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Brucellosis, caused by various Brucella species, poses a significant threat to global public health and livestock industries. This study aims to fill the knowledge gap concerning the presence of Brucella spp. in rodents on livestock farms in Iran. Both bacteriological and molecular surveys were conducted to assess the prevalence of Brucella spp. in these rodent populations. A total of 16 rodents were captured in four seropositive dairy cattle farms (n = 7) and two seropositive sheep farms (n = 9) and were then examined for the presence of the Brucella-infection. Five cow milk samples and 53 bovine lymph node samples from these farms were also tested for Brucella spp. Lymph node samples from dairy cattle farms contained 32 B. abortus biovar 3 isolates and one B. melitensis Rev1 vaccine isolate. The bacterial culture of rodents identified 12.5% of them (Mus musculus and Rattus norvegicus) harboring Brucella strains in dairy cattle farms. The rodents had B. abortus biovar 3 and B. melitensis biovar 1, suggesting a reservoir for these bacteria. A two-step molecular assay, utilizing the Omp28 sequences in tissue samples of rodents, demonstrated that 68.75% (n = 11) of the tested rodents yielded positive results. Bruce-ladder PCR and wboA typing on isolated bacteria revealed a close relationship to field strain of Brucella species. The study reveals that rodents on seropositive livestock farms in Iran harbor Brucella spp., indicating a potential reservoir for these bacteria. This highlights the importance of monitoring rodent populations through the molecular and bacterial methods to manage and control brucellosis in livestock., Competing Interests: Declaration of competing interest Authors confirm that they have no conflict of interest of any sort with any one., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Animal reproduction biotechniques are important tools for the technological advancement of livestock, as they allow the selection of the reproductive potential of superior quality females and males; however, infectious diseases that have a predilection for the reproductive system can be a hindrance for the use of these technologies. Therefore, the present study aimed to detect Brucella spp. in the ovarian follicular fluid of brucellosis-positive bovine cows. A total of 47 bovine ovarian follicular fluid aspirates from cows, positive in tests for brucellosis and from Brucella-positive herd, were submitted to PCR. The primers used in the PCR were specific to the genus Brucella (bcsp31 gene). All 47 bovine aspirates were negative for Brucella spp. 0.00% (95% CI: 0.00-4.00%). Our results demonstrated that Brucella spp. was absent in the ovarian follicular fluid from seropositive cows, which indicates that Brucella spp.-infected cows could be used for reproductive biotechnologies carried out with follicular aspirates. Future studies are needed to more precisely evaluate the feasibility and safety of using these oocytes from brucellosis-seropositive cows to transfer embryos to heifers/cows not infected by Brucella, aiming to produce calves free of the infection., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

Brucella abortus is a facultatively extracellular-intracellular pathogen that encounters a diversity of environments within the host cell. We report that bacteria extracted from infected cells at late stages (48 h postinfection) of the intracellular life cycle significantly increase their ability to multiply in new target cells. This increase depends on early interaction with the cell surface, since the bacteria become more adherent and penetrate more efficiently than in vitro -grown bacteria. At this late stage of infection, the bacterium locates within an autophagosome-like compartment, facing starvation and acidic conditions. At this point, the BvrR/BvrS two-component system becomes activated, and the expression of the transcriptional regulator VjbR and the type IV secretion system component VirB increases. Using bafilomycin to inhibit BvrR/BvrS activation and using specific inhibitors for VjbR and VirB, we showed that the BvrR/BvrS and VjbR systems correlate with increased interaction with new host cells, while the VirB system does not. Bacteria released from infected cells under natural conditions displayed the same phenotype as intracellular bacteria. We propose a model in which the B. abortus BvrR/BvrS system senses the transition from its replicative niche at the endoplasmic reticulum to the autophagosome-like exit compartment. This activation leads to the expression of VirB, which participates in the release of the bacterium from the cells, and an increase in VjbR expression that results in a more efficient interaction with new host cells.

Brucellosis is among the most important zoonotic infectious diseases worldwide affecting both humans and domestic animals. The present study aimed to determine and compare the seroprevalence of brucellosis among rural and periurban dairy cattle farms of four Iranian provinces from 2017 to 2019. We applied different serological tests, including RBT, SAT, and iELISA to evaluate the brucellosis prevalence among 2808 dairy cattle. Species-specific multiplex PCR and biotyping tests were also used to further identify the implicated Brucella species. Serological screening using RBT, SAT, and iELISA led to 157 (5.6%), 112 (3.9%), and 139 (4.9%) positive results among tested cattle, respectively. Brucella abortus biovars 1 (2 cases) and biovars 3 (42 cases) were identified by biotyping experiments and multiplex PCR in all 44 tested lymph node samples. Further, Cohen's kappa statistical analysis revealed that the best degree of agreement was seen between RBT and iELISA (99.4%), followed by SAT/iELISA (98.5%) and finally RBT/SAT (98.4%). Our results also showed a significantly lower seroprevalence of brucellosis in periurban dairy cattle when compared to rural dairy cattle population (p value= 0.01). These results reflect the need for better vaccine coverage using RB51 combined with an appropriate test-and-slaughter program in the rural dairy cattle population.

In South Africa, brucellosis testing and record-keeping are done by several laboratories, thus it is difficult to access any organized data to assess the status of the disease. This study evaluated the seropositivity for brucellosis using Rose Bengal test and complement fixation test in suspect cattle, sheep, goats and pigs sera submitted to Bacterial Serology Laboratory, Agricultural Research Council-Onderstepoort Veterinary Research (ARC-OVR) from nine provinces in the country during the period 2007-2015. This retrospective data analysis was conducted to estimate the occurrence of brucellosis in the country from the submitted samples, identify variables that affected seropositivity for brucellosis, investigate existing gaps in data recording and make recommendations on important variables to facilitate better data capture and inferences on brucellosis. Nine years of data were collated and analysed to detect association (seropositivity over time regarding animal species and location). Of the 764,276 animals tested, the distribution of samples was 90.50% (691,539/764,276), 5.19% (39,672/764,276), 3.92% (29,967/764,276) and 0.41% (3,098/764,276) for cattle, sheep, goats and pigs, respectively. The seropositivity for brucellosis by animal species was 6.31% (43,666/691,539, 95% CI: 6.26-6.37), 2.09% (828/39,672, 95% CI: 1.95-2.23), 0.63% (189/29,967, 95% CI: 0.55-0.73) and 0.13% (4/3,098, 95% CI: 0.05-0.33) in cattle, sheep, goats and pigs respectively. The data available did not capture information on the age, sex, breed and other host risk factors that would have been related to seropositivity for brucellosis. The data provide an understanding of the disease occurrence and confirm that brucellosis is enzootic in South Africa. Improved and standardized data collection can be used to pro-actively drive, monitor, change or formulate policies to mitigate the challenges brought about by brucellosis in the livestock sector in South Africa., (© 2020 The Authors. Veterinary Medicine and Science Published by John Wiley & Sons Ltd.)

Brucellosis is a neglected endemic zoonosis in Bangladesh and has a significant impact on public health and animal welfare of dairy farming as well as dairy farm economics. A cross-sectional study was conducted to evaluate the seroprevalence of and risk factors for brucellosis in dairy cattle in the Chittagong metropolitan area (CMA) of Chittagong, Bangladesh. We collected serum samples (n = 158) from six randomly selected dairy farms in the CMA between February and November, 2015. The Rose Bengal Plate Test (RBPT) and a competitive ELISA (cELISA) were used as the screening and confirmatory tests respectively. Farm level and animal level demographic and risk factor data were collected using a questionnaire. The risk factors were analysed using a multivariable logistic regression with random effects. The overall seroprevalences of antibodies against brucellosis in cattle were 21.5% (34/158) and 7.6% (12/158) based on parallel and serial interpretation of the two tests respectively. Our results revealed that 20.3% (32/158) samples were positive using the RBPT and 8.9% (14/158) were positive using the cELISA. The within-herd seroprevalence ranged from 10% to 26.3% and 5 to 20.7% using the RBPT and cELISA tests respectively. The odds of seropositivity were significantly higher in lactating cows (OR: 2.59; 95% CI: 1.02-6.55), cows producing less than 2 litres of milk (OR: 29.6; 95% CI: 4.3-353.8), cow producing 2-12 litres of milk (OR: 4.8; 95% CI: 1.1-33.4) and cows with reproductive disorders (OR: 3.2; 95% CI: 1.2-10.1). About 7.6% (12/158) and 1.3% (2/158) of cattle were found to be infected with acute and chronic brucellosis respectively. Based on these results, we suggest that cows that have reproductive disorders and are producing little milk should be prioritized for brucellosis screening in CMA. The screening tests should be used to control brucellosis in cattle in order to protect animal welfare, human health and to minimize the economic losses., (© 2020 The Authors. Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

