耐药结核分枝杆菌基因变异研究

Direct sequencing for katG and inhA genes of multi drug resistant isolates of mycobacterium tuberculosis

目的 研究耐多药结核分枝杆菌katG、inhA基因的变异机理。方法 用katG、inhA基因片段为引物 ,聚合酶链反应扩增产物 ,克隆后制备质粒 ,直接测定DNA序列。结果 15株耐异烟肼菌株均有katG基因突变 ,主要为点突变 ,其中 14株有 4 63位和 3 15位AA密码子改变。在 18个耐异烟肼菌株中 ,全部出现基因inhA基因变异 ,主要为点突变和缺失 ,以单个位点变异为主 ,各菌株变异不同。katG、inhA基因同时出现变异的比例高。结论 结核分枝杆菌耐药基因变异复杂 ,我国耐多药结核菌株与国外的菌株katG、inhA基因变异特点不同。

Objective To characterize the katG and inhA genetic variations in multi drug resistant isolates of M. tuberculosis. Methods The amplicons were cloned and sequenced by ABI 377 automatic sequencing system. Results All isolates of M. tuberculosis with MIC for INH ≥1 μg/ml had point mutations and insertion in katG gene. Fourteen of 15 isolates (86.7%) had 463 codon (Arg→Leu) and 315 (Ser→Asn/Thr) mutations. Nine of 13 mutation positions were not reported previously. All 18 isolates with MIC≥1 μg/ml had one or more variations in inhA. The variations consisted of one or more base pairs deletions or point mutation. Each strain had its unique mutation. Seven of 10 isolates had both katG and inhA mutations. Conclusion Genetic variants of M. tuberculosis are significantly different from those of foreign strains.