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Sample records for brucella

  1. Brucella

    Science.gov (United States)

    The genus Brucella encompasses a group of gram negative bacteria that survive almost exclusively in infected hosts with preference for localization in intracellular compartments of cells. The genus has traditionally been divided into species based on microbe characteristics and host preference, bu...

  2. Draft Genome Sequences of Brucella suis Biovar 4 Strain NCTC 10385, Brucella ceti Strain NCTC 12891T, Brucella inopinata Strain CAMP 6436T, and Brucella neotomae Strain ATCC 23459T

    National Research Council Canada - National Science Library

    Wahab, Tara; Ferrari, Sevinc; Lindberg, Martina; Bäckman, Stina; Kaden, Rene

    2014-01-01

    With the aim of developing quantitative PCR methods for the detection and differentiation of Brucella species, the genomes of Brucella ceti, Brucella inopinata, Brucella netotomae, and Brucella suis...

  3. Limited Genetic Diversity of Brucella spp.

    OpenAIRE

    Gándara, Benjamín; Merino, Ahidé López; Rogel, Marco Antonio; Martínez-Romero, Esperanza

    2001-01-01

    Multilocus enzyme electrophoresis (MLEE) of 99 Brucella isolates, including the type strains from all recognized species, revealed a very limited genetic diversity and supports the proposal of a monospecific genus. In MLEE-derived dendrograms, Brucella abortus and a marine Brucella sp. grouped into a single electrophoretic type related to Brucella neotomae and Brucella ovis. Brucella suis and Brucella canis formed another cluster linked to Brucella melitensis and related to Rhizobium tropici....

  4. Microbiology of Brucella.

    Science.gov (United States)

    Percin, Duygu

    2013-04-01

    The genus Brucella is a member of family Brucellaceae and includes ten species which are small, non-motile, non-sporing, aerobic, gram-negative intracellular coccobacilli. They are catalase, oxidase and urea positive bacteria. Members of the genus can grow on enriched media like blood agar or chocolate agar. Identification in species level can be done by agglutination with monospecific serum, cultivating the strains in the presence of dyes and/or with PCR methods. Antigenic structure of the Brucella is composed of surface, intracellular, and in vivo antigens. Thanks to various virulence factors that act as metabolic regulators, Brucella strains can protect themselves from immune system of the host, adapt easily to different environmental conditions, and multiply intracellular. Classification, epidemiological features, isolation and identification, antigenic structure and virulence factors of Brucella species along with the discussion of very few patents associated with Brucellosis have been reviewed in this paper.

  5. Brucella, nitrogen and virulence.

    Science.gov (United States)

    Ronneau, Severin; Moussa, Simon; Barbier, Thibault; Conde-Álvarez, Raquel; Zuniga-Ripa, Amaia; Moriyon, Ignacio; Letesson, Jean-Jacques

    2016-08-01

    The brucellae are α-Proteobacteria causing brucellosis, an important zoonosis. Although multiplying in endoplasmic reticulum-derived vacuoles, they cause no cell death, suggesting subtle but efficient use of host resources. Brucellae are amino-acid prototrophs able to grow with ammonium or use glutamate as the sole carbon-nitrogen source in vitro. They contain more than twice amino acid/peptide/polyamine uptake genes than the amino-acid auxotroph Legionella pneumophila, which multiplies in a similar vacuole, suggesting a different nutritional strategy. During these two last decades, many mutants of key actors in nitrogen metabolism (transporters, enzymes, regulators, etc.) have been described to be essential for full virulence of brucellae. Here, we review the genomic and experimental data on Brucella nitrogen metabolism and its connection with virulence. An analysis of various aspects of this metabolism (transport, assimilation, biosynthesis, catabolism, respiration and regulation) has highlighted differences and similarities in nitrogen metabolism with other α-Proteobacteria. Together, these data suggest that, during their intracellular life cycle, the brucellae use various nitrogen sources for biosynthesis, catabolism and respiration following a strategy that requires prototrophy and a tight regulation of nitrogen use.

  6. Draft Genome Sequences of Brucella suis Biovar 4 Strain NCTC 10385, Brucella ceti Strain NCTC 12891T, Brucella inopinata Strain CAMP 6436T, and Brucella neotomae Strain ATCC 23459T

    OpenAIRE

    Wahab, Tara; Ferrari, Sevinc; Lindberg, Martina; Bäckman, Stina; Kaden, Rene

    2014-01-01

    With the aim of developing quantitative PCR methods for the detection and differentiation of Brucella species, the genomes of Brucella ceti, Brucella inopinata, Brucella netotomae, and Brucella suis biovar 4 were sequenced and analyzed.

  7. Immunopathology of Brucella infection.

    Science.gov (United States)

    Baldi, Pablo C; Giambartolomei, Guillermo H

    2013-04-01

    In spite of the protean nature of the disease, inflammation is a hallmark of brucellosis and affected tissues usually exhibit inflammatory infiltrates. As Brucella lacks exotoxins, exoproteases or cytolysins, pathological findings in brucellosis probably arise from inflammation-driven processes. The cellular and molecular bases of immunopathological phenomena probably involved in Brucella pathogenesis have been unraveled in the last few years. Brucella-infected osteoblasts, either alone or in synergy with infected macrophages, produce cytokines, chemokines and matrixmetalloproteinases (MMPs), and similar phenomena are mounted by fibroblast-like synoviocytes. The released cytokines promote the secretion of MMPs and induce osteoclastogenesis. Altogether, these phenomena may contribute to the bone loss and cartilage degradation usually observed in brucellar arthritis and osteomyelitis. Proinflammatory cytokines may be also involved in the pathogenesis of neurobrucellosis. B. abortus and its lipoproteins elicit an inflammatory response in the CNS of mice, leading to astrogliosis, a characteristic feature of neurobrucellosis. Heat-killed bacteria (HKBA) and the L-Omp19 lipoprotein elicit astrocyte apoptosis and proliferation (two features of astrogliosis), and apoptosis depends on TNF-α signaling. Brucella also infects and replicates in human endothelial cells, inducing the production of chemokines and IL-6, and an increased expression of adhesion molecules. The sustained inflammatory process derived from the longlasting infection of the endothelium may be important for the development of endocarditis. Therefore, while Brucella induces a low grade inflammation as compared to other pathogens, its prolonged intracellular persistence in infected tissues supports a long-lasting inflammatory response that mediates different pathways of tissue damage. In this context, approaches to avoid the invasion of host cells or limit the intracellular survival of the bacterium may be

  8. [MOLECULAR ASPECTS OF BRUCELLA PERSISTENCE].

    Science.gov (United States)

    Kulakov Yu K

    2016-01-01

    Brucellosis is a dangerous zoonotic disease of animals and humans caused by bacteria of the genus Brucella, which are able to survive, multiply, and persist in host cells. The review is devoted to the Brucella species persistence connected to the molecular mechanisms of escape from innate and adaptive immunity of the host and active interaction of effector proteins of the type IV secretion system with the host's signaling pathways. Understanding of the molecular mechanisms used by Brucella for the intracellular persistence in the host organism can allow us to develop new and effective means for the prevention and treatment of chronic brucellosis infection.

  9. Brucella peritonitis and leucocytoclastic vasculitis due to Brucella melitensis

    Directory of Open Access Journals (Sweden)

    Murat Dizbay

    Full Text Available Brucellosis is a multisystemic disease that rarely leads to a fatal outcome. While reticuloendothelial system organs are mostly affected, peritonitis and posthepatitic cirrhosis are also complications of brucellosis, though they are very rare. Brucella spp. can also trigger immunological reactions. We report a case of brucellosis with peritonitis, renal failure and leucocytoclastic vasculitis caused by Brucella melitensis, which led to a fatal outcome. Brucellosis should be considered in the differential diagnosis of vasculitic diseases, especially in endemic areas.

  10. Activation of bovine neutrophils by Brucella spp.

    Science.gov (United States)

    Keleher, Lauren L; Skyberg, Jerod A

    2016-09-01

    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.

  11. ATP-Binding Cassette Systems of Brucella

    Directory of Open Access Journals (Sweden)

    Dominic C. Jenner

    2009-01-01

    Full Text Available Brucellosis is a prevalent zoonotic disease and is endemic in the Middle East, South America, and other areas of the world. In this study, complete inventories of putative functional ABC systems of five Brucella species have been compiled and compared. ABC systems of Brucella melitensis 16M, Brucella abortus 9-941, Brucella canis RM6/66, Brucella suis 1330, and Brucella ovis 63/290 were identified and aligned. High numbers of ABC systems, particularly nutrient importers, were found in all Brucella species. However, differences in the total numbers of ABC systems were identified (B. melitensis, 79; B. suis, 72; B. abortus 64; B. canis, 74; B. ovis, 59 as well as specific differences in the functional ABC systems of the Brucella species. Since B. ovis is not known to cause human brucellosis, functional ABC systems absent in the B. ovis genome may represent virulence factors in human brucellosis.

  12. Laboratory exposure to Brucella melitensis in Denmark

    DEFF Research Database (Denmark)

    Knudsen, A; Kronborg, G; Knudsen, Inge Jenny Dahl

    2013-01-01

    Brucella species are a frequent cause of laboratory-acquired infections. This report describes the handling of a laboratory exposure of 17 laboratory staff members exposed to Brucella melitensis in a large microbiology laboratory in a brucella-non-endemic area. We followed the US Centers...

  13. Epidemiology of brucellosis in domestic animals caused by Brucella melitensis, Brucella suis and Brucella abortus.

    Science.gov (United States)

    Díaz Aparicio, E

    2013-04-01

    Brucellosis is a disease that causes severe economic losses for livestock farms worldwide. Brucella melitensis, B. abortus and B. suis, which are transmitted between animals both vertically and horizontally, cause abortion and infertility in their primary natural hosts - goats and sheep (B. melitensis), cows (B. abortus) and sows (B. suis). Brucella spp. infect not only their preferred hosts but also other domestic and wild animal species, which in turn can act as reservoirs of the disease for other animal species and humans. Brucellosis is therefore considered to be a major zoonosis transmitted by direct contact with animals and/or their secretions, or by consuming milk and dairy products.

  14. Brucella Epididymoorchitis: A Report Of Two Cases

    OpenAIRE

    AYBEK, Z.; TUNCAY, L.; BOZBAY, C.; KALELİ, İ.

    2010-01-01

    Two brothers with Brucella epididimoorchitis is disscussed in terms of diagnosis and treatment of the disease, and, results obtained are compared with the literature. İki Olgu Nedeniyle Brucella Epididmoorşiti Brucella epididimoorşit tanısı konulan iki kardeş hasta sunularak ilgili literatür sonuçlarıyla tanı ve tedavi açısından tartışıldı

  15. Advancement of knowledge of Brucella over the past 50 years.

    Science.gov (United States)

    Olsen, S C; Palmer, M V

    2014-11-01

    Fifty years ago, bacteria in the genus Brucella were known to cause infertility and reproductive losses. At that time, the genus was considered to contain only 3 species: Brucella abortus, Brucella melitensis, and Brucella suis. Since the early 1960s, at least 7 new species have been identified as belonging to the Brucella genus (Brucella canis, Brucella ceti, Brucella inopinata, Brucella microti, Brucella neotomae, Brucella ovis, and Brucella pinnipedialis) with several additional new species under consideration for inclusion. Although molecular studies have found such high homology that some authors have proposed that all Brucella are actually 1 species, the epidemiologic and diagnostic benefits for separating the genus based on phenotypic characteristics are more compelling. Although pathogenic Brucella spp have preferred reservoir hosts, their ability to infect numerous mammalian hosts has been increasingly documented. The maintenance of infection in new reservoir hosts, such as wildlife, has become an issue for both public health and animal health regulatory personnel. Since the 1960s, new information on how Brucella enters host cells and modifies their intracellular environment has been gained. Although the pathogenesis and histologic lesions of B. abortus, B. melitensis, and B. suis in their preferred hosts have not changed, additional knowledge on the pathology of these brucellae in new hosts, or of new species of Brucella in their preferred hosts, has been obtained. To this day, brucellosis remains a significant human zoonosis that is emerging or reemerging in many parts of the world. © The Author(s) 2014.

  16. Renal abscess caused by Brucella

    Directory of Open Access Journals (Sweden)

    Jun Li

    2014-11-01

    Full Text Available Involvement of the renal parenchyma in the acute phase of brucellosis is very rare. Only two cases of renal brucelloma have been reported in the English language literature to date. We report a case of renal abscess caused by Brucella in the acute phase. A 45-year-old Chinese man presented with a high fever, urine occult blood, and a low density lesion in the right kidney. Ultrasound-guided aspiration was done. Brucella melitensis was isolated from both blood and puncture fluid culture. Minocycline combined with moxifloxacin was prescribed for 4 months. The infection relapsed at 6 months after discontinuation. Minocycline combined with rifampin was administered for another 2 months. The brucellosis had not relapsed at more than 20 months later. It is possible to cure renal brucelloma with antibiotics and ultrasound-guided aspiration. Treatment should not be discontinued until the abscess has disappeared and two consecutive blood cultures taken 1 month apart are negative.

  17. Molecular detection of Brucella spp. from broth culture of clinical ...

    African Journals Online (AJOL)

    PRECIOUS

    2009-11-02

    Nov 2, 2009 ... PCR was employed to detect Brucella spp. from broth cultures of clinical samples using a group ... studies, thus classifying Brucella as a monospecific ..... Bricker BJ, Halling SM (1995). Enhancement of the Brucella AMOS. PCR Assay for Differentiation of Brucella abortus vaccine strains S19 and RB51.

  18. 9 CFR 113.65 - Brucella Abortus Vaccine.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Brucella Abortus Vaccine. 113.65... Bacterial Vaccines § 113.65 Brucella Abortus Vaccine. Brucella Abortus Vaccine shall be prepared as a desiccated live culture bacterial vaccine from smooth colonial forms of the Brucella abortus organism...

  19. Characterization of ribonuclease III from Brucella.

    Science.gov (United States)

    Wu, Chang-Xian; Xu, Xian-Jin; Zheng, Ke; Liu, Fang; Yang, Xu-Dong; Chen, Chuang-Fu; Chen, Huan-Chun; Liu, Zheng-Fei

    2016-04-01

    Bacterial ribonuclease III (RNase III) is a highly conserved endonuclease, which plays pivotal roles in RNA maturation and decay pathways by cleaving double-stranded structure of RNAs. Here we cloned rncS gene from the genomic DNA of Brucella melitensis, and analyzed the cleavage properties of RNase III from Brucella. We identified Brucella-encoding small RNA (sRNA) by high-throughput sequencing and northern blot, and found that sRNA of Brucella and Homo miRNA precursor (pre-miRNA) can be bound and cleaved by B.melitensis ribonuclease III (Bm-RNase III). Cleavage activity of Bm-RNase III is bivalent metal cations- and alkaline buffer-dependent. We constructed several point mutations in Bm-RNase III, whose cleavage activity indicated that the 133th Glutamic acid residue was required for catalytic activity. Western blot revealed that Bm-RNase III was differently expressed in Brucella virulence strain 027 and vaccine strain M5-90. Collectively, our data suggest that Brucella RNase III can efficiently bind and cleave stem-loop structure of small RNA, and might participate in regulation of virulence in Brucella. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Detection of Brucella melitensis by the BacT/Alert automated system and Brucella broth culture.

    Science.gov (United States)

    Ozkurt, Z; Erol, S; Tasyaran, M A; Kaya, A

    2002-11-01

    This study was conducted to evaluate the ability of the BacT/Alert automated blood culture system to detect Brucella spp. in comparison with traditional Brucella broth culture. Overall, 100 (50 bone marrow and 50 blood samples) paired cultures were obtained, and 59 were positive by at least one method. The Brucella broth culture method detected all 59 positive cultures (100%), and the BacT/Alert system detected 30 (50.8%) (P broth culture (P > 0.05). There is no significant difference between the two methods with respect to growth time of the microorganism, but Brucella broth culture is more sensitive than the BacT/Alert system.

  1. Brucella spp. Virulence Factors and Immunity.

    Science.gov (United States)

    Byndloss, Mariana X; Tsolis, Renee M

    2016-01-01

    Brucellosis, caused by bacteria of the genus Brucella, is an important zoonotic infection that causes reproductive disease in domestic animals and chronic debilitating disease in humans. An intriguing aspect of Brucella infection is the ability of these bacteria to evade the host immune response, leading to pathogen persistence. Conversely, in the reproductive tract of infected animals, this stealthy pathogen is able to cause an acute severe inflammatory response. In this review, we discuss the different mechanisms used by Brucella to cause disease, with emphasis on its virulence factors and the dichotomy between chronic persistence and reproductive disease.

  2. Progress in Brucella vaccine development

    Science.gov (United States)

    YANG, Xinghong; SKYBERG, Jerod A.; CAO, Ling; CLAPP, Beata; THORNBURG, Theresa; PASCUAL, David W.

    2012-01-01

    Brucella spp. are zoonotic, facultative intracellular pathogens, which cause animal and human disease. Animal disease results in abortion of fetuses; in humans, it manifests flu-like symptoms with an undulant fever, with osteoarthritis as a common complication of infection. Antibiotic regimens for human brucellosis patients may last several months and are not always completely effective. While there are no vaccines for humans, several licensed live Brucella vaccines are available for use in livestock. The performance of these animal vaccines is dependent upon the host species, dose, and route of immunization. Newly engineered live vaccines, lacking well-defined virulence factors, retain low residual virulence, are highly protective, and may someday replace currently used animal vaccines. These also have possible human applications. Moreover, due to their enhanced safety and efficacy in animal models, subunit vaccines for brucellosis show great promise for their application in livestock and humans. This review summarizes the progress of brucellosis vaccine development and presents an overview of candidate vaccines. PMID:23730309

  3. Use of serology and bacterial culture to determine prevalence of Brucella spp. In feral Swine (sus scrofa) in proximity to a beef cattle herd positive for Brucella suis and Brucella abortus

    National Research Council Canada - National Science Library

    Musser, Jeffrey M B; Schwartz, Andy L; Srinath, Indumathi; Waldrup, Kenneth A

    2013-01-01

    .... and the antibody to Brucella spp. in a feral swine (Sus scrofa) population in proximity to a cattle herd that was culture positive for Brucella abortus and Brucella suis in north-central Texas, USA...

  4. Brucella melitensis Invades Murine Erythrocytes during Infection

    Science.gov (United States)

    Vitry, Marie-Alice; Hanot Mambres, Delphine; Deghelt, Michaël; Hack, Katrin; Machelart, Arnaud; Lhomme, Frédéric; Vanderwinden, Jean-Marie; Vermeersch, Marjorie; De Trez, Carl; Pérez-Morga, David; Letesson, Jean-Jacques

    2014-01-01

    Brucella spp. are facultative intracellular Gram-negative coccobacilli responsible for brucellosis, a worldwide zoonosis. We observed that Brucella melitensis is able to persist for several weeks in the blood of intraperitoneally infected mice and that transferred blood at any time point tested is able to induce infection in naive recipient mice. Bacterial persistence in the blood is dramatically impaired by specific antibodies induced following Brucella vaccination. In contrast to Bartonella, the type IV secretion system and flagellar expression are not critically required for the persistence of Brucella in blood. ImageStream analysis of blood cells showed that following a brief extracellular phase, Brucella is associated mainly with the erythrocytes. Examination by confocal microscopy and transmission electron microscopy formally demonstrated that B. melitensis is able to invade erythrocytes in vivo. The bacteria do not seem to multiply in erythrocytes and are found free in the cytoplasm. Our results open up new areas for investigation and should serve in the development of novel strategies for the treatment or prophylaxis of brucellosis. Invasion of erythrocytes could potentially protect the bacterial cells from the host's immune response and hamper antibiotic treatment and suggests possible Brucella transmission by bloodsucking insects in nature. PMID:25001604

  5. Infection of cattle in Kenya with Brucella abortus biovar 3 and Brucella melitensis biovar 1 genotypes

    NARCIS (Netherlands)

    Muendo, Esther N.; Mbatha, Peter M.; Macharia, Joseph; Abdoel, Theresia H.; Janszen, Paul V.; Pastoor, Rob; Smits, Henk L.

    2012-01-01

    Brucella melitensis biovar 1 was isolated from bovine milk samples from a herd in central Kenya, and Brucella abortus biovar 3 was isolated from aborted fetus materials and vaginal discharge fluids from cattle in central and eastern provinces of Kenya. All infections including those with B.

  6. Genetic Polymorphism Characteristics of Brucella canis Isolated in China

    OpenAIRE

    Di, Dongdong; Cui, Buyun; Wang, Heng; Zhao, Hongyan; Piao, Dongri; Tian, Lili; Tian, Guozhong; Kang, Jingli; Mao, Xiang; Zhang, Xiaojun; Du, Pengfei; Zhu, Lin; Zhao, Zhuo; Mao, Lingling; Yao, Wenqing

    2014-01-01

    In China, brucellosis is an endemic disease typically caused by Brucella melitensis infection (biovars 1 and 3). Brucella canis infection in dogs has not traditionally recognized as a major problem. In recent years however, brucellosis resulting from Brucella canis infection has also been reported, suggesting that infections from this species may be increasing. Data concerning the epidemiology of brucellosis resulting from Brucella canis infection is limited. Therefore, the purpose of this st...

  7. Isolation of Brucella abortus Using PCR-RFLP Analysis

    OpenAIRE

    M Salehi; E Pishva; R Salehi; R Rahmani

    2006-01-01

    Brucella transmission and epidemiology depend on infecting species and biovar. Therefore, exact identification of the Brucella is important to design correct control and treatment strategies. In this study, we examined presence of other Brucellae in Isfahan. One hundred twenty Brucella isolates were collected and genomic DNA was extracted from them. omp2a fragment of all isolates were amplified using a pair of specific primers and the PCR products were electrophoresed and stained with EtBr. T...

  8. 21 CFR 866.3085 - Brucella spp. serological reagents.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Brucella spp. serological reagents. 866.3085... (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3085 Brucella spp. serological reagents. (a) Identification. Brucella spp. serological reagents are devices that consist of...

  9. 9 CFR 113.32 - Detection of Brucella contamination.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Detection of Brucella contamination... REQUIREMENTS Standard Procedures § 113.32 Detection of Brucella contamination. The test for detection of Brucella contamination provided in this section shall be conducted when such a test is prescribed in an...

  10. Brucella Infection in HIV Infected Patients

    Directory of Open Access Journals (Sweden)

    SeyedAhmad SeyedAlinaghi

    2011-12-01

    Full Text Available The purpose of this study was to assess the possible correlation between Brucella and HIV infections. Iran is a country where HIV infection is expanding and Brucellosis is prevalent. In the present study, 184 HIV infected patients were assigned and for all of them HIV infection was confirmed by western blot test. In order to identify the prevalence rate of Brucella infection and systemic brucellosis in these subjects, sera samples were obtained and Brucella specific serological tests were performed to reveal antibody titers. Detailed history was taken and physical examination was carried out for all of patients. 11 (6% subjects had high titers but only 3 of them were symptomatic. Most of these subjects were injection drug user (IDU men and one was a rural woman. Considering both prevalence rates of Brucella infection (3% and symptomatic brucellosis (0.1% in Iran, our HIV positive patients show higher rates of Brucella infection and systemic brucellosis. Preserved cellular immunity of participants and retention of granulocytes activity may explain this poor association; whereas other explanations such as immunological state difference and non-overlapping geographical distribution of the 2 pathogens have been mentioned by various authors.

  11. Brucella epididymo-orchitis: a consideration in endemic area

    Directory of Open Access Journals (Sweden)

    Jaffar A. Al-Tawfiq

    2006-06-01

    Full Text Available Brucellosis is a zoonotic disease caused by Brucella sp. and may affect many parts of the body. Brucella epididymo-orchitis had been reported in up to 20% of patients with brucellosis. This is a case report of Brucella epididymo-orchitis in a Saudi male patient. He presented with a unilateral swelling of the left testicle. He had fever, arthralgia and night sweats. Ultrasound examination revealed enlarged left epididymis and testicle. Brucella serology was positive and the patient responded to treatment with doxycycline and gentamicin. Thus, brucella infection should be considered in the differential diagnosis of patients presenting with epididymo-orchitis from an endemic area.

  12. Genetic polymorphism characteristics of Brucella canis isolated in China.

    Science.gov (United States)

    Di, Dongdong; Cui, Buyun; Wang, Heng; Zhao, Hongyan; Piao, Dongri; Tian, Lili; Tian, Guozhong; Kang, Jingli; Mao, Xiang; Zhang, Xiaojun; Du, Pengfei; Zhu, Lin; Zhao, Zhuo; Mao, Lingling; Yao, Wenqing; Guan, Pingyuan; Fan, Weixing; Jiang, Hai

    2014-01-01

    In China, brucellosis is an endemic disease typically caused by Brucella melitensis infection (biovars 1 and 3). Brucella canis infection in dogs has not traditionally recognized as a major problem. In recent years however, brucellosis resulting from Brucella canis infection has also been reported, suggesting that infections from this species may be increasing. Data concerning the epidemiology of brucellosis resulting from Brucella canis infection is limited. Therefore, the purpose of this study was to assess the diversity among Chinese Brucella canis strains for epidemiological purposes. First, we employed a 16-marker VNTR assay (Brucella MLVA-16) to assess the diversity and epidemiological relationship of 29 Brucella canis isolates from diverse locations throughout China with 38 isolates from other countries. MLVA-16 analysis separated the 67 Brucella canis isolates into 57 genotypes that grouped into five clusters with genetic similarity coefficients ranging from 67.73 to 100%. Moreover, this analysis revealed a new genotype (2-3-9-11-3-1-5-1:118), which was present in two isolates recovered from Guangxi in 1986 and 1987. Second, multiplex PCR and sequencing analysis were used to determine whether the 29 Chinese Brucella canis isolates had the characteristic BMEI1435 gene deletion. Only two isolates had this deletion. Third, amplification of the omp25 gene revealed that 26 isolates from China had a T545C mutation. Collectively, this study reveals that considerable diversity exists among Brucella canis isolates in China and provides resources for studying the genetic variation and microevolution of Brucella.

  13. Genetic polymorphism characteristics of Brucella canis isolated in China.

    Directory of Open Access Journals (Sweden)

    Dongdong Di

    Full Text Available In China, brucellosis is an endemic disease typically caused by Brucella melitensis infection (biovars 1 and 3. Brucella canis infection in dogs has not traditionally recognized as a major problem. In recent years however, brucellosis resulting from Brucella canis infection has also been reported, suggesting that infections from this species may be increasing. Data concerning the epidemiology of brucellosis resulting from Brucella canis infection is limited. Therefore, the purpose of this study was to assess the diversity among Chinese Brucella canis strains for epidemiological purposes. First, we employed a 16-marker VNTR assay (Brucella MLVA-16 to assess the diversity and epidemiological relationship of 29 Brucella canis isolates from diverse locations throughout China with 38 isolates from other countries. MLVA-16 analysis separated the 67 Brucella canis isolates into 57 genotypes that grouped into five clusters with genetic similarity coefficients ranging from 67.73 to 100%. Moreover, this analysis revealed a new genotype (2-3-9-11-3-1-5-1:118, which was present in two isolates recovered from Guangxi in 1986 and 1987. Second, multiplex PCR and sequencing analysis were used to determine whether the 29 Chinese Brucella canis isolates had the characteristic BMEI1435 gene deletion. Only two isolates had this deletion. Third, amplification of the omp25 gene revealed that 26 isolates from China had a T545C mutation. Collectively, this study reveals that considerable diversity exists among Brucella canis isolates in China and provides resources for studying the genetic variation and microevolution of Brucella.

  14. Prevalence of Brucella spp in humans

    Directory of Open Access Journals (Sweden)

    Catharina de Paula Oliveira Cavalcanti Soares

    2015-10-01

    Full Text Available Objective: to determine the seroprevalence of Brucella spp in humans.Method: this is an observational study, developed with 455 individuals between 18 and 64 years old, who use the Estratégia de Saúde da Família (Brazil's family health strategy. The serum samples of volunteers underwent buffered acid antigen tests, such as screening, agar gel immunodiffusion and slow seroagglutination test in tubes and 2-Mercaptoethanol.Results: among the samples, 1.98% has responded to buffered-acid antigen, 2.85% to agar gel immunodiffusion test and 1.54% to the slow seroagglutination tests on tubes/2-Mercaptoethanol. The prevalence of Brucella spp was 4.4%, represented by the last two tests.Conclusion: the results of this research suggest that the studied population is exposed to Brucella spp infection.

  15. Metal acquisition and virulence in Brucella

    Science.gov (United States)

    Roop, R. Martin

    2013-01-01

    Similar to other bacteria, Brucella strains require several biologically essential metals for their survival in vitro and in vivo. Acquiring sufficient levels of some of these metals, particularly iron, manganese and zinc, is especially challenging in the mammalian host, where sequestration of these micronutrients is a well-documented component of both the innate and acquired immune responses. This review describes the Brucella metal transporters that have been shown to play critical roles in the virulence of these bacteria in experimental and natural hosts. PMID:22632611

  16. A modified test for Brucella agglutinins

    Science.gov (United States)

    Singh, J. N.

    1969-01-01

    0·1% Protamine sulphate in normal (0·85% w/v) saline has been used as a diluent in Brucella serology instead of saline as such. The use of protamine sulphate in this concentration has obviated the need for performing the modified Coombs test to detect non-agglutinating Brucella antibodies. This method has been used in the Microbiological Diagnostic Unit of the University of Melbourne in the routine titration of 721 sera and has proved to give reliable and reproducible results. PMID:4983354

  17. Brucella HTRA Protein and Pathogenesis: Brucella Delta HTRA Strains as Vaccines

    National Research Council Canada - National Science Library

    Roop

    1997-01-01

    .... The results of studies described in previous reports confirmed that the Brucella HtrA contributes to the resistance of these intracellular pathogens to killing by host neutrophils and macrophages...

  18. Brucella HTRA Protein and Pathogenesis: Brucella Delta HTA Strains as Vaccines

    National Research Council Canada - National Science Library

    Roop, Martin

    1998-01-01

    .... The results of studies funded by contract DAMD17-94-C-4054 confirmed that the Brucella HtrA contributes to the resistance of these intracellular pathogens to killing by host neutrophils and macrophages...

  19. Establishment of Chronic Infection: Brucella's Stealth Strategy

    Science.gov (United States)

    Ahmed, Waqas; Zheng, Ke; Liu, Zheng-Fei

    2016-01-01

    Brucella is a facultative intracellular pathogen that causes zoonotic infection known as brucellosis which results in abortion and infertility in natural host. Humans, especially in low income countries, can acquire infection by direct contact with infected animal or by consumption of animal products and show high morbidity, severe economic losses and public health problems. However for survival, host cells develop complex immune mechanisms to defeat and battle against attacking pathogens and maintain a balance between host resistance and Brucella virulence. On the other hand as a successful intracellular pathogen, Brucella has evolved multiple strategies to evade immune response mechanisms to establish persistent infection and replication within host. In this review, we mainly summarize the “Stealth” strategies employed by Brucella to modulate innate and the adaptive immune systems, autophagy, apoptosis and possible role of small noncoding RNA in the establishment of chronic infection. The purpose of this review is to give an overview for recent understanding how this pathogen evades immune response mechanisms of host, which will facilitate to understanding the pathogenesis of brucellosis and the development of novel, more effective therapeutic approaches to treat brucellosis. PMID:27014640

  20. Identification and characterization of Brucella effector proteins

    NARCIS (Netherlands)

    de Jong, Maarten Frederik

    2012-01-01

    Brucella-bacteriën gebruiken de eiwitten VceB en VceC om het immuunsysteem van humane gastheercellen te omzeilen, blijkt uit het promotieonderzoek van Maarten de Jong. Dit biedt nieuwe aanknopingspunten voor de bestrijding van deze gevaarlijke bacterie. Brucellose is een wereldwijd voorkomende

  1. Brucella neotomae Infection in Humans, Costa Rica.

    Science.gov (United States)

    Suárez-Esquivel, Marcela; Ruiz-Villalobos, Nazareth; Jiménez-Rojas, César; Barquero-Calvo, Elías; Chacón-Díaz, Carlos; Víquez-Ruiz, Eunice; Rojas-Campos, Norman; Baker, Kate S; Oviedo-Sánchez, Gerardo; Amuy, Ernesto; Chaves-Olarte, Esteban; Thomson, Nicholas R; Moreno, Edgardo; Guzmán-Verri, Caterina

    2017-06-01

    Several species of Brucella are known to be zoonotic, but B. neotomae infection has been thought to be limited to wood rats. In 2008 and 2011, however, B. neotomae was isolated from cerebrospinal fluid of 2 men with neurobrucellosis. The nonzoonotic status of B. neotomae should be reassessed.

  2. Isolation of Brucella microti from soil

    Czech Academy of Sciences Publication Activity Database

    Scholz, H. C.; Hubálek, Zdeněk; Nesvadbová, Jiřina; Tomaso, H.; Vergnaud, G.; Le Fleche, P.; Whatmore, A. M.; Al Dahouk, S.; Krüger, M.; Lodri, C.; Pfeffer, M.

    2008-01-01

    Roč. 14, č. 8 (2008), s. 1316-1317 ISSN 1080-6040 Institutional research plan: CEZ:AV0Z60930519 Keywords : Brucella microti * Microtus arvalis Subject RIV: EE - Microbiology, Virology Impact factor: 6.449, year: 2008

  3. Brucella Infection Associated with Complete Atrioventricular Block.

    Science.gov (United States)

    Bilici, Meki; Demir, Fikri; Yılmazer, Murat Muhtar; Bozkurt, Fatma; Tuzcu, Volkan

    2016-09-01

    The clinical spectrum of Brucella infection is quite diverse and characterized by multi-system involvement. Patients present with myocarditis, endocarditis, or pericarditis. Infective endocarditis is the most common cardiovascular complication in patients with brucellosis. Although conduction abnormalities are seen in cases with endocarditis, they are reported very rarely in the setting of cardiac Brucella infection. An eight and a half-year-old male patient was referred to our clinic due to inadequate response to cotrimaxazole plus streptomycin treatment at the 15th day of admission. Although local hospital records on the patient showed a heart rate of 80 bpm, we determined a heart rate of 46 bpm. The electrocardiogram showed complete atrioventricular (AV) block. The average heart rate was determined as 48 bpm with 24-hour Holter electrocardiogram (ECG) monitoring. The echocardiographic examination showed normal-sized heart chambers and the absence of valvular involvement. An agglutination test for brucellosis was found to be positive with a titer of 1/320. High fever, arthralgia, and splenomegaly regressed following doxycycline plus rifampicin therapy, but there was no improvement in the AV block. A permanent pacemaker was implanted because of the detection of an average heart rate of 48 bpm. Because cardiac failure and rhythm abnormalities are reported in the course of Brucella infection and may be associated with significant outcomes, cases with brucellosis should be evaluated carefully in terms of cardiac involvement. This report aims to draw attention to complete AV block as an extremely rare complication of Brucella infection.

  4. Molecular typing of Brucella species isolates from livestock and human.

    Science.gov (United States)

    Nagalingam, Mohandoss; Shome, Rajeswari; Balamurugan, Vinayagamurthy; Shome, Bibek Ranjan; NarayanaRao, Krishnamsetty; Vivekananda; Isloor, Shrikrishna; Prabhudas, Krishnamsetty

    2012-01-01

    Although host specificity has been observed in different species of Brucella, crossing the animal host boundary is likely to occur at any time. In this study, Bruce ladder PCR and abortus-melitensis-ovis-suis (AMOS) PCR assays were used to characterize 47 Brucella isolates from Indian origin in order to know exact species for understanding epidemiology of brucellosis. Out of them, 28, 14, and 5 isolates were found to be Brucella abortus, Brucella melitensis, and Brucella suis, respectively. Further analysis by AMOS PCR has identified that all the B. abortus isolates belong to any one of the biovar 1, 2, or 4; of the five B. suis isolates, three belong to biovar 1 and two belong to any one of the biovar 2, 3, 4, or 5. Although this multiplex Bruce ladder PCR is useful in differentiating all Brucella species, elaborate study is required to further characterize the isolates at exact biovar level.

  5. Antimicrobial Susceptibility of Brucella melitensis Isolates in Peru

    Science.gov (United States)

    2011-03-01

    2011, American Society for Microbiology. All Rights Reserved. Antin1icrobial Susceptibility of Brucella melitensis Isolates in Peru 9 Ryan C. Maves,1...48 human Brucella melitensis biotype 1 strains from Peru between 2000 and 2006. MICs of isolates to doxycycline, azithromycin, gentamicin, rifampin...of testing. Relapses did nut appear to be related tu drug resistance. Infection by Brucella species is a major cause of zoonotic disease

  6. Case report 469: Spondylitis (lumbar spine) due to Brucella abortus

    Energy Technology Data Exchange (ETDEWEB)

    Manaster, B.J.

    1988-03-01

    The current case is interesting in that, although the plain radiographs were diagnostic of infection and the patient's work history suggested brucellosis, both the negative serum antibody titers to brucella and the CT appearance of large calcified psoas abscesses made the diagnosis of tuberculous spondylitis most probable. Open biopsy with tissue culture proved brucella. From this experience it appears that the presence of large calcified psoas abscesses should not eliminate the diagnosis of brucella spondylitis in the proper clinical setting.

  7. Host response to Brucella infection: review and future perspective.

    Science.gov (United States)

    Elfaki, Mohamed G; Alaidan, Alwaleed Abdullah; Al-Hokail, Abdullah Abdulrahman

    2015-07-30

    Brucellosis is a zoonotic and contagious infectious disease caused by infection with Brucella species. The infecting brucellae are capable of causing a devastating multi-organ disease in humans with serious health complications. The pathogenesis of Brucella infection is influenced largely by host factors, Brucella species/strain, and the ability of invading brucellae to survive and replicate within mononuclear phagocytic cells, preferentially macrophages (Mf). Consequently, the course of human infection may appear as an acute fatal or progress into chronic debilitating infection with periodical episodes that leads to bacteremia and death. The existence of brucellae inside Mf represents one of the strategies used by Brucella to evade the host immune response and is responsible for treatment failure in certain human populations treated with anti-Brucella drugs. Moreover, the persistence of brucellae inside Mf complicates the diagnosis and may affect the host cell signaling pathways with consequent alterations in both innate and adaptive immune responses. Therefore, there is an urgent need to pursue the development of novel drugs and/or vaccine targets against human brucellosis using high throughput technologies in genomics, proteomics, and immunology.

  8. MLVA-16 typing of 295 marine mammal Brucella isolates from different animal and geographic origins identifies 7 major groups within Brucella ceti and Brucella pinnipedialis

    Directory of Open Access Journals (Sweden)

    Jacques Isabelle

    2009-07-01

    Full Text Available Abstract Background Since 1994, Brucella strains have been isolated from a wide range of marine mammals. They are currently recognized as two new Brucella species, B. pinnipedialis for the pinniped isolates and B. ceti for the cetacean isolates in agreement with host preference and specific phenotypic and molecular markers. In order to investigate the genetic relationships within the marine mammal Brucella isolates and with reference to terrestrial mammal Brucella isolates, we applied in this study the Multiple Loci VNTR (Variable Number of Tandem Repeats Analysis (MLVA approach. A previously published assay comprising 16 loci (MLVA-16 that has been shown to be highly relevant and efficient for typing and clustering Brucella strains from animal and human origin was used. Results 294 marine mammal Brucella strains collected in European waters from 173 animals and a human isolate from New Zealand presumably from marine origin were investigated by MLVA-16. Marine mammal Brucella isolates were shown to be different from the recognized terrestrial mammal Brucella species and biovars and corresponded to 3 major related groups, one specific of the B. ceti strains, one of the B. pinnipedialis strains and the last composed of the human isolate. In the B. ceti group, 3 subclusters were identified, distinguishing a cluster of dolphin, minke whale and porpoise isolates and two clusters mostly composed of dolphin isolates. These results were in accordance with published analyses using other phenotypic or molecular approaches, or different panels of VNTR loci. The B. pinnipedialis group could be similarly subdivided in 3 subclusters, one composed exclusively of isolates from hooded seals (Cystophora cristata and the two others comprising other seal species isolates. Conclusion The clustering analysis of a large collection of marine mammal Brucella isolates from European waters significantly strengthens the current view of the population structure of these two

  9. Brucella dissociation is essential for macrophage egress and bacterial dissemination.

    Science.gov (United States)

    Pei, Jianwu; Kahl-McDonagh, Melissa; Ficht, Thomas A

    2014-01-01

    It has long been observed that smooth Brucella can dissociate into rough mutants that are cytotoxic to macrophages. However, the in vivo biological significance and/or mechanistic details of Brucella dissociation and cytotoxicity remain incomplete. In the current report, a plaque assay was developed using Brucella strains exhibiting varying degrees of cytotoxicity. Infected monolayers were observed daily using phase contrast microscopy for plaque formation while Brucella uptake and replication were monitored using an immunofluorescence assay (IFA). Visible plaques were detected at 4-5 days post infection (p.i.) with cytotoxic Brucella 16MΔmanBA at an MOI of 0.1. IFA staining demonstrated that the plaques consisted of macrophages with replicating Brucella. Visible plaques were not detected in monolayers infected with non-cytotoxic 16MΔmanBAΔvirB2 at an MOI of 0.1. However, IFA staining did reveal small groups of macrophages (foci) with replicating Brucella in the monolayers infected with 16MΔmanBAΔvirB2. The size of the foci observed in macrophage monolayers infected with rough Brucella correlated directly with cytotoxicity measured in liquid culture, suggesting that cytotoxicity was essential for Brucella egress and dissemination. In monolayers infected with 16M, small and large foci were observed. Double antibody staining revealed spontaneous rough mutants within the large, but not the small foci in 16M infected monolayers. Furthermore, plaque formation was observed in the large foci derived from 16M infections. Finally, the addition of gentamicin to the culture medium inhibited plaque formation, suggesting that cell-to-cell spread occurred only following release of the organisms from the cells. Taken together, these results demonstrate that Brucella-induced cytotoxicity is critical for Brucella egress and dissemination.

  10. Brucella Dissociation Is Essential for Macrophage Egress and Bacterial Dissemination

    Directory of Open Access Journals (Sweden)

    Thomas A Ficht

    2014-03-01

    Full Text Available It has long been observed that smooth Brucella can dissociate into rough mutants that are cytotoxic to macrophages. However, the in vivo biological significance and/or mechanistic de-tails of Brucella dissociation and cytotoxicity remain incomplete. In the current report, a plaque assay was developed using Brucella strains exhibiting varying degrees of cytotoxicity. Infected monolayers were observed daily using phase contrast microscopy for plaque formation while Brucella uptake and replication were monitored using an immunofluorescence assay (IFA. Vis-ible plaques were detected at 4-5 days post infection (p.i. with cytotoxic Brucella 16M∆manBA at an MOI of 0.1. IFA staining demonstrated that the plaques consisted of macrophages with replicating Brucella. Visible plaques were not detected in monolayers infected with non-cytotoxic 16M∆manBA∆virB2 at an MOI of 0.1. However, IFA staining did reveal small groups of macrophages (foci with replicating Brucella in the monolayers infected with 16M∆manBA∆virB2. The size of the foci observed in macrophage monolayers infected with rough Brucella correlated directly with cytotoxicity measured in liquid culture, suggesting that cytotoxicity was essential for Brucella egress and dissemination. In monolayers infected with 16M, small and large foci were observed. Double antibody staining revealed spontaneous rough mutants within the large, but not the small foci in 16M infected monolayers. Furthermore, plaque formation was observed in the large foci derived from 16M infections. Finally, the addi-tion of gentamicin to the culture medium inhibited plaque formation, suggesting that the cell-to-cell spreading occurred only following release of the organisms from the cells. Taken together, these results demonstrate that Brucella induced cytotoxicity is critical for Brucella egress and dissemination.

  11. The new strains Brucella inopinata BO1 and Brucella species 83-210 behave biologically like classic infectious Brucella species and cause death in murine models of infection.

    Science.gov (United States)

    Jiménez de Bagüés, María P; Iturralde, María; Arias, Maykel A; Pardo, Julián; Cloeckaert, Axel; Zygmunt, Michel S

    2014-08-01

    Recently, novel atypical Brucella strains isolated from humans and wild rodents have been reported. They are phenotypically close to Ochrobactrum species but belong to the genus Brucella, based on genetic relatedness, although genetic diversity is higher among the atypical Brucella strains than between the classic species. They were classified within or close to the novel species Brucella inopinata. However, with the exception of Brucella microti, the virulence of these novel strains has not been investigated in experimental models of infection. The type species B. inopinata strain BO1 (isolated from a human) and Brucella species strain 83-210 (isolated from a wild Australian rodent) were investigated. A classic infectious Brucella reference strain, B. suis 1330, was also used. BALB/c, C57BL/6, and CD1 mice models and C57BL/6 mouse bone-marrow-derived macrophages (BMDMs) were used as infection models. Strains BO1 and 83-210 behaved similarly to reference strain 1330 in all mouse infection models: there were similar growth curves in spleens and livers of mice and similar intracellular replication rates in BMDMs. However, unlike strain 1330, strains BO1 and 83-210 showed lethality in the 3 mouse models. The novel atypical Brucella strains of this study behave like classic intracellular Brucella pathogens. In addition, they cause death in murine models of infection, as previously published for B. microti, another recently described environmental and wildlife species. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  12. Mean platelet volume in brucellosis: correlation between brucella ...

    African Journals Online (AJOL)

    Background: Brucellosis, a zoonotic infection, was most widely diagnosed by the Brucella standard serum agglutination test (SAT). No previous publication has demonstrated a correlation between the degree of Brucella SAT agglutination positivity and the severity of brucellosis infection. Objective: To contribute to the ...

  13. Knowledge of Brucella as a food-borne pathogen

    Science.gov (United States)

    Although Brucella spp. are known for causing reproductive losses in domestic livestock, they are also capable of infecting humans and causing clinical disease. Human infection with Brucella is almost exclusively a result of direct contact with infected animals or consumption of products made from un...

  14. A history of the development of Brucella vaccines.

    Science.gov (United States)

    Avila-Calderón, Eric Daniel; Lopez-Merino, Ahidé; Sriranganathan, Nammalwar; Boyle, Stephen M; Contreras-Rodríguez, Araceli

    2013-01-01

    Brucellosis is a worldwide zoonosis affecting animal and human health. In the last several decades, much research has been performed to develop safer Brucella vaccines to control the disease mainly in animals. Till now, no effective human vaccine is available. The aim of this paper is to review and discuss the importance of methodologies used to develop Brucella vaccines in pursuing this challenge.

  15. Bioinformatics analysis of Brucella vaccines and vaccine targets using VIOLIN.

    Science.gov (United States)

    He, Yongqun; Xiang, Zuoshuang

    2010-09-27

    Brucella spp. are Gram-negative, facultative intracellular bacteria that cause brucellosis, one of the commonest zoonotic diseases found worldwide in humans and a variety of animal species. While several animal vaccines are available, there is no effective and safe vaccine for prevention of brucellosis in humans. VIOLIN (http://www.violinet.org) is a web-based vaccine database and analysis system that curates, stores, and analyzes published data of commercialized vaccines, and vaccines in clinical trials or in research. VIOLIN contains information for 454 vaccines or vaccine candidates for 73 pathogens. VIOLIN also contains many bioinformatics tools for vaccine data analysis, data integration, and vaccine target prediction. To demonstrate the applicability of VIOLIN for vaccine research, VIOLIN was used for bioinformatics analysis of existing Brucella vaccines and prediction of new Brucella vaccine targets. VIOLIN contains many literature mining programs (e.g., Vaxmesh) that provide in-depth analysis of Brucella vaccine literature. As a result of manual literature curation, VIOLIN contains information for 38 Brucella vaccines or vaccine candidates, 14 protective Brucella antigens, and 68 host response studies to Brucella vaccines from 97 peer-reviewed articles. These Brucella vaccines are classified in the Vaccine Ontology (VO) system and used for different ontological applications. The web-based VIOLIN vaccine target prediction program Vaxign was used to predict new Brucella vaccine targets. Vaxign identified 14 outer membrane proteins that are conserved in six virulent strains from B. abortus, B. melitensis, and B. suis that are pathogenic in humans. Of the 14 membrane proteins, two proteins (Omp2b and Omp31-1) are not present in B. ovis, a Brucella species that is not pathogenic in humans. Brucella vaccine data stored in VIOLIN were compared and analyzed using the VIOLIN query system. Bioinformatics curation and ontological representation of Brucella vaccines

  16. Pathology of striped dolphins (Stenella coeruleoalba) infected with Brucella ceti.

    Science.gov (United States)

    González-Barrientos, R; Morales, J-A; Hernández-Mora, G; Barquero-Calvo, E; Guzmán-Verri, C; Chaves-Olarte, E; Moreno, E

    2010-05-01

    Seventeen striped dolphins (Stenella coeruleoalba) displaying swimming disorders compatible with neurological syndromes were investigated for Brucella infection. Sixteen dolphins had meningoencephalomyelitis. Serum antibody against Brucella antigen was detected in all 14 animals tested and Brucella ceti was isolated from eight out of nine animals. Brucella antigen was detected in the brain by immunofluorescence, but not by immunohistochemical labelling. By contrast, Brucella antigen was demonstrated by immunohistochemistry in the trophoblast of animals with severe placentitis and in the mitral valve of animals with myocarditis. The microscopical lesions observed in the tissues of the infected dolphins were similar to those of chronic brucellosis in man. The severity of brucellosis in S. coeruleoalba indicates that this dolphin species is highly susceptible to infection by B. ceti. Copyright 2009 Elsevier Ltd. All rights reserved.

  17. Nucleotide sequences specific to Brucella and methods for the detection of Brucella

    Energy Technology Data Exchange (ETDEWEB)

    McCready, Paula M [Tracy, CA; Radnedge, Lyndsay [San Mateo, CA; Andersen, Gary L [Berkeley, CA; Ott, Linda L [Livermore, CA; Slezak, Thomas R [Livermore, CA; Kuczmarski, Thomas A [Livermore, CA

    2009-02-24

    Nucleotide sequences specific to Brucella that serves as a marker or signature for identification of this bacterium were identified. In addition, forward and reverse primers and hybridization probes derived from these nucleotide sequences that are used in nucleotide detection methods to detect the presence of the bacterium are disclosed.

  18. The genome sequence of Brucella pinnipedialis B2/94 sheds light on the evolutionary history of the genus Brucella

    Directory of Open Access Journals (Sweden)

    Claverie Jean-Michel

    2011-07-01

    Full Text Available Abstract Background Since the discovery of the Malta fever agent, Brucella melitensis, in the 19th century, six terrestrial mammal-associated Brucella species were recognized over the next century. More recently the number of novel Brucella species has increased and among them, isolation of species B. pinnipedialis and B. ceti from marine mammals raised many questions about their origin as well as on the evolutionary history of the whole genus. Results We report here on the first complete genome sequence of a Brucella strain isolated from marine mammals, Brucella pinnipedialis strain B2/94. A whole gene-based phylogenetic analysis shows that five main groups of host-associated Brucella species rapidly diverged from a likely free-living ancestor close to the recently isolated B. microti. However, this tree lacks the resolution required to resolve the order of divergence of those groups. Comparative analyses focusing on a genome segments unshared between B. microti and B. pinnipedialis, b gene deletion/fusion events and c positions and numbers of Brucella specific IS711 elements in the available Brucella genomes provided enough information to propose a branching order for those five groups. Conclusions In this study, it appears that the closest relatives of marine mammal Brucella sp. are B. ovis and Brucella sp. NVSL 07-0026 isolated from a baboon, followed by B. melitensis and B. abortus strains, and finally the group consisting of B. suis strains, including B. canis and the group consisting of the single B. neotomae species. We were not able, however, to resolve the order of divergence of the two latter groups.

  19. The genome sequence of Brucella pinnipedialis B2/94 sheds light on the evolutionary history of the genus Brucella

    Science.gov (United States)

    2011-01-01

    Background Since the discovery of the Malta fever agent, Brucella melitensis, in the 19th century, six terrestrial mammal-associated Brucella species were recognized over the next century. More recently the number of novel Brucella species has increased and among them, isolation of species B. pinnipedialis and B. ceti from marine mammals raised many questions about their origin as well as on the evolutionary history of the whole genus. Results We report here on the first complete genome sequence of a Brucella strain isolated from marine mammals, Brucella pinnipedialis strain B2/94. A whole gene-based phylogenetic analysis shows that five main groups of host-associated Brucella species rapidly diverged from a likely free-living ancestor close to the recently isolated B. microti. However, this tree lacks the resolution required to resolve the order of divergence of those groups. Comparative analyses focusing on a) genome segments unshared between B. microti and B. pinnipedialis, b) gene deletion/fusion events and c) positions and numbers of Brucella specific IS711 elements in the available Brucella genomes provided enough information to propose a branching order for those five groups. Conclusions In this study, it appears that the closest relatives of marine mammal Brucella sp. are B. ovis and Brucella sp. NVSL 07-0026 isolated from a baboon, followed by B. melitensis and B. abortus strains, and finally the group consisting of B. suis strains, including B. canis and the group consisting of the single B. neotomae species. We were not able, however, to resolve the order of divergence of the two latter groups. PMID:21745361

  20. Demonstration of IS711 transposition in Brucella ovis and Brucella pinnipedialis

    Directory of Open Access Journals (Sweden)

    García-Lobo Juan M

    2008-01-01

    Full Text Available Abstract Background The Brucella genome contains an insertion sequence (IS element called IS711 or IS6501, which is specific to the genus. The copy number of IS711 varies in the genome of the different Brucella species, ranging from 7 in B. abortus, B. melitensis and B. suis to more than 30 in B. ovis and in Brucella strains isolated from marine mammals. At present, there is no experimental evidence of transposition of IS711, but the occurrence of this element with a high copy number in some species, and the isolation of Brucella strains with "ectopic" copies of IS711 suggested that this IS could still transpose. Results In this study we obtained evidence of transposition of IS711 from the B. ovis and B. pinnipedialis chromosomes by using the "transposon trap" plasmid pGBG1. This plasmid expresses resistance to tetracycline only if the repressor gene that it contains is inactivated. The strains B. melitensis 16 M, B. abortus RB51, B. ovis BOC22 (field strain and B. pinnipedialis B2/94, all containing the plasmid pGBG1, were grown in culture media with tetracycline until the appearance of tetracycline resistant mutants (TcR. TcR mutants due to IS711 transposition were only detected in B. ovis and B. pinnipedialis strains. Conclusion Four different copies of IS711 were found to transpose to the same target sequence in the plasmid pGBG1. This demonstrated that IS711 are active in vivo, specially in Brucella species with a high number of IS711 copies as B. ovis and B. pinnipedialis.

  1. Identification of Secondary Mutations Which Enhance and Stabilize the Attenuation of Brucella HTRA Mutants: Improving Brucella HTRA-Based Strains as Vaccine Candidates

    National Research Council Canada - National Science Library

    Roop, R

    2000-01-01

    Results to date from the studies funded under this contract suggest that Brucella genes involved in maintaining efficient stationary phase physiology and allowing the brucellae to make effective use...

  2. Characterisation of North American Brucella isolates from marine mammals.

    Science.gov (United States)

    Whatmore, Adrian M; Dawson, Claire; Muchowski, Jakub; Perrett, Lorraine L; Stubberfield, Emma; Koylass, Mark; Foster, Geoffrey; Davison, Nicholas J; Quance, Christine; Sidor, Inga F; Field, Cara L; St Leger, Judy

    2017-01-01

    Extension of known ecological niches of Brucella has included the description of two novel species from marine mammals. Brucella pinnipedialis is associated predominantly with seals, while two major Brucella ceti clades, most commonly associated with porpoises or dolphins respectively, have been identified. To date there has been limited characterisation of Brucella isolates obtained from marine mammals outside Northern European waters, including North American waters. To address this gap, and extend knowledge of the global population structure and host associations of these Brucella species, 61 isolates from marine mammals inhabiting North American waters were subject to molecular and phenotypic characterisation enabling comparison with existing European isolates. The majority of isolates represent genotypes previously described in Europe although novel genotypes were identified in both B. ceti clades. Harp seals were found to carry B. pinnipedialis genotypes previously confined to hooded seals among a diverse repertoire of sequence types (STs) associated with this species. For the first time Brucella isolates were characterised from beluga whales and found to represent a number of distinct B. pinnipedialis genotypes. In addition the known host range of ST27 was extended with the identification of this ST from California sea lion samples. Finally the performance of the frequently used diagnostic tool Bruce-ladder, in differentiating B. ceti and B. pinnipedialis, was critically assessed based on improved knowledge of the global population structure of Brucella associated with marine mammals.

  3. Characterisation of North American Brucella isolates from marine mammals.

    Directory of Open Access Journals (Sweden)

    Adrian M Whatmore

    Full Text Available Extension of known ecological niches of Brucella has included the description of two novel species from marine mammals. Brucella pinnipedialis is associated predominantly with seals, while two major Brucella ceti clades, most commonly associated with porpoises or dolphins respectively, have been identified. To date there has been limited characterisation of Brucella isolates obtained from marine mammals outside Northern European waters, including North American waters. To address this gap, and extend knowledge of the global population structure and host associations of these Brucella species, 61 isolates from marine mammals inhabiting North American waters were subject to molecular and phenotypic characterisation enabling comparison with existing European isolates. The majority of isolates represent genotypes previously described in Europe although novel genotypes were identified in both B. ceti clades. Harp seals were found to carry B. pinnipedialis genotypes previously confined to hooded seals among a diverse repertoire of sequence types (STs associated with this species. For the first time Brucella isolates were characterised from beluga whales and found to represent a number of distinct B. pinnipedialis genotypes. In addition the known host range of ST27 was extended with the identification of this ST from California sea lion samples. Finally the performance of the frequently used diagnostic tool Bruce-ladder, in differentiating B. ceti and B. pinnipedialis, was critically assessed based on improved knowledge of the global population structure of Brucella associated with marine mammals.

  4. Macrophage activation induced by Brucella DNA suppresses bacterial intracellular replication via enhancing NO production.

    Science.gov (United States)

    Liu, Ning; Wang, Lin; Sun, Changjiang; Yang, Li; Tang, Bin; Sun, Wanchun; Peng, Qisheng

    2015-12-01

    Brucella DNA can be sensed by TLR9 on endosomal membrane and by cytosolic AIM2-inflammasome to induce proinflammatory cytokine production that contributes to partially activate innate immunity. Additionally, Brucella DNA has been identified to be able to act as a major bacterial component to induce type I IFN. However, the role of Brucella DNA in Brucella intracellular growth remains unknown. Here, we showed that stimulation with Brucella DNA promote macrophage activation in TLR9-dependent manner. Activated macrophages can suppresses wild type Brucella intracellular replication at early stage of infection via enhancing NO production. We also reported that activated macrophage promotes bactericidal function of macrophages infected with VirB-deficient Brucella at the early or late stage of infection. This study uncovers a novel function of Brucella DNA, which can help us further elucidate the mechanism of Brucella intracellular survival. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Whole-genome analyses of speciation events in pathogenic Brucellae

    Energy Technology Data Exchange (ETDEWEB)

    Chain, Patrick S. G. [Lawrence Livermore National Laboratory (LLNL); Comerci, Diego J. [Universidad Nacional de General San Martin; Tolmasky, Marcelo E. [California State University; Larimer, Frank W [ORNL; Malfatti, Stephanie [Lawrence Livermore National Laboratory (LLNL); Vergez, Lisa [Lawrence Livermore National Laboratory (LLNL); Aguero, Fernan [Universidad Nacional de General San Martin; Land, Miriam L [ORNL; Ugalde, Rodolfo A. [Universidad Nacional de General San Martin; Garcia, Emilio [Lawrence Livermore National Laboratory (LLNL)

    2005-12-01

    Despite their high DNA identity and a proposal to group classical Brucella species as biovars of Brucella melitensis, the commonly recognized Brucella species can be distinguished by distinct biochemical and fatty acid characters, as well as by a marked host range (e.g., Brucella suis for swine, B. melitensis for sheep and goats, and Brucella abortus for cattle). Here we present the genome of B. abortus 2308, the virulent prototype biovar 1 strain, and its comparison to the two other human pathogenic Brucella species and to B. abortus field isolate 9-941. The global distribution of pseudogenes, deletions, and insertions supports previous indications that B. abortus and B. melitensis share a common ancestor that diverged from B. suis. With the exception of a dozen genes, the genetic complements of both B. abortus strains are identical, whereas the three species differ in gene content and pseudogenes. The pattern of species-specific gene inactivations affecting transcriptional regulators and outer membrane proteins suggests that these inactivations may play an important role in the establishment of host specificity and may have been a primary driver of speciation in the genus Brucella. Despite being nonmotile, the brucellae contain flagellum gene clusters and display species-specific flagellar gene inactivations, which lead to the putative generation of different versions of flagellum-derived structures and may contribute to differences in host specificity and virulence. Metabolic changes such as the lack of complete metabolic pathways for the synthesis of numerous compounds (e.g., glycogen, biotin, NAD, and choline) are consistent with adaptation of brucellae to an intracellular life-style.

  6. Brucella contamination in raw milk by polymerase chain reaction

    Directory of Open Access Journals (Sweden)

    Mohammad Khalili

    2016-10-01

    Full Text Available Background: Human brucellosis is a significant public health problem in many middle east countries including Iran. Brucella organisms, which are small aerobic, facultative intracellular coccobacilli, localize in the reproductive organs of host animals, causing abortions and sterility. They are shed in large numbers in the animal’s urine, milk, placental fluid, and other fluids. Dairy product from raw milk are a potential threat to public health in endemic developing countries. The gold standard for the diagnosis of brucellosis is isolation of Brucella species. However, isolation Brucella species is time consuming and needed to level 3 biocontainment facilities and highly skilled technical personnel to handle samples and live bacteria for eventual identification. Handling Brucella species increase risk of laboratory infection. Polymerase chain reaction (PCR with high sensitivity and specifity overcomed to these disadvantages. The aim of this study was to detect Brucella species in milk from dairy cattle farms in Kerman province, Iran by PCR technique. Methods: Forty and eight bulk tank milk (BTM were collected from October 2015 to March 2016 from 48 dairy cattle farm including 4200 cows. DNA of milk samples extracted by lysis buffer and proteinase K method. All milk samples were examined by PCR to detect Brucella-specific DNA targeting IS 711. Positive samples must be showed 317 bp amplified, corresponding to the expected size of the IS 711 genome region in all Brucella species. Results: Using IS711 primer were detected in 4 samples (8.3% Brucella spp. from 48 BTM samples in this area. Conclusion: The results indicate that brucellosis by Brucella species is endemic in the Kerman province dairy farms. Consumption of raw milk dairy products by individual farmers operating under poor hygienic conditions represents an high risk to public health. The need for implementing control measures and raising public awareness on zoonotic transmission of

  7. Research progress in live attenuated Brucella vaccine development.

    Science.gov (United States)

    Wang, Zhen; Wu, Qingmin

    2013-01-01

    Brucella spp. are facultative intracellular bacteria that cause brucellosis, which is a globally occurring zoonotic disease that is characterized by abortion in domestic animals and undulant fever, arthritis, endocarditis, and meningitis in humans. There are currently no licensed vaccines against brucellosis for human use, and only a few licensed live Brucella vaccines are available for use in animals. However, the available animal vaccines may cause abortion and are associated with lower protection rates in animals and higher virulence in humans. Much research has been performed recently to develop novel Brucella vaccines for the prevention and control of animal brucellosis. This article discusses the approaches and strategies for novel live attenuated vaccine development.

  8. Brucella papionis sp. nov., isolated from baboons (Papio spp.).

    Science.gov (United States)

    Whatmore, Adrian M; Davison, Nicholas; Cloeckaert, Axel; Al Dahouk, Sascha; Zygmunt, Michel S; Brew, Simon D; Perrett, Lorraine L; Koylass, Mark S; Vergnaud, Gilles; Quance, Christine; Scholz, Holger C; Dick, Edward J; Hubbard, Gene; Schlabritz-Loutsevitch, Natalia E

    2014-12-01

    Two Gram-negative, non-motile, non-spore-forming coccoid bacteria (strains F8/08-60(T) and F8/08-61) isolated from clinical specimens obtained from baboons (Papio spp.) that had delivered stillborn offspring were subjected to a polyphasic taxonomic study. On the basis of 16S rRNA gene sequence similarities, both strains, which possessed identical sequences, were assigned to the genus Brucella. This placement was confirmed by extended multilocus sequence analysis (MLSA), where both strains possessed identical sequences, and whole-genome sequencing of a representative isolate. All of the above analyses suggested that the two strains represent a novel lineage within the genus Brucella. The strains also possessed a unique profile when subjected to the phenotyping approach classically used to separate species of the genus Brucella, reacting only with Brucella A monospecific antiserum, being sensitive to the dyes thionin and fuchsin, being lysed by bacteriophage Wb, Bk2 and Fi phage at routine test dilution (RTD) but only partially sensitive to bacteriophage Tb, and with no requirement for CO2 and no production of H2S but strong urease activity. Biochemical profiling revealed a pattern of enzyme activity and metabolic capabilities distinct from existing species of the genus Brucella. Molecular analysis of the omp2 locus genes showed that both strains had a novel combination of two highly similar omp2b gene copies. The two strains shared a unique fingerprint profile of the multiple-copy Brucella-specific element IS711. Like MLSA, a multilocus variable number of tandem repeat analysis (MLVA) showed that the isolates clustered together very closely, but represent a distinct group within the genus Brucella. Isolates F8/08-60(T) and F8/08-61 could be distinguished clearly from all known species of the genus Brucella and their biovars by both phenotypic and molecular properties. Therefore, by applying the species concept for the genus Brucella suggested by the ICSP

  9. A Brucella spp. Isolate from a Pac-Man Frog (Ceratophrys ornata) Reveals Characteristics Departing from Classical Brucellae

    Science.gov (United States)

    Soler-Lloréns, Pedro F.; Quance, Chris R.; Lawhon, Sara D.; Stuber, Tod P.; Edwards, John F.; Ficht, Thomas A.; Robbe-Austerman, Suelee; O'Callaghan, David; Keriel, Anne

    2016-01-01

    Brucella are highly infectious bacterial pathogens responsible for brucellosis, a frequent worldwide zoonosis. The Brucella genus has recently expanded from 6 to 11 species, all of which were associated with mammals; The natural host range recently expanded to amphibians after some reports of atypical strains from frogs. Here we describe the first in depth phenotypic and genetic characterization of a Brucella strains isolated from a frog. Strain B13-0095 was isolated from a Pac-Man frog (Ceratophyrus ornate) at a veterinary hospital in Texas and was initially misidentified as Ochrobactrum anthropi. We found that B13-0095 belongs to a group of early-diverging brucellae that includes Brucella inopinata strain BO1 and the B. inopinata-like strain BO2, with traits that depart significantly from those of the “classical” Brucella spp. Analysis of B13-0095 genome sequence revealed several specific features that suggest that this isolate represents an intermediate between a soil associated ancestor and the host adapted “classical” species. Like strain BO2, B13-0095 does not possess the genes required to produce the perosamine based LPS found in classical Brucella, but has a set of genes that could encode a rhamnose based O-antigen. Despite this, B13-0095 has a very fast intracellular replication rate in both epithelial cells and macrophages. Finally, another major finding in this study is the bacterial motility observed for strains B13-0095, BO1, and BO2, which is remarkable for this bacterial genus. This study thus highlights several novel characteristics in strains belonging to an emerging group within the Brucella genus. Accurate identification tools for such atypical Brucella isolates and careful evaluation of their zoonotic potential, are urgently required. PMID:27734009

  10. A Brucella spp. Isolate from a Pac-Man Frog (Ceratophrys ornata Reveals Characteristics Departing from Classical Brucellae

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    Pedro Franscico Soler Llorens

    2016-09-01

    Full Text Available Brucella are highly infectious bacterial pathogens responsible for brucellosis, a frequent worldwide zoonosis. The Brucella genus has recently expanded from 6 to 11 species, all of which were associated with mammals; The natural host range recently expanded to amphibians after some reports of atypical strains from frogs. Here we describe the first in depth phenotypic and genetic characterization of a Brucella strains isolated from a frog. Strain B13-0095 was isolated from a Pac-Man frog (Ceratophyrus ornate at a veterinary hospital in Texas and was initially misidentified as Ochrobactrum anthropi. We found that B13-0095 belongs to a group of early-diverging brucellae that includes Brucella inopinata strain BO1 and the B. inopinata-like strain BO2, with traits that depart significantly from those of the ‘classical’ Brucella spp. Analysis of B13-0095 genome sequence revealed several specific features that suggest that this isolate represents an intermediate between a soil associated ancestor and the host adapted ‘classical’ species. Like strain BO2, B13-0095 does not possess the genes required to produce the perosamine based LPS found in classical Brucella, but has a set of genes that could encode a rhamnose based O-antigen. Despite this, B13-0095 has a very fast intracellular replication rate in both epithelial cells and macrophages. Finally, another major finding in this study is the bacterial motility observed for strains B13-0095, BO1 and BO2, which is remarkable for this bacterial genus.This study thus highlights several novel characteristics in strains belonging to an emerging group within the Brucella genus. Accurate identification tools for such atypical Brucella isolates and careful evaluation of their zoonotic potential, are urgently required.

  11. The changing nature of the Brucella-containing vacuole.

    Science.gov (United States)

    Celli, Jean

    2015-07-01

    Bacteria of the genus Brucella are intracellular vacuolar pathogens of mammals that cause the worldwide zoonosis brucellosis, and reside within phagocytes of infected hosts to promote their survival, persistence and proliferation. These traits are essential to the bacterium's ability to cause disease and have been the subject of much investigation to gain an understanding of Brucella pathogenic mechanisms. Although the endoplasmic reticulum-derived nature of the Brucella replicative niche has been long known, major strides have recently been made in deciphering the molecular mechanisms of its biogenesis, including the identification of bacterial determinants and host cellular pathways involved in this process. Here I will review and discuss the most recent advances in our knowledge of Brucella intracellular pathogenesis, with an emphasis on bacterial exploitation of the host endoplasmic reticulum-associated functions, and how autophagy-related processes contribute to the bacterium's intracellular cycle. © 2015 John Wiley & Sons Ltd.

  12. Serological Survey of Antibodies against Brucella Organisms in One ...

    African Journals Online (AJOL)

    Serological Survey of Antibodies against Brucella Organisms in One Humped Camel (Camelus dromedarius) Herds in the Lake Chad Area of Borno State, North Eastern Nigeria. MA Sadiq, I Ajogi, JOO Bale, FB Mosimabale, AN Tijjani, AA Kaikabo ...

  13. Brucella species survey in polar bears (ursus maritimus) of northern Alaska.

    Science.gov (United States)

    O'Hara, Todd M; Holcomb, Darce; Elzer, Philip; Estepp, Jessica; Perry, Quinesha; Hagius, Sue; Kirk, Cassandra

    2010-07-01

    We report on the presence of specific antibodies to Brucella spp. and Yersinia enterocolitica in polar bears (Ursus maritimus) from northern Alaska (southern Beaufort Sea) during 2003-2006. Based on numerous known stressors (e.g., climate change and loss of sea ice habitat, contaminants), there is increased concern regarding the status of polar bears. Considering these changes, it is important to assess exposure to potentially pathogenic organisms and to improve understanding of transmission pathways. Brucella or specific antibodies to Brucella spp. has been reported in marine mammals. Various assays were used to elucidate the pathway or source of exposure (e.g., "marine" vs. "terrestrial" Brucella spp.) of northern Alaska polar bears to Brucella spp. The standard plate test (SPT) and the buffered Brucella antigen card test (BBA) were used for initial screening for antibodies specific to Brucella. We then evaluated positive reactors (presence of serum antibody specific for Brucella spp.) using immunoblots and competitive enzyme-linked immunosorbent assay (cELISA; based on pinniped-derived Brucella spp. antigen). Annual prevalence of antibody (BBA and SPT) for Brucella spp. ranged from 6.8% to 18.5% over 2003-2006, with an overall prevalence of 10.2%. Prevalence of Brucella spp. antibody did vary by age class. Western blot analyses indicated 17 samples were positive for Brucella spp. antibody; of these, 13 were negative by marine (pinniped) derived Brucella antigen cELISA and four were positive by marine cELISA. Of the four samples positive for Brucella antibody by marine cELISA, three cross-reacted with Y. enterocolitica and Brucella spp. (one sample was Brucella negative and Y. enterocolitica positive). It appears the polar bear antibody does not react with the antigens used on the marine cELISA assay, potentially indicating a terrestrial (nonpinniped) source of Brucella spp.

  14. Genetic diversity of Brucella abortus and Brucella melitensis in Kazakhstan using MLVA-16.

    Science.gov (United States)

    Shevtsov, Alexandr; Ramanculov, Erlan; Shevtsova, Elena; Kairzhanova, Alma; Tarlykov, Pavel; Filipenko, Maxim; Dymova, Maya; Abisheva, Gulzada; Jailbekova, Aygul; Kamalova, Dinara; Chsherbakov, Andrei; Tulegenov, Samat; Akhmetova, Assel; Sytnik, Igor; Karibaev, Talgat; Mukanov, Kasim

    2015-08-01

    Brucellosis is an endemic disease in Central Asia characterized by high infection rates in humans and animals. Currently, little is known about the genetic diversity of Brucella spp. circulating in the region, despite the high prevalence of brucellosis. This study aimed to analyze the genetic diversity of Brucella melitensis and Brucella abortus strains circulating in the Republic of Kazakhstan. We genotyped 128 B. melitensis and 124 B. abortus strains collected in regions with the highest prevalence of brucellosis. Genotyping was performed using multi-locus variable-number tandem-repeat analysis (MLVA). Analysis of a subset of 8 loci (MLVA-8) of 128 B. melitensis strains identified genotypes 42 (n=108), 43 (n=2), and 63 (n=19) related to the 'East Mediterranean' group. An MLVA-16 assay sorted 128 B. melitensis strains into 25 different genotypes. Excluding one variable locus, MLVA-15 of B. melitensis was distinct from strains originating in the Mediterranean region; however, 77% of them were identical to strains isolated in China. A minimum spanning tree for B. melitensis using MLVA-15 analysis clustered the local strains together with strains previously collected in China. MLVA-8 analysis of 124 B. abortus strains identified them as genotype 36, suggesting Eurasian distribution of this lineage. Complete MLVA-16 assay analysis clustered the strains into five genotypes, revealing little diversity of B. abortus when compared on the global scale. A minimum spanning tree for B. abortus obtained using MLVA-15 analysis clustered the 2 most prevalent genotypes (n=117) together with strains previously collected in China. Thus, MLVA analysis was used to characterize 252 strains of Brucella collected in Kazakhstan. The analysis revealed genetic homogeneity among the strains. Interestingly, identical MLVA-15 profiles were found in seemingly unrelated outbreaks in China, Turkey, and Kazakhstan. Further analysis is needed for better understanding of the epidemiology of

  15. Analyzing the molecular mechanism of lipoprotein localization in Brucella

    Science.gov (United States)

    Goolab, Shivani; Roth, Robyn L.; van Heerden, Henriette; Crampton, Michael C.

    2015-01-01

    Bacterial lipoproteins possess diverse structure and functionality, ranging from bacterial physiology to pathogenic processes. As such many lipoproteins, originating from Brucella are exploited as potential vaccines to countermeasure brucellosis infection in the host. These membrane proteins are translocated from the cytoplasm to the cell membrane where they are anchored peripherally by a multifaceted targeting mechanism. Although much research has focused on the identification and classification of Brucella lipoproteins and their potential use as vaccine candidates for the treatment of Brucellosis, the underlying route for the translocation of these lipoproteins to the outer surface of the Brucella (and other pathogens) outer membrane (OM) remains mostly unknown. This is partly due to the complexity of the organism and evasive tactics used to escape the host immune system, the variation in biological structure and activity of lipoproteins, combined with the complex nature of the translocation machinery. The biosynthetic pathway of Brucella lipoproteins involves a distinct secretion system aiding translocation from the cytoplasm, where they are modified by lipidation, sorted by the lipoprotein localization machinery pathway and thereafter equipped for export to the OM. Surface localized lipoproteins in Brucella may employ a lipoprotein flippase or the β-barrel assembly complex for translocation. This review provides an overview of the characterized Brucella OM proteins that form part of the OM, including a handful of other characterized bacterial lipoproteins and their mechanisms of translocation. Lipoprotein localization pathways in gram negative bacteria will be used as a model to identify gaps in Brucella lipoprotein localization and infer a potential pathway. Of particular interest are the dual topology lipoproteins identified in Escherichia coli and Haemophilus influenza. The localization and topology of these lipoproteins from other gram negative bacteria

  16. A History of the Development of Brucella Vaccines

    OpenAIRE

    Eric Daniel Avila-Calderón; Ahidé Lopez-Merino; Nammalwar Sriranganathan; Boyle, Stephen M; Araceli Contreras-Rodríguez

    2013-01-01

    Brucellosis is a worldwide zoonosis affecting animal and human health. In the last several decades, much research has been performed to develop safer Brucella vaccines to control the disease mainly in animals. Till now, no effective human vaccine is available. The aim of this paper is to review and discuss the importance of methodologies used to develop Brucella vaccines in pursuing this challenge. CONACYT CB-2011-01, 169259 SIP-IPN 20110891, 20134610 ICYTDF-IPN (Project of Investiga...

