快速检测耐异烟肼结核分支杆菌方法的研究  

Study on the Method of Rapidly Detecting Isoniazid-resistant Isolates of Mycobacterium Tuberculosis

作　　者：程晓东[1] 别良峰[1] 苏明权[1] 张荣[1] 于文彬[1] 郝晓柯[1] 

机构地区：[1]第四军医大学附属西京医院检验科,陕西西安710032

出　　处：《医学临床研究》2006年第9期1393-1396,共4页Journal of Clinical Research

摘　　要：【目的】应用多重聚合酶链反应-单链构象多态性分析(multiple-Polymerase Chain Reaction-SingleStrand Conformation Polymorphism,multi-PCR-SSCP)方法,快速、特异地同时检出耐异烟肼(isoniazid,INH)结核分支杆菌aphC启动子i、nhA、katG基因的突变情况,并与直接测序(derict sequencing,DS)结果相比较,参照药敏试验,探讨该方法的可行性。【方法】根据结核分支杆菌的aphC启动子序列i、nhA序列、katG序列,分别设计出3对特异性寡聚核苷酸引物,采用multi-PCR-SSCP技术,同时检测对结核分支杆菌耐INH起作用的这三个基因的突变情况;同时进行耐药株的测序分析。【结果】对H37 Rv标准株、临床分离INH敏感株(23株)及INH耐药株(35株)进行multi-PCR-SSCP分析,突变检出率82.9%(29/35);耐药株测序分析突变检出率为85.7(30/35)。两种方法的符合率为97.1%[(29+5/)/35]。【结论】耐药基因检测指导治疗是一种新探索,multi-PCR-SSCP方法敏感、特异,能同时快速有效地检测结核分支杆菌aphC启动子i、nhA、katG三个INH耐药基因的突变,提高检验效率,有望成为临床指导用药的好方法。[Objective]To use the multi-PCR-SSCP system for detecting the aphC promoter.inhA, katG gene mutations of the M. Tuberculosis isoniazid resistance isolates in the single reaction, and exploring the feasibility of this technique in comparison with the direct sequencing results and in reference to drug resistant results. [Methods]According to aphC promoter.inhA.katG genes of the M. Tuberculosis, three pairs of oligonueleotide primer were designed to examine the isoniazid resistance with the multi-PCR-SSCP, and at the same time to detect the mutations of drug resistance isolates using DS. [Results]The isoniazid sensitivity and resistance isolates were analyzed with multi-PCR-SSCP,and to analyse the drug resistence isolates DS, H37 Rv was used as control. The mutation rates were 82.9% and 85.7% respectively. The coincidence rate for two techniques was 97.1% [（29＋5）/35]. [Conclusion] The Multi-PCR-SSCP is a sensititive and specific method for rapid detection of aphC promoter, inhA, katG gene mutations in the M. Tuberculosis isoniazid resistance isolates. Drug resistance genes detection may be clinically useful in the therapy of patients with Tuberculosis.

关 键 词：分支杆菌 结核 药物耐受性 异烟肿 

分 类 号：R378.911[医药卫生—病原生物学]