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Antimicrobial Agents and Chemotherapy, January 2002, p. 203-210, Vol. 46, No. 1
0066-4804/02/$04.00+0     DOI: 10.1128/AAC.46.1.203-210.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Pharmacodynamics of Trovafloxacin and Levofloxacin against Bacteroides fragilis in an In Vitro Pharmacodynamic Model

M. L. Peterson,1,2 L. B. Hovde,2 D. H. Wright,1,2 G. H. Brown,1,2 A. D. Hoang,1,2 and J. C. Rotschafer1,2*

University of Minnesota, College of Pharmacy, Minneapolis, Minnesota 55455,1 Antibiotic Pharmacodynamic Modeling Laboratory, Regions Hospital, St. Paul, Minnesota 551012

Received 19 June 2000/ Returned for modification 29 December 2000/ Accepted 16 October 2001


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ABSTRACT
 
An in vitro pharmacodynamic investigation was conducted to explore whether the area under the concentration time curve from 0 to 24 h (AUC0–24)/MIC ratio could predict fluoroquinolone performance against Bacteroides fragilis. An in vitro model was used to generate kill curves for trovafloxacin (TVA) and levofloxacin (LVX) at AUC0–24/MIC ratios of 1 to 406 against three strains of B. fragilis (ATCC 25285, ATCC 23745, and clinical isolate M97-117). TVA and LVX were bolused prior to the start of experiments to achieve the corresponding AUC0–24/MIC ratio. Experiments were performed in duplicate over 24 h and in an anaerobic environment. Analyses of antimicrobial performance were conducted by comparing the rates of bacterial kill (K) using nonlinear regression analysis with 95% confidence intervals. Statistical significance was defined as a lack of overlap in the 95% confidence limits generated from the slope of each kill curve. For both TVA and LVX, K was maximized once an AUC0–24/MIC ratio of ≥40 was achieved and was not further increased despite a 10-fold increase in AUC0–24/MIC from approximately 40 to 400 against all three strains of B. fragilis. No significant differences were found in K between AUC0–24/MIC ratios of approximately 40 to 200. In experiments where AUC0–24/MIC ratios that were ≥ 5 and ≤ 44 were conducted, 64% demonstrated regrowth at 24 h. Resistant strains were selected in 50% of those experiments, demonstrating regrowth, which resulted in increased MICs of two- to 16-fold for both TVA and LVX. Regrowth did not occur, nor were resistant strains selected in any studies with an AUC/MIC that was > 44. Our findings suggest that fluoroquinolones provide antibacterial effects against B. fragilis in a concentration-independent manner associated with an AUC0–24/MIC ratio of ≥40. Also, the potential for the selection of resistant strains of B. fragilis may increase with an AUC0–24/MIC ratio of ≤44.


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INTRODUCTION
 
Over the past several years, clinicians have proposed an alternative method to predict antibiotic performance. By the linking of antibiotic pharmacokinetic parameters to a bacterial MIC, pharmacodynamic parameters have been identified to describe antibiotic bacterial killing properties, whether concentration dependent or concentration independent (3–5, 14, 21, 22). These parameters include maximum concentration of drug in serum/MIC ratio (Cmax/MIC), area under the concentration-time curve/MIC ratio (AUC/MIC), and time that the antibiotic remains above the MIC (T > MIC). The Cmax/MIC ratio and, more recently, the AUC/MIC ratio have been identified as predictive of fluoroquinolone performance in the treatment of gram-negative and gram-positive infections (3, 5, 9, 10, 12, 19, 21, 22; M. L. Peterson, L. B. Hovde, D. H. Wright, A. D. Hoang, and J. C. Rotschafer, Abstr. 98th Gen. Meet. Am. Soc. Microbiol., abstr. A-85, p. 52, 1998; D. H. Wright, L. B. Hovde, M. L. Peterson, A. D. Hoang, and J. C. Rotschafer, Abstr. 98th Gen. Meet. Am. Soc. Microbiol., abstr. A-86, p. 53, 1998.) Both in vitro and in vivo studies have demonstrated an AUC from 0 to 24 h (AUC0–24)/MIC ratio of 125 as optimal for bactericidal activity against gram-negative bacteria (5, 12). The AUC/MIC ratio has also been identified as a predictive parameter for fluoroquinolones in the treatment of gram-positive infections. Recent studies for the treatment of Streptococcus pneumoniae suggest that an AUC0–24/MIC ratio of 30 to 60 may be the minimum requirement for optimal fluoroquinolone activity (9, 10; Wright et al., Abstr. 98th Gen. Meet. Am. Soc. Microbiol.), well below the suggested AUC0–24/MIC ratio of 125 for gram-negative infections. As a result of these data, the AUC/MIC ratio has been suggested as a generic pharmacodynamic predictor for fluoroquinolones in the treatment of infections caused by any bacterial species. However, optimal pharmacodynamic predictors for fluoroquinolones have not been thoroughly established for the treatment of anaerobes.

