结核分支杆菌耐药基因检测结果分析

Study of mycobacterium tuberculosis in patients with pulmonary tuberculosis by drug resistance gene detecting

目的分析肺结核患者结核分支杆菌获得性耐药情况，比较两种耐药性检测方法的检测效果。方法用药敏试验（绝对浓度法）检测结核分支杆菌株对利福平（RFP）、异烟肼（INH）、链霉素（SM）、吡嗪酰胺（PZA）和乙胺丁醇（EMB）的耐药情况，采用聚合酶链反应-单链构象多态性分析（PCR-SSCP）检测结核分支杆菌耐药rpoB、katG、rpsL、pncA和embB基因突变。结果400例肺结核耐药患者常规药敏试验耐药316例（79．8％）；耐药基因检测耐药株288株，突变率72．0％。RFP耐药例数和耐药率明显高于SM、PZA和EMB（耐药例数：F=2．45，2．56，2．69，P〈0．05；耐药率：F=2．55，2．66，2．79，P〈0．05），rpoB突变株和突变率明显高于katG、rpsL、pncA和embB（突变株：F=2．28，2．46，3．19，3．33，P〈0．05～0．01；突变率：F=2．36，2．61，3．25，3．45，P〈0．05～0．01），高浓度药物耐药菌株的突变株和突变率显著高于低浓度药物耐药菌株，随着不规律用药时间的延长，耐药率和突变率均逐渐上升（耐药率：F=2．77，2．88，P〈0．05；突变率：F=2．72，2．85，P〈0．05）。结论PCR．SSCP是一种快速检测结核杆菌rpoB、katG、rpsL、pncA和embB基因突变敏感、特异的方法。

Objective To investigate drug resistance of mycobacterium tuberculosis, compare detecting effect of two methods and evaluate their value of clinical application. Methods All of the strains of mycoba-cterium tuberculosis were tested for resistance to RFP, INH, SM, PZA and EMB by the absolute concentration method on lowenste in-jensen medium and the mutation of the rpoB,katG,rpsL, pncA and embB resistanc genes in mycobacterium. Tuberculosis was tested by polymerase chain reaction-single strand conformation polymorphism（ PCR-SSCP）. Results 400 cases of tuberculosis drug resistance in patients with conventional susceptibility test resistance 316 cases （79.8 % ） ; resistant strains resistant gene detection 288,the mutation rate（72.0 % ） ; the RFP cases of resistance and resistance was significantly higher than that of SM, PZA and the EMB （resistant cases ： F = 2.45,2.56,2.69, P 〈 0.05 ;Resistance： F = 2.55,2.66,2.79, P 〈 0.05）;the mutants and mutation rate of rpoB was higher than that of katG, rpsL, rpoB and embB （ mutant ： F = 2.28,2.46,3.19,3.33, P 〈 0.05-0.01 ; mutation rate ： F = 2.36,2.61, 3.25,3.45, P 〈 0.05-0.01 ）;the gene mutation rate was higher in strains isolated from high concentration resistance than those in strains isolated from low concentration resistance; the more irregular treatment was longer, the rate of drug-resistant was higher（resistance ： F = 2.77,2.88, P 〈 0.05 ; mutation rate ： F = 2.72,2.85, P 〈 0.05 ）. Conclusion PCR-SSCP is a sensitive and specific method for rapid detecting rpoB, katG, rpsL, pncA and embB gene mutations of myeobacterium tuberculosis.