GCAAGGCTTGATGACACAACGCGGCGAGCTTTGTTCAACGATCTTTTGGAGGTTTCGGCTTTCCCAGGCGAGAGTGAGATTCTCCG
CGCTATAGAAGTCACCATTGTCGTGCACGAAGACATTATGCCGTGGCGTTATCCAGCCAAGCGCGAACTGCAATTTGGAGAATGGC
AGCGCAATGACATTCTTGCGGGTATCTCCGAGCCAGCCATGATCGACGTTGATCTGGCTATCTTGCTGACAAAAGCAAGAGAACATA
GCGTTGCCTTGGTAGGTCCAGCTGCGGAGGAACTCTTTGATCCCGTTCCTGAACAGGATCTATTCGAGGCACTGAATGAAACCTTG
AAGCTATGGAACTCGCAGCCCGACTGGGCCGGCGATGAGCGAAATGTAGTGCTTACGTTGTCCCGCATTTGGTACAGCGCAATAAC
CGGCAAAATCGCGCCGAAGGATGTCGCTGCCGACTGGGCAATAAAACGCCTACCTGCCCAGTATCAGCCCGTCTTACTTGAAGCTA
AGCAAGCTTATCTGGGACAAAAAGAAGATCACTTGGCCTCACGCGCAGATCACTTGGAAGAATTTATTCGCTTTGTGAAAGGCGAGA
TCATCAAGTCAGTTGGTAAATGATGTCTAACAATTCGTTCAAGCCGACCGCGCTACGCGCGGCGGCTTAACTCCGGCG 

aadA29 

TTAGACATCATGAGTAACGCAGTACCCGCCGAGATTTCAGTACAGCTATCACAGGCACTCAACGTCATCAAGCGTCATCTGGAATCA
ACGTTGCTGGCCGTGCATTTGTACGGCTCCGCACTGGACGGCGGCCTGAAACCATACAGTGATATTGATTTGCTGGTTACTGTGGA
CACACGGCTCGATGAGACCGTAAGGCAAGCTCTGTTCGTCGATCTGTTGGAGATTTCGGCTTTCCCTGGCCAAAGCAAAGTGCTCC
GCGCCTTGGAAGTCACCATCGTCGTGTACAGCGACGTCGTCCCCTGGCGTTATCCGGCCAGACGGGAACTACAGTTCGGAGAGTG
GCAGCGCAAAGACATTCTTGCGGGTATCTTCGAGTCCGCCACAAATGATGTCGATCTGGCGATCCTACTAACAAAAGCAAGGCAAC
ATAACATCGCATTGGTGGGTTCGGCTGCGGAGGATTTCTTCAACCCAGTTCCGGAAAGCGATCTATTCAAGGCACTGGCCGGCACC
CTGAAACTATGGAACTCGCAGCCGGATTGGGAAGGTGACGAGCGTAATGTAGTGCTTACTTTGTCTCGTATTTGGTACAGCGCAGC
AACTGGTAAGATCGCGTCCAAGGATGTCGCTTCTAACTGGGCAATGGAGCGTCTGCCAGCTCAACATCAGCTTGTGCTATTTGAAG
CCCGGCAGGCTTATCTTGGAGATGGAGGAGATTGCTTGGCTTCGCGCGCAGATCAGTTGACGGCGTTCGTTCACTTTGTGAAGTAT
GAAGCCGGCAGACTCCTTGGCTCCACACCAATGATGTCTAACAGTACATTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAATTCAA
GCG 

aadA34 

TTAGACATCATGAGTAACGCAGTACCCGCCGAGATTTCGGTACAGCTATCACTGGCTCTCAACGCCATCGAGCGTCATCTGGAATCA
ACGTTGCTGGCCGTGCATTTGTACGGCTCTGCACTGGACGGTGGCCTGAAGCCATACAGTGATATTGATTTGCTGGTTACTGTGGC
TGCACGGCTCGATGAGACTGTCCGACAAGCCTTGGTCGTAGATCTCTTGGAAATTTCTGCCTCCCCTGGCCAAAGTGAAGCTCTCC
GCGCCTTGGAAGTTACCATCGTCGTGCATGGTGATGTTGTCCCTTGGCGTTATCCGGCCAGACGGGAACTGCAATTTGGAGAATGG
CAGCGCAATGACATTCTTGCAGGTATCTTCGAGCCAGCCACGATCGACATTGATCTGGCTATCTTGCTGACAAAAGCAAGAGAACAT
AGCGTTGCCTTGGTAGGTCCAGCGGCGGAGGAACTCTTTGATCCGGTTCCTGAACAGGATCTATTTGAGGCGCTAAATGAAACCTT
AACGCTATGGAACTCGCCGCCCGACTGGGCTGGCGATGAGCGAAATGTAGTGCTTACGTTGTCCCGCATTTGGTACAGCGCAGTAA
CCGGCAAAATCGCGCCGAAGGATGTCGCTGCCGACTGGGCAATGGAGCGCCTGCCGGCCCAGTATCAGCCCGTCATACTTGAAGC
TAGACAGGCTTATCTTGGACAAGAAGAAGATCGCTTGGCCTCCCGCGCAGATCAGTTGGAAGAATTTGTTCACTACGTGAAAGGCG
AGATCACCAAGGTAGTCGGCAAATAATGTCTAACAATTCGTTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAACTCAAGCG 

aadB 
ANT(2‘’)-Ia 

TTAGGCCGCATGGACACAACGCAGGTCACATTGATACACAAAATTCTAGCTGCGGCAGATGAGCGAAATCTGCCGCTCTGGATCGG
TGGGGGCTGGGCGATCGATGCACGGCTAGGGCGTGTAACACGCAAGCACGATGATATTGATCTGACGTTTCCCGGCGAGAGGCGC
GGCGAGCTCGAGGCAATAGTTGAAATGCTCGGCGGGCGCGTCATGGAGGAGTTGGACTATGGATTCTTAGCGGAGATCGGGGATG
AGTTACTTGACTGCGAACCTGCTTGGTGGGCAGACGAAGCGTATGAAATCGCGGAGGCTCCGCAGGGCTCGTGCCCAGAGGCGG
CTGAGGGCGTCATCGCCGGGCGGCCAGTCCGTTGTAACAGCTGGGAGGCGATCATCTGGGATTACTTTTACTATGCCGATGAAGTA
CCACCAGTGGACTGGCCTACAAAGCACATAGAGTCCTACAGGCTCGCATGCACCTCACTCGGGGCGGAAAAGGTTGAGGTCTTGC
GTGCCGCTTTCAGGTCGCGATATGCGGCCTAACAATTCGTCCAAGCCGACGCCGCTTCGCGGCGCGGCTTAACTCAGGTG 

aphA15 
APH(3’)-XV 

TTAGACCGCTATGACAGTCGCCCTCGACGAAGTATCTGAACTAAAGAATTTGCTTTCACCCTTGTTGGATGAATGCACTTTTGAAGAA
GTTGAGTATGGTCAGTCAGATGCTCGAGTGATTCGAGTTCTATTTCCTGATCGCAATACCGCGTATCTAAAGTACGCCTCCGGATCT
TCTGCTCAAGAAATTCTTCAAGAGCATCAGCGCACTAGATGGCTCAGAACACGAGCTCTCGTACCGGAAGTGATCTCATATGTCTCG
ACTTCAACTGTCACCATCCTGTTGACAAAAGCATTGATTGGCCACAATGCCGCTGACGCCGCAGATGCAGATCCAGTTATTGTTGTT
GCAGAGATGGCACGAGCGTTACGCGACCTCCATTCGATCTCGCCTGACGATTGCCCATTCGACGAAAGGCTCCACCTGCGACTGAA
GCTGGCTTCGGGCCGTTTGGAAGCCGGGTTAGTTGATGAGGAGGACTTTGATCACGCAAGGCAAGGCATGCTGGCGCGGGATGTT
TACGAGCAACTTTTTATACAAATGCCTGGAGCGGAGCAGCTGGTAGTCACACATGGCGACGCCTGTCCCGAGAACTTCATCTTCCAA
GGTAATGCCTTCGTCGGCTTCATAGACTGCGGTCGGGTCGGGCTTGCCGATAAGTATCAAGACCTGGCGCTTGCATCGAGAAACAT
TGACGCGGTATTTGGACCAGAACTCACTAACCAGTTCTTCATCGAGTATGGAGAGCCAAATCCGAACATAGCTAAGATTGAGTACTA
CCGGATCTTGGATGAGTTCTTCTAAGCGCGGTCTAACAATTCGTTCAAGCCGAGATCGCTTCGCGGCCGCGGAGTTGTTCTGTAAAT
TGTCACAACGCCGCGGCCGCAAAGCGCTCCGGCTTAACTCAGGCG 

aphA16 

TTAGCTTGACGCTCCGCGCAGGAAAGAGAAAATGGATAAACTTCCAAATTTCATATTTGAAAACTATAGTGACTTAAAAATTGAACGA
GATACTGAGGGTTGGTCTCCGGCTGAGGTATATAGTGTTACAACCAAGAAAAAAAGGTGGTTTCTAAAAAGAAGTCATACGAGATAC
AATAAAACTACATATAATGTAAGAAGAGAGAAAGAAATTATTGAATGGGCCTATCCAAAGTTTAGAGTACCACAAATAATACATTATGA
AGAAGCAAAAGAATATAATTCATTACTAATGAATCACATTGGCGGCTCTAGCCTTGAAATGCTTGGTCCATCAATTACTTTAGAGAAAT
ATATCGATTACTATGTACAATCCTTAAAATTAATGCAGTCCATTAACATCGAAAACTGTCCATACAATAATTGTATAAAGAATAGAATAA
TCGAATTAGAATATTTATTAGAGAATGATCTAGCAGATATTAATTCAAATAACTGGGAAGAAGATACGCGTGAGCATTTCAGAAACGG
TAAAGACTTATTTAATTATATAGTAAATAATAAGCCAAATGAAGACTTAGTTTTCTCACATGGAGATATGACAAACTCTAACATATTTAT
TGAAAACGAAGAAGTTGGATTTATAGATCTCGGTCGATGTGGATTAGCTGATAAGTGGGTTGATATTGCATTTTGTGTTAGAGATATA
AGAGAAATAAGTAATGAAAATAAATGGATAAAGATGTTATTTGATAAATTAGAAATAGTACCCAATTGGGATAAAATGAGATACTACAT
TTTGTTAGATGAGCTGTTCTAAGAATAAGCTAACAACCGCTTCAACCTGGCAACGCTACTGTCACGGTTTATGCTACTCGCTTCGCTC
GCTTACGCATAAACCGCGCCAGTTTACGCGTTGCAGGTTAAGCGAATG 

sat2 
TTAGGCGTCATATGAAGATTTCGGTGATCCCTGAGCAGGTGGCGGAAACATTGGATGCTGAGAACCATTTCATTGTTCGTGAAGTGT
TCGATGTGCACCTATCCGACCAAGGCTTTGAACTATCTACCAGAAGTGTGAGCCCCTACCGGAAGGATTACATCTCGGATGATGACT
CTGATGAAGACTCTGCTTGCTATGGCGCATTCATCGACCAAGAGCTTGTCGGGAAGATTGAACTCAACTCAACATGGAACGATCTAG



Supplementary Material 

 183 

CCTCTATCGAACACATTGTTGTGTCGCACACGCACCGAGGCAAAGGAGTCGCGCACAGTCTCATCGAATTTGCGAAAAAGTGGGCA
CTAAGCAGACAGCTCCTTGGCATACGATTAGAGACACAAACGAACAATGTACCTGCCTGCAATTTGTACGCAAAATGTGGCTTTACT
CTCGGCGGCATTGACCTGTTCACGTATAAAACTAGACCTCAAGTCTCGAACGAAACAGCGATGTACTGGTACTGGTTCTCGGGAGC
ACAGGATGACGCCTAACAATTCATTCAAGCCGACACCGCTTCGCGGCGCGGCTTAATTCAGGAG 

arr2 

TTATGCAGCCAAATCCCAACAATTAAGGGTCTTAAAATGGTAAAAGATTGGATTCCCATCTCTCATGATAATTACAAGCAGGTGCAAG
GACCGTTCTATCATGGAACCAAAGCCAATTTGGCGATTGGTGACTTGCTAACCACAGGGTTCATCTCTCATTTCGAGGACGGTCGTA
TTCTTAAGCACATCTACTTTTCAGCCTTGATGGAGCCAGCAGTTTGGGGAGCTGAACTTGCTATGTCACTGTCTGGCCTCGAGGGTC
GCGGCTACATATACATAGTTGAGCCAACAGGACCGTTCGAAGACGATCCGAATCTTACGAACAAAAAATTTCCCGGTAATCCAACAC
AGTCCTATAGAACCTGCGAACCCTTGAGAATTGTTGGCGTTGTTGAAGACTGGGAGGGGCATCCTGTTGAATTAATAAGGGGAATGT
TGGATTCGTTAGAGGACTTAAAGCGCCGTGGTTTACACGTCATTGAAGACTAGTCCTTTGCATAACAAAGCCATCAAACCGGACGCC
AGAGATTCCGCGCCTGTTGCGCATGGCTTCGCCATTTTATGCGCAATAGGCGCGCCACCCTGTCGCCGTTTATGGCGGCG 

arr5 

TTAGGCAACAAACCAATCTCTTTCGCTACATGGAACAAACGATGACGGTAGACTGGATCCCCATTTCGCACGACAACTACCATCAAG
TGCGTGGCCCGTTTTATCACGGAACAAAAGCCGAACTCGCCATTGGCGACTTAATTTCAACCGGATTTATTTCTCACTTTGAGCGGG
ACAGAGCACTAAAGCATGTGTACTTTTCCGCGCTGATGGAGCCAGCAATCTGGGGGGCCGAGCTCGCTGTAGCACTCTCTGGCTCT
GACGGGCCAGGCCATATTTACATCATTGAGCCAACCGGCCCGTTTGAAGACGACCCCAATCTCACAAACAAACGATTCCCTGGCAA
TCCAACACAGTCCTATCGCACATGCCACCCACTTAAAATTGTTGGCATACTGCGGGAGTGGGAGCGCCATTCTCCTGAAGCATTGAA
GACCATGCTAGATTCTCTGGCAGACCTCAAGCGACGCGGCTTGGCCATCATTGAAGAATGAAATTGTTGCCTAACAATTCATTCAAG
CCGACGCCGCTTCGCGGCGCGGCTTAATTCAGGCG 

arr6 

TTAGGCAGCAACCCCATTCTCGCCCCATGGAACCTAAGAAATGTCGAGTGACTGGACTCCCATCTCACATGAGAATTGCCAGCAGG
TGCGTGGGCCGTTCTATCACGGCACCAAAGCCCATCTATCGATTGGCGACTTGATAACAACTGGGCATCTCTCCCACTTTGAAGATG
GACGCGCTCTTAAACACGTCTACTTTTCAGCTTTGATGGAGCCTGCCATTTGGGGGGCGGAACTTGCAATGTCGTTGTCACGCCTA
GATGGCCGTGGCTACATATACATCGTCGAACCAACTGGACCGTTTGAGGACGACCCGAATCTTACGAACAAAAGATTTCCTGGAAAT
CCAACAAAGTCCTATCGCACGTGCGATCCGCTACGAATTGTCGGGTCAGTCGAAGACTGGCAAGGGCATCCCGCTGATGTGCTGCA
ACAGATGTTGGAGTCTTTAGAGGACCTAAAGCGCCGTGGTCTTGCCATCATCGAGGATTAGAACTGCTGCCTAACAATTCATTCAAG
CCGACGCCCCTTCGGGGCGCGGCTTAATTCAGGCG 

arr7 

TTAGGCAGCAAACCACCTTCCTCGCTAGACGGAACCAACGAATGCCGAATGACTGGATTCCCACCTCGCACGAAAACTGCTCGCTC
GTGCCGGGGCCGTTCTACCACGGCACCAAAGCAAAACTCGCAATAGGTGACTTGCTTTCGCCTGGACACCCGTCTCACTTTGAGCA
AGGCCGTAGGCTCAAACACATCTATTTTGCCGCACTGATGGAGCCAGCCATCTGGGGTGCTGAGCTTGCAATGTCATTGTCACGCC
AAGAGGGGCGCGGTTACATTTACATTGTTGAACCGCTCGGGCCGTTTGAGGACGACCCAAACCTTACAAACAAAAAATTTCCGGGC
AATCCAACCAAGTCCTACCGCACCAGTGAGTCGCTACGGATTGTGGAGGTAGTAGAGGACTGGCAAGGCCACTCACCGGATGTGC
TGCAGGGCATGTTGGCATCACTGGAGGATCTTCAGCGTCGCGGCCTCGCAATCATTGAGGACTAGAAATTGCTGCCTAACAATTCG
TTCAAGCCGAACTTGCTTCGTTCCACCAAAGCCATGGCAGATTAAGCTTGCCATGGCTTTGGCTCCACTACGCAAGTCGGCTTAACT
CAGGCG 

arr8b 

TTAGGCTGCCAAACCTTTTTCCAACCTTCGGAGTGAAGAATGATGAAAGATTGGGTTCCAATCACGCATGCAAATTGTAAAGATATGC
AGGGACCGTTTTATCATGGTACCAAAGCTAAATTATCGGTAGGTGAACTCCTAACAACTGGGTTCAACACTCATTTTGAAGAGGGTC
GCACACTCAAACACGTTTATTTTTCAGCTATGCTTGAGCCAGCAATTTGGGGCGCTGAACTTGCTGTTTCACTGTCAGGTCTAGATG
GCCGGGGATACATATACTTAGTTGAACCAACTGGACCTTTTGAGGATGACCCTAATCTTACCAATAAGAAATTTCCAGGGAATCCAAC
AATGTCCTATCGAACTTCCGAACCCCTCCAGATTGTTGGGCTCGTTGAAGAGTGGGAGGGACACTCTGCTGAAGCCCTGAAAACGA
TGCTGGATTCCTTGGAGAATCTAAAGCGCAATGGTCTTCATGTCATATATGATTAAGCGGAGCCTGCCTAACAAATCGTTCCAGCAA
CGTGCGCTTCGCGCACTCGACAGTCTGTAAGTCGCCTTTTTGTGGTTTTGCTACGCAAAAGTATTCCACAAAAAACCAACTTACAGA
CTGCGGCTGAACTTAGCG 

catB2 

TTAGGCGACGCGTGGAGTCGCTCTAGAATTTTCGGGTACAAATTTTATGACGAATTATTTTGAGAGTCCCTTCAAAGGGAAGCTTCT
GACTGAGCAGGTGAAGAATCCGAACATCAAGGTAGGGCGGTATAGCTACTATTCCGGCTATTACCATGGGCACTCGTTTGATGATT
GTGCTCGCTACCTTCTACCAGACCGTGATGACGTTGATCAGCTGATTATCGGCAGCTTCTGCTCCATCGGATCAGGCGCAGCTTTTA
TTATGGCTGGGAATCAAGGCCACCGATATGATTGGGTCTCTTCTTTCCCTTTCTTCTACATGAACGAGGAGCCCGCGTTTGCAAAAT
CAGTCGATGCATTCCAGCGGGCTGGCGACACAGTTATAGGAAGTGATGTGTGGATCGGTTCGGAGGCCATGATCATGCCCGGGAT
CAAGATCGGGCATGGAGCGGTGATAGGTAGCCGCGCTTTGGTTGCCAAAGACGTGGAACCCTACACCATAGTGGGGGGAAACCCT
GCAAAGTCGATTAGGAAGCGCTTTTCTGAAGAAGAAATTTCTATGCTTTTAGATATGGCTTGGTGGGATTGGCCGCTGGAACAAATC
AAGGAAGCAATGCCTTTTCTTTGTTCGTCTGGCATTGCCAGCCTGTATCGTCGCTGGCAAGGCACAAGCGCCTAACAATACGCTACA
CACGGACAAATTACTCGCTGCGCTCCTAATTTGCCGGTGAGCGTGGCG 

catB3 

TTAGACGGCAAAGTCACAGACCGCGGGATCTCTTATGACCAACTACTTTGATAGCCCCTTCAAAGGCAAGCTGCTTTCTGAGCAAGT
GAAGAACCCCAATATCAAAGTTGGGCGGTACAGCTATTACTCTGGCTACTATCATGGGCACTCATTCGATGACTGCGCACGGTATCT
GTTTCCGGACCGTGATGACGTTGATAAGTTGATCATCGGTAGTTTCTGCTCTATCGGGAGTGGGGCTTCCTTTATCATGGCTGGCAA
TCAGGGGCATCGGTACGACTGGGCATCATCTTTCCCGTTCTTTTATATGCAGGAAGAACCTGCATTCTCAAGCGCACTCGATGCCTT
CCAAAAAGCAGGTAATACTGTCATTGGCAATGACGTTTGGATCGGCTCTGAGGCAATGGTCATGCCCGGAATCAAGATCGGGCACG
GTGCGGTGATAGGCAGCCGCTCGTTGGTGACAAAAGATGTGGAGCCTTACGCTATCGTTGGCGGCAATCCCGCTAAGAAGATTAAG
AAACGCTTCACCGATGAGGAAATTTCATTGCTTCTGGAGATGGAGTGGTGGAATTGGTCACTGGAGAAGATCAAAGCGGCAATGCC
CATGCTGTGCTCGTCTAATATTGTTGGCCTGCACAAGTATTGGCTCGAGTTTGCCGTCTAACAATTCAATCAAGCCGATGCCGCTTC
GCGGCACGGCTTATTTCAGGCG 

catB5 

TTAGACGGCAAGAAAAGGTTCCACGAACTCTGATGAAAAACTACTTTGACAGCCCTTTCAAAGGGGAGCTTCTTTCTGAGCAAGTGA
AAAATCCAAACATCAAAGTAGGCCGTTATAGCTATTACTCTGGCTACTATCACGGCCACTCATTTGATGAATGCGCGCGATACTTGCA
TCCAGATCGTGATGACGTTGATAAATTGATCATTGGCAGCTTTTGTTCTATAGGAAGCGGGGCTTCCTTCATCATGGCTGGCAATCA
GGGGCATCGGCATGACTGGGCATCATCCTTCCCCTTCTTCTATATGCAAGAGGAACCTGCTTTCTCAAGCGCACTCGATGCCTTCCA
AAGAGCAGGTGATACCGCCATTGGCAATGATGTCTGGATAGGCTCGGAGGCAATGATTATGCCCGGAATCAAAATTGGAGACGGTG
CCGTGATAGGTAGTCGCTCGTTGGTGACAAAAGATGTAGTGCCTTATGCCATCATCGGAGGAAGTCCCGCAAAGCAAATTAAGAAG
CGCTTCTCCGATGAGGAAATCTCATTGCTCATGGAGATGGAGTGGTGGAACTGGCCACTGGATAAAATTAAGACAGCAATGCCTCT
GCTGTGCTCGTCAAATATTTTTGGTCTGCATAAGTATTGGCGCGAGTTTGTCGTCTAACAATTCATTCAAGCCGACGCCGCTTCGCG
GCACGGCTTAATTCTGGCG 

catB6 

TTAGACGGCAGAATAAATTTTTGCGATCTCTTATGGAAAATTACTTTGACAGTCCCTTCAAAGGGAAACTACTTTCAGAGCAAGTGAC
TAACCGCAACATCAAAGTTGGTCGGTACAGCTACTACTCTGGTTACTATCACGGGCATTCATTTGATGACTGCGCACGATACTTGCT
CCCAGACCGTGATGACGTTGACAAACTAATCATCGGCAGCTTTTGCTCCATCGGAAGCGGGGCTTCTTTCATCATGGCGGGCAATC
AGGGTCACCGGCATGACTGGGTAACATCTTTCCCTTTCTTCTACATGCAAGAAGAGCCAGCTTTTTCAAGTTCAACGGACGCCTTTC
AAAAGGCCGGTGACACCATCGTCGGCAATGATGTCTGGATAGGATCAGAGGCAATGATTATGCCCGGCATCAAGATTGGAGATGGC
GCGGTAATAGGCAGCCGATCGTTGGTGACGAGAGATGTAGAACCCTATACCATCATTGGCGGAAACCCTGCAAAGCAAATTAAAAA
GCGATTCTCTGACGAGGAGATTTCATTACTCATGGAAATGGAGTGGTGGAACTGGCCGTTAGATAAAATCAAAACAGCTATGCCCCT
TCTCTGCTCTTCAGACATTTTTGGTCTGCACAGGCATTGGCGTGGGATTGCCGTCTAACAAGCGCCAGCACCATCGCCTGCGGCGC
TGGACTCGCAACAAGTTGCTCGCCCGTGTGGCGGGCG 

catB10 

TTAGACGGCAAGAAATACGCTCCGTGAAAATCCCATGACCAACTATTTTGAAAGTCCATTTAAAGGCAAACTGCTGGCCGACCAGGT
AAAGAACCCGAACATCAAAGTCGGACGGTATAGCTATTATTCCGGCTATTACCATGGCCATTCGTTTGACGAGTGCGCTCGCTTTCT
CTTGCCAGATCGCGATGACATCGACCAACTGATCGTTGGTAGCTTCTGTTCCATCGGCACGGGCGCCTCCTTCATCATGGCCGGAA
ATCAGGGGCACCGTTATGACTGGGCGTCTTCTTTTCCCTTCTTCTACATGAAAGAGGAGCCAGCATTCTCGGGCGCACTTGATGCAT
TCCAAAAAGCCGGTGACACAGTCATCGGAAGTGATGTCTGGATAGGCTCTGAGGCCATGATCATGCCCGGCATCAACGTCGGTCAT
GGCGCTGTGATTGGAAGCCGCGCTTTGGTCACGAAAGATGTGGAGCCGTACACTATCGTTGGCGGAAATCCCGCCAAACCGATCA
AGAAACGCTTCTCCGACGAGGAGATCGCCATGCTTTTGAAAATGAATTGGTGGGATTGGCCAACTGAAAAAATTGAGGAAGCAATGC
CTTTGCTATGCTCATCCAACATCGTTGGGCTGCATCGATACTGGCAAGGCTTTGCCGTCTAACAATTTATTCAAGCCGACTCCGCTT
CGCGGCGCGGCTTAATTCAGGCG 



Supplementary Material 

  184 

catB11 

TTAGGCAACGGTGTTGTTGCAGATTTTTGGTGTGGAATTATATGAAGAACTATTTTGAGAGCCCGTTTAAGGGAAAACCTCTCGTCGA
ACAGGTAAAGAACCCAAATATCAAGGTGGGCCGTTATAGCTATTATTCAGGCTATTACCACGGGCATTCATTTGATGATTGCGCTCG
CTACCTCCTGCCTGATCGTGATGACGTTGATAAATTGATAATTGGAAGCTTTTGCTCCATAGGGACGGGTGCATCTTTTATCATGGCT
GGAAATCAAGGTCACCGATATGATTGGGTTACATCATTCCCTTTTTTCTATATGAATGAGGAACCGGCATTTTCGGAATCAGTTGATG
CTTTCCAGGCGGCAGGTAATACCGTCATAGGAAGCGACGTGNGGATTGGCTCTGAAGCAATGATTATGCCTGGAGTAAAGGTTGGC
CATGGAGCGGTAATTGGCAGCCGGGCTTTGGTTACCAAAGATATAGAGCCATACACAATTGTTGGTGGCAACCCTGCAAAAGAGAT
AAAGAAGCGCTTTTCAGAACAGGAAATTTCAATGTTGCTAGATATGAAGTGGTGGGATTGGCCGTTGGAGCAAATTAAAGAAGCAAT
GCCTCTTTTGTGCTCGTCTGATATTGCAGGCTTGTACCATTTTTGGCAGCGTTCAAGTGCCTAACAAACGGCTGTTGTCGCCCCTTC
GGGGCTGGGACGGCCTTTCCGCCGCTTTGCGGCTACAAGTCCGCCCCAAAGCCGGGCG 

cmlA1 

TTGGGCGCACAATAAGGCTCCTTGCAGAGTTGCTTGAAAGTTGTTACGATTCAAATTCAATCATGAGATAGTCAGCAGATGAGCACT
TCCAAGAACGCAGACAAGTAAGCCGCAGCAACCTTCATTTTTCGGTTGTTGCGGCGTTCTCATGAATCCTTTTGCTCTACGGGAGCG
CCGCCAAATCCTTTGTTCAAGGAGATGGTTTCGTGAGCTCAAAAAACTTTAGTTGGCGGTACTCCCTTGCCGCCACGGTGTTGTTGT
TATCACCGTTCGATTTATTGGCATCACTCGGCATGGACATGTACTTGCCAGCAGTGCCGTTTATGCCAAACGCGCTTGGTACGACAG
CGAGCACAATTCAGCTTACGCTGACAACGTACTTGGTCATGATTGGTGCCGGTCAGCTCTTGTTTGGACCGCTATCGGACCGACTG
GGGCGCCGCCCCGTTCTACTGGGAGGTGGCCTCGCCTACGTTGTGGCGTCAATGGGCCTCGCTCTTACGTCATCGGCTGAAGTCT
TTCTGGGGCTTCGGATTCTTCAGGCTTGTGGTGCCTCGGCGTGCCTTGTTTCCACATTTGCAACAGTACGTGACATTTACGCAGGTC
GCGAGGAAAGTAATGTCATTTACGGCATACTCGGATCCATGCTGGCCATGGTCCCGGCGGTAGGCCCATTGCTCGGAGCGCTCGT
CGACATGTGGCTTGGGTGGCGGGCTATCTTTGCGTTTCTAGGTTTGGGCATGATCGCTGCATCTGCAGCAGCGTGGCGATTCTGGC
CTGAAACCCGGGTGCAACGAGTTGCGGGCTTGCAATGGTCGCAGCTGCTACTCCCCGTTAAGTGCCTGAACTTCTGGTTGTACACG
TTGTGTTACGCCGCTGGAATGGGTAGCTTCTTCGTCTTTTTCTCCATTGCGCCCGGACTAATGATGGGCAGGCAAGGTGTGTCTCAG
CTTGGCTTCAGCCTGCTGTTCGCCACAGTGGCAATTGCCATGGTGTTTACGGCTCGTTTTATGGGGCGTGTGATACCCAAGTGGGG
CAGCCCAAGTGTCTTGCGAATGGGAATGGGATGCCTGATAGCTGGAGCAGTATTGCTTGCCATCACCGAAATATGGGCTTTGCAGT
CCGTGTTAGGCTTTATTGCTCCAATGTGGCTAGTGGGTATTGGTGTCGCCACAGCGGTATCTGTGGCGCCCAATGGCGCTCTTCGA
GGATTCGACCATGTTGCTGGAACGGTCACGGCAGTCTACTTCTGCTTGGGCGGTGTACTGCTAGGAAGCATCGGAACGTTGATCAT
TTCGCTGTTGCCGCGCAACACGGCTTGGCCGGTTGTCGTGTACTGTTTGACCCTTGCAACAGTCGTGCTCGGTCTGTCTTGTGTTTC
CCGAGTGAAGGGCTCTCGCGGCCAGGGGGAGCATGATGTGGTCGCGCTACAAAGTGCGGAAAGTACATCAAATCCCAATCGTTGA
GAGAATGTGGCAAGCTATCGCCCAACAAATCGCTGCAGCCGACCCAAAACCGCTACGCGGTTTCGGTCGGCTGAGCTCAGGCG 

cmlA2 

TTGGGCGAACAAAAAGATTCTTCGCGGAATTGCTTGAGAATTGTTACGATTCAAATTCAGCCATGAGATAGTCAGCAAATGAGCACTT
CCAAGAAAGCAGACAAGTAAGCCGCAGCAACCTTCATTTTTCGGTTGTTGCGGCGTTCTCATGAACCCTTTTGCTCTACGGGAGCGC
CGCCAAATCCTTCGTTCAAGGAGATGGTTTCGTGCGCTCAAAGAACTGTAATTGGCGGTATTCCCTTGCCGTCACTGTGTTGTTGTT
ATCACCTTTCGATTTACTGGCATCACTCGGCATGGACATGTACTTGCCAGCGGTGCCTTTCATGCCACATGCTCTTGGTACGACAGC
GGGCACAATTCAGCTTACGCTGACAACGTATTTGGTCATGATAGGGGCCGGTCAGCTTTTGTTTGGGCCACTGTCGGACCGGCTGG
GACGTCGTCCCGTGCTACTGGCGGGCGGTGCCGCCTACGTTGCGGCCTCAATCGGCCTCGTCGTCACGTCATCGGCTGGAGTATT
TCTGGGTTTTCGGATTCTCCAAGCTTGTGGTGCCTCGGCATGCCTTGTTGCCACATTTGCAACAGTGCGTGATATCTACGCAGGTCG
CAAGGAAAGTAACGTCATCTACGGCTTGCTTGGCTCTATGCTTGCTATGGTTCCGGCGATAGGCCCATTGCTGGGAGCGGTCATAG
ACACCTGGTTCGGGTGGCGGGCGATCTTTGCGTTCTTGGGATTGGGAATGATCGCTGCATTGACAGCAGCGTGGCGGCTCTGGCC
TGAGACCCGGGTGCAGCGACCAGCAGCTTTGCAATGGTCACAACTTCTGCTTCCCATCAAGCACCTTAACTTCTGGTTGTACACAGT
GTGTTATGCCGCAGGAATGGGCAGCTTCTTCGTCTTCTTCTCCATAGCGCCCGGATTGATGATGGGTAGGCAAGGCATGTCCCAGT
TTGGCTTCAGTCTGTTGTTCGCAACAGTGGCAATTGCGATGATGCTTGCGGCCCGCTTCATGGGGCGCGTAATCGCCAAGTGGGGC
AGCCTGAGTGCCTTGCGAATGGGGATGGGCTGCCTGATAGCAGGCGCAGTCTTGCTTGTCATCACCGAGCTATGGATTCCGCAGT
CCGTGTTGGGCTTTATTGCCCCAATGTGGCTAGTGGGCGTCGGCGTCGCGACAGCGGTATCCGTTGCACCCAATGGTGCGCTTCG
AGGGTTCGACCATATTGCAGGAGCCGTTACGGCAGTCTACTTCTGCTTGGGGGGGCTGCTGCTGGGGAGTGTTGGAACGCTCATC
ATTTCGCTGTTGCCGCGCGACACGGCCTGGCCAGTTATCGCGTATTGTTTGGTTCTTGCAACAATCGTGCTTGGACTGTCGTGTGTT
TCCCGAGCGAGAGACCTTCGCGGTCACGGGGAGTATGATGCGGTTGCACGCACATAGTGCGGAAAACACACCAAATCCCGATCAC
TGGGAATATGGCAAGTTGTCGCCCAACAAATCGCTGCAGCCGACCCAAAACCGCTACGCGGTTTCGGTCGGCTGAGCTCAAGCG 

ereA1 

TTATGCTCTGTGAGCCGGGTTATTGGCGAAGCGAACGTATGACGATTACAGCAATAAACGCAAAGGTAAAAAAATGACATGGAGAAC
GACCAGAACACTTTTACAGCCTCAAAAGCTGGACTTCAATGAGTTTGAGATTCTTACTTCCGTAATTGAGGGCGCCCGAATTGTCGG
CATTGGCGAGGGCGCTCATTTTGTCGCGGAGTTTTCACTGGCTAGAGCTAGTCTTATCCGCTATTTGGTCGAAAGGCATGAGTTTAA
TGCGATTGGTTTGGAATGTGGGGCGATTCAGGCATCCCGGTTATCTGAATGGCTCAACTCAACAGCCGGTGCTCATGAACTTGAGC
GATTTTCGGATACCCTGACCTTTTCTGTGTATGGCTCAGTGCTGATCTGGCTGAAATCATATCTCCGCGAATCAGGAAGAAAACTGC
AGTTAGTCGGAATCGACTTACCCAACACCCTGAACCCAAGGGACGACCTAGCGCAATTGGCCGAAATTATCCAGCTCATCGATCAC
CTCATGAAACCGCACGTTGATATGTTGACTCACTTGTTGGCGTCCATTGATGGCCAGTCGGCGGTTATTTCATCGGCAAAATGGGGG
GAGCTAGAAACGGCTCGGCAGGAGAAAGCTATCTCAGGGGTAACCAGATTGAAGCTCCGCTTGGCGTCGCTTGCCCCCGTCCTGA
AAAAACACGTCAACAGCGATTTGTTCCGAAAAGCCTCTGATCGAATAGAGTCGATAGAGTATACGTTGGAAACCTTGCGTATAATGA
AAACTTTCTTCGATGGTACCTCTCTTGAGGGAGATACTTCCGTACGTGACTCGTATATGGCGGGCGTAGTAGATGGAATGGTTCGAG
CGAATCCGGATGTGAAGATAATTCTGCTGGCGCACAACAATCATCTACAAAAAACTCCAGTCTCCTTTTCAGGCGAGCTTACGGCTG
TTCCCATGGGGCAGCACCTCGCAGAGAGGGTGAATTACCGTGCGATTGCATTCACCCATCTTGGACCCACCGTGCCGGAAATGCAT
TTCCCATCGCCAAAAAGTCCTCTTGGATTCTCTGTTGTGACCACGCCTGCCGATGCAATCCGTGAGGATAGTATGGAACAGTATGTC
ATCGACGCCTGTGGTACGGAGAATTCATGTCTGACATTGACAGATGCCCCCATGGAAGCAAAGCGAATGCGGTCTCAAAGCGCCTC
TGTAGAAACGAAATTGAGCGAGGCATTTGATGCCATCGTCTGTGTTACAAGCGCCGGCAAGGACAGCCTGGTTGCCCTATAGGAAA
CCGGAAATGAAAATGAGGGAGCATAACCTGCGAATCCACCGGACGGTTTTCAACCGCCGGTGATCAGCGCG 

ereA2 

TTATGCTCTGTGAGCCTGGTTATTGGCGAAGCGAAAGTATGACGATTTCAGCAATAAACGCAAAAGGATAAAAAAATGACATGGAGA
ACGACCAGAACACTTTTACAGCCTCAAAAGCTGGAGTTCAATGAGTTTGAGATTCTTAATCCCGTAGTTGAGGGCGCCCGAATTGTC
GGCATTGGCGAGGGTGCTCACTTTGTCGCGGAGTTCTCACTGGCTAGAGCTAGTCTTATTCGCTATTTTGTCGAGAGGCATGATTTT
AATGCGATTGGTTTGGAATGTGGGGCGATTCAGGCATCCCGGCTATCTGAATGGCTCAACTCAACAGCCGGTGCTCATGAACTTGA
GCGATTTTCGGATACCCTGACCTTTTCTTTGTATGGCTCAGTGCTGATTTGGGTTAAATCATATCTACGCGAATCAGGAAGAAAACTG
CAGTTAGTCGGAATCGATTTACCCAACACCTTGAATCCAAGGGACGACCTAGCGCAATTGGCCGAAATTATCCAGGTCATCGACCAC
CTCATGAAACCCCACGTTGATGCGCTGACTCAGTTGTTGACGTCCATTGATGGCCAGTCGGCGGTTATTTCATCGGCAAAATGGGG
GGAGTTGGAAACGGCTCAGCAGGAGAAAGCTATCTCAGGGGTAACCAGATTGAAGCTCCGTTTGGCGTCGCTTGCCCCTGTCCTGA
AAAATCACGTCAACAGCGATTTTTTCCGAAAAGCCTCTGATCGAATAGAGTCGATAGAGTATACGTTGGAAACCTTGCGTGTAATGAA
AGCTTTCTTCGATGGTACCTCTCTTGAGGGAGATACTTCCGTACGTGACTCGTATATGGCGGGCGTGGTGGATGGAATGGTTCGAG
CGAATCCGGATGTAAGGATAATTCTGCTGGCGCACAACAATCATTTACAAAAAACTCCAGTTTCCTTTTCAGGCGAGCTTACGGCTG
TTCCCATGGGACAGCATCTCGCAGAGAGGGAGGAGGGGGATTACCGTGCGATTGCATTCACCCATCTTGGACTCACCGTGCCGGA
AATGCATTTCCCATCGCCCGACAGTCCTCTTGGATTCTCTGTTGTGACCACGCCTGCCGATGCAATCCGTGAGGATAGTGTGGAACA
GTATGTCATCGATGCCTGTGGTAAGGAGGATTCATGCCTGACATTGACAGATGACCCCATGGAAGCAAAGCGAATGCGGTCCCAAA
GCGCCTCTGTAGAAACGAATTTGAGCGAGGCATTTGATGCCATCGTCTGCGTTCCCAGCGCCGGCAAGGACAGCCTGGTTGCCCTA
TAGGAAACCAGAAATGAAATGAAGGAGCATAACCTGCCAATCCACCGGACGGTTTTCAACCGCCGGTGATCAGCGCG 

ereA3 

TTAGAATCACTGAAAACTAAAAATAATTGCACAATCCGGCTCTACAGGTGGCAGAAGAAGTTATGACGGCATTGAATGGTTGAAAAAA
TTAAGACAATTCAGGACAATGAAATTATTGGCGAAGCGAACGCATGACAGCAATGAGCGCAAAGGCTAAAAAAATGACATGGAGAAC
TACCAGAACACTTTTACAGCCTCAAAAGCTGGACTTCAATGAGTTTGAGATTCTTACTCCCCTGGTTGAGGGCGCCCGAATTGTCGG
CCTTGGCGAGGGCGCTCACTTTGTCGCGGAGTTTTCACTGGCTAGAGCTAGTCTTATTCGCTATTTGGTCGAGAGGCATGATTTTAA
TGCGATTGGTTTGGAATGTGGGGCGATTCAGGCATCCCGGCTATCTGAATACCTCAACTCAACAGCCGGTGCTCATGAACTTGAGC
GATTTTCGGATCCACTGACCTTTTCTTTGTATGGCTCAGTGCTGATTTGGATTAAATCATATCTACGCGAATCAGGAAGAAAACTGCA
GTTAGTCGGAATCGATTTACCCAACACCTTGAATCCAAGGGACGACCTGGCACAATTGGCCGAAATTATCAAGGTCATCGATCACCT
CATTAAACCGCATGTTGATGAGCTGACTCACTTGTTGGCATCCATTGATGGTCAGTCGGCGGTTATTTCATCGGCAAAATGGGGGGA
GATGGAAACGGCTCAGCAGGAGAAAGCTATCTCAGGGGTAACCAGATTGAAGCTACGTTTGGCATCTCTTGCCCCTGTCCTGAAAA