Bovine brucellosis is an infectious bacterial disease caused by members of genus Brucella, affecting both animals and humans, and resulting in a serious economic loss in animal production sector and deterioration of public health. A cross-sectional study was conducted from November 2014 to April 2015 to determine the seroprevalence and associated risk factors of bovine brucellosis in Sendafa, Oromia Special Zone, Ethiopia. A total of 503 blood samples were collected using a simple random sampling technique from dairy cattle of above 6 months of age with no history of previous vaccination against brucellosis. All sera samples were subjected to both Rose Bengal Plate Test for screening and Complement Fixation Test for confirmation. Accordingly, the overall seroprevalence of bovine brucellosis in the study area was 0.40%. The result showed that the seroprevalence of bovine brucellosis in the study area was not statistically significant with all proposed risk factors. No reactors were observed in male animals. The seroprevalence was observed in animals without previous history of abortion. Moreover, information was gathered on individual animal and farm-level risk factors and other farm characteristics using a questionnaire. Awareness among society was poor, so the positive animals can be a potential hazard to animals and humans in the study area. Therefore, public education should be done to improve the awareness of the community on bovine brucellosis and its public health impact with due consideration on the safe consumption of food of animal origin., Competing Interests: The authors have declared that no competing interests exist.

Brucellosis is a zoonotic disease of importance to both public health and the livestock industry. The disease is likely to be endemic in Tanzania and little is reported on molecular characterization of Brucella species in pastoral settings. This study aimed at characterizing Brucella species (targeting genus Brucella) infecting humans, cattle and goat in Kagera region (Ngara and Karagwe districts) using real-time PCR, PCR amplification of 16S rRNA genes and Sanger sequencing. Brucella spp. were detected in 47 samples (19 sera and 28 milk) out of 125 samples (77 sera, 35 milk and 13 aborted materials) using real-time PCR. All aborted materials (13 samples) were negative to real-time PCR. Out of the 47 real-time PCR positive samples (28 milk and 19 sera), 20 samples (10 milk and 10 sera) showed an expected 16S rRNA gene PCR product. Sequence analysis and blasting confirmed the presence of Brucella spp. in pastoral areas of Kagera region. The Brucella spp. from Kagera were phylogenetically grouped in two clades and three branches all closer to B. melitensis, B. abortus and B. suis from USA, Sudan and Iran. However, they were distinct from other species isolated also in USA, New Zealand, Germany and Egypt. This was expected based on the distance between the geographical regions from which the data (nucleotides sequences from 16S gene sequencing) for the phylogeny reconstruction were obtained. This is the first study to report Brucella species identified using 16S rRNA gene sequencing in East and Central Africa. A livestock vaccination program re-inforced with a high index of Brucella diagnosis is needed to eradicate brucellosis in animals and minimize suffering from Brucella infections in humans in Tanzania., (© 2020 The Authors. Veterinary Medicine and Science Published by John Wiley & Sons Ltd.)

Bovine brucellosis, cause by infection with Brucella abortus , causes reproductive failure in cattle, has a major economic impact to producers, and as a zoonoses, it is a disease of public health concern. Characterization of the protective immune response against Brucella infection is important to our understanding of disease pathogenesis and for the development of diagnostic assays and vaccines. Most of the knowledge regarding protection against Brucella comes from studies in the murine model, but less is known about the immune responses in cattle. Assessment of antigen-specific T cell frequency and functional phenotype are critical to understand the immune status of the host, characterize mechanisms of protective immunity and immunopathology, and to predict immune protection. The frequency of circulating T cells specific for a particular pathogen is often very low, making analysis of such responses difficult. Our goal was to develop a flow-cytometry based approach to better track Brucella -specific T cell responses. Using peripheral blood mononuclear cells (PMBC) from Brucella abortus strain RB51-vaccinated cattle, we optimized an in vitro stimulation protocol based on a combination of antigen and pan-T cell stimulation. We then assessed RB51-specific T cell responses by concurrently measuring proliferation and cytokine production using flow-cytometry. This methodology enhances the detection of peripheral, Brucella -specific responses in cattle following RB51 vaccination. This protocol is versatile in that it can be modified to fit other in vitro stimulation systems and additional functional or phenotypic parameters can be added for flow cytometric detection and characterization of antigen-specific T cells., (Copyright © 2020 Boggiatto, Schaut and Olsen.)

Background: In the recent past (1997-2012), Northern Ireland in the United Kingdom suffered an outbreak of Brucella abortus, which at its height affected over 200 cattle herds. Initially, isolates were characterized using multi-locus variable number tandem repeats analysis (MLVA). While informative in this setting, hyper-variability in some loci limited the resolution necessary to infer fine-scale disease transmission networks. Consequently, we applied whole-genome sequencing to isolates from this outbreak to evaluate higher resolution markers for disease epizootiology., Results: Phylogenetic analysis revealed that the B. abortus outbreak in Northern Ireland was caused by two distinct pathogen lineages. One contained isolates consistent with the 1997-2012 outbreak being linked to a previous endemic infection thought eradicated. The dominant second lineage exhibited little genetic diversity throughout the recrudescent outbreak, with limited population sub-structure evident. This finding was inconsistent with prior MLVA molecular characterizations that suggested the presence of seven clonal complexes. Spatio-temporal modeling revealed a significant association of pairwise SNP differences between isolates and geographic distances. However, effect sizes were very small due to reduced pathogen diversity., Conclusions: Genome sequence data suggested that hyper-variability in some MLVA loci contributed to an overestimate of pathogen diversity in the most recent outbreak. The low diversity observed in our genomic dataset made it inappropriate to apply phylodynamic methods to these data. We conclude that maintaining data repositories of genome sequence data will be invaluable for source attribution/epizootiological inference should recrudescence ever re-occur. However genomic epizootiological methods may have limited utility in some settings, such as when applied to recrudescent/re-emergent infections of slowly-evolving bacterial pathogens., Competing Interests: Declaration of Competing Interest All authors declare no conflict of interest., (Crown Copyright © 2020. Published by Elsevier B.V. All rights reserved.)

Brucellosis, caused by Brucella abortus, is a major disease of cattle and humans worldwide distributed. Eradication and control of the disease has been difficult in Central and South America, Central Asia, the Mediterranean and the Middle East. Epidemiological strategies combined with phylogenetic methods provide the high-resolution power needed to study relationships between surveillance data and pathogen population dynamics, using genetic diversity and spatiotemporal distributions. This information is crucial for prevention and control of disease spreading at a local and worldwide level. In Costa Rica (CR), the disease was first reported at the beginning of the 20th century and has not been controlled despite many efforts. We characterized 188 B. abortus isolates from CR recovered from cattle, humans and water buffalo, from 2003 to 2018, and whole genome sequencing (WGS) was performed in 95 of them. They were also assessed based on geographic origin, date of introduction, and phylogenetic associations in a worldwide and national context. Our results show circulation of five B. abortus lineages (I to V) in CR, phylogenetically related to isolates from the United States, United Kingdom, and South America. Lineage I was dominant and probably introduced at the end of the 19th century. Lineage II, represented by a single isolate from a water buffalo, clustered with a Colombian sample, and was likely introduced after 1845. Lineages III and IV were likely introduced during the early 2000s. Fourteen isolates from humans were found within the same lineage (lineage I) regardless of their geographic origin within the country. The main CR lineages, introduced more than 100 years ago, are widely spread throughout the country, in contrast to new introductions that seemed to be more geographically restricted. Following the brucellosis prevalence and the farming practices of several middle- and low-income countries, similar scenarios could be found in other regions worldwide., Competing Interests: The authors have declared that no competing interests exist.

