RNA恒温扩增实时检测技术快速鉴定偶发分枝杆菌的研究

发布时间：2018-03-23 00:40

本文选题：RNA恒温扩增　切入点：鉴定　出处：《中国人兽共患病学报》2017年09期 　论文类型：期刊论文

【摘要】：目的建立RNA恒温扩增实时检测技术快速鉴定偶发分枝杆菌(RIARD-MF)的方法,评估其在鉴定分枝杆菌临床分离株中的应用效果。方法将偶发分枝杆菌的16SrRNA特异序列作为目的靶标进行检测,设计含T7启动子逆转录扩增引物和RNA探针,分别对5种非分枝杆菌细菌、20种分枝杆菌标准株和259株临床分枝杆菌分离株进行42℃恒温扩增实时检测,参考标准为PCR基因测序结果。结果RIARD-MF方法学检测灵敏度可达60CFU/mL。在25种细菌的RIARD-MF特异性检测中:只有偶发分枝杆菌检测结果为阳性,其余24种细菌均为阴性,与测序检测结果一致。在检测分枝杆菌临床分离菌株时,RIARD-MF鉴定偶发分枝杆菌5株,其余为非偶发分枝杆菌,与测序结果一致,检测灵敏度和特异度均为100%。结论 RIARD-MF鉴定偶发分枝杆菌具有较高的特异度、灵敏度和准确性,且检测快速,有望成为一种新的偶发分枝杆菌临床分离菌株鉴定方法。
[Abstract]:Objective to establish a real-time detection of RNA isothermal amplification technique for rapid identification of Mycobacterium fortuitum (RIARD-MF) method, and evaluate the application effect in the identification of mycobacteria in clinical isolates. Methods 16SrRNA specific sequence of Mycobacterium fortuitum as objective target detection, design containing T7 promoter transcription PCR primers and RNA probe respectively to 5 non Mycobacterium bacteria, 20 Mycobacterium standard strains and 259 clinical isolates of Mycobacterium were amplified at 42 C for the real-time detection of the reference standards for PCR gene sequencing. RIARD-MF specific test results RIARD-MF method detection sensitivity to 60CFU/mL. in 25 kinds of bacteria in Mycobacterium fortuitum: only the test result is positive, the rest 24 kinds of bacteria were negative, and the sequencing results were consistent. In the detection of Mycobacterium tuberculosis clinical isolates, RIARD-MF identified 5 strains of Mycobacterium fortuitum, the More than for the non Mycobacterium fortuitum, consistent with the sequencing results, specificity, sensitivity and specificity for detecting 100%. RIARD-MF identification conclusion Mycobacterium fortuitum has high sensitivity and accuracy, and fast detection, is expected to become a new kind of Mycobacterium tuberculosis clinical isolates and identification methods.

【作者单位】：中国海洋大学环境科学与工程学院;同济大学附属上海市肺科医院/上海市结核病(肺)重点实验室;
【基金】：国家十二五传染病重大专项课题(2013ZX10003001-003) 上海市自然基金课题(17ZR1423400)~~
【分类号】：R440;R516

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