AUTHOR=Zhang Wanhong , Jiao Yubing , Ding Chengying , Shen Lili , Li Ying , Yu Yanbi , Huang Kun , Li Bin , Wang Fenglong , Yang Jinguang 

TITLE=Rapid Detection of Tomato Spotted Wilt Virus With Cas13a in Tomato and Frankliniella occidentalis

JOURNAL=Frontiers in Microbiology

VOLUME=Volume 12 - 2021

YEAR=2021

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2021.745173

DOI=10.3389/fmicb.2021.745173

ISSN=1664-302X

ABSTRACT=As one of the top 10 plant viruses, the severity of losses to crop productivity caused by Tomato Spotted Wilt Virus (TSWV) has resulted in an urgent need to develop a more sensitive and rapid detection method. In this study, we developed a CRISPR/Cas13a-based detection system to diagnose TSWV in tomato and the western flower thrips (Frankliniella occidentalis). The detection system relies on recombinase polymerase amplification (RPA) and Cas13a-mediated collateral cleavage activity. Positive results can be distinguished by a significantly enhanced fluorescence signal after 20 min. We tested the sensitivity of CRISPR/Cas13a-based detection system. We found that the detection system that we developed has limits of detection that reach 2.26×102 copies/μL and a 10-fold increase compared with the sensitivity of using RT-PCR to detect the virus. Furthermore, the CRISPR/Cas13a-based detection system has a high selectivity for TSWV without interference from other viruses. The CRISPR/Cas13a-based detection system was utilized to detect TSWV in samples of tomato leaves and the transmission vector F. occidentalis that were in complete agreement with the results using RT-PCR to detect the virus.