7例皮肤结核分离菌株药敏试验及利福平、异烟肼耐药基因突变情况分析

Analysis of antimicrobial susceptibility and mutations of genes associated with rifampicin and isoniazid resistance in Mycobacteria tuberculosis isolates from 7 patients with cutaneous tuberculosis

目的 探讨皮肤结核中结核分枝杆菌耐药及rpoB、katG耐药相关基因的突变情况.方法 从皮肤结核患者皮损或分泌物中分离、鉴定出7株结核分枝杆菌,采用比例法的间接法分别检测7株结核分枝杆菌对利福平、异烟肼、链霉素和乙胺丁醇四种一线抗结核药物的耐药情况,并对利福平耐药基因rpoB和异烟肼耐药基因katG进行PCR扩增和DNA测序.结果 7株结核分枝杆菌中,6株对利福平、异烟肼、链霉素和乙胺丁醇均敏感,1株对利福平、异烟肼和乙胺丁醇同时耐药.7株结核分枝杆菌均扩增出rpoB和katG基因条带.DNA测序结果显示,临床耐多药菌株rpoB基因531位密码子由TCG突变为TTG,katG基因315位密码子由AGC突变为ACC;其余6株药物敏感株均未发现上述基因突变.结论 皮肤结核中已分离出耐多药菌株,耐多药菌株的形成可能与治疗不当相关.

Objective To assess the antimicrobial resistance and resistance-related mutations in rpoB and katG genes in Mycobacteria tuberculosis isolates from patients with cutaneous tuberculosis. Methods Seven strains of Mycobacteria were isolated from lesions or secretions of patients with cutaneous tuberculosis and identified as M. tuberculosis. Proportion method was used to test the susceptibility of M. tuberculosis isolates to rifampicin, isoniazid, streptomycin and ethambutol. PCR and sequencing were performed to analyze the mutations in rpoB and katG genes. Results Of the 7 isolates of M. tuberculosis, 1 was resistant to rifampicin,isoniazid and ethambutol simultaneously, and the other 6 were sensitive to rifampicin, isoniazid, streptomycin and ethambutol. All the 7 isolates were positive for the amplification of rpoB and katG genes by PCR. DNA sequencing revealed two mutations at codon 531 （TCG to TTG） and codon 315 （AGC to ACC） in the multi-drug resistant strain, which were absent in the other 6 strains. Conclusion Multi-drug resistance has emerged in M. tuberculosis isolates from patients with cutaneous tuberculosis, which is likely to be related to improper treatment.