Study on the diagnostic value of nanopore sequencing technology for non-tuberculous mycobacterial pulmonary disease

doi:10.19983/j.issn.2096-8493.20250024

Abstract

Abstract:

Objective: To explore the diagnostic value of nanopore sequencing technology for detecting non-tuberculous mycobacterial pulmonary disease (NTM-PD) in bronchoalveolar lavage fluid (BALF). Methods: A total of 155 patients with suspected NTM-PD admitted to the Tuberculosis Department of Nanning Fourth People’s Hospital from January 1, 2022, to December 31, 2024, were enrolled. BALF samples collected via bronchoscopy were subjected to mycobacterial culture and identification, fluorescence PCR melting curve method, and nanopore sequencing. The diagnostic efficiencies of these three methods were compared. Results: Among the patients, 96 were diagnosed with NTM-PD (NTM-PD group) and 59 with non-NTM pulmonary diseases (including 29 cases of pulmonary tuberculosis and 30 cases of other pneumonias). The sensitivity, diagnostic concordance rate, and Kappa value of nanopore sequencing for NTM-PD diagnosis were 92.71% (95%CI: 85.98%-96.75%), 86.45% (95%CI: 79.81%-91.56%), and 0.706 (95%CI: 0.586-0.808),The sensitivity of nanopore sequencing was significantly higher than that of mycobacterium identification by fluorescence PCR melting curve method (63.54%,95%CI:53.18%-73.09%;71.61%,95%CI:63.91%-78.49%;0.445,95%CI:0.350-0.618), mycobacterium culture and identification (63.54%,95%CI:53.18%-73.09%;74.84%,95%CI:67.35%-81.44%;0.515,95%CI:0.453-0.703), and the combination of the latter two methods (84.38%,95%CI:75.53%-90.94%;82.58%,95%CI:75.67%-88.23%;0.634,95%CI:0.523-0.761). However,the specificity of nanopore sequencing was 76.27%(95%CI:63.60%-86.37%), which was lower than that of mycobacterium identification by fluorescence PCR melting curve method (84.75%,95%CI:72.90%-92.71%), mycobacterium culture and identification (93.22%,95%CI:83.54%-98.01%), and the combination of the latter two methods (79.66%,95%CI:67.34%-89.14%). Conclusion: Nanopore sequencing technology has high diagnostic efficiency for NTM-PD and can directly identify bacterial species, but specificity issues should be considered in clinical application.

Key words: Nanopore sequencing, Atypical bacteria forms, Mycobacterium, Bronchoalveolar lavage fluid

CLC Number:

R517.9

Huang Gaoyan, Su Shenxiao, Zhao Juanjuan, Liang Baoyuan, Meng Xiayan, He Huawei. Study on the diagnostic value of nanopore sequencing technology for non-tuberculous mycobacterial pulmonary disease[J]. Journal of Tuberculosis and Lung Disease , 2025, 6(4): 413-419. doi: 10.19983/j.issn.2096-8493.20250024

Figures/Tables 3

特征	NTM肺病组(96例)	非NTM肺病组(59例)	统计检验值	P值
性别[例,构成比(%)]			χ²=1.310^a	0.253
男性	35(36.46)	27(45.76)
女性	61(63.54)	32(54.24)
年龄(岁, $x ˉ$ ±s)	59.37±12.04	58.82±12.95	t=-0.268	0.789
吸烟史[例,率(%)]	6(6.25)	2(3.39)	χ²=0.607^a	0.436
既往病史[例,率(%)]
高血压	9(9.38)	7(11.86)	χ²=0.243^a	0.622
糖尿病	4(4.17)	4(6.78)	χ²=0.506^a	0.477
慢性肾病	10(10.42)	6(10.17)	χ²=0.0137^a	0.907
艾滋病	2(2.08)	1(1.69)	χ²=0.0289^a	0.865
支气管扩张症	58(60.42)	19(32.20)	χ²=11.560^a	0.001

