荧光PCR探针熔解曲线法检测结核分枝杆菌耐利福平突变研究

Evaluation of the mutations detection of rpoB gene in Mycobacterium tuberculosis clinical isolates by probe melting analysis based real-time PCR

目的对荧光PCR熔解曲线法检测结核分枝杆菌耐利福平突变方法进行临床研究,评价检测能力及在国境口岸的应用价值。方法应用荧光PCR熔解曲线法检测结核病人临床分离结核分枝杆菌,以传统药物敏感性试验为标准,获得该方法的灵敏性、特异性。结果对347例结核病人临床分离培养样本,传统药物敏感性试验检出271例利福平敏感标本,76例利福平耐药标本。荧光PCR熔解曲线法检出269例利福平敏感标本,78例利福平耐药标本,灵敏性为93.42%,特异性为97.42%,符合率为96.54%。结论荧光PCR熔解曲线法检测速度快速、灵敏度高、特异性强,可用于结核分枝杆菌利福平耐药突变的快速检测,适于国境口岸对耐多药结核病的快速筛查。

Objective To evaluate the analysis abilities and valuation of the melting analysis based real-time PCR for detecting the mutations of ropB gene in mycobacterium tuberculosis from clinical samples at frontier port.Methods The probe melting analysis assay was adopted for the sensitivity and specificity in detecting the isolates from clinical samples against the standard of drug susceptibility testing（DST）.Results A total of 347 mycobacterium tuberculosis isolates were detected by two methods,in which,271 rifampin sensitivity isolates and 76 rifampin resistant isolates were gained by DST.Meanwhile,269 rifampin sensitivity isolates and 78 rifampin resistant isolates were gained by the probe melting analysis assay,the sensitivity and specificity of melting analysis assay was 93.42% and 97.42%,respectively.The concordance of the two methods was 96.54%.Conclusion The probe melting analysis assay own the characteristics of short-time,higher sensitivity and specificity for detecting rifampin resistance in tuberculosis using clinical sample and is available to use for rapid screening multidrug drug resistant tuberculosis at frontier ports.