Susceptibility of the Bacteroides fragilis group to newer quinolones and other standard anti-anaerobic agents

Ruth Horn,* and Hugh G. Robson

Department of Microbiology, Royal Victoria Hospital, McGill University Health Centre, 687 Pine Avenue West, L5.06, Montreal, Quebec, Canada H3A 1A1


    Abstract
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
The susceptibilities of 200 clinical isolates of the Bacteroides fragilis group to four quinolones (moxifloxacin, clinafloxacin, trovafloxacin and ciprofloxacin) were determined, as well as to cefoxitin, clindamycin, metronidazole, imipenem and ticarcillin–clavulanic acid. The results for the latter five agents were compared with those of a study on 200 isolates done 6 years previously. Clinafloxacin and trovafloxacin were the most active agents tested with MIC90s lower than all other antimicrobials except imipenem. Susceptibility rates for imipenem, ticarcillin– clavulanic and metronidazole continue to be high, although resistant strains are emerging. For ticarcillin–clavulanic acid and metronidazole, MIC90s increased four- to eight-fold for the B. fragilis species between the two study periods.


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Members of the Bacteroides fragilis group of organisms are the most common clinical anaerobic isolates. Antimicrobial resistance among these organisms has been known to vary between species, institutions and countries.1,2 Nationwide surveys in the USA and Canada have documented increases in antimicrobial resistance in these bacteria.2,3 Antibiotics with the best activity against anaerobes include metronidazole, ß-lactam–ß-lactamase inhibitor combinations and imipenem. Resistance to clindamycin and most cephalosporins is common.13 The development of new fluoroquinolone antimicrobials has provided a group of agents with good activity against a wide variety of organisms. Among these, moxifloxacin, clinafloxacin and trovafloxacin have been reported to have improved in vitro activity against anaerobes.46

Susceptibility testing of anaerobic bacteria is not performed routinely in most hospital laboratories and initial treatment of anaerobic infections is therefore usually empirical, based on knowledge of susceptibility results of recent clinical isolates. For this reason, as well as concern over emerging resistance, there is a need to test susceptibility to new agents and to document changing patterns of antimicrobial resistance among anaerobic bacteria. To achieve this, we collected 200 isolates of the B. fragilis group from January 1996 to December 1997 and compared their susceptibility with the same number of organisms isolated and tested in 1990–1991.1 In addition to cefoxitin, clindamycin, ticarcillin–clavulanic acid, metronidazole and imipenem, the more recent collection was also tested against ciprofloxacin, trovafloxacin, clinafloxacin and moxifloxacin.


    Materials and methods
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Non-duplicated clinical isolates of the B. fragilis group were collected and frozen in brain–heart infusion broth (Beckton Dickinson Diagnostic Systems, Sparks, MD, USA) with glycerol at –70°C. The sources of the organisms were surgical wounds (57), abdominal fluids (55), skin ulcers (33), blood (20), abscesses (15) and other sites (20). The anaerobes were thawed and subcultured on to blood agar plates twice. Plates were incubated at 37°C in Gas-Pak jars (BBL Microbiology Systems, Cockeysville, MD, USA) for 48 h. Determination of species was done by the Rapid Ana II system (Innovative Diagnostic Systems Inc., Atlanta, GA, USA). The strains from 1990–1991 had been speciated using the Minitek System (BBL). The following standard antimicrobial powders were kindly provided by the indicated manufacturers: cefoxitin and imipenem, Merck Frosst Canada Inc., Pointe-Claire, Quebec, Canada; clindamycin, The UpJohn Company of Canada, Don Mills, Ontario, Canada; metronidazole, Rhône-Poulenc Pharma Inc., Montreal, Quebec, Canada; ticarcillin–clavulanic acid, SmithKline Beecham Laboratories, Oakville, Ontario, Canada; ciprofloxacin and moxifloxacin, Bayer Inc., Etobicoke, Ontario, Canada; clinafloxacin, Parke-Davis, Scarborough, Ontario, Canada; and trovafloxacin, Pfizer Canada, Kirkland, Quebec, Canada. Antimicrobial susceptibility testing was performed by the broth microdilution method recommended by the NCCLS7 during both testing periods. With each susceptibility run, B. fragilis ATCC 25285 and Bacteroides thetaiotaomicron ATCC 29741 were included as controls.


