AUTHOR=Mi Shijiang , Wang Lihua , Li Hongwei , Bao Fei , Madera Rachel , Shi Xiju , Zhang Liying , Mao Yingying , Yan Renhe , Xia Xianzhu , Gong Wenjie , Shi Jishu , Tu Changchun 

TITLE=Characterization of monoclonal antibodies that specifically differentiate field isolates from vaccine strains of classical swine fever virus

JOURNAL=Frontiers in Immunology

VOLUME=Volume 13 - 2022

YEAR=2022

URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2022.930631

DOI=10.3389/fimmu.2022.930631

ISSN=1664-3224

ABSTRACT=Classical swine fever virus (CSFV) is a major animal pathogen threatening pig industry in the world. Up to date, numerous monoclonal antibodies (mAbs) against CSFV and their recognizing epitopes on the viral antigenic proteins have been reported, but few of them were systematically characterized for the capacity of differentiation, and molecular basis associated with antigenic differences between vaccines and field isolates are still largely unknown. In present study CSFV structural glycoproteins E2 of both virulent and vaccine strains and Erns of vaccine strain were expressed in eukaryotic cells and used to generate mAbs in mice. After serial screening and cloning of the hybridomas, the viral spectrum recognized by the mAbs were respectively identified by indirect fluorescent antibody assay (IFA) using 108 CSFVs, followed by western blot analysis using expressed glycoproteins of all CSFV sub-genotypes including vaccine strains. The antigenic structures recognized by the mAbs were characterized by epitope mapping using truncated, chimeric and site-directed mutated E2 and Erns proteins. Results showed that two vaccine-specific, one field isolate-specific and two universal CSFV-specific mAbs have been identified, and correspondingly 5 novel conformational epitopes have been defined with critical aa motifs being 213EPD215, 271RXGP274 and 37LXLNDG42 on E2, and 38CKGVP42,W81 and D100/V107 on Erns. Particularly noted is the identification of E213 of E2 as field isolate-specific amino acid (aa), while N40 of E2 and D100/V107 of Erns are vaccine-specific amino acids. The study further found that N40D field mutation on E2 was very likely produced under positive selection associated with long term mass vaccination, leading CSFV evasion of host immune response. Thereby D40 of E2 can be deemed as the marker residue confirming CSFV immune evasion under positive selection. Taking together, this study provides new insights into the antigenic structure of CSFV major glycoproteins and the differentiating mAbs will contribute to the development of diagnostic strategy to differentiate C-strain vaccination from natural infection of CSFV in terms of elimination of the disease in China.