Development of a Synthetic VP1 Protein Peptide-Based ELISA to Detect Antibodies Against Porcine Bocavirus Group 3.

Porcine bocavirus (PBoV), classified within the genus Bocaparvovirus, has been reported worldwide. PBoV has been divided into group 1, group 2, and group 3. PBoV group 3 (G3) viruses are the most prevalent in China. Currently, effective serological methods for the detection of antibodies against PBoV G3 are limited. In this study, we developed an indirect ELISA using a synthetic VP1 peptide designed on the basis of the conserved region of the PBoV VP1 protein as a coating antigen. Through matrix titration, the optimal coating concentration of the VP1 peptide (0.5 μg/mL), serum dilution (1:200), and working concentration of the secondary antibody (1:50,000) were determined. The cutoff value of this developed ELISA was set as 0.4239. Further investigations revealed that this developed ELISA had no cross-reactivity with positive serum antibodies against FMDV-O, FMDV-A, PRV, ASFV, SF, PCV2, PEDV, and TGEV. The detection limit of the method was a 1:1600 dilution of standard positive serum against PBoV G3. The coefficients of variation for both the intra- and interassay data were lower than 10%. A total of 1373 serum samples collected from 12 provinces in China between 2022 and 2023 were subjected to indirect ELISA. The results showed that 47.56% of the samples were PBoV G3 positive. These results reveal that peptide-based ELISA is a reliable and cost-effective method for detecting PBoV G3 antibodies. It also facilitates the investigation of the prevalence and distribution of PBoV G3.