Analysis on gene mutation characteristics of drug resistant Mycobacterium tuberculosis and its correlation with genotypes in Guangxi

doi:10.3969/j.issn.1000-6621.2021.06.013

Abstract

Abstract:

Objective To investigate the characteristics of Mycobacterium tuberculosis(MTB) gene mutation which are related with drug resistance to five first-line anti-tuberculosis drugs and their correlations with genotypes. Methods Whole-genome sequencing were performed on 721 MTB strains collected from drug resistance surveillance in Guangxi Zhuang Autonomous Region in 2017. Analyze the characteristics of drug resistance-related gene mutations and their correlation with genotypes. Results The total drug resistance-related gene mutation rate was 30.93% (223/721). The top 5 genes of mutation were katG (16.09%,116/721), rpoB (12.21%,88/721), embB (8.46%,61/721), rpsL (6.10%,44/721) and rrs (4.72%,34/721). The top 5 mutation loci were katG315 (12.34%,89/721), rpsL43 (5.27%,38/721), rpoB450 (4.85%,35/721), embB306 (4.02%,29/721) and rpoB445 (2.91%,21/721). RpsL, rpsL43 and rpoB445 mutation rate were higher in Beijing genotype strains than in non-Beijing genotype strains (10.51% (43/409) vs. 0.32% (1/312),χ²=32.090,P=0.000; 9.05% (37/409) vs. 0.32% (1/312),χ²=26.992,P=0.000; 4.40% (18/409) vs. 0.96% (3/312), χ²=7.404,P=0.007). Conclusion KatG,rpoB,embB,rpsL,rrs were the main drug resistance-related gene mutations in Guangxi MTB strains. The mutations in rpsL, rpsL43 and rpoB445 were correlated with Beijing genotype.

Key words: Mycobacterium tuberculosis, Drug resistance, Mutation, Genotype

LUO Dan, QIN Hui-fang, YE Jing, ZHAO Jin-ming, QIN Yi-xiang, LAN Ru-shu. Analysis on gene mutation characteristics of drug resistant Mycobacterium tuberculosis and its correlation with genotypes in Guangxi[J]. Chinese Journal of Antituberculosis, 2021, 43(6): 596-601. doi: 10.3969/j.issn.1000-6621.2021.06.013

