CRISPR detection of circulating cell-free Mycobacterium tuberculosis DNA in adults and children, including children with HIV: a molecular diagnostics study.
CRISPR detection of circulating cell-free Mycobacterium tuberculosis DNA in adults and children, including children with HIV: a molecular diagnostics study.
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CRISPR检测成人和儿童（包括HIV感染儿童）的循环无细胞结核分枝杆菌DNA：一项分子诊断研究
DOI：
10.1016/s2666-5247(22)00087-8
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发表时间：
2022-07
期刊：
影响因子：
38.2
通讯作者：
Hu, Tony Y.
中科院分区：
文献类型：
--
作者：
Huang, Zhen;LaCourse, Sylvia M.;Kay, Alexander W.;Stern, Joshua;Escudero, Jaclyn N.;Youngquist, Brady M.;Zheng, Wenshu;Vambe, Debrah;Dlamini, Muyalo;Mtetwa, Godwin;Cranmer, Lisa M.;Njuguna, Irene;Wamalwa, Dalton C.;Maleche-Obimbo, Elizabeth;Catanzaro, Donald G.;Lyon, Christopher J.;John-Stewart, Grace;DiNardo, Andrew;Mandalakas, Anna M.;Ning, Bo;Hu, Tony Y.
结核病仍然是全球死亡率的主要原因，特别是对于艾滋病毒感染者（CLHIV）的成人和儿童，基于结核病的检测诊断不足。需要非痰基检测来改善结核病诊断和结核病治疗监测。本研究的目的是确定血液中结核分枝杆菌无细胞DNA（Mtb-cfDNA）的超灵敏检测是否可以诊断结核病并评估结核病治疗反应。在这项分子诊断研究中，我们分析了来自斯威士兰和肯尼亚两个评估结核病的患者群体的存档血清，以检测Mtb-cfDNA，分析了来自具有足够血清体积和明确诊断结果的所有个体的血清。使用优化的CRISPR介导的结核病（CRISPR-TB）测定来检测在招募时来自患有推定结核病的成人和儿童及其无症状的家庭接触者的血清中的Mtb-cfDNA，以及在招募时和在结核病治疗期间来自结核病高风险的症状性CLHIV队列的血清中的Mtb-cfDNA，所述CLHIV队列在招募时和结核病治疗期间提供纵向血清。CRISPR-TB在主要为HIV阴性的斯威士兰成人队列中识别出微生物学和临床确诊的结核病例，灵敏度为96%（27/28 [96%]，95% CI 80-100）和94%特异性（17例中的16例[94%]，71-100），在儿科队列中敏感性为83%（6例中的5例[83%]，36-100），特异性为95%（22例中的21例[95%]，77-100），包括所有6例肺外结核病例。在肯尼亚CLHIV队列中，CRISPR-TB检测到了所有（13例中的13例[100%]，75-100例）确诊的结核病例和85%（46例中的39例[85%]，71-94例）通过非微生物临床结果诊断的未确诊结核病例。在入组时CRISPR-TB阳性的CLHIV患者在入组后6个月的死亡风险高出2.4倍。结核病治疗开始后Mtb-cfDNA信号降低，结核病治疗开始后6个月Mtb-cfDNA接近或完全清除。CRISPR介导的循环Mtb-cfDNA检测有望增加儿科结核病和艾滋病毒相关结核病的识别，以及早期诊断和快速监测结核病治疗反应的潜力。美国国防部、国家儿童健康和人类发展研究所、国家过敏和传染病研究所、华盛顿大学艾滋病研究中心和韦瑟黑德总统捐赠基金。
Tuberculosis remains a leading cause of global mortality, especially for adults and children living with HIV (CLHIV) underdiagnosed by sputum-based assays. Non-sputum-based assays are needed to improve tuberculosis diagnosis and tuberculosis treatment monitoring. Our aim in this study was to determine whether ultrasensitive detection of Mycobacterium tuberculosis cell-free DNA (Mtb-cfDNA) in blood can diagnose tuberculosis and evaluate tuberculosis treatment responses. In this molecular diagnostics study we analysed archived serum from two patient populations evaluated for tuberculosis in Eswatini and Kenya to detect Mtb-cfDNA, analysing serum from all individuals who had both sufficient serum volumes and clear diagnostic results. An optimised CRISPR-mediated tuberculosis (CRISPR-TB) assay was used to detect Mtb-cfDNA in serum at enrolment from adults and children with presumptive tuberculosis and their asymptomatic household contacts, and at enrolment and during tuberculosis treatment from a cohort of symptomatic CLHIV at high risk for tuberculosis, who provided longitudinal serum at enrolment and during tuberculosis treatment. CRISPR-TB identified microbiologically and clinically confirmed tuberculosis cases in the predominantly HIV-negative Eswatini adult cohort with 96% sensitivity (27 [96%] of 28, 95% CI 80–100) and 94% specificity (16 [94%] of 17, 71–100), and with 83% sensitivity (5 [83%] of 6, 36–100) and 95% specificity (21 [95%] of 22, 77–100) in the paediatric cohort, including all six cases of extrapulmonary tuberculosis. In the Kenyan CLHIV cohort, CRISPR-TB detected all (13 [100%] of 13, 75–100) confirmed tuberculosis cases and 85% (39 [85%] of 46, 71–94) of unconfirmed tuberculosis cases diagnosed by non-microbiological clinical findings. CLHIV who were CRISPR-TB positive at enrolment had a 2·4-times higher risk of mortality by 6 months after enrolment. Mtb-cfDNA signal decreased after tuberculosis treatment initiation, with near or complete Mtb-cfDNA clearance by 6 months after tuberculosis treatment initiation. CRISPR-mediated detection of circulating Mtb-cfDNA shows promise to increase the identification of paediatric tuberculosis and HIV-associated tuberculosis, and potential for early diagnosis and rapid monitoring of tuberculosis treatment responses. US Department of Defense, National Institute of Child Health and Human Development, National Institute of Allergy and Infectious Diseases, University of Washington Center for AIDS Research, and the Weatherhead Presidential Endowment fund.