Amplification-free CRISPR-Cas13a assay for detection of Taura syndrome virus

Abstract

Taura syndrome virus (TSV), which causes Taura syndrome, poses serious threats to the aquaculture industry. To date, early detection is still an important way to control its epidemic and spread. The clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) 13a (CRISPR-Cas13a) system, has been used to detect virus RNA. In this study, we developed an amplification-free CRISPR-Cas13a assay for directly detection of TSV from shrimp RNA. This assay could detect 107 copies/μL (16.6 pM) TSV in vitro transcribed (IVT) RNA in 5 min and 106 copies/μL (1.66 pM) within 15 min at 37 °C. It could distinguish TSV RNA specifically without any cross reaction with other shrimp pathogens. 56 Penaeus vannamei clinical samples were validated using this newly established assay, revealing 100 % consistency with the typical real-time RT-PCR (RT-qPCR) assay. These results demonstrate that this new assay for TSV is accurate, rapid and cost-effective, thus, it is promising for early diagnosis of RNA virus in aquaculture.

Keywords

• CRISPR-Cas13a
• Amplification-free detection
• Taura syndrome virus
• LwaCas13a