The activity of cefotaxime and desacetylcefotaxime against Bacteroides species compared to 7-methoxy cephems and other anti-anaerobe drugs

Transcription

1 Journal of Antimicrobial Chemotherapy (984) 4, Suppl. B, The activity of cefotaxime and desacetylcefotaxime against Bacteroides species compared to 7-methoxy cephems and other anti-anaerobe drugs Ronald N. Jones* Department of Pathology, Kaiser-Permanente Medical Care Program {Oregon Region) Clackamas, Oregon 9705, U.S.A. The combination of cefotaxime and desacetylcefotaxime was tested against 83 Bacteroides strains (seven species) isolated from several hospitals. The combination greatly enhanced the anti-anaerobic spectrum of cefotaxime: 79-7% of tested strains were synergistically killed by the combination. Although the combination appears clinically efficacious, it is only comparable to cefoxitin and still less active than clindamycin, chloramphenicol and metronidazole. Introduction Cefotaxime is a broad spectrum methoxy-imino aminothiazoyl-cephalosporin with potent antimicrobial activity against most clinically important aerobic and anaerobic bacteria (Jones & Thornsberry, 98). It is rapidly metabolized in vivo to a biologically active metabolite, desacetyl-cefotaxime (Chamberlain et ai, 980; Novick, 98). Cefotaxime and the desacetyl metabolite interact synergistically, usually producing enhanced killing or inhibition for more than 70% of tested facultative anaerobic strains (Jones, Barry & Thornsberry, 98; Chin & Neu, 984). Other older cephalosporins are metabolized to desacetyl forms (Wick, 966; Cabana, Van Harken & Hottendorff, 976; Nakai et ai, 976) and these /ff-lactams also have synergistic interactions with their desacetyl-metaboli te against Gram-positive and -negative species usually susceptible to the unmetabolized drug (Jones & Packer, 984). Only two studies have explored the possible synergistic activity of desacetylcefotaxime against anaerobic bacteria (Jones et al., 984; Aldridge, Sanders & Marier, 984). In this study we extend our previous synergy investigations on the Bacteroides species. Antibiotics Materials and methods Cefotaxime and desacetyl-cefotaxime were supplied by Hoechst-Roussel Pharmaceuticals, Inc. (Somerville, N.J.), and cefotetan by Stuart Pharmaceuticals, (Wilmington, Del.); cefoxitin by Merck, Sharp and Dohme (Rahway, N.J.); clindamycin by The Upjohn Company (Kalamazoo, Mich.); chloramphenicol by Warner-Lambert Company (Detroit, Mich.) and metronidazole by Searle Pharmaceuticals (Wilmington, Del.). Downloaded from at Pennsylvania State University on September 9, 06 Organisms The Bacteroides isolates were taken from recent patient specimens at the clinical microbiology laboratories of The Cleveland Clinic Foundation (Cleveland, Ohio), Kaiser Regional Laboratory, 000 S.E. Sunnyside Road, Clackamas, Oregon 9705, U.S.A /84/4B $0.00/0 984 The British Society for Antimicrobial Chemotherapy

2 40 R. N. Jones Northwestern Memorial Hospital (Chicago,.), Kaiser & Permanente Medical Care Program Regional Laboratory (Clackamas, Oreg.) and isolates from the cefotetan clinical trials in the United States, strains from six states. The organisms were held at 70 C with lysed rabbit blood until tested. Isolate identifications were confirmed with the API anaerobe test strip and other standard procedures. The strains tested were Bacteroides bivius (3), Bad. distasonis (3), Bact.fragilis (53), Bad. melaninogenicus (6), Bad. ovatus (4), Bad. thetaiotaomicron (0) and Bad. vulgatus (4). Susceptibility testing methods The MICs for the anaerobic isolates were determined by a broth microdilution procedure previously reported (Jones et ah, 978). The antibiotics were diluted in Wilkins-Chalgren broth with the following dilution schedules: cefotaxime, 3 to 003 mg/; cefotetan, 64 to 0-5 mg/; cefoxitin, 3 to 0-5 mg/; chloramphenicol, 3 to 0-5 mg/; clindamycin, 6 to 0 mg/; desacetyl-cefotaxime, 8 to 0 mg/ and metronidazole, 6 to 0 mg/. For the broth microdilution drug interaction studies, plastic microdilution trays were prepared (Dynatech Laboratories, Alexandria, Va) in which cefotaxime was combined with its desacetyl-metabolite. Equal part ratios (:) were used ranging from 3 to 0- mg/. Each test strain was inoculated into the plastic trays at a final concentration of approximately -0 x 