TY - JOUR

T1 - Development of antibiotic resistance and up-regulation of the antimutator gene pfpI in mutator Pseudomonas aeruginosa due to inactivation of two DNA oxidative repair genes (mutY, mutM)

AU - Mandsberg, Lotte Frigaard

AU - Maciá, Maria D

AU - Bergmann, Kirsten R

AU - Christiansen, Lasse

AU - Alhede, Morten

AU - Kirkby, Nikolai

AU - Høiby, Niels

AU - Oliver, Antonio

AU - Ciofu, Oana

N1 - © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

PY - 2011

Y1 - 2011

N2 - Prevention and correction of oxidative DNA lesions in Pseudomonas aeruginosa is ensured by the DNA oxidative repair system (GO). Single inactivation of mutT, mutY and mutM involved in GO led to elevated mutation rates (MRs) that correlated to increased development of resistance to antibiotics. In this study, we constructed a double mutant in mutY and mutM (PAOMY-Mgm) and characterized the phenotype and the gene expression profile using microarray and RT-PCR. PAOMY-Mgm presented 28-fold increases in MR compared with wild-type reference strain PAO1. In comparison, the PAOMYgm (mutY) single mutant showed only a fivefold increase, whereas the single mutant PAOMMgm (mutM) showed a nonsignificant increase in MR compared with PAO1 and the single mutants. Mutations in the regulator nfxB leading to hyperexpression of MexCD-OprJ efflux pump were found as the mechanism of resistance to ciprofloxacin in the double mutant. A better fitness of the mutator compared with PAO1 was found in growth competition experiments in the presence of ciprofloxacin at concentrations just below minimal inhibitory concentration. Up-regulation of the antimutator gene pfpI, that has been shown to provide protection to oxidative stress, was found in PAOMY-Mgm compared with PAO1. In conclusion, we showed that MutY and MutM are cooperating in the GO of P. aeruginosa, and that oxidative DNA lesions might represent an oxidative stress for the bacteria.

AB - Prevention and correction of oxidative DNA lesions in Pseudomonas aeruginosa is ensured by the DNA oxidative repair system (GO). Single inactivation of mutT, mutY and mutM involved in GO led to elevated mutation rates (MRs) that correlated to increased development of resistance to antibiotics. In this study, we constructed a double mutant in mutY and mutM (PAOMY-Mgm) and characterized the phenotype and the gene expression profile using microarray and RT-PCR. PAOMY-Mgm presented 28-fold increases in MR compared with wild-type reference strain PAO1. In comparison, the PAOMYgm (mutY) single mutant showed only a fivefold increase, whereas the single mutant PAOMMgm (mutM) showed a nonsignificant increase in MR compared with PAO1 and the single mutants. Mutations in the regulator nfxB leading to hyperexpression of MexCD-OprJ efflux pump were found as the mechanism of resistance to ciprofloxacin in the double mutant. A better fitness of the mutator compared with PAO1 was found in growth competition experiments in the presence of ciprofloxacin at concentrations just below minimal inhibitory concentration. Up-regulation of the antimutator gene pfpI, that has been shown to provide protection to oxidative stress, was found in PAOMY-Mgm compared with PAO1. In conclusion, we showed that MutY and MutM are cooperating in the GO of P. aeruginosa, and that oxidative DNA lesions might represent an oxidative stress for the bacteria.

KW - Anti-Bacterial Agents

KW - Bacterial Proteins

KW - DNA Glycosylases

KW - DNA-Formamidopyrimidine Glycosylase

KW - Drug Resistance, Bacterial

KW - Gene Expression Profiling

KW - Gene Expression Regulation, Bacterial

KW - Gene Knockout Techniques

KW - Microarray Analysis

KW - Mutation

KW - Pseudomonas aeruginosa

KW - Reverse Transcriptase Polymerase Chain Reaction

KW - Rifampin

U2 - 10.1111/j.1574-6968.2011.02383.x

DO - 10.1111/j.1574-6968.2011.02383.x

M3 - Journal article

C2 - 22092761

VL - 324

SP - 28

EP - 37

JO - F E M S Microbiology Letters

JF - F E M S Microbiology Letters

SN - 0378-1097

IS - 1

ER -