摘要
目的建立并评估快速鉴定结核分枝杆菌复合群(MTBC)与非结核分枝杆菌(NTM)的多重PCR方法。方法应用分别针对MTBC的oxyR-ahpC基因间隔区、MTBC和NTM的rpoB基因可变区的3对分枝杆菌特异性引物进行多重PCR扩增,扩增产物分别为473bp、235bp和136bp。应用该多重PCR方法对6株MTBC标准株、50株NTM标准株、312株MTBC临床分离株和300株NTM临床分离株进行了初步菌种鉴定。结果经多重PCR扩增后进行凝胶电泳,MTBC标准株473bp和235bp片段均可见,NTM标准株仅见136bp片段。312株MTBC临床分离株中,310株扩增出473bp和235bp片段,敏感度为99.36%(310/312),特异度为99.32%(294/296);300株NTM临床分离株中,294株扩增出136bp片段,敏感度为98.00%(294/300),特异度为100.00%(310/310)。结论该多重PCR方法可检测并鉴别MTBC及NTM,具有高度的特异度和敏感度,有可能作为鉴别MTBC与NTM的有价值的检测手段。
Objective To establish and evaluate a multiplex PCR technique to rapid differentiate Mycobac- terium tuberculosis complex （MTBC） from non-tuberculous mycobacteria （NTM）. Methods Three pairs of oligo nucleotide primers were used in the multiplex PCR reaction. A 473 bp DNA fragment encoding oxyR-ahpC inter- genic region in MTBC, a 235 bp and 136 bp DNA fragment encoding variable rpoB gene region from MTBC and NTM, respectively, were amplified. The multiplex PCR was assessed in 6 reference strains of MTBC, 50 reference strains of NTM, 312 clinical strains of MTBC and 300 clinical strains of NTM. Results The multiplex PCR pro- duced two DNA fragments at the size 473 bp and 235 bp for MTBC reference strains, and one DNA fragment with the size 136 bp for NTM reference strains. Among 312 MTBC clinical samples, the sensitivity was 99.36~ （310/312） and specificity was 99.32% （294/296）. Among 300 NTM clinical samples, the sensitivity and specificity were 98. 00% （294/300） and 100.00% （310/310） respectively. Conclusion The multiplex PCR can differentiate MTBC and NTM efficiently, and might be a valuable technique for clinical use.
出处
《中国防痨杂志》
CAS
2013年第9期686-689,共4页
Chinese Journal of Antituberculosis
基金
北京市自然科学基金(7132049)
关键词
分枝杆菌
结核
分枝杆菌属
多重聚合酶链反应
Mycobacterium tuberculosis
Mycobacterium
Multiplex polymerase chain reaction
