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ERA-CRISPR/Cas12a system: a rapid, highly sensitive and specific assay for Mycobacterium tuberculosis .
- Source :
-
Frontiers in cellular and infection microbiology [Front Cell Infect Microbiol] 2024 Aug 21; Vol. 14, pp. 1454076. Date of Electronic Publication: 2024 Aug 21 (Print Publication: 2024). - Publication Year :
- 2024
-
Abstract
- Introduction: Mycobacterium tuberculosis , the causative agent of human tuberculosis, poses a significant threat to global public health and imposes a considerable burden on the economy. However, existing laboratory diagnostic methods for M. tuberculosis are time-consuming and have limited sensitivity levels.<br />Methods: The CRISPR/Cas system, commonly known as the "gene scissors", demonstrates remarkable specificity and efficient signal amplification capabilities. Enzymatic recombinase amplification (ERA) was utilized to rapidly amplify trace DNA fragments at a consistent temperature without relying on thermal cyclers. By integrating of CRISPR/Cas12a with ERA, we successfully developed an ERA-CRISPR/Cas12a detection system that enables rapid identification of M. tuberculosis .<br />Results: The sensitivity of the ERA-CRISPR/Cas12a fluorescence and lateral flow systems was 9 copies/μL and 90 copies/μL, respectively. Simultaneously, the detection system exhibited no cross-reactivity with various of respiratory pathogens and non-tuberculosis mycobacteria, demonstrating a specificity of 100%. The positive concordance rate between the ERA-CRISPR/Cas12a fluorescence system and commercial qPCR was 100% in 60 clinical samples. Meanwhile, the lateral flow system showed a positive concordance rate of 93.8% when compared to commercial qPCR. Both methods demonstrated a negative concordance rate of 100%, and the test results can be obtained in 50 min at the earliest.<br />Discussion: The ERA-CRISPR/Cas12a system offers a rapid, sensitive, and specific method that presents a novel approach to laboratory diagnosis of M. tuberculosis .<br />Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.<br /> (Copyright © 2024 Gan, Yu, Deng and He.)
- Subjects :
- Humans
Tuberculosis diagnosis
Tuberculosis microbiology
Recombinases metabolism
Recombinases genetics
Molecular Diagnostic Techniques methods
Bacterial Proteins genetics
DNA, Bacterial genetics
CRISPR-Associated Proteins genetics
Endodeoxyribonucleases
Mycobacterium tuberculosis genetics
Mycobacterium tuberculosis isolation & purification
CRISPR-Cas Systems
Sensitivity and Specificity
Nucleic Acid Amplification Techniques methods
Subjects
Details
- Language :
- English
- ISSN :
- 2235-2988
- Volume :
- 14
- Database :
- MEDLINE
- Journal :
- Frontiers in cellular and infection microbiology
- Publication Type :
- Academic Journal
- Accession number :
- 39233906
- Full Text :
- https://doi.org/10.3389/fcimb.2024.1454076