Analysis of antibiotic resistance gene expression in Pseudomonas aeruginosa by quantitative real-time-PCR

• Published in: FEMS microbiology letters Vol. 254; no. 2; pp. 217 - 225
• Main Authors: Dumas, Jean-Luc, van Delden, Christian, Perron, Karl, Köhler, Thilo
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01-01-2006; Blackwell Science Ltd; Blackwell; Oxford University Press
• Subjects: Anti-Bacterial Agents - pharmacology; Antibiotic resistance; Antibiotics; Antimicrobial resistance; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteriology; beta-Lactamases - genetics; beta-Lactamases - metabolism; Biological and medical sciences; Drug Resistance, Bacterial - genetics; Efflux; Fundamental and applied biological sciences. Psychology; Gene expression; Gene Expression Regulation, Bacterial; Genes; Genetics; Humans; Laboratories; Microbiology; OprD protein; Pathogenicity, virulence, toxins, bacteriocins, pyrogens, host-bacteria relations, miscellaneous strains; Porins - genetics; Porins - metabolism; Pseudomonas aeruginosa; Pseudomonas aeruginosa - drug effects; Pseudomonas aeruginosa - genetics; Pseudomonas aeruginosa - metabolism; quantitative real‐time‐PCR (qRT‐PCR); Real time; Reverse Transcriptase Polymerase Chain Reaction - methods; antibiotic resistance; quantitative real-time-PCR (qRT-PCR); Pseudomonadales; Gene expression; Real time; Polymerase chain reaction; Resistance; Antibiotic; Pathogenic; Bacteria; Pseudomonadaceae; Pseudomonas aeruginosa; Quantitative analysis
• ISSN: 0378-1097; 1574-6968
• DOI: 10.1111/j.1574-6968.2005.00008.x

Abstract
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 β-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 β-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.