|ZHAO, JULIE - Department Of Fisheries And Oceans Canada|
|VENDRAMIN, NICCOLO - Technical University Of Denmark|
|CUENCA, ARGELIA - Technical University Of Denmark|
|HAWLEY, LAURA - Department Of Fisheries And Oceans Canada|
|GARVER, KYLE - Department Of Fisheries And Oceans Canada|
Submitted to: Pathogens
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/29/2021
Publication Date: 11/30/2021
Citation: Zhao, J., Vendramin, N., Cuenca, A., Polinski, M.P., Hawley, L., Garver, K. 2021. Pan-Piscine Orthoreovirus (PRV) detection using reverse transcription quantitative PCR. Pathogens. https://doi.org/10.3390/pathogens10121548.
Interpretive Summary: Piscine orthoreovirus (PRV) is a virus that infects salmon. There are multiple genetic forms of PRV circulating in salmon populations across the globe, each with unique host targets and disease causing potential. Previous methods used for viral detection were not able to identify all known genetic types (i.e., they were designed to detect specific subsets only). In this publication, we describe a method to universally detect all currently known forms of PRV in a single test and demonstrate its utility across multiple international testing laboratories.
Technical Abstract: Piscine orthoreovirus (PRV) infects farmed and wild salmon and trout species in North America, South America, Europe and East Asia. PRV groups into 3 distinct genotypes (PRV-1, PRV-2, PRV-3), that can vary in distribution, host specificity, and/or disease potential. Detection of the virus is currently restricted to genotype specific assays such that surveillance programs require the use of three assays to ensure universal detection of PRV. Consequently, herein we have developed, optimized, and validated a real-time reverse transcription quantitative PCR assay (RT-qPCR) that can detect all known PRV genotypes with high sensitivity and specificity. Targeting a conserved region at the 5’ terminus of the M2 segment, the pan-PRV assay reliably detected all PRV genotypes with as few as 5 copies of RNA. The assay exclusively amplifies PRV and does not cross-react with other salmonid viruses or salmonid host genomes and can be performed as either a one- or two-step RT-qPCR. The assay is highly reproducible and robust showing 100% agreement in test results from an inter-laboratory comparison between two laboratories in two countries. Overall, as the assay provides a single test to achieve highly sensitive pan-specific PRV detection, it is suitable for research, diagnostic, and surveillance purposes.