WorldWideScience

Sample records for brucella melitensis infection

  1. 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

  2. An influenza viral vector Brucella abortus vaccine induces good cross-protection against Brucella melitensis infection in pregnant heifers.

    Science.gov (United States)

    Tabynov, Kaissar; Ryskeldinova, Sholpan; Sansyzbay, Abylai

    2015-07-17

    Brucella melitensis can be transmitted and cause disease in cattle herds as a result of inadequate management of mixed livestock farms. Ideally, vaccines against Brucella abortus for cattle should also provide cross-protection against B. melitensis. Previously we created a novel influenza viral vector B. abortus (Flu-BA) vaccine expressing the Brucella ribosomal proteins L7/L12 or Omp16. This study demonstrated Flu-BA vaccine with adjuvant Montanide Gel01 provided 100% protection against abortion in vaccinated pregnant heifers and good cross-protection of the heifers and their calves or fetuses (90-100%) after challenge with B. melitensis 16M; the level of protection provided by Flu-BA was comparable to the commercial vaccine B. abortus S19. In terms of the index of infection and colonization of Brucella in tissues, both vaccines demonstrated significant (P=0.02 to P<0.0001) protection against B. melitensis 16M infection compared to the negative control group (PBS+Montanide Gel01). Thus, we conclude the Flu-BA vaccine provides cross-protection against B. melitensis infection in pregnant heifers.

  3. Evaluation of Brucella melitensis B115 as rough-phenotype vaccine against B. melitensis and B. ovis infections.

    Science.gov (United States)

    Adone, R; Francia, M; Ciuchini, F

    2008-09-08

    Brucella melitensis strain Rev1 is used as vaccine for the prophylaxis of brucellosis in sheep and goats. Because of its smooth phenotype, however, it induces antibodies directed to the O-polysaccharide (O-PS) of the lipopolysaccharide (LPS), thus unabling to distinguish between vaccinated and infected animals. It has been speculated that alternative vaccines could be live, attenuated Brucella rough strains, which are devoid of the O-PS. B. melitensis B115 is a natural, attenuated, rough strain. The O-PS is not exposed at the surface but is present in the cytoplasm. We tested the protective activity of B115 against B. melitensis and B. ovis infections in mice, in comparison with that of Rev1. The residual virulence and the humoral response following B115 vaccination were also evaluated. Vaccination with B115 conferred significant protective immunity against B. melitensis 16M and B. ovis challenge strains, equivalent to that provided by Rev1. No interfering antibodies to O-PS were detected, while the B115 vaccination was monitored by a specific B115-based complement fixation test. These promising features suggest further evaluation of B. melitensis B115 as vaccine for target animal hosts.

  4. Brucella abortus S19 vaccine protects dairy cattle against natural infection with Brucella melitensis.

    Science.gov (United States)

    van Straten, Michael; Bardenstein, Svetlana; Keningswald, Gaby; Banai, Menachem

    2016-11-21

    Brucellosis is a zoonotic disease that can cause severe illness in humans and considerable economic loss in the livestock industry. Although small ruminants are the preferential host for Brucella melitensis, this pathogen has emerged as a cause for Brucella outbreaks in cattle. S19 vaccination is implemented in many countries where B. abortus is endemic but its effectiveness against B. melitensis has not been validated. Here we show that vaccine effectiveness in preventing disease transmission between vaccinated and unvaccinated cohorts, as determined by seroconversion, was 87.2% (95% CI 69.5-94.6%). Furthermore, vaccination was associated with a reduced risk for abortion. Together, our data emphasize the role S19 vaccination could play in preventing B. melitensis outbreaks in areas where this pathogen is prevalent in small ruminant populations.

  5. Human infection by Brucella melitensis: an outbreak attributed to contact with infected goats.

    Science.gov (United States)

    Wallach, J C; Samartino, L E; Efron, A; Baldi, P C

    1997-12-01

    Although several outbreaks of Brucella melitensis infection have been reported among laboratory workers or goat cheese consumers, outbreaks related to rural labour have been rarely studied. An outbreak of human brucellosis among farm workers of Argentina was studied and revealed a close relationship with an epidemic of caprine abortions which occurred shortly before on the same farm. High rates of B. melitensis infection were found among goats. Active brucellosis was diagnosed in 33 subjects (14 with positive blood culture for B. melitensis), while other 27 did not show evidence of illness. While 25 of the brucellosis active patients were rural workers, only 5 of the healthy subjects were engaged in rural labour. Active brucellosis was diagnosed in 91.3% of the subjects in continuous contact with goats and in 32% of those having an occasional contact with the animals. All the 60 subjects denied consumption of goat cheese or milk. As shown here, epidemic human infections by B. melitensis may develop among people frequently in contact with infected goat herds or goat manure.

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

    OpenAIRE

    Gamal Wareth; Murat Eravci; Christoph Weise; Uwe Roesler; Falk Melzer; Sprague, Lisa D.; Heinrich Neubauer; Jayaseelan Murugaiyan

    2016-01-01

    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 ...

  7. 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.

  8. 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.

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

    Science.gov (United States)

    Wareth, Gamal; Eravci, Murat; Weise, Christoph; Roesler, Uwe; Melzer, Falk; Sprague, Lisa D; Neubauer, Heinrich; Murugaiyan, Jayaseelan

    2016-04-30

    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.

  10. A rare case of Brucella melitensis infection in an obstetrician during the delivery of a transplacentally infected infant.

    Science.gov (United States)

    Poulou, Aggeliki; Markou, Fani; Xipolitos, Ioannis; Skandalakis, Panayotis N

    2006-07-01

    We report a case of Brucella melitensis infection in an obstetrician who was infected during the delivery of an infant suffering from congenital brucellosis. The obstetrician was treated with doxycycline and rifampin and fully recovered. This is the first reported case of Brucella infection transmitted through infectious secretions at the delivery of a transplacentally infected newborn and emphasizes that, especially in endemic areas educational and technical measures are needed in order the obstetricians to avoid ingestion of secretions during clearance of the newborn's respiratory tract from saliva and amniotic fluid.

  11. 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.

  12. 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.

  13. Subcutaneous immunization with a novel immunogenic candidate (urease) confers protection against Brucella abortus and Brucella melitensis infections.

    Science.gov (United States)

    Abkar, Morteza; Amani, Jafar; Sahebghadam Lotfi, Abbas; Nikbakht Brujeni, Gholamreza; Alamian, Saeed; Kamali, Mehdi

    2015-08-01

    Brucellosis is a world prevalent endemic illness that is transmitted from domestic animals to humans. Brucella spp. exploits urease for survival in the harsh conditions of stomach during the gastrointestinal infection. In this study, we examined the immune response and the protection elicited by using recombinant Brucella urease (rUrease) vaccination in BALB/c mice. The urease gene was cloned in pET28a and the resulting recombinant protein was employed as subunit vaccine. Recombinant protein was administered subcutaneously and intraperitoneally. Dosage reduction was observed with subcutaneous (SC) vaccination when compared with intraperitoneal (IP) vaccination. rUrease induced mixed Th1-Th2 immune responses with high titers of specific IgG1 and IgG2a. In lymphocyte proliferation assay, splenocytes from IP and SC-vaccinated mice displayed a strong recall proliferative response with high amounts of IL-4, IL-12 and IFN-γ production. Vaccinated mice were challenged with virulent Brucella melitensis, B. abortus and B. suis. The SC vaccination route exhibited a higher degree of protection than IP vaccination (p value ≤ 0.05). Altogether, our results indicated that rUrease could be a useful antigen candidate for the development of subunit vaccines against brucellosis.

  14. Brucella Endocarditis Caused by Brucella Melitensis

    OpenAIRE

    Akdag, Serkan; Akyol, Aytac; Simsek, Hakki; Sahin,Musa; Yaman, Mehmet

    2014-01-01

    Abstract. We present a rare case of brucella endocarditis, forming a vegetation on the mitral valve. The definitive diagnosis has been made with clinical suspicion, positive serology, the demonstration of the vegetation with the echocardiography and with the production from the multiple blood culture of Brucella melitensis and from the excised valve. Our patient has been successfully treated with specific antibiotherapy and the surgery of replacement of mitral valve. Our aim in presenting the...

  15. Validation of a competitive ELISA for diagnosis of Brucella melitensis infection in sheep and goats.

    Science.gov (United States)

    Minas, Anastasios; Stournara, Athanasia; Christodoulopoulos, Georgios; Katsoulos, Panagiotis D

    2008-09-01

    A competitive enzyme-linked immunosorbent assay (cELISA) was validated for the serodiagnosis of Brucella melitensis infection in small ruminants using 2108 positive and 2154 negative reference sera from sheep and goats. The optimum cut-off values, offering the highest diagnostic sensitivity (DSn) and diagnostic specificity (DSp), determined by receiver operating characteristic analysis, were at 23.6%, 21.8% and 25.0% inhibition of the conjugate control for sheep, goats and both species, respectively. The DSns of the cELISA for sheep, goats and both species at these cut-off values were 89.2% (95% confidence interval 87.1-91.1%), 74.0% (95% CI 71.4-76.5%) and 77.9% (95% CI 76.1-79.7%), whereas DSps were 96.4% (95% CI 95.2-97.4%), 92.9% (95% CI 91.1-94.3%) and 97.2% (95% CI 96.4-97.8%), respectively. Compared to cELISA, indirect ELISA and fluorescence polarisation assay have higher DSns and DSps. However, the results obtained with the cELISA were in good agreement with those of the complement fixation test (CFT) under field conditions using 5735 sheep and goat sera. The cELISA can be used as an alternative to the CFT for diagnosing B. melitensis infection in small ruminants.

  16. Human to Human Transmission of Brucella Melitensis

    Directory of Open Access Journals (Sweden)

    Patrice Vigeant

    1995-01-01

    Full Text Available Human brucellosis is acquired mainly through contact with infected animal tissues, ingestion of unpasteurized dairy products or infected aerosols. Person to person transmission is still considered uncertain. The case of a woman diagnosed with proven brucellosis after her husband suffered a relapse of bacteremia with Brucella melitensis biotype 3, which was originally acquired abroad by eating goat cheese, is described. It was postulated that person to person spread of brucellosis is a likely mode of transmission in this case.

  17. Human to Human Transmission of Brucella Melitensis

    OpenAIRE

    Patrice Vigeant; Jack Mendelson; Miller, Mark A.

    1995-01-01

    Human brucellosis is acquired mainly through contact with infected animal tissues, ingestion of unpasteurized dairy products or infected aerosols. Person to person transmission is still considered uncertain. The case of a woman diagnosed with proven brucellosis after her husband suffered a relapse of bacteremia with Brucella melitensis biotype 3, which was originally acquired abroad by eating goat cheese, is described. It was postulated that person to person spread of brucellosis is a likely ...

  18. Deep-sequencing analysis of the mouse transcriptome response to infection with Brucella melitensis strains of differing virulence.

    Science.gov (United States)

    Wang, Fangkun; Hu, Sen; Liu, Wenxing; Qiao, Zujian; Gao, Yuzhe; Bu, Zhigao

    2011-01-01

    Brucella melitensis is an important zoonotic pathogen that causes brucellosis, a disease that affects sheep, cattle and occasionally humans. B. melitensis strain M5-90, a live attenuated vaccine cultured from B. melitensis strain M28, has been used as an effective tool in the control of brucellosis in goats and sheep in China. However, the molecular changes leading to attenuated virulence and pathogenicity in B. melitensis remain poorly understood. In this study we employed the Illumina Genome Analyzer platform to perform genome-wide digital gene expression (DGE) analysis of mouse peritoneal macrophage responses to B. melitensis infection. Many parallel changes in gene expression profiles were observed in M28- and M5-90-infected macrophages, suggesting that they employ similar survival strategies, notably the induction of anti-inflammatory and antiapoptotic factors. Moreover, 1019 differentially expressed macrophage transcripts were identified 4 h after infection with the different B. melitensis strains, and these differential transcripts notably identified genes involved in the lysosome and mitogen-activated protein kinase (MAPK) pathways. Further analysis employed gene ontology (GO) analysis: high-enrichment GOs identified endocytosis, inflammatory, apoptosis, and transport pathways. Path-Net and Signal-Net analysis highlighted the MAPK pathway as the key regulatory pathway. Moreover, the key differentially expressed genes of the significant pathways were apoptosis-related. These findings demonstrate previously unrecognized changes in gene transcription that are associated with B. melitensis infection of macrophages, and the central signaling pathways identified here merit further investigation. Our data provide new insights into the molecular attenuation mechanism of strain M5-90 and will facilitate the generation of new attenuated vaccine strains with enhanced efficacy.

  19. Deep-sequencing analysis of the mouse transcriptome response to infection with Brucella melitensis strains of differing virulence.

    Directory of Open Access Journals (Sweden)

    Fangkun Wang

    Full Text Available Brucella melitensis is an important zoonotic pathogen that causes brucellosis, a disease that affects sheep, cattle and occasionally humans. B. melitensis strain M5-90, a live attenuated vaccine cultured from B. melitensis strain M28, has been used as an effective tool in the control of brucellosis in goats and sheep in China. However, the molecular changes leading to attenuated virulence and pathogenicity in B. melitensis remain poorly understood. In this study we employed the Illumina Genome Analyzer platform to perform genome-wide digital gene expression (DGE analysis of mouse peritoneal macrophage responses to B. melitensis infection. Many parallel changes in gene expression profiles were observed in M28- and M5-90-infected macrophages, suggesting that they employ similar survival strategies, notably the induction of anti-inflammatory and antiapoptotic factors. Moreover, 1019 differentially expressed macrophage transcripts were identified 4 h after infection with the different B. melitensis strains, and these differential transcripts notably identified genes involved in the lysosome and mitogen-activated protein kinase (MAPK pathways. Further analysis employed gene ontology (GO analysis: high-enrichment GOs identified endocytosis, inflammatory, apoptosis, and transport pathways. Path-Net and Signal-Net analysis highlighted the MAPK pathway as the key regulatory pathway. Moreover, the key differentially expressed genes of the significant pathways were apoptosis-related. These findings demonstrate previously unrecognized changes in gene transcription that are associated with B. melitensis infection of macrophages, and the central signaling pathways identified here merit further investigation. Our data provide new insights into the molecular attenuation mechanism of strain M5-90 and will facilitate the generation of new attenuated vaccine strains with enhanced efficacy.

  20. 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.

  1. First evaluation of an influenza viral vector based Brucella abortus vaccine in sheep and goats: Assessment of safety, immunogenicity and protective efficacy against Brucella melitensis infection.

    Science.gov (United States)

    Tabynov, Kaissar; Yespembetov, Bolat; Matikhan, Nurali; Ryskeldinova, Sholpan; Zinina, Nadezhda; Kydyrbayev, Zhailaubay; Assanzhanova, Nurika; Tabynov, Kairat; Renukaradhya, Gourapura J; Mukhitdinova, Gulnara; Sansyzbay, Abylai

    2016-12-25

    Previously we developed and evaluated a candidate influenza viral vector based Brucella abortus vaccine (Flu-BA) administered with a potent adjuvant Montanide Gel01 in cattle, which was found safe and highly effective. This study was aimed to establish a proof-of-concept of the efficacy of Flu-BA vaccine formulation in sheep and goats. We vaccinated sheep and goats with Flu-BA vaccine and as a positive control vaccinated a group of animals with a commercial B. melitensis Rev.1 vaccine. Clinically, both Flu-BA and Rev.1 vaccines were found safe. Serological analysis showed the animals received Flu-BA vaccine did not induce antibody response against Brucella Omp16 and L7/L12 proteins during the period of our study (56days post-initial vaccination, PIV). But observed significant antigen-specific T cell response indicated by increased lymphocyte stimulation index and enhanced secretion of IFN-γ at day 56 PIV in Flu-BA group. The Flu-BA vaccinated animals completely protected 57.1% of sheep and 42.9% of goats against B. melitensis 16M challenge. The severity of brucellosis in terms of infection index and colonization of Brucella in tissues was significantly lower in the Flu-BA group compared to negative control animals group. Nevertheless, positive control commercial Rev.1 vaccine provided strong antigen-specific T cell immunity and protection against B. melitensis 16M infection. We conclude that the Flu-BA vaccine induces a significant antigen-specific T-cell response and provides complete protection in approximately 50% of sheep and goats against B. melitensis 16M infection. Further investigations are needed to improve the efficacy of Flu-BA and explore its practical application in small ruminants.

  2. Control and eradication of Brucella melitensis infection in sheep and goats.

    Science.gov (United States)

    Blasco, José M; Molina-Flores, Baldomero

    2011-03-01

    Brucella melitensis is the main etiological agent of brucellosis in sheep and goats, and is also the main agent responsible for human brucellosis, a predominantly occupational disease related to professions in direct contact with livestock. As there is currently no viable method of preventing human brucellosis to safeguard people attention must be directed toward effectively controlling the disease in sheep and goats. This review focuses on the different strategies in different socioeconomic and epidemiologic situations that can be applied to either control or eradicate brucellosis in sheep and goats.

  3. 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.

  4. Brucella melitensis Biovar 1 and Brucella abortus S19 Vaccine Strain Infections in Milkers Working at Cattle Farms in the Khartoum Area, Sudan.

    Directory of Open Access Journals (Sweden)

    Amira E F Osman

    Full Text Available Human brucellosis is a preventable zoonoses that may become persistent, causing, if left untreated, severe localized disease. Occupational exposure to infected animals or animal products and consumption of fresh contaminated dairy are main risk factors.One hundred farmworkers employed at two cattle farms one in Khartoum North and one in Omdurman were screened for the presence of specific antibodies and seropositive workers were invited to donate a blood sample for blood culture. Molecular typing was used to characterize Brucella isolates.Ten percent of farmworkers tested seropositive and while Brucella melitensis biovar 1 was isolated from the blood of three individuals, an isolate identical to the B. abortus S19 vaccine strain was isolated from a fourth person. All four bacteremic individuals were employed as milkers and did not have obvious disease.The isolation of the highly infectious pathogen B. melitensis from seropositive workers is consistent with the notion that the pathogen may persist in the blood without causing overt disease. While vaccination with strain S19 is essential for the control of bovine brucellosis the vaccine strain may be transmitted to the human population and protective measures remain important to prevent exposure also in view of the presence of B. melitensis. To create awareness for this potentially severe disease more information on the prevalence of the pathogen in different risk groups and in livestock in the Sudan is needed.

  5. In Situ Characterization of Splenic Brucella melitensis Reservoir Cells during the Chronic Phase of Infection in Susceptible Mice.

    Directory of Open Access Journals (Sweden)

    Delphine Hanot Mambres

    Full Text Available Brucella are facultative intracellular Gram-negative coccobacilli that chronically infect humans as well as domestic and wild-type mammals, and cause brucellosis. Alternatively activated macrophages (M2a induced by IL-4/IL-13 via STAT6 signaling pathways have been frequently described as a favorable niche for long-term persistence of intracellular pathogens. Based on the observation that M2a-like macrophages are induced in the spleen during the chronic phase of B. abortus infection in mice and are strongly infected in vitro, it has been suggested that M2a macrophages could be a potential in vivo niche for Brucella. In order to test this hypothesis, we used a model in which infected cells can be observed directly in situ and where the differentiation of M2a macrophages is favored by the absence of an IL-12-dependent Th1 response. We performed an in situ analysis by fluorescent microscopy of the phenotype of B. melitensis infected spleen cells from intranasally infected IL-12p40-/- BALB/c mice and the impact of STAT6 deficiency on this phenotype. Most of the infected spleen cells contained high levels of lipids and expressed CD11c and CD205 dendritic cell markers and Arginase1, but were negative for the M2a markers Fizz1 or CD301. Furthermore, STAT6 deficiency had no effect on bacterial growth or the reservoir cell phenotype in vivo, leading us to conclude that, in our model, the infected cells were not Th2-induced M2a macrophages. This characterization of B. melitensis reservoir cells could provide a better understanding of Brucella persistence in the host and lead to the design of more efficient therapeutic strategies.

  6. Characterization of possible correlates of protective response against Brucella ovis infection in rams immunized with the B. melitensis Rev 1 vaccine.

    Science.gov (United States)

    Galindo, Ruth C; Muñoz, Pilar M; de Miguel, María J; Marin, Clara M; Blasco, José M; Gortazar, Christian; Kocan, Katherine M; de la Fuente, José

    2009-05-18

    Vaccination with the live attenuated Brucella melitensis Rev 1 vaccine is used to control ovine brucellosis caused by Brucella ovis in sheep. The objective of this study was to identify possible correlates of protective response to B. ovis infection through the characterization by microarray hybridization and real-time RT-PCR of inflammatory and immune response genes differentially expressed in rams previously immunized with B. melitensis Rev 1 and experimentally challenged with B. ovis. Gene expression profiles were compared before and after challenge with B. ovis between rams protected and those vaccinated but found infected after challenge. The TLR10, Bak and ANXI genes were expressed at higher levels in vaccinated and protected rams. These genes provide possible correlates of protective response to B. ovis infection in rams immunized with the B. melitensis Rev 1 vaccine.

  7. Observations on brucellosis due to Brucella melitensis.

    Science.gov (United States)

    SPINK, W W

    1953-01-01

    A special study was made of the problem of brucellosis due to Brucella melitensis in visits to Mexico City in 1948, to the FAO/WHO Brucellosis Centres at Montpellier (France), Florence (Italy), and Rijeka (Yugoslavia) in 1951, and to Spain in 1952. Br. melitensis infection in human beings causes more severe illness than Br. abortus infection. It develops primarily in rural communities living in close contact with goats and sheep; cattle and swine may also harbour the infection.In diagnosis, the agglutination test has proved the most satisfactory procedure; testing would be more uniformly reliable if a single antigen were used. Lack of funds and technical assistance have in many instances limited the bacteriological studies upon which a more definitive diagnosis of brucellosis depends.Antibiotics, Brucella vaccines, and colloidal preparations of gold and silver-used separately and in combination-have proved of varying therapeutic value, although response to antibiotics is less favourable than in cases of Br. abortus infection.While the drastic measures-involving the slaughter of about 10,000 sheep-taken in Slovenia, Yugoslavia, in the late 1940's, against an outbreak of brucellosis, is an inspiring example of how the disease can be eradicated, the removal of all diseased animals is rarely feasible economically. It is hoped that future research will reveal a practicable alternative in the immunization of sheep and goats against the disease.

  8. Large scale immune profiling of infected humans and goats reveals differential recognition of Brucella melitensis antigens.

    Science.gov (United States)

    Liang, Li; Leng, Diana; Burk, Chad; Nakajima-Sasaki, Rie; Kayala, Matthew A; Atluri, Vidya L; Pablo, Jozelyn; Unal, Berkay; Ficht, Thomas A; Gotuzzo, Eduardo; Saito, Mayuko; Morrow, W John W; Liang, Xiaowu; Baldi, Pierre; Gilman, Robert H; Vinetz, Joseph M; Tsolis, Renée M; Felgner, Philip L

    2010-05-04

    Brucellosis is a widespread zoonotic disease that is also a potential agent of bioterrorism. Current serological assays to diagnose human brucellosis in clinical settings are based on detection of agglutinating anti-LPS antibodies. To better understand the universe of antibody responses that develop after B. melitensis infection, a protein microarray was fabricated containing 1,406 predicted B. melitensis proteins. The array was probed with sera from experimentally infected goats and naturally infected humans from an endemic region in Peru. The assay identified 18 antigens differentially recognized by infected and non-infected goats, and 13 serodiagnostic antigens that differentiate human patients proven to have acute brucellosis from syndromically similar patients. There were 31 cross-reactive antigens in healthy goats and 20 cross-reactive antigens in healthy humans. Only two of the serodiagnostic antigens and eight of the cross-reactive antigens overlap between humans and goats. Based on these results, a nitrocellulose line blot containing the human serodiagnostic antigens was fabricated and applied in a simple assay that validated the accuracy of the protein microarray results in the diagnosis of humans. These data demonstrate that an experimentally infected natural reservoir host produces a fundamentally different immune response than a naturally infected accidental human host.

  9. Establishment of systemic Brucella melitensis infection through the digestive tract requires urease, the type IV secretion system, and lipopolysaccharide O antigen.

    Science.gov (United States)

    Paixão, Tatiane A; Roux, Christelle M; den Hartigh, Andreas B; Sankaran-Walters, Sumathi; Dandekar, Satya; Santos, Renato L; Tsolis, Renée M

    2009-10-01

    Human brucellosis is caused mainly by Brucella melitensis, which is often acquired by ingesting contaminated goat or sheep milk and cheese. Bacterial factors required for food-borne infection of humans by B. melitensis are poorly understood. In this study, a mouse model of oral infection was characterized to assess the roles of urease, the VirB type IV secretion system, and lipopolysaccharide for establishing infection through the digestive tract. B. melitensis strain 16M was consistently recovered from the mesenteric lymph node (MLN), spleen, and liver beginning at 3 or 7 day postinfection (dpi). In the gut, persistence of the inoculum was observed up to 21 dpi. No inflammatory lesions were observed in the ileum or colon during infection. Mutant strains lacking the ureABC genes of the ure1 operon, virB2, or pmm encoding phosphomannomutase were constructed and compared to the wild-type strain for infectivity through the digestive tract. Mutants lacking the virB2 and pmm genes were attenuated in the spleen (P melitensis transited rapidly through polarized enterocyte monolayers containing M-like cells; however, transit through monolayers containing only enterocytes was reduced or absent. These results indicate that B. melitensis is able to spread systemically from the digestive tract after infection, most likely through M cells of the mucosa-associated lymphoid tissue.

  10. Erythritol triggers expression of virulence traits in Brucella melitensis.

    Science.gov (United States)

    Petersen, Erik; Rajashekara, Gireesh; Sanakkayala, Neelima; Eskra, Linda; Harms, Jerome; Splitter, Gary

    2013-06-01

    Erythritol is a four-carbon sugar preferentially utilized by Brucella spp. The presence of erythritol in the placentas of goats, cows, and pigs has been used to explain the localization of Brucella to these sites and the subsequent accumulation of large amounts of bacteria, eventually leading to abortion. Here we show that Brucella melitensis will also localize to an artificial site of erythritol within a mouse, providing a potential model system to study the pathogenesis of Brucella abortion. Immunohistological staining of the sites of erythritol within infected mice indicated a higher than expected proportion of extracellular bacteria. Ensuing experiments suggested intracellular B. melitensis was unable to replicate within macrophages in the presence of erythritol and that erythritol was able to reach the site of intracellular bacteria. The intracellular inhibition of growth was found to encourage the bacteria to replicate extracellularly rather than intracellularly, a particularly interesting development in Brucella pathogenesis. To determine the effect of erythritol on expression of B. melitensis genes, bacteria grown either with or without erythritol were analyzed by microarray. Two major virulence pathways were up-regulated in response to exposure to erythritol (the type IV secretion system VirB and flagellar proteins), suggesting a role for erythritol in virulence.

  11. Effects of Opsonization and Gamma Interferon on Growth of Brucella Melitensis 16M in Mouse Peritoneal Macrophages In Vitro

    Science.gov (United States)

    2000-01-01

    SUBTITLE Effects of Opsonization and Gamma Interferon on Growth of Brucella , melitensis 16M in Mouse Peritoneal Microphages rom In Vitro 3. REPORT...with Brucella melitensis 16M treated with complement- and/or antibody-rich serum. Mouse serum rich in antibody against Brucella lipopolysaccnaride...pathogens of humans and livestock. Brucella meli- tensis usually infects sheep, goats , and camels and is the most pathogenic species for humans (1). Like

  12. Development and application of dot-enzyme-linked immunosorbent (dot-ELISA) assay for detection of Brucella melitensis and evaluation of the shedding pattern in infected goats.

    Science.gov (United States)

    Onilude, Opeyemi Mayowa; Mohd Yusoff, Sabri; Emikpe, Benjamin Obukowho; Tanko, Polycarp; Shahrom, Salisi M; Effendy, Mohammed

    2017-01-01

    Early and accurate diagnosis of Brucella melitensis is essential for the treatment and control of brucellosis both in animals and humans. The thrust for the development of a rapid diagnostic technique to overcome the limitations of conventional microbiological and serological tests brought about this investigation on the development and application of dot-ELISA for antigen and antibody detection in infected goats. Fifteen apparently healthy Boer aged 2-3 years which tested negative for brucellosis using PCR and ELISA, were grouped into A (10 goats infected intraocularly with 10(7) CFU of B. melitensis) and B (5 goats) as control. Discharges (ocular, nasal, and vaginal) and blood were collected at days 3, 7, 10, 14, weekly until 42 post-infection (pi) for dot-ELISA, PCR, and RBPT. Dot-ELISA detected B. melitensis antigen and antibody in group A at day 3 and 7 pi, respectively with adequate sensitivity and specificity relative to PCR and RBPT. The bacteria shedding detected from discharges at day 3 pi in the nasal and ocular route with dot-ELISA. Group B were consistently negative. Values such as speed, simplicity, field adaptability, high sensitivity, and specificity make dot-ELISA a rapid and adequate technique for diagnosis of brucellosis in B. melitensis infected goats within few hours.

  13. Apyrexic Brucella melitensis aortic valve endocarditis.

    Science.gov (United States)

    al-Mudallal, D S; Mousa, A R; Marafie, A A

    1989-10-01

    The case of a young shepherd with Brucella melitensis aortic valve endocarditis is presented. His illness ran an afebrile course and was also complicated by disseminated intravascular coagulation (DIC), nephritis, hepatitis and peritonitis, all of which responded well to supportive measures and a combination of tetracycline, trimethoprim-sulphamethoxazole and amikacin sulphate. The fact that even the most severe case of brucellosis can present without fever is stressed.

  14. Brucella melitensis: a rarely suspected cause of infections of genitalia and the lower urinary tract

    Directory of Open Access Journals (Sweden)

    K. Stamatiou

    2009-04-01

    Full Text Available We examined the clinical presentation and outcome of Brucellar infections of genitalia and the lower urinary tract through a review of the medical records of 10 cases of male patients with brucellar infections of the genitalia and lower urinary tract. The mean age of the patients with brucellosis was 49.2, (median 52, range 15-77 years. Eleven out of 17 patients were rural residents, 15 reported that they might have consumed unpasteurized dairy products and four reported occupational exposure. Symptoms onset was acute in almost all cases. Scrotal pain, epidedimal swelling and fever were the most common symptoms. The Wright test was positive in 13 patients, while Brucella sp. was isolated from blood cultures in six cases. Only two patients were found with abnormal liver ultrasonography. All patients underwent treatment with doxycycline and aminoglycoside for seven days and doxycycline alone for two months. Most of them responded to antibiotic therapy with rapid regression of symptoms. One patient failed to respond to therapy and presented necrotizing orchitis, as well as abscesses, which required orchectomy. Brucellar infections of the genitalia and lower urinary tract have no specific clinical presentation; the usual laboratory examination is not sufficient to diagnose this kind of infection, therefore it could easily be misdiagnosed. An analytical medical history (including overall dietary habits and recent consumption of non-pasteurized dairy products could indicate Brucelosis as would the persistence of symptoms despite a one-week antibiotic treatment. In general, patients afflicted by brucellar epididymoorchitis respond to Brucellosis antibiotic therapy, except for some rare cases that present necrotizing orchitis and require surgical treatment.

  15. 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

  16. 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

    2014-01-01

    ABSTRACT 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. PMID:25028426

  17. Serial Kinetics of the Antibody Response against the Complete Brucella melitensis ORFeome in Focal Vertebral Brucellosis

    OpenAIRE

    Cannella, Anthony P.; Lin, Jennifer C.; Liang, Li; Atluri, Vidya; Gotuzzo, Eduardo; Felgner, Philip L; Tsolis, Renee M.; Vinetz, Joseph M.

    2012-01-01

    Human brucellosis is a common zoonosis worldwide. Here we present a case of focal vertebral brucellosis in a 71-year-old Mexican-American woman who contracted infection from unpasteurized goat milk. Standard agglutination serology was negative; the diagnosis was established by the isolation of Brucella melitensis from abscess fluid. A B. melitensis protein microarray comprised of nearly all proteins encoded by the bacterial genome was used to determine the kinetics of this patient's antibody ...

  18. Shedding Rates and SeroPrevalence of Brucella melitensis in Lactating Goats of Shahrekord, Iran

    OpenAIRE

    Ebrahimi, Azizollah; Milan, Jalal Sheykh kanluye; Mahzoonieh, Mohamad Reza; Khaksar, Khadijeh

    2014-01-01

    Background: Brucellosis remains a major worldwide zoonosis. Caprine brucellosis is a significant problem for both public health and animal production. Brucella melitensis causes disease in goats, sheep, humans, and occasionally cattle. Transmission is by ingestion or contact with infected materials, vaginal discharge, or milk. Objectives: The current study aimed to determine the rate of B. melitensis seropositives and its probable shedding in lactating goats from flocks in Shahrekord district...

  19. 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.

  20. Analysis of macrophage apoptosis induced by Brucella melitensis and the effects of caspases 3,8 and 9

    Institute of Scientific and Technical Information of China (English)

    任晓莉

    2013-01-01

    Objective To determine the difference of macrophage RAW264.7 apoptosis induced by Brucella melitensis virulent strain 16M and attenuated strain M5-90 and elucidate the regulatory role of caspases 3,8 and 9.Methods The best multiplicity of infection (MOI) was determined through kinetic analysis of Brucella melitensis strain 16M and M5-90 induced mouse macrophages apop-

  1. MLVA16 typing of Portuguese human and animal Brucella melitensis and Brucella abortus isolates.

    Science.gov (United States)

    Ferreira, Ana Cristina; Chambel, Lélia; Tenreiro, Tania; Cardoso, Regina; Flor, Lídia; Dias, Isabel Travassos; Pacheco, Teresa; Garin-Bastuji, Bruno; Le Flèche, Philippe; Vergnaud, Gilles; Tenreiro, Rogério; de Sá, Maria Inácia Corrêa

    2012-01-01

    To investigate the epidemiological relationship of isolates from different Portuguese geographical regions and to assess the diversity among isolates, the MLVA16(Orsay) assay (panels 1, 2A and 2B) was performed with a collection of 126 Brucella melitensis (46 human and 80 animal isolates) and 157 B. abortus field isolates, seven vaccine strains and the representative reference strains of each species. The MLVA16(Orsay) showed a similar high discriminatory power (HGDI 0.972 and 0.902) for both species but panel 1 and 2A markers displayed higher diversity (HGDI 0.693) in B. abortus compared to B. melitensis isolates (HGDI 0.342). The B. melitensis population belong to the "Americas" (17%) and "East Mediterranean" (83%) groups. No isolate belonged to the "West Mediterranean" group. Eighty-five percent of the human isolates (39 in 46) fit in the "East-Mediterranean" group where a single lineage known as MLVA11 genotype 116 is responsible for the vast majority of Brucella infections in humans. B. abortus isolates formed a consistent group with bv1 and bv3 isolates in different clusters. Four MLVA11 genotypes were observed for the first time in isolates from S. Jorge and Terceira islands from Azores. From the collection of isolates analysed in this study we conclude that MLVA16(Orsay) provided a clear view of Brucella spp. population, confirming epidemiological linkage in outbreak investigations. In particular, it suggests recent and ongoing colonisation of Portugal with one B. melitensis lineage usually associated with East Mediterranean countries.

  2. MLVA16 typing of Portuguese human and animal Brucella melitensis and Brucella abortus isolates.

    Directory of Open Access Journals (Sweden)

    Ana Cristina Ferreira

    Full Text Available To investigate the epidemiological relationship of isolates from different Portuguese geographical regions and to assess the diversity among isolates, the MLVA16(Orsay assay (panels 1, 2A and 2B was performed with a collection of 126 Brucella melitensis (46 human and 80 animal isolates and 157 B. abortus field isolates, seven vaccine strains and the representative reference strains of each species. The MLVA16(Orsay showed a similar high discriminatory power (HGDI 0.972 and 0.902 for both species but panel 1 and 2A markers displayed higher diversity (HGDI 0.693 in B. abortus compared to B. melitensis isolates (HGDI 0.342. The B. melitensis population belong to the "Americas" (17% and "East Mediterranean" (83% groups. No isolate belonged to the "West Mediterranean" group. Eighty-five percent of the human isolates (39 in 46 fit in the "East-Mediterranean" group where a single lineage known as MLVA11 genotype 116 is responsible for the vast majority of Brucella infections in humans. B. abortus isolates formed a consistent group with bv1 and bv3 isolates in different clusters. Four MLVA11 genotypes were observed for the first time in isolates from S. Jorge and Terceira islands from Azores. From the collection of isolates analysed in this study we conclude that MLVA16(Orsay provided a clear view of Brucella spp. population, confirming epidemiological linkage in outbreak investigations. In particular, it suggests recent and ongoing colonisation of Portugal with one B. melitensis lineage usually associated with East Mediterranean countries.

  3. 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.

  4. Draft Genome Sequences of Five Clinical Strains of Brucella melitensis Isolated from Patients Residing in Kuwait

    Science.gov (United States)

    Khan, Mohd Wasif; Habibi, Nazima; Shaheed, Faraz

    2016-01-01

    Human brucellosis is a neglected and underrecognized infection of widespread geographic distribution. Brucellosis is present on all inhabited continents and endemic in many areas of the world, including Kuwait and the Middle East. Here, we present draft genome assemblies of five Brucella melitensis strains isolated from brucellosis patients in Kuwait. PMID:27811090

  5. 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.

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

    Science.gov (United States)

    Azam, Sarwar; Rao, Sashi Bhushan; Jakka, Padmaja; NarasimhaRao, Veera; Bhargavi, Bindu; Gupta, Vivek Kumar

    2016-01-01

    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. PMID:27525259

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

    Science.gov (United States)

    Azam, Sarwar; Rao, Sashi Bhushan; Jakka, Padmaja; NarasimhaRao, Veera; Bhargavi, Bindu; Gupta, Vivek Kumar; Radhakrishnan, Girish

    2016-01-01

    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.

  8. The invA gene of Brucella melitensis is involved in intracellular invasion and is required to establish infection in a mouse model.

    Science.gov (United States)

    Alva-Pérez, Jorge; Arellano-Reynoso, Beatriz; Hernández-Castro, Rigoberto; Suárez-Güemes, Francisco

    2014-05-15

    Some of the mechanisms underlying the invasion and intracellular survival of B. melitensis are still unknown, including the role of a subfamily of NUDIX enzymes, which have been described in other bacterial species as invasins and are present in Brucella spp. We have generated a mutation in the coding gene of one of these proteins, the invA gene (BMEI0215) of B. melitensis strain 133, to understand its role in virulence. HeLa cell invasion results showed that mutant strain survival was decreased 5-fold compared with that of the parental strain at 2 h pi (Pgoat macrophage infection assay, mutant strain replication was 8-fold less than in the parental strain at 24 h pi (P<0.001); yet, at 48 h pi, no significant differences in intracellular replication were observed. Additionally, colocalization of the invA mutant with calregulin was significantly lower at 24 h pi compared with that of the parental strain. Furthermore, the mutant strain exhibited a low level of colocalization with cathepsin D, which was similar to the parental strain colocalization at 24 h pi. In vivo infection results demonstrated that spleen colonization was significantly lower with the mutant than with the parental strain. The immune response, measured in terms of antibody switching and IFN-γ transcription, was similar for Rev1 and infection with the mutant, although it was lower than the immune response elicited by the parental strain. Consequently, these results indicate that the invA gene is important during invasion but not for intracellular replication. Additionally, mutation of the invA gene results in in vivo attenuation.

  9. Immunogenicity and protective efficacy of Brucella abortus recombinant protein cocktail (rOmp19+rP39) against B. abortus 544 and B. melitensis 16M infection in murine model.

    Science.gov (United States)

    Tadepalli, Ganesh; Singh, Amit Kumar; Balakrishna, Konduru; Murali, Harishchandra Sripathy; Batra, Harsh Vardhan

    2016-03-01

    In this study, the immunogenicity and protective efficacy of recombinant proteins Omp19 (rO) and P39 (rP) from Brucella abortus were evaluated individually and compared with the cocktail protein (rO+rP) against B. abortus 544 and Brucella melitensis 16M infection in BALB/c mouse model. Intra-peritoneal (I.P.) immunization with rO+rP cocktail developed substantially higher antibody titers predominant with Th1 mediated isotypes (IgG2a/2b). Western blot analysis using anti-rO+rP antibodies showed specific reactivity with native Omp19 (19 kDa) and P39 (39 kDa) among whole cell proteins of B. abortus and B. melitensis. Splenocytes extracted from rO+rP immunized mice induced significantly (Pabortus 544 (72.27%) and B. melitensis 16M (68.57%). On the other hand, individual anti-rO and anti-rP polysera resulted in relatively lesser protection against the pathogens (64.79%, 54.45% and 47.13%, 45.11%, respectively). Immunized group of mice when I.P. challenged with 5 × 10(4) CFU of B. abortus 544 and B. melitensis 16M were found significantly (PBrucella vaccine.

  10. Protective properties of rifampin-resistant rough mutants of Brucella melitensis.

    Science.gov (United States)

    Adone, R; Ciuchini, F; Marianelli, C; Tarantino, M; Pistoia, C; Marcon, G; Petrucci, P; Francia, M; Riccardi, G; Pasquali, P

    2005-07-01

    Vaccination against Brucella infections in animals is usually performed by administration of live attenuated smooth B. abortus strain S19 and B. melitensis strain Rev1. They are proven effective vaccines against B. abortus in cattle and against B. melitensis and B. ovis in sheep and goats, respectively. However, both vaccines have the main drawback of inducing O-polysaccharide-specific antibodies that interfere with serologic diagnosis of disease. In addition, they retain residual virulence, being a cause of abortion in pregnant animals and infection in humans. To overcome these problems, one approach is to develop defined rough mutant Brucella strains lacking O antigen of lipopolysaccharide. B. abortus rough strain RB51, a rifampin-resistant mutant of virulent strain B. abortus 2308, is used as a vaccine against B. abortus infection in cattle in some countries. However, RB51 is not effective in sheep, and there is only preliminary evidence that it is effective in goats. In this study, we tested the efficacies of six rifampin-resistant rough strains of B. melitensis in protecting BALB/c mice exposed to B. melitensis infection. The protective properties, as well as both humoral and cellular immune responses, were assessed in comparison with those provided by B. melitensis Rev1 and B. abortus RB51 vaccines. The results indicated that these rough mutants were able to induce a very good level of protection against B. melitensis infection, similar to that provided by Rev1 and superior to that of RB51, without inducing antibodies to O antigen. In addition, all B. melitensis mutants were able to stimulate good production of gamma interferon. The characteristics of these strains encourage further evaluation of them as alternative vaccines to Rev1 in primary host species.

  11. Blocking the expression of syntaxin 4 interferes with initial phagocytosis of Brucella melitensis in macrophages.

    Science.gov (United States)

    Castañeda-Ramírez, Alfredo; González-Rodríguez, Diana; Hernández-Pineda, J Aide; Verdugo-Rodríguez, Antonio

    2015-01-01

    Brucella melitensis is the Brucella species most frequently associated with brucellosis in humans. It is also the causative agent of the disease in goats and other ruminants. Although significant aspects of the pathogenesis of infection by this intracellular pathogen have been clarified, several events during invasion of host cells remain to be elucidated. In this study, infections of human macrophages from the THP-1 monocyte cell line were conducted with B. melitensis Bm133 wild-type strain and a strain of Salmonella serovar Enteritidis as a control. A multiplicity of infection of 100 was used in trials focused on defining the relative expression of syntaxin 4 (STX4), a soluble N-ethylmaleimide-sensitive factor attachment protein receptor, in the early events of phagocytosis (at 15, 30, 45, and 60 min). Immunoblot assays were also done to visualize expression of the protein in cells infected with either bacterial strain. The expression of STX4 was not significantly different in cells infected with B. melitensis strain Bm133 compared to that observed in cells infected with S. Enteritidis. When the expression of STX4 mRNA was inhibited with short or small interfering, or silencing, RNA in the THP-1 cells, the survival of B. melitensis was significantly reduced at time 0, when gentamicin treatment of cultures was begun (after 1 h of phagocytosis), and also at 2 h and 12 h after infection.

  12. Evaluation of the shedding routes and serological patterns in experimentally-induced Brucella melitensis infection in dexamethasonetreated and transport-stressed goats

    Directory of Open Access Journals (Sweden)

    Polycarp Nwunuji Tanko

    2013-07-01

    Full Text Available Aim: To identify and evaluate the shedding routes and patterns following experimentally-induced Brucella melitensis infection in dexamethasone-treated and transport-stressed goats. Materials and Methods: Twenty four healthy, adult goats were divided into 4 groups: A, B, C and D respectively. Group A was treated with dexamethasone for 8 days prior to inoculation with 10 (7 Colony Forming Units of B. melitensis via the intraocular route. Group B was transported for 3 hours prior to inoculation with a similar dose. Group C was inoculated with a similar dose without subjecting the animals to any prior treatment, and this group served as our positive control. Group D was not inoculated with the infective dose and served as our negative control. Blood samples along with nasal, ocular, and vaginal swabs were collected on days 0, 3, 7, 10, 14, and weekly thereafter until day 63 post inoculation (pi and were analyzed by PCR, Rose Bengal Plate Test (RBPT, and indirect ELISAtechniques. Results: The nasal, ocular and vaginal swabs tested positive for Brucellosis with PCR from day 7, with nasal route being the first and most consistent route to reveal the positive results. Group B showed the earliest onset of shedding the bacterium (day 7 followed by group Awhich started from day 10 and shed relatively more positive of the bacterium via the routes examined. Blood samples tested positive with PCR from day 7 through 14 and the results were inconsistent subsequently. Sera samples tested positive with RBPT on day 14 in all the 3 infected groups but more consistent in group C. On the other hand, tests using ELISAshowed positive results from day 7 pi, with group C having a 100% seroconvertion –while groups Aand B showed only 50% seroconvertion. Conclusion: The consistent shedding via the nasal, ocular, and vaginal routes in groups A and B implied possible immunosuppression in the infected animals. We recommend that programs designed to control Brucellosis should

  13. Draft Genome Sequence of the Field Isolate Brucella melitensis Strain Bm IND1 from India.

    Science.gov (United States)

    Rao, Sashi Bhushan; Gupta, Vivek K; Kumar, Mukesh; Hegde, Nagendra R; Splitter, Gary A; Reddanna, Pallu; Radhakrishnan, Girish K

    2014-05-29

    Brucella spp. are facultative intracellular bacterial pathogens causing the zoonotic disease brucellosis. Here, we report the draft genome sequence of the Brucella melitensis strain from India designated Bm IND1, isolated from stomach contents of an aborted goat fetus.

  14. Draft Genome Sequence of the Field Isolate Brucella melitensis Strain Bm IND1 from India

    OpenAIRE

    Rao, Sashi Bhushan; Gupta, Vivek K.; Kumar, Mukesh; Hegde, Nagendra R; Splitter, Gary A.; Reddanna, Pallu; Radhakrishnan, Girish K.

    2014-01-01

    Brucella spp. are facultative intracellular bacterial pathogens causing the zoonotic disease brucellosis. Here, we report the draft genome sequence of the Brucella melitensis strain from India designated Bm IND1, isolated from stomach contents of an aborted goat fetus.

