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 Effects of human albumin and serum on the in vitro bactericidal activity of cefditoren against penicillin-resistant Streptococcus pneumoniae(Journal of Antimicrobial Chemotherapy, 2007) Sevillano Fernández, David; Giménez, María José; Alou Cervera, Luis; Aguilar, Lorenzo; Cafini, Fabio; Torrico, Martha; González Hidalgo, Natalia; Echeverría, Olatz; Coronel, Pilar; Prieto Prieto, JoséObjectives: Attempts to interpret antibiotic pharmacodynamics using reported protein binding may underestimate true activity. To elucidate this issue we examined bacterial killing kinetics at cefditoren concentrations equal to C(max) in the presence of 90% human serum or albumin at physiological concentrations. Methods: Killing curves (final inocula of approximately 10(7) cfu/mL, cefditoren concentration of 4.2 mg/L) were performed against Streptococcus pneumoniae strains exhibiting cefditoren MICs (mg/L) of 0.12 (strain 1), 0.25 (strain 2) and 0.5 (strain 3) in different media: (i) C(max)-MH, Mueller-Hinton broth plus 5% lysed horse blood (MH), (ii) C(max)-HS, MH broth with a final human serum concentration of 90%; and (iii) C(max)-HAlb, MH broth with 4 g/dL human albumin. Killing curves were also performed with a final cefditoren concentration of 0.5 mg/L (similar to free-drug C(max) considering 88% protein binding) in MH broth (12% C(max)). Results: No significant differences were found between the different media or concentrations with strain 1 (log(10) reductions >or=4 at 24 h). Against strains 2 and 3, we observed significantly higher initial inocula decreases at 24 h for C(max)-HS as compared with C(max)-HAlb or 12% C(max). Bactericidal activity (>or=3 log(10) reduction) was obtained at 24 h against the three strains only with C(max)-HS and C(max)-MH. Conclusions: The presence of physiological concentrations of human albumin, but not 90% human serum, limited bactericidal activity as did the use of concentrations similar to free-drug C(max), suggesting that extrapolation of active drug from total drug by using the protein binding rate is not an accurate method to study antibacterial activity.Item Is there a pharmacodynamic need for the use of continuous versus intermittent infusion with ceftazidime against Pseudomonas aeruginosa? An in vitro pharmacodynamic model(Journal of Antimicrobial Chemotherapy, 2005) Alou Cervera, Luis; Aguilar, Lorenzo; Sevillano Fernández, David; Giménez, María José; Echeverría, Olatz; Gómez-Lus Centelles, María Luisa; Prieto Prieto, JoséObjectives: In order to explore the pharmacodynamic need for continuous versus intermittent (three times a day) administration of ceftazidime in critically ill patients, a pharmacokinetic computerized device was used to simulate concentrations of ceftazidime in human serum after 6 g/day. Methods: Efficacy was measured as the capability of simulated concentrations over time to reduce initial inoculum against four strains of Pseudomonas aeruginosa. MICs of the strains matched NCCLS breakpoints: one susceptible strain (MIC = 8 mg/L), two intermediate strains (MIC = 16 mg/L) and one resistant strain (MIC = 32 mg/L). C(max) was 119.97+/-2.53 mg/L for intermittent bolus and C(ss) (steady-state concentration) was 40.38+/-0.16 mg/L for continuous infusion. AUC(0-24) was similar for both regimens (approximately 950 mg x h/L). Inhibitory quotients were three times higher for the intermittent administration whereas t > MIC was higher for continuous infusion (100%) versus intermittent administration (99.8%, 69% and 47.6% for the susceptible, intermediate and resistant strains, respectively). Results: Against the susceptible and intermediate strains, no differences were found between both regimens with > or = 3 log10 reduction from 8 to 24 h. Against the resistant strain, only the continuous infusion achieved