Bacteroides fragilis, an obligate anaerobe of the normal flora of the gastrointestinal tract, accounts for 30 to 60% of all anaerobic isolates obtained from cases of intra-abdominal infections (16). Historically, antimicrobial regimens for the treatment of B. fragilis have been limited to select beta-lactams, clindamycin, chloramphenicol, or metronidazole. However, newly developed fluoroquinolones, such as clinafloxacin, trovafloxacin, moxifloxacin, sitafloxacin, gatifloxacin, and gemifloxacin, have demonstrated favorable in vitro and in vivo activity against some species of anaerobic bacteria (2, 6, 7, 11, 13, 18, 24; Peterson et al., Abstr. 98th Gen. Meet. Am. Soc. Microbiol.; H. M. Wexler, D. Molitoris, and S. M. Finegold, Abstr. 39th Intersci. Conf. Antimicrob. Agents Chemother., abstr. E-361, 1999).

Newer fluoroquinolones also demonstrate activity against aerobic gram-negative and gram-positive bacteria, which potentially allows these agents to be used as single-antibiotic therapy for polymicrobial infections. Although the susceptibilities of these anaerobic bacteria to fluoroquinolones have been determined, the identification of appropriate pharmacodynamic properties, such as Cmax/MIC ratio, AUC/MIC ratio, and T > MIC for the treatment of anaerobic infections, has not been clearly characterized. Recently, a study by Ross et al. exploring levofloxacin, trovafloxacin, and sparfloxacin performance against Bacteroides thetaiotaomicron, in a in vitro pharmacodynamic model, found that fluoroquinolones provide antibacterial effects in a concentration-independent manner associated with an AUC0–24/MIC ratio of ≥11, where AUC/MIC ratios of 1 to 150 were compared (20).

The purpose of this pharmacodynamic analysis was to characterize the rate and extent in which fluoroquinolones (trovafloxacin and levofloxacin) kill B. fragilis and to determine whether the pharmacodynamic parameter AUC0–24/MIC ratio could predict fluoroquinolone performance against B. fragilis.

(Data were presented at the 98th General meeting of the American Society for Microbiology, Atlanta, Ga., 1998, and the 38th Interscience Conference on Antimicrobial Agents and Chemotherapy, San Diego, Calif., 1998.)


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MATERIALS AND METHODS
 
Bacteria. Experiments were performed using three strains of B. fragilis: ATCC 23745, ATCC 25285, and clinical isolate M97-117.

Antibiotics. Antibiotics provided by their respective manufacturers were as follows: trovafloxacin from Pfizer Inc. (Groton, Conn.) and levofloxacin from R. W. Johnson Pharmaceutical Research Institute (Spring House, Pa.). Antibiotic solutions were prepared in accordance with the manufacturers" specifications, and stock solutions were stored at –80°C.

Susceptibility testing. Susceptibility testing was performed both prior to antibiotic exposure and on any colonies visible on agar plates from 24-h time points. Pre- and postexposure MICs were determined using the NCCLS guidelines for antimicrobial susceptibility testing of anaerobic bacteria and the American Society for Microbiology Clinical Microbiology Procedures Handbook method for broth microdilution susceptibility testing of anaerobic bacteria (8, 15). MIC microtiter trays containing Anaerobe Broth MIC (Difco Laboratories, Detroit, Mich.) and antibiotics were inoculated at 106 CFU/ml and were read after 48 h of anaerobic incubation in a Bactron IV anaerobic chamber at 37°C. Reference strains Pseudomonas aeruginosa ATCC 27853 and Enterococcus faecalis ATCC 29212 were used as controls in all MIC determinations.

In vitro model. The in vitro pharmacodynamic system, previously described by Zabinski et. al., was used to conduct all time-kill experiments (28). An anaerobic environment was created by placing the in vitro pharmacodynamic model within a Bactron IV anaerobic chamber (Anaerobe Systems, Morgan Hill, Calif.) with an anaerobic gas mixture of 5% hydrogen, 5% carbon dioxide, and 90% nitrogen. Briefly, the model consisted of a sealed glass chamber with a total volume of 1 liter, representing the central compartment, which was filled with Anaerobe Broth MIC (Difco Laboratories) and fitted with input and output tubing. First-order elimination pharmacokinetics were created in the model by displacing an equal volume of antibiotic-containing broth media with antibiotic-free broth media using a peristaltic pump. Pump rates were calculated based on the desired clearance (monoexponential elimination rate constant [kel] · volume of distribution), which depended on the volume of the reaction vessel and desired half-life (t1/2) of each antibiotic (t1/2 = 0.693/kel). A range of AUC0–24/MIC ratios (AUC0–24/MIC = [Cmax at 0 h/kel – Cmin at 24 h/kel]/MIC) from 1 to 500 were attempted by varying the Cmax (trovafloxacin, 0.02 to 34.7 µg/ml; and levofloxacin, 0.2 to 174 µg/ml) while maintaining a constant t1/2 of 10 h for trovafloxacin and 8 h for levofloxacin. Peak concentrations were created by bolusing fluoroquinolone into the media at the start of each experiment. Further studies were conducted at AUC0–24/MIC ratios of approximately 1 to 10 to determine the maximum rate of kill for trovafloxacin and levofloxacin against B. fragilis (ATCC 25285 and M97-117), once no increase in the rate of bacterial kill was noted in experiments, despite a 10-fold increase in AUC0–24/MIC ratio from approximately 40 to 400.