    Results
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
The results obtained for the three new quinolones are compared with those for ciprofloxacin in Table IGo. Clinafloxacin was the most active, followed in order by trovafloxacin and moxifloxacin. As expected, ciprofloxacin showed little activity. Both clinafloxacin and trovafloxacin had MIC50 and MIC90 values lower than all the other antimicrobials tested except imipenem. For the entire B. fragilis group as well as for the B. fragilis species, the MIC90 results were: clinafloxacin 1 mg/L, trovafloxacin 4 mg/L and moxifloxacin 8 mg/L. Bacteroides vulgatus isolates were more resistant to all quinolones tested; the MIC90 of both clinafloxacin and trovafloxacin was 16 mg/L. B. thetaiotaomicron isolates were the most susceptible.


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Table I. Activities of antimicrobial agents
 
Susceptibility results for the other antimicrobials were compared with those of 200 strains isolated in 1990–1991.1 Speciation of the more recent isolates yielded a somewhat different strain distribution to that in the previous study. The B. fragilis species was more numerous (132 versus 100 strains) as was B. thethaiotaomicron (20 versus 13 strains), while Bacteroides distasonis strains were fewer (nine versus 41 in the previous study). These differences may be due to speciation methods as two different methods were used.

A comparison of activity for five antimicrobial agents against the most frequent isolates is given in Table IIGo. The most active agents in both studies were imipenem and ticarcillin–clavulanic acid. Whereas no resistance was found in 1990–1991, three strains were found to be resistant to imipenem (two B. fragilis sp., one B. thetaiotaomicron) and two strains were resistant to ticarcillin–clavulanic acid (one B. fragilis sp., one B. vulgatus) in 1996–1997. Four strains were resistant to metronidazole in the more recent study (three B. fragilis sp., one B. vulgatus) compared with only one previously (B. distasonis). Cefoxitin resistance remained virtually unchanged and, as seen previously, the MIC for many organisms fell on or within one dilution of the breakpoint. Overall, clindamycin resistance rose slightly from 6 to 10.5%; however, among B. fragilis sp., B. vulgatus and B. thetaiotaomicron, resistance rates doubled between 1990–1991 and 1996–1997. B. thetaiotaomicron and B. vulgatus resistance was 15% and 18%, respectively.


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Table II. Comparative susceptibility of B. fragilis organisms between 1990–1991 and 1996–1997a
 
The comparison of MIC90 results between the two study periods showed large shifts for ticarcillin–clavulanic acid and metronidazole, particularly among organisms of the B. fragilis species. MIC90s rose from 1 to 8 mg/L of ticarcillin–clavulanic acid and from 2 to 8 mg/L of metronidazole. MIC90s also rose from 1 to 64 mg/L of clindamycin for B. vulgatus species. Values for imipenem remained stable between the study periods.


    Discussion
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 Abstract
 Introduction
 Materials and methods
 Results
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 References
 
While imipenem, metronidazole and ticarcillin–clavulanic acid still appear active against members of the B. fragilis group as judged by percentage strain susceptibility at breakpoint concentrations, we note the emergence of some resistance indicators. Isolates with high MICs (>=64) for imipenem have begun to appear, as noted by others.8 To date, resistance of B. fragilis to metronidazole has rarely been reported. The resistance rate of 2% to metronidazole (up from 0.5%) and the rise in the MIC90 to 8 mg/L for isolates of the B. fragilis species are interesting. Since the susceptibility breakpoint for metronidazole is 8 mg/L, one may see a rise in resistance rate if the trend toward higher MICs continues. Although resistance to ticarcillin–clavulanic acid has remained low at 1%, the rise in MIC90 values is notable and has been seen by others.8 Since chart reviews were not done, it is not known whether the increase in MIC is correlated with antimicrobial use.

