COMPARISON OF MOLECULAR AND SEROLOGICAL TESTS FOR DETECTION OF BRUCELLA ABORTUS IN ASYMPTOMATIC BOVINE BREEDING BULLS

Brucellosis is endemic in Pakistan and a serious threat to the healthy, profitable development of livestock in the country. Artificial insemination with semen collected from animals affected with Brucellosis is a potential source for the spread of the disease. Brucellosis is a bacterial zoonotic disease, which usually remains asymptomatic in male animals and can be easy to misdiagnose. Present study was designed to detect Brucella abortus (B. abortus) infection in asymptomatic bovine semen donor bulls. Efficiencies of IS711 conventional polymerase chain reaction (PCR),WboA/IS711J PCR which can differentiate wild/virulent strains from vaccine strain RB51, rose Bengal plate test (RBPT) and indirect enzyme linked immunosorbent assay (i-ELISA) were compared for detection of Brucellosis in breeding bulls. Blood samples were collected from 258 asymptomatic bovine breeding bulls (cattle bulls=118, buffalo bulls=140) maintained at well -organized semen production units in Punjab, Pakistan and five doses of semen of a pedigree buffalo bull (cryopreserved in past on different dates),IS711PCR, RBPT, and i-ELISA were conducted for detection of B. abortus infection. Additionally, WboA PCR was conducted to substantiate that samples positive in IS711PCR contain DNA of B. abortus virulent strains but do not contain DNA of B. abortus vaccine strain. Out of 258 asymptomatic bulls, 26 were positive by IS711PCR, 5 by i-ELISA and RBPT declared all animals negative for B. abortus. All five doses of semen were found positive for B. abortus with PCR. A slight degree of agreement was observed between PCR and i-ELISA (K=0. 17). PCR detected more samples positive for B. abortus infection among the three tests used in this study. In conclusion, IS711conventional PCR can be routinely used for an accurate and efficient diagnosis of Brucella infection in asymptomatic bulls, because the chances of non -detection of an infected animal in PCR are minimal B. abortus positivity rate with IS711 conventional PCR was significantly higher in buffalo bulls (17. 86%) than cattle bulls (0. 88%) p -value < 0. 0001. WboA/IS711J PCR detected that all IS711PCR positive animals had DNA of B. abortus wild strains but not of B. abortus vaccine strain RB51. WboA/IS711J PCR detected 4 more animals positive for B. abortus than IS711PCR. Some amplicons of WboA/IS711J PCR were sequenced and BLAST analysis of these sequences revealed that they matched 100% with the DNA sequence of B. abortus wild strains registered in the GenBank database.