  17. A History of the Development of Brucella Vaccines

    Directory of Open Access Journals (Sweden)

    Eric Daniel Avila-Calderón

    2013-01-01

    Full Text Available Brucellosis is a worldwide zoonosis affecting animal and human health. In the last several decades, much research has been performed to develop safer Brucella vaccines to control the disease mainly in animals. Till now, no effective human vaccine is available. The aim of this paper is to review and discuss the importance of methodologies used to develop Brucella vaccines in pursuing this challenge.

  18. Analyzing the molecular mechanism of lipoprotein localization in Brucella

    CSIR Research Space (South Africa)

    Goolab, S

    2015-10-01

    Full Text Available (Comerci et al., 2001; Delrue et al., 2001; Martinez-Nunez et al., 2010). Furthermore, gene expression profiling used to define Brucella pathogenesis identified the Omps, lipoproteins, stress response proteins, Frontiers in Microbiology | www... al., 1984; Martinez de Tejada and Moriyon, 1993). Moreover, resistance to the bactericidal oxygen- independent systems of phagocytes characterizes the Brucella OM as a result of the resistance the LPS have to polycations (Martin and Hancock, 1990...

  19. Type IV secretion system of Brucella spp. and its effectors.

    Science.gov (United States)

    Ke, Yuehua; Wang, Yufei; Li, Wengfeng; Chen, Zeliang

    2015-01-01

    Brucella spp. are intracellular bacterial pathogens that cause infection in domestic and wild animals. They are often used as model organisms to study intracellular bacterial infections. Brucella VirB T4SS is a key virulence factor that plays important roles in mediating intracellular survival and manipulating host immune response to infection. In this review, we discuss the roles of Brucella VirB T4SS and 15 effectors that are proposed to be crucial for Brucella pathogenesis. VirB T4SS regulates the inflammation response and manipulates vesicle trafficking inside host cells. VirB T4SS also plays crucial roles in the inhibition of the host immune response and intracellular survival during infection. Here, we list the key molecular events in the intracellular life cycle of Brucella that are potentially targeted by the VirB T4SS effectors. Elucidating the functions of these effectors will help clarify the molecular role of T4SS during infection. Furthermore, studying the effectors secreted by Brucella spp. might provide insights into the mechanisms used by the bacteria to hijack the host signaling pathways and aid in the development of better vaccines and therapies against brucellosis.

  20. Thermostable cross-protective subunit vaccine against Brucella species.

    Science.gov (United States)

    Cherwonogrodzky, John W; Barabé, Nicole D; Grigat, Michelle L; Lee, William E; Poirier, Robert T; Jager, Scott J; Berger, Bradley J

    2014-12-01

    A subunit vaccine candidate was produced from Brucella suis 145 (biovar 4; expressing both the A antigen of Brucella abortus and the M antigen of Brucella melitensis). The preparation consisted mostly of polysaccharide (PS; >90% [wt/wt]; both cell-associated PS and exo-PS were combined) and a small amount of protein (1 to 3%) with no apparent nucleic acids. Vaccinated mice were protected (these had a statistically significant reduction in bacterial colonization compared to that of unvaccinated controls) when challenged with representative strains of three Brucella species most pathogenic for humans, i.e., B. abortus, B. melitensis, and B. suis. As little as 1 ng of the vaccine, without added adjuvant, protected mice against B. suis 145 infection (5 × 10(5) CFU), and a single injection of 1 μg of this subunit vaccine protected mice from B. suis 145 challenge for at least 14 months. A single immunization induced a serum IgG response to Brucella antigens that remained elevated for up to 9 weeks. The use of heat (i.e., boiling-water bath, autoclaving) in the vaccine preparation showed that it was thermostable. This method also ensured safety and security. The vaccine produced was immunogenic and highly protective against multiple strains of Brucella and represents a promising candidate for further evaluation. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  1. Contamination of bovine, sheep and goat meat with Brucella spp.

    Directory of Open Access Journals (Sweden)

    Francesco Casalinuovo

    2016-06-01

    Full Text Available A study was conducted in order to evaluate the contamination by Brucella spp. of meat from animals slaughtered because they had resulted positive for brucellosis at some time during their life. After slaughter and before delivery to market outlets, swab samples were taken from 307 carcasses of infected animals: 40 cattle, 60 sheep and 207 goats. The swabs were subsequently analysed by means of polymerase chain reaction (PCR tests. In addition, bacteriological tests were carried out on the lymph nodes and internal organs of the same animals. Brucella spp. was detected by means of PCR in 25/307 carcasses (8%: 1 bovine (2.5%, 9 sheep (15% and 15 goats (7.2% and was isolated by means of a cultural method in 136/307 carcasses (44%. Moreover, additional analysis, performed on lymph nodes from the same carcasses that had proved positive by PCR, allowed highlighting type 3 Brucella abortus in the bovine carcass and type 3 Brucella melitensis in the sheep and goat carcasses. The study shows that cattle, sheep and goats meat of animals slaughtered because they had tested positive for brucellosis may be contaminated by Brucella spp. As this could constitute a real risk of transmission to both butchery personnel and consumers, the meat of animals infected by Brucella spp. should be analysed before being marketed. In this respect, PCR technique performed on swabs proved to be more useful, practical and faster than the traditional bacteriological method.

  2. Antibody Reactivity to Omp31 from Brucella melitensis in Human and Animal Infections by Smooth and Rough Brucellae

    Science.gov (United States)

    Cassataro, Juliana; Pasquevich, Karina; Bruno, Laura; Wallach, Jorge C.; Fossati, Carlos A.; Baldi, Pablo C.

    2004-01-01

    Group 3 of outer membrane proteins (OMPs) of Brucella includes Omp25 and Omp31, which share 34% identity. Omp25 is highly conserved in Brucella species, and Omp31 is present in all Brucella species, except Brucella abortus. Antibodies to Brucella melitensis Omp31 have been sought only in infected sheep, and Western blotting of sera from infected sheep did not reveal anti-Omp31 reactivity. We obtained recombinant purified Omp31 (B. melitensis) and tested its recognition by sera from humans and animals suffering from brucellosis by an indirect enzyme-linked immunosorbent assay (ELISA). Serum samples from 74 patients, 57 sheep, and 47 dogs were analyzed; brucellosis was confirmed by bacteriological isolation in all ovine and canine cases and 31 human cases of brucellosis. Thirty-five patients (47%) were positive for antibodies to Omp31, including seven cases of Brucella suis infection, two cases of B. abortus infection, and three cases of B. melitensis infection. Of 39 sheep naturally infected with B. melitensis (biovars 1 and 3), 23 (59%) were positive for antibodies to Omp31. Anti-Omp31 antibodies were also detected in 12 of 18 rams (67%) in which Brucella ovis was isolated from semen. Antibodies to Omp31 were also found in 41 (87%) of the 47 dogs, including 13 with recent infection. These results suggest that an indirect ELISA using recombinant purified Omp31 from B. melitensis would be of limited value for the diagnosis of human and animal brucellosis. Nevertheless, the potential usefulness of this antigen in combination with other recombinant proteins from Brucella should not be dismissed.   PMID:14715555

  3. Oxidative metabolic profiles of Brucella strains isolated from marine mammals: contribution to their species classification.

    Science.gov (United States)

    Jacques, Isabelle; Grayon, Maggy; Verger, Jean-Michel

    2007-05-01

    Since the 1990s, Brucella strains not matching the characteristics of any of the six conventional species have been isolated worldwide from marine mammals. In this study, 31 Brucella strains isolated from various marine mammals were examined for their oxidative metabolic pattern on 12 amino-acid and carbohydrate substrates. Three main oxidative profiles different from those of the Brucella terrestrial mammal strains were identified for the marine mammal strains: one gathering strains isolated from pinnipeds and two gathering strains from cetaceans. Thus, both oxidative metabolism results and previous molecular studies are in agreement with the proposal of two new Brucella species, Brucella pinnipediae and Brucella cetaceae, to classify the Brucella strains isolated from marine mammals, and are also in accordance with a classification of species of the Brucella genus based on host preference.

  4. A review of Brucella infection in marine mammals, with special emphasis on Brucella pinnipedialis in the hooded seal (Cystophora cristata

    Directory of Open Access Journals (Sweden)

    Nymo Ingebjørg H

    2011-08-01

    Full Text Available Abstract Brucella spp. were isolated from marine mammals for the first time in 1994. Two novel species were later included in the genus; Brucella ceti and Brucella pinnipedialis, with cetaceans and seals as their preferred hosts, respectively. Brucella spp. have since been isolated from a variety of marine mammals. Pathological changes, including lesions of the reproductive organs and associated abortions, have only been registered in cetaceans. The zoonotic potential differs among the marine mammal Brucella strains. Many techniques, both classical typing and molecular microbiology, have been utilised for characterisation of the marine mammal Brucella spp. and the change from the band-based approaches to the sequence-based approaches has greatly increased our knowledge about these strains. Several clusters have been identified within the B. ceti and B. pinnipedialis species, and multiple studies have shown that the hooded seal isolates differ from other pinniped isolates. We describe how different molecular methods have contributed to species identification and differentiation of B. ceti and B. pinnipedialis, with special emphasis on the hooded seal isolates. We further discuss the potential role of B. pinnipedialis for the declining Northwest Atlantic hooded seal population.

  5. A review of Brucella infection in marine mammals, with special emphasis on Brucella pinnipedialis in the hooded seal (Cystophora cristata)

    Science.gov (United States)

    2011-01-01

    Brucella spp. were isolated from marine mammals for the first time in 1994. Two novel species were later included in the genus; Brucella ceti and Brucella pinnipedialis, with cetaceans and seals as their preferred hosts, respectively. Brucella spp. have since been isolated from a variety of marine mammals. Pathological changes, including lesions of the reproductive organs and associated abortions, have only been registered in cetaceans. The zoonotic potential differs among the marine mammal Brucella strains. Many techniques, both classical typing and molecular microbiology, have been utilised for characterisation of the marine mammal Brucella spp. and the change from the band-based approaches to the sequence-based approaches has greatly increased our knowledge about these strains. Several clusters have been identified within the B. ceti and B. pinnipedialis species, and multiple studies have shown that the hooded seal isolates differ from other pinniped isolates. We describe how different molecular methods have contributed to species identification and differentiation of B. ceti and B. pinnipedialis, with special emphasis on the hooded seal isolates. We further discuss the potential role of B. pinnipedialis for the declining Northwest Atlantic hooded seal population. PMID:21819589

  6. Brucella microti: the genome sequence of an emerging pathogen.

    Science.gov (United States)

    Audic, Stéphane; Lescot, Magali; Claverie, Jean-Michel; Scholz, Holger C

    2009-08-04

    Using a combination of pyrosequencing and conventional Sanger sequencing, the complete genome sequence of the recently described novel Brucella species, Brucella microti, was determined. B. microti is a member of the genus Brucella within the Alphaproteobacteria, which consists of medically important highly pathogenic facultative intracellular bacteria. In contrast to all other Brucella species, B. microti is a fast growing and biochemically very active microorganism with a phenotype more similar to that of Ochrobactrum, a facultative human pathogen. The atypical phenotype of B. microti prompted us to look for genomic differences compared to other Brucella species and to look for similarities with Ochrobactrum. The genome is composed of two circular chromosomes of 2,117,050 and 1,220,319 base pairs. Unexpectedly, we found that the genome sequence of B. microti is almost identical to that of Brucella suis 1330 with an overall sequence identity of 99.84% in aligned regions. The most significant structural difference between the two genomes is a bacteriophage-related 11,742 base pairs insert only present in B. microti. However, this insert is unlikely to have any phenotypical consequence. Only four protein coding genes are shared between B. microti and Ochrobactrum anthropi but impaired in other sequenced Brucella. The most noticeable difference between B. microti and other Brucella species was found in the sequence of the 23S ribosomal RNA gene. This unusual variation could have pleiotropic effects and explain the fast growth of B. microti. Contrary to expectations from the phenotypic analysis, the genome sequence of B. microti is highly similar to that of known Brucella species, and is remotely related to the one of O. anthropi. How the few differences in gene content between B. microti and B. suis 1330 could result in vastly different phenotypes remains to be elucidated. This unexpected finding will complicate the task of identifying virulence determinants in the

  7. Brucella microti: the genome sequence of an emerging pathogen

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    Scholz Holger C

    2009-08-01

    Full Text Available Abstract Background Using a combination of pyrosequencing and conventional Sanger sequencing, the complete genome sequence of the recently described novel Brucella species, Brucella microti, was determined. B. microti is a member of the genus Brucella within the Alphaproteobacteria, which consists of medically important highly pathogenic facultative intracellular bacteria. In contrast to all other Brucella species, B. microti is a fast growing and biochemically very active microorganism with a phenotype more similar to that of Ochrobactrum, a facultative human pathogen. The atypical phenotype of B. microti prompted us to look for genomic differences compared to other Brucella species and to look for similarities with Ochrobactrum. Results The genome is composed of two circular chromosomes of 2,117,050 and 1,220,319 base pairs. Unexpectedly, we found that the genome sequence of B. microti is almost identical to that of Brucella suis 1330 with an overall sequence identity of 99.84% in aligned regions. The most significant structural difference between the two genomes is a bacteriophage-related 11,742 base pairs insert only present in B. microti. However, this insert is unlikely to have any phenotypical consequence. Only four protein coding genes are shared between B. microti and Ochrobactrum anthropi but impaired in other sequenced Brucella. The most noticeable difference between B. microti and other Brucella species was found in the sequence of the 23S ribosomal RNA gene. This unusual variation could have pleiotropic effects and explain the fast growth of B. microti. Conclusion Contrary to expectations from the phenotypic analysis, the genome sequence of B. microti is highly similar to that of known Brucella species, and is remotely related to the one of O. anthropi. How the few differences in gene content between B. microti and B. suis 1330 could result in vastly different phenotypes remains to be elucidated. This unexpected finding will

  8. Brucella abortus RB51 and Hot Saline Extract from Brucella ovis as Antigens in a Complement Fixation Test Used To Detect Sheep Vaccinated with Brucella abortus RB51

    OpenAIRE

    Adone, Rosanna; Ciuchini, Franco

    2001-01-01

    The efficacy of Brucella abortus RB51 and hot saline extract (HSE) from Brucella ovis as antigens in complement fixation (CF) tests was comparatively evaluated in detecting immune responses of sheep vaccinated with B. abortus strain RB51. For this study, four 5-month-old sheep were vaccinated subcutaneously with 5 × 109 CFU of RB51, and two sheep received saline. Serum samples collected at different times after vaccination were tested for the presence of antibodies to RB51 by a CF test with R...

  9. Characterisation of the genetic diversity of Brucella by multilocus sequencing

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    MacMillan Alastair P

    2007-04-01

    Full Text Available Abstract Background Brucella species include economically important zoonotic pathogens that can infect a wide range of animals. There are currently six classically recognised species of Brucella although, as yet unnamed, isolates from various marine mammal species have been reported. In order to investigate genetic relationships within the group and identify potential diagnostic markers we have sequenced multiple genetic loci from a large sample of Brucella isolates representing the known diversity of the genus. Results Nine discrete genomic loci corresponding to 4,396 bp of sequence were examined from 160 Brucella isolates. By assigning each distinct allele at a locus an arbitrary numerical designation the population was found to represent 27 distinct sequence types (STs. Diversity at each locus ranged from 1.03–2.45% while overall genetic diversity equated to 1.5%. Most loci examined represent housekeeping gene loci and, in all but one case, the ratio of non-synonymous to synonymous change was substantially Brucella species, B. abortus, B. melitensis, B. ovis and B. neotomae correspond to well-separated clusters. With the exception of biovar 5, B. suis isolates cluster together, although they form a more diverse group than other classical species with a number of distinct STs corresponding to the remaining four biovars. B. canis isolates are located on the same branch very closely related to, but distinguishable from, B. suis biovar 3 and 4 isolates. Marine mammal isolates represent a distinct, though rather weakly supported, cluster within which individual STs display one of three clear host preferences. Conclusion The sequence database provides a powerful dataset for addressing ongoing controversies in Brucella taxonomy and a tool for unambiguously placing atypical, phenotypically discordant or newly emerging Brucella isolates. Furthermore, by using the phylogenetic backbone described here, robust and rationally selected markers for use in

  10. In vitro susceptibilities of Brucella melitensis isolates to eleven antibiotics

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    Loukaides Feidias

    2006-10-01

    Full Text Available Abstract Background Brucellosis is an endemic disease present in many countries worldwide, but it is rare in Europe and North America. Nevertheless brucella is included in the bacteria potentially used for bioterrorism. The aim of this study was the investigation of the antibiotic susceptibility profile of brucella isolates from areas of the eastern Mediterranean where it has been endemic. Methods The susceptibilities of 74 Brucella melitensis isolates derived from clinical samples (57 and animal products (17 were tested in vitro. The strains originate from Crete (59, Cyprus (10, and Syria (5. MICs of tetracycline, rifampicin, streptomycin, gentamicin, norfloxacin, ciprofloxacin, levofloxacin, trimethoprim/sulfamethoxazole, ampicillin, amoxicillin/clavulanic acid, and erythromycin were detected by E-test method. The NCCLS criteria for slow growing bacteria were considered to interpret the results. Results All the isolates were susceptible to tetracycline, streptomycin, gentamicin, ciprofloxacin, norfloxacin, and levofloxacin. Two isolates presented reduced susceptibility to rifampicin (MIC value: 1.5 mg/l and eight to SXT (MIC values: 0.75–1.5 mg/l. Erythromycin had the highest (4 mg/l MIC90value and both norfloxacin and erythromycin the highest (1.5 mg/l MIC50 value. Conclusion Brucella isolates remain susceptible in vitro to most antibiotics used for treatment of brucellosis. The establishment of a standardized antibiotic susceptibility method for Brucella spp would be useful for resistance determination in these bacteria and possible evaluation of bioterorism risks.

  11. Isolation of a novel 'atypical' Brucella strain from a bluespotted ribbontail ray (Taeniura lymma).

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    Eisenberg, Tobias; Riße, Karin; Schauerte, Nicole; Geiger, Christina; Blom, Jochen; Scholz, Holger C

    2017-02-01

    A pleomorphic Gram-negative, motile coccobacillus was isolated from the gills of a wild-caught bluespotted ribbontail ray after its sudden death during quarantine. Strain 141012304 was observed to grow aerobically, to be clearly positive for cytochrome oxidase, catalase, urease and was initially identified as "Brucella melitensis" or "Ochrobactrum anthropi" by Matrix-assisted laser desorption/ionization-time of flight mass spectrometry and VITEK2-compact ® , respectively. Affiliation to the genus Brucella was confirmed by bcsp31 and IS711 PCR as well as by Brucella species-specific multiplex PCR, therein displaying a characteristic banding pattern recently described for Brucella strains obtained from amphibian hosts. Likewise, based on recA sequencing, strain 141012304 was found to form a separate lineage, within the so called 'atypical' Brucella, consisting of genetically more distantly related strains. The closest similarity was detected to brucellae, which have recently been isolated from edible bull frogs. Subsequent next generation genome sequencing and phylogenetic analysis confirmed that the ray strain represents a novel Brucella lineage within the atypical group of Brucella and in vicinity to Brucella inopinata and Brucella strain BO2, both isolated from human patients. This is the first report of a natural Brucella infection in a saltwater fish extending the host range of this medically important genus.

  12. Identification and typing of Brucella spp. in stranded harbour porpoises (Phocoena phocoena) on the Dutch coast.

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    Maio, Elisa; Begeman, Lineke; Bisselink, Yvette; van Tulden, Peter; Wiersma, Lidewij; Hiemstra, Sjoukje; Ruuls, Robin; Gröne, Andrea; Roest, Hendrik-Ido-Jan; Willemsen, Peter; van der Giessen, Joke

    2014-09-17

    The presence of Brucella (B.) spp. in harbour porpoises stranded between 2008 and 2011 along the Dutch coast was studied. A selection of 265 tissue samples from 112 animals was analysed using conventional and molecular methods. In total, 4.5% (5/112) of the animals corresponding with 2.3% (6/265) Brucella positive tissue samples were Brucella positive by culture and these were all confirmed by real-time polymerase chain reaction (real-time PCR) based on the insertion element 711 (IS711). In addition, two more Brucella-positive tissue samples from two animals collected in 2011 were identified using real-time PCR resulting in an overall Brucella prevalence of 6.3% (7/112 animals). Brucella spp. were obtained from lungs (n=3), pulmonary lymph node (n=3) and lungworms (n=2). Multi Locus Variable Number of Tandem Repeats (VNTR) Analysis (MLVA) typing based on the MLVA-16 showed that the Brucella isolates were B. ceti. Additional in silico Multi Locus Sequence typing (MLST) after whole genome sequencing of the 6 Brucella isolates confirmed B. ceti ST 23. According to the Brucella 2010 MLVA database, the isolated Brucella strains encountered were of five genotypes, in two distinct subclusters divided in two different time periods of harbour porpoises collection. This study is the first population based analyses for Brucella spp. infections in cetaceans stranded along the Dutch coast. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  13. Importance of Lipopolysaccharide and Cyclic β-1,2-Glucans in Brucella-Mammalian Infections

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    Andreas F. Haag

    2010-01-01

    Full Text Available Brucella species are the causative agents of one of the most prevalent zoonotic diseases: brucellosis. Infections by Brucella species cause major economic losses in agriculture, leading to abortions in infected animals and resulting in a severe, although rarely lethal, debilitating disease in humans. Brucella species persist as intracellular pathogens that manage to effectively evade recognition by the host's immune system. Sugar-modified components in the Brucella cell envelope play an important role in their host interaction. Brucella lipopolysaccharide (LPS, unlike Escherichia coli LPS, does not trigger the host's innate immune system. Brucella produces cyclic β-1,2-glucans, which are important for targeting them to their replicative niche in the endoplasmic reticulum within the host cell. This paper will focus on the role of LPS and cyclic β-1,2-glucans in Brucella-mammalian infections and discuss the use of mutants, within the biosynthesis pathway of these cell envelope structures, in vaccine development.

  14. Brucella infection inhibits macrophages apoptosis via Nedd4-dependent degradation of calpain2.

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    Cui, Guimei; Wei, Pan; Zhao, Yuxi; Guan, Zhenhong; Yang, Li; Sun, Wanchun; Wang, Shuangxi; Peng, Qisheng

    2014-11-07

    The calcium-dependent protease calpain2 is involved in macrophages apoptosis. Brucella infection-induced up-regulation of intracellular calcium level is an essential factor for the intracellular survival of Brucella within macrophages. Here, we hypothesize that calcium-dependent E3 ubiquitin ligase Nedd4 ubiquitinates calpain2 and inhibits Brucella infection-induced macrophage apoptosis via degradation of calpain2.Our results reveal that Brucella infection induces increases in Nedd4 activity in an intracellular calcium dependent manner. Furthermore, Brucella infection-induced degradation of calpain2 is mediated by Nedd4 ubiquitination of calpain2. Brucella infection-induced calpain2 degradation inhibited macrophages apoptosis. Treatment of Brucella infected macrophages with calcium chelator BAPTA or Nedd4 knock-down decreased Nedd4 activity, prevented calpain2 degradation, and resulted in macrophages apoptosis. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. Duplex PCR for differentiation of the vaccine strain Brucella suis S2 and B. suis biovar 1 from other strains of Brucella spp.

    Science.gov (United States)

    Nan, Wenlong; Tan, Pengfei; Wang, Yong; Xu, Zouliang; Mao, Kairong; Peng, Daxin; Chen, Yiping

    2014-09-01

    Immunisation with attenuated Brucella spp. vaccines prevents brucellosis, but may also interfere with diagnosis. In this study, a duplex PCR was developed to distinguish Brucella suis vaccine strain S2 from field strains of B. suis biovar 1 and other Brucella spp. The PCR detected 60 fg genomic DNA of B. suis S2 or biovar 1 field strains and was able to distinguish B. suis S2 and wild-type strains of B. suis biovar 1 among 76 field isolates representing all the common species and biovars, as well as four vaccine strains, of Brucella. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Development and trial of vaccines against Brucella

    Science.gov (United States)

    Lalsiamthara, Jonathan

    2017-01-01

    The search for ideal brucellosis vaccines remains active today. Currently, no licensed human or canine anti-brucellosis vaccines are available. In bovines, the most successful vaccine (S19) is only used in calves, as adult vaccination results in orchitis in male, prolonged infection, and possible abortion complications in pregnant female cattle. Another widely deployed vaccine (RB51) has a low protective efficacy. An ideal vaccine should exhibit a safe profile as well as enhance protective efficacy. However, currently available vaccines exhibit one or more major drawbacks. Smooth live attenuated vaccines suffer shortcomings such as residual virulence and serodiagnostic interference. Inactivated vaccines, in general, confer relatively low levels of protection. Recent developments to improve brucellosis vaccines include generation of knockout mutants by targeting genes involved in metabolism, virulence, and the lipopolysaccharide synthesis pathway, as well as generation of DNA vaccines, mucosal vaccines, and live vectored vaccines, have all produced varying degrees of success. Herein, we briefly review the bacteriology, pathogenesis, immunological implications, candidate vaccines, vaccinations, and models related to Brucella. PMID:28859268

  17. Brucella ceti and Brucellosis in Cetaceans

    Science.gov (United States)

    Guzmán-Verri, Caterina; González-Barrientos, Rocío; Hernández-Mora, Gabriela; Morales, Juan-Alberto; Baquero-Calvo, Elías; Chaves-Olarte, Esteban; Moreno, Edgardo

    2012-01-01

    Since the first case of brucellosis detected in a dolphin aborted fetus, an increasing number of Brucella ceti isolates has been reported in members of the two suborders of cetaceans: Mysticeti and Odontoceti. Serological surveys have shown that cetacean brucellosis may be distributed worldwide in the oceans. Although all B. ceti isolates have been included within the same species, three different groups have been recognized according to their preferred host, bacteriological properties, and distinct genetic traits: B. ceti dolphin type, B. ceti porpoise type, and B. ceti human type. It seems that B. ceti porpoise type is more closely related to B. ceti human isolates and B. pinnipedialis group, while B. ceti dolphin type seems ancestral to them. Based on comparative phylogenetic analysis, it is feasible that the B. ceti ancestor radiated in a terrestrial artiodactyl host close to the Raoellidae family about 58 million years ago. The more likely mode of transmission of B. ceti seems to be through sexual intercourse, maternal feeding, aborted fetuses, placental tissues, vertical transmission from mother to the fetus or through fish or helminth reservoirs. The B. ceti dolphin and porpoise types seem to display variable virulence in land animal models and low infectivity for humans. However, brucellosis in some dolphins and porpoises has been demonstrated to be a severe chronic disease, displaying significant clinical and pathological signs related to abortions, male infertility, neurobrucellosis, cardiopathies, bone and skin lesions, strandings, and death. PMID:22919595

  18. Multivalent Fusion DNA Vaccine against Brucella abortus

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    Gómez, Leonardo; Llanos, Javiera; Escalona, Emilia; Sáez, Darwin; Álvarez, Francisco; Molina, Raúl; Flores, Manuel

    2017-01-01

    As an alternative brucellosis prevention method, we evaluated the immunogenicity induced by new multivalent DNA vaccines in BALB/c mice. We constructed the vaccines by fusion of BAB1_0273 and/or BAB1_0278 open reading frames (ORFs) from genomic island 3 (GI-3) and the Brucella abortus 2308 sodC gene with a link based on prolines and alanines (pV273-sod, pV278-sod, and pV273-278-sod, resp.). Results show that immunization with all tested multivalent DNA vaccines induced a specific humoral and cellular immune response. These novel multivalent vaccines significantly increased the production of IgM, IgG, and IgG2a antibodies as well as IFN-γ levels and the lymphoproliferative response of splenocytes. Although immunization with these multivalent vaccines induced a typical T-helper 1- (Th1-) dominated immune response, such immunogenicity conferred low protection levels in mice challenged with the B. abortus 2308 pathogenic strain. Our results demonstrated that the expression of BAB1_0273 and/or BABl_0278 antigens conjugated to SOD protein can polarize mice immunity to a Th1-type phenotype, conferring low levels of protection. PMID:29082252

  19. Multivalent Fusion DNA Vaccine against Brucella abortus

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    Leonardo Gómez

    2017-01-01

    Full Text Available As an alternative brucellosis prevention method, we evaluated the immunogenicity induced by new multivalent DNA vaccines in BALB/c mice. We constructed the vaccines by fusion of BAB1_0273 and/or BAB1_0278 open reading frames (ORFs from genomic island 3 (GI-3 and the Brucella abortus 2308 sodC gene with a link based on prolines and alanines (pV273-sod, pV278-sod, and pV273-278-sod, resp.. Results show that immunization with all tested multivalent DNA vaccines induced a specific humoral and cellular immune response. These novel multivalent vaccines significantly increased the production of IgM, IgG, and IgG2a antibodies as well as IFN-γ levels and the lymphoproliferative response of splenocytes. Although immunization with these multivalent vaccines induced a typical T-helper 1- (Th1- dominated immune response, such immunogenicity conferred low protection levels in mice challenged with the B. abortus 2308 pathogenic strain. Our results demonstrated that the expression of BAB1_0273 and/or BABl_0278 antigens conjugated to SOD protein can polarize mice immunity to a Th1-type phenotype, conferring low levels of protection.

  20. A Study of Brucella Infection in Humans

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    Hasanjani Roushan Mohammad Reza

    2014-07-01

    Full Text Available Objective: Brucellosis is the most usual zoonotic disease around the world especially in the Middle East, Mediterranean and Indian sub-continent areas. This bacterium has ten species that Brucella melitensis among them recognized as the most important cause of human brucellosis. This infection transfer ways to human include of wounds, bacteria inhalation and consumption of septic dairy such as raw milk, cream and butter. Brucellosis as a systemic disease can involve more organs of patients that have symptoms such as fever, night sweating, and backache. This infection can be divided as acute, sub-acute and chronic forms according to the manner of clinical presentation. Materials and Methods: This research is a review study and conducted by reviewing of the literature, which is related to this issue and also visiting, PubMed, and other linked websites. Results: In human brucellosis domestic animals are the main natural reservoir of infection. Whenever incidence rate of this infection in domestic and wild animals is reduced on the other hand incidence rate in human also will reduce. Conclusion: Blood cultures, serological tests and molecular tests are common laboratory methods of this infection. Diminution of relapse and therapeutic failure rates are as most important aim, which is researcher’s regards.

  1. Use of polymerase chain reaction to identify Brucella abortus strain RB51 among Brucella field isolates from cattle in Italy.

    Science.gov (United States)

    Adone, R; Ciuchini, F; La Rosa, G; Marianelli, C; Muscillo, M

    2001-03-01

    Brucella abortus strain RB51, a rough mutant of the B. abortus 2308 virulent strain, was recently approved in the United States as the official vaccine for brucellosis in cattle. Following recent evidence of unauthorized use of RB51 vaccine in Italy, where the use of vaccines for brucellosis is no longer allowed, the suitability of an RB51-specific polymerase chain reaction assay for identifying the RB51 strain among Brucella field isolates from cattle in Italy was investigated. The oligonucleotide primers used in this study, belonging to a six-primer cocktail for Brucella species previously described by other authors, allowed the amplification of a 364-base pair (bp) fragment specific for RB51 and its parent strain 2308, and a 498-bp product specific for B. abortus. In addition, unresolved bands ranging from 600 to 700 bp were observed from RB51 strain. Brucella abortus biovars 1, 2 and 4 have only one specific sensitive 498-bp band. The B. abortus biovars 3, 5 and 6 did not give any signal. The 498-bp product from a reference Brucella strain was sequenced and submitted to EMBL with the accession number AJ271969 while the 364-bp fragment from RB51 strain was submitted to EMBL database with accession number AJ271968. The sequence studies confirmed the specificity of the detected fragments. No amplification was obtained by testing DNA from strains antigenically related to Brucella, such as Yersinia enterocolitica O:9, Escherichia coli O:157, Salmonella urbana and Pasteurella multocida. The results of this study indicate that this technique, in combination with specific serological tests, could be a useful diagnostic method to verify the use of RB51 vaccine and can contribute to the creation of a databank of circulating strains.

  2. Evaluation of a New and Rapid Serologic Test for Detecting Brucellosis: Brucella Coombs Gel Test.

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    Hanci, Hayrunisa; Igan, Hakan; Uyanik, Muhammet Hamidullah

    2017-01-01

    Many serological tests have been used for the diagnosis of human brucellosis. A new serological method is identified as Brucella Coombs gel test based on the principle of centrifugation gel system similar to the gel system used in blood group determination. In this system, if Brucella antibodies were present in the serum, antigen and antibody would remain as a pink complex on the gel. Otherwise, the pink Brucella antigens would precipitate at the bottom of the gel card system. In this study, we aimed to compare the Brucella Coombs gel test, a new, rapid screen and titration method for detection of non-agglutinating IgG with the Brucella Coombs test. For this study, a total of 88 serum samples were obtained from 45 healthy persons and 43 individuals who had clinical signs and symptoms of brucellosis. For each specimen, Rose Bengal test, standard agglutination test, Coombs test and Brucella Coombs gel test were carried out. Sensitivity and specificity of Brucella Coombs gel test were found as 100.0 and 82.2%, respectively. Brucella Coombs gel test can be used as a screening test with high sensitivity. By the help of pink Brucella antigen precipitation, the tests' evaluation is simple and objective. In addition, determination of Brucella antibody by rapid titration offers another important advantage.

  3. A novel real-time PCR assay for specific detection of Brucella melitensis.

    Science.gov (United States)

    Kaden, Rene; Ferrari, Sevinc; Alm, Erik; Wahab, Tara

    2017-03-24

    Brucellosis is a zoonosis that occurs worldwide. The disease has been completely eradicated in livestock in Sweden in 1994, and all cases of confirmed human brucellosis are imported into Sweden from other countries. However, due to an increase in the number of refugees and asylum seekers from the middle-east to Sweden, there is a need to improve the current diagnostic methodology for Brucella melitensis. Whilst culture of Brucella species can be used as a diagnostic tool, real-time PCR approaches provide a much faster result. The aim of this study was to set up a species-specific real-time PCR for the detection of all biovars of Brucella melitensis, which could be used routinely in diagnostic laboratories. A Brucella melitensis real-time PCR assay was designed using all available genomes in the public database of Brucella (N = 96) including all complete genomes of Brucella melitensis (N = 17). The assay was validated with a collection of 37 Brucella species reference strains, 120 Brucella melitensis human clinical isolates, and 45 clinically relevant non-Brucella melitensis strains. In this study we developed a single real-time PCR for the specific detection of all biovars of Brucella melitensis. This new real-time PCR method shows a high specificity (100%) and a high sensitivity (1.25 GE/μl) and has been implemented in the laboratories of four governmental authorities across Sweden.

  4. Brucella Dysregulates Monocytes and Inhibits Macrophage Polarization through LC3-Dependent Autophagy

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    Yang Wang

    2017-06-01

    Full Text Available Brucellosis is caused by infection with Brucella species and exhibits diverse clinical manifestations in infected humans. Monocytes and macrophages are not only the first line of defense against Brucella infection but also a main reservoir for Brucella. In the present study, we examined the effects of Brucella infection on human peripheral monocytes and monocyte-derived polarized macrophages. We showed that Brucella infection led to an increase in the proportion of CD14++CD16− monocytes and the expression of the autophagy-related protein LC3B, and the effects of Brucella-induced monocytes are inhibited after 6 weeks of antibiotic treatment. Additionally, the production of IL-1β, IL-6, IL-10, and TNF-α from monocytes in patients with brucellosis was suppressed through the LC3-dependent autophagy pathway during Brucella infection. Moreover, Brucella infection inhibited macrophage polarization. Consistently, the addition of 3-MA, an inhibitor of LC3-related autophagy, partially restored macrophage polarization. Intriguingly, we also found that the upregulation of LC3B expression by rapamycin and heat-killed Brucella in vitro inhibits M2 macrophage polarization, which can be reversed partially by 3-MA. Taken together, these findings reveal that Brucella dysregulates monocyte and macrophage polarization through LC3-dependent autophagy. Thus, targeting this pathway may lead to the development of new therapeutics against Brucellosis.

  5. Brucella Infection in Asian Sea Otters (Enhydra lutris lutris) on Bering Island, Russia.

    Science.gov (United States)

    Burgess, Tristan L; Johnson, Christine Kreuder; Burdin, Alexander; Gill, Verena A; Doroff, Angela M; Tuomi, Pamela; Smith, Woutrina A; Goldstein, Tracey

    2017-10-01

    Infection with Brucella spp., long known as a cause of abortion, infertility, and reproductive loss in domestic livestock, has increasingly been documented in marine mammals over the past two decades. We report molecular evidence of Brucella infection in Asian sea otters (Enhydra lutris lutris). Brucella DNA was detected in 3 of 78 (4%) rectal swab samples collected between 2004 and 2006 on Bering Island, Russia. These 78 animals had previously been documented to have a Brucella seroprevalence of 28%, markedly higher than the prevalence documented in sea otters (Enhydra lutris) in North America. All of the DNA sequences amplified were identical to one or more previously isolated Brucella spp. including strains from both terrestrial and marine hosts. Phylogenetic analysis of this sequence suggested that one animal was shedding Brucella spp. DNA with a sequence matching a Brucella abortus strain, whereas two animals yielded a sequence matching a group of strains including isolates classified as Brucella pinnipedialis and Brucella melitensis. Our results highlight the diversity of Brucella spp. within a single sea otter population.

  6. Analyses of Brucella Pathogenesis, Host Immunity, and Vaccine Targets using Systems Biology and Bioinformatics

    Science.gov (United States)

    He, Yongqun

    2011-01-01

    Brucella is a Gram-negative, facultative intracellular bacterium that causes zoonotic brucellosis in humans and various animals. Out of 10 classified Brucella species, B. melitensis, B. abortus, B. suis, and B. canis are pathogenic to humans. In the past decade, the mechanisms of Brucella pathogenesis and host immunity have been extensively investigated using the cutting edge systems biology and bioinformatics approaches. This article provides a comprehensive review of the applications of Omics (including genomics, transcriptomics, and proteomics) and bioinformatics technologies for the analysis of Brucella pathogenesis, host immune responses, and vaccine targets. Based on more than 30 sequenced Brucella genomes, comparative genomics is able to identify gene variations among Brucella strains that help to explain host specificity and virulence differences among Brucella species. Diverse transcriptomics and proteomics gene expression studies have been conducted to analyze gene expression profiles of wild type Brucella strains and mutants under different laboratory conditions. High throughput Omics analyses of host responses to infections with virulent or attenuated Brucella strains have been focused on responses by mouse and cattle macrophages, bovine trophoblastic cells, mouse and boar splenocytes, and ram buffy coat. Differential serum responses in humans and rams to Brucella infections have been analyzed using high throughput serum antibody screening technology. The Vaxign reverse vaccinology has been used to predict many Brucella vaccine targets. More than 180 Brucella virulence factors and their gene interaction networks have been identified using advanced literature mining methods. The recent development of community-based Vaccine Ontology and Brucellosis Ontology provides an efficient way for Brucella data integration, exchange, and computer-assisted automated reasoning. PMID:22919594

  7. Analysis of ten Brucella genomes reveals evidence for horizontal gene transfer despite a preferred intracellular lifestyle.

    Science.gov (United States)

    Wattam, Alice R; Williams, Kelly P; Snyder, Eric E; Almeida, Nalvo F; Shukla, Maulik; Dickerman, A W; Crasta, O R; Kenyon, R; Lu, J; Shallom, J M; Yoo, H; Ficht, T A; Tsolis, R M; Munk, C; Tapia, R; Han, C S; Detter, J C; Bruce, D; Brettin, T S; Sobral, Bruno W; Boyle, Stephen M; Setubal, João C

    2009-06-01

    The facultative intracellular bacterial pathogen Brucella infects a wide range of warm-blooded land and marine vertebrates and causes brucellosis. Currently, there are nine recognized Brucella species based on host preferences and phenotypic differences. The availability of 10 different genomes consisting of two chromosomes and representing six of the species allowed for a detailed comparison among themselves and relatives in the order Rhizobiales. Phylogenomic analysis of ortholog families shows limited divergence but distinct radiations, producing four clades as follows: Brucella abortus-Brucella melitensis, Brucella suis-Brucella canis, Brucella ovis, and Brucella ceti. In addition, Brucella phylogeny does not appear to reflect the phylogeny of Brucella species' preferred hosts. About 4.6% of protein-coding genes seem to be pseudogenes, which is a relatively large fraction. Only B. suis 1330 appears to have an intact beta-ketoadipate pathway, responsible for utilization of plant-derived compounds. In contrast, this pathway in the other species is highly pseudogenized and consistent with the "domino theory" of gene death. There are distinct shared anomalous regions (SARs) found in both chromosomes as the result of horizontal gene transfer unique to Brucella and not shared with its closest relative Ochrobactrum, a soil bacterium, suggesting their acquisition occurred in spite of a predominantly intracellular lifestyle. In particular, SAR 2-5 appears to have been acquired by Brucella after it became intracellular. The SARs contain many genes, including those involved in O-polysaccharide synthesis and type IV secretion, which if mutated or absent significantly affect the ability of Brucella to survive intracellularly in the infected host.

  8. Brucella suis infection in domestic pigs in Sardinia (Italy).

    Science.gov (United States)

    Pilo, C; Tedde, M T; Orrù, G; Addis, G; Liciardi, M

    2015-07-01

    During a 4-year (2007-2010) survey, the presence of Brucella suis infection in domestic pigs in Sardinia was investigated. Serum samples were collected from breeding pigs located on 108 commercial farms with documented reproductive problems and analysed using the Rose Bengal (RBT) and complement fixation (CFT) tests for screening and confirmation of Brucella, respectively. Of the 1251 serum samples analysed by RBT, 406 sera, originating from 36 farms, were positive for B. suis. CFT was positive in 292/748 sera analysed, confirming positivity in all 36 pig herds. Pigs with international complement fixation test units per ml (ICFTU/ml) values ⩾160 were slaughtered, and their organs collected for bacteriological examination and testing by polymerase chain reaction (PCR). Brucella spp. strains were isolated in culture from 13/502 organs analysed, and subsequently identified as B. suis biovar 2. PCR detected positivity to Brucella spp. in 19/285 organs analysed. These results confirm the presence and emergence of B. suis infection in domestic pigs in Sardinia.

  9. [Brucella orchitis: A retrospective study of 69 cases].

    Science.gov (United States)

    Wang, Wen-qing; Guo, Zheng-yin

    2016-01-01

    To investigate the epidemiological characteristics, clinical manifestations, diagnosis, and treatment of Brucella orchitis, so as to provide reliable evidence for the prevention and treatment of the disease. We conducted retrospective statistical analyses on the medical records of 48 outpatients and 21 inpatients with Brucella orchitis. Brucella orchitis was diagnosed in 6.67% of the male patients with brucellosis (69/1 034). The disease exhibited typical epidemiological features, with a higher incidence rate among those in frequent contact with sheep and elderly people, in the period from April to July, and in the areas with sheep husbandry. All the Brucella orchitis patients had such local symptoms as testicular pain and swelling, more frequently involving both testes, and other most common symptoms included fever, chills, sweating, and painful joints. Based on IIEF-5, 45 of the patients suffered from severe erectile dysfunction, with their reproductive function temporarily affected in the course of the disease. Misdiagnosis easily occurred in the early stage of the disease. Therapeutic options mainly included doxycycline hydrochloride and rifampicin, administered orally or intravenously, which could effect a cure, though relapse might occur in some cases. Bru- cella orchitis has distinct epidemiological characteristics, with clinical manifestations of testicular pain and swelling. Though a transient disease, it affects the reproductive function of the patient before cured. It can be treated by combined oral and intravenous medication, with painkillers or ice bags for testicular pain and swelling.

  10. Prevalence of Brucella Antibodies in Migratory Fulani Cattle Herds ...

    African Journals Online (AJOL)

    Brucellosis is a major cause of economic losses such as abortion, infertility, low conception rate and low survival rate of neonates in the livestock industry and zoonoses of great public health significance. The prevalence of Brucella antibodies in migratory Fulani cattle in Kaduna State was determined using the Milk Ring ...

  11. Testing for Antibodies to Brucella abortus in Milk From Consumers ...

    African Journals Online (AJOL)

    Over 85% of all milk sales on Kenya pass through informal channels. The extent of the risk posed by the sale of this raw milk to human health in respect to brucellosis is unknown. This paper presents the results of a study on the occurrence of antibodies to Brucella abortus in milk from households consuming raw ...

  12. Seroprevalence of Brucella abortus in buffaloes and wildebeests in ...

    African Journals Online (AJOL)

    A sero-survey was conducted in buffalo and wildebeests in Ngorongoro Crater and Serengeti National Park (SNP) collectively known as Serengeti ecosystem to establish the level of exposure to Brucella arbortus. Rose Bengal Plate Agglutination test and Competitive ELISA were used serially in the analysis of 205 serum ...

  13. Molecular detection of Brucella spp. from broth culture of clinical ...

    African Journals Online (AJOL)

    PCR was employed to detect Brucella spp. from broth cultures of clinical samples using a group specific primer based on IS6501 sequence. The sensitivity and specificity of this assay was confirmed by Southern hybridization analysis using a digoxigenin-labeled DNA probe while reproducibility of the analysis was ...

  14. Brucella abortus agglutinins in dogs in Zaria, Nigeria | Osinubi ...

    African Journals Online (AJOL)

    Serum samples from dogs brought for routine physical examination, vaccination and other complaints at the Veterinary Teaching Hospital (VTH) of Ahmadu Bello University and other parts of Zaria were tested for Brucella abortus antibodies. Forty-three (21.5%) of the 200 dogs studied were positive for B. abortus agglutinins ...

  15. MLVA genotyping of human Brucella isolates from Peru

    NARCIS (Netherlands)

    Smits, Henk L.; Espinosa, Benjamin; Castillo, Rosa; Hall, Eric; Guillen, Alfredo; Zevaleta, Milagros; Gilman, Robert H.; Melendez, Paolo; Guerra, Carlos; Draeger, Angelika; Broglia, Alessandro; Nöckler, Karsten

    2009-01-01

    Recent human Brucella melitensis isolates from Peru were genotyped by multiple locus variable number repeat analysis. All 24 isolates originated from hospitalized patients living in the central part of Peru and consisted of six genomic groups comprising two to four isolates and nine unique

  16. Short Communication: Brucella Abortus Antibodies in The Sera of ...

    African Journals Online (AJOL)

    Short Communication: Brucella Abortus Antibodies in The Sera of Indigenous and Exotic Avian Species In Nigeria. SIB Cadmus, HK Adesokan, DO Oluwayelu, AO Idris, JA Stack. Abstract. No Abstract. Full Text: EMAIL FULL TEXT EMAIL FULL TEXT · DOWNLOAD FULL TEXT DOWNLOAD FULL TEXT. Article Metrics.

  17. Brucella bacteremia in patients with acute leukemia: a case series

    Directory of Open Access Journals (Sweden)

    Al-Anazi Khalid

    2007-11-01

    Full Text Available Abstract Background Brucellosis may cause serious infections in healthy individuals living in countries that are endemic for the infection. However, reports of brucella infections in immunocompromised hosts are relatively rare. Case Presentations Reported here are two patients with acute leukemia who developed Brucella melitensis bacteremia during their follow up at the Armed Forces Hospital in Riyadh. The first patient developed B. melitensis bacteremia during the transformation of his myelodysplasia into acute myeloid leukemia. The second patient developed B. melitensis bacteremia while his acute lymphoblastic leukemia was under control. Interestingly, he presented with acute cholecystitis during the brucella sepsis. Both brucella infections were associated with a marked reduction in the hematological parameters in addition to other complications. The bacteremic episodes were successfully treated with netilmicin, doxycycline and ciprofloxacin. Conclusion Brucellosis can cause systemic infections, complicated bacteremia and serious morbidity in patients with acute leukemia living in endemic areas. These infections may occur at the presentation of the leukemia or even when the leukemia is in remission. Nevertheless, the early diagnosis of brucellosis and the administration of appropriate antimicrobial therapy for sufficient duration usually improves the outcome in these immunocompromised patients.

  18. Effect of centrifugation and microagglutination techniques on Brucella agglutinin titers.

    OpenAIRE

    Klein, G C; Behan, K A; Brown, S L; Couch, E E

    1982-01-01

    The microagglutination technique without centrifugation was more effective than centrifugation of the standard tube test for increasing Brucella agglutinin titers of specimens with a titer greater than or equal to 160 but was less effective than centrifugation of the standard tube test for specimens with a titer less than 160.

  19. Identifikasi Brucella abortus Isolat Lokal dengan Brucella abortus Strain Specific-Polymerase Chain Reaction (IDENTIFICATION OF LOCAL ISOLATES OF BRUCELLA ABORTUS USING BRUCELLA ABORTUS STRAIN SPECIFIC-POLYMERASE CHAIN REACTION ASSAY

    Directory of Open Access Journals (Sweden)

    Susan Maphilindawati Noor

    2014-10-01

    Full Text Available Brucella abortus Strain Specific-Polymerase Chain Reaction (BaSS-PCR is a single multiplex PCRtechnique which able to identify and differentiate between Brucella abortus field strains (biovar 1, 2, and4, B. abortus vaccine strains, Brucella species, and non-Brucella species. In this study, BaSS-PCR wasapplied to identify local isolates of B. abortus in order to investigate the B. abortus strains that infectedcattle in Indonesia. Fifty local strains of B.abortus isolated from infected cattle in Java (Jakarta andBandung, South Sulawesi (Maros, East Nusa Tenggara (Kupang and Belu were used in this study. TheDNA bands were observed by agarose gel in the presence of ethidium bromide. Identification was performedbased on the size and number of DNA products amplified by PCR from each isolates. The results showedthat the 50 isolates were of B. abortus field strains. This finding showed that the cause of bovine brucellosisin Indonesia is B. abortus field strains.

  20. Genotyping of Brucella species using clade specific SNPs

    Directory of Open Access Journals (Sweden)

    Foster Jeffrey T

    2012-06-01

    Full Text Available Abstract Background Brucellosis is a worldwide disease of mammals caused by Alphaproteobacteria in the genus Brucella. The genus is genetically monomorphic, requiring extensive genotyping to differentiate isolates. We utilized two different genotyping strategies to characterize isolates. First, we developed a microarray-based assay based on 1000 single nucleotide polymorphisms (SNPs that were identified from whole genome comparisons of two B. abortus isolates , one B. melitensis, and one B. suis. We then genotyped a diverse collection of 85 Brucella strains at these SNP loci and generated a phylogenetic tree of relationships. Second, we developed a selective primer-extension assay system using capillary electrophoresis that targeted 17 high value SNPs across 8 major branches of the phylogeny and determined their genotypes in a large collection ( n = 340 of diverse isolates. Results Our 1000 SNP microarray readily distinguished B. abortus, B. melitensis, and B. suis, differentiating B. melitensis and B. suis into two clades each. Brucella abortus was divided into four major clades. Our capillary-based SNP genotyping confirmed all major branches from the microarray assay and assigned all samples to defined lineages. Isolates from these lineages and closely related isolates, among the most commonly encountered lineages worldwide, can now be quickly and easily identified and genetically characterized. Conclusions We have identified clade-specific SNPs in Brucella that can be used for rapid assignment into major groups below the species level in the three main Brucella species. Our assays represent SNP genotyping approaches that can reliably determine the evolutionary relationships of bacterial isolates without the need for whole genome sequencing of all isolates.

  1. Comparison of the efficacy of Brucella abortus strain RB51 and Brucella melitensis Rev. 1 live vaccines against experimental infection with Brucella melitensis in pregnant ewes.

    Science.gov (United States)

    el Idrissi, A H; Benkirane, A; el Maadoudi, M; Bouslikhane, M; Berrada, J; Zerouali, A

    2001-12-01

    To test the efficacy of rough Brucella strain vaccines in sheep, a vaccine recently developed in cattle (Brucella abortus strain RB51) was assessed in comparison with the conventional Rev. 1 vaccine. Forty-five ewes from twelve to fourteen months of age, from brucellosis-free flocks, were allotted to three groups of fifteen ewes each. Group one was vaccinated by the conjunctival route with 1.73 x 10(8) colony forming units (CFU) of Rev. 1 vaccine. Group two was vaccinated subcutaneously with 11 x 10(9) CFU of RB51 vaccine and group three was considered as a control. All sheep were challenged at two to three months of gestation with 5 x 10(7) CFU of virulent B. melitensis H38. Vaccination with RB51 vaccine did not result in the production of any antibodies against the O-side chain of lipopolysaccharide, as measured by conventional serological tests (Rose Bengal plate test and complement fixation test). Protection of sheep against abortion and excretion of virulent Brucella strain in vaginal fluid, aborted foetuses and/or non viable lambs at parturition and abortion was significantly lower than that afforded by Rev. 1 vaccine. The difference compared to the control group was not significant. Data from this study suggest that the RB51 vaccine used for cattle vaccination does not provide effective protection of sheep against abortion induced by B. melitensis.

  2. Brucella abortus RB51 and hot saline extract from Brucella ovis as antigens in a complement fixation test used To detect sheep vaccinated with Brucella abortus RB51.

    Science.gov (United States)

    Adone, R; Ciuchini, F

    2001-01-01

    The efficacy of Brucella abortus RB51 and hot saline extract (HSE) from Brucella ovis as antigens in complement fixation (CF) tests was comparatively evaluated in detecting immune responses of sheep vaccinated with B. abortus strain RB51. For this study, four 5-month-old sheep were vaccinated subcutaneously with 5 x 10(9) CFU of RB51, and two sheep received saline. Serum samples collected at different times after vaccination were tested for the presence of antibodies to RB51 by a CF test with RB51 as antigen, previously deprived of anticomplementary activity, and with HSE antigen, which already used as the official antigen to detect B. ovis-infected sheep. The results showed that vaccinated sheep developed antibodies which reacted weakly against HSE antigen and these antibodies were detectable for 30 days after vaccination. However, antibodies to RB51 could be detected for a longer period after vaccination by using homologous RB51 antigen in CF tests. In fact, high titers were still present at 110 days postvaccination with RB51 antigen. Sera from sheep naturally infected with B. ovis also reacted to RB51 but gave lower titers than those detected by HSE antigen. As expected, all sera from RB51-vaccinated sheep remained negative when tested with standard S-type Brucella standard antigens.

  3. Brucella ceti Infection in a Common Minke Whale ( Balaenoptera acutorostrata ) with Associated Pathology.

    Science.gov (United States)

    Davison, Nicholas J; Perrett, Lorraine L; Dawson, Claire; Dagleish, Mark P; Haskins, Gary; Muchowski, Jakub; Whatmore, Adrian M

    2017-07-01

    There are three major lineages of marine mammal strains of Brucella spp.: Brucella ceti ST23, found predominantly in porpoises; B. ceti ST26, in pelagic delphinids and ziphiids; and Brucella pinnipedialis ST24/25, predominantly in seals. The isolation of Brucella spp. in mysticetes has been described only in common minke whales ( Balaenoptera acutorostrata ) in Norway and Scotland. We report a third case of Brucella infection and isolation in a minke whale associated with a large abscess. In contrast to the two previous reports that involved isolates of B. pinnipedialis ST24 or the porpoise-associated B. ceti complex ST23, this case was associated with the dolphin-associated B. ceti ST26. Thus, minke whales can be infected naturally with members of all the distinct major lineages of Brucella associated with marine mammals. This report is unique in that the B. ceti ST26 did not originate from a pelagic delphinid or a beaked whale.

  4. A duplex PCR for rapid and simultaneous detection of Brucella spp. in human blood samples.

    Science.gov (United States)

    Mirnejad, Reza; Mohamadi, Mozafar; Piranfar, Vahbeh; Mortazavi, Seied Mojtaba; Kachuei, Reza

    2013-06-01

    To design a duplex PCR for rapid and simultaneous detection of Brucella species. in human blood samples. Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis. Following DNA extraction, PCR assay were performed, using three primers that could simultaneously identify and differentiate three major species of pathogenic Brucella in humans and animals. Of the 52 peripheral bloods samples tested, 25 sample (48%) showed positive reactions in PCR. Twelve samples were positive for Brucella abortus 39 (B. abortus 39) (23%), 13 for Brucella melitensis 39 (B. melitensis 39) (25%) and 0 for Brucella ovis 39 (B. ovis 39) (0%). This work demonstrates that in case where specific primers were utilized, duplex PCR has proved to be a simple, fast, and relatively inexpensive method for simultaneous detection of important species of Brucella in clinical samples. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  5. Highly Sensitive Bacteriophage-Based Detection of Brucella abortus in Mixed Culture and Spiked Blood

    OpenAIRE

    Sergueev, Kirill V.; Filippov, Andrey A.; Nikolich, Mikeljon P.

    2017-01-01

    For decades, bacteriophages (phages) have been used for Brucella species identification in the diagnosis and epidemiology of brucellosis. Traditional Brucella phage typing is a multi-day procedure including the isolation of a pure culture, a step that can take up to three weeks. In this study, we focused on the use of brucellaphages for sensitive detection of the pathogen in clinical and other complex samples, and developed an indirect method of Brucella detection using real-time quantitative...

  6. Molecular epidemiology and antibiotic susceptibility of livestock Brucella melitensis isolates from Naryn Oblast, Kyrgyzstan.

    OpenAIRE

    Kasymbekov, Joldoshbek; Imanseitov, Joldoshbek; Ballif, Marie; Schürch, Nadia; Paniga, Sandra; Pilo, Paola; Tonolla, Mauro; Benagli, Cinzia; Akylbekova, Kulyash; Jumakanova, Zarima; Schelling, Esther; Zinsstag, Jakob

    2013-01-01

    The incidence of human brucellosis in Kyrgyzstan has been increasing in the last years and was identified as a priority disease needing most urgent control measures in the livestock population. The latest species identification of Brucella isolates in Kyrgyzstan was carried out in the 1960s and investigated the circulation of Brucella abortus, B. melitensis, B. ovis, and B. suis. However, supporting data and documentation of that experience are lacking. Therefore, typing of Brucella spp. and ...

  7. Analyses of Brucella pathogenesis, host immunity, and vaccine targets using systems biology and bioinformatics

    OpenAIRE

    Yongqun eHe

    2012-01-01

    Brucella is a Gram-negative, facultative intracellular bacterium that causes zoonotic brucellosis in humans and various animals. Out of 10 classified Brucella species, B. melitensis, B. abortus, B. suis, and B. canis are pathogenic to humans. In the past decade, the mechanisms of Brucella pathogenesis and host immunity have been extensively investigated using the cutting edge systems biology and bioinformatics approaches. This article provides a comprehensive review of the applications of Omi...

  8. Observation of Serum Bactericidal Activity of Brucella abortus RB51 OMPs Combined with Brucella abortus RB51 Live Vaccine

    Directory of Open Access Journals (Sweden)

    Fahime Gholizadeh

    2013-06-01

    Full Text Available Background & objectives: vaccination is vital against brucellosis. Although current vaccines have low efficiency, some cell wall compartments such as Outer Membrane Proteins could be used as an immunogenic candidate in vaccine development. By this mean, our aim in this study was to evaluate the humoral immunity of the combination of Brucella abortus RB51 OMPs with the Brucella abortus RB51 live attenuated vaccine, by Serum Bactericidal Acitivity test. Materials and Methods: In this project, first Brucella abortus RB51 was cultivated in brucella agar. The OMPs were extracted by Sodium N-Lauryl Sarcosinate method, then added to the RB51 live attenuated vaccine. Immunization was done by injection of the vaccine to mice and rabbits. The blood was drawn on days 0, 15,30, and 45 from the rabbits and the sera were seperated. Brucella abortus 544 was also injected as challenge. Spleen colony count was also performed. Results: The data from Serum Bactericidal Assay has showed, there was a very high Humoral immunity and response as a bactericidal titre of the serum against Rb51 Live vaccine. There was a significant decrease of colonies in the group vaccinated with the combined vaccine in the Spleen colony count test. Statistical analysis of groups variances showed a significant difference between groups (P<0.05.Conclusions: The Serum Bactericidal Assay results showed despite previous studies, both the combine and live vaccine are capable to stimulate the Humoral immunity. greater activity of combined vaccine to boost the humoral activity might be due to the synergistic effect of this vaccine.

  9. A rare case of anterior mediastinal mass caused by Brucella infection.

    Science.gov (United States)

    Sabzi, Feridoun; Faraji, Reza

    2017-03-01

    A previously healthy man, who had undergone coronary artery bypass 10 years earlier and had been diagnosed with brucellosis due to Brucella septicemia after Brucella arthritis, presented with chest pain and high fever. Anti- Brucella antibiotics were started, but after 4 weeks, his high fever remained. An infected mass was confirmed by computed tomography, and surgical intervention was performed via a median sternotomy. A large amount of thick pus gushed from an abscess in the upper mediastinum. The abscess cavity had a thick granulation wall, and cultured pus was positive for Brucella only. The patient responded well to antibiotic therapy.

  10. MLVA and MLST typing of Brucella from Qinghai, China.

    Science.gov (United States)

    Ma, Jun-Ying; Wang, Hu; Zhang, Xue-Fei; Xu, Li-Qing; Hu, Gui-Ying; Jiang, Hai; Zhao, Fang; Zhao, Hong-Yan; Piao, Dong-Ri; Qin, Yu-Min; Cui, Bu-Yun; Lin, Gong-Hua

    2016-04-13

    The Qinghai-Tibet Plateau (QTP) of China is an extensive pastoral and semi-pastoral area, and because of poverty and bad hygiene conditions, Brucella is highly prevalent in this region. In order to adequately prevent this disease in the QTP region it is important to determine the identity of Brucella species that caused the infection. A total of 65 Brucella isolates were obtained from human, livestock and wild animals in Qinghai, a Chinese province in east of the QTP. Two molecular typing methods, MLVA (multi-locus variable-number tandem-repeat analysis) and MLST (multi locus sequence typing) were used to identify the species and genotypes of these isolates. Both MLVA and MLST typing methods classified the 65 isolates into three species, B. melitensis, B. abortus and B. suis, which included 60, 4 and 1 isolates respectively. The MLVA method uniquely detected 34 (Bm01 ~ Bm34), 3 (Ba01 ~ Ba03), and 1 (Bs01) MLVA-16 genotypes for B. melitensis, B. abortus and B. suis, respectively. However, none of these genotypes exactly matched any of the genotypes in the Brucella2012 MLVA database. The MLST method identified five known ST types: ST7 and ST8 (B. melitensis), ST2 and ST5 (B. abortus), and ST14 (B. suis). We also detected a strain with a mutant type (3-2-3-2-?-5-3-8-2) of ST8 (3-2-3-2-1-5-3-8-2). Extensive genotype-sharing events could be observed among isolates from different host species. There were at least three Brucella (B. melitensis, B. abortus and B. suis) species in Qinghai, of which B. melitensis was the predominant species in the area examined. The Brucella population in Qinghai was very different from other regions of the world, possibly owing to the unique geographical characteristics such as extremely high altitude in QTP. There were extensive genotype-sharing events between isolates obtained from humans and other animals. Yaks, sheep and blue sheep were important zoonotic reservoirs of brucellosis causing species found in humans.

  11. Brucella suis vaccine strain S2-infected immortalized caprine endometrial epithelial cell lines induce non-apoptotic ER-stress

    OpenAIRE

    Wang, Xiangguo; Lin, Pengfei; Yin, Yanlong; Zhou, Jinhua; Lei, Lanjie; Zhou, Xudong; Jin, Yaping; WANG, Aihua

    2015-01-01

    Brucella, which is regarded as an intracellular pathogen responsible for a zoonotic disease called brucellosis, survives and proliferates within several types of phagocytic and non-phagocytic cells. Brucella infects not only their preferred hosts but also other domestic and wild animal species, inducing abortion and infertility. Therefore, the interaction between uterine cells and Brucella is important for understanding the pathogenesis of this disease. In this study, we describe the Brucella...

  12. Complete Genome Sequence of Brucella abortus SKN 13 Isolated from Placenta of Aborted Cattle in Gujarat, India.

    Science.gov (United States)

    Chauhan, H C; Patel, Bharat K; Chandel, B S; Patel, Kirit B; Patel, A C; Shrimali, M D; Patel, S S; Bhagat, A G; Rajgor, Manish; Patel, Mitul A; Patel, Maulik; Kala, Jitendra; Patel, Bhumika

    2016-10-27

    Brucella abortus is generally known to cause brucellosis in cattle and buffalo. Here, we report the draft genome sequence of Brucella abortus SKN 13, isolated from aborted cattle placenta in the area of Gujarat, India, providing precious resources for comparative genomic analyses of Brucella field strains. Copyright © 2016 Chauhan et al.

  13. BRUCELLA ENDOCARDITIS IN IRANIAN PATIENTS: COMBINED MEDICAL AND SURGICAL TREATMENT

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    Ebrahim Nematipour

    1995-06-01

    Full Text Available Brucella endocarditis is a Tare but serious complication ofbrucellosis and is the main cause of death reuuedto thisdisease: Itis not rare in the endemic areas and aaualiy accounts for up to 8~lO% ofendocarditis infections: We report seven adult cases of brucella endocarditis in lmam-Khorneini Hospual: Contrary to previous independent reports, female patients were not rare in this study and accountedfor three out ofseven. Four patients were cared for by combined medical and surgical treatment and were recovered Three of the patients that did not receive the combined theraPl could not he saved This report confirms the necessity of prompt combined medical and surgical treatment ofbrucella endocarditis.

  14. Brucella suis in armadillos (Chaetophractus villosus) from La Pampa, Argentina.

    Science.gov (United States)

    Kin, Marta S; Fort, Marcelo; de Echaide, Susana T; Casanave, Emma B

    2014-06-04

    Brucellosis is a zoonotic disease transmitted from an animal reservoir to humans. Both, wildlife and domestic animals, contribute to the spreading of these zoonosis. The surveillance of the animal health status is strictly regulated for domestic animals, whereas disease monitoring in wildlife does not exist. The aim of the present study was to provide data on the prevalence of anti-Brucella antibodies in Chaetophractus villosus from a region of La Pampa, Argentina to assess public health risks. The C. villosus is endemic to South America, and in Argentina it represents a food resource for human consumption. A total of 150 sera of armadillos bleeding between 2007 and 2010 were tested using buffered plate antigen test (BPAT), serum agglutination test (SAT), 2-mercaptoethanol (2-ME) and complement fixation test (CFT), for the detection of anti-Brucella antibodies. Antibodies to Brucella sp. were found in 16% (24:150) of the armadillos tested using the BPAT test. All 24 positive samples were confirmed by the SAT, 2-ME and CFT tests. Strain isolation was attempted from liver and spleen samples of two animals with positive serology. Isolates were characterized by conventional biotyping and identification of specific DNA using polymerase chain reaction (PCR). A total of 2 isolates were recovered from spleen and liver. Both of them were identified as Brucella suis biovar 1. This preliminary study provides the first report on the seroprevalence of brucellosis and describes the first isolate of B. suis biovar 1 in C. villosus in Argentina. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. The first International Standard anti-Brucella melitensis Serum

    OpenAIRE

    McGiven, J.; Taylor, A.; Duncombe, L.; Sayers, R.; Albert, D.; Banai, M; Blasco Martínez, José María; Elena, S; Fretin, D.; Garin-Bastuji, B.; Melzer, F.; Muñoz Alvaro, Pilar María; Nielsen, K.; De Nicola, A.; Scacchia, M.

    2011-01-01

    The World Organisation for Animal Health (OIE) requested an International Standard anti-Brucella melitensis Serum (ISaBmS) to standardise diagnostic tests and reagents for sheep and goats. The agreed criteria were the highest dilution (in negative serum) of the standard which must give a positive result and the lowest dilution (in negative serum) which must simultaneously give a negative result. The two dilutions for each assay were, respectively: indirect enzyme-linked immu...

  16. New Features in the Lipid A Structure of Brucella suis and Brucella abortus Lipopolysaccharide

    Science.gov (United States)

    Casabuono, Adriana C.; Czibener, Cecilia; Del Giudice, Mariela G.; Valguarnera, Ezequiel; Ugalde, Juan E.; Couto, Alicia S.

    2017-12-01

    Brucellaceae are Gram-negative bacteria that cause brucellosis, one of the most distributed worldwide zoonosis, transmitted to humans by contact with either infected animals or their products. The lipopolysaccharide exposed on the cell surface has been intensively studied and is considered a major virulence factor of Brucella. In the last years, structural studies allowed the determination of new structures in the core oligosaccharide and the O-antigen of this lipopolysaccharide. In this work, we have reinvestigated the lipid A structure isolated from B. suis and B. abortus lipopolysaccharides. A detailed study by MALDI-TOF mass spectrometry in the positive and negative ion modes of the lipid A moieties purified from both species was performed. Interestingly, a new feature was detected: the presence of a pyrophosphorylethanolamine residue substituting the backbone. LID-MS/MS analysis of some of the detected ions allowed assurance that the Lipid A structure composed by the diGlcN3N disaccharide, mainly hexa-acylated and penta-acylated, bearing one phosphate and one pyrophosphorylethanolamine residue. [Figure not available: see fulltext.

  17. New Features in the Lipid A Structure of Brucella suis and Brucella abortus Lipopolysaccharide

    Science.gov (United States)

    Casabuono, Adriana C.; Czibener, Cecilia; Del Giudice, Mariela G.; Valguarnera, Ezequiel; Ugalde, Juan E.; Couto, Alicia S.

    2017-09-01

    Brucellaceae are Gram-negative bacteria that cause brucellosis, one of the most distributed worldwide zoonosis, transmitted to humans by contact with either infected animals or their products. The lipopolysaccharide exposed on the cell surface has been intensively studied and is considered a major virulence factor of Brucella. In the last years, structural studies allowed the determination of new structures in the core oligosaccharide and the O-antigen of this lipopolysaccharide. In this work, we have reinvestigated the lipid A structure isolated from B. suis and B. abortus lipopolysaccharides. A detailed study by MALDI-TOF mass spectrometry in the positive and negative ion modes of the lipid A moieties purified from both species was performed. Interestingly, a new feature was detected: the presence of a pyrophosphorylethanolamine residue substituting the backbone. LID-MS/MS analysis of some of the detected ions allowed assurance that the Lipid A structure composed by the diGlcN3N disaccharide, mainly hexa-acylated and penta-acylated, bearing one phosphate and one pyrophosphorylethanolamine residue. [Figure not available: see fulltext.

  18. Seroprevalence of Brucella antibodies in camels in Katsina State, Nigeria.

    Science.gov (United States)

    Salisu, U S; Kudi, C A; Bale, J O O; Babashani, M; Kaltungo, B Y; Saidu, S N A; Asambe, A; Baba, A Y

    2017-06-01

    A cross-sectional study was carried out to determine the status of Brucella infection in one-humped (Dromedary) camels in the North and Central senatorial districts of Katsina State, Nigeria. Nine hundred and eighty serum samples from live and slaughtered camels were tested. Modified Rose Bengal plate test (RBPT) and serum agglutination test (SAT) with ethylenediaminetetraacetic acid, (EDTA) were used as screening and standard tests, respectively. The prevalence of Brucella antibodies were 110 (11.2%) and 103 (10.5%) for RBPT and SAT, respectively. Of the 472 and 508 serum samples tested from the herds and abattoirs, respectively, 63 (13.3%) and 47 (9.3%) were positive by RBPT while 62 (13.1%) and 41 (8.1%) were positive by SAT, respectively. Based on the results, it was concluded that Brucella antibodies were present in camels in the study area. Poor management practices and mixing of camels with other species of livestock as well as unrestricted movement of camels were proposed to be the reasons for the prevalence of the disease in the study area. In view of the public health importance of the disease, it is recommended that there is the need to develop a strategic plan to decrease spread of brucellosis in the study area.

  19. Validation of the multiplex PCR for identification of Brucella spp.

    Directory of Open Access Journals (Sweden)

    Lívia de Lima Orzil

    2016-05-01

    Full Text Available ABSTRACT: A multiplex PCR technique for detection of Brucella spp. in samples of bacterial suspension was validated as a complementary tool in the diagnosis of the disease. This technique allows the characterization of the agent without performing biochemical tests, which greatly reduces the time for a final diagnosis, and provides more security for the analyst by reducing the time of exposure to microorganisms. The validation was performed in accordance with the Manual of Diagnostic Tests from OIE (2008 and following the requirements present in the ABNT NBR ISO/IEC 17025:2005. The mPCR validated in this study identified the different species of Brucella ( Brucella abortus , B. suis , B. ovis e B. melitensis of bacterial suspension obtained from the slaughterhouse samples, as well as distinguished the biovars (1, 2 e 4; 3b, 5, 6 e 9 of B. abortus in grouped form and differentiated the field strains from vaccine strains, as a quick, useful and less expensive technique in diagnosis of brucellosis in Brazil.

  20. Epidemiological survey for Brucella in wildlife and stray dogs, a cat and rodents captured on farms.

    Science.gov (United States)

    Truong, Lam Quang; Kim, Jung Taek; Yoon, Byung-Il; Her, Moon; Jung, Suk Chan; Hahn, Tae-Wook

    2011-12-01

    Brucella infections in wildlife originate either from contact with infected livestock or from a natural sustainable reservoir in wildlife populations. As South Korea has set a goal of brucellosis eradication by 2013, it is necessary to determine the prevalence of Brucella in wildlife and wild rodents. This information will play an important role in the control of brucellosis. Because of the absence of prominent clinical signs, direct and indirect laboratory tests are essential for diagnosing brucellosis. In this study, tissue and blood samples were taken from wild animals, abandoned dogs, a cat and wild rodents, and they were tested for Brucella or Brucella-specific antibodies by isolation, PCR and serology. Results showed that 18.6% (33/177) of blood samples were positive by PCR, and 5.7% (11/194) were positive by C-ELISA. However, none of these samples yielded culturable bacteria. Of the tissue samples, 9.7% (8/82) were positive by PCR. Brucella was isolated from only one tissue culture from a Chinese water deer carcass. This Brucella species was identified as Brucella abortus biovar 1 by biotyping, 16S rRNA PCR and the Bruce-ladder PCR assay. In this study, we reported the prevalence of Brucella in wildlife, dogs, a cat and rodents by using serological and molecular methods, and we report the first isolation of B. abortus in wild Chinese water deer in South Korea.

  1. Identification of Brucella spp. in feral swine (Sus scrofa) at abattoirs in Texas, USA

    Science.gov (United States)

    Various tissues, nasal swabs, urine, and blood samples were collected from 376 feral swine at two federally-inspected abattoirs in Texas during six separate sampling periods in 2015. Samples were tested for Brucella spp. by culture and serology. Brucella spp. were cultured from 13.0% of feral swin...

  2. Serological response of cattle to Brucella allergen after repeated intradermal applications of this allergen

    NARCIS (Netherlands)

    Muskens, J.A.M.; Bercovich, Z.; Damen, C.P.R.M.

    1996-01-01

    A study was conducted to determine whether an allergen that has been prepared from a mucoid strain of Brucella abortus triggers a serum antibody response that interferes with the interpretation of serologic tests results. Fifteen cattle seronegative for Brucella antigen were tested with the SDTH

  3. Pathogenesis and immunobiology of brucellosis: review of Brucella-host interactions.

    Science.gov (United States)

    de Figueiredo, Paul; Ficht, Thomas A; Rice-Ficht, Allison; Rossetti, Carlos A; Adams, L Garry

    2015-06-01

    This review of Brucella-host interactions and immunobiology discusses recent discoveries as the basis for pathogenesis-informed rationales to prevent or treat brucellosis. Brucella spp., as animal pathogens, cause human brucellosis, a zoonosis that results in worldwide economic losses, human morbidity, and poverty. Although Brucella spp. infect humans as an incidental host, 500,000 new human infections occur annually, and no patient-friendly treatments or approved human vaccines are reported. Brucellae display strong tissue tropism for lymphoreticular and reproductive systems with an intracellular lifestyle that limits exposure to innate and adaptive immune responses, sequesters the organism from the effects of antibiotics, and drives clinical disease manifestations and pathology. Stealthy brucellae exploit strategies to establish infection, including i) evasion of intracellular destruction by restricting fusion of type IV secretion system-dependent Brucella-containing vacuoles with lysosomal compartments, ii) inhibition of apoptosis of infected mononuclear cells, and iii) prevention of dendritic cell maturation, antigen presentation, and activation of naive T cells, pathogenesis lessons that may be informative for other intracellular pathogens. Data sets of next-generation sequences of Brucella and host time-series global expression fused with proteomics and metabolomics data from in vitro and in vivo experiments now inform interactive cellular pathways and gene regulatory networks enabling full-scale systems biology analysis. The newly identified effector proteins of Brucella may represent targets for improved, safer brucellosis vaccines and therapeutics. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  4. Serological Survey of Porcine Brucella Infection in SouthEast, Nigeria

    African Journals Online (AJOL)

    Porcine brucellosis, also called contagious abortion of swine is an infectious and zoonotic disease of swine caused by bacteria of the genus Brucella (Young, 1995). Brucella suis is the species found primarily in pigs. It is a zoonotic infection of domesticated and wild animals which humans (especially occupationally ...