Supplementary Material 

 185 

AACATGTCAACAGCGATTTGTTCCGAAAAGCCTCTGATCGAATAGAGTCGATAGAGTATACGTTGGAAACCTTGCGTATAATGAGAA
CTTTCTTCGATGGTACCTCTCTTGAGGGAGATACTTCCGTACGTGACTCGTATATGGCGGGCGTAGTGGATAGAATGGTTCGAGCAA
ATCCGGATGTGAAGATAATTCTGCTGGCGCACAACAATCATTTACAAAAAACTCCAGTCTCCTTTTCGGGCGAGCTTACGGCTGTTC
CCATGGGGCAGCACCTCGCAGAGAGGGAGGAGGAGGATTACCGTGCGATTGCATTCACCCATCTTGGATCCACCGTGCCGGAAAT
GCAATTCCCATCGCCCGGCAGTCCTCTTGGATTCTCTGTTGTGACCACGCCTGCCGATGCAATCCGTGAGGATAGTATGGAACAGT
ATATCATCGATGCCTGTGGTACGGAGGATTCATGTCTGACATTGACAGATGCCCCCATGAAAGCAAAGCGAATGCGGTCCCAAAGC
GCCTCTGTAGAAACGAATTTGAGCGAGGCATTTGATGCCATCGTCTGCGTCCCAAGCGCCGGCAAGGACGGCCTGGTTGACCTATA
GTAAACCGGTAGATGAAAATGATGGAGAATAACCTGCCAATCCACCGGACGGTTTTCAACCGCCGGTGATCAGCGCG 

fosC2 

TTATGTTCGCTGAGGAGAGTCAGTGTTACGAGGATTGAATCATATTACTATTGCAGTAAGTGACCTTGAACGTTCCGTGGAGTTCTAT
ACGCGTCTATTAGGAATGAAGGCACATGTCCGCTGGGATAGTGGGGCATATCTGAGCTTGGAGGCTACTTGGATTTGCTTGTCTTGT
GACGAAGTGCATCCGAGCCAAGATTACTGTCACATCGCGTTTGATGTTTCCGAAGAGAATTTCGAACCAGTTACTAAAAAGCTTCGC
GAAGCACATGTCGTTGAATGGAAACAAAATAGAAGCGAAGGACTTTCTTTATACTTGCTCGATCCTGACGGCCATAAATTGGAAATC
CATAGCGGTAGCCTACAAAGTCGTTTGGAATCGTTGAAGTCTAAACCCTATCAAGGGTTAGTATGGCTATAAGCAAACATAACAAGT
CAATTAACTACGCGCCTGCGGCGCCGGACGTGCTAGCGCACGCCGGTTATTTCGGGCG 

fosE 

TTAGCGCCCATGGAAGGTATCAGCCACATCACGCTTATTGTCCGCGACCTCTCGCGCATGACCACCTTCCTTTGCGATGGTCTCGG
TGCGCGAGAGGTTTATGACAGTGCTGGCCACAATTACTCGCTTTCCCGCGAGAAATTCTTTGTCCTTGGTGGCGTTTGGTTGGCCG
CTATGGAGGGAGTGCCGCCATCTGAGCGCTCCTATCAGCATGTCGCCTTTCGGGTGAGTGAGTCAGATCTTGCCGTATATCAGGCA
AGACTTGGGTCGCTGGGCGTGGAGATTCGCCCACCCAGGCCACGCGTGAATGGAGAGGGGCTGTCCCTGTACTTCTATGATTTTG
ACAACCATCTGTTTGAGCTGCACACCGGCACATTGGAGCAGCGCCTTGCCAGGTACGGAGCTGGGCGCTAACAATTCATTCAAGCC
GAAGCCGCTTCGCGGCTCGGCTTAATTCAGGCG 

fosF 

TTACGTTTTCTCAAGGTTCTCTGCATGATTACCGGCATCAATCACATCACATTTTCGGTTCGGGACCTGCGGGCATCGATTGAGTTCT
ACCGTGATCTTCTGGGAATGAAGTTGCACGTATTCTGGGACACAGGTGCTTATCTCACTGCAGGCAATACGTGGTTATGTTTGAGTT
TGGGGCAGCCCGAACCCGCCAAGGACTACACACACGTCGCTTTCAGTGTCCGCGAAGGGGAGCTCCTGGAGTTGCGAGCTAAACT
AAAGCAGGCTGGCGTTGAAGAGTGGAAGCAGAATACCAGTGAGGGTGACTCCATCTATTTACTTGACCCAAATGGGCATCGCCTTG
AACTGCATTGCGGAACACTGGCCACTCGCTTAGCTGAGCTGGAAAGCTCGCCTTATAAGGGGCTGGTGTGGAGCTGAAACGTAACA
ATCGGCTGTTGTCGCCCCTTCGGGGCTGGGACGGCCTTTTCGCCGCTGCGCGGCTACAAGTCCGCCCCAAAGCCGGGCG 

fosG 

TTATGTTTGTTAAGGTAGATTTGTGCTCCGAGGATTGAACCACATCACCATCGCTGTAAGCGATTTAGGCCGTTCTCTCGCCTTTTAT
ACTGATATCGTCGGTATGCTCGCTCACGTACGCTGGGATAACGGTGCTTACCTTAGTCTAGGCGGTGTTTGGTTTTGTCTTTCCTGT
GACAAGGTGATGCCAAGTAAGGATTATTCTCATATTGCCTTAGATATTTCAGAAGATGACTTTGCATCATTTTTGGAGAAACTGAGGA
GAGCCGATGTCACTGAGTGGAAACAAAATTCAAGTGAAGGCTATTCGGTGTATTTCTTAGATCCTGATGGAAATAAACTAGAAGCGC
ATAGCGGCTCGTTACAATCTCGTTTAAGTTCTTTAAAAGACAAACCTTATCCGGGCTTAGTATGGCTTTAAACAAACATAACAAGCGC
CATCAAAACGCCGCTTCGCGGCTCGACTCGCAACAAGTTGCTCGCGTTTGTGGCGGGCG 

fosH 

TTAAAGCTCATCATGGGAATTCTTGGCATTAGTCATCTGACATTCGTGGTCCGCGATGTAGAGCGCACTGCGAGACTGGTCTGCGAA
GGACTTGGGGCGGAAGAGGTATACGACAGCAAAGCCAAAAACTTTTCGCTGTCACGAGAGAAGTTTTTTCTCTTGGGCGGCGTATG
GCTTGCTTTCATGGAAGGAGTGCCATCGGAGCGGTCCTATCGGCACGTCGCTTTTGAAGTGACCGAAGAGGAAATTGCAAGATATG
AGGCTAGCCTTAGAAACCTCGGTGTGGAAGTCAGAGAGCCGCGGCCAAGAGTGGCTGGAGAGGGGCTGTCACTGTATTTCTACGA
CTATGACAACAACTTATTCGAGCTACATGCGGGAACACTGGCGCAACGACTTGAAAGGTATACGCAATGAGTTCTAACAATCCGCTC
CAGTGGACCGCCTGCGGCGTCCGCTGAGCTATCTCG 

fosI 

TTAGCTCCCATGAAAGGCATCAGCCACATCACATTCATCGTCCGCGACCTGAATCGTATGGCCGCACTTCTCTGTGAGGGACTGGG
TGCGCGTGAGGTGTATGACAGCTCAAACCAGAACTTCTCGTTGTCCCGCGAAAAGTTCTTTGTGCTTGGTAGTACGTGGCTAGCTG
CAATGGAAGGTGAACCGCCCGCCGAGCGTTCATATCAGCATGTTGCCTTTGCGGTGAGTGAGACGGACTTGCCTGCGTATCAAGCC
AGACTTGAGGCACTCGGCGTTGAGATTCGGCCACCGCGTAGCCGTGTTGACGGTGAGGGTCTCTCCCTGTACTTCTACGACTTTGA
CAATCATTTATTTGAACTTCATTCAGGTACTTTAGAGCAGCGCCTTGTCCGGTATCAGGCGGGGCGCTAACAATTCATTCAAGCTGA
CACCGCTTCGCAGCGCAGCTTAACTCAAGCG 

fosK 

TTACGTTTTAGGATCAGTCGTATGATCACTGGTATCAATCACATCACCTTTTCCGTCAGGGACTTGAGCTCTTCAATCGAGTTCTATC
GTGACTTGCTGGGAATGAGGCTGCACGTGACCTGGGAAGCAGGTGCTTATTTTACAGCGGGTGATACGTGGGTATGTCTGAGCGTC
GGGGAACCTAAACCCGCCAACGACTACACGCATGTGGCATTCAGTGTTGGCGAAAGAGAGCTTGTTGAGCTGCACGCTAGGCTAAA
AGAAGCCGGGGTTGAGGAGTGGAAGCAGAATACAAGTGAGGGTAACTCCGTGTATCTGCTTGATCCAAACGGCCATCGCATTGAGC
TTCACTGCGGAACGTTGGCAACCCGCTTAGCTGAGTTGGAGAAGTCGCCCTATAAAAGGTTGGTCTGGTGCTGAAACGTAACAATC
GGCTGTTGCCGCCCCTTCGGGGCTGGGACGGCCTTTCCGCCGCTCCGCGGCTACAAGTCCGCCCCAAAGCCGGGCG 

fosL 

TTAGCGCTCACTATGCAGATCGAGGGCATTAGTCACGTAACTTTCGTCGTAAAGGACCTAGAACGTGCATCCCGATTTTTCTGTCAG
GGTCTGGGAGCAACGGAGGTATACGACAGCAAGGGCAGCAACTTCTCTCTTTCACGAGAGAAGTTCTTTTTGGTTGGTGGTGTCTG
GGTTGCGGCAATGGAGGGCACACCACCCGCCACCGCCTCTTATCAGCACTTGGCGTTCAAGGTGGCGCCCGAAGACTTGCCGCAG
TTTGAGGCTCGTCTCCGCGCAATTGGTGTTGCCATCTCACCGCCGCGGGCGCGTGTCCAAGGCGAGGGCTTGTCGCTGTACTTCC
ACGACTTCGACAATCACCTCTTCGAGCTCCACACTGGAACGCTTACTGAGCGCCTTCAAGCCTATGCAGCCCCGCGCTAACAAGTC
CGTCAACGGGACGCCAAAATGCTGCGCATTTTGGTTCCCTCCGCTGCGCTCCGGCGCCCGTTACGTCCAACG 

fosM 

TTAGGCCCACATAAACAAGCGCCCAAGAATATGCCAGTCAGTGGGGTTAGCCATATCACGTTCATGTCGCGCGACTTGGCACGCAC
AGCAAGAATATGGACCTATGGATTGGGTGCCACTGAGGTCTACGACTCGGGTGAGACGACGTTCTCACTGTCTCAAGAGAAATTCTT
CTTACTTGGGAGCACATGGATTGCGGTCATGCTTGGCGAGCCGGCACCAAGGTCGTATCACCACGTCGCGTTTGAAGTAGACGACA
TTGACCTGCCCTCAATTGAGACGAAGCTAAAGGAGCTAGGTGTCGAGTTCATGCCCCCGCGGGCCCGAGTCGATGGAGAAGGCAA
ATCGCTTTACTTCTATGACTTCGATGACAACCTGCTCGAGCTCCATGCAGGAACCCTCCAACAGCGACTTGAGCGCTACAAGAAGG
GGAAAAATGTCTAGTACGGAACGTCCTATCCTCATGTCGCAGCGCCTAACCGTTTGGTCAAGCCGACGCCCACAAGCCTCGCTTGT
GGGTACCCTCCGTGCTACGCACTACGGCGCGGCTTACCGCGAGCG 

fosN 

TTACGTTTTTTGAGGGACTGCATGATCACCGGAATAAACCACATTACTTTTTCTGTCAGTGATCTGGACTCATCGATCCAGTTCTATC
GTGGTTTCCTGGGCATGAAGCTTCACGTTCTTTGGGACACGGGTGCGTATCTGACAGCCGGTGATACATGGCTTTGCTTGAGTTTG
GGAGAGCCCGACCCCGCTAAAGACTATACGCATGTGGCCTTTAGTATCAGTGAAAACGCGCTCTCGGAGCTACGCGCCAAACGGG
GCGAGATGGGATTCAAGGAATGGAAGCAAAACACTAGTGAGGGCGAATCGCTGTACCTACTTGATCCCAACGGCCATCGCCTAGAG
CTTCACTGTGGCACTTTGGCAACTCGCTTGGCGGAGTTGGAGAACTCACCTTACAAGGGGCTGGTATGGTGCTGAAACGTAACAAT
CGGCCGCACTGCGACCGCTTTTCCGCCGCTTCGCGCCTCCAAACCGGCGCGTGAGCCGGGCG 

foso 

TTAGCGCTCACGATGCAGATCGAAGGCATTAGTCACGTTACTTTTGTCGTTAAGAGCTTGGCACGTGCTGCCGAATTCTTCTGCCAA
GGTCTCGGAGCAACTGAGGTCTACGACAGCGGGGGGCAGAACTTCTCGCTCTCTCGAGAGAAGTTCTTCTTAGTAGGTGGTGTCTG
GGTGGCGGCAATGGAGGGCATCCCACCTGCTGCCCGTTCCTATCAACACTTGGCGTTCAAAGTCGCGCCCGAAGACTTGCCAAAG
TTTGAGGCTCGTCTCCGCGCAATCGGCGTTGAGATTTCCCCGCCTCGACCGCGGGTCCAAGGAGAGGGGTTGTCGCTGTACTTCC
ACGACTTCGACAACCACCTCTTCGAACTCCACACCGGAACGCTGACAGAGCGCCTGAACACCTATGCAGCCCCGCGCTAACAAGTC
CGTCAACGGGACGCCAAACTGCTGCGCAGTTTGGTTCCCTCCGCTGCGCTCCGGCGCCCGTTACGTCCAACG 

lnu(F)1 

TTGTGCGTAAGAATAAAACAAAAGCTGCGTATACCTTTCCGCAGAATCAGTGGCTTCAATAAAAGGATGTTTCAATGCTTCAGCAGAA
AATGATCGAACGCTTCAAGGAAGCTTGCCATGAGGATGCACGAATAATCGCGGCGCTGATGTTCGGCTCATTTGCTATCGGAGAGG
GTGACGAGTTCTCTGATATCGAATTTGCAGTGTTCATCCAGAATAATCATTTTGAAAATTTCGATCAGCGCTCGTGGCTTAATGCTGT
AAGTCCGGTTGCAGCTTACTTTCCGGATGACTTCGGCCACCACACCGCGCTTTTTGAAAACGGCATTCGCGGTGAATTCCATTTCAT
GCGAAAATCGGACATACCGGTCATTTCCACTTGGCAAGGCTACGGGTGGTTTCCCTCGCTTGAGGAGGCTGTTTTGTTGGACCGAT
CAGGAGAGTTGTCAAGGTACGCGAGTGCTCTCGTGGGCAGTCCCCCGAAACGTGAAGGCGCGCCGCTGGTGGAAGGACTTGTATT
GAACCTCATCAGCCTGATGCTCTTTGGGGCAAATCTTTTAAATCGGGGAGAGTATGCTCGCGCCTGGGCTTTGCTCAGCAAAGCAC
ATGAAAACTTACTCAAGTTGGTTCGCCTCCATGAAGGGGCAACAGACCACTGGCCGACACCTTCACGCGCGCTCGAAAAGGATGTC
TCGGAGGACTCGTATAATCGCTACCTGGCATGCACAGGCAGCGCGGAACCAAAAGCACTATGTGTAGCCTATCATGAAACGTGGAA
GTGGAGTCTCGAATTGTTCAGGAGTGTGGCTGGACCTCTGAATATCGAGCTTCCGAGAATTGTAATTGCGCAGACAAAAAGGTTGCT
AAATGAATCTGCGACGCCGCACAACAAGTAAATCCAGCGGACGCATAAAAGCGCGCCGCTGATTAACGCG 



Supplementary Material 

  186 

lnu(F)2 

TTGTGCTGACGAAAAAAAACAAAAATCTGCGTATACCTTTCCTCATAATCTGTGGCTTCAATAAAAGGATATTTCTATGCTTCAGCTGA
AAATGATCGAACTCTTCAAGGAAGGTTGTCATGAGGATGCACGAATAATCGCGGCATTGATGTTCGGCTCATTTGCTATCGGAGAGG
GTGACGAGTTCTCTGATATCGAATTCGCAGTGTTCATCCAGGATGACCATTTTGAAAATTTCGATCAGCGCTCGTGGCTTAATGCCG
TAAGTCCGGTTGCTGCTTACTTTCCGGACGACTTCGGCCACCACACCGCACTTTTTGAAAACGGCATTCGCGGTGAATTCCATTTCA
TGCGAAAATCGGACATACCGGTCATTTCCACTTGGCAAGGCTATGGGTGGTTTCCCTCGCTTGAGGCGGCTGTTTTGTTGGACCGA
TCAGGAGAGTTGTCAAGGTACGCAAGCGCTCTCGTGGGCGGTCCCCCGATACGTGAAGGCGCGCCGCTGGTGGAAGGGCTTGTG
TTGAACCTCATCAGCCTGATGCTCTTTGGGGCCAATCTTTTAAATCGGGGAGAGTACGCTCGCGCCTGGGCTTTGCTCAGCAAAGC
ACATGAAAACCTACTCAAGCTGGTTCGACTCCACGAAGGGGCAACAGACCACTGGCCGACACCTTCACGCGCGCTCGAAAAGGATA
TCTCGGAGGACTCGTATAATCGCTATCTGGCATGCACAAGCAGTGCAGAACCAAGAGCACTATGTGCAGCCTATCATCAAACGTGG
ACGTGGAGTCTCGAATTGTTCAAGAGCGTGACAGAACCTCTGAATATCGAGCTTCCGAGAACTGTAATTGCGCAGGCAAAAAGGTT
GCTCAATGAGTCTGCGACGCCGCACAACAAGTAAATCCAGCGGACGCATAAAAACGCGCCGCTGATTTTGACG 

smr1 
TTAGCCCCCACAATCGGAGATCGTTATGGGTTGGATATATCTCATTCTCGCTGGCGTCTTTGAAGTTGGTTGGCCAGTCGGGCTCAA
GATGGCGCAGACACCGGAGACTCGCTGGAGCGGCATCGGAGTGGCGGTTGCATTTATGACTGTGAGTGGGTTTTTACTCTGGCTT
GCGCAGCGGCAGATCCCCATTGGCACCGCATACGCGGTGTGGACAGGCATTGGTGCTGCCGGCACCTTTTTCGTAGGCGTGCTGT
ACTACGGCGATCCGACTTCGTTCTTTCGGTACATGGGCGTCGCCCTGATAATAGCGGGCGTTATTACCCTCAAGTTGGCTCATTAGA
GGTGTGCGCTAACAATTCATTCAAGCCCGACGCCCGCTTCGCGGCGCGGCTTAATTCAGGCG 

smr2 
TTAGCGCCCACGAACGGAGATCACGATGGCCTGGATATACCTGATCCTCGCTGGACTCTTTGAGATTGGCTGGCCCGTCGGACTGA
AGATGGCGCAGGTACCGGAAACCCGATGGAGTGGTGTTGGCATTGCGGTCGCGTTTATGGCCGTGAGTGGCTTTCTGCTATGGCT
GGCTCAGCGGCACATTCCCATCGGTACTGCCTACGCTGTTTGGACGGGCATCGGTGCGGCTGGCACCTTCCTCGTCGGCGTTCTT
TACTACGGAGATCCAACATCAGTTGCGCGGTACTTTGGCGTGGCGCTCATTGTTGCCGGGGTCATCACGCTCAAGCTCGCGCACTG
AGGTGTGGGCGCTAACAATTCATTCAAGCCGAAGCCGCTTCGCGGCGCGGCTTAATTCGGGCG 

smr3 
TTAGACGCCCACGGAACGGGAGATCACGAGGGCCTGGATGGACCTGATCCTCGCTGGACTCTTTGAGATTGGCTGGCCCGTCGGA
CTGAAGATGGCGCAGGTACCGGAAACCCGATGGAGTGGTGTTGGCATTGCGGTCGCGTTTATGGCCGTGAGTGGCTTTCTGCTAT
GGCTGGCTCAAGCGGCAACATTCCCATCGGTACTGCCTAACGCTGTTTGGACGGGCATTCGGTGCGGCTGGCACCTTCCTCGTCG
GCGTTCTTTACTACGGAGATCCAACATCAGTTGCGCGGTACTTTGGCGTGGCGCTCATTGTTGCCGGGGTCATCACGCTCAAGCTC
GCGCACTGAGGTGTGGGCGCTAACAATTCATTCAAGCCGAAAGCCGCTTCGCGGCGCGGCTTAATTCGGGCG 

qacE 

TTAGATGCACTAAGCACATAATTGCTCACAGCCAAACTATCAGGTCAAGTCTGCTTTTATTATTTTTAAGCGTGCATAATAAGCCCTAC
ACAAATTGGGAGATATATCATGAAAGGCTGGCTTTTTCTTGTTATCGCAATAGTTGGCGAAGTAATCGCAACATCCGCATTAAAATCT
AGCGAGGGCTTTACTAAGCTTGCCCCTTCCGCCGTTGTCATAATCGGTTATGGCATCGCATTTTATTTTCTTTCTCTGGTTCTGAAAT
CCATCCCTGTCGGTGTTGCTTATGCAGTCTGGTCGGGACTCGGCGTCGTCATAATTACAGCCATTGCCTGGTTGCTTCATGGGCAA
AAGCTTGATGCGTGGGGCTTTGTAGGTATGGGGCTCATAGTTAGTGGTGTAGTAGTTTTAAACTTGCTTTCCAAAGCAAGTGCCCAC
TAATAAACTCAGTCATCTAACAAGTCGTTGCAGCACCGCTCCAGCACTTCGTGCCTGCGCTGGACAGTTTTTAAGTCGCGGCTTTAT
GGTTTTGCTGCGCAAAAGTATTCCATAAAATCACAACTTAAAAACTGCCGCTGAACTCGGCG 

qacEDsul1 

TTAGATGCACTAAGCACATAATTGCTCACAGCCAAACTATCAGGTCAAGTCTGCTTTTATTATTTTTAAGCGTGCATAATAAGCCCTAC
ACAAATTGGGAGATATATCATGAAAGGCTGGCTTTTTCTTGTTATCGCAATAGTTGGCGAAGTAATCGCAACATCCGCATTAAAATCT
AGCGAGGGCTTTACTAAGCTTGCCCCTTCCGCCGTTGTCATAATCGGTTATGGCATCGCATTTTATTTTCTTTCTCTGGTTCTGAAAT
CCATCCCTGTCGGTGTTGCTTATGCAGTCTGGTCGGGACTCGGCGTCGTCATAATTACAGCCATTGCCTGGTTGCTTCATGGGCAA
AAGCTTGATGCGTGGGGCTTTGTAGGTATGGGGCTCATAATTGCTGCCTTTTTGCTCGCCCGATCCCCATCGTGGAAGTCGCTGCG
GAGGCCGACGCCATGGTGACGGTGTTCGGCATTCTGAATCTCACCGAGGACTCCTTCTTCGATGAGAGCCGGCGGCTAGACCCCG
CCGGCGCTGTCACCGCGGCGATCGAAATGCTGCGAGTCGGATCAGACGTCGTGGATGTCGGACCGGCCGCCAGCCATCCGGACG
CGAGGCCTGTATCGCCGGCCGATGAGATCAGACGTATTGCGCCGCTCTTAGACGCCCTGTCCGATCAGATGCACCGTGTTTCAATC
GACAGCTTCCAACCGGAAACCCAGCGCTATGCGCTCAAGCGCGGCGTGGGCTACCTGAACGATATCCAAGGATTTCCTGACCCTG
CGCTCTATCCCGATATTGCTGAGGCGGACTGCAGGCTGGTGGTTATGCACTCAGCGCAGCGGGATGGCATCGCCACCCGCACCGG
TCACCTTCGACCCGAAGACGCGCTCGACGAGATTGTGCGGTTCTTCGAGGCGCGGGTTTCCGCCTTGCGACGGAGCGGGGTCGCT
GCCGACCGGCTCATCCTCGATCCGGGGATGGGATTTTTCTTGAGCCCCGCACCGGAAACATCGCTGCACGTGCTGTCGAACCTTCA
AAAGCTGAAGTCGGCGTTGGGGCTTCCGCTATTGGTCTCGGTGTCGCGGAAATCCTTCTTGGGCGCCACCGTTGGCCTTCCTGTAA
AGGATCTGGGTCCAGCGAGCCTTGCGGCGGAACTTCACGCGATCGGCAATGGCGCTGACTACGTCCGCACCCACGCGCCTGGAG
ATCTGCGAAGCGCAATCACCTTCTCGGAAACCCTCGCGAAATTTCGCAGTCGCGACGCCAGAGACCGAGGGTTAGATCATGCCTAG
CATTCACCTTCCGGCCGCCCGCTAGCGGACCCTGGTCAGGTTCCGCGAAGGTGGGCGCAGACATGCTGGGCTCGTCAGGATCAAA
CTGCACTATGAGGCGGCGGTTCATACCGCGCCAGGGGAGCGAATGGACAGCGAGGAGCCTCCGAACGTTCGGGTCGCCTGCTCG
GGTGATATCGACGAGGTTGTGCGGCTGATGCACGACGCTGCGGCGTGGATGTCCGCCAAGGGAACGCCCGCCTGGGACGTCGCG
CGGATCGACCGGACATTCGCGGAGACCTTCGTCCTGAGATCCGAGCTCCTAGTCGCGAGTTGCAGCGACGGCATCGTCGGCTGTT
GCACCTTGTCGGCCGAGGATCCCGAGTTCTGGCCCGACGCCCTCAAGGGGGAGGCCGCATATCTGCACAAGCTCGCGGTGCGAC
GGACACATGCGGGCCGGGGTGTCAGCTCCGCGCTGATCGAGGCTTGCCGCCATGCCGCGCGAACGCAGGGGTGCGCCAAGCTG
CGGCTCGACTGCCACCCGAACCTGCGTGGCCTATACGAGCGGCTCGGATTCACCCACGTCGACACTTTCAATCCCGGCTGGGATC
CAACCTTCATCGCAGAACGCCTAGAACTCGAAATCTAA 

qacF 

TTAGATGCCAGATTTGGCGTTGCGTATCTCACAGAAAATCGATAGCCGCAAGACTGTTTTTGGCAACTCATAGCCACTACAATTTCTC
CTTCATACCGTAGAGGAGATTGCGCGTGAAGAACTGGATATTTCTGGCTGTTTCAATCTTTGGCGAGGTCATCGCAACTTCCGCACT
GAAGTCTAGCCATGGATTCACTAGGTTAGTTCCTTCCGTTGTAGTTGTGGCTGGCTACGGGCTTGCGTTCTATTTCTTGTCTCTCGC
GCTCAAGTCCATTCCGGTCGGTATTGCTTACGCTGTATGGGCTGGGCTTGGCATCGTGCTTGTGGCAGCTATTGCTTGGATTTTCCA
TGGCCAAAAACTAGACTTCTGGGCGTTCATTGGCATGGGACTTATCGTCAGTGGCGTCGCCGTTCTAAACCTGCTATCCAAGGTCA
GCGCACATTGACCGGGTTGGCATCTAACAATTCATTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAATTCAGGCG 

qacG 

TTAGATGCTTTGCTGTGCGCACAAATTTCGGCCAGCAACAAGACTGTTTTTTTTCTTAAATCGAACCTAAAATTTCTTCGCGGAACTC
CATGGAGAAATATTTTGAAAAATTGGTTATTTCTGGCTACGGCCATTATTTCTGAGGTCATTGCAACCTCTGCGCTCAAGTCTAGTGA
GGGCTTTACTAGGTTAGTACCGTCTTTTATCGTCGTAGCGGGATACGCTGCTGCTTTTTATTTCCTGTCGCTGACACTCAAATCGATT
CCTGTTGCAATCGCCTACGCAGTTTGGTCGGGCCTCGGGATCGTCTTGGTCACTGCGATTGCATGGGTTTTGCATGGTCAAAAACT
AGATATGTGGGGATTTGTTGGTGTCGGCTTCATTATCAGCGGCGTTGCTGTGCTCAACTTGCTATCTAAGGCAAGTGTTCACTAAAA
CGGTCGCATCTAACCATTCCGTCGAGAGGGACCGCCCACAAGCTGCGCTTGCGGGTTCCCTTCGCGGCTTCGCCGCTACGGCGGC
CCCTCACGTCAAACG 

qacH 

TTAGATGCCAGATTTGGCGTTGCGTATGCTCACAGAAAATCGACAGCCGCAAGACTGTTTTTGGCAACTCATAGCCACCACTATTTAT
CCTTCATACCGTAGAGGAGATTGCACGTGAAGAACTGGCTCTTTCTGGCTATTGCAATATTTGGTGAGGTCGTCGCAACTTCCGCAC
TGAAGTCCAGCCATGGATTCACCAAGTTAGTTCCTTCTGTTGTAGTTGTGGCTGGCTACGGGCTTGCGTTCTATTTCCTCTCTCTCG
CAATCAAGTCCATCCCGGTCGGCATTGCTTATGCTGTTTGGGCTGGCCTCGGCATCGTACTTGTGGCAGCTATCGCTTGGATCTTCC
ATGGCCAGAAACTAGACTTGTGGGCGTTCGTTGGCATGGGACTTATCGTTAGTGGCGTCGCCGTTCTAAATCTGCTATCCAAGGTCA
GCGCACATTGATCGGGCTGGCATCTAACAATTCATTCAAGCCGACGCCGCTTCGCGGCGCGGCTTAATTCAGGTG 

qacK 

TTAGCATGTAATTAACCGGAAAGTTGCGCACGAATAATTTTTTTCTTCAAGACTACAATTTTAAGAGAAAAAGACACATACTCCCTTCG
AATCATATTGGAGGCGGATCATGAAAAGCTGGTTATTTCTTACTATTTCGATTCTTGGAGAGGTAGTTGCAACATCTGCATTAAAGTC
CAGTGAAGGTTTTACGAAGCTCGTACCATCTTTCATCGTAATTGTTGGCTATGGAATTGCATTTTATTTCCTTTCATTGGTCTTGAAGT
CTATTCCTGTAGGGGTGGCCTATGCAGTTTGGTCTGGCCTTGGCGTAGTATTAGTCACTGCTTTTGCATGGGGGCTTTACGGGCAAA
AGATTGATGCATGGGGTTTCGTGGGTATTAGTTTAATAGTTTGCGGTGTATTGGTTTTAAATTTGCTTTCTAAGGCAAGCGTTCATTAA
GTGGCACTGGCATCTAACAATCGGCAGCAGGCGGACAGCCTACACTACGCGGTTTTGTGTTACTCGCTGCGCTCAAACATTAACAC
AAAACCGCTCCGTTACGGCTGCCGCTGTGCCGGGCG 

qacL 
TTGGGCGTCTCAATACTGAGGATCACCGCAAATGCACTGGATACTTCTGACCGCAGCGATTATTAGTGAGGTCATCGGCACATCGG
CCTTGAAGGCATCGGAAGGTTTCTCGCGTCTATGGCCTTCAGTTATTGTCACCATTGGCTACGCGCTGGCCTTCTACTTCCTCTCAC
TCACGCTCAAAACAATTCCAGTTGGGGTGGCCTACGCAATCTGGTCCGGAATTGGCATCGTTCTAATTGCATTGGTTGCGTGGGTTC



Supplementary Material 

 187 

TCTATGGGCAAGCTCTTGATCTTCCGGCAATCATAGGCATGTCGCTCATCGTGGCTGGCGTTGTTGTTCTCAATCTGTTTTCCAAGT
CCGTCTCTCACTCATGAACCGCCCAACCCATCATTCCACCGGACCTGCGCAAAAAGCCGCGCAGTCCGGTGAATTCAAACG 

qacM 

TTAGATGCCAGTGGTAGTTCCCGTAGGCGCACAGCTTTAGAGACAATACAGGCCTGCCGATTTTGAGATACAAGCCACATAATTGGC
CGCCCATTATTTTGGAGTGAACTCATGAAAGCTTGGATCTATCTTGCCGTCGCGATAACCGCCGAAGTGGTCGCAACTTCAGCGCTC
AAATCGAGTGAAGGATTCACCAAACTTGCCCCGTCTGCCGTAGTTGTCATCAGCTACGGTGTAGCTTTCTATTTTTTGTCGCTCGTCT
TGAAGACCGTTCCAGTCGGCGTTGCCTACGCTGTTTGGTCTGGCCTCGGCATCGTGCTGATCGCAGCAGTTGCGTGGCTCTTTCAT
GGACAAAAGCTCGACACATGGGCCTTCGTTGGCATGGGACTGATAGTTAGCGGAGTAGCGGTTCTCAACGTACTCTCCAAAACGAG
CGCCCACTGATCCGGTGCTGGCATCTAACAGTTCATTCAAGCCGATACCGCTTCGCGGCACGGCTTAACTCATTCG 

qnrVC1 

TTATGTGCTTTCTCTAAAACAAGCCAGATTCACTGAAACTTTCTTCTGGCTGGCCGCATCGGTCGTTTCGAATTAGCCACCTTTAAAA
TGCCGTTTTGATGCCTTATTTTGGCTAAAACGGGGTGTTTTAAGTTTCTGTTTTTATTGGTTTTGTTTTTGTTGAGAACTTTCAGGTAAA
TGATAGTCTTCAAATCAAATGTTTTTTGGAGCCACAGCATGGAAAAATCAAAGCAATTATATAATCAAGTGAACTTCTCACATCAGGAC
TTGCAAGAACATATCTTTAGCAATTGTACTTTTATACATTGTAATTTTAAGCGCTCAAACCTCCGAGATACACAGTTCATTAACTGTACT
TTCATAGAGCAGGGGGCATTGGAAGGGTGCGATTTTTCTTATGCTGATCTTCGAGATGCTTCATTTAAAAACTGTCAGCTTTCAATGT
CCCATTTTAAGGGGGCAAATTGCTTTGGTATTGAACTGAGAGATTGTGATCTTAAAGGAGCAAATTTTAGTCAAGTTAGTTTTGTAAAT
CAGGTTTCGAATAAAATGTACTTTTGTTCTGCATACATAACAGGTTGTAACTTATCCTATGCCAATTTTGAGCAGCAGCTTATTGAAAA
ATGTGACCTGTTCGAAAATAGATGGATTGGTGCAAATCTTCGAGGCGCTTCATTTAAAGAATCAGATTTAAGCCGTGGTGTTTTTTCG
GAAGACTGCTGGGAACAGTTTAGAGTACAAGGCTGTGATTTAAGCCATTCAGAGCTTTATGGTTTAGATCCTCGAAAGATTGATCTTA
CGGGTGTAAAAATATGCTCGTGGCAACAGGAACAGTTACTGGAGCAATTAGGGGTAATCATTGTTCCTGACTAAGCGCAAGATTCGG
CTACGCACATAACAAACGCTTTAAGACGGATTCGCAACGTTTGGCGGTTTTAGTTTGAATTGGCTTTTGTATTTACGGTGTAATAATT
GAGTGTTGTGGTAGCGTTGCTCACCACTTAAGCGGGCG 

qnrVC6 

TTATGTGCTTTCTCTAAAACTCACCAGGTTCACCAAAACTTTTTTCTAGCTGGCCGCATCGCTTATTTTGAATTAGCACCCTTTAAAAT
GCCATTTTGATGCCTTATCTTGGCTAAAACGGGGTGTTTTAAGTGTCTGTTTTTATTGGTTTTGTTTTTGTTGACAACTTTCAGGTAAA
TGGTAGTCTTCAAATCAAATGTTTTTTGGAGCTACAGCATGGAAAAATCAAAGCAATTATATAATCAAGTGAACTTCTCACATCAGGAC
TTGCAAGAACATATCTTTAGCAATTGTACTTTTATACATTGTAATTTTAAGCGCTCAAACCTTCGAGATACACAGTTCATTAACTGTACT
TTCATAGAGCAGGGGGCACTGGAAGGGTGCGATTTTTCTTATGCTGATCTTCGAGATGCTTCATTTAAAGATTGTCAGCTTTCAATGT
CCCATTTTAAGGGGGCAAATTGCTTTGGTATTGAACTGAGAGATTGTGATCTTAAAGGGGCAAATTTTAGCCAAGTTAGTTTTGTAAA
TCAGGTTTCGAATAAAATGTACTTTTGCTCTGCATACATAACAGGTTGTAACTTATCCTATGCCAATTTTGAGCAGCAGCTTATTGAAA
AATGTGACCTGTTCGAAAATAGATGGATTGGTGCAAATCTTCGAGGCGCTTCATTTAAAGAATCAGATTTAAGTCGTGGCGTTTTTTC
AGAAGACTGCTGGGAACAGTTTAGAGTACAAGGCTGTGATTTAAGTCATTCAGAGCTTTATGGTTTAGATCCTCGAAAGATTGATCTT
ACAGGTGTAAAAATATGCTCGTGGCAACAGGAGCAGTTACTGGAGCAATTAGGGGTAATCATTGTTCCTGACTAAGCGCAAGATTCG
GCTACGCACATAACAAAGCGTTTAAGACAGATTCGCAACGCTGGGCATTTTCGGTTTGCTTTGAATTTAGTGTTTACGGCACAATGGT
TTAGGTTGGGTGGCATGTTGCTCACTACTTAACGCGGCG 

sul4 

TTAGCTTGACCCTTCGGGCGGAGGAGAAAATGTCATCAATTTATAATAAAACTAAAAGTGGATTTTGTCGGATTTGTGGAAATTGTAA
GAAAGACAGCGTTTTTCTTCCAGTTTTGAAATTGGGCGTAAACCCACTTTTAAATGGGGGCCGCGCCTCATCGACATCGACATCCTT
TTCTATGACAACATGATCATGAACGACGATATCCTGGCTATTCCTCATCCCTTCGTCGATGAACGTGCTTTTGTCCTGGCACCCCTG
GCTGACATTGCGCCCGATTACGAACATCCCCAGACCCAAAAGTCTGTAGCCCAAATGCTGGCTGACGTATCCTCATCCGTAGATGA
GGCCACAACGGCCGTTTACCGTTTACCGGATCAACCAAACTTCATGGGCACTTAACCCCCACCCTAACCCTCCCCCTAAAATGGGG
AGGGAACATCTCCCTCTCCCGGTGGGAGAGGGCTGGGGAGAGGGTGAAAAAAATTTTCAAGGAACTGCTATTGATGTCAACCACAC
TAACCAGCTTCAAATGGGGTGAACGCACCTACATCATGGGCATCCTCAACGTCACTCCAGACAGCTTTTCTGGAGATGGCGTTATGG
TTGAAGAAGATGTCATCGCCAAAGCGGTAGCCCAGGCCAAACAATTTGTAGCCGACGGCGCAGACATCATCGACATTGGCGGCGA
GAGTACCCGCCCTGGCAGCTCACCTATAAGCGCAGAGGAAGAACTGGCGCGGGTGCTGCCGGTGGTGCAGGCCGTACGCCAGGC
TGTGGACGTCGTTATTTCCATCGACAGCTACCGCGCTTCCGTGGCCGAAGCGGCCCTGGCGGCAGGCGCCAGCTGGCTCAACGAC
GTCTGGGGGCTGCGCATGGACCCGGACATGGCCGGCCTGGCAGCACAAGCCGGCTGCCCCATCGTCCTTATGCACAACCGCAGC
AAACCAAAGAACATAGCGCAAGAAAAAAAGCTGGGCGGGCGCTTCATCGGGGTAAAATACGACGACCTCATCACCGACGTTAAACG
TGAATTACAAGAAAGCATCGACATCGCCTTAAAAGCCGGCGTAAAAGAGTCCCAAATTATCCTGGATCCCGGCATCGGCTTCGGTAA
AACCGTCGAGCAAAGTTTGCAACTGCTCGACCAGATTAATCAGTTCAAAACAATGGGATTTCCCATCTTAATAGGTCCGTCGCGCAA
ATCATTTATTGGCTATACGCTCGATTTGCCGCCAGACCAGCGCATAGAAGGAACGGCGGCCACCGTCGCCATTGGCATTGACCGAG
GAGCCGACGTTGTGCGCGTCCATGACGTCAAAGCAATCGTTCGGGTCGCCCGTATGACAGATGCAATCGTGAGACGTTAAAGGATT
TCAGTGCTATCAGTCACTAGTGAAATGCGTAAAACGTAATGCGTAAGGTTTATTACGCATTACGCATTACGAATCAAACCACAAACAC
ACTGAATTTAGACACATTTTATAGGAGCTGCACATGATCGATTTTGAACTAGACCAGGAACAAAAGATGCTAACCGATGCCATCCGC
CGCCTGGCGGAAGAGCGCATACGGAATCTGTAGCAACAAGGCAGGTGGTTCCGTTCTCAGTGCCACCTTCAAGTCAGGGAGTGAA
CATGAGGTTCGGCAGCTTCGTTCTCAGTGGCATCAACAAGTCCAGCTTGGTCATCAGCTTTGGCGCCCTGTCATGCTTCGTTTTGCC
TCCGGCTTCGCTTTCGCTCCGCAAATTTCGCTGCTTCAAGCTCAGTGTGCTGGCCCATAACAAGGGCAATCAGTTCGCGCCTGGCG
GCGCCGGACGCTCCCTGCGGTCGCGCCGCTGTTGCCGTCG 

  



Supplementary Material 

  188 

Supplementary Table S6. Transcriptomic analysis of pMBA containing E. coli MG1655. 
List of the DE genes showing log2fold change > 1 and padj < 0.05 between pMBA and E. coli MG1655. 
 

GeneID baseMean log2FoldChange p value padj Gene Product 

cds-NP_418663.1 10693.8904442378 -1.00187019607166 2.57782169890577e-27 4.89921797618355e-25 mgtA Mg(2(+)) importing P-
type ATPase 

cds-NP_418272.1 52.0910806549008 -1.00896369965551 0.0013172408829078 0.015393387793463 metR 
DNA-binding 

transcriptional dual 
regulator MetR 

cds-NP_416875.1 58.0916283169775 -1.01102292595806 0.00195839988792302 0.0204979188269277 frc formyl-CoA transferase 

cds-NP_417970.1 446.083518589937 -1.03686177543811 4.10136574207211e-17 4.23143762703496e-15 mdtE 
multidrug efflux pump 

membrane fusion protein 
MdtE 

cds-NP_417069.1 2485.94695662744 -1.05087078408895 4.19546189938973e-20 6.05992516747853e-18 nadB L-aspartate oxidase 

cds-YP_588479.2 35.077037599087 -1.06770199331554 0.00222542526789758 0.0228295756885743 ytjA DUF1328 domain-
containing protein YtjA 

cds-NP_418541.2 1602.55685511277 -1.07446338539299 3.95134934290832e-28 7.92684582068998e-26 adiA arginine decarboxylase 
degradative 

cds-NP_416891.1 325.504837731164 -1.09341197235169 2.43850117447425e-10 1.33415571833735e-08 ypeC DUF2502 domain-
containing protein YpeC 

cds-NP_415690.5 47.2068145589444 -1.09763047140475 0.00161439054843321 0.0178230830628353 ymgG PF13436 family protein 
YmgG 

cds-NP_417198.1 348.451721402939 -1.09908743444963 1.65503332805913e-10 9.63923443164762e-09 hycH formate hydrogenlyase 
assembly protein 

cds-NP_418734.1 7176.63990212688 -1.12856292851041 1.57314349580287e-33 4.73385096945346e-31 fimA type 1 fimbriae major 
subunit 

cds-NP_415125.1 144.45237445706 -1.13892334579405 1.48744209145397e-06 4.59072939507717e-05 entC isochorismate synthase 
EntC 

cds-NP_415689.2 324.101526267298 -1.16098304545022 6.73110270910539e-12 4.96041058828154e-10 ymgD PF16456 family protein 
YmgD 

cds-NP_416314.1 1175.93927497039 -1.18561882458864 1.52777873836164e-13 1.25382023277815e-11 dmlA 

D-malate/3-
isopropylmalate 
dehydrogenase 

(decarboxylating) 

cds-NP_418738.1 62.0951726085809 -1.18730439984642 3.7089408771689e-05 0.000769711810773384 fimF type 1 fimbriae minor 
subunit FimF 

cds-NP_417968.1 8185.58602351012 -1.20241857803249 1.26056005252381e-45 7.58647058277244e-43 hdeD acid-resistance 
membrane protein 

cds-NP_417966.4 4442.82625144058 -1.25888323493793 6.51195097558539e-47 4.70293099456777e-44 hdeB periplasmic acid stress 
chaperone HdeB 

cds-NP_417972.1 257.581871878644 -1.26709496136913 1.25508283680375e-14 1.13302603092458e-12 gadW 
DNA-binding 

transcriptional dual 
regulator GadW 

cds-NP_417967.1 25596.8816113662 -1.28217624096784 3.11087072230248e-47 2.80833854455856e-44 hdeA periplasmic acid stress 
chaperone HdeA 

cds-NP_417969.1 338.312828792651 -1.28635595342299 2.09204908496867e-18 2.36074663931933e-16 gadE 
DNA-binding 

transcriptional activator 
GadE 

cds-NP_417206.1 74.5310994938313 -1.29991775291447 2.85761324142033e-06 8.25507313181504e-05 hypA 
hydrogenase 3 nickel 
incorporation protein 

HypA 

cds-NP_414913.1 164.596952343568 -1.3237061134429 2.94541309601167e-11 1.93379757994512e-09 yaiY 
DUF2755 domain-

containing inner 
membrane protein YaiY 

cds-NP_418273.1 651.191211072339 -1.38186576369585 5.09007883207692e-21 7.99142376636076e-19 metE 
cobalamin-independent 

homocysteine 
transmethylase 

cds-NP_417964.1 118.29669531921 -1.40972625795003 2.49676273797498e-11 1.66959449015327e-09 dctR 
putative DNA-binding 

transcriptional regulator 
DctR 



Supplementary Material 

 189 

cds-
YP_001165313.1 25.8551598431996 -1.49550038523296 0.000581866104341478 0.00805026246274743 appX cytochrome bd-II 

accessory subunit AppX 

cds-NP_417963.4 1022.68724113612 -1.49706218936849 1.84656019716815e-30 4.44528591464947e-28 slp starvation lipoprotein 

cds-NP_417204.1 107.9875485457 -1.50463200583777 4.24713280549549e-10 2.16005585361186e-08 hycB formate hydrogenlyase 
subunit HycB 

cds-NP_416257.1 109.527698646828 -1.5115087489886 1.12858029666415e-10 6.79217241875705e-09 spy ATP-independent 
periplasmic chaperone 

cds-NP_417201.1 926.784117891567 -1.56854320792923 1.9470588036626e-31 5.4083302615582e-29 hycE formate hydrogenlyase 
subunit HycE 

cds-NP_417203.1 810.221299353247 -1.57093781469598 5.3907384108418e-21 8.11081516731239e-19 hycC formate hydrogenlyase 
subunit HycC 

cds-NP_418503.1 2203.73277275277 -1.5797159321735 2.24742324971757e-35 7.37767759520922e-33 fdhF formate dehydrogenase 
H 

cds-NP_417193.1 130.474119832831 -1.59941817702693 3.12299178074508e-15 2.96766403165013e-13 hydN putative electron 
transport protein HydN 

cds-NP_417205.1 91.228214770026 -1.61220742865463 4.48785081396124e-08 1.80062547657934e-06 hycA 
regulator of the 

transcriptional regulator 
FhlA 

cds-NP_417199.1 376.590610965942 -1.61436500829892 2.90749749359649e-19 3.49965781645898e-17 hycG formate hydrogenlyase 
subunit HycG 

cds-NP_417948.4 82.8261859132586 -1.648566414595 1.18646622675612e-09 5.86894458194021e-08 yhiM inner membrane protein 
YhiM 

cds-NP_417200.1 328.278566676392 -1.74084652611983 1.12322273232036e-28 2.38585722729931e-26 hycF formate hydrogenlyase 
subunit HycF 

cds-NP_417202.1 563.722546826826 -1.76741577752852 6.03747766254548e-31 1.55723798853227e-28 hycD formate hydrogenlyase 
subunit HycD 

cds-NP_416010.1 1987.91829646751 -1.84324309614455 2.3081259627586e-61 2.77821428384044e-58 gadB glutamate decarboxylase 
B 

cds-NP_417974.1 1703.80056588875 -1.85979209664822 4.35785969494246e-63 7.8681156792186e-60 gadA glutamate decarboxylase 
A 

cds-NP_416009.1 2133.8135129458 -1.92597354526408 1.35081147518026e-70 4.8777802368759e-67 gadC L-glutamate:4-
aminobutyrate antiporter 

cds-NP_416330.1 575.729018490171 1.00156857457841 1.7495276597497e-15 1.70744442685302e-13 yoaE TerC family inner 
membrane protein YoaE 

cds-NP_418773.1 268.860520730953 1.04583165807327 1.08321524415753e-09 5.43262534257339e-08 yjiX PF04328 family protein 
YjiX 

cds-NP_416070.1 45.9779416570507 1.07393586870842 0.00176038050631877 0.0189753253979615 cspI cold shock-like protein 
CspI 

cds-NP_416051.1 82.150638873981 1.09151841131902 1.24186900854371e-05 0.000300965704016867 ydeE dipeptide exporter 

cds-
YP_009518795.2 38.5599035136036 1.25274652798568 0.00026674359819901 0.00415177212541648 yoaL protein YoaL 

cds-YP_026180.1 4522.75607665035 1.42493316948833 5.26575900642921e-45 2.71637939603084e-42 srlA sorbitol-specific PTS 
enzyme IIC2 component 

cds-NP_418012.1 8648.70501940828 1.45560941318424 0.000121194668920872 0.00215583226341512 cspA cold shock protein CspA 

cds-NP_417184.1 1096.48071816521 1.49771860820237 1.37521370833922e-37 4.96589670081291e-35 srlB sorbitol-specific PTS 
enzyme IIA component 

cds-YP_026181.1 6638.65692040713 1.51989824764217 1.83466243237288e-42 8.28120755412311e-40 srlE 
sorbitol-specific PTS 

enzyme IIBC1 
component 

cds-NP_416448.1 18.0548919520912 1.59421555892762 0.00239947938589425 0.0242025700068831 fliF flagellar basal-body MS-
ring and collar protein 

cds-NP_417185.1 13570.2481440378 1.59712847138794 2.69769947622926e-38 1.08237697874043e-35 srlD sorbitol-6-phosphate 2-
dehydrogenase 

cds-NP_417186.1 288.828498583272 1.78289718949831 4.62530731416277e-29 1.04387404446511e-26 gutM 
DNA-binding 

transcriptional activator 
GutM 

cds-NP_418122.1 31.4981429368301 2.42429808868817 0.00490389169888549 0.0408959651840081 uhpT 
hexose-6-

phosphate:phosphate 
antiporter 

 



Supplementary Material 

  190 

Supplementary Table S7. Transcriptomic analysis of pMBA derivatives. 
List of the DE genes showing log2fold change > 1 and padj < 0.05 between pMBA derivatives (pMBAdfrA21, 
pMBAdfrA31, pMBAereA2, and pMBAereA3) and pMBA-containing strain. 
 

