Person: Sevillano Fernández, David
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First Name
David
Last Name
Sevillano Fernández
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Medicina
Department
Medicina
Area
Microbiología
Identifiers
36 results
Search Results
Now showing 1 - 10 of 36
Item Enhanced In Vivo Activity of Cefditoren in Pre-Immunized Mice against Penicillin-Resistant S. pneumoniae (Serotypes 6B, 19F and 23F) in a Sepsis Model(PLoS One, 2010) Cafini, Fabio; Yuste, Jose; Giménez, María José; Sevillano Fernández, David; Aguilar, Lorenzo; Alou Cervera, Luis; Ramos Sevillano, Elisa; Torrico, Martha; González Hidalgo, Natalia; García, Ernesto; Coronel, Pilar; Prieto Prieto, José; Michael OttoBackground: Specific antibodies are likely to be present before S. pneumoniae infection. We explored cefditoren (CDN) total and free values of serum concentrations exceeding the MIC (t>MIC) related to efficacy in a mice sepsis model, and the effect of specific gammaglobulins on in-vitro phagocytosis and in-vivo efficacy. Methodology/principal findings: We used three pneumococcal isolates (serotype, MIC OF CDN): Strain 1 (6B, 1 microg/ml), Strain 2 (19F, 2 microg/ml) and Strain 3 (23F, 4 microg/ml). Hyperimmune serum (HS) was obtained from mice immunized with heat-inactivated strains. In-vitro, phagocytosis by HS diluted 1/10 in presence/absence of sub-inhibitory concentrations was measured by flow cytometry including fluorescent bacteria and a neutrophil cell line. In-vivo dose-ranging experiments with HS (dilutions 1/2-1/16) and CDN (6.25 mg/kg-100 mg/kg tid for 48 h) were performed to determine the minimal protective dilution/dose (highest survival) and the non-protective highest dilution/dose (highest mortality: HS-np dilution and CDN-np dose) over 7 days. Efficacy of CDN-np in animals pre-immunized with HS-np (combined strategy) was explored and blood bacterial clearance determined. The CDN measured protein binding was 86.9%. In-vitro, CDN significantly increased phagocytosis (vs. HS 1/10). In non pre-immunized animals, t>MIC values for CDN of approximately 35% (total) and approximately 19% (free) were associated with 100% survival. Significant differences in survival were found between HS-np alone (< or = 20%) or CDN-np alone (< or = 20%) vs. the combined strategy (90%, 60% and 60% for Stains 1, 2 and 3), with t>MIC (total/free) of 22.8%/14.3%, 26.8%/16.0%, and 22.4%/12.7% for Strains 1, 2 and 3, respectively. Prior to the second dose (8 h), median bacterial counts were significantly lower in animals surviving vs. dead at day 7. Conclusions/significance: In mice (CDN protein binding similar to humans) total t>MIC values of approximately 35% (approximately 19% free) were efficacious, with a decrease in the required values in pre-immunized animals. This reinforces that immunoprotection to overcome resistance may provide lifesaving strategies.Item Efficacy of simulated cefditoren versus amoxicillin-clavulanate free concentrations in countering intrastrain ftsI gene diffusion in Haemophilus influenzae(Antimicrobial Agents and Chemotherapy, 2011) González Hidalgo, Natalia; Aguilar, Lorenzo; Sevillano Fernández, David; Giménez, María José; Alou Cervera, Luis; Cafini, Fabio; Torrico, Martha; López, Ana María; Coronel, Pilar; Prieto Prieto, JoséThis study explores the effects of cefditoren (CDN) versus amoxicillin-clavulanic acid (AMC) on the evolution (within a single strain) of total and recombined populations derived from intrastrain ftsI gene diffusion in β-lactamase-positive (BL+) and β-lactamase-negative (BL−) Haemophilus influenzae. DNA from β-lactamase-negative, ampicillin-resistant (BLNAR) isolates (DNABLNAR) and from β-lactamase-positive, amoxicillin-clavulanate-resistant (BLPACR) (DNABLPACR) isolates was extracted and added to a 107-CFU/ml suspension of one BL+ strain (CDN MIC, 0.007 μg/ml; AMC MIC, 1 μg/ml) or one BL− strain (CDN MIC, 0.015 μg/ml; AMC MIC, 0.5 μg/ml) in Haemophilus Test Medium (HTM). The mixture was incubated for 3 h and was then inoculated into a two-compartment computerized device simulating free concentrations of CDN (400 mg twice a day [b.i.d.]) or AMC (875 and 125 mg three times a day [t.i.d.]) in serum over 24 h. Controls were antibiotic-free simulations. Colony counts were performed; the total population and the recombined population were differentiated; and postsimulation MICs were determined. At time zero, the recombined population was 0.00095% of the total population. In controls, the BL− and BL+ total populations and the BL− recombined population increased (from ≈3 log10 to 4.5 to 5 log10), while the BL+ recombined population was maintained in simulations with DNABLPACR and was decreased by ≈2 log10 with DNABLNAR. CDN was bactericidal (percentage of the dosing interval for which experimental antibiotic concentrations exceeded the MIC [ft>MIC], >88%), and no recombined populations were detected from 4 h on. AMC was bactericidal against BL− strains (ft>MIC, 74.0%) in DNABLNAR and DNABLPACR simulations, with a small final recombined population (MIC, 4 μg/ml; ft>MIC, 30.7%) in DNABLPACR simulations. When AMC was used against the BL+ strain (in DNABLNAR or DNABLPACR simulations), the bacterial load was reduced ≈2 log10 (ft>MIC, 44.3%), but 6.3% and 32% of the total population corresponded to a recombined population (MIC, 16 μg/ml; ft>MIC, 0%) in DNABLNAR and DNABLPACR simulations, respectively. AMC, but not CDN, unmasked BL+ recombined populations obtained by transformation. ft>MIC values higher than those classically considered for bacteriological response are needed to counter intrastrain ftsI gene diffusion by covering recombined populations.Item Evaluating the optimal time for amikacin administration with respect to haemodialysis using an in vitro pharmacodynamic simulation against epidemic nosocomial OXA-48 producing Klebsiella pneumoniae ST405 strains(Journal of Global Antimicrobial Resistance, 2019) Carcas, Antonio J.; Sevillano Fernández, David; González Hidalgo, Natalia; Alou Cervera, Luis; Gómez Gil, Rosa; Muñoz, Mario; Llanos, Lucía; Sánchez Villanueva, Rafael J.; Gonzalez Parra, Emilio; Giménez, María José; Aguilar, LorenzoObjectives: Bacterial viability and enrichment of resistance resulting from three different amikacin administration schedules with respect to haemodialysis (HD) were assessed against three OXA-48-producing Klebsiella pneumoniae isolated during an outbreak in a Spanish hospital. Methods: A previously described two-compartment dynamic system was used. Three possible amikacin administration schedules were simulated: (i) haemodialysis immediately after amikacin infusion (pre-HD); (ii) infusion immediately after haemodialysis (post-HD); and (iii) infusion at 50% interdialytic period. Amikacin standard dose (SD) and double dose (DD) were simulated for each schedule. Values of Cmax/MIC, Cmax/MPC (mutant prevention concentration), AUC0-48h/MIC, AUC0-48h/MPC and %TMSW (percentage of time that the concentration was inside the mutant selection window) were determined with experimental data and were correlated with the area under the bacterial killing curve of the total population and the resistant subpopulation. Results: Both with SD and DD, the pre-HD schedule resulted in increases at 48h in bacterial counts of the total population at the expense of enrichment of pre-existing resistant subpopulations from 12h onwards for all strains. The estimated %TMSW that prevented enrichment of resistant mutants was <61.5%. The AUC0-48h/MPC (with values of ≈40 being associated with countering of increases in resistant subpopulations) was better than the %TMSW as a predictive parameter. Conclusion: This study showed that the longest times concentrations were above the MPC (i.e. highest AUC0-48h/MPC, lowest %TMSW), the lowest enrichment of resistant subpopulations. This implies use of the highest possible amikacin dose (limited by toxicity) and post-HD as the best administration schedule.Item Activity of simulated serum concentrations of daptomycin versus vancomycin during the first 24h of treatment in the presence of physiological albumin concentrations against vancomycin-susceptible, -tolerant or -intermediate-resistant Staphylococcus aureus(International Journal of Antimicrobial Agents, 2011) Torrico, Martha; Aguilar, Lorenzo; Sevillano Fernández, David; Giménez, María José; Alou Cervera, Luis; González