title: Characterization of a beta-lactamase from Clostridium clostridioforme.
abstract: A beta-lactamase-producing strain of Clostridium clostridioforme isolated from human peritoneal fluid was examined by MIC testing and enzyme characterization. MICs of penicillins (64-512 mg/L) were higher than those of cephalosporins (8-128 mg/L); the strain was susceptible to cefoxitin (8 mg/L) and imipenem (1 mg/L). No enhancement of cephalosporin activity occurred when clavulanate was also added, but a limited degree of enhancement of penicillin activity (resulting in beta-lactam MICs higher than available NCCLS breakpoints) occurred when clavulanate, sulbactam or tazobactam was added simultaneously. By contrast, addition of BRL 42715 with amoxycillin, ticarcillin or piperacillin led to a drop in beta-lactam MICs from 512 to < or = 1 mg/L, with a drop from 64 to 1 mg/L when BRL 42715 was added with cefotaxime. All inhibitors were added at fixed concentrations of 2 mg/L. As determined spectrophotometrically, the enzyme hydrolysed penicillin G, cloxacillin and piperacillin (Vmax values (%) 372, 1816, 1001, respectively relative to cephaloridine) more efficiently than cephalosporins (69-191, with cephaloridine as 100%). Km values (microM) varied between 30-308 microM (penicillins) and 2-20 microM (cephalosporins). Relative enzyme efficiency (relative Vmax/Km with cephaloridine as 100) varied from 21-100 (cephalosporins) and 8-77 (penicillins). IC50 values (microM) with nitrocefin, piperacillin and penicillin G substrates (concentrations 20, 100 and 20 microM, respectively) were > 1000, 7, 3.5 (clavulanate); > 1000, 300, 59 (sulbactam), > 1000, 29, 7.7 (tazobactam); 0.0004, 0.001, 0.0018 (BRL 42715). The enzyme was not inhibited by EDTA, cefoxitin, cloxacillin or aztreonam, but was inhibited by pCMB.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Hershey Medical Center, PA 17033.
authors.forename: P C
authors.initials: PC
authors.lastname: Appelbaum
authors.forename: S K
authors.initials: SK
authors.lastname: Spangler
authors.forename: G A
authors.initials: GA
authors.lastname: Pankuch
authors.forename: A
authors.initials: A
authors.lastname: Philippon
authors.forename: M R
authors.initials: MR
authors.lastname: Jacobs
authors.forename: R
authors.initials: R
authors.lastname: Shiman
authors.forename: E J
authors.initials: EJ
authors.lastname: Goldstein
authors.forename: D M
authors.initials: DM
authors.lastname: Citron
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001202:Ascitic Fluid / Q000382:microbiology; D001426:Bacterial Proteins / Q000378:metabolism; D003013:Clostridium / Q000187:drug effects / Q000201:enzymology*; D003015:Clostridium Infections / Q000382:microbiology; D004352:Drug Resistance, Microbial; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D013056:Spectrophotometry, Ultraviolet; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000032:analysis* / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases