Novel Soluble apxIVA-Truncated Protein and Its Application to Rapid Detection and Distinction of Actinobacillus pleuropneumoniae Wild-Strain-Infected Samples from Those Vaccinated with apxIV-Partially Deleted Vaccine

Actinobacillus pleuropneumoniae (APP) is a bacterial pathogen causing porcine pleuropneumonia, causing great economic loss to the global pig industry. Although natural apxIV contributes to the prevention and control of porcine pleuropneumonia, its isolation poses a great challenge, and recombinant soluble apxIV proteins tend to carry large molecular weight tags. The traditional serologic methods tend not to accurately detect the apxIV-partially deleted vaccine (GDV). In this study, we screened the soluble protein apxIVA N2 (756 bp) from six apxIV-truncated proteins and applied it to the enzyme-linked immunosorbent assay (ELISA) and colloidal gold immunochromatographic strip for detecting the samples vaccinated with APP GDV. The results indicate that N2 was close to the natural apxIV protein in terms of structure and function as it only contained a single His (0.86 kDa) tag and a single S (2 kDa) tag. Among the six candidate proteins, N2 exhibited the best performance in distinguishing APP-infected samples from those vaccinated with the APP GDV. Both ELISA and colloidal gold immunochromatographic strips based on this protein exhibited an excellent performance in detecting and distinguishing wild-strain-infected samples from those vaccinated with the subunit vaccine or the GDV. In addition, three monoclonal antibodies against different antigenic epitopes were identified using these truncated proteins. Our studies are of great significance for further research on APP, the differential diagnosis of wild strains and vaccine strains, and pig control breeding, exhibiting a broad application prospect in the on-site diagnosis of APP, particularly in remote areas lacking detection instruments and professionals.