title: Proteome-wide screening reveals immunodominance in the CD8 T cell response against classical swine fever virus with antigen-specificity dependent on MHC class I haplotype expression.
abstract: Vaccination with live attenuated classical swine fever virus (CSFV) vaccines induces a rapid onset of protection which has been associated with virus-specific CD8 T cell IFN-γ responses. In this study, we assessed the specificity of this response, by screening a peptide library spanning the CSFV C-strain vaccine polyprotein to identify and characterise CD8 T cell epitopes. Synthetic peptides were pooled to represent each of the 12 CSFV proteins and used to stimulate PBMC from four pigs rendered immune to CSFV by C-strain vaccination and subsequently challenged with the virulent Brescia strain. Significant IFN-γ expression by CD8 T cells, assessed by flow cytometry, was induced by peptide pools representing the core, E2, NS2, NS3 and NS5A proteins. Dissection of these antigenic peptide pools indicated that, in each instance, a single discrete antigenic peptide or pair of overlapping peptides was responsible for the IFN-γ induction. Screening and titration of antigenic peptides or truncated derivatives identified the following antigenic regions: core₂₄₁₋₂₅₅ PESRKKLEKALLAWA and NS3₁₉₀₂₋₁₉₁₂ VEYSFIFLDEY, or minimal length antigenic peptides: E2₉₉₆₋₁₀₀₃ YEPRDSYF, NS2₁₂₂₃₋₁₂₃₀ STVTGIFL and NS5A₃₀₇₀₋₃₀₇₈ RVDNALLKF. The epitopes are highly conserved across CSFV strains and variable sequence divergence was observed with related pestiviruses. Characterisation of epitope-specific CD8 T cells revealed evidence of cytotoxicity, as determined by CD107a mobilisation, and a significant proportion expressed TNF-α in addition to IFN-γ. Finally, the variability in the antigen-specificity of these immunodominant CD8 T cell responses was confirmed to be associated with expression of distinct MHC class I haplotypes. Moreover, recognition of NS₁₂₂₃₋₁₂₃₀ STVTGIFL and NS3₁₉₀₂₋₁₉₁₂ VEYSFIFLDEY by a larger group of C-strain vaccinated animals showed that these peptides could be restricted by additional haplotypes. Thus the antigenic regions and epitopes identified represent attractive targets for evaluation of their vaccine potential against CSFV.
authors.affiliation: Virology Department, Animal Health and Veterinary Laboratories Agency, Addlestone, United Kingdom ; Department of Microbial & Cellular Sciences, University of Surrey, Guildford, United Kingdom.
authors.forename: Giulia
authors.initials: G
authors.lastname: Franzoni
authors.affiliation: Virology Department, Animal Health and Veterinary Laboratories Agency, Addlestone, United Kingdom ; Nagpur Veterinary College, Maharashtra Animal & Fishery Sciences University, Nagpur, India.
authors.forename: Nitin V
authors.initials: NV
authors.lastname: Kurkure
authors.affiliation: Department of Pathobiology, University of Veterinary Medicine Vienna, Vienna, Austria.
authors.forename: Sabine E
authors.initials: SE
authors.lastname: Essler
authors.affiliation: Virology Department, Animal Health and Veterinary Laboratories Agency, Addlestone, United Kingdom.
authors.forename: Miriam
authors.initials: M
authors.lastname: Pedrera
authors.affiliation: Virology Department, Animal Health and Veterinary Laboratories Agency, Addlestone, United Kingdom.
authors.forename: Helen E
authors.initials: HE
authors.lastname: Everett
authors.affiliation: Department of Microbial & Cellular Sciences, University of Surrey, Guildford, United Kingdom.
authors.forename: Kikki B
authors.initials: KB
authors.lastname: Bodman-Smith
authors.affiliation: Virology Department, Animal Health and Veterinary Laboratories Agency, Addlestone, United Kingdom.
authors.forename: Helen R
authors.initials: HR
authors.lastname: Crooke
authors.affiliation: Virology Department, Animal Health and Veterinary Laboratories Agency, Addlestone, United Kingdom.
authors.forename: Simon P
authors.initials: SP
authors.lastname: Graham
mesh_terms: D000595:Amino Acid Sequence; D000704:Analysis of Variance; D000818:Animals; D001483:Base Sequence; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D006692:Classical Swine Fever Virus / Q000276:immunology*; D017931:DNA Primers / Q000235:genetics; D058501:Enzyme-Linked Immunospot Assay; D000939:Epitopes; D018984:Epitopes, T-Lymphocyte / Q000235:genetics; D005434:Flow Cytometry; D005805:Genes, MHC Class I / Q000276:immunology*; D016056:Immunodominant Epitopes / Q000235:genetics*; D008969:Molecular Sequence Data; D040901:Proteomics / Q000379:methods; D017422:Sequence Analysis, DNA; D034421:Sus scrofa / Q000276:immunology*; D014611:Vaccination / Q000379:methods
chemical_list: D017931:DNA Primers; D000939:Epitopes; D018984:Epitopes, T-Lymphocyte; D016056:Immunodominant Epitopes