Objective To develop and evaluate the multiplex allele - specific PCR ( MAS - PCR) for rapid detection of Mycobacterium tuberculosis (M. tuberculosis) resistannce to quinolone drugs. Methods Based on the sequence of gyrA gene in M. tuberculosis,four pairs of specific primers were designed for detecting the most commom mutations at the condon 90 and 94 of gyrA gene. PCR conditions were optimized. DNA direct sequencing and conventional proportion methods were used to evaluate the new method. Results 144 strains of quinolines - resistant and 56 strains of quinolines - sensitive clinical M. tubercutosis strains were detected by MAS - PCR. Compared with results of the proportion method and DNA sequencinS, the sensitivity af MAS - PCR were 53.57％ (30/56) , 71.43％ (30/ 42) reapectively, while the specificity were all 100％ . Conclusion MAS - PCR could be used to detect M. tuberculosis resistance to quinolones drugs rapidly. It is a aimple and specific method, but the sensitivity needs to be improved.%目的 建立等位基因特异性多重聚合酶链反应(multiplex allele-specific PCR,MAS-PCR)技术,以快速检测结核分枝杆菌对喹诺酮的耐药性.方法 根据结核分枝杆菌gyrA基因序列,分别设计4条特异性寡聚核苷酸引物,优化PCR条件,建立针对gyrA 基因90和94位点突变进行检测的MAS-PCR技术,对临床分离菌株进行喹诺酮耐药性检测,同时以比例法和DNA直接测序法做对照.结果 共对144株喹诺酮敏感株和56株喹诺酮耐药株进行了MAS-PCR检测.MAS-PCR与比例法比较,敏感性为53.57%(30/56);与DNA测序比较,敏感性为71.43%(30/42),特异性均为100%(144/144).结论 MAS-PCR技术检测结核分枝杆菌对喹诺酮耐药性简便、快速、特异性高,但敏感性需进一步提高.
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