title: Mutations in β-Lactamase AmpC Increase Resistance of Pseudomonas aeruginosa Isolates to Antipseudomonal Cephalosporins.
abstract: Mutation-dependent overproduction of intrinsic β-lactamase AmpC is considered the main cause of resistance of clinical strains of Pseudomonas aeruginosa to antipseudomonal penicillins and cephalosporins. Analysis of 31 AmpC-overproducing clinical isolates exhibiting a greater resistance to ceftazidime than to piperacillin-tazobactam revealed the presence of 17 mutations in the β-lactamase, combined with various polymorphic amino acid substitutions. When overexpressed in AmpC-deficient P. aeruginosa 4098, the genes coding for 20/23 of these AmpC variants were found to confer a higher (2-fold to >64-fold) resistance to ceftazidime and ceftolozane-tazobactam than did the gene from reference strain PAO1. The mutations had variable effects on the MICs of ticarcillin, piperacillin-tazobactam, aztreonam, and cefepime. Depending on their location in the AmpC structure and their impact on β-lactam MICs, they could be assigned to 4 distinct groups. Most of the mutations affecting the omega loop, the R2 domain, and the C-terminal end of the protein were shared with extended-spectrum AmpCs (ESACs) from other Gram-negative species. Interestingly, two new mutations (F121L and P154L) were predicted to enlarge the substrate binding pocket by disrupting the stacking between residues F121 and P154. We also found that the reported ESACs emerged locally in a variety of clones, some of which are epidemic and did not require hypermutability. Taken together, our results show that P. aeruginosa is able to adapt to efficacious β-lactams, including the newer cephalosporin ceftolozane, through a variety of mutations affecting its intrinsic β-lactamase, AmpC. Data suggest that the rates of ESAC-producing mutants are ≥1.5% in the clinical setting.
authors.affiliation: French National Reference Center for Antibiotic Resistance, University Hospital of Besançon, Besançon, France.
authors.forename: M
authors.initials: M
authors.lastname: Berrazeg
authors.affiliation: French National Reference Center for Antibiotic Resistance, University Hospital of Besançon, Besançon, France.
authors.forename: K
authors.initials: K
authors.lastname: Jeannot
authors.affiliation: Laboratoire de Cristallographie and RMN Biologiques, CNRS UMR8015 and Université Paris Descartes, Paris, France.
authors.forename: Véronique Yvette
authors.initials: VY
authors.lastname: Ntsogo Enguéné
authors.affiliation: Laboratoire de Cristallographie and RMN Biologiques, CNRS UMR8015 and Université Paris Descartes, Paris, France.
authors.forename: I
authors.initials: I
authors.lastname: Broutin
authors.affiliation: French National Reference Center for Antibiotic Resistance, University Hospital of Besançon, Besançon, France.
authors.forename: S
authors.initials: S
authors.lastname: Loeffert
authors.affiliation: French National Reference Center for Antibiotic Resistance, University Hospital of Besançon, Besançon, France.
authors.forename: D
authors.initials: D
authors.lastname: Fournier
authors.affiliation: French National Reference Center for Antibiotic Resistance, University Hospital of Besançon, Besançon, France patrick.plesiat@univ-fcomte.fr.
authors.forename: P
authors.initials: P
authors.lastname: Plésiat
mesh_terms: D000222:Adaptation, Physiological / Q000187:drug effects / Q000235:genetics*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001398:Aztreonam / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D000077723:Cefepime; D002442:Ceftazidime / Q000494:pharmacology; D002511:Cephalosporins / Q000494:pharmacology; D015870:Gene Expression; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009154:Mutation; D010397:Penicillanic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D010878:Piperacillin / Q000494:pharmacology; D000077725:Piperacillin, Tazobactam Drug Combination; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D000078142:Tazobactam; D013982:Ticarcillin / Q000494:pharmacology; D018440:beta-Lactam Resistance / Q000235:genetics*; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D002511:Cephalosporins; C000594038:ceftolozane, tazobactam drug combination; D000077725:Piperacillin, Tazobactam Drug Combination; D000077723:Cefepime; D010397:Penicillanic Acid; D002442:Ceftazidime; C048795:AmpC beta-lactamases; D001618:beta-Lactamases; D013982:Ticarcillin; D001398:Aztreonam; D000078142:Tazobactam; D010878:Piperacillin