The English surveillance system for bovine brucellosis was evaluated. The confidence in detecting at least one infected herd in the local population (surveillance system sensitivity or SSe), and the confidence in freedom from disease (PFree) adjusted (PFreeAdj) for the probability of disease introduction from abroad by imported animals (PIntro), were estimated for quarterly surveillance periods of 2016; because dairy herds were tested quarterly on bulk tank milk (BTM) with an antibody indirect ELISA. A stochastic model was developed and six surveillance components (representing also the local population strata), were evaluated. All English herds and their relative risk (RRs) of infection within each stratum were considered. The importance of each component was assessed using actual national data, which reflected non-random sampling. The contribution of the abortions testing was assessed with particular focus, because a decline in statutory submissions was observed in recent years. Beef herds without submissions (B-NoTest herds) at the laboratories were still considered as a population stratum, where infected cattle could be imported. Additionally, we evaluated the importance of different hypothetical design between-herds prevalence (Ph) values, at which the country could be classified as "infected". The potential negative effect on SSe due to the dilution of antibodies when individual samples are pooled within BTM and tested by the milk iELISA, was also investigated. The quarterly median SSe and PFreeAdj were both ≥ 95 % if at least four (0.008 %) herds were infected in the country due to independent import events. The system appeared able to substantiate Official Brucellosis Free (OBF) status frequently (on quarterly basis) using Ph=0.2 % (EU legislation). The component based only on BTM testing (M herds) showed the highest sensitivity; while the surveillance components based on abortions or post import calving (PIC) testing, had very low sensitivity at the (considered) Ph values lower than 0.2 %. In contrast, at Ph = 0.2 %, components based on abortion testing had median sensitivity between 91.3 % and 99.9 %, and the dilution effect on BTM testing did not change remarkably the SSe and PFreeAdj. When Ph was set to 1-2 infected herds (0.002-0.004 %), these were usually allocated by the model within the B-NoTest stratum (the largest stratum) and SSe reduced. Thus, if policy considers necessary increasing the SSe for low Phs (system's optimization as an early warning system); the cost efficiency of active risk based surveillance in beef herds (considering imports) could be investigated in the future., Competing Interests: Declaration of Competing Interest None., (Crown Copyright © 2020. Published by Elsevier B.V. All rights reserved.)

Background: Brucellosis is a zoonosis whose incidence is not declining worldwide despite the global effort to control the disease. Accurate and precise diagnosis is a crucial step in any prophylaxis program but single tests to unequivocally detect animals infected with Brucella spp. are currently unavailable. In Italy, serological diagnosis of bovine brucellosis is performed with two official tests: a rapid agglutination test (i.e., Rose Bengal Plate test, RBPT) and a complement fixation test (CFT) that detect antibodies directed mainly to the smooth lipopolysaccharide (S-LPS). Neither of the two tests is able to avoid the detection of false positive serological reactions (FPSRs) caused by bacteria sharing S-LPS components with Brucella spp. and responsible for the single reactors (SR) phenomenon. A B. melitensis R strain-based ELISA showed a good diagnostic performance in unravelling FP animals; however, since a limited number of animals were analyzed in that study, a large field study was conducted here to discriminate between Brucella-infected from FP animals, with the final aim of reducing the unnecessary slaughter of the latter. An ELISA based on a R strain of Brucella, i.e., Brucella melitensis B115, was employed to measure specific IgG responses in a collection of bovine sera (n = 648). Sera were obtained from 180 farms (either officially brucellosis-free or not brucellosis-free) recruited during an extended period of time (2014-2018) and were preliminarily assayed with the official tests by the Italian Reference Centers and then subjected to the ELISA., Results: Negative sera, when subjected to the ELISA, gave O.D. values below the cutoff; SR sera, i.e. RBPT positive and CFT negative, as well as double positive (DP) sera, i.e. RBPT and CFT positive, gave O.D. values that were below the cutoff. All positive sera, i.e. from Brucella-infected animals, were RBPT positive and CFT positive (ICFTU ranging from 20 to 1280) and gave ELISA O.D. values above the cutoff., Conclusions: The B. melitensis B115-based ELISA systematically unravelled all false positive (FP) sera while confirming the diagnosis in Brucella-infected animals. Thus, the test employed in the present study may complement the official assays to avoid the costly slaughter of FP animals.

To determine the prevalence of antibodies to Brucella melitensis, Brucella abortus and Coxiella burnetii in animals on Caribbean islands we obtained sera from convenience samples of cattle (C), sheep (S), goats (G) and cats (F) from Dominica (C, S, G), Grenada (C, S, G), Montserrat (C, S, G), Puerto Rico (C), Nevis (C, S, G), St Kitts (C, S, G, F) and St Lucia (C, G). The sera were tested for antibodies against the Brucella spp. using commercial ELISA kits. Some sera were also tested at 1/80 for antibodies to C. burnetii using an indirect fluorescent antibody test. Positive sera were also tested at 1/640. None of 599 cattle, 462 sheep or 434 goats were positive in the Brucella ELISAs. None of 230 cattle had antibodies against C. burnetii, but one of 299 sheep was positive at 1/80 (Dominica - 1/54, 2%, 95% CI (0%-5.6%)), as were two of 314 goats, at 1/80 (Grenada - 1/53, 2%, 95% CI (0%-7.5%)) and 1/640 (St Kitts - 1/18, 5.6%, 95% CI (0%-16.7%)), and one of 34 cats, at 1/80 (St Kitts - 1/34; 3%, 95% CI (0%-8.8%)). Our data suggests that there is a very low prevalence or absence of B. melitensis and B. abortus on Caribbean islands. Coxiella burnetii, however, is present but it appears to be present on only some islands and then only at low levels. Overall, there appears to be a low threat to human and animal health from these organisms in the Caribbean., (© 2019 The Authors. Veterinary Medicine and Science Published by John Wiley & Sons Ltd.)

Background: Brucellosis is a zoonotic disease caused by bacteria Brucella spp. belonging to the genus Brucella. It is endemic in domesticated animals in Bangladesh. Isolation, identification and genetic characterization of Brucella spp. in dairy cattle are essential to undertake appropriate control and preventive measures. The study was conducted to isolate and characterize the Brucella spp. circulating in dairy cattle., Methods: Uterine discharge (n = 45), milk (n = 115), vaginal swab (n = 71), placenta (n = 7) and aborted fetus (n = 2) were collected. Brucella selective agar plates were inoculated with samples and incubated at 37 ◦ C for 14 days under 5% CO 2 for isolation of Brucella spp. Brucella suspected colonies were recovered from samples were confirmed by genus and species specific PCR assays. Genetic characterization was performed by Multi Locus Variable number tandem-repeat Analysis-16 (MLVA-16)., Results: The isolates of Brucella recovered from samples were confirmed as B. abortus by AMOS-ERY PCR assay. The classical biotyping method confirmed all 10 B. abortus isolates belonged to the biovar 3. The MLVA-16 assay indicated all B. abortus isolates identical and the same genotype 40, based on panel 1 MLVA-8., Conclusion: Dendrogram analysis revealed all B. abortus isolates of the study were identical to three isolates from Brazil, one isolate of France and closely related to Chinese isolates. This is the first report of isolation and genetic characterization of B. abortus from the dairy cattle in Bangladesh., (© 2019 The Authors. Veterinary Medicine and Science Published by John Wiley & Sons Ltd.)

Background: Brucellosis is an infectious and contagious zoonotic bacterial disease of both humans and animals. In developing countries where brucellosis is endemic, baseline data on the prevalence of brucellosis, using abattoir facilities, is important., Objectives: The aim of this study was to determine the seroprevalence of antibodies against Brucella in slaughter cattle at Gauteng province, South Africa and to characterize isolates of Brucella spp., Methods: In this cross-sectional study, un-clotted blood samples with corresponding organ tissue samples were collected from slaughtered cattle. Serological [Rose Bengal test (RBT), complement fixation test (CFT) and indirect ELISA (iELISA)], molecular (PCR) and bacteriological methods were used to detect Brucella antibodies and Brucella spp. from 200 slaughtered cattle in 14 abattoirs., Results: The RBT revealed a seroprevalence of brucellosis as 11.0% (22 of 200) and iELISA confirmed 5.5% (11 of 200). The estimated seroprevalence from RBT and iELISA was 5.5% while RBT and CFT was 2.0% (4 of 200). Brucella melitensis (n = 6) and B. abortus (n = 5) were isolated from 11 cattle tissues (5.5%) as confirmed to species level with AMOS PCR and differentiated from vaccine strains with Bruce-ladder PCR. Seven of the 11 isolates originated from seropositive cattle of which five were biotyped as B. abortus bv 1 (n = 2) and B. melitensis bv 2 (n = 1) and B. melitensis bv 3 (n = 2)., Conclusions: This is the first documentation of B. melitensis in cattle in South Africa. The zoonotic risk of brucellosis posed by Brucella-infected slaughter cattle to abattoir workers and consumers of improperly cooked beef cannot be ignored., (© 2019 The Authors. Veterinary Medicine and Science Published by John Wiley & Sons Ltd.)

