A Novel Diagnostic Method For Discriminating Cattle Vaccinated With Brucella Suis Strain2from Those Naturally Infected With Brucella Abortus

Brucellosis, caused by Brucella spp., is a major zoonotic disease. The most commonways of diagnosing brucellosis in animals include the Rose-Bengal plate agglutination test,the serum agglutination test, the milk ring test, and the indirect enzyme linked immunosorbentassay. However, these methods cannot discriminate Brucella vaccine strains from naturallyacquired virulent strains. Therefore, development of a new diagnostic test that can effectivelydiscriminate between vaccine and non-vaccine strains will be useful for brucellosis diagnosisand control. To develop a method that can discriminate between cows inoculated with theBrucella suis vaccine strain2and those animals that had harbored natural Brucella infections,the gene sequences from six Brucella species (Br. melitensis, Br. abortus, Br. suis, Br. canis,Br. microti, and Br. ovis) were compared using Basic Local Alignment Search Tool software.This showed that one particular gene, the repA-related gene, discriminated Br. suis from Br.ovis and Br. abortus. The repA-related gene was PCR amplified and cloned into a pET32avector, heterologously expressed in Escherichia coli, affinity purified, and then used todevelop an indirect ELISA test. Serum from animals inoculated with the Br. suis S2vaccinestrain had positive repA-ELISA results. In contrast, serum from animals that had experiencednatural infections with Br. abortus or Br.melitensis had negative repA-ELISA results.Therefore, the repA-related protein-based indirect ELISA could distinguish between the Br.suis S2vaccinated cows and those that had experienced natural infections with Br. abortus;the serum samples for these animals also tested positive with the serum agglutination.