  15. Simultaneous detection and differentiates of Brucella abortus and Brucella melitensis by combinatorial PCR

    Institute of Scientific and Technical Information of China (English)

    Reza Mirnejad; Reza Hosseini Doust; Reza Kachuei; Seied Mojtaba Mortazavi; Mehdi Khoobdel; Ali Ahamadi

    2012-01-01

    Objective:To evaluate simultaneous detection and differentiates of Brucella abortus(B. abortus) and Brucella melitensis (B. melitensis) through the combinatorial PCR method. Methods:This study was designed using three primers that could simultaneously identify and differentiate two major species of pathogenic Brucella in humans and animals. Identification and differentiation of each species using the size of the PCR product were determined. To determine the specificity of the method, bacteria close to the genus Brucella were used. Finally, to confirm PCR products, In addition to the products sequence, RFLP was performed on PCR products using restriction enzymes. Results:The method of optimized combinatorial PCR in this study could simultaneously detect and differentiate B. abortus and B. melitensis with high specificity and sensitivity in clinical samples. Differentiation of species is based on the resulting bands;therefore, the band 494 bp for B. abortus and 733 bp for B. melitensis were obtained. RFLP and sequencing results confirmed PCR results. Conclusions:The results of this study shows that without routine diagnostic methods such as culture and serology tests, using the molecular method of combinatorial PCR, important species of Brucella can be simultaneously identified and differentiated in clinical samples.

  16. Selective amplification of Brucella melitensis mRNA from a mixed host-pathogen total RNA

    Directory of Open Access Journals (Sweden)

    Galindo Cristi L

    2010-09-01

    Full Text Available Abstract Background Brucellosis is a worldwide anthropozoonotic disease caused by an in vivo intracellular pathogen belonging to genus Brucella. The characterization of brucelae transcriptome's during host-pathogen interaction has been limited due to the difficulty of obtaining an adequate quantity of good quality eukaryotic RNA-free pathogen RNA for downstream applications. Findings Here, we describe a combined protocol to prepare RNA from intracellular B. melitensis in a quantity and quality suitable for pathogen gene expression analysis. Initially, B. melitensis total RNA was enriched from a host:pathogen mixed RNA sample by reducing the eukaryotic RNA..Then, to increase the Brucella RNA concentration and simultaneously minimize the contaminated host RNA in the mixed sample, a specific primer set designed to anneal to all B. melitensis ORF allows the selective linear amplification of sense-strand prokaryotic transcripts in a previously enriched RNA sample. Conclusion The novelty of the method we present here allows analysis of the gene expression profile of B. melitensis when limited amounts of pathogen RNA are present, and is potentially applicable to both in vivo and in vitro models of infection, even at early infection time points.

  17. Role of catalase in the virulence of Brucella melitensis in pregnant goats.

    Science.gov (United States)

    Gee, Jason M; Kovach, Michael E; Grippe, Vanessa K; Hagius, Sue; Walker, Joel V; Elzer, Philip H; Roop, R Martin

    2004-08-19

    An isogenic katE mutant derived from virulent Brucella melitensis 16M displays hypersensitivity to hydrogen peroxide in disk sensitivity assays but retains the capacity to colonize pregnant goats and induce abortion. These experimental findings indicate that although the sole periplasmic catalase of Brucella melitensis functions as an antioxidant, this enzyme does not play a critical role in virulence in the natural host.

  18. Genetic characterization of Brucella melitensis and Brucella abortus geographical clusters in Italy.

    Science.gov (United States)

    De Massis, Fabrizio; Garofolo, Giuliano; Cammà, Cesare; Ippoliti, Carla; Candeloro, Luca; Ancora, Massimo; Calistri, Paolo

    2015-01-01

    The genetic diversity of Brucella melitensis and Brucella abortus strains isolated in 199 cattle and sheep from 156 brucellosis outbreaks which occurred in 8 regions of Southern Italy in 2011, was determined using a Multiple-Locus Variable Number of Tandem Repeats Analysis approach. The existence of possible genetic clusters was verified through a hierarchical cluster analysis based on 'single link', which is closely related to the minimum spanning tree. The Hamming weighted distance matrix was adopted in the analysis. All calculations were performed using R and the additional libraries phangorn and Cluster. For a number of clusters, ranging from 2 to 15, the average silhouette width was calculated. The number of clusters adopted was identified according to the maximum average silhouette width. For B. abortus and B. melitensis, 6 and 11 genetic clusters were identified, respectively. Three out of 6 B. abortus clusters included the 96.7% of all B. abortus isolates. Clusters were clearly geographically separated, and this highlighted the known epidemiological links among them. Brucella melitensis genotypes resulted more heterogeneous; the 3 more representative genetic clusters included 79.7% of all B. melitensis isolates. A clear geographical clusterization of genotypes is recognizable only for 1 cluster, whereas the others are more widespread across Southern Italy. The genetic characterization of Brucella strains isolated from animals may be a useful tool to better understand the epidemiology and dissemination patterns of this pathogen through host populations.

  19. MLVA genotyping of Brucella melitensis and Brucella abortus isolates from different animal species and humans and identification of Brucella suis vaccine strain S2 from cattle in China.

    Directory of Open Access Journals (Sweden)

    Hai Jiang

    Full Text Available In China, brucellosis is an endemic disease and the main sources of brucellosis in animals and humans are infected sheep, cattle and swine. Brucella melitensis (biovars 1 and 3 is the predominant species, associated with sporadic cases and outbreak in humans. Isolates of B. abortus, primarily biovars 1 and 3, and B. suis biovars 1 and 3 are also associated with sporadic human brucellosis. In this study, the genetic profiles of B. melitensis and B. abortus isolates from humans and animals were analyzed and compared by multi-locus variable-number tandem-repeat analysis (MLVA. Among the B. melitensis isolates, the majority (74/82 belonged to MLVA8 genotype 42, clustering in the 'East Mediterranean' group. Two B. melitensis biovar 1 genotype 47 isolates, belonging to the 'Americas' group, were recovered; both were from the Himalayan blue sheep (Pseudois nayaur, a wild animal. The majority of B. abortus isolates (51/70 were biovar 3, genotype 36. Ten B. suis biovar 1 field isolates, including seven outbreak isolates recovered from a cattle farm in Inner Mongolia, were genetically indistinguishable from the vaccine strain S2, based on MLVA cluster analysis. MLVA analysis provided important information for epidemiological trace-back. To the best of our knowledge, this is the first report to associate Brucella cross-infection with the vaccine strain S2 based on molecular comparison of recovered isolates to the vaccine strain. MLVA typing could be an essential assay to improve brucellosis surveillance and control programs.

  20. Isolation and molecular characterization of Brucella melitensis from seropositive goats in Peninsula Malaysia.

    Science.gov (United States)

    Bamaiyi, P H; Hassan, L; Khairani-Bejo, S; Zainal Abidin, M; Ramlan, M; Krishnan, N; Adzhar, A; Abdullah, N; Hamidah N, H M; Norsuhanna, M M; Hashim, S N

    2012-12-01

    A study was carried out to isolate Brucella melitensis using established bacteriological and PCR techniques in Brucella seropositive goats in farms in Selangor, Negeri Sembilan, Melaka and Pulau Pinang. Brucella melitensis was isolated from 7 of 134 reactors with the highest isolation from the vaginal swabs (57.14%) followed by the spleen (28.57%), uterine fluid (14.29%). No Brucella was isolated from the lymph nodes. PCR confirmed all the seven isolates as B. melitensis and isolates were phylogenetically related to other isolates from India, Iran, and Israel but most closely related to isolates from Singapore.

  1. Brucella melitensis biotype 1 outbreak in goats in northern KwaZulu-Natal.

    Science.gov (United States)

    Reichel, R; Nel, J R; Emslie, R; Bishop, G C

    1996-06-01

    Brucella melitensis biotype 1 was confirmed in indigenous, outbred goats in three northern districts of the KwaZulu-Natal province following the diagnosis of human Malta fever in the same area. Six foci of infection were found during an extensive serological survey involving 6266 goats carried out in most of the districts of the KwaZulu-Natal province. The prevalence in the positive herds varied between 17% and 100%. The diagnosis was confirmed by culturing milk samples from serologically positive animals. Infected goats were found in only three districts (Ubombo, Ingwavuma and Pongola) and all infected herds fell within a 50-km radius.

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

    Directory of Open Access Journals (Sweden)

    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.

  3. Immunohistochemical detection of Brucella melitensis antigens in cases of naturally occurring abortions in sheep.

    Science.gov (United States)

    Ilhan, Fatma; Yener, Zabit

    2008-11-01

    Brucella melitensis, a worldwide zoonotic pathogen, is a significant cause of abortion in sheep and goats in some countries. The present study was carried out to determine, by immunohistochemistry, the presence of B. melitensis antigens in 110 naturally occurring aborted sheep fetuses. Sections of lung, liver, kidney, and spleen of each fetus were stained with immunoperoxidase to detect Brucella antigens. Brucella melitensis antigens were detected in 33 of 110 fetuses (30%). In the 33 positive cases, Brucella antigens were found in lung (25 [22.7%]), liver (21 [19%]), spleen (13 [11.8%]), and kidney (6 [5.4%]). Microscopic studies demonstrated that Brucella antigens were mainly located in the cytoplasm of macrophages and neutrophils of the lung, and in the cytoplasm of macrophages in the portal infiltrates and Kupffer cells of the liver. It was concluded that immunohistochemistry in formalin-fixed, paraffin-embedded tissues is a useful tool for the diagnosis of spontaneous ovine abortion caused by B. melitensis.

  4. Serial kinetics of the antibody response against the complete Brucella melitensis ORFeome in focal vertebral brucellosis.

    Science.gov (United States)

    Cannella, Anthony P; Lin, Jennifer C; Liang, Li; Atluri, Vidya; Gotuzzo, Eduardo; Felgner, Philip L; Tsolis, Renee M; Vinetz, Joseph M

    2012-03-01

    Human brucellosis is a common zoonosis worldwide. Here we present a case of focal vertebral brucellosis in a 71-year-old Mexican-American woman who contracted infection from unpasteurized goat milk. Standard agglutination serology was negative; the diagnosis was established by the isolation of Brucella melitensis from abscess fluid. A B. melitensis protein microarray comprised of nearly all proteins encoded by the bacterial genome was used to determine the kinetics of this patient's antibody responses to the complete collection of open reading frames existing in the genome (ORFeome). Three patterns of antibody responses against B. melitensis antigens were seen for serum samples obtained on days 0 (pretreatment), 14, 49, 100, and 180: (i) stable titers over time, (ii) a steady fall in titers, and (iii) an initial rise in titers followed by declining titers. Sera from this patient with chronic brucellosis recognized some of the same B. melitensis proteins as those recognized by sera from acute/subacute, blood culture-positive brucellosis patients but also recognized a distinct set of proteins. This study is the first to determine the kinetics of the human antibody responses to the complete repertoire of proteins encoded by a bacterial genome and demonstrates fundamentally different immunopathogenetic mechanisms between acute human brucellosis and chronic human brucellosis. While an extension of these findings to a larger patient population is necessary, these findings have important clinical and diagnostic implications and lead toward new insights into the fundamental immunopathogenesis of brucellosis.

  5. Pathogenicity and protective activity in pregnant goats of a Brucella melitensis Deltaomp25 deletion mutant.

    Science.gov (United States)

    Edmonds, M D; Cloeckaert, A; Hagius, S D; Samartino, L E; Fulton, W T; Walker, J V; Enright, F M; Booth, N J; Elzer, P H

    2002-06-01

    The Brucella melitensis mutant BM 25, which lacks the major 25 kDa outer membrane protein Omp25, has previously been found to be attenuated in the murine brucellosis model. In the present study, the capacity of the Deltaomp25 mutant to colonise and cause abortions in the caprine host was evaluated. The vaccine potential of BM 25 was also investigated in goats. Inoculation of nine pregnant goats in late gestation with the B. melitensis mutant resulted in 0/9 abortions, while the virulent parental strain, B. melitensis 16M, induced 6/6 dams to abort (Pgoats for two weeks post-infection. Owing to the ability of BM 25 to colonise both non-pregnant and pregnant adults without inducing abortions, a vaccine efficacy study was performed. Vaccination of goats prior to breeding with either BM 25 or the current caprine vaccine B. melitensis strain Rev. 1 resulted in 100 per cent protection against abortion following challenge in late gestation with virulent strain 16M (Pmelitensis Deltaomp25 mutant, BM 25, may be a safe and efficacious alternative to strain Rev. 1 when dealing with goat herds of mixed age and pregnancy status.

  6. Evaluation of novel Brucella melitensis unmarked deletion mutants for safety and efficacy in the goat model of brucellosis.

    Science.gov (United States)

    Kahl-McDonagh, Melissa M; Elzer, Philip H; Hagius, Sue D; Walker, Joel V; Perry, Quinesha L; Seabury, Christopher M; den Hartigh, Andreas B; Tsolis, Renee M; Adams, L Garry; Davis, Donald S; Ficht, Thomas A

    2006-06-12

    Pregnant goats were employed to assess unmarked deletion mutant vaccine candidates BMDeltaasp24, BMDeltacydBA, and BMDeltavirB2, as the target host species naturally infected with Brucella melitensis. Goats were assessed for the degree of pathology associated with the vaccine strains as well as the protective immunity afforded by each strain against abortion and infection after challenge with wild-type Brucella melitensis 16M. Both BMDeltaasp24 and BMDeltavirB2 were considered safe vaccine candidates in the pregnant goat model because they did not cause abortion or colonize fetal tissues. BMDeltaasp24 was isolated from the maternal tissues only, indicating a slower rate of clearance of the vaccine strain than for BMDeltavirB2, which was not isolated from any maternal or fetal tissues. Both strains were protective against abortion and against infection in the majority of pregnant goats, although BMDeltaasp24 was more efficacious than BMDeltavirB2 against challenge infection.

  7. Evaluating the virulence of a Brucella melitensis hemagglutinin gene in the caprine model.

    Science.gov (United States)

    Perry, Quinesha L; Hagius, Sue D; Walker, Joel V; Elzer, Philip H

    2010-10-01

    With the completion of the genomic sequence of Brucella melitensis 16M, a putative hemagglutinin gene was identified which is present in 16M and absent in Brucella abortus. The possibility of this hemagglutinin being a potential virulence factor was evaluated via gene replacement in B. melitensis yielding 16MΔE and expression in trans in B. abortus 2308-QAE. Utilizing the caprine brucellosis model, colonization and pathogenesis studies were performed to evaluate these strains. B. melitensis 16M hemagglutinin gene expression in trans in 2308-QAE revealed a significant (p≤0.05) increase in colonization and abortion rates when compared to B. abortus 2308, mimicking B. melitensis 16M virulence in pregnant goats. The B. melitensis disruption mutant's colonization and abortion rates demonstrated no attenuation in colonization but displayed a 28% reduction in abortions when compared to parental B. melitensis 16M.

  8. Brucella melitensis in France: persistence in wildlife and probable spillover from Alpine ibex to domestic animals.

    Directory of Open Access Journals (Sweden)

    Virginie Mick

    Full Text Available 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

  9. Asymptomatic Brucella bacteraemia and isolation of Brucella melitensis biovar 3 from human breast milk.

    Science.gov (United States)

    Celebi, Güven; Külah, Canan; Kiliç, Selçuk; Ustündağ, Gonca

    2007-01-01

    Brucellosis is a zoonotic disease and virtually all infections derived from exposure to animals or ingestion of unpasteurized dairy products. Brucellosis among family members has been reported. However, screening household members of an index case of acute brucellosis is not a routine procedure. A 10-y-old boy was diagnosed with acute brucellosis. Unpasteurized goat cheese commonly consumed within the family was thought to be the possible source of the bacteria. The family (parents, sister and brother) was screened with physical examination, serum tube agglutination test, blood cultures and routine laboratory tests. Three additional cases (parents and sister) of serological and culture proven brucellosis were detected. Two of them (mother and sister) were asymptomatic and had no clinical findings. Brucella melitensis biovar 3 was isolated from breast milk culture and from all blood cultures of 4 brucellosis cases. In conclusion, brucellosis, even with bacteraemia, can be completely asymptomatic. Consumption of raw milk products by household members is a common risk factor for brucellosis outbreak among family members. Thus, screening household members of an index case of brucellosis can expose new brucellosis cases.

  10. Indirect Enzyme-Linked Immunosorbent Assay for Detection of Brucella melitensis-Specific Antibodies in Goat Milk

    OpenAIRE

    Funk, N. D.; Tabatabai, L B; Elzer, P. H.; Hagius, S D; Martin, B. M.; Hoffman, L J

    2005-01-01

    Brucella melitensis is the cause of brucellosis in sheep and goats, which often results in abortion. Few cases of B. melitensis infection in goats have occurred in the United States over the last 25 years. However, vigilance must be maintained, as it is for the bovine milk industry, to ensure that brucellosis is not introduced into the U.S. goat population. The objective of this study was to develop a sensitive and specific indirect enzyme-linked immunosorbent assay (iELISA) for the detection...

  11. Immune reactivity of sera obtained from brucellosis patients and vaccinated-rabbits to a fusion protein from Brucella melitensis

    Directory of Open Access Journals (Sweden)

    Jafar Amani

    2015-04-01

    Full Text Available Objective(s: Brucella spp. are facultative intracellular pathogens which can stay alive and multiply in professional and nonprofessional phagocytes. Immunity against Brucella melitensis involves antigen-specific CD4+ and CD8+ T-cells activation and humoral immune responses. Due to negative aspects of live attenuated vaccines, much attention has been focused on finding Brucella-protective antigens to introduce them as potential subunit vaccine candidates. Materials and Methods: A chimeric gene encoding trigger factor (TF, Omp3148-74 and BP2687-111 fragments (TOB from B. melitensis was successfully cloned, expressed in Escherichia coliBL21-DE3 and purified by Ni-NTA agarose column. Antibodies to recombinant TOB (rTOB have been investigated in Brucella-infected human sera and a pool serum prepared from B. melitensis-vaccinated rabbits. Results: Our results showed that the immunized rabbit pool serum strongly reacted with rTOB. In addition, antibodies against rTOB were detectable in 76.5% of sera obtained from infected patients. Conclusion: These findings suggest that rTOB may provide a potential immunogenic candidate which could be considered in future vaccine studies.

  12. Brucella melitensis global gene expression study provides novel information on growth phase-specific gene regulation with potential insights for understanding Brucella:host initial interactions

    Directory of Open Access Journals (Sweden)

    Garner Harold R

    2009-05-01

    Full Text Available Abstract Background Brucella spp. are the etiological agents of brucellosis, a zoonotic infectious disease that causes abortion in animals and chronic debilitating illness in humans. Natural Brucella infections occur primarily through an incompletely defined mechanism of adhesion to and penetration of mucosal epithelium. In this study, we characterized changes in genome-wide transcript abundance of the most and the least invasive growth phases of B. melitensis cultures to HeLa cells, as a preliminary approach for identifying candidate pathogen genes involved in invasion of epithelial cells. Results B. melitensis at the late logarithmic phase of growth are more invasive to HeLa cells than mid-logarithmic or stationary growth phases. Microarray analysis of B. melitensis gene expression identified 414 up- and 40 down-regulated genes in late-log growth phase (the most invasive culture compared to the stationary growth phase (the least invasive culture. As expected, the majority of up-regulated genes in late-log phase cultures were those associated with growth, including DNA replication, transcription, translation, intermediate metabolism, energy production and conversion, membrane transport, and biogenesis of the cell envelope and outer membrane; while the down-regulated genes were distributed among several functional categories. Conclusion This Brucella global expression profile study provides novel information on growth phase-specific gene expression. Further characterization of some genes found differentially expressed in the most invasive culture will likely bring new insights into the initial molecular interactions between Brucella and its host.

  13. Identification of Brucella melitensis 16M genes required for bacterial survival in the caprine host.

    Science.gov (United States)

    Zygmunt, Michel S; Hagius, Sue D; Walker, Joel V; Elzer, Philip H

    2006-01-01

    Brucella species are gram-negative bacteria which belong to alpha-Proteobacteria family. These organisms are zoonotic pathogens that induce abortion and sterility in domestic mammals and chronic infections in humans known as Malta fever. The virulence of Brucella is dependent upon its ability to enter and colonize the cells in which it multiplies. The genetic basis of this aspect is poorly understood. Signature-tagged mutagenesis (STM) was used to identify potential Brucella virulence factors. PCR amplification has been used in place of DNA hybridization to identify the STM-generated attenuated mutants. A library of 288 Brucella melitensis 16M tagged mini-Tn5 Km2 mutants, in 24 pools, was screened for its ability to colonize spleen, lymph nodes and liver of goats at three weeks post-i.v. infection. This comparative screening identified 7 mutants (approximately 5%) which were not recovered from the output pool in goats. Some genes were known virulence genes involved in biosynthesis of LPS (lpsA gene) or in intracellular survival (the virB operon). Other mutants included ones which had a disrupted gene homologous to flgF, a gene coding for the basal-body rod of the flagellar apparatus, and another with a disruption in a gene homologous to ppk which is involved in the biosynthesis of inorganic polyphosphate (PolyP) from ATP. Other genes identified encoded factors involved in DNA metabolism and oxidoreduction metabolism. Using STM and the caprine host for screening, potential virulence determinants in B. melitensis have been identified.

  14. Skeletal Involvement of Brucella melitensis in Children: A Systematic Review

    Directory of Open Access Journals (Sweden)

    Anahita Sanaei Dashti

    2013-12-01

    Full Text Available Brucellosis is a protean disease and should be excluded in any febrile child with a constellation of symptoms such as fever, malaise, sweating, arthralgia, and joint swelling in endemic areas. Skeletal system involvement is the most common source of complaints in brucellosis. The frequency of skeletal involvement in children is 6.4% to 73.5%. There are some controversies regarding the most common sites of involvement: sacroiliac versus peripheral joints. In the vast majority of cases, peripheral joint involvement in pediatric brucellosis has a monoarticular pattern, although there is no agreement about the most commonly involved peripheral joint. In this systematic review, published articles that describe the bone involvement of Brucella melitensis, as the most prevalent kind of the microorganism in the region, in children are evaluated.

  15. CD8 Knockout Mice Are Protected from Challenge by Vaccination with WR201, a Live Attenuated Mutant of Brucella melitensis

    Directory of Open Access Journals (Sweden)

    Samuel L. Yingst

    2013-01-01

    Full Text Available CD8+ T cells have been reported to play an important role in defense against B. abortus infection in mouse models. In the present report, we use CD8 knockout mice to further elucidate the role of these cells in protection from B. melitensis infection. Mice were immunized orally by administration of B. melitensis WR201, a purine auxotrophic attenuated vaccine strain, then challenged intranasally with B. melitensis 16M. In some experiments, persistence of WR201 in the spleens of CD8 knockout mice was slightly longer than that in the spleens of normal mice. However, development of anti-LPS serum antibody, antigen-induced production of γ-interferon (IFN-γ by immune splenic lymphocytes, protection against intranasal challenge, and recovery of nonimmunized animals from intranasal challenge were similar between normal and knockout animals. Further, primary Brucella infection was not exacerbated in perforin knockout and Fas-deficient mice and these animals’ anti-Brucella immune responses were indistinguishable from those of normal mice. These results indicate that CD8+ T cells do not play an essential role as either cytotoxic cells or IFN-γ producers, yet they do participate in a specific immune response to immunization and challenge in this murine model of B. melitensis infection.

  16. Interferon-gamma responses in sheep exposed to virulent and attenuated Brucella melitensis strains.

    Science.gov (United States)

    Pérez-Sancho, Marta; Durán-Ferrer, Manuel; García-Seco, Teresa; Macías, Paula; García, Nerea; Martínez, Irene; Ruiz, Elena; Legaz, Emilio; Diez-Guerrier, Alberto; González, Sergio; Domínguez, Lucas; Álvarez, Julio

    2014-07-15

    Antibody detection is the basis of large-scale sheep brucellosis diagnosis because of its sensitivity and specificity. In contrast, information on the cellular mediated immune (CMI) response triggered after Brucella melitensis infection, a cornerstone in the protection against this pathogen, is more limited, particularly regarding the effect of the virulence of the infecting strain in the induced CMI reaction. Here, the interferon-gamma (IFN-γ) profiles evoked after exposure by different routes to virulent (H38) and attenuated (Rev.1) B. melitensis strains in 14 pregnant sheep and 87 ewe lambs, respectively, were characterized accounting for different host-related factors, and compared with their serological response and with the basal IFN-γ responses observed in 155 animals non exposed to Brucella. No significant differences in the IFN-γ response of Rev.1 vaccinated animals depending on the inoculation route was observed, in contrast with their serological results. Response in H38-challenged followed a similar trend although peaked later, and an effect of the abortion on the IFN-γ response was detected. This information could help to understand the interaction bacteria-host that leads to its intracellular survival and could be useful for the design of new diagnostic approaches.

  17. An overview of the eradication of Brucella melitensis from KwaZulu-Natal.

    Science.gov (United States)

    Emslie, F R; Nel, J R

    2002-06-01

    Brucella melitensis is a Gram-negative bacterium whose primary hosts are goats and sheep. Like the other BrucelIa spp., with the exception of Brucella ovis, it is not particularly host specific as it is pathogenic for a variety of other mammal species including humans. In humans the disease caused by it is rated as one of the most important zoonoses. Three outbreaks have been recorded in goats and sheep in South Africa; the first outbreak occurred in sheep in 1965 in the Mpumalanga and Northern Provinces (then both part of the Transvaal Province), the second occurred in sheep in 1989 near Pretoria, Gauteng Province, and the third and current outbreak was diagnosed in a flock of goats in northern KwaZulu-Natal in September 1994. Following the initial diagnosis of B. melitensis in north-eastern KwaZulu-Natal, a serological survey was conducted in order to identify foci of infection in the goat and sheep populations. Six positive foci were identified. In March 1996 a test-and-slaughter eradication campaign was initiated in these areas. Initial test results revealed a prevalence of between 1.23% and 4.02 %. All positive animals were identified and slaughtered. Eradication programmes were repeated between March 1996 and June 2000, in the populations at risk, and the disease prevalence was reduced in all the affected populations.

  18. Ovine and Caprine Brucellosis (Brucella melitensis) in Aborted Animals in Jordanian Sheep and Goat Flocks.

    Science.gov (United States)

    Samadi, Assadullah; Ababneh, M Mk; Giadinis, N D; Lafi, S Q

    2010-10-28

    Two hundred and fifty five biological samples were collected from 188 animals (81 sheep and 107 goats) during the lambing season from September 2009 to April 2010 from the Mafraq region of Jordan. Sampled animals belonged to 93 sheep and goat flocks that had abortion cases in the region. One hundred and seven (41.9%) biological samples were positive for the omp2 primers that were able to identify all Brucella species in the collected samples which were obtained from 86 aborted animals (86/188 = 45.7%). Using the B. melitensis insertion sequence 711 (IS711) primers on the 107 omp2 positive samples, only 61 confirmed to be positive for B. melitensis. These positive samples were obtained from 28 sheep and 33 goats. The prevalence rate of B. melitensis was 27.1% (51/188) among aborted animals. For differentiation between vaccine strain and field strain infection, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method using PstI endonuclease enzyme was used. Vaccination with Rev-1 in the last year (OR = 2.92, CI: 1.1-7.7) and grazing at common pasture (OR = 2.78, CI: 1.05-7.36) were statistically significant (P ≤ .05) risk factors positively associated with the occurrence of brucellosis in sheep and goat flocks.

  19. Ovine and Caprine Brucellosis (Brucella melitensis in Aborted Animals in Jordanian Sheep and Goat Flocks

    Directory of Open Access Journals (Sweden)

    Assadullah Samadi

    2010-01-01

    Full Text Available Two hundred and fifty five biological samples were collected from 188 animals (81 sheep and 107 goats during the lambing season from September 2009 to April 2010 from the Mafraq region of Jordan. Sampled animals belonged to 93 sheep and goat flocks that had abortion cases in the region. One hundred and seven (41.9% biological samples were positive for the omp2 primers that were able to identify all Brucella species in the collected samples which were obtained from 86 aborted animals (86/188=45.7%. Using the B. melitensis insertion sequence 711 (IS711 primers on the 107 omp2 positive samples, only 61 confirmed to be positive for B. melitensis. These positive samples were obtained from 28 sheep and 33 goats. The prevalence rate of B. melitensis was 27.1% (51/188 among aborted animals. For differentiation between vaccine strain and field strain infection, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP method using PstI endonuclease enzyme was used. Vaccination with Rev-1 in the last year (OR=2.92, CI: 1.1–7.7 and grazing at common pasture (OR=2.78, CI: 1.05–7.36 were statistically significant (P≤.05 risk factors positively associated with the occurrence of brucellosis in sheep and goat flocks.

  20. Indirect enzyme-linked immunosorbent assay for detection of Brucella melitensis-specific antibodies in goat milk.

    Science.gov (United States)

    Funk, N D; Tabatabai, L B; Elzer, P H; Hagius, S D; Martin, B M; Hoffman, L J

    2005-02-01

    Brucella melitensis is the cause of brucellosis in sheep and goats, which often results in abortion. Few cases of B. melitensis infection in goats have occurred in the United States over the last 25 years. However, vigilance must be maintained, as it is for the bovine milk industry, to ensure that brucellosis is not introduced into the U.S. goat population. The objective of this study was to develop a sensitive and specific indirect enzyme-linked immunosorbent assay (iELISA) for the detection of B. melitensis-specific antibodies in goat milk. Brucella salt-extractable protein extract was employed as an antigen, and a horseradish peroxidase-labeled polyclonal anti-goat antibody was used as an anti-species conjugate. Thirteen of 13 (100%) individual infected goat milk samples tested positive and 134 of 134 (100%) uninfected bulk milk samples tested negative by the developed iELISA. Three positive milk samples with high, medium, and low absorbance values were used to simulate one positive animal in an otherwise negative herd. By this estimation, one high-titer animal could be detected in a herd of >1,600 animals. Detection estimates for medium- and low-titer animals were one positive animal per herd of melitensis-specific antibodies in goat milk.

  1. The first International Standard anti-Brucella melitensis Serum.

    Science.gov (United States)

    McGiven, J; Taylor, A; Duncombe, L; Sayers, R; Albert, D; Banai, M; Blasco, J M; Elena, S; Fretin, D; Garin-Bastuji, B; Melzer, F; Muñoz, P M; Nielsen, K; Nicola, A; Scacchia, M; Tittarelli, M; Dias, I Travassos; Walravens, K; Stack, J

    2011-12-01

    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 immunosorbent assay (iELISA) 1/64 and 1/750, competitive ELISA (cELISA) 1/8 and 1/300, fluorescent polarisation assay (FPA) 1/16 and 1/200, Rose Bengal test (RBT) 1/16 and 1/200. The OIE International Standard Serum (OIEISS) will remain the primary standard for the RBT; the ISaBmS is an additional standard. It was impossible to set criteria for the complement fixation test, therefore the OIEISS will remain the primary standard. The ISaBmS can be used to standardise iELISA, cELISA and FPA to diagnose sheep and goat brucellosis. This standard should facilitate harmonisation of tests used for brucellosis surveillance and international trade in these species.

  2. 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.

  3. Comparison of serological tests for the detection of ovine and caprine antibody to Brucella melitensis.

    Science.gov (United States)

    Nielsen, K; Gall, D; Smith, P; Balsevicius, S; Garrido, F; Ferrer, M Durán; Biancifiori, F; Dajer, A; Luna, E; Samartino, L; Bermudez, R; Moreno, F; Renteria, T; Corral, A

    2004-12-01

    The indirect enzyme-linked immunosorbent assay (IELISA), the competitive enzyme-linked immunosorbent assay (CELISA) and the fluorescence polarisation assay (FPA) were evaluated with sera from sheep experimentally infected with Brucella melitensis and negative Canadian sheep. The sensitivity and specificity of the assays were as follows: IELISA: 91.7% and 97.6%, CELISA: 75.0% and 99.8% and FPA: 91.7% and 89.5%. Sera from the same experimental population were divided according to serological reaction in the rose bengal agglutination test (RBT) and the complement fixation test (CFT). Reactivity relative to the RBT positive and CFT positive sera were as follows: IELISA: 99.7%, CELISA: 93.2% and FPA: 99.1%. Since sera from goats with proven B. melitensis infection were not available, 699 sera from goats judged positive in the buffered antigen plate agglutination test (BPAT) and CFT and 982 BPAT/CFT negative Canadian goats were used. The sensitivity and specificity of the assays relative to the BPAT and CFT positive sera were: IELISA: 99.4% and 98.0%, CELISA: 95.4% and 97.1% and FPA: 92.7% and 99.8%.

  4. 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 (P<0.05) higher in the later part of the decade, in larger herd size and among the states located in the peninsula as compared to eastern Malaysia. 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.

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

    OpenAIRE

    Ayman Al-Mariri; Mazen Safi

    2013-01-01

    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 experi...

  6. Seroprevalence for Coxiella burnetii, Francisella tularensis, Brucella abortus and Brucella melitensis in Austrian adults: a cross-sectional survey among military personnel and civilians.

    Science.gov (United States)

    Tobudic, Selma; Nedomansky, Klara; Poeppl, Wolfgang; Müller, Maria; Faas, Angelus; Mooseder, Gerhard; Allerberger, Franz; Stanek, Gerold; Burgmann, Heinz

    2014-04-01

    The prevalence of Coxiella burnetii, Francisella tularensis, Brucella abortus, and Brucella melitensis infections in Austria and the exposure risk of military personnel were assessed in an exploratory nationwide cross-sectional seroprevalence survey in 526 healthy adult individuals, 222 of which were soldiers and 304 were civilians. Screening for IgA/IgG antibodies to C. burnetii (Phase I) and IgG/IgM antibodies to C. burnetii (Phase II), and to F. tularensis was done with commercial enzyme-linked immunosorbent assays. To detect antibodies against B. abortus and B. melitensis, an in-house complement fixation test was used. Overall, 11 individuals (2.0%) showed antibodies to C. burnetii, 3 individuals (0.5%) were seropositive for F. tularensis, and one (0.3%) individual was borderline positive. All individuals positive or borderline for F. tularensis tested negative for antibodies against C. burnetii. All individuals tested negative for antibodies against B. melitensis/B. abortus. There were no significant differences between the seroprevalence of C. burnetii and F. tularensis among military personnel and civilians. Our data demonstrate serological evidence of a low rate of exposure to C. burnetii and F. tularensis among the Austrian adult population and military personnel.

  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. Brucellosis vaccines: assessment of Brucella melitensis lipopolysaccharide rough mutants defective in core and O-polysaccharide synthesis and export.

    Directory of Open Access Journals (Sweden)

    David González

    Full Text Available BACKGROUND: The brucellae are facultative intracellular bacteria that cause brucellosis, one of the major neglected zoonoses. In endemic areas, vaccination is the only effective way to control this disease. Brucella melitensis Rev 1 is a vaccine effective against the brucellosis of sheep and goat caused by B. melitensis, the commonest source of human infection. However, Rev 1 carries a smooth lipopolysaccharide with an O-polysaccharide that elicits antibodies interfering in serodiagnosis, a major problem in eradication campaigns. Because of this, rough Brucella mutants lacking the O-polysaccharide have been proposed as vaccines. METHODOLOGY/PRINCIPAL FINDINGS: To examine the possibilities of rough vaccines, we screened B. melitensis for lipopolysaccharide genes and obtained mutants representing all main rough phenotypes with regard to core oligosaccharide and O-polysaccharide synthesis and export. Using the mouse model, mutants were classified into four attenuation patterns according to their multiplication and persistence in spleens at different doses. In macrophages, mutants belonging to three of these attenuation patterns reached the Brucella characteristic intracellular niche and multiplied intracellularly, suggesting that they could be suitable vaccine candidates. Virulence patterns, intracellular behavior and lipopolysaccharide defects roughly correlated with the degree of protection afforded by the mutants upon intraperitoneal vaccination of mice. However, when vaccination was applied by the subcutaneous route, only two mutants matched the protection obtained with Rev 1 albeit at doses one thousand fold higher than this reference vaccine. These mutants, which were blocked in O-polysaccharide export and accumulated internal O-polysaccharides, stimulated weak anti-smooth lipopolysaccharide antibodies. CONCLUSIONS/SIGNIFICANCE: The results demonstrate that no rough mutant is equal to Rev 1 in laboratory models and question the notion that

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

    Directory of Open Access Journals (Sweden)

    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.

  10. Brucella Melitensis 16M Regulates the Effect of AIR Domain on Inflammatory Factors, Autophagy, and Apoptosis in Mouse Macrophage through the ROS Signaling Pathway

    Science.gov (United States)

    Li, Tiansen; Xu, Yafang; Liu, Laizhen; Huang, Meiling; Wang, Zhen; Tong, Zhixia; Zhang, Hui; Guo, Fei; Chen, Chuangfu

    2016-01-01

    Brucellosis is a highly contagious zoonosis caused by Brucella. Brucella can invade and persist inside host cells, which results in chronic infection. We constructed AIR interference and overexpression lentiviruses to acquire AIR interference, overexpression, and rescue stable expression cell lines. We also established a Brucella melitensis 16M-infected macrophage model, which was treated with either the vehicle control or NAC (ROS scavenger N-acetylcysteine (NAC) for 0, 3, 6, 12, and 24 h. Confocal laser microscopy, transmission electron microscopy, fluorescence quantitative PCR, flow cytometry, ELISA, and Western blot were used to detect inflammation, cell autophagy and apoptosis-related protein expression levels, ROS levels, and the distribution of mitochondria. It was found that after interference and overexpression of AIR, ROS release was significantly changed, and mitochondria became abnormally aggregated. B. melitensis 16M activated the NLRP3/AIM2 inflammatory complex, and induced RAW264.7 cells to secrete IL-1β and IL-18 through the ROS pathway. B. melitensis 16M also altered autophagy-related gene expression, increased autophagy activity, and induced cell apoptosis through the ROS pathway. The results showed that after B. melitensis 16M infection, ROS induced apoptosis, inflammation, and autophagy while AIR inhibited autophagosome maturation and autophagy initiation. Autophagy negatively regulated the activation of inflammasomes and prevented inflammation from occurring. In addition, mitophagy could promote cell apoptosis. PMID:27907115

  11. Detecting brucella species in Ecuador

    OpenAIRE

    Luna Jarrín, Ligia Elizabeth

    2015-01-01

    Brucelosis is an emerging zoonotic disease in many countries around the world. There are some reports of Brucella abortus infections in cattle and humans in Ecuador, nevertheless, other Brucella species have not been identified. This study was designed to identify circulating Brucella species in 300 goat samples and one canine fetus from 8 different provinces of the highland Andes of the country. The results showed isolates from Brucella melitensis, Brucella suis, Brucella abortus y Brucella ...

  12. Immunoproteomic analysis of Brucella melitensis and identification of a new immunogenic candidate protein for the development of brucellosis subunit vaccine.

    Science.gov (United States)

    Yang, Yanling; Wang, Lin; Yin, Jigang; Wang, Xinglong; Cheng, Shipeng; Lang, Xulong; Wang, Xiuran; Qu, Hailong; Sun, Chunhui; Wang, Jinglong; Zhang, Rui

    2011-10-01

    In order to screen immunogenic candidate antigens for the development of a brucellosis subunit vaccine, an immunoproteomic assay was used to identify immunogenic proteins from Brucella melitensis 16 M soluble proteins. In this study, a total of 56 immunodominant proteins were identified from the two-dimensional electrophoresis immunoblot profiles by liquid chromatography tandem mass spectrometry (LC-MS/MS). Two proteins of interest, riboflavin synthase alpha chain (RS-α) and Loraine synthase (LS-2), which are both involved in riboflavin synthesis, were detected by two-dimensional immunoblots using antisera obtained from Brucella-infected human and goats. LS-2, however, is an already well-known vaccine candidate. Therefore, we focussed our studies on the novel vaccine candidate RS-α. B. melitensis RS-α and LS-2 were then expressed in Escherichia coli as fusion proteins with His tag. The humoral and cellular immune responses to the recombinant (r)RS-α was characterized. In response to in vitro stimulation by rRS-α, splenocytes from mice vaccinated with rRS-α were able to produce γ-interferon (IFN-γ) and interleukin (IL)-2 but not interleukin (IL)-4 and interleukin (IL)-10. Furthermore, rRS-α or rLS-2-vaccinated mice were partially protected against B. melitensis infection. Our results suggested that we have developed a high-throughout, accurate, rapid and highly efficient method for the identification of candidate antigens by a combination of immunoproteomics with immunisation and bacterial challenge and rRs-α could be a useful candidate for the development of subunit vaccines against B. melitensis.

  13. 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.

  14. Rifampicin resistance phenotyping of Brucella melitensis by rpoB gene analysis in clinical isolates.

    Science.gov (United States)

    Sayan, M; Yumuk, Z; Dündar, D; Bilenoglu, O; Erdenlig, S; Yaşar, E; Willke, A

    2008-08-01

    R Rifampicin resistance of Brucella melitensis by rpoB gene analysis has not yet been performed in Turkey, where brucellosis is endemic. In this study, we investigated the efficacy of E-test and single nucleotide polymorphism (SNP) analysis of the B. melitensis rpoB gene, for the detection of mutations conferring rifampicin resistance, by sequencing 21 human B. melitensis strains from the Southeast and Marmara regions of Turkey. On CLSI slow-growing bacteria standards, all isolates were sensitive to rifampicin except for 6 which showed intermediate resistance to rifampicin. MIC(50) and MIC(90)values were 1 microg/ml and 1.5 microg/ml respectively (range 0.50 -1.5 microg/ml). The rifampicin-resistant phenotype was investigated at Cd 154 (GTT/TTT), Cd 526 (GAC/TAC, GAC/AAC, GAC/GGC), Cd 536 (CAC/CTC, CAC/TAC), Cd 539 (CGC/AGC), Cd 541 (TCG/TTG) and Cd 574 (CCG/CTG) of the rpoB gene in B. melitensis 16M and B115 strains, and in clinical isolates. No missense mutations were found in any of the B. melitensis isolates, which indicates that all isolates were rifampicin-susceptible. In conclusion, SNP analysis was useful as a molecular tool for rifampin resistance testing. Although resistance to rifampicin was not detected in our strains of B. melitensis; the presence of strains with intermediate resistance to rifampicin indicates that susceptibility testing should be performed periodically.

  15. Improved performance of Brucella melitensis native hapten over Brucella abortus OPS tracer on goat antibody detection by the fluorescence polarization assay.

    Science.gov (United States)

    Ramírez-Pfeiffer, C; Díaz-Aparicio, E; Rodríguez-Padilla, C; Morales-Loredo, A; Alvarez-Ojeda, G; Gomez-Flores, R

    2008-06-15

    The current method for goat brucellosis diagnosis is based on the World Organization for Animal Health (OIE) using the screening card test (CT), with antigen at 8% (CT8) or 3% (CT3) of cell concentrations, and the confirmatory complement fixation test (CFT). However, these tests do not differentiate antibodies induced by vaccination from those derived from field infections by Brucella species or other bacterial agents; in places like Mexico, where the prevalence of brucellosis and the vaccination rates are high, there is a considerable percentage of false positive reactions that causes significant unnecessary slaughter of animals. Furthermore, results of the fluorescence polarization assay (FPA) using the Brucella abortus O-polysaccharide (OPS) tracer in goats are poorer than those with cattle. The present study was undertaken to investigate a tracer prepared from the native hapten (NH) of the Rev. 1 strain of Brucella melitensis to improve FPA performance on goat brucellosis diagnosis. Evaluation of 48 positive samples and 96 negative samples showed that the NH tracer was more accurate (pgoat sera samples selected by test series approved by the OIE (card test 3% and CFT). We demonstrated a new application for the NH lipopolysaccharide on detecting antibodies against Brucella using the FPA, which may yield faster results (minutes vs. 24-72h) than the immunodiagnosis assays frequently used in bovine brucellosis. In addition, NH tracer produces similar or better performance results than the conventional OPS tracer, using the FPA in goat sera samples.

  16. Evaluation of different enzyme-linked immunosorbent assays for the diagnosis of brucellosis due to Brucella melitensis in sheep.

    Science.gov (United States)

    García-Bocanegra, Ignacio; Allepuz, Alberto; Pérez, Julio José; Alba, Anna; Giovannini, Armando; Arenas, Antonio; Candeloro, Luca; Pacios, Alberto; Saez, José Luís; González, Miguel Ángel

    2014-03-01

    Six serological assays for the diagnosis of ovine brucellosis, due to Brucella melitensis were evaluated. Reference serum samples from sheep of known B. melitensis infection status (n=118) were assessed using the Rose Bengal test (RBT), complement fixation test (CFT) and four commercial enzyme-linked immunosorbent assays (ELISAs), including two indirect ELISAs (iELISAs), one competitive ELISA (cELISA) and one blocking ELISA (bELISA). The highest differential positive rates (DPR) were obtained with the cELISA and bELISA, while the lowest DPR was estimated using iELISAs. A latent class analysis was performed to estimate the accuracy of the CFT, RBT and bELISA using 1827 sera from sheep undergoing testing as part of a surveillance and control programme. Lower sensitivity and specificity were obtained for the three serological tests when the field samples were used. A higher DPR was achieved by the CFT, compared to bELISA and RBT. The results suggest that ELISAs, and particularly the bELISA, might be suitable for inclusion in the European Union legislation on intra-community trade for diagnosing B. melitensis infection in sheep, as it has a similar test performance compared to the RBT.

  17. Complete genome sequences of Brucella melitensis strains M28 and M5-90, with different virulence backgrounds.

    Science.gov (United States)

    Wang, Fangkun; Hu, Sen; Gao, Yuzhe; Qiao, Zujian; Liu, Wenxing; Bu, Zhigao

    2011-06-01

    Brucella melitensis is a Gram-negative coccobacillus bacteria belonging to the Alphaproteobacteria subclass. It is an important zoonotic pathogen that causes brucellosis, a disease affecting sheep, cattle, and sometimes humans. The B. melitensis strain M5-90, a live attenuated vaccine cultured from the B. melitensis virulent strain M28, has been an effective tool to control brucellosis in goats and sheep in China. Here we report the complete genome sequences of B. melitensis M28 and M5-90, strains with different virulence backgrounds, which will serve as a valuable reference for future studies.

  18. Isolation of Brucella melitensis from a human case of chronic additive polyarthritis.

    Science.gov (United States)

    Chahota, R; Dattal, A; Thakur, S D; Sharma, M

    2015-01-01

    Brucellar arthritis remains under diagnosed owing to non-specific clinical manifestations. Here, we report isolation of Brucella melitensis from synovial fluid of 5th metatarsophalangeal joint of a 39-year-old lady having unusually chronic asymmetric, additive, peripheral polyarthritis. This isolation was confirmed by Bruce-Ladder polymerase chain reaction (PCR). The patient had a history of contact with an aborted goat. Rose Bengal Plate Agglutination Test (RBPT) and Standard Tube Agglutination Test (SAT) were positive for Brucella-specific antibodies both for patient and in contact with sheep and goats. The patient was treated with doxycycline and rifampicin for 16 weeks and was recovered fully.

