Novel BPI3Vc-vectored surface displayed fusion and hemagglutinin-neuraminidase antigens elicit broadly neutralizing antibodies in cattle

Bovine parainfluenza-3 virus (BPI3V) contributes to Bovine Respiratory Disease Complex, causing severe pneumonia and death in cattle, leading to economic losses. Existing BPI3V commercial vaccines, based on genotype A strains, confer protection against some, but not all, genotype A strains and induce low neutralizing antibody titers against genotypes B and C. This study aimed to develop a live vaccine capable of inducing broad protection against diverse BPI3V strains using an attenuated BPI3V vaccine vector based on a genotype C strain. A rescued recombinant BPI3Vc mutant GFP virus exhibited a temperature-sensitive attenuated phenotype in vitro . Novel Fusion (designated F2) and Hemagglutinin-Neuraminidase (designated HN2) antigens, derived from consensus protein sequences of BPI3V genotypes A, B, and C, were used to develop a recombinant prototype vaccine, designated rBPI3Vc mut F2-HN2. The recombinant virus replicated efficiently, displayed the novel antigens on the surface of infected cells, and remained stable over nine in vitro passages. Intranasal vaccination of calves with the rBPI3Vc mut F2-HN2 virus induced strong systemic and mucosal IgG responses against BPI3V genotypes A, B, and C, which were significantly amplified upon boost, unlike the responses elicited by a commercial vaccine. Notably, sera from calves vaccinated with the rBPI3Vc mut F2-HN2 virus had significantly higher (p<0.0001) neutralizing antibodies against BPI3V genotypes A-C compared to the commercial vaccine. The F2-HN2 antigens were critical in eliciting neutralizing antibodies against wild-type BPI3Va and c (p< 0.0001), and BPI3Vb (p<0.001). Upon challenge with wild-type BPI3V genotype C virus, the rBPI3Vc mut F2-HN2-vaccinated calves shed the least amount of virus in nasal swabs, had lower viremia, and exhibited minimal pulmonary lesions. Therefore, rBPI3Vc mut F2-HN2 virus is a promising vaccine candidate that has potential to confer broad protection against multiple BPI3V strains.