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Ceftobiprole is a broad‐spectrum cephalosporin. The objective of this study was to test the hypothesis that the pharmacokinetics (PK) and exposure of ceftobiprole in Asian subjects are similar to those in non‐Asian subjects. Three approaches were followed. The first compared the individual PK estimates between the 2 subgroups derived from a population PK model previously built. Next, it was determined whether “Asian subject” was a significant covariate. Finally, a pharmacodynamic analysis was performed by comparing measures of exposure and target attainment. No significant differences were found between PK parameter estimates for Asian and non‐Asian subjects, with median values (range) for clearance of 4.82 L/h (2.12–10.47) and 4.97 L/h (0.493–20.6), respectively (P= .736). “Asian subject” was not a significant covariate in the population PK model. There were no significant differences between the measures of exposure. The geometric mean ratio for the fAUC was 1.022 (90%CI, 0.91–1.15), indicating bioequivalence. Taking a target of 60% coverage of the dose interval, more than 90% of the population in both subgroups was adequately exposed. This analysis demonstrated that there are no PK or pharmacodynamic differences between Asian and non‐Asian subjects for a ceftobiprole dose of 500 mg every 8 hours as a 2‐hour infusion.

ABSTRACTCeftolozane is a new broad-spectrum cephalosporin and is combined with tazobactam to broaden the activity of ceftolozane against strains producing extended-spectrum beta-lactamases (ESBLs). We determined the pharmacodynamics (PD) of the combination in the neutropenic mouse thigh model to determine the optimal exposure of tazobactam. Treatment of CD-1 neutropenic mice was started 2 h after infection with ceftolozane every 2 h (q2h) alone or in combination with tazobactam at different dosing frequencies for 24 h, and the number of CFU in the thighs was determined before and after treatment. The maximum effect model was fit to the dose-response and the pharmacokinetic/PD index (PDI)-response to determine the PDI values for ceftolozane alone and ceftolozane in combination with tazobactam resulting in a static effect and a 1-log kill. The effect of tazobactam was dependent on the percentage of time that the free drug concentration remained above the concentration threshold (percent fT>CT), whereby dosing q2h was more efficacious than dosing every 8 h (q8h), reducing the tazobactam daily dose by a factor 6.9 to 59.0 (n= 3 strains) to obtain a static effect. Using R2as an indicator of the best fit of the percent fT>CT-response relationships, the concentration threshold best correlating with the response varied from 0.5 to 2 mg/liter, depending on the strain. A similar result was obtained when the q2h and q8h regimens were analyzed. For all isolates tested, the mean fT>CTfor 0.5 mg/liter tazobactam was 28.2% (range, 17.5 to 45.8%) and 44.4% (range, 26.6 to 54.7%) for a static effect and a 1-log kill, respectively, at ceftolozane exposures that produced a ceftolozane concentration of 4 mg/liter (a concentration greater than the MIC) for 33.9 to 63.3% of a 24-h period under steady-state pharmacokinetic conditions. The main PDI that correlated with the effect of tazobactam was the fT>CTachieved with a CTof 0.5 mg/liter tazobactam.

ABSTRACTThe emergence of resistant strains among common and rare Candidaspecies necessitates continuous monitoring of the in vitrosusceptibilities of those isolates. We therefore assessed the in vitroactivities of micafungin against 1,099 molecularly identified isolates belonging to 5 common and 20 rare Candidaspecies by the EUCAST, CLSI, and Etest methods, assessing both the intralaboratory agreement and the interlaboratory agreement for two centers. The median micafungin EUCAST MICs were as follows, from the lowest to the highest: for Candida albicans, 0.004 mg/liter; for C. glabrata, 0.016 mg/liter; for C. tropicalis, 0.031 mg/liter; for C. krusei, 0.125 mg/liter; for C. parapsilosis, 2 mg/liter. Among rare Candidaspecies, high MICs were found for C. guilliermondii, C. lipolytica, C. orthopsilosis, C. metapsilosis, and C. fermentati.No resistant isolates were found by the CLSI method, whereas resistance rates of 1 to 2% were found by the EUCAST method. Overall, the EUCAST method resulted in MICs 1 to 2 dilutions higher than those found by the CLSI and Etest methods. The intra- and interlaboratory agreement between methods was >92%, except for the interlaboratory agreement between the EUCAST and CLSI methods (81%), where 17 to 31% of the differences were >2 2-fold dilutions for C. albicans, C. glabrata, C. tropicalis, and other rare Candidaspecies and <6% for C. parapsilosisand C. krusei. For the other interlaboratory comparisons, the EUCAST method resulted in higher MICs than the Etest method for all species, but <7% of these differences were >2 2-fold dilutions. Overall, the CLSI method resulted in lower MICs than the Etest method, with 11% of all isolates demonstrating >2 2-fold-dilution differences (6 to 20% for C. albicans, C. tropicalis, and rare Candidaspecies; <5% for C. glabrata, C. krusei, and C. parapsilosis) and smaller differences found after 24 h. Despite these differences, categorical agreement was excellent (>97%), with only 1 to 2% very major errors between the EUCAST method and the other two methods.

ABSTRACTThis study aimed to explore any mutation in the CYP51gene conferring azole resistance in Aspergillus flavus. Two voriconazole-resistant and 45 voriconazole-susceptible isolates were included in the study. Sequence analysis demonstrated a T1025C nucleotide change in CYP51C, resulting in the Y319H amino acid substitution in one resistant isolate. However, the earlier described T788G mutation in CYP51Cconferring voriconazole resistance in A. flavusisolates was present in all isolates, irrespective of their susceptibility status.

ABSTRACTCeftolozane is a new cephalosporin with activity against Gram-negative and Gram-positive microorganisms. However, the compound is susceptible to degradation by extended-spectrum beta-lactamases (ESBLs). Tazobactam is an ESBL inhibitor and is combined with ceftolozane to broaden its activity. Surprisingly, although tazobactam has been available for over 20 years, few if any reliable data exist on the tazobactam pharmacokinetic (PK) properties in mice. To evaluate the PK and pharmacodynamic (PD) relationships in mice, the PK properties of tazobactam and ceftolozane were extensively investigated. Thigh-infected neutropenic CD-1 mice were injected intraperitoneally with a single 0.1-ml dose containing ceftolozane, tazobactam, or both compounds. Ceftolozane was applied in 2-fold-increasing doses of 4 mg/kg of body weight to 64 mg/kg alone or in combination. Tazobactam was combined in reverse doses (thus, 64/4 mg/kg, 32/8 mg/kg, etc.) (n =2 per time point). In separate validation experiments, ceftolozane-tazobactam was given alone or in combination at 32/8 mg/kg and 8/32 mg/kg (n =4 per time point). Plasma samples (one per mouse) and bronchoalveolar lavage samples were collected at up to 12 time points until 6 h after administration. There were no significant differences in the ceftolozane and tazobactam PK alone versus combined, indicating no PK interaction. The PKs were linear and dose proportional for both compounds and showed a good penetration in the epithelial lining fluid. The estimated mean (standard deviation) half-life of ceftolozane was 0.287 h (0.031 h), and that of tazobactam was 0.176 h (0.026), and the Vwas 0.43 liter/kg and 1.14 liter/kg, respectively. The estimates of tazobactam parameters can also be used to (re)interpret PD data.

