Abstract: Objective  To investigate the function of the genes related with drug resistance and efflux pump in mono ofloxacin (Ofx)-resistant M. tuberculosis (Mtb) isolates. Methods  Seventeen mono Ofx-resistant Mtb clinical isolates were obtained from National Tuberculosis Reference Laboratory. Direct sequencing was performed to detect the mutations in the gyrA/B genes. The total RNA was extracted from the Ofx-resistant strains without gyrA/B mutations. After RNA reverse transcription, the expression levels of 20 drug efflux genes were detected by Real-time PCR. In comparison with the corresponding MIC results, the Ofx-related efflux pumps were screened from 20 drug efflux genes, and the function of candidates was identified among transgenic E.coli. Results  Of 17 mono Ofx-resistant strains, 4 (4/17) strains were found the mutations of gyrA, in which 1 isolates had mutation at codon 90 and 3 isolate had mutation at codon 94. In addition, the isolates harboring gyrA mutation displayed high-level of drug resistance, their MICs were more than 4 μg/ml. Among the strains without gyrA/B mutations, the expression levels of Rv0933 and Rv2938 were higher in 2 high-level resistant strains, which was 16 and 5 folds higher than that of H37Rv strain, respectively. Furthermore, E.coli BL21 transferred with pEASY-E1-Rv0933 showed higher MIC level, 2μg/ml, compared with the control E.coli. Conclusion  The mutations in gyrA are associated with high-level of Ofx resistance. Rv0933 may be the Ofx-specific efflux pumps in Mycobacterium tuberculosis. The high level of Rv0933 expression may correlated with high-level of Ofx resistance.

Key words: Mycobacterium tuberculosis, Fluoroquinolones, Drug resistance, bacterial, DNA Gyrase