title: Cephem Potentiation by Inactivation of Nonessential Genes Involved in Cell Wall Biogenesis of β-Lactamase-Producing Escherichia coli.
abstract: Reversal of antimicrobial resistance is an appealing and largely unexplored strategy in drug discovery. The objective of this study was to identify potential targets for "helper" drugs reversing cephem resistance inEscherichia colistrains producing β-lactamases. A CMY-2-encoding plasmid was transferred by conjugation to seven isogenic deletion mutants exhibiting cephem hypersusceptibility. The effect of each mutation was evaluated by comparing the MICs in the wild type and the mutant harboring the same plasmid. Mutation of two genes encoding proteins involved in cell wall biosynthesis,dapFandmrcB, restored susceptibility to cefoxitin (FOX) and reduced the MICs of cefotaxime and ceftazidime, respectively, from the resistant to the intermediate category according to clinical breakpoints. The same mutants harboring a CTX-M-1-encoding plasmid fell into the intermediate or susceptible category for all three drugs. Individual deletion ofdapFandmrcBin a clinical isolate of CTX-M-15-producingE. colisequence type 131 (ST131) resulted in partial reversal of ceftazidime and cefepime resistance but did not reduce MICs below susceptibility breakpoints. Growth curve analysis indicated no fitness cost in a ΔmrcBmutant, whereas a ΔdapFmutant had a 3-fold longer lag phase than the wild type, suggesting that drugs targeting DapF may display antimicrobial activity, in addition to synergizing with selected cephems. DapF appeared to be a potential FOX helper drug target candidate, sincedapFinactivation resulted in synergistic potentiation of FOX in the genetic backgrounds tested. The study showed that individual inactivation of two nonessential genes involved in cell wall biogenesis potentiates cephem activity according to drug- and strain-specific patterns.
authors.affiliation: Department of Veterinary Disease Biology, University of Copenhagen, Frederiksberg, Denmark KrBaker@rossvet.edu.kn BJana@rossvet.edu.kn LGuardabassi@rossvet.edu.kn.|Department of Biomedical Sciences, Ross University School of Veterinary Medicine, Basseterre, St. Kitts and Nevis.
authors.forename: Kristin R
authors.initials: KR
authors.lastname: Baker
authors.affiliation: Department of Veterinary Disease Biology, University of Copenhagen, Frederiksberg, Denmark.
authors.forename: Helga Høeg
authors.initials: HH
authors.lastname: Sigurðardóttir
authors.affiliation: Department of Veterinary Disease Biology, University of Copenhagen, Frederiksberg, Denmark KrBaker@rossvet.edu.kn BJana@rossvet.edu.kn LGuardabassi@rossvet.edu.kn.|Department of Biomedical Sciences, Ross University School of Veterinary Medicine, Basseterre, St. Kitts and Nevis.
authors.forename: Bimal
authors.initials: B
authors.lastname: Jana
authors.affiliation: Department of Veterinary Disease Biology, University of Copenhagen, Frederiksberg, Denmark KrBaker@rossvet.edu.kn BJana@rossvet.edu.kn LGuardabassi@rossvet.edu.kn.|Department of Biomedical Sciences, Ross University School of Veterinary Medicine, Basseterre, St. Kitts and Nevis.
authors.forename: Luca
authors.initials: L
authors.lastname: Guardabassi
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002473:Cell Wall / Q000187:drug effects / Q000378:metabolism; D002511:Cephalosporins / Q000494:pharmacology*; D003227:Conjugation, Genetic; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D020868:Gene Silencing; D008826:Microbial Sensitivity Tests; D046915:Penicillin-Binding Proteins / Q000172:deficiency / Q000235:genetics; D046928:Peptidoglycan Glycosyltransferase / Q000172:deficiency / Q000235:genetics; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism; D046929:Serine-Type D-Ala-D-Ala Carboxypeptidase / Q000172:deficiency / Q000235:genetics; D018440:beta-Lactam Resistance / Q000187:drug effects* / Q000235:genetics; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D002511:Cephalosporins; D029968:Escherichia coli Proteins; D046915:Penicillin-Binding Proteins; D046928:Peptidoglycan Glycosyltransferase; C478681:penicillin-binding protein 1B, E coli; D046929:Serine-Type D-Ala-D-Ala Carboxypeptidase; C473109:beta-lactamase CTX-M-15; C055184:beta-lactamase TEM-3; D001618:beta-Lactamases; C000605810:beta-lactamase CMY-2, E coli
keywords: cephem resistance; extended-spectrum β-lactamase (ESBL); helper drug