ABSTRACTSpecies of Verruconisand species of Ochroconisare dematiaceous fungi generally found in the environment but having the ability to infect humans, dogs, cats, poultry, and fish. This study presents the antifungal susceptibility patterns of these fungi at the species level. Forty strains originating from clinical and environmental sources were phylogenetically identified at the species level by using sequences of the ribosomal DNA internal transcribed spacer (rDNA ITS). In vitroantifungal susceptibility testing was performed against eight antifungals, using the Clinical and Laboratory Standards Institute (CLSI) broth microdilution method. The geometric mean MICs for amphotericin B (AMB), flucytosine (5FC), fluconazole (FLC), itraconazole (ITC), voriconazole (VRC), and posaconazole (POS) and minimum effective concentrations (MECs) for caspofungin (CAS) and anidulafungin (AFG) across the Ochroconisand Verruconisspecies were as follows, in increasing order. For Verruconisspecies, the values (μg/ml) were as follows: AFG, 0.04; POS, 0.25; ITC, 0.37; AMB, 0.50; CAS, 0.65; VRC, 0.96; 5FC, 10.45; and FLC, 47.25. For Ochroconisspecies, the values (μg/ml) were as follows: AFG, 0.06; POS, 0.11; CAS, 0.67; VRC, 2.76; ITC, 3.94; AMB, 5.68; 5FC, 34.48; and FLC, 61.33. Antifungal susceptibility of Ochroconisand Verruconiswas linked with phylogenetic distance and thermotolerance. Echinocandins and POS showed the greatest in vitroactivity, providing possible treatment options for Ochroconisand Verruconisinfections.

ABSTRACTThe percentage of the dosing interval that the non-protein-bound plasma concentration is above the MIC (%fT>MIC) for cephalosporins has been shown to correlate with microbiological outcomes in preclinical studies. However, clinical data are scarce. Using data from a randomized double-blind phase 3 clinical trial, we explored the relationship of ceftobiprole exposure with microbiological and clinical outcomes in patients with nosocomial pneumonia. The individual ceftobiprole exposure was determined for different pharmacokinetic (PK)/pharmacodynamic (PD) indices using individual pharmacokinetic data and a previously published population model. The MICs used in the analysis were the highest MICs for any bacterium cultured at baseline or the end of treatment (EOT). Outcomes were microbiological cure at EOT and clinical cure at test of cure (TOC). Multiple logistic regression (MLR) and classification and regression tree (CART) analyses were applied to determine the relationships among exposure, patient characteristics, and outcomes. MLR indicated that the %fT>MIC of ceftobiprole was the best predictor for both microbiological eradication and clinical cure. CART analysis showed a breakpoint value of 51.1% (n= 159; P= 0.0024) for clinical cure, whereas it was 62.2% (n= 251; P< 0.0001) for microbiological eradication. Other factors also contributed, particularly to clinical outcome. These included the difference between VAP and non-VAP patients, systemic inflammatory response syndrome (SIRS), creatinine clearance, the use of anti-Pseudomonascombination therapy, and Acute Physiology and Chronic Health Evaluation II (APACHE-II) score. There is a strong correlation between microbiological eradication and clinical cure with exposure to ceftobiprole. The %fT>MIC required to result in a favorable clinical outcome is >51% of the dosing interval, which is in line with the values found for microbiological eradication, the comparator ceftazidime, and preclinical models.

ABSTRACTAlthough conventional amphotericin B was for many years the drug of choice and remains an important agent against invasive aspergillosis, reliable susceptibility breakpoints are lacking. Three clinical Aspergillusisolates (Aspergillus fumigatus, Aspergillus flavus, and Aspergillus terreus) were tested in an in vitropharmacokinetic-pharmacodynamic model simulating the biphasic 24-h time-concentration profile of free amphotericin B concentrations in human serum with free peak concentrations (fCmax) of 0.1, 0.3, 0.6, 1.2, and 2.4 mg/liter administered once daily. Drug concentrations were measured with a bioassay, and fungal growth was monitored for 72 h with galactomannan production. The fCmax/MIC corresponding to half-maximal activity (P50) was determined for each species, and the percentage of target attainment was calculated for different MICs for the standard (1 mg/kg of body weight) and a lower (0.6-mg/kg) dose of amphotericin B with Monte Carlo simulation analysis. The fCmax/MICs (95% confidence intervals) corresponding to P50were 0.145 (0.133 to 0.158), 0.371 (0.283 to 0.486), and 0.41 (0.292 to 0.522) for A. fumigatus, A. flavus, and A. terreus, respectively. The median percentages of P50attainment were ≥88%, 47%, and 0% for A. fumigatusisolates with MICs of ≤0.5, 1, and ≥2 mg/liter, respectively, and ≥81%, 24%, and 0% and ≥75%, 15%, and 0% for A. flavusand A. terreusisolates with MICs of ≤0.25, 0.5, and ≥1 mg/liter, respectively. The lower dose of 0.6 mg/kg would retain efficacy for A. fumigatus, A. flavus, and A. terreusisolates with MICs of ≤0.25, ≤0.125, and ≤0.125 mg/liter, respectively. The susceptibility, intermediate susceptibility, and resistance breakpoints of ≤0.5, 1, and ≥2 mg/liter for A. fumigatusand ≤0.25, 0.5, and ≥1 mg/liter for A. flavusand A. terreuswere determined for conventional amphotericin B with a pharmacokinetic-pharmacodynamic model simulating free-drug serum concentrations.

