Emergence of fluoroquinolone-resistant Streptococcus pyogenes in Japan by a point mutation leading to a new amino acid substitution

Objectives: Streptococcus pyogenes causes various diseases in humans. While the prevalence of fluoroquinolone-resistant S. pyogenes isolates has been increasing since 2000 in the USA and Europe, it has remained very low in Japan. We isolated a fluoroquinolone-resistant S. pyogenes strain and analysed its genetics. Methods: TU-296, a strain of S. pyogenes resistant to levofloxacin (MIC 16 mg/L), was isolated from the throat of a patient in their thirties with pharyngitis in autumn 2007. We carried out susceptibility tests for various antimicrobial agents and PCR analysis of the genes gyrA, gyrB, parC and parE in the quinolone resistance-determining region, followed by sequencing of the PCR products to find mutation(s) and the resulting amino acid substitution(s). We then sequenced the PCR product of the emm gene and determined the emm genotype. Results: S. pyogenes TU-296 was found to have the following mutations and amino acid substitutions: adenine 476 to cytosine in gyrA and cytosine 367 to thymine in parC, resulting in Glu-85→Ala in GyrA and Ser-79→Phe in ParC. The genotype of the isolate was emm11. Conclusions: Amino acid substitutions in fluoroquinolone-resistant S. pyogenes have already been reported from Europe and the USA, including Ser-81→Phe or Tyr and Met-99→Leu in GyrA, as well as Ser-79→Phe, Tyr or Ala and others in ParC. Numerous point mutations were found in parC and parE of S. pyogenes TU-296. In addition, a new amino acid substitution was detected (Glu-85→Ala in GyrA). To our knowledge, there have been no previous reports of this substitution in a clinical isolate of S. pyogenes.