In vitro methods. The in vitro pharmacodynamic model was placed in a monitored 37°C water bath for the duration of each experiment within the Bactron IV anaerobic chamber. Each of the experiments was performed in duplicate for a duration of 24 h. Inocula were prepared by inoculating Anaerobic Broth MIC (Difco Laboratories) with B. fragilis and incubating in an anaerobic environment for 24 h at 37°C. Prior to the start of the experiments, 50 ml of an overnight inoculum was added to 200 ml of fresh broth and was reincubated for 1 h to achieve logarithmic growth. Each chemostat was inoculated by adding approximately 1/100 of the chemostat volume of the overnight suspension. A 0.5 McFarland standard was used as a guide to achieve a final inoculum of 106 CFU/ml. The final inoculum was verified by serial dilution and counting of visible colonies. Once daily dosing was simulated by bolusing the desired fluoroquinolone into each central compartment at time zero.

Samples were collected at time zero and after injection of the antibiotic at 1, 2, 3, 4, 6, 8, 12, and 24 h and were evaluated for bacterial load (numbers of CFU per milliliter). Antibiotic carryover was avoided by exposing 1-ml samples to 1 g of antimicrobial polymeric binding resin (Amberlite XAD-4; Supelco, Bellefonte, Pa.) (27). Following serial dilution, samples were plated onto Anaerobic Blood Agar (Remel, Lenexa, Kans.) and were incubated anaerobically for 48 h at 37°C. Following incubation, colonies were counted with a lower quantitative limit of accuracy for bacterial counts of 2.5 log10CFU/ml or 30 colonies on an agar plate containing 100 µl of undiluted sample.

Fluoroquinolone assay. Evaluation of fluoroquinolone concentration was conducted by obtaining 1-ml samples at 1, 8 and 24 h. A previously validated high-performance liquid chromatography assay was used for concentration determination (26). Briefly, reversed-phase, ion-paired chromatography was employed using an Alltech Adsorbosphere 7U HS C18 analytical column (150 by 4.6 mm [inside diameter]; Alltech Associates, Deerfield, Ill.) with ultraviolet detection at 280 nm. A mobile phase of acetonitrile and 0.02 M sodium phosphate buffer phase (pH 3) with 0.2% sodium dodecyl sulfate and triethylamine in a 40:60 (vol/vol) ratio was pumped at an isocratic flow rate of 1.75 ml/min. The lower limit of quantitation for all quinolones was 0.05 mg/liter. Intra and interday coefficients of variation were <10% for all assay runs.

Pharmacodynamic analysis. Time-kill curves were analyzed for the rate and extent of bacterial killing. Analyses of antimicrobial performance were performed by comparing rates of bacterial kill determined by nonlinear regression (curve fit) analysis with 95% confidence intervals using GraphPad Prism 3.0 (GraphPad Software, Inc., San Diego, Calif.). The rate of killing was determined from the start of the experiment to the time of maximal reduction in the log10CFU per milliliter. Statistical significance was defined as lack of overlap in the 95% confidence limits generated from the slope of each bacterial kill curve. The extent of bacterial killing was assessed by the presence or absence of regrowth at 24 h. Regrowth was only considered if preceded by a 3-log reduction in viable bacterial counts (99.9% kill of the initial inocula). A determination of the relationship between the rate of bacterial kill and AUC0–24/MIC was conducted and expressed as a correlation coefficient.


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RESULTS
 
Pharmacodynamic analysis. Time-kill curves for trovafloxacin and levofloxacin against two B. fragilis ATCC strains (25285 and 23745) and one clinical isolate of B. fragilis (M97-117) are presented in Fig. 1 and 2 with pharmacodynamic results summarized in Tables 1 1 and 2. All AUC/MIC ratios of ≥40 produced a 3-log kill by 12 h. The rate of bacterial kill for B. fragilis did not increase despite a 10-fold increase in AUC0–24/MIC from approximately 40 to 400 for both trovafloxacin and levofloxacin. For both trovafloxacin and levofloxacin, no significant difference was found for K between AUC0–24/MIC ratios of approximately 40 to 200 (Fig. 3). Statistical significance was defined as lack of overlap of 95% confidence limits for each rate of kill. K actually appeared to gradually decrease in many studies as AUC0–24/MIC increased from 200 to 400, as was represented by a negative correlation coefficient (Fig. 3).



