1: J Antimicrob Chemother. 2007 Mar;59(3):378-86. Epub 2007 Feb 8.

A combined phenotypic and genotypic method for the detection of Mex efflux pumps
in Pseudomonas aeruginosa.

Mesaros N, Glupczynski Y, Avrain L, Caceres NE, Tulkens PM, Van Bambeke F.

Unite de Pharmacologie cellulaire et moleculaire, Brussels, Universite
catholique de Louvain, UCL 7370 avenue E. Mounier 73, B-1200 Bruxelles, Belgium.

Background Mex efflux pumps contribute to multidrug resistance in Pseudomonas
aeruginosa. Evidencing their expression in clinical isolates would help in
rationalizing antibiotic selection. Methods We have developed a combined
phenotypic and genotypic approach for the differential diagnosis of resistance
mediated by four major transporters (MexAB-OprM, MexCD-OprJ, MexEF-OprN,
MexXY-OprM). The methodology was validated using reference strains harbouring
only one specific transporter and its applicability evaluated towards seven
selected clinical isolates, the resistance mechanisms of which could not be
assigned by conventional techniques. Phenotypic detection used MIC measurements
with reporter antibiotics [carbenicillin (MexAB-OprM); erythromycin
(MexCD-OprJ); norfloxacin and imipenem (MexEF-OprN); gentamicin (MexXY-OprM)]
with and without Phe-Arg-beta-naphthylamide. Genotypic detection was made by
semi-quantitative reverse transcription PCR (RT-PCR) for mexC and mexE, and by
quantitative competitive RT-PCR and real-time PCR for mexA and mexX (correlation
between both methods: > 88 % ; overexpression levels ranging between 4.8 and
8.1). Results Convergence between phenotypic and genotypic methods was observed
in control strains for all pumps. For clinical isolates, convergence was
obtained in 6 of 7 strains for MexXY-OprM and MexEF-OprM, and in 5 of 7 for
MexAB-OprM and MexCD-OprJ, mostly due to hard to interpret phenotypic data.
Conclusions The data plead for combining phenotypic and genotypic approaches in
the diagnosis of efflux-mediated resistance in P. aeruginosa.

PMID: 17289770 [PubMed - in process]

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