References 14

[1]	中华医学会结核病学分会,《中华结核和呼吸杂志》编辑委员会. 非结核分枝杆菌病诊断与治疗专家共识. 中华结核和呼吸杂志, 2020, 43(11):918-946. doi:10.3760/cma.j.cn112147-20200508-00570.
[2]	Kumar K, Ponnuswamy A, Capstick TG, et al. Non-tuberculous mycobacterial pulmonary disease (NTM-PD):Epidemiology,diagnosis and multidisciplinary management. Clin Med (Lond), 2024, 24(1):100017. doi:10.1016/j.clinme.2024.100017.
[3]	黄夏宁, 覃鑫, 周洁, 等. 基于纳米孔测序的病原基因组富集技术研究进展与应用前景. 中国口岸科学技术, 2024, 6(z2):28-34. doi:10.3969/j.issn.1002-4689.2024.z2.004.
[4]	张威, 全俊. 非结核分枝杆菌肺病治疗现状及新型治疗药物. 中国感染与化疗杂志, 2022, 22(1): 119-124. doi:10.16718/j.1009-7708.2022.01.024.
[5]	Zheng M, Chen X, Chen Q, et al. Employing Multicolor Melting Curve Analysis to Rapidly Identify Non-Tuberculous Mycobacteria in Patients with Bronchiectasis:A Study from a Pulmonary Hospital in the Fuzhou District of China, 2018—2022. Crit Rev Immunol, 2024, 44(4):41-49. doi:10.1615/CritRevImmunol.2024052213.
[6]	Tan Y, Deng Y, Yan X, et al. Nontuberculous mycobacterial pulmonary disease and associated risk factors in China:A prospective surveillance study. J Infect, 2021, 83(1):46-53. doi:10.1016/j.jinf.2021.05.019.
[7]	Kumar K, Loebinger MR. Nontuberculous Mycobacterial Pulmonary Disease: Clinical Epidemiologic Features, Risk Factors, and Diagnosis: The Nontuberculous Mycobacterial Series. Chest, 2022, 161(3):637-646. doi:10.1016/j.chest.2021.10.003.
[8]	Velagapudi M, Sanley MJ, Ased S, et al. Pharmacotherapy for nontuberculous mycobacterial pulmonary disease. Am J Health Syst Pharm, 2022, 79(6):437-445. doi:10.1093/ajhp/zxab422.
[9]	Nogueira LB, Garcia CN, Costa MSCD, et al. Non-tuberculous cutaneous mycobacterioses. An Bras Dermatol, 2021, 96(5):527-538. doi:10.1016/j.abd.2021.04.005. pmid: 34275692
[10]	Chapman R, Jones L, D’Angelo A, et al. Nanopore-Based Metagenomic Sequencing in Respiratory Tract Infection: A Developing Diagnostic Platform. Lung, 2023, 201(2):171-179. doi:10.1007/s00408-023-00612-y. pmid: 37009923
[11]	Lin WH, Tang F, Liu SS. Diagnostic value of nanopore sequencing technology in nontuberculous mycobacterial pulmonary disease. Am J Transl Res, 2024, 16(8):4208-4215. doi:10.62347/HHRH8076.
[12]	Liu Z, Yang Y, Wang Q, et al. Diagnostic value of a nanopore sequencing assay of bronchoalveolar lavage fluid in pulmonary tuberculosis. BMC Pulm Med, 2023, 23(1):77. doi:10.1186/s12890-023-02337-3. pmid: 36890507
[13]	杨晨, 高卫卫, 郭羿成, 等. 纳米孔靶向测序在分枝杆菌鉴定和耐药基因检测中的研究进展. 中国防痨杂志, 2024, 46(5):578-583. doi:10.19982/j.issn.1000-6621.20240049.
[14]	张述耀, 侯铁英, 黎小妍, 等. 纳米孔测序在病原微生物检测中的应用专家共识. 中国药房, 2024, 35(14):1673-1682. doi:10.6039/j.issn.1001-0408.2024.14.01.

特征	例数	率(%)	特征	例数	率(%)
临床症状			腰痛	2	2.08
咳嗽	64	66.67	气喘	5	5.21
咳痰	55	57.29	头痛、头晕	2	2.08
胸闷、气促	14	14.58	呕吐	1	1.04
咯血	10	10.42	皮肤疱疹	1	1.04
发热	5	5.21	颈淋巴结肿大并破溃感染	1	1.04
纳差	4	4.17	影像学征象
乏力	5	5.21	支气管扩张	58	60.42
消瘦	3	3.13	空洞影	23	23.96
胸痛	4	4.17	多发结节	47	48.96

检测方法	NTM肺病诊断		敏感度 (%)	特异度 (%)	阳性预测值(%)	阴性预测值(%)	一致率 (%)	Kappa值
检测方法	阳性(例)	阴性(例)	敏感度 (%)	特异度 (%)	阳性预测值(%)	阴性预测值(%)	一致率 (%)	Kappa值
纳米孔测序技术			92.71	76.27	86.41	86.54	86.45	0.706
阳性	89	14
阴性	7	45
分枝杆菌鉴定(荧光PCR熔解曲线法)			63.54	84.75	87.14	58.82	71.61	0.445
阳性	61	9
阴性	35	50
分枝杆菌培养及鉴定			63.54	93.22	93.85	61.11	74.84	0.515
阳性	61	4
阴性	35	55
联合检测方法^a			84.38	79.66	87.10	75.81	82.58	0.634
阳性	81	12
阴性	15	47