ABSTRACTEpidemiological cutoff values (ECV) are commonly used to separate wild-type isolates from isolates with reduced susceptibility to antifungal drugs, thus setting the foundation for establishing clinical breakpoints for Aspergillus fumigatus. However, ECVs are usually determined by eye, a method which lacks objectivity, sensitivity, and statistical robustness and may be difficult, in particular, for extended and complex MIC distributions. We therefore describe and evaluate a statistical method of MIC distribution analysis for posaconazole, itraconazole, and voriconazole for 296 A. fumigatusisolates utilizing nonlinear regression analysis, the normal plot technique, and recursive partitioning analysis incorporating cyp51Asequence data. MICs were determined by using the CLSI M38–A2 protocol (CLSI, CLSI document M38–A2, 2008) after incubation of the isolates for 48 h and were transformed into log2MICs. We found a wide distribution of MICs of all azoles, some ranging from 0.02 to 128 mg/liter, with median MICs of 32 mg/liter for itraconazole, 4 mg/liter for voriconazole, and 0.5 mg/liter for posaconazole. Of the isolates, 65% (192 of 296) had mutations in the cyp51Agene, and the majority of the mutants (90%) harbored tandem repeats in the promoter region combined with mutations in the cyp51Acoding region. MIC distributions deviated significantly from normal distribution (D'Agostino-Pearson omnibus normality test Pvalue, <0.001), and they were better described with a model of the sum of two Gaussian distributions (R2, 0.91 to 0.96). The normal plot technique revealed a mixture of two populations of MICs separated by MICs of 1 mg/liter for itraconazole, 1 mg/liter for voriconazole, and 0.125 mg/liter for posaconazole. Recursive partitioning analysis confirmed these ECVs, since the proportions of isolates harboring cyp51Amutations associated with azole resistance were less than 20%, 20 to 30%, and >70% when the MICs were lower than, equal to, and higher than the above-mentioned ECVs, respectively.

ABSTRACTOriginally, itraconazole for parenteral administration was licensed in a 40% hydroxypropyl-beta-cyclodextrin (HPBCD) solution for intravenous administration. A novel formulation, the NanoCrystal formulation (NCF), was prepared. NCF consists of drug particles of approximately 200 to 300 nm. The pharmacokinetics of itraconazole and its hydroxy metabolite in healthy subjects were evaluated after single and multiple doses of itraconazole as NCF. In the single-ascending-dose (SAD) study, itraconazole doses were planned to range from 50 to 500 mg, while in the multiple-ascending-dose (MAD) study, itraconazole doses of 100, 200, and 300 mg as NCF were studied, as was one dose level (200 mg) as an HBPCD solution. Samples were collected in heparinized tubes at various time points and were analyzed by high-performance liquid chromatography to allow full pharmacokinetic analysis both after the first dose and on day 7. The results of both the SAD and the MAD studies indicated that there was a dose dependency in the half-life of itraconazole from the novel formulation, increasing from 44 h (100 mg) to more than 150 h (300 mg) once steady state was achieved. Similar dose-dependent effects were observed for the hydroxy metabolite. The areas under the concentration-time curves for itraconazole and hydroxyitraconazole were also dose dependent. The pharmacokinetic profiles after 200-mg doses of itraconazole as NCF and HPBCD formulations were comparable with respect to the terminal half-life, both after a single dose and at steady state. NCF may provide an alternative to the HPBCD solution for the further optimization of antifungal treatment with itraconazole.

ABSTRACTThe therapeutic efficacy of flucytosine (5FC) monotherapy and the pharmacodynamic index predictive of efficacy were evaluated in a nonneutropenic mouse model of acute invasive aspergillosis. Mice were infected intravenously with an Aspergillus fumigatusisolate (the median MICs of 5FC were 128 μg/ml under the standard condition, 0.5 μg/ml at pH 6.0, and 0.031 μg/ml at pH 5.0) 2 h prior to the start of therapy and were treated for 7 days with different 5FC dosing regimens. The total doses ranged from 50 to 800 mg/kg of body weight/day and were administered at 6-, 12-, and 24-h intervals. The efficacy was assessed by means of survival. The survival rates of the treatment groups ranged from 40 to 90%, while the survival rate of the control group was 20%. The efficacy found depended primarily on the total daily dose. However, the power of our sample size may have been too low to exclude an effect of dose fractionation. The pharmacodynamic index that most strongly correlated with the efficacy was the area under the serum concentration-time curve and MIC ratio (R2= 0.86). We conclude that 5FC monotherapy is efficacious in a murine Aspergillus fumigatusinfection model.

ABSTRACTIn this study, we investigated the pH dependency of the in vitro activities of amphotericin B (AMB) and flucytosine (5FC) against Candidaspp., Cryptococcus neoformans, Aspergillus fumigatus, Rhizopusspp., and Scedosporium prolificansin RPMI 1640 buffered with citrate buffer (pH 4.0, 5.0, 5.4, and 6.0), citrate-phosphate buffer (pH 5.4, 6.0, 6.4, and 7.0), and 3-[N-morpholino]propanesulfonic acid (MOPS) (pH 6.4, 7.0, 7.4, and 7.9). For 5FC, no significant differences were found between MICs obtained with the different buffers, while for AMB, significant differences were found. The MICs obtained with citrate-phosphate buffer were approximately 1 twofold-dilution step higher than the MICs obtained with MOPS. We demonstrated that the in vitro activities of AMB and 5FC against yeast and mold isolates were pH dependent. The in vitro activity of AMB decreased when the pH was lowered, while the in vitro activity of 5FC increased. The effect of the pH on the in vitro activities was dependent not only on the antifungal agent tested but also on the microorganism. For AMB, there was a nonlinear relationship (median r2, 0.864) for Candidaspp., C. neoformans, A. fumigatus, and Rhizopusspp. over the pH range tested. The mean MICs ranged from 0.5 to 2.52 μg/ml at pH 7.0 and from 20.16 to 32 μg/ml at pH 5.0. For S. prolificans, there was no relationship. For 5FC, there was a linear relationship for Candidaspp. (median r2, 0.767) and a nonlinear relationship for C. neoformansand A. fumigatus(median r2, 0.882) over the pH range tested. The mean MIC values ranged from 0.125 to 1,024 μg/ml at pH 7.0 and from 0.02 to 4 μg/ml at pH 5.0. For Rhizopusspp. and S. prolificans, the relationship could not be determined, since the MIC was >1,024 μg/ml over a pH range of 4.0 to 7.9.

ABSTRACTThe in vitro susceptibilities of 21 Aspergillusisolates were tested against three antifungal agents in RPMI 1640 and yeast nitrogen base at pH 5.0 and 7.0 by a broth microdilution format of the NCCLS method. The MICs of amphotericin B and itraconazole were higher, while those of flucytosine were lower, at pH 5.0 than at pH 7.0. The poor correlation between in vitro results and clinical outcome could be due to a difference in pH between the in vitro susceptibility test and at the site of infection.

