PCR-SSCP法检测结核分枝杆菌耐药性

目的：探讨耐多药结核分枝杆菌耐药基因突变与耐药性的关系以及聚合酶链反应-单链构象多态性分析(poly merase chain reaction-single strand cinfomlation polymorphism，PCR-SSCP)方法的临床应用价值。方法：用PCR-SSCP方法检测58株结核分枝杆菌临床分离株katG，rpoB，rpsL基因突变并与常规药敏试验检测结果进行对比。结果：经常规药敏试验检测，58株结核分枝杆菌临床分离株中共有41株耐药，其中，耐异烟肼(INH)为35株，高耐药株27株；耐利福平(RFP)为31株，高耐药株24株；耐链霉素(SM)有31株，高耐药株26株。同时耐3种药物的有21株(51．2％)，耐2种药物的14株(34．1％)，单耐药株6株(14．6％)．PCR-SSCP方法对58株临床分离株katG，rpoB，rpsL基因突变的检测率为40％(23／58)，45％(26／58)，38％(22／58)，其中检出3个基因同时突变的有13株(32％)，2种基因突变的12株(29％)，1种基因突变的有10株(23．8％)．常规药敏试验与PCR-SSCP法检出结核分枝杆菌同时耐3种药物的符合率为61．9％(13／21)，检出耐2种药物的符合率为85．70k，(12／14)，检出耐1种药物的符合率为50％(3／6)．高耐药株中突变率为80．5％(62／77)，低耐药株中突变率为60％(12／20)．结论：PCR-SS-CP方法对耐2种以上药物的结核杆菌检出率较高，且耐药基因突变率随着耐药浓度增高而增高。将PCR-SSCP法与药敏试验联合应用可互相弥补，已成为临床指导用药的好方法。

Clinical application of PCR-SSCP in detecting Mycobacterium tuberculosis drug resistance

AIM To study the relationship of gene mutation and drug resistance in Mycobacterium tuberculosis and to evaluate the clinic application of PCR SSCP. METHODS: katG, rpoB, rpsL genes mutation in 58 clinical isolated strains of Mycobacterium tuberculosis were detecting by PCR SSCP and traditional drug susceptibility test, and the results were compared. RESULTS: The results of drug susceptibility test showed that there were 41 multi drug resistant strains in the 58 clinical isolated organisms. The number of three drug resistant strains was 21 (51.2%) and that of two drug resistant strains was 14 (34.1%), but only 6 (14.6%) strains were one drug resistant. The gene mutation rate of INH, RFP and SM was 40% (23/58), 45% (26/58) and 38% (22/58) by PCR SSCP in the 58 clinical isolated strains and the rate of one, two and three gene mutations was 32%, 29% and 23.8%, respectively. The coincidence rate of three drug resistance in isolates by susceptibility test and PCR SSCP was 61.9% (13/21), and those of two drug and of one drug were 85.7% (12/14) and 50% (3/6). The mutation rate of high resistant strains was 80.5% (62/77) and low resistant strains 60% (12/20). CONCLUSION: The detecting rate of two drug resistant strains and three drug resistant strains was higher and the mutation rate increases with the level of drug resistance. The combined application of PCR SSCP and traditional drug susceptibility test can enhance the detecting rate.