结核分枝杆菌临床分离株异烟肼耐药相关基因突变的分子特征

Molecular characterization of mutations associated with isoniazid resistance in Mycobacterium tuberculosis isolates

目的阐明结核分枝杆菌异烟肼(INH)耐药相关基因突变特征。方法对137株结核分枝杆菌临床分离株(耐异烟肼菌株87株,异烟肼敏感菌株50株)的9个结构基因furA、katG、inhA、kasA、Rv0340、iniB、iniA、iniC和efpA以及两个调控区oxyR-ahpC基因间隔区和mabA-inhA启动子进行DNA片段扩增及序列分析。结果82株(94．3％)INH耐药分离株的katG基因存在突变,其中katG Ser315Thr突变占优势(55．2％)。50株INH敏感的分离菌katG的463密码子没有突变。35株(40．2％)INH耐药的分离株katG的463有突变。87株INH耐药株中,20株(23．0％)的katG基因存在两重突变。13株(14．9％)分离菌inhA基因的启动子区存在突变,4．6％的分离菌有inhA结构基因突变,11．5％oxyR-ahpC基因间区存在突变。iniBAC区域和efpA中发现耐药性关联突变。结论研究证实多个基因突变与异烟肼耐药之间的关系,并且为阐明结核分枝杆菌耐药机制提供线索。

Objective To characterize the mutations associated with isoniazid （INH） resistance in clinical isolates. Methods Nine structural genes （furA, katG , inhA , kasA , Rv0340, iniB , iniA , iniC and efpA ） and two regulatory regions （the oxyR-ahpC intergenic region and the promoter of mabA-/nbA） in 87 INH-monoresistant and 50 INH-susceptible M. tuberculosis isolates were analyzed by sequencing. Resets 82 （94.3 % ） INH-resistant isolates had mutations in katG gene, with the katG Ser315Thr mutation predominant （55.2%）. No mutation at cedon 463 of katG was detected in 50 INH-susceptible isolates with different IS6110 fingerprints, and there were 35 （40.2%） INH-resistant isolates had mutation at the cedon 463 of katG. Among the 87 INH-resisrant strains, 20 （23.0%） isolates harbored double mutations at two separate loci of katG. Mutations in the inhA promoter region occurred in 13 （ 14.9% ） isolates; 4.6% of the isolates had inhA structural gene mutations, and 11.5% harbored mutations in the oxyR-ahpC intergenic region. Drug resistance associated mutations were detected in the iniBAC region and efpA. Conclusion These findings proved gene mutations associated with INH resistance and provide clues for elucidating the drug-reistance mechanisms of M. tuberculosis.