Leung Kenneth Siu-Sing, Tam Kingsley King-Gee, Ng Timothy Ting-Leung, Lao Hiu-Yin, Shek Raymond Chiu-Man, Ma Oliver Chiu Kit, Yu Shi-Hui, Chen Jing-Xian, Han Qi, Siu Gilman Kit-Hang, Yam Wing-Cheong
Department of Microbiology, Queen Mary Hospital, The University of Hong Kong, Pokfulam, Hong Kong SAR, China.
Department of Health Technology and Informatics, The Hong Kong Polytechnic University, Kowloon, Hong Kong SAR, China.
Front Microbiol. 2022 Sep 9;13:974428. doi: 10.3389/fmicb.2022.974428. eCollection 2022.
An in-house-developed target amplicon sequencing by next-generation sequencing technology (TB-NGS) enables simultaneous detection of resistance-related mutations in (MTB) against 8 anti-tuberculosis drug classes. In this multi-center study, we investigated the clinical utility of incorporating TB-NGS for rapid drug-resistant MTB detection in high endemic regions in southeast China. From January 2018 to November 2019, 4,047 respiratory specimens were available from patients suffering lower respiratory tract infections in Hong Kong and Guangzhou, among which 501 were TB-positive as detected by in-house IS6110-qPCR assay with diagnostic sensitivity and specificity of 97.9 and 99.2%, respectively. Preliminary resistance screening by GenoType MTBDR and MTBDR identified 25 drug-resistant specimens including 10 multidrug-resistant TB. TB-NGS was performed using MiSeq on all drug-resistant specimens alongside 67 pan-susceptible specimens, and demonstrated 100% concordance to phenotypic drug susceptibility test. All phenotypically resistant specimens with dominating resistance-related mutations exhibited a mutation frequency of over 60%. Three quasispecies were identified with mutation frequency of less than 35% among phenotypically susceptible specimens. They were well distinguished from phenotypically resistant cases and thus would not complicate TB-NGS results interpretations. This is the first large-scale study that explored the use of laboratory-developed NGS platforms for rapid TB diagnosis. By incorporating TB-NGS with our proposed diagnostic algorithm, the workflow would provide a user-friendly, cost-effective routine diagnostic solution for complicated TB cases with an average turnaround time of 6 working days. This is critical for timely management of drug resistant TB patients and expediting public health control on the emergence of drug-resistant TB.
通过下一代测序技术进行的内部开发的靶向扩增子测序（TB-NGS）能够同时检测结核分枝杆菌（MTB）对8类抗结核药物的耐药相关突变。在这项多中心研究中，我们调查了在中国东南部高流行地区将TB-NGS用于快速检测耐多药MTB的临床实用性。2018年1月至2019年11月，从香港和广州患有下呼吸道感染的患者中获得了4047份呼吸道标本，其中501份经内部IS6110-qPCR检测为TB阳性，诊断敏感性和特异性分别为97.9%和99.2%。通过GenoType MTBDR和MTBDR进行的初步耐药性筛查确定了25份耐药标本，包括10份耐多药结核病。对所有耐药标本以及67份全敏感标本使用MiSeq进行TB-NGS检测，结果显示与表型药物敏感性试验的一致性为100%。所有表型耐药且主要存在耐药相关突变的标本，其突变频率均超过60%。在表型敏感标本中鉴定出3个准种，其突变频率低于35%。它们与表型耐药病例有明显区别，因此不会使TB-NGS结果的解释复杂化。这是第一项探索使用实验室开发的NGS平台进行快速结核病诊断的大规模研究。通过将TB-NGS与我们提出的诊断算法相结合，该工作流程将为复杂结核病病例提供一种用户友好、具有成本效益的常规诊断解决方案，平均周转时间为6个工作日。这对于及时管理耐多药结核病患者以及加快对耐多药结核病出现的公共卫生控制至关重要。