Background: Drug pharmacokinetics may be altered during liver transplantation. Cefotaxime (CTX), used as perioperative prophylaxis, demonstrates time-dependent killing and therefore continuous infusion might have pharmacodynamic advantages. Objectives: To determine the pharmacokinetics of CTX and desacetylcefotaxime (DCTX) in serum, bile and urine during continuous and intermittent infusion when performing liver transplantation. Methods: Fifteen patients undergoing liver transplantation were studied after continuous infusion (CI) (4000 mg iv per 24 h following a loading dose of 1000 mg) and intermittent bolus infusion (BI) (1000 mg iv four times daily). Samples were collected during the first 48 h after liver transplantation. Concentrations of CTX and DCTX were determined by HPLC. Results: During surgery, the mean concentration in serum after CI was 18 mg/L. The lowest serum concentration was 5 mg/L in the CI group and levels were undetectable in the BI group. Target serum concentrations of ≥4 mg/L were reached for 100% of the dosing interval during CI and ∼60% during BI. Post-operatively, the mean concentration in serum after CI was 26 mg/L. The lowest serum concentration was 8 mg/L in the CI group and levels were undetectable after BI. The peroperative pharmacokinetics of CTX in this patient group were deranged and variable, mainly caused by an increased volume of distribution and decreased hepatic clearance. Metabolism was hampered, but DCTX area under the curve (AUC)/CTX AUC ratios varying between 0.7–0.9 were reached peroperatively. Post-operatively, DCTX AUC/CTX AUC ratios were higher (1.1–1.4). Unchanged CTX in bile was ∼0.1% of the administered dose, leading to concentrations >4 mg/L throughout the dosing interval for both regimens. Conclusion: Although an intermittent bolus infusion of CTX 1000 mg produces t > target concentration for 60% of the dosing interval during liver transplantation, serum concentrations may be insufficient during the reperfusion phase. Continuous infusion overcomes this. Post-operatively, CTX clearance is impaired by decreased metabolic clearance and there is substantial accumulation of DCTX. In bile, sufficient concentrations of CTX and its active metabolite are reached with both regimens.

ABSTRACTCombination therapy of flucytosine (5FC) with other antifungal agents could be of use for the treatment of invasive aspergillosis. However, interpretation of the results of in vitro interactions is problematic. The fractional inhibitory concentration (FIC) index is the most commonly used method, but it has several major drawbacks in characterizing antifungal drug interaction. Alternatively, a response surface approach using the concentration-effect relationship over the whole concentration range instead of just the MIC can be used. We determined the in vitro interactions between amphotericin B (AMB), itraconazole, and 5FC against 21 Aspergillusisolates with a broth microdilution checkerboard method that employs the dye MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide]. FIC indices based on three different MIC endpoints (MIC-0, MIC-1, and MIC-2) and the interaction coefficient alpha were determined, the latter by estimation from the response surface approach described by Greco et al. (W. R. Greco, G. Bravo, and J. C. Parsons, Pharmacol. Rev. 47:331-385, 1995). The value obtained for the FIC index was found to be dependent on the MIC endpoint used and could be either synergistic, indifferent, or antagonistic. The response surface approach gave more consistent results. Of the three combinations tested, the AMB-5FC combination was the most potent in vitro against Aspergillusspp. We conclude that the use of the response surface approach for the interpretation of in vitro interaction studies of antifungals may be helpful in order to predict the nature and intensity of the drug interaction. However, the correlation of these results with clinical outcome remains difficult and needs to be further investigated.

ABSTRACTDuring the last decade some studies have shown that the area under the curve (AUC)/MIC ratio is the pharmacodynamic index that best predicts the efficacies of quinolones, while other studies suggest that the predictive value of the peak concentration/MIC (peak/MIC) ratio is superior to the AUC/MIC ratio in explaining clinical and microbiological outcomes. In classical fractionated dose-response studies with animals, it is difficult to differentiate between the AUC/MIC ratio and the peak/MIC ratio because of colinearity. Three different levofloxacin and ciprofloxacin dosing regimens were studied in a neutropenic mouse pneumonia model. The different regimens were used with the aim of increasing the AUC/MIC ratio without changing the peak/MIC ratio and vice versa. The first regimen (RC) consisted of daily doses of 5 up to 160 mg/kg of body weight divided into one, two, or four doses. In the second regimen (R0), mice were given 1.25 mg/kg every hour from 1 to 23 h, while the dose given at 0 h was 2.5, 5, 10, 20, 40, or 80 mg/kg. In the third regimen (R11), mice also received 1.25 mg/kg every hour from 0 to 23 h; but in addition, they also received 2.5, 5, 10, 20, 40, or 80 mg/kg at 11 h. The level of protein binding was also evaluated. The results indicate that the unbound fraction (fu) was concentration dependent for both levofloxacin and ciprofloxacin and ranged from approximately 0.67 to 0.88 for both drugs between concentrations of 0.5 and 80 mg/liter. The relationships between the AUC/MIC ratio and the number of CFU were slightly better than those between the peak/MIC ratio and the number of CFU. There was no clear relationship between the amount of time that the concentration remained above the MIC and effect (R2< 0.1). For both drugs, the peak/MIC ratio that resulted in a 50% effective concentration was lower for the R0 and R11 dosing regimens, indicating the importance of the AUC/MIC ratio. The same was true for the static doses. Survival studies showed that for mice treated with the low doses the rate of survival was comparable to that for the controls, but with the higher doses the rate of survival was better for mice receiving the R0 regimen. We conclude that for quinolones the AUC/MIC ratio best correlates with efficacy against pneumococci and that the effect of the peak/MIC ratio found in some studies could be partly explained by concentration-dependent protein binding.

ABSTRACTCombination therapy could be of benefit for the treatment of invasive yeast infections. However, in vitro interaction studies are relatively scarce and the interpretation of the fractional inhibitory concentration (FIC) index can be contradictory due to various definitions used; not all information on the interaction study is used in the index, and different MIC end points exist for different classes of drugs. Fitting an interaction model to the whole response surface and estimation of an interaction coefficient alpha (ICα) would overcome these objections and has the additional advantage that confidence intervals of the interaction are obtained. The efficacy of flucytosine (5FC) in combination with amphotericin B (AB) and fluconazole (FCZ) was studied against 35 yeast isolates in triplicate (Candida albicans[n= 9], Candida glabrata[n= 9], Candida krusei[n= 9], and Cryptococcus neoformans[n= 8]) using a broth microdilution checkerboard method and measuring growth after 48 h by a spectrophotometer. The FIC index and ICα were determined, the latter by estimation from the response surface approach described by Greco et al. (W. R. Greco, G. Bravo, and J. C. Parsons, Pharmacol. Rev. 47:331-385, 1995) by using a computer program developed for that purpose. For the 5FC-FCZ combination, the interactions determined by the ICα generally were in concordance with the interactions determined by the FIC index, but large discrepancies were found between both methods for the 5FC-AB combination. These could mainly be explained by shortcomings in the FIC approach. The in vitro interaction of 5FC-AB demonstrated variable results depending on the tested Candidaisolate. In general, the 5FC-FCZ combination was antagonistic against Candidaspecies, but for some Candidaisolates synergism was found. For C. neoformansthe interaction for both combinations was highly dependent on the tested isolate and the method used. Response surface approach is an alternative method for determining the interaction between antifungal agents. By using this approach, some of the problems encountered with the FIC were overcome.

