分子杂交法快速鉴定结核分枝杆菌和非结核分枝杆菌的初步评价

Primary Evaluation of Molecular Hybridization Method for the Identification of Mycobacterium tuberculosis and Non-tuberculosis Mycobacterium

目的评价结核分枝杆菌鉴定试剂盒（分子杂交法）进行分枝杆菌菌种初筛的准确性，并探讨如何快速准确地鉴定结核分枝杆菌复合群（MTC）和非结核分枝杆菌（NTM）混合感染标本。方法采用分子杂交法对200株结核分枝杆菌临床分离株进行菌种鉴定。人工制备结核分枝杆菌和胞内分枝杆菌、结核分枝杆菌和脓肿分枝杆菌、结核分枝杆菌和偶发分枝杆菌的混合感染模型，评价基因测序法、芯片法和Haintest在检测分枝杆菌混合感染中的准确性。结果以对硝基苯甲酸（PNB）分离鉴定结果为参照，分子杂交法的敏感性和特异性分别为65．12％（65／86）和98．25％（112／114）。对于人工制备的13种不同比例混合的H37Rv和NTM，芯片法和Haintest均可以准确鉴定出混合感染标本中的混合菌种，而基因测序结果表明，H3RV：偶发分枝杆菌的比例在10％～90％范围内才能区分出混合感染的类型。结论分子杂交法对于结核分枝杆菌诊断有较高的特异性，一旦经该法鉴定为NTM，则具有很高的诊断意义。明确标本中存在混合感染后可以采用芯片法或Haintest对临床上高致病的分枝杆菌进行菌种水平的鉴定。

Objective To evaluate the accuracy of Mycobacterial species identification kit （ molecular hybridization method）, and to investigate the rapid and accurate approaches for the identification of mixed infection caused by Mycobacterium tuberculosis complex （ MTC ） and non-tuberculosis Mycobacterium （ NTM ）. Methods A total of 200 clinical Mycobacterium isolates were identified by molecular hybridization method. The mixed infection was simulated by the mixture of Mycobacterium tuberculosis and Mycobacterium intracellular , Mycobacterium tuberculosis and Mycobacterium kansasii, Mycobacterium tuberculosis and Mycobacterium fortuitum. The gene sequencing, gene chip array and Hain test were used to evaluate the accuracy of Mycobacterium-mixed infection. Results As reference to pnitrobenzoic acid （PNB） tests, the sensitivity and specificity of molecular hybridization method were 65.12 % （ 65/86 ） and 98.25% （112/114） , respectively. Among 13 different proportions of Mycobacterium tuberculosis and NTM, gene chip array and Hain test accurately detected all of the mixed infections. Gene sequencing can only detect the mixed infection caused by Mycobacterium tuberculosis and Mycobacterium fortuitum at a range from 10%to 90%. Conclusions The specificity of molecular hybridization method for the identification of Mycobacterium tuberculosis is high. The kit following the method for NTM has a significant diagnosis value. Once the mixed infection is identified in the culture, gene chip array and Hain test can identify the species of mixed culture for the common pathogenic Mycobacterium.