Real-Time PCR Assay for Detection of Fluoroquinolone Resistance Associated with grlA Mutations in Staphylococcus aureus

Example showing the real-time detection on a Smart Cycler of a fluoroquinolone-sensitive S. aureus strain (ATCC 25923) containing no mutation in the grlA gene (A) and a fluoroquinolone-resistant S. aureus strain (CCRI-12876) containing mutations at both codons 80 and 84 (B). For every sample tested, two separate PCRs, one containing probes S80-FAM and Sau-TET and one containing probes E84-FAM and Sau-TET, were performed. The left graphs show the fluorescence curves for the Sau-TET probe (hatched curve) and the S80-FAM probe (plain curve). The right graphs show the fluorescence curves for the Sau-TET probe (hatched curve) and the E84-FAM probe (plain curve). PCR assays were performed with crude DNA extracts prepared from bacterial suspensions whose turbidities were adjusted to that of a 0.5 McFarland standard.