Although the new quinolones clearly demonstrate antibacterial activity against anaerobes, trovafloxacin and moxifloxacin were not as highly active in our study as compared with other reports. Both Wexler et al.5 and Aldridge & Ashcraft4 reported lower MICs and higher susceptibility rates for these two drugs. Clinafloxacin showed the best activity of these agents with an MIC90 of 1.0 mg/L for the entire B. fragilis group as well as for the B. fragilis species. This is also higher than a MIC90 of 0.125 mg/L for the B. fragilis species reported by Wexler et al.6 The difference in MICs between that study and our study may be due to methodology and media, which have been shown to be important when testing quinolones.8

The role of fluoroquinolone antibiotics in the treatment of infection caused by anaerobic bacteria remains to be defined. Since the agents we tested showed important in vitro activity, it is probable that they would have also shown clinical efficacy. In serious infections involving anaerobes, experts agree that there is a correlation between in vitro susceptibility results and clinical response to treatment.9 With the withdrawal from the market of both trovafloxacin and clinafloxacin, there is a void in our armamentarium against anaerobic bacteria. These agents had the potential of being used as monotherapy for intra-abdominal and pelvic infections. There is now room for new antimicrobials to be developed to replace the agents within this class that were withdrawn due to severe secondary effects. This would be particularly welcome in view of the increase in resistance to older agents.


    Notes
 
* Corresponding author. Tel: +1-514-842-1231; Fax: +1-514-844-7526. Back


    References
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
1 . Horn, R., Lavallée, J. & Robson, H. G. (1992). Susceptibilities of members of the Bacteroides fragilis group to 11 antimicrobial agents. Antimicrobial Agents and Chemotherapy 36, 2051–3.[Abstract]

2 . Tally, F. P., Cuchural, G. J., Jr, Jacobus, N. V., Gorbach, S. L., Aldridge, K., Cleary, T. et al. (1985). Nationwide study of the susceptibility of the Bacteroides fragilis group in the United States. Antimicrobial Agents and Chemotherapy 28, 675–7.[ISI][Medline]

3 . Bourgault, A.-M., Lamothe, F., Hoban, D. J., Dalton, M. T., Kibsey, P. C., Harding, G. et al. (1992). Survey of Bacteroides fragilis group susceptibility patterns in Canada. Antimicrobial Agents and Chemotherapy 36, 343–7.[Abstract]

4 . Aldridge, K. E. & Ashcraft, D. S. (1997). Comparison of the in vitro activities of BAY 12-8039, a new quinolone, and other antimicrobials against clinically important anaerobes. Antimicrobial Agents and Chemotherapy 41, 709–11.[Abstract]

5 . Wexler, H. M., Molitoris, E., Molitoris, D. & Finegold, S. M. (1996). In vitro activities of trovafloxacin against 557 strains of anaerobic bacteria. Antimicrobial Agents and Chemotherapy 40, 2232–5.[Abstract]

6 . Wexler, H. M., Molitoris, E., Reeves, D. & Finegold, S. M. (1994). In-vitro activity of clinafloxacin (CI-960) and PD 131628-2 against anaerobic bacteria. Journal of Antimicrobial Chemotherapy 34, 579–84.[Abstract]

7 . National Committee for Clinical Laboratory Standards. (1997). Methods for Antimicrobial Susceptibility Testing of Anaerobic Bacteria: Approved Standard M11-A4. NCCLS, Villanova, PA.

8 . Snydman, D. R., McDermott, L., Cuchural, G. J., Jr, Hecht, D. W., Iannini, P. B., Harrell, L. J. et al. (1996). Analysis of trends in antimicrobial resistance patterns among clinical isolates of Bacteroides fragilis group species from 1990–1994. Clinical Infectious Diseases 23, Suppl.1, S54–65.[ISI][Medline]

9 . Rosenblatt, J. E. & Brook, I. (1993). Clinical relevance of susceptibility testing of anaerobic bacteria. Clinical Infectious Diseases 16, Suppl. 4, S446–8.[ISI][Medline]

Received 14 September 2000; returned 8 January 2001; revised 23 February 2001; accepted 30 March 2001