title: Genetic and antigenic characterization of recombinant nucleocapsid proteins derived from canine coronavirus and canine respiratory coronavirus in China.
abstract: To characterize the antigenicity of nucleocapsid proteins (NP) derived from canine coronavirus (CCoV) and canine respiratory coronavirus (CRCoV) in China, the N genes of CCoV (CCoV-BJ70) and CRCoV (CRCoV-BJ202) were cloned from swabs obtained from diseased pet dogs in Beijing and then sequenced. The recombinant NPs (rNPs) were expressed in Escherichia coli and purified by nickel-affinity column and size exclusion chromatography. Sequencing data indicated that the N genes of CCoV-BJ70 and CRCoV-BJ202 belonging to two distinctly different groups were relatively conserved within each subgroup. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) results showed that rNPs of CCoV and CRCoV were expressed efficiently and isolated with a final purity of over 95%. Western blot analysis revealed the rNP from CRCoV could cross-react with mice antisera against human coronavirus (HCoV-229E, NL63, OC43, HKU1), while rNP of CCoV had cross-reactivity with only anti-sera against viruses belonging to the same group (HCoV-229E and NL63). In summary, CCoV and CRCoV rNPs were successfully expressed in E. coli and showed antigenic cross-reactivity with antisera raised against human coronaviruses. These findings indicate that further serologic studies on coronavirus infections at the animal-human interface are needed.
authors.affiliation: Institute of Medical Virology, Wenzhou Medical University, Wenzhou, 325035, China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Lu
authors.affiliation: Institute of Medical Virology, Wenzhou Medical University, Wenzhou, 325035, China.
authors.forename: Yingzhu
authors.initials: Y
authors.lastname: Chen
authors.affiliation: National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, 102206, China.
authors.forename: Kun
authors.initials: K
authors.lastname: Qin
authors.affiliation: National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, 102206, China.
authors.forename: Jianfang
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Institute of Medical Virology, Wenzhou Medical University, Wenzhou, 325035, China.
authors.forename: Yongliang
authors.initials: Y
authors.lastname: Lou
authors.affiliation: Institute of Medical Virology, Wenzhou Medical University, Wenzhou, 325035, China. tanwj28@163.com.|National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, 102206, China. tanwj28@163.com.
authors.forename: Wenjie
authors.initials: W
authors.lastname: Tan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000956:Antigens, Viral / Q000276:immunology; D015153:Blotting, Western; D002681:China; D003001:Cloning, Molecular; D017939:Coronavirus, Canine / Q000235:genetics* / Q000276:immunology*; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D005814:Genes, Viral; D019590:Nucleocapsid Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid
chemical_list: D000956:Antigens, Viral; D019590:Nucleocapsid Proteins
keywords: canine coronavirus; canine respiratory coronavirus; cross-reactivity; expression; nucleocapsid protein