青海地区结核分枝杆菌检测与KatG、rpoB及gyrA的突变特征分析

Detection of mycobacterium tuberculosis in Qinghai area and analysis of gene mutation characteristics of KatG,rpoB and gyrA

目的探讨青海地区结核分枝杆菌(MTB)对利福平(RFP)、氟喹诺酮类药物、异烟肼(INH)等的耐受情况及与其相对应的rpoB、gyrA、katG突变基因关系。方法选择PCR和DNA测序法检测分析结核分枝杆菌分离株gyrA、katG以及rpoB的基因序列。结果经检测,耐INH菌株中katG基因突变率为70.9%,耐RFP菌株中rpoB基因突变率为83.33%,耐氟喹诺酮类药物菌株中gyrA基因突变率为77.27%。katG最常见的突变形式为Ser315Thr(86.67%),rpoB最常见的突变形式为526、531位氨基酸改变,总突变率为72.73%。氟喹诺酮类药物突变主要在gyrA基因耐药决定区(QRDR)发生,突变位点主要为94(65%)。结论 katG、gyrA、rpoB基因突变为青海地区结核分枝杆菌INH、氟喹诺酮类药物以及RFP耐药的主要分子机制,应用操作简单的PCR-DNA测序法,可作为临床结核分枝杆菌INH、氟喹诺酮类药物、RFP耐药性快速检测的辅助手段。

Objective To explore the drug resistance of mycobacterium tuberclosis （MTB）in Qinghai area to RFP,fluoroquinolones and INH and their relationship with the corresponding rpoB,gyrA,katG gene mutations.Methods PCR-DNA method was used to detect gene sequences of gyrA,katG and rpoB isolated by mycobacterium tuberculosis.Results After testing,katG gene mutation rate found in the resistant INH strain was70.9 ％.rpoB gene mutation rate in the resistant RFP strain was 83.33 ％.gyrA genemutation rate in fluoroquinolone resistant strains was 77.27 ％.KatG mutation was the most common form of Ser315Thr （86.67％）.The most common mutant forms of rpoB were amino acids 526 and 531,and the total mutation rate was 72.73％.Fluoroquinolones mutation occurred mainly in region decided by gyrA gene （QRDR） mutations,and the main point was in 94 （65 ％）.Conclusion Gene mutations of KatG,gyrA and rpoB are major molecular mechanism for drug resistance of mycobacterium tuberclosis （MTB） in Qinghai area to RFP,fluoroquinolones and INH.The PCR-DNA sequencing method is easy to apply and it can be used as assistant drug resistance in rapid detection of mycobacterium tuberculosis INH,fluoroquinolones and RFP in clinical practice.