  19. Isolation of Brucella melitensis from a human case of chronic additive polyarthritis

    Directory of Open Access Journals (Sweden)

    R Chahota

    2015-01-01

    Full Text Available Brucellar arthritis remains under diagnosed owing to non-specific clinical manifestations. Here, we report isolation of Brucella melitensis from synovial fluid of 5 th metatarsophalangeal joint of a 39-year-old lady having unusually chronic asymmetric, additive, peripheral polyarthritis. This isolation was confirmed by Bruce-Ladder polymerase chain reaction (PCR. The patient had a history of contact with an aborted goat. Rose Bengal Plate Agglutination Test (RBPT and Standard Tube Agglutination Test (SAT were positive for Brucella-specific antibodies both for patient and in contact with sheep and goats. The patient was treated with doxycycline and rifampicin for 16 weeks and was recovered fully.

  20. 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.

  1. Design and implementation of a database for Brucella melitensis genome annotation.

    Science.gov (United States)

    De Hertogh, Benoît; Lahlimi, Leïla; Lambert, Christophe; Letesson, Jean-Jacques; Depiereux, Eric

    2008-03-18

    The genome sequences of three Brucella biovars and of some species close to Brucella sp. have become available, leading to new relationship analysis. Moreover, the automatic genome annotation of the pathogenic bacteria Brucella melitensis has been manually corrected by a consortium of experts, leading to 899 modifications of start sites predictions among the 3198 open reading frames (ORFs) examined. This new annotation, coupled with the results of automatic annotation tools of the complete genome sequences of the B. melitensis genome (including BLASTs to 9 genomes close to Brucella), provides numerous data sets related to predicted functions, biochemical properties and phylogenic comparisons. To made these results available, alphaPAGe, a functional auto-updatable database of the corrected sequence genome of B. melitensis, has been built, using the entity-relationship (ER) approach and a multi-purpose database structure. A friendly graphical user interface has been designed, and users can carry out different kinds of information by three levels of queries: (1) the basic search use the classical keywords or sequence identifiers; (2) the original advanced search engine allows to combine (by using logical operators) numerous criteria: (a) keywords (textual comparison) related to the pCDS's function, family domains and cellular localization; (b) physico-chemical characteristics (numerical comparison) such as isoelectric point or molecular weight and structural criteria such as the nucleic length or the number of transmembrane helix (TMH); (c) similarity scores with Escherichia coli and 10 species phylogenetically close to B. melitensis; (3) complex queries can be performed by using a SQL field, which allows all queries respecting the database's structure. The database is publicly available through a Web server at the following url: http://www.fundp.ac.be/urbm/bioinfo/aPAGe.

  2. Septic arthritis caused by Brucella melitensis in urban Shenzhen, China: a case report

    OpenAIRE

    Wong, Tak Man; Lou, Nan; Jin, Wentao; Leung, Felix; To, Michael; Leung, Frankie

    2014-01-01

    Abstract Introduction: Brucellosis is a systemic infectious disease which is still a challenging medical problem in rural areas such as northern China. It rarely occurs in urban areas such as Shenzhen in southern China. Osteoarticular involvements are frequently seen in brucellosis, and rarely is arthritis the only clinical presentation. We report a case of hip septic arthritis caused by Brucella melitensis in an urban area of Shenzhen, China. Case presentation: A 29-year-old Chinese wom...

  3. Brucella melitensis Methionyl-tRNA-Synthetase (MetRS), a Potential Drug Target for Brucellosis

    Science.gov (United States)

    Ranade, Ranae M.; Zhang, Zhongsheng; Dranow, David M.; Myers, Janette B.; Choi, Ryan; Nakazawa Hewitt, Steve; Edwards, Thomas E.; Davies, Douglas R.; Lorimer, Donald; Boyle, Stephen M.; Barrett, Lynn K.; Buckner, Frederick S.; Fan, Erkang; Van Voorhis, Wesley C.

    2016-01-01

    We investigated Brucella melitensis methionyl-tRNA-synthetase (BmMetRS) with molecular, structural and phenotypic methods to learn if BmMetRS is a promising target for brucellosis drug development. Recombinant BmMetRS was expressed, purified from wild type Brucella melitensis biovar Abortus 2308 strain ATCC/CRP #DD-156 and screened by a thermal melt assay against a focused library of one hundred previously classified methionyl-tRNA-synthetase inhibitors of the blood stage form of Trypanosoma brucei. Three compounds showed appreciable shift of denaturation temperature and were selected for further studies on inhibition of the recombinant enzyme activity and cell viability against wild type B. melitensis strain 16M. BmMetRS protein complexed with these three inhibitors resolved into three-dimensional crystal structures and was analyzed. All three selected methionyl-tRNA-synthetase compounds inhibit recombinant BmMetRS enzymatic functions in an aminoacylation assay at varying concentrations. Furthermore, growth inhibition of B. melitensis strain 16M by the compounds was shown. Inhibitor-BmMetRS crystal structure models were used to illustrate the molecular basis of the enzyme inhibition. Our current data suggests that BmMetRS is a promising target for brucellosis drug development. However, further studies are needed to optimize lead compound potency, efficacy and safety as well as determine the pharmacokinetics, optimal dosage, and duration for effective treatment. PMID:27500735

  4. Assessment of milk ring test and some serological tests in the detection of Brucella melitensis in Syrian female sheep.

    Science.gov (United States)

    Al-Mariri, Ayman; Ramadan, Lila; Akel, Rand

    2011-04-01

    Brucella melitensis infection prevalence among Syrian female sheep, to evaluate a number of serological tests and to discuss some epidemiological aspects of brucellosis, was studied. A total of 2,580 unvaccinated Syrian female sheep sera samples were tested for B. melitensis antibodies detection using four serological methods: the Rose Bengal test (RBT), the serum agglutination test (SAT), the complement fixation test (CFT) and an indirect enzyme-linked immunosorbent assay (iELISA). In addition, 2,375 milk samples were collected, then milk ring test (MRT) and bacterial isolation test were employed to evaluate the natural organism shedding. The samples were considered positive in 66%, 64%, and 60% when we employed the RBT, SAT, and iELISA tests, respectively. Whereas, the CFT test revealed the smallest number of positive samples. By using the MRT, the total prevalence of brucellosis was nearly 38% of samples. A large variation was observed concerning the studied areas, ranging from 24% in Tartous to 44% in both Damascus and Damascus rural areas. Brucella was isolated from only 677 samples out of the 2,375 female sheep milk samples.

  5. OMP31 of Brucella melitensis 16M impairs the apoptosis of macrophages triggered by TNF-α

    Science.gov (United States)

    Zhang, Ke; Wang, Hui; Guo, Fei; Yuan, Li; Zhang, Wanjiang; Wang, Yuanzhi; Chen, Chuangfu

    2016-01-01

    Outer membrane proteins (OMPs) of microorganisms play important roles in directly interacting with host cells. Brucella species inhibit the apoptosis of host cells to benefit their own intracellular survival and replication. However, the association between OMP31 of Brucella and host cell apoptosis, and the underlying mechanism are unclear. In this study, an OMP31 gene deletion mutant based on B. melitensis 16M was constructed. Following the infection of RAW264.7 cells with B. melitensis 16M or the mutant strain, colony formation, apoptosis, tumor necrosis factor (TNF)-α levels and the levels of key downstream factors of the apoptosis pathways triggered by TNF-α, namely caspase-3, −8 and −9, cytochrome c, B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were detected. The mutant strain was shown to have the same phenotype as the parent strain using traditional microbiological tests. However, the mutant strain had impaired intracellular survival, with higher levels of apoptosis and TNF-α expression in infected RAW164.7 macrophages than the parent strain. The downstream factors of apoptosis triggered by TNF-α, including increased caspase-8, −3 and −9, cytochrome c and Bax, and decreased Bcl-2, indicated that the classical and mitochondrial cell death pathways were involved. It may be concluded that OMP31 from Brucella inhibited apoptosis and benefitted the intracellular survival of this microorganism. Furthermore, TNF-α may have served as a switch triggering classical death and mitochondrial cell death pathways. PMID:27698784

  6. Main functions and taxonomic distribution of virulence genes in Brucella melitensis 16 M.

    Directory of Open Access Journals (Sweden)

    Aniel Jessica Leticia Brambila-Tapia

    Full Text Available Many virulence genes have been detected in attenuated mutants of Brucella melitensis 16 M; nevertheless, a complete report of these genes, including the main Cluster of Orthologous Groups (COG represented as well as the taxonomical distribution among all complete bacterial and archaeal genomes, has not been analyzed. In this work a total of 160 virulence genes that have been reported in attenuated mutants in B. melitensis were included and analyzed. Additionally, we obtained 250 B. melitensis randomly selected genes as a reference group for the taxonomical comparisons. The COGs and the taxonomical distribution profile for 789 nonredundant bacterial and archaeal genomes were obtained and compared with the whole-genome COG distribution and with the 250 randomly selected genes, respectively. The main COGs associated with virulence genes corresponded to the following: intracellular trafficking, secretion and vesicular transport (U; cell motility (N; nucleotide transport and metabolism (F; transcription (K; and cell wall/membrane/envelope biogenesis (M. In addition, we found that virulence genes presented a higher proportion of orthologs in the Euryarchaeota and Proteobacteria phyla, with a significant decrease in Chlamydiae, Bacteroidetes, Tenericutes, Firmicutes and Thermotogae. In conclusion, we found that genes related to specific functions are more relevant to B. melitensis virulence, with the COG U the most significant. Additionally, the taxonomical distribution of virulence genes highlights the importance of these genes in the related Proteobacteria, being less relevant in distant groups of organisms with the exception of Euryarchaeota.

  7. Brucella suis strain 2 vaccine is safe and protective against heterologous Brucella spp. infections.

    Science.gov (United States)

    Zhu, Liangquan; Feng, Yu; Zhang, Ge; Jiang, Hui; Zhang, Zhen; Wang, Nan; Ding, Jiabo; Suo, Xun

    2016-01-12

    Brucellosis is a wide spread zoonotic disease that causes abortion and infertility in mammals and leads to debilitating, febrile illness in humans. Brucella abortus, Brucella melitensis and Brucella suis are the major pathogenic species to humans. Vaccination with live attenuated B. suis strain 2 (S2) vaccine is an essential and critical component in the control of brucellosis in China. The S2 vaccine is very effective in preventing brucellosis in goats, sheep, cattle and swine. However, there are still debates outside of China whether the S2 vaccine is able to provide protection against heterologous virulent Brucella species. We investigated the residual virulence, immunogenicity and protective efficacy of the S2 vaccine in BALB/c mice by determining bacteria persistence in spleen, serum antibody response, cellular immune response and protection against a heterologous virulent challenge. The S2 vaccine was of low virulence as there were no bacteria recovered in spleen four weeks post vaccination. The vaccinated mice developed Brucella-specific IgG in 2-3 weeks, and a burst production of IFN-γ at one week as well as a two-fold increase in TNF-α production. The S2 vaccine protected mice from a virulent challenge by B. melitensis M28, B. abortus 2308 and B. suis S1330, and the S2 vaccinated mice did not develop any clinical signs or tissue damage. Our study demonstrated that the S2 vaccine is of low virulence, stimulates good humoral and cellular immunity and protects animals against infection by heterologous, virulent Brucella species.

  8. 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.

  9. Surgical management of cervical spinal epidural abscess caused by Brucella melitensis : report of two cases and review of the literature.

    Science.gov (United States)

    Ekici, Mehmet Ali; Ozbek, Zühtü; Gökoğlu, Abdülkerim; Menkü, Ahmet

    2012-06-01

    Spinal epidural abscess, if especially caused by Brucellosis is a very rare disease which is usually a consequence of spondylodiscitis. The spinal column can be affected at any joint; however, the lumbar spine is the most common region, especially at the level of the L4-5 and L5-S1. The frequency of spinal involvement usually seen at the lumbar, thoracic and cervical spine respectively. As an occupational disease in farmers, veterinaries, butchers, laboratory staff and shepherds, brucellosis can also occur by direct contact to animals and infected materials or ingestion of raw cheese, milk or unpasteurized milk products. In this study, we presented two cases with cervical spinal epidural abscess caused by brucella melitensis, which was successfully treated by surgical approach. Initial treatment was combined with antibiotic therapy after the surgery for 3 months.

  10. Proteínas inmunodominantes de Brucella Melitensis evaluadas por Western Blot

    Directory of Open Access Journals (Sweden)

    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.

  11. Brucella melitensis survival during manufacture of ripened goat cheese at two temperatures.

    Science.gov (United States)

    Méndez-González, Karla Y; Hernández-Castro, Rigoberto; Carrillo-Casas, Erika M; Monroy, Jorge F; López-Merino, Ahide; Suárez-Güemes, Francisco

    2011-12-01

    The aim of the current work was to assess the influence of two temperatures, 4°C and 24°C, on pH and water activity and their association with Brucella melitensis survival during the traditional manufacture of ripened goat cheese. Raw milk from a brucellosis-free goat herd was used for the manufacture of ripened cheese. The cheese was inoculated with 5×10(9) of the B. melitensis 16M strain during the tempering stage. The cheeses were matured for 5, 20, and 50 days at both temperatures. To assess Brucella survival, the pH and a(w) were recorded at each stage of the process (curd cutting, draining whey, immersion in brine, ripening I, ripening II, and ripening III). B. melitensis was detected at ripening stage III (1×10(3) colony-forming unit [CFU]/mL) from cheeses matured at 4°C with a pH of 5.0 and a(w) of 0.90, and at a ripening stage II (1×10(4) CFU/mL) from cheeses ripened at 24°C with a pH of 4.0 and a(w) of 0.89. The remaining stages were free from the inoculated pathogen. In addition, viable B. melitensis was recovered in significant amounts (1-2×10(6) CFU/mL) from the whey fractions of both types of cheese ripened at 24°C and 4°C. These results revealed the effects of high temperature (24°C vs. 4°C) on the low pH (4) and a(w) (0.89) that appeared to be associated with the suppression of B. melitensis at the early stages of cheese ripening. In the ripened goat cheeses, B. melitensis survived under a precise combination of temperature during maturation, ripening time, and a(w) in the manufacturing process.

  12. Genotyping of Brucella melitensis strains from dromedary camels (Camelus dromedarius) from the United Arab Emirates with multiple-locus variable-number tandem repeat analysis.

    Science.gov (United States)

    Gyuranecz, Miklós; Wernery, Ulli; Kreizinger, Zsuzsa; Juhász, Judit; Felde, Orsolya; Nagy, Péter

    2016-04-15

    Camel brucellosis is a widespread zoonotic disease in camel-rearing countries caused by Brucella melitensis and Brucella abortus. The aim of this study was the first genetic analysis of B. melitensis strains isolated from dromedary camels (Camelus dromedarius) using multiple-locus variable-number tandem repeat analysis (MLVA). MLVA 16 and its MLVA 8 and MLVA11 subsets were used to determine the genotypes of 15 B. melitensis isolates from dromedary camels (11 strains) and other host species (4 strains) from the United Arab Emirates and the results were then compared to B. melitensis MLVA genotypes from other parts of the world. Five, including two novel genotypes were identified with MLVA 8. MLVA 16 further discriminated these five genotypes to ten variants. The eleven camel isolates clustered into four main genetic groups within the East-Mediterranean and African clades and this clustering correlated with the geographic origin of the hosts (United Arab Emirates, Kingdom of Saudi Arabia and Sudan) and the date of their isolation. The camel strains were also genetically related to strains isolated from wild and domestic ruminants from their close habitat or from other parts of the world. Although limited number of strains were analysed, based on our data imported animals from foreign countries, local small ruminants and wildlife species are hypothesized to be the main sources of camel brucellosis in the United Arab Emirates. MLVA was successfully applied to determine the epidemiological links between the different camel B. melitensis infections in the United Arab Emirates and it can be a beneficial tool in future disease control programs.

  13. Draft Genome Sequence of Brucella melitensis Strain ADMAS-G1, Isolated from Placental Fluids of an Aborted Goat.

    Science.gov (United States)

    Shome, Rajeswari; Krithiga, Natesan; Muttannagouda, Revanasiddappa Biradar; Veeregowda, Belamaranahalli Muniveerappa; Swati, Sahay; Shome, Bibek Ranjan; Vishnu, Udayakumar; Sankarasubramanian, Jagadesan; Sridhar, Jayavel; Gunasekaran, Paramasamy; Rahman, Habibur; Rajendhran, Jeyaprakash

    2013-10-10

    Here, we report the draft genome sequence and annotation of the Brucella melitensis strain designated ADMAS-G1, isolated from placental fluids of an aborted goat. The length of the genome is 3,284,982 bp, with a 57.3% GC content. A total of 3,325 protein-coding genes and 63 RNA genes were predicted.

  14. Draft Genome Sequence of Brucella melitensis Strain ADMAS-G1, Isolated from Placental Fluids of an Aborted Goat

    OpenAIRE

    Shome, Rajeswari; Krithiga, Natesan; Muttannagouda, Revanasiddappa Biradar; Veeregowda, Belamaranahalli Muniveerappa; Swati, Sahay; Shome, Bibek Ranjan; Vishnu, Udayakumar; Sankarasubramanian, Jagadesan; Sridhar, Jayavel; Gunasekaran, Paramasamy; Rahman, Habibur; Rajendhran, Jeyaprakash

    2013-01-01

    Here, we report the draft genome sequence and annotation of the Brucella melitensis strain designated ADMAS-G1, isolated from placental fluids of an aborted goat. The length of the genome is 3,284,982 bp, with a 57.3% GC content. A total of 3,325 protein-coding genes and 63 RNA genes were predicted.

  15. Evaluation of the immunogenicity and safety of Brucella melitensis B115 vaccination in pregnant sheep.

    Science.gov (United States)

    Pérez-Sancho, Marta; Adone, Rosanna; García-Seco, Teresa; Tarantino, Michaela; Diez-Guerrier, Alberto; Drumo, Rosanna; Francia, Massimiliano; Domínguez, Lucas; Pasquali, Paolo; Álvarez, Julio

    2014-04-01

    In spite of its limitations, Rev.1 is currently recognized as the most suitable vaccine against Brucella melitensis (the causative agent of ovine and caprine brucellosis). However, its use is limited to young animals when test-and-slaughter programs are in place because of the occurrence of false positive-reactions due to Rev.1 vaccination. The B. melitensis B115 rough strain has demonstrated its efficacy against B. melitensis virulent strains in the mouse model, but there is a lack of information regarding its potential use in small ruminants for brucellosis control. Here, the safety and immune response elicited by B115 strain inoculation were evaluated in pregnant ewes vaccinated at their midpregnancy. Vaccinated (n=8) and non-vaccinated (n=3) sheep were periodically sampled and analyzed for the 108 days following inoculations using tests designed for the detection of the response elicited by the B115 strain and routine serological tests for brucellosis [Rose Bengal Test (RBT), Complement Fixation Test (CFT) and blocking ELISA (ELISAb)]. Five out of the 8 vaccinated animals aborted, indicating a significant abortifacient effect of B115 inoculation at midpregnancy. In addition, a smooth strain was recovered from one vaccinated animal, suggesting the occurrence of an in vivo reversion phenomenon. Only one animal was positive in both RBT and CFT simultaneously (91 days after vaccination) confirming the lack of induction of cross-reacting antibody responses interfering with routine brucellosis diagnostic tests in most B115-vaccinated animals.

  16. Amplification, cloning and expression of Brucella melitensis bp26 gene (OMP28 isolated from Markazi province (Iran and purification of Bp26 Protein

    Directory of Open Access Journals (Sweden)

    Hosseini, S.D.

    2013-12-01

    Full Text Available Brucellosis is a debilitative disease that imposes costs on both economy and society. It is shown that although the vaccine can prevent abortion, it does not provide complete protection against infection. In Iran, Brucella melitensis is a common causative agent for brucellosis and BP26 protein of this bacterium having a good antigenesity and an important vaccine candidate. In this study B. melitensis bp26 gene was cloned first in to PTZ57R/T vector and accessed on the PET28a vector and sequenced. Recombinant vector transformed and expressed in to E. coli BL21 (DE3 and then recombinant protein was purified with Ni-NTA column of chromatography against His tag. Obtained rOmp28 could be used as a research experimental tool to find its potential as a detection kit and vaccine candidate.

  17. Isolation and Identification of Brucella melitensis Biovar 3 from Vaccinated Small Ruminants: A Public Health Threat in Kosovo.

    Science.gov (United States)

    Hamidi, A; Mayer-Scholl, A; Dreshaj, S; Robaj, A; Sylejmani, D; Ramadani, N; Al Dahouk, S; Nöckler, K

    2016-12-01

    In 2011, a human brucellosis case with severe clinical symptoms was reported at the University Clinic for Infectious Diseases in Prishtina, Kosovo. A trace-back investigation was conducted to find the source of human infection. A total of 49 blood samples and 15 corresponding milk samples from sheep and goats raised on the patient's farm were taken for serological and molecular analysis. Serology using RBT and CFT revealed 11 positive animals. Twelve milk samples were PCR positive. A Brucella strain isolated from a goat's milk sample was classified as Brucella melitensis biovar 3, indicating the first ever isolation and report in Kosovo. The use of the Bruce-ladder PCR provided differentiation between the field strain and the vaccine strain. Hence, the accidental transmission of the vaccine strain Rev 1 that was previously used for the vaccination of the farm animals could be excluded. The findings of this study show that brucellosis is still a public health threat in Kosovo despite control measures.

  18. Genotyping of Brucella melitensis and Brucella abortus strains currently circulating in Xinjiang, China.

    Science.gov (United States)

    Sun, Ming-Jun; Di, Dong-Dong; Li, Yan; Zhang, Zhi-Cheng; Yan, Hao; Tian, Li-Li; Jing, Zhi-Gang; Li, Jin-Ping; Jiang, Hai; Fan, Wei-Xing

    2016-10-01

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

  19. A Study of Brucella Infection in Humans

    Directory of Open Access Journals (Sweden)

    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.

  20. 羊布氏菌强毒株16M感染小鼠骨髓树突状细胞模型的建立%Establishment of Bone Marrow-derived Dendritic Cell Model of Mice Infected with Virulent Brucella melitensi 16M Strain

    Institute of Scientific and Technical Information of China (English)

    李晓艳; 王景龙; 张瑞; 孙春辉; 郎需龙; 杨艳玲; 屈海龙; 赵勇坤; 王兴龙

    2011-01-01

    Objective To establish a bone marrow-derived dendritic cell (DC) model of mice infected with virulent Brucella melitensi 16M strain. Methods Primary bone marrow cells of C57 mice were isolated in vitro, of which the differentiation was induced by granulocyte monocyte colony stimulating factor (GM-CSF) and recombinant human interleukin-4 (rhIL-4), then observed for morphology of DCs under invert microscope, and determined for differentiation level by flow cytometry. The DC model of mice infected with virulent B. melitensi 16M strain was established in vitro, identified by IFA and transmission electron microscopy, and subjected to re-culture and staining of B. melitensi 16M strain after infection. Results On the 5th day after culture, the morphology of cells was consistent with that of bone marrow-derived DCs, with long dendritic and pseudopod-like enations. On the 7th day, the cells were semi-suspended, with a large quantity of enations all around. The purity of DCs cultured in vitro reached about 70%,which met the requirements for test. DCs showed the strongest phagocytosis and the smallest amount of intracellular bacteria 12 h after infection. Typically bacterial morphology orB. melitensi 16M strain was observed by staining after re-culture. Conclusion The bone marrow-derived DC model of mice infected with virulent B. melitensi 16M strain was successfully constructed, which laid a foundation of further study on interaction of B. melitensi and DCs as well as the mechanism of intracellular parasitism.%目的 建立羊布氏菌强毒株16M感染小鼠骨髓树突状细胞(Dendritic cell,DC)的模型.方法 体外分离C57小鼠骨髓原代细胞,经粒细胞-巨噬细胞集落刺激因子(Granulocyte monocyte colony stimulating factor,GM-CSF)和重组人白细胞介素-4(Recombinant human interleukin-4,rhIL-4)诱导分化后,倒置显微镜观察DC形态,流式细胞术鉴定其分化程度.在体外建立羊布氏菌16M感染小鼠骨髓DC模型,间接免疫荧

  1. Immunogenic and invasive properties of Brucella melitensis 16M outer membrane protein vaccine candidates identified via a reverse vaccinology approach.

    Directory of Open Access Journals (Sweden)

    Gabriel Gomez

    Full Text Available Brucella is the etiologic agent of brucellosis, one of the most common and widely distributed zoonotic diseases. Its highly infectious nature, the insidious, systemic, chronic, debilitating aspects of the disease and the lack of an approved vaccine for human use in the United States are features that make Brucella a viable threat to public health. One of the main impediments to vaccine development is identification of suitable antigens. In order to identify antigens that could potentially be used in a vaccine formulation, we describe a multi-step antigen selection approach. We initially used an algorithm (Vaxign to predict ORF encoding outer membrane proteins with antigenic determinants. Differential gene expression during acute infection and published evidence for a role in virulence were used as criteria for down-selection of the candidate antigens that resulted from in silico prediction. This approach resulted in the identification of nine Brucella melitensis outer membrane proteins, 5 of which were recombinantly expressed and used for validation. Omp22 and Hia had the highest in silico scores for adhesin probability and also conferred invasive capacity to E. coli overexpressing recombinant proteins. With the exception of FlgK in the goat, all proteins reacted to pooled sera from exposed goats, mice, and humans. BtuB, Hia and FlgK stimulated a mixed Th1-Th2 response in splenocytes from immunized mice while BtuB and Hia elicited NO release from splenocytes of S19 immunized mice. The results support the applicability of the current approach to the identification of antigens with immunogenic and invasive properties. Studies to assess immunogenicity and protective efficacy of individual proteins in the mouse are currently underway.

  2. Immunogenic and invasive properties of Brucella melitensis 16M outer membrane protein vaccine candidates identified via a reverse vaccinology approach.

    Science.gov (United States)

    Gomez, Gabriel; Pei, Jianwu; Mwangi, Waithaka; Adams, L Garry; Rice-Ficht, Allison; Ficht, Thomas A

    2013-01-01

    Brucella is the etiologic agent of brucellosis, one of the most common and widely distributed zoonotic diseases. Its highly infectious nature, the insidious, systemic, chronic, debilitating aspects of the disease and the lack of an approved vaccine for human use in the United States are features that make Brucella a viable threat to public health. One of the main impediments to vaccine development is identification of suitable antigens. In order to identify antigens that could potentially be used in a vaccine formulation, we describe a multi-step antigen selection approach. We initially used an algorithm (Vaxign) to predict ORF encoding outer membrane proteins with antigenic determinants. Differential gene expression during acute infection and published evidence for a role in virulence were used as criteria for down-selection of the candidate antigens that resulted from in silico prediction. This approach resulted in the identification of nine Brucella melitensis outer membrane proteins, 5 of which were recombinantly expressed and used for validation. Omp22 and Hia had the highest in silico scores for adhesin probability and also conferred invasive capacity to E. coli overexpressing recombinant proteins. With the exception of FlgK in the goat, all proteins reacted to pooled sera from exposed goats, mice, and humans. BtuB, Hia and FlgK stimulated a mixed Th1-Th2 response in splenocytes from immunized mice while BtuB and Hia elicited NO release from splenocytes of S19 immunized mice. The results support the applicability of the current approach to the identification of antigens with immunogenic and invasive properties. Studies to assess immunogenicity and protective efficacy of individual proteins in the mouse are currently underway.

  3. Functional characterization of the incomplete phosphotransferase system (PTS of the intracellular pathogen Brucella melitensis.

    Directory of Open Access Journals (Sweden)

    Marie Dozot

    Full Text Available BACKGROUND: In many bacteria, the phosphotransferase system (PTS is a key player in the regulation of the assimilation of alternative carbon sources notably through catabolic repression. The intracellular pathogens Brucella spp. possess four PTS proteins (EINtr, NPr, EIIANtr and an EIIA of the mannose family but no PTS permease suggesting that this PTS might serve only regulatory functions. METHODOLOGY/PRINCIPAL FINDINGS: In vitro biochemical analyses and in vivo detection of two forms of EIIANtr (phosphorylated or not established that the four PTS proteins of Brucella melitensis form a functional phosphorelay. Moreover, in vitro the protein kinase HprK/P phosphorylates NPr on a conserved serine residue, providing an additional level of regulation to the B. melitensis PTS. This kinase activity was inhibited by inorganic phosphate and stimulated by fructose-1,6 bisphosphate. The genes encoding HprK/P, an EIIAMan-like protein and NPr are clustered in a locus conserved among α-proteobacteria and also contain the genes for the crucial two-component system BvrR-BvrS. RT-PCR revealed a transcriptional link between these genes suggesting an interaction between PTS and BvrR-BvrS. Mutations leading to the inactivation of EINtr or NPr significantly lowered the synthesis of VirB proteins, which form a type IV secretion system. These two mutants also exhibit a small colony phenotype on solid media. Finally, interaction partners of PTS proteins were identified using a yeast two hybrid screen against the whole B. melitensis ORFeome. Both NPr and HprK/P were shown to interact with an inorganic pyrophosphatase and the EIIAMan-like protein with the E1 component (SucA of 2-oxoglutarate dehydrogenase. CONCLUSIONS/SIGNIFICANCE: The B. melitensis can transfer the phosphoryl group from PEP to the EIIAs and a link between the PTS and the virulence of this organism could be established. Based on the protein interaction data a preliminary model is proposed in which

  4. Detection of Brucella melitensis DNA in the milk of sheep after abortion by PCR assay Detección de ADN de Brucella melitensis mediante la prueba de PCR en muestras de leche de ovejas postaborto

    Directory of Open Access Journals (Sweden)

    Z Ilhan

    2008-01-01

    Full Text Available Laboratory diagnosis of brucellosis is generally performed by microbiological and serological methods. PCR assay is a specific and sensitive choice for the detection of different bacterial agents. An evaluation of this test was carried out for the detection of Brucella melitensis DNA in sheep milk. 102 milk samples from sheep after abortion were taken and studied using bacteriological culture, PCR and milk ring test (MRT. PCR found B. melitensis DNA in 24 (23.5% out of 102 milk samples, while only 8 (7.8% of the samples were positive to B. melitensis through direct culture. MRT found 28 (27.4% positive milk samples. The detection limit for PCR in sheep milk inoculated with B. melitensis strain 16 M was 1.7x10³-1.7x10(4 cfu/ml. PCR and MRT coincidence was 96%. The diagnostic sensitivity and specificity were determined as 100% and 81.3% respectively for PCR assay and 75% and 75% for MRT. PCR is a useful tool for a fast diagnosis of B. melitensis in sheep milk.El diagnóstico de laboratorio de brucelosis es generalmente realizado por métodos microbiológicos y serológicos. La prueba PCR es reconocida como una alternativa específica y sensible para la detección de diferentes agentes bacterianos. Se realizó una evaluación de la prueba PCR para la detección de ADN de Brucella melitensis en leche de oveja. Ciento dos muestras tomadas de ovejas postaborto fueron analizadas por métodos de cultivo bacteriológico, prueba PCR y prueba del anillo en leche (MRT. El PCR detectó ADN de B. melitensis en 24 (23,5% de 102 muestras de leche, mientras que solamente 8 (7,8% muestras de leche fueron positivas a B. melitensis por cultivo directo. El MRT detectó 28 (27,4% muestras positivas de leche. El límite de detección de B. melitensis 16 M por PCR fue de 1,7x10³-1,7x10(4 ufc/ml en leche inoculada. La concordancia entre PCR y MRT fue 96%. La sensibilidad diagnóstica y la precisión fueron determinadas como el 100% y el 81,3% respectivamente para la

  5. Survey of Omp19 immunogenicity against Brucella abortus and Brucella melitensis: influence of nanoparticulation versus traditional immunization.

    Science.gov (United States)

    Abkar, Morteza; Lotfi, Abbas Sahebghadam; Amani, Jafar; Eskandari, Khadijeh; Ramandi, Mehdi Fasihi; Salimian, Jafar; Brujeni, Gholamreza Nikbakht; Alamian, Saeed; Kamali, Mehdi; Koushki, Hamid

    2015-12-01

    Brucellosis is the most common zoonotic bacterial disease. Prevention of human brucellosis is achieved through pasteurization of dairy products, appropriate sanitation and vaccination of domestic animals against the Brucella species. B. abortus unlipidated 19 kDa outer membrane protein (U-Omp19) is a promising candidate for a subunit vaccine against brucellosis. This study investigates immunogenicity of Omp19 alone and with Freund's adjuvant (Omp19-IFA) and N-trimethyl chitosan (TMC/Omp19) nanoparticles, as well as the effect of Omp19 administration route on immunological responses and protection. The omp19 gene was expressed in E. coli BL21 (DE3). After purification, the recombinant Omp19 was loaded onto TMC nanoparticles by ionic gelation with tripolyphosphate. Particle size and loading efficiency of the nanoparticles were determined. Omp19-IFA was administered intraperitoneally while TMC/Omp19 nanoparticles were administered orally and intraperitoneally. The results indicated that intraperitoneal (i.p.) immunization by Omp19-IFA and TMC/Omp19 nanoparticles induced Th1 and Th2 immune responses, respectively, whereas oral immunization of TMC/Omp19 nanoparticles induced a mixed Th1/Th17 immune response. Moreover, oral immunization increased IgA levels in feces. Immunized mice were challenged with virulent B. melitensis 16 M and B. abortus 544. Oral immunization with TMC/Omp19 nanoparticles induced a remarkably high protection level against B. melitensis and B. abortus. The results showed that immunization route has a pivotal role in immune response polarization and protective efficiency of Omp19 antigen. Also, it was deduced that the higher protection level achieved through oral administration of TMC/Omp19 nanoparticles may be due to the elicited Th17 response.

  6. 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.

  7. Yersinia enterocolitica serotype O:9 cultured from Swedish sheep showing serologically false-positive reactions for Brucella melitensis.

    Science.gov (United States)

    Chenais, Erika; Bagge, Elisabeth; Lambertz, Susanne Thisted; Artursson, Karin

    2012-01-01

    In a herd of 20 sheep in Sweden, a country where brucellosis has never been diagnosed in sheep or goats, a total of six sheep were found serologically positive to Brucella melitensis in two different rounds of sampling. Yersinia enterocolitica serotype O:9 could at the time of the second sampling be isolated from four sheep, one of them at the same time serologically positive for B. melitensis. The article describes the case and gives some background information on brucellosis and Y. enterocolitica in general as well as a more specific description of the Swedish surveillance program for B. melitensis and the test procedures used. The problem with false-positive reactions, in particular its implications for surveillance programs in low prevalence or officially brucellosis-free countries, is discussed.

  8. Seroprevalence of brucellosis and typing of Brucella melitensis biovar 2 in lactating cows in Kuwait

    Directory of Open Access Journals (Sweden)

    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

  9. Expression and regulation of the ery operon of Brucella melitensis in human trophoblast cells

    Science.gov (United States)

    Zhang, Hui; Dou, Xiaoxia; Li, Zhiqiang; Zhang, Yu; Zhang, Jing; Guo, Fei; Wang, Yuanzhi; Wang, Zhen; Li, Tiansen; Gu, Xinli; Chen, Chuangfu

    2016-01-01

    Brucellosis is primarily a disease of domestic animals in which the bacteria localizes to fetal tissues such as embryonic trophoblast cells and fluids containing erythritol, which stimulates Brucella spp. growth. The utilization of erythritol is a characteristic of the genus Brucella. The ery operon contains four genes (eryA, eryB, eryC and eryD) for the utilization of erythritol, and plays a major role in the survival and multiplication of Brucella spp. The objective of the present study was to conduct a preliminary characterization of differential genes expression of the ery operon at several time points after Brucella infected embryonic trophoblast cells (HPT-8 cells). The result showed that the ery operon expression was higher in HPT-8 cells compared with the medium. The relative expression of eryA, eryB and eryC peaked at 2 h post-infection in HPT-8 cells, and eryD expression peaked at 3 h post-infection. The expression of eryA, eryB and eryC may be inhibited by increased eryD expression. However, the expression of the ery operon was stable in the presence of erythritol in cells. 2308Δery and 027Δery mutants of the ery operon were successfully constructed by homologous recombination, which were attenuated in RAW 264.7 murine macrophages. The characterization of the ery operon genes and their expression profiles in response to Brucella infection further contributes to our understanding of the molecular mechanisms of infection and the pathogenesis of brucellosis. PMID:27698777

  10. Identification of Brucella melitensis and Molecular Cloning of Its BP26 Gene into p ET24a(+) Vector

    Institute of Scientific and Technical Information of China (English)

    Feng Ying; Wang Yu; Zhao Shihua; Gao Wa; Zhan Shubai; Chen Wei

    2015-01-01

    Brucella spp. are pathogenic to humans and domestic animal. Nowadays,there is no effective vaccine and control strategy in China. So,it is necessary to research effective vaccines for prevention and treatment of this disease. In order to deal with these,we isolated and identified the type of Brucella in Darhan Muminggan Joint Banner and Siziwang Banner of Inner Mongolia. Totally 26 samples of sheep blood which were positive in serological test,one sample of spleen from aborted and one sample of secretion from birth canal were isolated,and the 16 S r DNA genes of positive samples were sequenced. Phylogenetic analysis proved that there were four isolates similar to B. melitensis. As an important diagnostic antigens of B. melitensis,the BP26 gene was amplified. The BP26 gene was cloned into vector p ET24a( +) and conducted sequence analysis. The BP26 gene was 900 bp,with an open reading frame of 753 bp. The homology of BP26 gene with the vaccine strain M5 was 100%,and that with S2 and A19 vaccine strains was 99. 99%. These finding supported the development of BP26-based specific serodiagnostic test and vaccine for B. melitensis in China.

  11. Unusual presentation of neurobrucellosis: a solitary intracranial mass lesion mimicking a cerebral tumor : a case of encephalitis caused by Brucella melitensis.

    Science.gov (United States)

    Erdem, Mehtap; Namiduru, Mustafa; Karaoglan, Ilkay; Kecik, Vuslat Bosnak; Aydin, Abdullah; Tanriverdi, Mustafa

    2012-10-01

    Among the diverse presentations of neurobrucellosis, solitary intracranial mass lesions are extremely rare. To the best of our knowledge, we describe here the second case of neurobrucellosis mimicking a cerebral tumor caused by Brucella melitensis. The mass lesion was clinically and radiologically indistinguishable from a brain tumor. The diagnosis was established by isolating Brucella melitensis in a blood culture and a positive Wright's agglutination test on the cerebrospinal fluid at 1:320 titers. Paraffin sections of the cerebral mass showed nongranulomatous encephalitis. We suggest that patients with an isolated intraparenchymal mass lesion with nongranulomatous encephalitis should also be studied for brucellosis in endemic areas.

  12. Isolation and Identification of Brucella melitensis%羊布氏杆菌的分离与鉴定

    Institute of Scientific and Technical Information of China (English)

    欧燎原; 谈志祥; 何世成; 黄建龙; 张朝阳; 鲁杏华; 唐小明; 汪洪冰

    2011-01-01

    A gram negative Bacillus pumilus were isolated from a orchiditis goat, it was red in differential staining Coates Pavlovsky, and grew slowly in sheep's blood agar and chocolate agar. Its 16S rDNA sequence amplification by PCR was 100% identities with Brucella melitensis in GenBank by BLAST, and was strong positive in the agglutination test between the isolated bacterium with the reference Brucella's antiserum. The two test results shows the isolated bacterium were Brucella melitensis.%从一患睾丸炎公山羊中分离到一株革兰氏阴性短小杆菌,该菌经柯兹洛夫斯基鉴别染色红色,在普通绵羊血琼脂、巧克力平板上生长缓慢,对该菌的16S rDNA进行PCR扩增,测序,BLAST比较,结果显示该菌与羊布氏杆菌的16S rDNA有100%的同源性,菌体与布氏杆菌标准阳性血清起强凝集反应.结果显示,该分离菌为布氏杆菌.

  13. Protection against brucellosis in goats, five years after vaccination with reduced-dose Brucella melitensis Rev 1 vaccine.

    Science.gov (United States)

    Díaz-Aparicio, E; Hernández, L; Suárez-Güemes, F

    2004-02-01

    The protection conferred by the reduced-dose Rev 1 Brucella melitensis vaccine in goats that had been immunized 5 years previously was evaluated. Sixteen goats vaccinated 5 years before with Rev 1 (1 x 10(5) cfu) and 5 non-vaccinated goats were challenged with B. melitensis 16M (4 x 10(5) cfu) using the conjunctival route. After giving birth or aborting, the goats were sacrificed and tissue samples were taken for bacteriological study. The challenge strain was recovered in 12%, of the animals from the vaccinated group, and in (80% of the control group. It is concluded, therefore that the use of reduced-dose Rev 1 protects goats vaccinated in endemic areas for at least 5 years after immunization.

  14. Complete genome-wide screening and subtractive genomic approach revealed new virulence factors, potential drug targets against bio-war pathogen Brucella melitensis 16M

    Directory of Open Access Journals (Sweden)

    Pradeepkiran JA

    2015-03-01

    Full Text Available Jangampalli Adi Pradeepkiran,1* Sri Bhashyam Sainath,2,3* Konidala Kranthi Kumar,1 Matcha Bhaskar1 1Division of Animal Biotechnology, Department of Zoology, Sri Venkateswara University, Tirupati, India; 2CIMAR/CIIMAR, Centro Interdisciplinar de Investigação Marinha e Ambiental, Universidade do Porto, Rua dos Bragas, Porto, Portugal, 3Department of Biotechnology, Vikrama Simhapuri University, Nellore, Andhra Pradesh, India *These authors contributed equally to this work Abstract: Brucella melitensis 16M is a Gram-negative coccobacillus that infects both animals and humans. It causes a disease known as brucellosis, which is characterized by acute febrile illness in humans and causes abortions in livestock. To prevent and control brucellosis, identification of putative drug targets is crucial. The present study aimed to identify drug targets in B. melitensis 16M by using a subtractive genomic approach. We used available database repositories (Database of Essential Genes, Kyoto Encyclopedia of Genes and Genomes Automatic Annotation Server, and Kyoto Encyclopedia of Genes and Genomes to identify putative genes that are nonhomologous to humans and essential for pathogen B. melitensis 16M. The results revealed that among 3 Mb genome size of pathogen, 53 putative characterized and 13 uncharacterized hypothetical genes were identified; further, from Basic Local Alignment Search Tool protein analysis, one hypothetical protein showed a close resemblance (50% to Silicibacter pomeroyi DUF1285 family protein (2RE3. A further homology model of the target was constructed using MODELLER 9.12 and optimized through variable target function method by molecular dynamics optimization with simulating annealing. The stereochemical quality of the restrained model was evaluated by PROCHECK, VERIFY-3D, ERRAT, and WHATIF servers. Furthermore, structure-based virtual screening was carried out against the predicted active site of the respective protein using the

  15. Identification of Brucella melitensis Rev.1 vaccine-strain genetic markers: Towards understanding the molecular mechanism behind virulence attenuation.

    Science.gov (United States)

    Issa, Mohammad Nouh; Ashhab, Yaqoub

    2016-09-22

    Brucella melitensis Rev.1 is an avirulent strain that is widely used as a live vaccine to control brucellosis in small ruminants. Although an assembled draft version of Rev.1 genome has been available since 2009, this genome has not been investigated to characterize this important vaccine. In the present work, we used the draft genome of Rev.1 to perform a thorough genomic comparison and sequence analysis to identify and characterize the panel of its unique genetic markers. The draft genome of Rev.1 was compared with genome sequences of 36 different Brucella melitensis strains from the Brucella project of the Broad Institute of MIT and Harvard. The comparative analyses revealed 32 genetic alterations (30 SNPs, 1 single-bp insertion and 1 single-bp deletion) that are exclusively present in the Rev.1 genome. In silico analyses showed that 9 out of the 17 non-synonymous mutations are deleterious. Three ABC transporters are among the disrupted genes that can be linked to virulence attenuation. Out of the 32 mutations, 11 Rev.1 specific markers were selected to test their potential to discriminate Rev.1 using a bi-directional allele-specific PCR assay. Six markers were able to distinguish between Rev.1 and a set of control strains. We succeeded in identifying a panel of 32 genome-specific markers of the B. melitensis Rev.1 vaccine strain. Extensive in silico analysis showed that a considerable number of these mutations could severely affect the function of the associated genes. In addition, some of the discovered markers were able to discriminate Rev.1 strain from a group of control strains using practical PCR tests that can be applied in resource-limited settings.

  16. Preliminary results of sero-conversion of kids and lambs vaccinated with Brucella melitensis rev -1 strain. Current achievements and feature challenges on brucellosis

    Directory of Open Access Journals (Sweden)

    XHELIL KOLECI

    2014-06-01

    Full Text Available Sheep and goat brucellosis is an endemic and most important infectious disease of livestock in Albania. It continues to remain a frequent zoonotic disease and an important public health issue. Among available strategies, mass vaccination is an acceptable, cost effective approach, and is a widely used strategy in many countries including some neighbouring Balkan countries. Albanian veterinary services supported by the European Union-funded PAZA project (Protection Against Zoonotic diseases, Albania applied two successive annual mass vaccination campaigns that aimed to vaccinate all small ruminants in the country. These two campaigns aimed at significantly reducing disease spread, however, a small number of infection foci could remain and persist in some parts of country. Post-vaccination surveillance is essential for early detection and proper control of cases of brucellosis that might re-emerge. Limitation major complication arising from mass vaccination is the difficulty of interpretation of the results of serological tests conducted to diagnose the disease. The aim of this study was to evaluate the proportion of vaccinated animals that showed sero-conversion and the duration of detectable levels of agglutinins (antibody against brucellosis in vaccinated animals. Methods. In total, 69 individual animals, 23 lambs and 46 kids aged from 4 to 7 months, were sampled at monthly intervals. Jugular blood was collected before vaccination and at intervals thereafter and tested by means of the Rose Bengal test. All animals were serologically negative before vaccination with modified live Brucella melitensis Rev.1 strain vaccine. Rose Bengal test was performed before vaccination, 18 days, 2, 3 and 4 months after vaccination. Results. Eighteen days after vaccination, 63 out of 69 animals (91.3% 82.6% of lambs (19 out of 23 lambs and 95.6% of goat kids (44 out of 46 showed strong sero-conversion in Rose Bengal test. The proportion of positive vaccinated

  17. 快速检测牛羊猪种布鲁杆菌多重PCR的建立%Differentiation of Brucella abortus , Brucella melitensis , and Brucella suis by multiple primers PCR

    Institute of Scientific and Technical Information of China (English)

    刘锴; 王兴龙; 马鸣萧; 翟丽鹃

    2009-01-01

    Objective To establish a method for rapidly identifying Brucella abortus, Brucella melitensis and Brucella suis by multiple primers PCR. Methods According to Brncella abortus, Brucella melitensis and Brucella suis IS711 insertion sequences, a public primer and three specific primers(544A, 16M, 1330S) were designed to set up multiplex PCR detection method. Yersinia O : 9, Escherichia coli O157 : HT, Salmonella typhimurium 47729 were selected to undergo multiple PCR reactions to detect the specificity. The sensitivity of multiple primers PCR of Brucella abortus was detected using multiple proportion dilution method. Results The amplified fragment size of Brucella abortus was 485 bp, that of Brucella melitensis 731 bp, and that of Brucella suis 248 bp, but PCR for the DNA of Yersinia O : 9, Escherichia coli O157 : H7, Salmonella typhimurium 47729 was negative. A sensitivity of the multiple primers PCR with Brucella abortus DNA using multiple proportional dilution quantitative method was 0.0967 pg. Conclusions Multiple PCR amplification method for rapidly detecting Brucella abortus, Brucella melitensis and Brucella suis has been successfully established, resulting in good specificity and sensitivity.%目的 建立一种能同时快速检测并能鉴别牛、羊、猪种布鲁杆菌的多重PCR方法.方法 根据IS711插入序列设计1条公共引物和3条牛、羊、猪种布鲁杆菌(544A、16M、1330S)特有序列引物,进行多重PCR反应;选择耶尔森菌O:9、大肠埃希菌O157:H7、鼠伤寒沙门菌47729进行多重PCR反应的特异性检测;倍比稀释定量法观察牛种布鲁杆菌多重PCR反应的敏感性.结果 牛、羊、猪种布鲁杆菌多重PCR反应扩增片段产物长度分别为485、731、248 bp;耶尔森菌O:9、大肠埃希菌O157:H7、鼠伤寒沙门菌47729加入布鲁杆菌中进行多重PCR反应.扩增结果呈阴性;牛种布鲁杆菌多重PCR反应敏感性为0.0967 Pg.结论 成功建立快速检测牛、羊、猪种布鲁

  18. Complete genome-wide screening and subtractive genomic approach revealed new virulence factors, potential drug targets against bio-war pathogen Brucella melitensis 16M.

