Person: Domenech Lucas, Miriam
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First Name
Miriam
Last Name
Domenech Lucas
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Biológicas
Department
Genética, Fisiología y Microbiología
Area
Microbiología
Identifiers
18 results
Search Results
Now showing 1 - 10 of 18
Item Biofilm formation in Streptococcus pneumoniae(Microbial Biotechnology, 2011) Domenech Lucas, Miriam; García, Ernesto; Moscoso, MiriamBiofilm-grown bacteria are refractory to antimicrobial agents and show an increased capacity to evade the host immune system. In recent years, studies have begun on biofilm formation by Streptococcus pneumoniae, an important human pathogen, using a variety of in vitro model systems. The bacterial cells in these biofilms are held together by an extracellular matrix composed of DNA, proteins and, possibly, polysaccharide(s). Although neither the precise nature of these proteins nor the composition of the putative polysaccharide(s) is clear, it is known that choline-binding proteins are required for successful biofilm formation. Further, many genes appear to be involved, although the role of each appears to vary when biofilms are produced in batch or continuous culture. Prophylactic and therapeutic measures need to be developed to fight S. pneumoniae biofilm formation. However, much care needs to be taken when choosing strains for such studies because different S. pneumoniae isolates can show remarkable genomic differences. Multispecies and in vivo biofilm models must also be developed to provide a more complete understanding of biofilm formation and maintenance.Item Vancomycin tolerance in clinical and laboratory Streptococcus pneumoniae isolates depends on reduced enzyme activity of the major LytA autolysin or cooperation between CiaH histidine kinase and capsular polysaccharide(2010) Moscoso, Miriam; Miriam Domenech; Ernesto García; Domenech Lucas, MiriamVancomycin is frequently added to standard therapy for pneumococcal meningitis. Although vancomycin-resistant Streptococcus pneumoniae strains have not been isolated, reports on the emergence of vancomycin-tolerant pneumococci are a cause of concern. To date, the molecular basis of vancomycin tolerance in S. pneumoniae is essentially unknown. We examined two vancomycin-tolerant clinical isolates, i.e. a purported autolysin negative (LytA-), serotype 23F isolate (strain S3) and the serotype 14 strain ‘Tupelo’, which is considered a paradigm of vancomycin tolerance. S3 was characterized here as carrying a frameshift mutation in the lytA gene encoding the main pneumococcal autolysin. The vancomycin tolerance of strain S3 was abolished by transformation to the autolysin-proficient phenotype. The original Tupelo strain was discovered to be a mixture: a strain showing a vancomycin-tolerant phenotype (Tupelo_VT) and a vancomycin-nontolerant strain (Tupelo_VNT). The two strains differed only in terms of a single mutation in the ciaH gene present in the VT strain. Most interestingly, although the vancomycin tolerance of Tupelo_VT could be overcome by increasing the LytA dosage upon transformation by a multicopy plasmid or by externally adding the autolysin, we show that vancomycin tolerance in S. pneumoniae requires the simultaneous presence of a mutated CiaH histidine kinase and capsular polysaccharide.Item Vancomycin tolerance in Gram-positive cocci(Environmental Microbiology Reports, 2011) Moscoso, Miriam; Domenech Lucas, Miriam; García, ErnestoVancomycin, a glycopeptide antimicrobial agent, represents the last line of defence against a wide range of multi-resistant Gram-positive pathogens such as enterococci, staphylococci and streptococci. However, vancomycin-resistant enterococci and staphylococci, along with vancomycin-tolerant clinical isolates, are compromising the therapeutic efficacy of vancomycin. It is conceivable that tolerance may emerge during prolonged vancomycin use. It has not been until recently, however, that the molecular basis of this tolerance began to be understood. Superoxide anions might be involved in the bactericidal activity of vancomycin in enterococci, and recent evidence suggests that the stringent response is partly responsible for vancomycin tolerance