Rapid and sensitive diagnosis of live Mycobacterium tuberculosis using clustered regularly interspaced short palindromic repeat‐Cas13a point‐of‐care RNA testing
Yu Wang,
Huihuang Lin,
Anqi Yang,
Jiaming Huang,
Weicong Ren,
Jiajun Dong,
Shaojie Wang,
Wenxue Xu,
Yu Pang,
Jieming Qu,
Jia Liu
Affiliations
Yu Wang
Shanghai Institute for Advanced Immunochemical Studies and School of Life Science and Technology ShanghaiTech UniversityShanghai China
Huihuang Lin
Department of Pulmonary and Critical Care Medicine, Ruijin Hospital Shanghai Jiao Tong University School of Medicine Shanghai China
Anqi Yang
Department of Central Laboratory the Second Affiliated Hospital of Fujian Medical University Quanzhou China
Jiaming Huang
Department of Microbiology the Second Affiliated Hospital of Fujian Medical University Quanzhou China
Weicong Ren
Department of Bacteriology and Immunology Beijing Chest Hospital Capital Medical University/Beijing Tuberculosis & Thoracic Tumor Research Institute Beijing China
Jiajun Dong
Shanghai Institute for Advanced Immunochemical Studies and School of Life Science and Technology ShanghaiTech UniversityShanghai China
Shaojie Wang
Shanghai Institute for Advanced Immunochemical Studies and School of Life Science and Technology ShanghaiTech UniversityShanghai China
Wenxue Xu
Shanghai Institute for Advanced Immunochemical Studies and School of Life Science and Technology ShanghaiTech UniversityShanghai China
Yu Pang
Department of Bacteriology and Immunology Beijing Chest Hospital Capital Medical University/Beijing Tuberculosis & Thoracic Tumor Research Institute Beijing China
Jieming Qu
Department of Pulmonary and Critical Care Medicine, Ruijin Hospital Shanghai Jiao Tong University School of Medicine Shanghai China
Jia Liu
Shanghai Institute for Advanced Immunochemical Studies and School of Life Science and Technology ShanghaiTech UniversityShanghai China
Abstract Mycobacterium tuberculosis (MTB) is the causal pathogen of tuberculosis (TB). Rapid and accurate detection of live MTB is important for transmission control and patient treatment. Here, we described a clustered regularly interspaced short palindromic repeat (CRISPR)‐Cas13a‐based molecular diagnosis approach for rapid and specific detection of live MTB. This detection method, which we termed CRISPR‐Live‐MTB, contained two consecutive reactions including nuclear acid sequence‐based amplification (NASBA) and CRISPR‐Cas13a collateral cleavage reaction. CRISPR‐Live‐MTB could efficiently detect MTB single‐stranded RNA (ssRNA) in 2 hours with high specificity over double‐stranded DNA (dsDNA). Importantly, CRISPR‐Live‐MTB exhibited a limit of detection of 2.4 copies for MTB ssRNA, which was 1000 times lower than that of the clinically used NASBA method. Moreover, lateral flow was integrated into the CRISPR‐Live‐MTB method to enable point‐of‐care testing application with a sensitivity of 95% and a specificity of 100%. Overall, our study demonstrated the feasibility of CRISPR‐Live‐MTB as a rapid, sensitive, and specific approach for live MTB detection.
