Identification of Mycobacterium and analysis of drug resistance in Mycobacterium tuberculosis using nanopore targeted sequencing technology
HUANG Yinyan1, WANG Meng1, XU Xiang2
1. Hangzhou Center for Disease Control and Prevention (Hangzhou Health Supervision Institution), Hangzhou, Zhejiang 310021, China; 2. Hangzhou Shengting Medical Technology Co., Ltd, Hangzhou, Zhejiang 311113, China
Abstract：Objective To identify Mycobacterium and analyze the drug resistance of Mycobacterium tuberculosis by using nanopore targeted sequencing technology, so as to provide the methodological reference for the rapid diagnosis of tuberculosis. Methods Samples from patients aged 18 to 80 years old who were highly suspected of Mycobacterium tuberculosis infection and diagnosed with Mycobacterium tuberculosis infection in designated tuberculosis hospitals from March to November 2022 were collected for nanopore targeted sequencing technology. The detection status of Mycobacterium, types of Mycobacterium tuberculosis drug resistance, and gene mutation sites were analyzed. Results A total of 200 samples were collected, including 160 respiratory tract samples and 40 non-respiratory tract samples. Nanopore targeted sequencing technology detected 194 positive mycobacterial samples, with a detection sensitivity of 97.00% (95%CI: 93.28%-98.77%). There were 187 positive cases of Mycobacterium tuberculosis and 7 cases of nontuberculous mycobacteria, with detection rates of 93.50% and 3.50%, respectively. Nanopore targeted sequencing technology identified 11 mutations in the rifampicin resistance gene rpoB, 1 mutation in the isoniazid resistance gene katG, 2 mutations in the gene inhA, 3 mutations in the ethambutol resistance gene embB, 2 mutations in the streptomycin resistance gene rpsL, 1 mutation in the gene rrs, 11 mutations in the pyraz inamide resistance gene pncA, 6 mutations in the fluoroquinolone resistance gene gyrA, 2 mutations in the gene gyrB, and 3 mutations in the aminoglycoside resistance gene rrs. It took 5-6 hours to complete the detection of one sample using nanopore targeted sequencing technology. Conclusions The nanopore targeted sequencing technology has high sensitivity for detecting Mycobacterium tuberculosis and can identify 42 nucleic acid mutation sites in 9 drug resistance genes associated with 7 anti-tuberculosis drugs. It is time-efficient and has certain applicability in the rapid diagnosis of tuberculosis and the analysis of drug resistance genes.
黄银燕, 王勐, 徐翔. 纳米孔靶向测序技术鉴定分枝杆菌及分析结核分枝杆菌耐药性[J]. 预防医学, 2025, 37(6): 640-644,648.
HUANG Yinyan, WANG Meng, XU Xiang. Identification of Mycobacterium and analysis of drug resistance in Mycobacterium tuberculosis using nanopore targeted sequencing technology. Preventive Medicine, 2025, 37(6): 640-644,648.
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