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Brucella ovis infects sheep and causes a clinical or subclinical disease characterized by genital lesions and reduced fertility in rams, placentitis and abortions in ewes, and increased perinatal mortality in lambs. As part of this study, analyses were conducted on a sheep farm near Belgrade (Serbia). Of the serological tests, indirect ELISA was performed. A total of 94 blood sera were analyzed, 33 from rams and 61 from ewes. The results showed 23 (69.7%) positive findings in rams and 2 (3.3%) positive findings in ewes, with an overall prevalence of 25.4% bounded by a 95% confidence interval. Bruce-ladder multiplex PCR and Real time PCR were performed on 19 serologically positive rams and two serologically positive ewes. The results indicate a slightly higher sensitivity of Real time PCR compared to conventional PCR in diagnosing B. ovis from the reproductive tissues of rams. However, the differences in sensitivity between different nucleic acid extraction protocols were not significant. Most rams showed a positive PCR result in only one sample of reproductive tissue, suggesting the recommendation to take multiple samples from each animal. Further research is needed to bring the sensitivity of molecular tests in diagnosing ram epididymitis closer to the sensitivity of serological tests.

Brucella ovis causes non-zoonotic ovine brucellosis of worldwide distribution and is responsible for important economic losses mainly derived from male genital lesions and reproductive fails. Studies about the virulence mechanisms of this rough species (lacking lipopolysaccharide O-chains) are underrepresented when compared to the main zoonotic Brucella species that are smooth (with O-chains). Zinc intoxication constitutes a defense mechanism of the host against bacterial pathogens, which have developed efflux systems to counterbalance toxicity. In this study, we have characterized three potential B. ovis zinc exporters, including the ZntA ortholog previously studied in B. abortus. Despite an in-frame deletion removing 100 amino acids from B. ovis ZntA, the protein retained strong zinc efflux properties. Only indirect evidence suggested a higher exporter activity for B. abortus ZntA, which, together with differences in ZntR-mediated regulation of zntA expression between B. ovis and B. abortus, could contribute to explaining why the ΔzntR mutant of B. abortus is attenuated while that of B. ovis is virulent. Additionally, B. ovis ZntA was revealed as a powerful cadmium exporter contributing to cobalt, copper, and nickel detoxification, properties not previously described for the B. abortus ortholog. Deletion mutants for BOV_0501 and BOV_A1100, also identified as potential zinc exporters and pseudogenes in B. abortus, behaved as the B. ovis parental strain in all tests performed. However, their overexpression in the ΔzntA mutant allowed the detection of discrete zinc and cobalt efflux activity for BOV_0501 and BOV_ A1100, respectively. Nevertheless, considering their low expression levels and the stronger activity of ZntA as a zinc and cobalt exporter, the biological role of BOV_0501 and BOV_A1100 is questionable. Results presented in this study evidence heterogeneity among pathogenic Brucellae regarding zinc export and, considering the virulence of B. ovis ΔzntA, suggest that hostmediated zinc intoxication is not a relevant mechanism to control B. ovis infection. [ABSTRACT FROM AUTHOR]

Brucella ovis causes non-zoonotic ovine brucellosis of worldwide distribution and is responsible for important economic losses mainly derived from male genital lesions and reproductive fails. Studies about the virulence mechanisms of this rough species (lacking lipopolysaccharide O-chains) are underrepresented when compared to the main zoonotic Brucella species that are smooth (with O-chains). Zinc intoxication constitutes a defense mechanism of the host against bacterial pathogens, which have developed efflux systems to counterbalance toxicity. In this study, we have characterized three potential B. ovis zinc exporters, including the ZntA ortholog previously studied in B. abortus. Despite an in-frame deletion removing 100 amino acids from B. ovis ZntA, the protein retained strong zinc efflux properties. Only indirect evidence suggested a higher exporter activity for B. abortus ZntA, which, together with differences in ZntR-mediated regulation of zntA expression between B. ovis and B. abortus, could contribute to explaining why the ΔzntR mutant of B. abortus is attenuated while that of B. ovis is virulent. Additionally, B. ovis ZntA was revealed as a powerful cadmium exporter contributing to cobalt, copper, and nickel detoxification, properties not previously described for the B. abortus ortholog. Deletion mutants for BOV_0501 and BOV_A1100, also identified as potential zinc exporters and pseudogenes in B. abortus, behaved as the B. ovis parental strain in all tests performed. However, their overexpression in the ΔzntA mutant allowed the detection of discrete zinc and cobalt efflux activity for BOV_0501 and BOV_A1100, respectively. Nevertheless, considering their low expression levels and the stronger activity of ZntA as a zinc and cobalt exporter, the biological role of BOV_0501 and BOV_A1100 is questionable. Results presented in this study evidence heterogeneity among pathogenic Brucellae regarding zinc export and, considering the virulence of B. ovis ΔzntA, suggest that host-mediated zinc intoxication is not a relevant mechanism to control B. ovis infection.

Simple Summary: Since the beginning of the Cenozoic era, microorganisms have circulated worldwide, many of them cause significant morbidity and mortality in animals and humans. Ecological changes may favor transmission, and modifying of host–environment/pathogen interactions and leptospirosis is a good example of this, as it evolved from pathogens circulating in wildlife. Using data generated from an epidemiological survey and from the lab, the abortion burden of multiple microorganisms in sheep was predicted according to the artificial neural network approach and Generalized Linear Model (GLM) in a geographic area of the Mexican highlands. The results showed that the best GLM is integrated by the serological detection of Leptospira interrogans serovar Hardjo and Brucella ovis in animals on the slopes with elevation between 2600 and 2800 masl in the municipality of Xalatlaco. The sheep pen built with materials of metal grids and untreated wood, dirt and concrete floors, bed of straw, and the well water supply were also remained independently associated with infectious abortion. We suggest that sensitizing stakeholders on good agricultural practices could improve public health surveillance. Unidentified abortion, of which leptospirosis, brucellosis, and ovine enzootic abortion are important factors, is the main cause of disease spread between animals and humans in all agricultural systems in most developing countries. Although there are well-defined risk factors for these diseases, these characteristics do not represent the prevalence of the disease in different regions. This study predicts the unidentified abortion burden from multi-microorganisms in ewes based on an artificial neural networks approach and the GLM. Methods: A two-stage cluster survey design was conducted to estimate the seroprevalence of abortifacient microorganisms and to identify putative factors of infectious abortion. Results: The overall seroprevalence of Brucella was 70.7%, while Leptospira spp. was 55.2%, C. abortus was 21.9%, and B. ovis was 7.4%. Serological detection with four abortion-causing microorganisms was determined only in 0.87% of sheep sampled. The best GLM is integrated via serological detection of serovar Hardjo and Brucella ovis in animals of the slopes with elevation between 2600 and 2800 meters above sea level from the municipality of Xalatlaco. Other covariates included in the GLM, such as the sheep pen built with materials of metal grids and untreated wood, dirt and concrete floors, bed of straw, and the well water supply were also remained independently associated with infectious abortion. Approximately 80% of those respondents did not wear gloves or masks to prevent the transmission of the abortifacient zoonotic microorganisms. Conclusions: Sensitizing stakeholders on good agricultural practices could improve public health surveillance. Further studies on the effect of animal–human transmission in such a setting is worthwhile to further support the One Health initiative. [ABSTRACT FROM AUTHOR]