Figures/Tables 5

References 25

[1]	王前, 李涛, 杜昕, 等. 2015—2019年全国肺结核报告发病情况分析. 中国防痨杂志, 2021,43(2):107-112. doi: 10.3969/j.issn.1000-6621.2021.02.002. doi: 10.3969/j.issn.1000-6621.2021.02.002
[2]	Luo D, Zhao J, Lin M, et al. Drug resistance in newly presenting and previously treated tuberculosis patients in Guangxi Province, PR China. Asia Pac J Public Health, 2017,29(4):296-303. doi: 10.1177/1010539517700474. doi: 10.1177/1010539517700474 URL
[3]	赵雁林, 逄宇. 结核病实验室检验规程. 北京: 人民卫生出版社, 2015: 32.
[4]	De Almeida IN, Da Silva Carvalho W,Rossetti ML,et al.Evaluation of six different DNA extraction methods for detection of Mycobacterium tuberculosis by means of PCR-IS6110: preliminary study. BMC Res Notes, 2013,6:561. doi: 10.1186/1756-0500-6-561. doi: 10.1186/1756-0500-6-561 URL
[5]	洪创跃, 杨婷婷, 李金莉, 等. 深圳市耐多药结核分枝杆菌耐药基因突变特征分析. 中国防痨杂志, 2020,42(6):583-589. doi: 10.3969/j.issn.1000-6621.2020.06.009. doi: 10.3969/j.issn.1000-6621.2020.06.009
[6]	Walker TM, Kohl TA, Omar SV, et al. Whole-genome sequencing for prediction of Mycobacterium tuberculosis drug susceptibility and resistance: a retrospective cohort study. Lancet Infect Dis, 2015,15(10):1193-1202. doi: 10.1016/S1473-3099(15)00062-6. doi: 10.1016/S1473-3099(15)00062-6 URL
[7]	Liu Q, Luo T, Dong X, et al. Genetic features of Mycobacterium tuberculosis modern Beijing sublineage. Emerg Microbes Infect, 2016,5(4):e14. doi: 10.1038/emi.2016.14. doi: 10.1038/emi.2016.14
[8]	Sun H, Zhang C, Xiang L, et al. Characterization of mutations in streptomycin-resistant Mycobacterium tuberculosis isolates in Sichuan,China and theassociation between Beijing-lineage and dual-mutation in gidB. Tuberculosis (Edinb), 2016,96:102-106. doi: 10.1016/j.tube.2015.09.004. doi: 10.1016/j.tube.2015.09.004 URL
[9]	Martínez LM, Castro GP, Guerrero MI. A molecular platform for the diagnosis of multidrug-resistant and pre-extensively drug-resistant tuberculosis based on single nucleotide polymorphism mutations present in Colombian isolates of Mycobacterium. Mem Inst Oswaldo Cruz, 2016,111(2):93-100. doi: 10.1590/0074-02760150306. doi: 10.1590/0074-02760150306 URL
[10]	Ali A, Hasan Z, McNerney R,et al.Whole genome sequencing based characterization of extensively drug-resistant mycobacterium tuberculosis isolates from Pakistan. PLoS One, 2015,10(2):e0117771. doi: 10.1371/journal.pone.0117771. doi: 10.1371/journal.pone.0117771 URL
[11]	董苏荣, 徐娟, 徐祖龙. 泰州地区结核分枝杆菌一线抗结核药耐药基因检测结果分析. 微生物与感染, 2017,17(6):60-62.doi: 10.3969/j.issn.1671-0142.2017.06.019. doi: 10.3969/j.issn.1671-0142.2017.06.019
[12]	Ocheretina O, Escuyer VE, Mabou MM, et al. Correlation between genotypic and phenotypic testing for resistance to rifampin in Mycobacterium tuberculosis clinical isolates in Haiti: investigation of cases with discrepant susceptibility results. PLoS One, 2014,9(3):e90569. doi: 10.1371/journal.pone.0090569. doi: 10.1371/journal.pone.0090569 URL
[13]	Thirumurugan R, Kathirvel M, Vallayyachari K, et al. Molecular analysis of rpoB gene mutations in rifampicin resistant Mycobacterium tuberculosis isolates by multiple allele specific polymerase chain reaction in Puducherry, South India. N Engl J Med, 2013,369(3):290-292. doi: 10.1016/j.jiph.2015.05.003. doi: 10.1016/j.jiph.2015.05.003 URL
[14]	Mekonnen D, Admassu A, Mulu W, et al. Multidrug-resistant and heteroresistant Mycobacterium tuberculosis and associated gene mutations in Ethiopia. Int J Infect Dis, 2015,39:34-38.doi: 10.1016/j.ijid.2015.06.013. doi: 10.1016/j.ijid.2015.06.013 URL
[15]	杨建东, 陈阳贵, 马丽, 等. 乌鲁木齐市耐多药结核分枝杆菌基因突变特征分析. 中华疾病控制杂志, 2017,21(1):22-25.doi: 10.16462/j.cnki.zhjbkz.2017.01.005. doi: 10.16462/j.cnki.zhjbkz.2017.01.005
[16]	Zhang Y, Heym B, Young D, et al. The catalase-perox-idase gene and isoniazid resistance of Mycobacterium tuberculosis. Nature, 1992,358(6387):591-593. pmid: 1501713
[17]	Nwofor AC, Nyamngee A, Nwabuisi C, et al. Performance of Genotype MTBDRplus in the Detection of Resistance to Rifampicin and Isoniazid Among Clinical Mycobacteria Isolates in Ilorin,Nigeria. Curr HIV Res, 2015,13(4):308-314. doi: 10.2174/1570162x13666150407143147. doi: 10.2174/1570162x13666150407143147 URL
[18]	王先化. 新疆地区TB合并HIV感染流行病学分析及耐药结核分子机制研究. 青岛:青岛大学, 2015.
[19]	赛依旦·吐尔逊, 刘金宝, 阿尔泰, 等. 2013年新疆436例不同民族住院结核病患者的耐药情况分析. 中华疾病控制杂志, 2015,19(7):675-678. doi: 10.16462/j.cnki.zhjbkz.2015.07.008. doi: 10.16462/j.cnki.zhjbkz.2015.07.008
[20]	Zhao LL, Sun Q, Liu HC, et al. Analysis of embCAB mutations associated with ethambutol resistance in multidrug-resistant mycobacterium tuberculosis isolates from China. Antimicrob Agents Chemother, 2015,59(4):2045-2050. doi: 10.1128/AAC.04933-14. doi: 10.1128/AAC.04933-14 URL
[21]	Xu Y, Jia H, Huang H, et al. Mutations found in embCAB,em-bR,and ubiA genes of ethambutol-sensitive and -resistant myco-bacterium tuberculosis clinical isolates from China. Biomed Res Int, 2015,2015:951706.
[22]	杨亚滨, 博晓真, 高飞. 抗结核药物耐药分子机制的研究进展. 内蒙古医学杂志, 2015,47(2):195-198. doi: 10.16096/J.cnki.nmgyxzz.2015.47.02.023. doi: 10.16096/J.cnki.nmgyxzz.2015.47.02.023
[23]	Villellas C, Aristimuno L, Vitoria MA, et al. Analysis of mutations in streptomycin-resistant strains reveals a simple and reliable genetic marker for identification of the Mycobacterium tuberculosis Beijing genotype. J Clin Microbiol, 2013,51(7):2124-2130. doi: 10.1128/JCM.01944-12. doi: 10.1128/JCM.01944-12 URL
[24]	陈高瞻, 芦莲, 雷航, 等. 结核分枝杆菌链霉素耐药临床分离株rrs基因突变与其耐药水平相关性研究. 武汉轻工大学学报, 2014,33(3):27-31.
[25]	Honore N, Marchal G, Cole ST, et al. Novel mutation in 16s rRNA associated with streptomycin dependence in Mycobacterium tuberculosis. Antimicrob Agents Chemother, 1995,39(3):769-770. doi: 10.1128/aac.39.3.769. doi: 10.1128/aac.39.3.769 URL

