A novel 3A-based indirect enzyme-linked immunosorbent assay for serological testing of Senecavirus A

Senecavirus A (SVA) can cause a contagious porcine disease characterized by vesicular lesions on the mouth and feet. Its clinical symptoms are extremely similar to those of porcine foot-and-mouth disease, swine vesicular disease, and vesicular stomatitis, making it difficult to distinguish in clinical diagnosis. SVA can infect pigs at all stages and cause acute death of newborn piglets, causing serious economic losses to the pig industry. In this study, a novel indirect enzyme-linked immunosorbent assay (ELISA) was established based on the SVA 3A nonstructural protein, which was equally sensitive to the immunofluorescence assay (IFA). In addition, the indirect ELISA showed no cross-reaction with positive sera of porcine reproductive and respiratory syndrome virus, classical swine fever virus, porcine circovirus type 2, pseudorabies virus, porcine astrovirus, African swine fever virus, and porcine epidemic diarrhea virus, exhibiting good specificity. The maximum serum sample dilution for detecting SVA-positive serum sample is 1:4,096. The coefficients of variation obtained from both intra-assay and inter-assay repeated experiments were less than 5%. Additionally, 173 porcine serum samples were tested using both the indirect ELISA method and commercial kits, with immunofluorescence analysis (IFA) employed as an auxiliary verification method to further confirm the accuracy of the test results. The results showed that the compliance rate of the indirect ELISA (100%) was higher than that of the commercially available ELISA kit (89.60%). The indirect ELISA established in this study showed more efficient and reliable performance for SVA serological testing, making it more suitable for long-term monitoring and prevention of the SVA epidemic.

SVA mainly causes vesicular lesions in pigs, bringing economic losses to the porcine industry. Currently, no commercial vaccine is available, so effective diagnostic methods are vital for the prevention and control of the disease. Serological testing, as a diagnostic method to detect specific antibodies against pathogens, can also be used to confirm viral infection in non-vaccinated animals. Previous studies have shown that in Senecavirus A (SVA)-infected pigs, the antibody level of the 3AB protein is significantly higher than the structural VP1 protein. Considering the molecular weight of the 3B protein was only 2.4 kDa, it is speculated that the 3A protein is a potential diagnostic marker for SVA serological testing. In this study, a novel 3A-based indirect enzyme-linked immunosorbent assay (ELISA) was established, showing a 100.0% coincident rate compared with immunofluorescence assay for clinical serum. Since most studies of vaccine development focus on structural proteins, the 3A-based ELISA also provides a potential tool for distinguishing between infected and vaccinated animals.