Production of IFN-γ in feline whole blood after incubation with potential T-cell epitopes of the nucleocapsid protein of feline coronavirus

Interferon gamma (IFN-γ) plays an important role in cell mediated responses against mutated feline coronavirus strains (FCoV) involved in the pathogenesis of feline infectious peritonitis (FIP). The aim of this study was to establish a combined in silico and in vitro approach to assess feline leukocyte production of IFN-γ in response to selected peptides of the nucleocapside protein (N) of FCoVs. To this aim, we designed, through a bioinformatic approach, 8 potentially immunogenic peptides from the protein N corresponding to sequences of residues 14, 182, 198 detected only in FCoVs from FIP cats (virulent strains), only in FCoVs from healthy cats (avirulent strains) and both in FIP and in healthy cats (mixed strains).The peptides or a sham solution were incubated with whole blood from 16 cats (7 healthy and 9 with chronic diseases other than FIP) and IFN-γ concentration was measured on plasma using an ELISA system. RT-PCR expression of IFN-γ mRNA was also evaluated after incubation of the peptides or a sham solution with whole blood from 4 clinically healthy cats. The mean plasma concentration of IFN-γ in samples incubated with peptides decreased and the expression of IFN-γmRNA did not change compared with the sham solution, except for some cats with chronic diseases (which probably have a "pre-activated" immune response). These cats responded to "avirulent" or "mixed" peptides by increasing the concentration of IFN-γ and the expression of IFN-γ mRNA. The combined approach employed in this study allowed us to identify potentially immunogenic peptides of FCoV N protein that can modulate the production of IFN-γ especially in cats with a "pre-activated" cell mediated response