    Science.gov (United States)

    Pradeepkiran, Jangampalli Adi; Sainath, Sri Bhashyam; Kumar, Konidala Kranthi; Bhaskar, Matcha

    2015-01-01

    Brucella melitensis 16M is a Gram-negative coccobacillus that infects both animals and humans. It causes a disease known as brucellosis, which is characterized by acute febrile illness in humans and causes abortions in livestock. To prevent and control brucellosis, identification of putative drug targets is crucial. The present study aimed to identify drug targets in B. melitensis 16M by using a subtractive genomic approach. We used available database repositories (Database of Essential Genes, Kyoto Encyclopedia of Genes and Genomes Automatic Annotation Server, and Kyoto Encyclopedia of Genes and Genomes) to identify putative genes that are nonhomologous to humans and essential for pathogen B. melitensis 16M. The results revealed that among 3 Mb genome size of pathogen, 53 putative characterized and 13 uncharacterized hypothetical genes were identified; further, from Basic Local Alignment Search Tool protein analysis, one hypothetical protein showed a close resemblance (50%) to Silicibacter pomeroyi DUF1285 family protein (2RE3). A further homology model of the target was constructed using MODELLER 9.12 and optimized through variable target function method by molecular dynamics optimization with simulating annealing. The stereochemical quality of the restrained model was evaluated by PROCHECK, VERIFY-3D, ERRAT, and WHATIF servers. Furthermore, structure-based virtual screening was carried out against the predicted active site of the respective protein using the glycerol structural analogs from the PubChem database. We identified five best inhibitors with strong affinities, stable interactions, and also with reliable drug-like properties. Hence, these leads might be used as the most effective inhibitors of modeled protein. The outcome of the present work of virtual screening of putative gene targets might facilitate design of potential drugs for better treatment against brucellosis.

  19. 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.

  20. A DNA vaccine encoding p39 and sp41 of Brucella melitensis induces protective immunity in BALB/c mice

    Directory of Open Access Journals (Sweden)

    A Al-Mariri

    2014-01-01

    Full Text Available Brucella species are facultative intracellular gram-negative bacteria that can multiply within phagocytic and non-phagocytic cells of humans or animals as end hosts. B. melitensis causes abortion in pregnant animals and undulant fever in humans. A 41 kDa surface protein (sp41 is associated with bacterial adherence and invasion of HeLa cells. The role of this protein a is important for the interaction with host cells. Previously, the putative periplasmic binding protein p39 had been described as T-cell immunodominant Brucella antigens. Both vectors (pCIp39 and pCIsp41 induced antigen-specific serum immunoglobulin as well as a T-cell-proliferative response and a strong gamma interferon production upon re-stimulation with either the specific antigens or Brucella extract. The level of protection was significant in pCIp39 and pCIsp41 treated mice but it was lower than the required level.

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

    Directory of Open Access Journals (Sweden)

    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

  2. Use of mass vaccination with a reduced dose of REV 1 vaccine for Brucella melitensis control in a population of small ruminants.

    Science.gov (United States)

    Scharp, D W; al Khalaf, S A; al Muhanna, M W; Cheema, R A; Godana, W

    1999-06-01

    Mass vaccination with reduced dose 1/50 Rev 1 strain live vaccine (1-2 10(9) colony forming units), administered subcutaneously, over a four and a half year period reduced the prevalence of Brucella melitensis in Kuwait's small ruminant population from 5.8% in 1993 to 2.02% in 1997. Serological test results using the Rose Bengal Plate Test, Rivanol Agglutination Test and Complement Fixation showed no evidence of persistence of positive serology in animals nine or more months after vaccination. Questionnaires and post-vaccination flock inspections found that the effects on gestation (abortions) were minimal--and not proven to be due to the vaccine. The conclusion from these findings is that mass vaccination with reduced dose Rev 1 administered by the subcutaneous route is a practical field strategy for control of Brucella melitensis.

  3. Evaluation of humoral and cellular immune responses to BP26 and OMP31 epitopes in the attenuated Brucella melitensis vaccinated sheep.

    Science.gov (United States)

    Wang, Wenjing; Wu, Jingbo; Qiao, Jun; Weng, Yunceng; Zhang, Hui; Liao, Qingyu; Qiu, Jinlang; Chen, Chuangfu; Allain, Jean-Pierre; Li, Chengyao

    2014-02-07

    In recent years, the number of cases of human brucellosis has been increasing by approximately 10% per year in China. Most cases were caused by Brucella melitensis through contacts with infected sheep, goats or their products. An attenuated B. melitensis vaccine M5-90 is currently used to vaccinate both animals in China. This vaccine has not been investigated for critical parameters such as immune response and its association with protective efficacy. In this study, humoral and cellular immune response to the periplasmic protein BP26 and the outer membrane protein OMP31 were evaluated in M5-90 vaccinated Chinese merino and Kazak sheep. Antibodies to BP26 or OMP31 were detected at low levels, and specific IFN-γ response was quantified. Strongly reactive peptides derived from BP26 and OMP31 identified five T-cell epitopes (BP26-6, -8, -11, -12 and OMP31-23) common to both sheep species, five species-specific epitopes (BP26-10, -18, -21 and -22 and OMP31-12) and four animal-specific epitopes (BP26-15, -23, OMP31-6 and -21), which stimulated specific IFN-γ response in vaccinated sheep. Among those T-cell epitopes, reactivity to BP26-18 and -21 epitopes was significantly associated with MHC-I B allele (P=0.024). However, a specific T-cell response induced by the M5-90 vaccine was relatively week and did not sustain long enough, which might be suppressed by rapid activation of T-regulatory (Treg) cells following vaccination. These findings provide an insight in designing a safer and more effective vaccine for use in animals and in humans.

  4. Preliminary virulence evaluation on mutation of pgm attenuates Brucella melitensis 16 strain%布鲁杆菌M16株pgm基因突变活菌苗株毒力初步评价

    Institute of Scientific and Technical Information of China (English)

    李鹏; 潘玉焕; 罗德炎; 王希良

    2011-01-01

    目的 通过减毒性实验评价布鲁杆菌M16株pgm基因突变活菌苗株毒力减弱情况,为新型减毒活疫苗的研制和布鲁杆菌感染免疫防御的研究奠定基础.方法 进行布鲁杆菌侵袭实验等动物和细胞实验评价突变菌株的毒力减弱情况.结果 突变菌株在小鼠体内能生存,但较强毒株存活数量有很大减少(P<0.001);突变菌株较强毒株对多粘菌素B更为敏感,存活率与强毒株相比有明显下降(P<0.001);突变菌株LPS较强毒株LPS明显缺失;突变菌株在体外HeLa细胞中能存活但复制减弱.结论 布鲁杆菌M16株pgm基因突变菌株毒力减弱.可作为未来布鲁杆菌新型减毒活疫苗的候选者,有进一步研究的价值.%Objective Evaluate the effect of attenuation and lay the foundation for the research of immunological mechanism of brucella infection and the development of novel live attenuated vaccines. Methods The effect of attenuation is evaluated by brucella invasion experiment, and other animal and cell experiments. Results Compared with wild - type strains, the mutation strains have several aspects different from it: ( 1 ) though the mutation strains of Brucella melitensis 16. can survive in the mouse model, the survival rates in the spleens of the inoculated mice were significantly lower than the wild - type strain; ( 2 ) the mutation strains were more sensitive to PmB than the wild - type parental strain; ( 3 ) the amount of LPS on the mutation strain of Brucella melitensis 16. were seen less than on the wild - type parental strain; (4) the replication rate of mutatation stains in HeLa cell line decreased dramatically although it can survive in it in vitro. Conclusions We have confirmed the attenuated effect of the mutation strain of Brucella melitensis 16, it can be used as a candidate vaccine for the control of brucellosis.

  5. Evaluation of the currently used diagnostic procedures for the detection of Brucella melitensis in sheep

    NARCIS (Netherlands)

    Bercovich, Z.; Guler, L.; Baysal, T.; Schreuder, B.E.C.; Zijderveld, van F.G.

    1998-01-01

    A study was conducted to determine whether the use of the enzyme-linked immunosorbent assay (ELISA) improves detection of brucellosis in individual sheep. Sera from 132 sheep that aborted due to B. melitensis were used to assess the efficacy of the ELISA to detect brucellosis in sheep. ELISA results

  6. Molecular cloning of virB12 gene ofBrucella melitensis 16M strain in pET28a vector

    Institute of Scientific and Technical Information of China (English)

    Shiva Mirkalantari; Nour Amirmozafari; Bahram Kazemi; Gholamreza Irajian

    2012-01-01

    ABSTRACT Objective:To clone the virB12 gene in pET28a expression vector for production of recombinant protein to be used as antigenic component for future serological test development.Methods:Brucella melitensis (B. melitensis)16M strain was cultured and bacterialDNA was extracted by BioneerAccuPrep®GenomicDNAExtractionKit.Oligonucleotide primer pair was designed based onBrucella virB12 gene sequence withBamHI andHindIII restriction site at5´ end ofthe forward and reverse primers, respectively.DNA amplification was performed usingPrimSTAR®HSDNA polymerase and thePCR product was purified byDNAAccuPrep®GelPurificationKit.Purified DNA was cloned into pJET1.2 cloning vector.VirB12 gene fragment was excised from pJET1.2 usingBamHI/HindIII and subsequently subcloned into pET28a(+).Results: Brucella virB12 gene was successfully cloned in pJET1.2 and then in pET28a(+) plasmids.PCR and restriction enzyme digestion confirms the procedure.Conclusion:We cloned and expressed theBrucella virB12 gene which could be used as antigenic component for specific serological assay development.

  7. Immunization with Individual Proteins of the Lrp/AsnC Family Induces Protection Against Brucella melitensis 16M Challenges in Mice

    Directory of Open Access Journals (Sweden)

    Xinhui eWang

    2015-10-01

    Full Text Available Brucellosis is one of the most common zoonoses worldwide. Subunit vaccines are promising for the prevention of human brucellosis. In our previous protective antigen screening studies, we identified a new protective antigen, BMEI0357, which belongs to the Lrp/asnC protein family, a conserved transcriptional regulator in bacteria that is absent in eukaryotes. In the present study, the Brucella genome annotation was screened and a total of 6 proteins were identified as members of the Lrp/AsnC family. Lrp/AsnC proteins have 2 domains that are conserved among the family members. However, sequence similarities between these proteins ranged from 9% to 50%, indicating high sequence heterogeneity. To test whether proteins of this family have similar characteristics, all 6 proteins were cloned and expressed in Escherichia coli. The recombinant proteins were purified and their protective efficacy was evaluated in BALB/c mice challenged with Brucella melitensis 16M. The results show that all 6 Lrp/AsnC proteins could induce a protective immune response against Brucella melitensis 16M. Antibodies against the Lrp/AsnC proteins were detected in the immunized mice. However, levels of antibodies against these proteins were relatively variable in human brucellosis sera. Taken together, our results show that these 6 proteins of the Lrp/AsnC family in Brucella could induce protective immune responses in mice.

  8. Expression, purification, and improved antigenic specificity of a truncated recombinant bp26 protein of Brucella melitensis M5-90: a potential antigen for differential serodiagnosis of brucellosis in sheep and goats.

    Science.gov (United States)

    Liu, Wen-xing; Hu, Sen; Qiao, Zu-jian; Chen, Wei-ye; Liu, Lin-tao; Wang, Fang-kun; Hua, Rong-hong; Bu, Zhi-gao; Li, Xiang-rui

    2011-01-01

    Antibodies produced in animals vaccinated using live attenuated vaccines against Brucella spp. are indistinguishable using current conventional serological tests from those produced in infected animals. One potential approach is to develop marker vaccines in which specific genes have been deleted from parental vaccine strains that show good immunogenicity and vaccine efficacy. Corresponding methods of detection for antibodies raised by the marker vaccine should also be developed. A specific fragment of the bp26 gene of Brucella melitensis M5-90 was cloned into vector pQE32 to construct the recombinant plasmid (pQE32-rΔbp26). It was used to transform Escherichia coli M15 (pREP4) host cells, which expressed the rΔbp26 protein. Subsequently, the recombinant protein was purified by immobilized metal affinity chromatography and size-exclusion chromatography. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the purified rΔbp26 protein was represented by only one band, with a molecular weight of 14 kDa, and it showed good antigenic specificity on western blot and enzyme-linked immunosorbent assay (ELISA). The purified rΔbp26 protein was intended to be used as an antigen to develop a novel ELISA to differentiate animals vaccinated with bp26 mutants of Brucella spp. from those infected naturally and those vaccinated with the parental vaccine strains.

  9. Brucella Infection Associated with Complete Atrioventricular Block

    Science.gov (United States)

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

    2016-01-01

    Background: 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. Case Report: 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. Conclusion: 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. PMID:27761286

  10. 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...

  11. Latex agglutination using the periplasmic proteins antigen of Brucella melitensis is a successful, rapid, and specific serodiagnostic test for ovine brucellosis.

    Science.gov (United States)

    Ismael, Alaa Bassuny; Swelum, Ayman Abdel-Aziz; Mostafa, Salama A-H; Alhumiany, Abdel-Rahman A

    2016-09-01

    Brucellosis, especially caused by Brucella melitensis, is considered the most-widespread zoonosis in the world, particularly in developing countries. This study was planned to develop an accurate test for diagnosis of ovine brucellosis using a specific hot saline extracted soluble Brucella melitensis periplasmic proteins (SBPPs). The efficacy of the latex agglutination test (LAT) using SBPPs compared to the Rose Bengal test (RBT), buffered plate agglutination test (BPAT), serum agglutination test (SAT), and an indirect enzyme-linked immunosorbent assay (i-ELISA) was evaluated in the field diagnosis of ovine brucellosis. The test performance was evaluated by estimating sensitivity (Se), specificity (Sp), positive predictive value (PPV), negative predictive value (NPV), disease prevalence (DP), positive likelihood ratio (PLR), and negative likelihood ratio (NLR) using test agreement and bacteriological culture in 1777 samples. The false-positive result was significantly (P ⩽0.05) lower in LAT than RBT, BPAT, SAT, and i-ELISA. With reference to test agreement, the Se, Sp, PPV, and PLR were highest (P ⩽0.05) in LAT 99.33%, 99.88%, 98.68%, and 827.25%, respectively. With reference to bacteriological culture, the LAT and i-ELISA tests showed a significant difference in Se with SAT. However, no significant difference in specificity was detected. The DP was 8.44% in the five tests. In conclusion, LAT using SBPPs of B. melitensis could be a suitable serodiagnostic field test for ovine brucellosis, with high sensitivity and specificity.

  12. Safety and efficacy of reduced doses of Brucella melitensis strain Rev. 1 vaccine in pregnant Iranian fat-tailed ewes.

    Science.gov (United States)

    Ebrahimi, Mohammad; Nejad, Ramin Bagheri; Alamian, Saeed; Mokhberalsafa, Ladan; Abedini, Fatemeh; Ghaderi, Rainak; Jalali, Hamid Reza

    2012-01-01

    Brucellosis is one of the most important zoonotic diseases and is a significant cause of abortion in animals. Brucella melitensis strain Rev. 1 is recommended as the most effective vaccine for small ruminants but the application of full doses in adult animals is restricted. This study was conducted to determine a proper reduced dose of vaccine which confers protection but which is not abortifacient in Iranian fat-tailed sheep. A total of 51 non-vaccinated pregnant ewes were divided into three main groups and several subgroups. Ewes in different groups were vaccinated at different stages of pregnancy and various subgroups were subcutaneously immunised with different quantities of the micro-organism (7.5 × 10(6), 10(6), 5 × 10(5)). Ewes again became pregnant a year later and were challenged with the wild-type strain to evaluate the protection conferred. Results revealed that the proportion of vaccination-induced abortions was significantly higher in ewes immunised with 7.5 × 10(6) Rev. 1 organisms than in those which received 10(6) or 5 × 10(5) bacteria. While 80% of non-vaccinated ewes aborted after challenge, none of the vaccinated ewes aborted post-challenge. This study indicated that a reduced dose of Rev. 1 vaccine containing 10(6) or 5 × 10(5) live cells could be safely used to induce protection in Iranian fat-tailed sheep at various stages of pregnancy.

  13. Optimization and efficient purification of recombinant Omp28 protein of Brucella melitensis using Triton X-100 and β-mercaptoethanol.

    Science.gov (United States)

    Kumar, Ashu; Tiwari, Sapana; Thavaselvam, Duraipandian; Sathyaseelan, Kannusamy; Prakash, Archana; Barua, Anita; Arora, Sonia; Kameswara Rao, M

    2012-06-01

    The high level expression of recombinant proteins in Escherichia coli often leads to the formation of inclusion bodies that contain most of the expressed protein held together by non-covalent forces. The inclusion bodies are usually solubilized using strong denaturing agents like urea and guanidium hydrochloride. In this study recombinant Omp28 (rOmp28) protein of Brucella melitensis was expressed in two different vector systems and further efficient purification of the protein was done by modification in buffers to improve the yield and purity. Different concentrations of Triton X-100 and β-mercaptoethanol were optimized for the solubilization of inclusion bodies. The lysis buffer with 8M urea alone was not sufficient to solubilize the inclusion bodies. It was found that the use of 1% Triton X-100 and 20mM β-mercaptoethanol in lysis and wash buffers used at different purification steps under denaturing conditions increased the yield of purified rOmp28 protein. The final yield of purified protein obtained with modified purification protocol under denaturing conditions was 151 and 90mg/l of the culture or 11.8 and 9.37mg/g of wet weight of cells in pQE30UA and pET28a(+) vector respectively. Thus modified purification protocol yielded more than threefold increase of protein in pQE30UA as compared with purification by conventional methods.

  14. Safety and efficacy of reduced doses of Brucella melitensis strain Rev. 1 vaccine in pregnant Iranian fat-tailed ewes

    Directory of Open Access Journals (Sweden)

    Mohammad Ebrahimi

    2012-12-01

    Full Text Available Brucellosis is one of the most important zoonotic diseases and is a significant cause of abortion in animals. Brucella melitensis strain Rev. 1 is recommended as the most effective vaccine for small ruminants but the application of full doses in adult animals is restricted. This study was conducted to determine a proper reduced dose of vaccine which confers protection but which is not abortifacient in Iranian fat-tailed sheep. A total of 51 non-vaccinated pregnant ewes were divided into three main groups and several subgroups. Ewes in different groups were vaccinated at different stages of pregnancy and various subgroups were subcutaneously immunised with different quantities of the micro-organism (7.5 × 106, 106, 5 × 105. Ewes again became pregnant a year later and were challenged with the wild-type strain to evaluate the protection conferred. Results revealed that the proportion of vaccination-induced abortions was significantly higher in ewes immunised with 7.5 × 106 Rev. 1 organisms than in those which received 106 or 5 × 105 bacteria. While 80% of non-vaccinated ewes aborted after challenge, none of the vaccinated ewes aborted post-challenge. This study indicated that a reduced dose of Rev. 1 vaccine containing 106 or 5 × 105 live cells could be safely used to induce protection in Iranian fat-tailed sheep at various stages of pregnancy.

  15. Induction of immune response in mice with a DNA vaccine encoding outer membrane protein (omp31) of Brucella melitensis 16M.

    Science.gov (United States)

    Gupta, V K; Rout, P K; Vihan, V S

    2007-06-01

    Brucellosis causes serious economic losses to goat farmers by way of reproductive losses in the form of abortions and stillbirths. Nucleic acid vaccines provide an exciting approach for antigen presentation to the immune system. In this study, we evaluated the ability of DNA vaccine encoding the omp31 protein of Brucella melitensis 16M to induce cellular and humoral immune responses in mice. We constructed eukaryotic expression vectors called pTargeTomp31, encoding outer membrane protein (omp31) of B. melitensis 16M. pTargeTomp31 was injected intramuscularly three times, at 3-week intervals in groups of mice 6 weeks of age. pTargeTomp31 induced good antibody response in ELISA . pTargeTomp31 elicited a T-cell-proliferative response and also induced a strong gamma interferon production upon restimulation with either the omp31 antigen or B. melitensis 16M extract. We also demonstrate that animals immunized with this plasmid elicited a strong and long-lived memory immune response. Furthermore, pTargeTomp31 elicited a typical T-helper 1-dominated immune response in mice, as determined by immunoglobulin G isotype analysis. This vaccine also provided the moderate degree of protection to the mice. This study for the first time focuses on DNA immunization of a gene from B. melitensis. These results may lead to the development of a DNA-based vaccine for the control of brucellosis in goats.

  16. Modeling, molecular dynamics, and docking assessment of transcription factor rho: a potential drug target in Brucella melitensis 16M

    Directory of Open Access Journals (Sweden)

    Pradeepkiran JA

    2015-03-01

    Full Text Available Jangampalli Adi Pradeepkiran,1 Konidala Kranthi Kumar,1 Yellapu Nanda Kumar,2 Matcha Bhaskar11Division of Animal Biotechnology, Department of Zoology, Sri Venkateswara University, Tirupati, 2Biomedical Informatics Centre, Vector Control Research Centre, Indian Council of Medical Research, Pondicherry, India Abstract: The zoonotic disease brucellosis, a chronic condition in humans affecting renal and cardiac systems and causing osteoarthritis, is caused by Brucella, a genus of Gram-negative, facultative, intracellular pathogens. The mode of transmission and the virulence of the pathogens are still enigmatic. Transcription regulatory elements, such as rho proteins, play an important role in the termination of transcription and/or the selection of genes in Brucella. Adverse effects of the transcription inhibitors play a key role in the non-successive transcription challenges faced by the pathogens. In the investigation presented here, we computationally predicted the transcription termination factor rho (TtFRho inhibitors against Brucella melitensis 16M via a structure-based method. In view the unknown nature of its crystal structure, we constructed a robust three-dimensional homology model of TtFRho’s structure by comparative modeling with the crystal structure of the Escherichia coli TtFRho (Protein Data Bank ID: 1PVO as a template in MODELLER (v 9.10. The modeled structure was optimized by applying a molecular dynamics simulation for 2 ns with the CHARMM (Chemistry at HARvard Macromolecular Mechanics 27 force field in NAMD (NAnoscale Molecular Dynamics program; v 2.9 and then evaluated by calculating the stereochemical quality of the protein. The flexible docking for the interaction phenomenon of the template consists of ligand-related inhibitor molecules from the ZINC (ZINC Is Not Commercial database using a structure-based virtual screening strategy against minimized TtFRho. Docking simulations revealed two inhibitors compounds – ZINC

  17. Fed batch fermentation and purification strategy for high yield production of Brucella melitensis recombinant Omp 28 kDa protein and its application in disease diagnosis.

    Science.gov (United States)

    Karothia, B S; Athmaram, T N; D, Thavaselvam; Ashu, Kumar; Tiwari, Sapna; Singh, Anil K; Sathyaseelan, K; Gopalan, N

    2013-07-01

    Brucellosis is a disease caused by bacteria belonging to the genus Brucella. It affects cattle, goat, sheep, dog and humans. The serodiagnosis of brucellosis involves detection of antibodies generated against the LPS or whole cell bacterial extracts, however these tests lack sensitivity and specificity. The present study was performed to optimize the culture condition for the production of recombinant Brucella melitensis outer membrane protein 28 kDa protein in E.coli via fed batch fermentation. Expression was induced with 1.5mM isopropyl β thiogalactoside and the expressed recombinant protein was purified using Ni-NTA affinity chromatography. After fed-batch fermentation the dry cell weight of 17.81 g/L and a purified protein yield of 210.10 mg/L was obtained. The purified Brucella melitensis recombinant Omp 28 kDa protein was analyzed through SDS- poly acrylamide gel electrophoresis and western blotting. The obtained recombinant protein was evaluated for its diagnostic application through Indirect ELISA using brucellosis suspected human sera samples. Our results clearly indicate that recombinant Omp28 produced via fed batch fermentation has immense potential as a diagnostic reagent that could be employed in sero monitoring of brucellosis.

  18. Re-examination of the role of the Brucella melitensis HtrA stress response protease in virulence in pregnant goats.

    Science.gov (United States)

    Roop, R M; Phillips, R W; Hagius, S; Walker, J V; Booth, N J; Fulton, W T; Edmonds, M D; Elzer, P H

    2001-09-03

    Based on previously reported studies describing the experimental infection of pregnant goats with B. melitensis strain RWP5, we proposed that the HtrA protease plays an important role in the virulence of B. melitensis in its natural ruminant host. Subsequent studies, however, have shown that RWP5 is actually an htrA cycL double mutant. In order to definitively evaluate the role of the B. melitensis htrA in virulence, we constructed an authentic htrA mutant and examined this strain in pregnant goats. The findings of these studies indicate that the contribution of the htrA gene product to the virulence of B. melitensis in its natural host is not as great as was previously proposed.

  19. Isolation and biotyping of Brucella melitensis from aborted sheep and goat fetuses

    OpenAIRE

    BÜYÜKCANGAZ, Esra; ŞEN, Ayşin; KAHYA, Serpil

    2014-01-01

    The possible role of Brucella spp. in 65 abortion cases, 55 from sheep and 10 from goats, occurring in the birth seasons of 2004 and 2005 in northwestern Turkey was investigated. Colony morphology, agglutination by acriflavin, H2S production, CO2 requirement, dye sensitivity in thionin, basic fuchsin, growth characteristics in streptomycin, lysis with Tbilisi phage, and agglutination with monospecific A- and M antisera were examined for identification and biotyping. The isolates from 21 of 55...

  20. 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.

  1. MLVA Genotyping Characteristics of Human Brucella melitensis Isolated from Ulanqab of Inner Mongolia, China

    Science.gov (United States)

    Liu, Zhi-Guo; Di, Dong-Dong; Wang, Miao; Liu, Ri-Hong; Zhao, Hong-Yan; Piao, Dong-Ri; Tian, Guo-Zhong; Fan, Wei-Xing; Jiang, Hai; Cui, Bu-Yun; Xia, Xian-Zhu

    2017-01-01

    Brucellosis is a serious public health problem in Ulanqab, which is a region located in the middle of the Inner Mongolia Autonomous Region adjacent to Shanxi and Hebei provinces. The disease is prevalent in both the latter provinces and Ulanqab with the highest prevalence of brucellosis occurring in Inner Mongolia. The MLVA-16 scheme is a genotyping tool for assessing genetic diversity and relationships among isolates. Moreover, this genotyping tool can also be applied to epidemiological trace-back investigations. This study reports the occurrence of at least two B. melitensis biovars (1 and 3) in Ulanqab, encompassing 22 and 94 isolates, respectively. B. melitensis biovar 3 was the predominant biovar in the area examined. Panel 1 (MLVA-8) identified three genotypes (42, 63, and 114), with genotype 42 (n = 101) representing 87% of the tested strains. MLVA-11 identified eight genotypes (116, 111, 297, 291, and 342–345) from 116 of the analyzed isolates. All of these isolates were identified as belonging to the East Mediterranean group. Genotype 116 (n = 94) was the predominant genotype and represented 81% of the isolates. The isolates pertaining to this genotype were distributed throughout most of Ulanqab and neighboring regions. The MLVA-16 scheme showed the presence of 69 genotypes, with 46 genotypes being represented by single isolates. This analysis revealed that Ulanqab brucellosis cases had epidemiologically unrelated and sporadic characteristics. The remaining 23 genotypes were shared (between a total of 70 isolates) with each genotype being represented by two to eight isolates. These data indicate that these cases were epidemiologically related. MLVA genotyping confirmed the occurrence of a multipoint outbreak epidemic and intrafamilial brucellosis. Extensive genotype-sharing events were observed among isolates from different regions of Ulanqab and from other provinces of China. These findings suggest either a lack of control of animal movement between

  2. Ovine and Caprine Brucellosis (Brucella melitensis) in Aborted Animals in Jordanian Sheep and Goat Flocks

    OpenAIRE

    Assadullah Samadi; M. MK. Ababneh; Giadinis, N. D.; LAFI, S. Q.

    2010-01-01

    Two hundred and fifty five biological samples were collected from 188 animals (81 sheep and 107 goats) during the lambing season from September 2009 to April 2010 from the Mafraq region of Jordan. Sampled animals belonged to 93 sheep and goat flocks that had abortion cases in the region. One hundred and seven (41.9%) biological samples were positive for the omp2 primers that were able to identify all Brucella species in the collected samples which were obtained from 86 aborted animals (86/188...

  3. Multiplex fluorescence quantitative PCR detection on Brucella spp.,Brucella abortus, and Brucella melitensis%多重荧光定量PCR方法鉴定布鲁氏菌属及牛羊种布鲁氏菌研究

    Institute of Scientific and Technical Information of China (English)

    刘志国; 罗成旺; 张利; 姜海; 赵鸿雁; 朴东日; 田国忠; 崔步云

    2012-01-01

    verified by amplification of 97 local strains. Results from multiplex TaqMan real-time PCR detection of Brucella showed that only Brucella spp. , Brucella abortus (B. abortus) and Brucella melitensis (B. melitensis) had respective fluorescence signal for each of them; while there was no fluorescent signal for pathogens with closer homology. According to standard curve, the lower limit of detection for both single and multiplex fluorescent PCR was 102copy/μL (13-24 fg/μL) , 100 times higher than that of conventional PCR. It ' s suggested that with advantages of higher sensitivity and fast and easy operation, the new method could be used to conduct rapid identification of Brucella spp. , B. abortus and B. melitensis in a single test.

  4. Prevalence of the Most Common Virulence-Associated Genes among Brucella Melitensis Isolates from Human Blood Cultures in Hamadan Province, West of Iran

    Science.gov (United States)

    Naseri, Zahra; Alikhani, Mohammad Yousef; Hashemi, Seyed Hamid; Kamarehei, Farideh; Arabestani, Mohammad Reza

    2016-01-01

    Brucellosis is a widespread zoonotic disease causing considerable economic and public health problems. Despite animal vaccination, brucellosis remains endemic in some areas such as Iran, especially in the western Iranian province of Hamadan. We sought to detect some of the most common virulence-associated genes in Brucella isolated from human blood cultures to determine the prevalence of some virulence genes among Brucella isolates. Fifty-seven isolates were studied from patients with a clinical diagnosis of brucellosis who referred to the Infectious Diseases Ward of Sina Hospital in Hamadan Province, Iran, between April 2013 and July 2014. Blood samples were collected for the diagnosis of brucellosis using the BACTEC blood culture system. All of these isolates were confirmed by the bcsp31 Brucella-specific gene. We detected 11 virulence-associated genes of Brucella, namely cβg, virB, znuA, ure, bvfA, omp25, omp31, wbkA, mviN, manA, and manB, which are important for the pathogenesis of this bacterium in the intracellular environment by multiplex PCR. Totally, 149 patients with a clinical diagnosis of brucellosis were enrolled in this study. Fifty-seven (38.3%) patients had positive blood cultures. On biochemical and molecular testing, all of the isolates were Brucella melitensis. Ten of the virulence genes were detected among all of the 57 isolates, but the bvf gene was detected in 53 (93%) isolates. The high prevalence of virulence-associated genes among the Brucella isolates detected in Hamadan Province, Iran, underscores the pathogenicity of this bacterium in this region. PMID:27582592

  5. Prevalence of the Most Common Virulence-Associated Genes among Brucella Melitensis Isolates from Human Blood Cultures in Hamadan Province, West of Iran

    Directory of Open Access Journals (Sweden)

    Zahra Naseri

    2016-09-01

    Full Text Available Brucellosis is a widespread zoonotic disease causing considerable economic and public health problems. Despite animal vaccination, brucellosis remains endemic in some areas such as Iran, especially in the western Iranian province of Hamadan. We sought to detect some of the most common virulence-associated genes in Brucella isolated from human blood cultures to determine the prevalence of some virulence genes among Brucella isolates. Fifty-seven isolates were studied from patients with a clinical diagnosis of brucellosis who referred to the Infectious Diseases Ward of Sina Hospital in Hamadan Province, Iran, between April 2013 and July 2014. Blood samples were collected for the diagnosis of brucellosis using the BACTEC blood culture system. All of these isolates were confirmed by the bcsp31 Brucella-specific gene. We detected 11 virulence-associated genes of Brucella, namely cβg, virB, znuA, ure, bvfA, omp25, omp31, wbkA, mviN, manA, and manB, which are important for the pathogenesis of this bacterium in the intracellular environment by multiplex PCR. Totally, 149 patients with a clinical diagnosis of brucellosis were enrolled in this study. Fifty-seven (38.3% patients had positive blood cultures. On biochemical and molecular testing, all of the isolates were Brucella melitensis. Ten of the virulence genes were detected among all of the 57 isolates, but the bvf gene was detected in 53 (93% isolates. The high prevalence of virulence-associated genes among the Brucella isolates detected in Hamadan Province, Iran, underscores the pathogenicity of this bacterium in this region.

  6. Mass spectrometry data from proteomics-based screening of immunoreactive proteins of fully virulent Brucella strains using sera from naturally infected animals

    Science.gov (United States)

    Wareth, Gamal; Melzer, Falk; Weise, Christoph; Neubauer, Heinrich; Roesler, Uwe; Murugaiyan, Jayaseelan

    2015-01-01

    Here, we provide the dataset associated with our research article on comprehensive screening of Brucella immunoreactive proteins using sera of naturally infected hosts published in Biochemical and Biophysical Research Communications Wareth et al., 2015 [1]. Whole-cell protein extracts were prepared from Brucella abortus and Brucella melitensis, separated using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and subsequently western blotting was carried out using sera from bovines (cows and buffaloes) and small ruminants (goats and sheep). The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository [2] with the dataset identifiers PXD001270 and DOI:10.6019/PXD001270. PMID:26322324

  7. Mass spectrometry data from proteomics-based screening of immunoreactive proteins of fully virulent Brucella strains using sera from naturally infected animals

    Directory of Open Access Journals (Sweden)

    Gamal Wareth

    2015-09-01

    Full Text Available Here, we provide the dataset associated with our research article on comprehensive screening of Brucella immunoreactive proteins using sera of naturally infected hosts published in Biochemical and Biophysical Research Communications Wareth et al., 2015 [1]. Whole-cell protein extracts were prepared from Brucella abortus and Brucella melitensis, separated using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE and subsequently western blotting was carried out using sera from bovines (cows and buffaloes and small ruminants (goats and sheep. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org via the PRIDE partner repository [2] with the dataset identifiers PXD001270 and DOI:10.6019/PXD001270.

  8. Brucella infection with pancytopenia after pediatric liver transplantation.

    Science.gov (United States)

    Polat, K Y; Tosun, M S; Ertekin, V; Aydinli, B; Emre, S

    2012-06-01

    Brucellosis is considered the most widespread zoonosis in the world. It has been reported that the prevalence of seropositivity among the Turkish population varies from 3% to 14%. We present a case of brucellosis after pediatric liver transplantation. A 15-year-old boy with the diagnosis of neuro Wilson's disease underwent deceased-donor liver transplantation. The postoperative immunosuppressive protocol consisted of steroids and tacrolimus. Two months after the operation the patient experienced fever to 40°C. The patient complained of poor appetite, headache, and diarrhea. He had had pancytopenia. Despite administration of appropriate antibiotics, antiviral and antifungal agents, fever persisted for > 1 month. Multiple blood, urine, stool, and sputum cultures were negative. Bone marrow aspirate revealed hypocellularity. Liver biopsy was performed, but rejection was not observed on biopsy specimen. Brucella serology was positive and Brucella agglutination titer was 1:320. Bone marrow culture was positive for Brucella but blood culture was negative. The patient was then treated with oral doxycycline and rifampin for 8 weeks. No previous case report about Brucella infection after liver transplantation has appeared in the literature, to our knowledge; our case is presented as the first. Bone marrow hypoplasia is a rare feature of Brucella infection. Our patient with brucellosis and pancytopenia had had hypocellular bone marrow. The clinical and hematologic findings resolved with treatment of the infection. Brucella infection should be suspected in liver transplanted recipients with fever of unknown origin, especially in a recipient who has lived in an endemic area. Brucella also should be considered as a possible diagnosis in patients with pancytopenia.

  9. Polymorphisms at the 3' untranslated region of SLC11A1 gene are associated with protection to Brucella infection in goats.

    Science.gov (United States)

    Iacoboni, Paola A; Hasenauer, Flavia C; Caffaro, M Eugenia; Gaido, Analia; Rossetto, Cristina; Neumann, Roberto D; Salatin, Antonio; Bertoni, Emiliano; Poli, Mario A; Rossetti, Carlos A

    2014-08-15

    Goats are susceptible to brucellosis and the detection of Brucella-infected animals is carried out by serological tests. In other ruminant species, polymorphisms in microsatellites (Ms) of 3' untranslated region (3'UTR) of the solute carrier family 11 member A1 (SLC11A1) gene were associated with resistance to Brucella abortus infection. Goats present two polymorphic Ms at the 3'UTR end of SLC11A1 gene, called regions A and B. Here, we evaluated if polymorphisms in regions A and/or B are associated with Brucella infection in goats. Serum (for the detection of Brucella-specific antibodies) and hair samples (for DNA isolation and structure analysis of the SLC11A1 gene) were randomly collected from 229 adult native goats from the northwest of Argentina. Serological status was evaluated by buffer plate antigen test (BPAT) complemented by the fluorescent polarization assay (FPA), and the genotype of the 3'UTR of the SLC11A1 gene was determined by capillary electrophoresis and confirmed by sequence analysis. Polymorphisms in regions A and B of the 3'UTR SLC11A1 gene were found statistically significant associated with protection to Brucella infection. Specifically, the association study indicates statistical significance of the allele A15 and B7/B7 genotype with absence of Brucella-specific antibodies (p=0.0003 and 0.0088, respectively). These data open a promising opportunity for limiting goat brucellosis through selective breeding of animals based on genetic markers associated with natural resistance to B. melitensis infection.

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

    Science.gov (United States)

    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.

  11. Immune Reactivity of Brucella Melitensis–Vaccinated Rabbit Serum with Recombinant Omp31 and Dnak Proteins

    Directory of Open Access Journals (Sweden)

    Mahmood Jeddi-Tehrani

    2013-03-01

    Full Text Available Background and objectives: Brucella melitensis infection is still a major health problem for human and cattle in developing countries and the Middle East.Materials and Methods: In this study, in order to screen immunogenic candidate antigens for the development of a Brucella subunit vaccine, a cytoplasmic protein (DnaK and an outer membrane protein (Omp31 of B. melitensis were cloned, expressed in E.coli BL21 and then purified using Ni-NTA agarose. Immunized serum was prepared from a rabbit inoculated with attenuated B. melitensis.Results and Conclusion: It was proved that immunized serum contains antibodies against recombinant Omp31 (rOmp31 and DnaK (rDnaK by Western blot and ELISA assays. The results may suggest the importance of these proteins as subunit vaccines against B. melitensis as well as targets for immunotherapy.

  12. Use of the Brucella melitensis native hapten to diagnose brucellosis in goats by a rapid, simple, and specific fluorescence polarization assay.

    Science.gov (United States)

    Ramírez-Pfeiffer, Carlos; Díaz-Aparicio, Efrén; Gomez-Flores, Ricardo; Rodríguez-Padilla, Cristina; Morales-Loredo, Alberto; Alvarez-Ojeda, Genoveva

    2008-06-01

    The performance of the fluorescence polarization assay (FPA) using the recently described Brucella melitensis native hapten and the Brucella abortus O-polysaccharide tracer was evaluated and compared with those of The World Organization for Animal Health tests related to indirect and competitive enzyme-linked immunosorbent assays as classification variables for goat sera obtained from a high-prevalence area where vaccination was performed; test series were also evaluated to increase the final specificity of the tests. Our results showed that the respective relative sensitivity and specificity were 99.7% and 32.5% for the rose Bengal test with a 3% cell concentration (RBT3), 92.8% and 68.8% for the rose Bengal test with 8% cell concentration (RBT8), 98.4% and 84.9% for the Canadian complement fixation test (CFT), 83.7% and 65.5% for the Mexican CFT, 98.4% and 81.0% for the buffered plate agglutination test (BPAT), and 78.1% and 89.3% for the fluorescence polarization assay (FPA). The use of the FPA as the secondary test significantly increased the final specificities of test combinations; the screening tests BPAT, RBT3, and RBT8 plus FPA resulted in 90%, 91.2%, and 91.3% final specificities, respectively, whereas for the combinations RBT3 plus Mexican CFT, RBT8 plus Mexican CFT, and BPAT plus Canadian CFT, the specificities were 65.5%, 63.2%, and 91.7%, respectively. The results suggested that the FPA may be routinely applied as an adaptable screening test for diagnosis of goat brucellosis, since its cutoff can be adjusted to improve its sensitivity or specificity, it is a rapid and simple test, it can be the test of choice when specificity is relevant or when an alternative confirmatory test is not available, and it is not affected by vaccination, thus reducing the number of goats wrongly slaughtered due to misdiagnosis.

  13. Enzyme-Linked Immunosorbent Assay To Differentiate the Antibody Responses of Animals Infected with Brucella Species from Those of Animals Infected with Yersinia enterocolitica O9

    OpenAIRE

    Erdenebaatar, Janchivdorj; Bayarsaikhan, Balgan; Watarai, Masahisa; Makino, Sou-ichi; Shirahata, Toshikazu

    2003-01-01

    Enzyme-linked immunosorbent assays using antigens extracted from Brucella abortus with n-lauroylsarcosine differentiated natural Brucella-infected animals from Brucella-vaccinated or Yersinia enterocolitica O9-infected animals. A field trial in Mongolia showed cattle, sheep, goat, reindeer, camel, and human sera without infection could be distinguished from Brucella-infected animals by conventional serological tests.

  14. Enzyme-linked immunosorbent assay to differentiate the antibody responses of animals infected with Brucella species from those of animals infected with Yersinia enterocolitica O9.

    Science.gov (United States)

    Erdenebaatar, Janchivdorj; Bayarsaikhan, Balgan; Watarai, Masahisa; Makino, Sou-ichi; Shirahata, Toshikazu

    2003-07-01

    Enzyme-linked immunosorbent assays using antigens extracted from Brucella abortus with n-lauroylsarcosine differentiated natural Brucella-infected animals from Brucella-vaccinated or Yersinia enterocolitica O9-infected animals. A field trial in Mongolia showed cattle, sheep, goat, reindeer, camel, and human sera without infection could be distinguished from Brucella-infected animals by conventional serological tests.

  15. 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.

  16. [A meningitis case of Brucella and tuberculosis co-infection].

    Science.gov (United States)

    Karsen, Hasan; Karahocagil, Mustafa Kasim; Irmak, Hasan; Demiröz, Ali Pekcan

    2008-10-01

    Turkey is located at an endemic area for brusellosis and tuberculosis which are both important public health problems. Meningitis caused by Brucella and Mycobacterium spp. may be confused since the clinical and laboratory findings are similar. In this report, a meningitis case with Brucella and tuberculosis co-infection has been presented. A 19-years-old woman was admitted to our clinic with severe headache, fever, vomiting, meningeal irritation symptoms, confusion and diplopia. The patient was initially diagnosed as Brucella meningitis based on her history (stockbreeding, consuming raw milk products, clinical symptoms concordant to brucellosis lasting for 4-5 months), physical examination and laboratory findings of cerebrospinal fluid (CSF). Standard tube agglutination test for brucellosis was positive at 1/80 titer in CSF and at 1/640 titer in serum, whereas no growth of Brucella spp. was detected in CSF and blood cultures. Antibiotic therapy with ceftriaxone, rifampicin and doxycyclin was started, however, there was no clinical improvement and agitation and confusion of the patient continued by the end of second day of treatment. Repeated CSF examination yielded acid-fast bacteria. The patient was then diagnosed as meningitis with double etiology and the therapy was changed to ceftriaxone, streptomycin, morphozinamide, rifampicin and isoniazid for thirty days. Tuberculosis meningitis was confirmed with the growth of Mycobacterium tuberculosis on the 14th day of cultivation (BACTEC, Becton Dickinson, USA) of the CSF sample. On the 30th day of treatment she was discharged on anti-tuberculous treatment with isoniazid and rifampicin for 12 months. The follow-up of the patient on the first and third months of treatment revealed clinical and laboratory improvement. Since this was a rare case of Brucella and tuberculosis co-infection, this report emphasizes that such co-infections should be kept in mind especially in the endemic areas for tuberculosis and brucellosis.

  17. Characterization of periplasmic protein BP26 epitopes of Brucella melitensis reacting with murine monoclonal and sheep antibodies.

    Science.gov (United States)

    Qiu, Jinlang; Wang, Wenjing; Wu, Jingbo; Zhang, Hui; Wang, Yuanzhi; Qiao, Jun; Chen, Chuangfu; Gao, Goege F; Allain, Jean-Pierre; Li, Chengyao

    2012-01-01

    More than 35,000 new cases of human brucellosis were reported in 2010 by the Chinese Center for Disease Control and Prevention. An attenuated B. melitensis vaccine M5-90 is currently used for vaccination of sheep and goats in China. In the study, a periplasmic protein BP26 from M5-90 was characterized for its epitope reactivity with mouse monoclonal and sheep antibodies. A total of 29 monoclonal antibodies (mAbs) against recombinant BP26 (rBP26) were produced, which were tested for reactivity with a panel of BP26 peptides, three truncated rBP26 and native BP26 containing membrane protein extracts (NMP) of B. melitensis M5-90 in ELISA and Western-Blot. The linear, semi-conformational and conformational epitopes from native BP26 were identified. Two linear epitopes recognized by mAbs were revealed by 28 of 16mer overlapping peptides, which were accurately mapped as the core motif of amino acid residues ⁹³DRDLQTGGI¹⁰¹ (position 93 to 101) or residues ¹⁰⁴QPIYVYPD¹¹¹, respectively. The reactivity of linear epitope peptides, rBP26 and NMP was tested with 137 sheep sera by ELISAs, of which the two linear epitopes had 65-70% reactivity and NMP 90% consistent with the results of a combination of two standard serological tests. The results were helpful for evaluating the reactivity of BP26 antigen in M5-90.

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

    Directory of Open Access Journals (Sweden)

    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.