The objective of this study was to identify twelve Brucella abortus isolates of bovine origin from the department of Nariño in Colombia up to the biovar level. These isolates are included in the collection of the Germplasm Bank of Microorganisms of Animal Health Interest - Bacteria and Virus (BGSA-BV). The identification was carried out through conventional methods such as macro and microscopic morphological descriptions, enzymatic activity, biochemical profile, substrate use and sensitivity to dyes. Complementary genotypic characterization was carried out using multiplex PCR for B. abortus, Brucella melitensis, Brucella ovis, and Brucella suis-Erytritol (AMOS-ERY-PCR), RFLP-IS711, by southern blot hybridization, as well as by the multiple locus variable number of tandem repeat analysis (MLVA) using the ery gene and the insertion sequence IS711 and variable number of tandem repeats (VNTR) as molecular markers. The results of the phenotypic and molecular characterization allowed to identify twelve isolates as B. abortus biovar 4 as well as to differentiate field from vaccine strains. This is the first study on the phenotypic and molecular identification of B. abortus isolates in Colombia. It was concluded that the phenotypic and molecular identification of twelve isolates as B. abortus biovar 4 could be achieved using conventional and molecular techniques with enough resolution power. The identification of these isolates to the biovar level in taxonomic and epidemiological terms will allow the use of this genetic resource as reference strains in future research. This finding constitutes the basis for identifying biotypes not previously reported in the country that might be useful to support brucellosis survey programs in Colombia., (Copyright © 2018 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Objective: In the study, the laboratory results of 150 bovine abortion cases from 2018 (January-September) are presented., Material and Methods: Depending on the submitted sample material and the requested examination, serological, bacteriological and/or molecular biological investigations were performed to detect abortion-causing pathogens which need or do not need to be notified in Austria., Results: In addition to animal pathogens, the zoonotic pathogens Brucella melitensis and Salmonella Dublin were detected in 1 case each and Coxiella burnetii in 2 fetuses., Conclusion: The results show, that because of the zoonotic potential of some pathogens, care must be taken when handling abortion material to ensure that farmers, veterinary surgeons and laboratory staff are not at risk. Taking bovine brucellosis as an example, the reappearance of previously eradicated diseases has to be expected at any time., Clinical Relevance: For detailed diagnostics, fetus with placenta and blood samples from the dam should be submitted to the laboratory. According to the extensive pathogen spectrum, investigation of abortion cases is laborious and time consuming., Competing Interests: Die Autoren bestätigen, dass kein Interessenkonflikt besteht., (© Georg Thieme Verlag KG Stuttgart · New York.)

This paper describes the global distribution and temporal evolution of bovine brucellosis due to Brucella abortus during a 19-year period (1996-2014) using the information officially reported to the World Organisation for Animal Health (OIE) by veterinary services of 156 countries. Variables that can influence the health status of bovine brucellosis (i.e., year, per capita Gross Domestic Product (GDP), continent and bovine population) were also analysed. Countries were classified into three categories of health situations: ENZOOTIC: countries infected, which may have been free of brucellosis but for periods of fewer than 3 years; NON-ENZOOTIC: countries where the disease was present but that had at least a 3-year period without the disease; and FREE: countries where the disease remained absent during the whole period. The countries free from bovine brucellosis, or in the process of eradication, were located in Oceania and Europe, while the more affected regions were Central and South America, Africa and parts of Asia. Among the Non-Enzootic countries, the results showed that a very high proportion managed to control the disease during the period of study, with a sharp decline in the percentage of infected countries from 71% in 1996 to 10% in 2014. Among the Enzootic countries, a much smaller proportion managed to control the disease, with a slight drop in the percentage of infected countries from 92% in 1996 to 80% in 2014. A relationship was found between the status of the disease and the availability of economic resources; thus, countries with a high GDP per capita tended to be free from bovine brucellosis. On the other hand, countries with a larger bovine population showed a greater probability to have the disease present. An increase in surveillance programmes and implementation of control policies were observed during the period of study., (© 2019 Blackwell Verlag GmbH.)

Bovine brucellosis poses a risk to human health and causes serious economic losses for the animal industry. This report describes the use of different diagnostic methods for the diagnosis of brucellosis in cattle affected by cervical bursitis from a slaughterhouse located in São Luís, Maranhão, Brazil. Serum samples from a total of 47 cattle with bursitis were collected and submitted to the Rose Bengal Test (RBT), and RBT-positive samples were further confirmed by the 2-mercaptoethanol (2-ME) assay. RBT indicated 85.1% (40/47) of positive samples, from which 78.7% (37/47) were confirmed by 2-ME. Immunohistochemistry detected Brucella spp. in 34.0% (16/47) of tissues with bursitis. PCR and/or bacterial isolation demonstrated that 63.8% (30/47) of samples were positive and morphologically compatible with Brucella sp. All colonies suggestive of Brucella sp. were confirmed by PCR. Isolates were further characterized by PCR Multiplex AMOS-ENHANCED, which indicated that the isolates corresponded to biovar 1, 2, 4 (43.33%). This study evidences an association between cervical bursitis and Brucella spp. infection in cattle, and that different biovars of Brucella circulate in bovine herds in Maranhão.

A total of 27 seminal plasma samples from cattle-breeding farms or semen centres located in Minas Gerais, Brazil, previously negative by serological and tested positive for Brucella spp. with primer specific for the amplification of the gene virb5 by polymerase chain reaction (PCR) were analysed for the detection of Brucella abortus DNA by PCR. It was found that nine samples (33.33%) contained B. abortus B19 strain DNA, two (7.40%) contained B. abortus DNA and five (18.51%) contained both DNA. The larger number of samples with B. abortus B19 strain DNA would explained by the environmental contamination by vaccinated females with persistent excretion or some illegal vaccination process. It is first reported of male bovines detected with both DNA., (© 2017 Blackwell Verlag GmbH.)

Brucellosis is of worldwide economic and public health importance. Heifer vaccination with live attenuated Brucella abortus strain 19 (S19) is the cornerstone of control in low- and middle-income countries. Antibody persistence induced by S19 is directly correlated with the number of colony-forming units (CFU) per dose. There are two vaccination methods: a 'high' dose (5-8 × 1010 CFU) subcutaneously injected or one or two 'low' doses (5 × 109 CFU) through the conjunctival route. This study aimed to evaluate serological reactions to the 'high' dose and possible implications of the serological findings on disease control. This study included 58 female cases, vaccinated at Day 0, and 29 male controls. Serum was drawn repeatedly and tested for Brucella antibodies using the Rose Bengal Test (RBT) and an indirect enzyme-linked immunosorbent assay (iELISA). The cases showed a rapid antibody response with peak RBT positivity (98%) at 2 weeks and iELISA (95%) at 8 weeks, then decreased in an inverse logistic curve to 14% RBT and 32% iELISA positive at 59 weeks and at 4.5 years 57% (4/7 cases) demonstrated a persistent immune response (RBT, iELISA or Brucellin skin test) to Brucella spp. Our study is the first of its kind documenting the persistence of antibodies in an African communal farming setting for over a year to years after 'high' dose S19 vaccination, which can be difficult to differentiate from a response to infection with wild-type B. abortus. A recommendation could be using a 'low' dose or different route of vaccination.

Brucellosis is a worldwide extended zoonosis with a heavy economic and public health impact. Cattle, sheep and goats are infected by smooth Brucella abortus and Brucella melitensis, and represent a common source of the human disease. Brucellosis diagnosis in these animals is largely based on detection of a specific immunoresponse. We review here the immunological tests used for the diagnosis of cattle brucellosis. First, we discuss how the diagnostic sensitivity (DSe) and specificity (DSp), balance should be adjusted for brucellosis diagnosis, and the difficulties that brucellosis tests specifically present for the estimation of DSe/DSp in frequentistic (gold standard) and Bayesian analyses. Then, we present a systematic review (PubMed, GoogleScholar and CABdirect) of works (154 out of 991; years 1960-August 2017) identified (by title and Abstract content) as DSe and DSp studies of smooth lipopolysaccharide, O-polysaccharide-core, native hapten and protein diagnostic tests. We summarize data of gold standard studies (n = 23) complying with strict inclusion and exclusion criteria with regards to test methodology and definition of the animals studied (infected and S19 or RB51 vaccinated cattle, and Brucella-free cattle affected or not by false positive serological reactions). We also discuss some studies (smooth lipopolysaccharide tests, protein antibody and delayed type hypersensitivity [skin] tests) that do not meet the criteria and yet fill some of the gaps in information. We review Bayesian studies (n = 5) and report that in most cases priors and assumptions on conditional dependence/independence are not coherent with the variable serological picture of the disease in different epidemiological scenarios and the bases (antigen, isotype and immunoglobulin properties involved) of brucellosis tests, practical experience and the results of gold standard studies. We conclude that very useful lipopolysaccharide (buffered plate antigen and indirect ELISA) and native hapten polysaccharide and soluble protein tests exist, provided they are applied taking into account the means available and the epidemiological contexts of this disease: i) mass vaccination; ii) elimination based on vaccination combined with test-and-slaughter; and iii) surveillance and existence of false positive serological reactions. We also conclude that the insistence in recent literature on the lack of usefulness of all smooth lipopolysaccharide or native hapten polysaccharide tests in areas where S19 vaccination is implemented is a misinterpretation that overlooks scientific and practical evidence., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The health and well-being of cattle is an important issue in maintaining and increasing global agricultural output. In dairy production within low and middle income countries (LMICs), there is a significant biosensing challenge in detecting sexually transmitted infection (STI) pathogens during animal husbandry, due in part to difficulties associated with the limited infrastructure for veterinary medicine. Here we demonstrate low-cost, multiplexed, and sample-to-answer paper-origami tests for the detection of three bovine infectious reproductive diseases in semen samples, collected at a test site in rural India. Pathogen DNA from one viral pathogen, bovine herpes virus-1 (BoHV-1), and two bacteria (Brucella and Leptospira) was extracted, amplified (using loop-mediated isothermal amplification, LAMP), and detected fluorescently, enabling <1 pg (∼ from 115 to 274 copies per reaction) of target genomic DNA to be measured. Data was collected as a fluorescence signal either visually, using a low-cost hand-held torch, or digitally with a mobile-phone camera. Limits of detection and sensitivities of the paper-origami device for the three pathogens were also evaluated using pathogen-inoculated semen samples and were as few as 50 Leptospira organisms, 50 CFU Brucella, and 1 TCID 50 BoHV-1. Semen samples from elite bulls at a germplasm center were also tested in double-blind tests, as a demonstrator for a low-cost, user-friendly point-of-care sensing platform, for in-the-field resource-limited regions. The sensors showed excellent levels of sensitivity and specificity, and for the first time a demonstrated ability of the application of paper microfluidics devices for the diagnosis multiple infectious diseases from semen samples.

