Evaluation of a quintuple-gene-deleted PRV vector expressing PEDV S1: safety and immunogenicity in rabbits, mice, and...

Pseudorabies virus (PRV) and Porcine epidemic diarrhea virus (PEDV) are currently co-circulation among swine herd with multiple variant strains, which cause severe economic losses to the global swine industry. In the present study, a recombinant PRV-based vaccine candidate was constructed backbone of a PRV variant via deletion of five genes (gI, gE, US2, US9, and TK) and insertion of the S1 gene derived from a PEDV G2b strain, and the resulting recombinant virus was generated as rPRV-Δ5-S1. The safety of the quintuple-gene-deleted PRV vector (rPRV-Δ5) was firstly evaluated in rabbits and compared with the previously generated triple-gene-deleted vector (rPRV-Δ3). Rabbits inoculated with rPRV-Δ5 exhibited milder clinical manifestations, lower viral loads in tissues, and fewer histopathological lesions, indicating an improved safety profile. Subsequent immunization trials in mice demonstrated that rPRV-Δ5-S1 induced PEDV S1-specific antibodies and PEDV-neutralizing antibodies, which were higher than those observed in the commercial inactivated PEDV vaccine group. Furthermore, rPRV-Δ5-S1 elicited higher neutralizing antibody titers against PRV variants than the Bartha-K61 vaccine and provided complete protection against lethal challenge with the PRV variant. To further validate the immunogenicity and protective efficacy of rPRV-Δ5-S1, challenge experiments were conducted in piglets. The results showed that piglets immunized with rPRV-Δ5-S1 developed detectable antibody responses against both PRV and PEDV. After challenge with the PEDV G2b variant, rPRV-Δ5-S1-immunized piglets exhibited reduced diarrhea severity, decreased viral shedding, alleviated intestinal lesions, and improved weight gain compared with control animals. Collectively, these findings demonstrate that the quintuple-gene-deleted PRV vector exhibits an improved safety profile and can serve as an effective platform for heterologous antigen delivery. The recombinant virus rPRV-Δ5-S1 can induced immune responses against both PRV and PEDV in multiple animal models, which supports its potential as a bivalent vaccine candidate against these two economically critical swine pathogens.