Immunoprotective activity of a Salmonid Alphavirus Vaccine: comparison of the immune responses induced by inactivated whole virus antigen formulations based on CpG class B oligonucleotides and poly I:C alone or combined with an oil adjuvant.

CpG oligonucleotides and polyinosinic:polycytidylic acid (poly I:C) are toll-like receptor (TLR) agonists that mimic the immunostimulatory properties of bacterial DNA and double-stranded viral RNA respectively, and which have exhibited potential to serve as vaccine adjuvants in previous experiments. Here, a combination of CpGs and poly I:C together with water- or oil-formulated Salmonid Alphavirus (SAV) antigen preparations has been used for a vaccine in Atlantic salmon and tested for protection in SAV challenge trial. The results demonstrate that vaccination with a high dose of the SAV antigen induced protection against challenge with SAV which correlated with production of neutralizing antibodies (NAbs). As the high antigen dose alone induced full protection, no beneficial effect from the addition of CpG and poly I:C could be observed. Nevertheless, these TLR ligands significantly enhanced the levels of NAbs in serum of vaccinated fish. Interestingly, gene expression analysis demonstrated that while addition of oil suppressed the CpG/poly I:C-induced expression of IFN-γ, the upregulation of IFNa1 was substantially enhanced. A low dose of the SAV antigen combined with oil did not induce any detectable levels of NAbs either with or without TLR ligands present, however the addition of CpG and poly I:C to the low SAV antigen dose formulation significantly enhanced the protection against SAV suggesting that CpG/poly I:C may have enhanced a cytotoxic response - a process which is dependent on the up-regulation of type I IFN. These results highlight the immunostimulatory properties of the tested TLR ligands and will serve as a ground for further, more detailed studies aimed to investigate their capacity to serve as adjuvants in vaccine formulations for Atlantic salmon.