title: Two-way antigenic cross-reactivity between severe acute respiratory syndrome coronavirus (SARS-CoV) and group 1 animal CoVs is mediated through an antigenic site in the N-terminal region of the SARS-CoV nucleoprotein.
abstract: In 2002, severe acute respiratory syndrome-associated coronavirus (SARS-CoV) emerged in humans, causing a global epidemic. By phylogenetic analysis, SARS-CoV is distinct from known CoVs and most closely related to group 2 CoVs. However, no antigenic cross-reactivity between SARS-CoV and known CoVs was conclusively and consistently demonstrated except for group 1 animal CoVs. We analyzed this cross-reactivity by an enzyme-linked immunosorbent assay (ELISA) and Western blot analysis using specific antisera to animal CoVs and SARS-CoV and SARS patient convalescent-phase or negative sera. Moderate two-way cross-reactivity between SARS-CoV and porcine CoVs (transmissible gastroenteritis CoV [TGEV] and porcine respiratory CoV [PRCV]) was mediated through the N but not the spike protein, whereas weaker cross-reactivity occurred with feline (feline infectious peritonitis virus) and canine CoVs. Using Escherichia coli-expressed recombinant SARS-CoV N protein and fragments, the cross-reactive region was localized between amino acids (aa) 120 to 208. The N-protein fragments comprising aa 360 to 412 and aa 1 to 213 reacted specifically with SARS convalescent-phase sera but not with negative human sera in ELISA; the fragment comprising aa 1 to 213 cross-reacted with antisera to animal CoVs, whereas the fragment comprising aa 360 to 412 did not cross-react and could be a potential candidate for SARS diagnosis. Particularly noteworthy, a single substitution at aa 120 of PRCV N protein diminished the cross-reactivity. We also demonstrated that the cross-reactivity is not universal for all group 1 CoVs, because HCoV-NL63 did not cross-react with SARS-CoV. One-way cross-reactivity of HCoV-NL63 with group 1 CoVs was localized to aa 1 to 39 and at least one other antigenic site in the N-protein C terminus, differing from the cross-reactive region identified in SARS-CoV N protein. The observed cross-reactivity is not a consequence of a higher level of amino acid identity between SARS-CoV and porcine CoV nucleoproteins, because sequence comparisons indicated that SARS-CoV N protein has amino acid identity similar to that of infectious bronchitis virus N protein and shares a higher level of identity with bovine CoV N protein within the cross-reactive region. The TGEV and SARS-CoV N proteins are RNA chaperons with long disordered regions. We speculate that during natural infection, antibodies target similar short antigenic sites within the N proteins of SARS-CoV and porcine group 1 CoVs that are exposed to an immune response. Identification of the cross-reactive and non-cross-reactive N-protein regions allows development of SARS-CoV-specific antibody assays for screening animal and human sera.
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, Ohio Agricultural Research and Development Center, The Ohio State University, 1680 Madison Avenue, Wooster, OH 44691-4096, USA.
authors.forename: Anastasia N
authors.initials: AN
authors.lastname: Vlasova
authors.forename: Xinsheng
authors.initials: X
authors.lastname: Zhang
authors.forename: Mustafa
authors.initials: M
authors.lastname: Hasoksuz
authors.forename: Hadya S
authors.initials: HS
authors.lastname: Nagesha
authors.forename: Lia M
authors.initials: LM
authors.lastname: Haynes
authors.forename: Ying
authors.initials: Y
authors.lastname: Fang
authors.forename: Shan
authors.initials: S
authors.lastname: Lu
authors.forename: Linda J
authors.initials: LJ
authors.lastname: Saif
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D000956:Antigens, Viral / Q000097:blood / Q000276:immunology*; D002415:Cats; D002417:Cattle; D002460:Cell Line; D017934:Coronavirus / Q000145:classification / Q000276:immunology*; D018352:Coronavirus Infections / Q000662:veterinary* / Q000821:virology; D000086462:Coronavirus Nucleocapsid Proteins; D003429:Cross Reactions; D004285:Dogs; D006168:Guinea Pigs; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D019590:Nucleocapsid Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D045473:Severe acute respiratory syndrome-related coronavirus / Q000276:immunology*; D045169:Severe Acute Respiratory Syndrome / Q000821:virology
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D000086462:Coronavirus Nucleocapsid Proteins; D007106:Immune Sera; D019590:Nucleocapsid Proteins