Journal of Antimicrobial Chemotherapy (2006) 57, 714–719 doi:10.

1093/jac/dkl041 Advance Access publication 21 February 2006

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
´ ´ ´ Luis Alou, Lorenzo Aguilar, David Sevillano, Marıa-Jose Gimenez, Fabio Cafini, ´ ´ ˜ Eva Valero, Marıa-Teresa Relano and Jose Prieto*
Microbiology Department, School of Medicine, Universidad Complutense, Avda. Complutense s/n, 28040 Madrid, Spain
Received 29 September 2005; returned 6 December 2005; revised 25 January 2006; accepted 30 January 2006

Downloaded from by guest on January 28, 2011

Objectives: 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 £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 log10 reduction) was obtained against the susceptible (MIC £ 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 ‡ 32/16 mg/L), which exhibited a 2 log10 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. Keywords: pharmacodynamic modelling, E. coli, urine bactericidal activity, amoxicillin, clavulanic acid

The incidence of amoxicillin resistance in Escherichia coli clinical isolates exceeds 50% in many parts of the world,1 and in Spain only 42% of E. coli community urinary strains are susceptible to ampicillin.2,3 Most of these isolates are resistant due to production of b-lactamase, and addition of clavulanic acid increases the ampicillin/amoxicillin susceptibility rate to 90– 98%.3,4 Quinolone resistance in E. coli has spread in Spain, with resistance rates in the community reaching 22.8% in urinary isolates3 and 24% and 26% in faecal isolates from healthy adults and children, respectively.5

A new oral pharmacokinetically enhanced formulation of co-amoxiclav has been developed that prolongs the duration of adequate amoxicillin concentrations after dosing.6 In previous studies the pharmacodynamic potential of this new formulation against amoxicillin non-susceptible Streptococcus pneumoniae was shown,6 and ex vivo serum bactericidal activity was determined in healthy volunteers.7 The serum bactericidal activity was determined using kill curves, where fresh inoculum was added to the serum sample (containing antibiotic) at each time-point.7 However, in infections, the infecting organism is exposed to changing antibiotic concentrations, and for this reason in vitro pharmacodynamic simulations mimicking this in vivo situation


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USA). 4. respectively. Sequences obtained were compared with the NCBI BLAST database. However. Valencia. respectively.A. Samples (0. Spain) represented the infection site. respectively. coli clinical isolates from community-acquired cystitis were studied in this in vitro pharmacodynamic model. coli after co-amoxiclav treatment.10 This study evaluates the urine bactericidal activity of coamoxiclav against E. 8. CA.13 This approximation resulted in a biphasic clearance of norfloxacin from urine and was achieved by synchronization of the peristaltic pumps using the Win Lin software (Cole-Parmer Instrument Co. 10 and 12 h. the situation is more complex when considering urinary tract infections. SHV and CTX-M b-lactamase genes was performed by PCR as described previously. CA.125 and 0.12 The PCR products were purified using a QIAquick PCR purification kit (Qiagen.75 mg/L for norfloxacin and clavulanic acid and of 0.25% for norfloxacin. Cole-Parmer Instrument Co. USA) at rates of 3. The Antibiotics Amoxicillin trihydrate and lithium clavulanate laboratory reference standards were supplied by GlaxoSmithKline (Worthing.3% for clavulanic acid..9% killing) was obtained only up to 8 h for the susceptible and intermediate strains and at the 2– 4 h interval for the resistant strains. The intra-day and inter-day coefficients of variation between assays were 3. amoxicillin and clavulanic acid. 10 and 12 h. USA). Downloaded from jac. The norfloxacin laboratory reference standard was purchased from Sigma Chemical Co. and 7. The limit of detection was 0. This poorly reflects the in vivo situation. In this situation. The exponential decay of concentrations was obtained by a continuous dilution–elimination process using computerized peristaltic pumps (Masterflex. Villiersle-Bel.Co-amoxiclav urine bactericidal activity against E. 85 and 30 mg/L at 1. USA) and then sequenced using a 3730 DNA Analyser (Perkin Elmer Applied Biosystems. Four of them (FJ8. coli were performed against amoxicillin non-susceptible pneumococcal strains. tubing of the peripheral unit (FX50.3% and 4. Amplification of TEM.3 mg/L for amoxicillin. 