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  		FIG. 1. Activity of trovafloxacin against B. fragilis (ATCC 25285) at AUC0–24/MIC ratios of 1 (—{permzspch023}—), 6 (—{permzspch064}—), 9 (—{lozenge}—), 42 (—*—), 122 ({permzspch157}), 201 (—|—), 281 ({permzspch158}), and 407 ({permzspch154}) (A); against B. fragilis (M97-117) at AUC0–24/MIC ratios of 3 (—{permzspch023}—), 6 (—{permzspch064}—), 12 (—{lozenge}—), 40 (—*—), 122 (—{permzspch157}—), 204 (—|—), 283 (—{permzspch158}—), and 405 (—{permzspch157}—) (B); and against B. fragilis (ATCC 23745) at AUC0–24/MIC ratios of 41 (—*—), 122 (—{permzspch157}—), 202 (—|—), 284 (—{permzspch158}—), and 406 (—{permzspch154}—) (C). Growth controls are represented by ···{blacklozenge}···. Data points for each graph have been presented as mean ± standard deviation. The lower limit of detection is noted as 2.5 log10CFU/ml.



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  		FIG. 2. Activity of levofloxacin against B. fragilis (ATCC 25285) at AUC0–24/MIC ratios of 1 (—{permzspch023}—), 5 (—{permzspch064}—), 10 (—{lozenge}—), 62 (—*—), 131 (—{permzspch157}—), 217 (—|—), 308 (—{permzspch158}—), and 437 (—{permzspch154}—) (A); against B. fragilis (M97-117) at AUC0–24/MIC ratios of 1 (—{permzspch023}—), 5 (—{permzspch064}—), 10 (—{lozenge}—), 44 (—*—), 131 (—{permzspch157}—), 217 (—|—), 308 (—{permzspch158}—), and 369 (—{permzspch154}—) (B); and against B. fragilis (ATCC 23745) at AUC0–24/MIC ratios of 44 (—*—), 132 ({permzspch157}), 220 (—|—), 308 (—{permzspch158}—), and 439 ({permzspch154}) (C). Growth controls are represented by ···{blacklozenge}···. Data points for each graph have been presented as mean ± standard deviation. The lower limit of detection is noted as 2.5 log10CFU/ml.


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  		TABLE 1. Pharmacodynamic results for trovafloxacina


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  		TABLE 2. Pharmacodynamic results for levofloxacina



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  		FIG. 3. Rate of bacterial kill as correlated to AUC0–24/MIC ratios for levofloxacin against B. fragilis (ATCC 25285) (—{permzspch157}—), B. fragilis (M97-117) (—{blacksquare};—), and B. fragilis (ATCC 23745) (---{blacktriangleup}---) (A); and for trovafloxacin against B. fragilis (ATCC 25285) (—{permzspch157}—), B. fragilis (M97-117) (—{blacksquare}—), and B. fragilis (ATCC 23745) ···({blacktriangleup})··· (B). The rate of kill was determined by nonlinear regression (curve fit) analysis with error bars denoting 95% confidence intervals.

Bacterial regrowth at 24 h was observed in only two studies of 30 duplicate experiments (AUC0–24/MIC = 40 to 400) where levofloxacin was evaluated at an AUC0–24/MIC of 44 against B. fragilis M97-117 and where trovafloxacin was evaluated at an AUC0–24/MIC of 42 against B. fragilis ATCC 25285. In studies performed at an AUC0–24/MIC of approximately 125 to 400, concentrations of both levoflovacin and trovafloxacin remained above the MIC of the B. fragilis isolates for the entire 24-h dosing interval. T > MIC in studies conducted at AUC0–24/MIC ratios of 40 to 44 for trovafloxacin and levofloxacin were approximately 18 h or 75 to 79% of the dosing interval and 17 h or 71 to 88% of the dosing interval, respectively.

All experiments conducted at AUC0–24/MIC ratios of 9 to 12, where the Cmax/MIC = 0.8 to 1, revealed a 3-log decrease in viable bacterial counts followed by regrowth at 24 h. The exception was levofloxacin, which achieved a 2.8-log decrease in bacterial load at 8 h against B. fragilis M97-117. In experiments conducted at an AUC0–24/MIC ratio of 10 for levofloxacin against B. fragilis (ATCC 25285), the rate of bacterial kill was significantly different from AUC0–24/MIC ratios of 62 to 308. In experiments conducted at AUC0–24/MIC ratios of 9 for trovafloxacin against B. fragilis (ATCC 25285), the rate of bacterial kill was significantly different from AUC0–24/MIC ratios of 42 and 122. However, in experiments conducted at AUC0–24/MIC ratios of 12 for trovafloxacin against B. fragilis M97-117, the rate of bacterial kill was not significantly different from AUC0–24/MIC ratios of 40, 122, and 204. A 3-log decrease in initial bacterial load was not achieved in any of the studies where the AUC0–24/MIC ratio was <10.