Introduction: The diagnosis of brucellosis largely relies on tiger red plate agglutination test (RBPT). However, it is difficult to distinguish between natural infection antibody positive and vaccination antibody positive, nevertheless, the identification of specific Brucella species natural infection. Methods: Here, we analyzed the structure of main outer membrane proteins (OMPs), OMP25 and OMP31 from Brucella ovis (B. ovis) and Brucella melitensis (B. melitensis), which are the main pathogens of sheep brucellosis, and found the OMP25 and OMP31 could be used as the differential antigens for B. ovis and B. melitensis antibody. Then we expressed the OMP25 from B. ovis (OMP25o) and OMP31 from B. melitensis (OMP31m). Results: They have equally efficiency in antibody detection of vaccinated sheep serum, consistent with the RBPT results. However, through epidemiological investigations, we found some RBPT positive samples were negative by the OMP31m based serum antibody detection, but these samples gave positive results by the OMP25o. We verified these OMP31m negative but OMP25o positive samples by B. ovis and B. melitensis specific primers based PCR detection, and all these samples were B. melitensis negative. However, four out of six samples are B. ovis positive. These results showed that we could use the OMP25o and OMP31m to diagnose sheep brucellosis antibody, especially to discriminate the infection of the B. ovis. Discussion: Currently, China has not yet approved a vaccine based on B. ovis and B. ovis positive samples should be naturally infected. There should be some implicit transmission of B. ovis in Jilin province. Further epidemiological investigation should be conducted to monitor the B. ovis natural infection. [ABSTRACT FROM AUTHOR]

Objective: This study aims to compare the expression levels of tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β) in osteoblasts infiltrated with Mycobacterium tuberculosis H37Rv (H37Rv) and Brucella ovis to understand the differential bone destruction in spinal tuberculosis (STB) versus Brucella spondylitis (BS). Methods: Primary osteoblasts were isolated and cultured from the cranial bones of 2-5 days old mice and characterized by alkaline phosphatase (ALP) staining and alizarin red staining (ARS). H37Rv and B. ovis were cultured to the logarithmic phase, and transfection solutions were prepared. Osteoblasts were infiltrated with these bacteria at various multiplicities of infection (MOI) and time points. Cell survival post-infiltration was assessed using CCK-8 to determine optimal infection conditions. Osteoblasts were divided into three groups: the H37Rv group (infiltrated with optimal MOI H37Rv), the B. ovis group (infiltrated with optimal MOI B. ovis ), and a negative control group. TNF-α and IL-1β expression in the cytoplasm was observed using immunohistochemical staining, whereas their levels in cell supernatants were measured using enzyme-linked immunosorbent assay. Protein expression was analyzed by Western blot. Differences between groups were compared with using one-way analysis of variance and t-tests, with p < 0.05 indicating statistical significance. Results: Both H37Rv and B. ovis infiltrated osteoblasts, substantially increasing TNF-α and IL-1β expression. The H37Rv group showed substantially higher levels of TNF-α and IL-1β compared with the B. ovis group (p < 0.05). Conclusion: Infiltration of osteoblasts with H37Rv and B. ovis substantially increases TNF-α and IL-1β expression, with higher levels observed in H37Rv-infected osteoblasts. This overexpression may contribute to the more severe vertebral bone destruction seen in STB compared with BS.

In this study, it was aimed to develop an Enzyme-Linked Immunosorbent Assay (ELISA) prototype to evaluate humoral immune response against Brucella ovis by using different antigenic fractions from bacteria comparatively. For this purpose, four different antigens, namely Hot saline extract, Rough Lipopolysaccharide, Phage lysate 1, and Phage lysate 2 were used to test 84 sheep serum samples in ELISA. Seropositivity rates were, 14.3%, 13.1%, 10.7%, and 15.5%, respectively. Although the highest seropositivity rate was obtained by using the Phage lysate 2 antigen, it did not reach the level of statistical significance (χ²=0.89; P>0.05) difference between these antigens. It was concluded that in future studies, there would be a need for comparing the results of more serum samples tested in ELISA using Phage lysate 2 with the results of standard serological tests such as complement fixation test and agar gel immunodiffusion test or available commercial ELISA kit. Thus, the potential possibility of commercialization of a new in -house ELISA prototype will be evaluated more realistically. Given the seropositivity rate of Brucella ovis infection obtained by using a small number of serum samples, a large-scale serological study with greater serum samples from different parts of Turkey will reveal the real disease situation in our country. [ABSTRACT FROM AUTHOR]

• Brucella ovis is a suitable model for Brucella -induced fetal and placental lesions. • The candidate vaccine strain B. ovis Δ abcBA is safe for pregnant mice. • B. ovis Δ abcBA has protective potential in pregnant mice against virulent B. ovis. Ovine brucellosis caused by Brucella ovis is a major cause of reproductive failure in sheep. This study aimed to evaluate transplacental infection and pathogenicity of B. ovis wild type strain ATCC 25,840 (WT B. ovis) and the candidate vaccine strain B. ovis Δ abcBA in pregnant mice. A total of 40 BALB/c mice were equally divided into 4 groups: (i) non immunized and uninfected control mice (3/10 mice became pregnant); (ii) non immunized and challenged with WT B. ovis (5/10 pregnant); (iii) inoculated only with B. ovis Δ abcBA (6/10 pregnant); (iv) immunized with B. ovis Δ abcBA and challenged with WT B. ovis (5/10 pregnant). Female mice bred, and five days after visualization of the vaginal plug, they were inoculated intraperitoneally (ip) with 100 µL of sterile PBS, 100 µL of 1 × 106 CFU of B. ovis Δ abcBA , or 100 µL of 1 × 106 CFU of B. ovis WT, according to each group. At the 17th day of gestation, samples of spleen, liver, uterus, placenta, fetus and mammary gland were obtained for bacteriology, histopathology and immunohistochemistry. Non immunized mice challenged with B. ovis WT developed necrotizing placentitis as well as microgranulomas in the liver and spleen. These findings support the notion that B. ovis infection in pregnant mice induces lesions that are similar to those caused by B. abortus in the same animal model. B. ovis Δ abcBA was not recovered from any of the sampled organs, and it did not cause any gross or microscopic lesions, indicating that it is a safe and attenuated strain in this experimental model. In addition, B. ovis Δ abcBA was induced protective immunity as demonstrated by decreased numbers of B. ovis WT in the liver, uterus and fetuses of immunized mice after the challenge with B. ovis WT. [ABSTRACT FROM AUTHOR]

