Development of a VirB12 gene-deleted Brucella abortus A19 strain as a vaccine candidate for differentiating infected from vaccinated animals in cattle and establishment of a differentiating ELISA assay.

OBJECTIVE

To evaluate the application of a VirB12 gene-deleted Brucella abortus A19 strain (A19-ΔVirB12) as a vaccine for differentiating infected from vaccinated animals in cattle and to develop an indirect ELISA (i-ELISA) for serological differentiation between vaccinated and naturally infected animals.

METHODS

A total of 115 Holstein cattle were immunized with A19-ΔVirB12 vaccine at 6 X 1010 CFUs, and sera were collected at 15, 30, 60, 90, 120, 150, and 180 days after vaccination. To determine the optimal threshold, serum from 180 infected cattle and 204 cattle immunized with A19-ΔVirB12 vaccine were confirmed positive for brucellosis tested by serum agglutination tests and a commercial ELISA kit. A differential identification i-ELISA was assembled and tested.

RESULTS

The results showed that the Brucella antibody titers peaked at 15 days after immunization with a positive rate of 100%. The titers then continued to decrease over time until 180 days after immunization, when the positive rate was approximately 6.67%. No adverse reactions were observed in the herd after immunization. The differentiating i-ELISA kit had good differential efficiency to distinguish Brucella-positive antibodies induced by immunization with A19-ΔVirB12 vaccine from natural infection, with a sensitivity of 98.9%, specificity of 94.6%, positive predictive value of 94.2%, negative predictive value of 99%, and accuracy of 96.6%.

CONCLUSIONS

Application of A19-ΔVirB12 vaccine and differential identification kits would support brucellosis control and eradication efforts in dairy herds.

CLINICAL RELEVANCE: This study demonstrates that the application of A19‑ΔVirB12 vaccine and its differential identification kit in dairy farms can effectively distinguish infected animals from immunized, which support brucellosis control and eradication efforts.