Analysis of antibiotic resistance gene expression in Pseudomonas aeruginosa by quantitative real-time-PCR.
In Pseudomonas aeruginosa many of the clinically relevant resistance mechanisms result from changes in gene expression as exemplified by the Mex drug efflux pumps, the AmpC beta-lactamase and the carbapenem-specific porin OprD. We used quantitative real-time-PCR to analyze the expression of these genes in susceptible and antibiotic-resistant laboratory and clinical strains. In nalB mutants, which overexpress OprM, we observed a four- to eightfold increase in the expression of mexA, mexB, and oprM genes. MexX and mexY genes were induced eight to 12 times in the presence of 2 mg L(-1) tetracycline. The mexC/oprJ and mexE/oprN gene expression levels were increased 30- to 250-fold and 100- to 760-fold in nfxB and nfxC mutants, respectively. We further found that in defined laboratory strains expression levels of ampC and oprD genes paralleled beta-lactamase activity and OprD protein levels, respectively. Our data support the use of quantitative real-time-PCR chain reaction for the analysis of the antimicrobial resistance gene expression in P. aeruginosa.
Dumas
Jean-Luc
JL
Department of Microbiology and Molecular Medicine, University of Geneva, Geneva, Switzerland.
van Delden
Christian
C
Perron
Karl
K
Köhler
Thilo
T
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D018272:Porins / Q000235:genetics / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D018272:Porins; C077157:OprD protein, Pseudomonas aeruginosa; C048795:AmpC beta-lactamases; D001618:beta-Lactamases