2011 Materials and methods Bacterial strains Six E. E.9 Killing curves and the pharmacokinetic analysis Bacterial suspensions in Mueller–Hinton broth supplemented with Ca2+ (25 mg/L) and Mg2+ (50 mg/L) were allowed to grow to a density of 5 · 107 cfu/mL..14 Standards and samples were prepared and diluted in Mueller–Hinton broth as required. FJ16. All determinations were performed five times and modal values are presented. In vitro kinetic model The changing antibiotic urine concentrations were simulated in a model based on a two-compartment model described previously. IL. An aliquot of 60 mL of this inoculum was introduced into the peripheral compartment.8 The extra-capillary space and the intra-dialyser circulating 715 . The co-amoxiclav profile was simulated by the clearance of clavulanic acid from urine and supplementation of the amoxicillin concentration from time 3 to 10 h to mimic the amoxicillin concentrations obtained in urine with the new formulation. the reduction in bacterial density would result in a reduction in the b-lactamase level.oxfordjournals. Barcelona. The limit of detection was 5 · 10 cfu/mL.015. A computer-controlled syringe pump (402 Dilutor Dispenser. 0.25 mg/L for norfloxacin. 10 and 14 h. serially diluted in 0. This has been suggested in experiments using animal models to measure serum bactericidal activity against b-lactamase-producing E. 4.5 mg/L at 3 and 10 h.7 mL/min for the co-amoxiclav combination. Chicago.7.9 Results of the study may have been influenced by the fact that a fresh inoculum was exposed to each urine sample.5 mL) from the peripheral compartment were collected at 0. In addition.5 mL/min (period 0–3 h) and 1 mL/min (period 3–12 h) to simulate the clearance of norfloxacin and at a rate of 4. Kinetic simulations Pharmacokinetic urine profiles in healthy volunteers of co-amoxiclav corresponding to the new co-amoxiclav 2000/125 mg SR formulation6 and of 400 mg norfloxacin13 were simulated over 12 h. 1. FJ32 and FJ64) were those previously used in ex vivo determinations in a Phase I study. 2. and in turn prolongation of the in vivo bactericidal activity beyond that obtained ex vivo. Linear decay of both antimicrobials in urine was estimated using the mean concentration of each documented interval in Phase I studies. MO.3% for amoxicillin. if necessary. and particularly when b-lactamase-producing pathogens are being tested against co-amoxiclav. and colonies were counted after 24 h. Samples were assayed by bioassay using Micrococcus luteus ATCC 9341 and Klebsiella pneumoniae NCTC 11228 as indicator organisms for amoxicillin and clavulanic acid. for clavulanic acid. as measured by a UV-spectophotometer (Hitachi U-1100). Gilson S. MIC determination and b-lactamase gene sequencing MICs were determined by a microdilution method following CLSI methodology11 prior to and after the simulation process. France) allowed the simulation of concentrations in urine. 8. Additional pumps circulated the antibiotic-medium mixture at a 50 mL/min rate between the central and peripheral compartment and at a rate of 20 mL/min within the extra-capillary space through external tubing. by guest on January 28.1% and 4.9% sodium chloride. 3. USA). and stored at –50 C for the measurement of simulated urine concentrations. 141 and 21 mg/L at 3. >99.. respectively. 1.8% and 2. UK). (St Louis.8 Although it is relatively simple to undertake a pharmacodynamic analysis for systemic infections. by infusion of the antibiotic into the central compartment until the Cmax was reached. Foster City. and. amoxicillin and clavulanic acid. IL. Chicago. bactericidal activity (>3 log10 reduction. coli NCTC 10418 was used as the indicator organism for norfloxacin.A. 6. for norfloxacin. and 41 and 1. In a previous Phase I study. and each experiment was performed in triplicate. Figure 1 shows target and experimental concentrations of norfloxacin. where bacteria in the urine are contained in the bladder and exposed to changing antibiotic concentrations over time. for amoxicillin.9. At least four dilutions of each sample were spread onto Mueller–Hinton agar plates supplemented with 5% sheep blood and incubated at 37 C. with internal control concentrations of 0. 3. Fresenius Medical Care S. The target concentrations in urine were: 167. aliquots were taken from the central compartment at 0. coli in an in vitro model simulating urine concentrations obtained in humans up to 12 h after administration of a single dose in comparison with norfloxacin. respectively.9 significant antibacterial activity was obtained up to 12 h for the susceptible (8/4 mg/L) and intermediate (16/8 mg/L) strains and up to 8 h with the resistant (32/16 and 64/32 mg/L) strains with the new co-amoxiclav formulation. 3 and 12 h.