0 s cfu/ml. The trays were then incubated for 48 h, and MICs read and plotted as isobolograms. Minimal bactericidal concentrations (MBCs) were determined by subculture, after tray agitation, of at least 0% of the well volumes (00 ml), to drug-free 5% sheep blood agar plates. The MBC was defined as the lowest concentration resulting in a ^ 99-9% killing of the inoculum by the criteria established by Pearson et al. (980). All the synergy results presented represent the MBC endpoints. The criteria for the 83 synergy studies were the concavity of the MBC isobologram plots and a fourfold or more decrease in the MBC of one antibiotic when combined with a concentration equivalent to one-fourth of the MBC of the other. The additive interaction was defined as a fourfold or greater decrease in the MBC of one agent and a twofold reduction in the MBC of the other drug (also called partial synergy) or as a twofold decrease in the MBCs of both tested drugs (also referred to as additive). Indifference was defined as no significant decrease in the MIC or MBC of either compound or only a twofold decrease or increase in the MBC of one drug. Antagonism was defined as a fourfold or greater increase in the MBCs of either or both antimicrobial agents (Jones et al., 98, 984; Jones & Packer, 984). Clinically significant synergy would imply that the cefotaxime MBC must be, or be reduced to a drug level readily achieved in tissues (< 3 mg/) by an equal desacetylcefotaxime level. Downloaded from at Pennsylvania State University on September 9, 06 Results Table I shows the MIC results for the Bacteroides spp. using the MIC inhibiting 50% of tested strains (MIC 60 ) and the per cent of isolates susceptible by the criteria of the National Committee for Clinical Laboratory Standards (NCCLS) generally utilizing the susceptible and moderately susceptible categories for the /Mactams and only the susceptible breakpoint for the other drugs. Cefotaxime alone was two- to fourfold less active than the 7-methoxy cephalosporins against Bad. fragilis strains. However, cefotaxime was comparable or superior to cefotetan for all other six tested Bacteroides spp. The combination of cefotaxime and desacetyl-cefotaxime was superior to either drug

3 Cefotaxime and desacetyl-cefotaxime against Bacteroides 4 Table I. Minimum inhibitory concentrations inhibiting 50% of tested strains (MIC 60 ) for 83 strains of Bacteroides species determined by broth microdilution methods and Wilkins-Chalgren medium Bacteroides MIC S0 (% susceptible)* for Antibiotic fragilis (53)t thetaiotaomicron (0) melanino- Others* ( ) genicus (6) bivius (3) Cefotaxime 3(5) > Desacetyl-cefotaxime 8 (30) > Combination 6 (78) Cefoxitin 80 (9) Cefotetan 6(83) > Chloramphenicol 80 (00) Clindamycin 0-5 (96) Metronidazole 0 (00) 3 (0) > 8 (0) 3 (70) 3(40) 64(0) > 80 (00) 40 (90) 0 (00) 3(45) 8 (36) 3(64) 6(54) 64(8) 80 (00) 0(73) 0 (00) 0(83) 0(00) 0-5 (00) 0(00) 80 (83) 0 (00) 0(00) 0-5 (00) 6(00) 6(00) 40 (00) 40 (00) 6(67) 0 (00) 0(00) 0 (67) * Susceptible defined is < 3 mg/ for cefotaxime and cefotetan; < 6 mg/ for cefoxitin, < 8-0 mg/ for chloramphenicol and < 4.0 mg/ for clindamycin. t No. of strains tested in ( ). I Includes' Bad. fragilis group' species: Bad. distasonis (3 strains), Bad. ovatus (4 strains) and Bad. vulgatus (4 strains). Fixed : ratio combination of cefotaxime and desacetyl-cefotaxime. Table EL Results of antimicrobial interaction studies (synergy analysis) for fixed : combinations of cefotaxime and desacetyl-cefotaxime compared to the antimicrobial activity of the cephalosporins alone. Endpoints are minimum bactericidal concentration Bad. fragilis Bact. distasonis Bact. ovatus Bact. thetaiotaomicron Bact. vulgatus Bact. bivius Bact. melaninogenicus All strains Synergy 3 8(8)* Partial synergy (5-6) No. in ( ) = % of the interactions in that category. Drug interaction category Additive (3) Indifference 9 (7-) Indeterminate alone against the anaerobic Gram-negative bacilli. The combination was superior to cefoxitin (MIC 60 values) for Bact. melaninogenicus, equal for Bact. thetaiotaomicron and Bact. bivius and one log dilution inferior for Bact. fragilis, Bact. distasonis, Bact. ovatus and Bact. vulgatus. The overall spectrum of activity for the cefotaxime-desacetylcefotaxime combination included 4 of 53 Bact. fragilis strains and 3 of 30 non- Bact. fragilis, or 77- % of strains inhibited at < 3 mg/. The comparable in-vitro efficacy for cefoxitin was 80-7% (P = > 0-5, versus the combination) and cefotetan was 63-9% (P = < 005, versus cefoxitin and P = < 0- versus the combination). Clindamycin, chloramphenicol and metronidazole still have the highest potential clinical efficacy as measured by in-vitro tests of these Gram-negative anaerobes, e.g. > 95% strains were inhibited. 3 9 Downloaded from at Pennsylvania State University on September 9, 06

