新疆维吾尔自治区南疆地区结核分枝杆菌ESX-4分泌系统基因突变特征分析

杨淑柳; 向煜; 王泉; 武娅宁; 肖开提·米吉提; 王瑞欢; 高晓平; 赵秀芹; 李马超; 王冕; 万康林; 李桂莲; 刘海灿; 袁秀琴

doi:10.3784/jbjc.202311300656

新疆维吾尔自治区南疆地区结核分枝杆菌ESX-4分泌系统基因突变特征分析

Analysis on mutations in the genes of ESX-4 secretion system of Mycobacterium tuberculosis isolated from Southern Xinjiang

摘要

摘要:
目的比较不同家系及耐药性结核分枝杆菌临床分离株ESX-4分泌系统7个毒力基因突变的差异，探讨临床菌株中 ESX-4基因的多态性特征。
方法纳入新疆南疆地区807株结核分枝杆菌临床株，进行全基因组测序。利用TB-profiler软件及C++脚本程序检测不同临床菌株ESX-4基因的突变、家系和药物敏感性结果。对数据进行描述性分析、χ²检验及logistics回归分析。
结果在807株结核分枝杆菌中共发现119个单核苷酸多态性位点和12种不同的插入/缺失。非同义突变率排在前3位的基因是Rv3446c（43.12%）、esxU（40.02%）以及eccC4和eccD4基因（均为23.30%）。家系中Lineage 2（L2）占62.70%（506/807），L3占19.46%（157/807），L4占17.84%（144/807）；敏感株占39.41%（318/807），单耐药株占20.94%（169/807），多耐药株占18.59%（150/807），耐多药株占21.07%（170/807）；利福平耐药占34.08%（275/807），异烟肼耐药占28.50%（230/807），喹诺酮类耐药占12.14%（98/807）。 L3家系中eccC4和esxT基因的突变率均显著高于L2或L4家系。多因素logistics回归模型分析发现：L2家系和异烟肼耐药是esxU和Rv3446c基因发生突变的危险因素［均比值比（OR）>1，均P<0.001］；多耐药则是Rv3446c突变的又一危险因素［OR=1.94，95%置信区间（CI）：1.12～3.37］。
结论本研究发现了结核分枝杆菌ESX-4分泌系统中与L2、L3家系及耐药性相关的基因突变，为进一步研究结核分枝杆菌的传播、耐药和致病机制提供基础科学依据。

Abstract:
Objectives To compare the mutation differences of the virulence genes of ESX-4 system in clinical isolates of Mycobacterium tuberculosis in different lineages and with different drug resistance, and explore of ESX-4 system gene polymorphisms in clinical isolates.
Methods Whole-genome sequencing (WGS) was performed on 807 M. tuberculosis clinical isolates in southern Xinjiang. The mutation profiles of virulence genes of ESX-4 system as well as the lineages and drug susceptibility results were detected using TB-profiler software. Descriptive analysis method, and χ² test and logistics regression model were performed on the data and C++ Script programme.
Results A total of 119 single nucleotide polymorphic loci and 12 different insertions/deletions were found in 807 M. tuberculosis strains. The genes with top three non-synonymous mutation rates were Rv3446c (43.12%), esxU (40.02%), and eccC4 and eccD4 genes (both 23.30%). Lineage 2 strains accounted for 62.70% (506/807), lineage 3 strains accounted for 19.45% (157/807) and lineage 4 strains accounted for 17.84% (144/807). In addition, the sensitive strains accounted for 39.41% (318/807), mono-drug resistant strains accounted for 20.94% (169/807), poly-drug resistant strains accounted for 18.59% (150/807), multi-drug resistant (MDR) strains accounted for 21.07% (170/807); and the rifampicin resistant rate was 34.08% (275/807), isoniazid resistant rate was 28.50% (230/807), quinolones resistant rate was 12.14% (98/807). The mutation rates of both eccC4 and esxT genes were significantly higher in the lineage 3 strains than in lineage 2 or lineage 4 strains. Multivariate logistic regression model analysis revealed that lineage 2 and isoniazid resistance were risk factors for mutations in esxU and Rv3446c genes (all OR>1, all P<0.001), and MDR was an additional risk factor for Rv3446c mutation (OR=1.94, 95%CI: 1.12–3.37).
Conclusion Our results identified gene mutations associated with lineage 2 and 3 isolates and drug resistance in the ESX-4 secretion system of M. tuberculosis, which provided basic scientific evidence for further research of the transmission, drug resistance and pathogenic mechanism of M. tuberculosis.

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