Incidence of various gyrA mutants in 451 Staphylococcus aureus strains isolated in Japan and their susceptibilities to 10 fluoroquinolones

Abstract

Point mutations in the gyrA genes of 451 clinical strains of Staphylococcus aureus isolated in Japan were detected by a combination of nonradioisotopic single-strand conformation polymorphism analysis and restriction fragment length polymorphism analysis and by direct sequencing. Six types of gyrA mutations were observed in 149 of the 451 strains (33%), and ofloxacin MICs were greater than 6.25 g/ml for 147 of the 149 strains (98.7%). These mutations were localized between codons 84 and 88, and they were associated with fluoroquinolone resistance. Two types of silent mutations were also found. Among these eight types of mutations, three types are novel, i.e., the serine at position 84 (Ser-84)3Val (TCA3GTA), Ser-843Leu (TCA3TTA) plus Ile-86 (ATT3ATC, silent), and Phe-110 (TTT3TTC, silent). Among GyrA mutants, strains with a Ser-843Leu alteration and strains with a Glu-883Lys alteration were dominant. In contrast, few strains had Ser-843Ala and Glu-883Gly alterations. All fluoroquinolones tested showed greater than a fourfold decrease in their activities in terms of their MICs that inhibited 50% of strains tested for each GyrA mutant, in comparison with their MICs that inhibited 50% of strains tested for susceptible strains. Most of the currently available fluoroquinolones, such as norfloxacin, enoxacin, ofloxacin, ciprofloxacin, tosufloxacin, lomefloxacin, sparfloxacin, and fleroxacin, were ineffective against each mutant. Mutants containing a Ser843Leu or Val alteration showed high-level resistance to fluoroquinolones, and one containing a Ser-843Ala alteration showed relatively low-level resistance. Double mutations were associated with a higher level of resistance than single mutations.

Detection of Ciprofloxacin-Resistant Yersinia pestis by Fluorogenic PCR Using the LightCycler

“…Based on the results of our characterization of 65 Y. pestis Cp r mutants, the most common site of mutation in GyrA is Gly-81. Other studies have indicated that Ser-83 is the most common hotspot for changes in gyrA that result in Cp r (7,11,20,21,23,30). The fact that Gly-81 appears to be a hotspot for mutation in Y. pestis may reflect a difference in the organism's DNA repair capabilities or a difference in the tertiary structure of GyrA.…”

“…The substitutions in Y. pestis GyrA were Gly-81 to Asp or Cys and Ser-83 to Ile or Arg. All of the amino acid substitutions we identified in Y. pestis GyrA have been found in other organisms (4,15,20,23,25,30). Based on the results of our characterization of 65 Y. pestis Cp r mutants, the most common site of mutation in GyrA is Gly-81.…”

“…This method requires the isolation of the pathogen followed by an extra day of incubation with disks or strips. DNA-based methods for Cp r detection, such as mismatch amplification mutation assay (MAMA) combined with DNA sequencing (32) and single-stranded confirmation poly- morphism (23), have been developed. All of these methods require electrophoresis of reaction products to determine if a mutant allele of a cellular gyrase is encoded by the isolate.…”

Antibacterial-ResistantPseudomonas aeruginosa: Clinical Impact and Complex Regulation of Chromosomally Encoded Resistance Mechanisms

“…Similar to the case for other gram-negative bacteria, DNA gyrase is the primary target for the fluoroquinolones in P. aeruginosa (84). Therefore, the first target-specific mutations are typically observed within the quinolone resistance determining region (QRDR) of gyrA (6,116,165,249,283). The highest levels of resistance are observed in strains that have mutations in the QRDR of both gyrA and the topoisomerase IV gene parC (6,90,91,116,165).…”

Molecular Detection of Antimicrobial Resistance

“…Meanwhile, several investigators have applied SSCP analysis to the detection of mutations, mainly in gyrA, in several bacterial species (47,203,230,325,331,350). In the most recent study, Takenouchi et al (332) studied gyrA point mutations in 335 clinical P. aeruginosa isolates by nRI-SSCP analysis and direct sequencing.…”