this bactericidal activity in the same time period, minimizing the differences between resistant and susceptible strains. Significantly higher initial inoculum reduction at 32 h was obtained for the continuous versus the intermittent administration (83.35% versus 38.40%, respectively). Conclusions: These results stress the importance of optimizing t >MIC, even at peri-MIC concentrations, of ceftazidime against resistant strains. Local prevalence of resistance justifies, on a pharmacodynamic basis, electing for continuous infusion versus intermittent administration.Item β-Lactam Activity against Penicillin-Resistant Streptococcus pneumoniae Strains Exhibiting Higher Amoxicillin versus Penicillin Minimum Inhibitory Concentration Values: An in vitro Pharmacodynamic Simulation(Chemotherapy, 2008) Sevillano Fernández, David; Aguilar, Lorenzo; Alou Cervera, Luis; Giménez, María José; González Hidalgo, Natalia; Echeverría, Olatz; Torrico, Martha; Valdes, Laura; Coronel, Pilar; Prieto Prieto, JoséBackground: Activity of simulated serum concentrations after oral therapy with 400 mg cefditoren pivoxil b.i.d., 500 mg cefuroxime axetil b.i.d. and 875/125 mg amoxicillin/clavulanic acid b.i.d. and t.i.d. regimens was explored over 24 h against Streptococcus pneumoniae.Methods: Computerized pharmacodynamic simulations were performed against strains with penicillin/amoxicillin/cefuroxime/cefditoren minimum inhibitory concentrations (MICs, µg/ml) and serotypes: strain 1 (0.25/0.12/1/0.12; serotype 6A), strain 2 (2/4/ 2/0.25; serotype 6B), strain 3 (4/16/4/0.5; serotype 14), and strain 4 (4/16/8/1; serotype 14). Results: Bactericidal activity (≧3 log10 reduction) at 12 and 24 h was obtained against all strains with cefditoren, against strains 1 and 2 with cefuroxime and amoxicillin/clavulanic acid t.i.d., but only against strain 1 with amoxicillin/clavulanic acid b.i.d.. Bactericidal activity at 24 h was related to T > MIC of >30% dosing interval, 1.7–2.0 log10 reductions with T > MIC of 20–30%, and <1 log10 reduction or regrowth with T > MIC of 0%. Conclusions: It is difficult to achieve pharmacodynamic coverage and bactericidal activity by physiological concentrations of oral β-lactams against penicillin-resistant pneumococcal strains exhibiting higher amoxicillin versus penicillin MICs. Cefditoren may offer alternatives.Item Bactericidal activity of moxifloxacin against multidrug-resistant Streptoccocus pneumoniae at clinically achievable serum and epithelial lining fluid concentrations compared with three other antimicrobials(International Journal of Antimicrobial Agents, 2004) Cafini, Fabio; Alou Cervera, Luis; Sevillano Fernández, David; Valero, Eva; Prieto Prieto, JoséTime-kill studies compared the activities of moxifloxacin with those of levofloxacin, azithromycin and amoxicillin-clavulanate against 10 pneumococcal strains with various drug susceptibilities. Three Streptococcus pneumoniae strains were resistant to moxifloxacin: 6, 7 and 2 strains were resistant to levofloxacin, azithromycin and amoxicillin-clavulanate, respectively. Of these, 1 strain was resistant to all antimicrobial agents studied. Moxifloxacin and amoxicillin-clavulanate were bactericidal after 24h at serum Cmax levels against 9 and 8 strains, respectively, while levofloxacin and azithromycin were bactericidal against 3 and 2 strains, respectively. A higher activity was only observed for amoxicillin-clavulanate for logarithmic phase cultures at 1, 4 and 8h compared with stationary phase organisms. Amoxicillin-clavulanate and moxifloxacin were bactericidal at free serum levels (protein unbound) after 24h against 8 and 3 strains, respectively. Moxifloxacin was bactericidal at epithelial lining fluid levels against all strains at 24h, including one moxifloxacin, amoxicillin-clavulanate and azithromycin-resistant strain; lower levels of