[Real-time PCR for fast detection of plasmid-mediated qnr genes in extended spectrum beta-lactamase producing Enterobacteriaceae]

Abstract

Aim of the study: To develop a fast and reliable real time PCR technique for detecting plasmid-mediated quinolone resistance genes qnrA, qnrB and qnrS.

Methods: A real-time PCR assay using SYBR Green I and Roche LightCycler(®) was developed to detect qnr genes. Detection of qnr genes was based on comparison of melting temperature differences with a positive control of each qnr genes. This assay was performed to study 138 isolates collected from diagnostic and screening samples in the Champagne-Ardenne region in 2004 (France).

Results: In optimized conditions, the three positive controls tested alone and with isolates confirmed the specificity of the PCR primers. Each PCR assay was able to test 30 strains in 60min for 1 qnr gene. Out of 138 isolates screened, 3.6 % isolates were positive for a qnrA1, 1.5 % for qnrS1 and no qnrB-like gene. Prevalence of qnr determinants was 5 % and reached 9.5 % in clinical isolates.

Conclusion: Real-time PCR is a fast and reliable technique for screening of qnr-positive strains. This study shows a relatively high prevalence of qnr determinants (5 %) among ESBL-producing Enterobacteriaceae.