Hidalgo, Natalia; Cafini, Fabio; Cleeland, Roy; Prieto Prieto, JoséIn order to determine whether reduced susceptibility or tolerance to vancomycin in Staphylococcus aureus influences the activity of daptomycin by simulating serum concentrations in the first 24h of treatment in the presence of physiological concentrations of human albumin, a computerised pharmacodynamic simulation was performed using Mueller-Hinton broth with 4 g/dL human albumin concentrations. For daptomycin, the media was adjusted to physiological ionised calcium concentrations by adding 100 μg/mL Ca(2+). Protein binding was measured. Six S. aureus isolates were used, comprising one vancomycin-susceptible S. aureus (VSSA), three vancomycin-tolerant strains, one heteroresistant vancomycin-intermediate S. aureus (hVISA) and one homogeneous vancomycin-intermediate S. aureus (VISA). Minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of daptomycin increased eight times when determined in the presence of albumin (MIC(ALB) and MBC(ALB), respectively). Measured protein binding was 86.6% (C(max)) and 86.5% (C(min)) for daptomycin and 51.6% (C(max)) and 42.2% (C(min)) for vancomycin. Similar values were obtained for fAUC/MIC (where fAUC is the area under the concentration-time curve obtained with extrapolated concentrations using the highest protein binding rate experimentally obtained) and AUC/MIC(ALB) for each antibiotic. Daptomycin showed early (≤ 6 h) bactericidal activity [maximal effect (E(max)) >4 log(10) reductions in initial inocula] against all strains. Vancomycin produced an E(max) of 2.3 log(10) reductions at 8h against the VSSA and reductions ≤1.8 log(10) for the other strains in the 8-24h period. Pharmacodynamic parameters were fAUC/MBC from 8.0 to 15.6 (vancomycin) and from 56.0 to 111.6 (daptomycin) for tolerant strains, and fAUC/MIC of 126.8 and 63.3 for vancomycin and 222.6 and 113.2 for daptomycin against hVISA and VISA strains, respectively. Against the study strains (vancomycin-susceptible, -tolerant, heteroresistant or intermediate), daptomycin, in contrast to vancomycin, exhibited early bactericidal activity despite its high protein binding.Item Influence of the MBC/MIC ratio on the antibacterial activity of vancomycin versus linezolid against methicillin-resistant Staphylococcus aureus isolates in a pharmacodynamic model simulating serum and soft tissue interstitial fluid concentrations reported in diabetic patients(Journal of Antimicrobial Chemotherapy, 2013) González Hidalgo, Natalia; Sevillano Fernández, David; Alou Cervera, Luis; Cafini, Fabio; Gimenez, María José; Gómez-Lus Centelles, María Luisa; Prieto Prieto, José; Aguilar, LorenzoObjectives: To explore serum and tissue pharmacodynamics of linezolid versus vancomycin against methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates with different MBC/MIC ratios. Methods: Five strains (vancomycin MIC/MBCs, mg/L) were used: TOL-1 (2/≥64), TOL-2 (1/16), LT-1 and LT-2 (1/8) and NT (1/2). The linezolid MIC/MBC for all strains was 2/≥64 mg/L. A two-compartment dynamic computerized device was used (inocula 10(7) cfu/mL). Free concentrations obtained in serum and interstitial fluid with twice-daily regimens of 1 g of vancomycin or 600 mg of linezolid were simulated over 48 h. ABBCs (differences between control growth curves and killing curves of bacteria exposed to antibiotics; log10 cfu × h/mL) and log10 reductions in initial inocula were calculated. Results: In serum simulations, vancomycin (AUC0-24/MIC = 251.8 for TOL-1 and 503.6 for the remaining strains) was bacteriostatic against strains with MBC/MIC ≥8, but bactericidal against NT. In interstitial fluid simulations (AUC0-24/MIC = 54.6 for TOL-1 and 109.2 for the remaining strains), initial inocula grew in all cases. Linezolid, both in serum (AUC0-24/MIC = 87.0) and in interstitial fluid (AUC0-24/MIC = 130.6) simulations, reduced initial inocula ≥2.2 log10 for all strains (apart from LT-1 in serum simulations that showed a bacteriostatic profile). ABBCs were similar in serum and interstitial fluid with linezolid, but significantly lower in interstitial fluid simulations with vancomycin. Conclusions: From the pharmacodynamic