[1]	Tuberculosis Control Branch of Chinese Antituberculosis Association, Standardization Professional Branch of Chinese Antituberculosis Association, Elderly Tuberculosis Control Branch of Chinese Antituberculosis Association. Expert consensus on the application of Mycobacterium tuberculosis infection detection technologies [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(4): 365-381.
[2]	Tuberculosis Basic Professional Branch, Chinese Antituberculosis Association. Expert consensus on the standardization of broth microdilution method for drug susceptibility testing of Mycobacterium tuberculosis in China [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(4): 382-392.
[3]	Wu Yifei, Li Qingchun, Jia Qingjun, Huang Yinyan, Bai Xuexin, Cheng Qinglin. Identification and drug resistance analysis of non-tuberculous mycobacterial pulmonary disease pathogens in suburban areas of Hangzhou [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(4): 420-425.
[4]	Wei Daijue, Sun Jianjun, Chen Yan, Wang Shijun, Chang Yamei, Wei Nianhuan, Zhang Xin, Tong Chongxiang. Analysis of drug resistance of 2941 clinical isolates of Mycobacterium tuberculosis in Gansu Province [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(4): 444-448.
[5]	Qi Dan, Wang Sheng, Liu Min, Gao Meiqin, Shi Feng, Li Bing, Bai Jun, Hao Ruixia, Wang Dong. Analysis of the etiological positive rate among pulmonary tuberculosis patients in designated tuberculosis hospitals in Ordos,Inner Mongolia Autonomous Region, 2015—2024 [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(4): 456-463.
[6]	Chen Hua, Chen Pinru. 2025 Update interpretation of “T/CHATA-001-2025 non-tuberculous mycobacteria disease diagnosis” social organization standard [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(2): 135-142.
[7]	Chen Hao, Lin Sihan, Li Xiaofen, Liu Zhidong, Lin Yanwei. Analysis of the effectiveness of pulmonary tuberculosis screening for freshmen based on Markov model [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(2): 169-175.
[8]	Chen Yu, Li Xiaorui, Wang Miaoran, Zhang Yuqi, Liu Chang, Wang Zhaohua, Shi Jie, Fan Lichao, Yin Zhihua, Xie Jianping. The research progress on the role of metal ions in tuberculosis [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(1): 102-112.
[9]	Xu Yannan, Fang Zihao, Zhao Wenli, Zheng Jiaxiong, Liu Suyang, Lin Jianxiong, Ji Liwei, Chang Qiaocheng. Characterisation of isoniazid-resistant Mycobacterium tuberculosis mutations in China [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(1): 14-21.
[10]	Gu Jinhua, Zhang Panpan. Evaluation of the application value of three detection methods for Mycobacterium tuberculosis in a comprehensive hospital [J]. Journal of Tuberculosis and Lung Disease, 2025, 6(1): 68-72.
[11]	Wu Xiucen, Chen Guihua. Interpretation of the 2023 U.S. Preventive Clinical Services Guidelines Workgroup Statement of Recommendations for Screening Adults for Latent Tuberculosis Infection [J]. Journal of Tuberculosis and Lung Disease, 2024, 5(5): 398-403.
[12]	Li Xiaoxue, Xiao Xiao, Xu Chunhua, Dong Shulan, Wang Shanshan, Cao Jiayi, Wu Zheyuan, Hu Yi, Shen Xin. The prevalence of latent tuberculosis infection among close contacts of active tuberculosis patients: a Meta-analysis [J]. Journal of Tuberculosis and Lung Disease, 2024, 5(5): 404-414.
[13]	Xiong Yan, Xiao Yue, Chen Chuang, Xia Yong, Li Yunkui, Lu Jia, Xia Lan. Analysis of tuberculosis screening results among college freshmen in Sichuan Province in 2023 [J]. Journal of Tuberculosis and Lung Disease, 2024, 5(5): 422-429.
[14]	Sun Bo, Feng Liping, Teng Chong, Zhu Hanfang, Zhao Bing, Feng Tao, Wang Qingkui, Zhou Hao, Gao Xinghai, Ou Xichao. Analysis of features of drug resistance of Mycobacterium tuberculosis and risk factors of multidrug-resistance in Hinggan League of Inner Mongolia Autonomous Region, 2021—2023 [J]. Journal of Tuberculosis and Lung Disease, 2024, 5(5): 437-444.
[15]	Zhang Jie, Ding Beichuan, Ren Yixuan, Tian Lili, Yi Junli, Pang Mengdi, Yang Xinyu. Exploring the causes of recurrence and genetic characteristics of tuberculosis strains in Beijing based on genotypic analysis [J]. Journal of Tuberculosis and Lung Disease, 2024, 5(2): 128-134.