Pan-Piscine Orthoreovirus (PRV) Detection Using Reverse Transcription Quantitative PCR

Area of Science:

• Aquatic Animal Health
• Virology
• Molecular Diagnostics

Background:

• Piscine orthoreovirus (PRV) affects salmonid species globally, with three known genotypes (PRV-1, PRV-2, PRV-3).
• Current detection methods rely on genotype-specific assays, necessitating multiple tests for comprehensive PRV surveillance.
• This limits the efficiency and scope of diagnostic and research programs for PRV.

Purpose of the Study:

• To develop and validate a single, sensitive, and specific molecular assay for detecting all known PRV genotypes.
• To provide a universal tool for PRV surveillance, research, and diagnostics.

Main Methods:

• Development and optimization of a real-time reverse transcription quantitative PCR (RT-qPCR) assay targeting a conserved region of the PRV M2 segment.
• Validation of the assay's sensitivity, specificity, and reproducibility through rigorous testing.
• Inter-laboratory comparison to assess assay robustness across different settings.

Main Results:

• The developed pan-PRV RT-qPCR assay accurately detects all three PRV genotypes with high sensitivity (as few as five RNA copies).
• The assay demonstrates excellent specificity, with no cross-reactivity to other salmonid viruses or host genomes.
• 100% agreement was observed in an inter-laboratory comparison, confirming the assay's reproducibility and robustness.

Conclusions:

• A single, highly sensitive, and specific RT-qPCR assay for universal PRV detection has been successfully developed.
• This pan-PRV assay is suitable for diverse applications, including research, diagnostics, and large-scale surveillance programs.
• The assay simplifies PRV detection, improving efficiency and accuracy in monitoring fish health.