目的 初步了解四川省肺结核患者分枝杆菌菌种类型。 方法 收集267株分枝杆菌临床菌株,经PNB/TCH鉴别培养基进行培养鉴定后,采用聚合酶链反应(PCR)对16S rRNA、Rv0577、IS1561、Rv1510、Rv1970、Rv3877/8和Rv3120 基因位点进行扩增,鉴定至种,再经PRA-rpoB、hsp65基因测序进行验证。 结果 267株分枝杆菌临床分离株多位点PCR结果显示结核分枝杆菌262株,非洲分枝杆菌Ⅰ型3株,非结核分枝杆菌2株。PNB/TCH鉴别培养基培养鉴定结果为结核分枝杆菌复合群266株,非结核分枝杆菌1株。2株非结核分枝杆菌分别为鸟分枝杆菌(M. avium)和脓毒分枝杆菌(M. septicum)。多位点PCR结果与rpoB-PRA、hsp65基因测序结果一致。 结论 多位点PCR技术鉴定分枝杆菌菌种结果准确可靠,且具有简便和快速等优点,有较大的分子流行病学应用价值,且对于临床诊断和治疗都具有重要意义。
Objective To understand the species of Mycobacteria isolated from TB patients in Sichuan. Methods Multi-locus PCR was conducted to amplify 7 specific DNA fragments of 16S rRNA, Rv0577, IS1561, Rv1510, Rv1970, Rv3877/8 and Rv3120 in the genome of 267 Mycobacterium strains which had been identified by PNB/TCH previously. Their species were verified by PRA-rpoB. identification and hsp65 sequencing. Results The multi-locus PCR results showed that 262 strains belonged to Mycobacterium tuberculosis complex (MTBC), 3 belonged to Mycobacterium africanum type Ⅰ and 2 were non-tuberculosis mycobacterium (NTM) strains, and by PRA-rpoB identification and hsp65 sequencing, 2 NTM strains were Mycobacterium avium and Mycobacterium septicum respectively While PNB/TCH result showed that 266 strains belonged to MTBC, 1 was NTM. The result of multi-locus PCR was consistent with that of rpoB-PRA identification and hsp65. sequencing Conclusion Multi-locus PCR is a rapid and effective method for the species identification of Mycobacterium and can be used in molecular epidemiology research and clinical diagnosis.