Susceptibility testing. Susceptibilities of the three strains of B. fragilis prior to and following antibiotic exposure are listed in Tables 1 and 2. Changes in susceptibility of B. fragilis following exposure to levofloxacin and trovafloxacin were noted in studies where the AUC0–24/MIC was ≥6 and ≤44. Sixty-four percent of studies with AUC0–24/MIC ratios of ≥5 and ≤44 produced regrowth at 24 h, and 50% of those studies with regrowth selected for resistant isolates with 2- to 16-fold increases in MICs of trovafloxacin and levofloxacin (Tables 1 and 2). Further susceptibility tests with other fluoroquinolones (clinafloxacin, sparfloxacin, and ciprofloxacin), performed on all isolates of B. fragilis with decreased susceptibility to levofloxacin and trovafloxacin at 24 h, demonstrated similar increases in MICs of four- to 32-fold (data not shown). Susceptibility testing performed on 2-, 4-, 6-, 8-, 12-, and 24-h strains from a study selecting for resistance (trovafloxacin AUC0–24/MIC = 12 versus B. fragilis M97-117) revealed that resistance was selected by 12 h (data not shown).


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DISCUSSION
 
We performed an in vitro pharmacodynamic analysis in an attempt to characterize the rate and extent in which fluoroquinolones (trovafloxacin and levofloxacin) kill B. fragilis and to determine whether the pharmacodynamic parameter AUC/MIC ratio could predict fluoroquinolone performance against B. fragilis.

Our study identifies fluoroquinolones as concentration-independent killers of B. fragilis once an AUC0–24/MIC ratio of ≥40 is achieved. This is demonstrated by no further increase in bacterial kill when AUC0–24/MIC ratios were increased 10-fold from 40 to 400 and by no significant difference in the rate of kill between AUC0–24/MIC ratios of 40 to 200. In fact, a significant decrease in bacterial kill occurred when AUC0–24/MIC ratios increased from 200 to 400, as was demonstrated by a negative correlation coefficient. Experiments performed at AUC0–24/MIC ratios of ≤12 were inconclusive, as statistical differences from experiments performed at AUC0–24/MIC ratios of 40 to 200 were noted as well as regrowth at 24 h and selection of resistant strains.

Previous in vitro and in vivo studies have suggested that certain pharmacodynamic parameters correlate with the therapeutic efficacy of fluoroquinolones (5, 9, 10, 12, 19; Peterson et al., Abstr. 98th Gen. Meet. Am. Soc. Microbiol.; Wright et al., Abstr. 98th Gen. Meet. Am. Soc. Microbiol). Forrest et al. described the pharmacodynamic parameter area under the inhibitory time curve of ≥125 serum inhibitory titer (SIT)–1 · h to be predictive of both clinical and microbiological cures in a clinical study of ciprofloxacin in the treatment of nosocomial pneumonia (5). Similarly, Madaras-Kelly et al. produced equivalent antibacterial activity for ciprofloxacin and ofloxacin against P. aeruginosa when both were dosed at an AUC0–24/MIC ratio of 100 (12). In a report evaluating pharmacodynamic parameters of fluoroquinolones in the treatment of respiratory tract, skin, or urinary tract infections, a Cmax/MIC ratio of ≥12.2 was correlated with efficacy (19). A culmination of these data and others suggests the use of the AUC0–24/MIC ratio of 100 and Cmax/MIC ratio of ≥12.2 as generic pharmacodynamic predictors in comparisons of fluoroquinolone activity irrespective of bacterial species. However, data published concerning fluoroquinolones in the treatment of upper respiratory tract infections caused by S. pneumoniae suggest that an AUC0–24/MIC ratio of 30 to 60 appears to optimize the performance of fluoroquinolones against these pathogens (9, 10, Wright et al., Abstr. 98th Gen. Meet. Am. Soc. Microbiol.).

Surprisingly, the results of this study correspond closely with pharmacodynamic results published regarding gram-positive organisms, despite B. fragilis being a gram-negative anaerobe. Ross et al. recently reported similar findings in an investigation of fluoroquinolone pharmacodynamic parameters for B. thetaiotaomicron. Fluoroquinolones were found to provide antibacterial effects in a concentration-independent manner associated with an AUC0–24/MIC ratio of ≥11, where AUC/MIC ratios of 1 to 150 were compared. However, increases in MICs were noted following exposure at AUC/MIC ratios of 6 to 14 (20).

An unexpected finding of this study was the correlation between AUC0–24/MIC ratios of ≥6 and ≤44 and the selection for resistance. Corresponding Cmax values, which produced resistant isolates for trovafloxacin and levofloxacin, were 0.13 µg/ml to 1.7 µg/ml and 2.0 µg/ml to 8.7 µg/ml, respectively. Similar to clinically expected peak concentrations in humans for parenterally dosed alatrovafloxacin (doses of 200 to 300 mg; concentrations of 2 to 3 µg/ml) (Trovan package insert; Pfizer, Inc.) and for levofloxacin (dose of 500 mg; concentrations of 6.4 to 8.7 µg/ml) (Levaquin package insert; Ortho-McNeil Pharmaceutical, Inc.). The relationship of the pharmacodynamic parameter, AUC0–24/MIC ratio, and the production of resistance has been evaluated in a study by Thomas et al., where an AUC0–24/MIC ratio that was <100 was correlated with the selection of resistance of gram-negative organisms to fluoroquinolones (25). An AUC0–24/MIC ratio of <100 was correlated not only with antibacterial failure, whereas an AUC0–24/MIC ratio that was ≥ 100 has been shown to be predictive of fluoroquinolone efficacy in the treatment of gram-negative organisms, but was also correlated with the selection of bacterial resistance. The data of Thomas et al. correspond with our study, where an AUC0–24/MIC ratio that was ≤44 was correlated with the selection of bacterial resistance in B. fragilis isolates.

