Detection of mexA and mexX efflux genes in P. aeruginosa: correlation between QC-RT-PCR and real-time PCR
Abstract number: p1751
Mesaros N., Glupczynski Y., Pierard D., Dediste A., Van Laethem Y., Tulkens P., Mingeot-Leclercq M., Van Bambeke F.
Efflux systems are rarely identified as such in clinical microbiology laboratories. Yet, over expression of transporters such as MexAB-OprM and MexXY-OprM are likely to cause antibiotic multi- and cross-resistance in Pseudomonas aeruginosa, leading to potential clinical treatment failures because of their inducible character. We have previously developed and validated with reference strains a QC-RT-PCR method to quantify mexA and mexX expression levels (ECCMID 2005, P1731). In the present study, we have developed a Real-Time-PCR assay and present here the correlation between both methods using control strains and clinical isolates.
Expression levels of mexA and mexX were measured by both techniques in (i) 4 reference strains expressing only one of these efflux mechanisms [mexA (2) or mexX (2)]; and (ii) 8 clinical isolates, in comparison with the wild-type strain PAO1 (basal mexA and mexX expression levels).
Real-Time PCR showed an inter-day reproducibility of 95 ± 5.3% (triplicates of 10 strains). Among the clinical strains, 5 over expressed mexA and 3 mexX. The Table shows (i) the mean level of overexpression of mexA and mexX in comparison with the wild type strain PAO1 (set at 1), as detected by Real-time PCR for all strains; (ii) the ratio of these values to those observed by QC-RT-PCR for the corresponding transporters.
Both QC-RT-PCR and Real-Time-PCR are potentially useful in clinical laboratories as sensitive and rapid diagnostic tools to quantify the expression level of mexA and mexX in P. aeruginosa. Combined with phenotypic characterization, this approach may help in a better understanding of the resistance mechanisms and epidemiology of resistance in this difficult-to-treat nosocomial pathogen.
|Session name:||XXIst ISTH Congress|
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