Tissue Distribution of the Piscine Novirhabdovirus Genotype IVb in Muskellunge (Esox masquinongy)

Abstract

A novel sublineage of the piscine novirhabdovirus (synonym: viral hemorrhagic septicemia virus), genotype IVb, emerged in the Laurentian Great Lakes, causing serious losses in resident fish species as early as 2003. Experimentally infected juvenile muskellunge (Esox masquinongy) were challenged with VHSV-IVb at high (1 × 105 PFU mL−1), medium (4 × 103 PFU mL−1), and low (100 PFU mL−1) doses. Samples from spleen, kidneys, heart, liver, gills, pectoral fin, large intestine, and skin/muscle were collected simultaneously from four fish at each predetermined time point and processed for VHSV-IVb reisolaton on Epitheliosum papulosum cyprini cell lines and quantification by plaque assay. The earliest reisolation of VHSV-IVb occurred in one fish from pectoral fin samples at 24 h post-infection. By 6 days post-infection (dpi), all tissue types were positive for VHSV-IVb. Statistical analysis suggested that virus levels were highest in liver, heart, and skin/muscle samples. In contrast, the kidneys and spleen exhibited reduced probability of virus recovery. Virus distribution was further confirmed by an in situ hybridization assay using a VHSV-IVb specific riboprobe. Heart muscle fibers, hepatocytes, endothelia, smooth muscle cells, and fibroblast-like cells of the pectoral fin demonstrated riboprobe labeling, thus highlighting the broad cellular tropism of VHSV-IVb. Histopathologic lesions were observed in areas where the virus was visualized.