  19. Genome-Wide Profiling of Humoral Immune Response to Coxiella burnetii Infection by Protein Microarray

    Science.gov (United States)

    2010-01-01

    find reactivity to this antigen from Brucella melitensis infec- ted human sera (Fig. 3). This is likely due to the low sequence homology (39%) between...C. burnetii strain 493 and the two Brucella species (B. abortus strain 2308 and B. melitensis strain 16M, which share 100% amino acid identity for...global, with infections occurring in a variety of mammals, birds, reptiles, fish, and ticks [2]. Sheep, goats , and other livestock are the primary

  20. Resposta imunitária à vacinação conjuntival com a estirpe Rev.1 de Brucella melitensis em ovinos e caprinos Serological response of sheep and goats to conjunctival Brucella melitensis Rev.1 vaccine

    Directory of Open Access Journals (Sweden)

    P. Poeta

    2003-04-01

    Full Text Available The live B. melitensis Rev.1 strain is considered the best vaccine available for the prophylaxis of brucellosis in small ruminants, especially when used at the standard dose by the conjunctival route. In the present study a 1´ 10(9 CFU dose for both sheep and goats conjunctivally vaccinated was tested to evaluate the duration of serological responses. Conjunctival vaccination with Rev. 1 performed in adult animals induced a rapid rise in serological titres as measured by Rose Bengal Plate Test (RBPT, Complement Fixation Test (CF and Modified Rose Bengal Plate Test (MRBPT. Titres then decreased and became negative in most animals by four months after vaccination (except MRBPT. The goats responded better to the vaccination than the sheep as one month after vaccination 100% of the goats revealed positive results to RB and RBM and 93.4% to FC test. The RBM was the one that detected more positive animals along the study.

  1. Cloning, expression and molecular analysis of Iranian Brucella melitensis Omp25 gene for designing a subunit vaccine

    Science.gov (United States)

    Yousefi, Soheil; Tahmoorespur, Mojtaba; Sekhavati, Mohammad Hadi

    2016-01-01

    Brucellosis is a well-known domestic animal infectious disease, which is caused by Brucella bacterium. The outer membrane protein 25 kDa (Omp25) gene plays an important role in simulating of TNF-α, IFN-α, macrophage, and cytokines cells. In the current study molecular cloning and expression analysis of Omp25 gene for designing a subunit vaccine against Brucella was investigated. Amplifying the full length of candidate gene was performed using specific primers. Sub-cloning of this gene conducted using pTZ57R/T vector in TOP10F strain of Escherichia coli(E.coli) as the host. Also, pET32(a)+ vector used for expression in BL21 (DE3) strain of E.coli. Omp25 gene with 642 bp size was amplified and cloned successfully. The expression results were confirmed by sequencing and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analyses which showed 42 kDa protein band correctly. Also, phylogenic analysis showed this gene has a near genetic relation with other Brucella strains. According to our results we can propose this gene as a candidate useful for stimulation of cell-mediated and humoral immunity system in future study. PMID:27920824

  2. Infecção em cão por Brucella abortus: relato de caso Brucella abortus infection in dog: case report

    Directory of Open Access Journals (Sweden)

    J. Megid

    2007-12-01

    Full Text Available Brucella abortus infection is reported in a dog from a rural area that presented at clinical evaluation left testicular enlargement and right testicular decrease. Serum resulted negative to rapid agglutination test and agar gel immunodifusion with Brucella ovis antigen but positive to buffered plate agglutination test, tube agglutination test and 2- Mercapthoetanol with B. abortus antigen. Brucella isolation was negative in blood, testicular material, semen and urine. Brucella DNA was detected in PCR from urine and blood.

  3. In Vitro Brucella suis Infection Prevents the Programmed Cell Death of Human Monocytic Cells

    Science.gov (United States)

    Gross, Antoine; Terraza, Annie; Ouahrani-Bettache, Safia; Liautard, Jean-Pierre; Dornand, Jacques

    2000-01-01

    During the complex interaction between an infectious agent and a host organism, the pathogen can interfere with the host cell's programmed death to its own benefit. Induction or prevention of host cell apoptosis appears to be a critical step for determining the infection outcome. Members of the gram-negative bacterial genus Brucella are intracellular pathogens which preferentially invade monocytic cells and develop within these cells. We investigated the effect of Brucella suis infection on apoptosis of human monocytic phagocytes. The present study provides evidence that Brucella infection inhibited spontaneously occurring apoptosis in human monocytes. Prevention of monocyte apoptosis was not mediated by Brucella lipopolysaccharide and required bacterial survival within infected cells. Both invaded and noninvaded cells were protected, indicating that soluble mediators released during infection were involved in the phenomenon. Analysis of Brucella-infected monocytes revealed specific overexpression of the A1 gene, a member of the bcl-2 family implicated in the survival of hematopoietic cells. Brucella infection also rendered macrophage-like cells resistant to Fas ligand- or gamma interferon-induced apoptosis, suggesting that Brucella infection protected host cells from several cytotoxic processes occurring at different steps of the immune response. The present data clearly show that Brucella suis modulated the monocyte/macrophage's apoptotic response to the advantage of the pathogen, thus preventing host cell elimination. This might represent a strategy for Brucella development in infected hosts. PMID:10603407

  4. Detection of Brucella species in the milk of infected cattle, sheep, goats and camels by PCR.

    Science.gov (United States)

    Hamdy, Mahmoud E R; Amin, A S

    2002-05-01

    One hundred and three milk samples were collected from 52 cows, 21 ewes, 18 goats and 12 camels. The animals tested positive to at least one of the following: (1) standard tube agglutination test (SAT); (2) Rose Bengal plate test (RBPT); (3) milk ring test (MRT). All milk samples were examined by culture and single-step polymerase chain reaction (PCR) techniques for detection of Brucella species. The PCR assay amplified Brucella-DNA from 29 bovine milk samples, 10 from sheep, 13 from goats and one from a camel. The direct culture method detected Brucella organisms from 24 samples of cows' milk, 12 from sheep, 10 from goats and failed to detect any Brucella organisms from camels' milk. PCR detected up to 100 colony forming units (CFU) of B. abortus per millilitre of milk in 100% of diluted milk samples, and 1000 CFU of B. melitensis from 70% of milk samples. Although the overall sensitivity of the PCR was higher than the culture method, it should be possible to increase the sensitivity to detect lower numbers of Brucella organisms in field samples. The speed and sensitivity of the PCR assay suggest that this technique could be useful for detection of Brucella organisms in bovine milk, as well as in sheep, goat, and camels milk.

  5. 固有免疫应答在抗布鲁菌感染过程中的作用%The role of innate immune response during infection of Brucella

    Institute of Scientific and Technical Information of China (English)

    高微; 王梦; 曹志然

    2015-01-01

    Brucella is a Gram-negative, facultative intracellular bacteria, which includes Brucella. abortus(cattle), B.melitensis(goats), B.suis (swine), B.canis(dogs) and Brucella infected by marine mammals, et al. B.abortus, B.melitensis, B.suis and B.canis, et al can invade into humans and animals through a variety of ways and cause Brucellosis that is a severe zoonotic desease. The innate immune system is the ifrst line of defensing against the infection of pathogen for our organism, which plays a key role during the infection of Brucella. This paper overviews the role of innate immune response during the infection of Brucella.%布鲁菌是革兰氏阴性兼性胞内寄生菌,包括牛布鲁菌、羊布鲁菌、猪布鲁菌、犬布鲁菌和海洋哺乳动物感染的布鲁菌等,其中羊布鲁菌、牛布鲁菌、猪布鲁菌和犬布鲁菌等可以通过多种途径侵入人和动物体内,其引起的布鲁菌病是一种严重的人畜共患病。固有免疫系统是机体抵抗病原体感染的第一道防线,因此在布鲁菌感染过程中发挥重要作用。现综述布鲁菌感染过程中宿主固有免疫应答发挥的作用。

  6. Validation of a fluorescence polarization assay (FPA) performed in microplates and comparison with other tests used for diagnosing B. melitensis infection in sheep and goats.

    Science.gov (United States)

    Minas, A; Stournara, A; Minas, M; Stack, J; Petridou, E; Christodoulopoulos, G; Krikelis, V

    2007-03-30

    Fluorescence polarization assay (FPA) is a relatively new test for the serological diagnosis of Brucella spp. infection in animals. FPA, carried out in 96-well microplate format, was validated here for diagnosing B. melitensis infection in sheep and goats. This study included sera from 1933 sheep and goats, from animals reared in naturally infected flocks (verified by culture) and showing a positive reaction to two different tests conducted in parallel. In addition, 2154 sera originating from healthy sheep and goats, reared in areas where B. melitensis had never been isolated, were assayed. The optimum cut-off value offering the highest diagnostic sensitivity (DSn) and diagnostic specificity (DSp) was determined at 15 mP over the mean value of the buffer control used in each microplate as determined by receiver operating characteristic analysis. The DSn and DSp of the FPA for small ruminants carried out in microplates at this cut-off value were calculated to be 95.9% and 97.9% with 95% confidence intervals (95% CI) of 94.9-97.7% and 97.2-98.4%, respectively. The accuracy of the FPA, as expressed by determination of the area under the curve, was 0.991. Indirect ELISA and FPA tests offered the highest DSn when compared with the Rose Bengal test, the complement fixation test, the modified Rose Bengal test and competitive ELISA. The parallel or serial combination of FPA with indirect ELISA offers the highest DSn and DSp. As temperature can affect the results of the FPA, all reagents must be at the same temperature and the standard for comparison must always be read under the same conditions as the sera under test. FPA performed in microplates is a promising assay; the DSn and accuracy are better than those of the tests currently approved for diagnosing B. melitensis in small ruminants. Because of its simplicity, speed, and accuracy, this test can improve capacity for laboratory testing and the efficacy of an eradication program based on a test-and-slaughter policy.

  7. Immunotherapeutics to prevent the replication of Brucella in a treatment failure mouse model.

    Science.gov (United States)

    Jain-Gupta, N; Contreras-Rodriguez, A; Smith, G P; Garg, V K; Witonsky, S G; Isloor, S; Vemulapalli, R; Boyle, S M; Sriranganathan, N

    2014-02-12

    Outer membrane vesicles (OMVs) from Brucella melitensis and irradiated Brucella neotomae have been shown to be effective vaccines against a B. melitensis challenge in a mouse model. The present study evaluates the efficacy of these two vaccines as immuno-therapeutics in combination with conventional antibiotics against a B. melitensis infection. BALB/c mice chronically infected with B. melitensis were treated for 4 weeks with doxycycline and gentamicin and vaccinated twice during the course of therapy. Antibiotics in sub-therapeutic concentrations were chosen in such a way that the treatment would result in a therapeutic failure in mice. Although no additive effect of vaccines and antibiotics was seen on the clearance of B. melitensis, mice receiving vaccines along with antibiotics exhibited no Brucella replication post-treatment compared to mice treated only with antibiotics. Administration of irradiated B. neotomae along with antibiotics led to higher production of IFN-γ ex vivo by splenocytes upon stimulation with heat inactivated B. melitensis while no such effect was seen by splenocytes from mice vaccinated with OMVs. OMV vaccinated mice developed significantly higher anti-Brucella IgG antibody titers at the end of the treatment compared to the mice that received only antibiotics. The mice that received only vaccines did not show any significant clearance of Brucella from spleens and livers compared to non-treated control mice. This study suggests that incorporating OMVs or irradiated B. neotomae along with conventional antibiotics might be able to improve therapeutic efficacy and control the progression of disease in treatment failure cases.

  8. Brucella abortus Cell Cycle and Infection Are Coordinated.

    Science.gov (United States)

    De Bolle, Xavier; Crosson, Sean; Matroule, Jean-Yves; Letesson, Jean-Jacques

    2015-12-01

    Brucellae are facultative intracellular pathogens. The recent development of methods and genetically engineered strains allowed the description of cell-cycle progression of Brucella abortus, including unipolar growth and the ordered initiation of chromosomal replication. B. abortus cell-cycle progression is coordinated with intracellular trafficking in the endosomal compartments. Bacteria are first blocked at the G1 stage, growth and chromosome replication being resumed shortly before reaching the intracellular proliferation compartment. The control mechanisms of cell cycle are similar to those reported for the bacterium Caulobacter crescentus, and they are crucial for survival in the host cell. The development of single-cell analyses could also be applied to other bacterial pathogens to investigate their cell-cycle progression during infection.

  9. Brucella abortus-infected B cells induce osteoclastogenesis.

    Science.gov (United States)

    Pesce Viglietti, Ayelén Ivana; Arriola Benitez, Paula Constanza; Giambartolomei, Guillermo Hernán; Delpino, María Victoria

    2016-09-01

    Brucella abortus is an intracellular bacterium that establishes lifelong infections in livestock and humans although the mechanisms of its chronicity are poorly understood. Activated B cells have long lifespan and B. abortus infection activates B cells. Our results indicate that the direct infection of B cells with B. abortus induced matrix metalloproteinase-9 (MMP-9), receptor activator for NF κB ligand (RANKL), tumor necrosis factor (TNF)-α and interleukin (IL)-6 secretion. In addition, supernatants from B. abortus-infected B cells induced bone marrow-derived monocytes to undergo osteoclastogenesis. Using osteoprotegerin, RANKL's decoy receptor, we determined that RANKL is involved in osteoclastogenesis induced by supernatants from B. abortus-infected B cells. The results presented here shed light on how the interactions of B. abortus with B cells may have a role in the pathogenesis of brucellar osteoarticular disease.

  10. Brucellae through the food chain: the role of sheep, goats and springbok (Antidorcus marsupialis) as sources of human infections in Namibia.

    Science.gov (United States)

    Magwedere, K; Bishi, A; Tjipura-Zaire, G; Eberle, G; Hemberger, Y; Hoffman, L C; Dziva, F

    2011-12-01

    A confirmed case of human brucellosis motivated an investigation into the potential source of infection in Namibia. Since domestic animals are principal sources of Brucella infection in humans, 1692 serum samples were screened from sheep, goats and cattle from 4 presumably at-risk farms and 900 springbok (Antidorcas marsupialis) serum samples from 29 mixed farming units for Brucella antibodies by the Rose-Bengal test (RBT) and positive cases confirmed by complement fixation test (CFT). To assess the prevalence of human brucellosis, 137 abattoir employees were tested for Brucella antibodies using the standard tube agglutination test (STAT) and by enzyme linked immunosorbent assay (ELISA). Cattle and sheep from all 4 farms were negative by RBT and CFT but 2 of the 4 farms (Ba and C) had 26/42 and 12/285 seropositive goats, respectively. Post mortem examination of seropositive goats revealed no gross pathological lesions typical of brucellosis except enlarged mesenteric and iliac lymph nodes seen in a single buck. Culture for brucellae from organs of seropositive animals was negative. None of the wildlife sera tested positive by either RBT or CFT. Interviews revealed that besides the case that prompted the investigation, a family and another person from other farms with confirmed brucellosis shared a common history of consumption of unpasteurised goat milk, home-made goat cheese and coffee with raw milk and prior contact with goats, suggesting goats as the likely source of infection. All 137 abattoir employees tested negative by STAT, but 3 were positive by ELISA. The 3 abattoir workers were clinically normal and lacked historical connections with clinical cases. Although goats are often associated with B. melitensis, these studies could not explicitly implicate this species owing to cross-reactivity with B. abortus, which can also infect goats. Nevertheless, these data reinforce the need for a better National Control Programme for brucellosis in Namibia.

  11. Brucellae through the food chain : the role of sheep, goats and springbok (Antidorcus marsupialis as sources of human infections in Namibia

    Directory of Open Access Journals (Sweden)

    K. Magwedere

    2011-05-01

    Full Text Available A confirmed case of human brucellosis motivated an investigation into the potential source of infection in Namibia. Since domestic animals are principal sources of Brucella infection in humans, 1692 serum samples were screened from sheep, goats and cattle from 4 presumably at-risk farms and 900 springbok (Antidorcas marsupialis serum samples from 29 mixed farming units for Brucella antibodies by the Rose-Bengal test (RBT and positive cases confirmed by complement fixation test (CFT. To assess the prevalence of human brucellosis, 137 abattoir employees were tested for Brucella antibodies using the standard tube agglutination test (STAT and by enzyme linked immunosorbent assay (ELISA. Cattle and sheep from all 4 farms were negative by RBT and CFT but 2 of the 4 farms (Ba and C had 26/42 and 12/285 seropositive goats, respectively. Post mortem examination of seropositive goats revealed no gross pathological lesions typical of brucellosis except enlarged mesenteric and iliac lymph nodes seen in a single buck. Culture for brucellae from organs of seropositive animals was negative. None of the wildlife sera tested positive by either RBT or CFT. Interviews revealed that besides the case that prompted the investigation, a family and another person from other farms with confirmed brucellosis shared a common history of consumption of unpasteurised goat milk, home-made goat cheese and coffee with raw milk and prior contact with goats, suggesting goats as the likely source of infection. All 137 abattoir employees tested negative by STAT, but 3 were positive by ELISA. The 3 abattoir workers were clinically normal and lacked historical connections with clinical cases. Although goats are often associated with B. melitensis, these studies could not explicitly implicate this species owing to cross-reactivity with B. abortus, which can also infect goats. Nevertheless, these data reinforce the need for a better National Control Programme for brucellosis in Namibia.

  12. 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.

  13. Comparative assessment of passive surveillance in disease-free and endemic situation: Example of Brucella melitensis surveillance in Switzerland and in Bosnia and Herzegovina

    Directory of Open Access Journals (Sweden)

    Haracic Sabina

    2008-12-01

    Full Text Available Abstract Background Globalization and subsequent growth in international trade in animals and animal products has increased the importance of international disease reporting. Efficient and reliable surveillance systems are needed in order to document the disease status of a population at a given time. In this context, passive surveillance plays an important role in early warning systems. However, it is not yet routinely integrated in the assessment of disease surveillance systems because different factors like the disease awareness (DA of people reporting suspect cases influence the detection performance of passive surveillance. In this paper, we used scenario tree methodology in order to evaluate and compare the quality and benefit of abortion testing (ABT for Brucella melitensis (Bm between the disease free situation in Switzerland (CH and a hypothetical disease free situation in Bosnia and Herzegovina (BH, taking into account DA levels assumed for the current endemic situation in BH. Results The structure and input parameters of the scenario tree were identical for CH and BH with the exception of population data in small ruminants and the DA in farmers and veterinarians. The sensitivity analysis of the stochastic scenario tree model showed that the small ruminant population structure and the DA of farmers were important influential parameters with regard to the unit sensitivity of ABT in both CH and BH. The DA of both farmers and veterinarians was assumed to be higher in BH than in CH due to the current endemic situation in BH. Although the same DA cannot necessarily be assumed for the modelled hypothetical disease free situation as for the actual endemic situation, it shows the importance of the higher vigilance of people reporting suspect cases on the probability that an average unit processed in the ABT-component would test positive. Conclusion The actual sensitivity of passive surveillance approaches heavily depends on the context in

  14. A Pediatric Case of Concomitant Leishmania and Brucella Infection

    Directory of Open Access Journals (Sweden)

    Perihan Yasemen Canöz

    2014-08-01

    Full Text Available Visceral leishmaniasis (Kala-azar is a disease caused by protozoan parasites of the Leishmania Genus, and Brucellosis is a zoonotic disease infecting human host by infective animals. A sixteen year-old girl presented to our clinic with complaints of fatigue, myalgia, pallor, stomach ache, leg swelling, and diffuse body rash. Physical examination revealed findings of elevated body temperature, splenomegaly, upper and lower extremity edema, and diffuse erythema. Patients’ brucella agglutination test was positive at titers 1: 640. Since the treatment of Brucellosis was unsuccessful, other disease processes were investigated and extracellular and intracellular amastigots were detected in the bone marrow aspirate preparations. Kala-azar dipstick (rk-39 was also positive. We present a 16 year-old girl who was diagnosed with Kala-azar and Brucellosis together infection and successfully treated.

  15. 布鲁氏菌M5-90疫苗株磷酸葡萄糖变位酶基因(pgm)突变株的构建及免疫评价%Construction and Immune Evaluation on Mutation of phosphoglucomutase gene (pgm) Attenuates of Brucella Melitensis Vaccine M5-90

    Institute of Scientific and Technical Information of China (English)

    李天森; 张辉; 张艳; 孟茹; 张豫; 孙志华; 蒋攀文; 王震; 陈瑞花

    2013-01-01

    In order to obtain a Brucella vaccine candidate which can distinguish between natural infection and attenuated vaccine, this paper studied the phosphoglucomutase gene (pgm) influence on Brucella M5-90 vaccine strain virulence and immune evaluation. The recombinant plasmid pGEM-7zf-Δpgm was constructed, and which was electroporated into Brucella melitensis M5-90 competent cells, screening for Brucella vaccine strain M5-90 pgm gene deletion mutant strains (Δpgm) and to detect the M5-90 Δpgm strain genetic stability, and immunogenicity. The test results showed that the reverse mutation did not occur within 15 passages; the Δpgm antibody content and the parent strain M5-90 group had no significant difference; Δpgm had the ability to produce the IL-2 was stronger than that of M5-90, the ability to produce INF-γ was lower than the M5-90; the rose bengal plate agglutination test and tube agglutination test of this gene deletion strains was not agglutinated; the results of Western blot confirmed that the Δpgm could not induce the mice to produce anti-pgm protein antibody. The Δpgm constructed in this study, had good genetic stability and immunogenicity, provided basic data for the next step to more in-depth develop Brucella gene-deleted vaccine.%为获得可以区分自然感染和疫苗免疫且毒力较弱的布鲁氏菌候选疫苗株,本实验研究了磷酸葡萄糖变位酶基因(pgm)对布鲁氏菌(Brucella melitensis)M5-90疫苗株毒力的影响,并对其进行了免疫评价.本实验构建了重组质粒pGEM-7zf-△pgm,电转化布鲁氏菌M5-90感受态细胞,筛选获得布鲁氏菌疫苗株M5-90的pgm基因缺失株(△pgm),并对获得的M5-90 △pgm株进行遗传稳定性、免疫原性检测.结果显示,△pgm能稳定传15代;△pgm组的抗体含量较亲本株M5-90组差异不显著;△pgm在诱导机体产生IL-2的能力要强于M5-90,产生INF-γ的能力低于M5-90;该基因缺失株采用虎红平板凝集试验和试管凝集

  16. Important biology events and pathways in Brucella infection and implications for novel antibiotic drug targets.

    Science.gov (United States)

    Gao, Guangjun; Xu, Jie

    2013-01-01

    Brucellosis caused by Brucella spp. is a common zoonosis in many parts of the world. Humans are infected through contact with infected animals or their dirty products. Many mechanisms are needed for this successful infection, although the mechanisms are still unclear. Host immune response and some signaling molecules play an important role in the infection event. Bacterial pathogens operate by attacking crucial intracellular pathways or some important molecules in each of these pathways for survival in their hosts. The crucial components (molecules) of immunity or pathway play a critical role in the whole process of Brucella infection. Here we summarize the findings of the Brucella-host interactions' immune system and signaling molecular cascades involved in the TLR-initiated immune response to Brucella spp. infection. The paper serves to deepen our understanding of this complex process and to provide some clues regarding the discovery of drug targets for prevention and control.

  17. Isolation and Identification of Brucella melitensis firstly isolated from goat in Guizhou province%贵州省首次从山羊分离到布鲁氏菌及其种型鉴定

    Institute of Scientific and Technical Information of China (English)

    李世军; 陈贵春; 王月; 陈红; 田克诚; 唐光鹏; 王定明; 王仕香; 周敬祝; 刘昭兵

    2011-01-01

    To isolate Brucella strains from the blood of Brucella antibody positive goats in Guizhou province and identify the isolated suspicious strains with conventional and molecular techniques, blood culture method was used to isolate Brucella strains from the blood of anti-Brucella antibody positive goats detected with standard agglutination test (SAT). Conventional and molecular techniques were used to identify suspicious bacteria strains. The results showed that Brucella suspicious colonies were cultured in 4 (GZB03, GZB04, GZB09, GZB11 ) of the total 11 (GZB01-11) goat blood samples. 4 strains were identified as B. melitensis biotype 3 by conventional tests and were further identified as Brucella spp. by genus specific BCSP31-PCR. Strain GZB03, GZB04, and GZB11 were classified as B. melitensis by species-specific AMOS-PCR, while GZB09 was unidentifiable to species. It is the first time to confirm that B. melitensis exists in Guizhou province.%目的 从贵州布鲁氏菌抗体阳性的山羊血液分离布鲁氏菌病原体并对其进行种型鉴定.方法 采用血培养法对经试管凝集试验检测为布鲁氏菌抗体阳性的山羊血血液进行细菌分离培养,并应用传统方法 和和分子生物学方法 对可疑菌落进行种型鉴定.结果 11份(GZB 01-11)抗体阳性的山羊血标本中有4份(GZB03、GZB04、GZB09、GZB11)经血培养瓶培养出布鲁氏菌可疑菌落,可疑菌落经传统方法 鉴定为羊种生物3型布鲁氏菌,4株菌进一步采用布鲁氏菌属特异性PCR(BCSP31-PCR)鉴定为布鲁氏菌属细菌,GZB03,GZB04,GZB11经种/型特异性PCR(AMOS-PCR)将其定为羊种布鲁氏菌,而GZB09采用该方法 不能定种.结论 从11份贵州省布鲁氏菌抗体阳性山羊血样中分离到4株羊种布鲁氏菌,首次证实贵州省存在羊种布鲁氏菌.

  18. Immune response triggered by Brucella abortus following infection or vaccination.

    Science.gov (United States)

    Dorneles, Elaine M S; Teixeira-Carvalho, Andréa; Araújo, Márcio S S; Sriranganathan, Nammalwar; Lage, Andrey P

    2015-07-17

    Brucella abortus live vaccines have been used successfully to control bovine brucellosis worldwide for decades. However, due to some limitations of these live vaccines, efforts are being made for the development of new safer and more effective vaccines that could also be used in other susceptible species. In this context, understanding the protective immune responses triggered by B. abortus is critical for the development of new vaccines. Such understandings will enhance our knowledge of the host/pathogen interactions and enable to develop methods to evaluate potential vaccines and innovative treatments for animals or humans. At present, almost all the knowledge regarding B. abortus specific immunological responses comes from studies in mice. Active participation of macrophages, dendritic cells, IFN-γ producing CD4(+) T-cells and cytotoxic CD8(+) T-cells are vital to overcome the infection. In this review, we discuss the characteristics of the immune responses triggered by vaccination versus infection by B. abortus, in different hosts.

  19. Case-control study on risk factors associated with Brucella melitensis in goat farms in Peninsular Malaysia.

    Science.gov (United States)

    Bamaiyi, Pwaveno Huladeino; Hassan, Latiffah; Khairani-Bejo, Siti; ZainalAbidin, Mohamed; Ramlan, Mohamed; Krishnan, Nookaya; Adzhar, Azri; Abdullah, Nahariah; Hamidah, Nik Husin M; Norsuhanna, Mokthar M; Hashim, Siti N

    2014-06-01

    Caprine brucellosis is a bacterial zoonotic infection affecting goats especially in developing countries all over the world. In Malaysia, the risk factors associated with this infection in farms have not been studied. A case-control study was carried out in goat farms in four states of Malaysia to elucidate the risk factors associated with the infection on the farms using structured questionnaires and face-to-face interviews. Results indicate that the introduction of new animals (OR = 5.25; 90 % CI = 1.46, 18.88), younger age category of farms (OR = 5.53; 90 % CI = 1.09, 21.66), and farms with single breed of goats (OR = 8.50; 90 % CI = 1.27, 41.97) were significant risk factors for brucellosis. In order to control brucellosis or possibly eradicate it in goat farms, these factors need to be dealt with. Enforcing stringent importation protocols or complete ban of goat importation from brucellosis endemic countries will help reduce risk of introducing new infection into the country.

  20. Proinflammatory Response of Human Trophoblastic Cells to Brucella abortus Infection and upon Interactions with Infected Phagocytes.

    Science.gov (United States)

    Fernández, Andrea G; Ferrero, Mariana C; Hielpos, M Soledad; Fossati, Carlos A; Baldi, Pablo C

    2016-02-01

    Trophoblasts are targets of infection by Brucella spp. but their role in the pathophysiology of pregnancy complications of brucellosis is unknown. Here we show that Brucella abortus invades and replicates in the human trophoblastic cell line Swan-71 and that the intracellular survival of the bacterium depends on a functional virB operon. The infection elicited significant increments of interleukin 8 (IL8), monocyte chemotactic protein 1 (MCP-1), and IL6 secretion, but levels of IL1beta and tumor necrosis factor-alpha (TNF-alpha) did not vary significantly. Such proinflammatory response was not modified by the absence of the Brucella TIR domain-containing proteins BtpA and BtpB. The stimulation of Swan-71 cells with conditioned medium (CM) from B. abortus-infected human monocytes (THP-1 cells) or macrophages induced a significant increase of IL8, MCP-1 and IL6 as compared to stimulation with CM from non-infected cells. Similar results were obtained when stimulation was performed with CM from infected neutrophils. Neutralization studies showed that IL1beta and/or TNF-alpha mediated the stimulating effects of CM from infected phagocytes. Reciprocally, stimulation of monocytes and neutrophils with CM from Brucella-infected trophoblasts increased IL8 and/or IL6 secretion. These results suggest that human trophoblasts may provide a local inflammatory environment during B. abortus infections either through a direct response to the pathogen or through interactions with monocytes/macrophages or neutrophils, potentially contributing to the pregnancy complications of brucellosis.

  1. Investigation of Brucella Infection in Milk Collected from Burdur Province

    OpenAIRE

    TÜRÜTOĞLU, Hülya

    2014-01-01

    In this study, we aimed to investigate the regional profile of bovine and ovine Brucellosis in Burdur by isolating Brucella spp. in milk and by detecting antibodies to Brucellae in milk samples. For this purpose, 404 milk samples from 101 cows and 226 milk samples from 113 ewes located in 14 different areas of Burdur were collected. The samples were examined by bacteriological and serological methods. Brucella spp. were not isolated from the milk samples. Positive results by the milk ring te...

  2. Brucella Endocarditis Caused By Brucella Melitensis

    Directory of Open Access Journals (Sweden)

    Suzan Saçar

    2008-01-01

    Full Text Available Brucellosis is a zoonotic disease endemically seen in Turkey, which occurs with various clinical findings. It can lead to complications affecting many systems. Endocarditis is an infrequent, but serious complication of brucellosis.The aim of this case presentation is to remind that endocarditis can be a complication of brucellosis and if is undiagnosed or misdiagnosed, progresses fatal in a high rate.

  3. 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.

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

    Science.gov (United States)

    He, Yongqun

    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 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.

  5. 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.

  6. Brucella abortus uses a stealthy strategy to avoid activation of the innate immune system during the onset of infection.

    Directory of Open Access Journals (Sweden)

    Elías Barquero-Calvo

    Full Text Available BACKGROUND: To unravel the strategy by which Brucella abortus establishes chronic infections, we explored its early interaction with innate immunity. METHODOLOGY/PRINCIPAL FINDINGS: Brucella did not induce proinflammatory responses as demonstrated by the absence of leukocyte recruitment, humoral or cellular blood changes in mice. Brucella hampered neutrophil (PMN function and PMN depletion did not influence the course of infection. Brucella barely induced proinflammatory cytokines and consumed complement, and was strongly resistant to bactericidal peptides, PMN extracts and serum. Brucella LPS (BrLPS, NH-polysaccharides, cyclic glucans, outer membrane fragments or disrupted bacterial cells displayed low biological activity in mice and cells. The lack of proinflammatory responses was not due to conspicuous inhibitory mechanisms mediated by the invading Brucella or its products. When activated 24 h post-infection macrophages did not kill Brucella, indicating that the replication niche was not fusiogenic with lysosomes. Brucella intracellular replication did not interrupt the cell cycle or caused cytotoxicity in WT, TLR4 and TLR2 knockout cells. TNF-alpha-induction was TLR4- and TLR2-dependent for live but not for killed B. abortus. However, intracellular replication in TLR4, TLR2 and TLR4/2 knockout cells was not altered and the infection course and anti-Brucella immunity development upon BrLPS injection was unaffected in TLR4 mutant mice. CONCLUSION/SIGNIFICANCE: We propose that Brucella has developed a stealth strategy through PAMPs reduction, modification and hiding, ensuring by this manner low stimulatory activity and toxicity for cells. This strategy allows Brucella to reach its replication niche before activation of antimicrobial mechanisms by adaptive immunity. This model is consistent with clinical profiles observed in humans and natural hosts at the onset of infection and could be valid for those intracellular pathogens phylogenetically

  7. Contamination of Bovine, Sheep and Goat Meat with Brucella Spp.

    Science.gov (United States)

    Casalinuovo, Francesco; Ciambrone, Lucia; Cacia, Antonio; Rippa, Paola

    2016-01-01

    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. PMID:27853716

  8. Contamination of Bovine, Sheep and Goat Meat with Brucella Spp.

    Science.gov (United States)

    Casalinuovo, Francesco; Ciambrone, Lucia; Cacia, Antonio; Rippa, Paola

    2016-06-03

    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.

  9. 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.

  10. The role of NLRP3 and AIM2 in inflammasome activation during Brucella abortus infection

    Science.gov (United States)

    Gomes, Marco Tulio R.; Miraglia, Maria Cruz; Giambartolomei, Guillermo H.; Oliveira, Sergio C.

    2016-01-01

    The innate immune system is essential for detection and elimination of bacterial pathogens. Upon inflammasome activation, caspase-1 cleaves pro-IL-1β and pro-IL-18 to their mature forms IL-1β and IL-18, respectively, and the cell undergoes inflammatory death termed pyroptosis. Here we reviewed recent findings demonstrating that Brucella abortus ligands activate NLRP3 and AIM2 inflammasomes which leads to control of infection. This protective effect is due to inflammatory response caused by IL-1β and IL-18 rather than cell death. Brucella DNA is sensed by AIM2 and bacteria induced mitochondrial reactive oxygen species is detected by NLRP3. However, deregulation of proinflammatory cytokine production can lead to immunopathology. Nervous system invasion by bacteria of the genus Brucella results in an inflammatory disorder termed neurobrucellosis. Herein we discuss the mechanism of caspase-1 activation and IL-1β secretion in glial cells infected with B. abortus. Our results demonstrate that the ASC inflammasome is indispensable for inducing the activation of caspase-1 and secretion of IL-1β upon infection of astrocytes and microglia with Brucella. Moreover, our results demonstrate that secretion of IL-1β by Brucella-infected glial cells depends on NLRP3 and AIM2 and leads to neurobrucellosis. Further, the inhibition of the host cell inflammasome as an immune evasion strategy has been described for bacterial pathogens. We discuss here that the bacterial type IV secretion system VirB is required for inflammasome activation in host cells during infection. Taken together, our results indicate that Brucella is sensed by ASC inflammasomes mainly NLRP3 and AIM2 that collectively orchestrate a robust caspase-1 activation and proinflammatory response. PMID:27405866

  11. Niğde İlinde Satışa Sunulan Koyun-Keçi Sütü ve Peynirlerinde Brucella melitensis ve Biyotiplerinin Araştırılması

    OpenAIRE

    KARADAL, Fulden; ERTAŞ ONMAZ, Nurhan; BAĞCI, Cemalettin; YILDIRIM, Yeliz; AL, Serhat; ABAY, Seçil

    2016-01-01

    Bu çalışma Niğde ilinde satışa sunulan çiğ koyun ve keçi sütleri ile çiğ sütten yapılmış koyun ve keçi peynirlerindekiBrucella melitensis varlığını araştırmak amacıyla yapıldı. Bu amaçla, Niğde’nin çeşitli köy ve pazarlarındantoplanan 57 çiğ koyun sütü, 43 çiğ keçi sütü ve 50 koyun- keçi peyniri olmak üzere toplam 150 örnek materyal olarakkullanıldı. B. melitensis’in izolasyonunda Farrell yöntemi, şüpheli izolatların identifikasyonunda polimeraz zincir reaksiyonu(PZR) uygulandı. Çalışmada inc...

  12. Delta-pgm, a new live-attenuated vaccine against Brucella suis.

    Science.gov (United States)

    Czibener, Cecilia; Del Giudice, Mariela Giselda; Spera, Juan Manuel; Fulgenzi, Fabiana Rosa; Ugalde, Juan Esteban

    2016-03-18

    Brucellosis is one of the most widespread zoonosis in the world affecting many domestic and wild animals including bovines, goats, pigs and dogs. Each species of the Brucella genus has a particular tropism toward different mammals being the most relevant for human health Brucella abortus, Brucella melitensis and Brucella suis that infect bovines, goats/camelids and swine respectively. Although for B. abortus and B. melitensis there are vaccines available, there is no efficient vaccine to protect swine from B. suis infection so far. We describe here the construction of a novel vaccine strain that confers excellent protection against B. suis in a mouse model of infection. This strain is a clean deletion of the phosphoglucomutase (pgm) gene that codes for a protein that catalyzes the conversion of glucose-6-P to glucose-1-P, which is used as a precursor for the biosynthesis of many polysaccharides. The Delta-pgm strain lacks a complete lipopolysaccharide, is unable to synthesize cyclic beta glucans and is sensitive to several detergents and Polymyxin B. We show that this strain replicates in cultured cells, is completely avirulent in the mouse model of infection but protects against a challenge of the virulent strain inducing the production of pro-inflammatory cytokines. This novel strain could be an excellent candidate for the control of swine brucellosis, a disease of emerging concern in many parts of the world.

  13. Immunization of Mice with Recombinant Brucella abortus Organic Hydroperoxide Resistance (Ohr) Protein Protects Against a Virulent Brucella abortus 544 Infection.

    Science.gov (United States)

    Hop, Huynh Tan; Reyes, Alisha Wehdnesday Bernardo; Simborio, Hannah Leah Tadeja; Arayan, Lauren Togonon; Min, Won Gi; Lee, Hu Jang; Lee, Jin Ju; Chang, Hong Hee; Kim, Suk

    2016-01-01

    In this study, the Brucella abortus ohr gene coding for an organic hydroperoxide resistance protein (Ohr) was cloned into a maltose fusion protein expression system (pMAL), inserted into Escherichia coli, and purified, and its immunogenicity was evaluated by western blot analysis using Brucella-positive mouse sera. The purified recombinant Ohr (rOhr) was treated with adjuvant and injected intraperitoneally into BALB/c mice. A protective immune response analysis revealed that rOhr induced a significant increase in both the IgG1 and IgG2a titers, and IgG2a reached a higher level than IgG1 after the second and third immunizations. Additionally, immunization with rOhr induced high production of IFN-γ as well as proinflammatory cytokines such as TNF, MCP-1, IL-12p70, and IL-6, but a lesser amount of IL-10, suggesting that rOhr predominantly elicited a cell-mediated immune response. In addition, immunization with rOhr caused a significantly higher degree of protection against a virulent B. abortus infection compared with a positive control group consisting of mice immunized with maltose-binding protein. These findings showed that B. abortus rOhr was able to induce both humoral and cell-mediated immunity in mice, which suggested that this recombinant protein could be a potential vaccine candidate for animal brucellosis.

  14. Infection of California sea lions (Zalophus californianus) with terrestrial Brucella spp.

    Science.gov (United States)

    Avalos-Téllez, Rosalía; Ramírez-Pfeiffer, Carlos; Hernández-Castro, Rigoberto; Díaz-Aparicio, Efrén; Sánchez-Domínguez, Carlos; Zavala-Norzagaray, Alan; Arellano-Reynoso, Beatriz; Suárez-Güemes, Francisco; Aguirre, A Alonso; Aurioles-Gamboa, David

    2014-10-01

    Infections with Brucella ceti and pinnipedialis are prevalent in marine mammals worldwide. A total of 22 California sea lions (Zalophus californianus) were examined to determine their exposure to Brucella spp. at San Esteban Island in the Gulf of California, Mexico, in June and July 2011. Although samples of blood, vaginal mucus and milk cultured negative for these bacteria, the application of rose Bengal, agar gel immunodiffusion, PCR and modified fluorescence polarization assays found that five animals (22.7%) had evidence of exposure to Brucella strains. The data also suggested that in two of these five sea lions the strains involved were of terrestrial origin, a novel finding in marine mammals. Further work will be required to validate and determine the epidemiological significance of this finding.

  15. In Harm’s Way: Infections in Deployed American Military Forces

    Science.gov (United States)

    2006-05-23

    a US Army helicopter pilot after a 5-month deployment in Iraq (blood culture grew Brucella melitensis ); he had observed the slaughter of a sheep, but...This typically results from direct contact with the reservoir hosts (commonly cattle, goats , and sheep), but it may occur after exposure to...Brucellosis is a zoonotic disease endemic to the Middle East and transmitted to humans through contact with infected an- imals. Brucella bacteria may be

  16. Vaccination with recombinant L7/L12-truncated Omp31 protein induces protection against Brucella infection in BALB/c mice.

    Science.gov (United States)

    Golshani, Maryam; Rafati, Sima; Dashti, Amir; Gholami, Elham; Siadat, Seyed Davar; Oloomi, Mana; Jafari, Anis; Bouzari, Saeid

    2015-06-01

    Brucellosis is the most common bacterial zoonotic disease worldwide and no vaccine is available for the prevention of human brucellosis. In humans, brucellosis is mostly caused by Brucella melitensis and Brucella abortus. The Outer membrane protein 31 (Omp31) and L7/L12 are immunodominant and protective antigens conserved in human Brucella pathogens. In the present study, we evaluated the humoral and cellular immune responses induced by a fusion protein designed based on the Truncated form of Omp31 (TOmp31) and L7-L12 antigens. Vaccination of BALB/c mice with the recombinant fusion protein (rL7/L12-TOmp31) provided the significant protection level against B. melitensis and B. abortus challenge. Moreover, rL7/L12-TOmp31 elicited a strong specific IgG response (higher IgG2a titers) and significant IFN-γ/IL2 production and T-cell proliferation was also observed. The T helper1 (Th1) oriented response persisted for 12 weeks after the first immunization. The rL7/L12-TOmp31 could be a new potential antigen candidate for the development of a subunit vaccine against B. melitensis and B. abortus.

  17. Serological diagnosis of bovine brucellosis using B. melitensis strain B115.

    Science.gov (United States)

    Corrente, Marialaura; Desario, Costantina; Parisi, Antonio; Grandolfo, Erika; Scaltrito, Domenico; Vesco, Gesualdo; Colao, Valeriana; Buonavoglia, Domenico

    2015-12-01

    Bovine brucellosis is diagnosed by official tests, such as Rose Bengal plate test (RBPT) and Complement Fixation test (CFT). Both tests detect antibodies directed against the lipolysaccharide (LPS) of Brucella cell wall. Despite their good sensitivity, those tests do not discriminate between true positive and false positive serological reactions (FPSR), the latter being generated by animals infected with other Gram negative microorganisms that share components of Brucella LPS. In this study, an antigenic extract from whole Brucella melitensis B115 strain was used to set up an ELISA assay for the serological diagnosis of bovine brucellosis. A total of 148 serum samples from five different groups of animals were tested: Group A: 28 samples from two calves experimentally infected with Yersinia enterocolitica O:9; Group B: 30 samples from bovines infected with Brucella abortus; Group C: 50 samples from brucellosis-free herds; Group D: 20 samples RBPT positive and CFT negative; Group E: 20 samples both RBPT and CFT positive. Group D and Group E serum samples were from brucellosis-free herds. Positive reactions were detected only by RBPT and CFT in calves immunized with Y. enterocolitica O:9. Sera from Group B animals tested positive also in the ELISA assay, whereas sera from the remaining groups were all negative. The results obtained encourage the use of the ELISA assay to implement the serological diagnosis of brucellosis.

  18. 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

    cattle, sheep, goats , dogs, and camels. Important species of Brucella are: suis, abortus, ovis, melitensis , canis, and neotomae, each with certain...B. abortus is strain 19 and for protecting goats against B. melitensis is strain Rev 1. Vaccination with these strains leads to seroconversion...encoding the C-terminus portion of the Brucella protein. Western blot analysis of B. abortus and B. melitensis was performed using antibodies raised against

  19. 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.

  20. Serological differentiation of Brucella-vaccinated and -infected domesticated animals by the agar gel immunodiffusion test using Brucella polysaccharide in mongolia.

    Science.gov (United States)

    Erdenebaatar, Janchivdorj; Sugar, Sengee; Yondondorj, Agchbazar; Nagabayashi, Toshihiko; Syuto, Bunei; Watarai, Masahisa; Makino, Sou-Ichi; Shirahata, Toshikazu

    2002-09-01

    To investigate Brucella infection in cattle, sheep, goat, reindeer and yak in Mongolia, serological reactions of Brucella-infected and -vaccinated domestic animals were compared by the agar gel immunodiffusion (AGID) test with a polysaccharide (poly-B) of the B. Abortus strain S-19. The sensitivity and specificity were compared with conventional serological tests that are commonly used in Mongolia, such as the rose Bengal test, the tube agglutination test and the compliment fixation test. A total of 73.3, 100, 100, 95.8 and 61.9% of the sera from suspected cattle, yak, goat, sheep and reindeer, respectively, that were positive in the compliment fixation test, were also positive in the AGID test. Sera from vaccinated cattle, sheep and goat were positive over 90% by conventional tests 3 months after vaccination, but were negative by the AGID. These results suggest that the AGID test may be useful to differentiate infected and vaccinated animals in the field.

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

    Directory of Open Access Journals (Sweden)

    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

  2. Effects of gamma radiation and azathioprine on Brucella abortus infection in BALB/c mice

    Energy Technology Data Exchange (ETDEWEB)

    Elzer, P.H.; Rowe, G.E.; Enright, F.M.; Winter, A.J. (Department of Veterinary Microbiology, Immunology and Parasitology, College of Veterinary Medicine, Cornell University, Ithaca, NY (United States))

    1991-06-01

    Sublethal irradiation of BALB/c mice 4 hours prior to inoculation with 5 {times} 10(4) virulent Brucella abortus, caused significant (P less than 0.01) reductions in bacterial numbers in comparison with numbers in unirradiated controls. Numbers of brucellae in the spleen were significantly lower by 5 days after inoculation and decreased thereafter, so that at 2 and 3 weeks after inoculation, there were up to 1,000-fold fewer organisms in the spleen of irradiated mice. The number of brucellae in the spleen increased in irradiated mice thereafter. The course of events in the liver was similar, but developed more slowly, and peak differences in bacterial numbers were about 1 log less. These phenomena were not attributable to differences in implantation of brucellae in the liver or spleen, nor to an abnormal distribution of organisms in other organs of irradiated mice. Irradiation of mice during the plateau phase of infection also resulted in significant (P less than 0.05) reductions in bacterial counts in the spleen during the succeeding 4 weeks. Macrophage activation in the spleen, measured by a Listeria monocytogenes-killing assay, was significantly (P less than 0.01) increased by irradiation alone at 1 week after inoculation and at that time was significantly (P less than 0.01) greater in B abortus-infected, irradiated mice than in B abortus-infected controls. Histologic, cytologic, and immunologic studies revealed that the decrease in numbers of organisms between 1 and 2 weeks after inoculation in irradiated mice occurred at a time when their immune response to B abortus was suppressed and when numbers of neutrophils and monocytes infiltrating the spleen were significantly (P less than 0.01) diminished.

  3. Performance of skin tests with allergens from B. melitensis B115 and rough B. abortus mutants for diagnosing swine brucellosis.