7 – 27. taking into consideration the multiple comparisons performed in the analysis. and 152.03 for the strain MR61.15 Experimental Cmax (mg/L). Pharsight. from 7.12 for the strain FJ64. The P value was adjusted by the Bonferroni correction method. b-Lactamases and MICs (mg/L) for the six E. where t > MIC was 92%. clavulanic acid (b) and norfloxacin (c). and from 7.92 – 0.06 for the strain FJ32. 64 and 32 mg/L). from 7. 42.25 mg/L) according to CLSI breakpoints. Results Table 1 shows the b-lactamase profile and MICs of co-amoxiclav and norfloxacin for the E. In antibiotic-free simulations. Table 2 shows the initial inocula decrease over 12 h with the simulated urinary co-amoxiclav concentrations. (a) 1050 Concentration (mg/L) 800 550 300 50 40 20 0 0 (b) 100 Concentration (mg/L) 80 60 40 20 0 0 (c) 300 2 4 6 Time (h) 8 10 12 2 4 6 Time (h) 8 10 12 Table 1. Staircase maximum and minimum concentrations are shown as grey dotted lines.oxfordjournals.25 Strain MR110 MR61 FJ8 FJ16 FJ32 FJ64 b-Lactamase TEM-1 TEM-116 TEM-1 TEM-1 TEM-1 TEM-1 200 100 0 0 2 4 6 Time (h) 8 10 12 Figure 1. 42.5 for norfloxacin.81 – 0. 2011 Concentration (mg/L) 716 .25 £0.5.2.Alou et al.04 – 0.01 for the strain FJ8. Mountain View.0 – 85. one was intermediate (16/8 mg/L) and two were resistant (MICs of 32/16 and 64/32 mg/L). Table 3 shows the initial inocula decrease over 12 h with the simulated urinary norfloxacin concentrations. from 7.1. These concentrations gave mean values for t > MIC of 100% for all strains except for the most resistant strain (MIC of 64/32 mg/L).83 – 0. At least a 2 log10 reduction with respect to initial inocula (log10 cfu/mL initial – log10 cfu/mL at the corresponding sampling time) was obtained for all strains from the 3 h timepoint onwards.15 The three amoxicillin/clavulanic acid susceptible strains were resistant to norfloxacin (MICs of 256.98 – 0.2 – 3.7% and 0% for the resistant strains (MICs of 32.25 mg/L) and of 95.001 was considered statistically significant.8 – 19. 1 – 0 and 1038.69 – 0.66 – 0.1 for clavulanic acid.25 £0. 64 and 256 mg/L). However.09 to 10. Statistical analysis Differences in log10 colony counts at each sampling time with respect to initial inocula were calculated. with no significant differences between the strains. occuring when norfloxacin Downloaded from jac. Three strains were susceptible to amoxicillin/clavulanic acid (MICs of 4/2.5 – 13.75 – 0. CA.07 – 0. a ‡1 log10 increase in bacterial counts with respect to the previous sampling time-point occurred at 10 h (versus 8 h) and at 12 h (versus 10 h) with the resistant strains when the amoxicillin concentrations were lower than 3 · MIC (amoxicillin/clavulanic acid) and clavulanic acid concentrations were £1.64 – 0.04 to 10.8%. pharmacokinetic analysis was based on a non-compartmental approach (WinNonlin. 