title: Increased expression levels of chromosomal AmpC β-lactamase in clinical Escherichia coli isolates and their effect on susceptibility to extended-spectrum cephalosporins.
abstract: Forty-nine clinical Escherichia coli isolates, both extended-spectrum β-lactamase (ESBL) negative and ESBL positive, were studied to investigate whether increased AmpC expression is a mechanism involved in cefoxitin resistance and if this influences the third-generation cephalosporin activity. Nine of 33 (27.2%) cefoxitin-resistant (minimum inhibitory concentration [MIC] >8 mg/L) isolates showed hyperproduction of chromosomal AmpC (c-AmpC) based on (1) at least two positive tests using AmpC inhibitors, (2) mutations in the promoter/attenuator regions, and (3) a 6.1- to 163-fold increase in c-ampC expression by quantitative reverse transcription-polymerase chain reaction. In ESBL-negative isolates, MICs of ceftazidime and cefotaxime were mostly above the wild-type (WT) level, but below the S/I breakpoint (EUCAST guideline), except for one isolate with MICs of 4 mg/L. No plasmid-mediated AmpCs were found. Periplasmic extracts of nine c-AmpC hyperproducers were preincubated with or without cefuroxime or ceftazidime and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cefuroxime and ceftazidime were stable to hydrolysis but acted as inhibitors of the enzyme. None of these isolates showed loss of porins. Thus, cefoxitin resistance has low specificity for detecting upregulated c-AmpC production. c-AmpC hyperproducing E. coli is mostly still susceptible to third-generation cephalosporins but less than WT E. coli. Surveillance of cefoxitin-resistant E. coli to monitor developments in the activity of third-generation cephalosporins against c-AmpC hyperproducers is warranted.
authors.affiliation: 1 Department of Medical Microbiology, Leiden University Medical Center , Leiden, The Netherlands .
authors.forename: Sunita
authors.initials: S
authors.lastname: Paltansing
authors.forename: Margriet
authors.initials: M
authors.lastname: Kraakman
authors.forename: Ria
authors.initials: R
authors.lastname: van Boxtel
authors.forename: Ivo
authors.initials: I
authors.lastname: Kors
authors.forename: Els
authors.initials: E
authors.lastname: Wessels
authors.forename: Wil
authors.initials: W
authors.lastname: Goessens
authors.forename: Jan
authors.initials: J
authors.lastname: Tommassen
authors.forename: Alexandra
authors.initials: A
authors.lastname: Bernards
mesh_terms: D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology*; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D002511:Cephalosporins / Q000378:metabolism / Q000494:pharmacology*; D002876:Chromosomes, Bacterial / Q000235:genetics; D024901:Drug Resistance, Multiple, Bacterial; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D004927:Escherichia coli Infections / Q000382:microbiology*; D029968:Escherichia coli Proteins / Q000096:biosynthesis* / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006801:Humans; D008826:Microbial Sensitivity Tests; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity; D001618:beta-Lactamases / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002511:Cephalosporins; D029968:Escherichia coli Proteins; C048795:AmpC beta-lactamases; D001618:beta-Lactamases