    Science.gov (United States)

    Dieste-Pérez, L; Blasco, J M; De Miguel, M J; Marín, C M; Barberán, M; Conde-Álvarez, R; Moriyón, I; Muñoz, P M

    2014-01-10

    Swine brucellosis by Brucella suis biovar 2 is an emerging disease whose control is based on serological testing and culling. However, current serological tests detect antibodies to the O-polysaccharide (O/PS) moiety of Brucella smooth lipopolysaccharide (S-LPS), and thus lack specificity when infections by Yersinia enterocolitica O:9 and other gram-negative bacteria carrying cross-reacting O/PS occur. The skin test with the protein-rich brucellin extract obtained from rough B. melitensis B115 is assumed to be specific for discriminating these false positive serological reactions (FPSR). However, B115 strain, although unable to synthesize S-LPS, accumulates O/PS internally, which could cause diagnostic problems. Since the brucellin skin test has been seldom used in pigs and FPSR are common in these animals, we assessed its performance using cytosoluble protein extracts obtained from B. abortus rough mutants in manBcore or per genes (critical for O/PS biosynthesis) and B. melitensis B115. The diagnostic sensitivity and specificity were determined in B. suis biovar 2 culture positive and brucellosis free sows, and apparent prevalence in sows of unknown individual bacteriological and serological status belonging to B. suis biovar 2 naturally infected herds. Moreover, the specificity in discriminating brucellosis from FPSR was assessed in brucellosis free boars showing FPSR. The skin test with B. abortus ΔmanBcore and B. melitensis B115 allergens performed similarly, and the former one resulted in 100% specificity when testing animals showing FPSR in indirect ELISA, Rose Bengal and complement fixation serological tests. We conclude that O/PS-free genetically defined mutants represent an appropriate alternative to obtain Brucella protein extracts for diagnosing swine brucellosis.

  4. An unusual case of seronegative, 16S PCR positive Brucella infection

    Science.gov (United States)

    Backhouse, Lucy; Rawat, David; Naik, Sandhia; Millar, Michael

    2016-01-01

    Introduction: Brucella is a zoonotic infection commonly diagnosed by isolation of the organism from blood culture or positive serological testing. It is an uncommon cause of a pyrexia of unknown origin in the United Kingdom. Case presentation: We describe the case of a 14-year-old girl with no history of travel who presented with pyrexia, weight loss, arthralgia, multiple splenic abscesses and a subsequent pleural effusion, the latter of which isolated a Brucella species on 16S rRNA PCR. The patient responded well to initiation of treatment for brucellosis and on repeat imaging, after 3 months, the splenic abscesses had resolved. Conclusion: This unique case demonstrates uncommon complications of brucellosis and the challenges of diagnosing the organism, the latter of which can be alleviated by the utilization of molecularbased technologies. This patient had a negative serology result for brucellosis, which highlights the need to interpret serology results with caution in non-endemic regions for brucellosis.

  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.

  6. 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.

  7. Brucella DNA is not detected in in-vitro produced embryos derived from ovaries of naturally infected Brucella DNA is not detected in in-vitro produced embryos derived from ovaries of naturally infected buffaloes

    Directory of Open Access Journals (Sweden)

    L. Manna

    2010-02-01

    Full Text Available The aim of this study was to screen for Brucella spp. buffalo embryos produced in- vitro, by using cumulus oocytes complexes (COCs recovered from ovaries of slaughtered buffaloes naturally infected with Brucella spp. Ovaries were collected from 5 female pluriparous buffaloes slaughtered in a local abattoir. EDTA-blood samples and nasal swabs collected from each animal were used for Brucella spp. DNA detection by real-time PCR. Buffalo ovaries (n = 10 were transported to the laboratory and maintained strictly separated throughout laboratory processing. Recovered COCs were matured, fertilized and cultured in vitro until day 7. Some immature COCs, all uncleaved COCs, all blocked cleaved embryos (2 to 16 cells and all transferable embryos (tight morulae and blastocysts were separately analysed by real-time PCR assay. Brucella spp. DNA was detected in both blood and nasal mucus of all subjects, whereas no trace of DNA of Brucella spp. was found on either COCs or embryos. Currently, the infected or seropositive buffaloes have to be slaughtered for sanitary reasons. Interestingly, the results of this preliminary trial suggest a possible utilization of the COCs from the infected subjects of high genetic value to obtain safe embryos.

  8. 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...

  9. 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.

  10. 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.

  11. 羊种布氏杆菌基因外重复回文序列经Toll样受体9诱导IFN-α表达的研究%The effects of repetitive extragenic palindromic sequences from Brucella melitensis DNA on the toll-like receptor 9-mediated interferon-α production

    Institute of Scientific and Technical Information of China (English)

    白丽云; 张雅娴; 王占黎; 王英; 于慧

    2015-01-01

    Objective To screen the repetitive extragenic palindromic sequences with activation of toll-like receptor 9(TLR9) activity from Brucella melitensis DNA,providing new ideas and new targets for prevention and treatment of brucellosis.Methods Bioinformatics methods were used to detect repetitive extragenic palindromic(REP) sequences from Brucella melitensis DNA.The studied REPs were selected and synthesized.RAW264.7 was cultured and transfected with REPs mediated by lipofectamine 3000.Additionally,TLR9-siRNA was used to downregulate TLR9 expression.The content of interferon-α(IFN-α) in the supernatant was then measured by ELISA.Results A total of 2 200 REP sequences in Brucella melitensis DNA were identified.Twelve REP sequences were synthesized for further detecting of the TLR9 agonistic activity.IFN-α expression in RAW264.7 treated with M2,M3,M4,M5,M6,M7,M9,M12 were (26.944 ± 1.868),(46.461 ± 2.562),(34.980 ± 2.055),(43.016 ± 2.162),(62.533 ± 4.031),(67.125 ± 5.069),(18.908 ± 1.633),(39.572 ± 2.465) pg/ml respectively,which significantly increased when compared with the negative control group [(12.594 ± 1.338) pg/ml,t =10.817,20.295,15.812,20.724,20.365,18.016,5.180,16.660,all P < 0.05].Additionally,TLR9-siRNA can significantly decrease the levels of IFN-α in RAW264.7 treated with M6.Conclusion REP sequences presented in Brucella melitensis DNA are able to induce IFN-α expression through TLR9,which can be helpful for the understanding of pathogenesis and immunity of Brucella melitensis.%目的 筛选具有活化Toll样受体9(TLR9)活性的羊种布氏杆菌DNA中基因外重复回文序列(REPs),检测其经TLR9诱导的干扰素-α(IFN-α)表达,为羊种布氏杆菌病的防治提供新思路.方法 针对羊种布氏杆菌Brucella melitensis NI基因组序列,利用生物信息学技术识别其REPs后,合成序列.将合成的天然骨架脱氧寡核苷酸(ODNs)转染小鼠单核巨噬细胞株RAW264.7,酶联免疫吸附测定法(ELISA)检测IFN

  12. Prime-booster vaccination of cattle with an influenza viral vector Brucella abortus vaccine induces a long-term protective immune response against Brucella abortus infection.

    Science.gov (United States)

    Tabynov, Kaissar; Yespembetov, Bolat; Ryskeldinova, Sholpan; Zinina, Nadezhda; Kydyrbayev, Zhailaubay; Kozhamkulov, Yerken; Inkarbekov, Dulat; Sansyzbay, Abylai

    2016-01-20

    This study analyzed the duration of the antigen-specific humoral and T-cell immune responses and protectiveness of a recently-developed influenza viral vector Brucella abortus (Flu-BA) vaccine expressing Brucella proteins Omp16 and L7/L12 and containing the adjuvant Montadine Gel01 in cattle. At 1 month post-booster vaccination (BV), both humoral (up to 3 months post-BV; GMT IgG ELISA titer 214±55 to 857±136, with a prevalence of IgG2a over IgG1 isotype antibodies) and T-cell immune responses were observed in vaccinated heifers (n=35) compared to control animals (n=35, injected with adjuvant/PBS only). A pronounced T-cell immune response was induced and maintained for 12 months post-BV, as indicated by the lymphocyte stimulation index (2.7±0.4 to 10.1±0.9 cpm) and production of IFN-γ (13.7±1.7 to 40.0±3.0 ng/ml) at 3, 6, 9, and 12 months post-BV. Prime-boost vaccination provided significant protection against B. abortus infection at 3, 6, 9 and 12 months (study duration) post-BV (7 heifers per time point; alpha=0.03-0.01 vs. control group). Between 57.1 and 71.4% of vaccinated animals showed no signs of B. abortus infection (or Brucella isolation) at 3, 6, 9 and 12 months post-BV; the severity of infection, as indicated by the index of infection (P=0.0003 to Brucella colonization (P=0.03 to abortus infection was also observed among pregnant vaccinated heifers (alpha=0.03), as well as their fetuses and calves (alpha=0.01), for 12 months post-BV. Additionally, 71.4% of vaccinated heifers calved successfully whereas all pregnant control animals aborted (alpha=0.01). Prime-boost vaccination of cattle with Flu-BA induces an antigen-specific humoral and pronounced T cell immune response and most importantly provides good protectiveness, even in pregnant heifers, for at least 12 months post-BV.

  13. Association of Maedi Visna virus with Brucella ovis infection in rams

    Directory of Open Access Journals (Sweden)

    S Preziuso

    2009-06-01

    Full Text Available Maedi Visna Virus (MVV is the etiological agent of a systemic disease of sheep, which causes lesions in lungs, the central nervous system, joints, and mammary glands. It has been speculated that the association with Brucella ovis may lead to the venereal shedding of the virus. In this work, samples of epididymis from ten rams positive for MVV and infected experimentally with Brucella ovis, were subjected to liquid- phase PCR, immunohistochemistry (IHC and in situ PCR tests, aimed at identifying the pathogens in a tissue context. IHC was carried out using a monoclonal antibody raised against p28 MVV protein and a polyclonal antibody to B. ovis. Liquid phase- and in situ PCR were designed to amplify a portion of MVV proviral DNA Pol sequence. In the animals showing B. ovis-related histopathological changes, IHC clearly demonstrated a positivity for B. ovis and MVV in interstitial and epithelial ductal cells. In situ PCR assessed the presence of MVV proviral DNA in macrophages and elements inside the epithelium. The unaffected and reagent control samples constantly gave negative results. Taken together, these data demonstrate that MVV may affect ovine epididymis, apparently taking advantage of the concurrent infection by B. ovis. The tropism of MVV for the epididymal epithelial cells, may be responsible for its excretion with the semen.

  14. 5-Lipoxygenase negatively regulates Th1 response during Brucella abortus infection in mice.

    Science.gov (United States)

    Fahel, Júlia Silveira; de Souza, Mariana Bueno; Gomes, Marco Túlio Ribeiro; Corsetti, Patricia P; Carvalho, Natalia B; Marinho, Fabio A V; de Almeida, Leonardo A; Caliari, Marcelo V; Machado, Fabiana Simão; Oliveira, Sergio Costa

    2015-03-01

    Brucella abortus is a Gram-negative bacterium that infects humans and cattle, causing a chronic inflammatory disease known as brucellosis. A Th1-mediated immune response plays a critical role in host control of this pathogen. Recent findings indicate contrasting roles for lipid mediators in host responses against infections. 5-Lipoxygenase (5-LO) is an enzyme required for the production of the lipid mediators leukotrienes and lipoxins. To determine the involvement of 5-LO in host responses to B. abortus infection, we intraperitoneally infected wild-type and 5-LO-deficient mice and evaluated the progression of infection and concomitant expression of immune mediators. Here, we demonstrate that B. abortus induced the upregulation of 5-LO mRNA in wild-type mice. Moreover, this pathogen upregulated the production of the lipid mediators leukotriene B4 and lipoxin A4 in a 5-LO-dependent manner. 5-LO-deficient mice displayed lower bacterial burdens in the spleen and liver and less severe liver pathology, demonstrating an enhanced resistance to infection. Host resistance paralleled an increased expression of the proinflammatory mediators interleukin-12 (IL-12), gamma interferon (IFN-γ), and inducible nitric oxide synthase (iNOS) during the course of infection. Moreover, we demonstrated that 5-LO downregulated the expression of IL-12 in macrophages during B. abortus infection. Our results suggest that 5-LO has a major involvement in B. abortus infection, by functioning as a negative regulator of the protective Th1 immune responses against this pathogen.

  15. In vitro drug resistance of clinical isolated Brucella against antimicrobial agents

    Institute of Scientific and Technical Information of China (English)

    Xiu-Li Xu; Xiao Chen; Pei-Hong Yang; Jia-Yun Liu; Xiao-Ke Hao

    2013-01-01

    Objective:To explore the antibiotic resistance of Brucella melitensisand instruct rational use of antimicrobial agents in clinical treatment ofBrucella infection.Methods:Bacteria were cultured and identified byBACTEC9120 andVITEKⅡ automicrobic system.E-test was used to detect the minimal inhibitory concentration(MIC) of antimicrobial agents in the drug susceptivity experiment.Results:A total of19 brucella strains(allBrucella melitensis) wereisolated from19 patients, who had fever betweenJanuary2010 andJune2012, and17 samples were blood, one was bone marrow, the other sample was cerebrospinal fluid.TheMIC range of ceftazidime was2.0-8.0 mg/L, rifampicin was0.06-2.0 mg/L, amikacin was4.0-12.0 mg/L, levofloxacin was2.0-8.0 mg/L, doxycycline was8.0-32.0 mg/L, sulfamethoxazole-trimethoprim was4.0-16.0 mg/L, ampicillin was1.5-2.0 mg/L and gentamicin was0.50-0.75 mg/L.Conclusions:The drugs used in this experiment cover common drugs for treatingBrcella.Meanwhile, the results are consistent with clinical efficacy.It is suggested personalized regimen according to patients’ status in treatment of Brucella.

  16. [Immune response and reproductive consequences in experimentally infected ewes with Brucella ovis during late pregnancy].

    Science.gov (United States)

    Paolicchi, Fernando A; Nuñez, Marta; Fiorentino, María A; Malena, Rosana C; Trangoni, Marcos; Cravero, Silvio; Estein, Silvia M

    2013-01-01

    Ovine brucellosis by Brucella ovis is a highly prevalent disease in Argentina. This study aimed to evaluate the pathogenicity of B. ovis and the serological response in ewes during late pregnancy and in their offspring. Six adult ewes were distributed in two groupsG1 (pregnant females, n = 4) and G2 (nonpregnant females, n = 2). Three pregnant ewes at 15 days prepartum and one nonpregnant eve were inoculated with B. ovis. Sera of sheep and their offspring were analyzed by different serological tests. Samples of cervicovaginal mucus, placenta and milk were studied by bacteriology. A Brucella genus-specific PCR assay was carried out in placenta and milk samples. Placenta samples were hystopathologically processed. g1 females gave birth to live lambs, but one died hours postpartum. Serological techniques employed detected antibodies in serum of inoculated pregnant animal 5 days postchallenge. sera of female controls G1 and G2 remained negative throughout the study. Cervicovaginal mucus of infected ewes in G1 and G2 yielded negative results to bacteriology, but B. ovis was isolated from milk. The PCR assay was positive for the placenta and milk from inoculated pregnant ewes. Histopathology revealed necrotic suppurative placentitis in one placenta. However, although results demonstrated that B. ovis can invade the placenta and mammary gland, this bacterium did not cause abortion when it was inoculated intravenously at 15 days prepartum. B. ovis infection induced an early humoral response in pregnant ewes, but their lambs remained seronegative, indicating that there was no transfer of antibodies in infancy. Placenta colonization and milk excretion of B. ovis involves a potential source of infection for lambs, which could play a role as latent carriers of infection.

  17. 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.

  18. Murine and bovine γδ T cells enhance innate immunity against Brucella abortus infections.

    Directory of Open Access Journals (Sweden)

    Jerod A Skyberg

    Full Text Available γδ T cells have been postulated to act as a first line of defense against infectious agents, particularly intracellular pathogens, representing an important link between the innate and adaptive immune responses. Human γδ T cells expand in the blood of brucellosis patients and are active against Brucella in vitro. However, the role of γδ T cells in vivo during experimental brucellosis has not been studied. Here we report TCRδ(-/- mice are more susceptible to B. abortus infection than C57BL/6 mice at one week post-infection as measured by splenic colonization and splenomegaly. An increase in TCRγδ cells was observed in the spleens of B. abortus-infected C57BL/6 mice, which peaked at two weeks post-infection and occurred concomitantly with diminished brucellae. γδ T cells were the major source of IL-17 following infection and also produced IFN-γ. Depletion of γδ T cells from C57BL/6, IL-17Rα(-/-, and GMCSF(-/- mice enhanced susceptibility to B. abortus infection although this susceptibility was unaltered in the mutant mice; however, when γδ T cells were depleted from IFN-γ(-/- mice, enhanced susceptibility was observed. Neutralization of γδ T cells in the absence of TNF-α did not further impair immunity. In the absence of TNF-α or γδ T cells, B. abortus-infected mice showed enhanced IFN-γ, suggesting that they augmented production to compensate for the loss of γδ T cells and/or TNF-α. While the protective role of γδ T cells was TNF-α-dependent, γδ T cells were not the major source of TNF-α and activation of γδ T cells following B. abortus infection was TNF-α-independent. Additionally, bovine TCRγδ cells were found to respond rapidly to B. abortus infection upon co-culture with autologous macrophages and could impair the intramacrophage replication of B. abortus via IFN-γ. Collectively, these results demonstrate γδ T cells are important for early protection to B. abortus infections.

  19. Laboratory exposure to Brucella melitensis in Denmark

    DEFF Research Database (Denmark)

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

    2013-01-01

    for Disease Control and Prevention guidelines, but, of 14 staff members classified as high-risk exposure, none accepted post-exposure prophylaxis. However, in a period of 6 months of follow-up, none of the exposed laboratory workers developed brucellosis and all obtained sera were negative for antibrucella...

  20. Full Restoration of Brucella-Infected Dendritic Cell Functionality through Vγ9Vδ2 T Helper Type 1 Crosstalk

    OpenAIRE

    Ming Ni; Delphine Martire; Emmanuel Scotet; Marc Bonneville; Francoise Sanchez; Virginie Lafont

    2012-01-01

    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 dramatical...

  1. Serosurvey of Brucella spp. infection in the Kafue lechwe (Kobus leche kafuensis) of the Kafue flats in Zambia.

    Science.gov (United States)

    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.

  2. Milk Ring Test for spot identification of Brucella abortus infection in single cow herds

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  4. Brucellae through the food chain : the role of sheep, goats and springbok (Antidorcus marsupialis) as sources of human infections in Namibia

    OpenAIRE

    K. Magwedere; A. Bishi; G. Tjipura-Zaire; Eberle, G; Y. Hemberger; L. C. Hoffman; Dziva, F

    2011-01-01

    A confirmed case of human brucellosis motivated an investigation into the potential source of infection in Namibia. Since domestic animals are principal sources of Brucella infection in humans, 1692 serum samples were screened from sheep, goats and cattle from 4 presumably at-risk farms and 900 springbok (Antidorcas marsupialis) serum samples from 29 mixed farming units for Brucella antibodies by the Rose-Bengal test (RBT) and positive cases confirmed by complement fixation test (CFT). To ass...

  5. The changing Brucella ecology: novel reservoirs, new threats.

    Science.gov (United States)

    Pappas, Georgios

    2010-11-01

    Brucellosis is a zoonosis that preceded humans but continues to cause significant medical, veterinary and socioeconomic problems, mainly because its overall burden remains underestimated and neglected. Its ecology, or what we know of it, has evolved rapidly in recent years. Two novel species, Brucella ceti and B. pinnipedialis, with the potential for causing human disease have been isolated from marine mammals. Another novel species, B. microti, has been isolated from wildlife animals, whilst B. inopinata has been isolated from a human case. An active spillover of Brucella between domestic animals and wildlife is also being recognised, with elk transmitting B. abortus to cattle, and freshwater fish becoming infected with B. melitensis from waste meat. In recent years the global epidemiology of the disease has not altered drastically, apart from increased awareness of brucellosis in sub-Saharan Africa and a rapid expansion of disease endemicity in the Balkan Peninsula. Isolated stories and events underline that Brucella knows no borders. The modern world has offered the pathogen the ability to travel and manifest itself anywhere and has also offered scientists the ability to track these manifestations better than ever before. This may allow the disease to be neglected no longer, or at least to be recognised as neglected.

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

    Directory of Open Access Journals (Sweden)

    L.F. Costa

    2013-02-01

    Full Text Available The aim of the present study was to evaluate a species-specific nested PCR based on a previously described species-specific PCR for detection of B. ovis in semen and urine samples of experimentally infected rams. The performance of the species-specific nested PCR was compared with the results of a genus-specific PCR. Fourteen rams were experimentally infected with the Brucella ovis REO 198 strain and samples of semen and urine were collected every week up to 180 days post infection. Out of 83 semen samples collected, 42 (50.6% were positive for the species-specific nested PCR, and 23 (27.7% were positive for the genus-specific PCR. Out of 75 urine samples, 49 (65.3% were positive for the species-specific nested PCR, whereas 11 (14.6% were genus-specific PCR positive. Species-specific nested PCR was significantly more sensitive (P<0.001 than the genus-specific PCR in semen and urine from experimentally infected rams. In conclusion, the species-specific nested PCR developed in this study may be used as a diagnostic tool for the detection of B. ovis in semen and urine samples from suspected rams.

  7. The virB-encoded type IV secretion system is critical for establishment of infection and persistence of Brucella ovis infection in mice.

    Science.gov (United States)

    Sá, Joicy C; Silva, Teane M A; Costa, Erica A; Silva, Ana P C; Tsolis, Renée M; Paixão, Tatiane A; Carvalho Neta, Alcina V; Santos, Renato L

    2012-09-14

    Brucella spp. are gram-negative intracellular bacterial pathogens that cause chronic infections. Brucella virulence factors include a type IV secretion system (T4SS) and its lipopolysaccharide (LPS), which are essential for persistence. However, the role of the virB-encoded T4SS has not been investigated in naturally rough Brucella species such as Brucella ovis. In this study, male 6-week old BALBc mice were infected with B. ovis, Brucella abortus, and their respective ΔvirB2 mutant strains. During early infection, B. ovis and B. abortus wild type strains were similarly recovered from spleen. Interestingly, in contrast to ΔvirB2 B. abortus that was recovered at similar levels when compared to the wild type strain, the ΔvirB2 B. ovis was markedly attenuated as early as 24h post infection (hpi). The ΔvirB2 B. ovis was unable to survive and multiply in murine peritoneal macrophages and extracellularly within the peritoneal cavity at 12 and 24 hpi with lower splenic colonization than the parental strain at 6, 12 and 24 hpi. In contrast, wild type B. abortus and ΔvirB2 B. abortus had a similar kinetics of infection in this model. As expected, the T4SS was essential for intracellular replication of smooth and rough strains in RAW macrophages at 48 hpi. These results suggest that T4SS is important for survival of B. ovis in murine model, and that a T4SS deficient B. ovis strain is cleared at earlier stages of infection when compared to a similar B. abortus mutant.

  8. COX-2 Inhibition Reduces Brucella Bacterial Burden in Draining Lymph Nodes

    Science.gov (United States)

    Gagnaire, Aurélie; Gorvel, Laurent; Papadopoulos, Alexia; Von Bargen, Kristine; Mège, Jean-Louis; Gorvel, Jean-Pierre

    2016-01-01

    Brucella is a Gram-negative facultative intracellular bacterium responsible for a chronic disease known as brucellosis, the most widespread re-emerging zoonosis worldwide. Establishment of a Th1-mediated immune response characterized by the production of IL-12 and IFNγ is essential to control the disease. Leukotrienes derived from arachidonic acid (AA) metabolism are known to negatively regulate a protective Th1 immune response against bacterial infections. Here, using genomics approaches we demonstrate that Brucella abortus strongly stimulates the prostaglandin (PG) pathway in dendritic cells (DC). We also show an induction of AA production by infected cells. This correlates with the expression of Ptgs2, a gene encoding the downstream cyclooxygenase-2 (COX-2) enzyme in infected DC. By comparing different infection routes (oral, intradermal, intranasal and conjunctival), we identified the intradermal inoculation route as the more potent in inducing Ptgs2 expression but also in inducing a local inflammatory response in the draining cervical lymph nodes (CLN). NS-398, a specific inhibitor of COX-2 enzymatic activity decreased B. melitensis burden in the CLN after intradermal infection. This effect was accompanied by a decrease of Il10 and a concomitant increase of Ifng expression. Altogether, these results suggest that Brucella has evolved to take advantage of the PG pathway in the harsh environment of the CLN in order to persist and subvert immune responses. This work also proposes that novel strategies to control brucellosis may include the use of COX-2 inhibitors. PMID:28018318

  9. In situ repair of a primary Brucella-infected abdominal aortic aneurysm: long-term follow-up.

    Science.gov (United States)

    Goudard, Yvain; Pierret, Charles; de La Villéon, Bruno; Mlynski, Amélie; de Kerangal, Xavier

    2013-02-01

    Infected aortic aneurysms represent 0.85 to 1.3% of aortic aneurysms. Most often, the implicated bacteria species are Salmonella sp., Staphylococcus sp. and Streptococcus sp. Brucella-related infected aortic aneurysms are very rare. Most often, they result from endocarditis or from a local septic focus. Combined treatment by antibiotics and surgery is the standard for infectious aneurysms. In the absence of formal factual data, the surgical treatment is still discussed in the literature, especially since endovascular treatments have been in full expansion. We are reporting the case of a female patient presenting with a Brucella-related infra-renal abdominal aortic aneurysm, without primitive infectious source (area) or identified endocarditis. Surgical treatment with in situ prosthetic replacement and omentoplasty in association with adapted antibiotics allowed a favorable outcome with an excellent result after an 8-year follow up.

  10. Research Status and Prospect of Brucella Infection in Our Country%我国布鲁氏菌感染的研究现状及展望

    Institute of Scientific and Technical Information of China (English)

    成岩; 白靓; 张树军

    2012-01-01

    建国后,我国在布鲁氏菌感染的研究方面取得了显著成绩,建立了布鲁氏菌感染监测平台,研发、生产出了能够预防布鲁氏菌感染的疫苗,近几年还阐释了一些与布鲁氏菌致病相关的功能基因.但是由于一些原因导致近几年疫情急骤回升,引起国际社会广泛关注.作者综述了布鲁氏菌感染的研究现状、流行趋势及防控进展,旨在加深对布鲁氏菌感染的认识,为推动对布鲁氏菌的深入研究作贡献.%After the founding of China,our country has made remarkable achievements in Brucella infection research,such as establishment of the monitoring platform of Brucella infection,research and development,production of Brucella abortus vaccine to prevent infection.In recent years,some functional genes associate with Brucella virulence also has been illustrated.But because of some reasons,Chinese Brucella epidemic rapidly rise,this phenomenon has caused widespread concern of international community.The paper reviews the research status of Brucella infection,epidemic trend and prevention progress,aims to deepen the understanding of Brucella infection,to promote the further study of Brucella.

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

    Institute of Scientific and Technical Information of China (English)

    Reza Mirnejad; Mozafar mohamadi; Vahbeh Piranfar; Seied Mojtaba Mortazavi; Reza Kachuei

    2013-01-01

    Objective: To design a duplex PCR for rapid and simultaneous detection of Brucella species. in human blood samples. Methods: 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. Results: Of the 52 peripheral bloods samples tested, 25 sample (48%) showed positive reactions in PCR. Twelve samples were positive for Brucella abortus (B. abortus) (23%), 13 for Brucella melitensis (B. melitensis) (25%) and 0 for Brucella ovis (B. ovis) (0%). Conclusions: This work de=monstrates 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.

  12. Evidence for unapparent Brucella canis infections among adults with occupational exposure to dogs.

    Science.gov (United States)

    Krueger, W S; Lucero, N E; Brower, A; Heil, G L; Gray, G C

    2014-11-01

    Human serological assays designed to detect brucellosis will miss infections caused by Brucella canis, and low levels of periodic bacteremia limit diagnosis by blood culture. Recent B. canis outbreaks in dogs and concomitant illnesses in caretakers suggest that unapparent human infections may be occurring. With more than a quarter of a million persons in occupations involving dogs, and nearly 80 million dog owners in the United States, this pathogen is an under-recognized human health threat. To investigate occupational exposure to B. canis, we adapted a commercial canine serological assay and present the first controlled seroepidemiological study of human B. canis infections in recent years. 306 adults with occupational exposure to dogs and 101 non-matched, non-canine-exposed subjects were enrolled. Antibodies were detected using the canine D-Tec(®) CB rapid slide agglutination test (RSAT) kit with a secondary 2-mercaptoethanol (ME)-RSAT. Results were validated on a blinded subset of sera with an additional RSAT and indirect enzyme-linked immunoassay at the National Administration of Laboratories and Health Institutes (ANLIS) in Argentina. Seroprevalence ranged from 10.8% (RSAT) to 3.6% (ME-RSAT) among canine-exposed subjects. Kennel employees were more likely to test RSAT seropositive compared with other canine exposures (OR = 2.7; 95% CI, 1.3-5.8); however, low seroprevalence limited meaningful occupational risk factor analyses. Two seropositive participants reported experiencing symptoms consistent with brucellosis and having exposure to B. canis-infected dogs; however, temporality of symptom onset with reported exposure could not be determined. D-Tec(®) CB results had substantial agreement with ANLIS assays (Cohen's kappa = 0.60-0.68). These data add to a growing body of literature suggesting that people occupationally exposed to dogs may be at risk of unapparent B. canis infection. It seems prudent to consider B. canis as an occupational public health

  13. 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

    2017-01-09

    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.

  14. Infection of C57BL/6 mice by Trypanosoma musculi modulates host immune responses during Brucella abortus cocolonization.

    Science.gov (United States)

    Lowry, Jake E; Leonhardt, Jack A; Yao, Chaoqun; Belden, E Lee; Andrews, Gerard P

    2014-01-01

    Brucellosis, which results in fetal abortions in domestic and wildlife animal populations, is of major concern in the US and throughout much of the world. The disease, caused by Brucella abortus, poses an economic threat to agriculture-based communities. A moderately efficacious live attenuated vaccine (B. abortus strain RB51) exists. However, even with vaccine use, outbreaks occur. Evidence suggests that elk (Cervus canadensis), a wild host reservoir, are the source of recent outbreaks in domestic cattle herds in Wyoming, USA. Brucella abortus establishes a chronic, persistent infection in elk. The molecular mechanisms allowing the establishment of this persistent infective state are currently unknown. A potential mechanism could be that concurrent pathogen burdens contribute to persistence. In Wyoming, elk are chronically infected with Trypanosoma cervi, which may modulate host responses in a similar manner to that documented for other trypanosomes. To identify any synergistic relationship between the two pathogens, we simulated coinfection in the well-established murine brucellosis model using Trypanosoma musculi and B. abortus S19. Groups of C57BL/6 mice (Mus musculus) were infected with either B. abortus strain 19 (S19) or T. musculi or both. Sera were collected weekly; spleens from euthanized mice were tested to determine bacterial load near the end of normal brucellosis infection. Although changes in bacterial load were observed during the later stages of brucellosis in those mice coinfected with T. musculi, the most significant finding was the suppression of gamma interferon early during the infection along with an increase in interleukin-10 secretion compared with mice infected with either pathogen alone. These results suggest that immune modulatory events occur in the mouse during coinfection and that further experiments are warranted to determine if T. cervi impacts Brucella infection in elk.

  15. Epidemiological Survey of Brucella canis Infection in Different Breeds of Dogs in Fars Province, Iran

    Directory of Open Access Journals (Sweden)

    Mohammad Amin Behzadi and Asghar Mogheiseh1*

    2012-05-01

    Full Text Available This study was conducted to determine the prevalence of Brucella canis antibodies in different breeds, sex and ages of dogs in southern of Iran. A total of 113 whole blood samples were taken from different breeds based on exotic or native sources. The samples were examined with immunochromatography assay for detection of B. canis antibodies. Twelve dogs were serologically positive (10.62%. There was significant differences in ratio of infected dogs between breeds (exotic or native, ages (less, equal or more than 2 years old and the history of vaccination (against rabies, leptospirosis, parvovirus, adenovirus type 2, canine distemper, parainfluenza (P<0.001. However, the results were not significant statistically, among both sex (P=0.058 and the history of clinical signs (P=0.456 in seropositive dogs. Based on this study and the other investigation in companion dogs from southwest of Iran, it seems that the mixed and spray (native breeds are not infected with B. canis, yet. Conversely, the exotic breeds would be the source of bacterium in Iran. Therefore, preventive and control measures are strongly recommended.

  16. Immune Modulation of Recombinant OmpA against Brucella abortus 544 Infection in Mice.

    Science.gov (United States)

    Simborio, Hannah Leah Tadeja; Reyes, Alisha Wehdnesday Bernardo; Hop, Huynh Tan; Arayan, Lauren Togonon; Min, Wongi; Lee, Hu Jang; Lee, Jin Ju; Chang, Hong Hee; Kim, Suk

    2016-03-01

    Brucellosis affects a wide range of host species, including humans and many livestock animals. Chronic infections of the disease make antibiotic treatment costly, and the current vaccine used in livestock has not been approved for human use. This study investigated the possible use of the Brucella abortus outer membrane protein A (OmpA) as a candidate subunit vaccine in an infected mouse model. The ompA gene was cloned and overexpressed, and the recombinant OmpA (rOmpA) protein fused to maltose binding protein (MBP) was purified in Escherichia coli. Immunogenicity was verified through western blotting, and mice were immunized and challenged to evaluate its protective effect. Mice treated with rOmpA exhibited induced humoral and host cell-mediated responses, with a significant increase in immunoglobulin G (IgG1 and IgG2a) and cytokine levels, especially TNF-α and IL-12, compared with the control groups treated with either MBP or PBS. In conclusion, rOmpA should be highly considered as a future subunit vaccine for brucellosis, and further studies regarding rOmpA and its protective ability are suggested.

  17. Determinación de los tejidos y medios de elección para la confirmación microbiológica de los resultados serológicos de las campañas de control de Brucella melitensis en el ganado ovino Determination of best tissues and culture media for microbiological confirmation of the serological results of the control campaigns of Brucella melitensis in sheep

    Directory of Open Access Journals (Sweden)

    L Álvarez

    2010-01-01

    Full Text Available El sacrificio de animales seropositivos para el control de la brucelosis ovina requiere del aislamiento microbiológico como técnica de confirmación del proceso. Debido a la dificultad de realización de las técnicas microbiológicas en un elevado número de animales, así como de la gran cantidad de muestras, es necesario establecer qué tejidos son los más adecuados para ello. No existen datos al respecto obtenidos de animales infectados naturalmente y representativos de las distintas situaciones epidemiológicas tal y como recomienda la Organización Internacional de Epizootias. En nuestro trabajo realizamos la necropsia de 92 animales sospechosos de infección brucelar efectuando posteriormente el cultivo e identificación microbiológica de 10 tejidos diferentes. Los animales fueron clasificados en función de una encuesta epidemiológica como pertenecientes a Rebaños con Brucelosis Crónica, Activa o de muy Baja Prevalencia. Los resultados muestran diferencias en función del tipo de rebaño, así como indican que la toma de muestras de mama y linfonódulos mamarios es la más adecuada para la confirmación de la infección en un rebaño sospechoso. El uso combinado de los medios de cultivo Farrell y Thayer Martin mostró una mayor eficacia según los resultados obtenidos por nosotros.Slaughtering seropositive animals in order to control ovine brucellosis requires a microbiological culture as a confirmatory technique. It is necessary to establish the most adequate tissues, due to difficulties involved in performing microbiological techniques in a large number of animals and samples. In this sense, there is a lack of data obtained from naturally infected sheep representatives of the different epidemiological situations, as recommended by OIE. In our study we carried out the necropsy of 92 ewes suspicious to be infected by Brucella and we performed microbiological culture and identification of 10 tissues per animal. Animals were

  18. Molecular detection of Brucella species in patients suspicious of Brucellosis from Zanjan, Iran

    Directory of Open Access Journals (Sweden)

    Maryam Garshasbi

    2014-06-01

    Full Text Available Brucella is an intracellular pathogen capable of infecting animals and humans. The aim of this study was to identify Brucella spp in sera of high risk individuals by a polymerase chain reaction (PCR-based method. A total of 180 patients suspected to have Brucellosis were examined by serological tests. To establish a PCR protocol for diagnosis of active brucellosis, DNA was extracted from the serum samples by using a commercial kit. PCR amplification was done for detection of Brocella DNA using BCSP31 target gene and IS711 locus. The PCR assay showed that an amplicon of 223 bp was obtained in 73.8% (133/180 of the tested sera using primers (B4/B5 derived from a gene encoding the 31-kDa Brucella abortus antigen. In another PCR, an amplicon of 498 bp was obtained in 63.8% (115/180 of the samples using Brucella abortus-specific primers derived from a locus adjacent to the 3'-end of IS711, and also an amplicon of 731 bp was produced in 4.4% (8/180 of the tested samples using Brucella melitensis-specific primers. When the Wright method was used as a gold standard, the sensitivity and specificity of the PCR technique for genus identification were found to be 96 and 80.7%, respectively. However, the sensitivity value obtained with the species-specific PCR method was 82%, and specificity was similar to that previous reported. This is the first report of a high frequency of Brucella abortus in patients suspicious of Brucellosis from the Zanjan province.

  19. Immunoproteomic identification of immunodominant antigens independent of the time of infection in Brucella abortus 2308-challenged cattle.

    Science.gov (United States)

    Lee, Jin Ju; Simborio, Hannah Leah; Reyes, Alisha Wehdnesday Bernardo; Kim, Dae Geun; Hop, Huynh Tan; Min, Wongi; Her, Moon; Jung, Suk Chan; Yoo, Han Sang; Kim, Suk

    2015-03-01

    Brucellosis is a vital zoonotic disease caused by Brucella, which infects a wide range of animals and humans. Accurate diagnosis and reliable vaccination can control brucellosis in domestic animals. This study examined novel immunogenic proteins that can be used to detect Brucella abortus infection or as an effective subcellular vaccine. In an immunoproteomic assay, 55 immunodominant proteins from B. abortus 544 were observed using two dimensional electrophoresis (2DE) and immunoblot profiles with antisera from B. abortus-infected cattle at the early (week 3), middle (week 7), and late (week 10) periods, after excluding protein spots reacting with antisera from Yersinia enterocolitica O:9-infected and non-infected cattle. Twenty-three selected immunodominant proteins whose spots were observed at all three infection periods were identified using MALDI-MS/MS. Most of these proteins identified by immunoblot and mass spectrometry were determined by their subcellular localization and predicted function. We suggest that the detection of prominent immunogenic proteins during the infection period can support the development of advanced diagnostic methods with high specificity and accuracy; subsidiarily, these proteins can provide supporting data to aid in developing novel vaccine candidates.

  20. A polymerase chain reaction for detection of Brucella canis in vaginal swabs of naturally infected bitches.

    Science.gov (United States)

    Keid, L B; Soares, R M; Vasconcellos, S A; Chiebao, D P; Salgado, V R; Megid, J; Richtzenhain, L J

    2007-12-01

    A PCR assay for the detection of Brucella canis in canine vaginal swab samples was evaluated, comparing its performance with that of bacterial isolation, serological tests, and a blood PCR assay. One hundred and forty-four female dogs were clinically examined to detect reproductive problems and they were tested by the rapid slide agglutination test, with and without 2-mercaptoethanol (2ME-RSAT and RSAT, respectively). In addition, microbiological culture and PCR were performed on blood and vaginal swab samples. The results of the vaginal swab PCR were compared to those of the other tests using the Kappa coefficient and McNemar test. Of the 144 females that were examined, 66 (45.8%) were RSAT positive, 23 (15.9%) were 2ME-RSAT positive, 49 (34.02%) were blood culture positive, 6 (4.1%) were vaginal swab culture positive, 54 (37.5%) were blood PCR positive, 52 (36.2%) were vaginal swab PCR positive, and 50.69% (73/144) were positive by the combined PCR. The PCR was able to detect as few as 3.8 fg of B. canis DNA experimentally diluted in 54 ng of canine DNA, extracted from vaginal swab samples of non-infected bitches. In addition, the PCR assay amplified B. canis genetic sequences from vaginal swab samples containing 1.0 x 10(0) cfu/mL. In conclusion, vaginal swab PCR was a good candidate as a confirmatory test for brucellosis diagnosis in bitches suspected to be infected, especially those negative on blood culture or blood PCR; these animals may be important reservoirs of infection and could complicate attempts to eradicate the disease in confined populations.

  1. On the link between cell cycle and infection of the Alphaproteobacterium Brucella abortus

    Directory of Open Access Journals (Sweden)

    Michaël Deghelt

    2014-09-01

    Full Text Available Bacteria of the Brucella genus are responsible for brucellosis, a worldwide zoonosis. These bacteria are known to have a peculiar intracellular trafficking, with a first long and non-proliferative endosomal stage and a second proliferation stage, often associated with its localization of the bacteria in the endoplasmic reticulum (ER. However, the status of the bacterial cell cycle during the non-proliferative phase was still unknown. In a recent study [Nat. Communic. 5:4366], we followed the cell cycle of B. abortus in culture and inside the host cells. In culture, B. abortus initiates the replication of its large chromosome before the small chromosome. The origin and terminator regions of these two chromosomes display distinct localization and dynamics within B. abortus. In HeLa cells and RAW264.7 macrophages, the bacteria in G1 (i.e. before the initiation of chromosomes replication are preferentially found during the endosomal stage of the infection. During this period, growth is also arrested. The cell cycle arrest and resume during the B. abortus trafficking in host cell suggest that like the model Alphaproteobacterium Caulobacter crescentus, these bacteria are able to block their cell cycle at the G1 phase when starvation is sensed.

  2. 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.

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

    Science.gov (United States)

    Fifty years ago, bacteria in the genus Brucella were known to cause infertility and reproductive losses. The genus was considered to contain only three species, B. abortus, B. melitensis and B. suis. Since the early 1960’s, at least seven new species have been identified as belonging to the Brucell...

  4. MLVA genotyping of human Brucella isolates from Peru

    NARCIS (Netherlands)

    H.L. Smits; B. Espinosa; R. Castillo; E. Hall; A. Guillen; M. Zevaleta; R.H. Gilman; P. Melendez; C. Guerra; A. Draeger; A. Broglia; K. Nöckler

    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 genotypes

  5. Critical role of ASC inflammasomes and bacterial type IV secretion system in caspase-1 activation and host innate resistance to Brucella abortus infection.

    Science.gov (United States)

    Gomes, Marco Tulio R; Campos, Priscila C; Oliveira, Fernanda S; Corsetti, Patricia P; Bortoluci, Karina R; Cunha, Larissa D; Zamboni, Dario S; Oliveira, Sergio C

    2013-04-01

    Pathogens are detected by innate immune receptors that, upon activation, orchestrate an appropriate immune response. Recent studies revealed the intracellular signaling cascades involved in the TLR-initiated immune response to Brucella abortus infection. However, no report has elucidated the role of inflammasome receptors in Brucella recognition. Therefore, we decided to investigate the function of NLRC4, NLRP3, and AIM2 in sensing Brucella. In this study, we showed that NLRC4 is not required to induce caspase-1 activation and further secretion of IL-1β by B. abortus in macrophages. In contrast, we determined that AIM2, which senses Brucella DNA, and NLRP3 are partially required for caspase-1 activation and IL-1β secretion. Additionally, mitochondrial reactive oxygen species induced by Brucella were implicated in IL-1β production. Furthermore, AIM2, NLRP3, ASC, and caspase-1 knockout mice were more susceptible to B. abortus infection than were wild-type animals, suggesting that multiple ASC-dependent inflammasomes contribute to host protection against infection. This protective effect is due to the inflammatory response caused by IL-1β and IL-18 rather than pyroptosis, because we observed augmented bacterial burden in IL-1R and IL-18 knockout mice. Finally, we determined that bacterial type IV secretion system VirB and live, but not heat-killed, Brucella are required for full inflammasome activation in macrophages during infection. Taken together, our results indicate that Brucella is sensed by ASC inflammasomes that collectively orchestrate a robust caspase-1 activation and proinflammatory response.

  6. MyD88 and STING signaling pathways are required for IRF3-mediated IFN-β induction in response to Brucella abortus infection.

    Directory of Open Access Journals (Sweden)

    Leonardo A de Almeida

    Full Text Available Type I interferons (IFNs are cytokines that orchestrate diverse immune responses to viral and bacterial infections. Although typically considered to be most important molecules in response to viruses, type I IFNs are also induced by most, if not all, bacterial pathogens. In this study, we addressed the role of type I IFN signaling during Brucella abortus infection, a facultative intracellular bacterial pathogen that causes abortion in domestic animals and undulant fever in humans. Herein, we have shown that B. abortus induced IFN-β in macrophages and splenocytes. Further, IFN-β induction by Brucella was mediated by IRF3 signaling pathway and activates IFN-stimulated genes via STAT1 phosphorylation. In addition, IFN-β expression induced by Brucella is independent of TLRs and TRIF signaling but MyD88-dependent, a pathway not yet described for Gram-negative bacteria. Furthermore, we have identified Brucella DNA as the major bacterial component to induce IFN-β and our study revealed that this molecule operates through a mechanism dependent on RNA polymerase III to be sensed probably by an unknown receptor via the adaptor molecule STING. Finally, we have demonstrated that IFN-αβR KO mice are more resistant to infection suggesting that type I IFN signaling is detrimental to host control of Brucella. This resistance phenotype is accompanied by increased IFN-γ and NO production by IFN-αβR KO spleen cells and reduced apoptosis.

  7. Patterns of Hepatosplenic Brucella Abscesses on Cross-Sectional Imaging: A Review of Clinical and Imaging Features.

    Science.gov (United States)

    Heller, Tom; Bélard, Sabine; Wallrauch, Claudia; Carretto, Edoardo; Lissandrin, Raffaella; Filice, Carlo; Brunetti, Enrico

    2015-10-01

    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 abscesses caused by Brucella spp. are described and included in a review of 115 previously published cases. Clinical characteristics and patterns on ultrasound (US) and computed tomography imaging were analyzed. Furthermore, the proportion of patients with brucellosis affected by suppurative hepatosplenic lesions was estimated. Hepatosplenic abscesses were seen in 1.2% of patients with brucellosis and were mostly caused by Brucella melitensis. Imaging analysis revealed two main distinct patterns. Solitary abscesses involving liver more frequently than spleen, and showing characteristic central calcifications, characterize the first pattern. Multiple smaller abscesses, frequent spleen involvement, and absence of calcifications characterize the second pattern. Blood and aspirate cultures were frequently negative, however, the positivity rate increased over the past years. Indirect Coombs test was positive in 96%. Half of the patients were cured by antibiotic treatment; case fatality in this series was 1.9%. Hepatosplenic abscesses due to Brucella infections have characteristic imaging findings. Clinicians should be aware of these and the proactive use of cross-sectional imaging, particularly US, should be encouraged in endemic regions.