dfrA21 

GeneID baseMean log2FoldChange p value padj Gene Product 

cds-YP_026181.1 6487.289412203
97 -1.00594242247915 0.003461762326

11366 
0.025640023867

6883 srlE sorbitol-specific PTS enzyme 
IIBC1 component 

cds-NP_418343.1 42.41973398704
98 -1.00701228025525 0.001202075636

48017 
0.012331637994

9259 rhaT rhamnose/lyxose:H(+) symporter 

cds-NP_418491.1 111.8723506690
21 -1.00852598380315 7.720711109094

45e-05 
0.001621349332

90983 actP acetate/glycolate:cation symporter 

cds-NP_417248.1 93.39193253474
58 -1.00989548664724 0.000230477884

176258 
0.003546577786

67777 ygcP putative anti-terminator regulatory 
protein 

cds-NP_418773.1 269.3837662167
31 -1.01169318481721 2.216581929471

02e-07 
1.648582810044

07e-05 yjiX PF04328 family protein YjiX 

cds-NP_418652.2 101.9682905693
93 -1.02897837083554 2.561735671019

4e-07 
1.829079269107

85e-05 yjfF galactofuranose ABC transporter 
putative membrane subunit YjtF 

cds-NP_416051.1 117.8980186518
35 -1.03577429038046 7.858011366212

2e-07 
4.315861627288

85e-05 ydeE dipeptide exporter 

cds-YP_588462.1 23.60618649210
16 -1.04583171946196 0.006649593353

39796 
0.041867810002

876 arnE undecaprenyl-phosphate-alpha-L-
Ara4N flippase - ArnE subunit 

cds-NP_416844.1 1948.121368842
86 -1.07471274087035 0.004979156821

01308 
0.032978830892

4243 fadI 3-ketoacyl-CoA thiolase FadI 

cds-NP_418001.1 4577.791058463
04 -1.07727459072197 4.776601980038

47e-07 
2.991661240129

36e-05 dppA dipeptide ABC transporter 
periplasmic binding protein 

cds-NP_416750.2 44.35833300425
94 -1.08303472620099 0.000253564297

897925 
0.003835697218

20166 rhmD L-rhamnonate dehydratase 

cds-NP_418061.1 321.6017768166
38 -1.10564566663853 9.222508502515

8e-06 
0.000316580339

942129 lldR DNA-binding transcriptional dual 
regulator LldR 

cds-NP_418753.3 29.74886526843
95 -1.12072416313216 0.002876208581

19867 
0.022666809348

5193 yjiK PF06977 family protein YjiK 

cds-NP_415959.1 27.20740760740
21 -1.13567067360864 0.000998613707

047925 
0.010836021076

4775 ydcU putative ABC transporter 
membrane subunit YdcU 

cds-NP_418062.1 593.0085830577
56 -1.16523405377076 0.000744736820

647821 
0.008632176784

78157 lldD L-lactate dehydrogenase 

cds-NP_415662.2 21.82104927433
9 -1.23315732989422 0.001234581335

10432 
0.012432332406

0108 ymfJ uncharacterized protein YmfJ 

cds-NP_417186.1 296.4065670410
99 -1.24027268785217 0.002267692120

98597 
0.019093539792

2639 gutM DNA-binding transcriptional 
activator GutM 

cds-NP_414776.1 32.25585971117
02 -1.24090224552319 0.004660254918

19876 
0.031390773694

2822 phoE outer membrane porin PhoE 

cds-NP_418165.1 533.4896689973
93 -1.35892132253576 0.004850076804

18365 
0.032424670769

5423 tnaB tryptophan:H(+) symporter TnaB 

cds-NP_417999.1 96.30562456776
99 -1.45614484157938 2.188330864593

78e-06 
0.000105572178

197295 dppC dipeptide ABC transporter 
membrane subunit DppC 

cds-NP_416653.1 642.8599145638
75 -1.71751921264319 0.000157643530

888071 
0.002705708679

18469 mglC 
D-galactose/methyl-galactoside 

ABC transporter membrane 
subunit 

cds-NP_418122.1 27.53761613275
07 -1.78088837723667 0.002668865077

26699 
0.021556217931

7718 uhpT hexose-6-phosphate:phosphate 
antiporter 

cds-NP_416070.1 45.19894041913
33 -2.31086339396476 3.090336291760

75e-06 
0.000141442314

892127 cspI cold shock-like protein CspI 

cds-NP_417969.1 195.7063024033
97 1.00020304270268 6.862692931598

84e-05 
0.001493891083

28097 gadE DNA-binding transcriptional 
activator GadE 

cds-NP_416737.1 5317.389905439
69 1.00219005244976 7.470486650228

86e-11 
2.666963734131

7e-08 nrdA ribonucleoside-diphosphate 
reductase 1 subunit alpha 



Supplementary Material 

 191 

cds-NP_418541.2 912.2971050461
17 1.03495454621567 4.320197599552

36e-07 
2.754125969714

63e-05 adiA arginine decarboxylase 
degradative 

cds-NP_415125.1 70.23112095305
18 1.04335945990741 0.006750253768

77171 
0.042277905183

3597 entC isochorismate synthase EntC 

cds-NP_416257.1 65.99916141404
83 1.08130300153028 3.334208939173

38e-05 
0.000862545356

003549 spy ATP-independent periplasmic 
chaperone 

cds-NP_417948.4 48.36933548352
11 1.27521171276928 2.027252072158

99e-06 
0.000101933660

529684 yhiM inner membrane protein YhiM 

cds-NP_417974.1 574.8032304543
31 1.28701701002208 2.894404444490

86e-10 
6.888682577888

25e-08 gadA glutamate decarboxylase A 

cds-NP_416010.1 657.0809259000
64 1.31437149210185 7.524707410036

54e-14 
5.372641090766

09e-11 gadB glutamate decarboxylase B 

cds-NP_416009.1 668.1555314682
67 1.32045472392136 1.112536121562

e-12 
4.964692442470

41e-10 gadC L-glutamate:4-aminobutyrate 
antiporter 

cds-NP_418663.1 6541.461037171
63 1.33814138151681 4.832162063364

13e-19 
1.725081856620

99e-15 mgtA Mg(2(+)) importing P-type ATPase 

cds-NP_418503.1 930.1156076512
03 1.33855621541903 3.623797049788

55e-14 
4.312318489248

37e-11 fdhF formate dehydrogenase H 

cds-NP_417193.1 61.93734963142
93 1.37095893000914 4.860653032244

02e-07 
2.991815745708

82e-05 hydN putative electron transport protein 
HydN 

cds-NP_417206.1 35.14432160569
08 1.41367017686409 0.000228653934

850747 
0.003533742629

51155 hypA hydrogenase 3 nickel 
incorporation protein HypA 

cds-NP_417199.1 153.8249749384
79 1.4262226435346 9.355177060720

31e-10 
2.087373881673

22e-07 hycG formate hydrogenlyase subunit 
HycG 

cds-NP_417201.1 377.4558417983
58 1.43125977148256 6.091649578102

85e-13 
3.545952930736

98e-10 hycE formate hydrogenlyase subunit 
HycE 

cds-NP_417200.1 123.7834809112
14 1.45042159518709 1.794943684229

74e-10 
4.997430528998

42e-08 hycF formate hydrogenlyase subunit 
HycF 

cds-NP_417205.1 37.84374882569
24 1.49359830124189 0.000128431064

428747 
0.002327405583

81028 hycA regulator of the transcriptional 
regulator FhlA 

cds-NP_417203.1 340.6448002896
95 1.51239248936244 2.467474651518

75e-09 
4.636255003116

82e-07 hycC formate hydrogenlyase subunit 
HycC 

cds-NP_417204.1 47.86153764001
89 1.61683129301876 8.461304988063

2e-08 
7.551714701846

4e-06 hycB formate hydrogenlyase subunit 
HycB 

cds-NP_417202.1 235.1334728617
98 1.70587635235084 5.755600084511

95e-14 
5.136873075426

92e-11 hycD formate hydrogenlyase subunit 
HycD 

dfrA31 

GeneID baseMean log2FoldChange p value padj Gene Product 

cds-NP_416239.1 5106.710118226
48 -1.00295870039945 2.503728767267

39e-09 
2.478691479594

72e-08 yniA putative kinase YniA 

cds-YP_026205.1 245.7794095014
51 -1.00567705075414 5.265061015188

43e-09 
4.980747720368

26e-08 tdcE 2-ketobutyrate formate-
lyase/pyruvate formate-lyase 4 

cds-NP_417343.1 41.71798022171
61 -1.00961934022213 0.000524150598

476133 
0.001839496370

74435 xdhB putative xanthine dehydrogenase 
FAD-binding subunit XdhB 

cds-NP_418061.1 321.6017768166
38 -1.01127174471326 4.548058705578

29e-05 
0.000203159348

474782 lldR DNA-binding transcriptional dual 
regulator LldR 

cds-NP_415720.1 568.5762698326
72 -1.01884807812081 1.118592850917

92e-10 
1.322736046210

44e-09 ycgV putative autotransporter adhesin 
YcgV 

cds-NP_416167.1 682.3945380637
98 -1.03090823774436 5.851961922883

13e-24 
2.678688376958

44e-22 nemA N-ethylmaleimide reductase 

cds-NP_415824.1 149.0952484667
69 -1.03093038147068 2.613851600174

87e-07 
1.892022005826

31e-06 pspE thiosulfate sulfurtransferase PspE 

cds-NP_416513.1 278.5824715477
58 -1.03199797495722 2.524539771561

39e-10 
2.850654060354

6e-09 sbmC DNA gyrase inhibitor 

cds-NP_415568.1 133.6947209564
47 -1.03279106048107 5.481126466200

65e-09 
5.162202514738

09e-08 yceK DUF1375 domain-containing 
lipoprotein YceK 

cds-NP_416157.1 799.5974516422
7 -1.03283782253956 5.120535844569

27e-21 
1.801497610771

19e-19 anmK anhydro-N-acetylmuramic acid 
kinase 



Supplementary Material 

  192 

cds-NP_415304.1 257.2011126562
24 -1.0383233775689 2.171230166910

95e-08 
1.886311596028

55e-07 moaC cyclic pyranopterin 
monophosphate synthase 

cds-NP_416586.1 23.76694159372
34 -1.04085531619093 0.004393151890

62442 
0.011988235639

9924 ogrK prophage P2 late control protein 
OgrK 

cds-NP_417210.2 186.9453388508
84 -1.04261289809542 2.829028248089

81e-08 
2.423173692579

14e-07 hypE carbamoyl dehydratase HypE 

cds-NP_416686.1 1417.708348456
53 -1.04328560457147 2.663210334269

12e-06 
1.585634460557

15e-05 yejG protein YejG 

cds-NP_417221.1 9012.678720231
33 -1.04384801230758 2.363579754730

58e-22 
9.316443533229

72e-21 rpoS RNA polymerase sigma factor 
RpoS 

cds-NP_415638.3 85.91778971869
6 -1.04440369198083 3.501456726804

07e-06 
2.011565625641

69e-05 cobB 
protein-lysine 

deacetylase/desuccinylase/lipoami
dase 

cds-NP_417594.3 221.3297938878
87 -1.04523939241945 1.075977063532

88e-10 
1.275887008205

98e-09 garR tartronate semialdehyde reductase 

cds-NP_416925.2 719.3327943623
89 -1.04600135875511 2.778334060858

85e-07 
2.001246716933

35e-06 yfeW penicillin binding protein 4B 

cds-NP_414704.4 210.2124644616
22 -1.04679818450527 1.332687207850

96e-08 
1.196888068316

78e-07 cdaR DNA-binding transcriptional 
activator CdaR 

cds-NP_416368.1 6322.777934810
76 -1.0481206654344 8.149608613146

43e-31 
6.424608123363

77e-29 pykA pyruvate kinase 2 

cds-NP_415037.1 33.66845841882
11 -1.04975133245358 0.001582489708

80105 
0.004931668148

14499 allS DNA-binding transcriptional 
activator AllS 

cds-NP_415305.1 157.8553979102
35 -1.05501978926979 2.176440604122

75e-07 
1.597432353750

09e-06 moaD molybdopterin synthase sulfur 
carrier subunit 

cds-NP_415578.3 2906.281938542
92 -1.05814072766965 2.691519564861

97e-09 
2.640045803598

48e-08 bssS regulator of biofilm formation 

cds-NP_414984.1 141.0976264586
26 -1.05897879328065 0.000346950159

495555 
0.001268871846

19637 glnK nitrogen regulatory protein PII-2 

cds-NP_416673.1 9885.954842215
94 -1.06023256011612 1.881629893969

89e-07 
1.393060601500

84e-06 fruK 1-phosphofructokinase 

cds-NP_415772.1 198.9566705540
61 -1.06024407943794 1.779359721163

47e-07 
1.331236262388

91e-06 ompW outer membrane protein W 

cds-NP_416259.1 187.6958473081
95 -1.06119514746713 8.291273106240

92e-06 
4.395593677265

82e-05 astB N-succinylarginine dihydrolase 

cds-NP_417467.1 1976.068284783
07 -1.06367053364147 1.194317987950

9e-12 
1.933160332588

21e-11 hybD hydrogenase 2 maturation 
protease 

cds-YP_010051177.1 41.74690598712
22 -1.06375964971374 0.000437892080

70369 
0.001563847808

35202 ybgV protein YbgV 

cds-NP_416631.1 46.48461788895
06 -1.06653663488549 5.188163220268

87e-05 
0.000229107996

14818 mlrA DNA-binding transcriptional 
activator MlrA 

cds-NP_418739.1 98.71803536105
2 -1.07051479826251 2.860123179477

94e-06 
1.686363486847

31e-05 fimG type 1 fimbriae minor subunit 
FimG 

cds-NP_418573.1 143.0019959972
67 -1.07356218205448 2.383849776891

99e-09 
2.382171009443

48e-08 blc outer membrane lipoprotein Blc 

cds-NP_414879.3 966.7045638032
1 -1.07471797695453 6.746782319250

03e-16 
1.472874478617

81e-14 lacI DNA-binding transcriptional 
repressor LacI 

cds-NP_418206.1 5188.551436270
24 -1.08013934194146 0.000638934520

352716 
0.002216743482

59292 rbsC ribose ABC transporter membrane 
subunit 

cds-NP_416411.1 72.44210636033
13 -1.08015475106273 2.182235586013

68e-05 
0.000105445821

676053 otsB trehalose-6-phosphate 
phosphatase 

cds-NP_415978.1 23.45216553411
62 -1.08038588756133 0.002428679139

05951 
0.007145049823

75697 pptA tautomerase PptA 

cds-NP_417241.1 261.9041097727
2 -1.08262047047573 1.542524263912

68e-06 
9.628336937648

49e-06 cas3 CRISPR-associated 
endonuclease/helicase Cas3 

cds-NP_417533.1 20.07132044517
06 -1.08697741435459 0.003443410257

89049 
0.009707680442

27803 ttdA L(+)-tartrate dehydratase subunit 
alpha 

cds-NP_416635.1 92.99347007528
95 -1.08948551433167 1.687396994111

96e-06 
1.041050580280

38e-05 osmF glycine betaine ABC transporter 
periplasmic binding protein OsmF 



Supplementary Material 

 193 

cds-NP_416260.1 256.4657332673
36 -1.08976146235488 4.990172592603

62e-08 
4.100997051489

12e-07 astD aldehyde dehydrogenase 

cds-NP_416014.4 27.10154087823
16 -1.09135566025859 0.004297282631

32095 
0.011751751194

8764 ydeM putative anaerobic sulfatase 
maturation enzyme YdeM 

cds-NP_417187.1 1217.367136134
24 -1.09367662746772 6.308429105337

91e-10 
6.781561288238

25e-09 srlR DNA-binding transcriptional 
repressor SrlR 

cds-NP_416313.4 1003.581862203
9 -1.09441282673261 2.541656355231

86e-09 
2.498806259635

57e-08 dmlR DNA-binding transcriptional 
regulator DmlR 

cds-NP_417003.1 15925.24726083
58 -1.09499589338438 3.120397425758

53e-23 
1.289663285578

06e-21 guaB inosine 5'-monophosphate 
dehydrogenase 

cds-NP_416724.1 575.0254479405
89 -1.09570999188906 1.777483554017

2e-05 
8.798543592385

12e-05 atoC 
DNA-binding transcriptional 

activator/ornithine decarboxylase 
inhibitor AtoC 

cds-NP_418140.4 5157.213515986
6 -1.09637226524727 6.050211919444

72e-07 
4.056023959224

59e-06 yidE putative transport protein YidE 

cds-NP_415525.2 86.09650577585
67 -1.09962512787296 3.959890767179

21e-07 
2.745481269687

61e-06 ymdF 
stress-induced bacterial 
acidophilic repeat motifs-
containing protein YmdF 

cds-NP_418609.4 617.8697554591
33 -1.10038160421816 5.460867149682

59e-11 
6.757823097732

2e-10 yjfN protease activator YjfN 

cds-NP_417002.1 23485.00623114
03 -1.11119125973295 2.095637081955

95e-26 
1.173832507616

64e-24 guaA GMP synthetase 

cds-NP_416463.4 38.61272629585
9 -1.11249097960523 0.001767991381

27782 
0.005426344131

29032 yodD stress-induced protein 

cds-NP_416615.1 15.10829345050
99 -1.11445475989012 0.013879225653

2893 
0.032354598289

1286 yehE DUF2574 domain-containing 
protein YehE 

cds-NP_416799.1 9458.906099748 -1.11632180187817 1.270685147616
61e-20 

4.362344091454
76e-19 ackA acetate kinase 

cds-NP_417381.1 17473.64582219
78 -1.11989376602208 2.444677840040

43e-36 
2.738682517118

98e-34 gcvT aminomethyltransferase 

cds-NP_418207.1 4336.527227901
82 -1.12017581368549 2.894332516772

03e-09 
2.813053315958

57e-08 rbsB ribose ABC transporter 
periplasmic binding protein 

cds-NP_415176.1 2956.228690133
89 -1.12019602704911 3.866790645402

83e-09 
3.707416166099

06e-08 ybeL DUF1451 domain-containing 
protein YbeL 

cds-NP_415662.2 21.82104927433
9 -1.12150162358786 0.002356262740

09146 
0.006946406152

74885 ymfJ uncharacterized protein YmfJ 

cds-NP_418361.1 5621.031873293
01 -1.12162787858901 0.000272159935

392732 
0.001022581504

82512 glpK glycerol kinase 

cds-NP_416238.1 71.18474024730
11 -1.12327241334579 3.801922061201

15e-06 
2.163740937771

83e-05 ydiZ putative endoribonuclease YdiZ 

cds-NP_416744.1 3349.722942386
31 -1.12491485215334 0.008354106144

51554 
0.020848592783

997 glpA anaerobic glycerol-3-phosphate 
dehydrogenase subunit A 

cds-NP_415831.1 45.24715504767
3 -1.12563145998163 0.005124544372

25608 
0.013643017756

5317 ycjS D-glucoside 3-dehydrogenase 

cds-NP_416199.1 32.40192831087
39 -1.1287726079136 0.005818692618

37461 
0.015149953808

2084 sufA iron-sulfur cluster insertion protein 
SufA 

cds-NP_418614.4 79.12887774616
94 -1.13291458224687 2.251937955501

52e-07 
1.647164927245

7e-06 ulaA L-ascorbate specific PTS enzyme 
IIC component 

cds-NP_417401.1 51930.68061608
15 -1.13449152783854 4.933502100396

76e-18 
1.390855525919

8e-16 pgk phosphoglycerate kinase 

cds-NP_415303.1 682.8768824721
26 -1.13468893547921 1.246726420151

58e-16 
2.900171787205

07e-15 moaB protein MoaB 

cds-NP_418551.1 35.14754597381
19 -1.13963379409425 0.011186445884

4241 
0.026904350355

9285 yjdJ putative N-acetyltransferase YjdJ 

cds-NP_415261.2 97280.31316131
37 -1.14176259398373 2.360674657021

76e-17 
5.911407067612

73e-16 cydA cytochrome bd-I subunit 1 

cds-NP_418610.4 3538.733297104
39 -1.14236507132982 1.971011912402

77e-12 
3.107628781888

37e-11 bsmA DUF1471 domain-containing 
putative lipoprotein BsmA 

cds-NP_418578.1 9352.185496054
68 -1.14248907450796 7.393712811263

59e-11 
8.916440633866

6e-10 frdA fumarate reductase flavoprotein 
subunit 



Supplementary Material 

  194 

cds-NP_418445.1 278.1955669770
9 -1.14555507718808 1.141393658996

31e-07 
8.834386920631

46e-07 pepE peptidase E 

cds-NP_417248.1 93.39193253474
58 -1.14641464122472 2.670576705711

48e-05 
0.000125898616

126398 ygcP putative anti-terminator regulatory 
protein 

cds-NP_415319.1 230.3255643065
96 -1.14659764460984 7.784375429166

1e-07 
5.137687783249

63e-06 ybiA N-glycosidase YbiA 

cds-NP_418659.1 436.8717428998
15 -1.14811921236312 1.935804610275

29e-10 
2.221218389741

75e-09 nrdD anaerobic ribonucleoside-
triphosphate reductase 

cds-NP_414838.2 38.77872578528
53 -1.14930780730069 0.000161205745

703047 
0.000643764408

497067 rclA cupric reductase RclA 

cds-NP_415827.1 14.96763887057
43 -1.14974758692084 0.016379665299

0111 
0.037307776981

2146 ycjO putative ABC transporter 
membrane subunit YcjO 

cds-NP_416868.1 17.09430419649
67 -1.15079190584352 0.005964803331

34618 
0.015464170390

7069 emrY tripartite efflux pump membrane 
subunit EmrY 

cds-NP_418024.1 27.48973014421
93 -1.15477731540663 0.002353768136

61089 
0.006943860677

44461 xylG xylose ABC transporter ATP 
binding subunit 

cds-YP_009518834.1 1198.562760760
83 -1.15574088041292 6.408044859287

41e-13 
1.066122689606

97e-11 yjiT putative uncharacterized protein 
YjiT 

cds-NP_416660.1 174.4838897508
16 -1.15605817935643 1.372126186113

92e-06 
8.666381564223

99e-06 cirA iron-catecholate outer membrane 
transporter CirA 

cds-NP_416332.1 16404.00845694
86 -1.16391876089448 1.158509149028

38e-17 
3.121375599628

99e-16 manY mannose-specific PTS enzyme IIC 
component 

cds-NP_415915.1 208.1410929115
8 -1.16956176689738 3.666737500026

23e-11 
4.645625457622

52e-10 paaJ beta-ketoadipyl-CoA thiolase 

cds-NP_415902.1 274.2842712606
93 -1.17158835619866 4.145082579544

56e-07 
2.869205941645

73e-06 feaR DNA-binding transcriptional 
activator FeaR 

cds-NP_414606.1 562.6814189141
34 -1.17542038332598 1.395782208580

61e-10 
1.632374962068

03e-09 araC DNA-binding transcriptional dual 
regulator AraC 

cds-NP_416333.4 11655.74853020
7 -1.17577420824797 1.633967848014

66e-17 
4.190241643974

94e-16 manZ mannose-specific PTS enzyme IID 
component 

cds-NP_418562.4 32878.05796910
79 -1.17881921228055 2.073238931288

9e-11 
2.766701608306

22e-10 aspA aspartate ammonia-lyase 

cds-NP_417030.2 937.3342762888
82 -1.19089260007773 1.131092800752

26e-09 
1.177276785526

25e-08 csiE stationary phase-inducible protein 
CsiE 

cds-NP_414918.4 43.53544829229
31 -1.19305385074958 0.000991207877

859012 
0.003273523612

13795 psiF phosphate starvation-inducible 
protein PsiF 

cds-NP_417188.4 1510.471576791
31 -1.19340052283404 6.337995215173

74e-12 
9.368349177428

68e-11 gutQ D-arabinose 5-phosphate 
isomerase GutQ 

cds-NP_416853.1 777.7586469679
42 -1.1959897304615 3.213093783088

57e-14 
5.999184313424

59e-13 yfdI serotype-specific glucosyl 
transferase YfdI 

cds-NP_416396.1 13.42906539123
89 -1.20270379263497 0.014838636216

9692 
0.034203061428

4774 cheY chemotaxis protein CheY 

cds-NP_416955.1 13.86192615012
66 -1.20380137441467 0.014596709282

7418 
0.033748346338

9745 eutQ putative ethanolamine utilization 
acetate kinase EutQ 

cds-NP_416262.1 373.0213633040
12 -1.21348637544626 1.030463927692

71e-11 
1.433557169996

03e-10 astC succinylornithine transaminase 

cds-YP_026181.1 6487.289412203
97 -1.21368728498766 0.000419929387

472286 
0.001504746971

77569 srlE sorbitol-specific PTS enzyme 
IIBC1 component 

cds-NP_417207.1 174.7219722213
33 -1.21374060051732 1.123892642253

88e-11 
1.548353067338

12e-10 hypB hydrogenase isoenzymes nickel 
incorporation protein HypB 

cds-NP_415274.1 43.46978323178
75 -1.21593666861484 0.000178921994

366161 
0.000701354447

529235 ybgS PF13985 family protein YbgS 

cds-NP_415723.1 3835.079270237
1 -1.21806447511586 5.203080191357

59e-13 
8.754747974153

86e-12 ychH stress-induced protein 

cds-NP_417474.1 4465.605125667
21 -1.21818705674077 4.110379641665

92e-10 
4.521417605832

51e-09 gpr L-glyceraldehyde 3-phosphate 
reductase 

cds-NP_418550.1 62.28361581397
14 -1.23207127842491 2.582823811753

99e-06 
1.544252944752

35e-05 yjdI PF06902 family protein YjdI 



Supplementary Material 

 195 

cds-NP_417910.1 259.6815993429
53 -1.23701506326424 1.014075942032

79e-06 
6.590719519440

61e-06 ugpB 
sn-glycerol 3-phosphate ABC 

transporter periplasmic binding 
protein 

cds-NP_416331.1 20828.31250518
82 -1.24073332784845 1.240468201100

7e-17 
3.321178070494

13e-16 manX mannose-specific PTS enzyme 
IIAB component 

cds-NP_416290.1 131.2482778677
45 -1.24447553399045 2.024480282092

12e-10 
2.310512750902

46e-09 ydjL putative zinc-binding 
dehydrogenase YdjL 

cds-NP_415813.4 168.8135520493
78 -1.24526147023673 2.844810646327

19e-09 
2.771249181101

8e-08 puuA glutamate-putrescine ligase 

cds-NP_415401.1 738.2153891908
81 -1.24767489883394 1.194343072328

35e-16 
2.824621366056

54e-15 cspD DNA replication inhibitor CspD 

cds-NP_416737.1 5317.389905439
69 -1.24792899556057 6.979930766034

45e-16 
1.508302805634

96e-14 nrdA ribonucleoside-diphosphate 
reductase 1 subunit alpha 

cds-NP_417186.1 296.4065670410
99 -1.24799005581948 0.002091107917

40587 
0.006291057529

60904 gutM DNA-binding transcriptional 
activator GutM 

cds-NP_418120.2 29.95294747693
87 -1.25301382110682 0.000280181583

290811 
0.001047175592

68568 adeQ adenine transporter 

cds-NP_416813.1 929.3559354179
9 -1.25350242689953 3.522446049985

28e-10 
3.925406501253

23e-09 argT 
lysine/arginine/ornithine ABC 

transporter periplasmic binding 
protein 

cds-NP_418577.1 1955.887975843
96 -1.25857683799387 9.557719088264

88e-14 
1.724034328760

32e-12 frdB fumarate reductase iron-sulfur 
protein 

cds-NP_416258.1 115.8565405362
35 -1.26496136065647 1.331371487992

84e-05 
6.763303609051

92e-05 astE succinylglutamate desuccinylase 

cds-NP_415541.1 120.4350393545
01 -1.26589109870638 5.193862212538

35e-09 
4.935328446155

3e-08 pgaC poly-N-acetyl-D-glucosamine 
synthase subunit PgaC 

cds-NP_416129.1 6933.899450041
84 -1.27268584699376 1.818613027994

08e-19 
5.692526220713

82e-18 fumA fumarase A 

cds-NP_417468.1 4861.956797099
08 -1.27314301480207 3.703152649516

6e-25 
1.876704860594

31e-23 hybC hydrogenase 2 large subunit 

cds-NP_417466.1 540.8611092762
32 -1.27360376103198 3.844958812513

81e-17 
9.461265702237

75e-16 hybE hydrogenase 2-specific chaperone 

cds-NP_416800.1 16571.55940307
5 -1.27410365447281 4.533081012342

61e-34 
4.706664846229

87e-32 pta phosphate acetyltransferase 

cds-NP_418546.1 474.5779920442
9 -1.27562123359356 8.981959346139

33e-27 
5.098160124868

68e-25 fumB fumarase B 

cds-NP_417026.1 1788.774338029
92 -1.27626620804831 5.011685133476

03e-11 
6.220041869739

79e-10 iscR DNA-binding transcriptional dual 
regulator IscR 

cds-NP_417149.1 32.78277097914
28 -1.28053419743938 2.586540674882

2e-05 
0.000122545273

249933 gabP 4-aminobutanoate:H(+) symporter 

cds-NP_416191.1 32424.94133049
93 -1.2837988650999 1.286497579445

86e-29 
8.978065894591

83e-28 pykF pyruvate kinase 1 

cds-NP_416414.1 81.10592300982
62 -1.29146777571371 1.350482024829

84e-05 
6.835911985375

32e-05 araF arabinose ABC transporter 
periplasmic binding protein 

cds-YP_588442.1 227.7172345557
34 -1.29223009782238 1.110457763767

09e-07 
8.610598725603

83e-07 ybdD PF04328 family protein YbdD 

cds-NP_415558.1 29.06322715774
62 -1.30237610342337 0.002891831041

70692 
0.008317922124

69349 csgD DNA-binding transcriptional dual 
regulator CsgD 

cds-NP_415262.1 77375.89612436
68 -1.30569691747638 2.879717857651

74e-22 
1.114450810911

22e-20 cydB cytochrome bd-I subunit 2 

cds-NP_417680.1 3911.414688394
21 -1.30815748797476 2.047322471174

01e-30 
1.556330671390

67e-28 gltD glutamate synthase subunit GltD 

cds-NP_415799.1 30.53291223852
97 -1.30858280766198 0.000136864605

283961 
0.000554888213

994115 osmB osmotically-inducible lipoprotein 
OsmB 

cds-NP_416363.1 302.2304095865
41 -1.32029035161484 4.392122926191

64e-16 
9.840666998314

63e-15 purT phosphoribosylglycinamide 
formyltransferase 2 

cds-NP_414985.1 1226.598104308
66 -1.3211086921704 3.277324349155

13e-08 
2.795905762395

47e-07 amtB ammonium transporter 

cds-NP_417208.1 18.28745170197
42 -1.3265160775958 0.003424882608

09253 
0.009674668389

28329 hypC hydrogenase maturation factor 
HypC 



Supplementary Material 

  196 

cds-NP_415263.1 1619.863008047
01 -1.34336007388537 2.508221830655

28e-32 
2.266721244944

28e-30 ybgE PF09600 family protein YbgE 

cds-NP_414859.1 351.1374322143
61 -1.34523977447901 1.370487022033

53e-17 
3.604115778895

7e-16 yahK aldehyde reductase NADPH-
dependent 

cds-NP_415916.1 183.3848260917
63 -1.34751793786193 1.254310692538

05e-12 
2.022575991717

61e-11 paaK phenylacetate-CoA ligase 

cds-NP_416032.1 27.25490864909
74 -1.34772282670952 0.002095589737

02926 
0.006295642562

12672 lsrD Autoinducer-2 ABC transporter 
membrane subunit LsrD 

cds-NP_416598.1 101502.5985032
11 -1.34891990112405 1.399472812531

65e-27 
8.510793947067

45e-26 gatZ 
putative tagatose-1 

6-bisphosphate aldolase 2 
chaperone 

cds-NP_415984.1 67.65269911607
06 -1.3512814336931 3.677016188054

13e-08 
3.093489706036

84e-07 narY nitrate reductase Z subunit beta 

cds-NP_416651.1 1611.853427454
92 -1.35332103746303 5.069991386305

17e-05 
0.000224355024

235978 preT dihydropyrimidine dehydrogenase 
(NAD(+)) subunit PreT 

cds-NP_415894.4 289.5044262935
15 -1.35828806379464 2.017311800864

64e-15 
4.251334819940

98e-14 uspF universal stress protein F 

cds-NP_415580.1 666.8448517005
78 -1.3641434070577 4.880687447728

35e-22 
1.871809591439

6e-20 pyrC dihydroorotase 

cds-NP_416052.1 105.8830039422
92 -1.36484255133529 1.150234487959

26e-07 
8.870558360946

65e-07 dgcZ diguanylate cyclase DgcZ 

cds-NP_417471.1 5770.834001415
15 -1.36613967102646 6.611487919490

4e-19 
1.968189096032

91e-17 hybO hydrogenase 2 small subunit 

cds-NP_416875.1 31.31762786671
39 -1.36724639087724 0.000904364796

998111 
0.003024258398

13115 frc formyl-CoA transferase 

cds-NP_417079.1 3258.397975470
84 -1.36803828741877 6.331765137947

49e-17 
1.522843174702

96e-15 patZ peptidyl-lysine N-acetyltransferase 

cds-NP_415516.1 10.34102724638
83 -1.36861279038725 0.020976668614

4159 
0.046124833828

2895 torC cytochrome c menaquinol 
dehydrogenase TorC 

cds-YP_010051192.1 40.20926207756
39 -1.37246322781523 8.903113881014

32e-06 
4.702302207379

4e-05 pssL protein PssL 

cds-NP_415924.1 1323.162903397
06 -1.37811478189269 1.965522090434

89e-13 
3.415317554377

48e-12 pdxI pyridoxal reductase 

cds-NP_417147.1 228.3863598956
38 -1.37982405135344 3.340996350548

25e-19 
1.015901533163

13e-17 gabD succinate-semialdehyde 
dehydrogenase (NADP(+)) GabD 

cds-NP_418003.1 9691.420739557
79 -1.38486616781332 0.000148587380

437904 
0.000595608736

840071 yhjX putative transporter YhjX 

cds-NP_416337.1 8474.210851934
55 -1.40129517409156 4.345944638709

53e-17 
1.057182075827

8e-15 cspC transcription antiterminator and 
regulator of mRNA stability CspC 

cds-NP_416299.1 23.98909623075
97 -1.40494220360313 0.004469443643

43605 
0.012141941027

5094 cdgI putative c-di-GMP binding protein 
CdgI 

cds-YP_588444.1 953.9978810912
12 -1.405514757787 1.321761276766

91e-27 
8.154692398835

86e-26 cydX cytochrome bd-I accessory 
subunit CydX 

cds-NP_417469.1 3475.675228279
18 -1.40690352687789 2.191455664309

59e-17 
5.552992120813

06e-16 hybB hydrogenase 2 membrane subunit 

cds-NP_415707.1 9758.229291316
66 -1.41492042453985 2.967881162556

32e-13 
5.074004059840

26e-12 dadA D-amino acid dehydrogenase 

cds-NP_416590.1 77.93873644515
75 -1.41671593860382 2.417587687247

43e-09 
2.404595977713

16e-08 yegS lipid kinase YegS 

cds-NP_416745.1 2295.934344417
71 -1.41731825513213 0.000407453761

569358 
0.001464975222

12902 glpB anaerobic glycerol-3-phosphate 
dehydrogenase subunit B 

cds-NP_418688.1 96.20631949386
35 -1.41826867341653 2.298526737532

89e-08 
1.984752195066

43e-07 idnD L-idonate 5-dehydrogenase 

cds-NP_416738.1 2896.020145575
7 -1.4242783917545 1.872891327956

46e-30 
1.449617887838

3e-28 nrdB ribonucleoside-diphosphate 
reductase 1 subunit beta 

cds-NP_415522.1 4027.639569770
39 -1.4307434486983 3.252814764054

73e-14 
6.046826397633

63e-13 agp glucose-1-phosphatase 

cds-NP_418576.1 835.5496985218
53 -1.43181563761046 4.162048265863

14e-15 
8.600892945523

98e-14 frdC fumarate reductase membrane 
protein FrdC 



Supplementary Material 

 197 

cds-NP_415234.1 14.89793200718
96 -1.44003453070718 0.008848041702

78604 
0.021962748413

2712 ybfD H repeat-associated putative 
transposase YbfD 

cds-NP_418496.1 18.62581331652
65 -1.44252089976168 0.001288962196

35656 
0.004147476999

1609 nrfC putative menaquinol-cytochrome c 
reductase 4Fe-4S subunit 

cds-NP_415375.1 97.48999031012
12 -1.44325104827522 3.057224924422

73e-11 
3.980001988766

83e-10 potF putrescine ABC transporter 
periplasmic binding protein 

cds-NP_416217.1 719.2297168033
14 -1.44497396625818 3.867528018686

79e-11 
4.871025673239

54e-10 ppsA phosphoenolpyruvate synthetase 

cds-NP_415829.1 70.09575122615
63 -1.44762827406212 1.623770309291

14e-07 
1.221270354532

22e-06 ycjQ D-guloside 3-dehydrogenase 

cds-NP_417345.1 3403.658472650
74 -1.45118768111139 7.270685447145

52e-12 
1.049196879610

12e-10 ygeV putative sigma(54)-dependent 
transcriptional regulator YgeV 

cds-NP_416413.1 86.63409335729
54 -1.46348362209173 2.262845430745

77e-08 
1.959996163945

66e-07 araG arabinose ABC transporter ATP 
binding subunit 

cds-NP_417148.1 324.1012712985
72 -1.4641522664664 4.672107014840

68e-33 
4.419813236039

28e-31 gabT 4-aminobutyrate aminotransferase 
GabT 

cds-YP_009518815.1 111.4622372189
7 -1.46443234187093 3.301778332761

37e-08 
2.811134072513

03e-07 yghO putative DNA-binding 
transcriptional regulator YghO 

cds-NP_415523.1 131.0234259995
1 -1.46582493108766 4.640238115211

47e-15 
9.496871950218

86e-14 yccJ PF13993 family protein YccJ 

cds-NP_415828.1 10.13023396646
29 -1.46914845071769 0.012747303011

8723 
0.030147371623

078 ycjP putative ABC transporter 
membrane subunit YcjP 

cds-NP_415116.1 164.0030484256
81 -1.47773649894806 2.002100218899

43e-09 
2.010127507512

94e-08 fepA ferric enterobactin outer 
membrane transporter 

cds-NP_415960.1 29.94645991301
19 -1.47774370255434 4.846550963143

66e-05 
0.000215813467

051282 ydcV putative ABC transporter 
membrane subunit YdcV 

cds-NP_418459.1 36770.61368870
36 -1.48136684390103 3.518010850763

15e-11 
4.483883889730

16e-10 malK maltose ABC transporter ATP 
binding subunit 

cds-NP_416054.1 11.12270346325
59 -1.48670206365164 0.006145400321

58606 
0.015903324722

7914 ydeI BOF family protein YdeI 

cds-NP_415156.1 3991.736497560
06 -1.49179245758419 2.615415504024

48e-11 
3.436365370565

49e-10 cspE transcription antiterminator and 
regulator of RNA stability CspE 

cds-NP_415130.1 8084.358535077
8 -1.49564518170062 2.919156833312

16e-06 
1.716415834172

63e-05 cstA pyruvate transporter CstA 

cds-YP_026279.1 2634.248765980
45 -1.50429147566416 1.966915450671

48e-23 
8.290256508424

23e-22 thiS sulfur carrier protein ThiS 

cds-NP_418575.1 1717.551958134
4 -1.50784573582593 2.433315802834

69e-11 
3.226986097404

13e-10 frdD fumarate reductase membrane 
protein FrdD 

cds-NP_416033.1 35.93842467916
5 -1.50949533927872 3.786517172187

77e-06 
2.157858581258

81e-05 lsrB Autoinducer-2 ABC transporter 
periplasmic binding protein 

cds-NP_414863.1 111.8423928195
96 -1.50955497179511 6.411261652322

86e-13 
1.066122689606

97e-11 yahO DUF1471 domain-containing 
protein YahO 

cds-NP_416437.1 322.5467629267
07 -1.51549934855063 1.303988381415

05e-23 
5.571550190000

85e-22 amyA alpha-amylase 

cds-NP_417985.1 13427.96758268
61 -1.51820013110674 5.345427721968

44e-12 
7.984380986813

91e-11 dctA C4 dicarboxylate/orotate:H(+) 
symporter 

cds-NP_417470.1 5696.783358132
18 -1.51841400060919 1.461387538308

15e-21 
5.317202350921

19e-20 hybA hydrogenase 2 iron-sulfur protein 

cds-NP_415419.2 202.0631585022
07 -1.51958486924128 8.302472261290

71e-09 
7.683396612242

29e-08 ycaM putative transporter YcaM 

cds-NP_418418.2 13520.81366144
85 -1.5221907788976 1.772555194838

53e-24 
8.478390409469

25e-23 thiG 1-deoxy-D-xylulose 5-
phosphate:thiol sulfurtransferase 

cds-NP_415392.1 408.6096727860
41 -1.52688840588938 6.281711693046

56e-24 
2.840941102868

45e-22 poxB pyruvate oxidase 

cds-NP_418630.1 281.6870948308
53 -1.5317907927259 2.230104379429

48e-17 
5.617487777059

93e-16 ytfE iron-sulfur cluster repair protein 
YtfE 

cds-NP_418028.1 2005.264886299
17 -1.53307991911338 7.676313249377

82e-16 
1.650407348616

23e-14 malS alpha-amylase 

cds-NP_415979.2 89.56066582432
92 -1.53449842393434 4.216919764672

65e-08 
3.512999498671

52e-07 yddH flavin reductase-like protein YddH 



Supplementary Material 

  198 

cds-NP_415959.1 27.20740760740
21 -1.537468615424 1.216446798108

11e-05 
6.236838846005

1e-05 ydcU putative ABC transporter 
membrane subunit YdcU 

cds-NP_415160.1 298.7478976414
57 -1.53773231744432 8.232679863368

57e-26 
4.4931433562e-

24 tatE twin arginine protein translocation 
system - TatE protein 

cds-NP_416746.1 712.1982047490
08 -1.55015984143393 0.001367264602

35582 
0.004356620817

53646 glpC anaerobic glycerol-3-phosphate 
dehydrogenase subunit C 

cds-NP_416338.1 2871.365985262
08 -1.55221264231728 7.411951879402

79e-20 
2.408601461879

21e-18 yobF DUF2527 domain-containing 
protein YobF 

cds-NP_416589.2 16.39176500036
06 -1.55404892887271 0.001673780694

6704 
0.005159510801

74649 yegR uncharacterized protein YegR 

cds-NP_418663.1 6541.461037171
63 -1.55508484027996 6.265266292182

29e-25 
3.030822568843

18e-23 mgtA Mg(2(+)) importing P-type ATPase 

cds-NP_416874.1 46.23889790190
51 -1.5553355426768 2.167826646231

69e-06 
1.310857675143

23e-05 oxc oxalyl-CoA decarboxylase 

cds-NP_415524.1 255.0805192065
32 -1.5563381806635 2.037483641168

2e-20 
6.883784016232

57e-19 wrbA NAD(P)H:quinone oxidoreductase 

cds-NP_418719.1 67.64251113343
79 -1.55729245242559 2.366163387738

91e-06 
1.424718181273

63e-05 yjhI putative DNA-binding 
transcriptional regulator YjhI 

cds-NP_417379.1 17141.37247446
01 -1.55909631328641 2.120751033815

74e-54 
5.642523219346

e-52 gcvP glycine decarboxylase 

cds-NP_415123.1 77.19739867623
04 -1.55988974692558 1.476406860944

9e-07 
1.114373050894

05e-06 entS enterobactin exporter EntS 

cds-NP_414916.1 30.19636187530
97 -1.56174644252491 0.000140175655

499122 
0.000566693034

624655 iraP anti-adaptor protein IraP 

cds-NP_416294.4 4099.762591472
93 -1.56396078475529 8.284286326720

85e-24 
3.635691432252

64e-22 yeaD putative aldose 1-epimerase YeaD 

cds-NP_418420.4 11816.85395489
03 -1.56755780735969 6.621116995636

28e-24 
2.936051567752

46e-22 thiF sulfur carrier protein ThiS 
adenylyltransferase 

cds-NP_418547.1 424.9625391950
49 -1.57172163639723 1.935371391504

4e-27 
1.160405072342

85e-25 dcuB anaerobic C4-dicarboxylate 
transporter DcuB 

cds-NP_417380.1 5214.942753379
23 -1.57989703014498 5.195561497345

03e-32 
4.423501058839

56e-30 gcvH glycine cleavage system H protein 

cds-NP_416484.2 1680.865883606
65 -1.582856636032 3.791633209279

86e-11 
4.789609071781

71e-10 mtfA Mlc titration factor 

cds-NP_416743.1 8487.714267261
33 -1.58478910255369 1.574841323343

94e-05 
7.850434897679

84e-05 glpT sn-glycerol 3-
phosphate:phosphate antiporter 

cds-NP_418690.4 535.5232606703
39 -1.59269938400571 3.271622762632

55e-24 
1.530472318739

21e-22 ahr NADPH-dependent aldehyde 
reductase Ahr 

cds-NP_416298.1 366.9796454765
81 -1.59300697708171 4.067828019994

35e-16 
9.162298349796

8e-15 yeaH DUF444 domain-containing 
protein YeaH 

cds-NP_418651.3 97.51596354806
86 -1.5978239893898 3.790913436088

52e-10 
4.202582942038

76e-09 ytfT galactofuranose ABC transporter 
putative membrane subunit YtfT 

cds-NP_416652.4 1947.303593847
9 -1.59834518117784 3.424381478831

38e-07 
2.417511103712

31e-06 preA dihydropyrimidine dehydrogenase 
(NAD(+)) subunit PreA 

cds-NP_415127.1 96.89705054871
56 -1.59961248254003 1.999922358144

9e-05 
9.785826986922

83e-05 entB enterobactin synthase component 
B 

cds-NP_417454.1 4503.955395193
06 -1.60524581416851 1.965592672827

07e-13 
3.415317554377

48e-12 glcC DNA-binding transcriptional dual 
regulator GlcC 

cds-NP_417279.2 1159.857676732
36 -1.61342422823291 5.677803157215

64e-15 
1.156478853601

29e-13 fucO L-1 2-propanediol oxidoreductase 

cds-NP_417586.1 450.5274722313
33 -1.61703788321945 8.910442313530

9e-10 
9.482938232175

26e-09 tdcC threonine/serine:H(+) symporter 

cds-NP_417400.1 56193.92104051
45 -1.61951000090575 6.497775298988

6e-47 
1.317191878466

4e-44 fbaA fructose-bisphosphate aldolase 
class II 

cds-NP_416959.1 382.2317239732
55 -1.61995171930171 1.812911822718

31e-20 
6.174052503449

47e-19 talA transaldolase A 

cds-NP_415050.1 36.61474211646
82 -1.62313706581691 3.724540848040

83e-07 
2.594986970558

07e-06 allD ureidoglycolate dehydrogenase 

cds-NP_416656.1 3310.632448362
97 -1.62364481212583 1.618830851473

64e-07 
1.219710253933

32e-06 galS DNA-binding transcriptional dual 
regulator GalS 



Supplementary Material 

 199 

cds-NP_415708.1 2699.784343284
16 -1.62475670865603 2.618250774000

89e-15 
5.490587953163

45e-14 dadX alanine racemase 2 

cds-NP_417904.1 254.1344713643
81 -1.62733229474134 9.415313256038

35e-34 
9.543092507370

3e-32 ggt glutathione hydrolase proenzyme 

cds-NP_415333.1 2819.504070698
06 -1.62905300437556 2.198257323759

08e-32 
2.034343788530

95e-30 dps DNA protection during starvation 
protein 

cds-NP_418421.1 12476.22008345
07 -1.63203066951253 6.097450864483

86e-27 
3.555732647959

97e-25 thiE thiamine phosphate synthase 

cds-NP_416960.1 920.2591749111
42 -1.63772173599299 5.144107211919

01e-30 
3.649744066856

54e-28 tktB transketolase 2 

cds-NP_415019.1 283.4075056902
9 -1.63882842033046 1.430948815812

95e-17 
3.714359212753

49e-16 ybaT putative transporter YbaT 

cds-NP_418417.1 16453.22227020
08 -1.64077375700918 2.555852002756

06e-32 
2.266721244944

28e-30 thiH 2-iminoacetate synthase 

cds-NP_416597.1 21891.64529361
7 -1.64956295430051 2.847473568292

47e-42 
4.662190377008

09e-40 gatA galactitol-specific PTS enzyme IIA 
component 

cds-YP_026272.1 256.4157188948
6 -1.65407839167329 1.863674457415

03e-08 
1.635806632003

26e-07 fadA 3-ketoacyl-CoA thiolase 

cds-NP_417158.4 125.2957507662
29 -1.66008106276208 4.419557212823

89e-15 
9.088915485503

05e-14 ygaM DUF883 domain-containing 
protein YgaM 

cds-NP_415326.1 183.3177478553
15 -1.67536103899202 3.189277237765

45e-09 
3.078628843802

16e-08 fiu iron catecholate outer membrane 
transporter Fiu 

cds-NP_416902.1 9.488690515627
95 -1.67780175614881 0.005192918384

10668 
0.013816408475

7138 xapA xanthosine phosphorylase 

cds-NP_416288.1 58.40891750962
84 -1.67964759831768 4.188959718712

13e-08 
3.496549318148

53e-07 ydjJ putative zinc-binding 
dehydrogenase YdjJ 

cds-NP_418422.1 55115.07136252
65 -1.68267122672189 4.241062903480

14e-28 
2.735485572744

69e-26 thiC phosphomethylpyrimidine 
synthase 

cds-NP_418491.1 111.8723506690
21 -1.68306898101356 9.597649879527

75e-11 
1.144459258743

69e-09 actP acetate/glycolate:cation symporter 

cds-NP_414576.4 604.3057611172
95 -1.69257831083026 2.953307366539

49e-11 
3.856512104097

74e-10 caiF DNA-binding transcriptional 
activator CaiF 

cds-NP_416028.3 271.1867049016
04 -1.70133264889363 9.209813078942

89e-18 
2.513216299811

53e-16 lsrK autoinducer-2 kinase 

cds-NP_416843.1 1054.163081251
87 -1.70926589445209 5.255390493542

32e-13 
8.807951705121

91e-12 fadJ 3-hydroxyacyl-CoA 
dehydrogenase FadJ 

cds-NP_414605.1 80.22599670406
51 -1.72437152147369 3.632501007235

22e-10 
4.037482190026

19e-09 araB ribulokinase 

cds-NP_417378.1 502.5840908925
94 -1.72470084825245 6.862328517714

62e-28 
4.296019485281

05e-26 ygfF putative oxidoreductase YgfF 

cds-NP_414937.2 1423.865739964
02 -1.72590372074541 1.691408993680

97e-45 
3.130577428739

08e-43 malZ maltodextrin glucosidase 

cds-NP_416297.1 1072.966589731
6 -1.73298745581004 6.042953427416

4e-31 
4.853745800096

53e-29 yeaG protein kinase YeaG 

cds-NP_415540.1 45.10788943225
01 -1.73707449041295 5.222476508035

4e-09 
4.951466034455

83e-08 pgaD poly-N-acetyl-D-glucosamine 
synthase subunit PgaD 

cds-NP_416739.1 422.7478700901
68 -1.74634779841713 9.041586106917

6e-21 
3.129270898955

14e-19 yfaE 
ferredoxin-like diferric-tyrosyl 
radical cofactor maintenance 

protein YfaE 

cds-YP_009518758.1 8.945391040897
38 -1.74673274470853 0.002833758406

25047 
0.008172973939

97848 ybfQ inactive transposase YbfQ 

cds-NP_418754.2 148.6568375173
73 -1.75379024669244 2.311934326666

49e-13 
4.000774157975

3e-12 yjiL putative ATPase activator of (R)-
hydroxyglutaryl-CoA dehdratase 

cds-NP_418755.4 468.0353507779
45 -1.75568196019573 9.604503414900

59e-22 
3.555336611933

2e-20 yjiM putative dehydratase subunit 

cds-NP_415947.1 1208.764730867
06 -1.76790873552708 9.364109756141

e-22 
3.496755722095

81e-20 tehB tellurite methyltransferase 

cds-NP_417909.1 18.63615946405
51 -1.7766922530958 0.000187647001

908218 
0.000731513999

197148 ugpA 
sn-glycerol 3-phosphate ABC 
transporter membrane subunit 

UgpA 



Supplementary Material 

  200 

cds-NP_418661.1 6677.817284736
86 -1.7768336268528 2.777381834391

2e-07 
2.001246716933

35e-06 treB trehalose-specific PTS enzyme 
IIBC component 

cds-NP_416314.1 526.8863972574
59 -1.79364990532732 2.584241774236

35e-21 
9.322980705867

93e-20 dmlA D-malate/3-isopropylmalate 
dehydrogenase (decarboxylating) 

cds-NP_415706.1 301.9259522365
96 -1.82913004088183 1.970206165525

33e-31 
1.612916855123

33e-29 ycgB PF04293 family protein YcgB 

cds-NP_416596.1 7989.555437547
38 -1.83153347023635 1.184693244629

02e-35 
1.293138241637

36e-33 gatB galactitol-specific PTS enzyme IIB 
component 

cds-NP_415933.1 14740.81201545
98 -1.83438656445747 2.080213680928

94e-08 
1.810934486649

19e-07 aldA aldehyde dehydrogenase A 

cds-NP_415423.1 58905.98131048
88 -1.83481812309488 1.776804097544

29e-37 
2.101070845346

12e-35 pflB pyruvate formate-lyase 

cds-NP_415690.5 19.84347756703
91 -1.83593733200263 0.000348489336

929973 
0.001273406959

06515 ymgG PF13436 family protein YmgG 

cds-NP_415906.1 12.40186234853
46 -1.84051477934112 0.000312069420

653561 
0.001149203740

24413 paaA phenylacetyl-CoA 1 2-epoxidase 
monooxygenase subunit 

cds-NP_414864.1 77.21292756987
87 -1.84485256125833 4.952220152662

29e-17 
1.197818249425

19e-15 prpR DNA-binding transcriptional dual 
regulator PrpR 

cds-NP_414851.1 27.85630913240
53 -1.85609891395123 4.418688398158

e-06 
2.481577376115

91e-05 yahC uncharacterized protein YahC 

cds-NP_416655.1 4110.755856113
97 -1.86814868863919 4.515167140435

93e-16 
1.006338561090

88e-14 mglB 
D-galactose/methyl-galactoside 

ABC transporter periplasmic 
binding protein 

cds-NP_415417.1 50.16134356437
95 -1.88349170548725 5.798040351697

89e-12 
8.630160061950

33e-11 ycaC putative hydrolase YcaC 

cds-NP_416873.1 15.17828521221
03 -1.88384336322819 0.000306376080

224621 
0.001130977228

33761 yfdV putative transport protein YfdV 

cds-NP_417585.2 230.1403874086
64 -1.89682771245858 3.094863269960

46e-11 
4.016717359823

69e-10 tdcD propionate kinase 

cds-NP_417184.1 922.2386446009
12 -1.90678325553418 1.410959129570

16e-17 
3.684940499742

44e-16 srlB sorbitol-specific PTS enzyme IIA 
component 

cds-NP_418011.1 140.7192179270
07 -1.91585659148442 7.538244774375

e-20 
2.431083939735

94e-18 yiaG putative DNA-binding 
transcriptional regulator YiaG 

cds-NP_417280.1 101.6026107176
73 -1.92803804117236 2.793969952411

61e-14 
5.239616778597

46e-13 fucA L-fuculose-phosphate aldolase 

cds-NP_416415.1 872.9462849174
84 -1.93423705636296 1.167498134270

45e-08 
1.062393338293

14e-07 ftnB putative ferritin-like protein 

cds-NP_415557.1 7.006415684711
96 -1.93774755284353 0.013895824319

9213 
0.032360242959

4667 csgE curli assembly component CsgE 

cds-NP_417587.1 937.5163131029
54 -1.94049024994121 6.805182703741

38e-07 
4.540699493703

3e-06 tdcB catabolic threonine dehydratase 

cds-NP_415986.1 25.10866010688
66 -1.94989672747216 4.791710010852

21e-06 
2.676943506062

71e-05 narU nitrate/nitrite transporter NarU 

cds-YP_026286.1 217.0143679535
65 -1.95317488544804 1.868890210975

68e-25 
9.944832035154

36e-24 ytfR galactofuranose ABC transporter 
putative ATP binding subunit 

cds-NP_417193.1 61.93734963142
93 -1.95575406918133 2.536741768091

35e-09 
2.498806259635

57e-08 hydN putative electron transport protein 
HydN 

cds-NP_416844.1 1948.121368842
86 -1.96755593633448 2.791262806306

21e-07 
2.007163136223

91e-06 fadI 3-ketoacyl-CoA thiolase FadI 

cds-NP_415534.1 5654.354622804
36 -1.97279071882608 5.674047064121

63e-18 
1.589106470524

06e-16 putA 

fused DNA-binding transcriptional 
repressor/proline 

dehydrogenase/1-pyrroline-5-
carboxylate dehydrogenase PutA 

cds-NP_415826.1 23.75789389051
74 -1.98444325061853 3.404998173017

43e-05 
0.000156028818

326536 ycjN putative ABC transporter 
periplasmic binding protein YcjN 

cds-NP_416742.1 3822.886923093
93 -2.0057199727695 3.394946775873

3e-10 
3.793251555089

93e-09 glpQ glycerophosphoryl diester 
phosphodiesterase GlpQ 

cds-NP_418660.1 3167.705827070
12 -2.01090434892884 2.998509532599

26e-19 
9.249750058170

34e-18 treC trehalose-6-phosphate hydrolase 

cds-NP_416805.1 21.81344926117
78 -2.01176564050043 8.359330207839

89e-07 
5.483153882091

59e-06 yfcG disulfide bond oxidoreductase 
YfcG 



Supplementary Material 

 201 

cds-NP_417269.1 66.79334860565
83 -2.01441537149656 1.070946630015

2e-08 
9.825473715462

69e-08 gudP galactarate/D-glucarate 
transporter GudP 

cds-NP_415757.1 174154.2265167
24 -2.02663806061093 1.422255857813

87e-39 
1.953078447326

98e-37 adhE 

fused acetaldehyde-CoA 
dehydrogenase and iron-

dependent alcohol 
dehydrogenasealdehyde/alcohol 

dehydrogenase AdhE 

cds-YP_010051189.1 21.49831337399
85 -2.02911760001772 2.270486099419

56e-05 
0.000109090963

038703 yoaM protein YoaM 

cds-NP_416031.1 62.90744090621
13 -2.03815285222424 8.939185659692

19e-15 
1.745601529968

33e-13 lsrC Autoinducer-2 ABC transporter 
membrane subunit LsrC 

cds-NP_418541.2 912.2971050461
17 -2.03893761925043 2.760786267847

42e-22 
1.078226343323

53e-20 adiA arginine decarboxylase 
degradative 

cds-YP_026218.1 30845.38151732
85 -2.086564245994 7.033336056298

82e-39 
9.356534872395

03e-37 malP maltodextrin phosphorylase 

cds-NP_415713.1 16.81751762388
6 -2.10418555494769 6.922073866329

39e-05 
0.000299160085

776286 ymgE PF04226 family protein YmgE 

cds-NP_418023.1 42.48516130789
52 -2.11484183564336 6.808387950152

98e-09 
6.369957693143

13e-08 xylF xylose ABC transporter 
periplasmic binding protein 

cds-NP_416442.1 40.13785569700
21 -2.13417185890083 2.745919629039

14e-09 
2.681050426793

49e-08 yedL putative acetyltransferase YedL 

cds-NP_414913.1 65.95751657641
42 -2.14343422727279 2.724257106997

32e-12 
4.232541060031

97e-11 yaiY DUF2755 domain-containing inner 
membrane protein YaiY 

cds-NP_418493.1 1356.873904942
62 -2.16756569756791 2.375422947683

34e-10 
2.696580130210

13e-09 acs acetyl-CoA synthetase (AMP-
forming) 

cds-NP_416949.1 6.742384974609
63 -2.1697366266316 0.004172719434

53805 
0.011482396013

4638 eutJ putative ethanolamine utilization 
chaperonin EutJ 

cds-NP_417198.1 149.5084824214
43 -2.18143053503142 1.269014696895

45e-18 
3.751524697697

18e-17 hycH formate hydrogenlyase assembly 
protein 

cds-NP_415539.1 568.2235972416
96 -2.18280049900862 1.548991624574

81e-09 
1.570013653765

47e-08 phoH ATP-binding protein PhoH 

cds-NP_416289.1 33.57857141025
41 -2.19448257746658 4.092010327241

12e-10 
4.512872529291

57e-09 ydjK putative transporter YdjK 

cds-NP_416600.4 471.1007428743
54 -2.19548361604289 3.185084075385

77e-40 
4.519634302972

41e-38 fbaB fructose-bisphosphate aldolase 
class I 

cds-NP_418458.1 24975.49549012
95 -2.20382621177071 1.015572012070

88e-37 
1.235225730110

21e-35 malE maltose ABC transporter 
periplasmic binding protein 

cds-NP_415997.1 66.22036099466
53 -2.20943601630159 2.022222596028

22e-12 
3.176605753244

33e-11 sra ribosome-associated protein Sra 

cds-NP_416769.1 859.2862178381
8 -2.21058298626689 5.495726667207

e-25 
2.689115910609

22e-23 elaB tail-anchored inner membrane 
protein ElaB 

cds-NP_418648.1 326.7869946677
71 -2.22106508326753 1.570627733428

1e-29 
1.078413267936

04e-27 ytfQ galactofuranose ABC transporter 
periplasmic binding protein 

cds-NP_415126.1 95.07837122426
74 -2.23413332400979 4.469193883514

e-10 
4.890837625223

42e-09 entE 2 3-dihydroxybenzoate-AMP 
ligase 

cds-NP_415018.1 192.6334505764
87 -2.24795474749435 3.895761522454

86e-21 
1.393635025301

71e-19 glsA glutaminase 1 

cds-NP_417146.2 93.92346483914
8 -2.25164446068121 5.892801078791

96e-23 
2.389109923087

37e-21 lhgD L-2-hydroxyglutarate 
dehydrogenase 

cds-NP_414880.2 2339.413806853
21 -2.25605786936225 7.805674665159

27e-07 
5.143770441111

92e-06 mhpR DNA-binding transcriptional 
activator MhpR 

cds-NP_416319.1 2826.644070602
07 -2.26410777857501 8.955903910019

84e-10 
9.507551856597

12e-09 fadD long-chain-fatty-acid--CoA ligase 

cds-NP_418336.1 6.089181404117
42 -2.27993426808108 0.007341949389

72478 
0.018659509583

3184 frvA putative PTS enzyme IIA 
component FrvA 

cds-YP_001165313.1 9.550565847332
85 -2.29215895286888 0.001051869544

8644 
0.003449775541

20781 appX cytochrome bd-II accessory 
subunit AppX 

cds-NP_414874.1 10.19082624281
8 -2.2948030904916 0.000495980584

496088 
0.001752190330

4563 cynS cyanase 

cds-NP_418045.4 150.6307555668
12 -2.29608946447002 1.424604183231

73e-31 
1.189125491768

13e-29 aldB aldehyde dehydrogenase B 



Supplementary Material 

  202 

cds-NP_415125.1 70.23112095305
18 -2.29804061424974 5.485162520666

93e-08 
4.439227538113

9e-07 entC isochorismate synthase EntC 

cds-NP_417552.1 1410.582274923
43 -2.31182045633587 1.101380999661

6e-06 
7.082445491781

6e-06 fadH 2 4-dienoyl-CoA reductase 

cds-NP_415940.1 330.9489127904
68 -2.35608876333493 6.339896752936

4e-24 
2.840941102868

45e-22 ydcJ DUF1338 domain-containing 
protein YdcJ 

cds-YP_025308.1 181.4977481902
87 -2.39048332344961 9.229065137899

6e-35 
9.822032573009

65e-33 katE catalase HPII 

cds-NP_417145.4 117.6972761148
02 -2.39272623669237 3.199531731476

8e-18 
9.265582708093

01e-17 glaH glutarate dioxygenase GlaH 

cds-NP_417185.1 10286.95795060
23 -2.39596250030627 2.144792780822

9e-25 
1.127207761476

93e-23 srlD sorbitol-6-phosphate 2-
dehydrogenase 

cds-NP_417352.2 1987.218558287
23 -2.39690814868829 3.507941036221

35e-08 
2.962957339522

68e-07 yqeC uncharacterized protein YqeC 

cds-NP_418492.1 33.32285817848
2 -2.39717277132887 1.205694681610

47e-07 
9.264697219523

06e-07 yjcH DUF485 domain-containing inner 
membrane protein YjcH 

cds-NP_416654.1 1388.374083929
16 -2.43477532299453 3.887911275800

08e-09 
3.719289505860

89e-08 mglA 
D-galactose/methyl-galactoside 
ABC transporter ATP binding 

subunit 

cds-NP_415957.1 150.2590142156
61 -2.45328926178031 4.462214562951

48e-30 
3.219601253302

45e-28 ydcS 

putative ABC transporter 
periplasmic binding 

protein/polyhydroxybutyrate 
synthase 

cds-NP_417875.1 15701.06115001
4 -2.45993177439711 2.486962539157

4e-30 
1.857368338454

92e-28 malQ 4-alpha-glucanotransferase 

cds-NP_418457.1 10679.51232957
53 -2.4657790998801 3.906080885898

27e-32 
3.393507414544

68e-30 malF maltose ABC transporter 
membrane subunit MalF 

cds-NP_418503.1 930.1156076512
03 -2.48347119921441 3.830952689884

94e-41 
5.623574345117

31e-39 fdhF formate dehydrogenase H 

cds-YP_002791249.1 9.128037646036
4 -2.53419668107616 0.001088638785

13624 
0.003562133211

62565 yohP putative membrane protein YohP 

cds-NP_416030.1 187.4450760427
89 -2.56328087630518 3.454821808356

05e-20 
1.148998159232

17e-18 lsrA Autoinducer-2 ABC transporter 
ATP binding subunit 

cds-NP_417597.1 1035.388026483
83 -2.61772945583377 1.370631200495

85e-14 
2.604811169870

92e-13 garD GarD 

cds-NP_418288.1 1093.773214404
69 -2.67620727875303 6.834675247732

42e-07 
4.553241397433

01e-06 fadB 

multifunctional enoyl-CoA 
hydratase 3-hydroxyacyl-CoA 

epimerase Delta(3)-cis- Delta(2)-
trans-enoyl-CoA isomerase L-3-

hydroxyacyl-CoA dehydrogenase 

cds-NP_417351.1 2037.676974814
92 -2.72135590850613 1.891990972157

31e-14 
3.579646919321

64e-13 yqeB XdhC-CoxI family protein YqeB 

cds-NP_415958.1 60.50741919651
67 -2.74754406302167 6.145366953997

43e-18 
1.709857981906

34e-16 ydcT putative ABC transporter ATP-
binding protein YdcT 

cds-NP_417595.1 109.0990447232
26 -2.7478178937467 4.010733875235

13e-16 
9.081752184508

48e-15 garL alpha-dehydro-beta-deoxy-D-
glucarate aldolase 

cds-NP_414756.2 1532.269030477
09 -2.75747939358666 4.357478915362

22e-08 
3.615943029765

49e-07 fadE acyl-CoA dehydrogenase 

cds-NP_415500.1 474.6751741493
03 -2.80279015883576 1.761676121524

88e-41 
2.777576018270

89e-39 appA 
periplasmic phosphoanhydride 
phosphatase/multiple inositol-
polyphosphate phosphatase 

cds-NP_418460.1 27673.88358975
59 -2.83600540150556 3.948224674133

29e-46 
7.639814744447

92e-44 lamB 
maltose outer membrane 

channel/phage lambda receptor 
protein 

cds-NP_417971.1 1267.498407699
92 -2.86074827824636 1.124071984183

13e-65 
4.350158578788

71e-63 mdtF multidrug efflux pump RND 
permease MdtF 

cds-NP_416957.4 4.582142717906
72 -2.88764949042536 0.007434524500

66356 
0.018861007627

7263 eutS 
putative ethanolamine catabolic 
microcompartment shell protein 

EutS 

cds-NP_417199.1 153.8249749384
79 -2.9067838425433 3.014735108096

86e-25 
1.546232211466

06e-23 hycG formate hydrogenlyase subunit 
HycG 

cds-NP_418461.1 3816.451058259
29 -2.94384272113758 8.939958405022

74e-45 
1.522296117207

27e-42 malM maltose regulon periplasmic 
protein 



Supplementary Material 

 203 

cds-NP_418456.1 1959.601346768
86 -2.96208967253374 2.479380716339

68e-52 
5.555117741820

02e-50 malG maltose ABC transporter 
membrane subunit MalG 

cds-NP_417047.1 3030.237647276
76 -3.05910345296335 1.093038961947

27e-57 
3.323619186435

37e-55 hmp nitric oxide dioxygenase 

cds-NP_417206.1 35.14432160569
08 -3.06545114250494 2.411586539468

e-09 
2.404244472720

21e-08 hypA hydrogenase 3 nickel 
incorporation protein HypA 

cds-NP_417969.1 195.7063024033
97 -3.07330588132621 4.569447859013

27e-25 
2.288487004214

06e-23 gadE DNA-binding transcriptional 
activator GadE 

cds-NP_417596.1 316.6957065844
05 -3.10549306512647 6.720368590211

92e-15 
1.355858250641

33e-13 garP galactarate/D-glucarate 
transporter GarP 

cds-NP_417970.1 232.2298346067
79 -3.12295520264439 3.410775278854

02e-33 
3.376667526065

48e-31 mdtE multidrug efflux pump membrane 
fusion protein MdtE 

cds-NP_416653.1 642.8599145638
75 -3.15634499939536 4.849276269589

67e-12 
7.294476706587

71e-11 mglC 
D-galactose/methyl-galactoside 

ABC transporter membrane 
subunit 

cds-NP_417200.1 123.7834809112
14 -3.17254961505438 8.403615982293

64e-27 
4.834350437381

63e-25 hycF formate hydrogenlyase subunit 
HycF 

cds-NP_417963.4 427.4970093562
68 -3.29818394935767 4.364488291206

4e-38 
5.464596075195

77e-36 slp starvation lipoprotein 

cds-NP_418539.1 2082.538460230
01 -3.32186258941022 4.560259415877

34e-59 
1.493309564106

91e-56 adiC arginine:agmatine antiporter 

cds-NP_417974.1 574.8032304543
31 -3.45458365198746 3.892483507048

25e-53 
9.205723494169

12e-51 gadA glutamate decarboxylase A 

cds-NP_417964.1 48.92011024338
44 -3.4837717046488 1.300303022982

51e-16 
3.008364113498

13e-15 dctR putative DNA-binding 
transcriptional regulator DctR 

cds-NP_418164.4 3848.911659106
44 -3.60529816094719 1.896774644900

39e-28 
1.242241486667

84e-26 tnaA tryptophanase 

cds-NP_417205.1 37.84374882569
24 -3.61691231692054 1.492706816730

95e-10 
1.740946005157

17e-09 hycA regulator of the transcriptional 
regulator FhlA 

cds-NP_417202.1 235.1334728617
98 -3.62237291342426 4.573777491683

13e-37 
5.262316427593

26e-35 hycD formate hydrogenlyase subunit 
HycD 

cds-NP_417201.1 377.4558417983
58 -3.63264735268967 2.032825919400

46e-53 
5.090435258169

27e-51 hycE formate hydrogenlyase subunit 
HycE 

cds-NP_417203.1 340.6448002896
95 -3.81077032478555 7.563866276194

97e-39 
9.757387496291

51e-37 hycC formate hydrogenlyase subunit 
HycC 

cds-NP_417965.1 116.9775474242
67 -4.03770724457128 4.654833093287

48e-25 
2.304142381177

3e-23 yhiD inner membrane protein YhiD 

cds-NP_415495.1 193.5608316584
22 -4.06012629603409 1.358858771368

08e-13 
2.400274601541

05e-12 hyaE putative HyaA chaperone 

cds-NP_415497.1 1245.690279476
66 -4.0737132771598 8.905141859952

84e-57 
2.527279259854

61e-54 appC cytochrome bd-II subunit 1 

cds-NP_415498.1 1443.351029298
93 -4.1182068041067 6.180644103523

81e-83 
3.758714564100

12e-80 appB cytochrome bd-II subunit 2 

cds-NP_417204.1 47.86153764001
89 -4.21320969644875 8.697042835157

09e-15 
1.706143380150

4e-13 hycB formate hydrogenlyase subunit 
HycB 

cds-NP_417968.1 3240.549274029
7 -4.21426442219136 2.554502491636

41e-101 
3.624839035632

07e-98 hdeD acid-resistance membrane protein 

cds-NP_418163.1 115.2645123208
45 -4.24303181230724 1.780047877093

99e-48 
3.788831906394

56e-46 tnaC tnaAB operon leader peptide 

cds-NP_415496.1 345.9581840863
63 -4.25939749539409 5.147379847554

37e-66 
2.191239601103

9e-63 hyaF protein HyaF 

cds-NP_416010.1 657.0809259000
64 -4.34875843459001 1.490956710374

39e-92 
1.269400543212

76e-89 gadB glutamate decarboxylase B 

cds-NP_416009.1 668.1555314682
67 -4.48936542670871 1.588294723121

96e-89 
1.126895106055

03e-86 gadC L-glutamate:4-aminobutyrate 
antiporter 

cds-NP_418165.1 533.4896689973
93 -4.50889516152956 4.939288596125

05e-20 
1.617427042592

64e-18 tnaB tryptophan:H(+) symporter TnaB 

cds-NP_415492.1 2337.181442787
66 -4.56729215045044 2.120153433956

86e-45 
3.760622153480

99e-43 hyaB hydrogenase 1 large subunit 



Supplementary Material 

  204 

cds-NP_415491.1 1590.458375451
15 -4.65336765570616 7.717196138069

93e-81 
4.106512994970

46e-78 hyaA hydrogenase 1 small subunit 

cds-NP_415494.1 1319.596516223 -4.71942796520138 2.793400553598
28e-41 

4.246966484524
25e-39 hyaD putative hydrogenase 1 maturation 

protease HyaD 

cds-NP_415493.1 1132.650774507
86 -4.96515581661892 3.327046258885

2e-96 
3.540808981018

57e-93 hyaC hydrogenase 1 cytochrome b 
subunit 

cds-NP_417967.1 9328.477129582
23 -5.32613428879488 2.956096071124

97e-133 
6.292050487389

5e-130 hdeA periplasmic acid stress chaperone 
HdeA 

cds-NP_417966.4 1679.488522695
56 -5.46492044230663 4.957740844043

48e-182 
2.110510277309

31e-178 hdeB periplasmic acid stress chaperone 
HdeB 

cds-NP_417277.1 1718.462980645
61 1.00123741921778 4.445086264300

33e-11 
5.549188336400

74e-10 sdaB L-serine deaminase II 

cds-NP_416773.1 22.29720630161
33 1.0013847683626 0.005740766776

68458 
0.015002114283

8221 yfbK IPR002035/DUF3520 domain-
containing protein YfbK 

cds-NP_415295.1 165.7772617251
04 1.00370055623294 2.018563747709

95e-05 
9.865701347877

43e-05 bioA adenosylmethionine-8-amino-7-
oxononanoate aminotransferase 

cds-NP_417053.2 96.92744751379
27 1.00520919748888 7.096653952962

36e-06 
3.858295769828

96e-05 mltF membrane-bound lytic murein 
transglycosylase F 

cds-NP_415371.1 444.0526895702
16 1.00697791530529 1.917101497847

54e-08 
1.679238904596

09e-07 ybjC DUF1418 domain-containing 
protein YbjC 

cds-NP_414546.1 47.15430992097
92 1.010796482824 0.001237871475

31031 
0.004010364437

13547 yaaX DUF2502 domain-containing 
protein YaaX 

cds-NP_415596.1 133.5854476898
87 1.01208684778221 0.000447692509

541197 
0.001594834320

5999 flgG flagellar basal-body rod protein 
FlgG 

cds-NP_416879.4 1210.055228232
56 1.01337160375144 2.341157802071

17e-07 
1.704820883622

89e-06 lpxP palmitoleoyl acyltransferase 

cds-NP_417392.1 72.09928311560
07 1.01807903847024 1.790479717666

42e-06 
1.099866112280

8e-05 scpA methylmalonyl-CoA mutase 

cds-NP_417883.1 189.1926899135
36 1.02589705388234 7.069133338769

54e-11 
8.573589921123

06e-10 glpE thiosulfate sulfurtransferase GlpE 

cds-NP_416454.1 21.50275755406
16 1.02699887806083 0.022265974044

3832 
0.048434466789

4427 fliL flagellar protein FliL 

cds-NP_415874.1 200.1924996914
27 1.03770973208177 1.965965743365

91e-11 
2.631797537581

34e-10 racR DNA-binding transcriptional 
repressor RacR 

cds-NP_415546.2 28.65759898866
09 1.03829998133728 0.002226960736

25955 
0.006615611901

08645 insE4 IS3 element protein InsE 

cds-NP_418142.1 552.0981102414
1 1.04149747184673 2.850115170322

16e-06 
1.682793381423

23e-05 ibpA small heat shock protein IbpA 

cds-NP_417870.1 63.49153803960
14 1.04274423957403 1.295418768937

14e-05 
6.604308621994

49e-05 rpnA recombination-promoting nuclease 
RpnA 

cds-NP_417090.1 1193.795171921
42 1.04613548163361 1.049135477918

48e-14 
2.011788166440

99e-13 pheA fused chorismate 
mutase/prephenate dehydratase 

cds-NP_416453.1 27.35227925499
27 1.05242344874263 0.000546105928

771253 
0.001911819850

96976 fliK flagellar hook-length control 
protein 

cds-NP_418198.1 1063.266353422
84 1.0534249580296 8.285876160321

75e-12 
1.179698154330

76e-10 mioC flavoprotein MioC 

cds-NP_414655.1 1696.177986472
73 1.05621149766691 3.236863679216

8e-16 
7.408241227110

7e-15 pdhR DNA-binding transcriptional dual 
regulator PdhR 

cds-NP_417959.4 39.07376500042
24 1.06214695528566 0.000906035832

856288 
0.003027468242

12655 arsB arsenite/antimonite:H(+) antiporter 

cds-NP_417628.2 26.70273219621
74 1.06445145708504 0.019300978388

68 
0.043063031970

9699 ubiV ubiquinone biosynthesis protein 
UbiV 

cds-NP_417911.4 72.40172556080
62 1.07019742379872 7.004141503400

67e-06 
3.812868335035

38e-05 livF 

branched chain amino 
acid/phenylalanine ABC 

transporter ATP binding subunit 
LivF 

cds-NP_417515.1 13.11135614011
17 1.07434762332927 0.020648866526

0819 
0.045545194197

6843 ygiL putative fimbrial protein YgiL 

cds-NP_414924.1 162.5383638944
77 1.07460248566403 7.896530718408

75e-08 
6.259875468950

85e-07 aroM protein AroM 



Supplementary Material 

 205 

cds-NP_416145.1 177.4433800430
95 1.07817016760122 5.642558490021

05e-07 
3.806714974963

49e-06 rsxB SoxR [2Fe-2S] reducing system 
protein RsxB 

cds-NP_418359.1 1135.506260748
32 1.08110508767611 2.378918304255

35e-24 
1.125228357912

78e-22 fpr flavodoxin/ferredoxin-NADP(+) 
reductase 

cds-NP_418729.1 26.52374366241
58 1.08527203786662 0.019784724201

0155 
0.043935091770

3302 nanS N-acetyl-9-O-acetylneuraminate 
esterase 

cds-NP_415008.1 497.7414432454
82 1.08921321461179 1.082576305602

37e-25 
5.833578902467

45e-24 hemH ferrochelatase 

cds-NP_414790.1 278.8355330180
69 1.09477517941315 3.891349678178

69e-17 
9.520388264371

65e-16 insI1 IS30 family transposase 

cds-YP_009518795.2 84.64773598648
15 1.09552346829118 0.001941814402

68057 
0.005904502794

43655 yoaL protein YoaL 

cds-NP_414788.1 55.04036607967
91 1.09600194665676 1.512474146073

77e-05 
7.583748456815

14e-05 perR putative transcriptional regulator 
PerR 

cds-NP_417942.6 649.5910253241
45 1.097060812212 9.934114779991

03e-13 
1.620288376184

74e-11 yhhJ ABC transporter family protein 
YhhJ 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.09748823978663 0.002207387842
02241 

0.006562046119
75518 insO IS911B regulator fragment 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.09748823978663 0.002207387842
02241 

0.006562046119
75518 insO IS911B regulator fragment 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.09748823978663 0.002207387842
02241 

0.006562046119
75518 insO IS911B regulator fragment 

cds-NP_414746.1 89.91894266638
79 1.09963686059912 1.577114611444

72e-05 
7.852370644351

07e-05 yafE putative S-adenosylmethionine-
dependent methyltransferase 

cds-NP_416808.1 40.70327276883
25 1.10221011334146 0.000769097172

988954 
0.002610882508

30461 rpnB recombination-promoting nuclease 
RpnB 

cds-NP_415121.1 80.42888071154
42 1.10493192535106 0.003016701288

30299 
0.008630441790

5281 fepG 
ferric enterobactin ABC 

transporter membrane subunit 
FepG 

cds-NP_415355.1 69.38485796084
07 1.11059953455406 0.001496814556

87864 
0.004702538427

03495 dgcI putative diguanylate cyclase DgcI 

cds-NP_417785.1 26.19826490955
49 1.11225610084238 0.009339038386

06129 
0.023033769646

2705 gspE Type II secretion system protein 
GspE 

cds-NP_418388.1 33.48311635901
06 1.11371474863691 0.002866135281

81807 
0.008260756868

44924 frwD putative PTS enzyme IIB 
component FrwD 

cds-NP_415060.1 38.31916220756
06 1.1156579684183 0.001155155349

60833 
0.003762430239

69598 ybcI PF04307 family inner membrane 
protein YbcI 

cds-NP_415098.1 17.86704125140
09 1.11893487268066 0.009108237629

67001 
0.022516705917

2504 envY DNA-binding transcriptional 
activator EnvY 

cds-NP_418435.1 58.15345329252
62 1.11933483556655 1.520473369730

95e-05 
7.614888394052

51e-05 yjaA stress response protein 

cds-NP_417189.2 165.3897160296
51 1.12747970890308 9.374830757454

18e-13 
1.534948251326

25e-11 norR DNA-binding transcriptional dual 
regulator NorR 

cds-NP_415565.1 142.6254327754
88 1.13207443501821 2.704478547142

27e-08 
2.325851550542

35e-07 opgC 
osmoregulated periplasmic 
glucans (OPG) biosynthesis 

protein C 

cds-NP_416335.4 100.3404221915
42 1.14385540437436 5.797156719303

74e-06 
3.188436195617

06e-05 mntP Mn(2(+)) exporter 

cds-NP_416482.1 49.57070942379
01 1.14449458857379 1.099625530302

92e-05 
5.694776012773

17e-05 zinT metal-binding protein ZinT 

cds-NP_415211.2 92.17033505302
4 1.14550278946264 0.000126515456

500405 
0.000515384017

53323 ybfE ribbon-helix-helix domain-
containing protein YbfE 

cds-NP_416060.1 31.44819757929
8 1.1464851165341 0.000489190468

994841 
0.001732515662

65478 ydfI putative oxidoreductase YdfI 

cds-NP_414815.1 211.8271058913
96 1.16121092530275 7.059558605510

03e-13 
1.169359571348

49e-11 intF putative phage integrase 

cds-YP_026201.1 132.5820233937
82 1.16356271462219 3.565987958118

34e-08 
3.006021928259

36e-07 alx putative membrane-bound redox 
modulator Alx 



Supplementary Material 

  206 

cds-NP_417308.1 168.9602872980
83 1.1692730471782 1.166229171782

13e-07 
8.977644817859

93e-07 mutH DNA mismatch repair protein 
MutH 

cds-NP_417912.1 24.97980332682
07 1.1695731894421 0.001562976115

18254 
0.004881576905

59947 livG 

branched chain amino 
acid/phenylalanine ABC 

transporter ATP binding subunit 
LivG 

cds-NP_416649.1 91.39323008657
08 1.17053311417919 2.464738924101

67e-06 
1.477801915478

98e-05 sanA DUF218 domain-containing 
protein SanA 

cds-NP_415590.1 41.74239892692
45 1.17182579714242 3.016271272082

43e-05 
0.000140330784

756884 flgA flagellar basal body P-ring 
formation protein FlgA 

cds-NP_417915.1 37.87212534819
67 1.17980747712854 0.000252795178

098571 
0.000961706052

873654 livK 
L-leucine/L-phenylalanine ABC 
transporter periplasmic binding 

protein 

cds-YP_026277.1 182.6056226426
96 1.18119916362534 4.366254459067

49e-09 
4.167521352522

49e-08 cpxP periplasmic protein CpxP 

cds-NP_417393.1 22.05678284134
77 1.18580875097023 0.006539489426

01226 
0.016780353518

1038 argK methylmalonyl-CoA mutase-
interacting GTPase YgfD 

cds-NP_417708.1 52.63640027498
84 1.18632008579279 5.763862597371

54e-06 
3.174225495085

46e-05 aaeA aromatic carboxylic acid efflux 
pump membrane fusion protein 

cds-NP_418704.1 169.9628899818
2 1.19591480699752 1.031513061442

16e-14 
1.986946200253

06e-13 insI3 IS30 family transposase 

cds-NP_418487.1 40.47478463926
69 1.19713636012549 0.000656232918

134431 
0.002271206123

98233 soxR DNA-binding transcriptional dual 
regulator SoxR 

cds-YP_009518742.1 68.56582921918
12 1.19890864604691 4.188663995803

45e-07 
2.894666011385

6e-06 yagB orphan antitoxin YagB 

cds-NP_417980.1 80.95128890040
99 1.19942936253952 1.697699176415

05e-07 
1.274621762610

03e-06 yhjE putative transporter YhjE 

cds-NP_418271.1 165.7468525616
77 1.1999606882684 1.603103893508

03e-10 
1.864593790891

72e-09 bioP biotin transporter 

cds-NP_418041.1 14.37270433970
25 1.20838364785714 0.007470941168

60496 
0.018930831282

5901 yiaT outer membrane protein YiaT 

cds-NP_416403.1 12.29433717366
08 1.20890177433456 0.016130272362

9952 
0.036877856847

084 motB motility protein B 

cds-NP_418145.2 19.60761822366
58 1.21183243021276 0.010453534299

4325 
0.025385450948

4792 cbrA colicin M resistance protein 

cds-NP_416710.1 353.8969930754
41 1.21588761318332 4.017516376689

78e-12 
6.086322852515

44e-11 napA periplasmic nitrate reductase 
subunit NapA 

cds-NP_415011.1 2338.426994710
37 1.24464586993962 1.746089010251

08e-29 
1.179857288355

37e-27 ybaL putative transporter YbaL 

cds-YP_026223.1 207.6248164594
69 1.24499013353582 6.703327581880

98e-11 
8.198836749824

9e-10 livJ 

branched chain amino 
acid/phenylalanine ABC 

transporter periplasmic binding 
protein 

cds-NP_415968.1 269.9003341545
21 1.24635329491147 1.121620393895

21e-17 
3.041234405612

67e-16 yncD putative TonB-dependent outer 
membrane receptor 

cds-NP_415922.1 157.0370856146
52 1.24735177971814 7.507509459001

68e-15 
1.507522064479

72e-13 insI2 IS30 family transposase 

cds-NP_416707.4 18.93053048349
64 1.25283697852205 0.001824329293

63753 
0.005595223201

01944 napB periplasmic nitrate reductase 
cytochrome c550 protein 

cds-NP_416914.1 20.40291719345
07 1.25458865583603 0.002190967258

64482 
0.006524777538

23236 yfeK DUF5329 domain-containing 
protein YfeK 

cds-NP_415031.1 16.04684493373 1.25719987239908 0.012494632874
8394 

0.029632118188
4075 ybbC PF15631 family protein YbbC 

cds-NP_414684.1 73.93551205599
71 1.25764531489296 1.042841013124

08e-06 
6.746769290074

8e-06 folK 
2-amino-4-hydroxy-6-

hydroxymethyldihydropteridine 
diphosphokinase 

cds-NP_418625.1 23.28409438601
96 1.25842270308108 0.003888040417

98338 
0.010803778106

6288 yjfZ DUF2686 domain-containing 
protein YjfZ 

cds-NP_415354.1 118.9926249681
93 1.25910389235274 1.652260483340

64e-10 
1.911324151516

61e-09 pdeI putative c-di-GMP 
phosphodiesterase PdeI 

cds-NP_418305.1 2114.394713807
07 1.27693856674825 7.459448819118

79e-17 
1.783981664212

85e-15 glnL protein histidine kinase NtrB 



Supplementary Material 

 207 

cds-NP_415097.1 4805.989202823
66 1.27941204558076 3.431788608132

96e-30 
2.518814500831

38e-28 ompT omptin family outer membrane 
protease OmpT 

cds-NP_415507.2 13.56824978534
35 1.28929999344514 0.013223633239

0826 
0.031015430688

03 gfcA threonine-rich inner membrane 
protein GfcA 

cds-NP_416877.1 30.56633160617
53 1.30115391092176 0.000291107023

684989 
0.001080420749

63121 ypdI colanic acid synthesis putative 
lipoprotein YpdI 

cds-YP_026237.1 42.44481568606
18 1.30680811193368 2.148917972086

54e-06 
1.301272234306

18e-05 dgoD D-galactonate dehydratase 

cds-NP_417449.1 16.76433721224
82 1.34226998946754 0.005343300840

47055 
0.014154593452

323 glcA glycolate/lactate:H(+) symporter 
GlcA 

cds-NP_416221.1 274.0135590348 1.34271292174838 6.986255727930
02e-21 

2.437745133917
88e-19 selO protein adenylyltransferase SelO 

cds-NP_415220.1 15.63292225422
91 1.34473033527103 0.021562680298

2823 
0.047242578502

2069 speF inducible ornithine decarboxylase 

cds-NP_418633.4 28.63031426004
53 1.34539867888062 0.000943359249

577043 
0.003134957318

85205 ytfH putative transcriptional regulator 
YtfH 

cds-NP_416451.1 38.56110423085
87 1.35648166048973 0.000110700445

781123 
0.000456197287

212235 fliI flagellar export ATPase FliI 

cds-NP_417419.1 99.82983432080
4 1.35777988797199 1.069605014515

23e-05 
5.559595295227

53e-05 yggI protein YggI 

cds-YP_009518788.1 12.88853637358
79 1.36382236063089 0.008823462517

42145 
0.021914515715

6728 nohA putative prophage DNA-packaging 
protein NohA 

cds-NP_415594.1 179.0493645248
11 1.36637432491875 2.855385396976

88e-08 
2.440838480909

76e-07 flgE flagellar hook protein FlgE 

cds-NP_415254.1 9124.874914103
1 1.38225085828522 1.259286093127

01e-08 
1.138170042131

99e-07 sucA subunit of E1(0) component of 2-
oxoglutarate dehydrogenase 

cds-NP_416449.1 34.24669179818
44 1.40993495419924 0.000258208789

618216 
0.000980548454

419933 fliG flagellar motor switch protein FliG 

cds-NP_416778.2 51.27857766603
5 1.42079064384895 1.792627987186

48e-05 
8.863202487169

37e-05 yfbP uncharacterized protein YfbP 

cds-NP_418716.1 36.26806575717
54 1.43074215395088 5.036864355657

92e-05 
0.000223353453

771206 yjhF putative transporter YjhF 

cds-YP_009518818.1 21.98190121158
58 1.43191772599808 0.000595368202

132474 
0.002074044547

03596 yqiD protein YqiD 

cds-NP_418454.1 20.56208015689
65 1.43260221397805 0.000177062970

156748 
0.000694706971

389193 psiE putative phosphate starvation-
inducible protein PsiE 

cds-NP_416255.1 155.8784888530
94 1.43941143684895 6.151669862914

03e-16 
1.349879309609

54e-14 cho excinuclease Cho 

cds-NP_416608.1 24.67395496634
39 1.43992532534092 0.000220624368

612995 
0.000851494050

032203 rcnR DNA-binding transcriptional 
repressor RcnR 

cds-
gnl|b4580|CDS%3D3

93 
56.26462673832

47 1.44042841147638 3.346484597138
75e-06 

1.943517725787
13e-05 yaiT putative autotransporter YaiT 

cds-
gnl|b4580|CDS%3D3

93 
56.26462673832

47 1.44042841147638 3.346484597138
75e-06 

1.943517725787
13e-05 yaiT putative autotransporter YaiT 

cds-NP_418607.1 16.62222705907
86 1.44431798048833 0.001869395980

45448 
0.005716967448

84678 yjfC putative acid--amine ligase YjfC 

cds-NP_416455.1 48.37375885190
18 1.44486884249993 3.834607074864

99e-06 
2.179428880867

86e-05 fliM flagellar motor switch protein FliM 

cds-NP_418554.1 63.17356605423
84 1.44958024403997 3.457684500252

26e-07 
2.436668673933

72e-06 dtpC dipeptide/tripeptide:H(+) 
symporter DtpC 

cds-NP_416999.1 61.79703500479
02 1.45074320864606 1.510753972686

5e-05 
7.583748456815

14e-05 yfgG nickel/cobalt stress response 
protein YfgG 

cds-NP_415891.1 11.28631084495
63 1.45193247261466 0.008277711150

81597 
0.020705350288

7928 tfaR putative tail fiber assembly protein 
TfaR 

cds-NP_418178.1 101.9515860857
31 1.45664242021452 3.736116125470

81e-05 
0.000169740089

072884 bglF 
beta-glucoside specific PTS 
enzyme II/BglG kinase/BglG 

phosphatase 

cds-NP_418014.1 53.24318069765
57 1.46453409670841 4.628677454098

1e-07 
3.172991935925

22e-06 insJ insertion element IS150 protein 
InsA 



Supplementary Material 

  208 

cds-NP_416708.1 106.2857069182
24 1.48087512487458 4.167234069523

74e-10 
4.572143153083

13e-09 napH ferredoxin-type protein NapH 

cds-NP_416450.2 26.33745028425
78 1.49175748531025 3.334343710739

83e-05 
0.000153286189

812305 fliH flagellar biosynthesis protein FliH 

cds-NP_416456.1 22.89676509589
27 1.50398561318915 0.000386957725

314336 
0.001399557380

34251 fliN flagellar motor switch protein FliN 

cds-NP_415289.4 79.50780215137
68 1.50585103870361 4.713032032213

6e-10 
5.131298557834

6e-09 ybhD putative DNA-binding 
transcriptional regulator YbhD 

cds-NP_415296.1 274.9697529076
71 1.50717127013785 7.244086878467

79e-12 
1.048914212300

59e-10 bioB biotin synthase 

cds-NP_415297.1 164.5303195813
87 1.52746527278121 5.285416905124

06e-09 
4.988917908007

34e-08 bioF 8-amino-7-oxononanoate 
synthase 

cds-NP_415089.1 203.6718321920
13 1.55532547074602 1.969314052532

35e-12 
3.107628781888

37e-11 borD prophage lipoprotein BorD 

cds-NP_417978.2 99.12833624031
33 1.55853625676138 4.516210084624

45e-12 
6.817555436257

55e-11 rcdB putative DNA-binding 
transcriptional regulator YhjC 

cds-NP_418015.1 35.22050422382
51 1.56428776520572 1.948155933829

42e-07 
1.437313658632

9e-06 insK IS150 family conserved protein 
InsB 

cds-NP_415598.3 43.33008947289
16 1.59026884981434 3.055485303268

56e-05 
0.000141690642

004512 flgI flagellar P-ring protein 

cds-NP_415298.1 94.24245525215
32 1.59630844836499 3.608697218435

6e-11 
4.585738525038

91e-10 bioC malonyl-acyl carrier protein 
methyltransferase 

cds-NP_414993.1 399.8376879697
56 1.60006335808564 8.841339355068

74e-23 
3.550715248540

34e-21 hha hemolysin expression-modulating 
protein Hha 

cds-YP_002791259.1 21.47692468782
8 1.61329825742462 0.000102954447

402439 
0.000428004963

468927 ilvX uncharacterized protein IlvX 

cds-YP_588446.1 57.37993177778
82 1.61526218341104 3.194323558577

69e-10 
3.578482997069

8e-09 gnsA putative phosphatidylethanolamine 
synthesis regulator GnsA 

cds-NP_416116.1 136.3495821180
21 1.61800338489438 1.400762789058

3e-12 
2.250206487932

51e-11 mdtI multidrug/spermidine efflux pump 
membrane subunit MdtI 

cds-NP_414574.1 1835.095989063
44 1.61812465905999 3.277157876393

67e-23 
1.341428949981

52e-21 carB carbamoyl-phosphate synthetase 
large subunit 

cds-NP_415591.1 59.02754272356
72 1.63020410696099 1.311387157634

72e-05 
6.677721447429

41e-05 flgB flagellar basal-body rod protein 
FlgB 

cds-NP_416709.1 66.03098742297
19 1.63135116943174 6.721620920105

45e-11 
8.198836749824

9e-10 napG ferredoxin-type protein NapG 

cds-NP_415593.1 84.51190011749
51 1.64871186706882 3.469743895085

68e-07 
2.437409201547

81e-06 flgD flagellar biosynthesis  initiation of 
hook assembly 

cds-YP_009518761.1 34.13499990501
9 1.65795393264401 0.002756197084

67225 
0.007981721761

53044 ymcF protein YmcF 

cds-NP_416706.1 91.81856110797
73 1.65865728425874 1.172175178449

21e-15 
2.494974867329

15e-14 napC periplasmic nitrate reductase 
cytochrome c protein 

cds-NP_416076.1 27.67830259296
54 1.66709985498833 0.004154827100

06022 
0.011440555604

7583 cspF cold shock-like protein CspF 

cds-NP_416117.1 252.0627855961
59 1.67035546647242 1.213977586223

06e-10 
1.427597399047

39e-09 mdtJ multidrug/spermidine efflux pump 
membrane subunit MdtJ 

cds-NP_415633.1 13.27676564564
84 1.67587157197146 0.002191785736

35712 
0.006524777538

23236 ycfT inner membrane protein YcfT 

cds-YP_009518779.1 43.29010439386
31 1.67621139987125 0.018177497740

8053 
0.040748608679

6252 ynaM protein YnaM 

cds-NP_414768.1 47.40810799447
06 1.70211945315018 3.141548453761

95e-06 
1.834509158801

73e-05 yafO ribosome-dependent mRNA 
interferase toxin YafO 

cds-NP_415595.1 93.28277160496
82 1.70452965905239 3.391699266278

55e-07 
2.400831942172

34e-06 flgF flagellar basal-body rod protein 
FlgF 

cds-NP_416256.2 10.83360515263
15 1.7065783347887 0.005963877522

72942 
0.015464170390

7069 ves HutD family protein Ves 

cds-NP_415592.1 49.20470820356
51 1.70775714884493 0.008259219612

20249 
0.020682057581

8506 flgC flagellar basal-body rod protein 
FlgC 

cds-NP_417940.2 9.824233291134
86 1.71162273574325 0.003960324368

52242 
0.010975977107

2916 yhhH PF15631 family protein YhhH 



Supplementary Material 

 209 

cds-NP_414769.1 12.29142583880
67 1.75346263465221 0.002139971360

68275 
0.006397372248

89498 yafP putative N-acetyltransferase YafP 

cds-NP_416220.1 11.04744892277
67 1.75535202330475 0.003846359096

53909 
0.010715936304

9522 ydiE PF10636 family protein YdiE 

cds-NP_415896.1 2122.421755296
37 1.78962312274524 1.788940187947

04e-68 
8.461687088989

49e-66 ydbK putative pyruvate-flavodoxin 
oxidoreductase 

cds-NP_415599.1 33.16704176366
21 1.79654099236812 8.782947190275

12e-08 
6.911091717005

77e-07 flgJ putative peptidoglycan hydrolase 
FlgJ 

cds-YP_010051187.1 13.37409804699
16 1.85129729547891 0.000986812955

485667 
0.003261539403

34044 mdtU protein MdtU 

cds-NP_417472.1 6.902468537923
43 1.85410041627492 0.021362047229

8534 
0.046851228777

6846 yghW DUF2623 domain-containing 
protein YghW 

cds-NP_414573.1 673.4300652205
16 1.86787465097657 2.598115696166

89e-26 
1.436386820595

12e-24 carA carbamoyl-phosphate synthetase 
small subunit 

cds-NP_418200.1 4993.314500759
5 1.88432578676071 1.638550820114

73e-19 
5.166896919428

45e-18 asnA asparagine synthetase A 

cds-NP_418344.3 1774.789669426
83 1.88577183924192 7.741913163024

34e-15 
1.540061884812

83e-13 sodA superoxide dismutase (Mn) 

