title: [Detection of plasmid-mediated AmpC beta-lactamase in clinical isolates of cefoxitin-resistant Escherichia coli and Klebsiella pneumoniae].
abstract: Extended spectrum beta-lactamases (ESBL) or AmpC beta-lactamases may be associated with false cephalosporin susceptibility results. Although there are well-established methods for screening and confirmation of class A (ESBL) and class B metallobeta-lactamases (MBL), there is no current guideline for the detection of AmpC beta-lactamases. The aim of this study was to evaluate the performances of three different phenotypic tests [boronic acid (BA) - klavulanic acid (CA) inhibition test, AmpC disk test (TRISEDTA impregnanted), modified 3-dimensional test (M3DT)] for the detection of AmpC beta-lactamases. A total of 50 (42 were Escherichia coli and eight were Klebsiella pneumoniae) cefoxitin-insusceptible strains were collected during June-September 2009. Multiplex PCR was used as the genotypic confirmation test by using DHA, CIT, MOX, FOX and EBC primers. Twenty-five (50%) of 50 cefoxitin-insusceptible strains yielded positive result by BA-CA test. Twenty two (88%) of 25 BA-CA positive strains yielded positive result by multiplex PCR and all of them belonged to CIT family (CMY-2 to CMY-7, LAT-1 to LAT-4 and BIL-1 type) of AmpC beta-lactamases. One strain harboured both CIT and EBC family of AmpC beta- lactamases, one strain harboured CIT + MOX + ESBL. DHA and FOX family of AmpC beta-lactamases were not detected in this study. Both AmpC disk test and M3DT yielded positive test with 11 (52.3%) of 21 AmpC enzyme producing strains (except for one of the PCR positive strain which couldn't be screened by M3DT and AmpC disk test). When BA was added to CA inhibition test, the number of positive isolates of ESBL increased from 13 to 14 (63.6%) due to inhibition of AmpC with BA. Hovewer, the strains which yielded negative results by 3-aminophenylboronic acid (3-APB) were tested again by benzeneboronic acid. Three strains were also found to be positive with benzeneboronic acid. The results of this study indicated that BA test with benzene boronic acid or 3-APB warranted further study. In conclusion, for the phenotypic detection of AmpC beta-lactamases, BA-CA test is simple to perform and easy to interpret. Briefly, in order to prevent the masking effect of ESBL, CA must be added to the BA inhibition test. Also in order to prevent the masking effect of AmpC beta-lactamases on ESBL detection, BA must be added to the CA inhibition test.
authors.affiliation: Ankara University Faculty of Pharmacy, Department of Pharmaceutical Microbiology, Ankara, Turkey. srpl50star@gmail.com
authors.forename: Serpil
authors.initials: S
authors.lastname: Coşkun
authors.forename: Nurten
authors.initials: N
authors.lastname: Altanlar
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification*; D001897:Boronic Acids / Q000494:pharmacology; D002440:Cefoxitin / Q000494:pharmacology*; D019818:Clavulanic Acid / Q000494:pharmacology; D024881:Drug Resistance, Bacterial / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D007711:Klebsiella pneumoniae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D008826:Microbial Sensitivity Tests / Q000379:methods; D010641:Phenotype; D001618:beta-Lactamases / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D001897:Boronic Acids; D004791:Enzyme Inhibitors; D019818:Clavulanic Acid; D002440:Cefoxitin; C048795:AmpC beta-lactamases; D001618:beta-Lactamases; C010686:benzeneboronic acid