  8. The prevalence and impact of brucellosis in patients with hepatitis delta virus infection: inside the Brucella outbreak with cirrhosis

    Science.gov (United States)

    Dulger, Ahmet Cumhur; Suvak, Ozlem; Yesilyurt, Aysun Özel; Gultepe, Bilge; Guducuoglu, Huseyin

    2017-01-01

    Introduction Hepatitis D virus (HDV) infection is a serious health problem leading to cirrhosis and hepatocellular carcinoma (HCC). Despite evidence that zoonotic infections are associated with end-stage liver disease, brucellosis in patients with delta hepatitis related to liver disease has not been well characterized. So, we examined this relationship using recent hospital-based data. Material and methods We analyzed data from 96 delta hepatitis patients (mean age: 52.5 ±12.8 years; 50 male; 52 cirrhotics) and 117 (mean age: 50.4 ±7 years; 60 male) control subjects who were selected from patients with splenomegaly. The Brucella Wright test in connection with blood culture was used to detect active Brucella infection. Demographic features, laboratory data, results of ultrasonographic examination of the abdomen and Wright agglutination titers were compared between groups. Results There were 9 (9%) patients with active brucellosis in delta hepatitis patients. Compared to the control group, there was a statistically significant difference between groups in terms of having active brucellosis (9 vs. 2 patients; p brucellosis requiring hospitalization. Higher Wright titers among patients with more advanced liver disease may reflect a unique phenomenon that requires further investigation to determine underlying causative factors. PMID:28261291

  9. Prevalence of antibodies to Brucella spp. and individual risk factors of infection in traditional cattle, goats and sheep reared in livestock-wildlife interface areas of Zambia.

    Science.gov (United States)

    Muma, J B; Samui, K L; Siamudaala, V M; Oloya, J; Matop, G; Omer, M K; Munyeme, M; Mubita, C; Skjerve, E

    2006-04-01

    A cross-sectional study was performed in the livestock-wildlife interface areas of Lochinvar and Blue Lagoon National Parks and the non-interface area of Kazungula to determine the prevalence of antibodies to Brucella spp. in domestic ruminants and identify individual animal risk factors of infection. A total of 1245 cattle from 124 herds and 280 goats and sheep from 29 flocks were tested sequentially for Brucella antibodies using the Rose Bengal test (RBT) and competitive ELISA. In cattle, individual seroprevalence ranged from 14.1% to 28.1%, while herd sero-prevalence ranged from 46.2% to 74.0% in the three study areas. No goat or sheep tested positive for Brucella antibodies. Three types of cattle grazing strategies were encountered: locally grazed herds (LGH), transhumantly grazed herds (TGH) and river flood plain grazed herds (FGH). Brucella seroprevalence was seen to vary according to area and grazing strategy: Lochinvar and transhumant grazed herds recorded the highest figures, respectively. Age, sex and history of abortion were found to have independent effects on individual seroprevalence. This study establishes that brucellosis is endemic in domestic animals in the livestock-wildlife interface areas of Blue Lagoon and Lochinvar national parks and the disease is also present in Kazungula. We observed that type of grazing strategy had significant impact on cattle Brucella seroprevalence and that transhumant herds were at high risk of being infected.

  10. Infections and risk factors for livestock with species of Anaplasma, Babesia and Brucella under semi-nomadic rearing in Karamoja Region, Uganda.

    Science.gov (United States)

    Lolli, Chiara; Marenzoni, Maria Luisa; Strona, Paolo; Lappo, Pier Giorgio; Etiang, Patrick; Diverio, Silvana

    2016-03-01

    A survey was conducted to estimate the prevalence of Anaplasma, Babesia and Brucella spp. infections in cattle, goats and sheep in the Karamoja Region of Uganda and to identify possible risk factors existing in this semi-nomadic and pastoral area. Low cost laboratory tests were used to diagnose infections (Rose Bengal test for Brucella spp. antibodies and direct microscopic examination for Anaplasma and Babesia spp.). Multivariable logistic regression models were applied to identify possible risk factors linked to gender, animal species, age (only for cattle) and districts. A total of 3935 cattle, 729 goats and 306 sheep of five districts of the Karamoja Region were tested. Seroprevalence for Brucella was 9.2 % (CI, 95 %: 8.4-10), for Anaplasma 19.5 % (CI 95 %: 18.4-20.6) and for Babesia 16 % (CI 95 %: 15-17.1). Significant differences in infections prevalence were observed against risk factors associated with districts and species. Cattle were the species with higher risk of the infections. Female gender was identified as at risk only for Brucella spp. infection. Cattle more than one year old had greater likelihood to be Brucella seropositive. Co-infections of Anaplasma and Babesia spp. were statistically associated, especially in goats and sheep. Further studies to identify risk factors related to host species and geographical districts are needed. The influence on the semi-nomadic agro-pastoral system in Karamoja of animal raids and animal mixing should be further investigated. Findings were important to sensitize Karamojong undertaking measures on infection control, especially on cattle, which are their main source of food.

  11. Development of a dual vaccine for prevention of Brucella abortus infection and Escherichia coli O157:H7 intestinal colonization.

    Science.gov (United States)

    Iannino, Florencia; Herrmann, Claudia K; Roset, Mara S; Briones, Gabriel

    2015-05-05

    Zoonoses that affect human and animal health have an important economic impact. In the study now presented, a bivalent vaccine has been developed that has the potential for preventing the transmission from cattle to humans of two bacterial pathogens: Brucella abortus and Shiga toxin-producing Escherichia coli (STEC). A 66kDa chimeric antigen, composed by EspA, Intimin, Tir, and H7 flagellin (EITH7) from STEC, was constructed and expressed in B. abortus Δpgm vaccine strain (BabΔpgm). Mice orally immunized with BabΔpgm(EITH7) elicited an immune response with the induction of anti-EITH7 antibodies (IgA) that clears an intestinal infection of E. coli O157:H7 three times faster (t=4 days) than mice immunized with BabΔpgm carrier strain (t=12 days). As expected, mice immunized with BabΔpgm(EITH7) strain also elicited a protective immune response against B. abortus infection. A Brucella-based vaccine platform is described capable of eliciting a combined protective immune response against two bacterial pathogens with diverse lifestyles-the intracellular pathogen B. abortus and the intestinal extracellular pathogen STEC.

  12. 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.

  13. Doxycycline-rifampin versus doxycycline-streptomycin in treatment of human brucellosis due to Brucella melitensis. The GECMEI Group. Grupo de Estudio de Castilla-la Mancha de Enfermedades Infecciosas.

    Science.gov (United States)

    Solera, J; Rodríguez-Zapata, M; Geijo, P; Largo, J; Paulino, J; Sáez, L; Martínez-Alfaro, E; Sánchez, L; Sepulveda, M A; Ruiz-Ribó, M D

    1995-09-01

    Brucellosis is a common zoonosis in many parts of the world; the best regimen for the treatment of brucellosis has not been clearly determined. We have carried out a multicenter, open, controlled trial in five general hospitals in Spain to compare the efficacy and safety of doxycycline and rifampin (DR) versus doxycycline and streptomycin (DS) for the treatment of human brucellosis. The study included 194 ambulatory or hospitalized patients with acute brucellosis, without endocarditis or neurobrucellosis. The diagnostic criterion was isolation of Brucella species from blood or other tissues (n = 120) or a standard tube agglutination titer of 1/160 or more for anti-Brucella antibodies with compatible clinical findings (n = 74). Patients were randomly assigned to receive either 100 mg of doxycycline twice daily plus rifampin, 900 mg/day, in a single morning dose for 45 days (DR group) or the same dose of doxycycline for 45 days plus streptomycin, 1 g/day, intramuscularly for 14 days (DS group). A lack of therapeutic efficacy developed in 8 of the 100 patients in the DR group (8%) and in 2 of the 94 patients in the DS group (2%)(P = 0.10). Relapses occurred in 16 of the 100 patients in the DR group (16%) but in only 5 of the 94 patients in the DS group (5.3%) (P = 0.02). When relapse was considered in combination with initial lack of efficacy, 26 patients in the DR group (24%) and 7 patients in the DS group (7.45%) failed to respond to therapy (P = 0.0016). In general, therapy was well tolerated and only four patients (4%) in the DR group and two (2%) in the DS group had episodes of adverse effects necessitating discontinuation of treatment (P> 0.2). We conclude that a doxycycline-and-rifampin regimen is less effective than the doxycycline-and-streptomycin regimen in patients with acute brucellosis.

  14. Residual virulence and immunogenicity of CGV26 and CGV2631 B. melitensis Rev. 1 deletion mutant strains in sheep after subcutaneous or conjunctival vaccination.

    Science.gov (United States)

    Guilloteau, Laurence A; Laroucau, Karine; Olivier, Michel; Grillo, Maria Jesus; Marin, Clara M; Verger, Jean-Michel; Blasco, Jose-Maria

    2006-04-24

    The CGV26 and CGV2631 strains are novel engineered Brucella melitensis Rev.1 mutant strains deleted for the bp26 gene or for both bp26 and omp31 genes, respectively, coding for proteins of diagnostic significance. The residual virulence and immunogenicity of both mutants were compared to the parental Rev.1 strain in sheep after subcutaneous or conjunctival vaccination. The deletion of the bp26 gene or both bp26 and omp31 genes had no significant effect on the intracellular survival of the Rev.1 strain in ovine macrophage cultures. The kinetics of infection induced by both mutants in sheep was similar to the Rev.1 strain, and inoculation by the subcutaneous route produced wider and more generalized infections than the conjunctival route. All strains were cleared from lymph nodes and organs within 3 months after inoculation. The CGV26 and CGV2631 mutants induced both specific systemic antibody response and lymphoproliferation in sheep. The kinetics of the responses induced by the mutants was quite similar to that of the parental Rev.1 strain, except for the intensity of the lymphoproliferative response, which was attenuated for the CGV2631 mutant. In conclusion, the residual virulence of both CGV26 and CGV2631 mutants in sheep was similar to that of the parental Rev.1 vaccine strain. These mutants induced also significant specific antibody and cell-mediated immunity in sheep and are suitable to be evaluated as potential vaccine candidates against B. melitensis and B. ovis infections in sheep.

  15. Genome Sequences of 11 Brucella abortus Isolates from Persistently Infected Italian Regions

    OpenAIRE

    2015-01-01

    Bovine brucellosis, typically caused by Brucella abortus, has been eradicated from much of the developed world. However, the disease remains prevalent in southern Italy, persisting as a public and livestock health concern. We report here the whole-genome sequences of 11 isolates from cattle (Bos taurus) and water buffalo (Bubalus bubalis) that are representative of the current genetic diversity of B. abortus lineages circulating in Italy.

  16. Expression and immunogenicity of the recombinant protein omp25 of Brucella melitensis strain%羊种布鲁菌omp25的原核表达与免疫原性检测

    Institute of Scientific and Technical Information of China (English)

    吴杰; 吴树清; 李东兴; 刘艳琴; 张明月; 海岩

    2012-01-01

    To express omp25 gene of Brucella in E. Coli Rosella and detect the immunogenicity of the recombinant protein, the target fragment of omp25 gene was amplified by PCR from the B. Melilensis M306 strain and was cloned into expression vector PKT-32a. After transforming into competent E. Coli Rosella and inducing by IPTG, the recombinant protein was expressed. As detected by SDS-PAGK and western-blot, this recombinant protein has immunogenicity. It was confirmed DNA sequencing and restriction enzymes cleavage that the recombinant plasmid PET-32a/omp25 was constructed successfully. Furthermore, the result of SDS-PAGE and western-blot assay showed that PET-32a/omp25 was expressed successfully in E. Coli Rosella and the recombinant protein could react with B. Melilensis positive serum. This study provides a good foundation for the diagnosis of brucellosis and the preparation of new types of vaccines of Brucella.%目的 在大肠杆菌中表达羊种布鲁菌omp25基因并鉴定重组蛋白的免疫原性.方法 从羊种布鲁菌中用聚合酶链反应技术(PCR)扩增得到布鲁菌omp25基因片段,并将目的 基因插入原核表达载体PET-32a中,构建重组质粒PET-32a/omp25转入大肠杆菌E.coli Rosetta中并进行诱导表达,用SDS-PAGE和western-blot检测表达蛋白.结果 成功构建了重组质粒PET-32a/omp25,并在大肠杆菌Rosetta中获得了重组蛋白,重组蛋白与布鲁菌阳性血清发生特异性反应.结论 重组质粒PET-32a/omp25可以在大肠杆菌Rosetta中成功表达,并且重组蛋白可与布鲁菌阳性血清发生特异性反应,说明该重组蛋白有良好的免疫原性,该研究为疫苗的研制及布鲁菌病的检测打下良好的基础.

  17. Caspase-2-dependent dendritic cell death, maturation, and priming of T cells in response to Brucella abortus infection.

    Directory of Open Access Journals (Sweden)

    Xinna Li

    Full Text Available Smooth virulent Brucella abortus strain 2308 (S2308 causes zoonotic brucellosis in cattle and humans. Rough B. abortus strain RB51, derived from S2308, is a live attenuated cattle vaccine strain licensed in the USA and many other countries. Our previous report indicated that RB51, but not S2308, induces a caspase-2-dependent apoptotic and necrotic macrophage cell death. Dendritic cells (DCs are professional antigen presenting cells critical for bridging innate and adaptive immune responses. In contrast to Brucella-infected macrophages, here we report that S2308 induced higher levels of apoptotic and necrotic cell death in wild type bone marrow-derived DCs (WT BMDCs than RB51. The RB51 and S2308-induced BMDC cell death was regulated by caspase-2, indicated by the minimal cell death in RB51 and S2308-infected BMDCs isolated from caspase-2 knockout mice (Casp2KO BMDCs. More S2308 bacteria were taken up by Casp2KO BMDCs than wild type BMDCs. Higher levels of S2308 and RB51 cells were found in infected Casp2KO BMDCs compared to infected WT BMDCs at different time points. RB51-infected wild type BMDCs were mature and activated as shown by significantly up-regulated expression of CD40, CD80, CD86, MHC-I, and MHC-II. RB51 induced the production of cytokines TNF-α, IL-6, IFN-γ and IL12/IL23p40 in infected BMDCs. RB51-infected WT BMDCs also stimulated the proliferation of CD4(+ and CD8(+ T cells compared to uninfected WT BMDCs. However, the maturation, activation, and cytokine secretion are significantly impaired in Casp2KO BMDCs infected with RB51 or Salmonella (control. S2308-infected WT and Casp2KO BMDCs were not activated and could not induce cytokine production. These results demonstrated that virulent smooth strain S2308 induced more apoptotic and necrotic dendritic cell death than live attenuated rough vaccine strain RB51; however, RB51, but not its parent strain S2308, induced caspase-2-mediated DC maturation, cytokine production, antigen

  18. [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.

  19. Antigenic, immunologic and genetic characterization of rough strains B. abortus RB51, B. melitensis B115 and B. melitensis B18.

    Directory of Open Access Journals (Sweden)

    Rosanna Adone

    Full Text Available The lipopolysaccharide (LPS is considered the major virulent factor in Brucella spp. Several genes have been identified involved in the synthesis of the three LPS components: lipid A, core and O-PS. Usually, Brucella strains devoid of O-PS (rough mutants are less virulent than the wild type and do not induce undesirable interfering antibodies. Such of them proved to be protective against brucellosis in mice. Because of these favorable features, rough strains have been considered potential brucellosis vaccines. In this study, we evaluated the antigenic, immunologic and genetic characteristics of rough strains B. abortus RB51, B. melitensis B115 and B. melitensis B18. RB51 derived from B. abortus 2308 virulent strain and B115 is a natural rough strain in which the O-PS is present in the cytoplasm. B18 is a rough rifampin-resistan mutant isolated in our laboratory. The surface antigenicity of RB51, B115 and B18 was evaluated by testing their ability to bind antibodies induced by rough or smooth Brucella strains. The antibody response induced by each strain was evaluated in rabbits. Twenty-one genes, involved in the LPS-synthesis, were sequenced and compared with the B. melitensis 16M strain. The results indicated that RB51, B115 and B18 have differences in antigenicity, immunologic and genetic properties. Particularly, in B115 a nonsense mutation was detected in wzm gene, which could explain the intracellular localization of O-PS in this strain. Complementation studies to evaluate the precise role of each mutation in affecting Brucella morphology and its virulence, could provide useful information for the assessment of new, attenuated vaccines for brucellosis.

  20. Nucleotide-Binding Oligomerization Domain-1 and -2 Play No Role in Controlling Brucella abortus Infection in Mice

    Directory of Open Access Journals (Sweden)

    Fernanda S. Oliveira

    2012-01-01

    Full Text Available Nucleotide-binding oligomerization domain proteins (NODs are modular cytoplasmic proteins implicated in the recognition of peptidoglycan-derived molecules. Further, several in vivo studies have demonstrated a role for Nod1 and Nod2 in host defense against bacterial pathogens. Here, we demonstrated that macrophages from NOD1-, NOD2-, and Rip2-deficient mice produced lower levels of TNF-α following infection with live Brucella abortus compared to wild-type mice. Similar reduction on cytokine synthesis was not observed for IL-12 and IL-6. However, NOD1, NOD2, and Rip2 knockout mice were no more susceptible to infection with virulent B. abortus than wild-type mice. Additionally, spleen cells from NOD1-, NOD2-, and Rip2-deficient mice showed unaltered production of IFN-γ compared to C57BL/6 mice. Taken together, this study demonstrates that NOD1, NOD2 and Rip2 are dispensable for the control of B. abortus during in vivo infection.

  1. Association of Brucella Meningoencephalitis with Cerebrospinal Fluid Shunt in A Child: A Case Report

    Directory of Open Access Journals (Sweden)

    Babak ABDINIA

    2013-02-01

    melitensis in children. Clinical and epidemiological observations on 95 patients studied in Central Iran. Clin Pediatr 1978;17:904-7.5. Young EJ. Human brucellosis. Rev Infect Dis 1983; 5(5:821-42.6. Llorens-Terol J, Busquets RM. Brucellosis treated with rifampicin. Arch Dis Child 1980;55(6:486-8.7. FeizJ, Sabbaghian H, Miralai M. Brucellosis due to Brucella melitensis in children. Clinical and epidemiological observations on 95 patients studied in Central Iran. Clin Pediatr 1978;17:904-7.8. Wald SL, McLaurin RL. Cerebrospinal fluid antibiotic levels during treatment of shunt infections. J Neurosurg 1980;52(1:41-6. 

  2. Molecular Epidemiology of Brucella Genotypes in Patients at a Major Hospital in Central Peru

    NARCIS (Netherlands)

    K. Noeckler; R. Maves; D. Cepeda; A. Draeger; A. Mayer-Scholl; J. Chacaltana; M. Castaneda; B. Espinosa; R. Castillo; E. Hall; S. Al Dahouk; R.H. Gilman; F. Cabeza; H.L. Smits

    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 genotyp

  3. Toll-like receptors are critical for clearance of Brucella and play different roles in development of adaptive immunity following aerosol challenge in mice

    Directory of Open Access Journals (Sweden)

    Jianwu ePei

    2012-09-01

    Full Text Available Brucella spp. cause undulant fever in humans and brucellosis in variety of other animals. Both innate and adaptive immunity have been shown to be important in controlling Brucella infection. Toll-like receptors (TLRs represent a group of pattern recognition receptors that play critical roles in the host innate immune response, as well as development of adaptive immunity. In the current report, we investigated the role of TLR signaling in the clearance of Brucella and development of adaptive immunity in TLR2-/-, TLR4-/- or MyD88-/- mice following aerosol exposure to B. melitensis 16M. Consistent with previous reports, MyD88 is required for efficient clearance of Brucella from all three organs (lung, spleen and liver. The results reveal Th2-skewed immune responses in TLR2 / mice late in infection and support a TLR2 requirement for efficient clearance of Brucella from the lungs, but not from the spleen or liver. Similarly, TLR4 is required for efficient clearance of Brucella from the lung, but exhibits a minor contribution to clearance from the spleen and no demonstrable contribution to clearance from the liver. Lymphocyte proliferation assays suggest that the TLRs are not involved in the development of cell-mediated memory response to Brucella antigen. Antibody detection reveals that TLR2 and 4 are required to generate early antigen-specific IgG, but not during the late stages of infection. TLR2 and 4 are only transiently required for IgM production and not at all for IgA production. In contrast, MyD88 is essential for antigen specific IgG production late in infection, but is not required for IgM generation over the course of infection. Surprisingly, despite the prominent role for MyD88 in clearance from all tissues, MyD88-knockout mice express significantly higher levels of serum IgA. These results confirm the important role of MyD88 in controlling infection in the spleen while providing evidence of a prominent contribution to protection in other

  4. TLR7 and TLR3 Sense Brucella abortus RNA to Induce Proinflammatory Cytokine Production but They Are Dispensable for Host Control of Infection

    Science.gov (United States)

    Campos, Priscila C.; Gomes, Marco Túlio R.; Guimarães, Erika S.; Guimarães, Gabriela; Oliveira, Sergio C.

    2017-01-01

    Brucella abortus is a Gram-negative, facultative intracellular bacterium that causes brucellosis, a worldwide zoonotic disease leading to undulant fever in humans and abortion in cattle. The immune response against this bacterium relies on the recognition of microbial pathogen-associated molecular patterns, such as lipoproteins, lipopolysaccharides, and DNA; however, the immunostimulatory potential of B. abortus RNA remains to be elucidated. Here, we show that dendritic cells (DCs) produce significant amounts of IL-12, IL-6, and IP-10/CXCL10, when stimulated with purified B. abortus RNA. IL-12 secretion by DCs stimulated with RNA depends on TLR7 while IL-6 depends on TLR7 and partially on TLR3. Further, only TLR7 plays a role in IL-12 production induced by B. abortus infection. Moreover, cytokine production in DCs infected with B. abortus or stimulated with bacterial RNA was reduced upon pretreatment with MAPK/NF-κB inhibitors. By confocal microscopy, we demonstrated that TLR7 is colocalized with B. abortus in LAMP-1+ Brucella-containing vacuoles. Additionally, type I IFN expression and IP-10/CXCL10 secretion in DCs stimulated with bacterial RNA were dependent on TLR3 and TLR7. Our results suggest that TLR3 and TLR7 are not required to control Brucella infection in vivo, but they play an important role on sensing B. abortus RNA in vitro. PMID:28167945

  5. 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.

  6. High Incidence of Respiratory Involvement in a Cluster of Brucella suis-Infected Workers from a Pork Processing Plant in Argentina.

    Science.gov (United States)

    Wallach, J C; García, J L; Cardinali, P S; Seijo, A P; Benchetrit, A G; Echazarreta, S E; Garro, S L; Deodato, B; Baldi, P C

    2016-12-29

    Epidemiological and clinical aspects of Brucella suis infection in 17 workers from a pork processing plant in Argentina occurring between January 2014 and July 2015 are presented. All patients reported working 9 h daily without adequate personal protection garment. Blood cultures were positive for Brucella spp. in 14 of the 17 patients (82.3%). All isolates were identified as B. suis biovar 1. Although fever, sweats, asthenia, myalgia and hepatic involvement were the most frequent clinical manifestations, an unusually high incidence of respiratory involvement was found. From 13 patients in which chest radiography was performed, four (30%) had radiological abnormalities, including lobar pneumonia in two cases (one with pleural effusion) and interstitial involvement in other two. The high frequency of respiratory involvement in our series makes necessary to consider brucellosis in the differential diagnosis of respiratory diseases in pork processing plant employees.

  7. Adrenal steroids modulate the immune response during Brucella abortus infection by a mechanism that depends on the regulation of cytokine production.

    Science.gov (United States)

    Gentilini, María Virginia; Velásquez, Lis Noelia; Barrionuevo, Paula; Arriola Benitez, Paula Constanza; Giambartolomei, Guillermo Hernán; Delpino, María Victoria

    2015-05-01

    Human brucellosis is a protean disease with a diversity of clinical signs and symptoms resulting from infection with Brucella species. Recent reports suggest a cross-regulation between adrenal steroids (cortisol and dehydroepiandrosterone [DHEA]) and the immune system. Monocytes and macrophages are the main replication niche for Brucella. Therefore, we investigated the role of adrenal hormones on the modulation of the immune response mediated by macrophages in B. abortus infection. Cortisol treatment during B. abortus infection significantly inhibits cytokine, chemokine, and MMP-9 secretion. In contrast, DHEA treatment had no effect. However, DHEA treatment increases the expression of costimulatory molecules (CD40, CD86), the adhesion molecule CD54, and major histocompatibility complex class I (MHC-I) and MHC-II expression on the surface of B. abortus-infected monocytes. It is known that B. abortus infection inhibits MHC-I and MHC-II expression induced by gamma interferon (IFN-γ) treatment. DHEA reverses B. abortus downmodulation of the MHC-I and -II expression induced by IFN-γ. Taken together, our data indicate that DHEA immune intervention may positively affect monocyte activity during B. abortus infection.

  8. Variables Associated with Infections of Cattle by Brucella abortus., Leptospira spp. and Neospora spp. in Amazon Region in Brazil.

    Science.gov (United States)

    Chiebao, D P; Valadas, S Y O B; Minervino, A H H; Castro, V; Romaldini, A H C N; Calhau, A S; De Souza, R A B; Gennari, S M; Keid, L B; Soares, R M

    2015-10-01

    The frequency of Neospora spp., Leptospira spp. and Brucella abortus infections in adult cattle was determined in herds of the State of Pará, Brazil, which is an important region for cattle production located in the Amazon region. A total of 3466 adult female cattle from 176 herds were tested, leading to a frequency of seropositive animals of 14.7%, 3.7% and 65.5% and a herd positivity of 87.4%, 41.3% and 98.8% for infections caused by Neospora spp., B. abortus and Leptospira spp., respectively. The five most frequently diagnosed serologic responses to Leptospira spp. were those against serovars hardjo, wolfii, grippotyphosa, hebdomadis and shermani. The following associations were found: practice of artificial insemination, large farm size, large herd size, large number of dogs and high number of total abortions per year with the presence of antibodies against serovar hardjo; positive results to serovar grippotyphosa with the presence of dogs; inappropriate disposal of aborted foetuses with positivity to serovar hebdomadis. Serovar grippotyphosa was also associated with number of episodes of abortions. Neospora spp. positive herds were associated with episodes of abortion and B. abortus infection with the disposal of dead animals and aborted foetuses on pastures and with the use of artificial insemination. In conclusion, the high frequency of brucellosis, leptospirosis and neosporosis in the region may be a consequence of social, natural and raising conditions as: (i) climate conditions that favour the survival and spread of pathogens in the environment; (ii) farms located in regions bordering forest areas; (iii) farms in areas of difficult access to the veterinary service; (iv) extensive beef herds raised at pastures with different age and productive groups inter-mingled; and (v) minimal concerns regarding hygiene practices and disease prevention measures.

  9. 羊布鲁杆菌M5-90 BP26抗原单克隆抗体的制备及鉴定%Preparation and characterization of monoclonal antibodies against BP26 protein of Brucella melitensis M5-90

    Institute of Scientific and Technical Information of China (English)

    丘金浪; 吴静波; 黎诚耀; 王文敬

    2012-01-01

    Objective To prepare high specific monoclonal antibodies(mAbs) against BP26 of Brucella(B.)melitensis.Methods A recombinant plasmid pET-28a-BP26 was constructed and transformed into competent Escherichia coli BL21 (DE3),and then the bacteria were induced by 1 mmol/L isopropylthio-β-D-galactoside (IPTG).After induction,the recombinant BP26 protein (rBP26) was purified by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PGAE) and nickel ion affinity chromatography(Ni-NTA).Mice were inoculated with rBP26 antigens for three times at 2-week intervals.The first subcutaneous injection contained 100 μg rBP26 with 0.1 ml complete Freund adjuvant.The second subcutaneous injection was 50 μg rBP26 with 0.1 ml incomplete Freund adjuvant.The antibody titers to rBP26 were determined 2 weeks after each reimmunization.Three days before cell fusion,the mice with the highest titer were intraperitoneally injected with 50 μg rBP26 in 0.1 ml PBS.Pre- and post-immunization sera were collected and used as negative or positive controls for screening mAbs.Mice with the highest titer were sacrificed and spleen cells were isolated.The spleen cells of rBP26 immunized mice were fused with SP2/0 myeloma cells in a ratio of 5 ∶ 1 by polyethylene glycol(PEG) 1450.Antibody-producing hybridomas were primarily screened by an indirect enzyme-linked immunosorbnent assay(ELISA) with rBP26.Reactive hybridomas were subcloned for 3 times,then the strains of hybridoma cells secreting antibodies against BP26 were obtained.Supernatant of cloned hybridoma cultures was collected for mAb analyses.These mAbs were named by the hybridoma clone number and tested their reactivity to membrane proteins extracted(NMP) from B.melitensis vaccine strain(M5-90) by Western blotting and Dot-ELISA.mAbs isotyping and kappa(κ) or lambda(λ) light chain was identified by Mouse Monoclonal Antibody Isotyping Kit.Results A total of two mAbs reactive to rBP26 of B.melitensis were selected from antibody screening

  10. 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.

  11. Aspectos inmunológicos en el diagnóstico y control de la Epidemitis contagiosa del carnero por Brucella ovis Imunological aspects inthe diagnosis and control of contagious epidymitis of rams by Brucella ovis

    Directory of Open Access Journals (Sweden)

    S.M. ESTEIN

    1999-01-01

    de vacunas subcelulares que no presenten las desventajas de Rev. 1. Con ese objetivo se han desarrollado ensayos de inmunización pasiva con mezclas de anticuerpos monoclonales anti-proteínas de la membrana externa y anti-lipopolisacárido rugoso que han otorgado protección en el ratón contra B. ovisBrucella ovis is the aethiological agent of ram contagious epididymitis, an infectious disease causing reproductive failure in sheep. Control measures include elimination of rams found positive in serological tests and/or bacteriological culture of semen, and vaccination when the prevalence is high. The principal vaccine used against ovine brucellosis is B. melitensis Rev. 1, a live attenuated strain of B. melitensis. However Rev. 1 evokes antibodies interfering with the interpretation of serological tests used to diagnose infection by B. ovis and B. melitensis. B. ovis is a natural rough species so lacks O polisaccharide chain. The outer membrane proteins of B.ovis have been studied by several groups searching for antigens useful for diagnosis and protection. A hot saline extract fron B. ovis contains rough lipopolysaccharide and several proteins, including abundant group 3 outer membrane proteins The most efficient and widely used tests for serodiagnosis of B. ovis infection are double gel diffusion, complement fixation test and ELISA. Hot saline extract has provided best diagnostic results in all tests. However some cross-reactivity in these tests can be seen with sera of sheep naturally infected with B. melitensis or after B. melitensis Rev. 1 vaccination. Less information is available on internal B. ovis antigens. Recently, an indirect ELISA using BP26 (periplasmic protein has been developed for the differentiation of infected and vaccinated rams. The identification of B. ovis antigens able to elicit a protective immune response is of great interest for the development of subcellular vaccines avoiding the drawbacks of living attenuated vaccine. In the mouse model

  12. Herd-level risk factors for Campylobacter fetus infection, Brucella seropositivity and within-herd seroprevalence of brucellosis in cattle in northern Nigeria.

    Science.gov (United States)

    Mai, H M; Irons, P C; Kabir, J; Thompson, P N

    2013-09-01

    Brucellosis and campylobacteriosis are economically important diseases affecting bovine reproductive efficiency in Nigeria. A questionnaire-based survey was conducted in 271 cattle herds in Adamawa, Kaduna and Kano states of northern Nigeria using multistage cluster sampling. Serum from 4745 mature animals was tested for Brucella antibodies using the Rose-Bengal plate test and positives were confirmed in series-testing protocol using competitive enzyme-linked immunosorbent assay. Preputial scrapings from 602 bulls were tested using culture and identification for Campylobacter fetus. For each disease, a herd was classified as positive if one or more animals tested positive. For each herd, information on potential managemental and environmental risk factors was collected through a questionnaire administered during an interview with the manager, owner or herdsman. Multiple logistic regression models were used to model the odds of herd infection for each disease. A zero-inflated Poisson model was used to model the count of Brucella-positive animals within herds, with the number tested as an exposure variable. The presence of small ruminants (sheep and/or goats) on the same farm, and buying-in of >3 new animals in the previous year or failure to practice quarantine were associated with increased odds of herd-level campylobacteriosis and brucellosis, as well as increased within-herd counts of Brucella-positive animals. In addition, high rainfall, initial acquisition of animals from markets, practice of gynaecological examination and failure to practice herd prophylactic measures were positively associated with the odds of C. fetus infection in the herd. Herd size of >15, pastoral management system and presence of handling facility on the farm were associated with increased odds, and gynaecological examination with reduced odds of herd-level Brucella seropositivity. Furthermore, the zero-inflated Poisson model showed that borrowing or sharing of bulls was associated with

  13. Identification at biovar level of Brucella isolates causing abortion in small ruminants of iran.

    Science.gov (United States)

    Behroozikhah, Ali Mohammad; Bagheri Nejad, Ramin; Amiri, Karim; Bahonar, Ali Reza

    2012-01-01

    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.

  14. Identification at Biovar Level of Brucella Isolates Causing Abortion in Small Ruminants of Iran

    Directory of Open Access Journals (Sweden)

    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.

  15. 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.

  16. 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.

  17. Progress in the evolution and taxonomy of Brucella%布鲁菌进化和分类学研究进展

    Institute of Scientific and Technical Information of China (English)

    钟志军; 杜昕颖; 彭广能; 黄克和; 陈泽良; 于爽; 徐杰; 王玉飞; 白耀霞; 陈燕芬; 付思美; 王同坤; 汪舟佳

    2011-01-01

    Brucellae are Gram-negative,facultative intracellular bacteria that can infect many species of animals and man.Six species are currently recognized within the genus Brucella:B.melitensis,B.abortus,B.suis,B.neotomae,B.ovis,and B.canis.This classification is mainly based on differences in pathogenicity and in host preferences.Although the six species can be differentiated by conventional phenotypic tests,these species display a high degree of DNA homology in DNA-DNA hybridization assays(90% identity).Therefore it has been proposed that the Brucella genus should comprise only one species i.e.B.melitensis and that the other species should be considered as biovars.However,several molecular genotyping methods have shown of significant DNA polymorphism Brucella species allowing the species to be correctly differentiated.This is also true for the recent marine mammal Brucella isolates,which two new species names have been proposed,i.e.B.pinnipediae and B.cetaceae,according to the classical criteria of host preferentialism(pinnipeds and cetaceans respectively) and specific molecular markers.This article reviews the evolution and taxonomy of Brucella.%布鲁菌属革兰氏阴性兼性胞内寄生菌,能感染多种宿主动物和人。该属可分为6个典型种,包括羊种、牛种、猪种、沙林鼠种、绵羊附睾种以及犬种布鲁菌等。此分类是基于其致病性以及宿主偏好性的差异划分。尽管6个种通过传统表型试验能区分,但布鲁菌种内采用DNA-DNA杂交证明DNA同源性高度一致(相似性大于90%)。因此有人提议布鲁菌由单一种组成,即布鲁菌属中只有羊种布鲁菌,其他种都是羊种菌的生物亚型之一。然而基于其他分子技术的基因分型表明其DNA多态性表现明显,说明目前对这个种的分型还是比较准确。而最近分离的海洋种布鲁氏

  18. Understanding Virulence in the Brucellae and Francisellae: Towards Efficacious Treatments for Two Potential Biothreat Agents

    Energy Technology Data Exchange (ETDEWEB)

    Rasley, A; Parsons, D A; El-Etr, S; Roux, C; Tsolis, R

    2009-12-30

    Francisella tularensis, Yersinia pestis and Brucellae species are highly infectious pathogens classified as select agents by the Centers for Disease Control and Prevention (CDC) with the potential for use in bioterrorism attacks. These organisms are known to be facultative intracellular pathogens that preferentially infect human monocytes. As such, understanding how the host responds to infection with these organisms is paramount in detecting and combating human disease. We have compared the ability of fully virulent strains of each pathogen and their non-pathogenic near neighbors to enter and survive inside the human monocytic cell line THP-1 and have quantified the cellular response to infection with the goal of identifying both unique and common host response patterns. We expanded the scope of these studies to include experiments with pathogenic and non-pathogenic strains of Y. pestis, the causative agent of plague. Nonpathogenic strains of each organism were impaired in their ability to survive intracellularly compared with their pathogenic counterparts. Furthermore, infection of THP-1 cells with pathogenic strains of Y. pestis and F. tularensis resulted in marked increases in the secretion of the inflammatory chemokines IL-8, RANTES, and MIP-1{beta}. In contrast, B. melitensis infection failed to elicit any significant increases in a panel of cytokines tested. These differences may underscore distinct strategies in pathogenic mechanisms employed by these pathogens.

  19. 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...

  20. Antimicrobial Susceptibility of Brucella melitensis Isolates in Peru

    Science.gov (United States)

    2011-03-01

    and Prevmtive Medicine Unit Two. Norfolk, Virginia’ Hospital Naciorwl Daniel Alcides Carrion , Callao, Pent~; Hospital Arzobi5po Loayza, Lima, Peru5...Alcides Carrion (Callao, Peru). All cultures, species identification, and antimicrobial suscep- tibility tests were performed in the Bacteriology

  1. In silico design, cloning and high level expression of L7/L12-TOmp31 fusion protein of Brucella antigens

    OpenAIRE

    Golshani, Maryam; Rafati, Sima; Jahanian-Najafabadi, Ali; Nejati-Moheimani, Mehdi; Siadat, Seyed Davar; Shahcheraghi, Fereshteh; Bouzari, Saeid

    2015-01-01

    Globally, Brucella melitensis and B. abortus are the most common cause of human brucellosis. The outer membrane protein 31 (Omp31) and L7/L12 are immunodominant and protective antigens conserved in human Brucella pathogens which are considered as potential vaccine candidates. We aimed to design the fusion protein from Brucella L7/L12 and truncated Omp31proteins, in silico, clone the fusion in pET28a vector, and express it in Escherichia coli host. Two possible fusion forms, L7/L12-TOmp31 and ...

  2. Epidemiological aspects of an infection by Brucella abortus in risk occupational groups in the microregion of Araguaína, Tocantins

    Directory of Open Access Journals (Sweden)

    Taciana Rabelo Ramalho Ramos

    2008-04-01

    Full Text Available The aim of this paper was to study some epidemiological aspects of the infection by Brucella abortus in risk occupational groups in the microregion of Araguaína, Tocantins. For antibody research, 645 serum samples were analyzed by the complement fixation test (CF. A 4.0% frequency was found (26/645 in patients' serum and among those 4.1% (23/551 were slaughterhouses employees and 8.1% (3/37 rural workers. Of the total positive samples, three (2.0% were women and 23 (4.7% men; ten (2.9% were between the ages of 18 and 30, six (3.4% between 31 and 40, and nine (8.0% were above 41 years of age. Risk factors for brucellosis in the study groups were age, background (OR = 2.45; CI 95% = 0.98 to 6.10 and previous work conducted with production animals (OR 2.36; CI 95% = 0.95 to 6.02. It was concluded that the infection by Brucella abortus is found in some risk occupational groups in the microregion of Araguaína, Tocantins, and control and prophylactic measures must be implemented emphasizing risk factors identified in the study.

  3. Immunization of BALB/c mice with Brucella abortus 2308ΔwbkA confers protection against wild-type infection.

    Science.gov (United States)

    Li, Zhi-qiang; Gui, Dan; Sun, Zhi-hua; Zhang, Jun-bo; Zhang, Wen-zhi; Zhang, Hui; Guo, Fei; Chen, Chuang-fu

    2015-01-01

    Brucellosis is a zoonotic disease that causes animal and human diseases. Vaccination is a major measure for prevention of brucellosis, but it is currently not possible to distinguish vaccinated animals from those that have been naturally infected. Therefore, in this study, we constructed the Brucella (B.) abortus 2380 wbkA mutant (2308ΔwbkA) and evaluated its virulence. The survival of 2308ΔwbkA was attenuated in murine macrophage (RAW 264.7) and BALB/c mice, and it induced high protective immunity in mice. The wbkA mutant elicited an anti-Brucella-specific immunoglobulin G response and induced the secretion of gamma interferon. Antibodies to 2308ΔwbkA could be detected in sera from mice, implying the potential for use of this protein as a diagnostic antigen. The WbkA antigen would allow serological differentiation between infected and vaccinated animals. These results suggest that 2308ΔwbkA is a potential attenuated vaccine against 16M. This vaccine will be further evaluated in sheep.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  5. 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.

  6. 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.

  7. Characterization of some Brucella species from Zimbabwe by biochemical profiling and AMOS-PCR

    Directory of Open Access Journals (Sweden)

    Skjerve Eystein

    2009-12-01

    Full Text Available Abstract Background Bovine brucellosis caused by Brucella abortus is endemic in most large commercial and smallholder cattle farms of Zimbabwe, while brucellosis in other domestic animals is rare. The diagnosis of brucellosis is mainly accomplished using serological tests. However, some Brucella spp. have been isolated from clinical cases in the field and kept in culture collection but their biochemical profiles were not documented. We report biochemical profiling and AMOS-PCR characterization of some of these field isolates of Brucella originating from both commercial and smallholder cattle farming sectors of Zimbabwe. Findings Fourteen isolates of Brucella from culture collection were typed using biochemical profiles, agglutination by monospecific antisera, susceptibility to Brucella-specific bacteriophages and by AMOS-PCR that amplifies species- specific IS711. The results of the biochemical profiles for B. abortus biovar 1 (11 isolates and biovar 2 (2 isolates were consistent with those of reference strains. A single isolate from a goat originating from a smallholder mixed animal farm was identified as B. melitensis biovar 1. The AMOS-PCR produced DNA products of sizes 498 bp and 731 bp for B. abortus (biovar 1 and 2 and B. melitensis biovar 1, respectively. Conclusion We concluded that the biochemical profiles and AMOS-PCR characterization were consistent with their respective species and biovars. B. abortus biovar 1 is likely to be the predominant cause of brucellosis in both commercial and smallholder cattle farms in Zimbabwe.

  8. 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.

  9. The Assessment of Cytokine and Antibody Responses to Recombinant 31kDa Brucella Cell-Surface Protein in Brucella Abortus Infected Mouse Model

    Directory of Open Access Journals (Sweden)

    Nima Khoramabadi

    2014-06-01

    Conclusion: These findings suggest that specific humoral and cell-mediated responses to BCSP31 is formed during murine host infection with B. abortus. Based on these findings, rBCSP31 can be used in further design of immunogenic strategies for vaccination against brucellosis.

  10. Internal affairs: investigating the Brucella intracellular lifestyle.

    Science.gov (United States)

    von Bargen, Kristine; Gorvel, Jean-Pierre; Salcedo, Suzana P

    2012-05-01

    Bacteria of the genus Brucella are Gram-negative pathogens of several animal species that cause a zoonotic disease in humans known as brucellosis or Malta fever. Within their hosts, brucellae reside within different cell types where they establish a replicative niche and remain protected from the immune response. The aim of this article is to discuss recent advances in the field in the specific context of the Brucella intracellular 'lifestyle'. We initially discuss the different host cell targets and their relevance during infection. As it represents the key to intracellular replication, the focus is then set on the maturation of the Brucella phagosome, with particular emphasis on the Brucella factors that are directly implicated in intracellular trafficking and modulation of host cell signalling pathways. Recent data on the role of the type IV secretion system are discussed, novel effector molecules identified and how some of them impact on trafficking events. Current knowledge on Brucella gene regulation and control of host cell death are summarized, as they directly affect intracellular persistence. Understanding how Brucella molecules interplay with their host cell targets to modulate cellular functions and establish the intracellular niche will help unravel how this pathogen causes disease.

  11. Epidemiology and genetic characterization of BVDV, BHV-1, BHV-4, BHV-5 and Brucella spp. infections in cattle in Turkey.

    Science.gov (United States)

    Aslan, Muhammet Eren; Azkur, Ahmet Kursat; Gazyagci, Serkal

    2015-11-01

    The aim of the study was to determine the epidemiological data of bovine viral diarrhea virus (BVDV), bovine herpesvirus-1 (BHV-1), bovine herpesvirus-4 (BHV-4), bovine herpesvirus-5 (BHV-5) and Brucella-associated cattle that were previously reported to have abortion and infertility problems in Ankara, Corum, Kirikkale and Yozgat provinces, Turkey. Whole blood and sera samples were obtained from 656 cattle, and antibodies against Brucella spp. were detected in 45 (6.86%) and 41 (6.25%) animals by Rose Bengal plate and serum tube agglutination tests, respectively. The seropositivity rates against BVDV, BHV-1 and BHV-4 were 70.89%, 41.3% and 28.78%, respectively. RT-PCR and PCR were performed to detect RNA and DNA viruses in blood samples, respectively. The BVDV 5'-untranslated region and BHV-1 gB gene detected in this study were phylogenetically analyzed. The BVDV strains analyzed in this study were closely related to those previously reported from Turkey. The nucleotide sequence from the BHV-1 strain detected in this study is the first nucleotide sequence of BHV-1 circulating in this area of Turkey deposited in the GenBank. The presence of Brucella spp. and prevalence of BHV-1, BHV-4 and BVDV in cattle should be further investigated throughout these regions.

  12. Rapid serum agglutination and agar gel immunodiffusion tests associated to clinical signs in rams experimentally infected with Brucella ovis Teste de soro aglutinação rápida e do teste de imunodifusão em gel de ágar associados aos sinais clínicos em carneiros infectados experimentalmente com Brucella ovis

    Directory of Open Access Journals (Sweden)

    Cristiane Nakada Nozaki

    2011-08-01

    Full Text Available The purpose of this study was to evaluate the agar gel immunodiffusion and the rapid serum agglutination tests associated to clinical signs in rams experimentally infected with Brucella ovis. The serological profile during the 12 months of infection showed a large fluctuation of antibodies that favors the failure in the diagnostic. The evaluation of tests after the experimental infection allowed to suggest that none of the tests were able to detect the infection throughout the period of study. The study reinforces the importance of considering the clinical signs to support the diagnosis of Brucella ovis infection in rams.O objetivo deste estudo foi avaliar o uso do teste de imunodifusão em gel de ágar e o teste sorológico de aglutinação rápida comparados aos sinais clínicos em carneiros infectados experimentalmente com Brucella ovis para o diagnóstico confirmatório da brucelose ovina. O perfil sorológico durante os 12 meses pós-infecção mostrou flutuação da resposta por anticorpos, que favorece a falha no diagnóstico. A avaliação dos testes indicou que nenhum dos testes foi capaz de detectar a infecção durante todo o período de estudo. O estudo ressalta a importância de considerar os sinais clínicos para apoiar o diagnóstico confirmatório da infecção por Brucella ovis em carneiros.

