| To identify putative amino acid residues that could be involved in the virulence of infectious pancreatic necrosis virus (IPNV), overlapping cDNA clones of segments A and B of virulent and avirulent Sp (ALV 122 and ALV 122-10) and VR-299 (VR299-2 and VR299-25) isolates were constructed. The complete nucleotide and deduced amino acid sequences of Sp isolates revealed only five amino acid changes in segment A, which reside in VP2 region, whereas, 13 residues were different between VR299-2 and VR299-25 isolates of which, 12 were present in VP2 region, indicating that residues within VP2 play a major role in the virulence of IPNV. Analyses of the Sp isolates also showed the presence of two open reading frames (ORFs) in segment A that codes for a 12-15-kDa polypeptide, known as VP5, and a putative 25-kDa protein, containing a bipartite nuclear targeting sequence motif. The deduced amino acid sequence analysis of segment B of Sp and VR-299 isolates showed 3 and 5 amino acid substitutions, respectively.; To develop a cost-effective subunit vaccine, which could be used to immunize fry by immersion, all the structural protein genes, (encoded by segment A) of ALV122 strain were expressed in insect cells and cabbage looper larvae, using a baculovirus expression system. High yields of IPNV-specific proteins (0.7 mg/larvae) were obtained. The expressed structural proteins were processed correctly, which self-assembled and formed virus-like particles (VLPs). These VLPs were used to immunize rainbow trout fry with high (0.5 mg/mL) and low (0.01 mg/mL) dose by immersion. A relative percentage survival (RPS) of 43% was recorded between the vaccinated and control, indicating a partial cross-protection in fish that were vaccinated with a high dose and challenged with VR-299 after two weeks. No protection was evident in fish that were vaccinated with a low dose and challenged after three weeks. The VLPs were also used to vaccinate Atlantic salmon post-smolts by injection with high (200 μg/fish) and low (100 μg/fish) dose. A high RPS of 56% was obtained in fish that were vaccinated with a high dose, whereas, only 22% RPS was recorded in fish vaccinated with a low dose. These results indicate that protection was dose-dependent and it correlated with the high IPNV antibody titers in the vaccinated fish. |
