Construction and Evaluation of Recombinant Pseudorabies Virus Expressing African Swine Fever Virus Antigen Genes

Abstract

African swine fever (ASF) is a highly contact infectious disease caused by the African swine fever virus (ASFV). The extremely complex structure and infection mechanism make it difficult to control the spread of ASFV and develop the vaccine. The ASFV genome is huge with many antigenic genes. Among them, CP204L (p30), CP530R (pp62), E183L (p54), B646L (p72), and EP402R (CD2v) are involved in the process of the virus cycle, with strong immunogenicity and the ability to induce the body to produce neutralizing antibodies. In this study, the recombinant virus rBartha-K61-pASFV that expresses the above ASFV antigen genes was constructed by Red/ET recombineering technology using pseudorabies virus (PRV) vaccine strain Bartha-K61. Western blot analysis showed that the ASFV antigen gene was expressed and the recombinant virus showed good genetic stability and proliferation characteristics in 15 continuous generations on porcine kidney (PK15) cells. The results of immunoassay of piglets and mice showed that rBartha-K61-pASFV had good immunogenicity and could induce higher antibody levels in the body. Therefore, PRV was a promising viral vector for expressing the ASFV antigen gene, and all the experiments in this study laid a foundation for the further development of a new viral vector vaccine of ASFV.

An Intracellular Epitope of ASFV CD2v Protein Elicits Humoral and Cellular Immune Responses

“…Indeed, modified vaccinia virus Ankara (MVA) expressing CD2v, P72, and EP153R induced significant cellular immunity characterized by T cell proliferation and IFN-γ production [ 37 ]. For rBartha-K61-pASFV expressing CD2v, P72, P30, pp62, and P54, high production of antibodies was observed in piglets and mice [ 38 ]. Importantly, a baculovirus-based expression vector expressing the extracellular domains of CD2v, P54, and P30 induced specific T-cell immunity directly after in vivo immunization and partially protected pigs against a sublethal attack with ASFV [ 39 ].…”

Vaccines for African swine fever: an update

“…Pigs immunized and exposed to Benin97/1 using recombinant poxvirus for primary immunization and recombinant adenovirus for booster immunization represent the best combination of viral vector antigens for immunization to date ( Freitas et al, 2019 ). Chen et al assessed the immunoassay of piglets and mice using the recombinant virus rBartha-K61-pASFV ( Chen et al, 2022 ). Murgia et al used an alphavirus vector platform to deliver replicon particles (RPs) expressing ASFV antigens to swine ( Murgia et al, 2019 ).…”

Summary of the Current Status of African Swine Fever Vaccine Development in China

“…Once identified, the ASFV antigenic proteins were combined with other bacterial/viral proteins or functional proteins [49,50], so that the expressed ASFV antigenic proteins could self-assemble into a certain structure or improve the expression efficiency to improve the immunogenicity of the ASFV proteins. For the live vector vaccines, different vectors [51][52][53][54][55] and multiple gene expressions [56][57][58] were also evaluated. However, the major challenges to the development of the ASF subunit or live vector vaccines in China remain, with most of the candidates awaiting protective immunity validation and patent authorization.…”