Attenuation of a Brucella abortus mutant lacking a major 25 kDa outer membrane protein in cattle

To determine the virulence of a Brucella abortus mutant, BA25, lacking a major 25 kd outer membrane protein (Omp25) in cattle.

20 mixed-breed heifers in late gestation.

10 heifers were inoculated with 1 x 10(7) colony-forming units of the Omp25 mutant via the conjunctival sac, and an equal number were infected with the virulent parental strain B. abortus 2308. The delivery status of the dams was recorded, and colonization was assessed following necropsy. The ability of BA25 to replicate inside bovine phagocytes and chorionic trophoblasts was also evaluated in vitro because of the propensity of virulent brucellae to replicate inside these cells in vivo.

The parental strain induced abortions in 5 of 10 inoculated cattle, whereas only 1 of 10 dams exposed to BA25 aborted. Brucella abortus strain 2308 colonized all of the cow-calf pairs and induced Brucella-specific antibodies in 100% of the dams. In contrast, BA25 was isolated by bacteriologic cultural technique from 30% of the calves and 50% of the inoculated dams (n = 10). Of the 10 heifers inoculated with BA25, 4 did not develop Brucella-specific antibodies nor were they colonized by the mutant strain. In bovine macrophages and chorionic trophoblasts, BA25 replicated in significantly lower numbers than the virulent parental strain (n = 3).

The 25 kd outer membrane protein may be an important virulence factor for B. abortus in cattle. The attenuation of the Omp25 mutant in cattle may involve the inability of BA25 to replicate efficiently in bovine phagocytes and chorionic trophoblasts.