Putative ATP-Binding Cassette Transporter Is Essential for Brucella ovis Pathogenesis in Mice

Abstract

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...

Quorum-Sensing and BvrR/BvrS Regulation, the Type IV Secretion System, Cyclic Glucans, and BacA in the Virulence of Brucella ovis: Similarities to and Differences from Smooth Brucellae

“…Nevertheless, B. ovis is a naturally R Brucella species that is virulent in its natural host and in laboratory animal models (9,33,51). This ability of B. ovis to infect the host despite the absence of O-PS in the OM, together with its tropism for the male genital tract, is indicative of virulence mechanisms differing, at least in part, from those employed by S Brucella species, which are able to colonize the placenta and typically induce abortions.…”

“…Strains of the genus Brucella may be smooth (S) or rough (R), depending on the presence or absence, respectively, of O-polysaccharide (O-PS) chains in the LPS. O-PS masks other components of the bacterial surface (6), and since R mutant (M) strains derived from S strains are impaired in the interaction with phagocytes and are attenuated in animal models of infection (22,40,44), it is considered a key virulence factor in S Brucella species (such as B. melitensis, B. abortus, and B. suis).Nevertheless, B. ovis is a naturally R Brucella species that is virulent in its natural host and in laboratory animal models (9,33,51). This ability of B. ovis to infect the host despite the absence of O-PS in the OM, together with its tropism for the male genital tract, is indicative of virulence mechanisms differing, at least in part, from those employed by S Brucella species, which are able to colonize the placenta and typically induce abortions.…”

Brucella ovis: invasão, tráfego, fatores de virulência e resposta imune

“…Após a internalização da Brucella spp., o BCV precoce que é bem justo será o compartimento que permitirá a sobrevivência da bactéria, enquanto que o brucellassomo tardio que se apresenta mais espaçoso, será o compartimento da eliminação da bactéria (RITTING et al, 2001 Nas fases iniciais da infecção o sistema de transporte ABC é requerido para a sobrevivência intra e extracelular da B. ovis (SILVA et al, 2011). Devido sua quantidade de pseudogenes a B. ovis codifica poucas proteínas do sistema de transporte ABC (TSOLIS et al, 2009), o que resulta na falha do transporte de certos nutrientes.…”

“…Devido sua quantidade de pseudogenes a B. ovis codifica poucas proteínas do sistema de transporte ABC (TSOLIS et al, 2009), o que resulta na falha do transporte de certos nutrientes. Silva et al (2011) ressaltaram que esta deficiência no transporte de nutrientes poderia ser a responsável pela completa ausência de virulência da B. ovis em certos animais. Nos testes de biotipagem, a B. ovis é urease negativa, entretanto, o seu genoma possui dois clusters de urease com mutações, deleções e pseudogenes que sugerem a inatividade do teste de urease.…”

Species-specific nested PCR as a diagnostic tool for Brucella ovis infection in rams

“…Speciesspecific PCR used a primer pair targeting the ORF AO503 (F: 5'-ATCCCCCCATCACCATAACCGAAG-3'and R: 5'-GCCTACGCTGAAACTTGCTTTTG-3') located in the B. ovis-pathogenicity island 1 (BOPI-1) (Tsolis et al, 2009;Silva et al, 2011) as previously described (Xavier et al, 2010). PCR reaction and cycle parameters were performed as previously described (Xavier et al, 2010) and a volume of 2.5µL of template DNA was added per reaction.…”