3 – 0 and 155. P < 0. with bacterial regrowth. Antibacterial activity could be considered bactericidal (>3 log10 reduction with respect to initial inocula) from 3 to 12 h against the susceptible and intermediate strains and from 3 to 10 h against the two resistant strains.5 mg/L.17 to 10. with respect to the initial inocula. These produced mean values for t > MIC of 100% for all susceptible strains (MICs £ 0.66 – 0.7 for amoxicillin. from 7.08 to 10. Experimental (black continuous line) and target (black broken line) concentrations of amoxicillin (a). 3 – 0 and 4647. the bacterial load increased from initial inocula (log10 cfu/mL) to bacterial counts at 12 h (log10 cfu/mL) as follows: from 7.05 for the strain FJ16. Inter-strain differences at each time-point were determined using Tukey’s multiple comparison test.6 – 11. while the three amoxicillin/clavulanic acid non-susceptible strains (MICs ‡ 16/8 mg/L) were susceptible to norfloxacin (MICs £ 0. Tmax (h) and AUC0–24 (mg·h/L) values were 821.respectively. coli strains used in this study. USA).07 for the strain by guest on January 28. coli strains used in this study Amoxicillin/ clavulanic acid MIC 4/2 4/2 8/4 16/8 32/16 64/32 Norfloxacin MIC 256 64 32 £0.21 to 11.17 to 12.62 – 0. according to CLSI breakpoints. Significant interstrain differences were observed. 4/2 and 8/4 mg/L).

but was never observed for the strains with norfloxacin MICs of 64 and 256 mg/L.8 – – – – – – 10 h 0.0 0.1c –0.1 0. The AUC0–12/ MIC ratio was <24 in those strains that showed regrowth (norfloxacin MICs of 64 and 256 mg/L).3 – 11.3 0. this was at a time when resistance rates were not higher than 15% in Spain.3 1.0 0.5 3.6 – 0.1 – 0.8 1.3a 4h 3.8 – 0. c P < 0.8 3.8 2.7 1.3c 0.3 0.3 – 0.3 –1.3 6. 2011 Table 3. resistance rates in E. E.1 – 0. concentrations fell below the MIC for the isolate.6 5.08 3.3 4.4 5.2 0. and since then an 1% per annum increase in the resistance rate for E.9 –0. FJ16 and FJ8.5 2.6 – 13.8 – 0.12 It has been suggested that fluoroquinolones are a logical choice for empirical therapy of uncomplicated cystitis. Bactericidal activity (>3 log10 reduction) was generally observed from 4 to 12 h for the susceptible strains (norfloxacin MICs £ 0.7 0.8 3h 3.3 0.5 47.5 2. E.0 0.16.6 0. a Amoxicillin/clavulanic acid MIC.3 0.1 6.25 32 64 256 0h 0.4 6.0 821.0 0.9 4.3 2.4 – 0.9 3.02 AUC0–12 (mg·h/L) 773.4 4.3 – 7.8 0.5 – 13.19 and in early studies norfloxacin was associated with good efficacy in the treatment of uncomplicated cystitis.1 – 0.6 6.9 7.2 4.5 – – – – – – 0.3 Extended-spectrum b-lactamases are not often found in E. not determined.3a 0.3 by guest on January 28.25 £0.17 As a consequence.7 Negative values (given in bold) correspond to regrowth with respect to initial inocula.4 4.1 – 2.4 1.2 3.7 – 0.4 0.16 12.0 0.6 1.9 – 12 h 0.4 4. However.18 Susceptibility rates of E.1c 0.0 0.3 – 1.001 versus FJ64.1 0.1 0.0 – 17. Simulated norfloxacin urine concentrations (mean – SD) after a single 400 mg dose.1 – 0. coli initial inocula reduction over time (log10 cfu/mL initial inocula – log10 cfu/mL at the corresponding