Abstract Brucella ovis is a non-zoonotic rough Brucella that causes genital lesions, abortions and increased perinatal mortality in sheep and is responsible for important economic losses worldwide. Research on virulence factors of B. ovis is necessary for deciphering the mechanisms that enable this facultative intracellular pathogen to establish persistent infections and for developing a species-specific vaccine, a need in areas where the cross-protecting ovine smooth B. melitensis Rev1 vaccine is banned. Although several B. ovis virulence factors have been identified, there is little information on its metabolic abilities and their role in virulence. Here, we report that deletion of pyruvate phosphate dikinase (PpdK, catalyzing the bidirectional conversion pyruvate ⇌ phosphoenolpyruvate) in B. ovis PA (virulent and CO2-dependent) impaired growth in vitro. In cell infection experiments, although showing an initial survival higher than that of the parental strain, this ppdK mutant was unable to multiply. Moreover, when inoculated at high doses in mice, it displayed an initial spleen colonization higher than that of the parental strain followed by a marked comparative decrease, an unusual pattern of attenuation in mice. A homologous mutant was also obtained in a B. ovis PA CO2-independent construct previously proposed for developing B. ovis vaccines to solve the problem that CO2-dependence represents for large scale production. This CO2-independent ppdK mutant reproduced the growth defect in vitro and the multiplication/clearance pattern in mouse spleens, and is thus an interesting vaccine candidate for the immunoprophylaxis of B. ovis ovine brucellosis.

ABSTRACT: Brucella ovis causes economic and reproductive losses in sheep herds. The goal of this study was to characterize infection with B. ovis field isolates in a murine model, and to evaluate protection induced by the candidate vaccine strain B. ovis ΔabcBA in mice challenged with these field isolates. B. ovis field strains were able to colonize and cause lesions in the liver and spleen of infected mice. After an initial screening, two strains were selected for further characterization (B. ovis 94 AV and B. ovis 266 L). Both strains had in vitro growth kinetics that was similar to that of the reference strain B. ovis ATCC 25840. Vaccination with B. ovis ΔabcBA encapsulated with 1% alginate was protective against the challenge with field strains, with the following protection indexes: 0.751, 1.736, and 2.746, for mice challenged with B. ovis ATCC25840, B. ovis 94 AV, and B. ovis 266 L, respectively. In conclusion, these results demonstrated that B. ovis field strains were capable of infecting and inducing lesions in experimentally infected mice. The attenuated vaccine strain B. ovis ΔabcBA induced protection in mice challenged with different B. ovis field isolates, resulting in higher protection indexes against more pathogenic strains.

Brucella melitensis and Brucella ovis are the primary etiological agents of brucellosis in small domestic ruminants. B. melitensis was first isolated in 1887 by David Bruce in Malta Island from spleens of four soldiers, while B. ovis was originally isolated in Australia and New Zealand in early 1950's from ovine abortion and rams epididymitis. Today, both agents are distributed worldwide: B. melitensis remains endemic and associated with an extensive negative impact on the productivity of flocks in -some regions, and B. ovis is still present in most sheep-raising regions in the world. Despite being species of the same bacterial genus, B. melitensis and B. ovis have extensive differences in their cultural and biochemical characteristics (smooth vs. rough colonial phases, serum and CO2 dependence for in vitro growth, carbohydrate metabolism), host preference (female goat and sheep vs. rams), the outcome of infection (abortion vs. epididymitis), and their zoonotic potential. Some of these differences can be explained at the bacterial genomic level, but the role of the host genome in promoting or preventing interaction with pathogens is largely unknown. Diagnostic techniques and measures to prevent and control brucellosis in small ruminants vary, with B. melitensis having more available tools for detection and prevention than B. ovis. This review summarizes and analyzes current available information on: (1) the similarities and differences between these two etiological agents of brucellosis in small ruminants, (2) the outcomes after their interaction with different preferred hosts and current diagnostic methodologies, (3) the prevention and control measures, and (4) alerting animal producers about the disease and raise awareness in the research community for future innovative activities.

ABSTRACT Flavoproteins are a diverse class of proteins that are mostly enzymes and contain as cofactors flavin mononucleotide (FMN) and/or flavin adenine dinucleotide (FAD), which enable them to participate in a wide range of physiological reactions. We have compiled 78 potential proteins building the flavoproteome of Brucella ovis (B. ovis), the causative agent of ovine brucellosis. The curated list of flavoproteins here reported is based on (i) the analysis of sequence, structure and function of homologous proteins, and their classification according to their structural domains, clans, and expected enzymatic functions; (ii) the constructed phylogenetic trees of enzyme functional classes using 19 Brucella strains and 26 pathogenic and/or biotechnological relevant alphaproteobacteria together with B. ovis; and (iii) the evaluation of the genetic context for each entry. Candidates account for ∼2.7% of the B. ovis proteome, and 75% of them use FAD as cofactor. Only 55% of these flavoproteins belong to the core proteome of Brucella and contribute to B. ovis processes involved in maintenance activities, survival and response to stress, virulence, and/or infectivity. Several of the predicted flavoproteins are highly divergent in Brucella genus from revised proteins and for them it is difficult to envisage a clear function. This might indicate modified catalytic activities or even divergent processes and mechanisms still not identified. We have also detected the lack of some functional flavoenzymes in B. ovis, which might contribute to it being nonzoonotic. Finally, potentiality of B. ovis flavoproteome as the source of antimicrobial targets or biocatalyst is discussed. IMPORTANCE Some microorganisms depend heavily on flavin-dependent activities, but others maintain them at a minimum. Knowledge about flavoprotein content and functions in different microorganisms will help to identify their metabolic requirements, as well as to benefit either industry or health. Currently, most flavoproteins from the sheep pathogen Brucella ovis are only automatically annotated in databases, and only two have been experimentally studied. Indeed, certain homologues with unknown function are not characterized, and they might relate to still not identified mechanisms or processes. Our research has identified 78 members that comprise its flavoproteome, 76 of them flavoenzymes, which mainly relate to bacteria survival, virulence, and/or infectivity. The list of flavoproteins here presented allows us to better understand the peculiarities of Brucella ovis and can be applied as a tool to search for candidates as new biocatalyst or antimicrobial targets.

