Characterization of a novel porcine circovirus type 2a sequence and anti-viral evaluation of matrine and sanguinarine combined with network pharmacology and molecular docking.

Porcine circovirus type 2 (PCV2) remains an economically significant pathogen affecting global swine herds, with emerging genetic variants raising concerns about potential immune escape from existing PCV2a-based vaccines. In this study, we identified a novel PCV2a isolate carrying four nonsynonymous substitutions (S68A, M131T, V133A, and K191A) in the capsid (Cap) protein, with the M131T/V133A substitutions located within the well-characterized conformational neutralizing epitope at residues 132-137 (KATALT→KVPAQT). To assess the functional impact of this mutation on antibody recognition, we developed a liquid-phase competitive ELISA using synthetic peptides representing the wild-type (WT) and mutant epitope sequences, together with PCV2-positive reference serum. The WT peptide effectively competed for antibody binding (inhibition rate 89.40%), whereas the mutant peptide exhibited a markedly reduced inhibition rate (40.28%), indicating impaired antibody recognition at this epitope. Combined with three-dimensional structural modeling showing altered surface charge and spatial conformation within the epitope region, these findings provide direct evidence that the KVPAQT mutation reduces neutralizing antibody binding to this critical epitope. Network pharmacology and molecular docking were then applied to assess matrine and sanguinarine targeting the mutant Cap protein. Both compounds exhibited strong binding affinities, with binding energies of -8.2 and - 7.9 kcal/mol for matrine and sanguinarine, respectively. Furthermore, in vitro antiviral assays demonstrated that both compounds effectively suppressed PCV2 replication in PK-15 cells. Overall, these findings define a clinically isolated PCV2a antigenic variant with impaired antibody recognition and establish an integrated computational-experimental platform for efficiently screening natural products against emerging PCV2 variants. This study provides a molecular basis for understanding PCV2 antigenic drift and a practical strategy for antiviral development.