sampling time) and mean urine t > MIC Mean t > MICb (%) 100 100 100 100 100 92 Strain MIC (mg/L)a 0h 0.2 – 0.43 ‡3092.1 0.0 – 0.0 1h 0.8 3.0 – 0.1 0. Simulated co-amoxiclav urine concentrations (mean – SD) after a single 2000/125 mg dose. coli.001 versus all other strains.2 4.1 5.3 3. after norfloxacin exposure.7 – 0.3 4.6 – – – – – – 0.2 – – – – – – – – 4h 0.60 Strain FJ64 FJ16 FJ32 FJ8 MR61 MR110 MIC (mg/L) £0.5 40.4 – 19. b t > MIC = % dosing interval (12 h) where amoxicillin concentrations in urine exceed amoxicillin/clavulanic acid MIC.1 0.6 0. a P < 0.4 3. 717 .6 1.8 0. coli initial inocula reduction over time (log10 cfu/mL initial inocula – log10 cfu/mL at the corresponding sampling time) and mean urine AUC0–12/MIC Mean AUC0–12/MIC ‡3092.3 – 2.25 mg/L) and from 8 to 12 h for the strain with a norfloxacin MIC of 32 mg/L.17 their use in cystitis is not advocated without a b-lactamase inhibitor.9 462.5 – 0.4 – 8h 0.2 0.8 – 0. This occurred at 3 h in the case of the strain with an MIC of 256 mg/L and at 8 h in the case of the strain with an MIC of 64 mg/L.5 – 0.25 £0.0 0.0 0.3 0.4 5.5 5. b P < 0.2 583. Downloaded from jac.5 2.3 1.8 5. the strain FJ8 exhibited an MIC of 256 mg/L (three dilutions higher than the one pre-exposure: 32 mg/L) while the remaining strains exhibited the same MIC values.3 3. Isolates recovered at the end of the experiments exhibited amoxicillin/clavulanic acid MIC values equal to those prior to co-amoxiclav exposure. FJ32.43 ‡3092.4 ND – – – – – – – 3h 0.0 0.0 – 0.10 – 68. coli from outpatient urine culture in Spain (1.2c –0.1 252.6 – – – – – – 0. coli to fluoroquinolones has occurred.4 4.1 26.8 0.7 3.6 – 0.7 5.3 5.5 – 0.0 0.3 0.5 1.0 – 10 h 0.6 124.1 – 3.3 0.0 – Concentration (mg/L) 152.7 5.4 0.4 1.9 – 0.1 2.8 – 0.3 0.7 – 0.3 0.5 – 1. Bactericidal activity (against all strains with co-amoxiclav and against four strains with norfloxacin) was observed at earlier time-points with co-amoxiclav than with norfloxacin (3 h versus 4–8 h).0 0.0 0.0 0.5 79.0 0.0 – 0.1 3.5 6.19.43 24.9 1.0 – – – – – – – 0. coli to amoxicillin/clavulanic acid are 90% in Spain3 and have remained at this level over time because hyperproduction of TEM-1 b-lactamase (or its derivative IRT) has a low frequency in Spain.Co-amoxiclav urine bactericidal activity against E.4 2.2 – 0.oxfordjournals. Discussion Owing to the high prevalence of resistance to amoxicillin or ampicillin in E.20 However.3 – 0.4 4.5 – 1.3 0.1 71.7 – 1.0 – 3.3 4.3 4.5 – 12 h 0.0 – – – – – – – – 0.9 1.5 –2.0 1h 1.2 2.9 – 1. coli to norfloxacin now exceed the 20% level in many parts of Spain.7 0.4c –2.1 –2.9 – 0.9 3.0 0.0 0.2 3.0 0.8 42.0 0.5 – 0.4 0.1 0.001 versus FJ64 and FJ16.7 – 6.2 39.4%).0 – 1.1 2. coli Table 2.7 29.2 0.1 5.1 MR110 4/2 MR61 4/2 FJ8 8/4 FJ16 16/8 FJ32 32/16 FJ64 64/32 Amoxicillin concentration (mg/L) Clavulanate concentration (mg/L) ND.9 5.3c –3.3b 8h 6.0 0.0 0.0 0. and those that are found are not usually TEM derivates.

