Design and immunoinformatics characterization of a novel multi-epitope vaccine rPRRSV-35N targeting NADC30-like and NADC34-like porcine reproductive and respiratory syndrome virus variants

BACKGROUND: Porcine reproductive and respiratory syndrome (PRRS), caused by the genetically diverse porcine reproductive and respiratory syndrome virus (PRRSV), remains a major economic burden to the global swine industry. Current commercial vaccines, including modified live virus (MLV) and inactivated vaccines, have shown limited effectiveness against newly emerging PRRSV variants, particularly NADC30-like and NADC34-like strains. There is therefore a need for improved vaccine strategies targeting these circulating strains. In this study, we used immunoinformatics approaches to design a multi-epitope vaccine candidate, rPRRSV-35N, against PRRSV NADC30-like and NADC34-like strains, based on antigenic profiling and prediction of T- and B-cell epitopes. RESULTS: This multi-epitope vaccine candidate integrates six cytotoxic T lymphocyte (CTL) epitopes, three helper T lymphocyte (HTL) epitopes, and seven linear B-cell epitopes (LBE). The epitopes were linked sequentially by flexible linkers. Porcine β-defensin-2 (PBD-2) and PADRE epitopes were incorporated to potentially improve immunogenicity. In silico prediction showed an antigenicity score of 0.6597 and a solubility probability of 0.530. It showed no toxicity or allergenicity, with a molecular weight of 36.64 kDa and a theoretical isoelectric point (pI) of 9.81. Molecular docking and normal mode analysis indicated stable interactions with TLR2 and TLR4 receptors. Immunoinformatics simulations further revealed that the candidate has the potential to induce both humoral and cellular immune responses. The construct was designed for in silico cloning into pET28a(+), and the plasmid was commercially synthesized. Following transformation of the synthetic plasmid into E. coli BL21(DE3), protein expression was verified by Western blot analysis. CONCLUSIONS: This study constructed the multi-epitope vaccine candidate rPRRSV-35N targeting NADC30-like and NADC34-like PRRSV strains. Its expressibility was preliminarily verified via molecular cloning and protein expression assays. Further in vivo studies are required to assess its protective efficacy.