宁波地区耐多药结核分枝杆菌异烟肼与丙硫异烟胺交叉耐药基因分子特征及影响因素分析

doi:10.19982/j.issn.1000-6621.20250414

中国防痨杂志 ›› 2026, Vol. 48 ›› Issue (5): 576-585.doi: 10.19982/j.issn.1000-6621.20250414

宁波地区耐多药结核分枝杆菌异烟肼与丙硫异烟胺交叉耐药基因分子特征及影响因素分析

车洋¹, 张栋梁¹, 李相辰², 陆烨玮², 柳正卫³()

¹宁波市疾病预防控制中心结核病防制科, 宁波 315010
²杭州广科安德生物科技有限公司精准医学诊断与监测研究重点实验室, 杭州 310020
³浙江省疾病预防控制中心结核病防制所, 杭州 310051

收稿日期:2025-10-25 出版日期:2026-05-10 发布日期:2026-04-27
通信作者: 柳正卫 E-mail:zhwliu@cdc.zj.cn
基金资助:
宁波市公益性研究计划项目(2024S040);宁波市医疗卫生高端团队重大攻坚项目(2023020713);浙江省基础公益研究计划项目(LTGY23H190001)

Molecular characterization and risk factors of cross-resistance between isoniazid and prothionamide in multidrug-resistant Mycobacterium tuberculosis in Ningbo, China

Che Yang¹, Zhang Dongliang¹, Li Xiangchen², Lu Yewei², Liu Zhengwei³()

¹Institute of Tuberculosis Prevention and Control, Ningbo Municipal Center for Disease Control and Prevention, Zhejiang Province, Ningbo 315010, China
²Key Laboratory of Precision Medicine in Diagnosis and Monitoring Research, Cosmos Wisdom Biotechnology Co., Ltd, Hangzhou 310020, China
³The Institute of TB Control, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou 310051, China

Received:2025-10-25 Online:2026-05-10 Published:2026-04-27
Contact: Liu Zhengwei E-mail:zhwliu@cdc.zj.cn
Supported by:
Ningbo Public Welfare Science and Technology Program Project(2024S040);Ningbo Top Medical and Health Research Program(2023020713);Zhejiang Provincial Natural Science Foundation of China under Grant(LTGY23H190001)

摘要/Abstract

摘要：

目的: 基于全基因组测序(whole genome sequencing,WGS)技术分析宁波地区耐多药结核分枝杆菌(multidrug-resistant Mycobacterium tuberculosis,MDR-MTB)中异烟肼(isoniazid,INH)和丙硫异烟胺(prothionamide,Pto)的交叉耐药现况、耐药相关基因分子突变特征及交叉耐药影响因素。方法: 纳入宁波地区2021—2024年耐药监测期间134株具有左氧氟沙星(levofloxacin,Lfx)、贝达喹啉(bedaquiline,Bdq)、利奈唑胺(linezolid,Lzd)、德拉马尼(delamanid,Dlm)、氯法齐明(clofazimine,Cfz)、阿米卡星(amikacin,Am)、卷曲霉素(capreomycin,Cm)、对氨基水杨酸(para-aminosalicylic acid,PAS)及Pto表型药物敏感性试验(简称“表型药敏试验”)结果的MDR-MTB临床分离株作为研究对象,提取总DNA后进行WGS。应用TB-profiler耐药基因数据库确定INH和Pto耐药基因突变特征,同时结合多种抗结核药物的表型药敏试验结果及患者临床信息综合分析本地区MDR-MTB临床分离株INH和Pto交叉耐药现况及交叉耐药影响因素。结果: 纳入研究的134株MDR-MTB中129株发生INH耐药相关基因突变,占96.3%(129/134)。INH耐药相关基因katG、inhA和ahpC的突变均存在,各基因突变类型分别以katG Ser315Thr(89.1%,98/110)、inhA c-15t(50.0%,17/34)和ahpC c-81t (4/9)为主。134株MDR-MTB对Pto的总耐药率(INH和Pto的交叉耐药率)为20.2%(27/134),27株Pto耐药菌株中25株发生耐药相关基因突变,占92.6%(25/27)。Pto耐药相关的基因突变有ethA和inhA,突变占比分别为63.0%(17/27)和77.8%(21/27)。突变类型分别以ethA基因11_12insA(70.6%,12/17)和inhA基因 t-8c(66.7%,14/21)为主。纳入研究的MDR-MTB和准广泛耐药MTB(pre-XDR-MTB)中INH和Pto的交叉耐药率分别为20.2%(27/134)和43.6%(24/55)。多因素logistic回归分析结果显示,使用莫西沙星[OR(95%CI):4.123(1.207~14.080)]和耐Lfx[OR(95%CI):0.075(0.019~0.293)]是MDR-MTB中INH和Pto交叉耐药的独立影响因素。结论: 宁波地区MDR-MTB中INH和Pto交叉耐药现况较为严峻,耐药相关基因inhA突变是MDR-MTB中INH和Pto交叉耐药的主要分子机制,需进一步加强治疗前氟喹诺酮类药物的耐药检测并规范合理用药。

