A novel porcine interleukin-2-interferon-α-thymosin α1 tripartite fusion protein confers antiviral activity against pseudorabies virus.

Pseudorabies virus (PRV), the causative agent of Aujeszky's disease in pigs, threatens swine production and can infect other mammals, with rare zoonotic cases reported. Effective antiviral agents against PRV remain lacking. Here, we constructed and purified a recombinant fusion protein combining porcine Interleukin-2 (IL-2), interferon α (IFN-α), and Thymosin α1 (Tα1). The fusion protein showed no cytotoxicity or induction of apoptosis in PK-15 cells. In vitro, it significantly enhanced peripheral blood mononuclear cell (PBMC) metabolic activity and upregulated expression of key interferon-stimulated genes (ISGs) involved in antiviral defense, including IFIT1, ISG15, MX1, and OAS, which are critical for antiviral activity. Analysis of the viral life cycle indicated that Interleukin-2-Interferon-α-Thymosin α1 (IL-2-IFN-α-Tα1) inhibited PRV replication without affecting viral attachment, entry, assembly, or egress. Treatment with the fusion protein conferred marked therapeutic efficacy against PRV infection, including improved survival, reduced viral loads in the brain and spleen, and alleviated inflammatory pathology. These results demonstrate that IL-2-IFN-α-Tα1 fusion protein has potential as a novel immunomodulatory agent for controlling PRV infection in swine, offering a promising strategy for antiviral intervention in veterinary medicine.