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

Abstract

Classical swine fever virus (CSFV) is a major animal pathogen threatening the global pork industry. To date, numerous anti-CSFV monoclonal antibodies (mAbs) and their recognizing epitopes have been reported. However, few mAbs were systematically characterized for the capacity to differentiate field CSFV isolates from CSF vaccine strains, and the molecular basis associated with antigenic differences between vaccines and field isolates is still largely unknown. In the present study, recombinant CSFV structural glycoproteins E2 of both virulent and vaccine strains and Erns of vaccine strain were expressed using eukaryotic cells and murine mAbs generated against E2 and Erns. After serial screening and cloning of the hybridomas, the viral spectra of mAbs were respectively determined 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 these mAbs were characterized by epitope mapping using truncated, chimeric, and site-directed mutated E2 and Erns proteins. We have identified two vaccine-specific, one field isolate-specific, and two universal CSFV-specific mAbs and five novel conformational epitopes with critical amino acid (aa) motifs that are associated with these five mAbs: 213EPD215, 271RXGP274, and 37LXLNDG42 on E2 and 38CKGVP42, W81, and D100/V107 on Erns. Particularly, E213 of E2 is field isolate-specific, while N40 of E2 and D100/V107 of Erns are vaccine strain-specific. Results from our study further indicate that N40D of E2 mutation in field strains was likely produced under positive selection associated with long-term mass vaccination, leading to CSFV evasion of host immune response. Taking together, this study provides new insights into the antigenic structure of CSFV E2 and Erns and the differentiating mAbs will contribute to the development of a diagnostic strategy to differentiate C-strain vaccination from natural infection (DIVA) of CSFV in terms of elimination of CSF in China.

A Novel Competitive ELISA for Specifically Measuring and Differentiating Immune Responses to Classical Swine Fever C-Strain Vaccine in Pigs

“…Indirect fluorescent antibody assay (IFA) testing further confirmed that mAb 1504 lacks reaction to CSFV field isolates (genotypes 1.1, 2.1a, 2.1b, 2.1c, 2.1g, 2.1h, 2.1i, 2.1j, 2.2 and 2.3). Detailed data are described in our article for characterization of mAbs that specifically differentiate field isolates from vaccine strains of CSFV [ 25 ].…”

Editorial: Advances and insights in the diagnosis of viral infections and vaccines development in animals

“…Clinical signs and pathological findings in viral infected animals are highly variable due to both viral and host factors, and often confused with other diseases of animals (Murcia et al, 2009 ; Wang et al, 2020 ). Well-established laboratory techniques like virus isolation, real-time polymerase chain reaction (RT-PCR), and enzyme-linked immunosorbent assay (ELISA) have improved diagnostic accuracy (Wang et al, 2020 , 2022 ; Mi et al, 2022 ; Azam et al, 2023 ). Recent advancements have taken this progress a step further by offering faster, more efficient methods.…”

Correlation of the classical swine fever (CSF) antibody levels detected by serum neutralizaton and enzyme-linked immunosorbent assay (Elisa)

“…Recent study has discovered five monoclonal antibodies (mAb) against Erns, there are 1104, 1204, 1504, 1904, and 2004(Mi et al, 2022. In addition, the study has reported 8 epitopes in Erns protein successfully mapped by mAbs, there are 31 GIWPEKIC 38 , 65 NYTCCKLQ 72 , 73 RHEWNKHGW 81 , 88 DPWIQLMNR 96 , 116 YDKNTDVNV 124 , serum neutralization.…”

“…The broadly CSFV epitope 38 CKGVP 42 ,W 81 that recognized by mAb 1204 is completely conservative among different sub-genotypes. On the other hand, the mAb 1104 group has recognized highly c-strain-vaccine specific epitope with the critical determinant indicated by the amino acid combination of D 100 /V 107 , therefore, the elicits-mAb is able to differentiate vaccine strains from field isolates(Mi et al, 2022).Detection of CSFV-Erns antibodies can be conducted by antibody-based assay including ELISA coated with specific capture protein, western blot, SDS-page, or…”