Brucella abortus induces an inflammatory response that stimulates the endocrine system resulting in the secretion of cortisol and dehydroepiandrosterone (DHEA). Osteoarticular brucellosis is the most common presentation of the active disease in humans, and we have previously demonstrated that B. abortus infection inhibits osteoblast function. We aimed to evaluate the role of cortisol and DHEA on osteoblast during B. abortus infection. B. abortus infection induces apoptosis and inhibits osteoblast function. DHEA treatment reversed the effect of B. abortus infection on osteoblast by increasing their proliferation, inhibiting osteoblast apoptosis, and reversing the inhibitory effect of B. abortus on osteoblast differentiation and function. By contrast, cortisol increased the effect of B. abortus infection. Cortisol regulates target genes by binding to the glucocorticoid receptor (GR). B. abortus infection inhibited GRα expression. Cell responses to cortisol not only depend on GR expression but also on its intracellular bioavailability, that is, dependent on the activity of the isoenzymes 11β-hydroxysteroid dehydrogenase (HSD) type-1, 11β-HSD2 (which convert cortisone to cortisol and vice versa , respectively). Alterations in the expression of these isoenzymes in bone cells are associated with bone loss. B. abortus infection increased 11β-HSD1 expression but had no effect on 11β-HSD2. DHEA reversed the inhibitory effect induced by B. abortus infection on osteoblast matrix deposition in an estrogen receptor- and ERK1/2-dependent manner. We conclude that DHEA intervention improves osteoblast function during B. abortus infection making it a potential candidate to ameliorate the osteoarticular symptoms of brucellosis.

Bovine brucellosis is endemic in Colombia, and is a mandatory notifiable disease, subjected to a control program based on four surveillance procedures: passive surveillance, test-and-remove, certification of disease-free farms, and animal movements. The objective of this study is to estimate the evolution of bovine brucellosis in Colombia over a 7-year period (2006-2012) using data from the official control program. A total of 58 epidemiologic variables were analyzed for each year at the department level. Univariate descriptive analysis and principal components analysis (PCA) were performed to ascertain the behavior of the variables. These programs covered 3% of the census in 2006, increasing to 15% in 2012. The percentage of positive farms averaged 22% in 2006 and 23% in 2012. The highest proportion of positive farms was in the Orinoquía region (24.6 to 49.6%); the lowest was in the Amazon region, (17.9 to 32.7%). The percentage of positive animals presented certain differences between years but without any clear trend (4.7% in 2006 and 4.6% in 2012), indicating that the brucellosis control program had a low impact in Colombia in these years. The results for each surveillance procedure were 6.8% for passive surveillance, 5.9% for test-and-remove, and 4.4% both in disease-free farms and in animal movement tests. The results obtained by PCA led to finding three different clusters: geographic areas with low bovine production and low bovine brucellosis surveillance, areas with medium bovine production and medium surveillance for bovine brucellosis, and areas with a predominant bovine production, applying sanitary measures to control bovine brucellosis.

The present study aimed to estimate the herd-level sensitivity (Se) and specificity (Sp) of three commonly used serological tests in naturally-infected cattle and buffalo in smallholder farms in Pakistan. Between February and June 2015, a cross-sectional study was carried out in five districts of Punjab (Kasur, Okara, Pakpattan, Jhelum, and Bhakkar) and two districts of Sindh (Badin and Thatta). Serum samples from mixed farms of cattle (n=441) and buffalo (n=621) were collected and tested using the Rose Bengal test (RBT), indirect ELISA (I-ELISA) and competitive ELISA (C-ELISA). A Bayesian latent class analysis (LCA) approach was used to estimate the Se and Sp of these three serological tests and the true herd-level prevalence in each district. The model was fitted under the assumption of conditional independence between three tests and also conditional dependence by including covariances between the two ELISAs. In addition, the model was fitted using three different shapes of beta distributions to incorporate prior information in the model. The test with the highest Se was the C-ELISA, with a range from 76.3% (95% PCI (Posterior Credibility Interval), 62.6-88.2%) to 81.4% (95% PCI, 68.2-92.8%). The RBT was found to have the highest Sp (99.1-99.4%) of the tests. The highest estimated herd-level prevalence, 45% (95% PCI, 32-59%), was found in Jhelum district and the lowest in Thatta district, 1.1% (95% PCI 0.04-6.0%). The results of this study identified some discrepancy between the published literature on the level of Se of these tests, especially for RBT. It appears that RBT has lower Se and higher Sp when used in the field conditions of the present study. Consequently, it is recommended that none of the evaluated tests should be performed in isolation for the diagnosis of bovine brucellosis in the field conditions of Pakistan, but the use of tests in combination, with RBT and C-ELISA used in parallel returning optimal Se and Sp, is warranted., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The aim of the study was to investigate the farmers' socio-economic factors and their association with Brucella prevalence in the dry zone of Sri Lanka. A cross-sectional survey was planned and a total of 1,153 blood samples were collected from milking and dry animals of 155 farms from three selected veterinary ranges of Kalmunai, Navithanveli, and Mahaoya in the Ampara district, which is a multi-ethnic area. The 　Rose Bengal Test (RBT) and competitive enzyme-linked immunosorbent assay (c-ELISA) were used for the Brucella screening and confirmation, respectively. Socio-economic attributes such as family income, poverty, education, main job, ethnicity, parent farmer, farming experience, and training in animal husbandry were determined as potential farmer-level risk factors. Meanwhile, herd size, grazing practice, breeding method, animal brought-in to the farm, and abortions were considered as herd factors. The results revealed that the overall animal level sero-prevalence of brucellosis was 2.7% (35/1153; 95% confidence interval [CI]: 1.7, 3.7%) and the herd prevalence was 9.6% (15/155; 95% CI: 5.7, 15.7%) in the area of study. Brucellosis prevalence varies significantly (p<0.001) among the selected veterinary ranges with the highest herd prevalence in Kalmunai (20.0%) followed by Navithanveli (11.9%) and Mahaoya (2.7%). Disease prevalence showed variability (p<0.001) among ethnicities with the highest in Muslims (27.3%) followed by Tamils (8.1%) and Sinhalese (2.7%). Poverty was highly associated (OR=3.75; 95% CI: 1.43-10.00) with the disease. Free movement grazing practices (p<0.01) with OR=7.2 and animal brought-in from outside (p<0.06) with OR=3.06 were positively related to brucellosis. It was revealed that farmers' socio-economics, such as ethnicity and poverty, and animal movement patterns, such as grazing practices are significantly associated with epidemiology of brucellosis in the dry zone of Sri Lanka. Therefore, the "farmer factor" should be carefully considered in veterinary epidemiological studies and animal disease control plans in the future.　., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Brucellosis is highly contagious bacterial zoonoses affecting a wide range of domesticated and wild animals. In this study, Brucella (B.) abortus bv 1 was identified in uterine discharge of apparently healthy bitch and queen with open pyometra housed on a cattle farm. This study highlights the role of dogs and cats as symptomatic carriers and reservoirs for Brucella. To the best of our knowledge, this study represents the first report of feline infection with B. abortus bv 1 globally. These pet animals may contaminate the environment and infect both livestock and humans. Surveillance and control programmes of brucellosis have to include eradication of the disease in dogs, cats and companion animals., (© 2016 Blackwell Verlag GmbH.)