Brucella ovis is a non-zoonotic bacterium causing contagious epididymitis and other genital lesions in rams and responsible for significant economic losses in sheep-breeding areas. It is a naturally rough (without O-chains in the lipopolysaccharide) Brucella species whose virulence mechanisms have been less explored than those of zoonotic smooth brucellae (bearing O-chains that mask other outer membrane molecules). Considering the rough nature of Brucella ovis, the influence of surface components other than O-chains on its biological properties may be greater than in smooth Brucella species. Here we describe the construction and characterization of the mucR deletion mutant of virulent B. ovis PA, which is defective in a transcriptional regulator, affecting surface properties and virulence in smooth brucellae. This mutant showed increased amounts of three proteins identified as HdeA (acid-activated chaperone), Omp25d (outer membrane protein undetectable in the parental strain), and BOV_A0299 (hypothetical protein of unknown function). This observation correlated with the enhanced transcription of the corresponding genes and constitutes the first report on this type of proteome alteration in Brucella ΔmucR mutants. The upstream regions of the three genes contained AT rich domains with T-A steps described as binding sites for MucR in the Brucella abortus 2308 babR promoter (gene also upregulated in B. ovis ΔmucR), which suggests that hdeA, omp25d, and BOV_A0299 expression could be repressed by MucR through a direct binding to their promoter regions. Relative quantification of transcripts of several other genes selected according to the transcriptome of smooth brucellae ΔmucR mutants revealed not only similarities but also relevant differences among strains, such as those detected in flagellar and virB genes. Periplasmic HdeA has been related to the resistance of B. abortus to acidic pH, conditions encountered by Brucella inside phagocytes, but the deletion of hdeA in B. ovis PA and the ΔmucR mutant did not modify any of the evaluated properties of these strains. The B. ovis PA ΔmucR and ΔmucRΔhdeA mutants had defective in vitro growth and altered surface properties and architecture, exemplified by detectable amounts of Omp25d. Moreover, they showed virulence attenuation but established persistent splenic infection in mice, which encourages their evaluation as specifical attenuated vaccines against B. ovis.

Abstract Background Brucella ovis causes a sexually transmitted, infectious disease of domestic sheep characterized by genital lesions and epididymitis in rams, placentitis and rare abortions in ewes, and neonatal mortality in lambs. This study was designed to 1) estimate animal and flock seroprevalence of B. ovis in sheep across Wyoming, USA, and 2) describe epidemiologic risk factors associated with seropositive sheep and flocks. For the animal seroprevalence estimate, 2423 blood samples were collected from sheep on 18 producer-selected operations and a questionnaire about possible risk factors was distributed. For the flock seroprevalence estimate, blood samples from 82 operations were obtained, including samples from the previous 18 operations and 64 additional operations that sent samples to the Wyoming State Veterinary Laboratory for diagnostic testing. Categorical risk factors were created based on questionnaires and submission forms. Sera was analyzed using the B. ovis enzyme-linked immunosorbent assay. Results Estimated true animal and flock seroprevalence were 0.53% (95% CI: 0.21–1.01%; 22/2,423) and 22.5% (95% CI: 14–32%; 18/82), respectively. Using Fisher’s exact and Mid-p exact tests to compare apparent seroprevalence with respect to possible risk factors, increased age and breed type were risk factors associated with seropositive sheep, while region and large flock size were risk factors associated with seropositive flocks. Conclusions Results from this study suggest few sheep have been exposed to B. ovis, but many flocks contain at least one seropositive animal. Each region in Wyoming contained at least one seropositive animal and flock, emphasizing the importance of disease-free documentation before purchasing new sheep. Aged sheep (≥ 6 years of age) had the highest seroprevalence among age groups; hence, we propose the separation of young rams from older rams to help reduce disease spread outside the breeding season. Wool breeds (Rambouillet and Merino) may be less susceptible to B. ovis infection given they had the lowest animal seroprevalence of the breed types, and large flocks (> 100 breeding rams) had the highest seroprevalence of the flock size categories, likely due to more intensive management strategies that can contribute to the introduction and persistence of B. ovis infection in sheep and flocks.

Com objetivo de determinar a prevalência de anticorpos contra Brucella ovis foram examinados 95 carneiros provenientes de 23 cabanhas localizadas em 13 municípios de Santa Catarina. Foram realizados exames clínicos externos do aparelho reprodutor por palpação e coleta de sangue. Os soros desses animais foram submetidos à prova de gel difusão para pesquisa de anticorpos contra a Brucella ovis e reagiram negativamente. No exame clínico, não se observaram alterações nos órgãos reprodutores dos carneiros.

A serological survey was carried out to assess the frequency of leptospirosis, small ruminants lentivirus (SRLV), and brucellosis in small ruminant herds in the Recôncavo Baiano, State of Bahia, Brazil, from February to December 2017. In four goat herds, 125 animals were tested for SRLV and leptospirosis, while in five sheep herds, 378 animals were tested for leptospirosis, brucellosis, and SRLV. Regarding leptospirosis, MAT detected 80% of goats and 15.34% of sheep seroreactive. Reactivity was most frequent to serogroups Autumnalis and Grippotyphosa in goats and sheep, respectively. Regarding SRLV, 8.8% of goats and 0.79% of sheep were reactive. Search for anti-B. ovis antibodies revealed 0.52% reactivity. In sheep, three animals showed simultaneous seroreactivity for SRLV and leptospirosis, while one animal had a serological response for brucellosis and leptospirosis. In goats, simultaneous seroreactivity for SRLV and leptospirosis was observed in only one animal. Leptospirosis was the most frequent of the three infectious diseases in investigated herds.

Brucella ovis is a major cause of reproductive failure in rams and it is one of the few well-described Brucella species that is not zoonotic. Previous work showed that a B. ovis mutant lacking a species-specific ABC transporter (ΔabcBA) was attenuated in mice and was unable to survive in macrophages. The aim of this study was to evaluate the role of this ABC transporter during intracellular survival of B. ovis. In HeLa cells, B. ovis WT was able to survive and replicate at later time point (48 hpi), whereas an ΔabcBA mutant was attenuated at 24 hpi. The reduced survival of the ΔabcBA mutant was associated with a decreased ability to exclude the lysosomal marker LAMP1 from its vacuolar membrane, suggesting a failure to establish a replicative niche. The ΔabcBA mutant showed a reduced abundance of the Type IV secretion system (T4SS) proteins VirB8 and VirB11 in both rich and acid media, when compared to WT B. ovis. However, mRNA levels of virB1, virB8, hutC, and vjbR were similar in both strains. These results support the notion that the ABC transporter encoded by abcEDCBA or its transported substrate acts at a post-transcriptional level to promote the optimal expression of the B. ovis T4SS within infected host cells.