cds-NP_415597.1 33.82723420476
81 1.92896190491832 3.618295300106

66e-09 
3.476993926084

43e-08 flgH flagellar L-ring protein 

cds-NP_417705.2 554.7463352985
18 1.95430921277389 2.144021904024

31e-20 
7.186693894040

56e-19 yhcN DUF1471 domain-containing 
stress-induced protein YhcN 

cds-NP_416075.1 671.9271446626
15 1.96483102128886 1.505239289804

86e-13 
2.647852750702

19e-12 cspB cold shock-like protein CspB 

cds-NP_416458.1 11.76960298981
89 1.98991525072013 0.002094247722

03831 
0.006295642562

12672 fliP flagellar biosynthesis protein FliP 

cds-NP_414844.1 5.063122187826
22 2.00430696709959 0.021720655857

4323 
0.047477140393

8032 ykgH uncharacterized protein YkgH 

cds-YP_588452.1 8.283356601710
5 2.01155612503374 0.002954428463

67466 
0.008466080779

30556 rzoR putative prophage outer 
membrane lipoprotein RzoR 

cds-NP_415299.1 54.77169165425
99 2.04709687315041 6.370325783429

71e-12 
9.383556006941

27e-11 bioD dethiobiotin synthetase 

cds-NP_415255.1 10316.86885494
05 2.0629292970814 9.864315963244

31e-12 
1.385887559588

48e-10 sucB dihydrolipoyltranssuccinylase 

cds-NP_417610.1 9.234983398909
41 2.07313512326119 0.004596759275

92815 
0.012456017974

3005 agaI putative deaminase AgaI 

cds-NP_418441.1 26.81399071589
53 2.13035625973753 1.433655694481

3e-07 
1.084027049983

46e-06 arpA regulator of acetyl CoA synthetase 

cds-NP_416242.6 65.88491953164
94 2.13215288080372 3.132948804931

56e-17 
7.754048292205

62e-16 ydjM inner membrane protein YdjM 

cds-NP_415510.1 351.5100485675
8 2.15350759021128 1.407361060582

88e-05 
7.106922935825

98e-05 cspG cold shock protein CspG 

cds-NP_416404.1 7.398522387900
37 2.18566052747526 0.007462566115

66596 
0.018920871920

4229 motA motility protein A 

cds-NP_415511.1 17.92039650075
79 2.27125915491935 0.005283254407

17755 
0.014012968231

3737 ymcE protein YmcE 

cds-NP_416019.1 4.020939845723
76 2.28520236019401 0.018970532355

2038 
0.042370176409

2878 ydeQ putative fimbrial adhesin protein 
YdeQ 

cds-YP_009518791.1 40.57072597008
76 2.30521048169676 1.033784087850

61e-05 
5.399777744760

81e-05 ynfS protein YnfS 

cds-NP_415256.1 8848.675099583
82 2.32657703750501 1.501201394165

48e-11 
2.028766455543

63e-10 sucC succinyl-CoA synthetase subunit 
beta 

cds-NP_416867.1 873.7004048616
01 2.34208213389576 0.000161414997

05227 
0.000643995916

07452 dsdA D-serine ammonia-lyase 

cds-NP_415257.1 5422.091361455
16 2.40146736754531 3.106955795008

71e-11 
4.020155264240

76e-10 sucD succinyl-CoA synthetase subunit 
alpha 

cds-NP_417446.4 50.23042037861
56 2.46657834430919 6.706979695253

08e-12 
9.777949507771

36e-11 pppA prepilin peptidase 

cds-NP_418012.1 34841.40606166
52 2.47144708268749 4.154468803723

14e-08 
3.474572435648

21e-07 cspA cold shock protein CspA 



Supplementary Material 

  210 

cds-YP_010051199.1 24.46398060727
47 2.50871516241267 1.685140147285

69e-08 
1.491401581495

88e-07 yghF putative type II secretion system 
C-type protein YghF 

cds-NP_416647.1 543.5021968253
51 2.62820342834769 2.050081515719

93e-29 
1.363624533190

58e-27 yohK putative 3-hydroxypropanoate 
export protein YohK 

cds-YP_001165307.1 4.442179584178
87 2.64743608221851 0.006264126433

87819 
0.016132115081

0765 ykfM uncharacterized protein YkfM 

cds-NP_416646.1 156.8730078237
44 2.67466674263507 3.834826239120

76e-33 
3.710194386349

33e-31 yohJ putative 3-hydroxypropanoate 
export protein YohJ 

cds-YP_026189.1 1233.752131885
17 2.69685590674382 4.555581223591

42e-65 
1.616092439069

06e-62 yghJ putative lipoprotein YghJ 

cds-NP_415146.1 4.289641469002
07 2.72974697941154 0.007663530120

99554 
0.019361215267

1086 citG triphosphoribosyl-dephospho-CoA 
synthase 

cds-YP_588474.1 6.114890429299
78 2.7956222778709 0.001904390125

34949 
0.005807298541

2699 ghoT toxin of the GhoTS toxin-antitoxin 
system 

cds-YP_588447.1 4.006597305410
29 3.0236913102916 0.004577647731

0931 
0.012412131459

4034 ymgF inner membrane protein YmgF 

cds-NP_416459.1 3.426623192803
25 3.34505177339201 0.010741565808

5561 
0.025966408658

1621 fliQ flagellar biosynthesis protein FliQ 

cds-NP_417941.1 19.35419292973
96 3.38680006593795 1.046273118957

03e-07 
8.127709247080

41e-07 yhhI putative transposase 

cds-YP_002791243.1 3.159665811240
46 3.42053487591051 0.021384682257

859 
0.046876721097

6857 yoaJ uncharacterized protein YoaJ 

cds-NP_416837.1 4.895565727119
67 3.49857801826164 0.002678793185

42033 
0.007789359692

85134 yfcQ putative fimbrial protein YfcQ 

cds-YP_588478.1 4.357965276438
71 3.50455009726099 0.003803231376

85874 
0.010616626866

4181 yjjZ DUF1435 domain-containing 
protein YjjZ 

cds-NP_415509.1 20.58657693522 3.82221582034075 7.665274409530
73e-06 

4.130515590047
13e-05 cspH CspA family protein CspH 

cds-NP_417445.1 6.236953631470
38 4.15860513185643 0.000609502131

785353 
0.002117637411

52683 yghG lipoprotein YghG 

ereA2 

GeneID baseMean log2FoldChange p value padj Gene Product 

cds-NP_418191.1 7617.930136593
8 -1.00025290863863 1.902687158907

83e-29 
1.354650873956

83e-27 atpH ATP synthase F1 complex subunit 
delta 

cds-NP_416292.1 466.1232838420
85 -1.00191895491895 2.365365334652

79e-16 
6.714236371501

35e-15 msrB methionine sulfoxide reductase B 

cds-NP_415497.1 1245.690279476
66 -1.00498488996541 4.534002095896

2e-05 
0.000310097182

716176 appC cytochrome bd-II subunit 1 

cds-NP_416437.1 322.5467629267
07 -1.00602966113434 1.513531333442

2e-11 
2.738861075474

78e-10 amyA alpha-amylase 

cds-NP_416805.1 21.81344926117
78 -1.01091937240121 0.006316585806

55754 
0.023287718300

4069 yfcG disulfide bond oxidoreductase 
YfcG 

cds-NP_417721.1 4139.972542152
97 -1.01438336803834 1.539084952226

76e-15 
4.126077745383

23e-14 accB biotin carboxyl carrier protein 

cds-NP_417697.1 14431.84951614
62 -1.01606513020503 6.072701300388

63e-23 
2.866711059520

41e-21 rpsI 30S ribosomal subunit protein S9 

cds-NP_417756.1 33668.27539544
97 -1.01646595614325 2.765779088719

56e-27 
1.715968368901

29e-25 rpsK 30S ribosomal subunit protein S11 

cds-NP_415940.1 330.9489127904
68 -1.03471148914551 5.338124408750

67e-06 
4.441278602912

68e-05 ydcJ DUF1338 domain-containing 
protein YdcJ 

cds-NP_417758.1 4547.449245405
49 -1.03629373472411 6.022318209016

14e-12 
1.142136593089

83e-10 rpmJ 50S ribosomal subunit protein L36 

cds-NP_414692.1 790.4133834970
56 -1.03745087570298 3.877829776952

28e-05 
0.000269894466

68756 fhuA ferrichrome outer membrane 
transporter/phage receptor 

cds-NP_416285.1 55.75130898929
95 -1.04121148819328 0.000107076470

194358 
0.000679631519

068065 ydjG NADH-dependent methylglyoxal 
reductase 

cds-NP_416769.1 859.2862178381
8 -1.04688446920677 7.803057216179

77e-07 
7.632585020240

7e-06 elaB tail-anchored inner membrane 
protein ElaB 



Supplementary Material 

 211 

cds-NP_415305.1 157.8553979102
35 -1.05401517554306 3.228493423274

74e-07 
3.346383517251

12e-06 moaD molybdopterin synthase sulfur 
carrier subunit 

cds-NP_418414.1 56771.51988704
82 -1.05508373021547 5.198868811820

18e-28 
3.473644192266

93e-26 rpoB RNA polymerase subunit beta 

cds-NP_415538.1 179.4342590300
61 -1.05583482540722 8.065520737616

08e-07 
7.871585744599

24e-06 efeB heme-containing 
peroxidase/deferrochelatase 

cds-NP_415706.1 301.9259522365
96 -1.05917720399395 3.672302485922

15e-12 
7.216655971203

58e-11 ycgB PF04293 family protein YcgB 

cds-NP_417757.1 36668.11301657
59 -1.06475095552322 5.507698829749

75e-44 
6.834267433600

9e-42 rpsM 30S ribosomal subunit protein S13 

cds-NP_417185.1 10286.95795060
23 -1.06574783174568 3.552549015232

23e-06 
3.073450273536

57e-05 srlD sorbitol-6-phosphate 2-
dehydrogenase 

cds-NP_415156.1 3991.736497560
06 -1.06643607527747 1.865399728841

02e-06 
1.687798129657

62e-05 cspE transcription antiterminator and 
regulator of RNA stability CspE 

cds-NP_417351.1 2037.676974814
92 -1.06694859937831 0.002614001614

75338 
0.011011259954

2909 yqeB XdhC-CoxI family protein YqeB 

cds-NP_417400.1 56193.92104051
45 -1.06721432243542 2.528148677798

77e-21 
1.016643491451

86e-19 fbaA fructose-bisphosphate aldolase 
class II 

cds-NP_417764.1 45089.01910752
49 -1.06845905552486 1.322942333266

77e-24 
6.680858782997

21e-23 rplF 50S ribosomal subunit protein L6 

cds-NP_416538.1 1910.448887374
8 -1.07070978743745 4.496089210568

74e-19 
1.525509018867

19e-17 wbbI beta-1 6-
galactofuranosyltransferase WbbI 

cds-NP_416607.1 1653.482402597
6 -1.07557169658814 1.649774822991

34e-24 
8.142013700285

65e-23 thiM hydroxyethylthiazole kinase 

cds-NP_418688.1 96.20631949386
35 -1.07624832824323 1.976445442446

91e-05 
0.000144994975

617347 idnD L-idonate 5-dehydrogenase 

cds-NP_415303.1 682.8768824721
26 -1.07723058447583 5.568403508886

55e-15 
1.404301388262

15e-13 moaB protein MoaB 

cds-NP_415304.1 257.2011126562
24 -1.08049560669298 9.183726965134

37e-09 
1.191895756915

72e-07 moaC cyclic pyranopterin 
monophosphate synthase 

cds-NP_416606.1 2948.075299451
21 -1.08201010162524 1.337654155804

53e-26 
7.745909331545

44e-25 thiD 

bifunctional 
hydroxymethylpyrimidine 

kinase/phosphomethylpyrimidine 
kinase 

cds-NP_416289.1 33.57857141025
41 -1.08213519686362 0.000759652828

746019 
0.003836246785

16739 ydjK putative transporter YdjK 

cds-NP_414711.1 28571.08109291
15 -1.08433081955498 1.090021516602

27e-21 
4.596081016120

04e-20 rpsB 30S ribosomal subunit protein S2 

cds-NP_418517.1 44.34486684623
09 -1.08759310870917 0.000501844862

318213 
0.002651474740

9343 phnO aminoalkylphosphonate N-
acetyltransferase 

cds-NP_416199.1 32.40192831087
39 -1.09170471050013 0.008444180564

94861 
0.029670773619

3947 sufA iron-sulfur cluster insertion protein 
SufA 

cds-NP_416536.1 2733.514465891
87 -1.09392266655627 3.412494362570

79e-18 
1.089739927694

48e-16 wbbK putative glycosyltransferase WbbK 

cds-NP_417347.1 35.80060429703
12 -1.1010797843317 0.001513525806

43774 
0.006926493759

07178 ygeX 2 3-diaminopropionate ammonia-
lyase 

cds-NP_417754.1 69833.78449896
16 -1.10182277468351 3.534157202968

99e-24 
1.686686234340

04e-22 rpoA RNA polymerase subunit alpha 

cds-NP_418023.1 42.48516130789
52 -1.10901843797222 0.001184090684

40366 
0.005595762614

10781 xylF xylose ABC transporter 
periplasmic binding protein 

cds-NP_416868.1 17.09430419649
67 -1.10982035915796 0.009652019053

99486 
0.033058926460

1732 emrY tripartite efflux pump membrane 
subunit EmrY 

cds-NP_417755.1 40328.34631915
25 -1.11062061859468 3.162625915954

36e-19 
1.090101932776

97e-17 rpsD 30S ribosomal subunit protein S4 

cds-NP_415580.1 666.8448517005
78 -1.11064353766225 3.737366284003

54e-15 
9.604367912087

22e-14 pyrC dihydroorotase 

cds-NP_416539.1 2362.317064539
18 -1.11074032960491 1.137597788922

89e-17 
3.528990855208

65e-16 wbbH putative O-antigen polymerase 

cds-NP_418456.1 1959.601346768
86 -1.1174581263286 6.284565934186

75e-09 
8.476357096948

15e-08 malG maltose ABC transporter 
membrane subunit MalG 



Supplementary Material 

  212 

cds-NP_416030.1 187.4450760427
89 -1.12874122772444 2.333938160793

05e-05 
0.000168657128

657641 lsrA Autoinducer-2 ABC transporter 
ATP binding subunit 

cds-NP_418457.1 10679.51232957
53 -1.13127976702002 5.995895056627

11e-08 
6.962612895971

e-07 malF maltose ABC transporter 
membrane subunit MalF 

cds-NP_415375.1 97.48999031012
12 -1.13199546546719 1.556969356137

19e-07 
1.677895263946

36e-06 potF putrescine ABC transporter 
periplasmic binding protein 

cds-NP_418190.1 18779.26860812
2 -1.13430169419139 1.416431375270

78e-34 
1.308842864425

74e-32 atpA ATP synthase F1 complex subunit 
alpha 

cds-NP_418493.1 1356.873904942
62 -1.13539160658176 0.000884622323

575857 
0.004350979333

2842 acs acetyl-CoA synthetase (AMP-
forming) 

cds-NP_418410.1 12871.66249734
32 -1.14130221285177 2.451626411305

63e-15 
6.375696709161

9e-14 rplK 50S ribosomal subunit protein L11 

cds-NP_416653.1 642.8599145638
75 -1.14554202331963 0.011620895796

9495 
0.038379886270

8378 mglC 
D-galactose/methyl-galactoside 

ABC transporter membrane 
subunit 

cds-NP_414712.1 15542.32818836
86 -1.14564006966934 1.850915641984

51e-22 
8.461606982251

28e-21 tsf protein chain elongation factor EF-
Ts 

cds-NP_415957.1 150.2590142156
61 -1.15053347307065 7.990909236577

15e-09 
1.051652085286

5e-07 ydcS 

putative ABC transporter 
periplasmic binding 

protein/polyhydroxybutyrate 
synthase 

cds-NP_415689.2 177.1038013731
69 -1.15706482267046 3.205951288019

91e-09 
4.462643090984

13e-08 ymgD PF16456 family protein YmgD 

cds-NP_415641.1 2914.868683688 -1.15898612798284 7.818262310356
52e-48 

1.305950508226
09e-45 potD 

spermidine preferential ABC 
transporter periplasmic binding 

protein 

cds-NP_418411.1 46345.16937585
39 -1.16520294564925 7.043437686780

93e-28 
4.634795435407

51e-26 rplA 50S ribosomal subunit protein L1 

cds-NP_418623.1 3378.195545852
05 -1.16689254237262 6.050797382701

13e-16 
1.652742958054

78e-14 rpsR 30S ribosomal subunit protein S18 

cds-NP_418163.1 115.2645123208
45 -1.17212042241206 2.440341893358

27e-09 
3.441040533394

47e-08 tnaC tnaAB operon leader peptide 

cds-NP_417763.1 14704.90594751
4 -1.17490087708325 1.271573859014

61e-14 
3.102497342528

35e-13 rplR 50S ribosomal subunit protein L18 

cds-NP_418491.1 111.8723506690
21 -1.17901270261081 4.431629602394

29e-06 
3.751767517192

67e-05 actP acetate/glycolate:cation symporter 

cds-NP_416031.1 62.90744090621
13 -1.18107794217729 1.855060860850

29e-06 
1.687798129657

62e-05 lsrC Autoinducer-2 ABC transporter 
membrane subunit LsrC 

cds-NP_418621.5 14379.62872614
21 -1.18302056562258 7.845125503103

15e-13 
1.638047118268

12e-11 rpsF 30S ribosomal subunit protein S6 

cds-NP_418045.4 150.6307555668
12 -1.18667984542322 9.325455401046

4e-11 
1.551741486848

45e-09 aldB aldehyde dehydrogenase B 

cds-NP_417207.1 174.7219722213
33 -1.18686536101159 6.213683026164

04e-11 
1.041931481954

84e-09 hypB hydrogenase isoenzymes nickel 
incorporation protein HypB 

cds-NP_416540.1 2174.125525192
28 -1.19418798027666 1.266799513508

29e-14 
3.102497342528

35e-13 glf UDP-galactopyranose mutase 

cds-NP_418028.1 2005.264886299
17 -1.19605950935434 3.161519048017

68e-10 
5.029478837194

43e-09 malS alpha-amylase 

cds-NP_417753.1 10849.47776493
8 -1.19617390351974 2.089747823560

93e-15 
5.467334215497

05e-14 rplQ 50S ribosomal subunit protein L17 

cds-NP_418648.1 326.7869946677
71 -1.19701447375147 3.647519026917

77e-10 
5.781450778797

03e-09 ytfQ galactofuranose ABC transporter 
periplasmic binding protein 

cds-NP_418461.1 3816.451058259
29 -1.1988428714694 9.193761882725

42e-09 
1.191895756915

72e-07 malM maltose regulon periplasmic 
protein 

cds-NP_415524.1 255.0805192065
32 -1.2029290934819 5.577241548755

84e-13 
1.181559026646

18e-11 wrbA NAD(P)H:quinone oxidoreductase 

cds-NP_416753.4 126.3904626774
61 -1.20880774564736 9.029227233105

14e-11 
1.508228225899

06e-09 yfaZ putative porin YfaZ 

cds-NP_418458.1 24975.49549012
95 -1.21932718032965 1.198799306634

17e-12 
2.467481226877

82e-11 malE maltose ABC transporter 
periplasmic binding protein 



Supplementary Material 

 213 

cds-NP_415160.1 298.7478976414
57 -1.22141400609797 4.727115278136

16e-17 
1.415852527789

33e-15 tatE twin arginine protein translocation 
system - TatE protein 

cds-NP_418624.1 14617.62169489
83 -1.22229196196808 1.466498371515

42e-20 
5.586844234641

65e-19 rplI 50S ribosomal subunit protein L9 

cds-NP_415500.1 474.6751741493
03 -1.223396637489 7.356295980394

17e-10 
1.120996261152

7e-08 appA 
periplasmic phosphoanhydride 
phosphatase/multiple inositol-
polyphosphate phosphatase 

cds-NP_418622.1 14785.38934222
78 -1.233737970766 3.483179308188

71e-18 
1.104193265362

3e-16 priB primosomal replication protein N 

cds-NP_418189.1 13230.68494082
69 -1.24413145005397 3.013487268865

54e-54 
6.232178670801

45e-52 atpG ATP synthase F1 complex subunit 
gamma 

cds-NP_414713.1 4086.571032119
73 -1.24426460818396 6.571135155979

91e-32 
5.096149996860

85e-30 pyrH UMP kinase 

cds-NP_417761.1 7052.870443944
59 -1.24842093993999 9.980407156853

04e-30 
7.224151380368

79e-28 rpmD 50S ribosomal subunit protein L30 

cds-NP_418651.3 97.51596354806
86 -1.24979498346818 8.247495871176

76e-07 
8.013171044411

78e-06 ytfT galactofuranose ABC transporter 
putative membrane subunit YtfT 

cds-NP_417552.1 1410.582274923
43 -1.25210897119773 0.008246176932

67425 
0.029116379202

1173 fadH 2 4-dienoyl-CoA reductase 

cds-NP_418011.1 140.7192179270
07 -1.25499778706869 7.652176346817

39e-10 
1.157958253457

42e-08 yiaG putative DNA-binding 
transcriptional regulator YiaG 

cds-NP_416600.4 471.1007428743
54 -1.25719221175784 5.593924480067

96e-15 
1.404301388262

15e-13 fbaB fructose-bisphosphate aldolase 
class I 

cds-NP_417762.1 55161.10536528
58 -1.25845607972236 2.106918347769

14e-42 
2.473066590367

94e-40 rpsE 30S ribosomal subunit protein S5 

cds-NP_415498.1 1443.351029298
93 -1.25943043072433 6.040591232297

34e-10 
9.237425254178

64e-09 appB cytochrome bd-II subunit 2 

cds-NP_417722.1 7998.062315344
09 -1.25967352509558 1.651545631048

1e-18 
5.433835360334

79e-17 accC biotin carboxylase 

cds-NP_418420.4 11816.85395489
03 -1.26401919220762 4.265311801093

44e-16 
1.172420832414

48e-14 thiF sulfur carrier protein ThiS 
adenylyltransferase 

cds-NP_417904.1 254.1344713643
81 -1.26812586752012 1.203145714951

02e-21 
5.024290230800

28e-20 ggt glutathione hydrolase proenzyme 

cds-YP_026286.1 217.0143679535
65 -1.27464085009967 3.288958530811

31e-12 
6.522350182334

94e-11 ytfR galactofuranose ABC transporter 
putative ATP binding subunit 

cds-NP_418164.4 3848.911659106
44 -1.2787358548662 8.244302517996

25e-05 
0.000538421140

38583 tnaA tryptophanase 

cds-NP_418492.1 33.32285817848
2 -1.27944092013378 0.002711349240

77399 
0.011344306120

117 yjcH DUF485 domain-containing inner 
membrane protein YjcH 

cds-NP_415539.1 568.2235972416
96 -1.28120753612199 0.000374090696

6567 
0.002053951827

53483 phoH ATP-binding protein PhoH 

cds-NP_415534.1 5654.354622804
36 -1.28580235762931 1.763569635023

18e-08 
2.194608287938

58e-07 putA 

fused DNA-binding transcriptional 
repressor/proline 

dehydrogenase/1-pyrroline-5-
carboxylate dehydrogenase PutA 

cds-NP_416853.1 777.7586469679
42 -1.29207693635198 3.727389747038

99e-16 
1.037695748166

05e-14 yfdI serotype-specific glucosyl 
transferase YfdI 

cds-NP_418415.1 65063.92409335
12 -1.29646867669173 3.826132669054

81e-32 
3.077202626241

68e-30 rpoC RNA polymerase subunit beta' 

cds-NP_414917.2 142.8713446604
65 -1.30104718127119 2.246166777745

86e-11 
4.014445397428

1e-10 phoA alkaline phosphatase 

cds-YP_010051189.1 21.49831337399
85 -1.30654244897587 0.003756991269

89778 
0.014955649023

9836 yoaM protein YoaM 

cds-NP_418188.1 38351.76206693
08 -1.30795009289212 6.054509756026

12e-54 
1.195215266837

34e-51 atpD ATP synthase F1 complex subunit 
beta 

cds-NP_417759.1 101839.8797741
73 -1.31544965360239 6.108557056534

28e-29 
4.278945692988

45e-27 secY Sec translocon subunit SecY 

cds-NP_414937.2 1423.865739964
02 -1.3184955611695 1.867038766839

22e-27 
1.175152081794

6e-25 malZ maltodextrin glucosidase 

cds-NP_418497.1 32.10643244811
49 -1.31893560660205 5.783998084514

76e-05 
0.000387652834

584068 nrfD putative menaquinol-cytochrome c 
reductase subunit NrfD 



Supplementary Material 

  214 

cds-NP_416258.1 115.8565405362
35 -1.32249562300222 6.550095140719

e-06 
5.317208074045

35e-05 astE succinylglutamate desuccinylase 

cds-NP_417760.1 107998.2300433
81 -1.32299932362021 5.567349820310

91e-45 
7.111470667532

43e-43 rplO 50S ribosomal subunit protein L15 

cds-NP_418422.1 55115.07136252
65 -1.32417344155135 5.177228444756

61e-18 
1.617604542127

91e-16 thiC phosphomethylpyrimidine 
synthase 

cds-NP_415986.1 25.10866010688
66 -1.32946413501429 0.001074494969

21933 
0.005116811021

1837 narU nitrate/nitrite transporter NarU 

cds-NP_418421.1 12476.22008345
07 -1.33163343778015 1.785352464200

89e-18 
5.829914099266

52e-17 thiE thiamine phosphate synthase 

cds-NP_418417.1 16453.22227020
08 -1.33643012848219 5.358681758874

41e-22 
2.374770905999

14e-20 thiH 2-iminoacetate synthase 

cds-NP_418418.2 13520.81366144
85 -1.33807791460856 2.826992596473

27e-19 
9.822103077186

73e-18 thiG 1-deoxy-D-xylulose 5-
phosphate:thiol sulfurtransferase 

cds-NP_417003.1 15925.24726083
58 -1.34109847238139 5.938812506360

72e-34 
5.358989709106

62e-32 guaB inosine 5'-monophosphate 
dehydrogenase 

cds-NP_418187.1 9293.803867648
54 -1.34116670673509 1.978245574381

56e-32 
1.652215486449

83e-30 atpC ATP synthase F1 complex subunit 
epsilon 

cds-NP_417594.3 221.3297938878
87 -1.34542787623259 8.426319380867

01e-16 
2.273012737335

74e-14 garR tartronate semialdehyde reductase 

cds-NP_416319.1 2826.644070602
07 -1.35213568510488 0.000250511329

412408 
0.001442931967

68977 fadD long-chain-fatty-acid--CoA ligase 

cds-NP_415799.1 30.53291223852
97 -1.35811851671504 0.000130558464

985719 
0.000803137979

366825 osmB osmotically-inducible lipoprotein 
OsmB 

cds-YP_026279.1 2634.248765980
45 -1.36134175295367 1.856927287158

79e-19 
6.610356727975

94e-18 thiS sulfur carrier protein ThiS 

cds-NP_418288.1 1093.773214404
69 -1.36690107052709 0.011066111084

2033 
0.036855920581

8212 fadB 

multifunctional enoyl-CoA 
hydratase 3-hydroxyacyl-CoA 

epimerase Delta(3)-cis- Delta(2)-
trans-enoyl-CoA isomerase L-3-

hydroxyacyl-CoA dehydrogenase 

cds-NP_416874.1 46.23889790190
51 -1.3682181675165 3.279164209298

74e-05 
0.000232702780

408243 oxc oxalyl-CoA decarboxylase 

cds-NP_416450.2 26.33745028425
78 -1.37882235836589 0.002264312158

39266 
0.009775256166

89794 fliH flagellar biosynthesis protein FliH 

cds-NP_417936.1 30.10195989150
71 -1.37971056358008 0.000238464557

061547 
0.001382712378

26208 nikD Ni(2(+)) ABC transporter ATP 
binding subunit NikD 

cds-NP_417002.1 23485.00623114
03 -1.39005797370452 2.641058195357

21e-40 
2.867528935609

09e-38 guaA GMP synthetase 

cds-NP_414913.1 65.95751657641
42 -1.3980240413186 1.637862697235

91e-06 
1.503855749280

24e-05 yaiY DUF2755 domain-containing inner 
membrane protein YaiY 

cds-NP_418663.1 6541.461037171
63 -1.40116581447169 1.575741937542

18e-20 
5.950823682387

55e-19 mgtA Mg(2(+)) importing P-type ATPase 

cds-NP_416953.1 15.27938470381
1 -1.40142273361971 0.002581038138

55981 
0.010918052360

7495 eutD phosphate acetyltransferase EutD 

cds-NP_415958.1 60.50741919651
67 -1.4067945269203 1.167959649916

e-06 
1.110634471077

31e-05 ydcT putative ABC transporter ATP-
binding protein YdcT 

cds-NP_417780.1 11834.53862106
42 -1.40829184778513 4.394299450633

03e-17 
1.325308507923

56e-15 rpsJ 30S ribosomal subunit protein S10 

cds-NP_414838.2 38.77872578528
53 -1.4214659230084 8.319579208749

08e-06 
6.629712385981

15e-05 rclA cupric reductase RclA 

cds-NP_418165.1 533.4896689973
93 -1.42168983618025 0.003213516463

21865 
0.013141527306

7407 tnaB tryptophan:H(+) symporter TnaB 

cds-NP_417779.1 52347.66775977
84 -1.42202632021918 1.348413600664

1e-31 
1.027396538190

21e-29 rplC 50S ribosomal subunit protein L3 

cds-NP_418460.1 27673.88358975
59 -1.42297527629745 8.145553885670

46e-13 
1.692638302653

92e-11 lamB 
maltose outer membrane 

channel/phage lambda receptor 
protein 

cds-NP_414918.4 43.53544829229
31 -1.43657032092163 0.000118761876

620802 
0.000742133568

581504 psiF phosphate starvation-inducible 
protein PsiF 



Supplementary Material 

 215 

cds-NP_415127.1 96.89705054871
56 -1.4493495665581 0.000116210678

333631 
0.000730698598

353954 entB enterobactin synthase component 
B 

cds-NP_415116.1 164.0030484256
81 -1.46288393739985 3.976642580819

84e-09 
5.448125781861

38e-08 fepA ferric enterobactin outer 
membrane transporter 

cds-NP_417146.2 93.92346483914
8 -1.46289974682828 1.390077405902

87e-11 
2.536599232704

28e-10 lhgD L-2-hydroxyglutarate 
dehydrogenase 

cds-NP_418708.1 269.5269031672
71 -1.47217439843802 1.045616529990

48e-08 
1.343524434836

89e-07 fecD ferric citrate ABC transporter 
membrane subunit FecD 

cds-NP_416449.1 34.24669179818
44 -1.47570568880196 0.001221130860

6148 
0.005752029639

53369 fliG flagellar motor switch protein FliG 

cds-NP_417478.1 96.90464890985
35 -1.47597471293135 1.865308917526

41e-07 
1.985548193337

54e-06 exbD Ton complex subunit ExbD 

cds-YP_026218.1 30845.38151732
85 -1.47749019238518 2.523977172583

84e-20 
9.289519373331

86e-19 malP maltodextrin phosphorylase 

cds-NP_414756.2 1532.269030477
09 -1.48812541898114 0.003089553393

8508 
0.012694352307

9413 fadE acyl-CoA dehydrogenase 

cds-NP_417778.1 37471.06936255
64 -1.5273990579883 5.974716663031

52e-22 
2.621029744196

55e-20 rplD 50S ribosomal subunit protein L4 

cds-NP_418707.1 296.2178161011
54 -1.55511197143738 5.043860371966

6e-10 
7.823387712661

05e-09 fecE ferric citrate ABC transporter ATP 
binding subunit 

cds-NP_417193.1 61.93734963142
93 -1.56230518614811 1.211868973377

68e-06 
1.146655109232

95e-05 hydN putative electron transport protein 
HydN 

cds-NP_415123.1 77.19739867623
04 -1.56303829150945 2.283902431796

96e-07 
2.395890884370

58e-06 entS enterobactin exporter EntS 

cds-NP_415417.1 50.16134356437
95 -1.56779531893067 6.921554319715

49e-09 
9.249326280161

35e-08 ycaC putative hydrolase YcaC 

cds-NP_417875.1 15701.06115001
4 -1.58688052783797 1.501192691155

9e-13 
3.449566062269

89e-12 malQ 4-alpha-glucanotransferase 

cds-NP_417777.1 37003.97248350
76 -1.5926873200702 1.169682139591

54e-32 
9.960646141658

94e-31 rplW 50S ribosomal subunit protein L23 

cds-NP_415125.1 70.23112095305
18 -1.59512222542652 0.000105223798

397708 
0.000670068851

086873 entC isochorismate synthase EntC 

cds-NP_417541.1 74.80943488573
52 -1.6106752470405 5.615745980448

05e-10 
8.679425193268

99e-09 nfeF NADPH-dependent ferric-chelate 
reductase 

cds-NP_415126.1 95.07837122426
74 -1.65167703309721 2.738576367558

17e-06 
2.412502467404

69e-05 entE 2 3-dihydroxybenzoate-AMP 
ligase 

cds-NP_417047.1 3030.237647276
76 -1.66118749630172 1.817042262793

77e-18 
5.889115333815

93e-17 hmp nitric oxide dioxygenase 

cds-NP_415597.1 33.82723420476
81 -1.67799526895522 0.000201427035

677826 
0.001190200838

02558 flgH flagellar L-ring protein 

cds-NP_415537.1 134.5325020600
05 -1.68567058179516 1.195231927260

29e-11 
2.199530618682

81e-10 efeO ferrous iron transport system 
protein EfeO 

cds-NP_415556.1 10.13639077664
66 -1.69380236261667 0.008076116937

00222 
0.028702598901

3098 csgF curli assembly component CsgF 

cds-NP_417776.1 51031.72307326
07 -1.72440063028702 4.861220932257

22e-21 
1.902007433224

61e-19 rplB 50S ribosomal subunit protein L2 

cds-NP_416873.1 15.17828521221
03 -1.7718883524648 0.000810864155

18741 
0.004038512644

47124 yfdV putative transport protein YfdV 

cds-NP_416456.1 22.89676509589
27 -1.78505673235187 0.001235645423

01977 
0.005807800943

91218 fliN flagellar motor switch protein FliN 

cds-NP_417968.1 3240.549274029
7 -1.81576515265544 3.963749274292

29e-21 
1.564960281659

22e-19 hdeD acid-resistance membrane protein 

cds-YP_025308.1 181.4977481902
87 -1.81768084712021 4.174692001047

44e-22 
1.869143026860

73e-20 katE catalase HPII 

cds-NP_415596.1 133.5854476898
87 -1.82882997842255 1.044739676746

34e-08 
1.343524434836

89e-07 flgG flagellar basal-body rod protein 
FlgG 

cds-NP_417774.1 24570.38832739
97 -1.83585067845418 6.438762797390

33e-53 
1.215806383872

44e-50 rplV 50S ribosomal subunit protein L22 

cds-NP_417775.1 14135.83483927
52 -1.84386242306937 6.046292787156

91e-34 
5.358989709106

62e-32 rpsS 30S ribosomal subunit protein S19 



Supplementary Material 

  216 

cds-NP_417773.1 40911.91302582
74 -1.85619865990396 3.496634196543

2e-35 
3.301278764258

07e-33 rpsC 30S ribosomal subunit protein S3 

cds-NP_416432.3 10.57657096811
39 -1.86669022643928 0.011476621114

6424 
0.038048065267

8566 fliA RNA polymerase sigma factor FliA 

cds-NP_416442.1 40.13785569700
21 -1.88101312718122 1.487161124974

49e-07 
1.610658545078

36e-06 yedL putative acetyltransferase YedL 

cds-NP_418711.1 2498.566846138
09 -1.91846606174523 6.888853674325

61e-17 
2.007939027355

44e-15 fecA ferric citrate outer membrane 
transporter 

cds-NP_417770.1 4192.002234261
45 -1.93011381982074 6.065068481360

48e-17 
1.791877035003

3e-15 rpsQ 30S ribosomal subunit protein S17 

cds-NP_417771.1 57144.89870988
18 -1.94221429266863 4.416525659829

94e-49 
7.992071225267

26e-47 rpmC 50S ribosomal subunit protein L29 

cds-NP_417772.1 39157.55136943
39 -1.95712976027055 4.045385059578

36e-61 
1.098069207109

3e-58 rplP 50S ribosomal subunit protein L16 

cds-NP_416314.1 526.8863972574
59 -1.95765671061492 1.808668922939

76e-24 
8.727832369252

65e-23 dmlA D-malate/3-isopropylmalate 
dehydrogenase (decarboxylating) 

cds-NP_417145.4 117.6972761148
02 -1.97027258586629 4.017588420351

91e-13 
8.768033421903

7e-12 glaH glutarate dioxygenase GlaH 

cds-NP_417825.1 9.682547717162
64 -1.97259540322512 0.011493762171

5027 
0.038075826934

2764 nirD nitrite reductase (NADH) small 
subunit 

cds-NP_418503.1 930.1156076512
03 -1.99782162608551 1.288148557491

15e-27 
8.227101742917

73e-26 fdhF formate dehydrogenase H 

cds-NP_415224.1 155.2016512257
47 -2.08039551756212 4.755831049281

59e-11 
8.228914042641

41e-10 kdpC K(+) transporting P-type ATPase 
subunit KdpC 

cds-NP_418709.1 243.5409628728
09 -2.08587468844096 5.991886739339

6e-12 
1.141349303024

2e-10 fecC ferric citrate ABC transporter 
membrane subunit FecC 

cds-NP_417966.4 1679.488522695
56 -2.13122530215174 6.075006427540

45e-39 
6.281845932097

18e-37 hdeB periplasmic acid stress chaperone 
HdeB 

cds-NP_418787.1 188.0729906070
14 -2.15156173509937 3.806011927785

54e-12 
7.445725136203

88e-11 fhuF ferric-siderophore reductase FhuF 

cds-NP_415690.5 19.84347756703
91 -2.17685278790244 7.027974986715

92e-05 
0.000463867710

74935 ymgG PF13436 family protein YmgG 

cds-NP_416451.1 38.56110423085
87 -2.18808595097511 1.635378147716

75e-06 
1.503855749280

24e-05 fliI flagellar export ATPase FliI 

cds-NP_417974.1 574.8032304543
31 -2.1991897611389 1.010926376892

22e-24 
5.165239123344

6e-23 gadA glutamate decarboxylase A 

cds-NP_418710.4 1434.246756293
3 -2.20301506848223 9.247370594359

69e-25 
4.781110772774

3e-23 fecB ferric citrate ABC transporter 
periplasmic binding protein 

cds-NP_417269.1 66.79334860565
83 -2.33392793073627 1.829155467229

63e-10 
2.975289211302

73e-09 gudP galactarate/D-glucarate 
transporter GudP 

cds-NP_417205.1 37.84374882569
24 -2.40029241663967 6.688369202670

41e-07 
6.586754523174

05e-06 hycA regulator of the transcriptional 
regulator FhlA 

cds-NP_416448.1 23.21888947266
63 -2.44461700694058 8.797088862320

23e-06 
6.959154267587

76e-05 fliF flagellar basal-body MS-ring and 
collar protein 

cds-NP_415594.1 179.0493645248
11 -2.44946929421763 6.110584356798

32e-17 
1.793126206863

18e-15 flgE flagellar hook protein FlgE 

cds-NP_417967.1 9328.477129582
23 -2.52213416624339 3.378481237201

11e-32 
2.768442266634

79e-30 hdeA periplasmic acid stress chaperone 
HdeA 

cds-NP_417206.1 35.14432160569
08 -2.54649287087628 2.178688774721

15e-07 
2.302200814747

92e-06 hypA hydrogenase 3 nickel 
incorporation protein HypA 

cds-NP_415595.1 93.28277160496
82 -2.5576185913019 5.934964801512

64e-10 
9.107968951579

29e-09 flgF flagellar basal-body rod protein 
FlgF 

cds-NP_417597.1 1035.388026483
83 -2.55781124317962 5.672258467454

65e-14 
1.317359279366

61e-12 garD GarD 

cds-NP_415225.1 506.5134651539
28 -2.56776789503203 7.249819291409

17e-22 
3.117422295305

94e-20 kdpB K(+) transporting P-type ATPase 
subunit KdpB 

cds-NP_416010.1 657.0809259000
64 -2.73445177648057 8.874992703491

59e-47 
1.427559011528

3e-44 gadB glutamate decarboxylase B 

cds-NP_415226.1 572.0999757733
65 -2.74267750347421 1.761974360221

41e-33 
1.530450929288

32e-31 kdpA K(+) transporting P-type ATPase 
subunit KdpA 



Supplementary Material 

 217 

cds-NP_417199.1 153.8249749384
79 -2.75165527486185 1.929450969302

84e-22 
8.728755791335

68e-21 hycG formate hydrogenlyase subunit 
HycG 

cds-NP_417198.1 149.5084824214
43 -2.75676126528329 4.258943374070

23e-25 
2.283529762171

23e-23 hycH formate hydrogenlyase assembly 
protein 

cds-NP_415592.1 49.20470820356
51 -2.87724622363956 0.000127563708

109342 
0.000788064273

568805 flgC flagellar basal-body rod protein 
FlgC 

cds-NP_417203.1 340.6448002896
95 -3.05905570512471 1.220146834756

18e-27 
7.909101049770

26e-26 hycC formate hydrogenlyase subunit 
HycC 

cds-NP_417200.1 123.7834809112
14 -3.09049612872538 1.767689002042

98e-24 
8.625925096486

15e-23 hycF formate hydrogenlyase subunit 
HycF 

cds-NP_417595.1 109.0990447232
26 -3.11949361168915 5.676572491977

55e-19 
1.911112738965

77e-17 garL alpha-dehydro-beta-deoxy-D-
glucarate aldolase 

cds-NP_417201.1 377.4558417983
58 -3.14519624405024 1.677213616238

94e-42 
2.023371870923

82e-40 hycE formate hydrogenlyase subunit 
HycE 

cds-NP_416009.1 668.1555314682
67 -3.18968708460067 5.544534030763

03e-55 
1.203995564780

19e-52 gadC L-glutamate:4-aminobutyrate 
antiporter 

cds-NP_417202.1 235.1334728617
98 -3.22060841932529 6.726022809602

86e-31 
4.951036790187

33e-29 hycD formate hydrogenlyase subunit 
HycD 

cds-NP_415593.1 84.51190011749
51 -3.55915718399414 1.671061993919

36e-13 
3.779907416454

05e-12 flgD flagellar biosynthesis initiation of 
hook assembly 

cds-NP_417596.1 316.6957065844
05 -4.46965742932001 7.194580548203

16e-27 
4.400853988851

6e-25 garP galactarate/D-glucarate 
transporter GarP 

cds-NP_415591.1 59.02754272356
72 -4.54790863851073 5.764029636417

83e-11 
9.740537241619

7e-10 flgB flagellar basal-body rod protein 
FlgB 

cds-NP_417204.1 47.86153764001
89 -4.98096950708198 1.962714187552

42e-12 
3.946327646546

37e-11 hycB formate hydrogenlyase subunit 
HycB 

cds-NP_414674.1 79.98735589993
18 1.00268493406297 3.902303414500

7e-06 
3.324602704040

47e-05 rpnC recombination-promoting nuclease 
RpnC 

cds-NP_415353.1 417.6297922033
51 1.01270732939962 4.051585958728

79e-16 
1.120766740048

35e-14 gsiD glutathione ABC transporter 
membrane subunit GsiD 

cds-YP_026261.1 33.92884962003
93 1.01314357823549 0.013259993153

1268 
0.042563303964

5454 yigE DUF2233 domain-containing 
protein YigE 

cds-NP_415183.1 79.35411332617
16 1.01638685368055 0.000902309257

449669 
0.004432951476

36189 hscC chaperone protein HscC 

cds-YP_001165321.1 23.29445060968
19 1.01639165480622 0.005915518019

89982 
0.021995800308

583 cydH cytochrome bd-I accessory 
subunit CydH 

cds-NP_416482.1 49.57070942379
01 1.02522073753928 0.000110456212

604475 
0.000698269769

055656 zinT metal-binding protein ZinT 

cds-YP_010051179.1 26.59878055212
3 1.02544802384302 0.013971152154

3594 
0.044192799567

6495 yljB protein YljB 

cds-NP_415011.1 2338.426994710
37 1.02746116769765 1.685188424911

29e-20 
6.309287352922

2e-19 ybaL putative transporter YbaL 

cds-NP_415190.1 1587.730426104
48 1.02900205725677 6.399357787419

43e-32 
5.053165612865

92e-30 lnt apolipoprotein N-acyltransferase 

cds-NP_414672.1 308.6292918903
86 1.03074261397499 1.259488995441

33e-05 
9.504344240533

24e-05 yadE putative polysaccharide 
deacetylase lipoprotein YadE 

cds-NP_417645.1 18811.30989154
5 1.03317395749964 1.943238662366

77e-46 
2.910167417468

58e-44 ftsH ATP-dependent zinc 
metalloprotease FtsH 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.03394082670175 0.004354405826
38188 

0.016908394541
0644 insO IS911B regulator fragment 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.03394082670175 0.004354405826
38188 

0.016908394541
0644 insO IS911B regulator fragment 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.03394082670175 0.004354405826
38188 