9 where fresh E. 120: 361–4. coli inocula were exposed to fixed co-amoxiclav concentrations at different time-points (static experiments). Murbach V. Sevillano D. We believe that the difference between the two studies is due to the fact that in the Phase I study a fresh inoculum was exposed to the urine sample (containing the antibiotic) at each time-point. noncomparative clinical trial in healthy volunteers. 23: 578–84. bactericidal activity (>3 log10 reduction) was obtained for up to 8 h against the susceptible and intermediate strains. in this study. Alonso Sanz M. Xercavins M. Peter JD et al. In the assessment of an antibiotic for uncomplicated cystitis. 2011 References 1.9 In the earlier study. In this study the Mueller–Hinton broth used was supplemented with Ca2+ and Mg2+ to better simulate the urinary situation. J Antimicrob Chemother 2004. Rodriguez-Carballeira M et al. Effect of opsonophagocytosis mediated by specific antibodies on the co-amoxiclav serum bactericidal activity against Streptococcus pneumoniae after administration of a single oral dose of pharmacokinetically enhanced 2000/125 mg co-amoxiclav to healthy volunteers. Antimicrobial susceptibility of the pathogens of bacteraemia in the UK and Ireland 2001–2002: the BSAC Bacteraemia Resistance Surveillance Programme. 7. Moreover. 718 .28. where the bactericidal activity against four of the six strains used in this study (those strains with MICs of 8/4. In this study bactericidal activity with norfloxacin was obtained from 4 to 12 h against strains with MIC values of £32 mg/L. open-label. Alou L. This is a higher value than those found for strains with b-lactamase hyperproduction. Downloaded from jac. and only at the 2–4 h post-dosing interval against the two resistant strains.oxfordjournals. Clin Ther 2005. Garau J.21 One of the earlier attempts to use in vitro dynamic models to simulate bladder infections measured the effects of antibiotics by photometrically studying bacterial growth in artificial media and simulating micturation by alterations of the volume and flow rate of the medium. ´ 9. this bladder model correlated well with mouse infection protection models and provided realistic simulations of the response seen in patients in clinical trials. Estudio nacional multicentrico. quinolone treatment should be used with caution in countries such as Spain. ´ 2.. In previous in vitro27 and ex vivo studies in animal models10 and in humans. Enferm Infecc Microbiol Clı´n 2005. Fenotipos de resistencia en aislamientos urinarios de Escherichia coli en la comunidad: implica´ ciones terapeuticas. Ponte (Fundacion Jimenez Dıaz) ´ for the supply of the FJ E. 53: 1018–32.Alou et al. 27: 1043–9. categorization of this strain as resistant is correct taking into account the highly resistant subpopulation selected. this threshold was not observed. bactericidal activity was obtained up to 12 h (the whole dosing interval) against the susceptible and intermediate strains. Effects of following National Committee for Clinical Laboratory Standards and Deutsche Industrie Norm-Medizinische Mikrobiologie guidelines. as may occur in vivo. Sevillano D et al. which better mimics the in vivo situation. Andreu A. permitting bacterial regrowth that may result in therapeutic failure. J Clin Microbiol 1998. and up to 10 h against the resistant strains. oral. and the changes in bacterial counts resulted solely from the action of the antimicrobial agent. 23: 4–9. Emergence and dissemination of quinolone-resistant Escherichia coli in the community. Reynolds R. Gobernado M et al. Kaye CM. 36: 1361–5. 