ABSTRACTAlthough the fractional inhibitory concentration (FIC) index is most frequently used to define or to describe drug interactions, it has some important disadvantages when used for drugs against filamentous fungi. This includes observer bias in the determination of the MIC and no agreement on the endpoints (MIC-0, MIC-1, or MIC-2 [≥95, ≥75, and ≥50% growth inhibition, respectively]) when studying drug combinations. Furthermore, statistical analysis and comparisons are troublesome. The use of a spectrophotometric method to determine the effect of drug combinations yields quantitative data and permits the use of model fits to the whole response surface. We applied the response surface model described by Greco et al. (W. R. Greco, G. Bravo, and J. C. Parsons, Pharmacol. Rev. 47:331-385, 1995) to determine the interaction coefficient alpha (ICα) using a program developed for that purpose and compared the results with FIC indices. The susceptibilities of amphotericin B (AM), itraconazole (IT), and terbinafine (TB) were tested either alone or in combination against 10 IT-susceptible (IT-S) and 5 IT-resistant (IT-R) clinical strains of Aspergillus fumigatususing a modified checkerboard microdilution method that employs the dye MTT [3-(4,5-dimethyl-2-thiazyl)2,5-diphenyl-2H-tetrazolium bromide]. Growth in each well was determined by a spectrophotometer. FIC indices were determined and ICα values were estimated for each organism strain combination, and the latter included error estimates. Depending on the MIC endpoint used, the FIC index ranged from 1.016 to 2.077 for AM-IT, from 0.544 to 1.767 for AM-TB, and from 0.656 to 0.740 for IT-TB for the IT-S strains. For the IT-R strains the FIC index ranged from 0.308 to 1.767 for AM-IT, from 0.512 to 1.646 for AM-TB, and from 0.403 to 0.497 for IT-TB. The results indicate that the degree of interaction is not only determined by the agents themselves but also by the choice of the endpoint. Estimates of the ICα values showed more consistent results. Although the absolute FIC indices were difficult to interpret, there was a good correlation with the results obtained using the ICα values. The combination of AM with either IT or TB was antagonistic in vitro, whereas the combination of IT and TB was synergistic in vitro for both IT-S and IT-R strains. The use of response surface modeling to determine the interaction of drugs against filamentous fungi is promising, and more consistent results are obtained by this method than by using FIC indices.

Ceftazidime demonstrates time-dependent killing, which is maximal at 4 x or 5 x MIC for the organism, consequently continuous infusion (CI) has been proposed to ensure adequate ceftazidime concentrations for the entire course of therapy. Severe intra-abdominal infections (IAIs) require surgical or percutaneous drainage for management, and ceftazidime is frequently prescribed. Cardiovascular or metabolic changes and renal or liver dysfunction may alter drug pharmacokinetics during severe IAIs, and no data exist on concentrations of ceftazidime reached in the peritoneal fluid. The objectives here were to determine the pharmacokinetics of ceftazidime during continuous and intermittent administration in patients with severe IAIs, and to measure the concentrations of ceftazidime in the peritoneal exudate. Eighteen surgical patients with severe IAI and a creatinine clearance of >30 mL/min were studied. A non-randomized pilot study of six patients treated with CI alone was followed by a prospective, randomized comparative study of 12 patients. Pilot study patients received ceftazidime 1 g iv followed by a 4.5 g CI over 24 h. Randomized patients received either ceftazidime continuously as above or 1.5 g tds. Samples for pharmacokinetic analyses were collected on days 2 and 4. Ceftazidime concentrations were determined by high-performance liquid chromatography. CI resulted in a mean serum concentration >40 mg/L and a T> 4 x MIC for most pathogens encountered in severe IAIs for >90% of the course of therapy in both serum and peritoneal exudate. Eight-hourly administration resulted in T> 4 x MIC for most pathogens encountered in severe IAIs for >90% of the dosing interval, but in peritoneal exudate for only 44% of the dosing interval. During CI, AUCs in the peritoneal exudate were c. 60% of the concomitant serum AUCs. In critically ill surgical patients with severe IAIs, CI of ceftazidime resulted in more favourable concentrations in serum and peritoneal exudate than 8-hourly bolus infusion.

The pharmacokinetics of ceftazidime were investigated during intermittent (II) and continuous (CI) infusion in eight healthy male volunteers in a crossover fashion. The total daily dose was 75 mg/kg of body weight per 24 h in both regimens, given in three doses of 25 mg/kg/8 h (II) or 60 mg/kg/24 h with 15 mg/kg as a loading dose (CI). After the third dose (II) and during CI, serum and blister fluid samples were taken. Seven new blisters were raised for each timed sample by a suction blister technique. Blisters took 90 min to form. Samples were then taken from four blisters (A samples) and 1 h later were taken from the remaining three (B samples). The concentration of ceftazidime was determined using a high-performance liquid chromatography method. After II, the concentrations in serum immediately after infusion (t = 30 min) and 8 h after the start of the infusion were 137.9 (standard deviation [SD], 27.5) and 4.0 (SD, 0.7) micrograms/ml, respectively. The half-life at alpha phase (t1/2 alpha) was 9.6 min (SD, 4.6), t1/2 beta was 94.8 min (SD, 5.4), area under the concentration-time curve (AUC) was 285.4 micrograms.h/ml (SD, 22.7), total body clearance was 0.115 liter/h.kg (SD, 0.022), and volume of distribution at steady state was 0.178 liter/kg (SD, 0.023). The blister fluid (A) samples showed a decline in concentration parallel to that of the concentrations in serum during the elimination phase with a ratio of 1:1. The t1/2 of the A samples was 96.4 min (SD, 3.2). The concentration of ceftazidime in the B blister fluid samples was significantly higher (27%) than in the A samples over time. This shows that blisters may behave as a separate compartment and establishes the need to raise new blisters for each timed sample. The mean AUC/h during continuous infusion was 21.3 micrograms . h/ml (SD, 3.0). The total body clearance was 0.113 liter/h . kg (SD, 0.018), the urinary clearance was 0.105 liter/h . kg (SD, 0.012), and the ceftazidime/creatinine clearance ratio was 0.885. The mean AUC of blister fluid per hour was 84.5% (18.0 micrograms . h/liter; SD, 3.6) compared with that of serum. The A samples did not differ significantly from the B samples. The implications of continuous infusion of beta-lactams for treatment of serious infections are discussed.

Over a 12-month period, Corynebacterium striatum strains were isolated from clinical specimens from 14 patients admitted to a surgical intensive care unit. These isolates were identical by morphology and biotype and displayed the same antibiogram. Ten isolates were found to be the sole possible pathogen. These 10 isolates were from six patients, three of whom had signs of infection at the time of positive culture. Further typing was performed by random amplification of polymorphic DNA analysis, by which all strains were identical and were found to differ to various degrees from reference strains and from isolates found in clinical samples from other wards. In a case-control study the only independent risk factor for acquiring the strain was intubation for longer than 24 h (odds ratio, 20.09; 95% confidence interval, 2.29 to 176.09). The same strain was isolated from surfaces and from air sampled in the direct vicinity of infected patients but never from surfaces or air in other places of the ward. The strain was not isolated from the ventilators. The strain was cultured from the hands of personnel attending to infected patients, but no long-term carriers were found among members of the hospital personnel, suggesting transient carriage only. We conclude that C. striatum can cause serious nosocomial infections in surgical intensive care unit patients and may spread from patient to patient via the hands of attending personnel.

