Identification of a wild reservoir of salmonid alphavirus in common dab Limanda limanda, with emphasis on virus culture and sequencing

Common dab Limanda limanda from Scottish and international waters were examined by quantitative real-time RT-qPCR for evidence of viral RNA consistent with salmonid alphaviruses (SAV). SAV prevalence in heart tissue varied between sampling sites and reached up to 17% in fish collected near the Shetland Islands, Scotland. Raw Ct values ranging from 22.31 to 39.45 were obtained from the SAV-positive tissue material using the nsP1 RT-qPCR assay. Bayesian-, likelihood- and distance-based phylogenetic analyses performed with the amplified partial E2 gene sequence dataset suggest that the dab-derived virus belongs to SAV Subtypes I, II and V. A single SAV subtype was identified from the majority of sampling sites, apart from Shetland, where Subtypes II and V were also identified. The presence of SAV RNA from common dab in regions detached from salmon aquaculture lends support to the hypothesis that common dab are bone fide wild reservoirs of SAV, independent of fish farming activity. There was no link between the occurrence of viral RNA, length and sex of the dab, water depth, or health status as recorded using the International Council for the Exploration of the Sea (ICES) guidelines. In addition, the histological changes recorded in dab could not, with certainty, be attributed to infection with SAV. Finally, and for the first time, this study demonstrated that the dab-derived SAV Subtype V virus could be successfully cultured in a salmonid cell line.