perspective (serum concentrations), vancomycin tolerance should include MBC/MIC ≥8 since strains exhibiting this ratio showed bacteriostatic profiles similar to those obtained with isolates with MBC/MIC ratios of 16 or 32. Insufficient concentrations of vancomycin at the simulated infected site were linked to bacteriological failure. Free concentrations of linezolid at the infection site pharmacodynamically covered MRSA.Item Protection of Enterococcus faecalis in mixed cultures with carbapenemase-producing Escherichia coli and Bacteroide fragilis: effect of the bacterial load(Revista Española de Quimioterapia, 2013) Sevillano Fernández, David; Aguilar, Lorenzo; Alou Cervera, Luis; Gímenez, María José; Cafini, Fabio; González Hidalgo, Natalia; Prieto Prieto, JoséIntroduction: This study explores effects of pH and inoculum size on imipenem versus tigecycline activity against E. coli, B. fragilis and E. faecalis, both in individual and mixed cultures. Methods: MIC/MBCs (mg/L) of tigecycline and imipenem were 0.12/≥ 16 and 4/4 for E. coli, 0.12/0.5 and ≥ 16/≥ 16 for B. fragilis, and 0.12/≥ 16 and 2/≥ 16 for E. faecalis, respectively. Killing curves in supplemented Brucella broth were performed at pH 7 or 5.8, with two final inocula (≈ 105 or ≈ 107 cfu/ml) of each isolate (individual cultures) and with 1:1:1 mixed inocula. Tubes were 48 h incubated at 37 ºC in anaerobiosis. Final concentrations (estimated concentrations in colon) were 1.50 mg/L for tigecycline and 26.40 mg/L for imipenem, with antibiotic-free curves as controls. Experiments were performed in triplicate. Results: Imipenem showed inoculum effect against E.coli and B. fragilis, with reductions in initial inocula in experiments with standard inocula contrasting with increases in experiments with high inocula (both individual and mixed cultures). Against E. faecalis no inoculum effect for imipenem was observed in individual cultures, with marked reductions in initial inocula regardless inoculum size. However in mixed experiments the indirect protection of E. faecalis by the two gramnegatives resulted in bacterial regrowth. This protection was inoculum-dependant since it occurred with high but not with standard inocula. Tigecycline reduced initial inocula of the three isolates regardless culture type (individual/mixed) or experimental conditions (pH/inocula size), with lower reductions for the tolerant E. faecalis. Conclusion: Carbapenemase activity was inoculum-dependant for self-protection and indirect protection of E. faecalis.Item In Vitro Killing Activity of Crevicular Concentrations of Tinidazole Plus Common Oral Antibiotics Against High‐Density Mixed Inocula of Periodontal Pathogens in Strict Anaerobic Conditions(Journal of Periodontology, 2010) Alou Cervera, Luis; Giménez, María José; Manso, Francisco; Sevillano Fernández, David; Cafini, Fabio; Torrico, Martha; González Hidalgo, Natalia; Prieto Prieto, José; Alió, J.J.; Aguilar, LorenzoBackground: Odontogenic infections are polymicrobial. This study explores the in vitro killing activity by concentrations similar to those found in crevicular fluid of tinidazole in combination with amoxicillin/clavulanic acid, clindamycin and levofloxacin against four groups of high-density mixed inocula of anaerobes (Prevotella buccae, Fusobacterium nucleatum, and Veillonella spp.) and facultative (Capnocytophaga spp. and Streptococcus spp.) isolates of periodontal pathogens. Methods: Killing curves were assessed under strict anaerobic conditions with antibiotics alone and in combination with tinidazole at concentrations similar to those achieved in crevicular fluid against ∼107 colony forming units (CFU)/ml inoculum (1:1:1:1:1 proportion of the five bacterial isolates) of the four bacterial groups. Group 1 did not include β-lactamase–producing strains; groups 2, 3, and 4 included one, two, and three β-lactamase–producing strains, respectively. Results: In single-drug experiments, at 48 hours, tinidazole alone did not show significant killing of the entire bacterial population, whereas reductions in the initial inocula ≥2.09 log10 CFU/ml with clindamycin, ≥3.26 log10 CFU/ml with amoxicillin/clavulanic acid, and ≥3.83 log10 CFU/ml