Rapid Detection of Spring Viremia of Carp Virus (SVCV) Using RAA-CRISPR/Cas12a.
TL;DR
A recombinase-mediated isothermal amplification (RAA) in conjunction with the CRISPR/Cas12a system targeting the screened conserved region of the G gene in the genome to identify SVCV is established, offering a diagnosis project in resource-limited aquaculture environments and a field-applicable early warning of SVCV.
Abstract
Spring viremia of carp virus (SVCV), that can infect cyprinoid and koi causing an acute hemorrhagic symptom, with a feature of widespread epidemic, which poses a severe threat to the global aquaculture industry. Given the absence of specific effective medicine or vaccines, the early, accurate diagnosis is critical in the disease prevention. Many SVCV detection methods have been established, but most require specialized equipment and skilled operators, which limits their field application. In this study, we established a recombinase-mediated isothermal amplification (RAA) in conjunction with the CRISPR/Cas12a system targeting the screened conserved region of the G gene in the genome to identify SVCV. The optimal combination of the primer pairs and crRNA was screened via the fluorescence intensity. In addition, the test exhibited high sensitivity and specificity, which could detect at the limit of 1.243 × 101 copies/μL and show no cross-reactivity with other common freshwater fish pathogens. Furthermore, the result could be visible by excitation of fluorophores by blue light at 39°C within 30 min. The integrated method combines the advantages of RAA and CRISPR-based signal transduction, offering a diagnosis project in resource-limited aquaculture environments and a field-applicable early warning of SVCV.