关键词: 分枝杆菌, 结核, 抗药性, 多种, 细菌, 丙硫异烟胺, 突变, 基因

Abstract:

Objective: This study aimed to investigate the characterization, the drug resistance-associated gene mutations in multidrug-resistant Mycobacterium tuberculosis (MDR-MTB) strains based on whole-genome sequencing (WGS) and to analyze the risk factors of cross-resistance between isoniazid (INH) and prothionamide (Pto). Methods: A total of 134 MDR-MTB isolates selected in the drug resistance survey in Ningbo between 2021 and 2024 were enrolled in the study. WGS was conducted to detect the genomic DNA extracted from clinical isolates. The sequencing data were analyzed using an in-house pipeline and the gene mutations determined based on MTB drug resistance gene database. The conventional drug susceptibility test was also used to detect the drug-resistant profiles against 9 antibiotics (levofloxacin, bedaquiline, linezolid, delamanid, clofazimine, amikacin, capreomycin, para-aminosalicylic acid and Pto). Combined with phenotypic drug susceptibility testing results and clinical data, the cross-resistance profile and risk factors between INH and Pto among MDR-MTB isolates were comprehensively analyzed. Results: Among 134 MDR-MTB isolates, 129 (96.3%, 129/134) carried INH resistance-associated mutations. Mutations were present in all three INH resistance-associated genes, katG, inhA and ahpC. The main mutation types were katG Ser315Thr (89.1%, 98/110), inhA c-15t (50.0%, 17/34) and ahpC c-81t (4/9). Among 134 MDR-MTB isolates, the overall Pto resistance rate (INH-Pto cross-resistance rate) was 20.2% (27/134), with 92.6% (25/27) harboring mutations. The gene mutations associated with Pto resistance were ethA and inhA, with mutation rates of 63.0% (17/27) and 77.8% (21/27), respectively. The major mutation types were ethA 11_12insA (70.6%, 12/17) and inhA t-8C (66.7%, 14/21). Among MDR-MTB and Pre-XDR-MTB, the INH-Pto cross-resistance rates were 20.2% (27/134) and 43.6% (24/55). Multivariate logistic regression showed that moxifloxacin given (OR (95%CI): 4.123 (1.207-14.080)) and the resistance of levofloxacin (OR (95%CI): 0.075 (0.019-0.293)) were risk factors for INH-Pto cross-resistance. Conclusion: The prevalence of INH-Pto cross-resistance among MDR-MTB was serious in Ningbo and the mutation type inhA t-8C was closely with INH-Pto cross-resistance. The regimen for INH-Pto cross-resistant patients should be according to the drug susceptibility results of fluoroquinolones and choose optimal treatment.