  5. Analysis of pan-genome to identify the core genes and essential genes of Brucella spp.

    Science.gov (United States)

    Yang, Xiaowen; Li, Yajie; Zang, Juan; Li, Yexia; Bie, Pengfei; Lu, Yanli; Wu, Qingmin

    2016-04-01

    Brucella spp. are facultative intracellular pathogens, that cause a contagious zoonotic disease, that can result in such outcomes as abortion or sterility in susceptible animal hosts and grave, debilitating illness in humans. For deciphering the survival mechanism of Brucella spp. in vivo, 42 Brucella complete genomes from NCBI were analyzed for the pan-genome and core genome by identification of their composition and function of Brucella genomes. The results showed that the total 132,143 protein-coding genes in these genomes were divided into 5369 clusters. Among these, 1710 clusters were associated with the core genome, 1182 clusters with strain-specific genes and 2477 clusters with dispensable genomes. COG analysis indicated that 44 % of the core genes were devoted to metabolism, which were mainly responsible for energy production and conversion (COG category C), and amino acid transport and metabolism (COG category E). Meanwhile, approximately 35 % of the core genes were in positive selection. In addition, 1252 potential essential genes were predicted in the core genome by comparison with a prokaryote database of essential genes. The results suggested that the core genes in Brucella genomes are relatively conservation, and the energy and amino acid metabolism play a more important role in the process of growth and reproduction in Brucella spp. This study might help us to better understand the mechanisms of Brucella persistent infection and provide some clues for further exploring the gene modules of the intracellular survival in Brucella spp.

  6. Molecular Survey on Brucellosis in Rodents and Shrews - Natural Reservoirs of Novel Brucella Species in Germany?

    Science.gov (United States)

    Hammerl, J A; Ulrich, R G; Imholt, C; Scholz, H C; Jacob, J; Kratzmann, N; Nöckler, K; Al Dahouk, S

    2017-04-01

    Brucellosis is a widespread zoonotic disease introduced from animal reservoirs to humans. In Germany, bovine and ovine/caprine brucellosis were eradicated more than a decade ago and mandatory measures in livestock have been implemented to keep the officially brucellosis-free status. In contrast, surveillance of wildlife is still challenging, and reliable data on the prevalence of brucellae in small mammal populations do not exist. To assess the epidemiology of Brucella spp. in rodents and shrews, a molecular survey was carried out. A total of 537 rodents and shrews were trapped in four federal states located throughout Germany and investigated for the presence of Brucella. Using a two-step molecular assay based on the detection of the Brucella-specific bcsp31 and IS711 sequences in tissue samples, 14.2% (n = 76) of the tested animals were positive. These originated mainly from western and south-western Germany, where preliminary analyses indicate population density-dependent Brucella prevalence in voles (Myodes glareolus) and mice (Apodemus spp.). recA typing revealed a close relationship to a potentially novel Brucella species recently isolated from red foxes (Vulpes vulpes) in Austria. The molecular detection of brucellae in various rodent taxa and for the first time in shrew species shows that these animals may be naturally infected or at least have a history of exposure to Brucella spp. © 2015 Blackwell Verlag GmbH.

  7. Vector Development for the Expression of Foreign Proteins in the Vaccine Strain Brucella abortus S19

    Science.gov (United States)

    Comerci, Diego J.; Pollevick, Guido D.; Vigliocco, Ana M.; Frasch, Alberto C. C.; Ugalde, Rodolfo A.

    1998-01-01

    A vector for the expression of foreign antigens in the vaccine strain Brucella abortus S19 was developed by using a DNA fragment containing the regulatory sequences and the signal peptide of the Brucella bcsp31 gene. This fragment was cloned in broad-host-range plasmid pBBR4MCS, resulting in plasmid pBEV. As a reporter protein, a repetitive antigen of Trypanosoma cruzi was used. The recombinant fusion protein is stably expressed and secreted into the Brucella periplasmic space, inducing a good antibody response against the T. cruzi antigen. The expression of the repetitive antigen in Brucella neither altered its growth pattern nor generated a toxic or lethal effect during experimental infection. The application of this strategy for the generation of live recombinant vaccines and the tagging of B. abortus S19 vaccine is discussed. This is the first time that a recombinant protein has been expressed in the periplasm of brucellae. PMID:9673273

  8. Differential Stimulatory Activities of Smooth and Rough Brucella abortus Lipopolysaccharide in Murine Macrophages

    Directory of Open Access Journals (Sweden)

    Raheela Akhtar1,2*, Yongqun O. He2, Charles B. Larson2, Zafar I. Chaudhary3 and Mansur ud-Din Ahmad4

    2012-06-01

    Full Text Available Brucella abortus lipopolysaccharide (LPS was isolated and purified from rough (RB51 and smooth (S2308 strains of Brucella. The LPS preparations were used to treat murine (RAW 264.7 macrophages in order to study their differential effects. Treated macrophages were tested by lysozyme release test (LRT, nitroblue tetrazolium test (NBT and nitric oxide (NO assay, respectively. Rough Brucella LPS induced significantly higher levels of lysozyme release, oxidative stress, and nitric oxide in murine macrophages than smooth Brucella LPS or combined LPS (rough + smooth LPS. These responses were dose-dependent. Macrophages treated with rough LPS were more Brucellacidal than those treated with smooth LPS. The minimal stimulation of murine macrophages by Brucella smooth LPS may provide basis for less active immune responses against smooth strains.

  9. Genomic comparisons of Brucella spp. and closely related bacteria using base compositional and proteome based methods

    DEFF Research Database (Denmark)

    Bohlin, Jon; Snipen, Lars; Cloeckaert, Axel

    2010-01-01

    BACKGROUND: Classification of bacteria within the genus Brucella has been difficult due in part to considerable genomic homogeneity between the different species and biovars, in spite of clear differences in phenotypes. Therefore, many different methods have been used to assess Brucella taxonomy....... In the current work, we examine 32 sequenced genomes from genus Brucella representing the six classical species, as well as more recently described species, using bioinformatical methods. Comparisons were made at the level of genomic DNA using oligonucleotide based methods (Markov chain based genomic signatures...... between the oligonucleotide based methods used. Whilst the Markov chain based genomic signatures grouped the different species in genus Brucella according to host preference, the codon and amino acid frequencies based methods reflected small differences between the Brucella species. Only minor differences...

  10. Comparison of a New and Rapid Method: Brucella Coombs Gel Test With Other Diagnostic Tests.

    Science.gov (United States)

    Kalem, Fatma; Ergün, Ayşe Gül; Durmaz, Süleyman; Doğan, Metin; Ertuğrul, Ömür; Gündem, Seval

    2016-09-01

    The aim of this study was to detect reliability of Brucella Coombs gel test (BCGT) by comparing with with ELISA (IgG + IgM), Standard agglutination test, and Brucella immunocapture agglutination methods in serological diagnosis of brucellosis. Brucella Coombs gel test (BCGT), Brucella ELISA (IgG + IgM), Standard agglutination test, and Brucella immunocapture agglutination tests of 78 patients with presumptive diagnosis of brucellosis which were sent to Microbiology Laboratory of Konya Numune Hospital from various regions of Konya were studied. Of 78 patients with ELISA IgG and IgM, STA, BICA and BCGT; 26, 21, 10, 12 and 12 were positive. When compared with BICA, the sensitivity and specifity of BCGT were 100% and 100%, respectively. According to results BCGT can be used as a diagnostic test in routine laboratories after more comprehensive studies in control groups and patients. © 2016 Wiley Periodicals, Inc.

  11. Isolation of Brucella inopinata-Like Bacteria from White's and Denny's Tree Frogs.

    Science.gov (United States)

    Kimura, Masanobu; Une, Yumi; Suzuki, Michio; Park, Eun-Sil; Imaoka, Koichi; Morikawa, Shigeru

    2017-05-01

    Brucella inopinata strain BO1 and B. sp. strain BO2 isolated from human patients, respectively, are genetically different from classical Brucella species. We isolated bacteria of the genus Brucella from two species of wild-caught tropical frogs kept in the facilities in Japan: White's tree frog, which inhabits Oceania, and Denny's tree frog, which inhabits Southeast Asia. Phylogenetic analyses based on 16S rRNA and recA gene sequences and multilocus sequence analysis showed that two isolates of Brucella spp. showed significant similarity to BO1, BO2, and the isolates from other wild-caught frogs. These results suggest that a variety of frog species are susceptible to a novel clade of Brucella bacteria, including B. inopinata.

  12. Chronic meningoencephalitis associated with Brucella sp. infection in live-stranded striped dolphins (Stenella coeruleoalba).

    Science.gov (United States)

    González, L; Patterson, I A; Reid, R J; Foster, G; Barberán, M; Blasco, J M; Kennedy, S; Howie, F E; Godfroid, J; MacMillan, A P; Schock, A; Buxton, D

    2002-01-01

    A chronic, non-suppurative meningoencephalitis was found in three young striped dolphins (Stenella coeruleoalba) during routine neuropathological examination of marine mammals live-stranded on the Scottish coast. In all three dolphins the lesions were associated with the isolation of a Brucella sp. from the brain and with the immunohistochemical detection of brucella antigen. Moreover, antibodies to Brucella spp. were detected in the two dolphins that were subjected to serological examination. Immunohistochemical and serological examinations for morbillivirus antigen and antibodies, respectively, were negative in all cases. Although brucella infection of marine mammals has been extensively documented in recent years, its association with lesions and disease is less well recognized. The present report provides the first description of an association between Brucella sp. infection and neuropathological changes in a cetacean species. Copyright Harcourt Publishers Ltd.

  13. Identification of Secondary Mutations Which Enhance and Stabilize the Attenuation of Brucella HTRA Mutants: Improving Brucella HTRA-Based Strains as Vaccine

    National Research Council Canada - National Science Library

    Roop, R

    1999-01-01

    .... Studies completed to date under this contract indicate that although the Brucella HtrA contributes to resistance to oxidative killing in vitro and resistance to killing by cultured murine neutrophils...

  14. [Molecular typing of 12 Brucella strains isolated in Guizhou province in 2010-2013].

    Science.gov (United States)

    Wang, Yue; Chen, Hong; Liu, Ying; Zhou, Jingzhu; Li, Shijun; Hang, Yan; Tang, Guangpeng; Wang, Dingming; Chen, Guichun

    2015-09-01

    To identify and characterize the Brucella strains from Guizhou province in 2010-2013. A total of 12 strains of Brucella suspicious bacteria were isolated in Guizhou province from 2010 to 2013. Four strains (GZLL3, GZLL4, GZLL11 and SH2) were isolated from goat blood samples and eight strains (SH4, GZZY, GZSQ, GZZA, BR13001, BR13004, BR13005 and BR13006) were isolated from blood samples of patient 12 Brucella suspicious strains were identified and characterized using conventional methods. Brucella genus specific gene BCSP31-based PCR (BCSP31-PCR) was used to identify the genus of Brucella and IS711 insert sequence-based PCR (AMOS-PCR) was applied to identify the species of Brucella strains. Goats and patients originated Brucella strains were comparatively analysed using Pulse-field Gel Electrophoresis (PFGE). Both of conventional methods and PCR identified the 12 Brucella suspicious strains as B. melitensis biotype 3. BCSP31-PCR identification results showed that a specific DNA bands (223 bp) were detected in all the 12 strains and positive control samples with no DNA band in negative samples. AMOS-PCR amplified a 731 bp-DNA bands in all the 12 strains, with 731 bp, 498 bp and 275 bp in M5, S2 and A19 strains, respectively, and no DNA band was detected in the negative control samples. PFGE analysis showed that 12 Brucella isolates from patients and goats showed consistent PFGE patterns with the digestion of restriction enzyme Xba I. The epidemic species/type of Brucella in both human and animal in Guizhou province was B. melitensis biotype 3 and goat was the main animal source of infection of brucellosis in Guizhou province.

  15. Evaluation of PCR methods for detection of Brucella strains from culture and tissues.

    Science.gov (United States)

    Çiftci, Alper; İça, Tuba; Savaşan, Serap; Sareyyüpoğlu, Barış; Akan, Mehmet; Diker, Kadir Serdar

    2017-04-01

    The genus Brucella causes significant economic losses due to infertility, abortion, stillbirth or weak calves, and neonatal mortality in livestock. Brucellosis is still a zoonosis of public health importance worldwide. The study was aimed to optimize and evaluate PCR assays used for the diagnosis of Brucella infections. For this aim, several primers and PCR protocols were performed and compared with Brucella cultures and biological material inoculated with Brucella. In PCR assays, genus- or species-specific oligonucleotide primers derived from 16S rRNA sequences (F4/R2, Ba148/928, IS711, BruP6-P7) and OMPs (JPF/JPR, 31ter/sd) of Brucella were used. All primers except for BruP6-P7 detected the DNA from reference Brucella strains and field isolates. In spiked blood, milk, and semen samples, F4-R2 primer-oriented PCR assays detected minimal numbers of Brucella. In spiked serum and fetal stomach content, Ba148/928 primer-oriented PCR assays detected minimal numbers of Brucella. Field samples collected from sheep and cattle were examined by bacteriological methods and optimized PCR assays. Overall, sensitivity of PCR assays was found superior to conventional bacteriological isolation. Brucella DNA was detected in 35.1, 1.1, 24.8, 5.0, and 8.0% of aborted fetus, blood, milk, semen, and serum samples by PCR assays, respectively. In conclusion, PCR assay in optimized conditions was found to be valuable in sensitive and specific detection of Brucella infections of animals.

  16. Differences in two-component signal transduction proteins among the genus Brucella: implications for host preference and pathogenesis

    DEFF Research Database (Denmark)

    Binnewies, Tim Terence; Ussery, David; Lavín, JL

    2010-01-01

    Two-component systems (TCSs) are the predominant bacterial signal transduction mechanisms. Species of the genus Brucella are genetically highly related and differ mainly in mammalian host adaptation and pathogenesis. In this study, TCS proteins encoded in the available genome sequences of Brucella...... species have been identified using bioinformatic methods. All the Brucella species share an identical set of TCS proteins, and the number of TCS proteins in the closely related opportunistic human pathogen Ochrobactrum anthropi was higher than in Brucella species as expected from its lifestyle. O....... anthropi lacks orthologs of the Brucella TCSs NodVW, TceSR and TcfSR, suggesting that these TCS proteins could be necessary for the adaptation of Brucella as an intracellular pathogen. This genomic analysis revealed the presence of a differential distribution of TCS pseudogenes among Brucella species...

  17. Rapid detection of Brucella spp. using loop-mediated isothermal amplification (LAMP).

    Science.gov (United States)

    Chen, Shouyi; Li, Xunde; Li, Juntao; Atwill, Edward R

    2013-01-01

    Brucella spp. are facultative intracellular bacteria that cause zoonotic disease of brucellosis worldwide. Livestock that are most vulnerable to brucellosis include cattle, goats, and pigs. Brucella spp. cause serious health problems to humans and animals and economic losses to the livestock industry. Traditional methods for detection of Brucella spp. take 48-72 h (Kumar et al., J Commun Dis 29:131-137, 1997; Barrouin-Melo et al., Res Vet Sci 83:340-346, 2007) that do not meet the food industry's need of rapid detection. Therefore, there is an urgent need of fast, specific, sensitive, and inexpensive method for diagnosing of Brucella spp. Loop-mediated isothermal amplification (LAMP) is a method to amplify nucleic acid at constant temperatures. Amplification can be detected by visual detection, fluorescent stain, turbidity, and electrophoresis. We targeted at the Brucella-specific gene omp25 and designed LAMP primers for detection of Brucella spp. Amplification of DNA with Bst DNA polymerase can be completed at 65 °C in 60 min. Amplified products can be detected by SYBR Green I stain and 2.0% agarose gel electrophoresis. The LAMP method is feasible for detection of Brucella spp. from blood and milk samples.

  18. Isolation and identification of bovine Brucella isolates from Pakistan by biochemical tests and PCR.

    Science.gov (United States)

    Ali, Shahzad; Ali, Qurban; Melzer, Falk; Khan, Iahtasham; Akhter, Shamim; Neubauer, Heinrich; Jamal, Syed M

    2014-01-01

    Brucellosis is endemic in bovines in Pakistan. The Brucella species and biovars involved, however, are unknown. The objectives of the present study were to isolate and characterize brucellae from seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes which had recently aborted. The seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes were collected from the Potohar Plateau, Pakistan. Isolation of brucellae was done on modified Farrell's serum dextrose agar. Isolates were characterized by conventional biotyping methods, while molecular typing was done by genus (B4/B5) and species-specific (Brucella abortus, Brucella melitensis, Brucella ovis, and Brucella suis) polymerase chain reaction (PCR). A total of 30 isolates were recovered from milk (n = 5), aborted fetuses (n = 13), and vaginal swabs (n = 12). Most isolates were from cattle (56.7 %). All of them were identified as B. abortus biovar 1 based on conventional biotyping methods and genus and species-specific PCR. This preliminary study provides the first report on the prevalence of B. abortus biovar 1 in cattle and buffaloes in Pakistan.

  19. Construction of pTM series plasmids for gene expression in Brucella species.

    Science.gov (United States)

    Tian, Mingxing; Qu, Jing; Bao, Yanqing; Gao, Jianpeng; Liu, Jiameng; Wang, Shaohui; Sun, Yingjie; Ding, Chan; Yu, Shengqing

    2016-04-01

    Brucellosis, the most common widespread zoonotic disease, is caused by Brucella spp., which are facultative, intracellular, Gram-negative bacteria. With the development of molecular biology techniques, more and more virulence-associated factors have been identified in Brucella spp. A suitable plasmid system is an important tool to study virulence genes in Brucella. In this study, we constructed three constitutive replication plasmids (pTM1-Cm, pTM2-Amp, and pTM3-Km) using the replication origin (rep) region derived from the pBBR1-MCS vector. Also, a DNA fragment containing multiple cloning sites (MCSs) and a terminator sequence derived from the pCold vector were produced for complementation of the deleted genes. Besides pGH-6×His, a plasmid containing the groE promoter of Brucella spp. was constructed to express exogenous proteins in Brucella with high efficiency. Furthermore, we constructed the inducible expression plasmid pZT-6×His, containing the tetracycline-inducible promoter pzt1, which can induce expression by the addition of tetracycline in the Brucella culture medium. The constructed pTM series plasmids will play an important role in the functional investigation of Brucella spp. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Brucella antibody seroprevalence in Antarctic seals (Arctocephalus gazella, Leptonychotes weddellii and Mirounga leonina).

    Science.gov (United States)

    Jensen, Silje-Kristin; Nymo, Ingebjørg Helena; Forcada, Jaume; Hall, Ailsa; Godfroid, Jacques

    2013-09-03

    Brucellosis is a worldwide infectious zoonotic disease caused by Gram-negative bacteria of the genus Brucella, and Brucella infections in marine mammals were first reported in 1994. A serosurvey investigating the presence of anti-Brucella antibodies in 3 Antarctic pinniped species was undertaken with a protein A/G indirect enzyme-linked immunosorbent assay (iELISA) and the Rose Bengal test (RBT). Serum samples from 33 Weddell seals Leptonychotes weddelli were analysed, and antibodies were detected in 8 individuals (24.2%) with the iELISA and in 21 (65.6%) with the RBT. We tested 48 southern elephant seal Mirounga leonina sera and detected antibodies in 2 animals (4.7%) with both the iELISA and the RBT. None of the 21 Antarctic fur seals Arctocephalus gazella was found positive. This is the first report of anti-Brucella antibodies in southern elephant seals. The potential impact of Brucella infection in pinnipeds in Antarctica is not known, but Brucella spp. are known to cause abortion in terrestrial species and cetaceans. Our findings suggest that Brucella infection in pinnipeds is present in the Antarctic, but to date B. pinnipedialis has not been isolated from any Antarctic pinniped species, leaving the confirmation of infection pending.

  1. Brucella Intracellular Life Relies on the Transmembrane Protein CD98 Heavy Chain.

    Science.gov (United States)

    Keriel, Anne; Botella, Eric; Estrach, Soline; Bragagnolo, Gabriel; Vergunst, Annette C; Feral, Chloe C; O'Callaghan, David

    2015-06-01

    Brucella are intracellular bacterial pathogens that use a type IV secretion system (T4SS) to escape host defenses and create a niche in which they can multiply. Although the importance of Brucella T4SS is clear, little is known about its interactions with host cell structures. In this study, we identified the eukaryotic protein CD98hc as a partner for Brucella T4SS subunit VirB2. This transmembrane glycoprotein is involved in amino acid transport, modulation of integrin signaling, and cell-to-cell fusion. Knockdown of CD98hc expression in HeLa cells demonstrated that it is essential for Brucella infection. Using knockout dermal fibroblasts, we confirmed its role for Brucella but found that it is not required for Salmonella infection. CD98hc transiently accumulates around the bacteria during the early phases of infection and is required for both optimal bacterial uptake and intracellular multiplication of Brucella. These results provide new insights into the complex interplay between Brucella and its host. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  2. Brucella placentitis and seroprevalence in northern fur seals ( Callorhinus ursinus) of the Pribilof Islands, Alaska.

    Science.gov (United States)

    Duncan, Colleen G; Tiller, Rebekah; Mathis, Demetrius; Stoddard, Robyn; Kersh, Gilbert J; Dickerson, Bobette; Gelatt, Tom

    2014-07-01

    Brucella species infect a wide range of hosts with a broad spectrum of clinical manifestations. In mammals, one of the most significant consequences of Brucella infection is reproductive failure. There is evidence of Brucella exposure in many species of marine mammals, but the outcome of infection is often challenging to determine. The eastern Pacific stock of northern fur seals (NFSs, Callorhinus ursinus) has declined significantly, spawning research into potential causes for this trend, including investigation into reproductive health. The objective of the current study was to determine if NFSs on St. Paul Island, Alaska have evidence of Brucella exposure or infection. Archived DNA extracted from placentas ( n = 119) and serum ( n = 40) samples were available for testing by insertion sequence (IS) 711 polymerase chain reaction (PCR) and the Brucella microagglutination test (BMAT), respectively. As well, placental tissue was available for histologic examination. Six (5%) placentas were positive by PCR, and a single animal had severe placentitis. Multilocus variable number tandem repeat analysis profiles were highly clustered and closely related to other Brucella pinnipedialis isolates. A single animal was positive on BMAT, and 12 animals had titers within the borderline range; 1 borderline animal was positive by PCR on serum. The findings suggest that NFSs on the Pribilof Islands are exposed to Brucella and that the organism has the ability to cause severe placental disease. Given the population trend of the NFS, and the zoonotic nature of this pathogen, further investigation into the epidemiology of this disease is recommended.

  3. Detection and differentiation of the six Brucella species by polymerase chain reaction.

    Science.gov (United States)

    Sifuentes-Rincón, A M; Revol, A; Barrera-Saldaña, H A

    1997-11-01

    Brucelosis is a severe acute febrile disease caused by bacteria of the genus Brucella. Its current diagnosis is based on clinical observations that may be complemented by serology and microbiological culture tests; however, the former is limited in sensitivity and specificity, the latter is time consuming. To improve brucelosis diagnosis we developed a test which is specific and sensitive and is capable of differentiating the six species of Brucella. Four primers were designed from B. abortus sequences at the well-conserved Omp2 locus that are able to amplify the DNAs of all six species of Brucella. Our test detected all six species of Brucella. Their differentiation resulted directly from differences in the amplification patterns or was achieved indirectly using a RFLP present in one of the PCR products. The sensitivity and specificity of the new test were then determined; it was applied successfully in confirming the diagnosis of a patient whose clinical history and serology indicated infection with Brucella. The results make possible the use of a PCR test for Brucella detection and differentiation without relying on the measurement of the antibodies or microorganism culture. Our first results showed that the PCR test can confirm the presence of Brucella in blood samples of infected patients.

  4. Bison PRNP genotyping and potential association with Brucella spp. seroprevalence

    Science.gov (United States)

    Seabury, C.M.; Halbert, N.D.; Gogan, P.J.P.; Templeton, J.W.; Derr, J.N.

    2005-01-01

    The implication that host cellular prion protein (PrPC) may function as a cell surface receptor and/or portal protein for Brucella abortus in mice prompted an evaluation of nucleotide and amino acid variation within exon 3 of the prion protein gene (PRNP) for six US bison populations. A non-synonymous single nucleotide polymorphism (T50C), resulting in the predicted amino acid replacement M17T (Met ??? Thr), was identified in each population. To date, no variation (T50: Met) has been detected at the corresponding exon 3 nucleotide and/or amino acid position for domestic cattle. Notably, 80% (20 of 25) of the Yellowstone National Park bison possessing the C/C genotype were Brucella spp. seropositive, representing a significant (P = 0.021) association between seropositivity and the C/C genotypic class. Moreover, significant differences in the distribution of PRNP exon 3 alleles and genotypes were detected between Yellowstone National Park bison and three bison populations that were either founded from seronegative stock or previously subjected to test-and-slaughter management to eradicate brucellosis. Unlike domestic cattle, no indel polymorphisms were detected within the corresponding regions of the putative bison PRNP promoter, intron 1, octapeptide repeat region or 3???-untranslated region for any population examined. This study provides the first evidence of a potential association between nucleotide variation within PRNP exon 3 and the presence of Brucella spp. antibodies in bison, implicating PrPC in the natural resistance of bison to brucellosis infection. ?? 2005 International Society for Animal Genetics.

  5. The role of 'atypical' Brucella in amphibians: are we facing novel emerging pathogens?

    Science.gov (United States)

    Mühldorfer, K; Wibbelt, G; Szentiks, C A; Fischer, D; Scholz, H C; Zschöck, M; Eisenberg, T

    2017-01-01

    To discuss together the novel cases of Brucella infections in frogs with the results of published reports to extend our current knowledge on 'atypical' brucellae isolated from amphibians and to discuss the challenges we face on this extraordinary emerging group of pathogens. Since our first description, an additional 14 isolates from four different frog species were collected. Novel isolates and a subset of Brucella isolates previously cultured from African bullfrogs were characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), Fourier transform-infrared (FT-IR) spectroscopy and broth microdilution susceptibility testing. MALDI-TOF MS worked very efficiently for an accurate bacterial identification to the genus level. Within the cluster analysis, 'atypical' brucellae grouped distant from Brucella melitensis and were even more separated by FT-IR spectroscopy with respect to their geographical origin. Minimum inhibitory concentrations of 14 antimicrobial substances are provided as baseline data on antimicrobial susceptibility. The case history of Brucella infections in amphibians reveals a variety of pathologies ranging from localized manifestations to systemic infections. Some isolates seem to be capable of causing high mortality in zoological exhibitions putting higher demands on the management of endangered frog species. There is considerable risk in overlooking and misidentifying 'atypical' Brucella in routine diagnostics. Brucella have only recently been described in cold-blooded vertebrates. Their presence in frog species native to Africa, America and Australia indicates a more common occurrence in amphibians than previously thought. This study provides an extensive overview of amphibian brucellae by highlighting the main features of their clinical significance, diagnosis and zoonotic potential. © 2016 The Society for Applied Microbiology.

  6. Comparative Genomics of Early-Diverging Brucella Strains Reveals a Novel Lipopolysaccharide Biosynthesis Pathway

    Science.gov (United States)

    Wattam, Alice R.; Inzana, Thomas J.; Williams, Kelly P.; Mane, Shrinivasrao P.; Shukla, Maulik; Almeida, Nalvo F.; Dickerman, Allan W.; Mason, Steven; Moriyón, Ignacio; O’Callaghan, David; Whatmore, Adrian M.; Sobral, Bruno W.; Tiller, Rebekah V.; Hoffmaster, Alex R.; Frace, Michael A.; De Castro, Cristina; Molinaro, Antonio; Boyle, Stephen M.; De, Barun K.; Setubal, João C.

    2012-01-01

    ABSTRACT Brucella species are Gram-negative bacteria that infect mammals. Recently, two unusual strains (Brucella inopinata BO1T and B. inopinata-like BO2) have been isolated from human patients, and their similarity to some atypical brucellae isolated from Australian native rodent species was noted. Here we present a phylogenomic analysis of the draft genome sequences of BO1T and BO2 and of the Australian rodent strains 83-13 and NF2653 that shows that they form two groups well separated from the other sequenced Brucella spp. Several important differences were noted. Both BO1T and BO2 did not agglutinate significantly when live or inactivated cells were exposed to monospecific A and M antisera against O-side chain sugars composed of N-formyl-perosamine. While BO1T maintained the genes required to synthesize a typical Brucella O-antigen, BO2 lacked many of these genes but still produced a smooth LPS (lipopolysaccharide). Most missing genes were found in the wbk region involved in O-antigen synthesis in classic smooth Brucella spp. In their place, BO2 carries four genes that other bacteria use for making a rhamnose-based O-antigen. Electrophoretic, immunoblot, and chemical analyses showed that BO2 carries an antigenically different O-antigen made of repeating hexose-rich oligosaccharide units that made the LPS water-soluble, which contrasts with the homopolymeric O-antigen of other smooth brucellae that have a phenol-soluble LPS. The results demonstrate the existence of a group of early-diverging brucellae with traits that depart significantly from those of the Brucella species described thus far. PMID:22930339

  7. Marine Mammal Brucella Reference Strains Are Attenuated in a BALB/c Mouse Model.

    Science.gov (United States)

    Nymo, Ingebjørg H; Arias, Maykel A; Pardo, Julián; Álvarez, María Pilar; Alcaraz, Ana; Godfroid, Jacques; Jiménez de Bagüés, María Pilar

    2016-01-01

    Brucellosis is a zoonosis of worldwide distribution with numerous animal host species. Since the novel isolation of Brucella spp. from marine mammals in 1994 the bacteria have been isolated from various marine mammal hosts. The marine mammal reference strains Brucella pinnipedialis 12890 (harbour seal, Phoca vitulina) and Brucella ceti 12891 (harbour porpoise, Phocoena phocoena) were included in genus Brucella in 2007, however, their pathogenicity in the mouse model is pending. Herein this is evaluated in BALB/c mice with Brucella suis 1330 as a control. Both marine mammal strains were attenuated, however, B. ceti was present at higher levels than B. pinnipedialis in blood, spleen and liver throughout the infection, in addition B. suis and B. ceti were isolated from brains and faeces at times with high levels of bacteraemia. In B. suis-infected mice serum cytokines peaked at day 7. In B. pinnipedialis-infected mice, levels were similar, but peaked predominantly at day 3 and an earlier peak in spleen weight likewise implied an earlier response. The inflammatory response induced pathology in the spleen and liver. In B. ceti-infected mice, most serum cytokine levels were comparable to those in uninfected mice, consistent with a limited inflammatory response, which also was indicated by restricted spleen and liver pathology. Specific immune responses against all three strains were detected in vitro after stimulation of splenocytes from infected mice with the homologous heat-killed brucellae. Antibody responses in vivo were also induced by the three brucellae. The immunological pattern of B. ceti in combination with persistence in organs and limited pathology has heretofore not been described for other brucellae. These two marine mammal wildtype strains show an attenuated pattern in BALB/c mice only previously described for Brucella neotomea.

  8. ABrucellaspp. Isolate from a Pac-Man Frog (Ceratophrys ornata) Reveals Characteristics Departing from Classical Brucellae.

    Science.gov (United States)

    Soler-Lloréns, Pedro F; Quance, Chris R; Lawhon, Sara D; Stuber, Tod P; Edwards, John F; Ficht, Thomas A; Robbe-Austerman, Suelee; O'Callaghan, David; Keriel, Anne

    2016-01-01

    Brucella are highly infectious bacterial pathogens responsible for brucellosis, a frequent worldwide zoonosis. The Brucella genus has recently expanded from 6 to 11 species, all of which were associated with mammals; The natural host range recently expanded to amphibians after some reports of atypical strains from frogs. Here we describe the first in depth phenotypic and genetic characterization of a Brucella strains isolated from a frog. Strain B13-0095 was isolated from a Pac-Man frog ( Ceratophyrus ornate ) at a veterinary hospital in Texas and was initially misidentified as Ochrobactrum anthropi . We found that B13-0095 belongs to a group of early-diverging brucellae that includes Brucella inopinata strain BO1 and the B. inopinata -like strain BO2, with traits that depart significantly from those of the "classical" Brucella spp. Analysis of B13-0095 genome sequence revealed several specific features that suggest that this isolate represents an intermediate between a soil associated ancestor and the host adapted "classical" species. Like strain BO2, B13-0095 does not possess the genes required to produce the perosamine based LPS found in classical Brucella , but has a set of genes that could encode a rhamnose based O-antigen. Despite this, B13-0095 has a very fast intracellular replication rate in both epithelial cells and macrophages. Finally, another major finding in this study is the bacterial motility observed for strains B13-0095, BO1, and BO2, which is remarkable for this bacterial genus. This study thus highlights several novel characteristics in strains belonging to an emerging group within the Brucella genus. Accurate identification tools for such atypical Brucella isolates and careful evaluation of their zoonotic potential, are urgently required.

  9. Incidence of Brucella species in marine mammals of the German North Sea.

    Science.gov (United States)

    Prenger-Berninghoff, E; Siebert, U; Stede, M; König, A; Weiss, R; Baljer, G

    2008-08-19

    In this study, organ samples from 426 common seals Phoca vitulina, 298 harbour porpoises Phocoena phocoena, 34 grey seals Halichoerus grypus and 10 other marine mammals were assessed for the presence of Brucella species. Forty-seven common seals, 2 harbour porpoises and 1 grey seal were found to be positive for these bacteria. A total of 91 Brucella strains were successfully isolated, due to the fact that Brucella spp. were found in more than one organ sample in 15 animals. The primary organ in which the bacteria were present was the lung. In addition, 2 strains were isolated from lungworms (Parafilaroides spp.). Forty-nine of the isolated strains were selected for further analysis using conventional phenotyping methods. Molecular characterisation was carried out by analysing the IS711 and omp2 loci. With respect to the distribution of the IS711 loci in the genome, the 49 field isolates differed strongly from the terrestrial Brucella species and marginally from the marine Brucella reference strain NCTC12890. Based on the results of the PCR restriction fragment length polymorphism (PCR-RFLP) investigation of the omp2 locus, the majority of the Brucella field isolates were classified as B. pinnipediae, recently proposed B. pinnipedialis, possessing 1 omp2a gene and 1 omp2b gene. Two field isolates revealed the presence of 2 omp2a genes, as has been described for Brucella ovis. To our knowledge, these results confirm for the first time the presence of Brucella species in the marine mammal population of the German North Sea. These findings highlight the need for additional research on the relevance of these Brucella species for marine hosts and their environment.

  10. First isolation and characterization of Brucella microti from wild boar.

    Science.gov (United States)

    Rónai, Zsuzsanna; Kreizinger, Zsuzsa; Dán, Ádám; Drees, Kevin; Foster, Jeffrey T; Bányai, Krisztián; Marton, Szilvia; Szeredi, Levente; Jánosi, Szilárd; Gyuranecz, Miklós

    2015-07-11

    Brucella microti was first isolated from common vole (Microtus arvalis) in the Czech Republic in Central Europe in 2007. As B. microti is the only Brucella species known to live in soil, its distribution, ecology, zoonotic potential, and genomic organization is of particular interest. The present paper is the first to report the isolation of B. microti from a wild boar (Sus scrofa), which is also the first isolation of this bacterial species in Hungary. The B. microti isolate was cultured, after enrichment in Brucella-selective broth, from the submandibular lymph node of a female wild boar that was taken by hunters in Hungary near the Austrian border in September 2014. Histological and immunohistological examinations of the lymph node sections with B. abortus-, B. suis- and B. canis-specific sera gave negative results. The isolate did not require CO2 for growth, was oxidase, catalase, and urease positive, H2S negative, grew well in the presence of 20 μg/ml basic fuchsin and thionin, and had brownish pigmentation after three days of incubation. It gave strong positive agglutination with anti-A and anti-M but had a negative reaction with anti-R monospecific sera. The API 20 NE test identified it as Ochrobactrum anthropi with 99.9% identity, and it showed B. microti-specific banding pattern in the Bruce- and Suis-ladder multiplex PCR systems. Whole genome re-sequencing identified 30 SNPs in orthologous loci when compared to the B. microti reference genome available in GenBank, and the MLVA analysis yielded a unique profile. Given that the female wild boar did not develop any clinical disease, we hypothesize that this host species only harboured the bacterium, serving as a possible reservoir capable of maintaining and spreading this pathogen. The infectious source could have been either a rodent, a carcass that had been eaten or infection occurred via the boar rooting in soil. The low number of discovered SNPs suggests an unexpectedly high level of genetic homogeneity

  11. Brucella Endocarditis as a Late Onset Complication of Brucellosis

    Directory of Open Access Journals (Sweden)

    Panagiotis Andriopoulos

    2015-01-01

    Full Text Available Brucella endocarditis (BE is a rare but life threatening complication of brucellosis. We present a case report of a patient with relapsing brucellosis complicated with aortic valve endocarditis. The patient underwent valve replacement and required prolonged antibiotic treatment because of rupture of the noncoronary leaflet and development of congestive heart failure. Since the onset of endocarditis in patients with brucellosis is not known, proper follow-up is required in order to identify any late onset complications, especially in endemic areas.

  12. Molecular detection and characterization of Brucella species in raw informally marketed milk from Uganda

    OpenAIRE

    Hoffman, Tove; Rock, Kim; Mugizi, Denis Rwabiita; Muradrasoli, Shaman; Lindahl-Rajala, Elisabeth; Erume, Joseph; Magnusson, Ulf; Lundkvist, ?ke; Boqvist, Sofia

    2016-01-01

    This study identified and characterized Brucella species in the informal milk chain in Uganda. A total of 324 cattle bulk milk samples were screened for the genus Brucella by real-time PCR with primers targeting the bcsp31 gene and further characterized by the omp25 gene. Of the samples tested, 6.5% were positive for Brucella species. In the omp25 phylogeny, the study sequences were found to form a separate clade within the branch containing B. abortus sequences. The study shows that informal...

  13. Genomic comparisons of Brucella spp. and closely related bacteria using base compositional and proteome based methods

    Science.gov (United States)

    2010-01-01

    Background Classification of bacteria within the genus Brucella has been difficult due in part to considerable genomic homogeneity between the different species and biovars, in spite of clear differences in phenotypes. Therefore, many different methods have been used to assess Brucella taxonomy. In the current work, we examine 32 sequenced genomes from genus Brucella representing the six classical species, as well as more recently described species, using bioinformatical methods. Comparisons were made at the level of genomic DNA using oligonucleotide based methods (Markov chain based genomic signatures, genomic codon and amino acid frequencies based comparisons) and proteomes (all-against-all BLAST protein comparisons and pan-genomic analyses). Results We found that the oligonucleotide based methods gave different results compared to that of the proteome based methods. Differences were also found between the oligonucleotide based methods used. Whilst the Markov chain based genomic signatures grouped the different species in genus Brucella according to host preference, the codon and amino acid frequencies based methods reflected small differences between the Brucella species. Only minor differences could be detected between all genera included in this study using the codon and amino acid frequencies based methods. Proteome comparisons were found to be in strong accordance with current Brucella taxonomy indicating a remarkable association between gene gain or loss on one hand and mutations in marker genes on the other. The proteome based methods found greater similarity between Brucella species and Ochrobactrum species than between species within genus Agrobacterium compared to each other. In other words, proteome comparisons of species within genus Agrobacterium were found to be more diverse than proteome comparisons between species in genus Brucella and genus Ochrobactrum. Pan-genomic analyses indicated that uptake of DNA from outside genus Brucella appears to be

  14. Brucella and Coxiella; if you don't look, you don't find.

    Science.gov (United States)

    Lambourne, Jonathan R; Brooks, Tim

    2015-02-01

    Brucella and Coxiella are similar; both are obligate intracellular, zoonotic pathogens with a broad geographic distribution. Infection in animals is usually asymptomatic, but causes fetal loss and therefore has significant economic impact. Human infection may be asymptomatic or give rise to either organ-specific or multi-system disease. Organism culture is challenging for Coxiella and can lack sensitivity for Brucella. Therefore, infection is most commonly diagnosed by serology, but this may be negative in early infection and serology results may be challenging to interpret. Both Brucella and Coxiella are typically susceptible to a wide range of antimicrobials, but long courses may be needed. © 2015 Royal College of Physicians.

  15. Triad of infective endocarditis, splenic abscess, and septicemia caused by Brucella melitensis

    Directory of Open Access Journals (Sweden)

    Shashank Purwar

    2017-01-01

    Full Text Available A 40-year-old farmer from the district of North Karnataka who had received treatment for high fever of 8 days duration was admitted with fever, dyspnea, and poor general condition. Ultrasonography and echocardiogram revealed multiple splenic abscesses, vegetation on atrioventricular valve, aortic regurgitation (Grade I–II, and mitral valve regurgitation (Grade II–III, respectively. Brucella melitensis was detected in blood culture, and high titers of IgM and IgG anti-Brucella antibodies were observed in Brucella specific serological tests. The patient developed fulminant septicemia and succumbed due to multi-organ failure.

  16. African Lineage Brucella melitensis Isolates from Omani Livestock

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    Jeffrey T. Foster

    2018-01-01

    Full Text Available Brucellosis is a common livestock disease in the Middle East and North Africa, but remains poorly described in the region both genetically and epidemiologically. Traditionally found in goats and sheep, Brucella melitensis is increasingly recognized as infecting camels. Most studies of brucellosis in camels to date have focused on serological surveys, providing only limited understanding of the molecular epidemiology of circulating strains. We genotyped B. melitensis isolates from Omani camels using whole genome SNP assays and VNTRs to provide context for regional brucellosis cases. We identified a lineage of B. melitensis circulating in camels as well as in goats, sheep, and cattle in Oman. This lineage is genetically distinct from most genotypes from the Arabian Peninsula and from isolates from much of the rest of the Middle East. We then developed diagnostic assays that rapidly identify strains from this lineage. In analyses of genotypes from throughout the region, Omani isolates were genetically most closely related to strains from brucellosis cases in humans and livestock in North Africa. Our findings suggest an African origin for B. melitensis in Oman that has likely occurred through the trade of infected livestock. Moreover, African lineages of B. melitensis appear to be undersampled and consequently are underrepresented in genetic databases for Brucella. As we begin to more fully understand global genomic diversity of B. melitensis, finding and characterizing these unique but widespread lineages is essential. We predict that increased sampling of humans and livestock in Africa will reveal little known diversity in this important zoonotic pathogen.

  17. Brucella ceti infection in dolphins from the Western Mediterranean sea.

    Science.gov (United States)

    Isidoro-Ayza, Marcos; Ruiz-Villalobos, Nazareth; Pérez, Lola; Guzmán-Verri, Caterina; Muñoz, Pilar M; Alegre, Fernando; Barberán, Montserrat; Chacón-Díaz, Carlos; Chaves-Olarte, Esteban; González-Barrientos, Rocio; Moreno, Edgardo; Blasco, José María; Domingo, Mariano

    2014-09-17

    Brucella ceti infections have been increasingly reported in cetaceans. Brucellosis in these animals is associated with meningoencephalitis, abortion, discospondylitis', subcutaneous abscesses, endometritis and other pathological conditions B. ceti infections have been frequently described in dolphins from both, the Atlantic and Pacific Oceans. In the Mediterranean Sea, only two reports have been made: one from the Italian Tyrrhenian Sea and the other from the Adriatic Sea. We describe the clinical and pathological features of three cases of B. ceti infections in three dolphins stranded in the Mediterranean Catalonian coast. One striped dolphin had neurobrucellosis, showing lethargy, incoordination and lateral swimming due to meningoencephalitis, A B. ceti infected bottlenose dolphin had discospondylitis, and another striped dolphin did not show clinical signs or lesions related to Brucella infection. A detailed characterization of the three B. ceti isolates was performed by bacteriological, molecular, protein and fatty acid analyses. All the B. ceti strains originating from Mediterranean dolphins cluster together in a distinct phylogenetic clade, close to that formed by B. ceti isolates from dolphins inhabiting the Atlantic Ocean. Our study confirms the severity of pathological signs in stranded dolphins and the relevance of B. ceti as a pathogen in the Mediterranean Sea.

  18. Assay dependence of Brucella antibody prevalence in a declining Alaskan harbor seal (Phoca vitulina population

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    Hueffer Karsten

    2013-01-01

    Full Text Available Abstract Background Brucella is a group of bacteria that causes brucellosis, which can affect population health and reproductive success in many marine mammals. We investigated the serological prevalence of antibodies against Brucella bacteria in a declining harbor seal population in Glacier Bay National Park, Alaska. Results Prevalence ranged from 16 to 74 percent for those tests detecting antibodies, indicating that harbor seals in Glacier Bay have been exposed to Brucella bacteria. However, the actual level of serological prevalence could not be determined because results were strongly assay-dependent. Conclusions This study reinforces the need to carefully consider assay choice when comparing different studies on the prevalence of anti–Brucella antibodies in pinnipeds and further highlights the need for species- or taxon-specific assay validation for both pathogen and host species.

  19. Ontology-based Brucella vaccine literature indexing and systematic analysis of gene-vaccine association network

    Science.gov (United States)

    2011-01-01

    Background Vaccine literature indexing is poorly performed in PubMed due to limited hierarchy of Medical Subject Headings (MeSH) annotation in the vaccine field. Vaccine Ontology (VO) is a community-based biomedical ontology that represents various vaccines and their relations. SciMiner is an in-house literature mining system that supports literature indexing and gene name tagging. We hypothesize that application of VO in SciMiner will aid vaccine literature indexing and mining of vaccine-gene interaction networks. As a test case, we have examined vaccines for Brucella, the causative agent of brucellosis in humans and animals. Results The VO-based SciMiner (VO-SciMiner) was developed to incorporate a total of 67 Brucella vaccine terms. A set of rules for term expansion of VO terms were learned from training data, consisting of 90 biomedical articles related to Brucella vaccine terms. VO-SciMiner demonstrated high recall (91%) and precision (99%) from testing a separate set of 100 manually selected biomedical articles. VO-SciMiner indexing exhibited superior performance in retrieving Brucella vaccine-related papers over that obtained with MeSH-based PubMed literature search. For example, a VO-SciMiner search of "live attenuated Brucella vaccine" returned 922 hits as of April 20, 2011, while a PubMed search of the same query resulted in only 74 hits. Using the abstracts of 14,947 Brucella-related papers, VO-SciMiner identified 140 Brucella genes associated with Brucella vaccines. These genes included known protective antigens, virulence factors, and genes closely related to Brucella vaccines. These VO-interacting Brucella genes were significantly over-represented in biological functional categories, including metabolite transport and metabolism, replication and repair, cell wall biogenesis, intracellular trafficking and secretion, posttranslational modification, and chaperones. Furthermore, a comprehensive interaction network of Brucella vaccines and genes were

  20. Ontology-based Brucella vaccine literature indexing and systematic analysis of gene-vaccine association network.

    Science.gov (United States)

    Hur, Junguk; Xiang, Zuoshuang; Feldman, Eva L; He, Yongqun

    2011-08-26

    Vaccine literature indexing is poorly performed in PubMed due to limited hierarchy of Medical Subject Headings (MeSH) annotation in the vaccine field. Vaccine Ontology (VO) is a community-based biomedical ontology that represents various vaccines and their relations. SciMiner is an in-house literature mining system that supports literature indexing and gene name tagging. We hypothesize that application of VO in SciMiner will aid vaccine literature indexing and mining of vaccine-gene interaction networks. As a test case, we have examined vaccines for Brucella, the causative agent of brucellosis in humans and animals. The VO-based SciMiner (VO-SciMiner) was developed to incorporate a total of 67 Brucella vaccine terms. A set of rules for term expansion of VO terms were learned from training data, consisting of 90 biomedical articles related to Brucella vaccine terms. VO-SciMiner demonstrated high recall (91%) and precision (99%) from testing a separate set of 100 manually selected biomedical articles. VO-SciMiner indexing exhibited superior performance in retrieving Brucella vaccine-related papers over that obtained with MeSH-based PubMed literature search. For example, a VO-SciMiner search of "live attenuated Brucella vaccine" returned 922 hits as of April 20, 2011, while a PubMed search of the same query resulted in only 74 hits. Using the abstracts of 14,947 Brucella-related papers, VO-SciMiner identified 140 Brucella genes associated with Brucella vaccines. These genes included known protective antigens, virulence factors, and genes closely related to Brucella vaccines. These VO-interacting Brucella genes were significantly over-represented in biological functional categories, including metabolite transport and metabolism, replication and repair, cell wall biogenesis, intracellular trafficking and secretion, posttranslational modification, and chaperones. Furthermore, a comprehensive interaction network of Brucella vaccines and genes were identified. The asserted

  1. Ontology-based Brucella vaccine literature indexing and systematic analysis of gene-vaccine association network

    Directory of Open Access Journals (Sweden)

    Xiang Zuoshuang

    2011-08-01

    Full Text Available Abstract Background Vaccine literature indexing is poorly performed in PubMed due to limited hierarchy of Medical Subject Headings (MeSH annotation in the vaccine field. Vaccine Ontology (VO is a community-based biomedical ontology that represents various vaccines and their relations. SciMiner is an in-house literature mining system that supports literature indexing and gene name tagging. We hypothesize that application of VO in SciMiner will aid vaccine literature indexing and mining of vaccine-gene interaction networks. As a test case, we have examined vaccines for Brucella, the causative agent of brucellosis in humans and animals. Results The VO-based SciMiner (VO-SciMiner was developed to incorporate a total of 67 Brucella vaccine terms. A set of rules for term expansion of VO terms were learned from training data, consisting of 90 biomedical articles related to Brucella vaccine terms. VO-SciMiner demonstrated high recall (91% and precision (99% from testing a separate set of 100 manually selected biomedical articles. VO-SciMiner indexing exhibited superior performance in retrieving Brucella vaccine-related papers over that obtained with MeSH-based PubMed literature search. For example, a VO-SciMiner search of "live attenuated Brucella vaccine" returned 922 hits as of April 20, 2011, while a PubMed search of the same query resulted in only 74 hits. Using the abstracts of 14,947 Brucella-related papers, VO-SciMiner identified 140 Brucella genes associated with Brucella vaccines. These genes included known protective antigens, virulence factors, and genes closely related to Brucella vaccines. These VO-interacting Brucella genes were significantly over-represented in biological functional categories, including metabolite transport and metabolism, replication and repair, cell wall biogenesis, intracellular trafficking and secretion, posttranslational modification, and chaperones. Furthermore, a comprehensive interaction network of Brucella

  2. Brucella canis: inquéritos sorológico e bacteriológico em população felina Brucella canis: serological and bacteriological surveys in the feline population

    Directory of Open Access Journals (Sweden)

    Maria Helena Matiko Akao Larsson

    1984-02-01

    Full Text Available De 134 soros de felinos domésticos examinados pela prova de soroaglutinação lenta em tubos, 4 (3% foram positivos para Brucella canis, todos com título igual a 100. Não se obteve êxito na tentativa de isolamento de Brucella canis através de hemocultura desses animais.Of the 134 feline sera tested by tube agglutination test, 4 (3% were positive for Brucella canis antibodies, all with titer 100. It was not possible to isolate Brucella canis by blood culture in the case of these animals.

  3. Host-Brucella interactions and the Brucella genome as tools for subunit antigen discovery and immunization against brucellosis

    Science.gov (United States)

    Gomez, Gabriel; Adams, Leslie G.; Rice-Ficht, Allison; Ficht, Thomas A.

    2013-01-01

    Vaccination is the most important approach to counteract infectious diseases. Thus, the development of new and improved vaccines for existing, emerging, and re-emerging diseases is an area of great interest to the scientific community and general public. Traditional approaches to subunit antigen discovery and vaccine development lack consideration for the critical aspects of public safety and activation of relevant protective host immunity. The availability of genomic sequences for pathogenic Brucella spp. and their hosts have led to development of systems-wide analytical tools that have provided a better understanding of host and pathogen physiology while also beginning to unravel the intricacies at the host-pathogen interface. Advances in pathogen biology, host immunology, and host-agent interactions have the potential to serve as a platform for the design and implementation of better-targeted antigen discovery approaches. With emphasis on Brucella spp., we probe the biological aspects of host and pathogen that merit consideration in the targeted design of subunit antigen discovery and vaccine development. PMID:23720712

  4. First detection of Brucella canis infections in a breeding kennel in Austria.

    Science.gov (United States)

    Hofer, Erwin; Bag, Zolt N; Revilla-Fern Ndez, Sandra; Melzer, Falk; Tomaso, Herbert; L Pez-Go I, Igniacio; Fasching, Gerhard; Schmoll, Friedrich

    2012-10-01

    Brucella canis occurs almost worldwide and is a potential danger to the health of dogs and humans. The pathogen was detected in the placenta and fetuses of a Standard Poodle by direct culture and immunohistochemistry. Further, Brucellae were also isolated from the blood samples of two asymptomatic female Medium Poodles. The isolates were identified as B. canis by conventional microbiological methods and a novel Bruce-ladder multiplex PCR. Genotyping was performed by multiple locus variable number tandem repeats analysis (MLVA).

  5. Polymorphism in Brucella strains detected by studying distribution of two short repetitive DNA elements.

    OpenAIRE

    Mercier, E; Jumas-Bilak, E; Allardet-Servent, A; O'Callaghan, D; Ramuz, M

    1996-01-01

    Thirty-four Brucella reference or field strains representing all the species and biovars were studied by repetitive element sequence-based PCR, a PCR using primers complementary to two enterobacterial short repetitive sequences: repetitive extragenic palindromic and enterobacterial repetitive intergenic consensus sequences. All the stains showed a positive amplification, suggesting that the Brucella genome contains such sequences. Repetitive extragenic palindromic PCR was less discriminating ...

  6. Differential phenotyping of Brucella species using a newly developed semi-automated metabolic system

    Science.gov (United States)

    2010-01-01

    Background A commercial biotyping system (Taxa Profile™, Merlin Diagnostika) testing the metabolization of various substrates by bacteria was used to determine if a set of phenotypic features will allow the identification of members of the genus Brucella and their differentiation into species and biovars. Results A total of 191 different amines, amides, amino acids, other organic acids and heterocyclic and aromatic substrates (Taxa Profile™ A), 191 different mono-, di-, tri- and polysaccharides and sugar derivates (Taxa Profile™ C) and 95 amino peptidase- and protease-reactions, 76 glycosidase-, phosphatase- and other esterase-reactions, and 17 classic reactions (Taxa Profile™ E) were tested with the 23 reference strains representing the currently known species and biovars of Brucella and a collection of 60 field isolates. Based on specific and stable reactions a 96-well "Brucella identification and typing" plate (Micronaut™) was designed and re-tested in 113 Brucella isolates and a couple of closely related bacteria. Brucella species and biovars revealed characteristic metabolic profiles and each strain showed an individual pattern. Due to their typical metabolic profiles a differentiation of Brucella isolates to the species level could be achieved. The separation of B. canis from B. suis bv 3, however, failed. At the biovar level, B. abortus bv 4, 5, 7 and B. suis bv 1-5 could be discriminated with a specificity of 100%. B. melitensis isolates clustered in a very homogenous group and could not be resolved according to their assigned biovars. Conclusions The comprehensive testing of metabolic activity allows cluster analysis within the genus Brucella. The biotyping system developed for the identification of Brucella and differentiation of its species and biovars may replace or at least complement time-consuming tube testing especially in case of atypical strains. An easy to handle identification software facilitates the applicability of the Micronaut

  7. Caspase-2 mediated apoptotic and necrotic murine macrophage cell death induced by rough Brucella abortus.

    Directory of Open Access Journals (Sweden)

    Fang Chen

    Full Text Available Brucella species are Gram-negative, facultative intracellular bacteria that cause zoonotic brucellosis. Survival and replication inside macrophages is critical for establishment of chronic Brucella infection. Virulent smooth B. abortus strain 2308 inhibits programmed macrophage cell death and replicates inside macrophages. Cattle B. abortus vaccine strain RB51 is an attenuated rough, lipopolysaccharide O antigen-deficient mutant derived from smooth strain 2308. B. abortus rough mutant RA1 contains a single wboA gene mutation in strain 2308. Our studies demonstrated that live RB51 and RA1, but not strain 2308 or heat-killed Brucella, induced both apoptotic and necrotic cell death in murine RAW264.7 macrophages and bone marrow derived macrophages. The same phenomenon was also observed in primary mouse peritoneal macrophages from mice immunized intraperitoneally with vaccine strain RB51 using the same dose as regularly performed in protection studies. Programmed macrophage cell death induced by RB51 and RA1 was inhibited by a caspase-2 inhibitor (Z-VDVAD-FMK. Caspase-2 enzyme activation and cleavage were observed at the early infection stage in macrophages infected with RB51 and RA1 but not strain 2308. The inhibition of macrophage cell death promoted the survival of rough Brucella cells inside macrophages. The critical role of caspase-2 in mediating rough B. abortus induced macrophage cell death was confirmed using caspase-2 specific shRNA. The mitochondrial apoptosis pathway was activated in macrophages infected with rough B. abortus as demonstrated by increase in mitochondrial membrane permeability and the release of cytochrome c to cytoplasm in macrophages infected with rough Brucella. These results demonstrate that rough B. abortus strains RB51 and RA1 induce apoptotic and necrotic murine macrophage cell death that is mediated by caspase-2. The biological relevance of Brucella O antigen and caspase-2-mediated macrophage cell death in Brucella

  8. Caspase-2 mediated apoptotic and necrotic murine macrophage cell death induced by rough Brucella abortus.

    Science.gov (United States)

    Chen, Fang; He, Yongqun

    2009-08-28

    Brucella species are Gram-negative, facultative intracellular bacteria that cause zoonotic brucellosis. Survival and replication inside macrophages is critical for establishment of chronic Brucella infection. Virulent smooth B. abortus strain 2308 inhibits programmed macrophage cell death and replicates inside macrophages. Cattle B. abortus vaccine strain RB51 is an attenuated rough, lipopolysaccharide O antigen-deficient mutant derived from smooth strain 2308. B. abortus rough mutant RA1 contains a single wboA gene mutation in strain 2308. Our studies demonstrated that live RB51 and RA1, but not strain 2308 or heat-killed Brucella, induced both apoptotic and necrotic cell death in murine RAW264.7 macrophages and bone marrow derived macrophages. The same phenomenon was also observed in primary mouse peritoneal macrophages from mice immunized intraperitoneally with vaccine strain RB51 using the same dose as regularly performed in protection studies. Programmed macrophage cell death induced by RB51 and RA1 was inhibited by a caspase-2 inhibitor (Z-VDVAD-FMK). Caspase-2 enzyme activation and cleavage were observed at the early infection stage in macrophages infected with RB51 and RA1 but not strain 2308. The inhibition of macrophage cell death promoted the survival of rough Brucella cells inside macrophages. The critical role of caspase-2 in mediating rough B. abortus induced macrophage cell death was confirmed using caspase-2 specific shRNA. The mitochondrial apoptosis pathway was activated in macrophages infected with rough B. abortus as demonstrated by increase in mitochondrial membrane permeability and the release of cytochrome c to cytoplasm in macrophages infected with rough Brucella. These results demonstrate that rough B. abortus strains RB51 and RA1 induce apoptotic and necrotic murine macrophage cell death that is mediated by caspase-2. The biological relevance of Brucella O antigen and caspase-2-mediated macrophage cell death in Brucella pathogenesis and

  9. Brucella vulpis sp. nov., isolated from mandibular lymph nodes of red foxes (Vulpes vulpes).

    Science.gov (United States)

    Scholz, Holger C; Revilla-Fernández, Sandra; Al Dahouk, Sascha; Hammerl, Jens A; Zygmunt, Michel S; Cloeckaert, Axel; Koylass, Mark; Whatmore, Adrian M; Blom, Jochen; Vergnaud, Gilles; Witte, Angela; Aistleitner, Karin; Hofer, Erwin

    2016-05-01

    Two slow-growing, Gram-negative, non-motile, non-spore-forming, coccoid bacteria (strains F60T and F965), isolated in Austria from mandibular lymph nodes of two red foxes (Vulpes vulpes), were subjected to a polyphasic taxonomic analysis. In a recent study, both isolates were assigned to the genus Brucella but could not be attributed to any of the existing species. Hence, we have analysed both strains in further detail to determine their exact taxonomic position and genetic relatedness to other members of the genus Brucella. The genome sizes of F60T and F965 were 3 236 779 and 3 237 765 bp, respectively. Each genome consisted of two chromosomes, with a DNA G+C content of 57.2 %. A genome-to-genome distance of >80 %, an average nucleotide identity (ANI) of 97 % and an average amino acid identity (AAI) of 98 % compared with the type species Brucella melitensis confirmed affiliation to the genus. Remarkably, 5 % of the entire genetic information of both strains was of non-Brucella origin, including as-yet uncharacterized bacteriophages and insertion sequences as well as ABC transporters and other genes of metabolic function from various soil-living bacteria. Core-genome-based phylogenetic reconstructions placed the novel species well separated from all hitherto-described species of the genus Brucella, forming a long-branched sister clade to the classical species of Brucella. In summary, based on phenotypic and molecular data, we conclude that strains F60T and F965 are members of a novel species of the genus Brucella, for which the name Brucella vulpis sp. nov. is proposed, with the type strain F60T ( = BCCN 09-2T = DSM 101715T).

  10. Brucella discriminates between mouse dendritic cell subsets upon in vitro infection.

    Science.gov (United States)

    Papadopoulos, Alexia; Gagnaire, Aurélie; Degos, Clara; de Chastellier, Chantal; Gorvel, Jean-Pierre

    2016-01-01

    Brucella is a Gram-negative bacterium responsible for brucellosis, a worldwide re-emerging zoonosis. Brucella has been shown to infect and replicate within Granulocyte macrophage colony-stimulating factor (GMCSF) in vitro grown bone marrow-derived dendritic cells (BMDC). In this cell model, Brucella can efficiently control BMDC maturation. However, it has been shown that Brucella infection in vivo induces spleen dendritic cells (DC) migration and maturation. As DCs form a complex network composed by several subpopulations, differences observed may be due to different interactions between Brucella and DC subsets. Here, we compare Brucella interaction with several in vitro BMDC models. The present study shows that Brucella is capable of replicating in all the BMDC models tested with a high infection rate at early time points in GMCSF-IL15 DCs and Flt3l DCs. GMCSF-IL15 DCs and Flt3l DCs are more activated than the other studied DC models and consequently intracellular bacteria are not efficiently targeted to the ER replicative niche. Interestingly, GMCSF-DC and GMCSF-Flt3l DC response to infection is comparable. However, the key difference between these 2 models concerns IL10 secretion by GMCSF DCs observed at 48 h post-infection. IL10 secretion can explain the weak secretion of IL12p70 and TNFα in the GMCSF-DC model and the low level of maturation observed when compared to GMCSF-IL15 DCs and Flt3l DCs. These models provide good tools to understand how Brucella induce DC maturation in vivo and may lead to new therapeutic design using DCs as cellular vaccines capable of enhancing immune response against pathogens.

  11. Identification and effect decomposition of risk factors for Brucella contamination of raw whole milk in china.

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    Pengbo Ning

    Full Text Available BACKGROUND: Lack of clear risk factor identification is the main reason for the persistence of brucellosis infection in the Chinese population, and there has been little assessment of the factors contributing to Brucella contamination of raw whole milk. The purpose of this study was to identify risk factors affecting Brucella contamination of raw milk, and to evaluate effective measures for disease reduction in order to determine preventive strategies. METHODS AND FINDINGS: A nationwide survey was conducted and samples were obtained from 5211 cows corresponding to 25 sampling locations throughout 15 provinces in China. The prevalence of Brucella in the raw milk samples averaged 1.07% over the 15 Chinese provinces, while the prevalence of positive areas within these regions ranged from 0.23-3.84% among the nine provinces with positive samples. The survey examined factors that supposedly influence Brucella contamination of raw whole milk, such as management style, herd size, abortion rate, hygiene and disease control practices. A binary logistic regression analysis was carried out to determine the association between risk factors for Brucella and contamination of milk samples. Furthermore, a relative effect decomposition study was conducted to determine effective strategies for reducing the risk of Brucella contamination of raw whole milk. Our data indicate that disease prevention and control measures, abortion rate, and animal polyculture are the most important risk factors. Meanwhile, culling after quarantine was identified as an effective protective measure in the current Chinese dairy situation. CONCLUSIONS: These results indicate that, although there is a low risk of contamination of milk with Brucella nationwide in China, there are individual regions where contamination is a significant problem. Controlling three factors-culling after quarantine, maintaining a low abortion rate, and avoiding mixing groups of cattle and small ruminants

  12. Differential phenotyping of Brucella species using a newly developed semi-automated metabolic system

    Directory of Open Access Journals (Sweden)

    Appel Bernd

    2010-10-01

    Full Text Available Abstract Background A commercial biotyping system (Taxa Profile™, Merlin Diagnostika testing the metabolization of various substrates by bacteria was used to determine if a set of phenotypic features will allow the identification of members of the genus Brucella and their differentiation into species and biovars. Results A total of 191 different amines, amides, amino acids, other organic acids and heterocyclic and aromatic substrates (Taxa Profile™ A, 191 different mono-, di-, tri- and polysaccharides and sugar derivates (Taxa Profile™ C and 95 amino peptidase- and protease-reactions, 76 glycosidase-, phosphatase- and other esterase-reactions, and 17 classic reactions (Taxa Profile™ E were tested with the 23 reference strains representing the currently known species and biovars of Brucella and a collection of 60 field isolates. Based on specific and stable reactions a 96-well "Brucella identification and typing" plate (Micronaut™ was designed and re-tested in 113 Brucella isolates and a couple of closely related bacteria. Brucella species and biovars revealed characteristic metabolic profiles and each strain showed an individual pattern. Due to their typical metabolic profiles a differentiation of Brucella isolates to the species level could be achieved. The separation of B. canis from B. suis bv 3, however, failed. At the biovar level, B. abortus bv 4, 5, 7 and B. suis bv 1-5 could be discriminated with a specificity of 100%. B. melitensis isolates clustered in a very homogenous group and could not be resolved according to their assigned biovars. Conclusions The comprehensive testing of metabolic activity allows cluster analysis within the genus Brucella. The biotyping system developed for the identification of Brucella and differentiation of its species and biovars may replace or at least complement time-consuming tube testing especially in case of atypical strains. An easy to handle identification software facilitates the

  13. Development of a Selective Culture Medium for Primary Isolation of the Main Brucella Species▿

    OpenAIRE

    Miguel, María Jesús de; Marín, Clara M.; Muñoz, Pilar M.; Dieste, L.; Grilló, María Jesús; Blasco, José M

    2011-01-01

    Bacteriological diagnosis of brucellosis is performed by culturing animal samples directly on both Farrell medium (FM) and modified Thayer-Martin medium (mTM). However, despite inhibiting most contaminating microorganisms, FM also inhibits the growth of Brucella ovis and some B. melitensis and B. abortus strains. In contrast, mTM is adequate for growth of all Brucella species but only partially inhibitory for contaminants. Moreover, the performance of both culture media for isolating B. suis ...

  14. The Change of a Medically Important Genus: Worldwide Occurrence of Genetically Diverse Novel Brucella Species in Exotic Frogs.

    Science.gov (United States)

    Scholz, Holger C; Mühldorfer, Kristin; Shilton, Cathy; Benedict, Suresh; Whatmore, Adrian M; Blom, Jochen; Eisenberg, Tobias

    2016-01-01

    The genus Brucella comprises various species of both veterinary and human medical importance. All species are genetically highly related to each other, sharing intra-species average nucleotide identities (ANI) of > 99%. Infections occur among various warm-blooded animal species, marine mammals, and humans. Until recently, amphibians had not been recognized as a host for Brucella. In this study, however, we show that novel Brucella species are distributed among exotic frogs worldwide. Comparative recA gene analysis of 36 frog isolates from various continents and different frog species revealed an unexpected high genetic diversity, not observed among classical Brucella species. In phylogenetic reconstructions the isolates consequently formed various clusters and grouped together with atypical more distantly related brucellae, like B. inopinata, strain BO2, and Australian isolates from rodents, some of which were isolated as human pathogens. Of one frog isolate (10RB9215) the genome sequence was determined. Comparative genome analysis of this isolate and the classical Brucella species revealed additional genetic material, absent from classical Brucella species but present in Ochrobactrum, the closest genetic neighbor of Brucella, and in other soil associated genera of the Alphaproteobacteria. The presence of gene clusters encoding for additional metabolic functions, flanked by tRNAs and mobile genetic elements, as well as by bacteriophages is suggestive for a different ecology compared to classical Brucella species. Furthermore it suggests that amphibian isolates may represent a link between free living soil saprophytes and the pathogenic Brucella with a preferred intracellular habitat. We therefore assume that brucellae from frogs have a reservoir in soil and, in contrast to classical brucellae, undergo extensive horizontal gene transfer.

  15. Glutamate decarboxylase-dependent acid resistance in Brucella spp.: distribution and contribution to fitness under extremely acidic conditions.

    Science.gov (United States)

    Damiano, Maria Alessandra; Bastianelli, Daniela; Al Dahouk, Sascha; Köhler, Stephan; Cloeckaert, Axel; De Biase, Daniela; Occhialini, Alessandra

    2015-01-01

    Brucella is an expanding genus of major zoonotic pathogens, including at least 10 genetically very close species occupying a wide range of niches from soil to wildlife, livestock, and humans. Recently, we have shown that in the new species Brucella microti, the glutamate decarboxylase (Gad)-dependent system (GAD system) contributes to survival at a pH of 2.5 and also to infection in mice by the oral route. In order to study the functionality of the GAD system in the genus Brucella, 47 isolates, representative of all known species and strains of this genus, and 16 strains of the closest neighbor genus, Ochrobactrum, were studied using microbiological, biochemical, and genetic approaches. In agreement with the genome sequences, the GAD system of classical species was not functional, unlike that of most strains of Brucella ceti, Brucella pinnipedialis, and newly described species (B. microti, Brucella inopinata BO1, B. inopinata-like BO2, and Brucella sp. isolated from bullfrogs). In the presence of glutamate, these species were more acid resistant in vitro than classical terrestrial brucellae. Expression in trans of the gad locus from representative Brucella species in the Escherichia coli MG1655 mutant strain lacking the GAD system restored the acid-resistant phenotype. The highly conserved GAD system of the newly described or atypical Brucella species may play an important role in their adaptation to acidic external and host environments. Furthermore, the GAD phenotype was shown to be a useful diagnostic tool to distinguish these latter Brucella strains from Ochrobactrum and from classical terrestrial pathogenic Brucella species, which are GAD negative. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  16. The Change of a Medically Important Genus: Worldwide Occurrence of Genetically Diverse Novel Brucella Species in Exotic Frogs.

    Directory of Open Access Journals (Sweden)

    Holger C Scholz

    Full Text Available The genus Brucella comprises various species of both veterinary and human medical importance. All species are genetically highly related to each other, sharing intra-species average nucleotide identities (ANI of > 99%. Infections occur among various warm-blooded animal species, marine mammals, and humans. Until recently, amphibians had not been recognized as a host for Brucella. In this study, however, we show that novel Brucella species are distributed among exotic frogs worldwide. Comparative recA gene analysis of 36 frog isolates from various continents and different frog species revealed an unexpected high genetic diversity, not observed among classical Brucella species. In phylogenetic reconstructions the isolates consequently formed various clusters and grouped together with atypical more distantly related brucellae, like B. inopinata, strain BO2, and Australian isolates from rodents, some of which were isolated as human pathogens. Of one frog isolate (10RB9215 the genome sequence was determined. Comparative genome analysis of this isolate and the classical Brucella species revealed additional genetic material, absent from classical Brucella species but present in Ochrobactrum, the closest genetic neighbor of Brucella, and in other soil associated genera of the Alphaproteobacteria. The presence of gene clusters encoding for additional metabolic functions, flanked by tRNAs and mobile genetic elements, as well as by bacteriophages is suggestive for a different ecology compared to classical Brucella species. Furthermore it suggests that amphibian isolates may represent a link between free living soil saprophytes and the pathogenic Brucella with a preferred intracellular habitat. We therefore assume that brucellae from frogs have a reservoir in soil and, in contrast to classical brucellae, undergo extensive horizontal gene transfer.

  17. Highly Sensitive Bacteriophage-Based Detection of Brucella abortus in Mixed Culture and Spiked Blood

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    Kirill V. Sergueev

    2017-06-01

    Full Text Available For decades, bacteriophages (phages have been used for Brucella species identification in the diagnosis and epidemiology of brucellosis. Traditional Brucella phage typing is a multi-day procedure including the isolation of a pure culture, a step that can take up to three weeks. In this study, we focused on the use of brucellaphages for sensitive detection of the pathogen in clinical and other complex samples, and developed an indirect method of Brucella detection using real-time quantitative PCR monitoring of brucellaphage DNA amplification via replication on live Brucella cells. This assay allowed the detection of single bacteria (down to 1 colony-forming unit per milliliter within 72 h without DNA extraction and purification steps. The technique was equally efficient with Brucella abortus pure culture and with mixed cultures of B. abortus and α-proteobacterial near neighbors that can be misidentified as Brucella spp., Ochrobactrum anthropi and Afipia felis. The addition of a simple short sample preparation step enabled the indirect phage-based detection of B. abortus in spiked blood, with the same high sensitivity. This indirect phage-based detection assay enables the rapid and sensitive detection of live B. abortus in mixed cultures and in blood samples, and can potentially be applied for detection in other clinical samples and other complex sample types.

  18. Exposure of harbour seals Phoca vitulina to Brucella in declining populations across Scotland.

    Science.gov (United States)

    Kershaw, Joanna L; Stubberfield, Emma J; Foster, Geoffrey; Brownlow, Andrew; Hall, Ailsa J; Perrett, Lorraine L

    2017-09-20

    Since 2000 there has been a major decline in the abundance of Scottish harbour seals Phoca vitulina. The causes of the decline remain uncertain. The aim of this study was to establish the extent to which the seals in the regions of greatest decline have been exposed to Brucella, a bacterial pathogen that causes reproductive failure in terrestrial mammalian hosts. Tissues from dead seals collected between 1992 and 2013 were cultured for Brucella (n = 150). Serum samples collected from live capture-released seals (n = 343) between 1997 and 2012 were tested for Brucella antibodies using the Rose Bengal plate agglutination test (RBT) and a competitive enzyme-linked immunosorbent assay (cELISA). In total, 16% of seals cultured had Brucella isolated from one or more tissues, but there were no pathological signs of infection. The cELISA results were more sensitive than the RBT results, showing that overall 25.4% of seals were seropositive, with the highest seroprevalence in juveniles. As there was no evidence of either a higher seroprevalence or higher circulating antibody levels in seropositive animals in the areas with the greatest declines, it was concluded that Brucella infection is likely not a major contributing factor to recent declines. However, the consistently high proportion of seals exposed to Brucella indicates possible endemicity in these populations, likely due to B. pinnipedialis, which has demonstrated a preference for pinniped hosts. Importantly, given the close proximity between seals, humans and livestock in many areas, there is the potential for cross-species infections.

  19. Highly Sensitive Bacteriophage-Based Detection of Brucella abortus in Mixed Culture and Spiked Blood

    Science.gov (United States)

    Sergueev, Kirill V.; Filippov, Andrey A.; Nikolich, Mikeljon P.

    2017-01-01

    For decades, bacteriophages (phages) have been used for Brucella species identification in the diagnosis and epidemiology of brucellosis. Traditional Brucella phage typing is a multi-day procedure including the isolation of a pure culture, a step that can take up to three weeks. In this study, we focused on the use of brucellaphages for sensitive detection of the pathogen in clinical and other complex samples, and developed an indirect method of Brucella detection using real-time quantitative PCR monitoring of brucellaphage DNA amplification via replication on live Brucella cells. This assay allowed the detection of single bacteria (down to 1 colony-forming unit per milliliter) within 72 h without DNA extraction and purification steps. The technique was equally efficient with Brucella abortus pure culture and with mixed cultures of B. abortus and α-proteobacterial near neighbors that can be misidentified as Brucella spp., Ochrobactrum anthropi and Afipia felis. The addition of a simple short sample preparation step enabled the indirect phage-based detection of B. abortus in spiked blood, with the same high sensitivity. This indirect phage-based detection assay enables the rapid and sensitive detection of live B. abortus in mixed cultures and in blood samples, and can potentially be applied for detection in other clinical samples and other complex sample types. PMID:28604602

  20. Identification of Protective Brucella Antigens and their Expressions in Vaccinia Virus to Prevent Disease in Animals and Humans.

