Diagnostic application value of metagenomic next-generation sequencing technology in pathogen detection of mycobacterial infections_West China Medical Journal

Authors：

LI Xiaojie ¹ , Lü Xueyun ² , XU Yazhou ¹ , ZHANG Hongyan ³ ,  HU Bo ¹

1. Department of Laboratory Medicine, the Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510630, P. R. China;
2. Department of Laboratory Medicine, the Second People’s Hospital of Shaoguan City, Shaoguan, Guangdong 512026, P. R. China;
3. School of Clinical Laboratory Science, Guizhou Medical University, Guiyang, Guizhou 561113, P. R. China;

Corresponding?author：

HU Bo, Email: hubo@mail.sysu.edu.cn

Keywords：

Mycobacteria; metagenomic next-generation sequencing; infection; molecular diagnosis

DOI：

10.7507/1002-0179.202607006

Video：

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Abstract Full text Figures/Tables Video References Cited by

Objective To investigate the value of metagenomic next-generation sequencing (mNGS) in the rapid diagnosis of clinical mycobacterial infections. Methods The clinical data of 782 hospitalized patients with suspected mycobacterial infections from April 2023 to December 2025 were retrospectively analyzed. The diagnostic performance of mNGS was compared with that of conventional methods, including culture and acid-fast staining. Results Among the 782 patients, 465 were ultimately diagnosed with mycobacterial infections, including 388 cases of Mycobacterium tuberculosis infection and 77 cases of non-tuberculous mycobacterial infection. Using the final clinical diagnosis as the gold standard, the sensitivity, specificity, positive predictive value, and negative predictive value of mNGS for diagnosing mycobacterial infections in all specimens were 52.5%, 79.8%, 79.2%, and 53.4%, respectively; those of culture were 1.9%, 100.0%, 100.0%, and 41.0%, respectively; and those of acid-fast staining were 14.2%, 99.6%, 98.4%, and 41.1%, respectively. The areas under the receiver operating characteristic curves for mNGS, culture, and acid-fast staining in evaluating the diagnosis of mycobacterial infections were 0.66 [95% confidence interval (CI) (0.63, 0.69)], 0.51 [95%CI (0.50, 0.52)], and 0.57 [95%CI (0.55, 0.59)], respectively. Conclusion mNGS demonstrates certain sensitivity, specificity, and negative predictive value in the diagnosis of mycobacterial infections, and may serve as a reference for early clinical diagnosis.

Citation： LI Xiaojie, Lü Xueyun, XU Yazhou, ZHANG Hongyan, HU Bo. Diagnostic application value of metagenomic next-generation sequencing technology in pathogen detection of mycobacterial infections. West China Medical Journal, 2026, 41(8): 1253-1258. doi: 10.7507/1002-0179.202607006 Copy

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9.	中華醫學會檢驗醫學分會臨床微生物學組, 中華醫學會微生物學與免疫學分會臨床微生物學組, 中國醫療保健國際交流促進會臨床微生物與感染分會. 宏基因組高通量測序技術應用于感染性疾病病原檢測中國專家共識[J]. 中華檢驗醫學雜志, 2021, 44(2): 107-120.
10.	Chen H, Yin Y, Gao H, et al. Clinical utility of in-house metagenomic next-generation sequencing for the diagnosis of lower respiratory tract infections and analysis of the host immune response[J]. Clin Infect Dis, 2020, 71(Suppl 4): S416-S426.
11.	Zhang X, Chen H, Lin Y, et al. Diagnosis of non-tuberculous mycobacterial pulmonary disease by metagenomic next-generation sequencing on bronchoalveolar lavage fluid[J]. Infect Drug Resist, 2023, 16: 4137-4145.
12.	Wang L, Chen Y, Wang Q, et al. Comparison of molecular testing methods for diagnosing non-tuberculous mycobacterial infections[J]. Eur J Clin Microbiol Infect Dis, 2025, 44(1): 109-116.
13.	Coda R, Waller S, Vopat B. Nontuberculosis mycobacterium infections in orthopaedic surgery: review of the epidemiology, pathogenesis, diagnosis, and treatment guidelines[J]. J Am Acad Orthop Surg Glob Res Rev, 2025, 9(5): e24.00274.
14.	WHO announces first prequalification of a tuberculosis diagnostic test[J]. Saudi Med J, 2025, 46(1): 113.
15.	Xu P, Yang K, Yang L, et al. Next-generation metagenome sequencing shows superior diagnostic performance in acid-fast staining sputum smear-negative pulmonary tuberculosis and non-tuberculous mycobacterial pulmonary disease[J]. Front Microbiol, 2022, 13: 898195.
16.	Daley CL, Iaccarino JM, Lange C, et al. Treatment of nontuberculous mycobacterial pulmonary disease: an official ATS/ERS/ESCMID/IDSA clinical practice guideline[J]. Clin Infect Dis, 2020, 71(4): e1-e36.
17.	Wei W, Cao J, Wu XC, et al. Diagnostic performance of metagenomic next-generation sequencing in non-tuberculous mycobacterial pulmonary disease when applied to clinical practice[J]. Infection, 2023, 51(2): 397-405.
18.	Yu J, Lv X, Wang Q, et al. Metagenomic sequencing for identification of nontuberculous mycobacteria and other pathogens in patients with mixed infection of the lung[J]. Front Cell Infect Microbiol, 2025, 15: 1592216.
19.	中國防癆協會非結核分枝桿菌病分會, 孫勤, 程麗平. 非結核分枝桿菌病分子生物學診斷專家共識[J]. 中國防癆雜志, 2025, 47(8): 961-975.
20.	黃新蕾, 陳建保. 痰涂片、固體培養法、GeneXpert MTB/RIF 單獨及聯合檢測結核分枝桿菌能力的比較[J]. 臨床肺科雜志, 2025, 30(1): 88-91, 125.
21.	Li L, Chi Y, Kong Y, et al. Rapid species-level discrimination of pulmonary TB and NTM by metagenomic next-generation sequencing with concurrent respiratory microbiome profiling[J]. Diagn Microbiol Infect Dis, 2026, 116(1): 117442.