A cross-sectional study was conducted in three counties (Damxung, Maizhokunggar and Yadong) in Tibet in April and May 2015. A total of 1,523 yaks owned by 181 herders were randomly selected and blood sampled. Sera were tested using the rose bengal test (RBT) and a competitive immune-enzymatic assay (C-ELISA) and the test results interpreted in parallel. The individual yak prevalence was 2.8% (95% CI 2.0-3.7) with a herd prevalence of 18.2% (95% CI 12.9-24.6). At the individual level, two predictor variables, age and production system, were significantly associated with seropositivity by a binary logistic regression analysis. The odds of Brucella infection were significantly higher in older Yaks (3-5 years old, OR = 4.51; 95% CI 1.53-19.29; ≥6 years old, OR = 3.89; 95% CI 1.23-17.21) compared to those of younger yaks (≤2 years old). The odds of seropositivity for yaks managed under an agro-pastoral production system were 2.9 (95% CI 1.48-5.86) times higher compared to those managed under a pastoral production system. At the herd level, an association between the infection with Brucella and a history of abortions in the herd was observed (OR = 4.98, 95% CI 1.48-16.62). Surprisingly, vaccination was not associated with a lower level of infection (p = 0.49 and p = 0.99 for individual and herd level data, respectively). The results of the survey indicate that bovine brucellosis is endemic among the yak population in the plateau region of China, and the risk factors identified in the study should be considered in the epidemiology of the disease and when developing control programs for the disease.

Serology is primarily used in the diagnosis of bovine brucellosis. Bacterial culture and isolation is the gold standard in diagnosing brucellosis but, like serology, it does not offer complete (100%) diagnostic sensitivity and specificity. Polymerase chain reaction (PCR) has been suggested to offer better specificity and sensitivity. In this study, we evaluated the performance of Brucella abortus species specific (BaSS) PCR directly from different samples in the diagnosis of bovine brucellosis in naturally infected cattle in KwaZulu-Natal province of South Africa with known infectious status from culture. The BaSS PCR had a low diagnostic sensitivity (DSe) of 70%, but was able to identify vaccine strains using abomasal fluid from aborted foetuses and detect Brucella DNA from decomposing samples. The best sample for the BaSS PCR was abomasal fluid.

Surveillance systems of exotic infectious diseases aim to ensure transparency about the country-specific animal disease situation (i.e. demonstrate disease freedom) and to identify any introductions. In a context of decreasing resources, evaluation of surveillance efficiency is essential to help stakeholders make relevant decisions about prioritization of measures and funding allocation. This study evaluated the efficiency (sensitivity related to cost) of the French bovine brucellosis surveillance system using stochastic scenario tree models. Cattle herds were categorized into three risk groups based on the annual number of purchases, given that trading is considered as the main route of brucellosis introduction in cattle herds. The sensitivity in detecting the disease and the costs of the current surveillance system, which includes clinical (abortion) surveillance, programmed serological testing and introduction controls, were compared to those of 19 alternative surveillance scenarios. Surveillance costs included veterinary fees and laboratory analyses. The sensitivity over a year of the current surveillance system was predicted to be 91±7% at a design prevalence of 0.01% for a total cost of 14.9±1.8 million €. Several alternative surveillance scenarios, based on clinical surveillance and random or risk-based serological screening in a sample (20%) of the population, were predicted to be at least as sensitive but for a lower cost. Such changes would reduce whole surveillance costs by 20 to 61% annually, and the costs for farmers only would be decreased from about 12.0 million € presently to 5.3-9.0 million € (i.e. 25-56% decrease). Besides, fostering the evolution of the surveillance system in one of these directions would be in agreement with the European regulations and farmers perceptions on brucellosis risk and surveillance.

Brucellosis, caused by Brucella abortus is a major disease of cattle and a zoonosis. In order to estimate the bovine brucellosis prevalence in Costa Rica (CR), a total 765 herds (13078 bovines) from six regions of CR were randomly sampled during 2012-2013. A non-random sample of 7907 herds (532199 bovines) of the six regions, arriving for diagnoses during 2014-2016 to the Costa Rican Animal Health Service was also studied. The prevalence estimated by Rose Bengal test (RBT) ranged from 10.5%-11.4%; alternatively, the prevalence estimated by testing the RBT positives in iELISA, ranged from 4.1%-6.0%, respectively. However, cattle in CR are not vaccinated with B. abortus S19 but with RB51 (vaccination coverage close to 11%), and under these conditions the RBT displays 99% specificity and 99% sensitivity. Therefore, the RBT herd depicted in the random analysis stands as a feasible assessment and then, the recommended value in case of planning an eradication program in CR. Studies of three decades reveled that bovine brucellosis prevalence has increased in CR. B. abortus was identified by biochemical and molecular studies as the etiological agent of bovine brucellosis. Multiple locus variable-number tandem repeat analysis-16 revealed four B. abortus clusters. Cluster one and three are intertwined with isolates from other countries, while clusters two and four have only representatives from CR. Cluster one is widely distributed in all regions of the country and may be the primary B. abortus source. The other clusters seem to be restricted to specific areas in CR. The implications of our findings, in relation to the control of the disease in CR, are critically discussed.

Little is published on risk factors associated with bovine brucellosis in Pacific island communities. The 2009 re-emergence of bovine brucellosis in Fiji enabled us to do an interview-based questionnaire survey of 81 farms in the Wainivesi locality of the Tailevu province on the main island of Fiji to investigate what risk factors could have played a role in the re-emergence of the disease. The survey was conducted on 68 farms that had no positive cases of bovine brucellosis and on 13 farms in the same area where cattle had returned a positive result to the Brucella Rose Bengal test. Descriptive statistical methods were used to describe the demographic data while univariate analysis and multivariate logistic regression were used to evaluate the association between the selected risk factors and the presence of brucellosis on the farms at the time of the outbreak. The demographics of Fijian dairy farms are presented in the article and the biosecurity implications of those farming systems are discussed. Two risk factors were strongly associated with farms having brucellosis, and these were history of reactor cattle to brucellosis and or bovine tuberculosis on the farm (OR = 29, P ≤ 0.01) and farms that practised sharing of water sources for cattle within and with outside farms (OR = 39, P ≤ 0.01). Possible reasons why these were risk factors are also discussed. The potential risks for human health was also high as the use of personal protective equipment was low (15%). A high proportion of farmers (62%) could not recognise brucellosis thus contributing to the low frequency of disease reports (44%) made. The article also highlights other important risk factors which could be attributed to farming practices in the region and which could contribute to public health risks and the re-emergence of diseases.

Brucellosis remains as a major infectious disease of domestic animals and is considered a re-emerging zoonosis in several countries. B. abortus infections in bulls are related to reproductive tract infections, although infected animals show transient serological titers or nonreactor status. Thus, diagnosis of bovine brucellosis based exclusively on serological tests probably underestimates B. abortus infections in bulls. In this scenario, three hundred thirty-five serum samples from reproductively mature bovine bulls were subjected simultaneously to standard serodiagnosis using the rose Bengal test (RBT), 2-mercaptoethanol (2-ME), complement fixation (CFT), and fluorescence polarization assay (FPA). Furthermore, conventional semen plasma agglutination (SPA) and modified 2-ME, FC and, FPA were carried out in all bulls replaing serum by seminal plasma. Semen from all bulls was also analyzed for sperm viability, microbiological culture in Farrell media, and polymerase chain reaction (PCR). Only eight (2.38%) semen samples were considered improper for reproduction services (necrospermia and azoospermia), although none of these animals was positive in any of the diagnosis methods used. Five bulls (1.49%) were simultaneously positive in conventional RBT, 2-ME, SPA, modified 2-ME, microbiological culture in Farrell media, and in PCR for B. abortus strain 19. Two (1.67%) bulls were positive in PCR for B. abortus field strains and negative in all other tests, although semen was considered viable to reproduction service. The identification of B. abortus B19 strain in serum and semen of bulls occurred probably due to improper vaccination of males or infection by B19 strain shedding by vaccinated females that could to contaminated environment of farms. In addition, detection of B. abortus field strains only using PCR in bulls without sperm viability abnormalities indicate the need for including molecular methods to improve diagnosis of the disease in bovine bulls., (Copyright © 2017 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.)