Results of the potential for selection of resistance with an AUC0–24/MIC that was ≤44 from our study raise concerns regarding the clinical application of fluoroquinolones in the treatment of other infections caused by aerobic gram-negative and gram-positive organisms (i.e., pneumonia, skin and skin structure infections, and urinary tract infections) and the potential for the inadvertent selection of resistant B. fragilis isolates of the intestinal microflora. NCCLS-approved MIC breakpoints for susceptibility of Bacteroides spp. to fluoroquinolones are as follows: sensitive, ≤2 µg/ml, intermediate, 4 µg/ml, and resistant, ≥8 µg/ml (15). Preantibiotic exposure MICs in this study demonstrated that all strains of B. fragilis were sensitive to trovafloxacin, while strains of B. fragilis were either sensitive or intermediate to levofloxacin. Following 24 h of antibiotic exposure to levofloxacin (2 to 8.7 µg/ml) or trovafloxacin (0.13 to 1.7 µg/ml), strains of B. fragilis intermediately sensitive to trovafloxacin and intermediately sensistive or resistant to levofloxacin were selected with MICs that were ≥4 µg/ml. These resistant strains of B. fragilis were also found to be resistant to trovafloxacin, clinafloxacin, and sparfloxacin. A study recently published by Oh et al. provides clinical relevance to our concerns; fluoroquinolone-resistant B. fragilis isolates were obtained from 9 of 12 healthy volunteers following a 7-day exposure to clinafloxacin (200 mg) orally twice daily (17). Furthermore, a study published by Snydman et al. showed a 3.6% change in the susceptibility of the B. fragilis group isolated in 1995 and 1996 to trovafloxacin, from 3.7% in 1995 to 7.3% in 1996, although trovafloxacin was not approved by the U.S. Food and Drug Administration until December 1997 (23). Finally, a recently reported multicenter study by Aldridge et. al. reported that 93% of B. fragilis isolates were susceptible to trovafloxacin from 1998 to 1999 (1).

Clinically, protein binding my also be a factor in reducing the AUC/MIC ratio below 40, potentially resulting in an overall reduction in fluoroquinolone performance and an increase in selection of resistant isolates of B. fragilis. The predicted AUC for Trovan (trovafloxacin) (package insert; Pfizer, Inc.) with a 200-mg oral daily dose is 34.4. Protein binding is 76%; therefore, the free AUC is 8.3. The corresponding AUC/MIC ratios for organisms for which MICs were 0.25, 0.5, 1, and 2 µg/ml would be 33, 17, 8, and 4, respectively. Since the MICs of trovafloxacin were 0.25 to 1 µg/ml in this study, an AUC/MIC range of 8 to 33 would be expected. Likewise, for levofloxacin, the predicted AUC for Levaquin (package insert; Ortho-McNeil Pharmaceuticals, Inc.) with a 500-mg daily dose is 47.5. Protein binding is 24 to 38%; therefore, the free AUC is approximately 32.8. The corresponding AUC/MIC ratios for organisms for which the MICs were 0.5, 1, 2, and 4 µg/ml would be 66, 33, 16, and 8, respectively. Since the MICs in this study were 2 and 4 µg/ml, an AUC/MIC ratio range of 8 to 16 would be expected. The kill studies from this research suggest, with AUC/MIC ratios of 8 to 33, that a 3-log kill would likeliest be achieved with an AUC/MIC ratio of ≥10; however, an AUC/MIC ratio of ≤44 was found to select for resistance.

In conclusion, this study represents an attempt to further understand the appropriate use of fluoroquinolones in the treatment of anaerobic infections. Fluoroquinolones appeared to exhibit concentration-independent killing activity with optimization of bacterial kill at an AUC0–24/MIC ratio of ≥40 in the treatment of B. fragilis. Unfortunately, an AUC0–24/MIC that is ≤44 may be correlated with the selection of bacterial resistance and regrowth. Therefore, underdosing, limited antibiotic penetration at the site of infection, protein binding, and susceptibility of the infecting organism should be of consideration when making clinical decisions that may affect both antibiotic efficacy and the selection of bacterial resistance. More important, inadvertent exposure of gastrointestinal microflora, such as B. fragilis, to fluoroquinolones through their use in the treatment of infections caused by aerobic gram-positive or gram-negative organisms could potentially predispose anaerobes to select for resistance.


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ACKNOWLEDGMENTS
 
Work was supported by an educational grant from the American College of Clinical Pharmacy and a grant from Pfizer Inc., Groton, Conn.