The objective of the study was to estimate the prevalence of gonorrhoea and chlamydial infections and to determine sexual and demographic correlates for these sexually transmitted diseases (STD) among visitors of an STD clinic.

The most important pharmacodynamic parameter for beta-lactam antibiotics has been shown to be the time above the MIC, which is used as an argument to administer beta-lactam antibiotics by continuous infusion. Studies in vitro and in laboratory animals comparing efficacy of continuous and intermittent infusion of beta-lactam antibiotics generally show continuous infusion to be more efficacious. While comparative trials in humans are scarce and a significant difference was only found in subgroup analysis in one study, several case-reports support the use of continuous infusion. Arguments in favour and against continuous infusion are discussed. Although dose-ranging studies have not yet been performed in humans, the results from in-vitro and in-vivo experiments indicate that 4 x MIC for the infecting bacterium would be the target concentration. Pharmacokinetic studies which have been performed in humans during continuous infusion show that serum concentrations can be predicted from total clearance or, using population pharmacokinetic modelling, the elimination rate constant as obtained during intermittent infusion. A nomogram is presented which allows calculation of the daily dose to obtain the target steady state blood concentrations suggested by the susceptibility of the infecting bacterium, usually 4 x MIC. For bacteria with a low MIC, the daily dose may be substantially lower than that used in conventional dosing regimens, while in infections which are difficult to treat as a result of more resistant bacteria, continuous infusion may be more effective than an equivalent bolus dose.

Background:Few data are available on the pharmacokinetics of multiple enteral dosing of ciprofloxacin in critically ill intensive care patients and none for those with severe gram-negative intra-abdominal infections (GNIAI).

The in vitro activity of trovafloxacin (CP-99,219), a new fluoroquinolone, was compared with the in vitro activities of other commonly used quinolones and other antimicrobial agents against 445 gram-positive microorganisms isolated between 1986 and 1995 from patients with endocarditis and those with other bloodstream infections. The MICs at which 90% of the isolates are inhibited (MIC90) of trovafloxacin for methicillin-susceptible staphylococci, viridans group streptococci, and enterococci were 0.06, 0.25, and 0.5 mg/liter, respectively. The MIC90 of trovafloxacin for vancomycin-resistant enterococci as well as for methicillin-resistant Staphylococcus aureus and methicillin-susceptible and ciprofloxacin-resistant S. aureus, isolated from sources other than blood, was 1 mg/liter. For the quinolones the rank order of activity was trovafloxacin > sparfloxacin > ciprofloxacin = ofloxacin > pefloxacin. Depending on the species tested, trovafloxacin was 4- to 64-fold more active than ciprofloxacin. Further experimental and in vivo studies are warranted to evaluate the efficacy of trovafloxacin in the treatment of bacterial endocarditis and other infections caused by gram-positive organisms.

We developed and applied pharmacokinetic-pharmacodynamic (PK-PD) models to characterize in vitro bacterial rate of killing as a function of ceftazidime concentrations over time. For PK-PD modeling, data obtained during continuous and intermittent infusion of ceftazidime in Pseudomonas aeruginosa killing experiments with an in vitro pharmacokinetic model were used. The basic PK-PD model was a maximum-effect model which described the number of viable bacteria (N) as a function of the growth rate (lambda) and killing rate (epsilon) according to the equation dN/dt = [lambda - epsilon x [Cgamma(EC50gamma + Cgamma)]] N, where gamma is the Hill factor, C is the concentration of antibiotic, and EC50 is the concentration of antibiotic at which 50% of the maximum effect is obtained. Next, four different models with increasing complexity were analyzed by using the EDSIM program (MediWare, Groningen, The Netherlands). These models incorporated either an adaptation rate factor and a maximum number of bacteria (Nmax) factor or combinations of the two parameters. In addition, a two-population model was evaluated. Model discrimination was by Akaike's information criterion. The experimental data were best described by the model which included an Nmax term and a rate term for adaptation for a period up to 36 h. The absolute values for maximal growth rate and killing rate in this model were different from those in the original experiment, but net growth rates were comparable. It is concluded that the derived models can describe bacterial growth and killing in the presence of antibiotic concentrations mimicking human pharmacokinetics. Application of these models will eventually provide us with parameters which can be used for further dosage optimization.

Postinfusion data obtained from 17 patients with cystic fibrosis participating in two clinical trials were used to develop population models for ceftazidime pharmacokinetics during continuous infusion. Determinant (D)-optimal sampling strategy (OSS) was used to evaluate the benefits of merging four maximally informative sampling times with population modeling. Full and sparse D-optimal sampling data sets were analyzed with the nonparametric expectation maximization (NPEM) algorithm and compared with the model obtained by the traditional standard two-stage approach. Individual pharmacokinetic parameter estimates were calculated by weighted nonlinear least-squares regression and by maximum a posteriori probability Bayesian estimator. Individual parameter estimates obtained with four D-optimally timed serum samples (OSS4) showed excellent correlation with parameter estimates obtained by using full data sets. The parameters of interest, clearance and volume of distribution, showed excellent agreement (R2 = 0.89 and R2 = 0.86). The ceftazidime population models were described as two-compartment kslope models, relating elimination constants to renal function. The NPEM-OSS4 model was described by the equations kel = 0.06516+ (0.00708.CLCR) and V1 = 0.1773 +/- 0.0406 liter/kg where CLCR is creatinine clearance in milliliters per minute per 1.73 m2, V1 is the volume of distribution of the central compartment, and kel is the elimination rate constant. Predictive performance evaluation for 31 patients with data which were not part of the model data sets showed that the NPEM-ALL model performed best, with significantly better precision than that of the standard two-stage model (P < 0.001). Predictions with the NPEM-OSS4 model were as precise as those with the NPEM-ALL model but slightly biased (-2.2 mg/liter; P < 0.01). D-optimal monitoring strategies coupled with population modeling results in useful and cost-effective population models and will be of advantage in clinical practice, as it allows pharmacokinetic-pharmacodynamic modeling with sparse data, thus describing the relationship between ceftazidime exposure and response in the treatment of acute exacerbations in patients with cystic fibrosis.

