Brucella ovis is a major cause of reproductive failure in sheep, which is associated with epididymitis and infertility in rams. Importantly, B. ovis is one of the few Brucella species that is not zoonotic. Due to the scarcity of studies on B. ovis infection, a murine model of infection was developed. The roles of B. ovis genes encoding a putative hemagglutinin and an ABC transporter were investigated in the mouse model. The kinetics of B. ovis infection were similar in BALB/c and C57BL/6 mice, and both strains of mice developed multifocal microgranulomas in the liver and spleen, but only minimal colonization and histopathological changes were observed in the genital tract. Therefore, the mouse was considered a suitable infection model for B. ovis but not for B. ovis-induced genital disease. Two mutant strains were generated in this study (the ⌬abcAB and ⌬hmg strains). The B. ovis ⌬abcAB strain was attenuated in the spleens and livers of BALB/c mice compared to the wild-type (WT) strain (P < 0.001). Conversely, the ⌬hmg strain infected mice at the same level as WT B. ovis, suggesting that a putative hemagglutinin is not required for B. ovis pathogenesis. Additionally, the ⌬abcAB strain did not survive in peritoneal macrophages, extracellularly in the peritoneal cavity, or in RAW 264.7 macrophages. Moreover, infection with the ⌬abcAB strain was not lethal for male regulatory factor 1-knockout mice, whereas infection with the B. ovis WT strain was 100% lethal within 14 days postinfection. These results confirm that the predicted ABC transporter is required for the full virulence and survival of B. ovis in vivo.Brucella ovis is one of the main causes of reproductive failure in sheep (6). The disease is characterized by chronic epididymitis, orchitis, and infertility in sexually mature rams and occasional abortion and stillbirth in ewes (4,15,46). B. ovis has a worldwide distribution in economically important sheep-raising areas, with the exception of Great Britain (6). This organism may affect as much as 46% of a herd (41), leading to significant economic losses for the sheep industry due to infertility and early culling (8). B. ovis is a stable, rough, Gramnegative coccobacilli that belongs to the Alpha-2-Proteobacteriacea family (4, 17), and it is one of the few classical Brucella species that are not pathogenic to humans (4, 47).Brucella spp. are facultative intracellular bacteria that are able to survive and replicate in phagocytic and nonphagocytic cells and to establish chronic infections in animals and humans (18,46). Virulence factors are required for the invasion of host cells by Brucella spp. and for their survival and replication in host cells. Although classical virulence factors, such as capsules and fimbriae, are absent in Brucella species (18), pathogenic mechanisms specific to Brucella spp. have been identified (20,29,39,42). The mouse is often used as an animal model to investigate the pathogenesis of animal and human brucellosis (2, 13, 28). The murine model also allows studies that may reveal mech...