    Science.gov (United States)

    1996-05-01

    selected antigens is through fractionation of Brucella strain RB51 or E.coli recombinants expressing the appropriateBrucella antigen. Briefly, the method...animal species infected with Brucella spp. It is also able to induce the in vitro production of INF-y with lymphocytes of RB51 vaccinated mice (Table...SOD RB51 1IkDa 20 15 x0 0- 10 E 0- Uve Acetone Buffer Void 0-0.1 0.1-0-25 0.25->0.5 0.5-0.75 0.75->1.0 Klled 14 Preparation of new vaccinia/ Brucella

  1. Enzymatic, immunological and phylogenetic characterization of Brucella suis urease

    Directory of Open Access Journals (Sweden)

    Sriranganathan Nammalwar

    2008-07-01

    Full Text Available Abstract Background The sequenced genomes of the Brucella spp. have two urease operons, ure-1 and ure-2, but there is evidence that only one is responsible for encoding an active urease. The present work describes the purification and the enzymatic and phylogenomic characterization of urease from Brucella suis strain 1330. Additionally, the urease reactivity of sera from patients diagnosed with brucellosis was examined. Results Urease encoded by the ure-1 operon of Brucella suis strain 1330 was purified to homogeneity using ion exchange and hydrophobic interaction chromatographies. The urease was purified 51-fold with a recovery of 12% of the enzyme activity and 0.24% of the total protein. The enzyme had an isoelectric point of 5, and showed optimal activity at pH 7.0 and 28–35°C. The purified enzyme exhibited a Michaelis-Menten saturation kinetics with a Km of 5.60 ± 0.69 mM. Hydroxyurea and thiourea are competitive inhibitors of the enzyme with Ki of 1.04 ± 0.31 mM and 26.12 ± 2.30 mM, respectively. Acetohydroxamic acid also inhibits the enzyme in a competitive way. The molecular weight estimated for the native enzyme was between 130–135 kDa by gel filtration chromatography and 157 ± 7 kDa using 5–10% polyacrylamide gradient non-denaturing gel. Only three subunits in SDS-PAGE were identified: two small subunits of 14,000 Da and 15,500 Da, and a major subunit of 66,000 Da. The amino terminal sequence of the purified large subunit corresponded to the predicted amino acid sequence encoded by ureC1. The UreC1 subunit was recognized by sera from patients with acute and chronic brucellosis. By phylogenetic and cluster structure analyses, ureC1 was related to the ureC typically present in the Rhizobiales; in contrast, the ureC2 encoded in the ure-2 operon is more related to distant species. Conclusion We have for the first time purified and characterized an active urease from B. suis. The enzyme was characterized at the kinetic

  2. Risk factors and presence of antibodies to Brucella canis and smooth Brucella in dogs from the municipality of Araguaína, Tocantins, Brazil

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    Jordana Almeida Santana

    2013-12-01

    Full Text Available Canine brucellosis is an infectious disease of worldwide distribution that can affect dogs, wild canids and man. It is caused by Brucella canis, but dogs can also be infected by smooth Brucella such as B. abortus and B. suis. Due to the increasing importance of dogs in our society, to the scarcity of information about canine brucellosis in the country and its zoonotic character, the aims of the present study were (i to conduct a survey on the infection by B. canis and smooth Brucella in dogs from the municipality of Araguaína, Tocantins, Brazil, and (ii to evaluate the risk factors associated with these infections. Sera from 241 dogs were analyzed by agar gel immunodiffusion (AGID to detect B. canisantibodies, and Buffered Acidified Plate Antigen test (BAPA and fluorescence polarization assay (FPA to detect antibodies to smooth Brucella. From the 241 tested dogs, 132 reacted in the AGID and 128 reacted in the BAPA, but only two were positive in FPA. The seroprevalences of B. canis and smooth Brucella infections in dogs in Araguaína were 54.77% (95% CI: 48.25 to 61.17% and 0.83% (95% CI: 0.10 to 2.97%, respectively. The analysis of risk factors showed associations between B. canis infection and vaccination against leptospirosis, and between B. canis infection and use of manufactured food. In conclusion, data from the present study showed a low prevalence of infection by smooth Brucella and a widespread and high prevalence of infection by B. canis in the city of Araguaína, Tocantins, Brazil.

  3. Multiple Locus Variable-Number Tandem-Repeat and Single-Nucleotide Polymorphism-Based Brucella Typing Reveals Multiple Lineages in Brucella melitensis Currently Endemic in China

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    Mingjun Sun

    2017-12-01

    Full Text Available Brucellosis is a worldwide zoonotic disease caused by Brucella spp. In China, brucellosis is recognized as a reemerging disease mainly caused by Brucella melitensis specie. To better understand the currently endemic B. melitensis strains in China, three Brucella genotyping methods were applied to 110 B. melitensis strains obtained in past several years. By MLVA genotyping, five MLVA-8 genotypes were identified, among which genotypes 42 (1-5-3-13-2-2-3-2 was recognized as the predominant genotype, while genotype 63 (1-5-3-13-2-3-3-2 and a novel genotype of 1-5-3-13-2-4-3-2 were second frequently observed. MLVA-16 discerned a total of 57 MLVA-16 genotypes among these Brucella strains, with 41 genotypes being firstly detected and the other 16 genotypes being previously reported. By BruMLSA21 typing, six sequence types (STs were identified, among them ST8 is the most frequently seen in China while the other five STs were firstly detected and designated as ST137, ST138, ST139, ST140, and ST141 by international multilocus sequence typing database. Whole-genome sequence (WGS-single-nucleotide polymorphism (SNP-based typing and phylogenetic analysis resolved Chinese B. melitensis strains into five clusters, reflecting the existence of multiple lineages among these Chinese B. melitensis strains. In phylogeny, Chinese lineages are more closely related to strains collected from East Mediterranean and Middle East countries, such as Turkey, Kuwait, and Iraq. In the next few years, MLVA typing will certainly remain an important epidemiological tool for Brucella infection analysis, as it displays a high discriminatory ability and achieves result largely in agreement with WGS-SNP-based typing. However, WGS-SNP-based typing is found to be the most powerful and reliable method in discerning Brucella strains and will be popular used in the future.

  4. Multiple Locus Variable-Number Tandem-Repeat and Single-Nucleotide Polymorphism-Based Brucella Typing Reveals Multiple Lineages in Brucella melitensis Currently Endemic in China

    Science.gov (United States)

    Sun, Mingjun; Jing, Zhigang; Di, Dongdong; Yan, Hao; Zhang, Zhicheng; Xu, Quangang; Zhang, Xiyue; Wang, Xun; Ni, Bo; Sun, Xiangxiang; Yan, Chengxu; Yang, Zhen; Tian, Lili; Li, Jinping; Fan, Weixing

    2017-01-01

    Brucellosis is a worldwide zoonotic disease caused by Brucella spp. In China, brucellosis is recognized as a reemerging disease mainly caused by Brucella melitensis specie. To better understand the currently endemic B. melitensis strains in China, three Brucella genotyping methods were applied to 110 B. melitensis strains obtained in past several years. By MLVA genotyping, five MLVA-8 genotypes were identified, among which genotypes 42 (1-5-3-13-2-2-3-2) was recognized as the predominant genotype, while genotype 63 (1-5-3-13-2-3-3-2) and a novel genotype of 1-5-3-13-2-4-3-2 were second frequently observed. MLVA-16 discerned a total of 57 MLVA-16 genotypes among these Brucella strains, with 41 genotypes being firstly detected and the other 16 genotypes being previously reported. By BruMLSA21 typing, six sequence types (STs) were identified, among them ST8 is the most frequently seen in China while the other five STs were firstly detected and designated as ST137, ST138, ST139, ST140, and ST141 by international multilocus sequence typing database. Whole-genome sequence (WGS)-single-nucleotide polymorphism (SNP)-based typing and phylogenetic analysis resolved Chinese B. melitensis strains into five clusters, reflecting the existence of multiple lineages among these Chinese B. melitensis strains. In phylogeny, Chinese lineages are more closely related to strains collected from East Mediterranean and Middle East countries, such as Turkey, Kuwait, and Iraq. In the next few years, MLVA typing will certainly remain an important epidemiological tool for Brucella infection analysis, as it displays a high discriminatory ability and achieves result largely in agreement with WGS-SNP-based typing. However, WGS-SNP-based typing is found to be the most powerful and reliable method in discerning Brucella strains and will be popular used in the future. PMID:29312964

  5. A rapid minor groove binder PCR method for distinguishing the vaccine strain Brucella abortus 104M.

    Science.gov (United States)

    Nan, Wenlong; Qin, Lide; Wang, Yong; Zhang, Yueyong; Tan, Pengfei; Chen, Yuqi; Mao, Kairong; Chen, Yiping

    2018-01-24

    Brucellosis is a widespread zoonotic disease caused by Gram-negative Brucella bacteria. Immunisation with attenuated vaccine is an effective method of prevention, but it can interfere with diagnosis. Live, attenuated Brucella abortus strain 104M has been used for the prevention of human brucellosis in China since 1965. However, at present, no fast and reliable method exists that can distinguish this strain from field strains. Single nucleotide polymorphism (SNP)-based assays offer a new approach for such discrimination. SNP-based minor groove binder (MGB) and Cycleave assays have been used for rapid identification of four Brucella vaccine strains (B. abortus strains S19, A19 and RB51, and B. melitensis Rev1). The main objective of this study was to develop a PCR assay for rapid and specific detection of strain 104M. We developed a SNP-based MGB PCR assay that could successfully distinguish strain 104M from 18 representative strains of Brucella (B. abortus biovars 1, 2, 3, 4, 5, 6, 7 and 9, B. melitensis biovars 1, 2 and 3, B. suis biovars 1, 2, 3 and 4, B. canis, B. neotomae, and B. ovis), four Brucella vaccine strains (A19, S19, S2, M5), and 55 Brucella clinical field strains. The assay gave a negative reaction with four non-Brucella species (Escherichia coli, Pasteurella multocida, Streptococcus suis and Pseudomonas aeruginosa). The minimum sensitivity of the assay, evaluated using 10-fold dilutions of chromosomal DNA, was 220 fg for the 104M strain and 76 fg for the single non-104M Brucella strain tested (B. abortus A19). The assay was also reproducible (intra- and inter-assay coefficients of variation = 0.006-0.022 and 0.012-0.044, respectively). A SNP-based MGB PCR assay was developed that could straightforwardly and unambiguously distinguish B. abortus vaccine strain 104M from non-104M Brucella strains. Compared to the classical isolation and identification approaches of bacteriology, this real-time PCR assay has substantial advantages in terms of

  6. An ultrasensitive electrochemical genosensor for Brucella based on palladium nanoparticles.

    Science.gov (United States)

    Rahi, A; Sattarahmady, N; Heli, H

    2016-10-01

    Palladium nanoparticles were potentiostatically electrodeposited on a gold surface at a highly negative potential. The nanostructure, as a transducer, was utilized to immobilize a Brucella-specific probe and the process of immobilization and hybridization was detected by electrochemical methods. The proposed method for detection of the complementary sequence and a non-complementary sequence was applied. The fabricated genosensor was evaluated for the assay of the bacteria in the cultured and human samples with and without PCR. The genosensor could detect the complementary sequence with a sensitivity of 0.02 μA dm(3) mol(-1), a linear concentration range of 1.0 × 10(-12) to 1.0 × 10(-19) mol dm(-3), and a detection limit of 2.7 × 10(-20) mol dm(-3). Copyright © 2016 Elsevier Inc. All rights reserved.

  7. [Brucella abortus endocarditis: survival of a 74 year old patient].

    Science.gov (United States)

    Olea M, Pilar

    2010-02-01

    Brucellosis is not frequent in Chile but it may present with life threatening complications like endocarditis. The case reported refers to a 74 year old man admitted to the Infectious Diseases Hospital Dr. Lucio Córdova in Santiago. He had been febrile for 3 months with no specific symptoms. The trans-esophageal echocardiography confirmed multiple large vegetations and important involvement of the aortic valve. Blood cultures yielded Brucella abortus. The patient required cardiac surgery, along with antibiotics, and he had a satisfactory outcome, being alive at the moment of this report???. Brucellosis can be the responsible for prolonged fever of unknown origin. It is necessary to take in mind brucellosis to obtain the specific laboratory tests. For a best prognosis an early treatment with associated antibiotics for at least 4 a 6 weeks is important. If endocarditis is present valve replacement is often necessary.

  8. Risks of Brucella abortus spillover in the Greater Yellowstone area.

    Science.gov (United States)

    Schumaker, B

    2013-04-01

    Recurrent spillover of Brucella abortus from wildlife reservoirs to domestic cattle in the Greater Yellowstone Area (GYA) has prevented the United States from completely eradicating bovine brucellosis. Risks to cattle are a function of the size and location of wildlife and livestock populations, the degree and nature of spatio-temporal interactions between the various hosts, the level of disease in wildlife, and the susceptibility of livestock herds. While the brucellosis prevalence in wild, free-ranging GYA bison (Bison bison) is high, current management actions have successfully limited contact between bison and cattle. Under current management practices, the risks to cattle in the GYA are predominantly from wild elk (Cervus elaphus). Intra- and inter-species transmission events, while uncommon, are nevertheless crucial for the maintenance of brucellosis in the GYA. Future management actions should focus on decreasing elk herd densities and group sizes and on understanding the behavioural and environmental drivers that result in co-mingling that makes transmission possible.

  9. Vertical transmission of Brucella abortus causes sterility in pregnant mice.

    Science.gov (United States)

    Hashino, Masanori; Kim, Suk; Tachibana, Masato; Shimizu, Takashi; Watarai, Masahisa

    2012-08-01

    The mechanisms of abortion and sterility induced by bacterial infection are largely unknown. In the present study, we found that Brucella abortus, a causative agent of brucellosis and facultative intracellular pathogen, caused sterility in pregnant mice. We have recently established a mouse model for abortion induced by B. abortus infection and high rates of abortion are observed for bacterial infection on day 4.5 of gestation, but not for other days. Infected newborn (first generation) mice showed poor growth compared with uninfected newborn mice and bacterial replication in the spleen of the former was observed over a long period. When infected first generation female mice were mated to infected first generation male mice, the number of fetuses was significantly less than that in uninfected first generation mice. These infected second generation mice also showed poor growth. These results suggest that vertical transmission of B. abostus causes sterility in pregnant mice and our mouse model would be useful for the investigating of brucellosis.

  10. Recovery of a medieval Brucella melitensis genome using shotgun metagenomics.

    Science.gov (United States)

    Kay, Gemma L; Sergeant, Martin J; Giuffra, Valentina; Bandiera, Pasquale; Milanese, Marco; Bramanti, Barbara; Bianucci, Raffaella; Pallen, Mark J

    2014-07-15

    Shotgun metagenomics provides a powerful assumption-free approach to the recovery of pathogen genomes from contemporary and historical material. We sequenced the metagenome of a calcified nodule from the skeleton of a 14th-century middle-aged male excavated from the medieval Sardinian settlement of Geridu. We obtained 6.5-fold coverage of a Brucella melitensis genome. Sequence reads from this genome showed signatures typical of ancient or aged DNA. Despite the relatively low coverage, we were able to use information from single-nucleotide polymorphisms to place the medieval pathogen genome within a clade of B. melitensis strains that included the well-studied Ether strain and two other recent Italian isolates. We confirmed this placement using information from deletions and IS711 insertions. We conclude that metagenomics stands ready to document past and present infections, shedding light on the emergence, evolution, and spread of microbial pathogens. Importance: Infectious diseases have shaped human populations and societies throughout history. The recovery of pathogen DNA sequences from human remains provides an opportunity to identify and characterize the causes of individual and epidemic infections. By sequencing DNA extracted from medieval human remains through shotgun metagenomics, without target-specific capture or amplification, we have obtained a draft genome sequence of an ~700-year-old Brucella melitensis strain. Using a variety of bioinformatic approaches, we have shown that this historical strain is most closely related to recent strains isolated from Italy, confirming the continuity of this zoonotic infection, and even a specific lineage, in the Mediterranean region over the centuries. Copyright © 2014 Kay et al.

  11. ISOLATION AND CHARACTERIZATION OF A NOVEL MARINE BRUCELLA FROM A SOUTHERN SEA OTTER (ENHYDRA LUTRIS NEREIS), CALIFORNIA, USA.

    Science.gov (United States)

    Miller, Melissa A; Burgess, Tristan L; Dodd, Erin M; Rhyan, Jack C; Jang, Spencer S; Byrne, Barbara A; Gulland, Frances M D; Murray, Michael J; Toy-Choutka, Sharon; Conrad, Patricia A; Field, Cara L; Sidor, Inga F; Smith, Woutrina A

    2017-04-01

    We characterize Brucella infection in a wild southern sea otter ( Enhydra lutris nereis) with osteolytic lesions similar to those reported in other marine mammals and humans. This otter stranded twice along the central California coast, US over a 1-yr period and was handled extensively at two wildlife rehabilitation facilities, undergoing multiple surgeries and months of postsurgical care. Ultimately the otter was euthanized due to severe, progressive neurologic disease. Necropsy and postmortem radiographs revealed chronic, severe osteoarthritis spanning the proximal interphalangeal joint of the left hind fifth digit. Numerous coccobacilli within the joint were strongly positive on Brucella immunohistochemical labelling, and Brucella sp. was isolated in pure culture from this lesion. Sparse Brucella-immunopositive bacteria were also observed in the cytoplasm of a pulmonary vascular monocyte, and multifocal granulomas were observed in the spinal cord and liver on histopathology. Findings from biochemical characterization, 16S ribosomal DNA, and bp26 gene sequencing of the bacterial isolate were identical to those from marine-origin brucellae isolated from cetaceans and phocids. Although omp2a gene sequencing revealed 100% homology with marine Brucella spp. infecting pinnipeds, whales, and humans, omp2b gene sequences were identical only to pinniped-origin isolates. Multilocus sequence typing classified the sea otter isolate as ST26, a sequence type previously associated only with cetaceans. Our data suggest that the sea otter Brucella strain represents a novel marine lineage that is distinct from both Brucella pinnipedialis and Brucella ceti. Prior reports document the zoonotic potential of the marine brucellae. Isolation of Brucella sp. from a stranded sea otter highlights the importance of wearing personal protective equipment when handling sea otters and other marine mammals as part of wildlife conservation and rehabilitation efforts.

  12. Raman Spectroscopy as a Potential Tool for Detection of Brucella spp. in Milk

    Science.gov (United States)

    Meisel, Susann; Stöckel, Stephan; Elschner, Mandy; Melzer, Falk; Popp, Jürgen

    2012-01-01

    Detection of Brucella, causing brucellosis, is very challenging, since the applied techniques are mostly time-demanding and not standardized. While the common detection system relies on the cultivation of the bacteria, further classical typing up to the biotype level is mostly based on phenotypic or genotypic characteristics. The results of genotyping do not always fit the existing taxonomy, and misidentifications between genetically closely related genera cannot be avoided. This situation gets even worse, when detection from complex matrices, such as milk, is necessary. For these reasons, the availability of a method that allows early and reliable identification of possible Brucella isolates for both clinical and epidemiological reasons would be extremely useful. We evaluated micro-Raman spectroscopy in combination with chemometric analysis to identify Brucella from agar plates and directly from milk: prior to these studies, the samples were inactivated via formaldehyde treatment to ensure a higher working safety. The single-cell Raman spectra of different Brucella, Escherichia, Ochrobactrum, Pseudomonas, and Yersinia spp. were measured to create two independent databases for detection in media and milk. Identification accuracies of 92% for Brucella from medium and 94% for Brucella from milk were obtained while analyzing the single-cell Raman spectra via support vector machine. Even the identification of the other genera yielded sufficient results, with accuracies of >90%. In summary, micro-Raman spectroscopy is a promising alternative for detecting Brucella. The measurements we performed at the single-cell level thus allow fast identification within a few hours without a demanding process for sample preparation. PMID:22660699

  13. Comprehensive Identification of Immunodominant Proteins of Brucella abortus and Brucella melitensis Using Antibodies in the Sera from Naturally Infected Hosts

    Directory of Open Access Journals (Sweden)

    Gamal Wareth

    2016-04-01

    Full Text Available 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.

  14. Post-exposure serological and bacteriological responses of water buffalo (Bubalus bubalis) to Brucella abortus biovar 1 following vaccination with Brucella abortus strain RB51.

    Science.gov (United States)

    Diptee, M D; Asgarali, Z; Campbell, M; Fosgate, G; Adesiyun, A A

    2007-12-01

    Serological and bacteriological responses to Brucella abortus biovar 1 following vaccination with B. abortus strain RB51 (RB51) were evaluated in thirty domestic water buffalo (Bubalus bubalis) randomly divided into five treatment groups. Groups I to V received, respectively, the recommended dose (RD) of RB51 vaccine once, RD twice 4 weeks apart, double RD once, double RD twice 4 weeks apart, and saline once (control). Vaccination did not result in a serological response. Experimental animals released 27 weeks post initial inoculation (27 PIIW) into a brucellosis-positive herd failed to seroconvert after 29 weeks. Experimental challenge commenced at 57 PIIW. All animals received B. abortus biovar 1 intraconjunctivally at 0, 5 and 9 weeks post experimental exposure (PEEW). Serum samples collected at 4, 8 and 13 PEEW were negative. At 16 PEEW all animals received B. abortus biovar 1 subcutaneously (SC), and all seroconverted by 20 PEEW. Five of twenty-six animals were positive for Brucella infection on bacterial culture. Brucella abortus biovar 1 was isolated from three animals; B. abortus RB51 was isolated from two. Treatment group, age and sex had no effect on the isolation of Brucellae (P>0.05).

  15. Vaccination of elk (Cervus canadensis) with Brucella abortus strain RB51 overexpressing superoxide dismutase and glycosyltransferase genes does not induce adequate protection against experimental brucella abortus challenge

    Science.gov (United States)

    In recent years, elk (Cervus canadensis) have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area (GYA). In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the d...

  16. Anticorpos anti-Brucella canis e anti-Brucella abortus em cães de Araguaína, Tocantins

    Directory of Open Access Journals (Sweden)

    Elaine Maria Seles Dorneles

    2011-04-01

    Full Text Available The aims of the present study were to determine the seroprevalence of infection by Brucella canis and Brucella abortus and to evaluate possible risk factors for infection in dogs from Araguaína, Tocantins, Brazil. Sera from 374 dogs, of the urban zones of the municipality, from both sexes, were submitted to the agar-gel immunodiffusion for Brucella canis-antibodies and to rose Bengal test (AAT and fluorescence polarization assay (FPA for Brucella abortus-antibodies. From the 374 tested dogs, 21 reacted in the AAT, but no one was positive in the FPA. The seroprevalence of B. canis infection found in Araguaína, Tocantins, Brazil, was 44.53% (95% IC; 39.43 to 49.72. No association was found among seropositivity for B. canis and the risk factors studied. Thus, data from the present study showed that there was no infection by B. abortus among dogs in the sample and that infection by B. canis is widespread and at high prevalence in Araguaína, Tocantins, Brazil.

  17. Immunochromatographic Brucella-specific immunoglobulin M and G lateral flow assays for rapid serodiagnosis of human brucellosis

    NARCIS (Netherlands)

    Smits, Henk L.; Abdoel, Theresia H.; Solera, Javier; Clavijo, Encarnacion; Diaz, Ramon

    2003-01-01

    To fulfill the need for a simple and rapid diagnostic test for human brucellosis, we used the immunochromatographic lateral flow assay format to develop two assays, one for the detection of Brucella-specific immunoglobulin M (IgM) antibodies and one for the detection of Brucella-specific IgG

  18. Whole-genome sequence of the first sequence type 27 Brucella ceti strain isolated from European waters

    DEFF Research Database (Denmark)

    Duvnjak, Sanja; Spicic, Silvio; Kusar, Darja

    2017-01-01

    Brucella spp. that cause marine brucellosis are becoming more important, as the disease appears to be more widespread than originally thought. Here, we report a whole and annotated genome sequence of Brucella ceti CRO350, a sequence type 27 strain isolated from a bottlenose dolphin carcass found...

  19. Brucella glomerulonephritis resulting in end-stage renal disease: a case report and a brief review of the literature.

    Science.gov (United States)

    Bakri, Faris G; Wahbeh, Ayman; Mahafzah, Azmi; Tarawneh, Musleh

    2008-01-01

    Brucella glomerulonephritis is a rare condition with only a few reported cases. We review the literature, and describe a 24-year-old female who presented with edema and proteinuria. Blood grew Brucella melitensis. Renal biopsy showed diffuse proliferative glomerulonephritis. The patient progressed to end-stage renal disease despite antibiotic and steroid therapy.

  20. Typing of Brucella field strains isolated from livestock populations in Italy between 2001 and 2006

    Directory of Open Access Journals (Sweden)

    Elisabetta Di Giannatale

    2008-06-01

    Full Text Available The identification of species and biovars of Brucella field strains isolated in outbreaks is essential to fully understand the epidemiology of the disease and to trace sources of infection, thereby improving the outcome of brucellosis eradication programmes. It is important to identify the presence of Brucella strains in livestock populations and to determine the presence of new strains that might previously have been considered exotic. In this study, 732 Brucella strains isolated from livestock were submitted for typing by the Italian Istituti Zooprofilattici Sperimentali to the National Reference Laboratory for brucellosis between 2001 and 2006. Animal species affected, biovars typed and spatial distribution of isolates are discussed. Brucella field strains were identified using both conventional bacteriological methods and molecular techniques. Species identification was performed using the AMOS (AbortusMelitensisOvisSuis-polymerase chain reaction. For biovar identification, amplification of omp2a, omp2b and omp31 genes was performed, followed by restriction fragment length polymorphism. Final biovar identification was performed by growth in the presence of basic fuchsin and thionin, using the slide agglutination test with Brucella A- and M- specific antisera.

  1. Shedding of Brucella abortus rough mutant strain RB51 in milk of water buffalo (Bubalus bubalis).

    Science.gov (United States)

    Longo, Mariangela; Mallardo, Karina; Montagnaro, Serena; De Martino, Luisa; Gallo, Sergio; Fusco, Giovanna; Galiero, Giorgio; Guarino, Achille; Pagnini, Ugo; Iovane, Giuseppe

    2009-07-01

    The objective of this study was to determine if Brucella abortus rough mutant strain RB51 (SRB51) is eliminated in buffalo milk. Thirty Brucella-free female buffaloes were used in this study: ten 4-5 years old were inoculated with the triple of the recommended calfhood dose of SRB51 by subcutaneous route, ten 2-3 years old at the first lactation were previously vaccinated twice as calves with triple the recommended calf dose of RB51, while five 4-5 years old and five 2-3 years old not vaccinated Brucella-free female buffaloes served as controls. Milk samples were taken aseptically on a daily basis for the first 30 days and weekly for the second and third months. The samples were inoculated on selective media for isolation of SRB51 and incubated for 11 days. Moreover, PCR analysis was also performed directly on milk samples. SRB51 was isolated from milk samples only during the first week post-vaccination while RB51 DNA was detected during the first week till the fourth week post-vaccination only in water buffaloes vaccinated as adults. The identification of Brucella RB51 in milk samples, strongly suggests that this Brucella vaccine could be excreted in milk of buffalo cows vaccinated as adults, while our data demonstrate that the vaccine is safe for use in buffaloes vaccinated as calves in which it was not excreted in milk.

  2. Molecular strain typing of Brucella abortus isolates from Italy by two VNTR allele sizing technologies.

    Science.gov (United States)

    De Santis, Riccardo; Ancora, Massimo; De Massis, Fabrizio; Ciammaruconi, Andrea; Zilli, Katiuscia; Di Giannatale, Elisabetta; Pittiglio, Valentina; Fillo, Silvia; Lista, Florigio

    2013-10-01

    Brucellosis, one of the most important re-emerging zoonoses in many countries, is caused by bacteria belonging to the genus Brucella. Furthermore these bacteria represent potential biological warfare agents and the identification of species and biovars of field strains may be crucial for tracing back source of infection, allowing to discriminate naturally occurring outbreaks instead of bioterrorist events. In the last years, multiple-locus variable-number tandem repeat analysis (MLVA) has been proposed as complement of the classical biotyping methods and it has been applied for genotyping large collections of Brucella spp. At present, the MLVA band profiles may be resolved by automated or manual procedures. The Lab on a chip technology represents a valid alternative to standard genotyping techniques (as agarose gel electrophoresis) and it has been previously used for Brucella genotyping. Recently, a new high-throughput genotyping analysis system based on capillary gel electrophoresis, the QIAxcel, has been described. The aim of the study was to evaluate the ability of two DNA sizing equipments, the QIAxcel System and the Lab chip GX, to correctly call alleles at the sixteen loci including one frequently used MLVA assay for Brucella genotyping. The results confirmed that these technologies represent a meaningful advancement in high-throughput Brucella genotyping. Considering the accuracy required to confidently resolve loci discrimination, QIAxcel shows a better ability to measure VNTR allele sizes compared to LabChip GX.

  3. Meta-analysis of Brucella seroprevalence in dairy cattle of Ethiopia.

    Science.gov (United States)

    Asmare, Kassahun; Krontveit, Randi I; Ayelet, Gelagay; Sibhat, Berhanu; Godfroid, Jacques; Skjerve, Eystein

    2014-12-01

    This meta-analysis estimates a single-group summary (effect size) for seroprevalence of Brucella spp. exposure in dairy cattle of Ethiopia. It also attempts to identify study-level variables that could explain the variation in apparent seroprevalence. The literature search was restricted to studies published in English language from January 2000 to December 2013. A template was designed to retrieve the most biologically plausible and consistent variables from the articles. A total of 29 published papers containing 40 animal-level studies were used in the analyses. The single-group summary of Brucella seroprevalence in cattle was estimated to reach 3.3 % with 95 % confidence interval (CI) (2.6-4.2 %). Of all the variables considered, region was the only specific factor identified to explain about 20 % of between-study variation. Accordingly, the region-based meta-analysis forest plot revealed the highest prevalence in central Ethiopia followed by southern part. The lowest prevalence estimate was observed in the western part of the country. The visual inspection of the funnel plot demonstrated the presence of possible publication bias which might dictate shortage of studies with higher prevalences or variance inflation due to infectiousness of Brucella. In conclusion, the quantitative review showed the seroprevalence to be low but widely distributed. More importantly, the review underscores the need for isolation and characterization of the circulating Brucella spp. to capture the type of Brucella spp. involved and its distribution in cattle in Ethiopia.

  4. Neospora caninum versus Brucella spp. exposure among dairy cattle in Ethiopia: a case control study.

    Science.gov (United States)

    Asmare, Kassahun

    2014-08-01

    This case-control study aimed at assessing the relative association of Neospora caninum and Brucella species exposure with reproductive disorders. The study was carried out between October 2011 and June 2012 on 731 dairy cows sampled from 150 dairy farms in selected 17 conurbations of Ethiopia. Two hundred sixty-six of the cows were categorized as cases based on their history of abortion or stillbirth while the remaining 465 were controls. The presence of antibody to N. caninum was screened using indirect ELISA, while Brucella spp. exposure was assayed serially using Rose Bengal Plate Test and Complement Fixation Test. Exposure to N. caninum was more frequently observed among cases (23.8%) than controls (12.7%), while no significant difference (p > 0.05) was noted for Brucella exposure between the two groups. Moreover, the proportion of cows with disorders like retention of fetal membrane, endometritis and increased inter-calving period were significantly higher (p Brucella spp. exposure. However, neither N. caninum nor Brucella spp. could explain the majority (73.2%) of the reported abortions and stillbirths in cattle. Hence, this observation underscores the need for more intensive investigation on the identification of causes of the aforementioned disorders in dairy cattle of Ethiopia.

  5. Genome degradation in Brucella ovis corresponds with narrowing of its host range and tissue tropism.

    Directory of Open Access Journals (Sweden)

    Renee M Tsolis

    Full Text Available 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.

  6. Study of respiratory burst in patients with Brucella infection by using chemiluminescence method

    Directory of Open Access Journals (Sweden)

    Dabir M

    2000-08-01

    Full Text Available Although cellular immunity involving activated macrophage is important in resistance to Brucella infections, serum factors and polymorphonuclears (PMNs play some role in the initial responses to Brucella infections. In this research, we studied respiratory burst of PMNs against opsonized yeast and opsonized inactivated Brucella melitensis in chronic Brucellosis patients and controls with no previous history of Brucellosis. A group of 41 patients and another group of 20 blood donors as control, were included. The other 2 groups included 10 cases and 6 controls. Mean responses of PMNs of patients and controls to opsonized yeast were 110.3 and 129.3 milivolt respectively and the difference was not statistically significant. No statistically significant difference was observed between respiratory burst of PMNs exposed to inactivated Brucella in 10 patients with chronic Brucellosis (Mean 67.2 and 6 control blood donors (Mean 112.5, so we concluded that inactivated Brucella melitensis can't inhibit activity of myeloproxidase enzyme. 

  7. [Detection of Brucella with an automatic hemoculture system: Bact/Alert].

    Science.gov (United States)

    Casas, J; Partal, Y; Llosá, J; Leiva, J; Navarro, J M; de la Rosa, M

    1994-12-01

    The ability of in vitro and in vivo detection of Brucella spp. with the Bact/Alert system was studied. Three strains of Brucella melitensis and two of Brucella abortus were used. Different dilutions of the five strains were performed in trypticase soy broth (TSB), achieving concentrations of 1 cfu/ml, 5 cfu/ml, 10 cfu/ml and 100 cfu/ml. Ten ml of each dilution and strain were inoculated into 5 aerobic bottles Bact/Alert and 5 biphasic Hemóline bottles. Furthermore, over a 9 month period, 8,216 bottles of Bact/Alert bottles from hospitalized patients and from the emergency department were processed in the authors' laboratory. The mean detection time for Brucella growth was from 2 to 3 days with the Bact/Alert system, and 14 days in the biphasic bottles. Former bottles processed in the authors' laboratory, 11 aerobic bottles belonged to 5 patients in whom brucelosis was confirmed by bloodculture. The Bact/Alert system detected Brucella melitensis in only on bottle at 2.9 days of incubation. In 7 bottles Bact/Alert detected B. melitensis by a blind pass of these bottles at 10 to 20 days of incubation. These results suggest that the Bact/Alert system does not totally solve the diagnosis of brucellosis. Blind passes of the bloodcultures are required.

  8. DNA polymorphism analysis of Brucella lipopolysaccharide genes reveals marked differences in O-polysaccharide biosynthetic genes between smooth and rough Brucella species and novel species-specific markers

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    Cloeckaert Axel

    2009-05-01

    Full Text Available Abstract Background The lipopolysaccharide is a major antigen and virulence factor of Brucella, an important bacterial pathogen. In smooth brucellae, lipopolysaccharide is made of lipid A-core oligosaccharide and N-formylperosamine O-polysaccharide. B. ovis and B. canis (rough species lack the O-polysaccharide. Results The polymorphism of O-polysaccharide genes wbkE, manAO-Ag, manBO-Ag, manCO-Ag, wbkF and wbkD and wbo (wboA and wboB, and core genes manBcore and wa** was analyzed. Although most genes were highly conserved, species- and biovar-specific restriction patterns were found. There were no significant differences in putative N-formylperosamyl transferase genes, suggesting that Brucella A and M serotypes are not related to specific genes. In B. pinnipedialis and B. ceti (both smooth, manBO-Ag carried an IS711, confirming its dispensability for perosamine synthesis. Significant differences between smooth and rough species were found in wbkF and wbkD, two adjacent genes putatively related to bactoprenol priming for O-polysaccharide polymerization. B. ovis wbkF carried a frame-shift and B. canis had a long deletion partially encompassing both genes. In smooth brucellae, this region contains two direct repeats suggesting the deletion mechanism. Conclusion The results define species and biovar markers, confirm the dispensability of manBO-Ag for O-polysaccharide synthesis and contribute to explain the lipopolysaccharide structure of rough and smooth Brucella species.

  9. Genomic Island 2 Is an Unstable Genetic Element Contributing to Brucella Lipopolysaccharide Spontaneous Smooth-to-Rough Dissociation ▿ †

    Science.gov (United States)

    Mancilla, Marcos; López-Goñi, Ignacio; Moriyón, Ignacio; Zárraga, Ana María

    2010-01-01

    Brucella is a Gram-negative bacterium that causes a worldwide-distributed zoonosis. The genus includes smooth (S) and rough (R) species that differ in the presence or absence, respectively, of the O-polysaccharide of lipopolysaccharide. In S brucellae, the O-polysaccharide is a critical diagnostic antigen and a virulence determinant. However, S brucellae spontaneously dissociate into R forms, a problem in antigen and S vaccine production. Spontaneous R mutants of Brucella abortus, Brucella melitensis, and Brucella suis carried the chromosomal scar corresponding to genomic island 2 (GI-2) excision, an event causing the loss of the wboA and wboB O-polysaccharide genes, and the predicted excised circular intermediate was identified in B. abortus, B. melitensis, and B. suis cultures. Moreover, disruption of a putative phage integrase gene in B. abortus GI-2 caused a reduction in O-polysaccharide loss rates under conditions promoting S-R dissociation. However, spontaneous R mutants not carrying the GI-2 scar were also detected. These results demonstrate that the phage integrase-related GI-2 excision is a cause of S-R brucella dissociation and that other undescribed mechanisms must also be involved. In the R Brucella species, previous works have shown that Brucella ovis but not Brucella canis lacks GI-2, and a chromosomal scar identical to those in R mutants was observed. These results suggest that the phage integrase-promoted GI-2 excision played a role in B. ovis speciation and are consistent with other evidence, suggesting that this species and B. canis have emerged as two independent lineages. PMID:20952568

  10. Systems biology analysis of Brucella infected Peyer's patch reveals rapid invasion with modest transient perturbations of the host transcriptome.

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    Carlos A Rossetti

    Full Text Available Brucella melitensis causes the most severe and acute symptoms of all Brucella species in human beings and infects hosts primarily through the oral route. The epithelium covering domed villi of jejunal-ileal Peyer's patches is an important site of entry for several pathogens, including Brucella. Here, we use the calf ligated ileal loop model to study temporal in vivo Brucella-infected host molecular and morphological responses. Our results document Brucella bacteremia occurring within 30 min after intraluminal inoculation of the ileum without histopathologic traces of lesions. Based on a system biology Dynamic Bayesian Network modeling approach (DBN of microarray data, a very early transient perturbation of the host enteric transcriptome was associated with the initial host response to Brucella contact that is rapidly averted allowing invasion and dissemination. A detailed analysis revealed active expression of Syndecan 2, Integrin alpha L and Integrin beta 2 genes, which may favor initial Brucella adhesion. Also, two intestinal barrier-related pathways (Tight Junction and Trefoil Factors Initiated Mucosal Healing were significantly repressed in the early stage of infection, suggesting subversion of mucosal epithelial barrier function to facilitate Brucella transepithelial migration. Simultaneously, the strong activation of the innate immune response pathways would suggest that the host mounts an appropriate protective immune response; however, the expression of the two key genes that encode innate immunity anti-Brucella cytokines such as TNF-α and IL12p40 were not significantly changed throughout the study. Furthermore, the defective expression of Toll-Like Receptor Signaling pathways may partially explain the lack of proinflammatory cytokine production and consequently the absence of morphologically detectable inflammation at the site of infection. Cumulatively, our results indicate that the in vivo pathogenesis of the early infectious process

  11. Isolation and Molecular Characterization of Brucella Isolates in Cattle Milk in Uganda

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    Denis Rwabiita Mugizi

    2015-01-01

    Full Text Available Brucellosis is endemic in livestock and humans in Uganda and its transmission involves a multitude of risk factors like consumption of milk from infected cattle. To shed new light on the epidemiology of brucellosis in Uganda the present study used phenotypic and molecular approaches to delineate the Brucella species, biovars, and genotypes shed in cattle milk. Brucella abortus without a biovar designation was isolated from eleven out of 207 milk samples from cattle in Uganda. These isolates had a genomic monomorphism at 16 variable number tandem repeat (VNTR loci and showed in turn high levels of genetic variation when compared with other African strains or other B. abortus biovars from other parts of the world. This study further highlights the usefulness of MLVA as an epidemiological tool for investigation of Brucella infections.

  12. Molecular detection and characterization of Brucella species in raw informally marketed milk from Uganda

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    Tove Hoffman

    2016-11-01

    Full Text Available This study identified and characterized Brucella species in the informal milk chain in Uganda. A total of 324 cattle bulk milk samples were screened for the genus Brucella by real-time PCR with primers targeting the bcsp31 gene and further characterized by the omp25 gene. Of the samples tested, 6.5% were positive for Brucella species. In the omp25 phylogeny, the study sequences were found to form a separate clade within the branch containing B. abortus sequences. The study shows that informally marketed cattle milk in Uganda is a likely risk factor for human brucellosis and confirms that B. abortus is present in the cattle population. This information is important for potential future control measures, such as vaccination of cattle.

  13. Isolation and Molecular Characterization of Brucella Isolates in Cattle Milk in Uganda

    Science.gov (United States)

    Mugizi, Denis Rwabiita; Muradrasoli, Shaman; Erume, Joseph; Nasinyama, George William; Waiswa, Charles; Magnusson, Ulf

    2015-01-01

    Brucellosis is endemic in livestock and humans in Uganda and its transmission involves a multitude of risk factors like consumption of milk from infected cattle. To shed new light on the epidemiology of brucellosis in Uganda the present study used phenotypic and molecular approaches to delineate the Brucella species, biovars, and genotypes shed in cattle milk. Brucella abortus without a biovar designation was isolated from eleven out of 207 milk samples from cattle in Uganda. These isolates had a genomic monomorphism at 16 variable number tandem repeat (VNTR) loci and showed in turn high levels of genetic variation when compared with other African strains or other B. abortus biovars from other parts of the world. This study further highlights the usefulness of MLVA as an epidemiological tool for investigation of Brucella infections. PMID:25793204

  14. Leptospira and Brucella antibodies in collared anteaters (Tamandua tetradactyla) in Brazilian zoos.

    Science.gov (United States)

    Sales, Indiara dos Santos; Folly, Márcio Manhães; Garcia, Luize Néli Nunes; Ramos, Tatiane Mendes Varela; da Silva, Mariana Cristina; Pereira, Martha Maria

    2012-12-01

    The presence of Leptospira spp. and Brucella spp. antibodies was investigated in serum samples from 28 collared anteaters (Tamandua tetradactyla) kept in seven Brazilian zoos. Sera were tested against 19 Leptospira serovars using microscopic agglutination. Samples reacted to the following serovars: two (7.14%) to Patoc, three (10.71%) to Tarrasovi, three (10.71%) to both Patoc and Tarrasovi, two (7.14%) to Wolffi, and one (3.57%) to Australis. Two (7.14%) samples reacted to the buffered Brucella antigen test, but no confirmatory reaction occurred using the 2-mercaptoethanol slow slide agglutination test. No sample was reactive in the agar gel immunodiffusion test for rugose species of Brucella. The presence of anti-leptospira agglutinins in captive T. tetradactyla serum indicates that this species may be susceptible to infection by these bacteria.

  15. BtpB, a novel Brucella TIR-containing effector protein with immune modulatory functions

    Science.gov (United States)

    Salcedo, Suzana P.; Marchesini, María I.; Degos, Clara; Terwagne, Matthieu; Von Bargen, Kristine; Lepidi, Hubert; Herrmann, Claudia K.; Santos Lacerda, Thais L.; Imbert, Paul R. C.; Pierre, Philippe; Alexopoulou, Lena; Letesson, Jean-Jacques; Comerci, Diego J.; Gorvel, Jean-Pierre

    2013-01-01

    Several bacterial pathogens have TIR domain-containing proteins that contribute to their pathogenesis. We identified a second TIR-containing protein in Brucella spp. that we have designated BtpB. We show it is a potent inhibitor of TLR signaling, probably via MyD88. BtpB is a novel Brucella effector that is translocated into host cells and interferes with activation of dendritic cells. In vivo mouse studies revealed that BtpB is contributing to virulence and control of local inflammatory responses with relevance in the establishment of chronic brucellosis. Together, our results show that BtpB is a novel Brucella effector that plays a major role in the modulation of host innate immune response during infection. PMID:23847770

  16. Brucella alters the immune response in a prpA-dependent manner

    Science.gov (United States)

    Spera, Juan M.; Comerci, Diego J.; Ugalde, Juan E.

    2014-01-01

    Brucellosis, a disease caused by the gram-negative bacterium Brucella sp, is a widespread zoonosis that inflicts important animal and human health problems, especially in developing countries. One of the hallmarks of Brucella infection is its capacity to establish a chronic infection, characteristic that depends on a wide repertoire of virulence factors among which are immunomodulatory proteins such as PrpA (encoding the proline racemase protein A or hydroxyproline-2-epimerase), involved in the establishment of the chronic phase of the infectious process that we have previously identified and characterized. We report here that, in vivo, B. abortus prpA is responsible for an increment in the B-cell number and in the specific antibody response and that these antibodies promote cell infection. We additionally found that Brucella alters the cytokine levels of IFN-γ, IL-10, TGFβ1 and TNFα during the acute phase of the infectious process in a prpA dependent manner. PMID:24508400

  17. [Detection of Brucella canis by immunochromatography method in vague dogs captured in Temuco city, Chile, 2011].

    Science.gov (United States)

    Tuemmers, Christian; Lüders, Carlos; Rojas, Claudio; Serri, Michel; Castillo, Carolina; Espinoza, Rodrigo

    2013-08-01

    Brucella canis is responsible for brucellosis in dogs, causing reproductive disorders and is considered a zoonoses, as described in several countries. The epidemiological data are scarce in our country. To determine the prevalence of Brucella canis in vague dogs in Temuco city and housed in the Temuco Kennel. Quantitative and cross-section study. We used 400 samples of dogs of both sexes, different ages and mainly mixed race, which were tested by immunochromatography. Antibodies were detected in 4 samples Brucella canis which represented 1% of the population studied, 2 females (0.5%) and 2 males (0.5%). We conclude that dogs are infected by B. canis in a low range but remains a risk condition to the health of the human population if not maintained adequate sanitary control of pets, like vague dogs.

  18. Multipronged diagnostic approaches for monitoring the treatment of Brucella abortus infected patient: a case report

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    Rajeswari Shome

    2015-07-01

    Full Text Available Brucellosis caused by Brucella species is readily transmissible to humans, causing acute febrile illness and undulant fever which may progress to a more chronic form and can also produce serious complications affecting the musculoskeletal, cardiovascular, and central nervous systems. A veterinary livestock inspector presented to the institute with symptoms of intermittent fever, pain involving muscles and joints, loss of weight, anxiety and weakness for about three months has been investigated. The isolation, serological tests and PCR were performed for diagnosis of brucellosis. Based on history of constant professional association with animals, characteristic symptoms, hematological and biochemical, multiple serological and PCR assay results, the patient was diagnosed as brucellosis. Detection of Brucella abortus directly in the clinical samples by gel based PCRs were highly useful for diagnosis and monitoring of treatment. This diagnostic protocol will facilitate in a simple way to map major Brucella species infecting humans in a geographical region.

  19. Lab on a chip genotyping for Brucella spp. based on 15-loci multi locus VNTR analysis.

    Science.gov (United States)

    De Santis, Riccardo; Ciammaruconi, Andrea; Faggioni, Giovanni; D'Amelio, Raffaele; Marianelli, Cinzia; Lista, Florigio

    2009-04-07

    Brucellosis is an important zoonosis caused by the genus Brucella. In addition Brucella represents potential biological warfare agents due to the high contagious rates for humans and animals. Therefore, the strain typing epidemiological tool may be crucial for tracing back source of infection in outbreaks and discriminating naturally occurring outbreaks versus bioterroristic event. A Multiple Locus Variable-number tandem repeats (VNTR) Analysis (MLVA) assay based on 15 polymorphic markers was previously described. The obtained MLVA band profiles may be resolved by techniques ranging from low cost manual agarose gels to the more expensive capillary electrophoresis sequencing. In this paper a rapid, accurate and reproducible system, based on the Lab on a chip technology was set up for Brucella spp. genotyping. Seventeen DNA samples of Brucella strains isolated in Sicily, previously genotyped, and twelve DNA samples, provided by MLVA Brucella VNTR ring trial, were analyzed by MLVA-15 on Agilent 2100. The DNA fragment sizes produced by Agilent, compared with those expected, showed discrepancies; therefore, in order to assign the correct alleles to the Agilent DNA fragment sizes, a conversion table was produced. In order to validate the system twelve unknown DNA samples were analyzed by this method obtaining a full concordance with the VNTR ring trial results. In this paper we described a rapid and specific detection method for the characterization of Brucella isolates. The comparison of the MLVA typing data produced by Agilent system with the data obtained by standard sequencing or ethidium bromide slab gel electrophoresis showed a general concordance of the results. Therefore this platform represents a fair compromise among costs, speed and specificity compared to any conventional molecular typing technique.

  20. Exploring the Diversity of Field Strains of Brucella abortus Biovar 3 Isolated in West Africa

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    Moussa Sanogo

    2017-06-01

    Full Text Available Brucellosis is one of the most widespread bacterial zoonotic diseases in the world, affecting both humans and domestic and wild animals. Identification and biotyping of field strains of Brucella are of key importance for a better knowledge of the epidemiology of brucellosis, for identifying appropriate antigens, for managing disease outbreaks and for setting up efficient preventive and control programmes. Such data are required both at national and regional level to assess potential threats for public health. Highly discriminative genotyping methods such as the multiple locus variable number of tandem repeats analysis (MLVA allow the comparison and assessment of genetic relatedness between field strains of Brucella within the same geographical area. In this study, MLVA biotyping data retrieved from the literature using a systematic review were compared using a clustering analysis and the Hunter-Gaston diversity index (HGDI. Thus, the analysis of the 42 MLVA genotyping results found in the literature on West Africa [i.e., from Ivory Coast (1, Niger (1, Nigeria (34, The Gambia (3, and Togo (3] did not allow a complete assessment of the actual diversity among field strains of Brucella. However, it provided some preliminary indications on the co-existence of 25 distinct genotypes of Brucella abortus biovar 3 in this region with 19 genotypes from Nigeria, three from Togo and one from Ivory Coast, The Gambia, and Niger. The strong and urgent need for more sustainable molecular data on prevailing strains of Brucella in this sub-region of Africa and also on all susceptible species including humans is therefore highlighted. This remains a necessary stage to allow a comprehensive understanding of the relatedness between field strains of Brucella and the epidemiology of brucellosis within West Africa countries.

  1. Lab on a chip genotyping for Brucella spp. based on 15-loci multi locus VNTR analysis

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    Marianelli Cinzia

    2009-04-01

    Full Text Available Abstract Background Brucellosis is an important zoonosis caused by the genus Brucella. In addition Brucella represents potential biological warfare agents due to the high contagious rates for humans and animals. Therefore, the strain typing epidemiological tool may be crucial for tracing back source of infection in outbreaks and discriminating naturally occurring outbreaks versus bioterroristic event. A Multiple Locus Variable-number tandem repeats (VNTR Analysis (MLVA assay based on 15 polymorphic markers was previously described. The obtained MLVA band profiles may be resolved by techniques ranging from low cost manual agarose gels to the more expensive capillary electrophoresis sequencing. In this paper a rapid, accurate and reproducible system, based on the Lab on a chip technology was set up for Brucella spp. genotyping. Results Seventeen DNA samples of Brucella strains isolated in Sicily, previously genotyped, and twelve DNA samples, provided by MLVA Brucella VNTR ring trial, were analyzed by MLVA-15 on Agilent 2100. The DNA fragment sizes produced by Agilent, compared with those expected, showed discrepancies; therefore, in order to assign the correct alleles to the Agilent DNA fragment sizes, a conversion table was produced. In order to validate the system twelve unknown DNA samples were analyzed by this method obtaining a full concordance with the VNTR ring trial results. Conclusion In this paper we described a rapid and specific detection method for the characterization of Brucella isolates. The comparison of the MLVA typing data produced by Agilent system with the data obtained by standard sequencing or ethidium bromide slab gel electrophoresis showed a general concordance of the results. Therefore this platform represents a fair compromise among costs, speed and specificity compared to any conventional molecular typing technique.

  2. Brucella Modulates Secretory Trafficking via Multiple Type IV Secretion Effector Proteins

    Science.gov (United States)

    Myeni, Sebenzile; Child, Robert; Ng, Tony W.; Kupko, John J.; Wehrly, Tara D.; Porcella, Stephen F.; Knodler, Leigh A.; Celli, Jean

    2013-01-01

    The intracellular pathogenic bacterium Brucella generates a replicative vacuole (rBCV) derived from the endoplasmic reticulum via subversion of the host cell secretory pathway. rBCV biogenesis requires the expression of the Type IV secretion system (T4SS) VirB, which is thought to translocate effector proteins that modulate membrane trafficking along the endocytic and secretory pathways. To date, only a few T4SS substrates have been identified, whose molecular functions remain unknown. Here, we used an in silico screen to identify putative T4SS effector candidate proteins using criteria such as limited homology in other bacterial genera, the presence of features similar to known VirB T4SS effectors, GC content and presence of eukaryotic-like motifs. Using β-lactamase and CyaA adenylate cyclase reporter assays, we identified eleven proteins translocated into host cells by Brucella, five in a VirB T4SS-dependent manner, namely BAB1_0678 (BspA), BAB1_0712 (BspB), BAB1_0847 (BspC), BAB1_1671 (BspE) and BAB1_1948 (BspF). A subset of the translocated proteins targeted secretory pathway compartments when ectopically expressed in HeLa cells, and the VirB effectors BspA, BspB and BspF inhibited protein secretion. Brucella infection also impaired host protein secretion in a process requiring BspA, BspB and BspF. Single or combined deletions of bspA, bspB and bspF affected Brucella ability to replicate in macrophages and persist in the liver of infected mice. Taken together, these findings demonstrate that Brucella modulates secretory trafficking via multiple T4SS effector proteins that likely act coordinately to promote Brucella pathogenesis. PMID:23950720

  3. Whole-genome analyses of the speciation events in the pathogenic Brucellae

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    Chain, P; Comerci, D; Tolmasky, M; Larimer, F; Malfatti, S; Vergez, L; Aguero, F; Land, M; Ugalde, R; Garcia, E

    2005-07-14

    Despite their high DNA identity and a proposal to group classical Brucella species as biovars of B. melitensis, the commonly recognized Brucella species can be distinguished by distinct biochemical and fatty acid characters as well as by a marked host range (e.g. B. suis for swine, B. melitensis for sheep and goats, B. abortus for cattle). Here we present the genome of B. abortus 2308, the virulent prototype biovar 1 strain, and its comparison to the two other human pathogenic Brucellae species and to the B. abortus field isolate 9-941. The global distribution of pseudogenes, deletions and insertions support previous indications that B. abortus and B. melitensis share a common ancestor that diverged from B. suis. With the exception of a dozen genes, the genetic complement of both B. abortus strains is identical, whereas the three species differ in gene content and pseudogenes. The pattern of species-specific gene inactivations affecting transcriptional regulators and outer membrane proteins suggest that these inactivations may play an important role in the establishment of host-specificity and may have been a primary driver of speciation in the Brucellae. Despite being non-motile, the Brucellae contain flagellum gene clusters and display species-specific flagellar gene inactivations, which lead to the putative generation of different versions of flagellum-derived structures, and may contribute to differences in host-specificity and virulence. Metabolic changes such as the lack of complete metabolic pathways for the synthesis of numerous compounds (e.g. glycogen, biotin, NAD, and choline) are consistent with adaptation of Brucellae to an intracellular lifestyle.

  4. Expression, purification and immunochemical characterization of recombinant OMP28 protein of Brucella species

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    Y. Manat

    2016-05-01

    Full Text Available Brucellosis is the lion’s share of infectious disease of animals and it has a particular socio-economic importance for the Republic of Kazakhstan. Sixty percent of epizootic outbreaks of brucellosis identified in the Commonwealth of Independent States (CIS originated from Kazakhstan in recent years. Definitive diagnosis of brucellosis remains a difficult task. Precisely for this reason, we evaluated a purified recombinant out membrane protein 28 (rOMP28 of Brucella species (Brucella spp. produced in Escherichia coli (E. coli as a diagnostic antigen in an Indirect ELISA (I-ELISA for bovine brucellosis. The gene encoding OMP28 was synthesized using a two-round PCR procedure. In order to produce the rOMP28, the de novo synthesized DNA was cloned into the expression vector pET-22b(+. Then, the rOMP28 was expressed in E. coli system and characterized in the present study. We further estimated the diagnostic potential of purified rOMP28 of Brucella spp. for screening bovine sera. To determine if rOMP28 has a valuable benefit for use in the serodiagnosis of bovine brucellosis, rOMP28-based I-ELISA was performed. Brucella spp. positive (n=62 and Brucella spp. negative (n=28 samples from tube agglutination test (TAT were positive (n=59 and negative (n=27 by I-ELISA, respectively. These findings show that the rOMP28 of Brucella spp. could be a good candidate for improving serological diagnostic methods for bovine brucellosis.

  5. Medulloblastoma and Brucellosis - Molecular Evidence of Brucella sp in Association with Central Nervous System Cancer

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    Binxue Zhang, Mina Izadjoo, Iren Horkayne-Szakaly, Alan Morrison, Douglas J. Wear

    2011-01-01

    Full Text Available Neurobrucellosis has been reported to cause lesions in a number of different locations in the central nervous system. Histologically or radiologically, these lesions were consistent with an infection. In response to parents who believed their child's brain tumor, histologically typical of medulloblastoma, was in reality neurobrucellosis, formalin-fixed paraffin-embedded tumor tissue from the medulloblastoma was sectioned, DNA extracted, and tested by polymerase chain reaction (PCR. Specific primer/probe sets, designed in our laboratory to target Brucella species, B. melitensis, B. abortus and B. suis, and designated OMP31, B-m, B-a and B-s, respectively, were used in TaqMan real-time PCR to amplify those gene targets in two separate blocks of the child's tumor. Sections from two blocks were positive only for Brucella species. Although the patient grew up in a European country known to harbor brucella in foods, such as unpasturized milk and cheese, the patient was seronegative for B. mellitensis, B. suis, and B. abortus. In an effort to test whether a relationship existed between the presence of brucella and medulloblastoma, 20 medulloblastomas were retrieved from the tissue repository of the AFIP. The above four primer/probe sets were again used to amplify brucella DNA. Five of 20 tumors (25% contained Brucella species DNA by the OMP31 primer/probe set. None of the 20 medulloblastomas had specific sequences for B. mellitensis, B. suis, or B. abortus. Is chronic brucellosis similar to other infectious agents such as helicobacter that is associated with tumor formation?

  6. Risk Factors Associated with Brucella Seropositivity in Sheep and Goats in Duhok Province, Iraq

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    Ali. G. Alhamada

    2017-12-01

    Full Text Available Sera from 432 small ruminants (335 sheep and 97 goats from 72 farms in Duhok Province, northern Iraq, were collected to investigate risk factors associated with brucellosis seropositivity. Serum samples were tested using the Rose Bengal test (RBT and an indirect enzyme-linked immunosorbent assay (iELISA. Using parallel interpretation, RBT and iELISA results showed that 31.7% (95% confidence interval (CI: 26.1, 36.3 of sheep and 34.0% (95% CI: 24.7, 44.3 of goats had antibodies against Brucella in the study area. A random-effects multivariable logistic regression model indicated that a higher chance of being seropositive (odds ratio (OR = 1.7; 95% 1.4; 2.2 was associated with an increase in the age of animals. The odds of Brucella seropositivity in flocks where sheep and goats grazed together was 2.0 times higher (95% CI: 1.08; 3.9 compared to flocks where sheep and goats grazed separately. The odds of Brucella seropositivity in small ruminants was 2.2 higher (95% CI: 1.2; 4.3 for animals originating from farms with a history of goat abortion in the preceding 12 months. In contrast, for every 1000 Iraqi Dinars (~0.85 US Dollar spent by the farmers on control of Brucella in their flocks, the odds of Brucella seropositivity decreased significantly (OR = 0.9, p-value = 0.021. The final model also indicated significant differences in Brucella seropositivity between the different districts of Duhok Province. This study provides a contribution to the epidemiology of brucellosis in small ruminants in northern Iraq.

  7. Rol del metabolismo de flavinas en la virulencia de Brucella abortus

    OpenAIRE

    Bonomi, Hernán Ruy

    2010-01-01

    Brucella spp. son las bacterias causantes de la brucelosis, la enfermedad zoonótica con más incidencia en el mundo. Brucella es un patógeno exitoso que probablemente infecta animales desde hace millones de años, y se encuentra extremadamente adaptado al estilo de vida intracelular. La virulencia de este patógeno reside en su capacidad de invadir al hospedador, resistir los mecanismos de defensa y, establecerse y replicar en el inhóspito nicho replicativo. El nicho replicativo se caracteriza p...

  8. Rapid identification of Yersinia pestis and Brucella melitensis by chip-based continuous flow PCR

    Science.gov (United States)

    Dietzsch, Michael; Hlawatsch, Nadine; Melzer, Falk; Tomaso, Herbert; Gärtner, Claudia; Neubauer, Heinrich

    2012-06-01

    To combat the threat of biological agents like Yersinia pestis and Brucella melitensis in bioterroristic scenarios requires fast, easy-to-use and safe identification systems. In this study we describe a system for rapid amplification of specific genetic markers for the identification of Yersinia pestis and Brucella melitensis. Using chip based PCR and continuous flow technology we were able to amplify the targets simultaneously with a 2-step reaction profile within 20 minutes. The subsequent analysis of amplified fragments by standard gel electrophoresis requires another 45 minutes. We were able to detect both pathogens within 75 minutes being much faster than most other nucleic acid amplification technologies.

  9. Detection of Brucella species in organs of naturally infected cattle by polymerase chain reaction.

    Science.gov (United States)

    Gallien, P; Dorn, C; Alban, G; Staak, C; Protz, D

    1998-05-09

    A polymerase chain reaction (PCR) assay was used to detect Brucella species in the uterus, udder, spleen, lymph nodes, kidney and liver of three cows which had been naturally infected in an outbreak of brucellosis, and the results were compared with the results of bacteriological investigations. All 18 samples reacted positively in the PCR, but five samples had weak bands after the electrophoretic separation of PCR mixtures. No Brucella strains could be detected in these five samples by bacterial cultivation, but all the other samples gave positive results. A pre-enrichment procedure was necessary for the PCR. A PCR with DNA from eight Yersinia strains gave no amplification product.

  10. Evaluation of oxidative stress in brucella infected cows

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    N. Kataria

    2010-05-01

    Full Text Available Oxidative stress can influence the metabolism of cells in vital organs of the body. Oxidative stress is extremely dangerous as it does not exhibit any symptom and is recognisable with great difficulty by means of laboratory methods. It can be monitored with several biomarkers like antioxidants and pro-oxidants which can be assessed in serum. The inexorableness of exposure of cows to brucella infection makes oxidative stress associated with this infection an appropriate field of investigation. There is paucity of work to detect stress, which is essential to take timely corrective measures and to save the animal population. Therefore the investigation was carried out to evaluate oxidative stress in the cows suffering from brucellosis. For this serum iomarkers of oxidative stress viz. vitamin C, vitamin E, catalase, monoamine oxidase, glutathione reductase, superoxide dismutase, glutathione, xanthine oxidase, oxidase and peroxidase were determined. Results indicated that vitamin C, vitamin E and glutathione activity decreased significantly in affected cows as compared to healthy cows. Serum catalase, superoxide dismutase, monoamine oxidase, glutathione reductase, xanthine oxidase, oxidase and peroxidase activities increased significantly in affected cows as compared to healthy cows. Decreased activity of vitamin C, vitamin E and glutathione indicated towards their depletion which generally occurs in the oxidative stress to scavenge the free radicals. It was concluded that oxidative stress was there in the animals. This study recommends the use of antioxidants in affected cows

  11. Male rats transmit Brucella abortus biotype 1 through sexual intercourse.

    Science.gov (United States)

    Islam, Md Ariful; Khatun, Mst Minara; Baek, Byeong-Kirl

    2013-08-30

    The aim of this study was to evaluate transmission of Brucella abortus biotype 1 via sexual intercourse in rats. Male and female virgin Sprague-Dawley (SD) rats were experimentally infected intraperitoneally with 1×10(9)colony forming units (CFU) of B. abortus biotype 1, a Korean bovine isolate. At 14 days after infection, infected male rats (n=10) were housed with uninfected female rats (n=10) and infected female rats (n=10) were housed with uninfected male rats (n=10) for a period of one month. During this period all uninfected female rats became pregnant and 6 of 10 infected female rats became pregnant. Serum from two out of 10 female uninfected rats had positive reactions in the Rose Bengal Plate Agglutination Test (RBPAT), Tube Agglutination Test (TAT) or the Enzyme-Linked Immunosorbent Assay (ELISA); whereas none of the uninfected male rat had positive reactions in these tests. Using bacteriological culture and AMOS-PCR assay, B. abortus biotype 1 was isolated and identified from two uninfected female rats and all of the uninfected male rats were found negative for B. abortus biotype 1. It was concluded that transmission of B. abortus biotype 1 from infected male to uninfected female rats resulted from sexual intercourse. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Brucella abortus RB51 in milk of vaccinated adult cattle.

    Science.gov (United States)

    Miranda, Karina Leite; Poester, Fernando Padilla; Dorneles, Elaine Maria Seles; Resende, Thiago Magalhães; Vaz, Adil Knackfuss; Ferraz, Sandra Maria; Lage, Andrey Pereira

    2016-08-01

    The aim of this study was to evaluate the shedding of Brucella abortus in the milk of cows vaccinated with a full dose of RB51 during lactation. Eighteen cows, nine previously vaccinated with S19 as calves and nine non-vaccinated, were immunized subcutaneously with 1.3×10(10)CFU of B. abortus RB51, 30-60days after parturition. Milk samples from all animals were collected daily until day 7, and at weekly interval for the next 9 weeks after vaccination. To evaluate the shedding of B. abortus, milk samples were submitted for culture and PCR. No B. abortus was isolated from any sample tested. Only one sample, collected on first day after vaccination from a cow previously vaccinated, was faintly positive in the PCR. In conclusion, the public health hazard associated with milk consumption from cows vaccinated with RB51 in post-partum is very low, despite vaccination with the full dose and regardless of previous S19 vaccination. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Safety of Brucella abortus strain RB51 in deer mice.

    Science.gov (United States)

    Cook, W E; Williams, E S; Thorne, E T; Taylor, S K; Anderson, S

    2001-07-01

    Brucella abortus strain RB51 is an approved brucellosis vaccine for use in cattle that may have potential as an oral vaccine for use in elk (Cervus elaphus) and/or bison (Bison bison). This study was designed to determine effects of strain RB51 on deer mice (Peromyscus maniculatus), a nontarget species that could have access to treated baits in a field situation. In February 1994, 90 mice were orally dosed or intraperitoneally injected with 1 x 10(8) colony forming units strain RB51 and 77 controls were similarly dosed with sterile saline. At weekly intervals through early April 1994, 4 to 6 mice from each group were euthanized, gross necropsies performed, spleens and uteruses cultured, and tissues examined histologically. All orally inoculated mice cleared the infection by 6 wk post-inoculation (PI). While most of the injected mice cleared the infection by 7 wk PI, a few required 9 wk. There were minimal adverse effects attributable to strain RB51. Apparently, strain RB51 would not negatively impact P. maniculatus populations if it were used in a field situation. Also, deer mice appear to be able to clear the vaccine in 6 to 9 wk, thus the probability of these mice transmitting the vaccine to other animals is low.

  14. Seroprevalence of Brucella abortus and Leptospira hardjo in cattle

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    S. Jegaveera Pandian

    2015-02-01

    Full Text Available Aim: The aim was to assess the seroprevalence of B. abortus and Leptospira hardjo in the cattle population of Bihar, this work was carried out. Materials and Methods: Randomly selected 450 cattle from nine districts of Bihar were serologically screened for antibodies against L. hardjo and B. abortus. DAS-ELISA for leptospira and AB-ELISA for brucella were carried out. Based on the results prevalence in each district and the state are reported herewith. Results: In this study, it was found that the seroprevalence of L. hardjo was 9.11% and that of B. abortus was 12.2% in Bihar. Indigenous cattle were found to be less susceptible to leptospirosis and brucellosis even though they accounted for 83.11% of the study population. Conclusion: Although there was no acute disease, antibodies detected against L. hardjo and B. abortus in the cattle population indicated the presence of chronic and subclinical infection, which could challenge the fertility of the animals.

  15. ENCUESTA EXPLORATORIA DE INFECCION POR Brucella canis EN PERROS DE

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    Ana Pardo D

    2009-08-01

    Full Text Available Objetivo. Determinar la presencia de anticuerpos a B. canis en perros domésticos y callejeros del municipio de Villavicencio, Colombia. Materiales y métodos. Se utilizó la prueba de aglutinación rápida en placa con antígeno menos mucoide (M- en 201 muestras de suero. En dos animales seropositivos se realizó intento de aislamiento por hemocultivo en medio selectivo para Brucella (oxoid®. En un animal seropositivo, con crecimiento bacteriano con características morfológicas sugestivas a B. canis se realizó histopatología de testículo, bazo e hígado. Resultados. La seropositividad general fue de 1.49% y correspondió a tres caninos machos, dos de los cuales presentaron signos clínicos de epididimitis y orquitis (unilateral. El cultivo y la histopatología no fueron concluyentes para el diagnostico de B. canis. Conclusiones. La seropositividad fue baja y sugiere que la población estudiada no ha estado en contacto con la bacteria. La presencia de reactores puede estar asociado con falsos positivos. El no aislamiento de la bacteria no indica que la enfermedad no exista por lo que se requiere de nuevos estudios.

  16. Brucella abortus: pathogenicity and gene regulation of virulence

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    Olga Rivas-Solano

    2015-06-01

    Full Text Available Brucella abortus is a zoonotic intracellular facultative pathogen belonging to the subdivision α2 of class Proteobacteria. It causes a worldwide distributed zoonotic disease called brucellosis. The main symptoms are abortion and sterility in cattle, as well as an undulant febrile condition in humans. In endemic regions like Central America, brucellosis has a high socioeconomic impact. A basic research project was recently conducted at the ITCR with the purpose of studying gene regulation of virulence, structure and immunogenicity in B. abortus. The present review was written as part of this project. B. abortus virulence seems to be determined by its ability to invade, survive and replicate inside professional and non-professional phagocytes. It reaches its intracellular replicative niche without the activation of host antimicrobial mechanisms of innate immunity. It also has gene regulation mechanisms for a rapid adaptation to an intracellular environment such as the two-component signal transduction system BvrR/BvrS and the quorum sensing regulator called Vjbr, as well as other transcription factors. All of them integrate a complex gene regulation network.

  17. Brucella ovis: invasion, traffic, virulence factors and immune responseBrucella ovis: invasão, tráfego, fatores de virulência e resposta imune

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    João Marcelo Azevedo de Paula Antunes

    2013-06-01

    Full Text Available Brucellosis remains an economic problem in animals and public health. Worldwide ovine brucellosis caused by Brucella ovis is considered a major cause of infertility in sheep. The factors responsible for persistence of the agent in these locations are not known, as well as the mechanisms involved in immune defense and possibly the persistence of the agent. Brucella spp. induces moderate inflammatory response. The nature of the intracellular agent stimulates immune response of the type 1 helper T lymphocytes. Studies of the pathogenesis of ovine brucellosis are scarce. Recent developments have shown that the inflammatory response induced by moderate brucelas represent probably the result of an attempt to escape the immune response and suppression of host immune response. Were reviewed by the mechanisms described by brucelas and Brucella ovis for penetration into the host, escape of the immune response and the immune response generated by the infection. A brucelose permanece como problema econômico em animais e de saúde pública. Em todo o mundo a brucelose ovina ocasionada pela Brucella ovis é considerada uma das principais causas de infertilidade em ovinos. Os fatores responsáveis pela persistência do agente nestes locais não são conhecidos, bem como os mecanismos imunes envolvidos na defesa e eventualmente na persistência do agente. Brucella spp. induz resposta inflamatória moderada. A natureza intracelular do agente estimula resposta imune celular do tipo linfócito T helper 1. Os estudos de patogenia da brucelose ovina são escassos. Recentes avanços demonstraram que a resposta inflamatória moderada induzida pelas brucelas representam provavelmente o resultado de tentativa de escape da resposta imune e supressão da resposta imune hospedeira. Foram revisados os mecanismos descritos pelas brucelas e pela Brucella ovis para penetração no hospedeiro, escape da resposta imune, bem como a resposta imunológica gerada pela infecção.

  18. Effect of polymyxin B and environmental conditions on isolation of Brucella species and the vaccine strain RB51.

    Science.gov (United States)

    Jensen, Allen E; Halling, Shirley M

    2010-03-01

    Brucella are resistant to polymyxin B (PB), but their relative susceptibility to PB and its derivative, colistin (COL) has not been rigorously or systematically studied. Comparative susceptibility of Brucella reference strains, vaccine strain RB51, and Brucella isolates from marine mammals to these two cationic peptides were determined by Etest. Vast differences among Brucella species were found in susceptibility to both PB and COL. Brucella demonstrated similar pattern of relative susceptibility to PB as that of COL, but they were less susceptible to COL. Both B. melitensis and B. suis were the least susceptible to polymyxins and rough strains were more susceptible to both PB and COL than the smooth except for the BvrR mutant. Strains were generally less susceptible to PB when cultured in CO(2) rather than ambient air; some became more susceptible in acidified medium. Results show that environment cultural conditions must be considered when selecting for CO(2)-independent strains of Brucella especially the vaccine strain RB51 on selective media containing PB. Our observations extend basic knowledge of the differential resistance of Brucella to polymyxins. Published by Elsevier Ltd.

  19. The Bacterial Second Messenger Cyclic di-GMP Regulates Brucella Pathogenesis and Leads to Altered Host Immune Response.

    Science.gov (United States)

    Khan, Mike; Harms, Jerome S; Marim, Fernanda M; Armon, Leah; Hall, Cherisse L; Liu, Yi-Ping; Banai, Menachem; Oliveira, Sergio C; Splitter, Gary A; Smith, Judith A

    2016-12-01

    Brucella species are facultative intracellular bacteria that cause brucellosis, a chronic debilitating disease significantly impacting global health and prosperity. Much remains to be learned about how Brucella spp. succeed in sabotaging immune host cells and how Brucella spp. respond to environmental challenges. Multiple types of bacteria employ the prokaryotic second messenger cyclic di-GMP (c-di-GMP) to coordinate responses to shifting environments. To determine the role of c-di-GMP in Brucella physiology and in shaping host-Brucella interactions, we utilized c-di-GMP regulatory enzyme deletion mutants. Our results show that a ΔbpdA phosphodiesterase mutant producing excess c-di-GMP displays marked attenuation in vitro and in vivo during later infections. Although c-di-GMP is known to stimulate the innate sensor STING, surprisingly, the ΔbpdA mutant induced a weaker host immune response than did wild-type Brucella or the low-c-di-GMP guanylate cyclase ΔcgsB mutant. Proteomics analysis revealed that c-di-GMP regulates several processes critical for virulence, including cell wall and biofilm formation, nutrient acquisition, and the type IV secretion system. Finally, ΔbpdA mutants exhibited altered morphology and were hypersensitive to nutrient-limiting conditions. In summary, our results indicate a vital role for c-di-GMP in allowing Brucella to successfully navigate stressful and shifting environments to establish intracellular infection. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  20. Epidemiology of Brucella infection in the human, livestock and wildlife interface in the Katavi-Rukwa ecosystem, Tanzania.

    Science.gov (United States)

    Assenga, Justine A; Matemba, Lucas E; Muller, Shabani K; Malakalinga, Joseph J; Kazwala, Rudovick R

    2015-08-08

    Brucellosis is a zoonosis of public health importance worldwide. In Tanzania, the disease is underreported due to insufficient awareness, inadequate diagnostic protocols, including lack of appropriate reagents for diagnosis. Livestock and wildlife are considered potential sources of infection to humans; however, the role played by these carriers in the epidemiology of the disease in the ecosystems in Tanzania is not fully understood. The objective of this study was to establish the prevalence of anti-Brucella antibodies in humans, wildlife and livestock; and molecular prevalence of Brucella spp in cattle and goats in the Katavi- Rukwa ecosystem. Anti-Brucella antibodies were detected in humans at 0.6 % (95 % CI: 0.1, 2.1 %); cattle at 6.8 % (95 % CI: 5.4, 8.5 %), goats at 1.6 % (95 % CI: 0.4, 4.1 %) and buffaloes at 7.9 % (95 % CI: 1.7, 21.4 %). One of the two sampled lions tested positive. Cattle had a significantly higher prevalence of anti-Brucella antibodies as compared to goats (P Brucella infection. Eight (3.5 %) out of 231 milk samples tested were positive for Brucella spp on Polymerase Chain Reaction (PCR), and Brucella abortus biovar 1 was detected in cattle milk. However, no Brucella spp were detected in goat milk. This study has shown the presence of anti- Brucella antibodies in humans, livestock, and wildlife in the Katavi- Rukwa ecosystem. Transmission of the infection between wildlife, livestock and humans is likely to continue due to increasing human activities in the human wildlife interface. This information is an important contribution to public health policy development in the human wildlife interface of the Katavi- Rukwa ecosystem.

  1. Identification of species and biotypes of the brucella genus in apparently healthy and aborted ewes and goats in Egypt.

    Science.gov (United States)

    Abdel-Ghani, M; Abdel-Hamed, S; Nada, S M; Osman, K

    1984-12-01

    Brucella micro-organisms was absent in the vaginal swabs collected from apparently healthy ewes. Such incidence among the genital tract of aborted ewes was 4.84%. Among goats, brucella species represented 7.04% in aborted goats, while it was recovered in lower percentage (0.92%) from the vaginal swabs of apparently healthy goats. Br. melitensis biotype "3" was the sole species recovered from aborted ewes, while biotypes "3" and "2" could be obtained from clinically healthy and aborted foeti of goats. The predilection seats of brucella in the genital tracts of aborted animals and their foeti were discussed in details.