4 4 R. N. Jones Synergy or drug interaction tests were analysed by comparison of the : MIC ratio of cefotaxime-desacetyl-cefotaxime with the MIC recorded for each drug alone. Among the 53 Bact.fragilis isolates the combination of cefotaxime-desacetyl-cefotaxime, 3-7% were synergistically killed (MBC endpoints only) and 43-9% of strains showed partial synergy. No evidence of antagonism was observed and -6% of tests were indeterminate because the MIC for one or both of the drugs was above the tested concentration range. A slightly higher rate of synergy was found for the six other Bacteroiaes spp. The overall synergy rate (complete plus partial) was 870% primarily due to a marked increase in the partial synergy category. A comparable number of indeterminate tests, 3-3% was observed. The incidence of synergistic interactions for the 64 evaluable tests was 79-7%. Discussion Cefotaxime and its desacetyl metabolite have been shown to be synergistic in killing facultative bacteria (Jones et al., 98; Chin & Neu, 984) and strict anaerobes (Aldridge et al., 984; Jones, Packer & Barry, 984). This favourable interaction reduces the cefotaxime MICs in the presence of desacetyl-cefotaxime to a level most comparable to cefoxitin and other 7-methoxycephalosporins, with the MIC to 6 mg/ against Bad. fragilis. In this study equal concentrations of cefotaxime and the desacetyl-metabolite were used because of the near equal quantities found in cerebrospinal fluid, bile, urine and peritoneal fluids (Novick, 98; Okura et al., Abstract 87, nd ICAAC, Miami, 98). The rate or incidence ofsynergy in the study of 79-7% was nearly identical to that found earlier (800%), using a microdilution checkerboard technique and also testing Gram-positive strict anaerobes (Jones et al., 984). However, the incidence among Bad. fragilis strains was 6% lower in this report. The synergy contributed significantly to the reduction of the cephalosporins' MICs against some Bacteroides strains, particularly those with cefotaxime MICs < 3 mg/. The synergy increased the cefotaxime in-vitro efficacy (MICs ^ 3 mg/) from 48-% to 77- % inhibition of strains. The high incidence of partial synergy results, with fractional inhibitory concentration indices of > 0-50, but < 0-75, may have clinical relevance (Hall, Middleton & Westmacott, 983). The mechanism of the interaction may be related to the increased /ff-lactamase stability of the desacetyl form against Bacteroides enzymes (Limbert, Seibert & Schrinner, 98; Chin & Neu, 984). In addition desacetylated cephalosporins seem to have longer serum half-lives and penetrate more easily into animal tissues and into bacterial cells. Cefotaxime and desacetyl-cefotaxime could also compete favourably and co-operatively for the PDP a, b and 3 lethal target sites. Lastly, major reduction in /S-lactamase affinity is produced by desacetylation of cephalosporins at the 3'-position (Jones & Wilson, 98). This change may allow the drugs to interact in a co-operative manner in the presence of some /?-lactamase-producing strains. Although the actual mechanisms remains obscure, the fact also remains that cefotaxime and desacetyl-cefotaxime have synergistic antimicrobial activity against approximately 80% of anaerobic organisms. This action expands the cefotaxime spectrum to include > 75% of Bacteroides strains, a feature not appreciated on the basis of in-vitro testing of the parent compound alone (Jones & Thornsberry, 98). Downloaded from at Pennsylvania State University on September 9, 06 References Aldridge, K.. E., Sanders, C. V. & Marier, R. L. (984). In vitro synergy and potentiation between cefotaxime and desacetylcefotaxime against clinical isolates of Bacteroides. Diagnostic Microbiology of Infectious Disease (Suppl.), (in press).