We also thank Ronald Sawchuck and Cynthia Gross for their help in the statistical evaluation of these data.


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FOOTNOTES
 
* Corresponding author. Mailing address: Department of Clinical Pharmacy, Regions Hospital, 640 Jackson St., St. Paul, MN 55101. Phone: (651) 254-3896. Fax: (651) 292-4031. E-mail: rotsc001{at}tc.umn.edu.. Back


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REFERENCES
 
    1
  1. Aldridge, K. E., D. Ashcraft, K. Cambre, C. L. Pierson, S. G. Jenkins, and J. E. Rosenblatt. 2001. Multicenter survey of the changing in vitro antimicrobial susceptibilities of clinical isolates of Bacteroides fragilis group, Prevotella, Fusobacterium, Porphyromonas, and Peptostreptococcus species. Antimicrob. Agents Chemother. 45:1238–1243.[Abstract/Free Full Text]
    2. 2
  2. Citron, D. M., and M. D. Appleman. 1997. Comparative in vitro activities of trovafloxacin (CP-99,219) against 211 aerobic and 217 anaerobic bacteria isolated from patients with intra-abdominal infections. Antimicrob. Agents Chemother. 41:2312–2316.[Abstract/Free Full Text]
    3. 3
  3. Craig, W. A. 1998. Pharmacokinetic/pharmacodynamic parameters: rationale for antibacterial dosing of mice and men. Clin. Infect. Dis. 26:1–12.[Medline]
    4. 4
  4. Craig, W. A., and S. C. Ebert. 1990. Pharmacodynamic properties of antibiotics: application to drug monitoring and dosage regimen design. Infect. Control Hosp. Epidemiol. 11:319–326.[Medline]
    5. 5
  5. Forrest, A., D. E. Nix, C. H. Ballow, T. F. Goss, M. C. Birmingham, and J. J. Schentag. 1993. Pharmacodynamics of intravenous ciprofloxacin in seriously ill patients. Antimicrob. Agents Chemother. 37:1073–1081.[Abstract/Free Full Text]
    6. 6
  6. Goldstein, E. J. 1996. Possible role for the new fluoroquinolones (levofloxacin, grepafloxacin, trovafloxacin, clinafloxacin, sparfloxacin, and DU-6859a) in the treatment of anaerobic infections: review of current information on efficacy and safety. Clin. Infect. Dis. 23(Suppl. 1):S25–S30.[Medline]
    7. 7
  7. Goldstein, E. J. C., D. M. Citron, Y. Warren, K. Tyrrell, and C. V. Merriam. 1999. In vitro activity of gemifloxacin (SB 265805) against anaerobes. Antimicrob. Agents Chemother. 43:2231–2235.[Abstract/Free Full Text]
    8. 8
  8. Isenberg, H. D. (ed.). 1992. Clinical microbiology procedures handbook. American Society for Microbiology, Washington, D.C.
    9. 9
  9. Lacy, M. K., W. Lu, X. Xu, P. R. Tessier, D. P. Nicolau, R. Quintiliani, and C. H. Nightingale. 1999. Pharmacodynamic comparisons of levofloxacin, ciprofloxacin, and ampicillin against Streptococcus pneumoniae in an in vitro model of infection. Antimicrob. Agents Chemother. 43:672–677.[Abstract/Free Full Text]
    10. 10
  10. Lister, P. D., and C. C. Sanders. 1999. Pharmacodynamics of trovafloxacin, ofloxacin, and ciprofloxacin against Streptococcus pneumoniae in an in vitro pharmacokinetic model. Antimicrob. Agents Chemother. 43:1118–1123.[Abstract/Free Full Text]
    11. 11
  11. MacGowan, A. P., K. E. Bowker, H. A. Holt, M. Wootton, and S. S. Reeves. 1997. Bay 12-8039, a new 8-methoxyquinolone: comparative in-vitro activity with nine other antimicrobials against anaerobic bacteria. J. Antimicrob. Chemother. 40:503–509.[Abstract/Free Full Text]
    12. 12
  12. Madaras-Kelly, K. J., B. E. Ostergaard, L. B. Hovde, and J. C. Rotschafer. 1996 Twenty-four-hour area under the concentration-time curve/MIC ratio as a generic predictor of fluoroquinolone antimicrobial effect using three strains of Pseudomonas aeruginosa and an in vitro pharmacodynamic model. Antimicrob. Agents Chemother. 40:627–632.[Abstract/Free Full Text]
    13. 13
  13. Milatovic, D., F. J. Schmitz, S. Brisse, J. Verhoef, and A. C. Fluit. 2000. In vitro activities of sitafloxacin (DU-6859a) and six other fluoroquinolones against 8,796 clinical bacterial isolates. Antimicrob. Agents Chemother. 44:1102–1107.[Abstract/Free Full Text]
    14. 14
  14. Moore, R. D., P. S. Leitman, and C. R. Smith. 1987. Clinical response to aminoglycoside therapy: importance of the ratio of peak concentration to minimal inhibitory concentration. J. Infect. Dis. 155:93–99.[Medline]