1. Sarro YD, Kone B, Diarra B, et al. Simultaneous diagnosis of tuberculous and non-tuberculous mycobacterial diseases: time for a better patient management[J/OL]. Clin Microbiol Infect Dis, 2018, 3(3): 10.15761/CMID.1000144. DOI: 10.15761/CMID.1000144.
2. Nguyen MH, Haas MK, Kasperbauer SH, et al. Nontuberculous mycobacterial pulmonary disease: patients, principles, and prospects[J]. Clin Infect Dis, 2024, 79(4): e27-e47.
3. Gopalaswamy R, Shanmugam S, Mondal R, et al. Of tuberculosis and non-tuberculous mycobacterial infections - a comparative analysis of epidemiology, diagnosis and treatment[J]. J Biomed Sci, 2020, 27(1): 74.
4. Li Y, Bian W, Wu S, et al. Metagenomic next-generation sequencing for Mycobacterium tuberculosis complex detection: a meta-analysis[J]. Front Public Health, 2023, 11: 1224993.
5. Chen H, Huang Q, Wu W, et al. Assessment and clinical utility of metagenomic next-generation sequencing for suspected lower respiratory tract infections[J]. Eur J Med Res, 2024, 29(1): 213.
6. Sharma SK, Upadhyay V. Epidemiology, diagnosis & treatment of non-tuberculous mycobacterial diseases[J]. Indian J Med Res, 2020, 152(3): 185-226.
7. Hashimoto K, Fukushima K, Matsumoto Y, et al. Molecular epidemiological surveillance for non-tuberculous mycobacterial pulmonary disease: a single-center prospective cohort study[J]. Microbiol Spectr, 2025, 13(10): e0043625.
8. Gu W, Miller S, Chiu CY. Clinical metagenomic next-generation sequencing for pathogen detection[J]. Annu Rev Pathol, 2019, 14: 319-338.
9. 中華醫學會檢驗醫學分會臨床微生物學組, 中華醫學會微生物學與免疫學分會臨床微生物學組, 中國醫療保健國際交流促進會臨床微生物與感染分會. 宏基因組高通量測序技術應用于感染性疾病病原檢測中國專家共識[J]. 中華檢驗醫學雜志, 2021, 44(2): 107-120.
10. Chen H, Yin Y, Gao H, et al. Clinical utility of in-house metagenomic next-generation sequencing for the diagnosis of lower respiratory tract infections and analysis of the host immune response[J]. Clin Infect Dis, 2020, 71(Suppl 4): S416-S426.
11. Zhang X, Chen H, Lin Y, et al. Diagnosis of non-tuberculous mycobacterial pulmonary disease by metagenomic next-generation sequencing on bronchoalveolar lavage fluid[J]. Infect Drug Resist, 2023, 16: 4137-4145.
12. Wang L, Chen Y, Wang Q, et al. Comparison of molecular testing methods for diagnosing non-tuberculous mycobacterial infections[J]. Eur J Clin Microbiol Infect Dis, 2025, 44(1): 109-116.
13. Coda R, Waller S, Vopat B. Nontuberculosis mycobacterium infections in orthopaedic surgery: review of the epidemiology, pathogenesis, diagnosis, and treatment guidelines[J]. J Am Acad Orthop Surg Glob Res Rev, 2025, 9(5): e24.00274.
14. WHO announces first prequalification of a tuberculosis diagnostic test[J]. Saudi Med J, 2025, 46(1): 113.
15. Xu P, Yang K, Yang L, et al. Next-generation metagenome sequencing shows superior diagnostic performance in acid-fast staining sputum smear-negative pulmonary tuberculosis and non-tuberculous mycobacterial pulmonary disease[J]. Front Microbiol, 2022, 13: 898195.
16. Daley CL, Iaccarino JM, Lange C, et al. Treatment of nontuberculous mycobacterial pulmonary disease: an official ATS/ERS/ESCMID/IDSA clinical practice guideline[J]. Clin Infect Dis, 2020, 71(4): e1-e36.
17. Wei W, Cao J, Wu XC, et al. Diagnostic performance of metagenomic next-generation sequencing in non-tuberculous mycobacterial pulmonary disease when applied to clinical practice[J]. Infection, 2023, 51(2): 397-405.
18. Yu J, Lv X, Wang Q, et al. Metagenomic sequencing for identification of nontuberculous mycobacteria and other pathogens in patients with mixed infection of the lung[J]. Front Cell Infect Microbiol, 2025, 15: 1592216.
19. 中國防癆協會非結核分枝桿菌病分會, 孫勤, 程麗平. 非結核分枝桿菌病分子生物學診斷專家共識[J]. 中國防癆雜志, 2025, 47(8): 961-975.
20. 黃新蕾, 陳建保. 痰涂片、固體培養法、GeneXpert MTB/RIF 單獨及聯合檢測結核分枝桿菌能力的比較[J]. 臨床肺科雜志, 2025, 30(1): 88-91, 125.
21. Li L, Chi Y, Kong Y, et al. Rapid species-level discrimination of pulmonary TB and NTM by metagenomic next-generation sequencing with concurrent respiratory microbiome profiling[J]. Diagn Microbiol Infect Dis, 2026, 116(1): 117442.

West China Medical Journal

Diagnostic application value of metagenomic next-generation sequencing technology in pathogen detection of mycobacterial infections

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