with levofloxacin were obtained. When combined with tinidazole, reductions were significantly higher for all antibiotics: ≥5.28 log10 CFU/ml with clindamycin, ≥4.78 log10 CFU/ml with amoxicillin/clavulanic acid, and ≥6.17 log10 CFU/ml with levofloxacin. Conclusion: In addition to its high activity against anaerobic periodontal pathogens, tinidazole offered synergism with other antibiotics against the large strict anaerobic subpopulation and the small facultative subpopulation of a high-density mixed inocula of odontogenic pathogens under strict anaerobic conditions, similar to those of odontogenic infections.Item Influence of Media and Testing Methodology on Susceptibility to Tigecycline of Enterobacteriaceae with Reported High Tigecycline MIC(Journal of Clinical Microbiology, 2010) Torrico, Martha; González Hidalgo, Natalia; Giménez, María José; Alou Cervera, Luis; Sevillano Fernández, David; Navarro, David; Díaz Antolín, María Paz; Larrosa, Nieves; Aguilar, Lorenzo; GarcÍa Escribano, NataliaThe tigecycline susceptibility of six different Enterobacteriaceae strains with reported high tigecycline MICs was determined in quintuplicate by four methodologies using Mueller-Hinton agar and broth from six manufacturers. The MICs determined by Etest were a ≥1-fold dilution lower than those determined by broth microdilution and agar dilution, with the highest modal values given by agar dilution. The highest modal MICs were obtained using Oxoid medium, and the lowest inhibition zone values (disc diffusion) were obtained using Oxoid and bioMérieux media. The lowest MICs were obtained by Etest using Difco or Merck media.Item In vitro effect of physiological concentrations of human albumin on the antibacterial activity of tigecycline(Journal of Antimicrobial Chemotherapy, 2009) Alou Cervera, Luis; Gimenez, María José; Cafini, Fabio; Aguilar, Lorenzo; Sevillano Fernández, David; González Hidalgo, Natalia; Torrico, Martha; Prieto Prieto, José; Garcia-Rey, César; Garcia-Escribano, NuriaObjectives: To determine C(max) tigecycline activity in the presence/absence of physiological concentrations of human albumin with free fraction concentrations as controls. Methods: Killing curves (final inoculum: 1.0-5.0 x 10(7) cfu/mL) were performed with 0.88 mg/L final concentrations (serum C(max) after a 100 mg 1 h infusion) in Mueller-Hinton broth supplemented with Ca(2+) and Mg(2+) (MH) and in MH with 4 g/dL human albumin. Controls were curves in MH with concentrations similar to the free fraction (fC(max) = 0.17 mg/L) calculated using protein binding. Activity was measured as log(10) initial inoculum reduction (log(10) initial inoculum-log(10) at 12 h/24 h). Target strains (tigecycline MIC/MBC; mg/L) were: methicillin-resistant Staphylococcus aureus heteroresistant to vancomycin (0.12/0.25); Enterococcus faecium (0.12/0.25); Escherichia coli producing extended-spectrum beta-lactamase (0.12/0.25); and Acinetobacter baumannii (0.25/1). Results: At 24 h the fC(max) produced mean decreases of < or =0.1 cfu/mL for all strains, in contrast to the bactericidal activity (mean >3 log(10) reduction) provided by C(max) concentrations in the presence or absence of albumin for E. coli and E. faecium, and an activity nearly bactericidal for S. aureus (mean approximately 2.8 log(10) reduction). In the case of the A. baumannii isolate the C(max) in the presence or absence of albumin produced a mean reduction of 2.56 log(10) cfu/mL at 12 h (time of one dosing interval), with a bacteriostatic profile when considering 24 h colony counts (similar counts at 0 and 24 h). Conclusions: Correcting the total concentration for the reported literature binding values is unreliable since tigecycline antibacterial activity was greater than that suggested by the free fraction of the drug.Item Activity of serum concentrations of daptomycin vs. vancomycin against vancomycin-susceptible and resistant Enterococcus faecium in the presence of albumin physiological concentrations: an in vitro pharmacodynamic simulation(European Journal of Clinical Microbiology & Infectious Diseases, 2008) Alou Cervera, Luis; Aguilar, Lorenzo; Giménez, María José; Torrico, Martha; Sevillano Fernández, David