bactericidal activity was observed for levofloxacin, azithromycin and amoxicillin-clavulanate against 7, 2 and 4 strains, respectively. This demonstrated the importance of moxifloxacin tissue levels.Item Decrease in Bacterial Load Versus Resistance Selection of Pneumococcal Subpopulations by β-Lactam Physiological Concentrations over Time: An In Vitro Pharmacodynamic Simulation(Microbial Drug Resistance, 2008) Cafini, Fabio; Aguilar, Lorenzo; Sevillano Fernández, David; Giménez, María José; Alou Cervera, Luis; Fenoll, Asunción; Echevarría, Olatz; Torrico, Martha; González Hidalgo, Natalia; Coronel, Pilar; Prieto Prieto, JoséTo investigate β-lactam effects on Streptococcus pneumoniae–mixed cultures, a computerized pharmacodynamic model simulating over 24-hr concentrations obtained after several β-lactam regimens was used. Strain 1 (no penicillin binding protein [PBP] mutations) and strain 2 (mutation in pbp1a) were penicillin/amoxicillin susceptible. Strain 3 (mutations in pbp1a, pbp2x, and pbp2b) and strain 4 (mutations in pbp1a, pbp2x, and pbp2b [10 changes]) were penicillin/amoxicillin resistant. Initial inoculum was approximately 6 × 106 CFU (colony forming units)/ml (with a 1:1:1:1 proportion of each strain). Population analysis profile was performed pre- and postsimulations. Strain 1 exhibited the best fitness (growth over 24 hr) in individual cultures, and strain 2 did so in mixed cultures in antibiotic-free simulations. In antibiotic simulations with the mixed inocula, penicillin/amoxicillin-susceptible strains were eradicated with all study drugs (time that concentrations exceed the minimal inhibitory concentration [T > MIC ≥ 43%]). Penicillin-resistant strains showed different evolution depending on the antibiotic: (a) cefditoren produced >2 log10 reduction of initial inocula at 12–24 hr (T>MIC ≥45%), with a remaining population growing in plates with ≥4 mg/L amoxicillin; (b) cefuroxime, cefixime, and cefaclor did not decrease initial inocula at 12–24 hr (T > MIC = 0%), with minor subpopulations growing in plates with 4 mg/L amoxicillin; (c) amoxicillin produced 2.6 log10 decrease of initial inocula at 12 hr (T>MIC = 47.5%), but 1.1 log10 increase of initial inocula at 24 hr, with a significant population growing in plates with 4 mg/L amoxicillin. Antibiotic activity against mixed inocula (susceptible and resistant strains) depends on intrinsic activity (as well as its subsequent pharmacodynamic activity: T > MIC against resistant strains), and on possible selection of intra-strain-resistant subpopulations.Item Evaluation of two in vitro pharmacodynamic simulation models: microfiltration versus centrifugation–filtration(International Journal of Antimicrobial Agents, 2001) Alou Cervera, Luis; Sevillano Fernández, David; Bugella, Javier H.; Fuentes, Fernando; Gómez-Lus Centelles, María Luisa; Prieto Prieto, JoséPharmacodynamic in vitro models that simulate serum antimicrobial concentrations provide more information about the activity of an antibiotic than MICs or traditional time-kill methods. The aim of this study was to compare two pharmacodynamic simulation models using ATCC strains of five different species and five antibiotics. In the first model (Centriprep-10 system), a filtration-centrifugation process was used to eliminate the antibiotic; in the second model (microfiltration system) no centrifugation was necessary. The antibiotic concentrations tested were similar to those in serum after normal doses of cefuroxime, clarithromycin, ciprofloxacin, gentamicin and cefotaxime. No significant differences were observed in the killing rates between the models except in the case of Haemophilus influenzae and cefotaxime. The new microfiltration model had the following advantages: lack of the carry-over effect, the absence of centrifugation that could damage bacteria and the possibility