重庆市耐多药结核分枝杆菌gyrA与rrs基因突变特征分析

Characteristics of gyrA and rrs gene mutations in multidrug-resistant tuberculosis isolates in Chongqing,China

目的分析重庆市耐多药结核分枝杆菌gyrA、rrs基因突变特征及其与氧氟沙星(ofloxacin,Ofx)、卡那霉素耐药(kanamycin,Km)的关系。方法对89株耐多药结核分枝杆菌的Ofx耐药相关基因gyrA和Km耐药相关基因rrs进行序列测定,分析其基因突变特征。结果 89株MDR结核分枝杆菌中有50株对Ofx耐药,其中46株(92.00%,46/50)gyrA基因发生突变,突变位点包括74、89、90、91和94位密码子;22株耐Km菌株中,19株(86.36%,19/22)发生rrs基因突变,突变类型均为A1401G。Ofx、Km耐药株的gyrA、rrs基因突变率明显高于相应敏感株的基因突变率,两者之间的差异有统计学意义(χ2=62.937,P<0.001;Fisher双侧P<0.001)。结论 gyrA基因90、91、94位密码子突变是重庆市耐多药结核分枝杆菌Ofx耐药的主要机制;rrs基因A1401G突变则是Km耐药的主要原因。

We analysed the characteristics of gyrAgene mutation and rrs gene mutation of multidrug-resistant tuberculosis（MDR-TB）clinical isolates in Chongqing,and analysed the correlation between gene mutations and drug resistance to ofloxacin（Ofx）and kanamycin（Km）.The susceptibility of the MDR-TB clinical isolates to Ofx and Km was performed using the proportion method,and the gyrAand rrs gene mutations of MDR-TB strains were detected by DNA sequencing directly.Results showed that there were 50 strains resistant to Ofx among 89MDR-TB clinical isolates.Besides,the rest of the 39 strains which were sensitive to Ofx contained 3 gyrA mutation strains（7.69%,3/39）,and the mutation type was GAC94 GCC.While,the50 strains resistant to Ofx contained 46 gyrA mutation strains（92.00%,46/50）,and the mutation located at codon 74,89,90,91 and 94respectively.The highest mutation frequency was at codon 94,the rate was 58.00%（29/50）,followed by codon90（26.00%,13/50）.The mutation type of gyrAgene were mainly GAC94GGC（14strains）,GCG90GTG（13strains）and GAC94GCC（8strains）.There were 67 strains sensitve to Km among 89 MDR-TB clinical isolates and all of the sensitive strains had no rrs gene mutation.In addition,19 of 22Kmresistant isolates had an A-to-G transition at nucleotide position1 401 of rrs gene.Both the gyrAgene mutation rate of Ofx-resistant strains and the rrs gene mutation rate of Kmresistant strains were significantly higher than that of the corresponding sensitive strains（χ^2=62.937,P〈0.001;Fisher,P〈0.001）.In conclusion,mutations of gyrAcodons 90,91 and 94constitute the primary mechanism of Ofx resistanceamong MDR-TB clinical isolates in Chongqing,and an A1401 G mutation in the rrs gene is the main cause of Km resistance.Therefore,the gene mutation of gyrAand rrs could be used as the molecular biomarkers to detect the Ofx and Km resistance of MDR-TB clinical isolates in Chongqing.