title: Identification of vaccine candidate peptides in the NcSRS2 surface protein of Neospora caninum by using CD4+ cytotoxic T lymphocytes and gamma interferon-secreting T lymphocytes of infected holstein cattle.
abstract: Previously, our laboratory showed that Holstein cattle experimentally infected with Neospora caninum develop parasite-specific CD4+ cytotoxic T lymphocytes (CTL) that lyse infected, autologous target cells through a perforin-granzyme pathway. To identify specific parasite antigens inducing bovine CTL and helper T-lymphocyte responses for vaccine development against bovine neosporosis, the tachyzoite major surface proteins NcSAG1 and NcSRS2 were targeted. In whole tachyzoite antigen-expanded bovine T-lymphocyte lines, recombinant NcSRS2 induced potent memory CD4+- and CD8+-T-lymphocyte activation, as indicated by proliferation and gamma interferon (IFN-gamma) secretion, while recombinant NcSAG1 induced a minimal memory response. Subsequently, T-lymphocyte epitope-bearing peptides of NcSRS2 were mapped by using overlapping peptides covering the entire NcSRS2 sequence. Four experimentally infected cattle with six different major histocompatibility complex (MHC) class II haplotypes were the source of immune cells used to identify NcSRS2 peptides presented by Holstein MHC haplotypes. NcSRS2 peptides were mapped by using IFN-gamma secretion by rNcSRS2-stimulated, short-term T-lymphocyte cell lines, IFN-gamma enzyme-linked immunospot (ELISPOT) assay with peripheral blood mononuclear cells, and 51Cr release cytotoxicity assay of rNcSRS2-stimulated effector cells. Four N. caninum-infected Holstein cattle developed NcSRS2 peptide-specific T lymphocytes detected ex vivo in peripheral blood by IFN-gamma ELISPOT and in vitro by measuring T-lymphocyte IFN-gamma production and cytotoxicity. An immunodominant region of NcSRS2 spanning amino acids 133 to 155 was recognized by CD4+ T lymphocytes from the four cattle. These findings support investigation of subunit N. caninum vaccines incorporating NcSRS2 gene sequences or peptides for induction of NcSRS2 peptide-specific CTL and IFN-gamma-secreting T lymphocytes in cattle with varied MHC genotypes.
authors.affiliation: Department of Veterinary Microbiology and Pathology, Bustad Hall, Washington State University, Pullman, WA 99164-7040, USA.
authors.forename: Lauren M
authors.initials: LM
authors.lastname: Staska
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Davies
authors.forename: Wendy C
authors.initials: WC
authors.lastname: Brown
authors.forename: Travis C
authors.initials: TC
authors.lastname: McGuire
authors.forename: Carlos E
authors.initials: CE
authors.lastname: Suarez
authors.forename: Joo Youn
authors.initials: JY
authors.lastname: Park
authors.forename: Bruce A
authors.initials: BA
authors.lastname: Mathison
authors.forename: Jeffrey R
authors.initials: JR
authors.lastname: Abbott
authors.forename: Timothy V
authors.initials: TV
authors.lastname: Baszler
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000737:chemistry* / Q000276:immunology; D000954:Antigens, Surface / Q000737:chemistry* / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D002417:Cattle; D002418:Cattle Diseases / Q000276:immunology / Q000469:parasitology; D002460:Cell Line; D003048:Coccidiosis / Q000276:immunology / Q000469:parasitology / Q000662:veterinary; D018604:Epitope Mapping; D018984:Epitopes, T-Lymphocyte; D005260:Female; D007371:Interferon-gamma / Q000378:metabolism*; D008969:Molecular Sequence Data; D018688:Neospora / Q000276:immunology*; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D015800:Protozoan Proteins / Q000737:chemistry* / Q000276:immunology; D016052:Protozoan Vaccines / Q000737:chemistry / Q000276:immunology*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
chemical_list: D000953:Antigens, Protozoan; D000954:Antigens, Surface; D018984:Epitopes, T-Lymphocyte; D010455:Peptides; D015800:Protozoan Proteins; D016052:Protozoan Vaccines; C115389:SRS2 protein, Neospora caninum; D007371:Interferon-gamma
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