药品	单药突变		多药联合突变		合计 (株)
药品	菌株数	构成比(%)	菌株数	构成比(%)	合计 (株)
异烟肼	58	40.85	84	59.15	142
利福平	15	17.05	73	82.95	88
链霉素	33	44.00	42	56.00	75
吡嗪酰胺	9	33.33	18	66.67	27
乙胺丁醇	11	17.19	53	82.81	64

药品	耐药基因	突变菌株数	突变率(%)
异烟肼	ahpC	7	0.97
	fabG1	11	1.53
	katG	114	15.81
	kasA	4	0.55
	inhA+fabG1	3	0.42
	kasA+fabG1	1	0.14
	katG+fabG1	2	0.28
利福平	rpoB	88	12.21
乙胺丁醇	embA	2	0.28
	embB	31	4.30
	embC	1	0.14
	embB+embA	4	0.55
	embB+embB	8	1.11
	embC+embB	18	2.50
吡嗪酰胺	pnaD	1	0.14
	pncA	26	3.61
链霉素	rpsL	41	5.69
	rrs	31	4.30
	rpsL+rrs	3	0.42

耐药基因	北京基因型(409株)	非北京基因型(312株)	χ²值	P值
katG	65(15.89)	51(16.35)	0.270	0.870
rpoB	55(13.45)	33(10.58)	1.361	0.243
embB	29(7.09)	32(10.26)	2.291	0.130
rpsL	43(10.51)	1(0.32)	32.090	0.000
rrs	20(4.89)	14(4.49)	0.639	0.800

突变位点	北京基因型(409株)	非北京基因型(312株)	χ²值	P值
katG315	52(12.71)	37(11.86)	0.119	0.730
rpoB450	21(5.13)	14(4.49)	0.161	0.689
rpsL43	37(9.05)	1(0.32)	26.992	0.000
embB306	16(3.91)	13(4.17)	0.030	0.863
rpoB445	18(4.40)	3(0.96)	7.404	0.007