  2. Rapid and quantitative detection of Brucella by up-converting phosphor technology-based lateral-flow assay.

    Science.gov (United States)

    Qu, Qing; Zhu, Ziwen; Wang, Yufei; Zhong, Zhijun; Zhao, Jin; Qiao, Feng; Du, Xinying; Wang, Zhoujia; Yang, Ruifu; Huang, Liuyu; Yu, Yaqin; Zhou, Lei; Chen, Zeliang

    2009-10-01

    A rapid and quantitative up-converting phosphor technology-based later-flow assay (UPT-LF assay) was developed for on-site detection of Brucella. Different Brucella species both in pure cultures and in spiked samples could be quantitatively detected. The detection limit for pure culture was 5 x 10(6)CFU/ml and the sensitivity for different spiked samples ranged from 2.0 x 10(3) to 3.9 x 10(5)CFU/mg. The UPT-LF assay showed high specificity, reproducibility and stability, providing great potential for Brucella on-site detection.

  3. Characterization of novel Omp31 antigenic epitopes of Brucella melitensis by monoclonal antibodies.

    Science.gov (United States)

    Li, Jinfeng; Hu, Feihuan; Chen, Shouyi; Luo, Peifang; He, Zuoping; Wang, Wenjing; Allain, Jean-Pierre; Li, Chengyao

    2017-05-15

    Brucellosis is a severe zoonotic disease worldwide. Detection and identification of Brucella species are essential to prevent or treat brucellosis in humans and animals. The outer membrane protein-31 (Omp31) is a major protein of Brucellae except for B. abortus, while the Omp31 antigenic epitopes have not been extensively characterized yet. A total of 22 monoclonal antibodies (mAbs) were produced against Omp31 of Brucella (B.) melitensis, of which 13 recognized five linear epitopes, 7 reacted with semi-conformational epitopes and 2 reacted with conformational epitopes, respectively. The mAb isotypes were 11 (50%) IgG2a, 5 (23%) IgG1 and 6 (27%) IgM. On the basis of epitope recognition and reactivity levels, 8 mAbs including 3 IgM and 5 IgG clones were considered as highly reactive and potentially diagnostic antibodies. Among these mAbs, 7A3 (IgG1), 5B1 (IgG2a), 2C1 (IgG2a) and 5B3 (IgG2a) reacted with differently conserved linear epitopes of B. melitensis, B. ovis, B. suis and B. canis strains, while 5H3 (IgG2a) highly reacted with a conformational epitope of Omp31 when tested with several immunoassays. These potent monoclonal antibodies can be used for identifying Omp31 antigens or detecting B. melitensis and other Brucella species beyond B. abortus in vitro or in vivo.

  4. Molecular epidemiology and antibiotic susceptibility of livestock Brucella melitensis isolates from Naryn Oblast, Kyrgyzstan.

    Science.gov (United States)

    Kasymbekov, Joldoshbek; Imanseitov, Joldoshbek; Ballif, Marie; Schürch, Nadia; Paniga, Sandra; Pilo, Paola; Tonolla, Mauro; Benagli, Cinzia; Akylbekova, Kulyash; Jumakanova, Zarima; Schelling, Esther; Zinsstag, Jakob

    2013-01-01

    The incidence of human brucellosis in Kyrgyzstan has been increasing in the last years and was identified as a priority disease needing most urgent control measures in the livestock population. The latest species identification of Brucella isolates in Kyrgyzstan was carried out in the 1960s and investigated the circulation of Brucella abortus, B. melitensis, B. ovis, and B. suis. However, supporting data and documentation of that experience are lacking. Therefore, typing of Brucella spp. and identification of the most important host species are necessary for the understanding of the main transmission routes and to adopt an effective brucellosis control policy in Kyrgyzstan. Overall, 17 B. melitensis strains from aborted fetuses of sheep and cattle isolated in the province of Naryn were studied. All strains were susceptible to trimethoprim-sulfamethoxazole, gentamicin, rifampin, ofloxacin, streptomycin, doxycycline, and ciprofloxacin. Multilocus variable number tandem repeat analysis showed low genetic diversity. Kyrgyz strains seem to be genetically associated with the Eastern Mediterranean group of the Brucella global phylogeny. We identified and confirmed transmission of B. melitensis to cattle and a close genetic relationship between B. melitensis strains isolated from sheep sharing the same pasture.

  5. Detection of Brucella sp. and Leptospira sp. in dogs using conventional polymerase chain reaction

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    Khamesipour Faham

    2014-12-01

    Full Text Available The study was conducted to detect Brucella sp. and Leptospira sp. in blood samples of dogs in Isfahan and Shahrekord province in Iran. A total of 94 blood samples were collected from dogs of different breed, age, sex, and dogs’ type (stray or nonstray. The samples were examined using conventional polymerase chain reaction (PCR. Fourteen (14.89% dogs were positive for Brucella sp. and 18 (19.15%. dogs for Leptospira sp. There were no significant differences between the prevalence of the pathogens, provinces, sex, and age groups (P > 0.05. However, there was a statistically significant difference in prevalence of Brucella sp. and Leptospira sp. between stray and non-stray dogs (P < 0.0001; χ2 = 30.3767. The study also demonstrated that PCR was successfully used for the first time in Iran for the detection of Brucella sp. and Leptospira sp. in blood samples of dogs. Therefore, we recommend the PCR as a supplementary method with other commonly recognised methods (e.g. serological methods for the diagnosis of subclinical infections with the microorganisms. Strict measures for the control of stray dogs are also highly recommended.

  6. Molecular epidemiology and antibiotic susceptibility of livestock Brucella melitensis isolates from Naryn Oblast, Kyrgyzstan.

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    Joldoshbek Kasymbekov

    Full Text Available The incidence of human brucellosis in Kyrgyzstan has been increasing in the last years and was identified as a priority disease needing most urgent control measures in the livestock population. The latest species identification of Brucella isolates in Kyrgyzstan was carried out in the 1960s and investigated the circulation of Brucella abortus, B. melitensis, B. ovis, and B. suis. However, supporting data and documentation of that experience are lacking. Therefore, typing of Brucella spp. and identification of the most important host species are necessary for the understanding of the main transmission routes and to adopt an effective brucellosis control policy in Kyrgyzstan. Overall, 17 B. melitensis strains from aborted fetuses of sheep and cattle isolated in the province of Naryn were studied. All strains were susceptible to trimethoprim-sulfamethoxazole, gentamicin, rifampin, ofloxacin, streptomycin, doxycycline, and ciprofloxacin. Multilocus variable number tandem repeat analysis showed low genetic diversity. Kyrgyz strains seem to be genetically associated with the Eastern Mediterranean group of the Brucella global phylogeny. We identified and confirmed transmission of B. melitensis to cattle and a close genetic relationship between B. melitensis strains isolated from sheep sharing the same pasture.

  7. [Molecular-genetic characterization of canine and rangiferine brucella isolates from different regions of Russia].

    Science.gov (United States)

    Kulakov, Iu K; Tsirel'son, L E; Zheludkov, M M

    2012-01-01

    Comparative molecular-genetic characterization of Brucella isolates from dogs and reindeers in Russia by molecular-genetic typing methods. 19 canine and 2 rangiferine Brucella isolates were studied by molecular typing methods based on PCR for differential species and biovar specific molecular targets and MLVA (multiple locus variable number tandem repeats analysis) using primers to 12 known variable loci. Using PCR for differential molecular targets, canine Brucella isolates were characterized as B. canis and rangiferine isolates as B. suis biovar 4. MLVA revealed 5 identical and 7 variable MLVA loci. Using the dendrogram. all the isolates on the data of 12 loci were classified into the close related cluster. On the other hand, high discrimination power of MLVA with a resulting Hunter and Gaston discriminatory index (HGDI) of 0.9842 was shown to reveal genetic diversity for the isolates of 17 MLVA genotypes. B. canis and B. suis isolates from different geographical regions in Russia were genetically close related, thereby confirming known genetic relationship between these species. Related MLVA genotypes of isolates were connected to certain regions of preliminary isolation in Russia. To improve the system ofbrucellosis surveillance in Russia MLVA typing of more canine and rangiferine Brucella isolates having epidemiological danger for humans is required to be studied.

  8. Experimental Infection of Richardson's Ground Squirrels (Spermophilus richardsonii) with Attenuated and Virulent Strains of Brucella abortus

    Science.gov (United States)

    Exposure of non-target species to wildlife vaccines is an important concern when evaluating a candidate vaccine for use in the field. A previous investigation of the safety of Brucella abortus strain RB51 (sRB51) in various non-target species suggested that Richardson’s ground squirrels (Spermophil...

  9. Molecular Epidemiology and Antibiotic Susceptibility of Livestock Brucella melitensis Isolates from Naryn Oblast, Kyrgyzstan

    Science.gov (United States)

    Kasymbekov, Joldoshbek; Imanseitov, Joldoshbek; Ballif, Marie; Schürch, Nadia; Paniga, Sandra; Pilo, Paola; Tonolla, Mauro; Benagli, Cinzia; Akylbekova, Kulyash; Jumakanova, Zarima; Schelling, Esther; Zinsstag, Jakob

    2013-01-01

    The incidence of human brucellosis in Kyrgyzstan has been increasing in the last years and was identified as a priority disease needing most urgent control measures in the livestock population. The latest species identification of Brucella isolates in Kyrgyzstan was carried out in the 1960s and investigated the circulation of Brucella abortus, B. melitensis, B. ovis, and B. suis. However, supporting data and documentation of that experience are lacking. Therefore, typing of Brucella spp. and identification of the most important host species are necessary for the understanding of the main transmission routes and to adopt an effective brucellosis control policy in Kyrgyzstan. Overall, 17 B. melitensis strains from aborted fetuses of sheep and cattle isolated in the province of Naryn were studied. All strains were susceptible to trimethoprim-sulfamethoxazole, gentamicin, rifampin, ofloxacin, streptomycin, doxycycline, and ciprofloxacin. Multilocus variable number tandem repeat analysis showed low genetic diversity. Kyrgyz strains seem to be genetically associated with the Eastern Mediterranean group of the Brucella global phylogeny. We identified and confirmed transmission of B. melitensis to cattle and a close genetic relationship between B. melitensis strains isolated from sheep sharing the same pasture. PMID:23469294

  10. Brucella abortus is Prevalent in Both Humans and Animals in Bangladesh.

    Science.gov (United States)

    Rahman, A K M A; Saegerman, C; Berkvens, D; Melzer, F; Neubauer, H; Fretin, D; Abatih, E; Dhand, N; Ward, M P

    2017-08-01

    To determine the role of different Brucella (B.) spp. in Bangladesh, 62 animal samples and 500 human sera were tested. Animal samples from cattle, goats and sheep (including milk, bull semen, vaginal swabs and placentas) were cultured for Brucella spp. Three test-positive human sera and all animal samples were screened by Brucella genus-specific real-time PCR (RT-PCR), and positive samples were then tested by IS711 RT-PCR to detect B. abortus and B. melitensis DNA. Only B. abortus DNA was amplified from 13 human and six animal samples. This is the first report describing B. abortus as the aetiological agent of brucellosis in occupationally exposed humans in Bangladesh. Of note is failure to detect B. melitensis DNA, the species most often associated with human brucellosis worldwide. Further studies are required to explore the occurrence of Brucella melitensis in Bangladesh. © 2017 Blackwell Verlag GmbH.

  11. Meningoencephalitis and Listeria monocytogenes, Toxoplasma gondii and Brucella spp. coinfection in a dolphin in Italy.

    Science.gov (United States)

    Grattarola, Carla; Giorda, Federica; Iulini, Barbara; Pintore, Maria Domenica; Pautasso, Alessandra; Zoppi, Simona; Goria, Maria; Romano, Angelo; Peletto, Simone; Varello, Katia; Garibaldi, Fulvio; Garofolo, Giuliano; Di Francesco, Cristina Esmeralda; Marsili, Letizia; Bozzetta, Elena; Di Guardo, Giovanni; Dondo, Alessandro; Mignone, Walter; Casalone, Cristina

    2016-02-25

    Listeria monocytogenes, Toxoplasma gondii and Brucella spp. can infect a wide range of species, including humans. In cetaceans, meningoencephalitis has been associated with T. gondii and Brucella spp. infection, whereas to our knowledge, L. monocytogenes infection has not previously been reported. Meningoencephalitis and L. monocytogenes, T. gondii and Brucella spp. were identified by means of both direct and indirect laboratory techniques in an adult female striped dolphin Stenella coeruleoalba found stranded in January 2015 on the Ligurian Sea coast, northwestern Italy. The animal was emaciated, and histopathology disclosed severe meningoencephalitis. The nature of the inflammatory response and intra-lesional protozoa were consistent with a mixed infection by L. monocytogenes, T. gondii and Brucella spp. We believe this is an unprecedented case of infection by 3 zoonotic pathogens and also the first bacteriologically confirmed case report of neurolisteriosis in cetaceans. Cerebral toxoplasmosis and neurobrucellosis may have led to the animal's disorientation and stranding, with L. monocytogenes having likely exacerbated the coinfection leading to the demise of this dolphin.

  12. Immunogenicity and efficacy of a rough Brucella suis vaccine delivered orally or parenterally to feral swine

    Science.gov (United States)

    Brucella suis strain 353-1 is a stable vaccine strain that is clinically safe, does not cause positive serologic responses on conventional brucellosis surveillance tests, and induces humoral and cellular immunity in swine after vaccination. In this study, we evaluated tissue clearance and immunologi...

  13. Identification and typing of Brucella spp. in stranded harbour porpoises (Phocoena phocoena) on the Dutch coast.

    NARCIS (Netherlands)

    Maio, E.; Begeman, L.; Bisselink, Y.J.W.M.; Tulden, van P.W.; Wiersma, L.; Hiemstra, S.; Ruuls, R.; Gröne, A.; Roest, H.I.J.; Willemsen, P.T.J.; Giessen, van der J.

    2014-01-01

    The presence of Brucella (B.) spp. in harbour porpoises stranded between 2008 and 2011 along the Dutch coast was studied. A selection of 265 tissue samples from 112 animals was analysed using conventional and molecular methods. In total, 4.5% (5/112) of the animals corresponding with 2.3% (6/265)

  14. Et tilfælde af brucella spondylodiscitis efter rejse til Libanon

    DEFF Research Database (Denmark)

    Nielsen, Stig Lønberg; Johansen, Isik Somuncu

    2012-01-01

    Brucellosis is a widespread endemic zoonotic infection affecting more than 500,000 people per year. The disease is very uncommon in Denmark and almost always imported. We present a case of a 57 year-old male with blood culture and magnetic resonance imaging verified brucella spondylodiscitis. Prior...

  15. Molecular epidemiology of Brucella genotypes in patients at a major hospital in central Peru

    NARCIS (Netherlands)

    Nöckler, Karsten; Maves, Ryan; Cepeda, David; Draeger, Angelika; Mayer-Scholl, Anne; Chacaltana, Jesus; Castañeda, María; Espinosa, Benjamin; Castillo, Rosa; Hall, Eric; Al Dahouk, Sascha; Gilman, Robert H.; Cabeza, Franco; Smits, Henk L.

    2009-01-01

    The multiple-locus variable-number repeat analysis of 90 human Brucella melitensis isolates from a large urban area in central Peru revealed variations at 4 (Bruce07, Bruce09, Bruce18, and Bruce42) out of 16 loci investigated, of which 1 (Bruce42) also is used for species identification. Ten

  16. Modeling the survivability of brucella to exposure of Ultraviolet radiation and temperature

    Science.gov (United States)

    Howe, R.

    Accumulated summation of daily Ultra Violet-B (UV-B = 290? to 320 ? ) data? from The USDA Ultraviolet Radiation Monitoring Program show good correlation (R^2 = 77%) with daily temperature data during the five month period from February through June, 1998. Exposure of disease organisms, such as brucella to the effects of accumulated UV-B radiation, can be modeled for a 5 month period from February through June, 1998. Estimates of a lethal dosage for brucell of UV-B in the environment is dependent on minimum/maximum temperature and Solar Zenith Angle for the time period. The accumulated increase in temperature over this period also effects the decomposition of an aborted fetus containing brucella. Decomposition begins at some minimum daily temperature at 27 to 30 degrees C and peaks at 39 to 40C. It is useful to view the summation of temperature as a threshold for other bacteria growth, so that accumulated temperature greater than some value causes decomposition through competition with other bacteria and brucella die from the accumulated effects of UV-B, temperature and organism competition. Results of a study (Cook 1998) to determine survivability of brucellosis in the environment through exposure of aborted bovine fetuses show no one cause can be attributed to death of the disease agent. The combination of daily increase in temperature and accumulated UV-B radiation reveal an inverse correlation to survivability data and can be modeled as an indicator of brucella survivability in the environment in arid regions.

  17. Prevalence of Brucella,/i> antibodies in donkeys ( Equus asinus ) in ...

    African Journals Online (AJOL)

    A cross-sectional study was designed to study the serological prevalence of antibodies against Brucella spp by using Rose Bengal Plate Test (RBPT) and Microtitre Serum Agglutination Test (MSAT). A total of six hundred (600) adult donkeys comprising of 393 males and 207 females were sampled from three local ...

  18. Reliable identification at the species level of Brucella isolates with MALDI-TOF-MS

    NARCIS (Netherlands)

    Lista, F.; Reubsaet, F.A.G.; Santis, R. de; Parchen, R.R.; Jong, A.L. de; Kieboom, J.; Laaken, A.L. van der; Voskamp-Visser, I.A.I.; Fillo, S.; Jansen, H.J. de; Plas, J. van der; Paauw, A.

    2011-01-01

    Background: The genus Brucella contains highly infectious species that are classified as biological threat agents. The timely detection and identification of the microorganism involved is essential for an effective response not only to biological warfare attacks but also to natural outbreaks.

  19. Colony-level assessment of Brucella and Leptospira in the Guadalupe fur seal, Isla Guadalupe, Mexico.

    Science.gov (United States)

    Ziehl-Quirós, E Carolina; García-Aguilar, María C; Mellink, Eric

    2017-01-24

    The relatively small population size and restricted distribution of the Guadalupe fur seal Arctocephalus townsendi could make it highly vulnerable to infectious diseases. We performed a colony-level assessment in this species of the prevalence and presence of Brucella spp. and Leptospira spp., pathogenic bacteria that have been reported in several pinniped species worldwide. Forty-six serum samples were collected in 2014 from pups at Isla Guadalupe, the only place where the species effectively reproduces. Samples were tested for Brucella using 3 consecutive serological tests, and for Leptospira using the microscopic agglutination test. For each bacterium, a Bayesian approach was used to estimate prevalence to exposure, and an epidemiological model was used to test the null hypothesis that the bacterium was present in the colony. No serum sample tested positive for Brucella, and the statistical analyses concluded that the colony was bacterium-free with a 96.3% confidence level. However, a Brucella surveillance program would be highly recommendable. Twelve samples were positive (titers 1:50) to 1 or more serovars of Leptospira. The prevalence was calculated at 27.1% (95% credible interval: 15.6-40.3%), and the posterior analyses indicated that the colony was not Leptospira-free with a 100% confidence level. Serovars Icterohaemorrhagiae, Canicola, and Bratislava were detected, but only further research can unveil whether they affect the fur seal population.

  20. Brucella microti sp. nov., isolated from the common vole Microtus arvalis

    Czech Academy of Sciences Publication Activity Database

    Scholz, H. C.; Hubálek, Zdeněk; Sedláček, I.; Vergnaud, G.; Tomaso, H.; Al Dahouk, S.; Melzer, F.; Kämpfer, P.; Neubauer, H.; Cloeckaert, A.; Maquart, M.; Zygmunt, M. S.; Whatmore, A. M.; Falsen, E.; Bahn, P.; Göllner, C.; Pfeffer, M.; Huber, B.; Busse, H.-J.; Nöckler, K.

    2008-01-01

    Roč. 58, č. 2 (2008), s. 375-382 ISSN 1466-5026 Institutional research plan: CEZ:AV0Z60930519 Keywords : common vole * Brucella microti * rodent brucellosis Subject RIV: EE - Microbiology, Virology Impact factor: 2.222, year: 2008

  1. Identification and typing of Brucella spp. in stranded harbour porpoises (Phocoena phocoena) on the Dutch coast

    NARCIS (Netherlands)

    Maio, Elisa; Begeman, Lineke; Bisselink, Yvette; van Tulden, Peter; Wiersma, Lidewij; Hiemstra, Sjoukje; Ruuls, Robin; Gröne, Andrea; Roest, Hendrik-Ido-Jan; Willemsen, Peter; van der Giessen, Joke

    2014-01-01

    The presence of Brucella (B.) spp. in harbour porpoises stranded between 2008 and 2011 along the Dutch coast was studied. A selection of 265 tissue samples from 112 animals was analysed using conventional and molecular methods. In total, 4.5% (5/112) of the animals corresponding with 2.3% (6/265)

  2. In vitro bactericidal activity of aminoglycosides, including the next-generation drug plazomicin, against Brucella spp.

    Science.gov (United States)

    Plazomicin is a next-generation aminoglycoside with a potentially improved safety profile compared to other aminoglycosides. This study assessed plazomicin MICs and MBCs in four Brucella spp. reference strains. Like other aminoglycosides and aminocyclitols, plazomicin MBC values equaled MIC values ...

  3. Immuogenicity and safety of a natural rough mutant of Brucella suis as a vaccine for swine

    Science.gov (United States)

    The objective of the current study was to evaluate the safety, immunogenicity and clearance of the natural rough mutant of Brucella suis strain 353-1 (353-1) as a vaccine in domestic swine. In three studies encompassing 155 animals, pigs were inoculated with 353-1 by conjunctival (5 x 10**7 CFU), p...

  4. Seroprevalence of Brucella antibodies in harbor seals in Alaska, USA, with age, regional, and reproductive comparisons.

    Science.gov (United States)

    Hoover-Miller, A; Dunn, J L; Field, C L; Blundell, G; Atkinson, S

    2017-09-20

    Populations of harbor seal Phoca vitulina in the Gulf of Alaska have dramatically declined during the past 4 decades. Numbers of seals in Glacier Bay, in southeast Alaska, USA, have also declined despite extensive protection. Causes of the declines and slow recovery are poorly understood. Brucellosis is a zoonotic disease that adversely affects reproduction in many domestic species. We measured the seroprevalence of Brucella antibodies in 554 harbor seals in 3 Alaska locations: Prince William Sound (PWS), Glacier Bay (GB), and Tracy Arm Fords Terror (TAFT) Wilderness Area. Objectives included testing for regional, sex, age, and female reproductive state differences in Brucella antibody seroprevalence, persistence in titers in recaptured seals, and differences in titers between mother seals and their pups. Overall, 52% of adults (AD), 53% of subadults (SA), 77% of yearlings (YRL), and 26% of Brucella. Results show higher seroprevalence (64%) for AD and SA seals in the depressed and declining populations in PWS and GB than in TAFT (29%). Lactating females were less likely to be seropositive than other AD females, including pregnant females. Further research is needed to seek evidence of Brucella infection in Alaskan harbor seals, identify effects on neonatal viability, and assess zoonotic implications for Alaska Natives who rely on harbor seals for food.

  5. Brucella β 1,2 cyclic glucan is an activator of human and mouse dendritic cells.

    Directory of Open Access Journals (Sweden)

    Anna Martirosyan

    Full Text Available Bacterial cyclic glucans are glucose polymers that concentrate within the periplasm of alpha-proteobacteria. These molecules are necessary to maintain the homeostasis of the cell envelope by contributing to the osmolarity of Gram negative bacteria. Here, we demonstrate that Brucella β 1,2 cyclic glucans are potent activators of human and mouse dendritic cells. Dendritic cells activation by Brucella β 1,2 cyclic glucans requires TLR4, MyD88 and TRIF, but not CD14. The Brucella cyclic glucans showed neither toxicity nor immunogenicity compared to LPS and triggered antigen-specific CD8(+ T cell responses in vivo. These cyclic glucans also enhanced antigen-specific CD4(+ and CD8(+ T cell responses including cross-presentation by different human DC subsets. Brucella β 1,2 cyclic glucans increased the memory CD4(+ T cell responses of blood mononuclear cells exposed to recombinant fusion proteins composed of anti-CD40 antibody and antigens from both hepatitis C virus and Mycobacterium tuberculosis. Thus cyclic glucans represent a new class of adjuvants, which might contribute to the development of effective antimicrobial therapies.

  6. Clinical and Diagnostic Aspects of Brucellosis and Antimicrobial Susceptibility of Brucella Isolates in Hamedan, Iran.

    Science.gov (United States)

    Torkaman Asadi, Fatemeh; Hashemi, Seyyed Hamid; Alikhani, Mohammad Yousef; Moghimbeigi, Abbas; Naseri, Zahra

    2017-05-24

    Current drug regimens for brucellosis are associated with relatively high rates of therapeutic failure or relapse. Reduced antimicrobial susceptibility of Brucella spp. has been proposed recently as a potential cause of therapeutic failure. The aim of this study was to evaluate the antibiotic resistance pattern of Brucella melitensis clinical isolates by E-test method in Hamadan, west of Iran. In a 15-month period, all patients with suspected brucellosis were enrolled. Blood specimens were collected for diagnosis of brucellosis by BACTEC system and serological tests. Antimicrobial susceptibility of clinical isolates to 7 antibiotics was assessed by the E-test method. One hundred forty-nine patients with brucellosis were evaluated. 38.3% of cultures of clinical samples were positive for BACTEC system, of which 91.2% were associated with a positive serological test result. No significant associations were found between serology and the culture method. All Brucella isolates were susceptible to doxycycline, streptomycin, gentamicin, ciprofloxacin, and moxifloxacin. However, decreased sensitivity to rifampin and trimethoprim-sulfamethoxazole was found in 35.1% and 3.5% of isolates, respectively. Because of the high rates of intermediate sensitivity to rifampin among Brucella isolates, this drug should be prescribed with caution. We recommend restricting the use of rifampin for treatment of brucellosis except as an alternative drug for special situations.

  7. Molecular detection of Brucella melitensis in sheep and goat milk in ...

    African Journals Online (AJOL)

    Bolle X, Letesson JJ. A quorum sensing regulator controls expression of both the type IV secretion system and the flagellar apparatus of Brucella melitensis. Cell. Microbiol. 2005; 7: 1151-1161. 9. Comerci DJ, Martinez-Lorenzo MJ, Sieira R, Gorvel JP,. Ugalde RA. Essential role of the VirB machinery in the maturation of the ...

  8. DNA polymorphism in the omp25/omp31 family of Brucella spp.: identification of a 1.7-kb inversion in Brucella cetaceae and of a 15.1-kb genomic island, absent from Brucella ovis, related to the synthesis of smooth lipopolysaccharide.

    Science.gov (United States)

    Vizcaíno, Nieves; Caro-Hernández, Paola; Cloeckaert, Axel; Fernández-Lago, Luis

    2004-07-01

    Five genes homologous to the well-known omp25 and omp31 genes, that code for two major Brucella spp. outer membrane proteins (OMPs), have been detected in the genome of Brucella melitensis 16M and Brucella suis 1330. In this work we have determined the nucleotide sequence of these five genes, named omp31b, omp25b, omp25c, omp25d and omp22, in the six classical Brucella species reference strains and in representative strains of the recently proposed species Brucella cetaceae and Brucella pinnipediae that classify the Brucella strains isolated in the last years from marine mammals. Although these genes are quite conserved in the genus Brucella, several important differences have been found between species (i) omp31b contains a premature stop codon in B. canis and B. ovis truncating the encoded protein; (ii) the 5' end of omp31b is deleted in the three biovars of B. melitensis which probably prevents synthesis of Omp31b in this species; (iii) only B. melitensis, B. suis and B. neotomae would be able to synthesize the Omp25b protein with the characteristics shared by the Omp25/Omp31 group of proteins (characteristic signal sequence and C-terminal phenylalanine); (iv) a DNA inversion of 1747 bp including omp25b was detected in B. cetaceae strains; (v) a DNA deletion of about 15 kb was detected in all the six B. ovis strains tested. This deletion in B. ovis includes, among other genes, omp25b and wboA, a gene that has been shown to be required for the synthesis of the O-polysaccharide chain of the Brucella spp. smooth lipopolysaccharide. Several features of the DNA region absent from B. ovis suggest that this DNA fragment is a genomic island acquired by the Brucella ancestor by horizontal transfer and later deleted from B. ovis. The DNA polymorphism we have found in this work within the genus Brucella might be involved in the differences in pathogenicity and host preference displayed by the Brucella species.

  9. Origins and global context of Brucella abortus in Italy.

    Science.gov (United States)

    Garofolo, Giuliano; Di Giannatale, Elisabetta; Platone, Ilenia; Zilli, Katiuscia; Sacchini, Lorena; Abass, Anna; Ancora, Massimo; Cammà, Cesare; Di Donato, Guido; De Massis, Fabrizio; Calistri, Paolo; Drees, Kevin P; Foster, Jeffrey T

    2017-02-02

    Brucellosis is a common and chronic disease of cattle and other bovids that often causes reproductive disorders. Natural infection in cattle is caused by Brucella abortus and transmission typically occurs during abortions, calving, or nursing. Brucellosis is also a major zoonotic disease due to contamination of dairy products or contact with the tissues of infected animals. Brucellosis has been eradicated from most of the developed world in the last 40 years but persists in many regions-the disease remains prevalent in portions of Africa, the Middle East, Asia, and Central and South America, as well as in the Mediterranean basin. In Italy, B. abortus has persisted in southern regions in both cattle and water buffalo. Previous attempts at analyzing the phylogenetics of B. abortus in Italy have been challenging due to limited genetic variability and unresolved global population genetic structure of this pathogen. We conducted genome-wide phylogenetic analyses on 11 representative strains of B. abortus from Italy, and compared these sequences to a worldwide collection of publically available genomes. Italian isolates belong to three clades that are basal to the main and global B. abortus lineage. Using six SNP-based assays designed to identify substructure within the Italian clades, we surveyed a collection of 261 isolates and found that one clade predominates throughout endemic districts in the country, while the other two clades are more geographically restricted to portions of southern Italy. Although related strains exist worldwide, B. abortus isolates from Italy are substantially different than those found in much of the rest of Europe and North America, and are more closely related to strains from the Middle East and Asia. Our assays targeting genetic substructure within Italy allowed us to identify the major lineages quickly and inexpensively, without having to generate whole genome sequences for a large isolate collection. These findings highlight the

  10. High throughput MLVA-16 typing for Brucella based on the microfluidics technology

    Directory of Open Access Journals (Sweden)

    Di Giannatale Elisabetta

    2011-03-01

    Full Text Available Abstract Background Brucellosis, a zoonosis caused by the genus Brucella, has been eradicated in Northern Europe, Australia, the USA and Canada, but remains endemic in most areas of the world. The strain and biovar typing of Brucella field samples isolated in outbreaks is useful for tracing back source of infection and may be crucial for discriminating naturally occurring outbreaks versus bioterrorist events, being Brucella a potential biological warfare agent. In the last years MLVA-16 has been described for Brucella spp. genotyping. The MLVA band profiles may be resolved by different techniques i.e. the manual agarose gels, the capillary electrophoresis sequencing systems or the microfluidic Lab-on-Chip electrophoresis. In this paper we described a high throughput system of MLVA-16 typing for Brucella spp. by using of the microfluidics technology. Results The Caliper LabChip 90 equipment was evaluated for MLVA-16 typing of sixty-three Brucella samples. Furthermore, in order to validate the system, DNA samples previously resolved by sequencing system and Agilent technology, were de novo genotyped. The comparison of the MLVA typing data obtained by the Caliper equipment and those previously obtained by the other analysis methods showed a good correlation. However the outputs were not accurate as the Caliper DNA fragment sizes showed discrepancies compared with real data and a conversion table from observed to expected data was created. Conclusion In this paper we described the MLVA-16 using a rapid, sophisticated microfluidics technology for detection of amplification product sizes. The comparison of the MLVA typing data produced by Caliper LabChip 90 system with the data obtained by different techniques showed a general concordance of the results. Furthermore this platform represents a significant improvement in terms of handling, data acquiring, computational efficiency and rapidity, allowing to perform the strain genotyping in a time equal to

  11. Mutant Brucella abortus membrane fusogenic protein induces protection against challenge infection in mice.

    Science.gov (United States)

    de Souza Filho, Job Alves; de Paulo Martins, Vicente; Campos, Priscila Carneiro; Alves-Silva, Juliana; Santos, Nathalia V; de Oliveira, Fernanda Souza; Menezes, Gustavo B; Azevedo, Vasco; Cravero, Silvio Lorenzo; Oliveira, Sergio Costa

    2015-04-01

    Brucella species can cause brucellosis, a zoonotic disease that causes serious livestock economic losses and represents a public health threat. The mechanism of virulence of Brucella spp. is not yet fully understood. Therefore, it is crucial to identify new molecules that serve as virulence factors to better understand this host-pathogen interplay. Here, we evaluated the role of the Brucella membrane fusogenic protein (Mfp) and outer membrane protein 19 (Omp19) in bacterial pathogenesis. In this study, we showed that B. abortus Δmfp::kan and Δomp19::kan deletion mutant strains have reduced persistence in vivo in C57BL/6 and interferon regulatory factor 1 (IRF-1) knockout (KO) mice. Additionally, 24 h after macrophage infection with a Δmfp::kan or Δomp19::kan strain expressing green fluorescent protein (GFP) approximately 80% or 65% of Brucella-containing vacuoles (BCVs) retained the late endosomal/lysosomal marker LAMP-1, respectively, whereas around 60% of BCVs containing wild-type S2308 were found in LAMP-1-negative compartments. B. abortus Δomp19::kan was attenuated in vivo but had a residual virulence in C57BL/6 and IRF-1 KO mice, whereas the Δmfp::kan strain had a lower virulence in these same mouse models. Furthermore, Δmfp::kan and Δomp19::kan strains were used as live vaccines. Challenge experiments revealed that in C57BL/6 and IRF-1 KO mice, the Δmfp::kan strain induced greater protection than the vaccine RB51 and protection similar that of vaccine S19. However, a Δomp19::kan strain induced protection similar to that of RB51. Thus, these results demonstrate that Brucella Mfp and Omp19 are critical for full bacterial virulence and that the Δmfp::kan mutant may serve as a potential vaccine candidate in future studies. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  12. Serological survey of Brucella canis in dogs in urban Harare and selected rural communities in Zimbabwe

    Directory of Open Access Journals (Sweden)

    Simbarashe Chinyoka

    2014-02-01

    Full Text Available A cross-sectional study was conducted in order to detect antibodies for Brucella canis (B. canis in dogs from urban Harare and five selected rural communities in Zimbabwe. Sera from randomly selected dogs were tested for antibodies to B. canis using an enzyme-linked immunosorbent assay. Overall, 17.6% of sera samples tested (57/324, 95% CI: 13.5–21.7 were positive for B. canis antibodies. For rural dogs, seroprevalence varied from 11.7% – 37.9%. Rural dogs recorded a higher seroprevalence (20.7%, 95% CI: 15.0–26.4 compared with Harare urban dogs (12.7%, 95% CI: 6.9–18.5 but the difference was not significant (p = 0.07. Female dogs from both sectors had a higher seroprevalence compared with males, but the differences were not significant (p > 0.05. Five and two of the positive rural dogs had titres of 1:800 and 1:1600, respectively, whilst none of the positive urban dogs had a titre above 1:400. This study showed that brucellosis was present and could be considered a risk to dogs from the studied areas. Further studies are recommended in order to give insight into the epidemiology of brucellosis in dogs and its possible zoonotic consequences in Zimbabwe. Screening for other Brucella spp. (Brucella abortus, Brucella melitensis and Brucella suis other than B. canis is also recommended.

  13. Brucella Genetic Variability in Wildlife Marine Mammals Populations Relates to Host Preference and Ocean Distribution.

    Science.gov (United States)

    Suárez-Esquivel, Marcela; Baker, Kate S; Ruiz-Villalobos, Nazareth; Hernández-Mora, Gabriela; Barquero-Calvo, Elías; González-Barrientos, Rocío; Castillo-Zeledón, Amanda; Jiménez-Rojas, César; Chacón-Díaz, Carlos; Cloeckaert, Axel; Chaves-Olarte, Esteban; Thomson, Nicholas R; Moreno, Edgardo; Guzmán-Verri, Caterina

    2017-07-01

    Intracellular bacterial pathogens probably arose when their ancestor adapted from a free-living environment to an intracellular one, leading to clonal bacteria with smaller genomes and less sources of genetic plasticity. Still, this plasticity is needed to respond to the challenges posed by the host. Members of the Brucella genus are facultative-extracellular intracellular bacteria responsible for causing brucellosis in a variety of mammals. The various species keep different host preferences, virulence, and zoonotic potential despite having 97-99% similarity at genome level. Here, we describe elements of genetic variation in Brucella ceti isolated from wildlife dolphins inhabiting the Pacific Ocean and the Mediterranean Sea. Comparison with isolates obtained from marine mammals from the Atlantic Ocean and the broader Brucella genus showed distinctive traits according to oceanic distribution and preferred host. Marine mammal isolates display genetic variability, represented by an important number of IS711 elements as well as specific IS711 and SNPs genomic distribution clustering patterns. Extensive pseudogenization was found among isolates from marine mammals as compared with terrestrial ones, causing degradation in pathways related to energy, transport of metabolites, and regulation/transcription. Brucella ceti isolates infecting particularly dolphin hosts, showed further degradation of metabolite transport pathways as well as pathways related to cell wall/membrane/envelope biogenesis and motility. Thus, gene loss through pseudogenization is a source of genetic variation in Brucella, which in turn, relates to adaptation to different hosts. This is relevant to understand the natural history of bacterial diseases, their zoonotic potential, and the impact of human interventions such as domestication. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  14. Immunological characteristics of outer membrane protein omp31 of goat Brucella and its monoclonal antibody.

    Science.gov (United States)

    Zheng, W Y; Wang, Y; Zhang, Z C; Yan, F

    2015-10-05

    We examined the immunological characteristics of outer membrane protein omp31 of goat Brucella and its monoclonal antibody. Genomic DNA from the M5 strain of goat Brucella was amplified by polymerase chain reaction and cloned into the prokaryotic expression vector pGEX-4T-1. The expression and immunological characteristics of the fusion protein GST-omp31 were subjected to preliminary western blot detection with goat Brucella rabbit immune serum. The Brucella immunized BALB/c mouse serum was detected using purified protein. The high-potency mouse splenocytes and myeloma Sp2/0 cells were fused. Positive clones were screened by enzyme-linked immunosorbent assay to establish a hybridoma cell line. Mice were inoculated intraperitoneally with hybridoma cells to prepare ascites. The mAb was purified using the n-caprylic acid-ammonium sulfate method. The characteristics of mAb were examined using western blotting and enzyme-linked immunosorbent assay. A 680-base pair band was observed after polymerase chain reaction. Enzyme digestion identification and sequencing showed that the pGEX-4T-1-omp31 prokaryotic expression vector was successfully established; a target band of approximately 57 kDa with an apparent molecular weight consistent with the size of the target fusion protein. At 25°C, the expression of soluble expression increased significantly; the fusion protein GST-omp31 was detected by western blotting. Anti-omp31 protein mAb was obtained from 2 strains of Brucella. The antibody showed strong specificity and sensitivity and did not cross-react with Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Mycobacterium tuberculosis, or Bacillus pyocyaneus. The pGEX-4T-1-omp31 prokaryotic expression vector was successfully established and showed good immunogenicity. The antibody also showed strong specificity and good sensitivity.

  15. Promotion and Rescue of Intracellular Brucella neotomae Replication during Coinfection with Legionella pneumophila.

    Science.gov (United States)

    Kang, Yoon-Suk; Kirby, James E

    2017-05-01

    We established a new Brucella neotomae in vitro model system for study of type IV secretion system-dependent (T4SS) pathogenesis in the Brucella genus. Importantly, B. neotomae is a rodent pathogen, and unlike B. abortus , B. melitensis , and B. suis , B. neotomae has not been observed to infect humans. It therefore can be handled more facilely using biosafety level 2 practices. More particularly, using a series of novel fluorescent protein and lux operon reporter systems to differentially label pathogens and track intracellular replication, we confirmed T4SS-dependent intracellular growth of B. neotomae in macrophage cell lines. Furthermore, B. neotomae exhibited early endosomal (LAMP-1) and late endoplasmic reticulum (calreticulin)-associated phagosome maturation. These findings recapitulate prior observations for human-pathogenic Brucella spp. In addition, during coinfection experiments with Legionella pneumophila , we found that defective intracellular replication of a B. neotomae T4SS virB4 mutant was rescued and baseline levels of intracellular replication of wild-type B. neotomae were significantly stimulated by coinfection with wild-type but not T4SS mutant L. pneumophila Using confocal microscopy, it was determined that intracellular colocalization of B. neotomae and L. pneumophila was required for rescue and that colocalization came at a cost to L. pneumophila fitness. These findings were not completely expected based on known temporal and qualitative differences in the intracellular life cycles of these two pathogens. Taken together, we have developed a new system for studying in vitro Brucella pathogenesis and found a remarkable T4SS-dependent interplay between Brucella and Legionella during macrophage coinfection. Copyright © 2017 American Society for Microbiology.

  16. Genotyping of Indian antigenic, vaccine, and field Brucella spp. using multilocus sequence typing.

    Science.gov (United States)

    Shome, Rajeswari; Krithiga, Natesan; Shankaranarayana, Padmashree B; Jegadesan, Sankarasubramanian; Udayakumar S, Vishnu; Shome, Bibek Ranjan; Saikia, Girin Kumar; Sharma, Narendra Kumar; Chauhan, Harshad; Chandel, Bharat Singh; Jeyaprakash, Rajendhran; Rahman, Habibur

    2016-03-31

    Brucellosis is one of the most important zoonotic diseases that affects multiple livestock species and causes great economic losses. The highly conserved genomes of Brucella, with > 90% homology among species, makes it important to study the genetic diversity circulating in the country. A total of 26 Brucella spp. (4 reference strains and 22 field isolates) and 1 B. melitensis draft genome sequence from India (B. melitensis Bm IND1) were included for sequence typing. The field isolates were identified by biochemical tests and confirmed by both conventional and quantitative polymerase chain reaction (qPCR) targeting bcsp 31Brucella genus-specific marker. Brucella speciation and biotyping was done by Bruce ladder, probe qPCR, and AMOS PCRs, respectively, and genotyping was done by multilocus sequence typing (MLST). The MLST typing of 27 Brucella spp. revealed five distinct sequence types (STs); the B. abortus S99 reference strain and 21 B. abortus field isolates belonged to ST1. On the other hand, the vaccine strain B. abortus S19 was genotyped as ST5. Similarly, B. melitensis 16M reference strain and one B. melitensis field isolate were grouped into ST7. Another B. melitensis field isolate belonged to ST8 (draft genome sequence from India), and only B. suis 1330 reference strain was found to be ST14. The sequences revealed genetic similarity of the Indian strains to the global reference and field strains. The study highlights the usefulness of MLST for typing of field isolates and validation of reference strains used for diagnosis and vaccination against brucellosis.

  17. Evaluation and selection of tandem repeat loci for a Brucella MLVA typing assay

    Directory of Open Access Journals (Sweden)

    Denoeud France

    2006-02-01

    Full Text Available Abstract Background The classification of Brucella into species and biovars relies on phenotypic characteristics and sometimes raises difficulties in the interpretation of the results due to an absence of standardization of the typing reagents. In addition, the resolution of this biotyping is moderate and requires the manipulation of the living agent. More efficient DNA-based methods are needed, and this work explores the suitability of multiple locus variable number tandem repeats analysis (MLVA for both typing and species identification. Results Eighty tandem repeat loci predicted to be polymorphic by genome sequence analysis of three available Brucella genome sequences were tested for polymorphism by genotyping 21 Brucella strains (18 reference strains representing the six 'classical' species and all biovars as well as 3 marine mammal strains currently recognized as members of two new species. The MLVA data efficiently cluster the strains as expected according to their species and biovar. For practical use, a subset of 15 loci preserving this clustering was selected and applied to the typing of 236 isolates. Using this MLVA-15 assay, the clusters generated correspond to the classical biotyping scheme of Brucella spp. The 15 markers have been divided into two groups, one comprising 8 user-friendly minisatellite markers with a good species identification capability (panel 1 and another complementary group of 7 microsatellite markers with higher discriminatory power (panel 2. Conclusion The MLVA-15 assay can be applied to large collections of Brucella strains with automated or manual procedures, and can be proposed as a complement, or even a substitute, of classical biotyping methods. This is facilitated by the fact that MLVA is based on non-infectious material (DNA whereas the biotyping procedure itself requires the manipulation of the living agent. The data produced can be queried on a dedicated MLVA web service site.

  18. The Antibacterial Activity of Selected Labiatae (Lamiaceae) Essential Oils against Brucella melitensis.

    Science.gov (United States)

    Al-Mariri, Ayman; Safi, Mazen

    2013-03-01

    Brucellosis, a zoonosis caused by four species of brucella, has a high morbidity. The major cause of brucellosis worldwide is brucella melitensis. Medicinal plants are considered as new antibacterial sources that could replace conventional antibiotics in the treatment of antibiotic-resistant bacteria. The aim of this study was to evaluate the efficacy of some native plants, alone and in combination with some antibiotics, in the treatment of brucellosis. The present experimental in vitro study was carried out to evaluate the anti-brucella activities of essential oils of Rosmarinus officinalis L., Origanum syriacum, Thymus syriacus, Salvia palaestina Benth, Mentha piperia, and Lavandula stoechas L., alone and in combination with some antibiotics. The activity against 16 tetracycline-resistant B. melitensis isolates was determined by disc diffusion method incorporating a concentration of 5%. Antibiotic discs were also used as a control. Microdilution brucella broth susceptibility assay was used in order to determine the MICs of essential oils and five antibiotics. Among all the herbs evaluated, only the essential oils of O. syriacum and T. syriacus plants demonstrated most effective anti-brucella activity, and were then chosen for MIC study. The minimal inhibitory concentrations (MIC50) of essential oils of O. syriacum and T. syriacus against tetracycline-resistant B. melitensis were 3.125 µl/ml and 6.25 µl/ml, respectively. Among the essential oils studied, those of O. syriacum and T. syriacus were most effective. Since a combination of levofloxacin and Thymus syriacus essential oil increased the efficacy of this antibiotic, O. syriacum and T. syriacus are recommended to be used as bactericidal agents against B. melitensis.

  19. The Antibacterial Activity of Selected Labiatae (Lamiaceae Essential Oils against Brucella melitensis

    Directory of Open Access Journals (Sweden)

    Ayman Al-Mariri

    2013-03-01

    Full Text Available Background: Brucellosis, a zoonosis caused by four species of brucella, has a high morbidity. The major cause of brucellosis worldwide is brucella melitensis. Medicinal plants are considered as new antibacterial sources that could replace conventional antibiotics in the treatment of antibiotic-resistant bacteria. The aim of this study was to evaluate the efficacy of some native plants, alone and in combination with some antibiotics, in the treatment of brucellosis.Methods: The present experimental in vitro study was carried out to evaluate the anti-brucella activities of essential oils of Rosmarinus officinalis L., Origanum syriacum, Thymus syriacus, Salvia palaestina Benth, Mentha piperia, and Lavandula stoechas L., alone and in combination with some antibiotics. The activity against 16 tetracycline-resistant B. melitensis isolates was determined by disc diffusion method incorporating a concentration of 5%. Antibiotic discs were also used as a control. Microdilution brucella broth susceptibility assay was used in order to determine the MICs of essential oils and five antibiotics.Results: Among all the herbs evaluated, only the essential oils of O. syriacum and T. syriacus plants demonstrated most effective anti-brucella activity, and were then chosen for MIC study. The minimal inhibitory concentrations (MIC50 of essential oils of O. syriacum and T. syriacus against tetracycline-resistant B. melitensis were 3.125 µl/ml and 6.25 µl/ml, respectively. Conclusion: Among the essential oils studied, those of O. syriacum and T. syriacus were most effective. Since a combination of levofloxacin and Thymus syriacus essential oil increased the efficacy of this antibiotic, O. syriacum and T. syriacus are recommended to be used as bactericidal agents against B. melitensis.

  20. Molecular Epidemiological and Antibiotic Susceptibility Characterization of Brucella Isolates from Humans in Sicily, Italy▿

    Science.gov (United States)

    Marianelli, Cinzia; Graziani, Caterina; Santangelo, Carmela; Xibilia, Maria Teresa; Imbriani, Alida; Amato, Rosa; Neri, Domenico; Cuccia, Mario; Rinnone, Sebastiano; Di Marco, Vincenzo; Ciuchini, Franco

    2007-01-01

    Brucellosis is a serious problem in Sicily. Brucella melitensis was identified as the species most frequently isolated in humans in Italy. No data, however, are available about the molecular epidemiological characterization of Brucella isolates from humans. We have conducted this study to molecularly characterize clinical isolates of Brucella spp. and to evaluate their antimicrobial susceptibilities. Twenty Brucella isolates were studied. Differential growth characteristics and DNA polymorphisms such as the restriction patterns of the PCR-amplified omp2a and omp2b genes, rpoB nucleotide sequencing, and multiple-locus variable-number tandem repeat analysis of 16 loci (MLVA-16) were used to characterize the strains. In vitro antibiotic susceptibility was determined by the E-test method on two different agar media, and the results were compared. All isolates were identified as B. melitensis biovar 3. rpoB nucleotide sequence analysis allowed the identification of two different genotypes of B. melitensis biovar 3. On the other hand, the MLVA-16 typing assay recognized 17 distinct genotypes. All isolates were sensitive to all tested antibiotics (rifampin, doxycycline, ciprofloxacin, ceftriaxone, and trimethoprim-sulfamethoxazole), and the Mueller-Hinton agar plate is recommended for antibiotic susceptibility testing by the E-test method. Our findings identify B. melitensis biovar 3 as the etiological agent isolated in Sicily and encourage the use of both molecular methods, and in particular of the MLVA-16 assay, in epidemiological trace-back analysis. This study represents the first epidemiological data from molecular typing of Brucella strains circulating in Italy and, in particular, in eastern Sicily. PMID:17634297

  1. Cell-mediated immune responses differentiate infections with Brucella suis from Yersinia enterocolitica serotype O : 9 in pigs

    DEFF Research Database (Denmark)

    Riber, Ulla; Jungersen, Gregers

    2007-01-01

    Due to almost identical lipopolysaccharide (LPS) O-antigens, infections with Yersinia enterocolitica serotype 0:9 (YeO:9) cause false positive serological reactions (FPSR) in tests for Brucella and thus cause problems in National Brucella surveillance programs. As LPS are strong inducers...... of antibody responses it was hypothesized that cell-mediated immune responses to non-LPS antigens of the two bacteria can be used to separate immune responses to these two biologically very different infections. Following subclinical experimental infections with Brucella suis biovar 2, high interferon......-gamma (IFN-gamma) assay responses with a commercial Brucella melitensis antigen preparation (Brucellergene OCB) preceded the development of antibodies. High IFN-gamma responses in the seven B. suis inoculated pigs with serological evidence of infection were consistent throughout a 20-week postinoculation...

  2. Structural, functional and immunogenic insights on Cu,Zn Superoxide Dismutase pathogenic virulence factors from Neisseria meningitidis and Brucella abortus

    Science.gov (United States)

    Bacterial pathogens Neisseria meningitidis and Brucella abortus pose threats to human and animal health worldwide, causing meningococcal disease and brucellosis, respectively. Mortality from acute N. meningitidis infections remains high despite antibiotics, and brucellosis presents alimentary and he...

  3. Efficacy of Brucella abortus vaccine strain RB51 compared to the reference vaccine Brucella abortus strain 19 in water buffalo

    Directory of Open Access Journals (Sweden)

    Vincenzo Caporale

    2010-03-01

    Full Text Available Approximately 250 000 water buffalo (Bubalus bubalis live in the Campania region of southern Italy where the breeding of this species is very popular. Of these animals, almost 150 000 are concentrated in the Caserta province where the prevalence of Brucella abortus in this species represents approximately 20% at herd level. The Italian brucellosis eradication programme provides a slaughter and vaccination strategy for this province. B. abortus strain RB51 (RB51 has become the official vaccine for the prevention of brucellosis in cattle in several countries. The aim of this study was to evaluate the efficacy of RB51 in water buffalo compared to the B. abortus S19 vaccine (S19. The study was performed in accordance with a protocol described in mice. Female buffalo aged five months were inoculated. Five received a RB51 dosage on two occasions that was three times greater than that approved for use in cattle and a booster after one month, five received B. abortus S19 vaccine at the standard dosage and three controls received a phosphate buffer solution. Buffalo were then challenged with a virulent B. abortus strain 544 thirty days post vaccination. Antibodies that developed in the five animals vaccinated with RB51 were not detected by the Rose Bengal test or complement fixation test (CFT and were also tested by CFT prepared with RB51 antigen. After culling, B. abortus was cultured from the spleen, retropharyngeal and supra-mammary lymph nodes. A statistical evaluation was performed to assess the immunogenicity values obtained in buffalo vaccinated with S19, compared to those obtained in buffalo vaccinated with the RB51 vaccine and in the unvaccinated control group.

  4. Efficacy of Brucella abortus vaccine strain RB51 compared to the reference vaccine Brucella abortus strain 19 in water buffalo.

    Science.gov (United States)

    Caporale, Vincenzo; Bonfini, Barbara; Di Giannatale, Elisabetta; Di Provvido, Andrea; Forcella, Simona; Giovannini, Armando; Tittarelli, Manuela; Scacchia, Massimo

    2010-01-01

    Approximately 250,000 water buffalo (Bubalus bubalis) live in the Campania region of southern Italy where the breeding of this species is very popular. Of these animals, almost 150,000 are concentrated in the Caserta province where the prevalence of Brucella abortus in this species represents approximately 20% at herd level. The Italian brucellosis eradication programme provides a slaughter and vaccination strategy for this province. B. abortus strain RB51 (RB51) has become the official vaccine for the prevention of brucellosis in cattle in several countries. The aim of this study was to evaluate the efficacy of RB51 in water buffalo compared to the B. abortus S19 vaccine (S19). The study was performed in accordance with a protocol described in mice. Female buffalo aged five months were inoculated. Five received a RB51 dosage on two occasions that was three times greater than that approved for use in cattle and a booster after one month, five received B. abortus S19 vaccine at the standard dosage and three controls received a phosphate buffer solution. Buffalo were then challenged with a virulent B. abortus strain 544 thirty days post vaccination. Antibodies that developed in the five animals vaccinated with RB51 were not detected by the Rose Bengal test or complement fixation test (CFT) and were also tested by CFT prepared with RB51 antigen. After culling, B. abortus was cultured from the spleen, retropharyngeal and supra-mammary lymph nodes. A statistical evaluation was performed to assess the immunogenicity values obtained in buffalo vaccinated with S19, compared to those obtained in buffalo vaccinated with the RB51 vaccine and in the unvaccinated control group.

  5. Rapid identification of Brucella isolates to the species level by real time PCR based single nucleotide polymorphism (SNP analysis

    Directory of Open Access Journals (Sweden)

    Smith Catherine J

    2008-06-01

    Full Text Available Abstract Background Brucellosis, caused by members of the genus Brucella, remains one of the world's major zoonotic diseases. Six species have classically been recognised within the family Brucella largely based on a combination of classical microbiology and host specificity, although more recently additional isolations of novel Brucella have been reported from various marine mammals and voles. Classical identification to species level is based on a biotyping approach that is lengthy, requires extensive and hazardous culturing and can be difficult to interpret. Here we describe a simple and rapid approach to identification of Brucella isolates to the species level based on real-time PCR analysis of species-specific single nucleotide polymorphisms (SNPs that were identified following a robust and extensive phylogenetic analysis of the genus. Results Seven pairs of short sequence Minor Groove Binding (MGB probes were designed corresponding to SNPs shown to possess an allele specific for each of the six classical Brucella spp and the marine mammal Brucella. Assays were optimised to identical reaction parameters in order to give a multiple outcome assay that can differentiate all the classical species and Brucella isolated from marine mammals. The scope of the assay was confirmed by testing of over 300 isolates of Brucella, all of which typed as predicted when compared to other phenotypic and genotypic approaches. The assay is sensitive being capable of detecting and differentiating down to 15 genome equivalents. We further describe the design and testing of assays based on three additional SNPs located within the 16S rRNA gene that ensure positive discrimination of Brucella from close phylogenetic relatives on the same platform. Conclusion The multiple-outcome assay described represents a new tool for the rapid, simple and unambiguous characterisation of Brucella to the species level. Furthermore, being based on a robust phylogenetic framework, the

  6. Serologic evidence of Brucella infection in pinnipeds along the coast of Hokkaido, the northernmost main island of Japan.

    Science.gov (United States)

    Abe, Erika; Ohishi, Kazue; Ishinazaka, Tsuyoshi; Fujii, Kei; Maruyama, Tadashi

    2017-04-01

    Brucella infection in Hokkaido was serologically surveyed in four species of pinnipeds inhabiting Cape Erimo during 2008-2013 and the Shiretoko Peninsula in 1999 by ELISA using Brucella abortus and B. canis as antigens. Anti-Brucella positive sera showed higher absorbance to B. abortus than B. canis in almost all samples. Anti-B. abortus antibodies were detected in serum samples from 24% (n = 55) of Western Pacific harbor seals (Phoca vitulina stejnegeri) in Cape Erimo and from 66% (n = 41) of spotted seals (P. largha), 15% (n = 20) of ribbon seals (Histriophoca fasciata) and 18% (n = 17) of Western Steller's sea lions (Eumetopias jubatus jubatus) in the Shiretoko Peninsula. Anti-Brucella antibodies were detected at higher absorbance in 1- to 4-year-old harbor seals than in the pups and mature animals, suggesting either that Brucella infection mainly occurs after weaning or that it is maternally transmitted to pups with premature or suppressed immunity. Anti-Brucella antibodies were detected in both immature and mature spotted seals and ribbon seals, with higher absorbance in the former. The antibodies were detected only in mature Western Steller's sea lions. Western blot analysis of the serum samples showed some differences in band appearances, namely discrete versus smeary, and in the number of bands, indicating that multiple different Brucella may be prevalent in pinnipeds in Hokkaido. Alternatively, the Brucella of pinnipeds may have some intra-species diversity. © 2017 The Societies and John Wiley & Sons Australia, Ltd.

  7. Development of a bead-based Luminex assay using lipopolysaccharide specific monoclonal antibodies to detect biological threats from Brucella species.

    Science.gov (United States)

    Silbereisen, Angelika; Tamborrini, Marco; Wittwer, Matthias; Schürch, Nadia; Pluschke, Gerd

    2015-10-05

    Brucella, a Gram-negative bacterium, is classified as a potential bioterrorism agent mainly due to the low dose needed to cause infection and the ability to transmit the bacteria via aerosols. Goats/sheep, cattle, pigs, dogs, sheep and rodents are infected by B. melitensis, B. abortus, B. suis, B. canis, B. ovis and B. neotomae, respectively, the six classical Brucella species. Most human cases are caused by B. melitensis and B. abortus. Our aim was to specifically detect Brucellae with 'smooth' lipopolysaccharide (LPS) using a highly sensitive monoclonal antibody (mAb) based immunological assay. To complement molecular detection systems for potential bioterror agents, as required by international biodefense regulations, sets of mAbs were generated by B cell hybridoma technology and used to develop immunological assays. The combination of mAbs most suitable for an antigen capture assay format was identified and an immunoassay using the Luminex xMAP technology was developed. MAbs specific for the LPS O-antigen of Brucella spp. were generated by immunising mice with inactivated B. melitensis or B. abortus cells. Most mAbs recognised both B. melitensis and B. abortus and antigen binding was not impeded by inactivation of the bacterial cells by γ irradiation, formalin or heat treatment, a step required to analyse the samples immunologically under biosafety level two conditions. The Luminex assay recognised all tested Brucella species with 'smooth' LPS with detection limits of 2×10(2) to 8×10(4) cells per mL, depending on the species tested. Milk samples spiked with Brucella spp. cells were identified successfully using the Luminex assay. In addition, the bead-based immunoassay was integrated into a multiplex format, allowing for simultaneous, rapid and specific detection of Brucella spp., Bacillus anthracis, Francisella tularensis and Yersinia pestis within a single sample. Overall, the robust Luminex assay should allow detection of Brucella spp. in both natural

  8. Real-time PCR assays for detection of Brucella spp. and the identification of genotype ST27 in bottlenose dolphins (Tursiops truncatus).

    Science.gov (United States)

    Wu, Qingzhong; McFee, Wayne E; Goldstein, Tracey; Tiller, Rebekah V; Schwacke, Lori

    2014-05-01

    Rapid detection of Brucella spp. in marine mammals is challenging. Microbiologic culture is used for definitive diagnosis of brucellosis, but is time consuming, has low sensitivity and can be hazardous to laboratory personnel. Serological methods can aid in diagnosis, but may not differentiate prior exposure versus current active infection and may cross-react with unrelated Gram-negative bacteria. This study reports a real-time PCR assay for the detection of Brucella spp. and application to screen clinical samples from bottlenose dolphins stranded along the coast of South Carolina, USA. The assay was found to be 100% sensitive for the Brucella strains tested, and the limit of detection was 0.27fg of genomic DNA from Brucella ceti B1/94 per PCR volume. No amplification was detected for the non-Brucella pathogens tested. Brucella DNA was detected in 31% (55/178) of clinical samples tested. These studies indicate that the real-time PCR assay is highly sensitive and specific for the detection of Brucella spp. in bottlenose dolphins. We also developed a second real-time PCR assay for rapid identification of Brucella ST27, a genotype that is associated with human zoonotic infection. Positive results were obtained for Brucella strains which had been identified as ST27 by multilocus sequence typing. No amplification was found for other Brucella strains included in this study. ST27 was identified in 33% (18/54) of Brucella spp. DNA-positive clinical samples. To our knowledge, this is the first report on the use of a real-time PCR assay for identification of Brucella genotype ST27 in marine mammals. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Major Outer Membrane Protein Omp25 of Brucella suis Is Involved in Inhibition of Tumor Necrosis Factor Alpha Production during Infection of Human Macrophages

    OpenAIRE

    Jubier-Maurin, Véronique; Boigegrain, Rose-Anne; Cloeckaert, Axel; Gross, Antoine; Alvarez-Martinez, Maria-Teresa; Terraza, Annie; Liautard, Janny; Köhler, Stephan; Rouot, Bruno; Dornand, Jacques; Liautard, Jean Pierre

    2001-01-01

    Brucella spp. can establish themselves and cause disease in humans and animals. The mechanisms by which Brucella spp. evade the antibacterial defenses of their host, however, remain largely unknown. We have previously reported that live brucellae failed to induce tumor necrosis factor alpha (TNF-α) production upon human macrophage infection. This inhibition is associated with a nonidentified protein that is released into culture medium. Outer membrane proteins (OMPs) of gram-negative bacteria...

  10. Brucella melitensis prosthetic joint infection in a traveller returning to the UK from Thailand: Case report and review of the literature

    OpenAIRE

    Lewis, Joseph M; Folb, Jonathan; Kalra, Sanjay; Squire, S. Bertel; Taegtmeyer, Miriam; Beeching, Nick J.

    2016-01-01

    BACKGROUND\\ud Brucella spp. prosthetic joint infections are infrequently reported in the literature, particularly in returning travellers, and optimal treatment is unknown.\\ud \\ud METHOD\\ud We describe a prosthetic joint infection (PJI) caused by Brucella melitensis in a traveller returning to the UK from Thailand, which we believe to be the first detailed report of brucellosis in a traveller returning from this area. The 23 patients with Brucella-related PJI reported in the literature are su...

  11. Brucella invasion of human intestinal epithelial cells elicits a weak proinflammatory response but a significant CCL20 secretion.

    Science.gov (United States)

    Ferrero, Mariana C; Fossati, Carlos A; Rumbo, Martín; Baldi, Pablo C

    2012-10-01

    In spite of the frequent acquisition of Brucella infection by the oral route in humans, the interaction of the bacterium with cells of the intestinal mucosa has been poorly studied. Here, we show that different Brucella species can invade human colonic epithelial cell lines (Caco-2 and HT-29), in which only smooth species can replicate efficiently. Infection with smooth strains did not produce a significant cytotoxicity, while the rough strain RB51 was more cytotoxic. Infection of Caco-2 cells or HT-29 cells with either smooth or rough strains of Brucella did not result in an increased secretion of TNF-α, IL-1β, MCP-1, IL-10 or TGF-β as compared with uninfected controls, whereas all the infections induced the secretion of IL-8 and CCL20 by both cell types. The MCP-1 response to flagellin from Salmonella typhimurium was similar in Brucella-infected or uninfected cells, ruling out a bacterial inhibitory mechanism as a reason for the weak proinflammatory response. Infection did not modify ICAM-1 expression levels in Caco-2 cells, but increased them in HT-29 cells. These results suggest that Brucella induces only a weak proinflammatory response in gut epithelial cells, but produces a significant CCL20 secretion. The latter may be important for bacterial dissemination given the known ability of Brucella to survive in dendritic cells. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  12. The presence of Brucella ceti ST26 in a striped dolphin (Stenella coeruleoalba) with meningoencephalitis from the Mediterranean Sea.

    Science.gov (United States)

    Alba, Patricia; Terracciano, Giuliana; Franco, Alessia; Lorenzetti, Serena; Cocumelli, Cristiano; Fichi, Gianluca; Eleni, Claudia; Zygmunt, Michel S; Cloeckaert, Axel; Battisti, Antonio

    2013-05-31

    Brucella spp. was isolated from brain, lung and intestinal lymph nodes of a dead adult male striped dolphin (Stenella coeruleoalba) found stranded on the Tyrrhenian coast (Tuscany, Italy) of the Mediterranean Sea in February 2012. Brucella spp. was associated with moderate to severe lesions of meningoencephalitis. A co-infection by Toxoplasma gondii was also demonstrated at brain level by means of molecular and histopathologic methods. The Brucella isolate was further characterized based on a fragment-specific polymerase chain reaction (PCR) approach, consisting of a set of five specific PCRs, targeting specific chromosomal IS711 locations for marine mammal Brucellae, as described previously. The isolate was thus classified as Brucella ceti I; V fragment-positive (or B. ceti dolphin type), according to previous studies. Multi Locus Sequence Analysis demonstrated that the isolate belongs to Sequence Type 26, while omp2 (omp2a and omp2b genes) sequence analysis further confirmed the isolate belonged to this group of strains. This is the first report of Brucella spp. from marine mammals in the Mediterranean Sea, and represents a further observation that this strain group is associated with hosts of the Family Delphinidae, and particularly with the striped dolphins, also in the Mediterranean area, thus constituting a further biological hazard of concern for this vulnerable subpopulation. Copyright © 2013 Elsevier B.V. All rights reserved.

  13. Estandarización de una prueba de PCR para la detección de Brucella sp.

    Directory of Open Access Journals (Sweden)

    Carlos Padilla R

    2003-04-01

    Full Text Available Objetivo: Estandarizar una prueba de PCR para la detección de Brucella spp. Materiales y métodos: Se usó oligonucleótidos reportados que amplifican la secuencia de 16S rRNA de Brucella spp. Fueron evaluados dos métodos de extracción de ADN: fenol-cloroformo-alcohol isoamílico y un kit comercial basado en columnas con afinidad. Para determinar la sensibilidad de la prueba se usó 8 cepas peruanas de Brucella y para determinar la especificidad de la prueba se usó otras cepas bacterianas peruanas de E. coli, Shigella, Proteus mirabilis, Salmonella aratyphi, Salmonella typhi, Citrobacter freundii y Vibrio cholerae. Resultados: Los 2 métodos de extracción de ADN evaluados fueron efectivos. La sensibilidad analítica de la prueba es alta, lográndose detectar 80 femtogramos de ADN de Brucella spp. purificado. Todas las cepas peruanas de Brucella spp. fueron detectadas por la prueba. Además, la prueba es negativa para cepas peruanas de otras especies bacterianas. Conclusión: Se ha estandarizado las condiciones de una prueba de PCR para la detección de cepas peruanas de Brucella spp., la cual es muy sensible y específica en el laboratorio.

  14. Comparison of brucella and non-specific epididymorchitis: gray scale and color Doppler ultrasonographic features

    Energy Technology Data Exchange (ETDEWEB)

    Ozturk, Adil [Department of Radiology, Harran University School of Medicine, Arastirma ve Uygulama Hastanesi, TR-63100 Sanliurfa (Turkey)]. E-mail: ozturka26@hotmail.com; Ozturk, Ebru [Department of Radiology, Harran University School of Medicine, Arastirma ve Uygulama Hastanesi, TR-63100 Sanliurfa (Turkey); Zeyrek, Fadile [Department of Microbiology, Harran University School of Medicine, Sanliurfa (Turkey); Onur, Kahraman [Department of Urology, SSK Sanliurfa Hastanesi, Sanliurfa (Turkey); Sirmatel, Ocal [Department of Radiology, Harran University School of Medicine, Arastirma ve Uygulama Hastanesi, TR-63100 Sanliurfa (Turkey); Kat, Nurcan [Department of Radiology, Harran University School of Medicine, Arastirma ve Uygulama Hastanesi, TR-63100 Sanliurfa (Turkey)

    2005-11-01

    Objective: The aim of this study is to find out if it is possible to differentiate between brucellar and non-specific epididymorchitis by comparing ultrasonography (US) and color Doppler ultrasonography (CDUS) findings. Material and methods: Fifty-six patients diagnosed to have epididymorchitis both clinically and ultrasonographically were included to study. All of the patients were investigated serologically for brucella. Twenty-eight of those patients were admitted brucella epididymorchitis because of high agglutinations titers for brucella. The other 28 patients were admitted non-specific epididymorchitis because of normal agglutinations titers for brucella. Testicular size, echogenicity, hydrocele, internal echoes and/or septations within hydrocele, and scrotal skin thickness of normal and involved testis were compared by ultrasonography. Besides, pick systolic velocity, end diastolic velocity, resistive index and pick systolic velocity ratio values were measured by bilateral testicular color Doppler ultrasonography in both groups. When the p-value is <0.05, the difference between groups is accepted as statistically significant. Results: Thickening of scrotal skin was seen in 17 of 28 patients with brucella epididymorchitis (BEPO) (67%) and in 25 of 28 patients with non-specific epididymorchitis (NEPO) (89.2%) (p < 0.01). There was no difference between groups regarding presence of hydrocele. However hydrocele seen in all patients was anechoic except for two patients (8.6%). Hydrocele seen in 18 of 22 patients with BEPO and hydrocele had internal echogenicity or septation (p < 0.001). Sizes of testes and epididymis were found to be increased in involved testis compared to normal testis. Testes of all patients with NEPO were homogenous with decreased echogenicity except for five patients (17.8%). However, 23 patients with BEPO (82%) found to have heterogenous testis (p < 0.001). Spectral measurements showed increased PSV and EDV values and decreased RI values in

  15. Brucella BioR Regulator Defines a Complex Regulatory Mechanism for Bacterial Biotin Metabolism

    Science.gov (United States)

    Xu, Jie; Zhang, Huimin; Srinivas, Swaminath

    2013-01-01

    The enzyme cofactor biotin (vitamin H or B7) is an energetically expensive molecule whose de novo biosynthesis requires 20 ATP equivalents. It seems quite likely that diverse mechanisms have evolved to tightly regulate its biosynthesis. Unlike the model regulator BirA, a bifunctional biotin protein ligase with the capability of repressing the biotin biosynthetic pathway, BioR has been recently reported by us as an alternative machinery and a new type of GntR family transcriptional factor that can repress the expression of the bioBFDAZ operon in the plant pathogen Agrobacterium tumefaciens. However, quite unusually, a closely related human pathogen, Brucella melitensis, has four putative BioR-binding sites (both bioR and bioY possess one site in the promoter region, whereas the bioBFDAZ [bio] operon contains two tandem BioR boxes). This raised the question of whether BioR mediates the complex regulatory network of biotin metabolism. Here, we report that this is the case. The B. melitensis BioR ortholog was overexpressed and purified to homogeneity, and its solution structure was found to be dimeric. Functional complementation in a bioR isogenic mutant of A. tumefaciens elucidated that Brucella BioR is a functional repressor. Electrophoretic mobility shift assays demonstrated that the four predicted BioR sites of Brucella plus the BioR site of A. tumefaciens can all interact with the Brucella BioR protein. In a reporter strain that we developed on the basis of a double mutant of A. tumefaciens (the ΔbioR ΔbioBFDA mutant), the β-galactosidase (β-Gal) activity of three plasmid-borne transcriptional fusions (bioBbme-lacZ, bioYbme-lacZ, and bioRbme-lacZ) was dramatically decreased upon overexpression of Brucella bioR. Real-time quantitative PCR analyses showed that the expression of bioBFDA and bioY is significantly elevated upon removal of bioR from B. melitensis. Together, we conclude that Brucella BioR is not only a negative autoregulator but also a repressor of

  16. Improved influenza viral vector based Brucella abortus vaccine induces robust B and T-cell responses and protection against Brucella melitensis infection in pregnant sheep and goats.

    Science.gov (United States)

    Mailybayeva, Aigerim; Yespembetov, Bolat; Ryskeldinova, Sholpan; Zinina, Nadezhda; Sansyzbay, Abylai; Renukaradhya, Gourapura J; Petrovsky, Nikolai; Tabynov, Kaissar

    2017-01-01

    We previously developed a potent candidate vaccine against bovine brucellosis caused by Brucella abortus using the influenza viral vector expressing Brucella Omp16 and L7/L12 proteins (Flu-BA). Our success in the Flu-BA vaccine trial in cattle and results of a pilot study in non-pregnant small ruminants prompted us in the current study to test its efficacy against B. melitensis infection in pregnant sheep and goats. In this study, we improved the Flu-BA vaccine formulation and immunization method to achieve maximum efficacy and safety. The Flu-BA vaccine formulation had two additional proteins Omp19 and SOD, and administered thrice with 20% Montanide Gel01 adjuvant, simultaneously by both subcutaneous and conjunctival routes at 21 days intervals in pregnant sheep and goats. At 42 days post-vaccination (DPV) we detected antigen-specific IgG antibodies predominantly of IgG2a isotype but also IgG1, and also detected a strong lymphocyte recall response with IFN-γ production. Importantly, our candidate vaccine prevented abortion in 66.7% and 77.8% of pregnant sheep and goats, respectively. Furthermore, complete protection (absence of live B. melitensis 16M) was observed in 55.6% and 66.7% of challenged sheep and goats, and 72.7% and 90.0% of their fetuses (lambs/yeanlings), respectively. The severity of B. melitensis 16M infection in vaccinated sheep and goats and their fetuses (index of infection and rates of Brucella colonization in tissues) was significantly lower than in control groups. None of the protection parameters after vaccination with Flu-BA vaccine were statistically inferior to protection seen with the commercial B. melitensis Rev.1 vaccine (protection against abortion and vaccination efficacy, alpha = 0.18-0.34, infection index, P = 0.37-0.77, Brucella colonization, P = 0.16 to P > 0.99). In conclusion, our improved Flu-BA vaccine formulation and delivery method were found safe and effective in protecting pregnant sheep and goats against adverse

  17. Descripción de características reproductivas en tres perros seropositivos a Brucella canis Description of reproductive characteristics of three Brucella canis seropositive dogs

    OpenAIRE

    C. BORIE; R. CEPEDA; M. VILLARROEL; M. DE LOS REYES

    2002-01-01

    Se describen las características reproductivas a nivel histológico y seminal en tres perros seropositivos a Brucella canis. A nivel seminal se observaron alteraciones en volumen y en la morfología espermática, encontrándose en un perro ausencia total de espermatozoides. Esta situación concordó con los estudios histológicos, donde se encontró alteración de la línea espermatogénica, además de infiltración eritrocitaria tubular indicativo de alteración de la barrera hematotesticular. Los resulta...

  18. Brucella abortus: inmunidad, vacunas y estrategias de prevención basadas en ácidos nucleicos Brucella abortus: immunity, vaccines and prevention strategies based on nucleic acids

    OpenAIRE

    R Rivers; E Andrews; A González-Smith; G Donoso; A Oñate

    2006-01-01

    Brucelosis, enfermedad causada por la bacteria intracelular facultativa Brucella abortus, es una zoonosis ampliamente distribuida a nivel mundial que afecta principalmente al ganado bovino, causando esterilidad en machos y abortos en hembras en gestación. La resistencia depende del desarrollo de una inmunidad mediada por células, con la participación de células T CD4+ de tipo Th1, que secreten interferón gama (-INF), citosina que estimula la actividad bactericida por macrófagos y la actividad...