Brucella ovis is a facultative intracellular bacterium that causes a non-zoonotic ovine brucellosis mainly characterized by male genital lesions and is responsible for important economic losses in sheep farming areas. Studies about the virulence mechanisms of Brucella have been mostly performed with smooth (bearing O-polysaccharide in lipopolysaccharide) zoonotic species, and those performed with B. ovis have revealed similarities but also relevant differences. Except for few strains recently isolated from unconventional hosts, Brucella species are non-motile but contain the genes required to assemble a flagellum, which are organized in three main loci of about 18.5, 6.4, and 7.8 kb. Although these loci contain different pseudogenes depending on the non-motile Brucella species, smooth B. melitensis 16M builds a sheathed flagellum under particular culture conditions and requires flagellar genes for virulence. However, nothing is known in this respect regarding other Brucella strains. In this work, we have constructed a panel of B. ovis PA mutants defective in one, two or the three flagellar loci in order to assess their role in virulence of this rough (lacking O-polysaccharide) Brucella species. No relevant differences in growth, outer membrane-related properties or intracellular behavior in cellular models were observed between flagellar mutants and the parental strain, which is in accordance with previous results with B. melitensis 16M single-gene mutants. However, contrary to these B. melitensis mutants, unable to establish a chronic infection in mice, removal of the three flagellar loci in B. ovis did not affect virulence in the mouse model. These results evidence new relevant differences between B. ovis and B. melitensis, two species highly homologous at the DNA level and that cause ovine brucellosis, but that exhibit differences in the zoonotic potential, pathogenicity and tissue tropism.

Brucella ovis infection results in genital damage and epididymitis in rams, placental inflammation and rare abortion in ewes, and neonatal mortality in lambs. However, the mechanism underlying B. ovis infection remains unclear. In the present study, we used prokaryotic transcriptome sequencing to identify the differentially expressed genes (DEGs) between wild-type B. ovis and intracellular B. ovis in RAW264.7 macrophages. Gene ontology (GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed, and quantitative reverse transcriptase PCR (qRT-PCR) was used to validate the top 10 upregulated and downregulated DEGs. The results showed that 212 genes were differentially expressed, including 68 upregulated and 144 downregulated genes, which were mainly enriched in 30 GO terms linked to biological process, cellular component, and molecular function. KEGG analysis showed that the DEGs were enriched in the hypoxia-inducible factor 1 (HIF-1) signaling pathway, mitogen-activated protein kinase (MAPK) signaling pathway, beta-alanine metabolism, and quorum sensing pathway. BME_RS01160, BME_RS04270, BME_RS08185, BME_RS12880, BME_RS25875, predicted_RNA865, and predicted_RNA953 were confirmed with the transcriptome sequencing data. Hence, our findings not only reveal the intracellular parasitism of B. ovis in the macrophage immune system, but also help to understand the mechanism of chronic B. ovis infection. [ABSTRACT FROM AUTHOR]

As the molecular mechanisms of Brucella ovis pathogenicity are not completely clear, we have applied a transcriptome approach to identify the differentially expressed genes (DEGs) in RAW264.7 macrophage infected with B. ovis. The DEGs related to immune pathway were identified by Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) functional enrichment analysis. Quantitative real-time PCR (qRT-PCR) was performed to validate the transcriptome sequencing data. In total, we identified 337 up-regulated and 264 down-regulated DEGs in B. ovis -infected group versus mock group. Top 20 pathways were enriched by KEGG analysis and 20 GO by functional enrichment analysis in DEGs involved in the molecular function, cellular component, and biological process and so on, which revealed multiple immunological pathways in RAW264.7 macrophage cells in response to B. ovis infection, including inflammatory response, immune system process, immune response, cytokine activity, chemotaxis, chemokine-mediated signaling pathway, chemokine activity, and CCR chemokine receptor binding. qRT-PCR results showed Ccl2 (ENSMUST00000000193), Ccl2 (ENSMUST00000124479), Ccl3 (ENSMUST00000001008), Hmox1 (ENSMUST00000005548), Hmox1 (ENSMUST00000159631), Cxcl2 (ENSMUST00000075433), Cxcl2 (ENSMUST00000200681), Cxcl2 (ENSMUST00000200919), and Cxcl2 (ENSMUST00000202317). Our findings firstly elucidate the pathways involved in B. ovis -induced host immune response, which may lay the foundation for revealing the bacteria–host interaction and demonstrating the pathogenic mechanism of B. ovis. [ABSTRACT FROM AUTHOR]

Control of Brucella ovis infection in sheep flocks in the United States depends on early detection of B. ovis antibodies via serologic testing. We used 2,276 sheep sera and various cutoff values to compare seroprevalence and agreement between 2 ELISAs: the National Veterinary Services Laboratories (NVSL) B. ovis indirect ELISA and the IDEXX B. ovis ELISA kit. A subset of 295 sera was used to compare agreement and evaluate relative sensitivity and specificity of the 2 ELISAs with an agar gel immunodiffusion (AGID) test kit. There was no significant difference in B. ovis seroprevalence between the ELISAs; however, there was poor agreement between them. When the AGID test was used as the reference test, the IDEXX ELISA with a moderate cutoff value (S/P ratio = 45%) had the highest relative sensitivity of 38.1% and specificity of 92.0%. The NVSL ELISA with a lax cutoff value (S/P ratio = 0.75) had relative sensitivity of 19.1% and specificity of 94.6%. Receiver operating characteristic analysis revealed that optimal cutoff values for the NVSL and IDEXX ELISAs were 0.091 and 16.5%, respectively. This results in sensitivity and specificity of 85.7% and 31.8% for the NVSL ELISA, and sensitivity and specificity of 81.0% and 53.6% for the IDEXX ELISA, respectively. [ABSTRACT FROM AUTHOR]

Brucella ovis is a veterinary pathogen associated with epididymitis in sheep. Despite its genetic similarity to the zoonotic pathogens B. abortus, B. melitensis and B. suis, B. ovis does not cause zoonotic disease. Genomic analysis of the type strain ATCC25840 revealed a high percentage of pseudogenes and increased numbers of transposable elements compared to the zoonotic Brucella species, suggesting that genome degradation has occurred concomitant with narrowing of the host range of B. ovis. The absence of genomic island 2, encoding functions required for lipopolysaccharide biosynthesis, as well as inactivation of genes encoding urease, nutrient uptake and utilization, and outer membrane proteins may be factors contributing to the avirulence of B. ovis for humans. A 26.5 kb region of B. ovis ATCC25840 Chromosome II was absent from all the sequenced human pathogenic Brucella genomes, but was present in all of 17 B. ovis isolates tested and in three B. ceti isolates, suggesting that this DNA region may be of use for differentiating B. ovis from other Brucella spp. This is the first genomic analysis of a non-zoonotic Brucella species. The results suggest that inactivation of genes involved in nutrient acquisition and utilization, cell envelope structure and urease may have played a role in narrowing of the tissue tropism and host range of B. ovis.