0.016908394541
0644 insO IS911B regulator fragment 

cds-NP_414776.1 32.25585971117
02 1.03680459881756 0.004645757654

47186 
0.017799050561

3802 phoE outer membrane porin PhoE 

cds-NP_416778.2 51.27857766603
5 1.04571214130933 0.002022953525

37703 
0.008904276453

22023 yfbP uncharacterized protein YfbP 



Supplementary Material 

  218 

cds-YP_588467.1 33.54052743296
41 1.05815989189551 0.003251666239

51889 
0.013260081200

2164 arfA alternative ribosome-rescue factor 
A 

cds-NP_418306.1 24796.04273014
36 1.06856524328365 2.057145871787

82e-09 
2.929240826614

59e-08 glnA glutamine synthetase 

cds-NP_416914.1 20.40291719345
07 1.06996910762782 0.010804994692

8048 
0.036260632402

093 yfeK DUF5329 domain-containing 
protein YfeK 

cds-NP_416826.1 18427.00105588
54 1.07751473713238 6.871131027744

64e-23 
3.174608729095

21e-21 fabB 3-oxoacyl-[acyl carrier protein] 
synthase 1 

cds-YP_588445.1 41.97181210256
99 1.07943610085259 0.000188568245

637294 
0.001120317224

08039 insA4 IS1 family protein InsA 

cds-NP_414992.1 97.56845814048
65 1.08031230102381 0.000211222089

441849 
0.001239645316

81885 maa maltose O-acetyltransferase 

cds-NP_415244.4 20.91364380693
49 1.08238641120814 0.006549464612

074 
0.023943034352

0517 ybgO putative fimbrial protein YbgO 

cds-NP_415632.1 3715.879450211
28 1.08317700261552 1.143896405558

16e-28 
7.885622364030

29e-27 mfd transcription-repair coupling factor 

cds-YP_026189.1 1233.752131885
17 1.08366259388887 1.447101406270

3e-11 
2.629607283444

32e-10 yghJ putative lipoprotein YghJ 

cds-NP_417978.2 99.12833624031
33 1.10699528051636 1.599059811965

99e-06 
1.480749842935

67e-05 rcdB putative DNA-binding 
transcriptional regulator YhjC 

cds-NP_416255.1 155.8784888530
94 1.10845705317475 1.087185007146

75e-09 
1.606001525863

37e-08 cho excinuclease Cho 

cds-NP_417393.1 22.05678284134
77 1.11162752074332 0.011965126914

916 
0.039248146670

3023 argK methylmalonyl-CoA mutase-
interacting GTPase YgfD 

cds-NP_418736.3 89.20910703503
14 1.11219189573814 3.678073156628

96e-06 
3.163142914700

91e-05 fimC type 1 fimbriae periplasmic 
chaperone 

cds-NP_416650.1 24.78795173660
89 1.11320476061029 0.007278934346

48218 
0.026299172919

9033 yeiS DUF2542 domain-containing 
protein YeiS 

cds-NP_416761.4 87.85953999989
41 1.11581584274149 8.400072774890

04e-05 
0.000546949266

287068 arnF undecaprenyl-phosphate-alpha-L-
Ara4N flippase - ArnF subunit 

cds-NP_418554.1 63.17356605423
84 1.11750114110578 0.000116603062

233201 
0.000730746175

00547 dtpC dipeptide/tripeptide:H(+) 
symporter DtpC 

cds-YP_588446.1 57.37993177778
82 1.12991717312052 2.058597691047

39e-05 
0.000150767112

516337 gnsA putative phosphatidylethanolamine 
synthesis regulator GnsA 

cds-NP_415207.1 75.31040183062
54 1.13821866755718 0.001641293091

0274 
0.007402010274

48805 chiP chitooligosaccharide outer 
membrane channel 

cds-NP_415354.1 118.9926249681
93 1.13993725573957 1.167333294018

1e-08 
1.495495131540

01e-07 pdeI putative c-di-GMP 
phosphodiesterase PdeI 

cds-YP_588463.1 27.70653292056
9 1.14159629574642 0.004491896647

39357 
0.017299000253

158 torI 
prophage CPS-53 recombination 
directionality factor and response 

regulator inhibitor 

cds-NP_416608.1 24.67395496634
39 1.14466901238029 0.004218052048

0169 
0.016518485161

8913 rcnR DNA-binding transcriptional 
repressor RcnR 

cds-NP_416304.1 152.2072573335
35 1.14620157113302 4.139437265726

24e-12 
8.025703591539

75e-11 nimR DNA-binding transcriptional 
repressor NimR 

cds-NP_415191.1 3196.834043719
81 1.14678701797604 9.974870844468

57e-38 
9.845650926710

68e-36 ybeX CorC-HlyC family protein YbeX 

cds-NP_414924.1 162.5383638944
77 1.15666251817249 8.847730133487

06e-09 
1.153924683775

81e-07 aroM protein AroM 

cds-YP_588471.1 13.35832895928
48 1.16078879033091 0.009177995226

47666 
0.031811678586

2635 yicS uncharacterized protein YicS 

cds-NP_416299.1 23.98909623075
97 1.174012152535 0.005300299574

38702 
0.019947314602

7408 cdgI putative c-di-GMP binding protein 
CdgI 

cds-NP_416999.1 61.79703500479
02 1.18262242379944 0.000499858889

556138 
0.002644198730

01499 yfgG nickel/cobalt stress response 
protein YfgG 

cds-NP_416649.1 91.39323008657
08 1.18546208322962 2.194084171826

64e-06 
1.972858707710

79e-05 sanA DUF218 domain-containing 
protein SanA 

cds-NP_417628.2 26.70273219621
74 1.18834395924203 0.009263691126

28344 
0.032006531870

6834 ubiV ubiquinone biosynthesis protein 
UbiV 



Supplementary Material 

 219 

cds-NP_415289.4 79.50780215137
68 1.19692503229618 1.237501090561

45e-06 
1.168362442675

74e-05 ybhD putative DNA-binding 
transcriptional regulator YbhD 

cds-NP_417873.1 1964.490037176
57 1.19747359870764 1.371042906750

15e-21 
5.670894613348

48e-20 nfuA iron-sulfur cluster carrier protein 
NfuA 

cds-NP_417446.4 50.23042037861
56 1.19779877849073 0.001525613745

33419 
0.006974463679

98565 pppA prepilin peptidase 

cds-NP_418200.1 4993.314500759
5 1.21186213327494 6.402771780045

44e-09 
8.609051963076

57e-08 asnA asparagine synthetase A 

cds-NP_415508.1 406.3412469790
03 1.2185614796058 1.530260392827

48e-08 
1.920786383251

37e-07 insB4 IS1 family protein InsB 

cds-YP_010051200.1 11.76742723905
53 1.22743924006478 0.012616363768

857 
0.040890199886

6759 yqiM protein YqiM 

cds-NP_416895.2 589.5674590721
4 1.23956734869858 6.713049587434

4e-22 
2.915477435822

76e-20 insL3 putative IS186/IS421 transposase 

cds-NP_418565.1 162.9283525053
78 1.25181473705068 7.683710605218

69e-09 
1.014296509375

83e-07 yjeH L-methionine/branched chain 
amino acid exporter 

cds-NP_418178.1 101.9515860857
31 1.25457327605787 0.000420063245

255476 
0.002280418342

68066 bglF 
beta-glucoside specific PTS 
enzyme II/BglG kinase/BglG 

phosphatase 

cds-NP_417752.1 201.3814992450
56 1.25476627136627 6.355582643901

72e-15 
1.577274024140

87e-13 yhdN DUF1992 domain-containing 
protein YhdN 

cds-NP_417783.1 11.48203502349
11 1.26264207681261 0.015822850355

2708 
0.048771212982

925 gspC Type II secretion system protein 
GspC 

cds-NP_417074.1 131.3512653872
55 1.27088363472802 4.069607892468

92e-06 
3.458768508217

71e-05 grcA stress-induced alternate pyruvate 
formate-lyase subunit 

cds-NP_417065.1 936.9139784418
68 1.2711153546787 8.384527355032

42e-27 
5.057500319848

03e-25 rseC protein RseC 

cds-NP_415355.1 69.38485796084
07 1.271465253493 0.000283915250

942663 
0.001613931851

89003 dgcI putative diguanylate cyclase DgcI 

cds-
gnl|b4580|CDS%3D3

93 
56.26462673832

47 1.28271461617629 4.408657233838
73e-05 

0.000303322851
043347 yaiT putative autotransporter YaiT 

cds-
gnl|b4580|CDS%3D3

93 
56.26462673832

47 1.28271461617629 4.408657233838
73e-05 

0.000303322851
043347 yaiT putative autotransporter YaiT 

cds-NP_418014.1 53.24318069765
57 1.29734741191224 1.103260974595

44e-05 
8.490828874328

74e-05 insJ insertion element IS150 protein 
InsA 

cds-NP_414993.1 399.8376879697
56 1.30934730348542 1.614280697445

67e-15 
4.278040416602

16e-14 hha hemolysin expression-modulating 
protein Hha 

cds-NP_418388.1 33.48311635901
06 1.31252986566686 0.000448304850

155348 
0.002409520220

03817 frwD putative PTS enzyme IIB 
component FrwD 

cds-NP_417902.1 13.10737706372
63 1.31900418027287 0.010130930376

4808 
0.034508729902

0047 insB6 IS1 family protein InsB 

cds-NP_415031.1 16.04684493373 1.32820237157265 0.008763253893
07731 

0.030593900046
3302 ybbC PF15631 family protein YbbC 

cds-NP_416646.1 156.8730078237
44 1.3306655453555 8.175397293435

27e-09 
1.072681282338

05e-07 yohJ putative 3-hydroxypropanoate 
export protein YohJ 

cds-NP_418716.1 36.26806575717
54 1.33324426061591 0.000190376951

923512 
0.001129517899

18554 yjhF putative transporter YjhF 

cds-NP_414746.1 89.91894266638
79 1.34300039558894 1.325563654625

85e-07 
1.446463053276

4e-06 yafE putative S-adenosylmethionine-
dependent methyltransferase 

cds-NP_414841.1 912.5979546853
23 1.35583387598887 3.493613439321

13e-12 
6.896710530441

68e-11 ykgF 

putative amino acid 
dehydrogenase with NAD(P)-

binding domain and ferridoxin-like 
domain 

cds-NP_418304.1 3935.605930174
47 1.36161113595132 1.600683047191

28e-19 
5.745261548720

44e-18 glnG DNA-binding transcriptional dual 
regulator NtrC 

cds-NP_417954.1 743.3995509171
86 1.36794587421769 8.358666219470

79e-39 
8.442252881665

5e-37 rsmJ 16S rRNA m(2)G1516 
methyltransferase 

cds-NP_415054.1 19.64457867772
02 1.36872188126339 0.002098205809

01962 
0.009158299325

19819 ybcF putative carbamate kinase 



Supplementary Material 

  220 

cds-NP_414573.1 673.4300652205
16 1.3721033358656 9.415774935720

74e-15 
2.323449462831

54e-13 carA carbamoyl-phosphate synthetase 
small subunit 

cds-NP_418735.2 248.1532755165
23 1.3742536154512 9.927682123729

56e-20 
3.592993621946

46e-18 fimI putative fimbrial protein FimI 

cds-NP_415114.1 566.5219506042
56 1.39268922112135 3.091281170481

34e-21 
1.231691203981

69e-19 insL2 IS186/IS421 family transposase 

cds-NP_418511.1 42.63006800106
37 1.40171342793169 5.105879278980

97e-08 
5.977044126311

15e-07 alsA D-allose ABC transporter ATP 
binding subunit 

cds-NP_418012.1 34841.40606166
52 1.41335848129883 0.001711812156

57954 
0.007672239624

38074 cspA cold shock protein CspA 

cds-NP_415098.1 17.86704125140
09 1.41338893149258 0.000943124254

982763 
0.004597069180

01138 envY DNA-binding transcriptional 
activator EnvY 

cds-NP_417955.1 3903.808357406
6 1.41513898611235 6.295944072525

34e-58 
1.439120268788

29e-55 prlC oligopeptidase A 

cds-NP_418015.1 35.22050422382
51 1.41651446521509 3.771006191831

97e-06 
3.236656104965

66e-05 insK IS150 family conserved protein 
InsB 

cds-NP_416242.6 65.88491953164
94 1.42349323569878 5.531616780869

1e-08 
6.458013892288

85e-07 ydjM inner membrane protein YdjM 

cds-NP_418365.1 313.1483386385
6 1.42618605044512 2.529792793185

66e-19 
8.893153760436

22e-18 menA 1 4-dihydroxy-2-naphthoate 
octaprenyltransferase 

cds-NP_418000.1 104.4324927768
59 1.43998261040133 9.627227292118

36e-07 
9.270742379084

27e-06 dppB dipeptide ABC transporter 
membrane subunit DppB 

cds-NP_416492.1 65.08891664272
58 1.44555558142941 1.460956520534

86e-06 
1.361573855940

54e-05 cbl DNA-binding transcriptional 
activator Cbl 

cds-NP_417646.1 3297.229786281
05 1.44592769395651 1.112854787589

81e-58 
2.843016672060

32e-56 rlmE 23S rRNA 2'-O-ribose U2552 
methyltransferase 

cds-NP_414768.1 47.40810799447
06 1.44774013489693 9.251215803972

5e-05 
0.000596113208

25894 yafO ribosome-dependent mRNA 
interferase toxin YafO 

cds-YP_009518769.1 9.282449830567
93 1.45672476470571 0.012328289517

9599 
0.040136252905

9219 ychS putative uncharacterized protein 
YchS 

cds-NP_416335.4 100.3404221915
42 1.48236103435601 3.702351994948

04e-09 
5.104544353669

63e-08 mntP Mn(2(+)) exporter 

cds-NP_416111.1 1904.616960475
3 1.48735859813175 4.509145570429

41e-25 
2.359424001490

95e-23 mlc DNA-binding transcriptional 
repressor Mlc 

cds-NP_417419.1 99.82983432080
4 1.4915398878005 1.393572201344

97e-06 
1.304371566905

43e-05 yggI protein YggI 

cds-NP_416647.1 543.5021968253
51 1.50505565712558 1.585174346957

73e-10 
2.607731889711

15e-09 yohK putative 3-hydroxypropanoate 
export protein YohK 

cds-NP_415097.1 4805.989202823
66 1.52223917337474 4.943394423832

54e-42 
5.649779469132

82e-40 ompT omptin family outer membrane 
protease OmpT 

cds-NP_418633.4 28.63031426004
53 1.525306918252 0.000182240453

984168 
0.001084205878

97705 ytfH putative transcriptional regulator 
YtfH 

cds-NP_415193.2 2457.726498602
44 1.52924566464311 8.251698306699

12e-91 
3.583712574599

43e-88 ybeZ PhoH-like protein 

cds-NP_417751.1 223.8323402823
22 1.53542311014203 1.459644998155

45e-24 
7.286480720677

13e-23 zntR DNA-binding transcriptional 
activator ZntR 

cds-NP_418487.1 40.47478463926
69 1.53849933466445 1.098037809566

5e-05 
8.470298769000

56e-05 soxR DNA-binding transcriptional dual 
regulator SoxR 

cds-NP_414840.1 457.7445069435
93 1.54568503202701 5.235525468932

34e-11 
8.916818475126

73e-10 ykgE putative lactate utilization 
oxidoreductase YkgE 

cds-NP_418092.1 282.2690357581
44 1.55591898276194 2.945148180806

34e-26 
1.639843403748

97e-24 mutM DNA-formamidopyrimidine 
glycosylase 

cds-NP_415192.1 842.1525848052
43 1.57194826594345 3.055163936158

74e-58 
7.371431652631

9e-56 ybeY endoribonuclease YbeY 

cds-NP_416808.1 40.70327276883
25 1.57623139146486 1.150012248169

84e-06 
1.097693009626

73e-05 rpnB recombination-promoting nuclease 
RpnB 

cds-NP_415510.1 351.5100485675
8 1.59880780740446 0.001286690992

11255 
0.006021658382

26809 cspG cold shock protein CspG 

cds-NP_418556.1 450.4023204511
75 1.63169610321649 0.006901002979

51289 
0.025080381539

7694 cadB lysine:cadaverine antiporter 



Supplementary Material 

 221 

cds-NP_417860.2 1316.627641117
39 1.66087491448982 2.825126010087

56e-46 
3.957910407035

57e-44 hslO molecular chaperone Hsp33 

cds-NP_415891.1 11.28631084495
63 1.66482796574269 0.002483889168

1913 
0.010576010448

4851 tfaR putative tail fiber assembly protein 
TfaR 

cds-NP_414557.1 627.1975114961
7 1.69304999990861 1.566291390446

02e-26 
8.950530932509

32e-25 insL1 IS186/IS421 family transposase 

cds-NP_415025.4 2522.585859251
35 1.70344024583744 6.020716165830

52e-63 
1.743198020546

8e-60 cnoX chaperedoxin 

cds-NP_418592.1 6758.253784722
78 1.71467933185072 1.124833165706

38e-69 
3.757808029740

61e-67 miaA tRNA dimethylallyltransferase 

cds-NP_416117.1 252.0627855961
59 1.72268214620149 3.394198006682

58e-11 
5.968016980980

75e-10 mdtJ multidrug/spermidine efflux pump 
membrane subunit MdtJ 

cds-YP_009518777.1 9.330592992853
66 1.72308374177426 0.005807519670

36874 
0.021631267520

0784 ydaY putative uncharacterized protein 
YdaY 

cds-NP_416116.1 136.3495821180
21 1.73262781670921 3.849882536254

34e-14 
9.136633800520

55e-13 mdtI multidrug/spermidine efflux pump 
membrane subunit MdtI 

cds-NP_414842.4 724.9604012807
69 1.73918717833177 3.083490368171

1e-15 
7.971189683908

98e-14 ykgG DUF162 domain-containing lactate 
utilization protein YkgG 

cds-NP_415507.2 13.56824978534
35 1.74609550746891 0.000685650507

982354 
0.003499154120

05566 gfcA threonine-rich inner membrane 
protein GfcA 

cds-NP_415633.1 13.27676564564
84 1.76071652128192 0.001393024319

36078 
0.006436068743

59986 ycfT inner membrane protein YcfT 

cds-NP_416256.2 10.83360515263
15 1.7631785299902 0.004815051536

33161 
0.018343656861

6563 ves HutD family protein Ves 

cds-NP_418555.1 674.9255446437
39 1.76866863725081 0.006384800114

82657 
0.023465309539

8666 cadA lysine decarboxylase 1 

cds-YP_588452.1 8.283356601710
5 1.79489508767621 0.009317086024

05384 
0.032165424962

2145 rzoR putative prophage outer 
membrane lipoprotein RzoR 

cds-YP_009518791.1 40.57072597008
76 1.79823505553143 0.000680643629

513194 
0.003477688568

20683 ynfS protein YnfS 

cds-YP_009518787.1 16.27977968490
59 1.83510647261052 0.000763033802

966135 
0.003848845303

46333 ynfT protein YnfT 

cds-YP_009518761.1 34.13499990501
9 1.83628295903632 0.000923979936

331758 
0.004524064107

65369 ymcF protein YmcF 

cds-NP_414973.1 22244.68729894
82 1.88981203457347 2.227762624902

9e-123 
3.225057693317

77e-120 lon Lon protease 

cds-NP_418340.1 9.839581373527
08 1.90692505474556 0.003858158800

76585 
0.015330268684

1044 rhaB rhamnulokinase 

cds-YP_009518818.1 21.98190121158
58 1.93435810571 2.584551431655

72e-06 
2.295441077235

34e-05 yqiD protein YqiD 

cds-NP_418305.1 2114.394713807
07 1.94088957828788 6.208214706582

97e-37 
5.991616993486

63e-35 glnL protein histidine kinase NtrB 

cds-NP_417104.1 5428.605549201
47 1.9502369103212 1.542963880646

43e-46 
2.393247190588

38e-44 grpE nucleotide exchange factor GrpE 

cds-NP_418441.1 26.81399071589
53 1.97061387769163 1.585836628134

63e-06 
1.471642836749

72e-05 arpA regulator of acetyl CoA synthetase 

cds-NP_415486.4 1853.581680395
75 2.03082094050477 2.192125536500

16e-45 
2.884970062127

33e-43 hspQ heat shock protein HspQ 

cds-NP_417859.1 1010.610415453
52 2.03187661582355 1.975634980165

33e-70 
7.150152265715

01e-68 hslR heat shock protein Hsp15 

cds-NP_416877.1 30.56633160617
53 2.03714957800872 6.156867059737

6e-09 
8.329991788299

19e-08 ypdI colanic acid synthesis putative 
lipoprotein YpdI 

cds-NP_415400.1 882.3079552676
02 2.0474223216713 2.035300541497

77e-48 
3.535724100689

92e-46 macB ABC-type tripartite efflux pump 
ATP binding/membrane subunit 

cds-NP_416075.1 671.9271446626
15 2.08032853595053 5.377415157002

34e-15 
1.365737662389

54e-13 cspB cold shock-like protein CspB 

cds-YP_010051187.1 13.37409804699
16 2.22398285884966 6.848561688511

07e-05 
0.000453404015

445176 mdtU protein MdtU 

cds-NP_416404.1 7.398522387900
37 2.24736339053479 0.006258388776

513 
0.023110100309

1596 motA motility protein A 



Supplementary Material 

  222 

cds-NP_415893.2 28.07718439716
09 2.28074756705748 0.007284804287

96769 
0.026299172919

9033 ynaE uncharacterized protein YnaE 

cds-NP_416059.1 6.440986321228
33 2.28376390117825 0.001823526952

87789 
0.008114321266

7507 ydfZ putative selenoprotein YdfZ 

cds-NP_416727.1 6.323526475029
93 2.35466280246021 0.000846696806

222406 
0.004183395027

78602 atoE short chain fatty acid transporter 

cds-YP_009518779.1 43.29010439386
31 2.40539088213583 0.000666180516

795785 
0.003411818377

88219 ynaM protein YnaM 

cds-YP_588474.1 6.114890429299
78 2.44612189460234 0.007757857178

66053 
0.027844936964

3989 ghoT toxin of the GhoTS toxin-antitoxin 
system 

cds-NP_416062.2 21.52596879825
84 2.4876499619846 0.000146273383

368985 
0.000889727316

486698 ydfK cold shock protein YdfK 

cds-NP_415164.1 188.2664250160
82 2.76861052275006 1.023856663468

28e-40 
1.140156279344

29e-38 ybeD DUF493 domain-containing 
protein YbeD 

cds-NP_418366.1 6347.994467281
74 2.79075506673962 3.241531053130

93e-153 
7.038984681873

82e-150 hslU ATPase component of the HslVU 
protease 

cds-NP_417941.1 19.35419292973
96 2.86875278826751 9.370477622361

1e-06 
7.378584635777

77e-05 yhhI putative transposase 

cds-NP_418567.1 24347.88890688
78 2.88342516396534 6.008358098960

21e-68 
1.863878515984

58e-65 groL chaperonin GroEL 

cds-NP_414556.1 3799.557353581
61 2.97684140354745 6.369619825263

46e-95 
3.073695433457

69e-92 dnaJ chaperone protein DnaJ 

cds-NP_418367.1 2990.402902396
6 3.0410720509163 1.123355044120

73e-185 
4.878730956616

34e-182 hslV peptidase component of the 
HslVU protease 

cds-NP_415837.1 819.3407186710
41 3.04783421708749 4.779156358963

95e-88 
1.886897824270

95e-85 ycjX DUF463 domain-containing 
protein YcjX 

cds-NP_415511.1 17.92039650075
79 3.13237277749794 0.000103930540

126697 
0.000662805192

026792 ymcE protein YmcE 

cds-NP_415838.1 594.1904410295
7 3.13363966694361 5.343818390393

86e-102 
2.901025408685

07e-99 ycjF DUF697 domain-containing inner 
membrane protein YcjF 

cds-NP_418566.1 3990.305475406
23 3.16345364912059 2.602799353553

29e-46 
3.767985864160

64e-44 groS cochaperonin GroES 

cds-NP_417445.1 6.236953631470
38 3.30869741539388 0.007476810545

66268 
0.026947542074

5336 yghG lipoprotein YghG 

cds-NP_415509.1 20.58657693522 3.36466841792476 9.357827073913
54e-05 

0.000601198860
68057 cspH CspA family protein CspH 

cds-NP_418564.2 466.9887948491
61 3.57557576506118 3.293421658025

76e-112 
2.043332894400

84e-109 fxsA protein FxsA 

cds-NP_414555.1 26343.06470085
49 3.59600851869751 4.361018926998

13e-114 
3.156650866658

81e-111 dnaK chaperone protein DnaK 

cds-NP_415006.1 10418.01854438
76 3.66449470634312 6.878922860890

6e-116 
5.975032396969

58e-113 htpG chaperone protein HtpG 

cds-NP_416244.2 3.061229871938
06 3.75902058334061 0.009177503376

69668 
0.031811678586

2635 ydjO protein YdjO 

cds-NP_416837.1 4.895565727119
67 3.80902391002518 0.001059383902

37438 
0.005067075207

06162 yfcQ putative fimbrial protein YfcQ 

cds-NP_417083.1 1529.174392278
8 4.01806373696876 4.685932625633

67e-28 
3.179844592676

1e-26 clpB chaperone protein ClpB 

cds-NP_416774.2 2.139213662055
67 4.08752086705629 0.008859879378

58316 
0.030906390474

8487 yfbL putative peptidase YfbL 

cds-NP_418141.2 101.3819031325
39 4.33284105777532 1.196251291465

18e-45 
1.623537299635

39e-43 ibpB small heat shock protein IbpB 

cds-NP_415090.2 1.709846115818
21 4.35446676867609 0.007006959557

99446 
0.025401690618

005 ybcV DUF1398 domain-containing 
protein YbcV 

cds-YP_002791243.1 3.159665811240
46 4.40791239458981 0.002604260797

12473 
0.010991549700

5954 yoaJ uncharacterized protein YoaJ 

cds-NP_418142.1 552.0981102414
1 5.0348500900724 3.285993304511

87e-123 

3.567767230373
76e-120 

 
 

ibpA small heat shock protein IbpA 



Supplementary Material 

 223 

ereA3 

GeneID baseMean log2FoldChange p value padj Gene Product 

cds-NP_418708.1 269.5269031672
71 -1.00398727528514 7.219931586200

64e-05 
0.000321054989

912476 fecD ferric citrate ABC transporter 
membrane subunit FecD 

cds-NP_418793.1 292.2100409263
11 -1.00416594321017 6.219858928911

49e-10 
7.099936906223

97e-09 osmY periplasmic chaperone OsmY 

cds-NP_417757.1 36668.11301657
59 -1.00551684738293 1.821718990746

36e-39 
4.034196705007

81e-37 rpsM 30S ribosomal subunit protein S13 

cds-NP_415261.2 97280.31316131
37 -1.00645538277051 7.985828283611

1e-14 
1.443642182371

98e-12 cydA cytochrome bd-I subunit 1 

cds-NP_418189.1 13230.68494082
69 -1.00684890507773 2.595268613632

15e-36 
4.257201736954

36e-34 atpG ATP synthase F1 complex subunit 
gamma 

cds-NP_416739.1 422.7478700901
68 -1.00748936176478 3.048824372110

43e-08 
2.637352176577

56e-07 yfaE 
ferredoxin-like diferric-tyrosyl 
radical cofactor maintenance 

protein YfaE 

cds-NP_414605.1 80.22599670406
51 -1.0082788943345 0.000110744621

539428 
0.000466243278

325215 araB ribulokinase 

cds-NP_417594.3 221.3297938878
87 -1.01120636512006 3.280756535705

32e-10 
3.927154242334

83e-09 garR tartronate semialdehyde reductase 

cds-NP_415155.1 39.09389694631
64 -1.01159732325816 0.002057670874

56251 
0.006547000217

98662 pagP Lipid A palmitoyltransferase 

cds-NP_416738.1 2896.020145575
7 -1.01266297520612 2.512682219935

08e-16 
6.114653600050

81e-15 nrdB ribonucleoside-diphosphate 
reductase 1 subunit beta 

cds-YP_026170.1 2087.809903968
86 -1.01370896022826 4.109012440712

7e-22 
1.685075564807

09e-20 purL phosphoribosylformylglycinamide 
synthetase 

cds-NP_417759.1 101839.8797741
73 -1.01519642857827 6.846386351433

69e-18 
1.981872232058

81e-16 secY Sec translocon subunit SecY 

cds-NP_415763.1 16841.15712143
42 -1.01769604398676 7.194690220171

74e-28 
4.978950466428

23e-26 oppF 

murein tripeptide ABC 
transporter/oligopeptide ABC 

transporter ATP binding subunit 
OppF 

cds-NP_418488.1 605.7891960190
62 -1.0198775514987 5.601402982272

94e-11 
7.540612099844

02e-10 ghxP guanine/hypoxanthine transporter 
GhxP 

cds-NP_415638.3 85.91778971869
6 -1.02251710081071 4.675728173947

87e-06 
2.672103236440

66e-05 cobB 
protein-lysine 

deacetylase/desuccinylase/lipoami
dase 

cds-NP_417470.1 5696.783358132
18 -1.02551450319265 1.103772231881

72e-10 
1.421106748547

71e-09 hybA hydrogenase 2 iron-sulfur protein 

cds-NP_418661.1 6677.817284736
86 -1.02743431177947 0.002957191462

3976 
0.008983128248

94304 treB trehalose-specific PTS enzyme 
IIBC component 

cds-NP_416363.1 302.2304095865
41 -1.02758006768493 1.212339891011

98e-10 
1.547392904118

74e-09 purT phosphoribosylglycinamide 
formyltransferase 2 

cds-NP_416167.1 682.3945380637
98 -1.02977227936841 4.221383824892

52e-24 
2.124603290960

11e-22 nemA N-ethylmaleimide reductase 

cds-NP_415419.2 202.0631585022
07 -1.03047366694851 7.329420824805

8e-05 
0.000325270589

509668 ycaM putative transporter YcaM 

cds-NP_415961.1 130.6439713271
53 -1.03211944402501 5.017426446740

76e-09 
4.949260964947

63e-08 patD gamma-aminobutyraldehyde 
dehydrogenase 

cds-NP_417221.1 9012.678720231
33 -1.03342902156195 5.913192456763

6e-22 
2.380866308273

27e-20 rpoS RNA polymerase sigma factor 
RpoS 

cds-NP_414606.1 562.6814189141
34 -1.03555234849513 1.384772100704

4e-08 
1.291190659793

64e-07 araC DNA-binding transcriptional dual 
regulator AraC 

cds-NP_417533.1 20.07132044517
06 -1.03575326488323 0.004545483540

76462 
0.012996737638

5065 ttdA L(+)-tartrate dehydratase subunit 
alpha 

cds-NP_414917.2 142.8713446604
65 -1.03671396150791 3.005963930491

06e-08 
2.610473381989

2e-07 phoA alkaline phosphatase 

cds-NP_416015.2 88.72185799460
83 -1.03695690145363 5.231259177406

96e-05 
0.000242102893

382815 ydeN putative sulfatase YdeN 

cds-NP_417468.1 4861.956797099
08 -1.0424299303697 1.921542431107

12e-17 
5.096114627169

71e-16 hybC hydrogenase 2 large subunit 



Supplementary Material 

  224 

cds-NP_416586.1 23.76694159372
34 -1.04507911241314 0.003922045573

34995 
0.011413101080

3988 ogrK prophage P2 late control protein 
OgrK 

cds-NP_416606.1 2948.075299451
21 -1.04897773009639 2.607724715699

84e-25 
1.461976299472

73e-23 thiD 

bifunctional 
hydroxymethylpyrimidine 

kinase/phosphomethylpyrimidine 
kinase 

cds-NP_416542.1 1659.491393326
42 -1.04908265424204 1.086724968095

68e-17 
3.065671900443

17e-16 rfbC dTDP-4-dehydrorhamnose 3 5-
epimerase 

cds-NP_415525.2 86.09650577585
67 -1.05365760600607 8.761461970373

93e-07 
5.825233659975

5e-06 ymdF 
stress-induced bacterial 
acidophilic repeat motifs-
containing protein YmdF 