  19. SEROLOGICAL SURVEY FOR BRUCELLA OVIS DISSEMINATION AMONG GOATS (Capra aegagrus hircus

    Directory of Open Access Journals (Sweden)

    Atanas ARNAUDOV

    2012-03-01

    Full Text Available By complement fixation test 230 blood samples from goats and he goats were examined for presence of antibodies against Brucella ovis. 134 blood samples were from goats (23 of them were from slipping goats and 96- from he goats. The animals come from private farms in the Plovdiv and Pazardzhik regions (Southern Bulgaria. 10.87% of all tested blood samples contain antibodies against Brucella ovis. Differences in the percentage of the positive reagents of different goat categories were found. The highest percentage was among slipping goats (39.13% towards to 11.71% among the healthy goats and 3.13% among he goats. It can be concluded that goats play an important role in the epidemiology of the disease. The greatest risk is slipping goats bred together with sheep flocks.

  20. Seroprevalence of Brucella abortus and B. canis in household dogs in southwestern Nigeria: a preliminary report

    Directory of Open Access Journals (Sweden)

    S. I. B. Cadmus

    2011-04-01

    Full Text Available A preliminary serological study of 366 household dogs in Lagos and Ibadan, southwestern Nigeria, was carried out to determine antibodies due to exposure to Brucella abortus and B. canis, using the rose bengal test (RBT and the rapid slide agglutination (RSA test, respectively. Results showed that 5.46 % (20/366 and 0.27 % (1/366 of the dogs screened were seropositive to B. abortus and B. canis, respectively.Of all dogs, 36 had a history of being fed foetuses from cows and 11 (30.6 % of these tested positive in the RBT. Our findings, although based on a limited sample size and a dearth of clinical details, revealed that dogs in Nigeria may be infected with Brucella spp. given the wide range of risk factors. Further studies are recommended to elucidate the epidemiology of brucellosis in dogs and its possible zoonotic consequences in the country.

  1. Immune responses of bison and efficacy after booster vaccination with Brucella abortus strain RB51.

    Science.gov (United States)

    Olsen, S C; McGill, J L; Sacco, R E; Hennager, S G

    2015-04-01

    Thirty-one bison heifers were randomly assigned to receive saline or a single vaccination with 10(10) CFU of Brucella abortus strain RB51. Some vaccinated bison were randomly selected for booster vaccination with RB51 at 11 months after the initial vaccination. Mean antibody responses to RB51 were greater (P RB51-vaccinated bison at 8, 16, and 24 weeks after the initial vaccination and at 8 weeks after the booster vaccination. The vaccinated bison had greater (P Brucella organisms in all tissues, except in retropharyngeal and supramammary lymph nodes. Our study suggests that RB51 booster vaccination is an effective vaccination strategy for enhancing herd immunity against brucellosis in bison. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  2. Evaluation of Brucella abortus strain RB51 and strain 19 in pronghorn antelope

    Science.gov (United States)

    Elzer, P.H.; Smith, J.; Roffe, T.; Kreeger, T.; Edwards, J.; Davis, D.

    2002-01-01

    Free-roaming elk and bison in the Greater Yellowstone Area remain the only wildlife reservoirs for Brucella abortus in the United States, and the large number of animals and a lack of holding facilities make it unreasonable to individually vaccinate each animal. Therefore, oral delivery is being proposed as a possible option to vaccinate these wild ungulates. One of the main problems associated with oral vaccination is the potential exposure of nontarget species to the vaccines. The purpose of this study was to determine the effects of two Brucella vaccines, strain 19 (S19) and the rough strain RB51 (SRB51), in pregnant pronghorn antelope. We conclude that S19 and SRB51 rarely colonize maternal and fetal tissues of pregnant pronghorn and were not associated with fetal death. Oral delivery of either vaccine at this dose appears to be nonhazardous to pregnant pronghorn.

  3. Genetic Characterization and Comparative Genome Analysis of Brucella melitensis Isolates from India

    Directory of Open Access Journals (Sweden)

    Sarwar Azam

    2016-01-01

    Full Text Available Brucellosis is the most frequent zoonotic disease worldwide, with over 500,000 new human infections every year. Brucella melitensis, the most virulent species in humans, primarily affects goats and the zoonotic transmission occurs by ingestion of unpasteurized milk products or through direct contact with fetal tissues. Brucellosis is endemic in India but no information is available on population structure and genetic diversity of Brucella spp. in India. We performed multilocus sequence typing of four B. melitensis strains isolated from naturally infected goats from India. For more detailed genetic characterization, we carried out whole genome sequencing and comparative genome analysis of one of the B. melitensis isolates, Bm IND1. Genome analysis identified 141 unique SNPs, 78 VNTRs, 51 Indels, and 2 putative prophage integrations in the Bm IND1 genome. Our data may help to develop improved epidemiological typing tools and efficient preventive strategies to control brucellosis.

  4. A review of Brucella sp. infection of sea mammals with particular emphasis on isolates from Scotland.

    Science.gov (United States)

    Foster, G; MacMillan, A P; Godfroid, J; Howie, F; Ross, H M; Cloeckaert, A; Reid, R J; Brew, S; Patterson, I A P

    2002-12-20

    Brucellae recovered from sea mammals were first reported in 1994. In the years since both culture and serological analysis have demonstrated that the infection occurs in a wide range of species of marine mammals inhabiting a vast amount of the world's oceans. Molecular studies have demonstrated that the isolates differ from those found amongst terrestrial animals and also distinguish between strains which have seals and cetaceans as their preferred hosts. At the phenotypic level seal and cetacean strains can also be differed with respect to their CO(2) requirement, primary growth on Farrells medium and metabolic activity on galactose. Two new species B. cetaceae and B. pinnipediae have been proposed as a result. This paper provides a review of Brucella in sea mammals and updates findings from the study of sea mammals from around the coast of Scotland.

  5. Treatment of a subdural empyema complicated by intracerebral abscess due to Brucella infection

    Directory of Open Access Journals (Sweden)

    J. Zhang

    Full Text Available A 55-year-old male presented with fever, stupor, aphasia, and left hemiparesis. A history of head trauma 3 months before was also reported. Cranial magnetic resonance imaging revealed slight contrast enhancement of lesions under the right frontal skull plate and right frontal lobe. Because of deterioration in nutritional status and intracranial hypertension, the patient was prepared for burr hole surgery. A subdural empyema (SDE recurred after simple drainage. After detection of Brucella species in SDE, craniotomy combined with antibiotic treatment was undertaken. The patient received antibiotic therapy for 6 months (two doses of 2 g ceftriaxone, two doses of 100 mg doxycycline, and 700 mg rifapentine for 6 months that resulted in complete cure of the infection. Thus, it was speculated that the preexisting subdural hematoma was formed after head trauma, which was followed by a hematogenous infection caused by Brucella species.

  6. Study of Nitric Oxide production by murine peritoneal macrophages induced by Brucella Lipopolysaccharide

    Directory of Open Access Journals (Sweden)

    Kavoosi G

    2001-07-01

    Full Text Available Brueclla is a gram negative bacteria that causes Brucellosis. Lipopolysaccharide (LPS ", the pathogenic agent of Brucella is composed of O-chain, core oligosaccharide and lipid A. in addition, the structural and biological properties of different LPS extracted from different strains are not identical. The first defense system against LPS is nonspecific immunity that causes macrophage activation. Activated macrophages produce oxygen and nitrogen radicals that enhance the protection against intracellular pathogens.In this experiment LPS was extracted by hot phenol- water procedure and the effect of various LPSs on nitric oxide prodution by peritoneal mouse macrophages was examined.Our results demonstrated that the effect of LPS on nitric oxide production is concentration-dependent we observed the maximum response in concentration of 10-20 microgram per milliliter. Also our results demonstrate that LPS extracted from vaccine Brucella abortus (S 19 had a highe effect on nitric oxide production than the LPS from other strains

  7. Diverse Genetic Regulon of the Virulence-Associated Transcriptional Regulator MucR in Brucella abortus 2308

    Science.gov (United States)

    Caswell, Clayton C.; Elhassanny, Ahmed E. M.; Planchin, Emilie E.; Roux, Christelle M.; Weeks-Gorospe, Jenni N.; Ficht, Thomas A.; Dunman, Paul M.

    2013-01-01

    The Ros-type regulator MucR is one of the few transcriptional regulators that have been linked to virulence in Brucella. Here, we show that a Brucella abortus in-frame mucR deletion strain exhibits a pronounced growth defect during in vitro cultivation and, more importantly, that the mucR mutant is attenuated in cultured macrophages and in mice. The genetic basis for the attenuation of Brucella mucR mutants has not been defined previously, but in the present study the genes regulated by MucR in B. abortus have been elucidated using microarray analysis and real-time reverse transcription-PCR (RT-PCR). In B. abortus 2308, MucR regulates a wide variety of genes whose products may function in establishing and maintaining cell envelope integrity, polysaccharide biosynthesis, iron homeostasis, genome plasticity, and transcriptional regulation. Particularly notable among the MucR-regulated genes identified is arsR6 (nolR), which encodes a transcriptional regulator previously linked to virulence in Brucella melitensis 16 M. Importantly, electrophoretic mobility shift assays (EMSAs) determined that a recombinant MucR protein binds directly to the promoter regions of several genes repressed by MucR (including arsR6 [nolR]), and in Brucella, as in other alphaproteobacteria, MucR binds to its own promoter to repress expression of the gene that encodes it. Overall, these studies have uncovered the diverse genetic regulon of MucR in Brucella, and in doing so this work has begun to define the MucR-controlled genetic circuitry whose misregulation contributes to the virulence defect of Brucella mucR mutants. PMID:23319565

  8. Serologic response in bottlenose dolphins Tursiops truncatus infected with Brucella sp. using a dolphin-specific indirect ELISA.

    Science.gov (United States)

    Meegan, Jenny; Dunn, J Lawrence; Venn-Watson, Stephanie K; Smith, Cynthia R; Sidor, Inga; Jensen, Eric D; Van Bonn, William G; Pugh, Roberta; Ficht, Thomas; Adams, L Garry; Nielsen, Klaus; Romano, Tracy A

    2012-12-03

    Marine-origin Brucella infections and serologic evidence of exposure have been documented in multiple cetacean species. A dolphin-specific indirect enzyme-linked immunosorbent assay (ELISA) was developed to screen bottlenose dolphin sera for anti-Brucella antibodies. A total of 131 serum samples collected over a 2 to 18 yr period from 6 bottlenose dolphins Tursiops truncatus with confirmed Brucella infections were analyzed for the presence and magnitude of antibody titers against marine-origin Brucella to compare individual antibody responses to various disease manifestations. Additionally, an epidemiologic serologic survey of a managed population of 64 bottlenose dolphins was performed to evaluate for the presence of antibodies and to determine whether there were any clinical pathology predictors for exposure or infection. The serologic results revealed that the dolphins with Brucella-associated abortions were seronegative for 7 to 18 yr until after the abortion and maintained positive titers for several years, with 2 of 3 animals returning to seronegative status. In contrast, the dolphins with Brucella-associated pulmonary or bone lesions maintained persistent positive titers for 2 to 18 yr. The population serosurvey revealed no significant differences in antibody levels among males and females, and dolphins between the ages of 17 and 25 yr were 6.8 times more likely to be Brucella antibody positive compared to those that were younger or older. Seropositive dolphins did not have significant inflammation compared to seronegative dolphins but were more likely to have higher levels of aspartate aminotransferase and gamma-glutamyl transpeptidase. Among 16 dolphins that tested seropositive, 13 (81.3%) had previously been seropositive for at least 3 to 5 yr.

  9. Changes of predominant species/biovars and sequence types of Brucella isolates, Inner Mongolia, China.

    Science.gov (United States)

    Chen, Yanfen; Ke, Yuehua; Wang, Yufei; Yuan, Xitong; Zhou, Xiaoyan; Jiang, Hai; Wang, Zhoujia; Zhen, Qing; Yu, Yaqin; Huang, Liuyu; Cui, Buyun; Chen, Zeliang

    2013-11-01

    Human brucellosis incidence in China was divided into 3 stages, high incidence (1950-1960s), decline (1970-1980s) and re-emergence (1990-2000s). Human brucellosis has been reported in all the 32 provinces, of which Inner Mongolia has the highest prevalence, accounting for over 40% of the cases in China. To investigate the etiology alteration of human brucellosis in Inner Mongolia, the species, biovars and genotypes of 60 Brucella isolates from this province were analyzed. Species and biovars of the Brucella strains isolated from outbreaks were determined based on classical identification procedures. Strains were genotyped by multi locus sequence typing (MLST). Sequences of 9 housekeeping genes were obtained and sequence types were defined. The distribution of species, biovars and sequence types (STs) among the three incidence stages were analyzed and compared. The three stages of high incidence, decline and re-emergence were predominated by B. melitensis biovar 2 and 3, B. abortus biovar 3, and B. melitensis biovar 1, respectively, implying changes in the predominant biovars. Genotyping by MLST revealed a total of 14 STs. Nine STs (from ST28 to ST36), accounting for 64.3% of all the STs, were newly defined and different from those observed in other countries. Different STs were distributed among the three stages. ST8 was the most common ST in 1950-1960s and 1990-2000s, while ST2 was the most common in 1970-1980s. The prevalence of biovars and sequence types of Brucella strains from Inner Mongolia has changed over time in the three stages. Compared with those from other countries, new sequence types of Brucella strains exist in China.

  10. Release of outer membrane fragments by exponentially growing Brucella melitensis cells.

    OpenAIRE

    Gamazo, C.; Moriyon, I

    1987-01-01

    Rough and smooth strains of Brucella melitensis released a membranous material that was devoid of detectable NADH oxidase and succinic dehydrogenase activity (cytoplasmic membrane markers) but that contained lipopolysaccharide, proteins, and phospholipids. This material was composed of two fractions that had similar chemical compositions but that were of different sizes which were separated by differential ultracentrifugation. Electron microscopy showed that both fractions are made of unit me...

  11. Brucella melitensis and Mycobacterium tuberculosis depict overlapping gene expression patterns induced in infected THP-1 macrophages.

    Science.gov (United States)

    Masoudian, M; Derakhshandeh, A; Ghahramani Seno, M M

    2015-01-01

    Pathogens infecting mammalian cells have developed various strategies to suppress and evade their hosts' defensive mechanisms. In this line, the intracellular bacteria that are able to survive and propagate within their host cells must have developed strategies to avert their host's killing attitude. Studying the interface of host-pathogen confrontation can provide valuable information for defining therapeutic approaches. Brucellosis, caused by the Brucella strains, is a zoonotic bacterial disease that affects thousands of humans and animals around the world inflicting discomfort and huge economic losses. Similar to many other intracellular dwelling bacteria, infections caused by Brucella are difficult to treat, and hence any attempt at identifying new and common therapeutic targets would prove beneficial for the purpose of curing infections caused by the intracellular bacteria. In THP-1 macrophage infected with Brucella melitensis we studied the expression levels of four host's genes, i.e. EMP2, ST8SIA4, HCP5 and FRMD5 known to be involved in pathogenesis of Mycobacterium tuberculosis. Our data showed that at this molecular level, except for FRMD5 that was downregulated, the other three genes were upregulated by B. melitensis. Brucella melitensis and M. tuberculosis go through similar intracellular processes and interestingly two of the investigated genes, i.e. EMP2 and ST4SIA8 were upregulated in THP-1 cell infected with B. melitensis similar to that reported for THP-1 cells infected with M. tuberculosis. At the host-pathogen interaction interface, this study depicts overlapping changes for different bacteria with common survival strategies; a fact that implies designing therapeutic approaches based on common targets may be possible.

  12. Virulence Effects and Signaling Partners Modulated by Brucella melitensis Light-sensing Histidine Kinase

    Science.gov (United States)

    Gourley, Christopher R.

    The facultative intracellular pathogen Brucella melitensis utilizes diverse virulence factors. A Brucella light sensing histidine kinase can influence in vitro virulence of the bacteria during intracellular infection. First, we demonstrated that the B. melitensis light sensing kinase (BM-LOV-HK) affects virulence in an IRF-1-/- mouse model of infection. Infection with a Δ BM-LOV-HK strain resulted in less bacterial colonization of IRF-1-/- spleens and extended survivorship compared to mice infected with wild type B. melitensis 16M. Second, using PCR arrays, we observed less expression of innate and adaptive immune system activation markers in ΔBM-LOV-HK infected mouse spleens than wild type B. melitensis 16M infected mouse spleens 6 days after infection. Third, we demonstrated by microarray analysis of B. melitensis that deletion of BM-LOV-HK alters bacterial gene expression. Downregulation of genes involved in control of the general stress response system included the alternative sigma factor RpoE1 and its anti-anti sigma factor PhyR. Conversely, genes involved in flagella production, quorum sensing, and the type IV secretion system (VirB operon) were upregulated in the Δ BM-LOV-HK strain compared to the wild type B. melitensis 16M. Analysis of genes differentially regulated in Δ BM-LOV-HK versus the wild type strain indicated an overlap of 110 genes with data from previous quorum sensing regulator studies of Δ vjbR and/ΔblxR(babR) strains. Also, several predicted RpoE1 binding sites located upstream of genes were differentially regulated in the ΔBM-LOV-HK strain. Our results suggest BM-LOV-HK is important for in vivo Brucella virulence, and reveals that BM-LOV-HK directly or indirect regulates members of the Brucella quorum sensing, type IV secretion, and general stress systems.

  13. Entrance and Survival of Brucella pinnipedialis Hooded Seal Strain in Human Macrophages and Epithelial Cells

    Science.gov (United States)

    Briquemont, Benjamin; Sørensen, Karen K.; Godfroid, Jacques

    2013-01-01

    Marine mammal Brucella spp. have been isolated from pinnipeds (B. pinnipedialis) and cetaceans (B. ceti) from around the world. Although the zoonotic potential of marine mammal brucellae is largely unknown, reports of human disease exist. There are few studies of the mechanisms of bacterial intracellular invasion and multiplication involving the marine mammal Brucella spp. We examined the infective capacity of two genetically different B. pinnipedialis strains (reference strain; NTCT 12890 and a hooded seal isolate; B17) by measuring the ability of the bacteria to enter and replicate in cultured phagocytes and epithelial cells. Human macrophage-like cells (THP-1), two murine macrophage cell lines (RAW264.7 and J774A.1), and a human malignant epithelial cell line (HeLa S3) were challenged with bacteria in a gentamicin protection assay. Our results show that B. pinnipedialis is internalized, but is then gradually eliminated during the next 72 – 96 hours. Confocal microscopy revealed that intracellular B. pinnipedialis hooded seal strain colocalized with lysosomal compartments at 1.5 and 24 hours after infection. Intracellular presence of B. pinnipedialis hooded seal strain was verified by transmission electron microscopy. By using a cholesterol-scavenging lipid inhibitor, entrance of B. pinnipedialis hooded seal strain in human macrophages was significantly reduced by 65.8 % (± 17.3), suggesting involvement of lipid-rafts in intracellular entry. Murine macrophages invaded by B. pinnipedialis do not release nitric oxide (NO) and intracellular bacterial presence does not induce cell death. In summary, B. pinnipedialis hooded seal strain can enter human and murine macrophages, as well as human epithelial cells. Intracellular entry of B. pinnipedialis hooded seal strain involves, but seems not to be limited to, lipid-rafts in human macrophages. Brucella pinnipedialis does not multiply or survive for prolonged periods intracellulary. PMID:24376851

  14. Innocuity and immune response to Brucella melitensis Rev.1 vaccine in camels (Camelus dromedarius

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    A. Benkirane

    2014-10-01

    Full Text Available A field trial was conducted in a camel brucellosis-free herd to evaluate antibody response to the Brucella melitensis Rev.1 vaccine in camels and assess shedding of the vaccine strain in milk. Twenty eight camels were divided into four groups according to their age and vaccination route. Groups A (n=3 and B (n=3 consisted of non-pregnant lactating female camels, vaccinated through subcutaneous and conjunctival routes, respectively. Groups C (n=10 consisted of 8-11 months old calves vaccinated through conjunctival route. The rest of the herd (n=12 composed of female and young camels were not vaccinated and were considered as the control group. Each animal from groups A, B and C was given the recommended dose of 2 x 109 colony forming units of Rev.1 vaccine irrespective of age or route of vaccination. Blood samples were collected from all the animals at the time of vaccination and at weekly, bi-weekly and monthly interval until 32 weeks post vaccination and from controls at weeks 8 and 24. The serological tests used were modified Rose Bengal Test, sero-agglutination test, and an indirect Enzyme Linked Immunosorbent Assay. Milk samples were collected from all vaccinated female camels and tested for the presence of Rev.1 vaccine strain. Most vaccinated animals started to show an antibody response at week 2 and remained positive until week 16. By week 20 post-vaccination all animals in the three groups were tested negative for Brucella antibodies. Bacteriological analysis of milk samples did not allow any isolation of Brucella melitensis. All samples were found Brucella negative in PCR analysis. The results of this study indicate that the Rev.1 vaccine induces seroconversion in camels. Rev.1 vaccine strain is not excreted in the milk of camels. These findings are promising as to the safe use of the Rev.1 vaccine in camels.

  15. The Complete Genome of Brucella Suis 019 Provides Insights on Cross-Species Infection

    OpenAIRE

    Yuanzhi Wang; Zhen Wang; Xin Chen; Hui Zhang; Fei Guo; Ke Zhang; Hanping Feng; Wenyi Gu; Changxin Wu; Lei Ma; Tiansen Li; Chuangfu Chen; Shan Gao

    2016-01-01

    Brucella species are the most important zoonotic pathogens worldwide and cause considerable harm to humans and animals. In this study, we presented the complete genome of B. suis 019 isolated from sheep (ovine) with epididymitis. B. suis 019 has a rough phenotype and can infect sheep, rhesus monkeys and possibly humans. The comparative genome analysis demonstrated that B. suis 019 is closest to the vaccine strain B. suis bv. 1 str. S2. Further analysis associated the rsh gene to the pathogeni...

  16. Isolation & characterization of Brucella melitensis isolated from patients suspected for human brucellosis in India.

    Science.gov (United States)

    Barua, Anita; Kumar, Ashu; Thavaselvam, Duraipandian; Mangalgi, Smita; Prakash, Archana; Tiwari, Sapana; Arora, Sonia; Sathyaseelan, Kannusamy

    2016-05-01

    Brucellosis is endemic in the southern part of India. A combination of biochemical, serological and molecular methods is required for identification and biotyping of Brucella. The present study describes the isolation and biochemical, molecular characterization of Brucella melitensis from patients suspected for human brucellosis. The blood samples were collected from febrile patients suspected to have brucellosis. A total of 18 isolates were obtained from 102 blood samples subjected to culture. The characterization of these 18 isolates was done by growth on Brucella specific medium, biochemical reactions, CO2 requirement, H2S production, agglutination with A and M mono-specific antiserum, dye sensitivity to basic fuchsin and thionin. Further, molecular characterization of the isolates was done by amplification of B. melitensis species specific IS 711 repetitive DNA fragment and 16S (rRNA) sequence analysis. PCR-restriction fragment length polymorphism (RFLP) analysis of omp2 locus and IS711 gene was also done for molecular characterization. All 102 suspected samples were subjected to bacteria isolation and of these, 18 isolates could be recovered on blood culture. The biochemical, PCR and PCR-RFLP and 16s rRNA sequencing revealed that all isolates were of B. melitensis and matched exactly with reference strain B. melitensis 16M. The present study showed an overall isolation rate of 17.64 per cent for B. melitensis. There is a need to establish facilities for isolation and characterization of Brucella species for effective clinical management of the disease among patients as well as surveillance and control of infection in domestic animals. Further studies are needed from different geographical areas of the country with different level of endemicity to plan and execute control strategies against human brucellosis.

  17. Sacroiliitis as a sole manifestation of Brucella melitensis infection in a child

    Energy Technology Data Exchange (ETDEWEB)

    Miron, D.; Garty, I.; Tal, I.; Horovitz, Y.; Kedar, A.

    1987-06-01

    A case of a 12-year-old boy with sacroiliitis documented by positive Tc-99m MDP and Ga-67 scans is described. Isolation of brucella melitensis from the blood and bone marrow established the diagnosis. He responded promptly to docycycline therapy. Throughout the course of his disease this boy had neither fever nor other signs of brucellosis, and x-ray was normal.

  18. Comparative Brucella abortus antibody prevalence in cattle under contrasting husbandry practices in Uganda

    Directory of Open Access Journals (Sweden)

    Gerald Nizeyimana

    2013-02-01

    Full Text Available A study was conducted in the Luwero and Nakasongola districts in central Uganda to determine and compare the prevalence and distribution of antibodies against Brucella abortus in cattle under contrasting husbandry practices, using two serological tests. Three hundred and fifteen serum samples were systematically sampled from 29 farms and subsequently tested using the Rose Bengal plate test (RBPT and Indirect Antibody Enzyme Linked Immunosorbent Assay (I-ELISA. The overall prevalence of antibodies against Brucella abortus in the Nakasongola and Luwero districts was 2.4% and 4.7% on RBPT, compared with 1.2% and 3.34 % on I-ELISA. There was no significant difference between the results obtained by RBPT and indirect antibody ELISA (p > 0.05. It was noted that antibodies against Brucella abortus were widely spread over different farms regardless of the cattle grazing system (p > 0.05. Based on the findings, it is feasible to use RBPT as a cheaper screening alternative for brucellosis. A comprehensive national brucellosis study should be undertaken to study the epidemiology and prevalence of brucellosis in Uganda.

  19. A genome-wide SNP-based phylogenetic analysis distinguishes different biovars of Brucella suis.

    Science.gov (United States)

    Sankarasubramanian, Jagadesan; Vishnu, Udayakumar S; Gunasekaran, Paramasamy; Rajendhran, Jeyaprakash

    2016-07-01

    Brucellosis is an important zoonotic disease caused by Brucella spp. Brucella suis is the etiological agent of porcine brucellosis. B. suis is the most genetically diverged species within the genus Brucella. We present the first large-scale B. suis phylogenetic analysis based on an alignment-free k-mer approach of gathering polymorphic sites from whole genome sequences. Genome-wide core-SNP based phylogenetic tree clearly differentiated and discriminated the B. suis biovars and the vaccine strain into different clades. A total of 16,756 SNPs were identified from the genome sequences of 54 B. suis strains. Also, biovar-specific SNPs were identified. The vaccine strain B. suis S2-30 is extensively used in China, which was discriminated from all biovars with the accumulation of the highest number of SNPs. We have also identified the SNPs between B. suis vaccine strain S2-30 and its closest homolog, B. suis biovar 513UK. The highest number of mutations (22) was observed in the phosphomannomutase (pmm) gene essential for the synthesis of O-antigen. Also, mutations were identified in several virulent genes including genes coding for type IV secretion system and the effector proteins, which could be responsible for the attenuated virulence of B. suis S2-30. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Safety of revaccination of pregnant bison with Brucella abortus strain RB51.

    Science.gov (United States)

    Olsen, S C; Holland, S D

    2003-10-01

    From December 1998 through February 1999, a study was conducted in a Brucella-infected bison herd to evaluate the safety of booster vaccination of adult bison (Bison bison) with 6 x 10(9) colony forming units (CFU) of Brucella abortus strain RB51 (SRB51) that had previously been vaccinated as yearlings with 1 x 10(10) CFU of SRB51. Abortions or other adverse effects were not observed after SRB51 booster vaccination. At 10 wk after adult vaccination, pregnant and nonpregnant bison (n = 65) were randomly selected for bacteriologic sampling of targeted maternal tissues during abattoir processing. Fetal tissues were also sampled in pregnant bison. The SR351 recovered from tissue samples of eight of 48 pregnant bison and none of 17 nonpregnant bison. In three of the eight culture-positive bison, SRB51 was recovered from fetal tissues. In three additional bison, one pregnant and two nonpregnant, B. abortus biovar 1 field strain was recovered from internal iliac or supramammary lymphatic tissues. Results of this study suggest the possibility that the SRB51 vaccine can be safely used to booster vaccinate pregnant bison in a Brucella-infected bison herd. Our data also reaffirms the potential for B. abortus field strains to persist in bison until attainment of reproductive age, despite extensive use of vaccination and serologic testing.

  1. Efficacy of vaccination strategies against intranasal challenge with Brucella abortus in BALB/c mice.

    Science.gov (United States)

    Surendran, Naveen; Sriranganathan, Nammalwar; Lawler, Heather; Boyle, Stephen M; Hiltbold, Elizabeth M; Heid, Bettina; Zimmerman, Kurt; Witonsky, Sharon G

    2011-03-24

    Brucellosis is a zoonotic disease affecting 500,000 people worldwide annually. Inhalation of aerosol containing a pathogen is one of the major routes of disease transmission in humans. Currently there are no licensed human vaccines available. Brucella abortus strain RB51 is a USDA approved live attenuated vaccine against cattle brucellosis. In a mouse model, strain RB51 over-expressing superoxide dismutase (SOD) administered intraperitoneally (IP) has been shown to be more protective than strain RB51 against an IP challenge with B. abortus pathogenic strain 2308. However, there is lack of information on the ability of these vaccine strains to protect against intranasal challenge. With the long-term goal of developing a protective vaccine for animals and people against respiratory challenge of Brucella spp., we tested a number of different vaccination strategies against intranasal infection with strain 2308. We employed strains RB51 and RB51SOD to assess the efficacy of route, dose, and prime-boost strategies against strain 2308 challenge. Despite using multiple protocols to enhance mucosal and systemic protection, neither rough RB51 vaccine strains provided respiratory protection against intranasal pathogenic Brucella infection. However, intranasal (IN) administration of B. abortus vaccine strain 19 induced significant (p≤0.05) pulmonary clearance of strain 2308 upon IN challenge infection compared to saline. Further studies are necessary to address host-pathogen interaction in the lung microenvironment and elucidate immune mechanisms to enhance protection against aerosol infection. Published by Elsevier Ltd.

  2. Characterization of Brucella abortus mutant strain Δ22915, a potential vaccine candidate.

    Science.gov (United States)

    Bao, Yanqing; Tian, Mingxing; Li, Peng; Liu, Jiameng; Ding, Chan; Yu, Shengqing

    2017-04-04

    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_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.

  3. Identification of Brucella ovis exclusive genes in field isolates from Argentina.

    Science.gov (United States)

    Alvarez, Lucía Paula; García-Effrón, Guillermo; Robles, Carlos Alejandro

    2016-03-01

    Brucellosis caused by Brucella ovis is one of the most important infectious diseases of sheep. The aim of this study was to determine the presence of genes both inside and outside the specific B. ovis pathogenicity island 1 (BOPI-1) in a large collection of field isolates of B. ovis and other Brucella spp. from Argentina. The BOV_A0500 gene from B. ovis BOPI-1 was identified in all 104 B. ovis isolates studied. The BOPI-1 complete sequence was found to be conserved in 10 B. ovis strains from the collection, for which whole genome sequencing was performed. The BOV_0198 gene, which is outside BOPI-1 and considered exclusive to B. ovis, showed 90-100% identity with genomic regions of B. ovis, B. melitensis, B. abortus, B. canis, B. suis, B. microti, B. ceti and B. pinnipedialis. The results demonstrate that BOPI-1 is the only exclusive genetic region of B. ovis and marine Brucella spp. and that it is highly conserved in B. ovis field isolates from Argentina. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Evidence of Brucella strain ST27 in bottlenose dolphin (Tursiops truncatus) in Europe.

    Science.gov (United States)

    Cvetnić, Željko; Duvnjak, Sanja; Đuras, Martina; Gomerčić, Tomislav; Reil, Irena; Zdelar-Tuk, Maja; Špičić, Silvio

    2016-11-30

    Marine mammal brucellosis has been known for more than 20 years, but recent work suggests it is more widespread than originally thought. Brucella (B.) pinnipedialis has been isolated from pinnipeds, while B. ceti strains have been associated with cetaceans. Here we report a Brucella strain isolated from multiple lymph nodes of one bottlenose dolphin (Tursiops truncatus) during routine examination of dolphin carcasses found in the Croatian part of the northern Adriatic Sea during the summer of 2015. Classical bacteriological biotyping, PCR-based techniques (single, multiplex, PCR-RFLP) and 16S rRNA DNA sequencing were used to identify Brucella spp. Multiple-locus variable number tandem repeat analysis of 16 loci and multilocus sequence typing of 9 loci were used for genotyping and species determination. The combination of bacteriological, molecular and genotyping techniques identified our strain as ST27, previously identified as a human pathogen. This report provides, to our knowledge, the first evidence of ST27 in the Adriatic Sea in particular and in European waters in general. The zoonotic nature of the strain and its presence in the Adriatic, which is inhabited by bottlenose dolphins, suggest that the strain may pose a significant threat to human health. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Experimental infection of chicken embryos with recently described Brucella microti: Pathogenicity and pathological findings.

    Science.gov (United States)

    Wareth, Gamal; Böttcher, Denny; Melzer, Falk; Shehata, Awad Ali; Roesler, Uwe; Neubauer, Heinrich; Schoon, Heinz-Adolf

    2015-08-01

    Brucellae are facultative intracellular pathogens causing disease in a wide range of domestic and wild animals as well as in humans. Brucella (B.) microti is a recently recognized species and was isolated from common voles (Microtus arvalis), red foxes and soil in Austria and the Czech Republic. Its pathogenicity for livestock and its zoonotic potential has not been confirmed yet. In the present study 25 SPF chicken embryos were inoculated at day 11 of age with 1.6×10(3) and 1.6×10(5)B. microti by yolk sac and allantoic sac routes. Re-isolation of B. microti indicated rapid multiplication of bacteria (up to 1.7×10(12)CFU). B. microti provoked marked gross lesions, i.e. hemorrhages and necroses. All inoculated embryos were dead (100% mortality) in between 2nd and 4th day post inoculation. The predominant histopathological lesion was necroses in liver, kidneys, lungs, spleen, gastrointestinal tract, spinal meninges, yolk sac and chorioallantoic membrane. Immunohistochemical examination showed the presence of Brucella antigen in nearly all of these organs, with infection being mainly restricted to non-epithelial cells or tissues. This study provides the first results on the multiplication and pathogenicity of the mouse pathogenic B. microti in chicken embryos. These data suggest that, even though chicken are not mammals, they could provide a useful tool for understanding the pathogenesis of B. microti associated disease. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Brucellosis caused by the wood rat pathogen Brucella neotomae: two case reports.

    Science.gov (United States)

    Villalobos-Vindas, Juan M; Amuy, Ernesto; Barquero-Calvo, Elías; Rojas, Norman; Chacón-Díaz, Carlos; Chaves-Olarte, Esteban; Guzman-Verri, Caterina; Moreno, Edgardo

    2017-12-19

    Brucellosis is a chronic bacterial disease caused by members of the genus Brucella. Among the classical species stands Brucella neotomae, until now, a pathogen limited to wood rats. However, we have identified two brucellosis human cases caused by B. neotomae, demonstrating that this species has zoonotic potential. Within almost 4 years of each other, a 64-year-old Costa Rican white Hispanic man and a 51-year-old Costa Rican white Hispanic man required medical care at public hospitals of Costa Rica. Their hematological and biochemical parameters were within normal limits. No adenopathies or visceral abnormalities were found. Both patients showed intermittent fever, disorientation, and general malaise and a positive Rose Bengal test compatible with Brucella infection. Blood and cerebrospinal fluid cultures rendered Gram-negative coccobacilli identified by genomic analysis as B. neotomae. After antibiotic treatment, the patients recovered with normal mental activities. This is the first report describing in detail the clinical disease caused by B. neotomae in two unrelated patients. In spite of previous claims, this bacterium keeps zoonotic potential. Proposals to generate vaccines by using B. neotomae as an immunogen must be reexamined and countries housing the natural reservoir must consider the zoonotic risk.

  7. Endovascular treatment of Brucella-infected abdominal aortic aneurysm: A case report.

    Science.gov (United States)

    Zhang, Tao; Ji, Donghua; Wang, Feng

    2017-10-01

    In very rare cases, a primary infected abdominal aortic aneurysm (IAAA) is caused by a species of Brucella. In this report, we report such a case that was successfully treated with a novel approach. To the best of our knowledge, this was the first case occurring in China, in which an infection of the abdominal aortic aneurysm was caused by a Brucella species. The clinical findings included high fever, fatigue, and abdominal pain. The diagnosis was confirmed by computed tomography angiography and by bacteriologic isolation from the patient's blood culture. The patient was given endovascular aneurysm repair (EVAR) and Brucella-sensitive antibiotics for 6 weeks. During the 10-month follow-up, the patient's clinical course remained uneventful. Our case study supports the premise that endovascular aneurysm repair is an appropriate alternative strategy to treat an infected abdominal aortic aneurysm. Compared with conventional surgical treatment, EVAR with long-term oral antibiotics is a simpler, less traumatic, and more efficient procedure. However, this needs to be further evaluated through long-term follow-up.

  8. [A case with tricuspid valve brucella endocarditis presenting with acute right heart failure].

    Science.gov (United States)

    Yazıcı, Hüseyin Uğur; Mert, Kadir Uğur; Senol, Utku; Ulus, Taner

    2012-06-01

    Although the presence of brucella endocarditis is encountered rarely, it is the most fetal complication of brucellosis, which is shown to affect the aortic valve primarily and the mitral valve secondarily. Involvement of the tricuspid valve is extremely rare. A 62-year-old female was admitted with complaints of fever, fatigue, difficulty in breathing, and swellings in her legs. A transthoracic echocardiogram was performed since acute right heart failure was considered due to her symptoms. The echocardiogram showed enlarged right heart chambers, serious tricuspid valve insufficiency, and a mass on the tricuspid valve compatible with a vegetation moving in and out of the right ventricle. Although no growths were observed in the blood culture, antibody titration for brucellosis was found to be 1/640 (+) in the serological examination. The patient was diagnosed with brucella endocarditis and placed on doxycycline, rifampicin, and ceftriaxone treatment for eight weeks. At the end of the eight-week treatment, the symptoms of right heart failure receded and the patient recovered from the endocarditis. Tricuspid valve brucella endocarditis should be considered in patients suffering from acute right heart failure accompanied by systemic infection findings since brucellosis is presently endemic in Turkey.

  9. [Assessment of polymerase chain reaction (PCR) to diagnose brucellosis in a Brucella infected herd].

    Science.gov (United States)

    Lavaroni, O; Aguirre, N; Vanzini, V; Lugaresi, C; Torioni de Echaide, S

    2004-01-01

    The diagnosis of bovine brucellosis using PCR in blood and milk samples from two dairy herds were compared to in vitro isolation, complement fixation test (CF), competitive ELISA (C-ELISA) in serum, and indirect ELISA (I-ELISA) in milk. Samples were obtained from 99 cows vaccinated with Brucella abortus strain 19, from a naturally infected herd (A), whose cows were also vaccinated with B. abortus strain RB51 as adults, and 100 from brucellosis free herd (B). In herd A, PCR identified 14 B. abortus infected cows: nine infected with wild type, and five with wild type and RB51, B. abortus S 19 was not identified. B. abortus biotype 1 was isolated from one cow. All cows infected with a wild strain of B. abortus were positive in serologic tests. Brucella was not found in herd B using PCR. Serological test showed 100% sensitivity related to PCR. The specificity for CF, C-ELISA and I-ELISA was 100%, 99% and 95% respectively. PCR could be useful to identify Brucella biotypes and to complement serologic tests.

  10. [Does reactive arthritis caused by Brucella exist? Apropos of 4 cases].

    Science.gov (United States)

    Dubost, J J; Constantin, A; Soubrier, M; Ristori, J M; Cantagrel, A; Bussière, J L

    1997-02-22

    Arthritis observed in patients with Brucella infection is usually considered to result from live micro-organisms invading the synovia. We observed four cases of brucellosis in which the clinical and laboratory findings suggested a different mechanism: reactive arthritis. The diagnosis of brucellosis was made on the basis of serology tests in 3 patients and blood cultures in 1. All 4 patients presented oligoarthritis. The synovial fluid was sterile in 3. Antibiotics were ineffective in reducing joint pain and inflammation whereas local and systemic anti-inflammatory drugs were effective. Three patients also had other manifestations (sausage-shaped toes, talalgia, sacroiliitis) and fulfilled the diagnostic criteria for spondylarthropathy. All patients were positive for antigen HLA-B27. These observations suggest that Brucella should be added to the list of intracellular infectious agents capable of inducing reactive arthritis, despite the lack of all the diagnostic criteria. For some, such as the uretritis or diarrhea observed before joint involvement, it would be difficult to implicate the germ. Brucella serology should be part of the etiology work-up for reactive arthritis in endemic areas.

  11. Brucella abortus choloylglycine hydrolase affects cell envelope composition and host cell internalization.

    Science.gov (United States)

    Marchesini, María Inés; Connolly, Joseph; Delpino, María Victoria; Baldi, Pablo C; Mujer, Cesar V; DelVecchio, Vito G; Comerci, Diego J

    2011-01-01

    Choloylglycine hydrolase (CGH, E.C. 3.5.1.24) is a conjugated bile salt hydrolase that catalyses the hydrolysis of the amide bond in conjugated bile acids. Bile salt hydrolases are expressed by gastrointestinal bacteria, and they presumably decrease the toxicity of host's conjugated bile salts. Brucella species are the causative agents of brucellosis, a disease affecting livestock and humans. CGH confers Brucella the ability to deconjugate and resist the antimicrobial action of bile salts, contributing to the establishment of a successful infection through the oral route in mice. Additionally, cgh-deletion mutant was also attenuated in intraperitoneally inoculated mice, which suggests that CGH may play a role during systemic infection other than hydrolyzing conjugated bile acids. To understand the role CGH plays in B. abortus virulence, we infected phagocytic and epithelial cells with a cgh-deletion mutant (Δcgh) and found that it is defective in the internalization process. This defect along with the increased resistance of Δcgh to the antimicrobial action of polymyxin B, prompted an analysis of the cell envelope of this mutant. Two-dimensional electrophoretic profiles of Δcgh cell envelope-associated proteins showed an altered expression of Omp2b and different members of the Omp25/31 family. These results were confirmed by Western blot analysis with monoclonal antibodies. Altogether, the results indicate that Brucella CGH not only participates in deconjugation of bile salts but also affects overall membrane composition and host cell internalization.

  12. Purification and properties of Cu-Zn superoxide dismutase extracted from Brucella abortus strain 19

    Energy Technology Data Exchange (ETDEWEB)

    Tabatabai, L.B. (ARS-USDA, Ames, IA (United States))

    1991-03-11

    Recent work showed that a recombinant 20 kDa protein from Brucella abortus expressed in E. coli is a Cu-Zn superoxide dismutase (SOD). Western blot and ELISA results indicated that cattle with brucellosis have antibody to SOD. Here the authors report the purification and properties of the native B. abortus Cu-Zn SOD. SOD was extracted from methanol-killed Brucella abortus strain 19 with 0.1 M sodium citrate-1.0 M sodium chloride solution. The extract was dialyzed and protein precipitated by ammonium sulfate at 70-100% saturation was collected. The SOD was purified by HPLC anion exchange chromatography. SOD activity was assayed with a coupled enzyme assay using xanthine oxidase-cytochrome C reduction assay. The authors determined that the Brucella SOD is present in two molecular forms both inhibitable with KCN with Ki's of 0.32 mM and 4.98 mM, respectively. No other form of SOD was identified in the extract. Polyclonal antibody to SOD and polyclonal antibody to SOD synthetic peptide residues 134-143 inhibited SOD activity by 50% and 13%, respectively. Both SOD and the synthetic peptide inhibited binding of anti-SOD antibody to SOD by 60% and 20%, respectively. Based on these results the SOD and its amphipathic peptide will be considered as candidates for the design of synthetic multiple peptide vaccines and diagnostic reagents for bovine brucellosis.

  13. Goat farm management and Brucella serological test among goat keepers and livestock officers, 2011–2012, Nakhon Si Thammarat Province, southern Thailand

    Directory of Open Access Journals (Sweden)

    Thanidtha Te-Chaniyom

    2016-12-01

    Several goat farming management practices in the study area may increase the risk of Brucella infection in animals. Livestock officers in the area have a high risk of being infected with Brucella. Improving goat farm biosecurity practices in needed to reduce the risk of brucellosis in this area.

  14. The efficacy of the skin delayed-type hypersensitivity using a brucellin prepared from a mucoid strain of Brucella abortus to detect brucellosis

    NARCIS (Netherlands)

    Bercovich, Z.; Muskens, J.A.M.

    1998-01-01

    Eight-hundred-and-ninety-six cattle belonging to herds officially designated Brucella-free, and 190 cattle belonging to infected herds were tested with the skin delayed-type hypersensitivity (SDTH) test, using brucellin (273) prepared from a rnucoid strain of Brucella abortus. An increase in

  15. Maintenance of Brucella abortus free herds; a review with emphasis on the epidemiology and the problems in diagnosing brucellosis in areas of low prevalence

    NARCIS (Netherlands)

    Bercovich, Z.

    1998-01-01

    This review covers some epidemiological aspects that allow Brucella to survive, spread, and maintain itself in the environment. Because the success of maintaining Brucella-free herds is determined by the efficiency of the serological tests to detect a single infected animal the limitations of the

  16. Evaluation of a Multiplex PCR Assay (Bruce-ladder) for Molecular Typing of All Brucella Species, Including the Vaccine Strains▿ †

    Science.gov (United States)

    López-Goñi, I.; García-Yoldi, D.; Marín, C. M.; de Miguel, M. J.; Muñoz, P. M.; Blasco, J. M.; Jacques, I.; Grayon, M.; Cloeckaert, A.; Ferreira, A. C.; Cardoso, R.; Corrêa de Sá, M. I.; Walravens, K.; Albert, D.; Garin-Bastuji, B.

    2008-01-01

    An evaluation of a multiplex PCR assay (Bruce-ladder) was performed in seven laboratories using 625 Brucella strains from different animal and geographical origins. This robust test can differentiate in a single step all of the classical Brucella species, including those found in marine mammals and the S19, RB51, and Rev.1 vaccine strains. PMID:18716225

  17. Complete Genome Sequence of a Brucella ceti ST26 Strain Isolated from a Striped Dolphin (Stenella coeruleoalba) on the Coast of Italy.

    Science.gov (United States)

    Ancora, Massimo; Marcacci, Maurilia; Orsini, Massimiliano; Zilli, Katiuscia; Di Giannatale, Elisabetta; Garofolo, Giuliano; Cammà, Cesare

    2014-03-06

    Brucella spp. are important pathogens affecting a wide range of terrestrial and aquatic animals. We report the complete and annotated genome sequence of Brucella ceti ST26 strain TE10759-12, isolated from a striped dolphin (Stenella coeruleoalba) stranded along the Italian shoreline in March of 2012.

  18. Complete Genome Sequence of a Brucella ceti ST26 Strain Isolated from a Striped Dolphin (Stenella coeruleoalba) on the Coast of Italy

    OpenAIRE

    Ancora, Massimo; Marcacci, Maurilia; Orsini, Massimiliano; Zilli, Katiuscia; Di Giannatale, Elisabetta; Garofolo, Giuliano; Cammà, Cesare

    2014-01-01

    Brucella spp. are important pathogens affecting a wide range of terrestrial and aquatic animals. We report the complete and annotated genome sequence of Brucella ceti ST26 strain TE10759-12, isolated from a striped dolphin (Stenella coeruleoalba) stranded along the Italian shoreline in March of 2012.

  19. Meta-analysis of variables affecting mouse protection efficacy of whole organism Brucella vaccines and vaccine candidates.

    Science.gov (United States)

    Todd, Thomas E; Tibi, Omar; Lin, Yu; Sayers, Samantha; Bronner, Denise N; Xiang, Zuoshuang; He, Yongqun

    2013-01-01

    Vaccine protection investigation includes three processes: vaccination, pathogen challenge, and vaccine protection efficacy assessment. Many variables can affect the results of vaccine protection. Brucella, a genus of facultative intracellular bacteria, is the etiologic agent of brucellosis in humans and multiple animal species. Extensive research has been conducted in developing effective live attenuated Brucella vaccines. We hypothesized that some variables play a more important role than others in determining vaccine protective efficacy. Using Brucella vaccines and vaccine candidates as study models, this hypothesis was tested by meta-analysis of Brucella vaccine studies reported in the literature. Nineteen variables related to vaccine-induced protection of mice against infection with virulent brucellae were selected based on modeling investigation of the vaccine protection processes. The variable "vaccine protection efficacy" was set as a dependent variable while the other eighteen were set as independent variables. Discrete or continuous values were collected from papers for each variable of each data set. In total, 401 experimental groups were manually annotated from 74 peer-reviewed publications containing mouse protection data for live attenuated Brucella vaccines or vaccine candidates. Our ANOVA analysis indicated that nine variables contributed significantly (P-value Brucella vaccine protection efficacy: vaccine strain, vaccination host (mouse) strain, vaccination dose, vaccination route, challenge pathogen strain, challenge route, challenge-killing interval, colony forming units (CFUs) in mouse spleen, and CFU reduction compared to control group. The other 10 variables (e.g., mouse age, vaccination-challenge interval, and challenge dose) were not found to be statistically significant (P-value > 0.05). The protection level of RB51 was sacrificed when the values of several variables (e.g., vaccination route, vaccine viability, and challenge pathogen strain

  20. [Determination of in vitro susceptibilities of Brucella spp. strains against 11 different antibacterial gents isolated from blood cultures].

    Science.gov (United States)

    Keşli, Recep; Bilgin, Hüseyin; Yılmaz, Halim

    2017-07-01

    Brucellosis is a worldwide zoonotic disease and still continuous to be a major public health problem. In this study, it was aimed to identify the Brucella strains to the species level isolated from blood cultures, and to determine the rate of antimicrobial susceptibility against eleven antibacterial agents. A total of 106 Brucella spp. strains were included in the study, which were isolated from blood cultures in University of Health Sciences, Konya Training and Research Hospital, Medical Microbiology Laboratory between January 2011 and June 2013. Identification of the isolated strains were mainly based on conventional methods. In vitro antibacterial susceptibilities of azithromycin, ciprofloxacin, doxycycline, gentamicin, levofloxacin, moxifloxacin, rifampicin, streptomycin, tetracycline, tigecycline, and trimethoprim/sulfamethoxazole, were evaluated by using the gradient (E-test, bioMerieux, France) strip method. The bacterial suspensions adjusted to 0.5 McFarland turbidity was inoculated to Mueller Hinton agar plates, supplemented with 5% sheep blood, and E-test strips of selected antibacterial were applied. The plates were incubated in ambient air 48 hours at 37ºC and Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 29213 were used as quality control strains for antimicrobial susceptibility testing. Minimum inhibitors concentration (MIC) values were interpreted according to Clinical and Laboratory Standards Institute (CLSI) guidelines for slow-growing bacteria such as Haemophilus spp. Of the 106 Brucella spp. strains included in to the study, 90 were identified as Brucella melitensis, and 16 were Brucella abortus. MIC90 values of azithromycin, ciprofloxacin, doxycycline, gentamicin, levofloxacin, moxifloxacin, rifampicin, streptomycin, tetracycline, tigecycline, and trimethoprim/sulfamethoxazole were determined as 1 µg/ml, 0.25 µg/ml, 0.19 µg/ml, 0.25 µg/ml, 0.19 µg/ml, 0.75 µg/ml, 0.25 µg/ml, 0.75 µg/ml, 0.38 µg/ml, 0.64 µg/ml, and 0

  1. Meta-analysis of variables affecting mouse protection efficacy of whole organism Brucella vaccines and vaccine candidates

    Science.gov (United States)

    2013-01-01

    Background Vaccine protection investigation includes three processes: vaccination, pathogen challenge, and vaccine protection efficacy assessment. Many variables can affect the results of vaccine protection. Brucella, a genus of facultative intracellular bacteria, is the etiologic agent of brucellosis in humans and multiple animal species. Extensive research has been conducted in developing effective live attenuated Brucella vaccines. We hypothesized that some variables play a more important role than others in determining vaccine protective efficacy. Using Brucella vaccines and vaccine candidates as study models, this hypothesis was tested by meta-analysis of Brucella vaccine studies reported in the literature. Results Nineteen variables related to vaccine-induced protection of mice against infection with virulent brucellae were selected based on modeling investigation of the vaccine protection processes. The variable "vaccine protection efficacy" was set as a dependent variable while the other eighteen were set as independent variables. Discrete or continuous values were collected from papers for each variable of each data set. In total, 401 experimental groups were manually annotated from 74 peer-reviewed publications containing mouse protection data for live attenuated Brucella vaccines or vaccine candidates. Our ANOVA analysis indicated that nine variables contributed significantly (P-value Brucella vaccine protection efficacy: vaccine strain, vaccination host (mouse) strain, vaccination dose, vaccination route, challenge pathogen strain, challenge route, challenge-killing interval, colony forming units (CFUs) in mouse spleen, and CFU reduction compared to control group. The other 10 variables (e.g., mouse age, vaccination-challenge interval, and challenge dose) were not found to be statistically significant (P-value > 0.05). The protection level of RB51 was sacrificed when the values of several variables (e.g., vaccination route, vaccine viability, and

  2. Novel vector vaccine against Brucella abortus based on influenza A viruses expressing Brucella L7/L12 or Omp16 proteins: evaluation of protection in pregnant heifers.

    Science.gov (United States)

    Tabynov, Kaissar; Yespembetov, Bolat; Sansyzbay, Abylai

    2014-10-14

    The present study provides the first information about the protection of a novel influenza viral vector vaccine expressing the Brucella proteins ribosomal L7/L12 or Omp16 containing the adjuvant Montanide Gel01 in pregnant heifers. Immunization of pregnant heifers was conducted via the conjunctival (n=10) or subcutaneous (n=10) route using cross prime and booster vaccination schedules at an interval of 28 days. The vector vaccine was evaluated in comparison with positive control groups vaccinated with Brucella abortus S19 (n=10) or B. abortus RB51 (n=10) and a negative (PBS+Montanide Gel01; n=10) control group. Via both the conjunctival or subcutaneous route, evaluation of protectiveness against abortion, effectiveness of vaccination and index of infection (in heifers and their fetuses or calves) demonstrated the vector vaccine provided good protection against B. abortus 544 infection compared to the negative control group (PBS+Montanide Gel01) and comparable protection to commercial vaccines B. abortus S19 or B. abortus RB51. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Comparison of an indirect ELISA with the Brucella milk ring test for detection of antibodies to Brucella abortus in bulk milk samples.

    Science.gov (United States)

    Vanzini, V R; Aguirre, N P; Valentini, B S; Torioni de Echaide, S; Lugaresi, C I; Marchesino, M D; Nielsen, K

    2001-09-03

    An indirect enzyme-linked immunosorbent assay (ELISA) for the detection of Brucella abortus antibodies in bovine bulk milk samples was evaluated. About 31 individual milk samples from B. abortus infected cows were diluted into bulk milk from a brucellosis free herd. Individual milk samples obtained from 96 negative or positive herds to ELISA or Brucella ring test (BRT), were tested by ELISA. All positive cows were bled and serum samples were tested by the complement-fixation test (CFT) which was considered the definitive test. A herd was considered infected if at least, one cow was positive in the CFT. Four samples were negative in the BRT at the dilution 1:10 but positive in the ELISA. For samples positive in both tests, BRT titers ranged from 1:10 to 1:480 while ELISA titers ranged from 1:10 to 1:3200. Using bulk milk samples, the sensitivity of the ELISA (98.1%) was higher than the BRT (72.2%) but the specificity of BRT (90.5%) was not statistically different (P=1.0) from the ELISA (88.1%). The implications of the results for brucellosis control are discussed.

  4. Relative Quantitative Proteomic Analysis of Brucella abortus Reveals Metabolic Adaptation to Multiple Environmental Stresses

    Directory of Open Access Journals (Sweden)

    Xiaodong Zai

    2017-11-01

    Full Text Available Brucella spp. are facultative intracellular pathogens that cause chronic brucellosis in humans and animals. The virulence of Brucella primarily depends on its successful survival and replication in host cells. During invasion of the host tissue, Brucella is simultaneously subjected to a variety of harsh conditions, including nutrient limitation, low pH, antimicrobial defenses, and extreme levels of reactive oxygen species (ROS via the host immune response. This suggests that Brucella may be able to regulate its metabolic adaptation in response to the distinct stresses encountered during its intracellular infection of the host. An investigation into the differential proteome expression patterns of Brucella grown under the relevant stress conditions may contribute toward a better understanding of its pathogenesis and adaptive response. Here, we utilized a mass spectrometry-based label-free relative quantitative proteomics approach to investigate and compare global proteomic changes in B. abortus in response to eight different stress treatments. The 3 h short-term in vitro single-stress and multi-stress conditions mimicked the in vivo conditions of B. abortus under intracellular infection, with survival rates ranging from 3.17 to 73.17%. The proteomic analysis identified and quantified a total of 2,272 proteins and 74% of the theoretical proteome, thereby providing wide coverage of the B. abortus proteome. By including eight distinct growth conditions and comparing these with a control condition, we identified a total of 1,221 differentially expressed proteins (DEPs that were significantly changed under the stress treatments. Pathway analysis revealed that most of the proteins were involved in oxidative phosphorylation, ABC transporters, two-component systems, biosynthesis of secondary metabolites, the citrate cycle, thiamine metabolism, and nitrogen metabolism; constituting major response mechanisms toward the reconstruction of cellular

  5. Relative Quantitative Proteomic Analysis of Brucella abortus Reveals Metabolic Adaptation to Multiple Environmental Stresses.

    Science.gov (United States)

    Zai, Xiaodong; Yang, Qiaoling; Yin, Ying; Li, Ruihua; Qian, Mengying; Zhao, Taoran; Li, Yaohui; Zhang, Jun; Fu, Ling; Xu, Junjie; Chen, Wei

    2017-01-01

    Brucella spp. are facultative intracellular pathogens that cause chronic brucellosis in humans and animals. The virulence of Brucella primarily depends on its successful survival and replication in host cells. During invasion of the host tissue, Brucella is simultaneously subjected to a variety of harsh conditions, including nutrient limitation, low pH, antimicrobial defenses, and extreme levels of reactive oxygen species (ROS) via the host immune response. This suggests that Brucella may be able to regulate its metabolic adaptation in response to the distinct stresses encountered during its intracellular infection of the host. An investigation into the differential proteome expression patterns of Brucella grown under the relevant stress conditions may contribute toward a better understanding of its pathogenesis and adaptive response. Here, we utilized a mass spectrometry-based label-free relative quantitative proteomics approach to investigate and compare global proteomic changes in B. abortus in response to eight different stress treatments. The 3 h short-term in vitro single-stress and multi-stress conditions mimicked the in vivo conditions of B. abortus under intracellular infection, with survival rates ranging from 3.17 to 73.17%. The proteomic analysis identified and quantified a total of 2,272 proteins and 74% of the theoretical proteome, thereby providing wide coverage of the B. abortus proteome. By including eight distinct growth conditions and comparing these with a control condition, we identified a total of 1,221 differentially expressed proteins (DEPs) that were significantly changed under the stress treatments. Pathway analysis revealed that most of the proteins were involved in oxidative phosphorylation, ABC transporters, two-component systems, biosynthesis of secondary metabolites, the citrate cycle, thiamine metabolism, and nitrogen metabolism; constituting major response mechanisms toward the reconstruction of cellular homeostasis and metabolic

  6. The Prevalence of Brucella Biotypes Isolated From Sterile Body Fluids of Patients With Brucellosis in Kashan, Iran in 2013

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    Erami

    2016-07-01

    Full Text Available Background Brucella species are classified based on their pathogenic and genetic properties and hosts. Considering the significance of identifying different biotypes of Brucella from the epidemiological point of view and lack of such information in the city of Kashan, Iran. Objectives This study was designed to determine the biotypes and strains of Brucella isolated from patients with brucellosis. Methods This was a descriptive study of 206 samples obtained from patients with suspected brucellosis in 2013 in Kashan. BACTEC 9050 culture media was employed to test the samples. Suspected colonies of Brucella were identified through morphology, staining, and biochemical tests. The biotypes were identified by the Razi Research Institute. Lysis tests with the Tbilisi (Tb phage were performed, the need for CO2, SH2 production, sensitivity to basic fuchsin and thionin stains, and the reaction of all the samples to specific antiserum A and M (monospecific were tested. Results Fifty (24.3% of the 206 samples were culture positive. SH3 production was not detected in any of the isolates, and none of the isolated strains required CO2. The results of the sensitivity test to basic fuchsin and thionin staining and specific agglutination and phage lysis (phage typing tests indicated that all the isolated strains were biotype 1 B. melitansis. Conclusions The cause of human brucellosis in Kashan and its suburbs was biotype 1 B. melitensis. The identification of various biotypes of Brucella is important. Similar studies should be performed to detect the presence of new biotypes originating from neighboring countries.

  7. Structural Insights into the HWE Histidine Kinase Family: The Brucella Blue Light-Activated Histidine Kinase Domain.

    Science.gov (United States)

    Rinaldi, Jimena; Arrar, Mehrnoosh; Sycz, Gabriela; Cerutti, María Laura; Berguer, Paula M; Paris, Gastón; Estrín, Darío Ariel; Martí, Marcelo Adrián; Klinke, Sebastián; Goldbaum, Fernando Alberto

    2016-03-27

    In response to light, as part of a two-component system, the Brucella blue light-activated histidine kinase (LOV-HK) increases its autophosphorylation, modulating the virulence of this microorganism. The Brucella histidine kinase (HK) domain belongs to the HWE family, for which there is no structural information. The HWE family is exclusively present in proteobacteria and usually coupled to a wide diversity of light sensor domains. This work reports the crystal structure of the Brucella HK domain, which presents two different dimeric assemblies in the asymmetric unit: one similar to the already described canonical parallel homodimers (C) and the other, an antiparallel non-canonical (NC) dimer, each with distinct relative subdomain orientations and dimerization interfaces. Contrary to these crystallographic structures and unlike other HKs, in solution, the Brucella HK domain is monomeric and still active, showing an astonishing instability of the dimeric interface. Despite this instability, using cross-linking experiments, we show that the C dimer is the functionally relevant species. Mutational analysis demonstrates that the autophosphorylation activity occurs in cis. The different relative subdomain orientations observed for the NC and C states highlight the large conformational flexibility of the HK domain. Through the analysis of these alternative conformations by means of molecular dynamics simulations, we also propose a catalytic mechanism for Brucella LOV-HK. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Evaluation of three formulations of culture media for isolation of Brucella spp. regarding their ability to inhibit the growth of contaminating organisms.

    Science.gov (United States)

    Vicente, Acácia F; Antunes, João M A P; Lara, Gustavo H B; Mioni, Mateus S R; Allendorf, Susan D; Peres, Marina G; Appolinário, Camila M; Listoni, Fernando J P; Ribeiro, Marcio G; Megid, Jane

    2014-01-01

    Three culture media (Brucella agar, Farrell medium, and CITA) were compared for their effectiveness in inhibiting contamination and for isolating Brucella spp. One hundred lymph nodes from pigs (n = 50) and wild boars (n = 50) with lymphadenitis were collected in slaughterhouses in the State of São Paulo and were assessed on these three selective media for Brucella spp. All of the samples were negative for Brucella spp. on the three culture media. On the agar medium, fungal (70 plates) and Gram-positive bacterial (59 plates) contaminants were observed; in the CITA medium, the absence of fungal and Gram-positive bacteria on 15 plates was observed; no bacterial or fungal growth was observed on the Farrell media. The results demonstrated that the CITA and Farrell media inhibited the growth of contaminants better than the Brucella agar.

  9. Detection and characterization of Brucella spp. in bovine milk in small-scale urban and peri-urban farming in Tajikistan.

    Science.gov (United States)

    Lindahl-Rajala, Elisabeth; Hoffman, Tove; Fretin, David; Godfroid, Jacques; Sattorov, Nosirjon; Boqvist, Sofia; Lundkvist, Åke; Magnusson, Ulf

    2017-03-01

    Brucellosis is one of the most common zoonoses globally, and Central Asia remains a Brucella hotspot. The World Health Organization classifies brucellosis as a neglected zoonotic disease that is rarely in the spotlight for research and mainly affects poor, marginalized people. Urban and peri-urban farming is a common practice in many low-income countries, and it increases the incomes of families that are often restrained by limited economic resources. However, there is a concern that the growing number of people and livestock living close together in these areas will increase the transmission of zoonotic pathogens such as Brucella. This study investigates the presence of Brucella DNA in bovine milk in the urban and peri-urban area of Dushanbe, Tajikistan. Brucella DNA was detected in 10.3% of 564 cow milk samples by IS711-based real-time PCR. This finding is concerning because consumption of unpasteurized dairy products is common in the region. Furthermore, Brucella DNA was detected in the milk of all seropositive cows, but 8.3% of the seronegative cows also showed the presence of Brucella DNA. In addition, sequence analysis of the rpoB gene suggests that one cow was infected with B. abortus and another cow was most likely infected with B. melitensis. The discrepancies between the serology and real-time PCR results highlight the need to further investigate whether there is a need for implementing complementary diagnostic strategies to detect false serological negative individuals in Brucella surveillance, control, and eradication programmes. Furthermore, vaccination of cattle with S19 in addition to vaccination of small ruminants with Rev 1 might be needed in order to control Brucella infections in the livestock population but further research focusing on the isolation of Brucella is required to obtain a comprehensive understanding of the Brucella spp. circulating among the livestock in this region.

  10. Detection and characterization of Brucella spp. in bovine milk in small-scale urban and peri-urban farming in Tajikistan.

    Directory of Open Access Journals (Sweden)

    Elisabeth Lindahl-Rajala

    2017-03-01

    Full Text Available Brucellosis is one of the most common zoonoses globally, and Central Asia remains a Brucella hotspot. The World Health Organization classifies brucellosis as a neglected zoonotic disease that is rarely in the spotlight for research and mainly affects poor, marginalized people. Urban and peri-urban farming is a common practice in many low-income countries, and it increases the incomes of families that are often restrained by limited economic resources. However, there is a concern that the growing number of people and livestock living close together in these areas will increase the transmission of zoonotic pathogens such as Brucella. This study investigates the presence of Brucella DNA in bovine milk in the urban and peri-urban area of Dushanbe, Tajikistan. Brucella DNA was detected in 10.3% of 564 cow milk samples by IS711-based real-time PCR. This finding is concerning because consumption of unpasteurized dairy products is common in the region. Furthermore, Brucella DNA was detected in the milk of all seropositive cows, but 8.3% of the seronegative cows also showed the presence of Brucella DNA. In addition, sequence analysis of the rpoB gene suggests that one cow was infected with B. abortus and another cow was most likely infected with B. melitensis. The discrepancies between the serology and real-time PCR results highlight the need to further investigate whether there is a need for implementing complementary diagnostic strategies to detect false serological negative individuals in Brucella surveillance, control, and eradication programmes. Furthermore, vaccination of cattle with S19 in addition to vaccination of small ruminants with Rev 1 might be needed in order to control Brucella infections in the livestock population but further research focusing on the isolation of Brucella is required to obtain a comprehensive understanding of the Brucella spp. circulating among the livestock in this region.

  11. Epidemiological investigation of the first human brucellosis case in Spain due to Brucella suis biovar 1 strain 1330.

    Science.gov (United States)

    Compés Dea, Cecilia; Guimbao Bescós, Joaquín; Alonso Pérez de Ágreda, Juan Pablo; Muñoz Álvaro, Pilar María; Blasco Martínez, José María; Villuendas Usón, María Cruz

    2017-03-01

    No cases of human brucellosis caused by Brucella suis has been reported in Spain. This study involved interviews with the case and his co-workers, inspection of their workplace, checking infection control measures, and typing the Brucella strain isolated in the blood culture. Brucella suis biovar 1 strain 1330 was isolated from a patient who worked in a waste treatment plant. Food borne transmission, contact with animals, and risk jobs were ruled out. An accidental inoculation with a contaminated needle from a research laboratory waste container was identified as the most probable mode of transmission. There should be controls to ensure that waste containers are sealed. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  12. Reporte de primer caso humano de aislamiento y tipificación de Brucella abortus RB 51: first report in Chile Isolation and identification of Brucella abortus RB 51 in human

    OpenAIRE

    M. Villarroel; M. Grell; R. Saenz

    2000-01-01

    En el Laboratorio de Microbiología del Hospital Base de Osorno, se aisló una cepa de Brucella sp, que posteriormente se tipificó en el Laboratorio Regional de Diagnósticos del Servicio Agrícola y Ganadero, SAG Osorno. Esta cepa fue aislada de un hemocultivo realizado a un paciente con sintomatología clínica compatible con Brucelosis. El resultado de la tipificación fue Brucella abortus RB 51, cepa vaccinal que se presumía apatógena para el hombre. Este es el primer reporte a nivel mundial de ...

  13. Genotipos de aislados de campo de Brucella abortus de distintas regiones geográficas de Chile Genotypes of Brucella abortus field isolates from different geographical regions of Chile

    OpenAIRE

    M Mancilla; M Villarroel; ME Saldías; J Soto; AM Zárraga

    2008-01-01

    La brucelosis bovina es una enfermedad zoonótica, endémica de alto impacto económico. La identificación genética de las cepas prevalentes de Brucella abortus, el patógeno, es clave para establecer estrategias epidemiológicas de control de la enfermedad. La secuencia de inserción IS711 ha sido utilizada como un marcador genético para diferenciar entre especies de Brucella, miembros de una misma especie y dentro de un mismo biovar. Hemos analizado los perfiles de IS711-RFLP de 46 aislados de B....

  14. Detección de anticuerpos Anti-brucella en focas de Weddell (Leptonychotes weddellii) de Cabo Shirref, Antártica Detection of anti-brucella antibodies in Weddell seals (Leptonychotes weddellii) from cape Shirref, Antarctica

    OpenAIRE

    O. BLANK; P. RETAMAL; P. ABALOS; D. TORRES

    2002-01-01

    Posterior al hallazgo de anticuerpos anti-Brucella en muestras de lobo fino antártico (Arctocephalus gazella), el estudio serológico en Pinnipedia Antárticos se continuó con el fin de determinar la presencia de anticuerpos anti-Brucella en otras especies. Se colectaron muestras de sangre y de fluido extravascular de 12 ejemplares de foca de Weddell (Leptonychotes weddellii) encontrados en el Sitio de Especial Interés Científico (SEIC) Nº 32 y sitio CCAMLR Ecosystem Monitoring Program (CEMP) N...

  15. Chronic Brucella Infection Induces Selective and Persistent Interferon Gamma-Dependent Alterations of Marginal Zone Macrophages in the Spleen.

    Science.gov (United States)

    Machelart, Arnaud; Khadrawi, Abir; Demars, Aurore; Willemart, Kevin; De Trez, Carl; Letesson, Jean-Jacques; Muraille, Eric

    2017-11-01

    The spleen is known as an important filter for blood-borne pathogens that are trapped by specialized macrophages in the marginal zone (MZ): the CD209(+) MZ macrophages (MZMs) and the CD169(+) marginal metallophilic macrophages (MMMs). Acute systemic infection strongly impacts MZ populations and the location of T and B lymphocytes. This phenomenon has been linked to reduced chemokine secretion by stromal cells. Brucella spp. are the causative agent of brucellosis, a widespread zoonotic disease. Here, we used Brucella melitensis infection as a model to investigate the impact of chronic stealth infection on splenic MZ macrophage populations. During the late phase of Brucella infection, we observed a loss of both MZMs and MMMs, with a durable disappearance of MZMs, leading to a reduction of the ability of the spleen to take up soluble antigens, beads, and unrelated bacteria. This effect appears to be selective as every other lymphoid and myeloid population analyzed increased during infection, which was also observed following Brucella abortus and Brucella suis infection. Comparison of wild-type and deficient mice suggested that MZ macrophage population loss is dependent on interferon gamma (IFN-γ) receptor but independent of T cells or tumor necrosis factor alpha receptor 1 (TNF-αR1) signaling pathways and is not correlated to an alteration of CCL19, CCL21, and CXCL13 chemokine mRNA expression. Our results suggest that MZ macrophage populations are particularly sensitive to persistent low-level IFN-γ-mediated inflammation and that Brucella infection could reduce the ability of the spleen to perform certain MZM- and MMM-dependent tasks, such as antigen delivery to lymphocytes and control of systemic infection. Copyright © 2017 American Society for Microbiology.

  16. Role of TLRs in Brucella mediated murine DC activation in vitro and clearance of pulmonary infection in vivo.

    Science.gov (United States)

    Surendran, Naveen; Hiltbold, Elizabeth M; Heid, Bettina; Akira, Shizuo; Standiford, Theodore J; Sriranganathan, Nammalwar; Boyle, Stephen M; Zimmerman, Kurt L; Makris, Melissa R; Witonsky, Sharon G

    2012-02-14

    Brucellosis is worldwide zoonoses affecting 500,000 people annually with no approved human vaccines available. Live attenuated Brucella abortus vaccine strain RB51 protects cattle through CD4 and CD8 T-cell mediated responses. However, limited information is known regarding how Brucella stimulate innate immunity. Although the most critical toll like receptors (TLRs) involved in the recognition of Brucella are TLR2, TLR4 and TLR9, it is important to identify the essential TLRs that induce DC activation/function in response to Brucella, to be able to upregulate both vaccine strain RB51-mediated protection, and clearance of pathogenic strain 2308. Furthermore, in spite of the importance of aerosol transmission of Brucella, no published studies have addressed the role of TLRs in the clearance of strain 2308 or strain RB51 from intranasally infected mice. Therefore, we used a (a) bone marrow derived dendritic cell model in TLRKO and control mice to assess the differential role of pathogenic and vaccine strains to induce DC activation and function in vitro, and (b) respiratory model in TLRKO and control mice to assess the critical roles for TLRs in clearance of strains in vivo. In support of the essential TLRs in clearance and protection, we performed challenge experiments to identify if these critical TLRs (as agonists) could enhance vaccine induced protection against pathogenic strain 2308 in a respiratory model. We determined: vaccine strain RB51 induced significant (p≤0.05) DC activation vs. strain 2308 which was not dependent on a specific TLR; strain RB51 induced TNF-α production was TLR2 and TLR9 dependent, and IL-12 production was TLR2 and TLR4 dependent; TLR4 and TLR2 were critical for clearance of vaccine and pathogenic Brucella strains respectively; and TLR2 (pRB51-mediated protection against respiratory challenge with strain 2308 in the lung. Published by Elsevier Ltd.

  17. Brucella suis Vaccine Strain 2 Induces Endoplasmic Reticulum Stress that Affects Intracellular Replication in Goat Trophoblast Cells In vitro.

    Science.gov (United States)

    Wang, Xiangguo; Lin, Pengfei; Li, Yang; Xiang, Caixia; Yin, Yanlong; Chen, Zhi; Du, Yue; Zhou, Dong; Jin, Yaping; Wang, Aihua

    2016-01-01

    Brucella has been reported to impair placental trophoblasts, a cellular target where Brucella efficiently replicates in association with the endoplasmic reticulum (ER), and ultimately trigger abortion in pregnant animals. However, the precise effects of Brucella on trophoblast cells remain unclear. Here, we describe the infection and replication of Brucella suis vaccine strain 2 (B.suis.S2) in goat trophoblast cells (GTCs) and the cellular and molecular responses induced in vitro. Our studies demonstrated that B.suis.S2 was able to infect and proliferate to high titers, hamper the proliferation of GTCs and induce apoptosis due to ER stress. Tunicamycin (Tm), a pharmacological chaperone that strongly mounts ER stress-induced apoptosis, inhibited B.suis.S2 replication in GTCs. In addition, 4 phenyl butyric acid (4-PBA), a pharmacological chaperone that alleviates ER stress-induced apoptosis, significantly enhanced B.suis.S2 replication in GTCs. The Unfolded Protein Response (UPR) chaperone molecule GRP78 also promoted B.suis.S2 proliferation in GTCs by inhibiting ER stress-induced apoptosis. We also discovered that the IRE1 pathway, but not the PERK or ATF6 pathway, was activated in the process. However, decreasing the expression of phosphoIRE1α and IRE1α proteins with Irestatin 9389 (IRE1 antagonist) in GTCs did not affect the proliferation of B.suis.S2. Although GTC implantation was not affected upon B.suis.S2 infection, progesterone secretion was suppressed, and prolactin and estrogen secretion increased; these effects were accompanied by changes in the expression of genes encoding key steroidogenic enzymes. This study systematically explored the mechanisms of abortion in Brucella infection from the viewpoint of pathogen invasion, ER stress and reproductive endocrinology. Our findings may provide new insight for understanding the mechanisms involved in goat abortions caused by Brucella infection.

