Mutations in the gyrA and gyrB genes of fluoroquinolone-resistant Mycobacterium tuberculosis from TB patients in Thailand.

ABSTRACT

Among fluoroquinolone-resistant Mycobacterium tuberculosis (FQr-MTB) isolates, mutation at positions 90, 91, and 94 in gyrA gene and at positions 495, 516, and 533 in gyrB gene have been frequently reported. In this study, 35 isolates of FQr-MTB were collected from Siriraj Hospital and Chest Disease Institute. The quinolone-resistance-determining regions (QRDR) of gyrA and gyrB genes in all 35 FQr-MTB isolates and from the H37Ra MTB strain were amplified using polymerase chain reaction (PCR). DNA-sequencing and single-strand conformation polymorphism (SSCP) were further utilized for characterization of the mutations in the QRDR of gyrA and gyrB genes and mutation screening, respectively. From DNA-sequencing, 21 of 35 (60%) exhibited single-point mutations in different positions, at Ala90Val, Ser91Pro, and Asp94(Gly/Ala/His/Asn); and one novel mutation position at Gly88Cys in the gyrA gene and Asp495Asn in the gyrB gene. These positions were previously frequently reported to be responsible for FQr-MTB. The other 14 FQr-MTB isolates (40%) had no mutation. This study is the first report of mutation occurring only in the QRDR of the gyrB gene, without prior mutation in the gyrA QRDR among FQr-MTB isolates. By SSCP analysis for screening of the mutant FQr-MTB, the SSCP patterns of mutated FQr-MTB isolates were clearly differentiated from the SSCP patterns of FQs-MTB.