An in vitro pharmacokinetic model mimicking human serum drug concentrations, based on a dialyzer unit, was developed to study the efficacies of continuous infusion and intermittent administration of ceftazidime over a period of 36 h. The daily dose of ceftazidime was 300 mg/liter/24 h given either as a continuous infusion or as three bolus doses. The intermittent dosing regimen yielded peak and trough concentrations after the fourth dose of 92.3 (standard deviation, 8.0) and 1.4 (standard deviation, 0.9) mg/liter, respectively. Continuous administration yielded concentrations of approximately 20 mg/liter. To study efficacy, three Pseudomonas aeruginosa strains, ATCC 27853, CF4, and CF16, were used. The MICs of ceftazidime for these strains were 1, 4, and 16 mg/liter, respectively. Strain CF16 was killed initially during both regimens and then started to regrow. At the end of the fourth dosing interval, i.e., after 32 h, viable counts showed no difference between the regimens. Strains ATCC 27853 and CF4 were killed initially during both dosing schedules, and after the first dosing interval viable counts were similar. However, after the fourth interval, there was a marked difference between bacterial counts during continuous and intermittent infusion, being 2.2 and 2.8 log10, respectively, demonstrating a greater efficacy during continuous infusion. The results indicate that, in the absence of other factors, a sustained level of ceftazidime around or slightly above the MIC is not high enough to maintain efficacy over more than one (8-h) dosing interval. When sustained concentrations higher than four times the MIC are employed, continuous administration in this model is more efficacious than intermittent dosing.

The Gen-Probe PACE 2 assay (GP), a nonisotopic DNA probe, was evaluated by using cell culture as the method of reference. Specimens were collected from 260 men and 482 women visiting the outpatient department for sexually transmitted diseases at the University Hospital in Rotterdam, The Netherlands. The prevalences of Chlamydia culture-positive men and women were 13.2 and 8.6%, respectively. Sensitivity values for the male and female patients were 70.6 and 92.7%, respectively. Specificity values for these groups were 98.2 and 97.7%, respectively. Sensitivity was significantly lower when the number of inclusions in cell culture was low. Samples which showed a discordance between cell culture and GP results were retested by the polymerase chain reaction. If the results of the polymerase chain reaction were considered as the points of reference, the sensitivity and specificity of both GP and cell culture could be calculated. The performance of GP for females was comparable to that of cell culture. In males, the sensitivity of GP was considerably lower than that of cell culture (77.2 versus 91.4%), while specificity was somewhat higher (99.6 versus 99.1%). Compared with cell culture, the GP is a relatively simple and rapid test that is suitable for diagnosing Chlamydia infections in urogenital specimens.

We compared the Gen-Probe transcription-mediated amplification assay (AMP CT), the Abbott LCx assay, and the Roche COBAS AMPLICOR assay for the detection of Chlamydia trachomatis in a mixed population in urine samples. First-void urine, urethral specimens, and cervical specimens in females were obtained from 1,000 patients (544 males and 456 females) visiting the outpatient sexually transmitted disease clinic of our hospital. The prevalence of C. trachomatis infection was 7.7% as determined by tissue culture of urethral and cervical specimens. The sensitivities of LCx, COBAS AMPLICOR, and AMP CT compared to cell culture were 79, 86, and 78%, respectively. Sensitivity and specificity were recalculated by using a new "gold standard", i.e., a sample was considered to be true positive if two or more techniques yielded positive results. Specimens positive only by cell culture or positive in only one commercial amplification technique were retested by a previously described in-house PCR. After discordance analysis the sensitivities of LCx, COBAS AMPLICOR, and AMP CT were 84, 93, and 85%, respectively. Specificity exceeded 99% for all three assays. With each method the sensitivity was lower for urine samples from females compared to urine samples from males. By application of this new gold standard, existing differences between methods are highlighted; future evaluations of new techniques should be validated against two or more amplification assays.

The amplified Chlamydia trachomatis test (AMP-CT; Gen-Probe), a new diagnostic test for the detection of Chlamydia trachomatis, was evaluated with urine specimens from 1,000 patients visiting the outpatient department for sexually transmitted diseases at the University Hospital Rotterdam, Rotterdam, The Netherlands, by comparing the results to those of cell culture. From February 1996 to July 1996, urine samples for the AMP-CT test and urethral swabs for cell culture were collected from 544 men, while cervical swabs from 456 women were also taken for cell culture. Positive test results were obtained for 130 (13%) of the patients. AMP-CT test and cell culture results were discordant for 70 (7%) specimens. Analysis of the samples with discordant results was performed by an in-house PCR. After resolution of the discordant results, the sensitivity, specificity, and positive and negative predictive values of the AMP-CT test were 84.3, 98.8, 89.6, and 98%, respectively, for samples from females and 100, 99.2, 93.1, and 100%, respectively, for samples from males, while for cell culture these values were 72.5, 99.2, 92.5, and 98%, respectively, for samples from females and 57.4, 99.0, 86.1, and 95.4%, respectively, for samples from males. We conclude that the AMP-CT test is a fast and reliable test for the detection of C. trachomatis in urine specimens from females and, in particular, males.

Objective: To determine the effect of peak inspiratory pressure (PIP) and positive end-expiratory pressure (PEEP) on the development of bacteremia with Klebsiella pneumoniaeafter mechanical ventilation of intratracheally inoculated rats.

Synergism between two antibiotics is usually tested by a checkerboard titration technique, or by time-kill methods. Both methods have the disadvantage that synergism is determined at constant concentrations of the antibiotics, which do not reflect reality in vivo. In the present study we determined whether synergism between tobramycin and ceftazidime can be found at declining concentrations below the MIC, and whether change in dosing sequence of the antibiotics would result in differences in killing. Three monotherapy and six combination therapy schedules were tested in an in vitro pharmacokinetic model, using a Pseudomonas aeruginosa resistant to both antibiotics. During all q8h dosing schedules the peak concentration (Cmax) was adjusted to the MIC for the strain of both antibiotics. During all monotherapy regimens bacterial growth was present, while all six combination therapy schedules showed significant killing. At t = 24 h there were no differences between all combination therapy schedules, but at t = 8 h the two combination therapy schedules with administration of tobramycin once daily showed a significantly faster killing. By using the area under the killing curve (AUKC) as a parameter for synergistic killing, simultaneous combination therapy starting with tobramycin once daily was significantly better than all other regimens. We conclude that there is synergism between tobramycin and ceftazidime at declining antibiotic concentrations below the MIC, resulting in a pronounced killing of a resistant Pseudomonas strain. Infections due to resistant Pseudomonas strains could possibly be treated by a synergistic combination of these drugs.