cds-NP_415522.1 4027.639569770
39 -1.05445804328695 2.160422889722

72e-08 
1.921388148309

62e-07 agp glucose-1-phosphatase 

cds-NP_417032.1 367.0757467530
86 -1.06247571190419 1.641553159939

89e-08 
1.502156806895

41e-07 hcaR DNA-binding transcriptional dual 
regulator HcaR 

cds-NP_418547.1 424.9625391950
49 -1.06473386249637 4.101909385480

92e-14 
7.710302296156

23e-13 dcuB anaerobic C4-dicarboxylate 
transporter DcuB 

cds-NP_417722.1 7998.062315344
09 -1.06478351939193 1.082265420298

33e-13 
1.925041584940

29e-12 accC biotin carboxylase 

cds-NP_416742.1 3822.886923093
93 -1.06538929828266 0.000841394810

471559 
0.002934281587

06971 glpQ glycerophosphoryl diester 
phosphodiesterase GlpQ 

cds-NP_418623.1 3378.195545852
05 -1.06836612058525 1.145317697304

75e-13 
2.029044832545

09e-12 rpsR 30S ribosomal subunit protein S18 

cds-NP_418576.1 835.5496985218
53 -1.06961908168645 3.694951842815

69e-09 
3.727777155314

5e-08 frdC fumarate reductase membrane 
protein FrdC 

cds-NP_418787.1 188.0729906070
14 -1.08207161131566 0.000292764269

551237 
0.001127523143

81788 fhuF ferric-siderophore reductase FhuF 

cds-NP_415829.1 70.09575122615
63 -1.08421518081101 5.154280738307

59e-05 
0.000240047136

995752 ycjQ D-guloside 3-dehydrogenase 

cds-NP_418755.4 468.0353507779
45 -1.08519949554972 1.564368925325

55e-09 
1.689899992748

01e-08 yjiM putative dehydratase subunit 

cds-NP_416805.1 21.81344926117
78 -1.08564052617139 0.002655405278

14413 
0.008189556215

48561 yfcG disulfide bond oxidoreductase 
YfcG 

cds-NP_418754.2 148.6568375173
73 -1.08674788294869 2.743398912117

01e-06 
1.635331599161

e-05 yjiL putative ATPase activator of (R)-
hydroxyglutaryl-CoA dehdratase 

cds-NP_417079.1 3258.397975470
84 -1.08733794033664 2.864584915633

05e-11 
3.977193288820

93e-10 patZ peptidyl-lysine N-acetyltransferase 

cds-NP_418433.1 774.2333670130
63 -1.0881718130494 7.471534891087

7e-17 
1.923920234455

08e-15 purD phosphoribosylamine--glycine 
ligase 

cds-NP_418497.1 32.10643244811
49 -1.08817992542051 0.000444474438

396904 
0.001653594249

61522 nrfD putative menaquinol-cytochrome c 
reductase subunit NrfD 

cds-NP_417760.1 107998.2300433
81 -1.08930633683834 4.637600503950

84e-31 
4.191822986122

1e-29 rplO 50S ribosomal subunit protein L15 

cds-NP_416288.1 58.40891750962
84 -1.09095323660077 0.000177399029

055748 
0.000714272999

716278 ydjJ putative zinc-binding 
dehydrogenase YdjJ 

cds-YP_010051192.1 40.20926207756
39 -1.09370955341606 0.000252912875

341144 
0.000983451382

69177 pssL protein PssL 

cds-NP_417186.1 296.4065670410
99 -1.09675442978413 0.006760672893

36051 
0.018392518577

8217 gutM DNA-binding transcriptional 
activator GutM 

cds-NP_417026.1 1788.774338029
92 -1.09772336916517 1.520834092563

18e-08 
1.399578207064

54e-07 iscR DNA-binding transcriptional dual 
regulator IscR 

cds-NP_417207.1 174.7219722213
33 -1.09809616390158 4.682100986638

12e-10 
5.410631527582

52e-09 hypB hydrogenase isoenzymes nickel 
incorporation protein HypB 

cds-NP_418719.1 67.64251113343
79 -1.10029884508553 0.000596924796

004598 
0.002156427342

17322 yjhI putative DNA-binding 
transcriptional regulator YjhI 

cds-NP_417763.1 14704.90594751
4 -1.10567502730511 3.932298234939

65e-13 
6.647385069674

7e-12 rplR 50S ribosomal subunit protein L18 

cds-NP_418445.1 278.1955669770
9 -1.10678454348185 2.759485528356

69e-07 
1.952358051931

6e-06 pepE peptidase E 

cds-NP_416032.1 27.25490864909
74 -1.10691898418768 0.010003387345

3516 
0.025924518755

1565 lsrD Autoinducer-2 ABC transporter 
membrane subunit LsrD 



Supplementary Material 

 225 

cds-NP_414576.4 604.3057611172
95 -1.11005223017894 1.130147945368

51e-05 
5.987350777556

35e-05 caiF DNA-binding transcriptional 
activator CaiF 

cds-NP_416129.1 6933.899450041
84 -1.11109647249843 3.208262859643

19e-15 
6.897765148232

86e-14 fumA fumarase A 

cds-NP_416285.1 55.75130898929
95 -1.11150568662618 2.362597229605

99e-05 
0.000116008238

690964 ydjG NADH-dependent methylglyoxal 
reductase 

cds-NP_415326.1 183.3177478553
15 -1.11232794896262 6.119413285536

95e-05 
0.000276560014

710644 fiu iron catecholate outer membrane 
transporter Fiu 

cds-NP_417762.1 55161.10536528
58 -1.1141713167163 1.235233507675

48e-33 
1.334353464754

8e-31 rpsE 30S ribosomal subunit protein S5 

cds-NP_416128.1 1136.858836764
4 -1.11586020636087 1.990895058659

71e-07 
1.450275364276

95e-06 fumC fumarase C 

cds-NP_415535.1 3090.752710727
34 -1.11880730712113 1.593602145476

42e-15 
3.676074949122

43e-14 putP proline:Na(+) symporter 

cds-NP_417773.1 40911.91302582
74 -1.12364976069596 6.560451969546

58e-14 
1.205653185606

71e-12 rpsC 30S ribosomal subunit protein S3 

cds-NP_417764.1 45089.01910752
49 -1.12573418254509 3.819981620247

22e-27 
2.488043911187

49e-25 rplF 50S ribosomal subunit protein L6 

cds-NP_417539.1 7687.733417197
25 -1.12995973412731 4.794075717824

13e-40 
1.117524281802

27e-37 rpoD RNA polymerase sigma factor 
RpoD 

cds-NP_416413.1 86.63409335729
54 -1.13543626736427 8.862243008662

29e-06 
4.775045533499

42e-05 araG arabinose ABC transporter ATP 
binding subunit 

cds-NP_415924.1 1323.162903397
06 -1.13693729716796 1.206077062244

16e-09 
1.332098580718

05e-08 pdxI pyridoxal reductase 

cds-NP_418621.5 14379.62872614
21 -1.138099427429 5.409620481050

29e-12 
8.121765800193

81e-11 rpsF 30S ribosomal subunit protein S6 

cds-YP_009518815.1 111.4622372189
7 -1.13947024001288 1.205068954793

64e-05 
6.346314388562

46e-05 yghO putative DNA-binding 
transcriptional regulator YghO 

cds-NP_416338.1 2871.365985262
08 -1.14101964029865 1.814754100038

03e-11 
2.576136509316

8e-10 yobF DUF2527 domain-containing 
protein YobF 

cds-NP_417003.1 15925.24726083
58 -1.16142199899032 6.176482061948

64e-26 
3.799394312829

24e-24 guaB inosine 5'-monophosphate 
dehydrogenase 

cds-NP_417038.4 96.72280690700
31 -1.16255214243881 1.484851602796

22e-07 
1.109006365730

94e-06 yphA putative inner membrane protein 

cds-NP_417771.1 57144.89870988
18 -1.16767134928437 8.155259018633

15e-19 
2.579974442394

73e-17 rpmC 50S ribosomal subunit protein L29 

cds-NP_415263.1 1619.863008047
01 -1.17133076531888 3.404845027925

24e-25 
1.839031540083

03e-23 ybgE PF09600 family protein YbgE 

cds-NP_418541.2 912.2971050461
17 -1.17454839507786 1.409931290884

94e-08 
1.309137460656

06e-07 adiA arginine decarboxylase 
degradative 

cds-NP_415984.1 67.65269911607
06 -1.1792856285124 8.605499574449

64e-07 
5.731392122592

1e-06 narY nitrate reductase Z subunit beta 

cds-NP_415915.1 208.1410929115
8 -1.18263775855217 1.848039444849

66e-11 
2.606677293388

26e-10 paaJ beta-ketoadipyl-CoA thiolase 

cds-NP_416191.1 32424.94133049
93 -1.18627711251832 1.544830939456

64e-25 
9.122741641137

95e-24 pykF pyruvate kinase 1 

cds-NP_415262.1 77375.89612436
68 -1.18687428943553 1.124067177859

15e-18 
3.505981359674

77e-17 cydB cytochrome bd-I subunit 2 

cds-NP_417985.1 13427.96758268
61 -1.1884171237923 6.688190129157

82e-08 
5.395627337347

91e-07 dctA C4 dicarboxylate/orotate:H(+) 
symporter 

cds-NP_415894.4 289.5044262935
15 -1.1899742734645 2.040812966280

44e-12 
3.228128795591

46e-11 uspF universal stress protein F 

cds-NP_417772.1 39157.55136943
39 -1.196858688908 5.720789579651

43e-24 
2.815264116475

13e-22 rplP 50S ribosomal subunit protein L16 

cds-NP_417466.1 540.8611092762
32 -1.19879792928746 1.644325733641

93e-15 
3.773429364922

33e-14 hybE hydrogenase 2-specific chaperone 

cds-NP_418624.1 14617.62169489
83 -1.20414201560569 5.066613120893

92e-20 
1.780954723209

46e-18 rplI 50S ribosomal subunit protein L9 

cds-NP_415018.1 192.6334505764
87 -1.20479227063682 6.421941401644

7e-08 
5.209300085693

11e-07 glsA glutaminase 1 



Supplementary Material 

  226 

cds-NP_417030.2 937.3342762888
82 -1.20839886257399 6.262437065013

39e-10 
7.130162920551

23e-09 csiE stationary phase-inducible protein 
CsiE 

cds-NP_418711.1 2498.566846138
09 -1.20860161160257 1.289277909939

75e-07 
9.794531497638

32e-07 fecA ferric citrate outer membrane 
transporter 

cds-NP_415916.1 183.3848260917
63 -1.20987341103537 1.134168732016

47e-10 
1.454926600211

3e-09 paaK phenylacetate-CoA ligase 

cds-NP_414713.1 4086.571032119
73 -1.21179090865633 1.537823929575

46e-30 
1.335494545899

95e-28 pyrH UMP kinase 

cds-NP_417345.1 3403.658472650
74 -1.21297093185742 9.935224645112

85e-09 
9.483428869225

18e-08 ygeV putative sigma(54)-dependent 
transcriptional regulator YgeV 

cds-NP_415160.1 298.7478976414
57 -1.21397753326828 1.744254301480

38e-17 
4.682001394700

98e-16 tatE twin arginine protein translocation 
system - TatE protein 

cds-NP_415707.1 9758.229291316
66 -1.21548504638249 3.629735283191

98e-10 
4.284976935547

54e-09 dadA D-amino acid dehydrogenase 

cds-NP_415538.1 179.4342590300
61 -1.21826325022853 9.879232734822

36e-09 
9.450350277003

93e-08 efeB heme-containing 
peroxidase/deferrochelatase 

cds-NP_415019.1 283.4075056902
9 -1.21878511785182 8.008133843618

65e-11 
1.046254418683

98e-09 ybaT putative transporter YbaT 

cds-NP_415580.1 666.8448517005
78 -1.24493485362969 7.546141916695

47e-19 
2.404450543096

71e-17 pyrC dihydroorotase 

cds-NP_414916.1 30.19636187530
97 -1.25851904131057 0.001549318587

29844 
0.005049250936

82473 iraP anti-adaptor protein IraP 

cds-NP_418622.1 14785.38934222
78 -1.26710776437698 4.132958321209

94e-19 
1.345946500341

09e-17 priB primosomal replication protein N 

cds-NP_415127.1 96.89705054871
56 -1.26797192246828 0.000622388796

180176 
0.002237467514

83929 entB enterobactin synthase component 
B 

cds-NP_415333.1 2819.504070698
06 -1.26942275843245 1.926988742672

16e-20 
6.995600935487

71e-19 dps DNA protection during starvation 
protein 

cds-NP_417002.1 23485.00623114
03 -1.27130734089282 4.842425549090

98e-34 
5.499257117159

99e-32 guaA GMP synthetase 

cds-NP_415830.2 24.05780930655
88 -1.27416024283264 0.001328262865

40841 
0.004406648862

09276 ycjR 3-dehydro-D-guloside 4-
epimerase 

cds-NP_418709.1 243.5409628728
09 -1.28404621198547 1.419959826864

e-05 
7.346965038762

45e-05 fecC ferric citrate ABC transporter 
membrane subunit FecC 

cds-NP_415375.1 97.48999031012
12 -1.2896410496931 1.450215813355

32e-09 
1.582021142204

61e-08 potF putrescine ABC transporter 
periplasmic binding protein 

cds-NP_416813.1 929.3559354179
9 -1.30096337428862 7.313677511949

83e-11 
9.669336626992

78e-10 argT 
lysine/arginine/ornithine ABC 

transporter periplasmic binding 
protein 

cds-NP_417454.1 4503.955395193
06 -1.30542777086074 2.201669275029

99e-09 
2.316197914277

4e-08 glcC DNA-binding transcriptional dual 
regulator GlcC 

cds-NP_415757.1 174154.2265167
24 -1.31591546468816 1.251805038369

08e-17 
3.465152821835

41e-16 adhE 

fused acetaldehyde-CoA 
dehydrogenase and iron-

dependent alcohol 
dehydrogenasealdehyde/alcohol 

dehydrogenase AdhE 

cds-NP_414984.1 141.0976264586
26 -1.31661719056753 9.786412123610

27e-06 
5.253820520663

01e-05 glnK nitrogen regulatory protein PII-2 

cds-NP_415377.1 25.22390215902
87 -1.31816462637766 6.831199816244

4e-05 
0.000305609939

254005 potH putrescine ABC transporter 
membrane subunit PotH 

cds-NP_415979.2 89.56066582432
92 -1.32336287507204 1.574643656642

02e-06 
9.934610762489

3e-06 yddH flavin reductase-like protein YddH 

cds-NP_415813.4 168.8135520493
78 -1.32551438002469 2.510997018795

83e-10 
3.055276317650

2e-09 puuA glutamate-putrescine ligase 

cds-NP_416843.1 1054.163081251
87 -1.32560634618478 1.970552516252

07e-08 
1.766716011028

42e-07 fadJ 3-hydroxyacyl-CoA 
dehydrogenase FadJ 

cds-NP_417780.1 11834.53862106
42 -1.32936569954894 2.099248036814

6e-15 
4.695742199521

14e-14 rpsJ 30S ribosomal subunit protein S10 

cds-NP_415902.1 274.2842712606
93 -1.33565028046341 8.317540080241

4e-09 
8.008344568562

86e-08 feaR DNA-binding transcriptional 
activator FeaR 



Supplementary Material 

 227 

cds-NP_414851.1 27.85630913240
53 -1.33607677934968 0.000434836368

271337 
0.001622485488

68892 yahC uncharacterized protein YahC 

cds-NP_415708.1 2699.784343284
16 -1.33779316940357 7.092964112440

2e-11 
9.405610195807

68e-10 dadX alanine racemase 2 

cds-NP_416656.1 3310.632448362
97 -1.34074304266892 1.506639863601

27e-05 
7.768228120943

e-05 galS DNA-binding transcriptional dual 
regulator GalS 

cds-NP_417910.1 259.6815993429
53 -1.34886429567586 9.892113089358

98e-08 
7.659470082652

26e-07 ugpB 
sn-glycerol 3-phosphate ABC 

transporter periplasmic binding 
protein 

cds-YP_026181.1 6487.289412203
97 -1.35670756955325 8.053913970850

08e-05 
0.000352478112

419911 srlE sorbitol-specific PTS enzyme 
IIBC1 component 

cds-NP_417774.1 24570.38832739
97 -1.35723339204484 9.122460580187

46e-30 
7.623278850877

41e-28 rplV 50S ribosomal subunit protein L22 

cds-NP_415376.4 36.52850564761
92 -1.35813275729649 1.551795974625

96e-05 
7.978586395369

56e-05 potG putrescine ABC transporter ATP 
binding subunit 

cds-NP_416655.1 4110.755856113
97 -1.36866930086832 2.549046939634

35e-09 
2.662671909349

18e-08 mglB 
D-galactose/methyl-galactoside 

ABC transporter periplasmic 
binding protein 

cds-NP_417775.1 14135.83483927
52 -1.36894057675729 1.744000521731

75e-19 
5.941675623653

78e-18 rpsS 30S ribosomal subunit protein S19 

cds-NP_416005.1 11.81890553997
58 -1.37264890926847 0.011471152039

2713 
0.029092394365

4133 ddpX D-alanyl-D-alanine dipeptidase 

cds-NP_415541.1 120.4350393545
01 -1.37965354741997 2.083974032652

6e-10 
2.571008632484

22e-09 pgaC poly-N-acetyl-D-glucosamine 
synthase subunit PgaC 

cds-NP_415537.1 134.5325020600
05 -1.38129866846422 7.809995547408

84e-09 
7.562293290725

37e-08 efeO ferrous iron transport system 
protein EfeO 

cds-NP_416052.1 105.8830039422
92 -1.38641733366817 6.441217321554

93e-08 
5.215384189610

01e-07 dgcZ diguanylate cyclase DgcZ 

cds-YP_588444.1 953.9978810912
12 -1.39365911655369 2.654607581755

85e-27 
1.808808766091

79e-25 cydX cytochrome bd-I accessory 
subunit CydX 

cds-NP_416294.4 4099.762591472
93 -1.39538952194908 2.573595886740

05e-19 
8.570267806294

5e-18 yeaD putative aldose 1-epimerase YeaD 

cds-NP_416959.1 382.2317239732
55 -1.3977287057878 5.887876980077

57e-16 
1.402011136815

25e-14 talA transaldolase A 

cds-NP_418546.1 474.5779920442
9 -1.39806363446056 1.029086451408

95e-31 
9.908312811500

48e-30 fumB fumarase B 

cds-NP_415772.1 198.9566705540
61 -1.39854142629035 9.106362627109

87e-12 
1.326713160377

29e-10 ompW outer membrane protein W 

cds-NP_414985.1 1226.598104308
66 -1.4014033275332 4.566327443384

06e-09 
4.565296669694

8e-08 amtB ammonium transporter 

cds-NP_417776.1 51031.72307326
07 -1.40286432780516 1.879949826548

04e-14 
3.667972591093

06e-13 rplB 50S ribosomal subunit protein L2 

cds-NP_418630.1 281.6870948308
53 -1.41117218551463 2.928494231693

79e-15 
6.326976074230

14e-14 ytfE iron-sulfur cluster repair protein 
YtfE 

cds-NP_414913.1 65.95751657641
42 -1.41139954296567 7.172829717190

54e-07 
4.820707559550

36e-06 yaiY DUF2755 domain-containing inner 
membrane protein YaiY 

cds-NP_418688.1 96.20631949386
35 -1.41281495590082 2.177843344952

17e-08 
1.929133634958

63e-07 idnD L-idonate 5-dehydrogenase 

cds-NP_415524.1 255.0805192065
32 -1.4135602389629 1.527546904663

97e-17 
4.125308073632

16e-16 wrbA NAD(P)H:quinone oxidoreductase 

cds-NP_416262.1 373.0213633040
12 -1.42018267973725 2.185799839104

86e-15 
4.840453743697

71e-14 astC succinylornithine transaminase 

cds-NP_416853.1 777.7586469679
42 -1.42165831251508 2.267927095378

94e-19 
7.667671072849

88e-18 yfdI serotype-specific glucosyl 
transferase YfdI 

cds-NP_416960.1 920.2591749111
42 -1.4298295046359 1.452498560736

23e-23 
6.843740559043

36e-22 tktB transketolase 2 

cds-NP_415156.1 3991.736497560
06 -1.43101276799944 1.585802870822

33e-10 
1.984045456178

56e-09 cspE transcription antiterminator and 
regulator of RNA stability CspE 

cds-NP_415978.1 23.45216553411
62 -1.4315286100784 9.630672379174

04e-05 
0.000414118912

304484 pptA tautomerase PptA 



Supplementary Material 

  228 

cds-NP_415997.1 66.22036099466
53 -1.43797995013853 9.942151581264

43e-07 
6.533203168163

23e-06 sra ribosome-associated protein Sra 

cds-NP_417269.1 66.79334860565
83 -1.43879447836546 2.410335647837

65e-05 
0.000118221224

632037 gudP galactarate/D-glucarate 
transporter GudP 

cds-NP_416484.2 1680.865883606
65 -1.43996497300794 1.736462775792

99e-09 
1.857679621736

02e-08 mtfA Mlc titration factor 

cds-NP_415130.1 8084.358535077
8 -1.45611977053065 5.285747067814

83e-06 
2.989856163901

9e-05 cstA pyruvate transporter CstA 

cds-NP_417147.1 228.3863598956
38 -1.45638643731633 2.971138981035

78e-21 
1.144276047565

87e-19 gabD succinate-semialdehyde 
dehydrogenase (NADP(+)) GabD 

cds-NP_416217.1 719.2297168033
14 -1.47367109489204 1.523132357650

46e-11 
2.176113939365

76e-10 ppsA phosphoenolpyruvate synthetase 

cds-NP_415986.1 25.10866010688
66 -1.48875555227674 0.000195600670

923755 
0.000783995811

331502 narU nitrate/nitrite transporter NarU 

cds-NP_417778.1 37471.06936255
64 -1.49442869616763 4.348662612587

01e-21 
1.646173223175

03e-19 rplD 50S ribosomal subunit protein L4 

cds-NP_415540.1 45.10788943225
01 -1.49529869229857 1.697120244716

14e-07 
1.252757593974

63e-06 pgaD poly-N-acetyl-D-glucosamine 
synthase subunit PgaD 

cds-NP_414918.4 43.53544829229
31 -1.4982838683664 4.426049438945

3e-05 
0.000207658611

918313 psiF phosphate starvation-inducible 
protein PsiF 

cds-NP_417279.2 1159.857676732
36 -1.50131340501496 3.333160727688

83e-13 
5.721925915865

82e-12 fucO L-1 2-propanediol oxidoreductase 

cds-NP_415828.1 10.13023396646
29 -1.50287726545317 0.010368083730

7299 
0.026775651803

7333 ycjP putative ABC transporter 
membrane subunit YcjP 

cds-NP_415959.1 27.20740760740
21 -1.5073749519075 1.415055350212

98e-05 
7.330152217652

97e-05 ydcU putative ABC transporter 
membrane subunit YdcU 

cds-NP_418122.1 27.53761613275
07 -1.50997901242599 0.009141397474

49762 
0.023886282840

4424 uhpT hexose-6-phosphate:phosphate 
antiporter 

cds-NP_417148.1 324.1012712985
72 -1.51587620815309 1.583237249096

82e-35 
2.261986379435

43e-33 gabT 4-aminobutyrate aminotransferase 
GabT 

cds-NP_417779.1 52347.66775977
84 -1.52038125849136 6.940421802857

19e-36 
1.070964887125

25e-33 rplC 50S ribosomal subunit protein L3 

cds-NP_416639.2 43.84577408936
07 -1.52397915870105 6.580775901708

68e-08 
5.318659939537

91e-07 yohC putative inner membrane protein 

cds-NP_416414.1 81.10592300982
62 -1.52568892978142 3.337415146253

83e-07 
2.342537509153

44e-06 araF arabinose ABC transporter 
periplasmic binding protein 

cds-NP_418024.1 27.48973014421
93 -1.53483915619741 7.955290177607

18e-05 
0.000349197028

707851 xylG xylose ABC transporter ATP 
binding subunit 

cds-NP_418459.1 36770.61368870
36 -1.5366812802466 6.400825989694

4e-12 
9.481357293764

71e-11 malK maltose ABC transporter ATP 
binding subunit 

cds-NP_417400.1 56193.92104051
45 -1.54274385332985 9.629324539499

04e-43 
2.843218559029

42e-40 fbaA fructose-bisphosphate aldolase 
class II 

cds-NP_417777.1 37003.97248350
76 -1.54328690220625 8.790290811919

62e-31 
7.786439601198

4e-29 rplW 50S ribosomal subunit protein L23 

cds-NP_418651.3 97.51596354806
86 -1.54619059452117 1.020334970809

75e-09 
1.135443111989

04e-08 ytfT galactofuranose ABC transporter 
putative membrane subunit YtfT 

cds-NP_415123.1 77.19739867623
04 -1.5472561862653 1.518627100725

81e-07 
1.128259407389

52e-06 entS enterobactin exporter EntS 

cds-NP_418418.2 13520.81366144
85 -1.54833432721519 2.836480650152

72e-25 
1.570346599940

8e-23 thiG 1-deoxy-D-xylulose 5-
phosphate:thiol sulfurtransferase 

cds-NP_415423.1 58905.98131048
88 -1.54866933398299 3.436259304761

05e-27 
2.305938251634

35e-25 pflB pyruvate formate-lyase 

cds-NP_415960.1 29.94645991301
19 -1.55292013038087 1.905130889867

58e-05 
9.588437171844

89e-05 ydcV putative ABC transporter 
membrane subunit YdcV 

cds-NP_415116.1 164.0030484256
81 -1.55408586964459 2.768687131436

85e-10 
3.359593234283

24e-09 fepA ferric enterobactin outer 
membrane transporter 

cds-NP_415392.1 408.6096727860
41 -1.55515196179851 6.733616380819

35e-25 
3.593155054295

05e-23 poxB pyruvate oxidase 

cds-NP_418690.4 535.5232606703
39 -1.56082001265435 1.735219265708

69e-23 
8.005506383149

77e-22 ahr NADPH-dependent aldehyde 
reductase Ahr 



Supplementary Material 

 229 

cds-NP_418648.1 326.7869946677
71 -1.57386626524827 1.897368264495

97e-16 
4.721035979467

79e-15 ytfQ galactofuranose ABC transporter 
periplasmic binding protein 

cds-NP_418420.4 11816.85395489
03 -1.57522243010381 3.952624040148

5e-24 
2.012203663657

21e-22 thiF sulfur carrier protein ThiS 
adenylyltransferase 

cds-NP_416874.1 46.23889790190
51 -1.58162374689593 1.286835010593

42e-06 
8.308151985303

6e-06 oxc oxalyl-CoA decarboxylase 

cds-NP_416589.2 16.39176500036
06 -1.58316568886659 0.001282909163

73646 
0.004268974219

52577 yegR uncharacterized protein YegR 

cds-NP_416437.1 322.5467629267
07 -1.58326845344897 1.099697073734

57e-25 
6.581835594014

06e-24 amyA alpha-amylase 

cds-NP_416199.1 32.40192831087
39 -1.5920140715731 0.000150266637

650288 
0.000612263972

541974 sufA iron-sulfur cluster insertion protein 
SufA 

cds-NP_418710.4 1434.246756293
3 -1.59621110040304 6.270055987822

49e-14 
1.157086582086

08e-12 fecB ferric citrate ABC transporter 
periplasmic binding protein 

cds-NP_418422.1 55115.07136252
65 -1.59877131268573 1.573875179658

06e-25 
9.171964698296

76e-24 thiC phosphomethylpyrimidine 
synthase 

cds-NP_416031.1 62.90744090621
13 -1.60186008559673 1.420036960133

15e-10 
1.796955341837

06e-09 lsrC Autoinducer-2 ABC transporter 
membrane subunit LsrC 

cds-NP_415947.1 1208.764730867
06 -1.60335174123035 2.855117234549

38e-18 
8.602254579468

84e-17 tehB tellurite methyltransferase 

cds-YP_026286.1 217.0143679535
65 -1.61341176451071 1.263371645645

83e-18 
3.885745151781

5e-17 ytfR galactofuranose ABC transporter 
putative ATP binding subunit 

cds-YP_026272.1 256.4157188948
6 -1.61661571948358 3.620898615849

68e-08 
3.084030763384

27e-07 fadA 3-ketoacyl-CoA thiolase 

cds-NP_416415.1 872.9462849174
84 -1.61861591807057 1.733357433552

33e-06 
1.085861396492

68e-05 ftnB putative ferritin-like protein 

cds-NP_414863.1 111.8423928195
96 -1.63294094025225 8.562030974536

06e-15 
1.739506201202

76e-13 yahO DUF1471 domain-containing 
protein YahO 

cds-NP_416337.1 8474.210851934
55 -1.63502632604007 1.099095262859

83e-22 
4.726109630297

26e-21 cspC transcription antiterminator and 
regulator of mRNA stability CspC 

cds-NP_415050.1 36.61474211646
82 -1.6421037958099 2.224667152203

68e-07 
1.612610608364

99e-06 allD ureidoglycolate dehydrogenase 

cds-NP_415706.1 301.9259522365
96 -1.64470754921051 1.685385136534

14e-26 
1.066367252815

67e-24 ycgB PF04293 family protein YcgB 

cds-YP_026279.1 2634.248765980
45 -1.65486824940241 5.395112743744

97e-28 
3.854024893878

47e-26 thiS sulfur carrier protein ThiS 

cds-NP_416259.1 187.6958473081
95 -1.65551236638015 8.317410766745

22e-12 
1.215769382373

42e-10 astB N-succinylarginine dihydrolase 

cds-NP_418417.1 16453.22227020
08 -1.65766535121605 5.853159775296

29e-33 
6.028754568555

18e-31 thiH 2-iminoacetate synthase 

cds-NP_416844.1 1948.121368842
86 -1.68781886758267 1.038268514560

02e-05 
5.540350904802

79e-05 fadI 3-ketoacyl-CoA thiolase FadI 

cds-NP_414864.1 77.21292756987
87 -1.71032674839068 1.221701948356

99e-15 
2.847851541722

69e-14 prpR DNA-binding transcriptional dual 
regulator PrpR 

cds-NP_418421.1 12476.22008345
07 -1.7118427983611 1.780300392137

77e-29 
1.408026863710

39e-27 thiE thiamine phosphate synthase 

cds-NP_416007.3 27.68545759393
92 -1.7211553216194 3.977964668853

56e-05 
0.000187829483

138085 dosC diguanylate cyclase DosC 

cds-NP_414838.2 38.77872578528
53 -1.72756210709832 5.843922473448

68e-08 
4.784238934363

07e-07 rclA cupric reductase RclA 

cds-NP_416298.1 366.9796454765
81 -1.73448599194035 9.241738497715

51e-19 
2.902954596197

3e-17 yeaH DUF444 domain-containing 
protein YeaH 

cds-NP_416590.1 77.93873644515
75 -1.7367240127676 9.529156269454

27e-13 
1.540315077277

85e-11 yegS lipid kinase YegS 

cds-NP_416289.1 33.57857141025
41 -1.741918021012 1.424111776565

4e-07 
1.065437678785

16e-06 ydjK putative transporter YdjK 

cds-NP_418491.1 111.8723506690
21 -1.74854875887915 1.657921669465

08e-11 
2.361072371080

65e-10 actP acetate/glycolate:cation symporter 

cds-NP_415933.1 14740.81201545
98 -1.75216568033436 8.593265244667

23e-08 
6.724305966189

25e-07 aldA aldehyde dehydrogenase A 



Supplementary Material 

  230 

cds-NP_416654.1 1388.374083929
16 -1.79932546540117 1.310711876843

61e-05 
6.837624149046

33e-05 mglA 
D-galactose/methyl-galactoside 
ABC transporter ATP binding 

subunit 

cds-NP_418011.1 140.7192179270
07 -1.81194205666485 2.173331296337

86e-18 
6.592934459918

08e-17 yiaG putative DNA-binding 
transcriptional regulator YiaG 

cds-NP_416297.1 1072.966589731
6 -1.81969043179445 5.934537473510

78e-34 
6.571016617544

81e-32 yeaG protein kinase YeaG 

cds-NP_417904.1 254.1344713643
81 -1.82091114779697 5.563370956951

45e-41 
1.368898331574

33e-38 ggt glutathione hydrolase proenzyme 

cds-NP_417158.4 125.2957507662
29 -1.82854242606367 9.311989221211

57e-18 
2.660825823273

94e-16 ygaM DUF883 domain-containing 
protein YgaM 

cds-YP_009518758.1 8.945391040897
38 -1.83701970939545 0.001693597333

79477 
0.005475140577

64747 ybfQ inactive transposase YbfQ 

cds-NP_417352.2 1987.218558287
23 -1.84875721916633 2.074990906134

39e-05 
0.000103144048

52154 yqeC uncharacterized protein YqeC 

cds-NP_418663.1 6541.461037171
63 -1.86104199081314 6.345803151892

44e-35 
8.266341811685

76e-33 mgtA Mg(2(+)) importing P-type ATPase 

cds-NP_417146.2 93.92346483914
8 -1.86724772034827 1.070717854444

52e-17 
3.039877805983

83e-16 lhgD L-2-hydroxyglutarate 
dehydrogenase 

cds-NP_416030.1 187.4450760427
89 -1.87478318300287 3.773025516288

27e-12 
5.742518904343

89e-11 lsrA Autoinducer-2 ABC transporter 
ATP binding subunit 

cds-NP_417280.1 101.6026107176
73 -1.88554514905175 5.992509470247

18e-14 
1.110494746599

36e-12 fucA L-fuculose-phosphate aldolase 

cds-NP_416873.1 15.17828521221
03 -1.89483107005971 0.000248582400

554724 
0.000970874296

346449 yfdV putative transport protein YfdV 

cds-NP_416314.1 526.8863972574
59 -1.90396379419673 8.668168455909

83e-24 
4.172969357741

81e-22 dmlA D-malate/3-isopropylmalate 
dehydrogenase (decarboxylating) 

cds-NP_417963.4 427.4970093562
68 -1.91585133320556 1.681316800410

17e-15 
3.818744671290

6e-14 slp starvation lipoprotein 

cds-NP_415940.1 330.9489127904
68 -1.92130312838202 6.659748107127

82e-17 
1.724913705641

47e-15 ydcJ DUF1338 domain-containing 
protein YdcJ 

cds-NP_415126.1 95.07837122426
74 -1.92443243697237 4.367489803489

5e-08 
3.684497588505

71e-07 entE 2 3-dihydroxybenzoate-AMP 
ligase 

cds-YP_002791249.1 9.128037646036
4 -1.97423535838039 0.006046257765

9808 
0.016695059629

3821 yohP putative membrane protein YohP 

cds-NP_416261.1 208.2128169504
08 -1.98292808513435 2.243579975733

38e-22 
9.463634011926

79e-21 astA arginine N-succinyltransferase 

cds-NP_418028.1 2005.264886299
17 -1.98881507210275 1.728354931048

69e-25 
9.941407778720

3e-24 malS alpha-amylase 

cds-NP_415417.1 50.16134356437
95 -1.99437889174653 3.571814780777

23e-13 
6.084449101562

44e-12 ycaC putative hydrolase YcaC 

cds-NP_417964.1 48.92011024338
44 -1.99484320948663 7.078468096957

37e-10 
7.977235420209

72e-09 dctR putative DNA-binding 
transcriptional regulator DctR 

cds-NP_414937.2 1423.865739964
02 -1.99615580967704 9.256866457838

02e-60 
5.856951648823

51e-57 malZ maltodextrin glucosidase 

cds-NP_418539.1 2082.538460230
01 -2.0149230133895 1.504993783205

89e-23 
7.016439437704

08e-22 adiC arginine:agmatine antiporter 

cds-NP_416260.1 256.4657332673
36 -2.01951742500989 1.765747675431

58e-22 
7.519708129313

92e-21 astD aldehyde dehydrogenase 

cds-NP_417971.1 1267.498407699
92 -2.02808526466951 1.580591827438

35e-35 
2.261986379435

43e-33 mdtF multidrug efflux pump RND 
permease MdtF 

cds-NP_416600.4 471.1007428743
54 -2.03032132368186 2.115015518501

05e-35 
2.927313666075

36e-33 fbaB fructose-bisphosphate aldolase 
class I 

cds-NP_417970.1 232.2298346067
79 -2.04057096819338 1.429719140642

63e-17 
3.884801272335

11e-16 mdtE multidrug efflux pump membrane 
fusion protein MdtE 

cds-NP_418045.4 150.6307555668
12 -2.05793117924609 3.602667460756

09e-27 
2.381524505028

17e-25 aldB aldehyde dehydrogenase B 

cds-NP_416769.1 859.2862178381
8 -2.0678780833484 3.360691559914

61e-22 
1.404198388571

87e-20 elaB tail-anchored inner membrane 
protein ElaB 



Supplementary Material 

 231 

cds-NP_417351.1 2037.676974814
92 -2.07100989217866 5.313986835030

46e-09 
5.218547160166

27e-08 yqeB XdhC-CoxI family protein YqeB 

cds-NP_415906.1 12.40186234853
46 -2.07228044566435 6.960885271101

2e-05 
0.000311003860

932382 paaA phenylacetyl-CoA 12-epoxidase 
monooxygenase subunit 

cds-NP_415671.1 5.052920625930
23 -2.07843570606774 0.018174101262

6594 
0.043021429445

3867 ymfQ DUF2313 domain-containing 
protein YmfQ 

cds-NP_415539.1 568.2235972416
96 -2.07860138821017 8.507189891014

62e-09 
8.173176578590

83e-08 phoH ATP-binding protein PhoH 

cds-NP_415125.1 70.23112095305
18 -2.11411978135396 3.665383381531

28e-07 
2.556532755401

9e-06 entC isochorismate synthase EntC 

cds-NP_417184.1 922.2386446009
12 -2.12724894332225 1.943574788928

68e-21 
7.685797089433

13e-20 srlB sorbitol-specific PTS enzyme IIA 
component 

cds-NP_414880.2 2339.413806853
21 -2.13896213774937 2.802311036270

62e-06 
1.668203706941

21e-05 mhpR DNA-binding transcriptional 
activator MhpR 

cds-NP_418493.1 1356.873904942
62 -2.24034340092758 5.832030683618

41e-11 
7.803644682098

48e-10 acs acetyl-CoA synthetase (AMP-
forming) 

cds-NP_416258.1 115.8565405362
35 -2.26052684385935 6.865923671130

75e-14 
1.256577518158

6e-12 astE succinylglutamate desuccinylase 

cds-YP_025308.1 181.4977481902
87 -2.27661630726541 6.197539879681

07e-33 
6.238387301615

33e-31 katE catalase HPII 

cds-NP_417552.1 1410.582274923
43 -2.2921686188879 1.351365988875

82e-06 
8.661649731882

76e-06 fadH 2 4-dienoyl-CoA reductase 

cds-NP_415958.1 60.50741919651
67 -2.29347264375567 2.949478904570

22e-14 
5.679670464496

31e-13 ydcT putative ABC transporter ATP-
binding protein YdcT 

cds-NP_418492.1 33.32285817848
2 -2.298104813442 2.572225486936

13e-07 
1.837481722845

18e-06 yjcH DUF485 domain-containing inner 
membrane protein YjcH 

cds-NP_416319.1 2826.644070602
07 -2.32267036799557 3.262778219465

62e-10 
3.916218085098

44e-09 fadD long-chain-fatty-acid--CoA ligase 

cds-NP_415534.1 5654.354622804
36 -2.33377071419633 1.692251623579

51e-24 
8.817626400980

74e-23 putA 

fused DNA-binding transcriptional 
repressor/proline 

dehydrogenase/1-pyrroline-5-
carboxylate dehydrogenase PutA 

cds-NP_418457.1 10679.51232957
53 -2.34737747176128 2.711752954030

97e-29 
2.107079619895

29e-27 malF maltose ABC transporter 
membrane subunit MalF 

cds-NP_418458.1 24975.49549012
95 -2.36001636669004 5.360793343648

04e-43 
1.695925265644

08e-40 malE maltose ABC transporter 
periplasmic binding protein 

cds-NP_418023.1 42.48516130789
52 -2.38124874834165 1.498983137384

32e-10 
1.886078498714

53e-09 xylF xylose ABC transporter 
periplasmic binding protein 

cds-NP_418503.1 930.1156076512
03 -2.38433563024141 1.735362434645

87e-38 
3.202466759602

74e-36 fdhF formate dehydrogenase H 

cds-NP_417965.1 116.9775474242
67 -2.3956092060003 5.986399471854

34e-13 
9.930248412300

7e-12 yhiD inner membrane protein YhiD 

cds-NP_415957.1 150.2590142156
61 -2.44583286115698 2.043949102707

8e-30 
1.740894341517

86e-28 ydcS 

putative ABC transporter 
periplasmic binding 

protein/polyhydroxybutyrate 
synthase 

cds-NP_417597.1 1035.388026483
83 -2.45176071929932 5.191523200284

23e-13 
8.709566762901

08e-12 garD GarD 

cds-YP_026218.1 30845.38151732
85 -2.45597465116317 3.662424408626

1e-53 
1.802319745089

44e-50 malP maltodextrin phosphorylase 

cds-NP_417047.1 3030.237647276
76 -2.47758844736536 7.181900501415

21e-39 
1.445847150944

e-36 hmp nitric oxide dioxygenase 

cds-NP_417595.1 109.0990447232
26 -2.48129581421372 7.771196095517

7e-14 
1.410599487993

77e-12 garL alpha-dehydro-beta-deoxy-D-
glucarate aldolase 

cds-NP_415556.1 10.13639077664
66 -2.51004680750125 0.000212972134

767646 
0.000848249626

695959 csgF curli assembly component CsgF 

cds-NP_418456.1 1959.601346768
86 -2.51459630363246 1.364854231454

09e-38 
2.628234517873

98e-36 malG maltose ABC transporter 
membrane subunit MalG 

cds-NP_415500.1 474.6751741493
03 -2.51798872392528 6.955568489655

58e-35 
8.801775097338

44e-33 appA 
periplasmic phosphoanhydride 
phosphatase/multiple inositol-
polyphosphate phosphatase 



Supplementary Material 

  232 

cds-NP_418288.1 1093.773214404
69 -2.53040317661987 2.631400642973

52e-06 
1.577060006458

69e-05 fadB 

multifunctional enoyl-CoA 
hydratase 3-hydroxyacyl-CoA 

epimerase Delta(3)-cis- Delta(2)-
trans-enoyl-CoA isomerase L-3-

hydroxyacyl-CoA dehydrogenase 

cds-NP_417185.1 10286.95795060
23 -2.53567346611233 3.044435971120

37e-28 
2.210460150179

03e-26 srlD sorbitol-6-phosphate 2-
dehydrogenase 

cds-NP_417198.1 149.5084824214
43 -2.5367802068753 1.095032156327

18e-23 
5.214943462766

76e-22 hycH formate hydrogenlyase assembly 
protein 

cds-NP_416653.1 642.8599145638
75 -2.58132100789343 1.433201331828

61e-08 
1.327959978801

02e-07 mglC 
D-galactose/methyl-galactoside 

ABC transporter membrane 
subunit 

cds-NP_418461.1 3816.451058259
29 -2.59153843953063 3.209543577631

41e-35 
4.307596516766

51e-33 malM maltose regulon periplasmic 
protein 

cds-NP_417145.4 117.6972761148
02 -2.64832709844364 1.661931658014

63e-21 
6.631257039051

15e-20 glaH glutarate dioxygenase GlaH 

cds-NP_418460.1 27673.88358975
59 -2.68500931297178 1.523668143492

91e-41 
3.969603651488

29e-39 lamB 
maltose outer membrane 

channel/phage lambda receptor 
protein 

cds-NP_418163.1 115.2645123208
45 -2.68709722136575 1.711414711407

25e-34 
1.994698883374

4e-32 tnaC tnaAB operon leader peptide 

cds-NP_415835.1 3.742832444530
45 -2.71556580390073 0.015951628488

134 
0.038727948443

5494 ompG outer membrane porin G 

cds-NP_417875.1 15701.06115001
4 -2.71556986985641 1.401000998681

99e-36 
2.386551316600

97e-34 malQ 4-alpha-glucanotransferase 

cds-YP_001165313.1 9.550565847332
85 -2.76026628278807 0.000219400768

790861 
0.000870722226

679859 appX cytochrome bd-II accessory 
subunit AppX 

cds-NP_416442.1 40.13785569700
21 -2.78466099389415 3.769533567704

21e-13 
6.396652939219

14e-12 yedL putative acetyltransferase YedL 

cds-NP_418164.4 3848.911659106
44 -2.82156303209921 4.116576741991

01e-18 
1.223645529548

87e-16 tnaA tryptophanase 

cds-NP_418165.1 533.4896689973
93 -2.83301821675624 4.893396636981

4e-09 
4.837690559194

34e-08 tnaB tryptophan:H(+) symporter TnaB 

cds-NP_415497.1 1245.690279476
66 -2.89567179063701 4.282925876204

58e-31 
3.951891397022

94e-29 appC cytochrome bd-II subunit 1 

cds-NP_414756.2 1532.269030477
09 -2.90331138665979 8.176083996519

9e-09 
7.889297607970

94e-08 fadE acyl-CoA dehydrogenase 

cds-NP_415491.1 1590.458375451
15 -2.92084251142389 7.012414027236

91e-36 
1.070964887125

25e-33 hyaA hydrogenase 1 small subunit 

cds-NP_417199.1 153.8249749384
79 -3.01866249146698 5.633238709206

35e-27 
3.615886122184

77e-25 hycG formate hydrogenlyase subunit 
HycG 

cds-NP_417596.1 316.6957065844
05 -3.0592753166386 1.475545760596

89e-14 
2.930579450082

34e-13 garP galactarate/D-glucarate 
transporter GarP 

cds-NP_415492.1 2337.181442787
66 -3.07214414601514 4.673582045380

74e-22 
1.899017878806

54e-20 hyaB hydrogenase 1 large subunit 

cds-NP_415496.1 345.9581840863
63 -3.16817350198279 1.358939072091

15e-47 
4.629800884839

76e-45 hyaF protein HyaF 

cds-NP_415498.1 1443.351029298
93 -3.19045199672676 2.343931824036

88e-53 
1.297659256082

42e-50 appB cytochrome bd-II subunit 2 

cds-NP_415494.1 1319.596516223 -3.19419869688116 1.075574299237
79e-20 

4.003124849852
23e-19 hyaD putative hydrogenase 1 maturation 

protease HyaD 

cds-NP_415493.1 1132.650774507
86 -3.25865464822726 8.218175653763

37e-50 
3.033191664209

83e-47 hyaC hydrogenase 1 cytochrome b 
subunit 

cds-NP_417206.1 35.14432160569
08 -3.33093762954418 3.041499409280

02e-10 
3.670517951962

18e-09 hypA hydrogenase 3 nickel 
incorporation protein HypA 

cds-NP_417974.1 574.8032304543
31 -3.34807218898754 3.396481868811

47e-51 
1.504301819696

6e-48 gadA glutamate decarboxylase A 

cds-NP_417201.1 377.4558417983
58 -3.41767029661653 2.814289976116

58e-50 
1.133135482201

85e-47 hycE formate hydrogenlyase subunit 
HycE 

cds-NP_415495.1 193.5608316584
22 -3.55805192524107 3.439650772332

6e-11 
4.716474696799

1e-10 hyaE putative HyaA chaperone 



Supplementary Material 

 233 

cds-NP_417202.1 235.1334728617
98 -3.63230276857347 5.360621017272

18e-38 
9.496876194199

39e-36 hycD formate hydrogenlyase subunit 
HycD 

cds-NP_417200.1 123.7834809112
14 -3.65993369152525 4.196989211517

94e-31 
3.951891397022

94e-29 hycF formate hydrogenlyase subunit 
HycF 

cds-NP_417968.1 3240.549274029
7 -3.71038432270214 1.363941860252

26e-80 
1.208179699811

45e-77 hdeD acid-resistance membrane protein 

cds-NP_416114.2 1637.039358775
68 -3.7594800651994 0.000755723125

314543 
0.002660650017

50247 asr periplasmic chaperone Asr 

cds-NP_417203.1 340.6448002896
95 -3.77327088467965 6.300391470219

81e-39 
1.328782562933

5e-36 hycC formate hydrogenlyase subunit 
HycC 

cds-NP_417205.1 37.84374882569
24 -3.82960241642539 3.471124646152

35e-11 
4.744941684508

87e-10 hycA regulator of the transcriptional 
regulator FhlA 

cds-NP_417966.4 1679.488522695
56 -3.97330696726928 1.681383496427

42e-120 
7.446847505677

03e-117 hdeB periplasmic acid stress chaperone 
HdeB 

cds-NP_417204.1 47.86153764001
89 -4.29904097664917 2.349192266670

5e-15 
5.150778489645

36e-14 hycB formate hydrogenlyase subunit 
HycB 

cds-NP_416009.1 668.1555314682
67 -4.31314324886979 2.201968918822

24e-87 
2.438130085365

93e-84 gadC L-glutamate:4-aminobutyrate 
antiporter 

cds-NP_416010.1 657.0809259000
64 -4.32748479897044 6.509045082386

58e-94 
9.609520223296

73e-91 gadB glutamate decarboxylase B 

cds-NP_417967.1 9328.477129582
23 -4.61412644913442 4.741752811373

59e-102 
1.050061160078

68e-98 hdeA periplasmic acid stress chaperone 
HdeA 

cds-NP_418177.1 18.76269640110
98 1.00798259499972 0.013151378665

9528 
0.032796991053

7753 bglB 6-phospho-beta-glucosidase B 

cds-NP_415504.1 52.86783007341
66 1.00906118732302 0.000447089004

809706 
0.001661205706

62935 gfcD putative lipoprotein GfcD 

cds-NP_418271.1 165.7468525616
77 1.0125064479844 7.389194112880

5e-08 
5.875536934640

53e-07 bioP biotin transporter 

cds-YP_026273.1 1643.577325566
29 1.0132703342139 1.428679538345

73e-34 
1.710168020360

34e-32 trkH K(+) transporter TrkH 

cds-NP_414788.1 55.04036607967
91 1.01552102035051 6.042082922824

27e-05 
0.000273903636

286476 perR putative transcriptional regulator 
PerR 

cds-NP_418355.1 122.9398095469
53 1.01826336511737 6.077470622096

21e-07 
4.153876139701

26e-06 yiiQ DUF1454 domain-containing 
protein YiiQ 

cds-NP_415742.1 84.18296536103
44 1.01887068700073 2.551319872115

02e-05 
0.000123765560

93754 narG nitrate reductase A subunit alpha 

cds-NP_415102.1 130.9254563585
41 1.02024355529118 3.190334835468

15e-07 
2.249998883166

95e-06 cusS sensor histidine kinase CusS 

cds-NP_415011.1 2338.426994710
37 1.0210262243121 2.415177914205

61e-20 
8.696604050420

02e-19 ybaL putative transporter YbaL 

cds-YP_026261.1 33.92884962003
93 1.02237613584601 0.011500301529

3827 
0.029138921895

6727 yigE DUF2233 domain-containing 
protein YigE 

cds-YP_588445.1 41.97181210256
99 1.02336757735155 0.000337916488

218629 
0.001285766431

54666 insA4 IS1 family protein InsA 

cds-NP_417707.1 81.95134904818
18 1.0244974413743 4.791455265013

19e-05 
0.000224564607

07665 aaeB aromatic carboxylic acid efflux 
pump subunit AaeB 

cds-NP_416492.1 65.08891664272
58 1.0248492582847 0.000660919943

713002 
0.002360656798

95555 cbl DNA-binding transcriptional 
activator Cbl 

cds-NP_415352.1 470.8859974034
94 1.02755805800909 1.428996313550

22e-18 
4.364844601871

67e-17 gsiC glutathione ABC transporter 
membrane subunit GsiC 

cds-NP_415656.1 35.00228370798
56 1.02896926292454 0.005906868426

09012 
0.016371414430

0082 ymfE uncharacterized protein YmfE 

cds-NP_417053.2 96.92744751379
27 1.02962509398857 3.880326219544

52e-06 
2.252420029667

46e-05 mltF membrane-bound lytic murein 
transglycosylase F 

cds-NP_416060.1 31.44819757929
8 1.03302753313791 0.001700330359

7675 
0.005492898003

94622 ydfI putative oxidoreductase YdfI 

cds-NP_418290.4 889.5105469945
46 1.03596699712105 1.446774751634

27e-11 
2.080443303567

59e-10 yigZ IMPACT family member YigZ 

cds-NP_418388.1 33.48311635901
06 1.0398772910638 0.005337967957

96257 
0.015001180257

4976 frwD putative PTS enzyme IIB 
component FrwD 



Supplementary Material 

  234 

cds-NP_414790.1 278.8355330180
69 1.04209469776709 1.044954172225

82e-15 
2.461756398291

56e-14 insI1 IS30 family transposase 

cds-NP_417393.1 22.05678284134
77 1.04227828484145 0.017087231694

7267 
0.040996397170

0674 argK methylmalonyl-CoA mutase-
interacting GTPase YgfD 

cds-NP_418633.4 28.63031426004
53 1.04427830743771 0.010874080536

8009 
0.027855004451

9902 ytfH putative transcriptional regulator 
YtfH 

cds-NP_416933.4 150.7991593207
81 1.04766182666408 3.641120472479

86e-12 
5.560869852625

28e-11 eutK 
putative structural protein 
ethanolamine utilization 

microcompartment 

cds-NP_418487.1 40.47478463926
69 1.06666540014456 0.002434982103

976 
0.007584061700

7804 soxR DNA-binding transcriptional dual 
regulator SoxR 

cds-NP_416592.2 25.40545610052
29 1.06704616097959 0.005546100233

48265 
0.015536798187

2831 insE5 IS3 element protein InsE 

cds-NP_418599.1 325.3238602074
18 1.07089115109254 2.920255593131

12e-06 
1.733754962731

6e-05 nsrR DNA-binding transcriptional 
regulator NsrR 

cds-NP_414798.1 55.06597263619
23 1.07183996325242 0.000100929801

631001 
0.000431901537

607443 insB2 IS1 family protein InsB 

cds-NP_414799.1 53.50670933429
12 1.0763069049935 1.977708749476

03e-05 
9.919900397994

73e-05 insA2 IS1 family protein InsA 

cds-NP_415314.1 378.1529560199
04 1.07836248111912 4.132772670967

07e-15 
8.800024115246

71e-14 ybhS ABC exporter membrane subunit 
YbhS 

cds-NP_415696.1 185.2281440760
08 1.07964871389736 1.128244820237

93e-08 
1.065457635145

8e-07 pliG inhibitor of g-type lysozyme 

cds-NP_415380.1 335.0345086543
13 1.08168361288951 1.290506594022

95e-09 
1.414765768546

45e-08 rlmC 23S rRNA m(5)U747 
methyltransferase 

cds-NP_416627.2 21.08863116833
55 1.0829867270876 0.005458630432

85074 
0.015311129947

4958 yehR DUF1307 domain-containing 
lipoprotein YehR 

cds-NP_418197.1 2923.082586016
15 1.08428400826736 4.407491867904

81e-32 
4.337951440655

64e-30 mnmG 
5-

carboxymethylaminomethyluridine
-tRNA synthase subunit MnmG 

cds-NP_414841.1 912.5979546853
23 1.08568446761277 2.524275454386

65e-08 
2.218257140372

71e-07 ykgF 

putative amino acid 
dehydrogenase with NAD(P)-

binding domain and ferridoxin-like 
domain 

cds-NP_418607.1 16.62222705907
86 1.08955841944942 0.020419223868

9829 
0.047611844245

167 yjfC putative acid--amine ligase YjfC 

cds-NP_415874.1 200.1924996914
27 1.09878852664158 9.315555467343

25e-13 
1.516860116355

27e-11 racR DNA-binding transcriptional 
repressor RacR 

cds-NP_415628.1 1101.590976348
8 1.10313617966822 1.142382660501

01e-14 
2.299824001526

8e-13 ycfJ PF05433 family protein YcfJ 

cds-NP_416482.1 49.57070942379
01 1.10767706641591 1.989801358822

62e-05 
9.958000246582

35e-05 zinT metal-binding protein ZinT 

cds-NP_418554.1 63.17356605423
84 1.10784595210329 0.000111245970

422686 
0.000467909214

626853 dtpC dipeptide/tripeptide:H(+) 
symporter DtpC 

cds-NP_417980.1 80.95128890040
99 1.10984785464409 1.301887052908

52e-06 
8.393097172244

27e-06 yhjE putative transporter YhjE 

cds-YP_026202.1 644.3917040946
89 1.11055023548307 1.422663413520

72e-06 
9.066152889903

98e-06 cyuA putative L-cysteine desulfidase 
CyuA 

cds-NP_417641.4 19.16179350363
86 1.11352047681276 0.021446461235

9755 
0.049653098177

8022 yhbX putative hydrolase YhbX 

cds-NP_416276.4 132.8818999160
08 1.1179931232993 6.641694450356

07e-06 
3.672417568118

23e-05 ynjI DUF1266 domain-containing 
protein YnjI 

cds-NP_415715.1 341.0853353219
6 1.12789442353723 1.728417984936

68e-09 
1.853577490962

71e-08 treA periplasmic trehalase 

cds-NP_414746.1 89.91894266638
79 1.13986330922486 7.026409873558

6e-06 
3.851481352721

66e-05 yafE putative S-adenosylmethionine-
dependent methyltransferase 

cds-NP_415353.1 417.6297922033
51 1.14586318165661 1.129100850409

53e-20 
4.167323055386

5e-19 gsiD glutathione ABC transporter 
membrane subunit GsiD 

cds-NP_416998.1 58.15032697893
87 1.14782304048239 1.169126874052

71e-05 
6.179072703078

08e-05 pdeF cyclic di-GMP phosphodiesterase 
PdeF 



Supplementary Material 

 235 

cds-NP_414655.1 1696.177986472
73 1.15170825343185 5.007291820472

4e-19 
1.618780691450

53e-17 pdhR DNA-binding transcriptional dual 
regulator PdhR 

cds-NP_417705.2 554.7463352985
18 1.15793615440624 4.697504139067

02e-08 
3.925518081495

82e-07 yhcN DUF1471 domain-containing 
stress-induced protein YhcN 

cds-NP_417975.1 36.32441700245
43 1.16485148011067 0.000684493228

864685 
0.002436993979

61551 ccp cytochrome c peroxidase 

cds-NP_417420.1 54.58979586975
38 1.16688193254083 8.320868527561

91e-05 
0.000362727625

084367 endA DNA-specific endonuclease I 

cds-NP_418379.1 34.25876031584
42 1.1682014066059 0.000933672516

618977 
0.003233178714

70324 yijF DUF1287 domain-containing 
protein YijF 

cds-NP_418352.1 53.44130581491
63 1.16863814920423 5.974971653354

59e-06 
3.349765753507

28e-05 sbp sulfate/thiosulfate ABC transporter 
periplasmic binding protein Sbp 

cds-NP_418129.2 1147.598343166
8 1.17088301705211 4.064791842006

45e-16 
9.784219058829

65e-15 emrD multidrug efflux pump EmrD 

cds-NP_417902.1 13.10737706372
63 1.17143200040919 0.021275921501

875 
0.049361475291

6733 insB6 IS1 family protein InsB 

cds-NP_417911.4 72.40172556080
62 1.17758607596612 6.315729490987

74e-07 
4.303440910089

96e-06 livF 

branched chain amino 
acid/phenylalanine ABC 

transporter ATP binding subunit 
LivF 

cds-NP_418015.1 35.22050422382
51 1.18939219513524 9.539665234420

97e-05 
0.000411003670

459635 insK IS150 family conserved protein 
InsB 

cds-NP_418716.1 36.26806575717
54 1.18951282748992 0.000800863940

637504 
0.002806191766

67999 yjhF putative transporter YjhF 

cds-YP_009518778.1 15.49323298406
05 1.19305174684976 0.008437190365

11689 
0.022296131340

7534 ynaA 
putative prophage tail length tape 

measure domain-containing 
protein YnaA 

cds-NP_415215.1 36.31092680535
16 1.19454170094853 5.954251868142

62e-05 
0.000270198581

188562 ybfP lipoprotein YbfP 

cds-NP_414868.1 13.91352880164
95 1.195366756555 0.017652915332

3918 
0.042012231062

4197 prpD 2-methylcitrate dehydratase 

cds-NP_417978.2 99.12833624031
33 1.19883607926296 1.298156725410

83e-07 
9.845096124733

84e-07 rcdB putative DNA-binding 
transcriptional regulator YhjC 

cds-YP_025303.1 99.98963851365
19 1.20294617417253 6.169806125301

06e-08 
5.032425659108

36e-07 blr beta-lactam resistance protein 

cds-NP_415308.1 49.83009960445
55 1.2090869675259 0.000184929027

664461 
0.000743240166

538928 ybhM Bax1-I family protein YbhM 

cds-NP_418566.1 3990.305475406
23 1.21057650434544 4.756258020802

26e-08 
3.959674205664

14e-07 groS cochaperonin GroES 

cds-NP_414815.1 211.8271058913
96 1.21825722973678 3.958620860258

35e-14 
7.492620423112

91e-13 intF putative phage integrase 

cds-NP_414869.1 45.30984500639
3 1.22197510697539 0.000122094575

992224 
0.000508708256

885759 prpE propionyl-CoA synthetase 

cds-NP_415089.1 203.6718321920
13 1.22693004894377 3.136876937657

97e-08 
2.687278134794

42e-07 borD prophage lipoprotein BorD 

cds-NP_416462.1 13.69192123517
48 1.2343531779132 0.008678143638

4634 
0.022840329647

6645 dsrB protein DsrB 

cds-NP_416403.1 12.29433717366
08 1.23449630880773 0.013448439246

275 
0.033387408868

6951 motB motility protein B 

cds-NP_415712.1 19.66557500206
32 1.2349787035729 0.003778477966

26611 
0.011031561577

1869 ycgR flagellar brake protein YcgR 

cds-YP_009518788.1 12.88853637358
79 1.23653289889415 0.017762529723

1659 
0.042227720957

5425 nohA putative prophage DNA-packaging 
protein NohA 

cds-NP_414562.1 27.21856646034
85 1.24208666188148 0.000485573085

080603 
0.001787700077

99002 insB1 IS1 family protein InsB 

cds-NP_417074.1 131.3512653872
55 1.24574239465879 5.667045248288

29e-06 
3.185195863536

66e-05 grcA stress-induced alternate pyruvate 
formate-lyase subunit 

cds-NP_418299.1 66.72855561951
83 1.24748075784684 7.839605673155

54e-06 
4.255099696863

46e-05 yihG 1-acylglycerol-3-phosphate O-
acyltransferase YihG 

cds-NP_417628.2 26.70273219621
74 1.24767293457218 0.005670198743

44232 
0.015804474659

9786 ubiV ubiquinone biosynthesis protein 
UbiV 



Supplementary Material 

  236 

cds-NP_416710.1 353.8969930754
41 1.24841097496982 9.781196090934

37e-13 
1.569598459664

79e-11 napA periplasmic nitrate reductase 
subunit NapA 

cds-NP_416145.1 177.4433800430
95 1.24864132710542 5.745484622122

78e-09 
5.605011319687

62e-08 rsxB SoxR [2Fe-2S] reducing system 
protein RsxB 

cds-NP_416117.1 252.0627855961
59 1.25104735760524 1.536972357317

58e-06 
9.738555894935

02e-06 mdtJ multidrug/spermidine efflux pump 
membrane subunit MdtJ 

cds-YP_009518795.2 84.64773598648
15 1.25767030381097 0.000356584149

624325 
0.001350993326

50653 yoaL protein YoaL 

cds-NP_417901.1 9.083663743775
04 1.25794400342875 0.020601711839

9986 
0.047973176519

1134 insA6 IS1 family protein InsA 

cds-NP_415296.1 274.9697529076
71 1.25936278720485 1.090539591629

45e-08 
1.034261210134

22e-07 bioB biotin synthase 

cds-NP_415896.1 2122.421755296
37 1.25980670401305 1.207559249894

63e-34 
1.485633310495

37e-32 ydbK putative pyruvate-flavodoxin 
oxidoreductase 

cds-NP_414992.1 97.56845814048
65 1.2603231213097 1.250010361443

41e-05 
6.559592287716

68e-05 maa maltose O-acetyltransferase 

cds-NP_416142.1 12.27658807899
73 1.26777708938481 0.010992569407

9808 
0.028142248501

7034 cnu H-NS- and StpA-binding protein 

cds-NP_414808.1 97.94866145680
99 1.27507659045896 2.691686746809

07e-07 
1.916636752671

6e-06 insB3 IS1 family protein InsB 

cds-NP_415653.4 132.9153806989
54 1.27519367703135 1.819533559713

38e-08 
1.651375847534

95e-07 rluE 23S rRNA pseudouridine(2457) 
synthase 

cds-NP_414923.1 67.63688617031
15 1.27519966527257 3.676332708307

47e-06 
2.145253961145

43e-05 yaiA protein YaiA 

cds-NP_416708.1 106.2857069182
24 1.28483560158129 6.367342208135

69e-08 
5.174487823822

57e-07 napH ferredoxin-type protein NapH 

cds-NP_417942.6 649.5910253241
45 1.28741950668767 4.758929883739

07e-17 
1.254601217564

31e-15 yhhJ ABC transporter family protein 
YhhJ 

cds-NP_414840.1 457.7445069435
93 1.28813814859705 4.436793913225

44e-08 
3.732871912721

98e-07 ykgE putative lactate utilization 
oxidoreductase YkgE 

cds-NP_418567.1 24347.88890688
78 1.29378057947902 5.578500429637

61e-15 
1.159961427364

55e-13 groL chaperonin GroEL 

cds-YP_026277.1 182.6056226426
96 1.29566771502734 1.006504245401

5e-10 
1.299652274893

07e-09 cpxP periplasmic protein CpxP 

cds-YP_026229.1 16.29106542472
83 1.30179473730278 0.002491863782

9771 
0.007734032722

3585 hokA small toxic polypeptide 

cds-NP_417065.1 936.9139784418
68 1.30343520672179 2.429973296352

43e-28 
1.793725288257

49e-26 rseC protein RseC 

cds-NP_414842.4 724.9604012807
69 1.30984573829809 2.890471385331

13e-09 
2.970277903858

83e-08 ykgG DUF162 domain-containing lactate 
utilization protein YkgG 

cds-NP_418344.3 1774.789669426
83 1.31015744032725 6.810043247717

36e-08 
5.473989390951

04e-07 sodA superoxide dismutase (Mn) 

cds-NP_415565.1 142.6254327754
88 1.31533817360399 7.577406786016

56e-11 
9.988194837877

18e-10 opgC 
osmoregulated periplasmic 
glucans (OPG) biosynthesis 

protein C 

cds-NP_417520.1 28.77181544715
35 1.32545432259091 6.095866046871

11e-05 
0.000275777229

025456 yqiI putative fimbrial protein YqiI 

cds-NP_416608.1 24.67395496634
39 1.34169229439423 0.000579344174

652231 
0.002099767061

81238 rcnR DNA-binding transcriptional 
repressor RcnR 

cds-NP_415289.4 79.50780215137
68 1.34349476900522 2.958530323833

7e-08 
2.574328252310

3e-07 ybhD putative DNA-binding 
transcriptional regulator YbhD 

cds-NP_415336.1 311.9254114929
02 1.34501951979671 1.150217949760

7e-08 
1.083896872231

94e-07 opgE phosphoethanolamine transferase 
OpgE 

cds-NP_415508.1 406.3412469790
03 1.34589559108923 3.504232563031

68e-10 
4.172109145609

49e-09 insB4 IS1 family protein InsB 

cds-NP_415355.1 69.38485796084
07 1.34775267807086 0.000105132482

434497 
0.000447292761

481641 dgcI putative diguanylate cyclase DgcI 

cds-NP_418625.1 23.28409438601
96 1.35108591962044 0.001801794796

20675 
0.005791109689

69498 yjfZ DUF2686 domain-containing 
protein YjfZ 



Supplementary Material 

 237 

cds-YP_010051200.1 11.76742723905
53 1.35621467639943 0.004744789754

10035 
0.013531663761

0499 yqiM protein YqiM 

cds-NP_418729.1 26.52374366241
58 1.35956868669724 0.003190631923

91415 
0.009561102023

69131 nanS N-acetyl-9-O-acetylneuraminate 
esterase 

cds-NP_415242.1 447.5105494509
63 1.36085650138449 2.554898837300

46e-33 
2.694201654858

03e-31 nei endonuclease VIII 

cds-
gnl|b4580|CDS%3D3

93 
56.26462673832

47 1.36335451046513 1.074917213841
83e-05 

5.715256110570
78e-05 yaiT putative autotransporter YaiT 

cds-
gnl|b4580|CDS%3D3

93 
56.26462673832

47 1.36335451046513 1.074917213841
83e-05 

5.715256110570
78e-05 yaiT putative autotransporter YaiT 

cds-YP_026237.1 42.44481568606
18 1.36389294616358 6.296360573075

13e-07 
4.296853771671

77e-06 dgoD D-galactonate dehydratase 

cds-NP_418251.1 36.25970157588
45 1.3686814593136 4.580855749349

82e-06 
2.621267456572

4e-05 cyaY frataxin CyaY 

cds-NP_416761.4 87.85953999989
41 1.37549330918186 8.946141702234

24e-07 
5.913800238685

89e-06 arnF undecaprenyl-phosphate-alpha-L-
Ara4N flippase - ArnF subunit 

cds-YP_588477.1 43.95983431426
98 1.38915811134117 2.743341304449

77e-06 
1.635331599161

e-05 topAI toxin of the TopAI-YjhQ toxin-
antitoxin system TopA inhibitor 

cds-NP_415207.1 75.31040183062
54 1.3967451346223 9.567083398637

76e-05 
0.000411784376

79851 chiP chitooligosaccharide outer 
membrane channel 

cds-NP_416914.1 20.40291719345
07 1.40402497008817 0.000523218308

502994 
0.001913570510

61912 yfeK DUF5329 domain-containing 
protein YfeK 

cds-NP_416255.1 155.8784888530
94 1.40424543235839 2.731221380249

38e-15 
5.929695829962

99e-14 cho excinuclease Cho 

cds-NP_416706.1 91.81856110797
73 1.40578229706932 1.474786048826

84e-11 
2.113860003318

47e-10 napC periplasmic nitrate reductase 
cytochrome c protein 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.40608625116431 7.174944148869
59e-05 