Brucellosis, caused by Brucella spp., is an important zoonosis worldwide. Vaccination is an effective strategy for protection against Brucella infection in livestock in developing countries and in wildlife in developed countries. However, current vaccine strains including S19 and RB51 are pathogenic to humans and pregnant animals, limiting their use. In this study, we constructed the Brucella abortus (B. abortus) S2308 mutant strain Δ22915, in which the putative lytic transglycosylase gene BAB&#95;RS22915 was deleted. The biological properties of mutant strain Δ22915 were characterized and protection of mice against virulent S2308 challenge was evaluated. The mutant strain Δ22915 showed reduced survival within RAW264.7 cells and survival in vivo in mice. In addition, the mutant strain Δ22915 failed to escape fusion with lysosomes within host cells, and caused no observable pathological damage. RNA-seq analysis indicated that four genes associated with amino acid/nucleotide transport and metabolism were significantly upregulated in mutant strain Δ22915. Furthermore, inoculation of ∆22915 at 10 5 colony forming units induced effective host immune responses and long-term protection of BALB/c mice. Therefore, mutant strain ∆22915 could be used as a novel vaccine candidate in the future to protect animals against B. abortus infection.

Knowing the status of Bovine brucellosis and associated risk factors is a crucial step in formulating evidence based control scheme. In this study, a total of 967 dairy cows from 307 dairy farms in eastern Ethiopia were serologically tested for Brucella antibodies. The screening was done first using RBPT and positive samples were subsequently subjected to CFT for confirmation. A pre-tested structured questionnaire was used to collect relevant data from 307 dairy cattle owners or attendants to assess their awareness and routine practice. The data were run using univariable logistic regression analysis using STATA version 11.0 for Windows. Accordingly, herd and individual animal seroprevalence were found to be 6.8% (95% CI = 4.28-10.28) and 1.3% (95% CI = 0.72-2.29), respectively. The prevalence of sero-reactors among local breeds was observed to be higher compared to cross breed (p < 0.05). Herd level analysis of the risk factors indicated that in farms with large herd size (>20 animals), the odds ratio (OR) = 9.13, p = 0.00, CI = 3.01-27.69 of having brucellosis was 9.13 times higher than smaller size herds (<20 animals). Intensively managed herds had shown the highest seroprevalence (20.8%) than extensive (6.7%) and semi-intensive (4.2%). Experience of dairy handlers about the disease that cause abortion in late pregnancy was significantly associated (p < 0.001) with the occurrence of brucellosis in the herds. However, about 91% of the dairy cattle owners/attendants lack awareness about disease(s) that causes abortion in late pregnancy. Similarly risk of having brucellosis in those herds experiencing abortion was 6.3 times higher (OR = 6.3, p < 0.001, CI = 2.50-15.92). This study identified some of the handling practices for aborted and retained fetal materials to be risky. Therefore, the study highlights the need of comprehensive brucellosis surveillance in animal and human and institutions of public education and on farm biosafety measures in shaping proper disease control scheme.

Bovine brucellosis is endemic in Egypt in spite of application of surveillance and control measures. An increase of abortions was reported in a Holstein dairy cattle herd with 600 animals in Damietta governorate in Egypt after immunisation with Brucella (B.) abortus RB51 vaccine. Twenty one (10.6%) of 197 vaccinated cows aborted after 3 months. All aborted cows had been tested seronegative for brucellosis in the past 3 years. B. abortus was isolated from four foetuses. Conventional biochemical and bacteriological identification and polymerase chain reaction (PCR) confirmed two B. abortus biovar (bv.) 1 smooth and two B. abortus rough strains. None of the B. abortus isolates were identified as RB51. Genotyping analysis by multiple locus of variable number tandem repeats analysis based on 16 markers (MLVA-16) revealed two different profiles with low genetic diversity. B. abortus bv1 was introduced in the herd and caused abortions., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Bovine brucellosis is an important zoonosis caused by Brucella abortus that negatively impacts livestock productivity. In 2001, Brazil launched a new national program aimed at eradicating animal brucellosis that included large-scale studies of the prevalence and risk factors to support strategic decision-making. These studies were implemented by the animal health authorities and were underpinned by the scientific coordination of the University of São Paulo and the University of Brasília. The state-level results were published and revealed important differences in herd prevalence among regions. The risk factors varied across states and did not clearly explain the observed spatial disease spread. This study used a consolidated herd-level database of 14 states and 17,100 herds, from the prevalence surveys' data, to gain insights into herd profiles and cattle production practices that might be associated with the risk of brucellosis. At the time of data collection, the study area comprised just over 56 million bovine females aged over 24 months and approximately 1.8 million herds. After an exploratory univariable analysis, all factors with p≤0.20 were included in a multiple logistic regression model, using the design-based method in order to take herd sampling weights into account. The number of females in the herd markedly increased the risk of infection; compared with smaller herds (less than 30 females), the odds ratio was 3.42 [CI 95% 2.98-3.91] for herds with 31 to 100 females, 5.68 [4.92-6.55] for herds with 101 to 400 females, and 13.14 [10.94-15.78] for herds with more than 400 females. The risk was higher for extensive cattle production farms (OR=1.23 [1.07-1.42]) and for farms that purchased replacement stock from cattle traders (OR=1.27 [1.08-1.47]) or directly from other farms (OR=1.19 [1.07-1.32]). The exclusive use of artificial insemination (OR=0.57 [0.4-0.81]) and regular veterinary support (OR=0.68 [0.6-0.77]) appeared to be protective factors. These findings are consistent with the regional prevalence trends observed in the study and provide key guidance for the planning of the national effort to control and eradicate brucellosis. High vaccination coverage of heifers is recommended, especially when targeted to areas where large-scale extensive cattle production predominates. The smaller, more intensive herds, are good candidates for disease accreditation schemes., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Brucellosis is a well-known zoonotic disease that can cause severe economic and healthcare losses. Xinjiang, one of the biggest livestock husbandry sectors in China, has gone through increasing incidence of brucellosis in cattle and small ruminants recently. In this paper, 50 B. melitensis strains and 9 B. abortus strains collected from across Xinjiang area (from 2010 to 2015) were genotyped using multiple locus variable-number tandem-repeat (VNTR) analysis (MLVA) and multi-locus sequence typing (MLST). Based on 8 loci (MLVA-8), 50 B. melitensis strains were classified into three genotypes. Genotypes 42 (n=38, 76%) and 63 (n=11, 22%) were part of the East Mediterranean group, and one genotype with pattern of 1-5-3-13-2-4-3-2 represents a single-locus variant from genotype 63. MLVA-16 resolved 50 B. melitensis strains into 28 genotypes, of which 15 are unique to Xinjiang and 10 are in common with those in adjacent country Kazakhstan and neighboring provinces of China. Minimum Spanning Tree (MST) analysis implies that B. melitensis strains collected from across Kazakhstan, Xinjiang and China areas may share a common origin. Nine B. abortus strains were sorted into three genotypes by MLVA-8, genotypes 36 (n=7, 77.8%), 86 (n=1, 11.1%) and a new genotype with pattern of 4-5-3-13-2-2-3-1. Each B. abortus strain showed distinct MLVA-16 genotypes, suggesting that B. abortus species may possess more genetic diversity than B. melitensis. Using MLST, most B. melitensis strains (n=49) were identified as sequence type ST8, and most B. abortus strains (n=8) were recognized as ST2. Two new sequence types, ST37 and ST38, represented by single strain from B. melitensis and B. abortus species respectively, were also detected in this study. These results could facilitate the pathogen surveillance in the forthcoming eradication programs and serve as a guide in source tracking in case of new outbreaks occur., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Bovine brucellosis is an infectious disease of worldwide public health and economic importance. The usual tests for the diagnosis of this disease include the Rose-Bengal test (RBT), complement fixation test (CFT), serum agglutination test (SAT) and indirect ELISA. New tests such as competitive ELISAs (C-ELISA) and fluorescence polarisation assay (FPA) have been developed. However, C-ELISA may correspond to different protocols and a wide variation may exist in their diagnostic performance. The aim of this study was to evaluate three commercially available C-ELISA kits (C-ELISA1-3) and FPA for the diagnosis of bovine brucellosis and compare test performance with RBT, CFT, indirect ELISA and FPA. Sera submitted to EU laboratories in 2011 from 5111 adult cattle were tested. Individual test sensitivities (Se) and specificities (Sp) were estimated. Threshold assessment using the receiver operating characteristic method was also performed. The most sensitive tests were FPA (99.0%; 95% confidence interval [CI], 97.9-100%), C-ELISA1 (98.4%; 95% CI, 97.0-99.8%) and RBT (97.7%; 95% CI, 95.9-99.3%). The most specific tests were CFT (99.98%; 95% CI, 99.93-100%), SAT (99.98%; 95% CI, 99.93-100%) and RBT (99.89%; 95% CI, 99.79-99.99%). Among the new tests, none of the three C-ELISA kits studied could be recommended as a single screening test because of their low specificity, especially when used in a herd. C-ELISA3 could not be recommended as confirmatory test on individual animals to determine whether false positive serological test results had occurred., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Brucellosis is a globally important zoonotic infectious disease caused by gram negative bacteria of the genus Brucella. While many species of Brucella exist, Brucella melitensis, Brucella abortus, and Brucella suis are the most common pathogens of humans and livestock. The virulence of Brucella is largely influenced by its ability to evade host factors, including phagocytic killing mechanisms, which are critical for the host response to infection. The aim of this study was to characterize the bovine neutrophil response to virulent Brucella spp. Here, we found that virulent strains of smooth B. abortus, B. melitensis, B. suis, and virulent, rough, strains of Brucella canis possess similar abilities to resist killing by resting, or IFN-γ-activated, bovine neutrophils. Bovine neutrophils responded to infection with a time-dependent oxidative burst that varied little between Brucella spp. Inhibition of TAK1, or SYK kinase blunted the oxidative burst of neutrophils in response to Brucella infection. Interestingly, Brucella spp. did not induce robust death of bovine neutrophils. These results indicate that bovine neutrophils respond similarly to virulent Brucella spp. In addition, virulent Brucella spp., including naturally rough strains of B. canis, have a conserved ability to resist killing by bovine neutrophils., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Brucellosis is a highly contagious zoonotic infection affecting livestock and human beings. The disease has been reported worldwide except in few countries where it has been eradicated. The prevalence of brucellosis among cattle from 11 farms having a history of abortions was studied. A total of 481 samples comprising of blood, milk, vaginal swabs, vaginal discharges, placental tissues and fetal tissues were collected from 296 animals. Clinical samples were processed for the isolation of Brucella. Serum samples (n=296) were tested by Rose Bengal Plate Test (RBPT) and indirect ELISA. A total of 90 (30.40%) and 123 (41.55%) samples were positive by RBPT and indirect ELISA, respectively. Also 27.02% samples were positive by both the tests. Brucella isolates (n= 8) were recovered from clinical samples using Brucella selective media. All the isolates demonstrated PCR amplification for the bcsp31 and IS711 genes. Amplification of Brucella abortus specific primer was demonstrated by all the isolates in AMOS PCR indicating isolates to be of either B. abortus biotype 1, 2 or 4. Risk factors for transmission of brucellosis among cattle population were studied by field surveys. It was observed that lack of awareness about brucellosis (OR=8.739, P=0.138) and inadequate floor space (OR=0.278, P=0.128) were crucial risk factors for transmission of bovine brucellosis., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Targeting threonyl-tRNA synthetase (ThrRS) of Brucella abortus is a promising approach to developing small-molecule drugs against bovine brucellosis. Using the BLASTp algorithm, we identified ThrRS from Escherichia coli (EThrRS, PDB ID 1QF6), which is 51% identical to ThrRS from Brucella abortus (BaThrRS) at the amino acid sequence level. EThrRS was used as the template to construct a BaThrRS homology model which was optimized using molecular dynamics simulations. To determine the residues important for substrate ATP binding, we identified the ATP-binding regions of BaThrRS, docked ATP to the protein, and identified the residues whose side chains surrounded bound ATP. We then used the binding site of ATP to virtually screen for BaThrRS inhibitors and got seven leads. We further characterized the BaThrRS-binding site of the compound with the highest predicted inhibitory activity. Our results should facilitate future experimental effects to find novel drugs for use against bovine brucellosis.

