Revealing novel CD8⁺ T-cell epitopes from the H5N1 avian influenza virus in HBW/B1 haplotype ducks

The duck CD8⁺ T-cell response effectively defends against H5N1 highly pathogenic avian influenza virus (HPAIV) infection, but the recognized peptide is rarely identified. Here, we found that the ratio of CD8⁺ T cells and the expression of IFN-γ and cytotoxicity-associated genes, including granzyme A/K, perforin and IL2, at 7 days post-infection in peripheral blood mononuclear cells (PBMCs) from B1 haplotype ducks significantly increased in the context of defending against H5N1 AIV infection in vivo. Moreover, similar results were observed in cultured and sorted H5N1 AIV-stimulated duck CD8⁺ T cells in vitro. Next, we selected 109 epitopes as candidate epitopes on the basis of the MHC-I restriction binding peptide prediction website database and further identified twelve CD8⁺ T-cell epitopes that significantly increased IFN-γ gene expression after stimulating B1 haplotype duck memory PBMCs. In particular, NP_338-346, NP_473-481, M_2-10, PB1_540-548 and PA_80-88 were highly conserved in H5N1, H5N6, H5N8, H7N9, and H9N2 AIVs. These findings provide directions for the development of universal T-cell epitope vaccines for AIV in ducks.