Brucella ovis is the causative agent of ovine brucellosis, which is an important infectious disease in sheep farming worldwide and is responsible for economic losses because of its negative effect on the reproductive system of rams and ewes. Serologic tests are the main tools for detection of infection; however, these tests commonly yield a high frequency of false-negative results. We compared 2 serologic tests, agar gel immunodiffusion (AGID) and ELISA, for the detection of anti– B. ovis antibodies in naturally infected sheep. Of the 728 serum samples analyzed, 0.3% were positive by AGID and 9.2% by ELISA. Positive results were obtained for different animals and flocks. There was no statistical difference between the detection frequency of the 2 methods (p = 0.674), and the kappa test indicated low concordance (κ = 0.005). The lack of agreement between results obtained using AGID and ELISA, associated with the absence of clinical signs, makes it difficult to detect ovine brucellosis efficiently, and demonstrates the need for effective tests for the definitive detection of B. ovis infection. [ABSTRACT FROM AUTHOR]

Aim of this study is to report a laboratory investigation performed following the isolation of Brucella ovis, causing ovine epididymitis, in a traditional sheep farm in Sicily (South Italy). This disease represents a newly emerging risk for Italian livestock and is listed among diseases of EU priority (EU Reg 2016/429). Blood samples from 56 rams and 143 ewes were analyzed by both Enzyme-Linked Immunosorbent Assay (ELISA) and Complement Fixation Test (CFT). Genital swabs from all rams and 15 lactating ewes were collected to perform real-time PCR. Eighteen serologically positive rams were slaughtered and postmortem-inspected. Samples of testicle, epididymis, lymph nodes, and urine were also collected in order to perform microbiological, molecular, and histopathological analysis. Twelve slaughtered rams showed anatomo-pathological lesions. Real-time PCR for B. ovis BOV_A0504 gene was positive for 13 testicles and epididymis and 11 urine while B. ovis was isolated from epididymis and testicles of 7 slaughtered rams. This is the first exhaustive laboratory report of a microbiological, molecular, and serological pattern of the disease in sheep in Italy. Despite the impact on health and animal welfare, the epidemiology of B. ovis infection is still unknown, particularly in our country where the disease is considered endemic.

El objetivo de este estudio fue caracterizar los hallazgos patológicos y su relación con los resultados clínicos y serológicos en carneros infectados experimentalmente (IE) con Brucella ovis. Se utilizaron 18 carneros de 1 a 4 años y libres de B. ovis; que se distribuyeron en tres grupos: Grupo 1 (n= 6): inoculación en mucosas (B. ovis); Grupo 2 (n= 6), inoculación vía intravenosa y Grupo 3 (n= 6) Control. Se llevó a cabo el seguimiento de un carnero -1 grupo durante 189 días (d) post-IE. Se obtuvieron muestras de 2 mL de suero sanguíneo y semen entre los días 0 y 189 post-IE para realizar pruebas serológicas de Inmunodifusión en Gel de Agar (IGDA) y ELISA. El semen se procesó por la técnica de reacción en cadena de la polimerasa PCR. El análisis estadístico se realizó mediante modelos categóricos y mediciones repetidas. ELISA mostró mayor sensibilidad (P

Abstract Subcellular vaccines against ovine contagious epididymitis due Brucella ovis can solve some shortcomings associated with the use of Brucella melitensis Rev 1. We have demonstrated that the parenteral immunization with polymeric antigen BLSOmp31 emulsified in oil adjuvant conferred significant protection against B. ovis in rams. In our previous studies, we have characterized chitosan microspheres (ChMs) and a thermoresponsive and mucoadhesive in situ gel (Poloxamer 407-Ch) as two novel formulation strategies for the delivery of BLSOmp31 in nasal as well as conjunctival mucosa. In the present work, we evaluated the immunogenicity and protection conferred by the intranasal and conjunctival immunization with these two mucosal delivery systems against B. ovis in rams. BLSOmp31-ChM administered by intranasal route and BLSOmp31-P407-Ch applied by intranasal or conjunctival routes induced systemic, local and preputial IgG and IgA antibody response. Neither formulation showed interference in the serological diagnosis. Thus, mucosal immunization using either formulation induced significant specific cellular immune responses (in vitro and in vivo) and it prevented the excretion of B. ovis in semen. Although these vaccines did not prevent infection in immunized rams, colonization reduction of infected organs and bacterial distribution differed significantly between vaccinated and unvaccinated rams. [ABSTRACT FROM AUTHOR]

Sheep breeding has been important in agribusiness, transforming the Brazilian productive scenario. However, it is still deficient due to the damages caused by infectious diseases. Leptospirosis is a severe disease with global distribution, caused by bacteria from the Leptospira genre affecting both humans and animals. The general infection is unapparent, or its clinical signs, when present, are similar to other infections. Brucellosis is an infectious disease caused by bacteria from the Brucella genre responsible for reproductive disorders in animals, especially ruminants. The purpose of this paper was to seroepidemiological study of Leptospira spp. and Brucella ovis in sheep and dogs from nonmechanized rural properties from the northwestern region in the state of Paraná, Brazil. In order to detect anti-Leptospira antibodies, microscopic agglutination (MAT) was performed. For anti-Brucella antibodies, the agar gel immunodiffusion assay (AGID) was performed. From the total 542 samples from sheep sera analyzed, 11.25% were considered reagent to Leptospira spp. and 18.26% to Brucella ovis. From the 36 dog samples, 25% were reagent to MAT and AGID. From the 32 properties analyzed, 75% were considered positive for leptospirosis and 56.25% for brucellosis. Antibodies against the most probable serovars were Hardjo (34.42%) and Butembo (44.44%) in sheep and dogs, respectively, and the variable exchange of animals among properties was associated to leptospiric infection (p=0.028) in sheep. Leptospirosis and brucellosis are present in the sheep herd and dogs in the rural properties studied, and such result is a warning of the zoonotic importance and the need to establish sanitary programs directed to these animal species.

Ovine brucellosis can cause epididymitis and reduce fertility in rams and therefore is an important disease in sheep and rams from economic point of view. Sometimes it causes abortions and increased perinatal mortality in sheep as well, but it primarily affects male animals. Brucella ovis was first identified in northern region of Serbia, in 2008-2009 during a research study. Specific antibodies against Brucella ovis were detected in the sheep that originated from Becej and Titel municipalities in the South Bačka district. The prevalence was low, only 0.89%. For the purpose of this study, a total number of 6,407 serum samples from rams and sheep were used. All of the samples were from the northern part of Serbia, belonging to South Bačka and Srem districts. Serological test was performed in order to determine the presence of specific antibodies against Brucella ovis in rams and sheep, by ELISA test (indirect ELISA - iELISA). The prevalence varied from 0% - 19.3% during the years. If we look into the municipalities of South Bačka and Srem district, we can see that the numbers of positive samples are different, and the number of analyzed samples varies. The overall seroprevalence from 2014-2018 in South Bačka and Srem district is 6.15%, but it varies significantly among different municipalities from 0 to 26%. The majority of analyzed sample were from rams and all of the positive findings are in rams. Only a small number of sheep (female) has been examined so far, so the prevalence among females is still unknown.