  18. Multiple-locus variable-number tandem-repeat analysis (MLVA) genotyping of human Brucella isolates in Malaysia.

    Science.gov (United States)

    Tay, Bee Yong; Ahmad, Norazah; Hashim, Rohaidah; Mohamed Zahidi, Jama'ayah; Thong, Kwai Lin; Koh, Xiu Pei; Mohd Noor, Azura

    2015-06-02

    Brucellosis is one of the most common zoonotic diseases worldwide. It can cause acute febrile illness in human and is a major health problem. Studies in human brucellosis in Malaysia is limited and so far no genotyping studies has been done on Brucella isolates. The aim of the study was to determine the genetic diversity among Brucella species isolated from human brucellosis, obtained over a 6-year period (2009-2014). In this study, the genotypic characteristics of 43 human Brucella melitensis isolates were analysed using multiple-locus variable-number tandem-repeat analysis (MLVA) which consisted of eight minisatellite loci (panel 1) and eight microsatellite loci; panels 2A (3 microsatellite loci) and panel 2B (5 microsatellite loci). Two human Brucella suis isolates were also investigated using the MLVA assay. Using panel 1 (MLVA8), two genotypes namely genotype 43 and 44 were obtained from the 43 B. melitensis isolates. Using the combination of panels 1 and 2A loci (MLVA11), two genotypes were obtained while using the complete panels 1, 2A and 2B, nine genotypes were obtained. The polymorphisms in using the complete panels (MLVA16) were observed in three loci from panel 2B, which showed a diversity index higher than 0.17. All B. melitensis isolates were closely related to the East Mediterranean group. For B. suis isolates, only genotype 6 and genotype 33 were obtained using panel 1 and MLVA11 respectively. In conclusion, the results of the present study showed a low genetic diversity among B. melitensis and B. suis isolates from human patients. Based on the MLVA16 assay, B. melitensis belonging to the East Mediterranean group is responsible for the vast majority of Brucella infections in our Malaysian patients. To our knowledge, this is the first genotyping study of human Brucella isolates in Malaysia.

  19. Endocarditis por Brucella canis: primer caso documentado en un paciente adulto en Argentina Brucella canis endocarditis: first documented case in Argentina

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    Valeria Manias

    2013-03-01

    Full Text Available Se describe el primer caso documentado de endocarditis por Brucella canis en Argentina. El paciente fue un varón adulto que consultó por edemas en miembros inferiores, registros febriles aislados de 2 meses de evolución y dolor precordial opresivo que irradiaba a brazo izquierdo. Negaba contacto con animales de cría o consumo de productos sin pasteurización. Estudios cardiológicos constataron endocarditis infecciosa. Se resuelve cirugía de recambio valvular ante fracaso terapéutico empírico con cefalotina, ampicilina y gentamicina. Los hemocultivos fueron positivos (4 de 4 muestras con bacilos gram negativos. Se realizó la identificación con técnica API 20 NE (bioMérieux, el método automatizado Phoenix (BD y las pruebas bioquímicas convencionales, sin concluir género ni especie. Se derivó la cepa al departamento de Bacteriología Especial INEI-ANLIS "Dr. Carlos G. Malbrán" donde se identificó al aislamiento como Brucella canis. Se rotó el esquema terapéutico a doxiciclina, rifampicina y trimetoprima-sulfametoxazol con buena evolución. La importancia del caso radica en la posible falla del tratamiento antimicrobiano empírico administrado para endocarditis, ya que B. canis no responde a los antimicrobianos convencionales para esta patología.We herein present the case of an adult male patient who consulted for lower extremity edema, a 2- month history of fever and oppressive chest pain radiating to the left arm. He referred neither contact with breeding animals nor consumption of unpasteurized dairy products. A diagnosis of endocarditis was confirmed by cardiac studies. Since the empirical treatment with cephalotin, ampicillin and gentamicin failed, the patient underwent aortic valve replacement. A total of four blood cultures were positive with a gram-negative rod. Bacterial identification was performed using the API 20 NE technique (bioMerieux, the Phoenix automated method (BD and conventional biochemical tests which were

  20. Brucella-Salmonella lipopolysaccharide chimeras are less permeable to hydrophobic probes and more sensitive to cationic peptides and EDTA than are their native Brucella sp. counterparts.

    Science.gov (United States)

    Freer, E; Moreno, E; Moriyón, I; Pizarro-Cerdá, J; Weintraub, A; Gorvel, J P

    1996-01-01

    A rough (R) Brucella abortus 45/20 mutant was more sensitive to the bactericidal activity of polymyxin B and lactoferricin B than was its smooth (S) counterpart but considerably more resistant than Salmonella montevideo. The outer membrane (OM) and isolated lipopolysaccharide (LPS) of S. montevideo showed a higher affinity for these cationic peptides than did the corresponding B. abortus OM and LPS. We took advantage of the moderate sensitivity of R B. abortus to cationic peptides to construct live R B. abortus-S-LPS chimeras to test the activities of polymyxin B, lactoferricin B, and EDTA. Homogeneous and abundant peripheral distribution of the heterologous S-LPS was observed on the surface of the chimeras, and this coating had no effect on the viability or morphology of the cells. When the heterologous LPS corresponded to the less sensitive bacterium S B. abortus S19, the chimeras were more resistant to cationic peptides; in contrast, when the S-LPS was from the more sensitive bacterium S. montevideo, the chimeras were more susceptible to the action of peptides and EDTA. A direct correlation between the amount of heterologous S-LPS on the surface of chimeric Brucella cells and peptide sensitivity was observed. Whereas the damage produced by polymyxin B in S. montevideo and B. abortus-S. montevideo S-LPS chimeras was manifested mainly as OM blebbing and inner membrane rolling, lactoferricin B caused inner membrane detachment, vacuolization, and the formation of internal electron-dense granules in these cells. Native S and R B. abortus strains were permeable to the hydrophobic probe N-phenyl-1-naphthylamine (NPN). In contrast, only reduced amounts of NPN partitioned into the OMs of the S. montevideo and B. abortus-S. montevideo S-LPS chimeras. Following peptide exposure, accelerated NPN uptake similar to that observed for S. montevideo was detected for the B. abortus-S. montevideo LPS chimeras. The partition of NPN into native or EDTA-, polymyxin B-, or

  1. First report of Brucella ceti-associated meningoencephalitis in a long-finned pilot whale Globicephala melas.

    Science.gov (United States)

    Davison, Nicholas J; Brownlow, Andrew; McGovern, Barry; Dagleish, Mark P; Perrett, Lorraine L; Dale, Emma-Jane; Koylass, Mark; Foster, Geoffrey

    2015-10-27

    Fatal Brucella ceti infection with histological lesions specific to the central nervous system has been described in only 3 species of cetaceans: striped dolphins Stenella coeruleoalba, Atlantic white-sided dolphins Lagenorhynchus acutus and short-beaked common dolphins Delphinus delphis. This paper describes the first report of a B. ceti-associated meningoencephalitis in a long-finned pilot whale Globicephala melas, showing the increasing range of species susceptibility. Brucella was recovered in larger numbers from cerebrospinal fluid than from brain tissue and is the sample of choice for isolation.

  2. Detection of Brucella melitensis in bovine milk and milk products from apparently healthy animals in Egypt by real-time PCR.

    Science.gov (United States)

    Wareth, Gamal; Melzer, Falk; Elschner, Mandy C; Neubauer, Heinrich; Roesler, Uwe

    2014-10-15

    Brucellosis in Egypt is an endemic disease among animals and humans. In endemic developing countries, dairy products produced from untreated milk are a potential threat to public health. The aim of this study was to detect brucellae in milk and milk products produced from apparently healthy animals to estimate the prevalence of contamination. Two hundred and fifteen unpasteurized milk samples were collected from apparently healthy cattle (n = 72) and buffaloes (n = 128) reared on small farms, and from milk shops (n = 15) producing dairy products for human consumption. All milk samples were examined by indirect enzyme-linked immunosorbent assay (iELISA) and real-time PCR (RT-PCR) to detect Brucella antibodies and Brucella-specific DNA, respectively. Using iELISA, anti-Brucella antibodies were detected in 34 samples (16%), while RT-PCR amplified Brucella-specific DNA from 17 milk samples (7.9%). Species-specific IS711 RT-PCR identified 16 of the RT-PCR-positive samples as containing B. melitensis DNA; 1 RT-PCR-positive sample was identified as containing B. abortus DNA. The detection of Brucella DNA in milk or milk products sold for human consumption, especially the highly pathogenic species B. melitensis, is of obvious concern. The shedding of Brucella spp. in milk poses an increasing threat to consumers in Egypt. Consumption of dairy products produced from non-pasteurized milk by individual farmers operating under poor hygienic conditions represents an unacceptable risk to public health.

  3. Anti-Brucella Antibodies in Moose (Alces alces gigas), Muskoxen (Ovibos moschatus), and Plains Bison (Bison bison bison) in Alaska, USA.

    Science.gov (United States)

    Nymo, Ingebjørg Helena; Beckmen, Kimberlee; Godfroid, Jacques

    2016-01-01

    We used an indirect enzyme-linked immunosorbent assay (iELISA) and the rose bengal test (RBT) to test for anti-Brucella antibodies in moose (Alces alces gigas), muskoxen (Ovibos moschatus), and plains bison (Bison bison bison) from various game management units (GMUs) in Alaska, US, sampled from 1982 to 2010. A portion of the sera had previously been tested with the standard plate test (SPT), the buffered Brucella antigen (BBA) card test, and the card test (CARD). No antibody-positive plains bison were identified. Anti-Brucella antibodies were detected in moose (iELISA, n=4/87; RBT, n=4/87; SPT, n=4/5; BBA, n=4/4) from GMU 23 captured in 1992, 1993, and 1995 and in muskoxen (iELISA, n=4/52; RBT, n=4/52; CARD, n=4/35) from GMUs 26A and 26B captured in 2004, 2006, and 2007. A negative effect of infection on the health of individuals of these species is probable. The presence of antibody-positive animals from 1992 to 2007 suggests presence of brucellae over time. The antibody-positive animals were found in northern Alaska, an area with a historically higher prevalence of Brucella-positive caribou, and a spillover of Brucella suis biovar 4 from caribou may have occurred. Brucella suis biovar 4 causes human brucellosis, and transmission from consumption of moose and muskoxen is possible.

  4. Vaccination of Elk (Cervus canadensis) with Brucella abortus Strain RB51 Overexpressing Superoxide Dismutase and Glycosyltransferase Genes Does Not Induce Adequate Protection against Experimental Brucella abortus Challenge.

    Science.gov (United States)

    Nol, Pauline; Olsen, Steven C; Rhyan, Jack C; Sriranganathan, Nammalwar; McCollum, Matthew P; Hennager, Steven G; Pavuk, Alana A; Sprino, Phillip J; Boyle, Stephen M; Berrier, Randall J; Salman, Mo D

    2016-01-01

    In recent years, elk (Cervus canadensis) have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area. In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the development of effective disease management strategies for wild elk herds is of utmost importance, not only for the prevention of reintroduction of brucellosis to cattle, but also for the overall health of the Greater Yellowstone Area elk populations. In two studies, we evaluated the efficacy of B. abortus strain RB51 over-expressing superoxide dismutase and glycosyltransferase for protecting elk from infection and disease caused by B. abortus after experimental infection with a virulent B. abortus strain. Our data indicate that the recombinant vaccine does not protect elk against brucellosis. Further, work is needed for development of an effective brucellosis vaccine for use in elk.

  5. Vaccination of elk (Cervus canadensis with Brucella abortus strain RB51 overexpressing superoxide dismutase and glycosyltransferase genes does not induce adequate protection against experimental Brucella abortus challenge

    Directory of Open Access Journals (Sweden)

    Pauline eNol

    2016-02-01

    Full Text Available In recent years, elk (Cervus canadensis have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area. In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the development of effective disease management strategies for wild elk herds is of utmost importance, not only for the prevention of reintroduction of brucellosis to cattle, but also for the overall health of the Greater Yellowstone Area elk populations. In two studies, we evaluated the efficacy of B. abortus strain RB51 over-expressing superoxide dismutase and glycosytransferase for protecting elk from infection and disease caused by B. abortus after experimental infection with a virulent B. abortus strain. Our data indicate that the recombinant vaccine does not protect elk against brucellosis. Further work is needed for development of an effective brucellosis vaccine for use in elk

  6. Molecular typing of isolates obtained from aborted foetuses in Brucella-free Holstein dairy cattle herd after immunisation with Brucella abortus RB51 vaccine in Egypt.

    Science.gov (United States)

    Wareth, Gamal; Melzer, Falk; Böttcher, Denny; El-Diasty, Mohamed; El-Beskawy, Mohamed; Rasheed, Nesma; Schmoock, Gernot; Roesler, Uwe; Sprague, Lisa D; Neubauer, Heinrich

    2016-12-01

    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.

  7. Identification of new IS711 insertion sites in Brucella abortus field isolates

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    Moriyón Ignacio

    2011-08-01

    Full Text Available Abstract Background Brucellosis is a zoonosis caused by Brucella spp., a group of highly homogeneous bacteria. The insertion sequence IS711 is characteristic of these bacteria, and occurs in variable numbers and positions, but always constant within a given species. This species-associated polymorphism is used in molecular typing and identification. Field isolates of B. abortus, the most common species infecting cattle, typically carry seven IS711 copies (one truncated. Thus far, IS711 transposition has only been shown in vitro and only for B. ovis and B. pinnipedialis, two species carrying a high number of IS711 copies, but never in other Brucella species, neither in vitro nor in field strains. Results We found several B. abortus strains isolated from milk and aborted fetuses that carried additional IS711 copies in two hitherto undescribed insertion sites: one in an intergenic region near to the 3' end of a putative lactate permease gene and the other interrupting the sequence of a marR transcriptional regulator gene. Interestingly, the second type of insertion was identified in isolates obtained repeatedly from the same herd after successive brucellosis outbreaks, an observation that proves the stability and virulence of the new genotype under natural conditions. Sequence analyses revealed that the new copies probably resulted from the transposition of a single IS711 copy common to all Brucella species sequenced so far. Conclusions Our results show that the replicative transposition of IS711 can occur under field conditions. Therefore, it represents an active mechanism for the emergence of genetic diversity in B. abortus thus contributing to intra-species genetic polymorphism.

  8. Species identification and molecular typing of human Brucella isolates from Kuwait.

    Science.gov (United States)

    Mustafa, Abu S; Habibi, Nazima; Osman, Amr; Shaheed, Faraz; Khan, Mohd W

    2017-01-01

    Brucellosis is a zoonotic disease of major concern in Kuwait and the Middle East. Human brucellosis can be caused by several Brucella species with varying degree of pathogenesis, and relapses are common after apparently successful therapy. The classical biochemical methods for identification of Brucella are time-consuming, cumbersome, and provide information limited to the species level only. In contrast, molecular methods are rapid and provide differentiation at intra-species level. In this study, four molecular methods [16S rRNA gene sequencing, real-time PCR, enterobacterial repetitive intergenic consensus (ERIC)-PCR and multilocus variable-number tandem-repeat analysis (MLVA)-8, MLVA-11 and MLVA-16 were evaluated for the identification and typing of 75 strains of Brucella isolated in Kuwait. 16S rRNA gene sequencing of all isolates showed 90-99% sequence identity with B. melitensis and real-time PCR with genus- and species- specific primers identified all isolates as B. melitensis. The results of ERIC-PCR suggested the existence of 75 ERIC genotypes of B. melitensis with a discriminatory index of 0.997. Cluster classification of these genotypes divided them into two clusters, A and B, diverging at ~25%. The maximum number of genotypes (n = 51) were found in cluster B5. MLVA-8 analysis identified all isolates as B. melitensis, and MLVA-8, MLVA-11 and MLVA-16 typing divided the isolates into 10, 32 and 71 MLVA types, respectively. Furthermore, the combined minimum spanning tree analysis demonstrated that, compared to MLVA types discovered all over the world, the Kuwaiti isolates were a distinct group of MLVA-11 and MLVA-16 types in the East Mediterranean Region.

  9. Transcriptome analysis of the Brucella abortus BvrR/BvrS two-component regulatory system.

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    Cristina Viadas

    Full Text Available BACKGROUND: The two-component BvrR/BvrS system is essential for Brucella abortus virulence. It was shown previously that its dysfunction alters the expression of some major outer membrane proteins and the pattern of lipid A acylation. To determine the genes regulated by BvrR/BvrS, we performed a whole-genome microarray analysis using B. abortus RNA obtained from wild type and bvrR mutant cells grown in the same conditions. METHODOLOGY/PRINCIPAL FINDINGS: A total of 127 differentially expressed genes were found: 83 were over expressed and 44 were less expressed in the bvrR mutant. Two operons, the phosphotransferase system and the maltose transport system, were down-regulated. Several genes involved in cell envelope or outer membrane biogenesis were differentially expressed: genes for outer membrane proteins (omp25a, omp25d, lipoproteins, LPS and fatty acid biosynthesis, stress response proteins, chaperones, flagellar genes, and twelve genes encoding ABC transport systems. Ten genes related with carbon metabolism (pckA and fumB among others were up-regulated in the bvrR mutant, and denitrification genes (nirK, norC and nosZ were also regulated. Notably, seven transcriptional regulators were affected, including VjbR, ExoR and OmpR that were less expressed in the bvrR mutant. Finally, the expression of eleven genes which have been previously related with Brucella virulence was also altered. CONCLUSIONS/SIGNIFICANCE: All these data corroborate the impact of BvrR/BvrS on cell envelope modulation, confirm that this system controls the carbon and nitrogen metabolism, and suggest a cross-talk among some regulators to adjust the Brucella physiology to the shift expected to occur during the transit from the extracellular to the intracellular niche.

  10. Serological survey for Brucella antibodies in donkeys of north-eastern Nigeria.

    Science.gov (United States)

    Tijjani, A O; Junaidu, A U; Salihu, M D; Farouq, A A; Faleke, O O; Adamu, S G; Musa, H I; Hambali, I U

    2017-08-01

    A cross-sectional epidemiological study was conducted to determine seroprevalence and risk factors influencing the presence of Brucella antibodies in donkeys of Borno State, north-eastern Nigeria. The study aimed at providing baseline information that may be used in planning a control policy against equine brucellosis. Blood samples were collected from 601 donkeys, comprised of 374 males and 227 females from the six agricultural zones of the state between March 2013 and September 2014. The sera obtained were tested for Brucella antibodies using Rose Bengal plate test (RBPT) and competitive enzyme-linked immunosorbent assay (cELISA). Of the 601 donkeys tested, 43 (7.2%) and 40 (6.7%) were seropositive by RBPT and cELISA, respectively. A seroprevalence of 8.6% was obtained in male and 3.5% in female donkeys. According to age, the highest seroprevalence of 9.6% was obtained from donkeys of age group 4-6 years. With respect to pregnancy status, a higher seroprevalence (6.8%) was obtained from pregnant donkeys compared to 3.8% obtained from the non-pregnant ones. There were statistically significant associations between the presence of antibodies and sex (p  0.05) was observed between the pregnancy status and presence of antibodies. The study concludes that Brucella infection is present in donkeys in all the agricultural zones of the state. The relatively high seroprevalence (7.2%) obtained is of public health concern because of the close interaction between donkeys, ruminants, and humans in the study area.

  11. Efficacies of gentamicin-loaded magnetite block ionomer complexes against chronic Brucella melitensis infection

    Energy Technology Data Exchange (ETDEWEB)

    Jain-Gupta, Neeta [Virginia-Maryland Regional College of Veterinary Medicine, Virginia Polytechnic Institute and State University, Department of Biomedical Sciences and Pathobiology (United States); Pothayee, Nipon; Pothayee, Nikorn [Virginia Polytechnic Institute and State University, Macromolecules and Interfaces Institute (United States); Tyler, Ronald; Caudell, David L. [Virginia-Maryland Regional College of Veterinary Medicine, Virginia Polytechnic Institute and State University, Department of Biomedical Sciences and Pathobiology (United States); Balasubramaniam, Sharavanan; Hu, Nan; Davis, Richey M.; Riffle, Judy S. [Virginia Polytechnic Institute and State University, Macromolecules and Interfaces Institute (United States); Sriranganathan, Nammalwar, E-mail: nathans@vt.edu [Virginia-Maryland Regional College of Veterinary Medicine, Virginia Polytechnic Institute and State University, Department of Biomedical Sciences and Pathobiology (United States)

    2013-11-15

    Anionic copolymers can enable intracellular delivery of cationic drugs which otherwise cannot cross cell membrane barriers. We tested the efficacy of gentamicin-loaded magnetite block ionomer complexes (MBICs) against intracellular Brucella melitensis. Anionic block copolymers were used to coat nanomagnetite through adsorption of a portion of anions on the particle surfaces, then the remaining anions were complexed with 30–32 weight percentage of gentamicin. The zeta potential changed from −39 to −13 mV after encapsulation of the drug with complementary charge. The gentamicin-loaded MBICs had intensity average hydrodynamic diameters of 62 nm, while the polymer-coated nanomagnetite particles without drug were 34 nm in size. No toxicity as measured by a MTS assay was observed upon incubation of the MBICs with J774A.1 murine macrophage-like cells. Confocal microscopic images showed that the MBICs were taken up by the macrophages and distributed in the cell cytoplasm and endosomal/lysosomal compartments. Upon treatment with gentamicin-loaded MBICs (3.5 Log{sub 10}), B. melitensis-infected macrophages showed significantly higher clearance of Brucella compared to the treatment with free g (0.9 Log{sub 10}). Compared to doxycycline alone, a combination of doxycycline and gentamicin (either free or encapsulated in MBICs) showed significantly higher clearance of B.melitensis from chronically infected mice. Histopathological examination of kidneys from the MBICs-treated mice revealed multifocal infiltration of macrophages containing intracytoplasmic iron (MBICs) in peri-renal adipose. Although MBICs showed similar efficacy as free gentamicin against Brucella in mice, our strategy presents an effective way to deliver higher loads of drugs intracellularly and ability to study the bio-distribution of drug carriers.

  12. Efficacies of gentamicin-loaded magnetite block ionomer complexes against chronic Brucella melitensis infection

    Science.gov (United States)

    Jain-Gupta, Neeta; Pothayee, Nipon; Pothayee, Nikorn; Tyler, Ronald; Caudell, David L.; Balasubramaniam, Sharavanan; Hu, Nan; Davis, Richey M.; Riffle, Judy S.; Sriranganathan, Nammalwar

    2013-11-01

    Anionic copolymers can enable intracellular delivery of cationic drugs which otherwise cannot cross cell membrane barriers. We tested the efficacy of gentamicin-loaded magnetite block ionomer complexes (MBICs) against intracellular Brucella melitensis. Anionic block copolymers were used to coat nanomagnetite through adsorption of a portion of anions on the particle surfaces, then the remaining anions were complexed with 30-32 weight percentage of gentamicin. The zeta potential changed from -39 to -13 mV after encapsulation of the drug with complementary charge. The gentamicin-loaded MBICs had intensity average hydrodynamic diameters of 62 nm, while the polymer-coated nanomagnetite particles without drug were 34 nm in size. No toxicity as measured by a MTS assay was observed upon incubation of the MBICs with J774A.1 murine macrophage-like cells. Confocal microscopic images showed that the MBICs were taken up by the macrophages and distributed in the cell cytoplasm and endosomal/lysosomal compartments. Upon treatment with gentamicin-loaded MBICs (3.5 Log10), B. melitensis-infected macrophages showed significantly higher clearance of Brucella compared to the treatment with free g (0.9 Log10). Compared to doxycycline alone, a combination of doxycycline and gentamicin (either free or encapsulated in MBICs) showed significantly higher clearance of B. melitensis from chronically infected mice. Histopathological examination of kidneys from the MBICs-treated mice revealed multifocal infiltration of macrophages containing intracytoplasmic iron (MBICs) in peri-renal adipose. Although MBICs showed similar efficacy as free gentamicin against Brucella in mice, our strategy presents an effective way to deliver higher loads of drugs intracellularly and ability to study the bio-distribution of drug carriers.

  13. Cervical Lymph Nodes as a Selective Niche for Brucella during Oral Infections

    Science.gov (United States)

    von Bargen, Kristine; Gagnaire, Aurélie; Arce-Gorvel, Vilma; de Bovis, Béatrice; Baudimont, Fannie; Chasson, Lionel; Bosilkovski, Mile; Papadopoulos, Alexia; Martirosyan, Anna; Henri, Sandrine; Mège, Jean-Louis; Malissen, Bernard; Gorvel, Jean-Pierre

    2015-01-01

    Cervical lymph nodes (CLN) are the first lymph nodes encountered by material taking the oral route. To study their role in orally acquired infections, we analyzed 307 patients of up to 14 years treated in the university clinic of Skopje, Macedonia, for brucellosis, a zoonotic bacterial disease frequently acquired by ingestion of contaminated dairy products. From these children, 36% had lymphadenopathy. Among orally infected children, lymphadenopathy with CLN being the only lymph nodes affected was significantly more frequent as compared to those infected by contact with animals (83% vs. 63%), suggesting a possible involvement of CLN during orally acquired human brucellosis. Using a murine model where bacteria are delivered into the oral cavity, we show that Brucella quickly and selectively colonize the CLN where they proliferate and persist over long periods of time for up to 50 days post-infection. A similar efficient though less specific drainage to CLN was found for Brucella, Salmonella typhimurium and fluorescent microspheres delivered by gavage, a pathway likely representing a mixed infection mode of intragastric and oral infection, suggesting a central pathway of drained material. Microspheres as well as bacteria drained to CLN predominately reside in cells expressing CD68 and no or low levels of CD11c. Even though no systemic response could be detected, Brucella induced a locally restricted inflammatory reaction with increased expression levels of interferon γ, interleukin (IL)-6, IL-12, granzyme B and a delayed induction of Nos2. Inflammation led to pronounced lymphadenopathy, infiltration of macrophages/monocytes expressing high levels of major histocompatibility complex II and to formation of epitheloid granulomas. Together, these results highlight the role of CLN in oral infections as both, an initial and efficient trap for bacterial invaders and as possible reservoir for chronic pathogens. They likewise cast a new light on the significance of oral

  14. Brucella melitensis in France: persistence in wildlife and probable spillover from Alpine ibex to domestic animals.

    Science.gov (United States)

    Mick, Virginie; Le Carrou, Gilles; Corde, Yannick; Game, Yvette; Jay, Maryne; Garin-Bastuji, Bruno

    2014-01-01

    Bovine brucellosis is a major zoonosis, mainly caused by Brucella abortus, more rarely by Brucella melitensis. France has been bovine brucellosis officially-free since 2005 with no cases reported in domestic/wild ruminants since 2003. In 2012, bovine and autochthonous human cases due to B. melitensis biovar 3 (Bmel3) occurred in the French Alps. Epidemiological investigations implemented in wild and domestic ruminants evidenced a high seroprevalence (>45%) in Alpine ibex (Capra ibex); no cases were disclosed in other domestic or wild ruminants, except for one isolated case in a chamois (Rupicapra rupicapra). These results raised the question of a possible persistence/emergence of Brucella in wildlife. The purpose of this study was to assess genetic relationships among the Bmel3 strains historically isolated in humans, domestic and wild ruminants in Southeastern France, over two decades, by the MLVA-panel2B assay, and to propose a possible explanation for the origin of the recent bovine and human infections. Indeed, this genotyping strategy proved to be efficient for this microepidemiological investigation using an interpretation cut-off established for a fine-scale setting. The isolates, from the 2012 domestic/human outbreak harbored an identical genotype, confirming a recent and direct contamination from cattle to human. Interestingly, they clustered not only with isolates from wildlife in 2012, but also with local historical domestic isolates, in particular with the 1999 last bovine case in the same massif. Altogether, our results suggest that the recent bovine outbreak could have originated from the Alpine ibex population. This is the first report of a B. melitensis spillover from wildlife to domestic ruminants and the sustainability of the infection in Alpine ibex. However, this wild population, reintroduced in the 1970s in an almost closed massif, might be considered as a semi-domestic free-ranging herd. Anthropogenic factors could therefore account with the

  15. Using real-time polymerase chain reaction as an alternative rapid method for enumeration of colony count in live Brucella vaccines

    Directory of Open Access Journals (Sweden)

    Waleed S. Shell

    2017-06-01

    Full Text Available Aim: Brucellosis is a major bacterial zoonosis of global importance affecting a range of animal species and man worldwide. It has economic, public health, and bio-risk importance. Control and prevention of animal brucellosis mainly depend on accurate diagnostic tools and implementation of effective and safe animal vaccination program. There are three types of animal Brucella live vaccines - Brucella melitensis Rev-1 vaccine, Brucella abortus S19, and B. abortus RB51. Evaluation of these vaccines depends mainly on enumeration of Brucella viable count. At present, used colony count method is time consuming, costly and requires especial skills. Hence, the aim of this study is to use and standardize real-time polymerase chain reaction (RT-PCR as an alternative, quantitative, sensitive, and rapid method to detect the colony count of Brucella in live Brucella vaccine. Materials and Methods: Four batches of different live Brucella vaccines were evaluated using of conventional bacterial count and RT-quantitative PCR (RT-qPCR using BSCP31 gene specific primers and probe. Standard curve was generated from DNA template extracted from 10-fold serial dilution of living B. abortus RB51 vaccine to evaluate the sensitivity of RT-qPCR. Results: Results revealed that three batches of living Brucella vaccines were acceptable for Brucella colony count when traditional bacterial enumeration method was used. Results of RT-qPCR were identical to that of conventional bacterial count. Conclusion: Results concluded that RT-qPCR was relatively sensitive compared to traditional bacterial colony count of these vaccines.

  16. Unusual Case of Occult Brucella Osteomyelitis in the Skull Detected by Bone Scintigraphy

    Energy Technology Data Exchange (ETDEWEB)

    Sohn, Myung Hee; Lim, Seok Tae; Jeong, Young Jin; Kim, Dong Wook; Jeong, Hwan Jeong; Lee, Chang Seob [Chonbuk National University Medical School, Jeonju (Korea, Republic of)

    2010-06-15

    Brucellosis is a worldwide infectious disease of animals that can be transmitted to humans. Osteoarticular involvement is the most common complication of brucellosis. A 47-year-old man, who was a stock breeder, complained of myalgia with fever and chills for 2 weeks. The serology titers and blood cultures for brucellosis were positive. Bone scintigraphy demonstrated a focally increased uptake in the left supra orbital area. Plain radiographs showed an osteolytic lesion, and an MRI revealed signal abnormalities in the corresponding site. We present an unusual case of occult Brucella osteomyelitis in the frontal bone of the skull detected by done scintigraphy.

  17. Gamma radiation grafted polymers for immobilization of Brucella antigen in diagnostic test studies

    Science.gov (United States)

    Docters, E. H.; Smolko, E. E.; Suarez, C. E.

    The radiation grafting process has a wide field of industrial applications, and in the recent years the immobilization of biocomponents in grafted polymeric materials obtained by means of ionizing radiations is a new and important contribution to biotechnology. In the present work, gamma preirradiation grafting method was employed to produce acrylics hydrogels onto polyethylene (PE), polyvinyl chloride (PVC) and polystyrene (PS). Two monomers were used to graft the previously mentioned polymers: methacrylic acid (MAAc) and acrylamide (AAm), and several working conditions were considered as influencing the degree of grafting. All this grafted polymers were used to study the possibility of a subsequent immobilization of Brucella antigen (BAg) in diagnostic test studies (ELISA).

  18. Et tilfælde af brucella spondylodiscitis efter rejse til Libanon

    DEFF Research Database (Denmark)

    Nielsen, Stig Lønberg; Johansen, Isik Somuncu

    2012-01-01

    Brucellosis is a widespread endemic zoonotic infection affecting more than 500,000 people per year. The disease is very uncommon in Denmark and almost always imported. We present a case of a 57 year-old male with blood culture and magnetic resonance imaging verified brucella spondylodiscitis. Pri...... to debut of symptoms the patient had visited Lebanon where he had ingested unpasteurized goat milk. The patient was initially treated with an antimicrobial chemotherapy regimen for 12 weeks, which was prolonged due to inadequate radiological response....

  19. Proteínas inmunodominantes de Brucella Melitensis evaluadas por Western Blot

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    Elizabeth Anaya

    1997-01-01

    Full Text Available Se separaron extractos de proteínas totales de Brucella melitensis en gel 15% SDS-PAGE. Su seroreactividad fue analizada por Western Blot con resultados satisfactorios. Para éste propósito sueros controles negativos (n=03, sueros de pacientes con brucelosis (n=34, cólera (n=12, tifoidea (n=02 y tuberculosis (n=02 fueron usados. Esta prueba inmunodiagnóstica detectó bandas seroreactivas altamente específicas (100% correspondientes a 8,14,18, un complejo de 25-48 y 58kDa. La sensibilidad del test fue del 90% usando los sueros antes mencionados.

  20. Seroprevalencia de Brucella spp en estudiantes de Medicina Veterinaria, Bogotá, Colombia

    OpenAIRE

    Iván Alberto Méndez R; Diego Mauricio Trujillo C; Cristian Camilo Duque S; Edward Javier Acero M; Luis Ángel Cabrera; Diana Patricia Pachón B

    2013-01-01

    Introducción: La brucelosis es un enfermedad zoonotica relacionada al contacto con perros y con ganado vacuno, porcino y caprino, el agente etiológico pertenece al género Brucella con diez especies de importancia para el ser humano; su capacidad para evadir el sistema inmune y sobrevivir en el fago-lisosoma genera un cuadro clínico crónico en el ser humano caracterizado por fiebre ondulante y afectación osteoarticular. Su amplia variedad de expresión clínica genera un reto al momento del diag...

  1. Outbreak of laboratory-acquired Brucella abortus in Brazil: a case report

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    Ana Luisa Calixto Rodrigues

    2013-12-01

    Full Text Available Human brucellosis is an occupational disease affecting workers in slaughterhouses, butcher shops and the milk and dairy product industry as well as individuals who work in clinical or research laboratories. We report the first outbreak of a Brucella abortus infection in a Brazilian laboratory and compare the data obtained with reports available in the literature. Exposure was a result of damage to a biological safety cabinet and failure of the unidirectional airflow ventilation system. An epidemiological investigation identified 3 seroconverted individuals, 1 of whom had clinical manifestations and laboratory results compatible with infection at the time of exposure (n=11; attack rate=9.1%.

  2. Differential Requirements for VirB1 and VirB2 during Brucella abortus Infection

    OpenAIRE

    den Hartigh, Andreas B.; Sun, Yao-Hui; Sondervan, David; Heuvelmans, Niki; Reinders, Marjolein O.; Ficht, Thomas A.; Tsolis, Renée M.

    2004-01-01

    The Brucella abortus virB operon, encoding a type IV secretion system (T4SS), is required for intracellular replication and persistent infection in the mouse model. The products of the first two genes of the virB operon, virB1 and virB2, are predicted to be localized at the bacterial surface, where they could potentially interact with host cells. Studies to date have focused on characterization of transposon mutations in these genes, which are expected to exert polar effects on downstream gen...

  3. Brucella Rough Mutant Induce Macrophage Death via Activating IRE1α Pathway of Endoplasmic Reticulum Stress by Enhanced T4SS Secretion

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    Peng Li

    2017-09-01

    Full Text Available Brucella is a Gram-negative facultative intracellular pathogen that causes the worldwide zoonosis, known as brucellosis. Brucella virulence relies mostly on its ability to invade and replicate within phagocytic cells. The type IV secretion system (T4SS and lipopolysaccharide are two major Brucella virulence factors. Brucella rough mutants reportedly induce the death of infected macrophages, which is T4SS dependent. However, the underlying molecular mechanism remains unclear. In this study, the T4SS secretion capacities of Brucella rough mutant and its smooth wild-type strain were comparatively investigated, by constructing the firefly luciferase fused T4SS effector, BPE123 and VceC. In addition, quantitative real-time PCR and western blotting were used to analyze the T4SS expression. The results showed that T4SS expression and secretion were enhanced significantly in the Brucella rough mutant. We also found that the activity of the T4SS virB operon promoter was notably increased in the Brucella rough mutant, which depends on quorum sensing-related regulators of VjbR upregulation. Cell infection and cell death assays revealed that deletion of vjbR in the Brucella rough mutant absolutely abolished cytotoxicity within macrophages by downregulating T4SS expression. This suggests that up-regulation of T4SS promoted by VjbR in rough mutant ΔrfbE contribute to macrophage death. In addition, we found that the Brucella rough mutant induce macrophage death via activating IRE1α pathway of endoplasmic reticulum stress. Taken together, our study provide evidence that in comparison to the Brucella smooth wild-type strain, VjbR upregulation in the Brucella rough mutant increases transcription of the virB operon, resulting in overexpression of the T4SS gene, accompanied by the over-secretion of effecter proteins, thereby causing the death of infected macrophages via activating IRE1α pathway of endoplasmic reticulum stress, suggesting novel insights into the

  4. Cloning and Expression of 31kDa Outer Membrane Protein of Brucella melitansis in E.coli

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    Sayeneh Khodadadi

    2012-04-01

    Full Text Available Background & Objectives: The identification of Brucella spp. antigens with the capacity to elicit a protective immune response is of the great interest for the researchers. So, characterization and assessment of diverse antigens of Brucella need to be evaluated. In this study, we report the cloning and expression of the gene coding for 31 KDa OMP (OMP31 of Brucella melitensis 16M.   Methods: Brucella melitensis Omp31 gene was amplified with specific primers, cloned into pJET1/2 and subsequently subcloned in pET28a (+ vector. Both these recombinant plasmids were sequenced and then after, expression of recombinant protein was induced by 1mM IPTG. Western blot analysis was also performed by polyclonal rabbit antiserum.   Results: Omp31 successfully was cloned in both plasmid vectors. The recombinant Omp31 was expressed in E.coli host and purified with significant yield. Western blot results along with those of sequencing ensured accurate production of recombinant omp31 and retaining of its partial epitopes.   Conclusion: Our results show that, an expression host such as E. coli is suitable for omp31 production.

  5. MLVA as an Epidemiological Tool To Trace Back Brucella melitensis Biovar 1 Re-Emergence in Italy.

    Science.gov (United States)

    De Massis, F; Ancora, M; Atzeni, M; Rolesu, S; Bandino, E; Danzetta, M L; Zilli, K; Di Giannatale, E; Scacchia, M

    2015-10-01

    Brucellosis is an important zoonosis caused by Brucella spp., still prevalent in most areas of the world. Brucellosis control in animals is the key to protect humans. The knowledge of Brucella spp. prevailing genotypes in a territory represents an important epidemiological tool to formulate policies and strategies for disease control and to trace back the introduction of new strains previously considered as exotic. In the last years, multiple-locus variable number tandem repeat analysis (MLVA) has been proposed as complementary to classical biotyping methods. MLVA may add important information to the classical epidemiological investigation techniques, to help in tracing back sources of infection in brucellosis outbreaks. Sardinia is an Italian region officially free from sheep and goats brucellosis since 1998. In 2011, Brucella melitensis biovar 1, a biotype not reported in Italy since 1995, was isolated in one flock in the region. The genotyping MLVA-16 showed that isolates belonged to a rare American lineage, confirming it was introduced from other countries. The strain was considered as probably originating from Spain, where this lineage is endemic. BrucellaMLVA-16 has been proved to be useful to analyse the epidemiological correlation of strains enabling to trace its geographic origin by comparing their previously reported genetic patterns. © 2015 The Authors. Transboundary and Emerging Diseases Published by Blackwell Verlag GmbH.

  6. Literatuuronderzoek naar gegevens betreffende de betekenis van een aantal verwekkers van zoonosen in verband met de vleesconsumptie. VIII: Brucella abortus

    NARCIS (Netherlands)

    Bos; J.M.; Engel; H.W.B.; Groothuis; D.G.; Knapen; F. van; Weiss; J.W.

    1986-01-01

    Brucellose bij de mens heeft meestal een weinig specifiek, griepachtig beeld. In Nederland is de voornaamste verwekker Brucella abortus, waar toe het literatuuronderzoek zich dan ook beperkt. Bij dieren is het rund de voornaamste gastheer en leidt een infectie bij drachtige dieren tot abortus.

  7. Brucella abortus strain RB51 leucine auxotroph as an environmentally safe vaccine for plasmid maintenance and antigen overexpression.

    Science.gov (United States)

    Rajasekaran, Parthiban; Seleem, Mohamed N; Contreras, Andrea; Purwantini, Endang; Schurig, Gerhardt G; Sriranganathan, Nammalwar; Boyle, Stephen M

    2008-11-01

    To avoid potentiating the spread of an antibiotic resistance marker, a plasmid expressing a leuB gene and a heterologous antigen, green fluorescent protein (GFP), was shown to complement a leucine auxotroph of cattle vaccine strain Brucella abortus RB51, which protected CD1 mice from virulent B. abortus 2308 and elicited GFP antibodies.

  8. Brucella pinnipedialis hooded seal (Cystophora cristata) strain in the mouse model with concurrent exposure to PCB 153.

    Science.gov (United States)

    Nymo, Ingebjørg H; das Neves, Carlos G; Tryland, Morten; Bårdsen, Bård-Jørgen; Santos, Renato Lima; Turchetti, Andreia Pereira; Janczak, Andrew M; Djønne, Berit; Lie, Elisabeth; Berg, Vidar; Godfroid, Jacques

    2014-05-01

    Brucellosis, a worldwide zoonosis, is linked to reproductive problems in primary hosts. A high proportion of Brucella-positive hooded seals (Cystophora cristata) have been detected in the declined Northeast Atlantic stock. High concentrations of polychlorinated biphenyls (PCBs) have also been discovered in top predators in the Arctic, including the hooded seal, PCB 153 being most abundant. The aim of this study was to assess the pathogenicity of Brucella pinnipedialis hooded seal strain in the mouse model and to evaluate the outcome of Brucella spp. infection after exposure of mice to PCB 153. BALB/c mice were infected with B. pinnipedialis hooded seal strain or Brucella suis 1330, and half from each group was exposed to PCB 153 through the diet. B. pinnipedialis showed a reduced pathogenicity in the mouse model as compared to B. suis 1330. Exposure to PCB 153 affected neither the immunological parameters, nor the outcome of the infection. Altogether this indicates that it is unlikely that B. pinnipedialis contribute to the decline of hooded seals in the Northeast Atlantic. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. "HOOF-Print" Genotyping and Haplotype Inference Discriminates among Brucella spp Isolates From a Small Spatial Scale

    Science.gov (United States)

    We demonstrate that the “HOOF-Print” assay provides high power to discriminate among Brucella isolates collected on a small spatial scale (within Portugal). Additionally, we illustrate how haplotype identification using non-random association among markers allows resolution of B. melitensis biovars ...

  10. Patterns of Hepatosplenic Brucella Abscesses on Cross-Sectional Imaging: A Review of Clinical and Imaging Features

    NARCIS (Netherlands)

    Heller, Tom; Bélard, Sabine; Wallrauch, Claudia; Carretto, Edoardo; Lissandrin, Raffaella; Filice, Carlo; Brunetti, Enrico

    2015-01-01

    While diffuse involvement of liver and spleen is frequently seen in brucellosis, suppurative abscesses caused by Brucella are less common but well described. With the increased availability of cross-sectional imaging techniques, reports have become more frequent. Four patients with hepatosplenic

  11. Coinfection and vertical transmission of Brucella and Morbillivirus in a neonatal sperm whale (Physeter macrocephalus) in Hawaii, USA.

    Science.gov (United States)

    West, Kristi L; Levine, Gregg; Jacob, Jessica; Jensen, Brenda; Sanchez, Susan; Colegrove, Kathleen; Rotstein, David

    2015-01-01

    The viral genus Morbillivirus and the bacterial genus Brucella have emerged as important groups of pathogens that are known to affect cetacean health on a global scale, but neither pathogen has previously been reported from endangered sperm whales (Physeter macrocephalus). A female neonate sperm whale stranded alive and died near Laie on the island of Oahu, Hawaii, US, in May of 2011. Congestion of the cerebrum and enlarged lymph nodes were noted on the gross necropsy. Microscopic findings included lymphoid depletion, chronic meningitis, and pneumonia, suggesting an in utero infection. Cerebrum, lung, umbilicus, and select lymph nodes (tracheobronchial and mediastinal) were positive for Brucella by PCR. Brucella sp. was also cultured from the cerebrum and from mediastinal and tracheobronchial lymph nodes. Twelve different tissues were screened for Morbillivirus by reverse-transcriptase (RT)-PCR and select tissues by immunohistochemistry, but only the tracheobronchial lymph node and spleen were positive by RT-PCR. Pathologic findings observed were likely a result of Brucella, but Morbillivirus may have played a key role in immune suppression of the mother and calf. The in utero infection in this individual strongly supports vertical transmission of both pathogens.

  12. A Live Vaccine from Brucella abortus Strain 82 for Control of Cattle Brucellosis in the Russian Federation

    Science.gov (United States)

    During the first half of the 20th century, widespread regulatory efforts to control cattle brucellosis (Brucella abortus) in the Union of Soviet Socialist Republics were essentially nonexistent, and control was limited to selective test and slaughter of serologic agglutination reactors. By the 1950...

  13. Immune Responses and Protection against Experimental Brucella suis biovar 1 Challenge in Non-vaccinated or RB51-Vaccinated Cattle

    Science.gov (United States)

    Twenty Hereford heifers, approximately 9 months of age, were vaccinated with saline (control) or 2 x 10**10 CFU of Brucella abortus strain RB51 (RB51) vaccine. Immunologic responses after inoculation demonstrated significantly greater (P<0.05) antibody and proliferative responses to RB51 antigens i...

  14. Differentiation between serological responses to Brucella suis and Yersinia enterocolitica serotype O : 9 after natural or experimental infection in pigs

    DEFF Research Database (Denmark)

    Jungersen, Gregers; Sørensen, Vibeke; Giese, Steen Bjørck

    2006-01-01

    with responses of B. suis biovar 2-inoculated pigs. FPSR were limited to 2-9 weeks post-YeO:9 inoculation, while B. suis-infected pigs were test-positive throughout the 21-week period of investigation. Although YeO:9-inoculated pigs exhibited FPSR in Brucella tests for a limited period of time, the serological...

  15. Outer Membrane Protein 25 of Brucella Activates Mitogen-Activated Protein Kinase Signal Pathway in Human Trophoblast Cells

    Directory of Open Access Journals (Sweden)

    Jing Zhang

    2017-12-01

    Full Text Available Outer membrane protein 25 (OMP25, a virulence factor from Brucella, plays an important role in maintaining the structural stability of Brucella. Mitogen-activated protein kinase (MAPK signal pathway widely exists in eukaryotic cells. In this study, human trophoblast cell line HPT-8 and BALB/c mice were infected with Brucella abortus 2308 strain (S2308 and 2308ΔOmp25 mutant strain. The expression of cytokines and activation of MAPK signal pathway were detected. We found that the expressions of tumor necrosis factor-α, interleukin-1, and interleukin-10 (IL-10 were increased in HPT-8 cells infected with S2308 and 2308ΔOmp25 mutant. S2308 also activated p38 phosphorylation protein, extracellular-regulated protein kinases (ERK, and Jun-N-terminal kinase (JNK from MAPK signal pathway. 2308ΔOmp25 could not activate p38, ERK, and JNK branches. Immunohistochemistry experiments showed that S2308 was able to activate phosphorylation of p38 and ERK in BABL/c mice. However, 2308ΔOmp25 could weakly activate phosphorylation of p38 and ERK. These results suggest that Omp25 played an important role in the process of Brucella activation of the MAPK signal pathway.

  16. Efficacy of dart or booster vaccination with strain RB51 in protecting bison against experimental Brucella abortus challenge

    Science.gov (United States)

    Vaccination is an effective tool for reducing the prevalence of brucellosis in natural hosts. In this study, we characterized the efficacy of the Brucella abortus strain RB51 (RB51) vaccine in bison when delivered by single intramuscular vaccination (Hand RB51), single pneumatic dart delivery (Dart ...

  17. Comparison of abortion and infection after experimental challenge of pregnant bison and cattle with Brucella abortus strain 2308

    Science.gov (United States)

    A comparative study was conducted using data from naive bison (n=45) and cattle (n=46) from 8 and 6 studies, respectively, in which a standardized Brucella abortus strain 2308 experimental challenge was administered. The incidence of abortion, fetal infection, uterine or mammary infection, or infec...

  18. In Situ Microscopy Analysis Reveals Local Innate Immune Response Developed around Brucella Infected Cells in Resistant and Susceptible Mice

    Science.gov (United States)

    Copin, Richard; Vitry, Marie-Alice; Hanot Mambres, Delphine; Machelart, Arnaud; De Trez, Carl; Vanderwinden, Jean-Marie; Magez, Stefan; Akira, Shizuo; Ryffel, Bernhard; Carlier, Yves; Letesson, Jean-Jacques; Muraille, Eric

    2012-01-01

    Brucella are facultative intracellular bacteria that chronically infect humans and animals causing brucellosis. Brucella are able to invade and replicate in a broad range of cell lines in vitro, however the cells supporting bacterial growth in vivo are largely unknown. In order to identify these, we used a Brucella melitensis strain stably expressing mCherry fluorescent protein to determine the phenotype of infected cells in spleen and liver, two major sites of B. melitensis growth in mice. In both tissues, the majority of primary infected cells expressed the F4/80 myeloid marker. The peak of infection correlated with granuloma development. These structures were mainly composed of CD11b+ F4/80+ MHC-II+ cells expressing iNOS/NOS2 enzyme. A fraction of these cells also expressed CD11c marker and appeared similar to inflammatory dendritic cells (DCs). Analysis of genetically deficient mice revealed that differentiation of iNOS+ inflammatory DC, granuloma formation and control of bacterial growth were deeply affected by the absence of MyD88, IL-12p35 and IFN-γ molecules. During chronic phase of infection in susceptible mice, we identified a particular subset of DC expressing both CD11c and CD205, serving as a reservoir for the bacteria. Taken together, our results describe the cellular nature of immune effectors involved during Brucella infection and reveal a previously unappreciated role for DC subsets, both as effectors and reservoir cells, in the pathogenesis of brucellosis. PMID:22479178

  19. Full restoration of Brucella-infected dendritic cell functionality through Vγ9Vδ2 T helper type 1 crosstalk.

    Directory of Open Access Journals (Sweden)

    Ming Ni

    Full Text Available Vγ9Vδ2 T cells play an important role in the immune response to infectious agents but the mechanisms contributing to this immune process remain to be better characterized. Following their activation, Vγ9Vδ2 T cells develop cytotoxic activity against infected cells, secrete large amounts of cytokines and influence the function of other effectors of immunity, notably cells playing a key role in the initiation of the adaptive immune response such as dendritic cells. Brucella infection dramatically impairs dendritic cell maturation and their capacity to present antigens to T cells. Herein, we investigated whether V T cells have the ability to restore the full functional capacities of Brucella-infected dendritic cells. Using an in vitro multicellular infection model, we showed that: 1/Brucella-infected dendritic cells activate Vγ9Vδ2 T cells through contact-dependent mechanisms, 2/activated Vγ9Vδ2 T cells induce full differentiation into IL-12 producing cells of Brucella-infected dendritic cells with functional antigen presentation activity. Furthermore, phosphoantigen-activated Vγ9Vδ2 T cells also play a role in triggering the maturation process of dendritic cells already infected for 24 h. This suggests that activated Vγ9Vδ2 T cells could be used to modulate the outcome of infectious diseases by promoting an adjuvant effect in dendritic cell-based cellular therapies.

  20. Prevalence of Brucella antibodies in rural and suburban communities in three provinces of Turkey: need for improved diagnosis and prevention

    NARCIS (Netherlands)

    Kose, Sukran; Smits, Henk L.; Abdoel, Theresia H.; Ozbel, Yusuf

    2006-01-01

    To determine the seroprevalence of Brucella-specific antibodies in rural and suburban communities in different provinces of Anatolia. Cross-sectional seroepidemiological study on serum samples collected in communities in two relatively developed provinces in west Anatolia with an official low

  1. A Rapid Detection Method of Brucella with Quantum Dots and Magnetic Beads Conjugated with Different Polyclonal Antibodies

    Science.gov (United States)

    Song, Dandan; Qu, Xiaofeng; Liu, Yushen; Li, Li; Yin, Dehui; Li, Juan; Xu, Kun; Xie, Renguo; Zhai, Yue; Zhang, Huiwen; Bao, Hao; Zhao, Chao; Wang, Juan; Song, Xiuling; Song, Wenzhi

    2017-03-01

    Brucella spp. are facultative intracellular bacteria that cause zoonotic disease of brucellosis worldwide. Traditional methods for detection of Brucella spp. take 48-72 h that does not meet the need of rapid detection. Herein, a new rapid detection method of Brucella was developed based on polyclonal antibody-conjugating quantum dots and antibody-modified magnetic beads. First, polyclonal antibodies IgG and IgY were prepared and then the antibody conjugated with quantum dots (QDs) and immunomagnetic beads (IMB), respectively, which were activated by N-(3-dimethylaminopropyl)- N'-ethylcar-bodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) to form probes. We used the IMB probe to separate the Brucella and labeled by the QD probe, and then detected the fluorescence intensity with a fluorescence spectrometer. The detection method takes 105 min with a limit of detection of 103 CFU/mL and ranges from 10 to 105 CFU/mL ( R 2 = 0.9983), and it can be well used in real samples.

  2. LOV Histidine Kinase Modulates the General Stress Response System and Affects the virB Operon Expression in Brucella abortus.

    Directory of Open Access Journals (Sweden)

    Gabriela Sycz

    Full Text Available Brucella is the causative agent of the zoonotic disease brucellosis, and its success as an intracellular pathogen relies on its ability to adapt to the harsh environmental conditions that it encounters inside the host. The Brucella genome encodes a sensor histidine kinase containing a LOV domain upstream from the kinase, LOVHK, which plays an important role in light-regulated Brucella virulence. In this report we study the intracellular signaling pathway initiated by the light sensor LOVHK using an integrated biochemical and genetic approach. From results of bacterial two-hybrid assays and phosphotransfer experiments we demonstrate that LOVHK functionally interacts with two response regulators: PhyR and LovR, constituting a functional two-component signal-transduction system. LOVHK contributes to the activation of the General Stress Response (GSR system in Brucella via PhyR, while LovR is proposed to be a phosphate-sink for LOVHK, decreasing its phosphorylation state. We also show that in the absence of LOVHK the expression of the virB operon is down-regulated. In conclusion, our results suggest that LOVHK positively regulates the GSR system in vivo, and has an effect on the expression of the virB operon. The proposed regulatory network suggests a similar role for LOVHK in other microorganisms.

  3. Assessment of genetic diversity of zoonotic Brucella spp. recovered from livestock in Egypt using multiple locus VNTR analysis.

    Science.gov (United States)

    Menshawy, Ahmed M S; Perez-Sancho, Marta; Garcia-Seco, Teresa; Hosein, Hosein I; García, Nerea; Martinez, Irene; Sayour, Ashraf E; Goyache, Joaquín; Azzam, Ragab A A; Dominguez, Lucas; Alvarez, Julio

    2014-01-01

    Brucellosis is endemic in most parts of Egypt, where it is caused mainly by Brucella melitensis biovar 3, and affects cattle and small ruminants in spite of ongoing efforts devoted to its control. Knowledge of the predominant Brucella species/strains circulating in a region is a prerequisite of a brucellosis control strategy. For this reason a study aiming at the evaluation of the phenotypic and genetic heterogeneity of a panel of 17 Brucella spp. isolates recovered from domestic ruminants (cattle, buffalo, sheep, and goat) from four governorates during a period of five years (2002-2007) was carried out using microbiological tests and molecular biology techniques (PCR, MLVA-15, and sequencing). Thirteen strains were identified as B. melitensis biovar 3 while all phenotypic and genetic techniques classified the remaining isolates as B. abortus (n = 2) and B. suis biovar 1 (n = 2). MLVA-15 yielded a high discriminatory power (h = 0.801), indicating a high genetic diversity among the B. melitensis strains circulating among domestic ruminants in Egypt. This is the first report of the isolation of B. suis from cattle in Egypt which, coupled with the finding of B. abortus, suggests a potential role of livestock as reservoirs of several zoonotic Brucella species in the region.

  4. Assessment of Genetic Diversity of Zoonotic Brucella spp. Recovered from Livestock in Egypt Using Multiple Locus VNTR Analysis

    Directory of Open Access Journals (Sweden)

    Ahmed M. S. Menshawy

    2014-01-01

    Full Text Available Brucellosis is endemic in most parts of Egypt, where it is caused mainly by Brucella melitensis biovar 3, and affects cattle and small ruminants in spite of ongoing efforts devoted to its control. Knowledge of the predominant Brucella species/strains circulating in a region is a prerequisite of a brucellosis control strategy. For this reason a study aiming at the evaluation of the phenotypic and genetic heterogeneity of a panel of 17 Brucella spp. isolates recovered from domestic ruminants (cattle, buffalo, sheep, and goat from four governorates during a period of five years (2002–2007 was carried out using microbiological tests and molecular biology techniques (PCR, MLVA-15, and sequencing. Thirteen strains were identified as B. melitensis biovar 3 while all phenotypic and genetic techniques classified the remaining isolates as B. abortus (n=2 and B. suis biovar 1 (n=2. MLVA-15 yielded a high discriminatory power (h=0.801, indicating a high genetic diversity among the B. melitensis strains circulating among domestic ruminants in Egypt. This is the first report of the isolation of B. suis from cattle in Egypt which, coupled with the finding of B. abortus, suggests a potential role of livestock as reservoirs of several zoonotic Brucella species in the region.

  5. Brucella 'HOOF-Prints': strain typing by multi-locus analysis of variable number tandem repeats (VNTRs

    Directory of Open Access Journals (Sweden)

    Halling Shirley M

    2003-07-01

    Full Text Available Abstract Background Currently, there are very few tools available for subtyping Brucella isolates for epidemiological trace-back. Subtyping is difficult because of the genetic homogeneity within the genus. Sequencing of the genomes from three Brucella species has facilitated the search for DNA sequence variability. Recently, hypervariability among short tandem repeat sequences has been exploited for strain-typing of several bacterial pathogens. Results An eight-base pair tandem repeat sequence was discovered in nine genomic loci of the B. abortus genome. Eight loci were hypervariable among the three Brucella species. A PCR-based method was developed to identify the number of repeat units (alleles at each locus, generating strain-specific fingerprints. None of the loci exhibited species- or biovar-specific alleles. Sometimes, a species or biovar contained a specific allele at one or more loci, but the allele also occurred in other species or biovars. The technique successfully differentiated the type strains for all Brucella species and biovars, among unrelated B. abortus biovar 1 field isolates in cattle, and among B. abortus strains isolated from bison and elk. Isolates from the same herd or from short-term in vitro passage exhibited little or no variability in fingerprint pattern. Sometimes, isolates from an animal would have multiple alleles at a locus, possibly from mixed infections in enzootic areas, residual disease from incomplete depopulation of an infected herd or molecular evolution within the strain. Therefore, a mixed population or a pool of colonies from each animal and/or tissue was tested. Conclusion This paper describes a new method for fingerprinting Brucella isolates based on multi-locus characterization of a variable number, eight-base pair, tandem repeat. We have named this technique "HOOF-Prints" for Hypervariable Octameric Oligonucleotide Finger-Prints. The technique is highly discriminatory among Brucella species, among

  6. The Glutaminase-Dependent System Confers Extreme Acid Resistance to New Species and Atypical Strains ofBrucella.

    Science.gov (United States)

    Freddi, Luca; Damiano, Maria A; Chaloin, Laurent; Pennacchietti, Eugenia; Al Dahouk, Sascha; Köhler, Stephan; De Biase, Daniela; Occhialini, Alessandra

    2017-01-01

    Neutralophilic bacteria have developed specific mechanisms to cope with the acid stress encountered in environments such as soil, fermented foods, and host compartments. In Escherichia coli , the glutamate decarboxylase (Gad)-dependent system is extremely efficient: it requires the concerted action of glutamate decarboxylase (GadA/GadB) and of the glutamate (Glu)/γ-aminobutyrate antiporter, GadC. Notably, this system is operative also in new strains/species of Brucella , among which Brucella microti , but not in the "classical" species, with the exception of marine mammals strains. Recently, the glutaminase-dependent system (named AR2_Q), relying on the deamination of glutamine (Gln) into Glu and on GadC activity, was described in E. coli . In Brucella genomes, a putative glutaminase ( glsA )-coding gene is located downstream of the gadBC genes. We found that in B. microti these genes are expressed as a polycistronic transcript. Moreover, using a panel of Brucella genus-representative strains, we show that the AR2_Q system protects from extreme acid stress (pH ≤2.5), in the sole presence of Gln, only the Brucella species/strains predicted to have functional glsA and gadC . Indeed, mutagenesis approaches confirmed the involvement of glsA and gadC of B. microti in AR2_Q and that the acid-sensitive phenotype of B. abortus can be ascribed to a Ser248Leu substitution in GlsA, leading to loss of glutaminase activity. Furthermore, we found that the gene BMI_II339, of unknown function and downstream of the gadBC-glsA operon, positively affects Gad- and GlsA-dependent AR. Thus, we identified novel determinants that allow newly discovered and marine mammals Brucella strains to be better adapted to face hostile acidic environments. As for significance, this work may contribute to the understanding of the host preferences of Brucella species and opens the way to alternative diagnostic targets in epidemiological surveillance of brucellosis.

  7. The Glutaminase-Dependent System Confers Extreme Acid Resistance to New Species and Atypical Strains of Brucella

    Directory of Open Access Journals (Sweden)

    Luca Freddi

    2017-11-01

    Full Text Available Neutralophilic bacteria have developed specific mechanisms to cope with the acid stress encountered in environments such as soil, fermented foods, and host compartments. In Escherichia coli, the glutamate decarboxylase (Gad-dependent system is extremely efficient: it requires the concerted action of glutamate decarboxylase (GadA/GadB and of the glutamate (Glu/γ-aminobutyrate antiporter, GadC. Notably, this system is operative also in new strains/species of Brucella, among which Brucella microti, but not in the “classical” species, with the exception of marine mammals strains. Recently, the glutaminase-dependent system (named AR2_Q, relying on the deamination of glutamine (Gln into Glu and on GadC activity, was described in E. coli. In Brucella genomes, a putative glutaminase (glsA-coding gene is located downstream of the gadBC genes. We found that in B. microti these genes are expressed as a polycistronic transcript. Moreover, using a panel of Brucella genus-representative strains, we show that the AR2_Q system protects from extreme acid stress (pH ≤2.5, in the sole presence of Gln, only the Brucella species/strains predicted to have functional glsA and gadC. Indeed, mutagenesis approaches confirmed the involvement of glsA and gadC of B. microti in AR2_Q and that the acid-sensitive phenotype of B. abortus can be ascribed to a Ser248Leu substitution in GlsA, leading to loss of glutaminase activity. Furthermore, we found that the gene BMI_II339, of unknown function and downstream of the gadBC–glsA operon, positively affects Gad- and GlsA-dependent AR. Thus, we identified novel determinants that allow newly discovered and marine mammals Brucella strains to be better adapted to face hostile acidic environments. As for significance, this work may contribute to the understanding of the host preferences of Brucella species and opens the way to alternative diagnostic targets in epidemiological surveillance of brucellosis.

  8. Within-host evolution of Brucella canis during a canine brucellosis outbreak in a kennel.

    Science.gov (United States)

    Gyuranecz, Miklós; Rannals, Brandy D; Allen, Christina A; Jánosi, Szilárd; Keim, Paul S; Foster, Jeffrey T

    2013-04-12

    Little is currently known about Brucella evolution within the host during infection. The current study is the first to employ fine-scale genotyping on an isolate collection derived from a Brucella canis outbreak. Eight isolates of B. canis, cultured from different tissues of three dogs (female, stud dog, puppy of another female) from a single kennel over three months were genetically characterized with a 15-marker multi-locus, variable-number tandem repeat (VNTR) analysis (MLVA) to assess the genetic relatedness of isolates and potential rapid mutational changes. MLVA discriminated among the otherwise indistinguishable isolates from different animals and from isolates collected at different time points within each host, with different VNTR alleles being detected at multiple dates and tissue sites. We suspect that all isolates cultured from the female, puppy, and stud dogs originated from the same strain, with subsequent rapid in vivo mutations. However, high mutation rates and apparent in several of the loci prevented making definitive epidemiological relationships among isolates. This investigation highlights the rapid in vivo genetic mutations of several VNTRs of B. canis over a short time period in the host and the emergence of alternate alleles. However, this work also suggests the challenges of using highly mutable VNTRs to infer epidemiological relationships of strains within a short duration outbreak.

  9. Molecular Epidemiology of Brucella abortus in Northern Ireland-1991 to 2012.

    Directory of Open Access Journals (Sweden)

    Adrian Allen

    Full Text Available Brucellosis is the most common bacterial zoonoses worldwide. Bovine brucellosis caused by Brucella abortus has far reaching animal health and economic impacts at both the local and national levels. Alongside traditional veterinary epidemiology, the use of molecular typing has recently been applied to inform on bacterial population structure and identify epidemiologically-linked cases of infection. Multi-locus variable number tandem repeat VNTR analysis (MLVA was used to investigate the molecular epidemiology of a well-characterised Brucella abortus epidemic in Northern Ireland involving 387 herds between 1991 and 2012.MLVA identified 98 unique B. abortus genotypes from disclosing isolates in the 387 herds involved in the epidemic. Clustering algorithms revealed the relatedness of many of these genotypes. Combined with epidemiological information on chronology of infection and geographic location, these genotype data helped to identify 7 clonal complexes which underpinned the outbreak over the defined period. Hyper-variability of some VNTR loci both within herds and individual animals led to detection of multiple genotypes associated with single outbreaks. However with dense sampling, these genotypes could still be associated with specific clonal complexes thereby permitting inference of epidemiological links. MLVA- based epidemiological monitoring data were congruent with an independent classical veterinary epidemiology study carried out in the same territory.MLVA is a useful tool in ongoing disease surveillance of B. abortus outbreaks, especially when combined with accurate epidemiological information on disease tracings, geographical clustering of cases and chronology of infection.

  10. Brucella ceti from two striped dolphins stranded on the Apulia coastline, Italy.

    Science.gov (United States)

    Garofolo, Giuliano; Zilli, Katiuscia; Troiano, Pasquale; Petrella, Antonio; Marotta, Francesca; Di Serafino, Gabriella; Ancora, Massimo; Di Giannatale, Elisabetta

    2014-02-01

    Since 1994, when Brucella ceti was first isolated from an aborted dolphin fetus, several cases have been reported worldwide. The first case of B. ceti in the Mediterranean (and in Italy), however, was recorded only in 2012, off the coast of Tuscany. Extensive studies, using serological and microbiological methods, have documented this bacterium in dolphins and demonstrated its zoonotic potential. We describe the typing of two B. ceti strains isolated from striped dolphins (Stenella coeruleoalba) stranded on the southern Apulia coastline. B. ceti isolates were conventionally typed, and then genotyped by both the multilocus sequence typing (MLST) and the multilocus variable number of tandem repeats typing (MLVA) methodologies to infer phylogeny and potential epidemiological links between the two cases. The two isolates were identified through MLST analysis as belonging to the common sequence type 26 (ST26), while MLVA analysis, having established that the two isolates have identical profiles, assigned them to a novel genotype within cluster A - a unique representative of a new Mediterranean subcluster. The results thus revealed a link between the two cases studied, demonstrating the usefulness of MLST and MLVA for the epidemiological investigation of brucellae among marine mammals.

  11. Molecular Epidemiology of Brucella abortus in Northern Ireland-1991 to 2012.

    Science.gov (United States)

    Allen, Adrian; Breadon, Eleanor; Byrne, Andrew; Mallon, Thomas; Skuce, Robin; Groussaud, Pauline; Dainty, Amanda; Graham, Judith; Jones, Kerri; Pollock, Lorraine; Whatmore, Adrian

    2015-01-01

    Brucellosis is the most common bacterial zoonoses worldwide. Bovine brucellosis caused by Brucella abortus has far reaching animal health and economic impacts at both the local and national levels. Alongside traditional veterinary epidemiology, the use of molecular typing has recently been applied to inform on bacterial population structure and identify epidemiologically-linked cases of infection. Multi-locus variable number tandem repeat VNTR analysis (MLVA) was used to investigate the molecular epidemiology of a well-characterised Brucella abortus epidemic in Northern Ireland involving 387 herds between 1991 and 2012. MLVA identified 98 unique B. abortus genotypes from disclosing isolates in the 387 herds involved in the epidemic. Clustering algorithms revealed the relatedness of many of these genotypes. Combined with epidemiological information on chronology of infection and geographic location, these genotype data helped to identify 7 clonal complexes which underpinned the outbreak over the defined period. Hyper-variability of some VNTR loci both within herds and individual animals led to detection of multiple genotypes associated with single outbreaks. However with dense sampling, these genotypes could still be associated with specific clonal complexes thereby permitting inference of epidemiological links. MLVA- based epidemiological monitoring data were congruent with an independent classical veterinary epidemiology study carried out in the same territory. MLVA is a useful tool in ongoing disease surveillance of B. abortus outbreaks, especially when combined with accurate epidemiological information on disease tracings, geographical clustering of cases and chronology of infection.

  12. Molecular Epidemiology of Brucella abortus in Northern Ireland—1991 to 2012

    Science.gov (United States)

    Byrne, Andrew; Mallon, Thomas; Skuce, Robin; Groussaud, Pauline; Dainty, Amanda; Graham, Judith; Jones, Kerri; Pollock, Lorraine; Whatmore, Adrian

    2015-01-01

    Background Brucellosis is the most common bacterial zoonoses worldwide. Bovine brucellosis caused by Brucella abortus has far reaching animal health and economic impacts at both the local and national levels. Alongside traditional veterinary epidemiology, the use of molecular typing has recently been applied to inform on bacterial population structure and identify epidemiologically-linked cases of infection. Multi-locus variable number tandem repeat VNTR analysis (MLVA) was used to investigate the molecular epidemiology of a well-characterised Brucella abortus epidemic in Northern Ireland involving 387 herds between 1991 and 2012. Results MLVA identified 98 unique B. abortus genotypes from disclosing isolates in the 387 herds involved in the epidemic. Clustering algorithms revealed the relatedness of many of these genotypes. Combined with epidemiological information on chronology of infection and geographic location, these genotype data helped to identify 7 clonal complexes which underpinned the outbreak over the defined period. Hyper-variability of some VNTR loci both within herds and individual animals led to detection of multiple genotypes associated with single outbreaks. However with dense sampling, these genotypes could still be associated with specific clonal complexes thereby permitting inference of epidemiological links. MLVA- based epidemiological monitoring data were congruent with an independent classical veterinary epidemiology study carried out in the same territory. Conclusions MLVA is a useful tool in ongoing disease surveillance of B. abortus outbreaks, especially when combined with accurate epidemiological information on disease tracings, geographical clustering of cases and chronology of infection. PMID:26325586

  13. Antibodies to Brucella in marine mammals around the coast of England and Wales.

    Science.gov (United States)

    Jepson, P D; Brew, S; MacMillan, A P; Baker, J R; Barnett, J; Kirkwood, J K; Kuiken, T; Robinson, I R; Simpson, V R

    1997-11-15

    Following the isolation of previously unrecognised species of Brucella from stranded seals and cetaceans in Scotland and northern England, a serological survey was carried out to investigate the range of marine mammal species which may have been exposed to Brucella species around the coasts of England and Wales, the prevalence of infection and the temporal and geographical distribution of seropositive animals. Serum collected from 153 stranded marine mammals from the coasts of England and Wales between 1989 and 1995 were tested by competitive and indirect ELISA. Positive titres were recorded for six of 62 (10 per cent) grey seals (Halichoerus grypus), one of 12 (8 per cent) common seals (Phoca vitulina), 11 of 35 (31 per cent) harbour porpoises (Phocoena phocoena) and nine of 29 (31 per cent) common dolphins (Delphinus delphis) tested. Positive titres were also found in a striped dolphin (Stenella coeruleoalba), a bottlenose dolphin (Tursiops truncatus), a killer whale (Orcinus orca) and a pilot whale (Globicephala melas). The seropositive animals were from all around the coasts of England and Wales and the first seropositive sample was from a common dolphin in 1990.

  14. Abortion and premature birth in cattle following vaccination with Brucella abortus strain RB51.

    Science.gov (United States)

    Fluegel Dougherty, Amanda M; Cornish, Todd E; O'Toole, Donal; Boerger-Fields, Amy M; Henderson, Owen L; Mills, Ken W

    2013-09-01

    Brucella abortus RB51 is the vaccine strain currently licensed for immunizing cattle against brucellosis in the United States. Most cattle are vaccinated as heifer calves at 4-12 months of age. Adult cattle may be vaccinated in selected high-risk situations. Two herds of pregnant adult cattle in the brucellosis-endemic area of Wyoming were vaccinated with a standard label dose (1.0-3.4 × 10(10) organisms) of RB51. Reproductive losses in the vaccinated herds were 5.3% (herd A) and 0.6% (herd B) and included abortions, stillbirths, premature calves, and unbred cows (presumed early abortion). Brucella abortus was cultured from multiple tissues of aborted and premature calves (7/9), and from placenta. Isolates were identified as B. abortus strain RB51 by standard strain typing procedures and a species-specific polymerase chain reaction. Bronchopneumonia with intralesional bacteria and placentitis were observed microscopically. There was no evidence of involvement of other infectious or toxic causes of abortion. Producers, veterinarians, and laboratory staff should be alert to the risk of abortion when pregnant cattle are vaccinated with RB51, to potential human exposure, and to the importance of distinguishing field from vaccinal strains of B. abortus.

  15. Brucella abortus vaccine strain RB51 produces low levels of M-like O-antigen.

    Science.gov (United States)

    Cloeckaert, Axel; Zygmunt, Michel S; Guilloteau, Laurence A

    2002-03-15

    Brucella abortus RB51 is a rough (R) stable vaccine strain used in cattle and is believed to be devoid of O-side chain. We analyzed by use of a panel of monoclonal antibodies (MAbs) directed against seven previously defined O-polysaccharide (O-PS) epitopes the O-chain expression in strain RB51. Two MAbs specific for the C/Y (A=M) and C (M>A) epitopes showed low bindings in ELISA to strain RB51. O-chain expression was further confirmed by Western blot after SDS-PAGE of strain RB51. In particular, the MAb of C (M>A) specificity, showing preferential binding to M-dominant smooth (S) Brucella strains, revealed in strain RB51 a typical smooth-lipopolysaccharide (S-LPS) pattern which resembled that of M-dominant S-LPS. Thus, the results clearly show that strain RB51 produces low levels of M-like O-antigen.

  16. Brucella abortus RB51: enhancing vaccine efficacy and developing multivalent vaccines.

    Science.gov (United States)

    Vemulapalli, Ramesh; He, Yongqun; Sriranganathan, Nammalwar; Boyle, Stephen M; Schurig, Gerhardt G

    2002-12-20

    Brucella abortus vaccine strain RB51 is an attenuated, stable rough mutant that is being used in many countries to control bovine brucellosis. Our earlier study demonstrated that the protective efficacy of strain RB51 can be significantly enhanced by overexpressing Cu-Zn superoxide dismutase (SOD), a homologous protective antigen. We have also previously demonstrated that strain RB51 can be engineered to express heterologous proteins and mice vaccinated with such recombinant RB51 strains develop a strong Th1 type of immune response to the foreign proteins. The present study is aimed at combining these two characteristics to generate new recombinant RB51 vaccines with enhanced abilities to protect against brucellosis and simultaneously able to protect against infections by Mycobacterium spp. We constructed two recombinant RB51 strains, RB51SOD/85A which overexpresses SOD with simultaneous expression of the 85A, a protective protein of Mycobacterium spp., and RB51ESAT which expresses ESAT-6, another protective protein of M. bovis, as a fusion protein with the signal sequence and few additional amino terminal amino acids of SOD. Mice vaccinated with these recombinant strains developed specific immune responses to the mycobacterial proteins and significantly enhanced protection against Brucella challenge compared to the mice vaccinated with strain RB51 alone. Copyright 2002 Elsevier Science B.V.

  17. Identification at Biovar Level of Brucella Isolates Causing Abortion in Small Ruminants of Iran

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    Ali Mohammad Behroozikhah

    2012-01-01

    Full Text Available To determine the most prevalent biovar responsible for brucellosis in sheep and goat populations of Iran, a cross-sectional study was carried out over 2 years in six provinces selected based on geography and disease prevalence. Specimens obtained from referred aborted sheep and goat fetuses were cultured on Brucella selective media for microbiological isolation. Brucellae were isolated from 265 fetuses and examined for biovar identification using standard microbiological methods. Results showed that 246 isolates (92.8% were B. melitensis biovar 1, 18 isolates (6.8% were B. melitensis biovar 2, and, interestingly, one isolate (0.4% obtained from Mazandaran province was B. abortus biovar 3. In this study, B. melitensis biovar 3 was isolated in none of the selected provinces, and all isolates from 3 provinces (i.e., Chehar-mahal Bakhtiari, Markazi, and Ilam were identified only as B. melitensis biovar 1. In conclusion, we found that B. melitensis biovar 1 remains the most prevalent cause of small ruminant brucellosis in various provinces of Iran.

  18. Smooth to Rough Dissociation in Brucella: The Missing Link to Virulence.

    Science.gov (United States)

    Mancilla, Marcos

    2015-01-01

    Dissociation encompasses changes in a series of phenotypes: colony and cell morphology, inmunological and biochemical reactions and virulence. The concept is generally associated to the in vitro transition between smooth (S) and rough (R) colonies, a phenotypic observation in Gram-negative bacteria commonly made since the beginning of microbiology as a science. It is also well known that the loss of the O-polysaccharide, the most external lipopolysaccharide (LPS) moiety, triggers the change in the colony phenotype. Although dissociation is related to one of the most basic features used to distinguish between species, i.e., colony morphology, and, in the case of pathogens, predict their virulence behavior, it has been considered a laboratory artifact and thus did not gain further attention. However, recent insights into genetics and pathogenesis of members of Brucella, causative agents of brucellosis, have brought a new outlook on this experimental fact, suggesting that it plays a role beyond the laboratory observations. In this perspective article, the current knowledge on Brucella LPS genetics and its connection with dissociation in the frame of evolution is discussed. Latest reports support the notion that, by means of a better understanding of genetic pathways linked to R phenotype and the biological impact of this intriguing "old" phenomenon, unexpected applications can be achieved.

