title: Molecular characterizations of carbapenem and ciprofloxacin resistance in clinical isolates of Pseudomonas putida.
abstract: To analyze the genetic mechanisms of carbapenem and ciprofloxacin resistance in clinical isolates of Pseudomonas putida, 27 clinical isolates (comprising 11 carbapenem- and ciprofloxacin-resistant strains, 13 carbapenem-resistant and ciprofloxacin-susceptible strains, and 3 carbapenem- and ciprofloxacin-susceptible strains) were collected from different patients. Carbapenem resistance was examined by polymerase chain reaction (PCR) and DNA sequencing for metallo-beta-lactamase (MBL) and integrase genes (IntI-1 and IntI-3), and by reverse transcriptase-PCR (RT-PCR) for expression of the porin gene (oprD). Ciprofloxacin resistance was characterized by PCR and DNA sequencing for mutations in the quinoloneresistance determining regions of the gyrA and parC genes. The blaIMP-1 MBL and intI-1 and/or intI-3 genes were detected in all carbapenem-resistant strains, and decreased expression of the oprD gene as compared to carbapenemsusceptible strains was observed in several strains. All the 11 strains with ciprofloxacin minimal inhibitory concentrations (MICs) of > or =64 mg/l had substitution in GyrA (Thr83Ile), and one (ciprofloxacin MIC of 512 mg/l) of these strains also had substitution in ParC (Ser87Leu). Overproduction of the efflux pump was observed in 10 of the 11 ciprofloxacin-resistant strains. We concluded that the production of IMP-1 type MBL was the most critical factor in developing high-level resistance to carbapenems, and mutations in the target proteins and overproduction of the efflux pump synergistically contribute to the acquisition of high-level resistance to ciprofloxacin in clinical isolates of P. putida.
authors.affiliation: Department of Microbiology and Immunology, Graduate School of Health Sciences, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan. wakako@topaz.plala.or.jp
authors.forename: Wakako
authors.initials: W
authors.lastname: Kumita
authors.forename: Ryoichi
authors.initials: R
authors.lastname: Saito
authors.forename: Kenya
authors.initials: K
authors.lastname: Sato
authors.forename: Takashi
authors.initials: T
authors.lastname: Ode
authors.forename: Kyoji
authors.initials: K
authors.lastname: Moriya
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Koike
authors.forename: Toshio
authors.initials: T
authors.lastname: Chida
authors.forename: Noboru
authors.initials: N
authors.lastname: Okamura
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics; D015780:Carbapenems / Q000494:pharmacology*; D002939:Ciprofloxacin / Q000494:pharmacology*; D027101:DNA Topoisomerase IV / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D016521:Electrophoresis, Gel, Pulsed-Field; D006801:Humans; D019426:Integrases / Q000235:genetics; D008826:Microbial Sensitivity Tests; D016133:Polymerase Chain Reaction; D018272:Porins / Q000235:genetics; D016958:Pseudomonas putida / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics*; D018440:beta-Lactam Resistance / Q000235:genetics; D001618:beta-Lactamases / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D015780:Carbapenems; D018272:Porins; D002939:Ciprofloxacin; D019426:Integrases; D001618:beta-Lactamases; D027101:DNA Topoisomerase IV