Study on gyrA and grlA genes in stepwise-selected enrofloxcin-resistant mutants of Staphylococcus aureus

Fluoroquinolone-resistant mutants (MIC: 4-≥128#mu#g/ml) were obtained in vitro from Staphylococcus aureus 28001 by stepwise selection on increasing concentrations of enrofloxacin. The chromosomal DNAs of three (MIC: 4, 16 and ≥128#mu#g/ml) of these strains were obtained, PCR was employed to amplify the gyrA and grlA genes; the PCR product were aligned to pMD18-T vector; transformation in E. coli JM109 competent cell were performed; positive clones were chosen and sequenced. The sequence analysis showed that grlA Ser80 (TCC)→Phe (TGC) or grlA, Ser80 (TCC)→Tyr (TAC) point mutation was involved in low resistant mutation strains (MIC≤16#mu#g/ml); grlA Ser80 (TCC)→Phe (TGC) and gyrA Ser84 (TCA)→Leu (TTA) multiple point mutation were involved in high resistant strains (MIC≥128#mu#g/ml). All three resistant mutation have grlA Ser80→Tyr (Phe) point mutation. Results suggested that the primary target enzyme to Staphylococcus aureus of enrofloxacin was topoisomerase IV.