in Enterococcus faecalis. The mechanism of vancomycin tolerance in Staphylococcus aureus and Streptococcus pneumoniae is sometimes associated with a reduction of autolysin activity. Vancomycin tolerance in S. aureus and S. pneumoniae also appears to be somehow related with the two-component regulatory systems linked to cell envelope stress, although the precise molecular regulatory pathways remain poorly defined.Item In vitro biofilm development of Streptococcus pneumoniae and formation of choline‐binding protein-DNA complexes(Environmental Microbiology Reports, 2015) Domenech Lucas, Miriam; Ruiz, Susana; Moscoso, Miriam; García, ErnestoExtracellular deoxyribonucleic acid (eDNA) is an essential component of bacterial biofilm matrices, and is required in their formation and maintenance. Extracellular DNA binds to exopolysaccharides or extracellular proteins, affording biofilms greater structural integrity. Recently, we reported evidence of intercellular eDNA−LytC complexes in pneumococcal biofilms. The LytC lysozyme is a member of the choline-binding family of proteins (CBPs) located on the pneumococcal surface. The present work shows that other CBPs, i.e. LytA, LytB, Pce, PspC and CbpF, which have a pI between 5 and 6, can bind DNA in vitro. This process requires the presence of divalent cations other than Mg2+. This DNA binding capacity of CBPs appears to be independent of their enzymatic activity and, at least in the case of LytA, does not require the choline-binding domain characteristic of CBPs. Positively charged, surface-exposed, 25 amino acid-long peptides derived from the catalytic domain of LytB, were also found capable of DNA binding through electrostatic interactions. Confocal laser scanning microcopy revealed the existence of cell-associated LytB−eDNA complexes in Streptococcus pneumoniae biofilms. These and other findings suggest that these surface-located proteins of S. pneumoniae could play roles of varying importance in the colonization and/or invasion of human host where different environmental conditions exist.Item Ecophysiological characterization of Penicillium expansum population in lleida (Spain)(2008) Morales, Hector; Marín, Sonia; Obea, Laura; Domenech Lucas, Miriam; Patiño Álvarez, Aurora Belén; Ramos, Antonio J.; Sanchis, VicentePenicillium expansum, a patulin producer fungus, is the most important fungus causing decay in cold stored both apples and pears. This can lead to patulin contaminated by-products. The aim of this assay was to evaluate the phenotypical and physiological variability in the population of P. expansum that cause fruit spoilage in post-harvest stages in Lleida (Spain). In total, 101 isolates of P. expansum from the 2004 and the 2005 seasons were obtained from decayed fruits. Significant differences were found in the observations from both seasons. Variability of the isolates in each season seemed to be partially explained by differences in growth in media, patulin accumulation and resistance to fungicides. Patulin production was detected in almost 100% of the isolates. Variability existing in P. expansum population could not be totally explained, but the above mentioned variables explained up to 74% of the diversity in some cases. The results obtained point to the existence of different populations of P. expansum in each season and may explain the differences in fungicide resistance observed between both seasons. The capacity to colonize apple flesh and some variables involved in fruit colonization were not a source of variation neither in each season nor when both seasons were compared. As storage rooms are cleaned and disinfected each season, this suggests that each season, the populations in storage rooms develop only from strains capable to colonize apple flesh. This may lead to rapid sporulation and spreading of spores.Item Versatility of the capsular genes during biofilm formation by Streptococcus pneumoniae(Environmental Microbiology, 2009) Domenech Lucas, Miriam; Ernesto García; Miriam MoscosoStreptococcus pneumoniae forms part of the natural microbiota of the nasopharynx. For the pneumococcus to cause infection, colonization needs to occur and this process is mediated by adherence of bacteria to the respiratory