  19. Smooth to rough dissociation in Brucella: the missing link to virulence

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    Marcos eMancilla

    2016-01-01

    Full Text Available Dissociation encompasses changes in a series of phenotypes: colony and cell morphology, inmunological and biochemical reactions and virulence. The concept is generally associated to the in vitro transition between smooth (S and rough (R colonies, a phenotypic observation in Gram-negative bacteria commonly made since the beginning of microbiology as a science. It is also well known that the loss of the O-polysaccharide, the most external lipopolysaccharide (LPS moiety, triggers the change in the colony phenotype. Although dissociation is related to one of the most basic features used to distinguish between species, i.e. colony morphology, and, in the case of pathogens, predict their virulence behaviour, it has been considered a laboratory artifact and thus did not gain further attention. However, recent insights into genetics and pathogenesis of members of Brucella, causative agents of brucellosis, have brought a new outlook on this experimental fact, suggesting that it plays a role beyond the laboratory observations.In this perspective article, the current knowledge on Brucella LPS genetics and its connection with dissociation in the frame of evolution is discussed. Latest reports support the notion that, by means of a better understanding of genetic pathways linked to R phenotype and the biological impact of this intriguing old phenomenon, unexpected applications can be achieved.

  20. Seroprevalence of brucellosis and typing of Brucella melitensis biovar 2 in lactating cows in Kuwait

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    Adel El-Gohary

    2016-09-01

    Results: The results showed that the overall seroprevalence of bovine brucellosis was 339 (7.25% by BAPAT, 332 (7.1% by RBPT, and 329 (7.04% by CFT. The results revealed that, 42 (8.6%, 5 (1.4% and 292 (7.6% sera were positive for brucellosis by BAPAT in the cows of Al-Wafra, Al-Kabed and Al-Salebia areas, respectively. Whereas, their respective number and seroreactive cases by RBPT were 39 (8.02%, 5 (1.4% and 288 (7.4%. Similarly, as confirmatory test by CFT, the number and seroreactive cases in these areas were 39 (8.02%, 5 (1.4% and 285 (7.46%. MRT revealed that the average positive case was 61.67% (59.46% in Al-Wafra; 60% in Al-Kabed and 66.6% in Al-Salebia. Two Brucella isolates could be recovered from the stomach content of the two aborted feti and typed as Brucella melitensis biovar 2. Conclusion: Brucellosis is prevalent among lactating cows in Kuwait. This indicates the potential role of these dairy animals in disseminating and spread of such zoonosis to human. Considering public health significance, appropriate preventive measures are suggestive for combating brucellosis in Kuwait. [J Adv Vet Anim Res 2016; 3(3.000: 229-235

  1. Detection of Antibodies to Brucella Cytoplasmic Proteins in the Cerebrospinal Fluid of Patients with Neurobrucellosis

    Science.gov (United States)

    Baldi, Pablo C.; Araj, George F.; Racaro, Graciela C.; Wallach, Jorge C.; Fossati, Carlos A.

    1999-01-01

    The diagnosis of human neurobrucellosis usually relies on the detection of antibodies to Brucella lipopolysaccharide (LPS) in cerebrospinal fluid (CSF) by agglutination tests or enzyme-linked immunosorbent assay (ELISA). Here we describe the detection of immunoglobulin G (IgG) to cytoplasmic proteins (CP) of Brucella spp. by ELISA and Western blotting in seven CSF samples from five patients with neurobrucellosis. While IgG to CP (titers of 200 to 12,800) and IgG to LPS (800 to 6,400) were found in the CSF of these patients, these antibodies were not detected in CSF samples from two patients who had systemic brucellosis without neurological involvement. The latter, however, had serum IgG and IgM to both LPS and CP. No reactivity to these antigens was found in CSF samples from 14 and 20 patients suffering from nonbrucellar meningitis and noninfectious diseases, respectively. These findings suggest that, in addition to its usefulness in the serological diagnosis of human systemic brucellosis, the ELISA with CP antigen can be used for the specific diagnosis of human neurobrucellosis. PMID:10473531

  2. First evidence of Brucella ovis infection in rams in the Pirot Municipality, Serbia

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    Miloš Petrović

    2014-12-01

    Full Text Available 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.

  3. First evidence of Brucella ovis infection in rams in the Pirot Municipality, Serbia.

    Science.gov (United States)

    Petrović, Miloš; Špičić, Silvio; Potkonjak, Aleksandar; Lako, Branislav; Kostov, Miloš; Cvetnić, Željko

    2014-12-29

    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. Onesided 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.

  4. Neutrophils exert a suppressive effect on Th1 responses to intracellular pathogen Brucella abortus.

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    Elías Barquero-Calvo

    2013-02-01

    Full Text Available Polymorphonuclear neutrophils (PMNs are the first line of defense against microbial pathogens. In addition to their role in innate immunity, PMNs may also regulate events related to adaptive immunity. To investigate the influence of PMNs in the immune response during chronic bacterial infections, we explored the course of brucellosis in antibody PMN-depleted C57BL/6 mice and in neutropenic mutant Genista mouse model. We demonstrate that at later times of infection, Brucella abortus is killed more efficiently in the absence of PMNs than in their presence. The higher bacterial removal was concomitant to the: i comparatively reduced spleen swelling; ii augmented infiltration of epithelioid histiocytes corresponding to macrophages/dendritic cells (DCs; iii higher recruitment of monocytes and monocyte/DCs phenotype; iv significant activation of B and T lymphocytes, and v increased levels of INF-γ and negligible levels of IL4 indicating a balance of Th1 over Th2 response. These results reveal that PMNs have an unexpected influence in dampening the immune response against intracellular Brucella infection and strengthen the notion that PMNs actively participate in regulatory circuits shaping both innate and adaptive immunity.

  5. Microscopy-based Assays for High-throughput Screening of Host Factors Involved in Brucella Infection of Hela Cells.

    Science.gov (United States)

    Casanova, Alain; Low, Shyan H; Emmenlauer, Mario; Conde-Alvarez, Raquel; Salcedo, Suzana P; Gorvel, Jean-Pierre; Dehio, Christoph

    2016-08-05

    Brucella species are facultative intracellular pathogens that infect animals as their natural hosts. Transmission to humans is most commonly caused by direct contact with infected animals or by ingestion of contaminated food and can lead to severe chronic infections. Brucella can invade professional and non-professional phagocytic cells and replicates within endoplasmic reticulum (ER)-derived vacuoles. The host factors required for Brucella entry into host cells, avoidance of lysosomal degradation, and replication in the ER-like compartment remain largely unknown. Here we describe two assays to identify host factors involved in Brucella entry and replication in HeLa cells. The protocols describe the use of RNA interference, while alternative screening methods could be applied. The assays are based on the detection of fluorescently labeled bacteria in fluorescently labeled host cells using automated wide-field microscopy. The fluorescent images are analyzed using a standardized image analysis pipeline in CellProfiler which allows single cell-based infection scoring. In the endpoint assay, intracellular replication is measured two days after infection. This allows bacteria to traffic to their replicative niche where proliferation is initiated around 12 hr after bacterial entry. Brucella which have successfully established an intracellular niche will thus have strongly proliferated inside host cells. Since intracellular bacteria will greatly outnumber individual extracellular or intracellular non-replicative bacteria, a strain constitutively expressing GFP can be used. The strong GFP signal is then used to identify infected cells. In contrast, for the entry assay it is essential to differentiate between intracellular and extracellular bacteria. Here, a strain encoding for a tetracycline-inducible GFP is used. Induction of GFP with simultaneous inactivation of extracellular bacteria by gentamicin enables the differentiation between intracellular and extracellular

  6. Proteomic Analysis of Detergent Resistant Membrane Domains during Early Interaction of Macrophages with Rough and Smooth Brucella melitensis

    Science.gov (United States)

    Lauer, Sabine A.; Iyer, Srinivas; Sanchez, Timothy; Forst, Christian V.; Bowden, Brent; Carlson, Kay; Sriranganathan, Nammalwar; Boyle, Stephen M.

    2014-01-01

    The plasma membrane contains discrete nanometer-sized domains that are resistant to non-ionic detergents, and which are called detergent resistant membrane domains (DRMDs) or lipid rafts. Exposure of host cells to pathogenic bacteria has been shown to induce the re-distribution of specific host proteins between DRMDs and detergent soluble membranes, which leads to the initiation of cell signaling that enable pathogens to access host cells. DRMDs have been shown to play a role in the invasion of Brucella into host macrophages and the formation of replicative phagosomes called Brucella-containing vacuoles (BCVs). In this study we sought to characterize changes to the protein expression profiles in DRMDs and to respective cellular pathways and networks of Mono Mac 6 cells in response to the adherence of rough VTRM1 and smooth 16 M B. melitensis strains. DRMDs were extracted from Mono Mac 6 cells exposed for 2 minutes at 4°C to Brucella (no infection occurs) and from unexposed control cells. Protein expression was determined using the non-gel based quantitative iTRAQ (Isobaric Tags for Relative and Absolute Quantitation) mass spectrometry technique. Using the identified iTRAQ proteins we performed enrichment analyses and probed constructed human biochemical networks for interactions and metabolic reactions. We identified 149 proteins, which either became enriched, depleted or whose amounts did not change in DRMDs upon Brucella exposure. Several of these proteins were distinctly enriched or depleted in DRMDs upon exposure to rough and smooth B. melitensis strains which results in the differential engagement of cellular pathways and networks immediately upon Brucella encounter. For some of the proteins such as myosin 9, small G protein signaling modulator 3, lysine-specific demethylase 5D, erlin-2, and voltage-dependent anion-selective channel protein 2, we observed extreme differential depletion or enrichment in DRMDs. The identified proteins and pathways could provide

  7. Temporal Role for MyD88 in a Model of Brucella-Induced Arthritis and Musculoskeletal Inflammation.

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    Lacey, Carolyn A; Mitchell, William J; Brown, Charles R; Skyberg, Jerod A

    2017-03-01

    Brucella spp. are facultative intracellular Gram-negative bacteria that cause the zoonotic disease brucellosis, one of the most common global zoonoses. Osteomyelitis, arthritis, and musculoskeletal inflammation are common focal complications of brucellosis in humans; however, wild-type (WT) mice infected systemically with conventional doses of Brucella do not develop these complications. Here we report C57BL/6 WT mice infected via the footpad with 10 3 to 10 6 CFU of Brucella spp. display neutrophil and monocyte infiltration of the joint space and surrounding musculoskeletal tissue. Joint inflammation is detectable as early as 1 day postinfection and peaks 1 to 2 weeks later, after which WT mice are able to slowly resolve inflammation. B and T cells were dispensable for the onset of swelling but required for resolution of joint inflammation and infection. At early time points, MyD88 -/- mice display decreased joint inflammation, swelling, and proinflammatory cytokine levels relative to WT mice. Subsequently, swelling of MyD88 -/- joints surpassed WT joint swelling, and resolution of joint inflammation was prolonged. Joint bacterial loads in MyD88 -/- mice were significantly greater than those in WT mice by day 3 postinfection and at all time points thereafter. In addition, MyD88 -/- joint inflammatory cytokine levels on day 3 and beyond were similar to WT levels. Collectively these data demonstrate MyD88 signaling mediates early inflammatory responses in the joint but also contributes to subsequent clearance of Brucella and resolution of inflammation. This work also establishes a mouse model for studying Brucella -induced arthritis, musculoskeletal complications, and systemic responses, which will lead to a better understanding of focal complications of brucellosis. Copyright © 2017 American Society for Microbiology.

  8. Serosurvey of Brucella spp. infection in the Kafue lechwe (Kobus leche kafuensis) of the Kafue flats in Zambia.

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    Muma, John Bwalya; Lund, Arve; Siamudaala, Victor M; Munang'andu, Hetron Mweemba; Munyeme, Musso; Matope, Gift; Nielsen, Klaus; DJønne, Berit; Godfroid, Jacques; Tryland, Morten; Skjerve, Eystein

    2010-10-01

    One of the diseases of veterinary and public health importance affecting the Kafue lechwe (Kobus leche kafuensis) on the Kafue flats is brucellosis, for which only scant information is available. During the 2003 (October), 2004 (December), and 2008 (July-December) hunting seasons in the Kafue flats, we conducted a study to determine the seroprevalence of Brucella spp. in the Kafue lechwe and to evaluate serologic tests for detection of Brucella spp. antibodies in lechwe. The Rose Bengal Test (RBT), competitive enzyme-linked immunosorbent assay (cELISA), and fluorescence polarization assay (FPA) were used. A total of 121 Kafue lechwe were hunted for disease investigations in 2003, 2004, and 2008 in the Kafue Flat Game Management Area. Of these, 21.6%, (95% confidence interval [CI]: 14.2-29.1%) had detectable antibodies to Brucella spp. The Kafue lechwe in Lochnivar National Park had higher antibody results than those in Blue Lagoon National Park (odds ratio=3.0; 95% CI: 0.94-9.4). Infection levels were similar in females (21.6%) and males (21.7%). Results were similar among RBT, FPA, cELISA tests, suggesting that these could effectively be used in diagnosing brucellosis in the Kafue lechwe. Our study demonstrates the presence of Brucella infections in the Kafue lechwe in two national parks located in the Kafue flats and further highlights the suitability of serologic assays for testing the Kafue lechwe. Because the Kafue lechwe is the most hunted wildlife species in Zambia, hunters need to be informed of the public health risk of Brucella spp. infection.

  9. Circulating strains of Brucella abortus in cattle in the province of Santo Domingo de los Tsáchilas - Ecuador.

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    Richar Ivan Rodríguez Hidalgo

    2015-03-01

    Full Text Available In Ecuador, the Province of Santo Domingo de los Tsáchilas represents the largest informal market of livestock due to its strategic position in the country; thus, given the high mobility of cattle in this region, the aim of this study was to determine the strain variation of Brucella sp.. Part of the study was the isolation, biotyping and genotyping of Brucella species isolated from milk and supra-mammary lymph nodes of sero-positive bovines. Protocols used for isolation, biotyping and genotyping of Brucella species were selective Farrell medium, biochemical assays and IS711-PCR, AMOS-PCR and HOOF-Prints techniques, respectively. In total, 656 animals from sero-positive dairy herds and from the slaughterhouse of the province were diagnosed by Rose Bengal and Wright’s Slow Agglutination test with EDTA. From these animals, 50 animals were found sero-positive for brucellosis. Twenty-five lymph nodes and 25 milk samples from each group of positive reactors were cultured and growth. Isolation was possible in 4 (16% and 9 (36%, respectively; and of these, 10 isolates were diagnosed as Brucella sp. All 4 isolates of lymphatic tissue corresponded to Brucella abortus biotype 1, confirmed as field strains by molecular analysis. Milk isolations, biochemically showed a more dispersed pattern in which B. abortus biotypes 1 and 4 were found; yet four samples gave a pattern similar to B. abortus biotype 2; however, only biotypes 1 and 4 were confirmed by molecular analysis. The concordance between biochemical and molecular diagnostic tests reached 76.9%.

  10. Cloning and expression of Brucella outer membrane protein 36kDa (OMP2b in E. coli

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    Nazanin Behshti

    2011-09-01

    Full Text Available Background & Objective: Brucellosis is an important zoonotic disease of economic significance. Brucella species are gram-negative, facultative intracellular bacteria, and are capable of replicating in the phagosomes of macrophages. They cause infection in several animal species and humans. Prevention of new diseases and diagnosis of cases infected with the organism are both essential for eradication of the disease. Characterization and evaluation of different antigens of Brucella cells has a key role in progression of prevention and diagnosis programs. Here, we report the production and purification of recombinant 31kDa outer membrane protein Brucella abortus (Omp2b. Materials & Methods: Brucella abortus 36kDa outer membrane protein gene was amplified with PrimeSTAR® HS DNA polymerase, cloned in pJET1.2. The target gene was subcloned in pET28a (+. Recombinant pET28a vectors were transformed into E coli BL21 (DE3. Expression of recombinant protein was induced with 1mM IPTG. Proteins were absorbed to Ni-NTA agarose resins and Recombinant proteins were eluted with 250mM imidazol. Imidazol removed by dialysis. Proteins were assayed by Western-blotting and rOmp2b was probed by Brucella rabbit anti serum. Result: Appearance of a golden brown band at the site of reaction, in Western blotting confirmed successfully clone and expression. We purified Omp2b by affinity chromatography and this method prepared refolds proteins on the column. Conclusion: Omp2b were successfully cloned, expressed and purified. The recombinant proteins were recognized by polyclonal antiserum which suggests the accuracy of procedure.

  11. Entry and elimination of marine mammal Brucella spp. by hooded seal (Cystophora cristata alveolar macrophages in vitro.

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    Anett K Larsen

    Full Text Available A high prevalence of Brucellapinnipedialis serology and bacteriology positive animals has been found in the Northeast Atlantic stock of hooded seal (Cystophoracristata; however no associated gross pathological changes have been identified. Marine mammal brucellae have previously displayed different infection patterns in human and murine macrophages. To investigate if marine mammal Brucella spp. are able to invade and multiply in cells originating from a presumed host species, we infected alveolar macrophages from hooded seal with a B. pinnipedialis hooded seal isolate. Hooded seal alveolar macrophages were also challenged with B. pinnipedialis reference strain (NCTC 12890 from harbor seal (Phocavitulina, B. ceti reference strain (NCTC 12891 from harbor porpoise (Phocoenaphocoena and a B. ceti Atlantic white-sided dolphin (Lagenorhynchusacutus isolate (M83/07/1, to evaluate possible species-specific differences. Brucella suis 1330 was included as a positive control. Alveolar macrophages were obtained by post mortem bronchoalveolar lavage of euthanized hooded seals. Phenotyping of cells in the lavage fluid was executed by flow cytometry using the surface markers CD14 and CD18. Cultured lavage cells were identified as alveolar macrophages based on morphology, expression of surface markers and phagocytic ability. Alveolar macrophages were challenged with Brucella spp. in a gentamicin protection assay. Following infection, cell lysates from different time points were plated and evaluated quantitatively for colony forming units. Intracellular presence of B. pinnipedialis hooded seal isolate was verified by immunocytochemistry. Our results show that the marine mammal brucellae were able to enter hooded seal alveolar macrophages; however, they did not multiply intracellularly and were eliminated within 48 hours, to the contrary of B. suis that showed the classical pattern of a pathogenic strain. In conclusion, none of the four marine mammal strains

  12. Microarray-based identification of differentially expressed genes in intracellular Brucella abortus within RAW264.7 cells.

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    Mingxing Tian

    Full Text Available Brucella spp. is a species of facultative intracellular Gram-negative bacteria that induces abortion and causes sterility in domesticated mammals and chronic undulant fever in humans. Important determinants of Brucella's virulence and potential for chronic infection include the ability to circumvent the host cell's internal surveillance system and the capability to proliferate within dedicated and non-dedicated phagocytes. Hence, identifying genes necessary for intracellular survival may hold the key to understanding Brucella infection. In the present study, microarray analysis reveals that 7.82% (244/3334 of all Brucella abortus genes were up-regulated and 5.4% (180/3334 were down-regulated in RAW264.7 cells, compared to free-living cells in TSB. qRT-PCR verification further confirmed a >5-fold up-regulation for fourteen genes. Functional analysis classified araC, ddp, and eryD as to partake in information storage and processing, alp, flgF and virB9 to be involved in cellular processes, hpcd and aldh to play a role in metabolism, mfs and nikC to be involved in both cellular processes and metabolism, and four hypothetical genes (bruAb1_1814, bruAb1_0475, bruAb1_1926, and bruAb1_0292 had unknown functions. Furthermore, we constructed a B. abortus 2308 mutant Δddp where the ddp gene is deleted in order to evaluate the role of ddp in intracellular survival. Infection assay indicated significantly higher adherence and invasion abilities of the Δddp mutant, however it does not survive well in RAW264.7 cells. Brucella may survive in hostile intracellular environment by modulating gene expression.

  13. Seroprevalence of Brucella spp. in Cattle, Molecular Characterization in Milk, and the Analysis of Associated Risk Factors with Seroprevalence in Humans, Egypt.

    Science.gov (United States)

    El-Diasty, Mohamed M; Ahmed, Heba A; Sayour, Ashraf E; El Hofy, Fatma I; Tahoun, Asmaa B M B; Shafik, Saleh M

    2016-12-01

    The objective of the present study was to estimate the seroprevalence of Brucella spp. in humans and cattle at Sharkia Governorate, Egypt. In addition, identification of Brucella spp. in milk samples by PCR and culture with the evaluation of the risk factors associated with Brucella spp. seroprevalence in humans were carried out. Overall, the seroprevalence of Brucella antibodies in the examined cattle was 23.8%, while in human participants it was 21%. The examination of 205 milk samples using PCR revealed that 6.3% were positive for B. abortus biovar 1 and the results were confirmed by culture methods. Multivariate logistic regression revealed that consumption of unpasteurized dairy products, occupational contact with animals, and knowledge about the disease are risk factors associated with infection in humans. This study documented the endemic status of brucellosis in Egypt. Hygienic measures and increased awareness about the disease are recommended to minimize the spread of infection from animals to humans.

  14. Computational prediction of secretion systems and secretomes of Brucella: identification of novel type IV effectors and their interaction with the host.

    Science.gov (United States)

    Sankarasubramanian, Jagadesan; Vishnu, Udayakumar S; Dinakaran, Vasudevan; Sridhar, Jayavel; Gunasekaran, Paramasamy; Rajendhran, Jeyaprakash

    2016-01-01

    Brucella spp. are facultative intracellular pathogens that cause brucellosis in various mammals including humans. Brucella survive inside the host cells by forming vacuoles and subverting host defence systems. This study was aimed to predict the secretion systems and the secretomes of Brucella spp. from 39 complete genome sequences available in the databases. Furthermore, an attempt was made to identify the type IV secretion effectors and their interactions with host proteins. We predicted the secretion systems of Brucella by the KEGG pathway and SecReT4. Brucella secretomes and type IV effectors (T4SEs) were predicted through genome-wide screening using JVirGel and S4TE, respectively. Protein-protein interactions of Brucella T4SEs with their hosts were analyzed by HPIDB 2.0. Genes coding for Sec and Tat pathways of secretion and type I (T1SS), type IV (T4SS) and type V (T5SS) secretion systems were identified and they are conserved in all the species of Brucella. In addition to the well-known VirB operon coding for the type IV secretion system (T4SS), we have identified the presence of additional genes showing homology with T4SS of other organisms. On the whole, 10.26 to 14.94% of total proteomes were found to be either secreted (secretome) or membrane associated (membrane proteome). Approximately, 1.7 to 3.0% of total proteomes were identified as type IV secretion effectors (T4SEs). Prediction of protein-protein interactions showed 29 and 36 host-pathogen specific interactions between Bos taurus (cattle)-B. abortus and Ovis aries (sheep)-B. melitensis, respectively. Functional characterization of the predicted T4SEs and their interactions with their respective hosts may reveal the secrets of host specificity of Brucella.

  15. Bacteriemia por Brucella canis: Aislamiento con el Sistema Bact-Alert Bacteriemia by Brucella canis: Isolation with the Bact-Alert System

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    R. Soloaga

    2004-06-01

    Full Text Available B.canis puede causar enfermedad en humanos, pero esto es poco frecuente aún en países donde la infección es común en perros. Se presenta el caso de un paciente de 15 años de edad que ingresa por fiebre de quince días de evolución sin foco aparente. El exámen físico mostró dolor hepático a la palpación y adenomegalias axilares, cervicales e inguinales. La ecografía abdominal evidenció esplenomegalia, la radiografía de tóraxy el ecocardiograma transtorácico no presentaron anormalidades. Se tomaron 5 hemocultivos (sistema Bact-Alert, Biomerieux, Marcy, l’Etolie, Francia los cuales resultaron positivos; los mismos correspondieron a 2 botellas estándares (tiempo de positivización de 72 y 64,8 horas y a 3 pediátricas FAN (tiempo de positivización de 74,5; 72 y 67,2 hs. El microorganismo fue identificado en forma presuntiva por pruebas bioquímicascomo B. canis y la identificación fue confirmada en el Instituto Malbrán. Luego de 14 días de tratamiento con ceftriaxona el paciente evolucionó afebril;al tener documentación bacteriológica de B.canis, se realizó tratamiento ambulatorio durante 21 días con doxiciclina (100 mg cada 12 hs y rifampicina (600 mg/día durante 6 semanas. Se descartó compromiso óseo y la evolución fue favorable desapareciendo todos los síntomas.Brucella canis and other species of the genus Brucella can cause human disease. However, this species infrequently cause human disease, including in countries where dogs population is highly infected. A 15 years old male was admitted to the hospital with 15 days history of fever without visible focus. Physical examination revealed pain at liver palpation and axillar, cervical and inguinal lymphoadenomegalies. Abdominal ultrasonography showed spleenomegally, the chest Rx and the trans thoracic echocardiogram were normal. Five blood samples were obtained and cultured in 2 standards bottles (time of positivization 72 - 64,8 hours, and 3 pediatric FAN bottles

  16. [Comparison of effectiveness between two surgical methods in treatment of thoracolumbar brucella spondylitis].

    Science.gov (United States)

    Yang, Xinming; Zuo, Xianhong; Jia, Yongli; Chang, Yuefei; Zhang, Peng; Ren, Yixing

    2014-10-01

    To compare the effectiveness between the method of simple posterior debridement combined with bone grafting and fusion and internal fixation and the method of one-stage anterior radical debridement combined with bone grafting and fusion and posterior internal fixation in the treatment of thoracolumbar brucella spondylitis so as to provide the reference for the clinical treatment. A retrospective analysis was made on the clinical data of 148 cases of thoracolumbar brucella spondylitis between January 2002 and January 2012. Simple posterior debridement combined with bone grafting and fusion and internal fixation was used in 78 cases (group A), and one-stage anterior radical debridement combined with bone grafting and fusion and posterior internal fixation in 70 cases (group B). There was no significant difference in gender, age, disease duration, involved vertebral segments, erythrocyte sedimentation rate (ESR), visual analogue scale (VAS) score, neural function grade of America Spinal Injury Association (ASIA), and kyphosis Cobb angle before operation between 2 groups (P > 0.05). The peri operation period indexes (hospitalization time, operation time, and intraoperative blood loss) and the clinical effectiveness indexes (VAS score, ASIA grade, Cobb angle, and ESR) were compared; the bone fusion and the internal fixation were observed. Incision infection and paravertebral and/or psoas abscess occurred in 2 and 3 cases of group A respectively. All incisions healed by first intention and 2 cases had pneumothorax in group B. The operation time and the hospitalization time of group A were significantly shorter than those of group B (P average. The VAS, ESR, and Cobb angle were significantly decreased at each time point after operation when compared with preoperative ones in 2 groups (P 0.05). The neurological function was significantly improved at 3 months after operation; there were 1 case of ASIA grade C, 14 cases of grade D, and 63 cases of grade E in group A, and

  17. Systems Biology Analysis of Temporal In vivo Brucella melitensis and Bovine Transcriptomes Predicts host:Pathogen Protein–Protein Interactions

    Science.gov (United States)

    Rossetti, Carlos A.; Drake, Kenneth L.; Lawhon, Sara D.; Nunes, Jairo S.; Gull, Tamara; Khare, Sangeeta; Adams, Leslie G.

    2017-01-01

    To date, fewer than 200 gene-products have been identified as Brucella virulence factors, and most were characterized individually without considering how they are temporally and coordinately expressed or secreted during the infection process. Here, we describe and analyze the in vivo temporal transcriptional profile of Brucella melitensis during the initial 4 h interaction with cattle. Pathway analysis revealed an activation of the “Two component system” providing evidence that the in vivo Brucella sense and actively regulate their metabolism through the transition to an intracellular lifestyle. Contrarily, other Brucella pathways involved in virulence such as “ABC transporters” and “T4SS system” were repressed suggesting a silencing strategy to avoid stimulation of the host innate immune response very early in the infection process. Also, three flagellum-encoded loci (BMEII0150-0168, BMEII1080-1089, and BMEII1105-1114), the “flagellar assembly” pathway and the cell components “bacterial-type flagellum hook” and “bacterial-type flagellum” were repressed in the tissue-associated B. melitensis, while RopE1 sigma factor, a flagellar repressor, was activated throughout the experiment. These results support the idea that Brucella employ a stealthy strategy at the onset of the infection of susceptible hosts. Further, through systems-level in silico host:pathogen protein–protein interactions simulation and correlation of pathogen gene expression with the host gene perturbations, we identified unanticipated interactions such as VirB11::MAPK8IP1; BtaE::NFKBIA, and 22 kDa OMP precursor::BAD and MAP2K3. These findings are suggestive of new virulence factors and mechanisms responsible for Brucella evasion of the host's protective immune response and the capability to maintain a dormant state. The predicted protein–protein interactions and the points of disruption provide novel insights that will stimulate advanced hypothesis-driven approaches

  18. Prevalencia de anticuerpos séricos contra la bacteria Brucella sp. en una población humana tropical

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    Grettel Chanto

    2007-06-01

    Full Text Available La brucelosis es una enfermedad bacteriana generalizada con tendencia a la cronicidad si no se trata adecuadamente. El diagnóstico de laboratorio se realiza por el aislamiento e identificación del agente o en la mayoría de los casos mediante pruebas serológicas. En este estudio se informa la prevalencia de anticuerpos séricos contra Brucella sp. en habitantes de comunidades rurales de Cartago dedicadas principalmente a la ganadería de leche. Se analizó un total de 714 muestras representativas de la población mediante la técnica de aglutinación Rosa de Bengala en lámina (RBT y la prueba de aglutinación en microplaca (SAT. Se encontró una prevalencia de 0.87 %, y en ninguna persona seropositiva se demostró seroconversión. Aún, cuando el 71 % de la población refirió consumir leche no pasteurizada, no se encontró asociación estadísticamente significativa (1.09 %, IC 0.30-2.77 entre el consumo de leche no pasteurizada y la presencia de anticuerpos sugestivos de infección por Brucella sp. A pesar de ser una zona de alta prevalencia de brucelosis bovina por Brucella abortus, la ausencia de síntomas en la población y los bajos títulos de anticuerpos observados sugieren una escasa exposición previa al agente.Prevalence of serum antibodies for the bacterium Brucella sp. in a tropical human population. Brucellosis is a systemic bacterial disease that can last years if not treated properly. The laboratory diagnosis is made by serological tests or isolation of the agent. We studied the prevalence of antibodies against Brucella sp. in inhabitants of rural communities of Cartago, Costa Rica, who are dedicated mainly to dairy cattle activities. We analyzed 714 representative samples with the Rose Bengal plate agglutination test (RBT and the standard agglutination test in microplate (SAT. The overall prevalence was 0.87 %. No seroconversion was demonstrated for seropositive individuals. Even though 71 % of the population mentioned

  19. Systems Biology Analysis of Temporal In vivo Brucella melitensis and Bovine Transcriptomes Predicts host:Pathogen Protein–Protein Interactions

    Directory of Open Access Journals (Sweden)

    Carlos A. Rossetti

    2017-07-01

    Full Text Available To date, fewer than 200 gene-products have been identified as Brucella virulence factors, and most were characterized individually without considering how they are temporally and coordinately expressed or secreted during the infection process. Here, we describe and analyze the in vivo temporal transcriptional profile of Brucella melitensis during the initial 4 h interaction with cattle. Pathway analysis revealed an activation of the “Two component system” providing evidence that the in vivo Brucella sense and actively regulate their metabolism through the transition to an intracellular lifestyle. Contrarily, other Brucella pathways involved in virulence such as “ABC transporters” and “T4SS system” were repressed suggesting a silencing strategy to avoid stimulation of the host innate immune response very early in the infection process. Also, three flagellum-encoded loci (BMEII0150-0168, BMEII1080-1089, and BMEII1105-1114, the “flagellar assembly” pathway and the cell components “bacterial-type flagellum hook” and “bacterial-type flagellum” were repressed in the tissue-associated B. melitensis, while RopE1 sigma factor, a flagellar repressor, was activated throughout the experiment. These results support the idea that Brucella employ a stealthy strategy at the onset of the infection of susceptible hosts. Further, through systems-level in silico host:pathogen protein–protein interactions simulation and correlation of pathogen gene expression with the host gene perturbations, we identified unanticipated interactions such as VirB11::MAPK8IP1; BtaE::NFKBIA, and 22 kDa OMP precursor::BAD and MAP2K3. These findings are suggestive of new virulence factors and mechanisms responsible for Brucella evasion of the host's protective immune response and the capability to maintain a dormant state. The predicted protein–protein interactions and the points of disruption provide novel insights that will stimulate advanced hypothesis

  20. The prevalence and distribution of Brucella melitensis in goats in Malaysia from 2000 to 2009.

    Science.gov (United States)

    Bamaiyi, P H; Hassan, L; Khairani-Bejo, S; ZainalAbidin, M; Ramlan, M; Adzhar, A; Abdullah, N; Hamidah, N H M; Norsuhanna, M M; Hashim, S N

    2015-05-01

    A study was conducted to describe the prevalence and distribution of zoonotic Brucella melitensis in goats in Peninsular Malaysia. Using serosurveillance data of the last decade (2000-2009) involving 119,799 goats and 3555 farms, the seroprevalence of brucellosis among goats was 0.91% (95% CI=0.86-0.96) and among farms was 7.09% (95% CI=6.27-7.98). The odds of brucellosis was significantly (PMalaysia. The infection was detected throughout Malaysia but at generally low seroprevalences with states like Perlis that border neighbouring countries having higher seroprevalence of brucellosis than other non-border states. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. Milk Ring Test for spot identification of Brucella abortus infection in single cow herds

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    Najibullah Mohamand

    2014-06-01

    Full Text Available In this study, milk samples were collected from 109 dairy cows to detect antibodies against Brucella (B. using Milk Ring Test (MRT. Overall, 18.35% (n=20/109 of the milk samples were positive by MRT. The cows were divided into three groups based on lactation number viz., 1st, 2nd to 4th and ≥5th lactations; the prevalence of brucellosis in the groups were found to be 0.92% (n=1/109, 15.60% (n=17/109 and 1.83% (n=2/109, respectively. Considering simplicity and cost effectiveness, the MRT can be used for the preliminary screening of B. abortus infection especially in single cow herds.

  2. Biotyping and genotyping (MLVA16 of Brucella abortus isolated from cattle in Brazil, 1977 to 2008.

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    Sílvia Minharro

    Full Text Available Brucellosis is a worldwide distributed zoonosis that causes important economic losses to animal production. In Brazil, information on the distribution of biovars and genotypes of Brucella spp. is scarce or unavailable. This study aimed (i to biotype and genotype 137 Brazilian cattle isolates (from 1977 to 2008 of B. abortus and (ii to analyze their distribution. B. abortus biovars 1, 2 and 3 (subgroup 3b were confirmed and biovars 4 and 6 were first described in Brazil. Genotyping by the panel 1 revealed two groups, one clustering around genotype 40 and another around genotype 28. Panels 2A and 2B disclosed a high diversity among Brazilian B. abortus strains. Eighty-nine genotypes were found by MLVA16. MLVA16 panel 1 and 2 showed geographic clustering of some genotypes. Biotyping and MLVA16 genotyping of Brazilian B. abortus isolates were useful to better understand the epidemiology of bovine brucellosis in the region.

  3. Isolation of Brucella melitensis from a RB51-vaccinated seronegative goat.

    Science.gov (United States)

    Herrera, Enrique; Rivera, Aldo; Palomares, E Gabriela; Hernández-Castro, Rigoberto; Díaz-Aparicio, Efrén

    2011-08-01

    The purpose of this study is to determine the etiology of abortions presented in a goat herd declared as free of brucellosis and vaccinated with RB51 located in Mexico. The serological diagnosis of brucellosis in 33 animals was performed. The study included three goats that aborted in the last third of gestation and 15 goats that gave birth normally; samples of milk and vaginal exudate were subjected to bacteriological study. All animals were negative for serological diagnosis, and isolation of Brucella melitensis was achieved in a single goat from vaginal exudate. However, the particularity is that this goat was negative to the card, indirect ELISA, and radial immunodiffusion tests. Isolation of a field strain was confirmed by biochemical test resistance to rifampicin and PCR. It is concluded that a goat which aborted in the last third of gestation was found spreading B. melitensis through vaginal discharge despite being vaccinated with RB51 and seronegative for brucellosis.

  4. The Complete Genome of Brucella Suis 019 Provides Insights on Cross-Species Infection

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    Yuanzhi Wang

    2016-01-01

    Full Text Available Brucella species are the most important zoonotic pathogens worldwide and cause considerable harm to humans and animals. In this study, we presented the complete genome of B. suis 019 isolated from sheep (ovine with epididymitis. B. suis 019 has a rough phenotype and can infect sheep, rhesus monkeys and possibly humans. The comparative genome analysis demonstrated that B. suis 019 is closest to the vaccine strain B. suis bv. 1 str. S2. Further analysis associated the rsh gene to the pathogenicity of B. suis 019, and the WbkA gene to the rough phenotype of B. suis 019. The 019 complete genome data was deposited in the GenBank database with ID PRJNA308608.

  5. An ecological perspective on the changing face of Brucella abortus in the western United States

    Science.gov (United States)

    Cross, Paul C.; Maichak, Eric J.; Brennan, Angela; Scurlock, Brandon M.; Henningsen, John C.; Luikart, Gordon

    2013-01-01

    After a hiatus during the 1990s, outbreaks of Brucella abortus in cattle are occurring more frequently in some of the western states of the United States, namely, Montana, Wyoming and Idaho. This increase is coincident with increasing brucellosis seroprevalence in elk (Cervus elaphus), which is correlated with elk density. Vaccines are a seductive solution, but their use in wildlife systems remains limited by logistical, financial, and scientific constraints. Cattle vaccination is ongoing in the region. Livestock regulations, however, tend to be based on serological tests that test for previous exposure and available vaccines do not protect against seroconversion. The authors review recent ecological studies of brucellosis, with particular emphasis on the Greater Yellowstone Area, and highlight the management options and implications of this work, including the potential utility of habitat modifications and targeted hunts, as well as scavengers and predators. Finally, the authors discuss future research directions that will help us to understand and manage brucellosis in wildlife.

  6. Proteomic Profile of Brucella abortus-Infected Bovine Chorioallantoic Membrane Explants

    Science.gov (United States)

    Mol, Juliana P. S.; Pires, Simone F.; Chapeaurouge, Alexander D.; Perales, Jonas; Santos, Renato L.; Andrade, Hélida M.; Lage, Andrey P.

    2016-01-01

    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. PMID:27104343

  7. The feasibility of using antigens prepared with rough Brucella strains for diagnosis of canine brucellosis Utilidad de los antígenos preparados con cepas rugosas de Brucella en el diagnóstico de brucelosis canina

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    G. I. Escobar

    2010-02-01

    Full Text Available Clinical diagnosis of canine brucellosis is not sensitive enough and a negative blood culture cannot rule out the disease. Indirect methods of serological testing such as agar gel immunodiffusion (AGID, rapid slide agglutination test (RSAT and indirect enzyme linked immunoassay (IELISA are preferred for routine diagnosis. Since Brucella canis shares antigenic components with the Brucella ovis and Brucella abortus RB51 strain, it would seem that either strain could be used as antigen. We present data on AGID and IELISA tests using the B. ovis antigen, RSAT and IELISA using the B. canis antigen and IELISA using the B. abortus RB51 antigen. The cut-off values were adjusted by the ROC analysis; the IELISA-B. ovis cut-off value was 23 (%P and the IELISA-B. abortus RB51, 24 (%P, with 100% sensitivity and 98.8% specificity. RSAT detected 100% of positive cases, while AGID was less sensitive. The sera from dogs treated with antibiotic showed that %P correlated well with the clinical course. Sera from dogs presumptively infected with B. suis were negative in all tests performed with the rough Brucella strains. RSAT is a very sensitive screening test and IELISA-B. canis, B. ovis and B. abortus RB51 could be used as confirmatory tests, since they show good specificity and sensitivity.Las técnicas más usadas en el diagnóstico de brucelosis canina son la inmunodifusión en gel de agar (AGID, la microaglutinación en portaobjetos (RSAT y el ELISA indirecto ya que el diagnóstico clínico es poco sensible y el bacteriológico no excluye la enfermedad. Como Brucella canis comparte componentes antigénicos con Brucella ovis y Brucella abortus RB51, estas cepas podrían ser usadas indistintamente como antígenos. En este trabajo presentamos datos sobre las pruebas de AGID e IELISA con antígeno B. ovis, RSAT e IELISA con antígeno B. canis e IELISA con antígeno B. abortus RB51. Los puntos de corte ajustados con la curva ROC fueron (%P 23 para IELISA-B. ovis y

  8. Identification of an IS711 Element Interrupting the wboA Gene of Brucella abortus Vaccine Strain RB51 and a PCR Assay To Distinguish Strain RB51 from Other Brucella Species and Strains

    Science.gov (United States)

    Vemulapalli, Ramesh; McQuiston, John R.; Schurig, Gerhardt G.; Sriranganathan, Nammalwar; Halling, Shirley M.; Boyle, Stephen M.

    1999-01-01

    Brucella abortus vaccine strain RB51 is a natural stable attenuated rough mutant derived from the virulent strain 2308. The genetic mutations that are responsible for the roughness and the attenuation of strain RB51 have not been identified until now. Also, except for an assay based on pulsed-field gel electrophoresis, no other simple method to differentiate strain RB51 from its parent strain 2308 is available. In the present study, we demonstrate that the wboA gene encoding a glycosyltransferase, an enzyme essential for the synthesis of O antigen, is disrupted by an IS711 element in B. abortus vaccine strain RB51. Exploiting this feature, we developed a PCR assay that distinguishes strain RB51 from all other Brucella species and strains tested. PMID:10473532

  9. Pesquisa de aglutininas anti Brucella canis em soros humanos na cidade de São Paulo, Brasil Research on agglutinins for Brucella canis in human sera in the city of S. Paulo, Brazil

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    Maria Helena Matiko Akao Larsson

    1980-09-01

    Full Text Available De 330 soros humanos examinados pela prova de soroaglutinação lenta em tubos, 4(1,21% apresentaram aglutininas anti Brucella canis em diluição 1:100 (1 reagente com título 100, 2 reagentes com título 200 e 1 reagente com título 400.Of the 330 human sera tested by tube agglutination test, 4 (1.21% were positive for Brucella canis antibodies with tilers 1:100 or higher (1 reagent with titer of 1:100, 2 reagents with titer of 1:200, and 1 reagent with tiler of 1:400.

  10. Detection of Brucella canis-induced reproductive diseases in a kennel.

    Science.gov (United States)

    Gyuranecz, Miklós; Szeredi, Levente; Rónai, Zsuzsanna; Dénes, Béla; Dencso, László; Dán, Ádám; Pálmai, Nimród; Hauser, Zsófia; Lami, Erzsébet; Makrai, László; Erdélyi, Károly; Jánosi, Szilárd

    2011-01-01

    Brucella spp. were isolated from an abortion case submitted for laboratory examination 8 months after the first clinical symptoms appeared in a kennel consisting of 31 dogs. Pathological investigations revealed the parallel presence of necrotic placentitis and the strong immunostaining of trophoblast cells by immunohistochemistry (IHC) using hyperimmune rabbit anti-Brucella canis primary antibodies. The rapid slide agglutination test was positive in 7 of 31 (23%) cases. The organism B. canis was successfully cultured from the blood, tissues, or vaginal swabs of only 3 of 31 (10%) cases. The isolated strains were identified as B. canis based on their colony morphology and agglutination with R sera. The strains were initially misidentified as B. suis with the "Bruce-ladder" method, and were subsequently correctly identified as B. canis with a single nucleotide polymorphism (SNP) typing test. Three culture-positive cases and 3 culture-negative cases with histories of reproductive disorders were selected and examined for the presence of B. canis infection using histopathology, IHC, and polymerase chain reaction (PCR) assays. Characteristic histologic lesions were found in all of the 6 animals, whereas IHC and PCR yielded positive results only in single cases from both groups. The results imply that all cases of canine abortion should be examined for brucellosis by bacterial culture of aborted fetuses and placentas. Immunohistochemical examination of placentas is also recommended because it is a quick and sensitive technique compared with bacterial culture. Multiple methods (i.e., serology, blood, and genital bacterial cultures) should be applied simultaneously and repeatedly for the reliable screening of B. canis infection in live individuals.

  11. Efficacy of single calfhood vaccination of elk with Brucella abortus strain 19

    Science.gov (United States)

    Roffe, T.J.; Jones, L.C.; Coffin, K.; Drew, M.L.; Sweeney, Steven J.; Hagius, S.D.; Elzer, P.H.; Davis, D.

    2004-01-01

    Brucellosis has been eradicated from cattle in the states of Wyoming, Montana, and Idaho, USA. However, free-ranging elk (Cervus elaphus) that use feedgrounds in the Greater Yellowstone Area (GYA) and bison (Bison bison) in Yellowstone and Grand Teton national parks still have high seroprevalence to the disease and have caused loss of brucellosis-free status in Wyoming. Management tools to control or eliminate the disease are limited; however, wildlife vaccination is among the methods currently used by wildlife managers in Wyoming. We conducted a controlled challenge study of single calfhood vaccination. Elk calves, caught in January and February of 1999 and 2000 and acclimated to captivity for 3 weeks, were randomly assigned to control or vaccinate groups. The vaccinate groups received Brucetta abortus vaccine strain 19 (S19) by hand-delivered intramuscular injection. Calves were raised to adulthood and bred at either 2.5 or 3.5 years of age for 2000 and 1999 captures, respectively. Eighty-nine (44 controls, 45 vaccinates) pregnant elk entered the challenge portion of the study. We challenged elk at mid-gestation with pathogenic B. abortus strain 2308 by intraconjunctival instillation. Abortion occurred in significantly more (P = 0.002) controls (42; 93%) than vaccinates (32; 71%), and vaccine protected 25% of the vaccinate group. We used Brucella culture of fetus/calf tissues to determine the efficacy of vaccination for preventing infection, and we found that the number of infected fetuses/calves did not differ between controls and vaccinates (P = 0.14). Based on these data, single calfhood vaccination with S19 has low efficacy, will likely have only little to moderate effect on Brucella prevalence in elk, and is unlikely to eradicate the disease in wildlife of the GYA.

  12. Human brucellosis in northwest Ecuador: typifying Brucella spp., seroprevalence, and associated risk factors.

    Science.gov (United States)

    Ron-Román, Jorge; Ron-Garrido, Lenin; Abatih, Emmanuel; Celi-Erazo, Maritza; Vizcaíno-Ordóñez, Laura; Calva-Pacheco, Jaime; González-Andrade, Pablo; Berkvens, Dirk; Benítez-Ortíz, Washington; Brandt, Jef; Fretin, David; Saegerman, Claude

    2014-02-01

    Human brucellosis in Ecuador is underreported and based only on passive surveillance. Since 2008, brucellosis was removed from the list of communicable diseases in the country. Until now, the true human brucellosis picture has not yet been determined. The aim of this study was to determine the seroprevalence of the disease, identify risk factors associated with brucellosis seropositivity in humans, and isolate circulating strains of Brucella spp. in the northwestern part of Ecuador. Between 2006 and 2008, a large transect survey was conducted, based on blood sampling of people from the northwestern part of Ecuador (n=3733) together with an epidemiological inquiry. On the basis of three diagnostic tests used in parallel, the overall seroprevalence was estimated as 1.88% (95% confidence interval [CI] 1.48-2.38). Based on a multivariable random effects logistic regression analysis, the main risk factors associated with human brucellosis seropositivity were contact with livestock (odds ratio [OR]=3.0; CI 1.25-7.08), consumption of fetus and placenta (OR=2.5; CI 1.18-5.22), and involvement in activities at risk for brucellosis infection (OR=1.8; CI 1.00-3.35). Noticeable variation in brucellosis seropositivity among humans within cantons was observed. The circulating strain was Brucella abortus biotype 4. This study emphasized that contact with livestock, consumption of fetus and placenta, and occupational hazard group were all significant risk factors for the transmission of brucellosis among individuals in the northwestern part of Ecuador. Alongside encouraging the launching of educational campaigns against brucellosis, especially in rural areas where 36% of the population lives, controlling this zoonotic disease in animals will directly benefit its prevention in humans, especially because there is no safe and efficacious vaccine against brucellosis in humans.

  13. Protection to respiratory challenge of Brucella abortus strain 2308 in the lung.

    Science.gov (United States)

    Surendran, Naveen; Sriranganathan, Nammalwar; Boyle, Stephen M; Hiltbold, Elizabeth M; Tenpenny, Nancy; Walker, Michelle; Zimmerman, Kurt; Werre, Stephen; Witonsky, Sharon G

    2013-08-28

    Brucella is amongst the top 5 causes of zoonotic disease worldwide. Infection is through ingestion, inhalation or contact exposure. Brucella is characterized as a class B pathogen by Centers of Disease Control and Prevention (CDC). Currently, there are no efficacious vaccines available in people. Currently available USDA approved vaccines for animals include B. abortus strain RB51 and B. melitensis Rev1. Protection is mediated by a strong innate and CD4 Th1, CD8 Tc1 immune response. If protective vaccines can be developed, disease in people and animals can be controlled. While strain RB51 protects in cattle, and against intraperitoneal challenge in mice, it does not protect against respiratory challenge. Therefore, we assessed the efficacy of strain RB51 combined with different TLR agonists, and O-side chain from LPS, to enhance protection against respiratory challenge with strain 2308. We hypothesized that TLR agonists and O-side chain would enhance protection. Strains RB51 with TLR2 agonist, RB51 with TLR4 agonist and strain 19 provided significant protection in the lung. Protection using strain RB51 with TLR agonists was associated with increased IgG2a and IgG1 in the (bronchoalveolar lavage) BAL and serum, and increased IgA (serum). Splenocytes from strain RB51 with TLR2 vaccinated mice up-regulated antigen specific interferon-gamma and TNF-alpha production. Vaccination and challenge resulted in significant increases in activated dendritic cells (DCs), and increased CD4 and CD8 cells in the BAL. Overall, this study demonstrates the ability of TLR agonists 2 and 4 to up-regulate strain RB51 mediated protection in the lung to respiratory challenge against strain 2308. Copyright © 2013. Published by Elsevier Ltd.

  14. Parenteral vaccination of domestic pigs with Brucella abortus strain RB51.

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    Stoffregen, William C; Olsen, Steven C; Bricker, Betsy J

    2006-10-01

    To determine the immunogenicity and efficacy of Brucella abortus strain RB51 (SRB51) as a vaccine in domestic pigs. Sixty-eight 6-week-old crossbred domestic pigs and twenty-four 4-month-old gilts. In experiment 1, pigs were vaccinated IM (n = 51) with 2 x 10(10) CFUs of SRB51 or sham inoculated (17). Periodic blood samples were obtained to perform blood cultures, serologic evaluations, and cell-mediated immunity assays. Necropsies were performed at selected times between weeks 1 and 23 after vaccination to determine vaccine clearance. In experiment 2, gilts were similarly vaccinated (n = 18) or sham inoculated (8) and similar samples were obtained after vaccination. Gilts were bred and challenged conjunctivally with 5.0 x 10(7) CFUs of virulent Brucella suis strain 3B. Necropsies were performed on gilts and on fetuses or neonates after abortion or parturition, respectively. Bacterial cultures and serologic evaluations were performed on samples obtained at necropsy to determine vaccine efficacy. Humoral and cell-mediated immune responses did not differ between vaccinates and controls. After vaccination, SRB51 was not isolated from blood cultures of either group and was isolated from lymphoid tissues of 3 pigs at 2 weeks (n = 2) and 4 weeks (1) after vaccination. No differences were found in isolation of B suis or in seroconversion between vaccinated and control gilts and between their neonates or aborted fetuses. Parenteral vaccination with SRB51 does not induce humoral or cell-mediated immune responses. Vaccination with SRB51 did not protect gilts or their neonates and fetuses from virulent challenge with B suis.

  15. Enhanced efficacy of recombinant Brucella abortus RB51 vaccines against B. melitensis infection in mice.

    Science.gov (United States)

    Vemulapalli, Ramesh; Contreras, Andrea; Sanakkayala, Neelima; Sriranganathan, Nammalwar; Boyle, Stephen M; Schurig, Gerhardt G

    2004-09-08

    Brucella abortus strain RB51 is an attenuated rough strain, currently being used as the official live vaccine for bovine brucellosis in the USA and several other countries. In strain RB51, the wboA gene, encoding a glycosyltransferase required for the O-side chain synthesis, is disrupted by an IS711 element. Recently, we have demonstrated that strain RB51WboA, RB51 complemented with a functional wboA gene, remains rough but expresses low quantities of O-side chain in the cytoplasm. Mice vaccinated with strain RB51WboA develop greatly enhanced resistance against challenge with B. abortus virulent strain 2308. We have also demonstrated that overexpression of Cu/Zn superoxide dismutase (SOD) in strain RB51 (RB51SOD) significantly increases its vaccine efficacy against strain 2308 challenge. In this study, we constructed a new recombinant strain, RB51SOD/WboA, that over expresses SOD with simultaneous expression of O-side chain in the cytoplasm. We tested the vaccine potential of strains RB51SOD, RB51WboA, RB51SOD/WboA against challenge with virulent Brucella melitensis 16M and B. abortus 2308 in mice. In comparison with strain RB51, strain RB51SOD induced better protection against strain 2308, but not strain 16M, challenge. Similar to strain RB51WboA, vaccination with strain RB51SOD/WboA resulted in complete protection of the mice from infection with strain 2308. When challenged with strain 16M, mice vaccinated with either strain RB51WboA or strain RB51SOD/WboA were significantly better protected than those vaccinated with strain RB51 or RB51SOD. These results suggest that strains RB51WboA and RB51SOD/WboA are good vaccine candidates for inducing enhanced protection against B. melitensis infection.

  16. Brucella abortus strain RB51 vaccination in elk. I. Efficacy of reduced dosage.

    Science.gov (United States)

    Cook, Walter E; Williams, Elizabeth S; Thorne, E Tom; Kreeger, Terry J; Stout, Glen; Bardsley, Katie; Edwards, Hank; Schurig, Gerhardt; Colby, Lesley A; Enright, Fred; Elzer, Philip H

    2002-01-01

    Bovine brucellosis is a serious zoonotic disease affecting some populations of Rocky Mountain elk (Cervus elaphus nelsoni) and bison (Bison bison) in the Greater Yellowstone Area, USA. The fear that elk and/or bison may spread Brucella abortus to livestock has prompted efforts to reduce or eliminate the disease in wildlife. Brucella abortus strain RB51 (RB51) vaccine has recently been approved for use in cattle. Unlike strain 19 vaccine, RB51 does not cause false positive reactions on standard brucellosis serologic tests. If effective, it may become the vaccine of choice for wildlife. In February 1995, 45 serologically negative female elk calves were trapped and taken to the Sybille Wildlife Research and Conservation Education Unit near Wheatland, Wyoming, USA. In May 1995, 16 of these elk calves were hand-vaccinated with 1 x 10(9) colony forming units (CFU) of RB51, 16 were vaccinated with 1 x 10(8) CFU RB51 by biobullet, and 13 were given a saline placebo. The elk were bred in fall of 1996 and they were challenged with 1 x 10(7) CFU of B. abortus strain 2308 by intraconjunctival inoculation in March 1997. Thirteen (100%) control elk aborted, 14 (88%) hand-vaccinated elk aborted, and 12 (75%) biobullet vaccinated elk aborted or produced nonviable calves. These results suggest that a single dose of 1 x 10(8) to 1 x 10(9) CFU RB51 does not provide significant protection against B. abortus induced abortion in elk. However, the vaccine appears to be safe at this dose and additional study may reveal a more effective RB51 vaccine regimen for elk.

  17. The Lipopolysaccharide Core of Brucella abortus Acts as a Shield Against Innate Immunity Recognition

    Science.gov (United States)

    Iriarte, Maite; Manček-Keber, Mateja; Barquero-Calvo, Elías; Palacios-Chaves, Leyre; Chacón-Díaz, Carlos; Chaves-Olarte, Esteban; Martirosyan, Anna; von Bargen, Kristine; Grilló, María-Jesús; Jerala, Roman; Brandenburg, Klaus; Llobet, Enrique; Bengoechea, José A.; Moreno, Edgardo

    2012-01-01

    Innate immunity recognizes bacterial molecules bearing pathogen-associated molecular patterns to launch inflammatory responses leading to the activation of adaptive immunity. However, the lipopolysaccharide (LPS) of the gram-negative bacterium Brucella lacks a marked pathogen-associated molecular pattern, and it has been postulated that this delays the development of immunity, creating a gap that is critical for the bacterium to reach the intracellular replicative niche. We found that a B. abortus mutant in the wadC gene displayed a disrupted LPS core while keeping both the LPS O-polysaccharide and lipid A. In mice, the wadC mutant induced proinflammatory responses and was attenuated. In addition, it was sensitive to killing by non-immune serum and bactericidal peptides and did not multiply in dendritic cells being targeted to lysosomal compartments. In contrast to wild type B. abortus, the wadC mutant induced dendritic cell maturation and secretion of pro-inflammatory cytokines. All these properties were reproduced by the wadC mutant purified LPS in a TLR4-dependent manner. Moreover, the core-mutated LPS displayed an increased binding to MD-2, the TLR4 co-receptor leading to subsequent increase in intracellular signaling. Here we show that Brucella escapes recognition in early stages of infection by expressing a shield against recognition by innate immunity in its LPS core and identify a novel virulence mechanism in intracellular pathogenic gram-negative bacteria. These results also encourage for an improvement in the generation of novel bacterial vaccines. PMID:22589715

  18. DETECCION DE Brucella abortus POR PCR EN MUESTRAS DE SANGRE Y LECHE DE VACUNOS

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    Xiomara Mosquera C

    2008-12-01

    Full Text Available Objetivo. Evaluar el uso de la Reacción en Cadena de la Polimerasa (PCR para la detección de Brucella abortus en muestras de sangre y leche de vacunos. Materiales y métodos. Este estudio de tipo descriptivo fue realizado durante los años 2004 y 2005. Se analizaron 136 animales de tres fincas localizadas en el municipio de Durania, Norte de Santander, Colombia. Se evaluó la presencia de anticuerpos en la leche mediante la prueba del anillo (PAL. Se amplificó el fragmento de 223pb del gen BCSP31. Se emplearon los cebadores B4 y B5 de la región interna de la secuencia del gen BCSP31 (GenBank, número M20404. Resultados. En aquellos animales positivos se obtuvo una muestra de sangre y leche para el análisis por PCR, la sangre no fue analizada por serología. Se evaluaron diferentes métodos de extracción de ADN. Se encontró que un 13.2% (18/136 de las muestras de leche fueron positivas a la PAL. Se analizaron 33 muestras de leche negativas por PAL de las cuales el 30.3% (10/33 resultaron positivas por PCR. Al analizar las muestras de sangre de los animales positivos por PAL el 94.1% (16/17 fueron positivas por PCR, mientras que el 47% (8/17 de las muestras de leche positivas por PAL, fueron positivas por PCR. Conclusiones. Se demostró la amplificación de un fragmento de ADN de Brucella abortus en muestras de sangre y leche de vacunos. Los resultados preliminares demostraron que es posible usar PCR como prueba diagnóstica de brucelosis en Colombia.

  19. Seroprevalencia de Brucella spp en estudiantes de Medicina Veterinaria, Bogotá, Colombia

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    Iván Alberto Méndez R

    2013-08-01

    Full Text Available Introducción: La brucelosis es un enfermedad zoonotica relacionada al contacto con perros y con ganado vacuno, porcino y caprino, el agente etiológico pertenece al género Brucella con diez especies de importancia para el ser humano; su capacidad para evadir el sistema inmune y sobrevivir en el fago-lisosoma genera un cuadro clínico crónico en el ser humano caracterizado por fiebre ondulante y afectación osteoarticular. Su amplia variedad de expresión clínica genera un reto al momento del diagnóstico y tratamiento; por tal razón es relevante el reconocimiento de las pruebas diagnósticas que permitan establecer el posible contacto con el microorganismo. Objetivo: Establecer la seroprevalencia a Brucella spp en una población de estudiantes de Medicina Veterinaria con posible exposición ocupacional. Materiales y Métodos: Estudio descriptivo observacional donde se estudiaron 272 sueros obtenidos de estudiantes de medicina veterinaria mediante la técnica de rosa de bengala. Resultados: El 18,4% (50/272 de los sujetos presento positividad por rosa de bengala; 28 hombres (56% y 22 mujeres (44%. Conclusiones: La serología mediante rosa de bengala en esta población de riesgo evidenció un 18,5% de sujetos seropositivos. Se sugiere el empleo de barreras efectivas de protección durante el contacto con animales portadores del microorganismo durante el entrenamiento como estudiantes de Medicina veterinaria.

  20. MLVA and LPS Characteristics of Brucella canis Isolated from Humans and Dogs in Zhejiang, China

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    Dongri Piao

    2017-12-01

    Full Text Available BackgroundBrucella canis is a pathogenic bacterium that causes brucellosis in dogs, and its zoonotic potential has been increasing in recent years. B. canis is a rare source of human brucellosis in China, where Brucella melitensis has been the major pathogen associated with human brucellosis outbreaks. In late 2011, a case of a B. canis infection was detected in a human patient in Zhejiang Province, China. To compare the genotypes between strains of B. canis isolated from the patient and from dogs, a multiple-locus variable-number tandem-repeat analysis (MLVA-16 was performed. In addition, the lipopolysaccharide-synthesis-related genes were analyzed with the B. canis reference strain RM6/66.Results32 B. canis strains were divided into 26 genotypes using MLVA-16 [Hunter-Gaston Diversity Index (HGDI = 0.976]. The HGDI indexes for various loci ranged between 0.000 and 0.865. All four Hangzhou isolates were indistinguishable using panel 1 (genotype 3 and panel 2A (genotype 28. However, these strains were distinctly different from other isolates from Beijing, Jiangsu, Liaoning, and Inner Mongolia at Bruce 09. The emergence of a human B. canis infection was limited to an area. Comparative analysis indicated B. canis from canines and humans have no differences in lipopolysaccharide-synthesis locus.ConclusionThe comprehensive approaches have been used to analyze human and canine B. canis isolates, including molecular epidemiological and LPS genetic characteristics. Further detailed analysis of the whole genomic sequencing will contribute to understanding of the pathogenicity of B. canis in humans.

  1. Retrospective seroepidemiological investigations against Morbillivirus, Toxoplasma gondii and Brucella spp. in cetaceans stranded along the Italian coastline (1998-2014).

    Science.gov (United States)

    Profeta, Francesca; Di Francesco, Cristina Esmeralda; Marsilio, Fulvio; Mignone, Walter; Di Nocera, Fabio; De Carlo, Esterina; Lucifora, Giuseppe; Pietroluongo, Guido; Baffoni, Marina; Cocumelli, Cristiano; Eleni, Claudia; Terracciano, Giuliana; Ferri, Nicola; Di Francesco, Gabriella; Casalone, Cristina; Pautasso, Alessandra; Mazzariol, Sandro; Centelleghe, Cinzia; Di Guardo, Giovanni

    2015-08-01

    This study reports the results of seroepidemiological investigations carried out against Morbillivirus, Toxoplasma gondii and Brucella spp. on blood serum samples collected from 70 cetacean specimens found stranded along the Italian coastline between 1998 and 2014. A total number of 23 serum samples (32.8%) obtained from Stenella coeruleoalba, Tursiops truncatus, Balaenoptera physalus and Globicephala melas harboured anti-Morbillivirus neutralizing antibodies. Ten sera (16%) collected from S. coeruleoalba and T. truncatus were found positive against T. gondii, while no antibodies against Brucella spp. were found. These data reveal that stranded cetaceans provide a unique opportunity for monitoring the health status of free-ranging animals living in the Mediterranean Sea, in order to investigate the level of exposure of cetacean populations to selected infectious agents representing a serious threat for aquatic mammals. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. THE INVESTIGATION OF BRUCELLA ANTIBODY WITH MILK RING TEST AND AGGLUTINATION TEST IN MILK COLLECTED FROM SAMSUN REGION

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    Goknur TERZI

    2006-06-01

    Full Text Available In this study Brucella antibodies were investigated with agglutination test (Whey-AT and Milk Ring Test (MRT in a total of 100 milk samples as 50 of cow milk and 50 of goat milk collected from center and villages of Samsun. According to MRT Brucella antibodies was positive at 10 samples (20 % of cow milk and 6 samples (12 % of goat milk. In cow milk, 4 (8 % positive, 3 (6 % suspicious and 43 (86 % negative samples; in goat milk 3 (6 % positive, 2 (4 % suspicious and 45 (90 % negative samples were determined according to antibodies titre of serum agglutination test (Whey-AT. [TAF Prev Med Bull 2006; 5(3.000: 196-203

  3. Evaluation of bison (Bison bison) semen from Yellowstone National Park, Montana, USA, bulls for Brucella abortus shedding.

    Science.gov (United States)

    Frey, Rebecca K; Clarke, P Ryan; McCollum, Matt P; Nol, Pauline; Johnson, Kammy R; Thompson, Brent D; Ramsey, Jennifer M; Anderson, Neil J; Rhyan, Jack C

    2013-07-01

    To determine if bison (Bison bison) bulls from Yellowstone National Park (YNP), Montana, USA, shed an infective dose of Brucella abortus in semen, 50 YNP bulls were captured on public lands in Montana during the winter and early spring (April-May) of 2010 and 2011. The bulls were immobilized, and blood and semen samples were collected for serology and Brucella culture. Thirty-five bulls (70%) were antibody-positive, and B. abortus was cultured from semen in three (9%) of the 35 antibody-positive or suspect bulls, though not at concentrations considered an infective dose. Eight bulls (six antibody-positive, two negative) had palpable lesions of the testes, epididymides, or seminal vesicles consistent with B. abortus infection. Breeding soundness exams and semen analysis suggested that antibody-positive bulls were more likely to have nonviable ejaculate (8/35; 23%) than bulls without detectable antibody (2/15; 13%).

  4. Efecto de la inoculacion intracutanea de brucellas en el cobayo. La multiplicación intracelular de los germenes

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    Javier Arias-Stella

    1951-01-01

    Full Text Available Se hace un estudio de la sucesión de fenómenos desencadenados en el cobayo por la inyección intracutánea de brucellas, dando los caracteres macro y microscópicos escalonados de la reacción. Se logra demostrar la multiplicación intracelular del germen habiéndose seguido cuantitativamente, día a día, el incremento del número de Brucellas desarrollándose en los macrófagos en el foco inflamatorio. Entre el 9° y 13° día el 100 % de los histiocitos estaban parasitados.

  5. Comparison of Abortion and Infection after Experimental Challenge of Pregnant Bison and Cattle with Brucella abortus Strain 2308▿

    Science.gov (United States)

    Olsen, S. C.; Johnson, C.

    2011-01-01

    A comparative study was conducted using data from naive bison (n = 45) and cattle (n = 46) from 8 and 6 studies, respectively, in which a standardized Brucella abortus strain 2308 experimental challenge was administered during midgestation. The incidence of abortion, fetal infection, uterine or mammary infection, or infection in maternal tissues after experimental challenge was greater (P abort, the time between experimental challenge and abortion was shorter (P abortion did not differ (P > 0.05) between cattle and bison. The results of our study suggest that naive bison and cattle have similarities and differences after experimental exposure to a virulent B. abortus strain. Although our data suggest that bison may be more susceptible to infection with Brucella, some pathogenic characteristics of brucellosis were similar between bison and cattle. PMID:21976222

  6. Development of multiplex real-time PCR assay for the detection of Brucella spp., Leptospira spp. and Campylobacter foetus

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    Abdelfattah M. Selim

    2014-12-01

    Full Text Available Abortion among dairy cattle is one of the major causes of economic losses in the livestock industry. This study describes a 1-step multiplex real-time polymerase chain reaction (PCR to detect Brucella spp., Leptospira spp. and Campylobacter foetus, these are significant bacteria commonly implicated in bovine abortion. ß-actin was added to the same PCR reaction as an internal control to detect any extraction failure or PCR inhibition. The detection limit of multiplex real-time PCR using purified DNA from cultured organisms was set to 5 fg for Leptospira spp. and C. foetus and to 50 fg for Brucella spp. The multiplex real-time PCR did not produce any non-specific amplification when tested with different strains of the 3 pathogens. This multiplex real-time PCR provides a valuable tool for diagnosis, simultaneous and rapid detection for the 3 pathogens causing abortion in bovine.

  7. A 6-Nucleotide Regulatory Motif within the AbcR Small RNAs of Brucella abortus Mediates Host-Pathogen Interactions

    Science.gov (United States)

    Sheehan, Lauren M.

    2017-01-01

    ABSTRACT In Brucella abortus, two small RNAs (sRNAs), AbcR1 and AbcR2, are responsible for regulating transcripts encoding ABC-type transport systems. AbcR1 and AbcR2 are required for Brucella virulence, as a double chromosomal deletion of both sRNAs results in attenuation in mice. Although these sRNAs are responsible for targeting transcripts for degradation, the mechanism utilized by the AbcR sRNAs to regulate mRNA in Brucella has not been described. Here, two motifs (M1 and M2) were identified in AbcR1 and AbcR2, and complementary motif sequences were defined in AbcR-regulated transcripts. Site-directed mutagenesis of M1 or M2 or of both M1 and M2 in the sRNAs revealed transcripts to be targeted by one or both motifs. Electrophoretic mobility shift assays revealed direct, concentration-dependent binding of both AbcR sRNAs to a target mRNA sequence. These experiments genetically and biochemically characterized two indispensable motifs within the AbcR sRNAs that bind to and regulate transcripts. Additionally, cellular and animal models of infection demonstrated that only M2 in the AbcR sRNAs is required for Brucella virulence. Furthermore, one of the M2-regulated targets, BAB2_0612, was found to be critical for the virulence of B. abortus in a mouse model of infection. Although these sRNAs are highly conserved among Alphaproteobacteria, the present report displays how gene regulation mediated by the AbcR sRNAs has diverged to meet the intricate regulatory requirements of each particular organism and its unique biological niche. PMID:28588127

  8. A 6-Nucleotide Regulatory Motif within the AbcR Small RNAs of Brucella abortus Mediates Host-Pathogen Interactions

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    Lauren M. Sheehan

    2017-06-01

    Full Text Available In Brucella abortus, two small RNAs (sRNAs, AbcR1 and AbcR2, are responsible for regulating transcripts encoding ABC-type transport systems. AbcR1 and AbcR2 are required for Brucella virulence, as a double chromosomal deletion of both sRNAs results in attenuation in mice. Although these sRNAs are responsible for targeting transcripts for degradation, the mechanism utilized by the AbcR sRNAs to regulate mRNA in Brucella has not been described. Here, two motifs (M1 and M2 were identified in AbcR1 and AbcR2, and complementary motif sequences were defined in AbcR-regulated transcripts. Site-directed mutagenesis of M1 or M2 or of both M1 and M2 in the sRNAs revealed transcripts to be targeted by one or both motifs. Electrophoretic mobility shift assays revealed direct, concentration-dependent binding of both AbcR sRNAs to a target mRNA sequence. These experiments genetically and biochemically characterized two indispensable motifs within the AbcR sRNAs that bind to and regulate transcripts. Additionally, cellular and animal models of infection demonstrated that only M2 in the AbcR sRNAs is required for Brucella virulence. Furthermore, one of the M2-regulated targets, BAB2_0612, was found to be critical for the virulence of B. abortus in a mouse model of infection. Although these sRNAs are highly conserved among Alphaproteobacteria, the present report displays how gene regulation mediated by the AbcR sRNAs has diverged to meet the intricate regulatory requirements of each particular organism and its unique biological niche.

  9. Geographic pattern of serum antibody prevalence for Brucella spp. in caribou, grizzly bears, and wolves from Alaska, 1975-1998.

    Science.gov (United States)

    Zarnke, Randall L; Ver Hoef, Jay M; DeLong, Robert A

    2006-07-01

    Blood samples were collected from 2,635 caribou (Rangifer tarandus), 1,238 grizzly bears (Ursus arctos), and 930 wolves (Canis lupus) from throughout mainland Alaska during 1975-98. Sera were tested for evidence of exposure to Brucella spp. Serum antibody prevalences were highest in the northwestern region of the state. In any specific area, antibody prevalences for caribou and wolves were of a similar magnitude, whereas antibody prevalence for bears in these same areas were two to three times higher.

  10. Complement Fixation Test To Assess Humoral Immunity in Cattle and Sheep Vaccinated with Brucella abortus RB51

    OpenAIRE

    Adone, Rosanna; Ciuchini, Franco

    1999-01-01

    The live attenuated Brucella abortus strain RB51 is a rifampin-resistant, lipopolysaccharide (LPS) O-chain-deficient mutant of virulent B. abortus 2308. The reduced O-chain content in RB51 prevents this bacterium from inducing antibodies detectable by the conventional serologic tests for bovine brucellosis diagnosis that mainly identify antibodies to LPS. The absence of available serologic tests for RB51 also complicates the diagnosis of possible RB51 infections in...

  11. Efficacy of Dart or Booster Vaccination with Strain RB51 in Protecting Bison against Experimental Brucella abortus Challenge

    OpenAIRE

    Olsen, S. C.; Johnson, C. S.

    2012-01-01

    This study characterized the efficacy of the Brucella abortus strain RB51 vaccine in bison when delivered by single intramuscular vaccination (hand RB51), by single pneumatic dart delivery (dart RB51), or as two vaccinations approximately 13 months apart (booster RB51) in comparison to control bison. All bison were challenged intraconjunctivally in midgestation with 107 CFU of B. abortus strain 2308 (S2308). Bison were necropsied and sampled within 72 h of abortion or delivery of a live calf....

  12. Multi-locus variable-number tandem repeat analysis of Chinese Brucella strains isolated from 1953 to 2013.

    Science.gov (United States)

    Tian, Guo-Zhong; Cui, Bu-Yun; Piao, Dong-Ri; Zhao, Hong-Yan; Li, Lan-Yu; Liu, Xi; Xiao, Pei; Zhao, Zhong-Zhi; Xu, Li-Qing; Jiang, Hai; Li, Zhen-Jun

    2017-05-02

    Brucellosis was a common human and livestock disease caused by Brucella strains, the category B priority pathogens by the US Center for Disease Control (CDC). Identified as a priority disease in human and livestock populations, the increasing incidence in recent years in China needs urgent control measures for this disease but the molecular background important for monitoring the epidemiology of Brucella strains at the national level is still lacking. A total of 600 Brucella isolates collected during 60 years (from 1953 to 2013) in China were genotyped by multiple locus variable-number tandem repeat analysis (MLVA) and the variation degree of MLVA11 loci was calculated by the Hunter Gaston Diversity Index (HGDI) values. The charts and map were processed by Excel 2013, and cluster analysis and epidemiological distribution was performed using BioNumerics (version 5.1). The 600 representative Brucella isolates fell into 104 genotypes with 58 singleton genotypes by the MLVA11 assay, including B. melitensis biovars 2 and 3 (five main genotypes), B. abortus biovars 1 and 3 (two main genotypes), B. suis biovars 1 and 3 (three main genotypes), and B. canis (two main genotypes) respectively. While most B. suis biovar 1 and biovar 3 were respectively found in northern provinces and southern provinces, B. melitensis and B. abortus strains were dominant in China. Canine Brucellosis was only found in animals without any human cases reported. Eight Brucellosis epidemic peaks emerged during the 60 years between 1953 and 2013: 1955 - 1959, 1962 - 1969, 1971 - 1975, 1977 - 1983, 1985 - 1989, 1992 - 1997, 2000 - 2008 and 2010 - 2013 in China. Brucellosis has its unique molecular epidemiological patterns with specific spatial and temporal distribution according to MLVA. IDOP-D-16-00101.

  13. The prevalence of antibodies of Brucella abortus, Dermatophilus congolensis and bovine leukaemia virus in Nigerian slaughter cattle.

    Science.gov (United States)

    Oyejide, A; Adu, F D; Makinde, A A; Ezeh, E N

    1987-01-01

    In a pilot survey to compare the relative prevalence of three diseases in apparently healthy White Fulani Zebu (WFZ) cattle slaughtered in Nigeria, sera from 80 randomly selected animals with no significant gross lesions on ante mortem and post mortem inspection were examined for antibodies to Brucella abortus, Dermatophilus congolensis and bovine leukaemia virus. Of the samples screened, 5.0, 8.8 and 2.0% showed serological evidence for brucellosis, cutaneous streptothricosis and bovine leukosis respectively.

  14. Brucella suis vaccine strain S2-infected immortalized caprine endometrial epithelial cell lines induce non-apoptotic ER-stress.

    Science.gov (United States)

    Wang, Xiangguo; Lin, Pengfei; Yin, Yanlong