Comparative conventional and molecular tools for detection and differentiation of brucella field and vaccinal strains

Abstract EN

The current study was applied on 111 cows (78 Brucella suspected, 13 vaccinated with S19 and 20 with RB51) for differentiation of Brucella wild and vaccinal strains using conventional and molecular methods (PCR-RFLP, AMOS and real-time PCR).

Serum samples were examined using rose Bengal (RBT), tube agglutination and rivanol tests.

Milk samples were subjected to milk ring test (MRT), isolation and PCR assays.

The diagnostic sensitivity and specificity of RBT were 100% and 75% respectively, while MRT were 60% and 71% respectively.

The highest diagnostic sensitivity and specificity were recorded for PCR (100% and 82%) respectively.

Serological tests of vaccinated cows with S19 and RB51 revealed different seropositive and seronegative results respectively for all applied tests.

Five isolates of suspected cows were obtained and identified as B.melitensis biovar-3, two isolates of S19 vaccinated cows were obtained and identified as S19, and two isolates of RB51 vaccinated cows (one was identified as RB51 and the other as B.melitensis biovar-3).

PCR-RFLP assay revealed two patterns (P1 with 238bp and P2 with bands 282, 238bp) were obtained for field B.melitensis and Rev.1 vaccine respectively.

AMOS and real-time PCR revealed three different amplicons and three different dissociation peaks respectively specific for different Brucella species.

Conclusively, PCR-RFLP can differentiate B.meletensis (wild and vaccinal) and appropriate for application in mixed farms, while AMOS-PCR assay is recommended to distinguish between S19 and RB51 vaccinal strains and B.melitensis.

The use of more than one method provides a better reliable diagnostic approach for potent improvement of brucellosis control program