Key words: Mycobacterium tuberculosis, Drug resistance, multiple, bacterial, Prothionamide, Mutation, Genes

中图分类号:

车洋, 张栋梁, 李相辰, 陆烨玮, 柳正卫. 宁波地区耐多药结核分枝杆菌异烟肼与丙硫异烟胺交叉耐药基因分子特征及影响因素分析[J]. 中国防痨杂志, 2026, 48(5): 576-585. doi: 10.19982/j.issn.1000-6621.20250414

Che Yang, Zhang Dongliang, Li Xiangchen, Lu Yewei, Liu Zhengwei. Molecular characterization and risk factors of cross-resistance between isoniazid and prothionamide in multidrug-resistant Mycobacterium tuberculosis in Ningbo, China[J]. Chinese Journal of Antituberculosis, 2026, 48(5): 576-585. doi: 10.19982/j.issn.1000-6621.20250414

图/表 5

表1

表2

表3

表4

耐多药结核病患者中INH和Pto交叉耐药组和非交叉耐药组的特征比较 [例(构成比,%)]

特征		非交叉耐药组(107例)			交叉耐药组(27例)			χ²值			P值
性别								5.048			0.025
男性		77(72.0)			25(92.6)
女性		30(28.0)			2(7.4)
年龄组(岁)								3.679			0.159
15~29		21(19.6)			10(37.0)
30~59		60(56.1)			12(44.4)
≥60		26(24.3)			5(18.6)
户籍								0.091			0.763
本地		52(48.6)			14(51.9)
非本地		55(51.4)			13(48.1)
吸烟								0.518			0.472
是		21(19.6)			7(25.9)
否		86(80.4)			20(74.1)
饮酒									0.790			0.374
是			14(13.1)			6(22.2)
否			93(86.9)			21(77.8)
合并糖尿病									0.036			0.850
是			22(20.6)			6(22.2)
否			85(79.4)			21(77.8)
合并高血压									0.076			0.783
是			14(13.1)			3(11.1)
否			93(86.9)			24(88.9)
合并慢性阻塞性肺疾病									0.313			0.576
是			3(2.8)			2(7.4)
否			104(97.2)			25(92.6)
结核病治疗史									3.452			0.063
有			46(43.0)			17(63.0)
无			61(57.0)			10(37.0)
空洞									1.570			0.210
有			57(53.3)			18(66.7)
无			50(46.7)			9(33.3)
涂片镜检									0.223			0.636
阳性			53(49.5)			12(44.4)
阴性			54(50.5)			15(55.6)
诊断时生化指标
A/G									3.622			0.057
正常			69(64.5)			12(44.4)
异常			38(35.5)			15(55.6)
AST/ALT									0.017			0.895
正常			50(46.7)			13(48.1)
异常			57(53.3)			14(51.9)
CRP水平									0.831			0.362
正常			61(57.0)			18(66.7)
异常			46(43.0)			9(33.3)
CRE水平									2.720			0.099
正常			65(60.7)			21(77.8)
异常			42(39.3)			6(22.2)
BUN水平									1.269			0.260
正常			85(79.4)			24(88.9)
异常			22(20.6)			3(11.1)
TC水平									1.014			0.314
正常			100(93.5)			23(85.2)
异常			7(6.5)			4(14.8)
TG水平								0.158			0.691
正常		100(93.5)			24(88.9)
异常		7(6.5)			3(11.1)
用药情况
使用Bdq								0.347			0.556
是		34(31.8)			7(25.9)
否		73(68.2)			20(74.1)
使用Lzd								0.019			0.892
是		61(57.0)			15(55.6)
否		46(43.0)			12(44.4)
使用Mfx								4.497			0.034
是		52(48.6)			7(25.9)
否		55(51.4)			20(74.1)
使用Lfx								0.016			0.900
是		25(23.4)			6(22.2)
否		82(76.6)			21(77.8)
使用Cfz								0.094			0.759
是		48(44.9)			13(48.1)
否		59(55.1)			14(51.9)
使用Cm								0.008			0.931
是		5(4.7)			2(7.4)
否		102(95.3)			25(92.6)
使用Cs								0.005			0.946
是		76(71.0)			19(70.4)
否		31(29.0)			8(29.6)
使用PAS								5.343			0.021
是		4(3.7)			5(18.5)
否		103(96.3)			22(81.5)
使用Pto								0.078			0.780
是		25(23.4)			7(25.9)
否		82(76.6)			20(74.1)
使用PZA								0.461			0.497
是		72(67.3)			20(74.1)
否		35(32.7)			7(25.9)
菌株基因型								2.969			0.085
Lineage 2		92(86.0)			27(100.0)
Lineage 4		15(14.0)			0(0.0)
菌株耐药谱
耐Lfx								31.986			<0.010
是		31(29.0)			24(88.9)
否		76(71.0)			3(11.1)
耐Am							17.184			<0.010
是	3(2.8)			8(29.6)
否	104(97.2)			19(70.4)
耐Cm							5.343			0.021
是	4(3.7)			5(18.5)
否	103(96.3)			22(81.5)
耐PAS							0.420			0.517
是	20(18.7)			3(11.1)
否	87(81.3)			24(88.9)