54: 1148–51. Ex vivo pharmacodynamics of co-amoxiclav against b-lactamase-producing Escherichia coli in a yucatan miniature pig model that mimics human by guest on January 28. Simpson I. Although generally bacteria grow more quickly in Mueller–Hinton broth than in urine. Clin Ther 2001. This study was supported by an unrestricted grant from GlaxoSmithKline S. Durodie J. Acknowledgements ´ ´ ´ We thank F. Gimenez MJ et al. 8. where resistance rates are higher than 15%. Alos JI. the system used was closed (bacteria were not eliminated with alterations in the flow rate). Urinary concentrations and bactericidal activity against amoxicillin-nonsusceptible strains of Escherichia coli with single-dose. and site of infection on susceptibility of Escherichia coli to co-amoxiclav (Augmentin).22 Despite criticisms that such spectrophotometric measurements included both non-viable and viable bacteria and that they were not reliable for dense bacterial populations (106–109 cfu/mL). coli strains and O. Madrid. fluoroquinolone activity can be decreased in urine due to magnesium concentrations present. In the present in vitro simulation. Colman M et al. a clavulanic acid threshold for the activity of amoxicillin was observed. bactericidal activity in urine is a relevant pharmacodynamic parameter because the gradual decrease in antibiotic concentration (which is not possible to mimic in standard in vitro testing) could lead to concentrations below the MIC. 5. Abad Becquer MI. Ponte C. ´ 3. Potz N. 16/8.26 The results obtained in this study with co-amoxiclav contrast with those obtained previously in a Phase I study in healthy volunteers. J Antimicrob Chemother 2004. Soriano and M.23–25 In the present pharmacodynamic simulation. but regrowth (with respect to the initial inoculum) occurred when concentrations were below the MIC for resistant strains with MICs ‡ 64 mg/L. Perry S et al. 46: 3782–9.29 In contrast. Etiologıa y sensibilidad a ´ ´ los antimicrobianos de los uropatogenos causantes de la infeccion urinaria ´ baja adquirida en la comunidad. The clinical pharmacokinetics of a new pharmacokinetically enhanced formulation of amoxicillin/ clavulanate. country of isolate origin. Gracia M. Bronner S. 4. Aguilar L. Antimicrob Agents Chemother 2002. while in this simulation the same inoculum is exposed to changing concentrations over time. From this perspective the results of this study suggest that clavulanic acid can be given twice daily (to protect amoxicillin activity) in the treatment of uncomplicated cystitis due to E. Med Clin (Barc) 2003. 55: 742–7. with the strain with an MIC of 32 mg/L the survival bacterial population at the end of the simulation exhibited an increased MIC (256 mg/L). Echeverrıa for her technical assistance in this study. In contrast.A. 32/16 and 64/32 mg/L) was assessed. C. where exposure of the same inocula to changing co-amoxiclav concentrations was used. coli exhibiting amoxicillin/ clavulanic acid MICs up to 64/32 mg/L. Spain. Allen A. Gimenez MJ et al. 43: 2736–41. 10. Knott S et al. 6. sustained-release amoxicillin/ clavulanic acid: a phase I. This suggests that despite the original bactericidal activity obtained against this strain with norfloxacin. Antimicrob Agents Chemother 1999. Bactericidal activity of amoxicillin against non-susceptible Streptococcus pneumoniae in an in vitro pharmacodynamic model simulating the concentrations obtained with the 2000/125 mg sustained-release co-amoxiclav formulation. J Antimicrob Chemother 2005. Calvo A. Transparency declarations None to declare.

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