Engineering a recombinant VP2-Based neutralizing epitope vaccine candidate against canine parvovirus: a preliminary immunogenicity assessment.

Background: Canine parvovirus (CPV) poses a severe threat to canine health, necessitating the development of safer and more effective vaccines. While traditional vaccines carry risks of virulence reversion and environmental contamination, subunit vaccines-especially neutralizing epitope vaccines-offer promising alternatives by eliciting targeted immune responses with enhanced safety.

Methods: We employed bacterial display technology to express 11 overlapping CPV VP2 gene fragments on the periplasmic membrane of E. coli. Key neutralizing antigenic epitopes of CPV VP2 were mapped using four neutralizing single-chain antibodies in combination with FCM. The identified epitopes were concatenated via GS linkers and expressed as a recombinant VP2D protein. Then, animal experiments were conducted to evaluate the immunogenicity of the epitope-based vaccine in mice.

Results: The data shows that the antigenic epitopes of the four single-chain antibodies are located in V2-V3, V8, V10 and V11 respectively. After linking five antigenic epitope fragments, we constructed a novel chimeric antigen protein, VP2D, and achieved the efficient expression of VP2D. Animal immunization evaluation results demonstrated that the VP2D vaccine exhibited superior immunological properties compared to both the VP2 vaccine and the commercial vaccine. The VP2D vaccine significantly enhanced the host's capacity to mount both humoral and cellular immune responses. Notably, the antibody titers in the VP2D vaccine group were consistently 1.2- to 1.5-fold higher than those in the VP2 vaccine group throughout the experimental period. Furthermore, the proportions of CD4⁺ T cells and F4/80⁺ cells were increased by up to 12% and 8%, respectively, in the VP2D group. Additionally, both the protein levels and mRNA transcription of inflammatory cytokines were elevated in the VP2D vaccine group relative to both the VP2 and commercial vaccine groups, indicating a consistently enhanced inflammatory response.

Conclusions: Identified CPV-VP2 neutralizing antigenic epitopes (70-150, 410-440, 510-585 aa), and an effective candidate vaccine for the prevention of CPV was provided.