Brucella ovis is a non-zoonotic Brucella species lacking specific vaccine. It presents a narrow host range, a unique biology relative to other Brucella species, and important distinct surface properties. To increase our knowledge on its peculiar surface and virulence features, and seeking to develop a specific vaccine, multiple mutants for nine relevant cell-envelope-related genes were investigated. Mutants lacking Omp10 plus Omp19 could not be obtained, suggesting that at least one of these lipoproteins is required for viability. A similar result was obtained for the double deletion of omp31 and omp25 that encode two major surface proteins. Conversely, the absence of major Omp25c (proved essential for internalization in HeLa cells) together with Omp25 or Omp31 was tolerated by the bacterium. Although showing important in vitro and in vivo defects, the Δomp10Δomp31Δomp25c mutant was obtained, demonstrating that B. ovis PA survives to the simultaneous absence of Omp10 and four out seven proteins of the Omp25/Omp31 family (i.e., Omp31, Omp25c, Omp25b, and Omp31b, the two latter naturally absent in B. ovis). Three multiple mutants were selected for a detailed analysis of virulence in the mouse model. The Δomp31Δcgs and Δomp10Δomp31Δomp25c mutants were highly attenuated when inoculated at 106 colony forming units/mouse but they established a persistent infection when the infection dose was increased 100-fold. The Δomp10ΔugpBΔomp31 mutant showed a similar behavior until week 3 post-infection but was then totally cleared from spleen. Accordingly, it was retained as vaccine candidate for mice protection assays. When compared to classical B. melitensis Rev1 heterologous vaccine, the triple mutant induced limited splenomegaly, a significantly higher antibody response against whole B. ovis PA cells, an equivalent memory cellular response and, according to spleen colonization measurements, better protection against a challenge with virulent B. ovis PA. Therefore, it would be a good candidate to be evaluated in the natural host as a specific vaccine against B. ovis that would avoid the drawbacks of B. melitensis Rev1. In addition, the lack in this attenuated strain of Omp31, recognized as a highly immunogenic protein during B. ovis infection, would favor the differentiation between infected and vaccinated animals using Omp31 as diagnostic target.

Brucella ovis is a bacterial pathogen present in most major sheep-producing regions of the world. The pathogen is associated with ram infertility, decreased ewe conception rates, and premature lambs. Twenty ELISA seropositive or indeterminate rams were culled from a B. ovis–positive flock, and donated to the Wyoming State Veterinary Laboratory for evaluation of infection. Tissues from each ram were collected at autopsy for additional testing, including bacterial culture, PCR, and histopathology. Of 17 seropositive rams, 11 rams were also positive by culture and PCR, and had evidence of mild histologic lesions; 1 seropositive ram was positive by culture with mild histologic lesions, but negative by PCR. Five seropositive rams were negative by culture and PCR and had no histologic lesions. Three indeterminate rams were negative by culture and by PCR and had no histologic lesions. The tissues in which B. ovis was most often detected included the epididymis, vesicular gland, and ampulla. Although this was a small study, the observation that 5 of 17 (29%) rams that were initially seropositive had no evidence of infection is interesting. A convalescent test for valuable seropositive animals prior to culling may be useful, and reproductive tissues may be evaluated postmortem if confirmatory testing is desired. [ABSTRACT FROM AUTHOR]

Brucella ovis is a non-zoonotic Brucella species lacking specific vaccine. It presents a narrow host range, a unique biology relative to other Brucella species, and important distinct surface properties. To increase our knowledge on its peculiar surface and virulence features, and seeking to develop a specific vaccine, multiple mutants for nine relevant cell-envelope-related genes were investigated. Mutants lacking Omp10 plus Omp19 could not be obtained, suggesting that at least one of these lipoproteins is required for viability. A similar result was obtained for the double deletion of omp31 and omp25 that encode two major surface proteins. Conversely, the absence of major Omp25c (proved essential for internalization in HeLa cells) together with Omp25 or Omp31 was tolerated by the bacterium. Although showing important in vitro and in vivo defects, the Δ omp10 Δ omp31 Δ omp25c mutant was obtained, demonstrating that B. ovis PA survives to the simultaneous absence of Omp10 and four out seven proteins of the Omp25/Omp31 family (i.e., Omp31, Omp25c, Omp25b, and Omp31b, the two latter naturally absent in B. ovis). Three multiple mutants were selected for a detailed analysis of virulence in the mouse model. The Δ omp31 Δ cgs and Δ omp10 Δ omp31 Δ omp25c mutants were highly attenuated when inoculated at 106 colony forming units/mouse but they established a persistent infection when the infection dose was increased 100-fold. The Δ omp10 Δ ugpB Δ omp31 mutant showed a similar behavior until week 3 post-infection but was then totally cleared from spleen. Accordingly, it was retained as vaccine candidate for mice protection assays. When compared to classical B. melitensis Rev1 heterologous vaccine, the triple mutant induced limited splenomegaly, a significantly higher antibody response against whole B. ovis PA cells, an equivalent memory cellular response and, according to spleen colonization measurements, better protection against a challenge with virulent B. ovis PA. Therefore, it would be a good candidate to be evaluated in the natural host as a specific vaccine against B. ovis that would avoid the drawbacks of B. melitensis Rev1. In addition, the lack in this attenuated strain of Omp31, recognized as a highly immunogenic protein during B. ovis infection, would favor the differentiation between infected and vaccinated animals using Omp31 as diagnostic target. [ABSTRACT FROM AUTHOR]

Copyright of Archives of Veterinary Medicine / Arhiv Veterinarske Medicine is the property of Scientific Veterinary Institute Novi Sad and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

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Ovine brucellosis is often neglected contagious bacterial disease causing enormous economic losses to sheep industry. India is recognized as geographical hotspot for brucellosis and there is only one seroprevalence report of Brucella ovis infection in sheep. Ovine brucellosis is caused by both rough and smooth strains of Brucella sp. In the present study, 300 sheep serum samples collected from 9 different districts of Karnataka by 3 stage random sampling approach were screened by antibody and DNA detection tests (RBPT and iELISA) and PCR. Antibodies specific to B. melitensis and B. ovis infection were 8.67% and 5.34%, respectively. In the same set of samples, 5% (15/300) showed Brucella DNA amplification by bcsp31 and IS711 genus specific PCRs. In B. melitensis and B. ovis species specific PCRs, 3.34% and 1.67%, respectively were detected positive indicating presence of both Brucella species in sheep population. Among the 7 districts, Yadgir, Tumkur, Raichur and Bagalkot showed higher prevalence of antibodies against rough and smooth strains of Brucella. This emphasizes the need for sensitization of national system in designing surveillance and control strategies for both B. ovis and B. melitensis infections in sheep. [ABSTRACT FROM AUTHOR]

