肺结核患者结核分枝杆菌利福平基因突变特征及耐药情况

Mutation characteristics and drug resistance of rifampicin gene of Mycobacterium tuberculosis in patients with pulmonary tuberculosis

目的探讨肺结核患者当中的结核分枝杆菌利福平基因发生突变的特征,同时分析其耐药情况,为耐药结核病的预防和治疗提供科学依据。方法选取2016年6月—2018年12月杭州师范大学附属医院收治的肺结核患者532例,依据其痰液中的结核分枝杆菌耐药性分为耐药株(218株)及敏感株(314株)。测定2组菌株的rpoB基因序列及最低抑菌浓度(MIC)值水平。结果218株耐药菌株中180株为单重突变耐药菌株,rpoB基因结构分析发现,发生比率最高的突变密码子分别为526位(13.2%)、531位(74.4%)。206株(94.5%)耐药菌株出现至少1个突变位点,且均位于耐利福平决定区(RRDR),12株(5.5%)耐药菌株突变位点在RRDR之外。有4株耐药菌株在RRDR之内发生同义突变,4株耐药菌株在RRDR之外发生同义突变。H37Rv密码子未发生任何突变。有8株敏感菌株发生同义突变在RRDR之外。单重突变菌株的RIF平均MIC值水平为(47.34±2.04)μg/mL,显著低于多种突变菌株平均MIC值水平[(118.24±3.95)μg/mL,t=107.636,P<0.001]。526单密码子平均MIC值水平与531单密码子突变菌株的MIC值水平比较差异无统计学意义(P>0.05)。多重密码子突变菌株的MIC值水平为(116.03±5.06)μg/mL高于单重密码子平均MIC值水平[(47.08±1.31)μg/mL,t=85.530,P<0.001]。结论利福平耐药性机制可能与rpoB基因的起始端531位、526位等突变相关,rpoB基因序列可用于预测结核分枝杆菌中的利福平耐药情况及表型。

Objective To explore the mutation characteristics of rifampin gene of Mycobacterium tuberculosis in patients with pulmonary tuberculosis,and analyze the drug resistance,so as to provide scientific basis for the prevention and treatment of drug-resistant tuberculosis.Methods Total 532 patients with pulmonary tuberculosis admitted to the Affiliated Hospital of Hangzhou Normal University from June 2016 to December 2018 were divided into drug-resistant strains(n=218)and sensitive(n=314)strains according to the drug resistance of Mycobacterium tuberculosis in sputum.The rpoB gene sequence and minimal inhibitory concentration(MIC)value were determined.Results Among the 218 resistant strains,180 were single mutation resistant strains.The mutation codon with the highest rate of rpoB gene structure analysis was 526(13.2%)and 531(74.4%).Total 206(94.5%)resistant strains showed at least one mutation site,and all of them were located in the rifampicin resistant decision region(RRDR),and 12(5.5%)resistant strains were outside RRDR.Four resistant strains had synonymous mutation in RRDR and 4 resistant strains had synonymous mutation outside RRDR.There was no mutation in H37 Rv codon.Eight sensitive strains had synonymous mutations outside RRDR.The RIF MIC level of the single-mutated strain was(47.34±2.04)μg/mL,which was significantly lower than that of the mutant strain(118.24±3.95)μg/mL(t=107.636,P<0.001).There was no significant difference between the MIC level of single codon 526 and single codon 531 mutant strains(P>0.05).The MIC level of the multicodon mutant was(116.03±5.06)μg/mL higher than that of the single codon(47.08±1.31)μg/mL(t=85.530,P<0.001).Conclusion The mechanism of rifampicin resistance may be related to the mutations at 531 and 526 of the start of rpoB gene.The rpoB gene sequence can be used to predict the resistance and phenotype of rifampicin in Mycobacterium tuberculosis.