目的:通过比较分析不同耐药的结核杆菌临床分离株PhoP基因和PhoR基因的表达水平差异,探讨研究结核杆菌PhoPR双组分系统与新疆地区广泛流行的耐药结核杆菌耐药性是否具有相关性。方法以新疆地区广泛流行的耐药结核杆菌临床分离株为研究对象,分别提取结核分枝杆菌临床分离药物敏感菌株、单纯耐异烟肼的结核杆菌临床分离株( INH-MTB)、单纯耐利福平的结核杆菌临床分离株( RFP-MTB)、单纯耐链霉素的结核杆菌临床分离株( SM-MTB)、单纯耐乙胺丁醇的结核杆菌临床分离株( EB-MTB)、耐多药结核杆菌临床分离株( MDR)总RNA,并进行纯度鉴定,运用SYBR GreenⅠ实时荧光定量PCR的方法,分别检测各组结核杆菌菌株的PhoP基因和PhoR基因的表达,比较分析不同耐药的结核杆菌临床分离株PhoP基因和PhoR基因的表达水平的差异。结果与结核分枝杆菌临床分离药物敏感菌株相比,PhoP基因在RFP-MTB中的表达上调1.48倍,在MDR中的表达上调2.74倍,差异有统计学意义(P<0.05);与MDR相比,PhoP基因在INH-MTB、RFP-MTB、SM-MTB和EB-MTB中的表达下调0.70、0.50、0.25和0.21倍,差异有统计学意义(P<0.05)。与结核分枝杆菌临床分离药物敏感菌株相比,PhoR基因在INH-MTB、RFP-MTB、SM-MTB和EB-MTB中的表达分别上调6.33、4.56、2.34和1.85倍,在MDR中的表达上调9.06倍,差异有统计学意义( P<0.05);与 MDR相比,PhoR基因在INH-MTB、RFP-MTB、SM-MTB 和EB-MTB 中的表达下调0.36、0.54、0.35和0.19倍,差异有统计学意义(P<0.05)。结论在结核分枝杆菌临床分离药物敏感菌株、INH-MTB、RFP-MTB、SM-MTB、EB-MTB和MDR中,结核杆菌PhoP和PhoR基因的表达有差异,结核杆菌PhoPR双组分系统与新疆地区广泛流行的耐药结核杆菌的耐药性具有相关性。
Objective To explore the correlation between Mycobacterium tuberculosis ( MTB ) PhoPR two-component system and drug resistance of MTB clinical isolates widespread in Xinjiang region by analyzing the expression of PhoP gene and PhoR gene among different isolates .Methods Total RNA of MTB was extracted from drug-susceptible strains , the strains only resistant to a single first-line anti-TB drugs (INH, RFP, SM and EB) and multidrug-resistant (MDR) strains, respectively.The purity of total RNA was checked by agarose gel electrophoresis .The expressions of PhoP gene and PhoR gene were quantified by using SYBR Green I qRT-PCR and the differences of their gene expression in different isolates were ana-lyzed.Results Compared with the drug-susceptible strains of MTB, the expression of PhoP gene was up-regulated for about 1.48 times in MTB strains resistant to RFP (RFP-MTB) and 2.74 times in MDR strain (P〈0.05).Compared with MDR strain, the expressions of PhoP gene in the isolates resistant to INH (IN-HMTB), RFP (RFP-MTB), SM (SM-MTB) and EB (EB-MTB) were down-regulated for 0.70, 0.50, 0.25 and 0.21 times respectively.The expressions of PhoR gene were down-regulated for 0.36, 0.54, 0.35 and 0.19 times, respectively (P〈0.05).The expressions of PhoR gene in the isolates of INH-MTB, RFP-MTB, SM-MTB and EB-MTB were up-regulated for 6.33, 4.56, 2.34, 1.85 and 9.06 times, respectively as compared with the drug-susceptible strains (P〈0.05).Conclusion Significant differences of PhoR gene and PhoP gene expressions were observed among drug-susceptible strains , INH-MTB, RFP-MTB, SM-MTB, EB-MTB and MDR strains.Therefore, the Mycobacterium tuberculosis PhoPR two-component system is asso-ciated with the drug resistance of MTB strains prevalent in Xinjiang region .