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Specific Detection of Influenza A and B Viruses by CRISPR-Cas12a-Based Assay

Authors
Park, Bum JuPark, Man SeongLee, Jae MyunSong, Yoon Jae
Issue Date
3월-2021
Publisher
MDPI
Keywords
influenza virus; diagnosis; CRISPR-Cas12a; DETECTR
Citation
BIOSENSORS-BASEL, v.11, no.3
Indexed
SCIE
SCOPUS
Journal Title
BIOSENSORS-BASEL
Volume
11
Number
3
URI
https://scholar.korea.ac.kr/handle/2021.sw.korea/128493
DOI
10.3390/bios11030088
ISSN
2079-6374
Abstract
A rapid and accurate on-site diagnostic test for pathogens including influenza viruses is critical for preventing the spread of infectious diseases. Two types of influenza virus, A and B cause seasonal flu epidemics, whereas type A can cause influenza pandemics. To specifically detect influenza A (IAV) and B (IBV) viruses, we developed a clustered, regularly interspaced, short palindromic repeats (CRISPR) and CRISPR-associated (Cas) system-based assay. By coupling reverse transcription recombinase polymerase amplification (RT-RPA) and reverse transcription loop-mediated isothermal amplification (RT-LAMP), a CRISPR-Cas12a DNA endonuclease-targeted CRISPR trans-reporter (DETECTR) detected IAV and IBV titers as low as 1 x 10(0) plaque forming units (PFUs) per reaction without exhibiting cross-reactivity. Only 75 to 85 min were required to detect IAV and IBV, depending on isothermal nucleic acid amplification methods, and results were verified using a lateral flow strip assay that does not require additional analytic equipment. Taken together, our findings establish RT-RPA and RT-LAMP-coupled DETECTR-based diagnostic tests for rapid, specific and high-sensitivity detection of IAV and IBV using fluorescence and lateral flow assays. The diagnostic test developed in this study can be used to distinguish IAV and IBV infections, a capability that is necessary for monitoring and preventing the spread of influenza epidemics and pandemics.
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