of increasing the number of incubation periods to give a better fit of the kinetic profile of man.Item Urine bactericidal activity against resistant Escherichia coli in an in vitro pharmacodynamic model simulating urine concentrations obtained after 2000/125 mg sustained-release co-amoxiclav and 400 mg norfloxacin administration(Journal of Antimicrobial Chemotherapy, 2006) Alou Cervera, Luis; Aguilar, Lorenzo; Sevillano Fernández, David; Giménez, María José; Cafini, Fabio; Valero, Eva; Relaño, María Teresa; Prieto Prieto, JoséObjectives: To explore the urine bactericidal activity of co-amoxiclav and norfloxacin against Escherichia coli in an in vitro pharmacodynamic model simulating the human urinary concentrations observed after administration of a single oral dose of 2000/125 mg sustained-release co-amoxiclav and 400 mg norfloxacin. Methods: Six E. coli isolates exhibiting amoxicillin/clavulanic acid MICs of 4/2 (two strains), 8/4, 16/8, 32/16 and 64/32 mg/L and norfloxacin MICs of < or =0.25 mg/L (three strains), 32, 64 and 256 mg/L were used. Colony counts were determined over 12 h and differences between the bacterial counts of initial inocula and the bacterial counts at each sampling time-point were calculated. Results: With co-amoxiclav, bactericidal activity (>3 log(10) reduction) was obtained against the susceptible (MIC < or = 8/4 mg/L) and intermediate (MIC = 16/8 mg/L) strains from 3 to 12 h, and from 3 to 10 h against the resistant strains (MIC > or = 32/16 mg/L), which exhibited a 2 log(10) reduction at 12 h. With norfloxacin, bactericidal activity was obtained against the susceptible strains from 4 to 12 h and from 8 to 12 h against the resistant strain with an MIC of 32 mg/L. Regrowth, with respect to initial inocula, occurred from 8 h onwards with the strain with MIC = 64 mg/L and from 3 h onwards with the strain with MIC = 256 mg/L. Conclusions: While regrowth occurs after exposure of high norfloxacin-resistant E. coli to urine physiological concentrations of norfloxacin, this study suggests that clavulanic acid can be given twice daily (to protect amoxicillin activity) with respect to uncomplicated cystitis due to E. coli exhibiting amoxicillin/clavulanic acid MICs up to 64/32 mg/L.Item β-Lactam Effects on Mixed Cultures of Common Respiratory Isolates as an Approach to Treatment Effects on Nasopharyngeal Bacterial Population Dynamics(PLOS ONE, 2008) Sevillano Fernández, David; Aguilar, Lorenzo; Alou Cervera, Luis; Giménez, María José; González Hidalgo, Natalia; Torrico, Martha; Cafini, Fabio; Coronel, Pilar; Prieto Prieto, José; Keertan DhedaBackground: Streptococcus pneumoniae, Streptococcus pyogenes and Haemophilus influenzae are bacteria present in the nasopharynx as part of normal flora. The ecological equilibrium in the nasopharynx can be disrupted by the presence of antibiotics. Methodology/principal findings: A computerized two-compartment pharmacodynamic model was used to explore beta-lactam effects on the evolution over time of a bacterial load containing common pharyngeal isolates by simulating free serum concentrations obtained with amoxicillin (AMX) 875 mg tid, amoxicillin/clavulanic acid (AMC) 875/125 mg tid and cefditoren (CDN) 400 mg bid regimens over 24 h. Strains and MICs (microg/ml) of AMX, AMC and CDN were: S. pyogenes (0.03, 0.03 and 0.015), S. pneumoniae (2, 2 and 0.25), a beta-lactamase positive H. influenzae (BL(+); >16, 2 and 0.06) and a beta-lactamase positive AMC-resistant H. influenzae (BLPACR, >16, 8 and 0.06). Mixture of identical 1:1:1:1 volumes of each bacterial suspension were prepared yielding an inocula of approximately 4 x 10(6) cfu/ml. Antibiotic concentrations were measured both in bacterial and in bacteria-free antibiotic simulations. beta-lactamase production decreased AMX concentrations and fT(>MIC) against S. pneumoniae (from 43.2% to 