Between 2005 and 2006, a cross-sectional survey was carried out in domestic ruminants in agropastoral communities of Serengeti district, Tanzania to determine the seroprevalence of brucellosis in domestic-wildlife interface villages. Both the Rose Bengal Plate Test (RBPT) and Competitive Enzyme Linked-immunosorbent Assay (c-ELISA) were used to analyse 82 human and 413 livestock sera from four randomly selected villages located along game reserve areas of Serengeti National Park. Although both cattle (288) and small ruminants (125) were screened, seropositivity was detected only in cattle. The overall seroprevalence based on c-ELISA as a confirmatory test was 5.6%. In cattle both age and sex were not statistically associated with brucellosis seropositivity (P = 0.63; 95% CI = 0.03, 0.8 and 0.33; 95% CI = 0.6, 3.7, respectively). Overall herd level seropositivity was 46.7% (n = 7), ranging from 25% to 66.7% (n = 4-10). Each village had at least one brucellosis seropositive herd. None of the 82 humans tested with both RBPT and c-ELISA were seropositive. Detecting Brucella infection in cattle in such areas warrants further investigation to establish the circulating strains for eventual appropriate control interventions in domestic animals.

Bovine brucellosis is endemic in Nigeria; however, limited data exist on nationwide studies and risk factors associated with the disease. Using a cross-sectional sero-epidemiological survey, we determined the prevalence of and risk factors for brucellosis in slaughtered cattle in three geographical regions of Nigeria. Serum samples from randomly selected unvaccinated cattle slaughtered over a period of 3 years (between December 2010 and September 2013) from northern, southern and south-western Nigeria were tested for antibodies to Brucella abortus using the Rose Bengal test. Data associated with risk factors of brucellosis were analysed by Stata Version 12. In all, 8105 cattle were screened. An overall seroprevalence of 3.9% (315/8105) was recorded by the Rose Bengal test, with 3.8%, 3.4% and 4.0% from the northern, southern and south-western regions, respectively. Bivariate analysis showed that cattle screened in northern Nigeria were less likely to be seropositive for antibodies to Brucella spp. than those from south-western Nigeria (odds ratio = 0.94; 95% confidence interval: 0.73-1.22). However, logistic regression analysis revealed that breed ( p = 0.04) and sex ( p £ 0.0001) of cattle were statistically significant for seropositivity to Brucella spp. The study found that brucellosis was endemic at a low prevalence among slaughtered cattle in Nigeria, with sex and breed of cattle being significant risk factors. Considering the public health implications of brucellosis, we advocate coordinated surveillance for the disease among diverse cattle populations in Nigeria, as is carried out in most developed countries.

Brucellosis is a debilitating zoonotic disease that affects humans and animals. The diagnosis of brucellosis is challenging, as accurate species level identification is not possible with any of the currently available serology-based diagnostic methods. The present study aimed at identifying Brucella (B.) species-specific proteins from the closely related species B. abortus and B. melitensis using sera collected from naturally infected host species. Unlike earlier reported investigations with either laboratory-grown species or vaccine strains, in the present study, field strains were utilized for analysis. The label-free quantitative proteomic analysis of the naturally isolated strains of these two closely related species revealed 402 differentially expressed proteins, among which 63 and 103 proteins were found exclusively in the whole cell extracts of B. abortus and B. melitensis field strains, respectively. The sera from four different naturally infected host species, i.e., cattle, buffalo, sheep, and goat were applied to identify the immune-binding protein spots present in the whole protein extracts from the isolated B. abortus and B. melitensis field strains and resolved on two-dimensional gel electrophoresis. Comprehensive analysis revealed that 25 proteins of B. abortus and 20 proteins of B. melitensis were distinctly immunoreactive. Dihydrodipicolinate synthase, glyceraldehyde-3-phosphate dehydrogenase and lactate/malate dehydrogenase from B. abortus, amino acid ABC transporter substrate-binding protein from B. melitensis and fumarylacetoacetate hydrolase from both species were reactive with the sera of all the tested naturally infected host species. The identified proteins could be used for the design of serological assays capable of detecting pan-Brucella, B. abortus- and B. melitensis-specific antibodies.

Brucella abortus is the etiological agent of bovine brucellosis, a zoonotic disease that causes significant economic losses worldwide. The differential proteomic profile of bovine chorioallantoic membrane (CAM) explants at early stages of infection with B. abortus (0.5, 2, 4, and 8 h) was determined. Analysis of CAM explants at 0.5 and 4 h showed the highest differences between uninfected and infected CAM explants, and therefore were used for the Differential Gel Electrophoresis (DIGE). A total of 103 spots were present in only one experimental group and were selected for identification by mass spectrometry (MALDI/ToF-ToF). Proteins only identified in extracts of CAM explants infected with B. abortus were related to recognition of PAMPs by TLR, production of reactive oxygen species, intracellular trafficking, and inflammation.

Brucellosis is one of the most widespread zoonoses in the world caused by several species of the genus Brucella. The disease, eradicated in many developed countries, is a re-emerging neglected zoonosis endemic in several zones especially in the Mediterranean region, impacting on human health and livestock production. A One Health approach could address brucellosis control in Morocco but scarcity of reliable epidemiological data, as well as underreporting, hinders the implementation of sustainable control strategies. Surveillance and control policies implemented by the Moroccan government in domestic animals (cattle and small ruminants) in the last few decades are assessed for disease impact. This study considers the origins of animal brucellosis in Morocco and the potential for emergence of brucellosis during a shift from extensive to intensive livestock production.