表4

表5

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因素	β值	s${}_{\overline{x}}$值	Wald χ²值	P值	OR(95%CI)值
性别	-1.389	0.889	2.439	0.118	0.249(0.044~1.425)
使用Mfx	1.416	0.627	5.109	0.024	4.123(1.207~14.080)
使用PAS	-0.694	1.002	0.480	0.489	0.499(0.070~3.562)
耐Lfx	-2.588	0.693	13.929	<0.010	0.075(0.019~0.293)
耐Am	-2.420	1.430	2.862	0.091	0.089(0.005~1.467)
耐Cm	0.249	1.669	0.022	0.882	1.282(0.049~33.799)

宁波地区耐多药结核分枝杆菌异烟肼与丙硫异烟胺交叉耐药基因分子特征及影响因素分析

Molecular characterization and risk factors of cross-resistance between isoniazid and prothionamide in multidrug-resistant Mycobacterium tuberculosis in Ningbo, China

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药敏结果	耐药基因	突变类型	菌株数	构成比(%)
INH耐药且Pto敏感(107株)	katG	Val1Ala	1	0.9
		Trp91Arg	1	0.9
		Asn138Ser	1	0.9
		Ala172Thr	1	0.9
		Trp191Arg	1	0.9
		Ser315Asn	1	0.9
		Ser315Gly	2	1.9
		Ser315Thr	79	73.8
		Thr380Ile	1	0.9
		Gly629Ser	1	0.9
	ahpC	c-81t	3	2.8
		c-57t	3	2.8
		c-52t	1	0.9
	ethA	-2170_1075del	1	0.9
		11_12insA	2	1.9
		61delA	1	0.9
		403_424del	1	0.9
		752insG	1	0.9
		851delC	1	0.9
	inhA	g-154a	1	0.9
		c-15t	10	9.3
		t-8c	3	2.8
		Ile21Val	1	0.9
		Ile194Thr	2	1.9
INH和Pto均耐药(27株)	katG	Ser315Asn	1	3.7
		Ser315Thr	19	70.4
		Asp419His	1	3.7
	ahpC	c-81t	1	3.7
	ethA	11_12insA	12	44.4
		13_14insC	3	11.1
		Tyr84Asp	1	3.7
		1112_1113insG	1	3.7
	inhA	t-8c	14	51.9
		c-15t	7	25.9
		Ser94Ala	3	11.1

药敏结果	耐药基因	突变类型	菌株数	构成比(%)
INH耐药且Pto敏感(31株)	katG	Ala172Thr	1	3.2
		Trp191Arg	1	3.2
		Ser315Gly	1	3.2
		Ser315Thr	19	61.3
		Thr380Ile	1	3.2
	ahpC	c-81t	1	3.2
	ethA	11_12insA	1	3.2
		752insG	1	3.2
		851delC	1	3.2
INH和Pto均耐药 (24株)	inhA	g-154a	1	4.2
		c-15t	5	20.8
		t-8c	2	8.3
		Ile194Thr	2	8.3
	katG	Ser315Thr	18	75.0
		Ser315Asn	1	4.2
	inhA	c-15t	5	20.8
		t-8c	14	58.3

突变类型	菌株数	L2基因型		L4基因型
突变类型	菌株数	菌株数	构成比(%)	菌株数	构成比(%)
katG Ser315Thr	98	87	88.8	11	11.2
inhA c-15t	17	16	94.1	1	5.9
inhA t-8c	17	17	100.0	0	0.0
ahpC c-81t	4	4	4/4	0	0.0
ethA 11_12insA	14	14	100.0	0	0.0

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