Rapid detection of quinolone resistance-associated gyrA mutations in Neisseria gonorrhoeae with a LightCycler

Abstract

 A fluorometric real-time polymerase chain-reaction (PCR)-hybridization system, the LightCycler was developed for the detection of Neisseria gonorrhoeae in clinical samples and the analysis of point mutations associated with quinolone resistance in the gyrA gene. This system allowed us to amplify the N. gonorrhoeae-specific gyrA gene from an amount of DNA corresponding to five genome copies per reaction. We were able to detect N. gonorrhoeae in either 55 control strains or 36 nonisolated clinical urethral swab specimens, and to analyze the presence of mutations in codons Ser-91 and Asp-95 of the gyrA gene within 1 h. The mutation status in the gyrA gene assessed by the LightCycler assay was completely in agreement with the results of our previous conventional sequencing analysis, and was associated with the susceptibility to ciprofloxacin.