GYRA MUTATIONS IN HIGH-LEVEL FLUOROQUINOLONE-RESISTANT CLINICAL ISOLATES OF ESCHERICHIA-COLI

Double mutations in the quinolone resistance determining region of the gyrase A gene (gyrA) have recently been reported to be associated wit h high-level resistance to fluoroquinolones in clinical isolates of Es cherichia coli. We examined the type and frequency of such mutations i n a large number of clinical isolates that were obtained from ten diff erent geographical locations and had been genotypically characterized by pulsed field gel electrophoresis (PFGE) of chromosomal DNA digests. Of 36 isolates with ofloxacin MICs greater than or equal to 4 mg/L th at represented at least 24 distinct genotypes, 35 had double mutations at amino acid codons 83 and 87 of gyrA, while two isolates with oflox acin MICs of 0.5 and 4 mg/L, respectively, each had a single mutation at codon 83. Mutations at codon Ser-83 were uniform, resulting in subs titution by Leu. The additional mutations at amino acid codon 87 in th e 35 double-mutants were diverse, resulting in Asp-87 substitutions by residues Asn (23 isolates), Gly (7 isolates), Tyr (4 isolates), or Hi s (1 isolate) without a discernable correlation with fluoroquinolone M ICs or with phenotypic resistance to chemically unrelated antibacteria l agents. Maximal differences between MICs of double-mutants with the same amino acid substitution were eight-fold. The changes of amino aci d residues at codon Asp-87 differed between individual patient isolate s with the same genotype (and similar MICs), suggesting that the amino acid codon 87 mutations (and possibly the development of high-level f luoroquinolone resistance) might have occurred after the transmission and sharing of a precursor strain carrying the Ser-83-->Leu mutation.