MUTATIONS IN TOPOISOMERASE-IV AND DNA GYRASE OF STAPHYLOCOCCUS-AUREUS- NOVEL PLEIOTROPIC EFFECTS AN QUINOLONE AND COUMARIN ACTIVITY

Previous studies have shown that topoisomerase IV and DNA gyrase inter act with quinolones and coumarins in different ways. The MICs of couma rins (novobiocin and coumermycin) for MT5, a Staphylococcus aureus nov mutant, are higher than those for wild-type strains. Sequencing the g yrB gene encoding one subunit of the DNA gyrase revealed the presence of a double mutation likely to be responsible for this resistance: at codon 102 (Ile to Ser) and at codon 144 (Arg to Ile). For single-step flqA mutant MT5224c9, previously selected on ciprofloxacin, the fluoro quinolone MTC was higher and the coumarin MIC was lower than those for its parent, MT5. Sequencing the grlB and grlA genes of topoisomerase IV of MT5224c9 showed a single Asn-470-to-Asp mutation in GrlB. Geneti c outcrosses by. transformation with chromosomal DNA and introduction of plasmids carrying either the wild-type or the mutated grlB gene ind icated that this mutation causes both increased MICs of fluoroquinolon es and decreased MICs of coumarins and that the mutant grlB allele is codominant for both phenotypes with multicopy alleles. Integration of these plasmids into the chromosome confirmed the codominance of fluoro quinolone resistance, but grlB(+) appeared dominant over grlB (Asp-470 ) for coumarin resistance. Finally, the gyrA (Leu-84) mutation previou sly described as silent for fluoroquinolone resistance increased the M IC of nalidixic acid, a nonfluorinated quinolone. Combining the grlA ( Phe-80) and grlB (Asp-47O) mutations with this gyrA mutation also had differing effects. The findings indicate that alterations in topoisome rases may have pleiotropic effects on different classes of inhibitors as well as on inhibitors within the same class. A full understanding o f drug action and resistance at the molecular level must take into acc ount both inhibitor structure-activity relationships and the effects o f different classes of topoisomerase mutants.