epithelium. Although the capsular polysaccharide (CPS) of S. pneumoniae is known to be important for infection to occur, its role in colonization is controversial. Biofilm models are starting to emerge as a promising tool to investigate the role of CPS during nasopharyngeal carriage, which is the first step in the dissemination and initiation of a pneumococcal infection. Using a well-defined model system to analyse in vitro biofilm formation in pneumococcus, here we explore the molecular changes underlying the appearance of capsular mutants using type 3 S. pneumoniae cells. Spontaneous colony phase variants show promoter mutations, as well as duplications, deletions and point mutations in the cap3A gene, which codes for a UDP-glucose dehydrogenase (UDP-GlcDH). Increased biofilm-forming capacity could usually be correlated with a reduction both in colony size and in the relative amount of CPS present on the cell surface of each colony variant. However, a mutation in Cap3A Thr83Ile (a strictly conserved residue in bacterial UDP-GlcDHs) that resulted in very low CPS production also led to impaired biofilm formation. We propose that non-encapsulated mutants of pneumococcal type 3 strains are essentially involved in the initial stages (the attachment stage) of biofilm formation during colonization/pathogenesis.Item A national longitudinal study evaluating the activity of cefditoren and other antibiotics against non-susceptible Streptococcus pneumoniae strains during the period 2004–20 in Spain(Journal of Antimicrobial Chemotherapy, 2022) Sempere, Julio; González-Camacho, Fernando; Domenech Lucas, Miriam; Llamosí, Mirella; Del Río, Idoia; López-Ruiz, Beatriz; Gimeno, Mercedes; Coronel, Pilar; Yuste, JoséBackground Surveillance studies including antibiotic resistance and evolution of pneumococcal serotypes are critical to evaluate the susceptibility of commonly used antibiotics and the contribution of conjugate vaccines against resistant strains. Objectives To determine the susceptibility of clinical isolates of Streptococcus pneumoniae with reduced susceptibility to penicillin to a panel of antibiotics during the period 2004–20 and characterize the impact of pneumococcal conjugate vaccines in the evolution of resistant serotypes. Methods We selected 3017 clinical isolates in order to determine the minimal inhibitory concentration to penicillin, amoxicillin, cefotaxime, erythromycin, levofloxacin and oral cephalosporins, including cefditoren, cefixime and cefpodoxime. Results The antibiotics with the lowest proportion of resistant strains from 2004 to 2020 were cefditoren (<0.4%), followed by cefotaxime (<5%), penicillin (<6.5%) and levofloxacin (<7%). Among oral cephalosporins, cefixime was the cephalosporin with the highest MIC90 (32 mg/L) and MIC50 (8–16 mg/L) throughout the study, followed by cefpodoxime with highest values of MIC90 (4 mg/L) and MIC50 (2 mg/L) for the majority of the study period. In contrast, cefditoren was the cephalosporin with the lowest MIC90 (1 mg/L) and MIC50 (0.25–0.5 mg/L). Conclusions Cefditoren was the antibiotic with the highest proportion of susceptible strains. Hence, more than 80% of the clinical strains were susceptible to cefditoren throughout the period 2004–20. The proportion of resistant isolates to cefditoren and cefotaxime was scarce, being less than 0.4% for cefditoren and lower than 5% for cefotaxime, despite the increased rates of serotypes not covered by the 13-valent pneumococcal conjugate vaccine.Item Biofilm Formation Avoids Complement Immunity and Phagocytosis of Streptococcus pneumoniae(Infection and Immunity, 2013) Domenech Lucas, Miriam; Ramos-Sevillano, Elisa; García, Ernesto; Moscoso, Miriam; Yuste, JoseStreptococcus pneumoniae is a frequent member of the microbiota of the human nasopharynx. Colonization of the nasopharyngeal tract is a first and necessary step in the infectious process and often involves the formation of sessile microbial communities by this human pathogen. The ability to grow and persist as biofilms is an advantage for many microorganisms, because biofilm-grown bacteria show reduced susceptibility to antimicrobial agents and hinder recognition by the immune system. The extent of host protection against