    15. 15
  15. National Committee for Clinical Laboratory Standards. 2001. Methods for antimicrobial susceptibility testing of anaerobic bacteria, 5th ed. Approved standard. NCCLS document M11-A5. National Committee for Clinical Laboratory Standards, Wayne, Pa.
    16. 16
  16. Nichols, R. L., and J. W. Smith. 1993. Wound and intraabdominal infections: microbiological considerations and approaches to treatment. Clin. Infect. Dis. 16:S266–S272.[Medline]
    17. 17
  17. Oh, H. C. E. Nord, L. Barkholt, M. Hedberg, and C. Edlund. 2000. Ecological disturbances in intestinal microflora caused by clinafloxacin, an extended-spectrum quinolone. Infection 28:272–277.[CrossRef][Medline]
    18. 18
  18. Peterson, M. L., L. B. Hovde, D. H. Wright, A. D. Hoang, J. K. Raddatz, P. J. Boysen, and J. C. Rotschafer. 1999. Fluoroquinolone resistance in Bacteroides fragilis following sparfloxacin exposure. Antimicrob. Agents Chemother. 43:2251–2255.[Abstract/Free Full Text]
    19. 19
  19. Preston, S. L., G. L. Drusano, A. L. Berman, C. L. Fowler, A. T. Chow, B. Dornseif, V. Reichl, J. Natarajan, and M. Corrado. 1998. Pharmacodynamics of levofloxacin: a new paradigm for early clinical trials. JAMA 279:125–129.[Abstract/Free Full Text]
    20. 20
  20. Ross, G. H., D. H. Wright, L. B. Hovde, M. L. Peterson, and J. C. Rotschafer.2001. Fluoroquinolone resistance in anaerobic bacteria following exposure to levofloxacin, trovafloxacin, and sparfloxacin in an in vitro pharmacodynamic model. Antimicrob. Agents Chemother. 45:2136–2140.
    21. 21
  21. Schentag, J. J., K. K. Gilliland, and J. A. Paladino. 2001. What have we learned from pharmacokinetic and pharmacodynamic theories? Clin. Infect. Dis. 32(Suppl. 1):S39–S46.
    22. 22
  22. Schentag, J. J., D. E. Nix, and M. H. Adelman. 1991. Mathematical examination of dual individualization principles: relationships between AUC above MIC and area under the inhibitory curve for cefmenoxime, ciprofloxacin, and tobramycin. Ann. Pharmacother. 25:1050–1057.[Abstract]
    23. 23
  23. Snydman, D. R., N. V. Jacobus, L. A. McDermott, S. Supran, G. J. Cuchural, Jr., S. Finegold, L. Harrell, D. W. Hecht, P. Iannini, S. Jenkins, C. Pierson, J. Rihs, and S. L. Gorbach. 1999. Multicenter study of in vitro susceptibility of the Bacteroides fragilis group, 1995 to 1996, with comparison of resistance trends from 1990 to 1996. Antimicrob. Agents Chemother. 43:2417–2422.[Abstract/Free Full Text]
    24. 24
  24. Spangler, S. K., M. R. Jacobs, and P. C. Appelbaum. 1994. Activity of CP 99,219 compared with those of ciprofloxacin, grepafloxacin, metronidazole, cefoxitin, piperacillin, and piperacillin-tazobactam against 489 anaerobes. Antimicrob. Agents Chemother. 38:2471–2476.[Abstract/Free Full Text]
    25. 25
  25. Thomas, J. K., A. Forrest, S. Bhavnani, J. M. Hyatt, A. Cheng, C. H. Ballow, and J. J. Schentag. 1998. Pharmacodynamic evaluation of factors associated with the development of bacterial resistance in acutely ill patient during therapy. Antimicrob. Agents Chemother. 42:521–527.[Abstract/Free Full Text]
    26. 26
  26. Wright, D. H., V. K. Herman, F. N. Konstantinides, and J. C. Rotschafer. 1998. Determination of quinolone antibiotics in growth media by reversed-phase high-performance liquid chromatography. J. Chromatog. 709:97–104.
    27. 27
  27. Zabinski, R. A., A. J. Larsson, K. J. Walker, S. S. Gilliland, and J. C. Rotschafer. 1993. Elimination of quinolone antibiotic carryover through use of antibiotic-removal beads. Antimicrob. Agents Chemother. 37:1377–1379.[Abstract/Free Full Text]
    28. 28
  28. Zabinski, R. A., K. Vance-Bryan, A. J. Krinke, K. J. Walker, J. A. Moody, and J. C. Rotschafer. 1993. Evaluation of the activity of temafloxacin versus Bacteroides fragilis using an in vitro pharmacodynamic system. Antimicrob. Agents Chemother. 37:2454–2458.[Abstract/Free Full Text]

Antimicrobial Agents and Chemotherapy, January 2002, p. 203-210, Vol. 46, No. 1
0066-4804/02/$04.00+0     DOI: 10.1128/AAC.46.1.203-210.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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