5 Cefotaxime and desacetyl-cefotaxime against Bacteroides 43 Cabana, B. E., Van Harken, D. R. & Hottendorf, G. H. (976). Comparative pharmacokinetics and metabolism of cephapirin in laboratory animals and humans. Antimicrobial Agents Chemotherapy 0, Chamberlain, J., Coombes, J. D., Dell, D., Fromson, J. M., Ings, R. J., Macdonald, C. M. & McEwen, J. (980). Metabolism of cefotaxime in animals and man. Journal of Antimicrobial Chemotherapy 6 (Suppl. A), Chin, N.-X. & Neu, H. C. (984). Cefotaxime and desacetylcefotaxime: an example of advantageous antimicrobial metabolism. Diagnostic Microbiology of Infectious Disease (Suppl.), (in press). Hall, M. J., Middleton, R. F. & Westmacott, D. (983). The fractional inhibitory concentration (FIC) index as a measure of synergy. Journal Antimicrobial Chemotherapy, Jones, R. N., Barry, A. L. & Thornsberry, C. (98). Antimicrobial activity of desacetylcefotaxime alone and in combination with cefotaxime: evidence of synergy. Review of Infectious Disease 4, S Jones, R. N., Fuchs, P. C, Thornsberry, C. & Rhodes, N. (978). Antimicrobial susceptibility test for anaerobic bacteria. Comparison of Wilkins-Chalgren agar reference method and a microdilution method, and determination of stability of antimicrobics frozen in broth. Current Microbiology, 8-3. Jones, R. N. & Packer, R. R. (984). Cefotaxime, cephalothin and cephapirin: antimicrobial activity and synergy studies of cephalosporins with significant in vivo desacetyl-metabolite concentrations. Diagnostic Microbiology of Infectious Disease, Jones, R. N., Packer, R. R. & Barry, A. L. (984). The activity of cefotaxime and desacetylcefotaxime alone and in combination against anaerobes and staphylococci. Diagnostic Microbiology of Infectious Disease (Suppl.), (in press). Jones, R. N. & Thornsberry, C. (98). Cefotaxime: a review of in vitro antimicrobial properties and spectrum of activity. Review of Infectious Disease 4, S3OO-5. Jones, R. N. & Wilson, H. W. (98). Comparative /?-lactamase hydrolysis of and inhibition by 7-aminothiazolyl alpha-methoxyiminocephalosporins. Infection 0, Limbert, M., Seibert. G. & Schrinner, E. (98). Cooperation of cefotaxime and desacetylcefotaxime. Infection 0, Nakai, Y., Kanai, Y., Fugono, T. & Tanayama, S. (976). Metabolic fate of cephacetrile after parenteral administration in rats and rabbits. Journal of Antibiotics 6, Novick, W. J. (98). Levels of cefotaxime in bodyfluidsand tissues: a review. Review of Infectious Disease 4, S Pearson, R. D., Steigbigel, R. T., Davis, H. T. & Chapman, S. W. (980). Method for reliable determination of minimal lethal antibiotic concentrations. Antimicrobial Agents and Chemotherapy 8, Wick, W. E. (966). In vitro and in vivo laboratory comparison of cephalothin and desacetylcephalothin. Antimocrobial Agents and Chemotherapy, Downloaded from at Pennsylvania State University on September 9, 06