0.000319696455
083938 insO IS911B regulator fragment 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.40608625116431 7.174944148869
59e-05 

0.000319696455
083938 insO IS911B regulator fragment 

cds-
gnl|b4623|CDS%3D4

503 
42.23515573558

37 1.40608625116431 7.174944148869
59e-05 

0.000319696455
083938 insO IS911B regulator fragment 

cds-NP_418014.1 53.24318069765
57 1.40698130846236 1.242767931636

61e-06 
8.035356451413

93e-06 insJ insertion element IS150 protein 
InsA 

cds-YP_588446.1 57.37993177778
82 1.41454713811506 4.136937288830

15e-08 
3.496659399280

29e-07 gnsA putative phosphatidylethanolamine 
synthesis regulator GnsA 

cds-NP_415354.1 118.9926249681
93 1.4300951501499 2.428713546743

33e-13 
4.251688655543

96e-12 pdeI putative c-di-GMP 
phosphodiesterase PdeI 

cds-NP_417083.1 1529.174392278
8 1.43224551831605 9.345369959750

52e-05 
0.000403811156

602293 clpB chaperone protein ClpB 

cds-NP_416116.1 136.3495821180
21 1.43678747065379 3.429850272436

04e-10 
4.094557104210

03e-09 mdtI multidrug/spermidine efflux pump 
membrane subunit MdtI 

cds-NP_414897.1 10.18452702833
53 1.4437793401756 0.016015411750

5317 
0.038803204946

9939 yaiP putative glucosyltransferase 

cds-NP_415255.1 10316.86885494
05 1.45470359764682 1.580733782693

24e-06 
9.958847686413

04e-06 sucB dihydrolipoyltranssuccinylase 

cds-YP_009518777.1 9.330592992853
66 1.46243314214388 0.018878308137

0004 
0.044498151537

4001 ydaY putative uncharacterized protein 
YdaY 

cds-NP_415630.1 18.98054559660
19 1.47207742962856 0.007463688059

59825 
0.020070840568

2821 bhsA 
DUF1471 domain-containing 

multiple stress resistance outer 
membrane protein BhsA 

cds-NP_418435.1 58.15345329252
62 1.47268940244275 6.751219565804

16e-09 
6.557270056347

95e-08 yjaA stress response protein 

cds-NP_416650.1 24.78795173660
89 1.47931329674478 0.000252045948

551001 
0.000982180796

439726 yeiS DUF2542 domain-containing 
protein YeiS 

cds-NP_415298.1 94.24245525215
32 1.4912203837612 6.347230884915

17e-10 
7.208175792125

46e-09 bioC malonyl-acyl carrier protein 
methyltransferase 



Supplementary Material 

  238 

cds-NP_416709.1 66.03098742297
19 1.49931141881437 2.087595588531

66e-09 
2.206673236660

31e-08 napG ferredoxin-type protein NapG 

cds-NP_416999.1 61.79703500479
02 1.49941348655624 7.260586160709

81e-06 
3.970016803183

18e-05 yfgG nickel/cobalt stress response 
protein YfgG 

cds-NP_415060.1 38.31916220756
06 1.5064655424725 8.020940567186

33e-06 
4.342878456243

06e-05 ybcI PF04307 family inner membrane 
protein YbcI 

cds-NP_418305.1 2114.394713807
07 1.51092327017641 5.077922190696

38e-23 
2.249011738259

43e-21 glnL protein histidine kinase NtrB 

cds-NP_415054.1 19.64457867772
02 1.5114668358808 0.000537400002

117821 
0.001962196710

12352 ybcF putative carbamate kinase 

cds-NP_418198.1 1063.266353422
84 1.51245774371108 7.131839939740

51e-23 
3.096756773834

38e-21 mioC flavoprotein MioC 

cds-NP_415097.1 4805.989202823
66 1.52107391352721 4.846148360695

78e-42 
1.341474443095

1e-39 ompT omptin family outer membrane 
protease OmpT 

cds-NP_415211.2 92.17033505302
4 1.52276955473755 2.727288826799

44e-07 
1.938870339308

94e-06 ybfE ribbon-helix-helix domain-
containing protein YbfE 

cds-NP_414993.1 399.8376879697
56 1.52561353207227 7.278843100927

51e-21 
2.732033567288

81e-19 hha hemolysin expression-modulating 
protein Hha 

cds-NP_416903.1 10.75910637642
74 1.5575994893727 0.003235518067

27715 
0.009675968615

78022 yfeN putative outer membrane porin 
YfeN 

cds-NP_415507.2 13.56824978534
35 1.5608631365583 0.002257668547

30505 
0.007096674234

21863 gfcA threonine-rich inner membrane 
protein GfcA 

cds-NP_414924.1 162.5383638944
77 1.56354344574511 2.148652796037

52e-15 
4.782102127462

41e-14 aroM protein AroM 

cds-NP_416646.1 156.8730078237
44 1.56525014156797 5.479538459918

4e-12 
8.198944540195

48e-11 yohJ putative 3-hydroxypropanoate 
export protein YohJ 

cds-YP_002791259.1 21.47692468782
8 1.57119199262285 0.000151015685

762786 
0.000614185924

925051 ilvX uncharacterized protein IlvX 

cds-NP_414769.1 12.29142583880
67 1.58194348672882 0.005738774752

0645 
0.015945441265

3034 yafP putative N-acetyltransferase YafP 

cds-NP_414684.1 73.93551205599
71 1.58409216673939 4.493939782978

55e-10 
5.224057558743

3e-09 folK 
2-amino-4-hydroxy-6-

hydroxymethyldihydropteridine 
diphosphokinase 

cds-NP_415297.1 164.5303195813
87 1.59868032683366 9.183708116132

49e-10 
1.027137455715

93e-08 bioF 8-amino-7-oxononanoate 
synthase 

cds-NP_416220.1 11.04744892277
67 1.6012865719542 0.008508542284

35906 
0.022457886637

322 ydiE PF10636 family protein YdiE 

cds-NP_418340.1 9.839581373527
08 1.61616236216875 0.014221359890

1733 
0.035129059092

9044 rhaB rhamnulokinase 

cds-NP_416649.1 91.39323008657
08 1.63069349412122 2.633489958828

13e-11 
3.679409156987

32e-10 sanA DUF218 domain-containing 
protein SanA 

cds-NP_415633.1 13.27676564564
84 1.63077935176162 0.002820495361

09318 
0.008615154451

22874 ycfT inner membrane protein YcfT 

cds-NP_414573.1 673.4300652205
16 1.63644396610209 1.560855559850

32e-20 
5.713247334361

21e-19 carA carbamoyl-phosphate synthetase 
small subunit 

cds-NP_417708.1 52.63640027498
84 1.63690331937343 1.484005472046

71e-10 
1.872552773702

24e-09 aaeA aromatic carboxylic acid efflux 
pump membrane fusion protein 

cds-NP_415006.1 10418.01854438
76 1.64705959507874 9.144040904736

4e-25 
4.821304424652

09e-23 htpG chaperone protein HtpG 

cds-NP_416707.4 18.93053048349
64 1.67984943238161 1.891725379295

27e-05 
9.542655700340

25e-05 napB periplasmic nitrate reductase 
cytochrome c550 protein 

cds-NP_417214.1 7.511245656732
79 1.6822623253462 0.021093589581

9821 
0.048964102860

9008 pphB phosphoprotein phosphatase 2 

cds-NP_416242.6 65.88491953164
94 1.68856310197779 3.847003670248

37e-11 
5.242578232470

77e-10 ydjM inner membrane protein YdjM 

cds-NP_416808.1 40.70327276883
25 1.71179697616877 7.793099074605

91e-08 
6.163506393112

43e-07 rpnB recombination-promoting nuclease 
RpnB 

cds-NP_416778.2 51.27857766603
5 1.72176538052024 1.492909298471

7e-07 
1.113147354028

81e-06 yfbP uncharacterized protein YfbP 



Supplementary Material 

 239 

cds-NP_417610.1 9.234983398909
41 1.75258132047211 0.017480304701

5254 
0.041691044438

91 agaI putative deaminase AgaI 

cds-NP_414768.1 47.40810799447
06 1.77151344495762 1.117131915063

58e-06 
7.276143017377

34e-06 yafO ribosome-dependent mRNA 
interferase toxin YafO 

cds-NP_416062.2 21.52596879825
84 1.77755342965181 0.006983543612

7837 
0.018940670337

4274 ydfK cold shock protein YdfK 

cds-YP_001165332.1 4.935429242969
8 1.79251108444304 0.021337891802

3103 
0.049479331304

9385 yjbT PF17089 family protein YjbT 

cds-NP_416647.1 543.5021968253
51 1.82762085707818 5.954697374158

21e-15 
1.226667659076

59e-13 yohK putative 3-hydroxypropanoate 
export protein YohK 

cds-NP_415257.1 5422.091361455
16 1.82776071729034 4.318148212444

74e-07 
2.997661196382

09e-06 sucD succinyl-CoA synthetase subunit 
alpha 

cds-NP_415256.1 8848.675099583
82 1.83223524382477 1.073662619349

75e-07 
8.284410698780

6e-07 sucC succinyl-CoA synthetase subunit 
beta 

cds-NP_417289.2 17.03062787607
94 1.83799327808656 3.436811649114

74e-05 
0.000165093696

246521 ygdI DUF903 domain-containing 
lipoprotein YgdI 

cds-NP_418200.1 4993.314500759
5 1.85456662111002 5.960448638980

73e-19 
1.912958479858

38e-17 asnA asparagine synthetase A 

cds-YP_588452.1 8.283356601710
5 1.86457403741711 0.005974787952

05406 
0.016538959899

7796 rzoR putative prophage outer 
membrane lipoprotein RzoR 

cds-NP_418178.1 101.9515860857
31 1.86553662057153 1.088663245428

05e-07 
8.385546980871

03e-07 bglF 
beta-glucoside specific PTS 
enzyme II/BglG kinase/BglG 

phosphatase 

cds-NP_418441.1 26.81399071589
53 1.87329442009067 4.205318623512

06e-06 
2.422022910732

76e-05 arpA regulator of acetyl CoA synthetase 

cds-NP_418012.1 34841.40606166
52 1.93124288388787 1.824062585075

04e-05 
9.241677611534

46e-05 cspA cold shock protein CspA 

cds-NP_416877.1 30.56633160617
53 1.9448270081038 2.170879259660

72e-08 
1.926818485177

82e-07 ypdI colanic acid synthesis putative 
lipoprotein YpdI 

cds-NP_416256.2 10.83360515263
15 1.95966017845059 0.001377133589

4381 
0.004544951317

15451 ves HutD family protein Ves 

cds-NP_416075.1 671.9271446626
15 2.06662997516882 7.597520236023

95e-15 
1.557843385432

87e-13 cspB cold shock-like protein CspB 

cds-YP_009518761.1 34.13499990501
9 2.09061057493477 0.000142778239

579607 
0.000587153967

59339 ymcF protein YmcF 

cds-NP_416335.4 100.3404221915
42 2.10067455396216 1.201546637674

06e-17 
3.346949722175

1e-16 mntP Mn(2(+)) exporter 

cds-NP_415893.2 28.07718439716
09 2.11189281019349 0.012871447233

8936 
0.032171354288

3265 ynaE uncharacterized protein YnaE 

cds-NP_417419.1 99.82983432080
4 2.12041539693794 3.103123445595

39e-12 
4.822362715979

64e-11 yggI protein YggI 

cds-NP_415510.1 351.5100485675
8 2.13197099102969 1.709996915747

89e-05 
8.695265602580

25e-05 cspG cold shock protein CspG 

cds-NP_416404.1 7.398522387900
37 2.14382656784402 0.008569775798

54815 
0.022579141589

3931 motA motility protein A 

cds-YP_009518818.1 21.98190121158
58 2.15980133017082 8.293601792440

38e-08 
6.512830201900

43e-07 yqiD protein YqiD 

cds-YP_009518779.1 43.29010439386
31 2.28841867330812 0.001187435247

74092 
0.003996315130

88489 ynaM protein YnaM 

cds-YP_009518791.1 40.57072597008
76 2.32354809782248 8.534673067464

22e-06 
4.608069869201

72e-05 ynfS protein YnfS 

cds-NP_415299.1 54.77169165425
99 2.36557925484608 1.018372030805

98e-15 
2.411962419486

46e-14 bioD dethiobiotin synthetase 

cds-YP_010051187.1 13.37409804699
16 2.46316291581668 6.970665880457

91e-06 
3.830406846718

13e-05 mdtU protein MdtU 

cds-NP_415679.1 5.057430647547
49 2.5476157217157 0.010597483073

3955 
0.027241005532

2511 ycgX DUF1398 domain-containing 
protein YcgX 

cds-NP_416076.1 27.67830259296
54 2.58323843472484 6.639112737646

e-06 
3.672417568118

23e-05 cspF cold shock-like protein CspF 



Supplementary Material 

  240 

cds-YP_010051199.1 24.46398060727
47 2.59125485407825 4.874227835788

38e-09 
4.829520153178

24e-08 yghF putative type II secretion system 
C-type protein YghF 

cds-YP_026189.1 1233.752131885
17 2.63534562036893 3.136819814673

02e-62 
2.315495826531

14e-59 yghJ putative lipoprotein YghJ 

cds-NP_415086.1 3.842130974781
42 2.67785175475451 0.017686592045

0618 
0.042069772377

8619 essD putative phage lysis protein 

cds-NP_417446.4 50.23042037861
56 2.74275063346281 1.558765458763

37e-14 
3.082041168242

39e-13 pppA prepilin peptidase 

cds-NP_415506.1 6.472748132001
35 2.81448121050485 0.001370790690

51825 
0.004530770125

60101 gfcB lipoprotein GfcB 

cds-NP_415511.1 17.92039650075
79 2.91890168472461 0.000293366187

335756 
0.001127880940

72054 ymcE protein YmcE 

cds-NP_416837.1 4.895565727119
67 2.91904103909601 0.013327228357

0102 
0.033138679932

1514 yfcQ putative fimbrial protein YfcQ 

cds-YP_009518755.1 2.610625363053
52 3.00343209921787 0.019718301963

5196 
0.046183162028

7827 tfaD putative tail fiber assembly protein 
TfaD 

cds-YP_588474.1 6.114890429299
78 3.09342751292085 0.000520662915

757265 
0.001905798391

64374 ghoT toxin of the GhoTS toxin-antitoxin 
system 

cds-NP_417941.1 19.35419292973
96 3.20163725734292 5.182842990070

73e-07 
3.569955148215

13e-06 yhhI putative transposase 

cds-NP_417729.1 6.983882354189
38 3.30081181916733 0.000868893051

529721 
0.003015930505

66233 yhdU DUF2556 domain-containing 
protein YhdU 

cds-YP_588478.1 4.357965276438
71 3.5138201211375 0.003646531381

41875 
0.010716948319

4102 yjjZ DUF1435 domain-containing 
protein YjjZ 

cds-NP_415872.2 3.693415097400
92 3.73973720127324 0.004534017287

77206 
0.012972327240

0145 ydaF DUF1391 domain-containing 
protein YdaF 

cds-NP_415090.2 1.709846115818
21 3.80131019372962 0.019345718114

0143 
0.045454740332

6097 ybcV DUF1398 domain-containing 
protein YbcV 

cds-NP_417445.1 6.236953631470
38 3.99602037378283 0.001000365704

7311 
0.003439650986

24402 yghG lipoprotein YghG 

cds-NP_416774.2 2.139213662055
67 4.09513182323563 0.008091822874

20843 
0.021511814831

8542 yfbL putative peptidase YfbL 

cds-NP_416244.2 3.061229871938
06 4.1075469612904 0.003902153267

41303 
0.011370155803

5344 ydjO protein YdjO 

cds-NP_415509.1 20.58657693522 4.15346393752686 1.067085936267
93e-06 

6.980980224122
07e-06 cspH CspA family protein CspH 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 



Supplementary Material 

 241 

Supplementary Table S8. Sequences of all integron cassette 5’UTRs used in this work. 
GTT crossover point is marked in blue, and GFP start codon in green. Related to Figure 50. Table 
from 107 
 

ARC 5’ UTR Sequence 

aacA1 GTTAGGGCGACGCCGCTAATG 
aacA2 GTTAGGCGTCATG 
aacA3 GTTAGGCAGCACAGAGCGACCATTTCATG 
aacA4 GTTAGGCATCACAAAGTACAGCATCGTGACCAACAGCAACGATTCCGTCACAATG 
aacA5 GTTAGGCAGCACGGAGACACTTCAGCATG 
aacA7 GTTAGGCACCAATG 
aacA8 GTTAGGCAGCACAAACTCCGTCCTCATG 
aacA16 GTTGGGCTGATTGATTTGTTTGTTCTAGCATTACCTATATG 
aacA17 GTTGGGCTGATTGATTTGTTTGTTCTAGTATTACCTATATG 
aacA27 GTTAGGCCCGCACGGAATCAACATCTCATG 
aacA28 GTTAGCCGGACGCTGCGCGCGAAGAGGTTTTATG 
aacA29 GTTAGACGGCTATG 
aacA30 GTTAGGCTGGCGCGCTTCGCGCGGAAGACTTTATGGCTACTCGGAGACCTTAAATG 
aacA31 GTTAGGCAGCACAAAGACCGTTCTCATG 
aacA32 GTTAGGCAGCACAAAAGGACCGTCCCATG 
aacA34 GTTAGAAGGCCCAGGCTATG 
aacA35 GTTAGGCAGCACAGGGCCACCTTCTTATG 
aacA37 GTTAACCGCGGCTATG 
aacA38 GTTAGGCAGCACATAACCACCGTCACACCATG 
aacA39 GTTAGCCGGACGCTTCGCGCAAAGAGGTATTATG 
aacA40 GTTAGGCAGCACAGTCCAGACTCCGCATG 
aacA42 GTTAGGCTGACGCGCTTCGCGCGGAAGACTTTATGGCTACTCGGAGACTTTGAATG 
aacA43 GTTAGCCAGACGCTTCGCGCCGAGGACAAATG 

aacA44 GTTAGGCAGCACAAGACCACCTGTTCATGCCCGCGAACGAAAACACCGTAACCCTACGTCTGATGACTGA
GCACAATTGGTGATTAAATG 

aacA45 GTTAAAAGGCTCAGCCAATG 
aacA46 GTTAGGGCGACGCCGCATTCGCGGCGCGTGAAGAAAGAGGATCTTATG 
aacA47 GTTATG 
aacA48 GTTAGGTCCCACTAAACCTGCACCGAGCATG 
aacA49 GTTAGCTTGACGCTTCGCGCAGAGGAGAGTTTCAATG 
aacA50 GTTAGACAGCACAAAGACAATTCTCATG 
aacA51 GTTAGGCCACAAGGAACCGTCCCAGTATG 
aacA52 GTTAGGCAGCACAAGATG 
aacA54 GTTAGGCCGCACAAAATCAACGCCTTATG 
aacA56 GTTAGCCGGACGCTTCGCGCAGGAGTAAGAATG 
aacA59 GTTAGGCAGCACAGAAGCCGCATCCCATG 
aacA61 GTTAGGCAGCACAGGGCCACCGCTTATG 
aacA64 GTTAGCCGGACGCCTTCGGCGCTAGGAATAAAATG 

aacAX GTTATGCATACAAATCATCACCGTGATTTACTCTTACCGGAAAGCTGAAGAAACAGATAGAGAAGCCATCT
ACCAAATG 

aacC1 GTTAGGTGGCTCAAGTATG 
aacC2 GTTAGGTGGCTCAATG 
aacC3 GTTAGGCAGCAGCAGCTAAGATG 
aacC4 GTTAGGTGGCTCACGTATG 
aacC5 GTTAGGCATCAGGAGCAGACGAGTGTCAGTCGAAATCATCCATCTCACTGGAAACGATGTTGCGATG 
aacC6 GTTAGGTGGCTCAATG 
aacC11 GTTAGGTGGCTCACGTATG 
aacC13 GTTAGGCATTAGGAGCCGATGAATG 



Supplementary Material 

  242 

aadA1 GTTAAACATCATG 
aadA2 GTTAGACATCATG 
aadA4 GTTAGGCATCTTCATG 
aadA5 GTTAGGCATCATG 
aadA6 GTTAGACATCATG 
aadA7 GTTAGACATCATG 
aadA9 GTTAGACATG 
aadA10 GTTAGACATCATG 
aadA11 GTTAGACATCATG 
aadA13 GTTAGACATCATG 
aadA16 GTTAGACATCATG 
aadA24 GTTAGACATCATG 
aadA28 GTTAGACATCATG 
aadA29 GTTAGACATCATG 
aadA34 GTTAGACATCATG 
aadB GTTAGGCCGCATG 

aphA15 GTTAGACCGCTATG 
aphA16 GTTAGCTTGACGCTCCGCGCAGGAAAGAGAAAATG 
blaOXA-9 GTTATGCACCTATTAAGCGCACAGCGGAGCAATG 
dfrA5 GTTAACCCGGAACCAAAATTGTGAAAGTATCATTAATG 
fosG GTTATGTTTGTTAAGGTAGATTTGTGCTCCGAGGAATG 

aacA1 alt GTTAGGGCGACGCCGCTATTGCGGCGCGAATACAAAGAGGAAGAGATG 
aacA4 alt GTTAGGCATCACAAAGTACAGCATCATG 
aacA43 alt GTTAGCCAGACGCTTCGCGCCGAGGACAATTGATG 
aacA47 alt GTTATGCATCACAGAACCACCATACCTATG 

aacAX alt GTTATGCATACAAATCATCACCGTGATTTACTCTTACCGGAAAGCTGAAGAAACAGATAGAGAAGCCATCT
ACCAATTGTATTGCTTGGTAATG  

aacC5 alt GTTAGGCATCAGGAGCAGACGAATG 
aadA9 alt GTTAGACATGATG 

 
  



Supplementary Material 

 243 

Supplementary Figure S1. Resistance characterisation of aa ARCs by agar diffusion test.  
Antimicrobial resistance to several beta-lactams is shown as reduction in growth inhibition halo (mm.) in 
comparison with the parental strain in the presence of these antibiotic discs. Dark red correlates with higher 
resistance (inhibition halo decrease), while light blue denotes lower resistance. Blank cells represent a 
variation in inhibition halo £ 5 mm. (also found when unrelated antibiotics such as ampicillin and ciprofloxacin 
are tested). A phylogenetic tree showing sequence homology between the encoded proteins is shown in 
the left side of the graph. Figure from 122. 
 
 

Kan
am

yc
in

To
bra

myc
in

Amika
cin

Gen
tam

ici
n

Stre
pto

myc
in

Apra
myc

in

Neo
myc

in

sat2

aacAX

aphA15

aphA16

aacA49

aacA43

aacA64

aacA28

aacA56

aacA16

aacA17

aacA30

aacA42

aacA7

aacA2

aacA48

aacA29

aacA37

aacA34

aacA45

aacA4

aacA50

aacA8

aacA31

aacA51

aacA59

aacA38

aacA27

aacA54

aacA61

aacA3

aacA47

aacA35

aacA52

aacC5

aacC13

aacC3

aacC2

aacC6

aacC4

aacC1

aacC11

aadB

aadA4

aadA5

aadA29

aadA7

aadA6

aadA10

aadA16

aadA34

aadA1

aadA13

aadA2

aadA28

aadA11

aadA24

Antibiotics

A
R
C
s

-20 mm.

-15 mm.

-10 mm.

-5 mm.

Inhibition halo reduction

Kan
am

yc
in

To
bra

myc
in

Amika
cin

Gen
tam

ici
n

Stre
pto

myc
in

Apra
myc

in

Neo
myc

in

sat2

aacAX

aphA15

aphA16

aacA49

aacA43

aacA64

aacA28

aacA56

aacA16

aacA17

aacA30

aacA42

aacA7

aacA2

aacA48

aacA29

aacA37

aacA34

aacA45

aacA4

aacA50

aacA8

aacA31

aacA51

aacA59

aacA38

aacA27

aacA54

aacA61

aacA3

aacA47

aacA35

aacA52

aacC5

aacC13

aacC3

aacC2

aacC6

aacC4

aacC1

aacC11

aadB

aadA4

aadA5

aadA29

aadA7

aadA6

aadA10

aadA16

aadA34

aadA1

aadA13

aadA2

aadA28

aadA11

aadA24

Antibiotics

-20 mm.

-15 mm.

-10 mm.

-5 mm.

Inhibition halo reduction



Supplementary Material 

  244 

Supplementary Figure S2. Resistance characterisation of bla ARCs by agar diffusion test. 
Antimicrobial resistance to several beta-lactams is shown as reduction in growth inhibition halo (mm.) in 
comparison with the parental strain in the presence of these antibiotic discs. Dark blue correlates with higher 
resistance (inhibition halo decrease), while light blue denotes lower resistance. Blank cells represent a 
variation in inhibition halo £ 4 mm. (as observed for control antibiotic discs of kanamycin and ciprofloxacin). 
A phylogenetic tree showing sequence homology between the encoded proteins is shown in the left side of 
the graph. Figure from 122. 
 
  

Ampic
illin

Amox
ici

llin

Amox
. C

lav
ula

nic

Cefa
clo

r

Cefo
tax

im
e 

Cefo
xit

in 

Ceft
az

idi
me

Im
ipe

ne
m

Mero
pe

ne
m

Erta
pe

ne
m

Aztr
eo

na
m 

OXA-2

OXA-21

OXA-46

OXA-118

OXA-20

OXA-5

OXA-129

OXA-10

OXA-198

OXA-1

OXA-9

VIM-1

VIM-2

VIM-7

IMP-2

IMP-31

BEL-1

GES-1

PBL-1

Antibiotics

bl
a 

A
R

C
s

-25 mm.

-20 mm.

-15 mm.

-10 mm.

-5 mm.

Inhibition halo reduction

Ampic
illin

Amox
ici

llin

Amox
. C

lav
ula

nic

Cefa
clo

r

Cefo
tax

im
e 

Cefo
xit

in 

Ceft
az

idi
me

Im
ipe

ne
m

Mero
pe

ne
m

Erta
pe

ne
m

Aztr
eo

na
m 

OXA-2

OXA-21

OXA-46

OXA-118

OXA-20

OXA-5

OXA-129

OXA-10

OXA-198

OXA-1

OXA-9

VIM-1

VIM-2

VIM-7

IMP-2

IMP-31

BEL-1

GES-1

PBL-1

Antibiotics

bl
a 

A
R

C
s

-25 mm.

-20 mm.

-15 mm.

-10 mm.

-5 mm.

Inhibition halo reduction



Supplementary Material 

 245 

 
Supplementary Figure S3. pMBAARCs fluorescence do not change in stationary growth phase.  
A) GFP fluorescence normalized to pMBA levels of each pMBA derived strain in stationary phase. B) 
Normalised GFP fluorescence levels of pMBAARCs both in exponential and stationary growth phases. ARCs 
are ordered by families according to its nomenclature. Data is expressed as the mean of three independent 
replicates, error bars represent the standard error of the mean (SEM). C) Correlation between pMBAARCs 
GFP normalised fluorescence in exponential and stationary growth phases. Pearson correlation coefficient 
(r) and p-value are indicated. 
 
  

dfr
A1
2

bla
OX
A-
20

aa
cA
54

aa
cA
59

aa
cA
35

aa
cA
38arr

7

bla
OX
A-
1

dfr
A2
1

qa
cE
Δs
ul1
dfr
B7

dfr
A2
2

aa
cA
27

ap
hA
15

aa
dA
16
fos
M

bla
OX
A-
10

bla
IM
P-
31

bla
OX
A-
46

aa
cA
45arr

6

bla
PB
L-1

aa
cC
11

aa
cA
48
aa
dA
5
aa
cA
3

aa
cA
61
aa
dA
4
aa
cA
2
qa
cK

bla
BE
L-1arr

5

dfr
A2
9

aa
cA
31

bla
IM
P-
2

bla
OX
A-
19
8
fos
L

dfr
A3
4
dfr
B6
qa
cF
dfr
B1

aa
dA
34

bla
OX
A-
2

aa
cA
7
aa
dA
6
qa
cL

aa
cA
29
aa
cC
4
sa
t2

aa
cA
64
aa
cC
3
aa
cC
2

aa
cA
X
aa
cC
1
qa
cG
dfr
B5

dfr
A1
4

aa
cA
51
aa
dA
1
fos
G
fos
E

aa
dA
29

ca
tB
10
dfr
B9

bla
OX
A-
21

aa
cA
42

aa
cA
50

aa
cA
37

bla
VI
M-
7

aa
cC
6
dfr
A5
aa
cA
4

aa
dA
11
aa
cA
8
fos
I

dfr
A2
5

dfr
A3
5
aa
dA
7

dfr
A3
0
sm
r2

bla
OX
A-
12
9

bla
GE
S-
1

aa
dA
10
qa
cE
dfr
B2

aa
cA
17
ere
A3
sm
r3
dfr
B3

aa
cA
28

dfr
A1
7

dfr
A2
7
qa
cH

aa
cA
47

bla
OX
A-
11
8

ca
tB
6
ere
A2

aa
cA
52

aa
cA
30

dfr
A1
6

aa
cA
34
qa
cM
dfr
A1
5

bla
OX
A-
5
dfr
A1

aa
cA
16fos

F

bla
VI
M-
1

ap
hA
16
dfr
B8fos

H
ca
tB
3
dfr
A6
aa
dA
2

aa
dA
28

bla
OX
A-
9
fos
N
aa
cC
5
dfr
A7

aa
cA
49

aa
cA
56

bla
VI
M-
2

ca
tB
2
fos
K

aa
dA
24
ca
tB
5

dfr
A3
1

aa
cC
13
fos
C2arr

2
sm
r1

aa
dA
13
dfr
B4
aa
dB
arr
8b

aa
cA
43

0.01

0.1

1

10

ARCs

G
FP

 fl
uo

re
sc

en
ce

 (n
or

m
. p

M
B

A
)

aa
cA
2
aa
cA
3
aa
cA
4
aa
cA
7
aa
cA
8

aa
cA
16

aa
cA
17

aa
cA
27

aa
cA
28

aa
cA
29

aa
cA
30

aa
cA
31

aa
cA
34

aa
cA
35

aa
cA
37

aa
cA
38

aa
cA
42

aa
cA
43

aa
cA
45

aa
cA
47

aa
cA
48

aa
cA
49

aa
cA
50

aa
cA
51

aa
cA
52

aa
cA
54

aa
cA
56

aa
cA
59

aa
cA
61

aa
cA
64

aa
cA
X
aa
cC
1
aa
cC
2
aa
cC
3
aa
cC
4
aa
cC
5
aa
cC
6

aa
cC
11

aa
cC
13
aa
dA
1
aa
dA
2
aa
dA
4
aa
dA
5
aa
dA
6
aa
dA
7

aa
dA
10

aa
dA
11

aa
dA
13

aa
dA
16

aa
dA
24

aa
dA
28

aa
dA
29

aa
dA
34
aa
dB

ap
hA
15

ap
hA
16sa

t2
arr
2
arr
5
arr
6
arr
7
arr
8b

bla
BE
L-1

bla
GE
S-
1

bla
IM
P-
2

bla
I MP

-31

bla
OX
A-
1

bla
OX
A-
2

bla
OX
A-
5

bla
OX
A-
9

bla
OX
A-
10

bla
OX
A-
20

bla
OX
A-
21

bla
OX
A-
46

bla
OX
A-
11
8

bla
OX
A-
12
9

bla
OX
A-
19
8

bla
PB
L-1

bla
VI
M-
1

bla
VI
M-
2

bla
VI
M-
7

ca
tB
2
ca
tB
3
ca
tB
5
ca
tB
6

ca
tB
10
dfr
A1
dfr
A5
dfr
A6
dfr
A7

dfr
A1
2

dfr
A1
4

dfr
A1
5

dfr
A1
6

dfr
A1
7

dfr
A2
1

dfr
A2
2

dfr
A2
5

dfr
A2
7

dfr
A2
9

dfr
A3
0

dfr
A3
1

dfr
A3
4

dfr
A3
5
dfr
B1
dfr
B2
dfr
B3
dfr
B4
dfr
B5
dfr
B6
dfr
B7
dfr
B8
dfr
B9
ere
A2
ere
A3
fos
C2fos

E
fos
F
fos
G
fos
H fos

I
fos
K
fos
L
fos
M
fos
N
qa
cE

qa
cE
Δs
ul1qa
cF
qa
cG
qa
cH
qa
cK
qa
cL
qa
cMsm

r1
sm
r2
sm
r3

0.01

0.1

1

10

ARCs

G
FP

 fl
uo

re
sc

en
ce

 (n
or

m
. p

M
B

A
) GFP fluorescence expo.

GFP fluorescence stat. 

0.0 0.5 1.0 1.5 2.0 2.5
0.0

0.5

1.0

1.5

2.0

2.5

Correlation GFP fluorescence
stat. vs. expo.

GFP fluorescence expo. (norm. pMBA)

G
FP

 fl
uo

re
sc

en
ce

 s
ta

t. 
(n

or
m

. p
M

B
A

)

r = 0,8950
p-value <0.0001

A 

B 

C 



Supplementary Material 

  246 

 
pMBAARC GROWTH CURVES 

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

pMBA

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA2

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA3

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA4

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA7

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA8

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA16

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA17

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA27

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA28

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA29

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA30

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA31

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA34

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA35

0 5 10 15 20 25 30 35
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA37

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA38

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA42

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA43

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA45

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA47

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA48

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA49

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA50

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA51

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA52

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA54

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA56

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA59

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA61

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacA64

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacAX

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
D

60
0

aacC1

0 2 4 6 8 10 12 14 16 18 20 22 24
0.0

0.2

0.4

0.6

0.8

1.0

1.2

time (h)

O
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sat2



Supplementary Material 

 247 

 
  

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arr2

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arr7

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blaBEL-1

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blaGES-1

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blaIMP-2

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blaIMP-31

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blaOXA-1

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blaOXA-2

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blaOXA-9

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blaOXA-10

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blaOXA-20

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blaOXA-21

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blaOXA-46

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blaOXA-118

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blaOXA-129

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blaOXA-198

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blaPBL-1

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blaVIM-1

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blaVIM-2

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blaVIM-7

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catB2

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catB3

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catB5

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catB6

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catB10

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dfrA1

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dfrA21

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dfrA22

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dfrA25

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dfrB1

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dfrB2

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dfrB3

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dfrB4

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dfrB6

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dfrB7

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dfrB8

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dfrB9

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ereA2

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ereA3



Supplementary Material 

  248 

 

 
Supplementary Figure S4. Growth curves of all pMBA derivatives. 
Growth (OD600) was measured along 24h in MH medium supplemented with zeocin. Growth curves are 
represented as the mean of three independent replicates, the standard error of the mean (SEM) is 
represented as a shadow in lighter colour. 
  

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fosK

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fosL

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fosM

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fosN

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qacE

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qacEΔsul1

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qacK

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qacM

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smr1

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Supplementary Material 

 249 

Supplementary Figure S5. Relative abundance of each OMM12 member in vivo competitions. 
Relative abundances of every member of OMM12 consortia in mice before (day 0) and after (day 7 and 
caecum) in vivo competitions of pMBAARC DGFP (ereA2, aacA7, and blaOXA-10) vs. pMBA DGFP. OMM12 
members and E. coli are represented in different colours following the figure legend pattern. 
 

0

5

10

20

40

60

80

100

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re
la

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e 

ab
un

da
nc

e 
(%

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f t

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al

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en

e 
co

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es

)

0

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60

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E.coli

YL45

KB1

YL32

YL31

I49

I46

YL44

YL27

I48

YL58

0

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60

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100

 Day 7 caecum

re
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en

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co

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es

)

ereA2 aacA7 blaOXA-10



Supplementary Material 

  250 

 
Supplementary Figure S6. Volcano plots showing DE genes of pMBA and pMBA derivatives. 
A) pMBA DE genes in comparison with E. coli MG1655. B) pMBAereA3  DE genes in comparison with pMBA. 
C) pMBAdfrA31 DE genes in comparison with pMBA. D) pMBAdfrA21 DE genes in comparison with pMBA. Gene 
expression was measured by RNA-seq in every strain. Dotted-lines represent significance and fold change 
thresholds (padj <0.05 and fold change >2 respectively). Genes with a similar transcription in both strains 
are coloured in blue, while upregulated and downregulated genes are represented in green and red 
respectively. Data is represented as the mean of three independent replicates. 
 
 
 
 
 

-6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6

0

5

10

15

20

log2 (FoldChange)

-lo
g 10

(p
ad

j)

pMBAdfrA21 vs. pMBA

-6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6

0

50

100

150

200

log2 (FoldChange)

-lo
g 10

(p
ad

j)

pMBAdfrA31 vs. pMBA

-6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6

0

50

100

150

log2 (FoldChange)

-lo
g 10

(p
ad

j)

pMBAereA3 vs pMBA

-6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6

0

20

40

60

80

log2 (FoldChange)

-lo
g 10

(p
ad

j)
pMBA vs MG1655

A B 

C D 



Supplementary Material 

 251 

 

 
Supplementary Figure S7. Non-corrected recombination frequency of each ARC.  
Graph showing recombination rates of all ARCs present in pMBA recombination collection previous to 
integron-dependent checking and correction. Bars represents the mean of at least 3 biological replicates 
and error bars correspond to the standard error of the mean (SEM).  
 
 
 
 
 
 
 
 

  

aa
dA7 Δ

int

qac
EΔ

su
l1
dfrA

5

dfrA
16

dfrA
15

dfrA
35

dfrA
25
fosN

dfrA
14

fosF
aa

dA4

dfrA
12

dfrA
34

dfrA
30
fosK

dfrA
29

ap
ha1

6

bla OXA-5

blaI
MP31

aa
dA16

dfrA
27
qac

K
sm

r2

bla OXA-1

aa
dA10

ere
A3

aa
dA13

qac
G

ap
ha1

5
ca

tB
6
dfrB

6

bla OXA-2

bla OXA-12
9

bla OXA-21

aa
dA5

fosG

aa
cC

11

bla VIM
-7

ca
tB

2
arr

8b

aa
cC

13
dfrB

3
aa

dB
arr

2

bla OXA-46

aa
dA11

aa
dA24

sm
r3
dfrB

5
dfrA

7

aa
dA28

aa
cC

6
fosC

2
dfrB

8

bla OXA-19
8

dfrB
7

dfrA
17
ca

tB
3
dfrB

2
dfrB

1

bla OXA-20

ca
tB

5

bla OXA-10

aa
cA

49

aa
cA

16
aa

dA2

bla OXA-9

ca
tB

10

bla PBL-1

ere
A2

fosL
dfrB

4
qac

L
dfrA

1

aa
dA34

fosM
qac

E

bla OXA-11
8

bla VIM
-1
fosH

dfrA
31
aa

cC
2

bla VIM
-2

dfrB
9
qac

H
qac

MfosI

aa
dA1

arr
6
arr

5

aa
cA

30

aa
cA

42arr
7

dfrA
22sa

t2

bla IM
P-2

bla BEL-1
qac

F
sm

r1
aa

cC
5
dfrA

6

aa
cA

38

aa
cA

43
aa

cC
3
fosE

dfrA
21
aa

cA
3

aa
cA

27
aa

cC
1

bla GES-1

aa
cA

54

aa
dA29

aa
cA

17

aa
cA

56

aa
cA

28

aa
cA

47
aa

dA6

aa
cA

48
aa

dA7

aa
cA

35
aa

cA
4

aa
cC

4
aa

cA
8

aa
cA

59
aa

cA
2

aa
cA

31

aa
cA

34

aa
cA

64

aa
cA

45

aa
cA

29

aa
cA

51

aa
cA

37

aa
cA

X

aa
cA

50

aa
cA

61
aa

cA
7

aa
cA

52

10-8

10-7

10-6

10-5

10-4

10-3

10-2

10-1

100

ARCs

R
. F

.



Supplementary Material 

  252 

 

 
Supplementary Figure S8. Cassette expression correlates with the presence of SD-like sequences.  
Correlation between GFP fluorescence and the number of matches to AGGAGG at a distance of A) 6 bp from 
the start codon (positions -7 to -12); and B) 6 or 7 bp from the start codon (positions -7 to -13). Curves were 
fitted using a nonlinear (semilog) regression. 5’-UTRs that are too short to encode an RBS at these positions 
were excluded from the analysis. Related to Figure 50. Figure modified  from 107. 
 
  

A B 



Supplementary Material 

 253 
 



Supplementary Material 

  254 
 



Supplementary Material 

 255 
 



Supplementary Material 

  256 

 
Supplementary Figure S9. Antibiotic concentrations used do not affect growth. 
Growth curves of all constructions in media used to deliver “riboswitch” induction experiments. Curves are 
the mean of three biological replicates. The order of 5´UTRs is conserved among panels. Figure modified 
from 107. Related to Figures 51, 53, and 54. 
 
  



Supplementary Material 

 257 

 
Supplementary Figure S10. Alternative 5´UTRs are not induced by aminoglycosides. 
A) Sequence of main alternative 5´UTRs used in case of dubious annotations. B) Fluorescence of main and 
alternative sequences fused to the GFP gene. C) Induction ratios of main and alternative 5´UTRs for all Ag. 
Induction is not observed for any sequence. Figure from 107. 



Supplementary Material 

  258 

 
Supplementary Figure S11. Results from statistical models applied to induction assays. 
Change in fluorescence readings in the presence of A) aminoglycosides and B) all antibiotics, as determined 
by a linear mixed regression model considering gene, batch, and sample as random effects. C) Expected 
change in florescence readings after the addition of antibiotics is estimated by a linear mixed model including 
genes and batches as random effects. Related to figures 51 and 53. Figure from 107. 



Supplementary Material 

 259 

 
 
Supplementary Figure S12. Hierarchical clustering tree of all attC sites present in ARCs. 
Phylogenetic relation between nucleotidic sequences of all attC sites harboured in ARCs.  
 





 

  

 


	Tesis Alberto Hipólito Carrillo de Albornoz
	PORTADA
	ACKNOWLEDGEMENTS
	TABLE OF CONTENTS
	LIST OF ABBREVIATIONS
	RESUMEN/ SUMMARY
	RESUMEN
	SUMMARY

	INTRODUCTION
	OBJECTIVES
	MATERIALS AND METHODS
	RESULTS
	PART 1. EVOLUTIONARY DYNAMICS OF INTEGRON RESISTANCE CASSETTES
	PART 2. A DEEPER LOOK INTO THE INTEGRON MODEL

	DISCUSSION
	CONCLUSIONS
	BIBLIOGRAPHY
	ANNEX. SUPPLEMENTARY MATERIAL