17.7%) or S. pyogenes (from 99.9% to 24.9%), and eradication was precluded. The presence of clavulanic acid countered this effect of co-pathogenicity, and S. pyogenes (but not BL(+) and S. pneumoniae) was eradicated. Resistance of CDN to TEM beta-lactamase avoided this co-pathogenicity effect, and CDN eradicated S. pyogenes and H. influenzae strains (fT(>MIC) >58%), and reduced in 94% S. pneumoniae counts (fT(>MIC) approximately 25%). Conclusions/significance: Co-pathogenicity seems to be gradual since clavulanic acid countered this effect for strains very susceptible to AMX as S. pyogenes but not for strains with AMX MIC values in the limit of susceptibility as S. pneumoniae. There is a potential therapeutic advantage for beta-lactamase resistant cephalosporins with high activity against streptococci.Item Effects of antimicrobials on the competitive growth of Streptococcus pneumoniae: a pharmacodynamic in vitro model approach to selection of resistant populations(Journal of Antimicrobial Chemotherapy, 2006) Sevillano Fernández, David; Aguilar, Lorenzo; Alou Cervera, Luis; Giménez, María José; Echevarría, Olatz; Cafini, Fabio; Valero, Eva; Fenoll, Asunción; Prieto Prieto, JoséObjectives: To investigate antimicrobial effects on a mixed culture of five Streptococcus pneumoniae serotypes (S) as an approach to ecology of population dynamics. Methods: A computerized pharmacodynamic model simulating concentrations obtained after levofloxacin, ciprofloxacin and azithromycin doses was used. Resistance patterns were S12, susceptible to study drugs; S31, low-level macrolide-resistant (efflux phenotype); S11, high-level macrolide-resistant (erm genotype); S9V, low-level quinolone-resistant; and S3, high-level quinolone-resistant. Initial mixed inocula (time 0) included similar percentages of each serotype. Results: At 24 h of control drug-free experiments, dominant strains were S9V (57.4%) and S12 (41.8%) with marginal populations of S31, S3 and S11. Azithromycin selected to a much higher extent the strain with low-level resistance to macrolides (S31) rather than the strain with high-level resistance (S11) (accounting for 99.9% versus 0.1% of the total population at 24 h). Ciprofloxacin selected to a higher extent low-level (S9V) rather than high-level (S3) quinolone resistance (72.4% versus 27.6%). Levofloxacin decreased the proportion of the predominant S9V in controls to 22.2% (an intermediate-resistant strain with MIC = 4 mg/L) and unmasked the high-level resistant strain (MIC = 32 mg/L) up to 77.8%. Conclusions: Strain distribution in an antibiotic-free environment depends on bacterial fitness in mono- and multi-strain niches. Selective pressure of antimicrobial regimens eradicate some populations and unmask minor populations, thus redistributing the whole population. Selective potential only for resistance phenotypes with very low prevalence (such as high-level quinolone resistance) in the community should be preferred to that selecting more prevalent resistance phenotypes.Item In vitro activity of mupirocin and amoxicillin-clavulanate alone and in combination against staphylococci including those resistant to methicillin(International Journal of Antimicrobial Agents, 2004) Alou Cervera, Luis; Cafini, Fabio; Sevillano Fernández, David; Unzueta, Irune; Prieto Prieto, JoséMupirocin and amoxicillin-clavulanate were synergistic against 9 of 49 (18%) strains of methicillin-resistant and methicillin-susceptible Staphylococcus aureus and coagulase-negative staphylococci (CNS). A pattern of enhanced killing was also found using time-kill studies. Time-kill assays were more discriminatory than chequerboard titration assays in demonstrating synergy. These results suggest that combinations of amoxicillin-clavulanate and mupirocin may have therapeutic benefits in prophylaxis against staphylococcal infections.