Multiplex allele-specific PCR combined with PCR-RFLP analysis for rapid detection of gyrA gene fluoroquinolone resistance mutations in Mycobacterium tuberculosis

A combined use of MAS-PCR (multiplex allele-specific PCR) and PCR-RFLP (PCR restriction fragment length polymorphism), was established to detect mutations in codons 90, 91 and 94 of the gyrA gene in Mycobacterium tuberculosis (M. tuberculosis). With conventional phenotypic drug susceptibility testing as a reference standard, the sensitivity, specificity and accuracy of the modified method for gyrA gene mutation detection were 70.8%, 100% and 84.8% respectively.

► We established MAS-PCR combined with RFLP analysis to detect fluoroquinolone resistance in M. tuberculosis.
► The method could find out the possible mutations in codons 90, 91 and 94 of the gyrA gene in M. tuberculosis.
► The performance of the method was evaluated with 125 clinical isolates of M. tuberculosis from China.
► The method is sensitive, specific and accurate enough to permit rapid detection of fluoroquinolone resistant M. tuberculosis.