biofilm-related pneumococcal disease has not been determined yet. Using pneumococcal strains growing as planktonic cultures or as biofilms, we have investigated the recognition of S. pneumoniae by the complement system and its interactions with human neutrophils. Deposition of C3b, the key complement component, was impaired on S. pneumoniae biofilms. In addition, binding of C-reactive protein and the complement component C1q to the pneumococcal surface was reduced in biofilm bacteria, demonstrating that pneumococcal biofilms avoid the activation of the classical complement pathway. In addition, recruitment of factor H, the downregulator of the alternative pathway, was enhanced by S. pneumoniae growing as biofilms. Our results also show that biofilm formation diverts the alternative complement pathway activation by a PspC-mediated mechanism. Furthermore, phagocytosis of pneumococcal biofilms was also impaired. The present study confirms that biofilm formation in S. pneumoniae is an efficient means of evading both the classical and the PspC-dependent alternative complement pathways the host immune system.Item Nationwide Trends of Invasive Pneumococcal Disease in Spain From 2009 Through 2019 in Children and Adults During the Pneumococcal Conjugate Vaccine Era(Clinical Infectious Diseases, 2020) Miguel, Sara de; González-Camacho, Fernando; Domenech Lucas, Miriam; Sempere, Julio; Vicioso, Dolores; Sanz, Juan Carlos; García Comas, Luis; Ardanuy, Carmen; Fenoll, Asunción; Yuste, JoséBackground: Introduction of pneumococcal conjugate vaccines (PCVs) has reduced the disease caused by vaccine serotypes in children, providing herd protection to adults. However, the emergence of nonvaccine serotypes is of great concern worldwide. Methods: This study includes national laboratory data from invasive pneumococcal disease (IPD) cases that affected pediatric and adult populations during 2009–2019. The impact of implementing different vaccine strategies for immunocompetent adults by comparing Spanish regions that used the 13-valent PCV (PCV13) vs regions that used the 23-valent pneumococcal polysaccharide vaccine (PPV23) was also analyzed for 2017−2019. Results: The overall reductions in IPD cases by PCV13 serotypes in children and adults were 88% and 59%, respectively, during 2009–2019, with a constant increase in serotype 8 in adults since 2015. IPD cases by additional serotypes covered by PPV23 increased from 20% in 2009 to 52% in 2019. In children, serotype 24F was the most frequent in 2019, whereas serotypes 3 and 8 accounted for 36% of IPD cases in adults. Introduction of PCV13 or PPV23 in the adult calendar of certain Spanish regions reduced the IPD cases by PCV13 serotypes by up to 25% and 11%, respectively, showing a decrease of serotype 3 when PCV13 was used. Conclusions: Use of PCV13 in children has affected the epidemiology, reducing the burden of IPD in children but also in adults by herd protection; however, the increase in serotype 8 in adults is worrisome. Vaccination with PCV13 in adults seems to control IPD cases by PCV13 serotypes including serotype 3.Item In Vitro Destruction of Streptococcus pneumoniae Biofilms with Bacterial and Phage Peptidoglycan Hydrolases(Antimicrobial Agents and Chemotherapy, 2011) Domenech Lucas, Miriam; García, Ernesto; Moscoso, MiriamHost- and phage-coded cell wall hydrolases have been used to fight Streptococcus pneumoniae growing as planktonic cells in vitro as well as in animal models. Until now, however, the usefulness of these enzymes in biofilm-grown pneumococci has gone untested. The antipneumococcal activity of different cell wall hydrolases produced by S. pneumoniae and a number of its phages was examined in an in vitro biofilm model. The major pneumococcal autolysin LytA, an N-acetylmuramoyl-l-alanine amidase, showed the greatest efficiency in disintegrating S. pneumoniae biofilms. The phage-encoded lysozymes Cpl-1 and Cpl-7 were also very efficient. Biofilms formed by the close pneumococcal relatives Streptococcus pseudopneumoniae and Streptococcus oralis were also destroyed by the phage endolysins but not by the S. pneumoniae autolysin LytA. A cooperative effect of LytA and Cpl-1 in the disintegration of S. pneumoniae biofilms was recorded.