Copyright of Revista Cientifica de la Facultade de Veterinaria is the property of Universidad del Zulia, Facultad de Ciencias Veterinarias and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Abstract Ovine epididymitis (Brucella ovis) has been assessed according to the criteria of the Animal Health Law (AHL), in particular criteria of Article 7 on disease profile and impacts, Article 5 on the eligibility of ovine epididymitis to be listed, Article 9 for the categorisation of ovine epididymitis according to disease prevention and control rules as in Annex IV and Article 8 on the list of animal species related to ovine epididymitis. The assessment has been performed following a methodology composed of information collection and compilation, expert judgement on each criterion at individual and, if no consensus was reached before, also at collective level. The output is composed of the categorical answer, and for the questions where no consensus was reached, the different supporting views are reported. Details on the methodology used for this assessment are explained in a separate opinion. According to the assessment performed, ovine epididymitis can be considered eligible to be listed for Union intervention as laid down in Article 5(3) of the AHL. The disease would comply with the criteria as in Sections 3, 4 and 5 of Annex IV of the AHL, for the application of the disease prevention and control rules referred to in points (c), (d) and (e) of Article 9(1). The animal species to be listed for ovine epididymitis according to Article 8(3) criteria are mainly sheep and other species of the families Bovidae and Cervidae as susceptible and sheep and deer as reservoirs.

Brucella ovis is a major cause of epididymitis in sexually mature rams, resulting in subfertility, infertility, and economic losses for the sheep industry worldwide. The aim of this study was to develop an indirect ELISA (iELISA) using recombinant proteins, namely rBoP59 and rBP26, as antigens for serological diagnosis of B. ovis infection. The BoP59 and BP26 recombinant proteins were expressed in E. coli and purified by affinity chromatography. Antigenicity was tested by Western blot and iELISA. Standardization of iELISA was performed with 500ng and 1µg BoP59 and rBP26 per well, testing serum from uninfected and experimentally infected rams. rBP26 was effective in distinguishing positive from negative rams. The rBP26 iELISA developed in this study is the first to use a completely purified rBP26 as antigen resulting in high sensitivity (100%) and specificity (90.2%), and an overall accuracy equal to 1.0.

This paper describes a research on Brucella ovis infection in rams in the Pirot Municipality of South Serbia. A positive result with indirect immunoenzyme test (i-ELISA) was confirmed in 67 (29.8%) and suspicious in 31 (13.8%) out of 225 tested rams. Complement fixation test (CFT) was used as a confirmation test on 67 ELISA positive sera and gave positive reaction in 41 (61.2%) ram serum samples. Rams originated from 113 flocks with 4751 sheep, from 28 villages in the Pirot Municipality of southern Serbia. Clinical examination was performed on epididymis and testes of 12 rams from 7 seropositive flocks by inspection and palpation. The examination showed scrotum asymmetry and unilateral increase of the epididymistail in 5 (41.7%) out of 12 seropositive rams. Pathomorphological examination of testes and epididymis confirmed pathological changes in 7 (58.3%) of the 12 examined rams. One-sided epididymitis with pronounced hypertrophy of the epididymitis was also confirmed. Twelve rams were tested for the presence of bacteria, i.e. 21 epididymis, testes and lymph nodes samples. We isolated 20 Brucella strains from 11 (91.7%) of the 12 examined animals. All isolates were identified with bacteriological and molecular techniques as B. ovis. This is the first evidence of ovine epididymitis (B. ovis) in Republic of Serbia.

The distribution of infection by the species Brucella ovis in rams and abortion cases in sheep in Croatia was investigated in the period from 2011 to 2015. The investigation relied on serological testing in rams and sheep after abortion conducted using a complement fixation test and immunosorbent assay conducted with bacteriological testing of the testes in seropositive rams and aborted material. Seropositive reactions were confirmed in 1100 (6%) of rams samples and in 12 (2.3%) of ewe samples. Isolated strains were also confirmed on the molecular level. In 10 (45.5%) animals asymmetry of the scrotum and unilateral enlargement of the tail of the epididymis were confirmed. Pathomorphological examination revealed visible changes such as granulomas, fibrosis, and atrophy of the testes and epididymis in 12 (54.5%) ram testes. In bacteriological investigation of samples from 22 rams and fetuses following abortion in ewes, 13 (59.1%) and 2 (14.3%) isolations were performed, respectively. All 15 isolates were identified as Brucella ovis using a molecular method. The results show that infectious epididymitis caused by Brucella ovis is distributed throughout the study area in Croatia. The disease most often spreads through uncontrolled trade or contacts of sheep between flocks, which significantly hinders eradication measures. [ABSTRACT FROM AUTHOR]

Ovine epididymitis (Brucella ovis) has been assessed according to the criteria of the Animal Health Law (AHL), in particular criteria of Article 7 on disease profile and impacts, Article 5 on the eligibility of ovine epididymitis to be listed, Article 9 for the categorisation of ovine epididymitis according to disease prevention and control rules as in Annex IV and Article 8 on the list of animal species related to ovine epididymitis. The assessment has been performed following a methodology composed of information collection and compilation, expert judgement on each criterion at individual and, if no consensus was reached before, also at collective level. The output is composed of the categorical answer, and for the questions where no consensus was reached, the different supporting views are reported. Details on the methodology used for this assessment are explained in a separate opinion. According to the assessment performed, ovine epididymitis can be considered eligible to be listed for Union intervention as laid down in Article 5(3) of the AHL. The disease would comply with the criteria as in Sections 3, 4 and 5 of Annex IV of the AHL, for the application of the disease prevention and control rules referred to in points (c), (d) and (e) of Article 9(1). The animal species to be listed for ovine epididymitis according to Article 8(3) criteria are mainly sheep and other species of the families Bovidae and Cervidae as susceptible and sheep and deer as reservoirs. [ABSTRACT FROM AUTHOR]

Infectious epididymitis is a reproductive disease of great sanitary and economic importance to the sheep industry. This study aimed to evaluate the occurrence of infectious epididymitis in the State of Mato Grosso (Brazil), and to identify the causative agents including: Brucella ovis , Actinobacillus seminis , and Histophilus somni . Antibodies anti- B. ovis were detected in 31.6% (152/480) of sheep, including rams (n = 83), by AGID and ELISA. Clinical changes, inflammatory cells in the semen and/or azoospermia were observed in 77.1% (64/83) of rams, and detection of B. ovis , A. seminis , and H. somni in urine and semen samples was associated with evidences of genital inflammation in some cases. These data demonstrated the presence of B. ovis , A. seminis and/or H. somni involved in reproductive tract infection in sheep flocks of Mato Grosso. [ABSTRACT FROM AUTHOR]