Rapid and Highly Sensitive Detection of Utilizing the Recombinase Aided Amplification-Based CRISPR-Cas13a System

Overview

Journal Microorganisms

Specialty Microbiology

Date 2024 Aug 29

PMID 39203350

Authors

Qiao Li

Nenhan Wang

Mengdi Pang

Honghao Miao

Xiaowei Dai

Bo Li

Xinyu Yang

Chuanyou Li

Yi Liu

Affiliations

Soon will be listed here.

Abstract

Tuberculosis (TB), a disease caused by (MTB) infection, remains a major threat to global public health. To facilitate early TB diagnosis, an IS6110 gene-based recombinase aided amplification (RAA) assay was coupled to a clustered, regularly interspaced short palindromic repeats (CRISPR)-Cas13a fluorescence assay to create a rapid MTB detection assay (named RAA-CRISPR-MTB). Its diagnostic efficacy was evaluated for sensitivity and specificity through sequential testing of recombinant plasmids, mycobacterium strains, and clinical specimens. RAA-CRISPR detected IS6110 genes at levels approaching 1 copy/μL with pUC57-6110 as the template and 10 copies/μL with H37Rv as the template. There was no observed cross detection of non-tuberculosis mycobacteria (NTM) with either template. Furthermore, RAA-CRISPR testing of 151 clinical specimens yielded a diagnostic specificity rate of 100% and a diagnostic sensitivity rate of 69% that exceeded the corresponding Xpert MTB/RIF assay rate (60%). In conclusion, we established a novel RAA-CRISPR assay that achieved highly sensitive and specific MTB detection for use as a clinical TB diagnostic tool in resource-poor settings.

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