利用荧光定量RT-PCR和PCR方法检测结核分枝杆菌复合群

Rapid detection of Mycobacterium tuberculosis complex by combining real-time RT-PCR and real-time PCR assay

目的建立一种新的检测结核分枝杆菌复合群的荧光定量试验方法(R/P分析),探讨R/P分析检测临床标本中结核分枝杆菌复合群的应用价值。方法根据结核分枝杆菌复合群基因保守序列设计引物和探针构建质粒标准品。运用R/P分析检测54例确诊结核病人临床样本,检测的结果同时与培养法、荧光定量PCR法进行比较。结果不同临床标本用R/P分析诊断结核病,其敏感性高于荧光定量PCR法和培养法,阳性检出率分别为96.3%、83.3%和55.6%。结论 R/P方法是一种快速、特异的直接检测方法,它可以区分结核分枝杆菌复合群的死菌与活菌,因此可以更好的指导医生进行治疗,更准确地做出公共卫生决策。

Objective To develop a new real-time assay（R/P assay） for detection of Mycobacterium tuber- culosis complex and evaluate the application value of R/P assay in detection of Mycobacterium tuberculosis complex （MTBC） in clinical specimens. Methods The primers and probe were designed and constituted standard sample. The R/P assay was applied to 54 specimens of the clinical tuberculosis cases and the results were compared with those of culture and real-time PCR. Results The positive rate of R/P assay, Real time PCR,culture by the different clinical specimens were 96. 3% ,83.3% and 55.6% respectively. Conclusion The R/P assay is a rapid and specific method for direct detection of MTBC, which also can distinguish viable and nonviable MTBC, thus may guide patient therapy, and public health decisions.