The influence of a natural pulmonary surfactant on antibiotic activity was investigated to assess the possible use of exogenous surfactant as a vehicle for antibiotic delivery to the lung. The influence of surfactant on the bactericidal activity of amoxicillin was tested against Staphylococcus aureus and Streptococcus pneumoniae, and the influence of surfactant on the activities of ceftazidime and tobramycin was tested against Klebsiella pneumoniae, Pseudomonas aeruginosa, S. aureus, and S. pneumoniae. In vitro antibiotic activity was determined by killing curve studies in media with and without surfactant. Amoxicillin and ceftazidime activities were not changed in the presence of surfactant, except for a decreased killing rate of S. pneumoniae by ceftazidime in medium with additional rabbit serum. In contrast, killing curves with low concentrations of tobramycin (0.25x and 1x the MIC) showed a decreased level of activity of tobramycin against all pathogens tested in the presence of surfactant. With higher tobramycin concentrations (4x the MIC) killing rates were decreased less or were unchanged in the presence of surfactant. Concluding from the results of the study, both amoxicillin and ceftazidime can be combined with surfactant without the loss of activity. For mixing surfactant with tobramycin, dosages should be adjusted to overcome the partial inactivation of tobramycin by surfactant.

ABSTRACTWe assessed the effect of increasing manganese concentrations in test media (0.001 to 1,024 mg/liter) on MICs of tigecycline. For both broth microdilution (BMD) and Etests, this effect was negligible for physiological concentrations, but MICs increased when concentrations exceeded 8 mg/liter. Susceptibility testing should be performed on media with standardized low manganese content.

The kinetics of the postantibiotic effect (PAE) during one dosing interval of tobramycin against Staphylococcus aureus and Pseudomonas aeruginosa was investigated. We determined the PAE at different time points during this dosing interval of 12 h in an in vitro pharmacokinetic model simulating human pharmacokinetics in which the half-life of tobramycin was adjusted to 2.4 +/- 0.2 h. Using an enzymatic method to inactivate tobramycin, we determined PAEs in samples extracted from the model at 1, 5, 8, and 12 h, corresponding with tobramycin concentrations of 20, 5, 2, and 1 times the MIC for the test organism. The PAE decreased significantly from 2.5 h at 1 h to 0 h at 12 h. No change in MIC was observed for the strains during the experiments. We conclude that the PAE decreases with decreasing tobramycin concentrations during a 12-h dosing interval and completely disappears after the concentration has reached the MIC for the test organism. On the basis of these observations, the emphasis that is placed on the PAE in discussions about the optimal dosing interval in aminoglycoside therapy is questionable.

To determine the postantibiotic effect of aminoglycosides, two methods are currently being used to remove the test drug: repeated washing and dilution. An enzymatic inactivation method of removing gentamicin and tobramycin was developed and compared with the dilution method. This enzymatic method provides a rapid and simple alternative method of removing aminoglycosides which results in reliable postantibiotic-effect values.

Murepavadin (POL7080) represents the first member of a novel class of outer membrane protein-targeting antibiotics. It specifically interacts with LptD and inhibits lipopolysaccharide (LPS) transport.

ABSTRACTAlthough antibacterial therapy has an impact on human intestinal flora and the emergence of resistant bacteria, its role in the amplification of antimicrobial resistance and the quantitative exposure-effect relationship is not clear. An observational prospective study was conducted to determine whether and how ceftriaxone exposure is related to amplification of resistance in non-intensive care unit (non-ICU) patients. Serial stool samples from 122 extended-spectrum β-lactamase-positive (ESBL+) hospitalized patients were analyzed by quantitative real-time PCR to quantify the resistant gene blaCTX-M. Drug exposure was calculated for each patient by using a population pharmacokinetic model. Multi- and univariate regression and classification regression tree (CART) analyses were used to explore relationships between measures of exposure and amplification of blaCTX-Mgenes. Amplification of blaCTX-Mwas observed in 0% (0/11) of patients with no treatment and 33% (20/61) of patients treated with ceftriaxone. Stepwise regression analysis showed a significant association between amplification of blaCTX-Mand the plasma area under the concentration-time curve from 0 to 24 h for the unbound fraction of the drug (fAUC0–24), the maximum concentration of drug in serum for the unbound fraction of the drug (fCmax), and the duration of ceftriaxone therapy. Using CART analysis, amplification of blaCTX-Mwas observed in 11/16 (69%) patients treated for >14 days and in 9/40 (23%) patients treated for ≤14 days (P= 0.0019). In the latter group, amplification was observed in 5/7 (71%) patients with an fAUC0–24of ≥222 mg · h/liter and in 4/33 (12%) patients with lower drug exposures (P= 0.0033). A similar association was found for an fCmaxof ≥30 mg/liter (63% versus 13%, P= 0.0079). A significant association was found between the amplification of blaCTX-Mresistance genes and exposure to ceftriaxone. Both duration of treatment and degree of ceftriaxone exposure have a significant impact on the amplification of resistance genes. (The project described in this paper has been registered at ClinicalTrials.gov under identifier NCT01208519.)

ABSTRACTThe in vitrosusceptibilities of 1,099 molecularly identified clinical Candidaisolates against 8 antifungal drugs were determined using the EUCAST microdilution method. A new simple, objective, and mathematically solid method for determining epidemiological cutoff values (ECOFFs) was developed by derivatizing the MIC distribution and determining the derivatized ECOFF (dECOFF) as the highest MIC with the maximum second derivative. The dECOFFs were similar (95% agreement within 1 dilution) to the EUCAST ECOFFs. Overall, low non-wild-type/resistance rates were found. The highest rates were found for azoles with C. parapsilosis(2.7 to 9.8%), C. albicans(7%), and C. glabrata(1.7 to 2.3%) and for echinocandins with C. krusei(3.3%), C. albicans(1%), and C. tropicalis(1.7%).

Studies of two types of small liposomes, differing with respect to their lipid composition in terms of bilayer fluidity, charge, and hydrophilicity of the liposomal surface, were done to evaluate their usefulness for delivery of encapsulated therapeutic agents to sites of infection. The liposomes showed substantial localization in infected lung tissue after intravenous administration. This was demonstrated in a model of unilateral Klebsiella pneumoniae pneumonia in rats, in which the left lung was infected but the right lung of the same animal developed no infection. The degree of localization in the infected left lung was different for the two types of liposomes. For the liposome type with the longer blood residence time, containing a surface coating of polyethylene glycol, localization in the infected left lung was dependent on the liposomal dose, correlated with the intensity of infection, and reached 9% of the injected liposomal dose in severely infected rats.

To evaluate the importance of nasal carriage of Staphylococcus aureus as a risk factor for the development ofwound infection at the sternotomy site after cardiac surgery, a case-control study was done. The study population consisted of 1980 consecutive patients. Cases were all patients who developed a sternal wound infection from which S. aureus was cultured. Forty cases were identified, and 120 controls were selected. Preoperative nasal carriage of S. aureus, insulin-dependent diabetes mellitus, and younger age were identified as significant risk factors. The crude odds ratio of nasal carriage was 9.6 (95% confidence interval, 3.9–23.7). The median postoperative length of hospital stay for cases was 30 days longer than for controls. Mortality was also significantly higher for cases than for controls (10.0% and 0.8%, respectively).