基于新疆维吾尔自治区南疆地区169株结核分枝杆菌探讨基因缺失对贝达喹啉的耐药机制  

Mechanism of gene deletion induced resistance to bedaquiline based on 169 strains of Mycobacterium tuberculosis in southern Xinjiang

作　　者：向煜 杨淑柳 武娅宁 王泉 曹滨 高晓平 于晋杰 肖开提·米吉提 赵秀芹[2] 李马超[2] 赵丽丽[2] 万康林[2] 刘海灿[2] 李桂莲[2] 袁秀琴[1] Xiang Yu;Yang Shuliu;Wu Yaning;Wang Quan;Cao Bin;Gao Xiaoping;Yu Jinjie;Xiaokaiti·mijiti;Zhao Xiuqin;Li Machao;Zhao Lili;Wan Kanglin;Liu Haican;Li Guilian;Yuan Xiuqin(School of Public Health,University of South China,Hengyang 421002,Hunan,China;National Key Laboratory of Intelligent Tracking and Forecasting for Communicable Diseases,National Institute for Communicable Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China;The Eighth Affiliated Hospital of Xinjiang Medical University,Urumqi 830000,Xinjiang,China)

机构地区：[1]南华大学公共卫生学院,湖南衡阳421001 [2]传染病溯源预警与智能决策全国重点实验室,中国疾病预防控制中心传染病预防控制所,北京102206 [3]新疆医科大学第八附属医院,新疆乌鲁木齐830017

出　　处：《疾病监测》2024年第5期639-646,共8页Disease Surveillance

基　　金：北京市自然科学基金面上项目(No.7242189);中国疾病预防控制中心传染病预防控制所青年基金项目(No.08064;0104327);新疆科技重大专项(No.2017A03006-3);“十三五”国家科技重大专项(No.2018ZX10103001-003-012)。

摘　　要：目的挖掘与结核分枝杆菌(MTB)对贝达喹啉耐药相关的基因缺失片段。方法2017—2020年期间从新疆维吾尔自治区南疆地区收集807株MTB分离株,对菌株进行全基因组测序,采用TB-Profiler 4.1.2检测菌株的家系和预测异烟肼、利福平、氟喹诺酮类等7种药物的敏感性;选择44株准广泛耐药(Pre-XDR)菌株和125株随机菌株,采用Delly 0.9.1软件检测基因缺失情况,采用C++脚本程序检测Rv0678、atpE和pepQ基因突变情况,采用微量肉汤稀释微孔板法检测菌株对贝达喹啉的最小抑菌浓度(MIC)及耐药情况。采用χ^(2)检验和秩和检验等方法分析基因缺失与贝达喹啉耐药的相关性及不同家系菌株对贝达喹啉耐药性及敏感程度的差异。结果44株Pre-XDR株中共鉴定出23株贝达喹啉耐药株(MIC≥0.25μg/mL)和49种基因缺失类型,发生缺失菌株数>5株的缺失类型共有8种,其中2729618 bp~2729833 bp(Rv2434c)缺失组贝达喹啉耐药率和MIC水平低于未缺失组(χ^(2)=6.145,P=0.013和Z=−2.224,P=0.026);2235169 bp~2235233 bp(ctpG)、1710766 bp~1711557 bp(Rv1519-Rv1520)和3952924 bp~3954223 bp(echA19-Rv3517)多基因缺失组贝达喹啉耐药率和MIC水平均高于未缺失组(χ^(2)=4.322,P=0.038和Z=−2.526,P=0.012)。但是在125株验证用菌株(含贝达喹啉耐药株23株)中均未发现差异具有统计学意义(χ^(2)=1.660,P=0.198和Z=−1.152,P=0.249)。将44株菌株和125株菌株合并分析,仅发现2235169 bp~2235233 bp(ctpG)、1710766 bp~1711557 bp(Rv1519-Rv1520)和3952924 bp~3954223 bp(echA19-Rv3517)多基因缺失组贝达喹啉耐药率高于未缺失组(χ^(2)=6.396,P=0.011),该多基因缺失仅发生于L3家系菌株,同时两两比较发现L3家系菌株的贝达喹啉耐药率显著高于L4家系(P<0.017)。结论2235169 bp~2235233 bp(ctpG)、1710766 bp~1711557 bp(Rv1519-Rv1520)和3952924 bp~3954223 bp(echA19-Rv3517)多基因缺失为南疆地区L3家系菌株特有的基因缺失类型(与该地其他家系相比),其与贝�Objective To understand gene deletion associated bedaquiline resistance in Mycobacterium tuberculosis.Methods During 2017−2020,a total of 807 strains of M.tuberculosis were collected from the southern area of Xinjiang Uygur autonomous region for whole genome sequencing,then TB-Profiler was used to detect the strain lineages and predict the strains’susceptibilities to 7 antibiotics,including isoniazid,rifampicin,fluoroquinolone,and so on.In the 807 isolates,44 preextensively drug resistant(Pre-XDR)strains were selected and 125 isolates were randomly selected for the tests of gene deletion by using Delly 0.9.1 software,the mutations in Rv0678,atpE and pepQ by using C++script program,and the minimum inhibitory concentrations(MICs)of bedaquiline to the strains by using the micro-broth dilution microplate method.The association between gene deletion and bedaquiline resistance and the difference in bedaquiline resistance rates and MIC levels among the strains in different lineages were analyzed byχ^(2) test and rank-sum test.Results A total of 49 gene deletion types,including 8 types involving>5 strains,and 23 bedaquiline resistant strains were identified in 44 Pre-XDR strains.In the 44 Pre-XDR isolates,the bedaquiline resistance rate and MIC level of the deletion group of 2729618 bp–2729833 bp(Rv2434c)were lower than those of the non-deletion group(χ^(2)=6.145,P=0.013 and Z=−2.224,P=0.026);multi-gene deletion group of 2235169 bp–2235233 bp(ctpG),1710766 bp–1711557 bp(Rv1519–Rv1520)and 3952924 bp–3954223 bp(echA19–Rv3517)showed higher bedaquiline resistance rate and MIC level compared with non-deletion group(χ^(2)=4.322,P=0.038 and Z=−2.526,P=0.012).However,no significant difference was found among the 125 strains(including 23 bedaquiline resistant strains)used for validation(χ^(2)=1.660,P=0.198 and Z=−1.152,P=0.249).Combined analysis on the 44 strains and 125 strains revealed that only the multi-gene deletion group of 2235169 bp–2235233 bp(ctpG),1710766 bp–1711557 bp(Rv1519–Rv1520)and 3

关 键 词：结核分枝杆菌 耐药 基因缺失 贝达喹啉 L3家系 

分 类 号：R211[医药卫生—中医学] R521