  13. 布鲁氏菌M5-90△WboA基因缺失株的构建及免疫效果的初步评价%Construction and preliminary evaluation of immune effects of deletion mutant of WboA gene of Brucella melitensis vaccine M5-90

    Institute of Scientific and Technical Information of China (English)

    张艳; 陈创夫; 张辉; 张沾; 李志强; 张俊波; 孟仁; 王震

    2011-01-01

    Vaccination is a major measure for prevention of brucellosis, but it is unable to be distinguished from the vaccinated to natural infected animals. In this study, the WboA gene was knocked out of the genomic DNA of Brucelh melitensis vaccine M5-90 strain to construct the recombinant M5-90 A WboA by homologous recombination. The test results showed that M5-90 A WboA was less virulent than that of the parent M5-90 strain (p<0.01). Humoral immunity and cellular immunity tests showed that there was no significant difference between M5-90A WboA and M5-90 parent strain (p<0.05). BALB/c mice were immunized with M5-90 A WboA or M5-90 and challenged with virulent strain 16M and the survival rate were 10% and 20%, respectively, indicating that the M5-90 A WboA provided the similar protection of M5-90 strain. Agglutination test and western blot showed that the serum response of M5-90 △ WboA in vaccinated mice were negative. These results indicated that M5-90 A WboA strain might be a promising vaccine against brucellosis, and could be distinguished from the vaccine immunization to natural infection in animals by serum test.%为获得毒力较弱并能区分自然感染和疫苗免疫的布鲁氏菌候选疫苗株,本研究用PCR方法扩增WboA基因的上下游同源臂序列,构建重组质粒pGEM-7zf-△WboA-Sac,电转化布鲁氏菌M5-90感受态细胞,筛选布鲁氏菌疫苗株M5-90的WboA基因缺失株,并对获得的M5-90△WboA遗传稳定性、毒力、免疫保护性、抗体水平等指标进行检测.实验结果表明M5-90△WboA株的毒力比M5-90株明显减弱,差异极显著(p<0.01),体液免疫和细胞免疫结果表明M5-90△WboA株与亲本M5-90株相比差异不显著(p<0.05),M5-90△WboA株和亲本株的保护率分别为10%和20%,表明M5-90△WboA株与M5-90株具有相似的保护性.凝集试验和western blot试验显示M5-90△WboA株免疫小鼠的血清反应结果为阴性.本研究构建的布鲁氏菌基因缺失株M5-90△WboA

  14. 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.

  15. 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.

  16. 16S rRNA and omp31 gene based molecular characterization of field strains of B. melitensis from aborted foetus of goats in India.

    Science.gov (United States)

    Singh, Ajay; Gupta, Vivek Kumar; Kumar, Amit; Singh, Vikas Kumar; Nayakwadi, Shivasharanappa

    2013-01-01

    Brucellosis is a reemerging infectious zoonotic disease of worldwide importance. In human, it is mainly caused by Brucella melitensis, a natural pathogen for goats. In India, a large number of goats are reared in semi-intensive to intensive system within the close vicinity of human being. At present, there is no vaccination and control strategy for caprine brucellosis in the country. Thus, to formulate an effective control strategy, the status of etiological agent is essential. To cope up with these, the present study was conducted to isolate and identify the prevalent Brucella species in caprine brucellosis in India. The 30 samples (fetal membrane, fetal stomach content and vaginal swabs) collected throughout India from the aborted fetus of goats revealed the isolation of 05 isolates all belonging to Brucella melitensis biovars 3. All the isolates produced amplification products of 1412 and 720 bp in polymerase chain reaction with genus and species specific 16S rRNA and omp31 gene based primers, respectively. Moreover, the amplification of omp31 gene in all the isolates confirmed the presence of immuno dominant outer membrane protein (31 kDa omp) in all the field isolates of B. melitensis in aborted foetus of goats in India. These findings can support the development of omp31 based specific serodiagnostic test as well as vaccine for the control of caprine brucellosis in India.

  17. 16S rRNA and Omp31 Gene Based Molecular Characterization of Field Strains of B. melitensis from Aborted Foetus of Goats in India

    Directory of Open Access Journals (Sweden)

    Ajay Singh

    2013-01-01

    Full Text Available Brucellosis is a reemerging infectious zoonotic disease of worldwide importance. In human, it is mainly caused by Brucella melitensis, a natural pathogen for goats. In India, a large number of goats are reared in semi-intensive to intensive system within the close vicinity of human being. At present, there is no vaccination and control strategy for caprine brucellosis in the country. Thus, to formulate an effective control strategy, the status of etiological agent is essential. To cope up with these, the present study was conducted to isolate and identify the prevalent Brucella species in caprine brucellosis in India. The 30 samples (fetal membrane, fetal stomach content and vaginal swabs collected throughout India from the aborted fetus of goats revealed the isolation of 05 isolates all belonging to Brucella melitensis biovars 3. All the isolates produced amplification products of 1412 and 720 bp in polymerase chain reaction with genus and species specific 16S rRNA and omp31 gene based primers, respectively. Moreover, the amplification of omp31 gene in all the isolates confirmed the presence of immuno dominant outer membrane protein (31 kDa omp in all the field isolates of B. melitensis in aborted foetus of goats in India. These findings can support the development of omp31 based specific serodiagnostic test as well as vaccine for the control of caprine brucellosis in India.

  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. Investigation and Analysis of Brucella Infections in Livestock in Guiyang%贵阳市家畜布鲁菌感染情况调查与分析

    Institute of Scientific and Technical Information of China (English)

    徐春志; 蔺俐仲; 方英; 白璧; 袁翠霞; 景书灏; 范伟兴

    2016-01-01

    为了解贵阳市2015年家畜布鲁菌的感染情况,以指导全市家畜布鲁菌病的防控工作,采用虎红平板凝集试验和试管凝集试验对全市8381份牛、羊、猪血清开展布鲁菌感染情况调查。结果表明,牛、羊和猪布鲁菌病阳性率依次为0.29%(13/4530)、0.39%(11/2848)和0%(0/1003)。说明贵阳市家畜布鲁菌病的流行率在国家控制标准以内,今后宜实行检疫和扑杀相结合的防控策略。%In order to find out Brucella infection of livestock in 2015,and guide the prevention and control of brucellosis in Guiyang,8 381 serum samples were detected by RBT and SAT.The results showed that the positive rates of cattle,goats and swine were 0.29%,0.39% and 0%,respectively.The infection rates of Brucella were within national control standards,and should carry out the prevention and control strategy of quarantine and slaughter in the future.

  20. Osteoarticular tissue infection and development of skeletal pathology in murine brucellosis

    Directory of Open Access Journals (Sweden)

    Diogo M. Magnani

    2013-05-01

    Brucellosis, a frequent bacterial zoonosis, can produce debilitating chronic disease with involvement of multiple organs in human patients. Whereas acute brucellosis is well studied using the murine animal model, long-term complications of host-pathogen interaction remain largely elusive. Human brucellosis frequently results in persistent, chronic osteoarticular system involvement, with complications such as arthritis, spondylitis and sacroiliitis. Here, we focused on identifying infectious sites in the mouse that parallel Brucella melitensis foci observed in patients. In vivo imaging showed rapid bacterial dispersal to multiple sites of the murine axial skeleton. In agreement with these findings, immunohistochemistry revealed the presence of bacteria in bones and limbs, and in the lower spine vertebrae of the axial skeleton where they were preferentially located in the bone marrow. Surprisingly, some animals developed arthritis in paws and spine after infection, but without obvious bacteria in these sites. The identification of Brucella in the bones of mice corroborates the findings in humans that these osteoarticular sites are important niches for the persistence of Brucella in the host, but the mechanisms that mediate pathological manifestations in these sites remain unclear. Future studies addressing the immune responses within osteoarticular tissue foci could elucidate important tissue injury mediators and Brucella survival strategies.

  1. CCL20 and Beta-Defensin 2 Production by Human Lung Epithelial Cells and Macrophages in Response to Brucella abortus Infection

    Science.gov (United States)

    Fernández, Andrea G.; Bonetto, Josefina; Giambartolomei, Guillermo H.; Fossati, Carlos A.; Baldi, Pablo C.

    2015-01-01

    Both CCL20 and human β-defensin 2 (hBD2) interact with the same membrane receptor and display chemotactic and antimicrobial activities. They are produced by airway epithelia in response to infectious agents and proinflammatory cytokines. Whereas Brucella spp. can infect humans through inhalation, their ability to induce CCL20 and hBD2 in lung cells is unknown. Here we show that B. abortus induces CCL20 expression in human alveolar (A549) or bronchial (Calu-6) epithelial cell lines, primary alveolar epithelial cells, primary human monocytes, monocyte-derived macrophages and the monocytic cell line THP-1. CCL20 expression was mainly mediated by JNK1/2 and NF-kB in both Calu-6 and THP-1 cells. CCL20 secretion was markedly induced in A549, Calu-6 and THP-1 cells by heat-killed B. abortus or a model Brucella lipoprotein (L-Omp19) but not by the B. abortus lipopolysaccharide (LPS). Accordingly, CCL20 production by B. abortus-infected cells was strongly TLR2-dependent. Whereas hBD2 expression was not induced by B. abortus infection, it was significantly induced in A549 cells by conditioned media from B. abortus-infected THP-1 monocytes (CMB). A similar inducing effect was observed on CCL20 secretion. Experiments using blocking agents revealed that IL-1β, but not TNF-α, was involved in the induction of hBD2 and CCL20 secretion by CMB. In the in vitro antimicrobial assay, the lethal dose (LD) 50 of CCL20 for B. abortus (>50 μg/ml) was markedly higher than that against E. coli (1.5 μg/ml) or a B. abortus mutant lacking the O polysaccharide in its LPS (8.7 ug/ml). hBD2 did not kill any of the B. abortus strains at the tested concentrations. These results show that human lung epithelial cells secrete CCL20 and hBD2 in response to B. abortus and/or to cytokines produced by infected monocytes. Whereas these molecules do not seem to exert antimicrobial activity against this pathogen, they could recruit immune cells to the infection site. PMID:26448160

  2. CCL20 and Beta-Defensin 2 Production by Human Lung Epithelial Cells and Macrophages in Response to Brucella abortus Infection.

    Science.gov (United States)

    Hielpos, M Soledad; Ferrero, Mariana C; Fernández, Andrea G; Bonetto, Josefina; Giambartolomei, Guillermo H; Fossati, Carlos A; Baldi, Pablo C

    2015-01-01

    Both CCL20 and human β-defensin 2 (hBD2) interact with the same membrane receptor and display chemotactic and antimicrobial activities. They are produced by airway epithelia in response to infectious agents and proinflammatory cytokines. Whereas Brucella spp. can infect humans through inhalation, their ability to induce CCL20 and hBD2 in lung cells is unknown. Here we show that B. abortus induces CCL20 expression in human alveolar (A549) or bronchial (Calu-6) epithelial cell lines, primary alveolar epithelial cells, primary human monocytes, monocyte-derived macrophages and the monocytic cell line THP-1. CCL20 expression was mainly mediated by JNK1/2 and NF-kB in both Calu-6 and THP-1 cells. CCL20 secretion was markedly induced in A549, Calu-6 and THP-1 cells by heat-killed B. abortus or a model Brucella lipoprotein (L-Omp19) but not by the B. abortus lipopolysaccharide (LPS). Accordingly, CCL20 production by B. abortus-infected cells was strongly TLR2-dependent. Whereas hBD2 expression was not induced by B. abortus infection, it was significantly induced in A549 cells by conditioned media from B. abortus-infected THP-1 monocytes (CMB). A similar inducing effect was observed on CCL20 secretion. Experiments using blocking agents revealed that IL-1β, but not TNF-α, was involved in the induction of hBD2 and CCL20 secretion by CMB. In the in vitro antimicrobial assay, the lethal dose (LD) 50 of CCL20 for B. abortus (>50 μg/ml) was markedly higher than that against E. coli (1.5 μg/ml) or a B. abortus mutant lacking the O polysaccharide in its LPS (8.7 ug/ml). hBD2 did not kill any of the B. abortus strains at the tested concentrations. These results show that human lung epithelial cells secrete CCL20 and hBD2 in response to B. abortus and/or to cytokines produced by infected monocytes. Whereas these molecules do not seem to exert antimicrobial activity against this pathogen, they could recruit immune cells to the infection site.

  3. Glial Cell-Elicited Activation of Brain Microvasculature in Response to Brucella abortus Infection Requires ASC Inflammasome-Dependent IL-1β Production.

    Science.gov (United States)

    Miraglia, M Cruz; Costa Franco, Miriam M; Rodriguez, Ana M; Bellozi, Paula M Q; Ferrari, Carina C; Farias, Maria I; Dennis, Vida A; Barrionuevo, Paula; de Oliveira, Antonio C P; Pitossi, Fernando; Kim, Kwang Sik; Delpino, M Victoria; Oliveira, Sergio Costa; Giambartolomei, Guillermo H

    2016-05-01

    Blood-brain barrier activation and/or dysfunction are a common feature of human neurobrucellosis, but the underlying pathogenic mechanisms are largely unknown. In this article, we describe an immune mechanism for inflammatory activation of human brain microvascular endothelial cells (HBMEC) in response to infection with Brucella abortus Infection of HBMEC with B. abortus induced the secretion of IL-6, IL-8, and MCP-1, and the upregulation of CD54 (ICAM-1), consistent with a state of activation. Culture supernatants (CS) from glial cells (astrocytes and microglia) infected with B. abortus also induced activation of HBMEC, but to a greater extent. Although B. abortus-infected glial cells secreted IL-1β and TNF-α, activation of HBMEC was dependent on IL-1β because CS from B. abortus-infected astrocytes and microglia deficient in caspase-1 and apoptosis-associated speck-like protein containing a CARD failed to induce HBMEC activation. Consistently, treatment of CS with neutralizing anti-IL-1β inhibited HBMEC activation. Both absent in melanoma 2 and Nod-like receptor containing a pyrin domain 3 are partially required for caspase-1 activation and IL-1β secretion, suggesting that multiple apoptosis-associated speck-like protein containing CARD-dependent inflammasomes contribute to IL-1β-induced activation of the brain microvasculature. Inflammasome-mediated IL-1β secretion in glial cells depends on TLR2 and MyD88 adapter-like/TIRAP. Finally, neutrophil and monocyte migration across HBMEC monolayers was increased by CS from Brucella-infected glial cells in an IL-1β-dependent fashion, and the infiltration of neutrophils into the brain parenchyma upon intracranial injection of B. abortus was diminished in the absence of Nod-like receptor containing a pyrin domain 3 and absent in melanoma 2. Our results indicate that innate immunity of the CNS set in motion by B. abortus contributes to the activation of the blood-brain barrier in neurobrucellosis and IL-1β mediates

  4. Six cases of Brucella infection in children and review of literatures%儿童布鲁杆菌病六例报告并文献复习

    Institute of Scientific and Technical Information of China (English)

    朱丹; 张艳玲; 钟雪梅; 马昕; 宁慧娟; 杨洋

    2015-01-01

    Objective To present six cases of Brucella infection in children,analyze the characteristics of the disease,diagnostic and therapeutic process.Method The clinical manifestations,laboratory test results and diagnostic process of 6 confirmed cases of brucellosis seen between 2011-2012 were retrospectively analyzed and domestic and foreign literature was reviewed.Result All the 6 children had a history of either exposure to,travelling to endemic area,or consuming infected lamb/beef.After the relevant examinations for these children,either positive etiologic or serologic evidence of brucellosis infection was obtained.The main clinical manifestation was fever in all cases,the peak body temperature was 37.5-38.0 ℃ in 3 cases,38.1-39.0 ℃ in 2 cases,39.1-41 ℃ in 1 case.Except for 1 case whose fever type was undulant fever,all the rest had irregular fever.Joint pain existed in 3 cases,orchitis in 1 case,cervical lymphadenopathy in 3 cases,hepatosplenomegaly in 2 cases,and impaired liver function in 4 cases.The Brucella agglutination test was positive in 5 cases.The blood culture was positive for all cases.In 4 cases the sulfamethoxazle and rifampicin were used for treatment,1 case was treated with rifampicin and erythromycin,parents of 1 case refused to use the drug.The "brucellosis in children" was used to search literature at Wanfang database,Pubmed database for literature of recent 10 years,and a total of 13 articles including 15 cases were retrieved.All the patients had fever,6 cases had joint swelling and pain,10 cases had hepatosplenomegaly,6 cases had cervical lymphadenopathy,4 cases were complicated with central nervous system infection.Brucella agglutination test was positive in 9 cases and blood culture was positive for Brucella infection in all cases.Conclusion Childhood Brucella infections are usually presented with various clinical manifestations,and are often accompanied by symptoms of systemic infection.For fever of unknown origin,one should include

  5. The effects of red ginseng saponin fraction-A (RGSF-A) on phagocytosis and intracellular signaling in Brucella abortus infected RAW 264.7 cells.

    Science.gov (United States)

    Arayan, Lauren Togonon; Simborio, Hannah Leah; Reyes, Alisha Wehdnesday Bernardo; Hop, Huynh Tan; Min, WonGi; Lee, Hu Jang; Rhee, Man Hee; Chang, Hong Hee; Kim, Suk

    2015-06-01

    This study indicated that RGSF-A caused a marked reduction in the adherence, internalization and intracellular growth of Brucella abortus in RGSF-A-treated cells. Furthermore, a decline in the intensity of F-actin fluorescence was observed in RGSF-A-treated cells compared with untreated B. abortus-infected cells. In addition, an evaluation of phagocytic signaling proteins by Western blot analysis revealed an apparent reduction of ERK and p38α phosphorylation levels in B. abortus-infected RGSF-A-treated cells compared with the control. Upon intracellular trafficking of the pathogen, a higher number of B. abortus-containing phagosomes colocalized with LAMP-1 in RGSF-A-treated cells compared with control cells. These results strongly suggest that inhibition of B. abortus uptake could be mediated by suppression in the activation of MAPKs signaling proteins phospho-ERK 1/2, and p38 levels. On the other hand, inhibition of intracellular replication results from the enhancement of phagolysosome fusion in host macrophages. This study highlights the phagocytic and intracellular modulating effect of RGSF-A and its potential as an alternative remedy to control B. abortus infection.

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

    Science.gov (United States)

    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

    2009-10-01

    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 genotypes were identified, separated by the number of Bruce42 repeats into two groups that may have distinct phenotypic characteristics. Whereas genotypes with five or six Bruce42 repeats were cultured mainly from adult patients, genotypes with three Bruce42 repeats were isolated from children and young adolescents as well as from adults. In addition, the isolates with three Bruce42 repeats were obtained more often from patients with splenomegaly (P = 0.02) or hepatomegaly (P = 0.006). An annual variation in the diversity of genotypes was observed, possibly reflecting changes in sources of fresh dairy products, supply routes to city shops and markets, and the movement of infected dairy goat herds.

  7. PLGA (85:15) nanoparticle based delivery of rL7/L12 ribosomal protein in mice protects against Brucella abortus 544 infection: A promising alternate to traditional adjuvants.

    Science.gov (United States)

    Singh, Damini; Somani, Vikas Kumar; Aggarwal, Somya; Bhatnagar, Rakesh

    2015-12-01

    There is a compelling need for the development of suitable adjuvants for human use to enhance the efficacy of the upcoming vaccines for the prevention of life threatening infections. In the current study, we have tried to explore the immunogenic potential of nanoparticles (NPs) made of PLGA (poly lactic-co-glycolic acid), a biodegradable and biocompatible polymer approved by FDA for human use after entrapping rL7/L12 protein, an immunodominant antigen of Brucella. Adjuvant properties were exhibited by the formulation as it elicited high IgG antibody titers just after first immunization which increased significantly after the booster administration. A good elicitation of the Th1 cytokines especially IFN-γ was recorded. Amongst the IgG antibody subclasses, IgG1 remained the predominant subclass to be elicited in mice serum after immunization; however IgG1/2a ratio showed a mixed profile of Th1/Th2 response. Lymphocyte proliferation assay as a marker of amplification in cellular immunity demonstrated that the splenocytes of the immunized mice had a high proliferation index with reference to the control, revealing that L7/L12 entrapping PLGA nanoparticles are potent inducer of inflammatory cell response indispensable to combat Brucella infection. Enumeration of splenic CFU after 14 days of infection with Brucella abortus 544 showed a significant reduction in log CFU of splenic bacteria in the vaccinated mice as compared to the control group. Therefore it is evident that PLGA nano formulations delivering the entrapped vaccine candidate in mice elicit specific humoral as well as cellular responses specific to the entrapped Brucella antigen. So there is much promise in this approach and this work by highlighting the adjuvant properties of the PLGA nanospheres will accelerate the development of improved vaccines safe for human as well as veterinary use.

  8. Brucella spp noncanonical LPS: structure, biosynthesis, and interaction with host immune system

    Directory of Open Access Journals (Sweden)

    Oliveira Sergio

    2006-03-01

    Full Text Available Abstract Brucella spp. are facultative intracellular pathogens that have the ability to survive and multiply in professional and non-professional phagocytes, and cause abortion in domestic animals and undulant fever in humans. Several species are recognized within the genus Brucella and this classification is mainly based on the difference in pathogenicity and in host preference. Brucella strains may occur as either smooth or rough, expressing smooth LPS (S-LPS or rough LPS (R-LPS as major surface antigen. This bacterium possesses an unconventional non-endotoxic lipopolysaccharide that confers resistance to anti-microbial attacks and modulates the host immune response. The strains that are pathogenic for humans (B. abortus, B. suis, B. melitensis carry a smooth LPS involved in the virulence of these bacteria. The LPS O-chain protects the bacteria from cellular cationic peptides, oxygen metabolites and complement-mediated lysis and it is a key molecule for Brucella survival and replication in the host. Here, we review i Brucella LPS structure; ii Brucella genome, iii genes involved in LPS biosynthesis; iv the interaction between LPS and innate immunity.

  9. Evaluation of four DNA extraction protocols for Brucella abortus detection by PCR in tissues from experimentally infected cows with the 2308 strain.

    Science.gov (United States)

    Vejarano, M P; Matrone, M; Keid, L B; Rocha, V C M; Ikuta, C Y; Rodriguez, C A R; Salgado, V R; Ferreira, F; Dias, R A; Telles, E O; Ferreira Neto, J S

    2013-04-01

    This study compared 4 protocols for DNA extraction from homogenates of 6 different organs of cows infected with the Brucella abortus 2308 strain. The extraction protocols compared were as follows: GT (guanidine isothiocyanate lysis), Boom (GT lysis with the carrying suspension diatomaceous earth), PK (proteinase K lysis), and Santos (lysis by boiling and freezing with liquid nitrogen). Positive and negative gold standard reference groups were generated by classical bacteriological methods. All samples were processed with the 4 DNA extraction protocols and amplified with the B4 and B5 primers. The number of positive samples in the placental cotyledons was higher than that in the other organs. The cumulated results showed that the Santos protocol was more sensitive than the Boom (p=0.003) and GT (p=0.0506) methods and was similar to the PK method (p=0.2969). All of the DNA extraction protocols resulted in false-negative results for PCR. In conclusion, despite the disadvantages of classical bacteriological methods, the best approach for direct diagnosis of B. abortus in organs of infected cows includes the isolation associated with PCR of DNA extracted from the cotyledon by the Santos or PK methods.

  10. Serological activity of white-tail deer against several species of Brucella.

    Science.gov (United States)

    Salinas-Meléndez, J A; Martínez-Muñoz, A; Avalos-Ramírez, R; Cerutti-Pereyra, N; Riojas-Valdés, V M

    1998-01-01

    In Mexico, brucellosis is a widely distributed disease of domesticated ruminants, but its frequency in wild ruminants has not been documented. Since northeast Mexico is the main distribution area of white-tailed deer and has been reported as an area positive for brucellosis in domesticated species, the present study was conducted in order to determine serological activity against several species of the genus Brucella in white-tailed deer. A total of 208 sera of white-tailed deer were collected during the springs of 1994 and 1995 in the north part of the states of Nuevo León and Coahuila. Each serum was analyzed for the detection of antibodies against two smooth (B. abortus and B. melitensis) and one rough (B. ovis) species of the genus Brucella. The serological tests used for the determination of the presence of antibodies against Brucella were card and plate agglutination for B. abortus, plate agglutination and rivanol precipitation for B. melitensis, and agar gel immunodiffusion for B. ovis. Each assay had positive and negative controls. None of the analyzed samples was found to be positive, and only two sera showed partial plate agglutination against B. melitensis at a dilution of 1:25; however, at higher dilutions and to the rivanol precipitation test the same samples were negative. Therefore, the percentage of positive sera was estimated at 0% (0/208). This result makes evident the absence of positive white-tailed deer against Brucella in the sampled area, despite that this disease is considered present in domesticated species. Therefore, white-tailed deer does not have, at the present time, an important role for the dispersion of the disease. The same result has been reported in other countries.

  11. Evalution of immunogical response of live Brucella vaccine for injection%注射用布氏菌活疫苗免疫学评价

    Institute of Scientific and Technical Information of China (English)

    陈成; 魏东; 李恪梅; 付丽丽; 解晓露; 黄长江; 王国治

    2014-01-01

    Objective To evaluate the immune response of live Brucella vaccine for intradermal injection, which detect humoral immunity and cellular immune responses in mice. Methods Mice were randomly divided into 3 groups with intradermal immune live Brucella vaccine, After 4 weeks, blood was sampled and spleen lymphocytes were separated. Then serum ELISA was used to detect the immune animal total IgG antibody, IgG1, IgG2a The number of spleen lymphocytes with secretion of IFN-γ, IL-4 was examined by ELISPOT. ELISA method was adopted to detect the level of cytokines IFN-γ, IL-4 from spleen cells after restimulation in vitro. After Brucella melitensis M5 infected animals, the spleen were isolated in sterile. The acquired immune protection of immune Brucella vaccine was evaluated through the bacteria count of spleen. Results The ELISA results showed that there exerted specific antibodies in mice infected by immune live Brucella vaccine. The number of spleen lymphocytes with secretion of INF-γwas increased through ELISPOT assay. The level of IFN-γwas higher than the control after detection of ELISA method. From this, live Brucella vaccine for intradermal injection mainly induced type TH1 immune response. The results from analysis of protection force of live Brucella vaccine demonstrated that the vaccine produced strong immune protection. Conclusion The method of live Brucella vaccine for intradermal injection is feasible, which can get intensively immunogenicity.%目的以小鼠为动物模型,通过检测布氏菌活疫苗免疫小鼠产生的体液免疫、细胞免疫应答及保护力,对其进行免疫学评价。  方法将30只小鼠随机分为3组,皮下免疫布氏活疫苗,免疫4周后分离血清及脾脏淋巴细胞。ELISA法检测免疫动物血清中总 IgG 抗体,抗体亚类 IgG1及 IgG2a;ELISPOT 法检测分泌 IFN-γ、IL-4的脾脏淋巴细胞数目,以及用 ELISA 方法检测体外再刺激后小鼠脾细胞分泌IFN-γ、IL-4的

  12. Immunogenicity and protective effect of recombinant Brucella abortus Ndk (rNdk) against a virulent strain B. abortus 544 infection in BALB/c mice.

    Science.gov (United States)

    Hop, Huynh Tan; Simborio, Hannah Leah; Reyes, Alisha Wehdnesday Bernardo; Arayan, Lauren Togonon; Min, WonGi; Lee, Hu Jang; Kim, Dong Hee; Chang, Hong Hee; Kim, Suk

    2015-02-01

    In this study, we particularly evaluated the protective effect of recombinant protein encoded by Brucella abortus 544 ndk (nucleoside diphosphate kinase) gene against B. abortus infection in the BALB/c mice. Cloning and expression of B. abortus Ndk was accomplished by PCR amplification into a pMAL expression system, and purification of a recombinant Ndk (rNdk). As for the determination of IgG responses, rNdk induced vigorous IgG production, especially higher in IgG2a compared to IgG1 with titers of 5.2 and 4.8, respectively, whereas titers of these in mice immunized with MBP were 2.4 of IgG2a and 2.6 of IgG1. The analysis of cytokine has revealed that rNdk can strongly induce production of IFN-γ as well as proinflammatory cytokines (TNF, MCP1 and IL-6) but not much IL-10, suggesting rNdk elicited predominantly cell-mediated immune responses. Furthermore, the spleen proliferation and bacterial burden in the spleen of rNdk immunized mice were significantly lower than those of MBP-immunized mice against virulent B. abortus challenge (P abortus might be a useful candidate for subunit vaccine for brucellosis in animals.

  13. Brucella as a potential agent of bioterrorism.

    Science.gov (United States)

    Doganay, Gizem D; Doganay, Mehmet

    2013-04-01

    Perception on bioterrorism has changed after the deliberate release of anthrax by the postal system in the United States of America in 2001. Potential bioterrorism agents have been reclassified based on their dissemination, expected rate of mortality, availability, stability, and ability to lead a public panic. Brucella species can be easily cultured from infected animals and human materials. Also, it can be transferred, stored and disseminated easily. An intentional contamination of food with Brucella species could pose a threat with low mortality rate. Brucella spp. is highly infectious through aerosol route, making it an attractive pathogen to be used as a potential agent for biological warfare purposes. Recently, many studies have been concentrated on appropriate sampling of Brucella spp. from environment including finding ways for its early detection and development of new decontamination procedures such as new drugs and vaccines. There are many ongoing vaccine development studies; some of which recently received patents for detection and therapy of Brucella spp. However, there is still no available vaccine for humans. In this paper, recent developments and recent patents on brucellosis are reviewed and discussed.

  14. 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

  15. 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).

  16. 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.

  17. Cloning and Sequence Analysis of omp31 Genes of Brucella Strain Isolated from Inner Mongolia%布鲁氏菌内蒙古分离株omp31基因的克隆与序列分析

    Institute of Scientific and Technical Information of China (English)

    陆静; 关平原; 乌日罕; 特木尔巴根; 马立峰; 菊花

    2009-01-01

    对三株布鲁氏菌内蒙古分离株的omp31基因进行克隆与序列分析.参照GenBank中布鲁氏菌的omp31基因序列,设计一对特异性引物,用PCR方法扩增布鲁氏菌omp31基因,对PCR产物进行克隆、测序,将测定序列拼接后与GenBank中获得的参考毒株omp31基因序列进行了比较.结果显示,三株布鲁氏菌分离株的omp31基因开放阅读框核苷酸序列同源性均为100.0%;与Brucella ceti B14/94、Brucella neotomae 5K33、Brucella suis 1330三个参考毒株omp31基因核苷酸序列同源性最高,为100.0%;与Brucella canis RM6/66、Brucella melitensis 183、Brucella melitensis 16M三个参考毒株omp31基因核苷酸序列同源性为99.9%;与Brucella melitensis Rev1参考毒株omp31基因核苷酸序列同源性为99.6%;与Brucella ovis ATCC 25840参考毒株omp31基因核苷酸序列同源性最低,为98.8%.

  18. Genotyping of human Brucella isolated by multiple locus variable numbers of tandem repeats analysis%布鲁菌多位点可变数目串联重复序列分析的分型研究

    Institute of Scientific and Technical Information of China (English)

    杨红霞; 张秋香; 郝瑞娥; 姚素霞; 张凡非; 李虹; 崔步云; 姜海

    2016-01-01

    目的 采用多位点可变数目串联重复序列分析(MLVA)方法对山西省分离的布鲁菌进行基因型分析.方法 2012、2013年,在山西省布鲁菌病(简称布病)监测点,采集已确诊的布病患者血液,进行血培养,对培养出的布鲁菌进行传统的生物分型鉴定.选取MLVA的16个位点[分为2组:panel 1、panel 2(包括panel 2A、panel 2B)],对分离的布鲁菌进行MLVA分型,并对分型结果进行聚类分析.结果 2012、2013年,共分离47株羊种3型布鲁菌;MLVA分型后进行聚类分析,发现分离的菌株高度同源,均为东地中海型;菌株可分为A群和B群,A群为优势基因群;panel 2B的Bruce04 、Bruce16及Bruce30位点的变异度较高.结论 在同一年份,相邻地区的布鲁菌菌株MLVA分型结果一致或相近,对追溯传染源、分析布病的流行和爆发有重要意义.panel 2B的Bruce04、Bruce16及Bruce30位点的变异度高,提示这3个位点对分析研究同一生物型菌株的变异情况有价值.%Objective To analyze the genotypes of Brucella isolated in Shanxi Province using multiple locus variable numbers of tandem repeats analysis (MLVA).Methods In brucellosis monitoring points in Shanxi Province in 2012 and 2013,the blood samples were collected from brucellosis patients,and blood culture was done,then the traditional identification of biological classification was carried out.MLVA-t6 [two groups:panel 1,panel 2(including panel 2A,panel 2B)] was used for typing of Brucella,then genotyping results were cluster analyzed.Results In 2012 and 2013,a total of 47 Brucella were isolated,all of the strains were Brucella melitensis type 3 and highly homologous,clustering in panel 1 to the"East Mediterranean" Brucella melitensis group.The strains were divided into groups A and B,and group A was an advantageous gene group;the variabilities of Bruce04,Bruce16and Bruce30 in panel 2B were higher.Conclusions The result of MLVA for Brucella is identical or similar in the same

  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. 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.

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

    Science.gov (United States)

    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.

  2. Microbiological study and antimicrobial susceptibilities of brucella isolates in serologic diagnosed cases

    Directory of Open Access Journals (Sweden)

    Abtin Heidarzadeh

    2009-01-01

    Full Text Available (Received 25 Nov, 2008 ; Accepted 4 Mar, 2009AbstractBackground and purpose: Brucellosis is a zoonotic disease with worldwide distribution that is endemic in Iran. Worldwide, brucellosis remains a major cause of morbidity in humans and domesticated animals. The disease has a wide spectrum of clinical manifestation and can affect a variety of organs and systems. This study focused on blood culture of serologic diagnosed brucellosis and antimicrobial susceptibility test.Materials and methods: In this cross sectional study, microbiologic survey was done on a total of 30 serum samples with STA titer of 1:160 or greater and 2ME titer of 1:40 or greater, which were presumptive for brucellosis. Blood cultures were done by lysis centrifugation and antimicrobial susceptibility test, against 9 antimicrobial agents by disk method. The data was analyzed by stata V8.0 software.Results: At the end this study, the blood culture isolation rate was 23.3 %( 7 cases out of 30 patients and all of the isolates were brucella melitensis. Antimicrobial susceptibility tests showed high in vitro activity of ofloxacin, ciprofloxacin and doxycycline and also, low in vitro activity of streptomycin and cotrimoxazole.Conclusion: Brucellosis is endemic in Iran. Brucella melitensis was the most common strain of brucella in our patients. Except cotrimoxazole and streptomycin, high in vitro activity was found with other antibrucella agents, especially with ofloxacin, ciprofloxacin and doxycycline. J Mazand Univ Med Sci 2009; 19(68: 74-78 (Persian

  3. 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

    responses in a YeO:9-purified O-antigen indirect ELISA did not decrease accordingly. Analysis of available cross-sectional serum samples from pig herds naturally infected with YeO: 9 or B. suis biovar 2 confirmed that the observed difference in the duration of the serological responses between the two...... infections could be used to discriminate between herds infected with B. suis biovar 2 and YeO:9....

  4. 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.

  5. Type IV Secretion System of Brucella spp. and its Effectors

    Directory of Open Access Journals (Sweden)

    Yuehua eKe

    2015-10-01

    Full Text Available Brucella spp. cause brucellosis in domestic and wild animals. They are intracellular bacterial pathogens and 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 will discuss roles of Brucella VirB T4SS and in more detail of all 15 identified effectors, which may be crucial for Brucella pathogenesis. VirB T4SS regulates the inflammation response and manipulates vesicle trafficking inside host cells, suggesting that it plays crucial roles in the inhibition of the host immune response and intracellular survival during infection. So, we listed some key molecular events in the intracellular life cycle of Brucella potentially targeted by the VirB T4SS effectors. Elucidating functions of the effectors secreted will be crucial to clarifying mechanism of T4SS during infection. Studying the effectors secreted by Brucella spp. might provide insights into the mechanisms by which the bacteria hijack the host signaling pathways, which help us to develop better vaccines and therapies against brucellosis.

  6. Brucella suis urease encoded by ure1 but not ure2 is necessary for intestinal infection of BALB/c mice

    Directory of Open Access Journals (Sweden)

    Sriranganathan Nammalwar

    2007-06-01

    Full Text Available Abstract Background In prokaryotes, the ureases are multi-subunit, nickel-containing enzymes that catalyze the hydrolysis of urea to carbon dioxide and ammonia. The Brucella genomes contain two urease operons designated as ure1 and ure2. We investigated the role of the two Brucella suis urease operons on the infection, intracellular persistence, growth, and resistance to low-pH killing. Results The deduced amino acid sequence of urease-α subunits of operons-1 and -2 exhibited substantial identity with the structural ureases of alpha- and beta-proteobacteria, Gram-positive and Gram-negative bacteria, fungi, and higher plants. Four ure deficient strains were generated by deleting one or more of the genes encoding urease subunits of B. suis strain 1330 by allelic exchange: strain 1330Δure1K (generated by deleting ureD and ureA in ure1 operon, strain 1330Δure2K (ureB and ureC in ure2 operon, strain 1330Δure2C (ureA, ureB, and ureC in ure2 operon, and strain 1330Δure1KΔure2C (ureD and ureA in ure1 operon and ureA, ureB, and ureC in ure2 operon. When grown in urease test broth, strains 1330, 1330Δure2K and 1330Δure2C displayed maximal urease enzyme activity within 24 hours, whereas, strains 1330Δure1K and 1330Δure1KΔure2C exhibited zero urease activity even 96 h after inoculation. Strains 1330Δure1K and 1330Δure1KΔure2C exhibited slower growth rates in tryptic soy broth relative to the wild type strain 1330. When the BALB/c mice were infected intraperitoneally with the strains, six weeks after inoculation, the splenic recovery of the ure deficient strains did not differ from the wild type. In contrast, when the mice were inoculated by gavage, one week after inoculation, strain 1330Δure1KΔure2C was cleared from livers and spleens while the wild type strain 1330 was still present. All B. suis strains were killed when they were incubated in-vitro at pH 2.0. When the strains were incubated at pH 2.0 supplemented with 10 mM urea, strain

  7. 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.

  8. Extended Multilocus Sequence Analysis to Describe the Global Population Structure of the Genus Brucella: Phylogeography and Relationship to Biovars

    Science.gov (United States)

    Whatmore, Adrian M.; Koylass, Mark S.; Muchowski, Jakub; Edwards-Smallbone, James; Gopaul, Krishna K.; Perrett, Lorraine L.

    2016-01-01

    An extended multilocus sequence analysis (MLSA) scheme applicable to the Brucella, an expanding genus that includes zoonotic pathogens that severely impact animal and human health across large parts of the globe, was developed. The scheme, which extends a previously described nine locus scheme by examining sequences at 21 independent genetic loci in order to increase discriminatory power, was applied to a globally and temporally diverse collection of over 500 isolates representing all 12 known Brucella species providing an expanded and detailed understanding of the population genetic structure of the group. Over 100 sequence types (STs) were identified and analysis of data provided insights into both the global evolutionary history of the genus, suggesting that early emerging Brucella abortus lineages might be confined to Africa while some later lineages have spread worldwide, and further evidence of the existence of lineages with restricted host or geographical ranges. The relationship between biovar, long used as a crude epidemiological marker, and genotype was also examined and showed decreasing congruence in the order Brucella suis > B. abortus > Brucella melitensis. Both the previously described nine locus scheme and the extended 21 locus scheme have been made available at http://pubmlst.org/brucella/ to allow the community to interrogate existing data and compare with newly generated data. PMID:28066370

  9. 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

  10. A comparison of two agar gel immunodiffusion methods and a complement fixation test for serologic diagnosis of Brucella ovis infection in experimentally infected rams

    Directory of Open Access Journals (Sweden)

    M.N. Xavier

    2011-08-01

    Full Text Available A infecção por Brucella ovis é considerada uma das principais causas de epididimite e infertilidade em carneiros, resultando em falhas reprodutivas e perdas econômicas significativas em rebanhos ovinos ao redor do mundo. O estudo teve o objetivo de avaliar três testes sorológicos disponíveis para o diagnóstico da brucelose ovina por B. ovis, utilizando 181 soros ovinos. Amostras de soro provenientes de carneiros experimentalmente infectados foram coletadas ao longo de 192 dias pós-infecção (n=117 e durante o período pré-infecção (n=9. Adicionalmente, amostras de soro foram obtidas de ovinos provenientes de um rebanho livre para B. ovis (n=55. As técnicas de imunodifusão em gel de agar (IDGA, utilizando dois antígenos disponíveis comercialmente, e de fixação de complemento foram comparadas (FC. Foram obtidos resultados de sensibilidade especificidade semelhantes para ambos os métodos de IDGA e ainda, a técnica de IDGA foi mais eficiente do que a da FC para o diagnóstico sorológico da infecção por B. ovis.

  11. Assessment of the diagnostic sensitivity and specificity of an indirect ELISA kit for the diagnosis of Brucella ovis infection in rams

    Directory of Open Access Journals (Sweden)

    Praud Anne

    2012-07-01

    Full Text Available Abstract Background Brucella ovis causes an infectious disease responsible for infertility and subsequent economic losses in sheep production. The standard serological test to detect B. ovis infection in rams is the complement fixation test (CFT, which has imperfect sensitivity and specificity in addition to technical drawbacks. Other available tests include the indirect enzyme-linked immunosorbent assays (I-ELISA but no I-ELISA kit has been fully evaluated. The study aimed to compare an I-ELISA kit and the standard CFT. Our study was carried out on serum samples from 4599 rams from the South of France where the disease is enzootic. A Bayesian approach was used to estimate tests characteristics (diagnostic sensitivity, Se and diagnostic specificity, Sp. The tests were then studied together in order to optimise testing strategies to detect B. ovis. Results After optimising the cut-off values in order to avoid doubtful results without deteriorating the concordance between the results of the two tests, the I-ELISA appeared to be slightly more sensitive than CFT (Se I-ELISA = 0.917 [0.822; 0.992], 95% Credibility Interval (CrI compared to Se CFT = 0.860 [0.740; 0.967], 95% CrI. However, CFT was slightly more specific than I-ELISA (Sp CFT = 0.988 [0.947; 1.0], 95% CrI compared to Sp I-ELISA =0.952 [0.901; 1.0], 95% CrI. The tests were then associated with two different interpretation schemes. The series association increased the specificity of screening and could be used for pre-movement testing in rams from uninfected flocks. The parallel association increased sequence sensitivity, thus appearing more suitable for eradicating the disease in infected flocks. Conclusions The high sensitivity and acceptable specificity of this I-ELISA kit support its potential interest to avoid the limitations of CFT. The two tests could also be used together or combined with other diagnostic methods such as semen culture to improve the testing strategy

  12. Intrinsic and selected resistance to antibiotics binding the ribosome: analyses of Brucella 23S rrn, L4, L22, EF-Tu1, EF-Tu2, efflux and phylogenetic implications

    Directory of Open Access Journals (Sweden)

    Jensen Allen E

    2006-10-01

    Full Text Available Abstract Background Brucella spp. are highly similar, having identical 16S RNA. However, they have important phenotypic differences such as differential susceptibility to antibiotics binding the ribosome. Neither the differential susceptibility nor its basis has been rigorously studied. Differences found among other conserved ribosomal loci could further define the relationships among the classical Brucella spp. Results Minimum inhibitory concentration (MIC values of Brucella reference strains and three marine isolates to antibiotics binding the ribosome ranged from 0.032 to >256 μg/ml for the macrolides erythromycin, clarithromycin, and azithromycin and 2 to >256 μg/ml for the lincosamide, clindamycin. Though sequence polymorphisms were identified among ribosome associated loci 23S rrn, rplV, tuf-1 and tuf-2 but not rplD, they did not correlate with antibiotic resistance phenotypes. When spontaneous erythromycin resistant (eryR mutants were examined, mutation of the peptidyl transferase center (A2058G Ec correlated with increased resistance to both erythromycin and clindamycin. Brucella efflux was examined as an alternative antibiotic resistance mechanism by use of the inhibitor L-phenylalanine-L-arginine β-naphthylamide (PAβN. Erythromycin MIC values of reference and all eryR strains, except the B. suis eryR mutants, were lowered variably by PAβN. A phylogenetic tree based on concatenated ribosomal associated loci supported separate evolutionary paths for B. abortus, B. melitensis, and B. suis/B. canis, clustering marine Brucella and B. neotomae with B. melitensis. Though Brucella ovis was clustered with B. abortus, the bootstrap value was low. Conclusion Polymorphisms among ribosomal loci from the reference Brucella do not correlate with their highly differential susceptibility to erythromycin. Efflux plays an important role in Brucella sensitivity to erythromycin. Polymorphisms identified among ribosome associated loci construct a

  13. Molecular cloning, purification and immunogenicity of recombinant Brucella abortus 544 malate dehydrogenase protein.

    Science.gov (United States)

    Reyes, Alisha Wehdnesday Bernardo; Simborio, Hannah Leah Tadeja; Hop, Huynh Tan; Arayan, Lauren Togonon; Kim, Suk

    2016-03-01

    The Brucella mdh gene was successfully cloned and expressed in E. coli. The purified recombinant malate dehydrogenase protein (rMDH) was reactive to Brucella-positive bovine serum in the early stage, but not reactive in the middle or late stage, and was reactive to Brucella-positive mouse serum in the late stage, but not in the early or middle stage of infection. In addition, rMDH did not react with Brucella-negative bovine or mouse sera. These results suggest that rMDH has the potential for use as a specific antigen in serological diagnosis for early detection of bovine brucellosis.

  14. 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.

  15. Epidemiology of Brucella Infection in the Human, Livestock and Wildlife Interface in the Katavi-Rukwa Ecosystem, Tanzania.

    OpenAIRE

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

    2015-01-01

    Background 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...

  16. 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.

  17. 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.

  18. 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.

  19. 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.

  20. Risk factors for Brucella seropositivity in goat herds in eastern and western Uganda.

    Science.gov (United States)

    Kabagambe, E K; Elzer, P H; Geaghan, J P; Opuda-Asibo, J; Scholl, D T; Miller, J E

    2001-12-03

    Cross-sectional prevalences and risk factors for Brucella seropositivity in goats in eastern and western Uganda were investigated. Serum was collected from 1518 goats randomly selected from 145 herds which had been identified using multistage sampling. The brucellosis card test (CT) and the Brucella melitensis tube-agglutination test (TAT) were used in parallel to detect antibodies against B. abortus and B. melitensis, respectively. Interviewer-administered questionnaires were used to collect information on goat health and management. This information was used in multivariable logistic-regression models to determine the risk factors for Brucella seropositivity in goat herds. For each analysis, a herd was considered positive if at least one goat in the herd tested positive for antibodies against Brucella and negative if none was positive. Four percent (55/1480) of the goats screened with the CT had antibodies against Brucella. The reactors were distributed in 13% (19/145) of the herds. The most-important herd-level risk factors identified were use of a hired caretaker as the primary manager of the operation compared to owner/family members (adjusted odds ratio (OR)=8.1; 95% CI 1.6, 39.7), keeping sheep in addition to goats (OR=6.0; CI 1.5, 23.7) compared to having no sheep, and free browsing (OR=4.7; 95% CI 1.0, 20.7) when compared to tethering or zero-grazing. Using the TAT, 10% (141/1446) of the goats tested positive. The positives were distributed in 43% (63/145) of the herds. Free browsing (OR=6.7; 95% CI 2.7, 16.9) when compared to tethering or zero-grazing and lack of veterinary care (OR=2.9; CI 1.3, 6.7) were the most-important factors identified in the multivariable model for B. melitensis herd seropositivity. To explore/reduce the risk of misclassification in a secondary analysis, herds were reclassified as positive if at least one goat tested positive on both tests and negative if none of the goats was positive on any of the two tests. Using this

  1. Inquérito soro-epidemiológico e fatores de risco associados à infecção por Brucella ovis em carneiros deslanados do semi-árido da Paraíba Sero-epidemiological survey and risk factors for Brucella ovis infection in rams of the semiarid region of Paraíba, northeastern Brazil

    Directory of Open Access Journals (Sweden)

    Inácio J. Clementino

    2007-04-01

    Full Text Available Foi realizado um levantamento soro-epidemiológico da brucelose por Brucella ovis em reprodutores ovinos deslanados na Paraíba com os objetivos de verificar a prevalência e distribuição da infecção por B. ovis em propriedades rurais e analisar os possíveis fatores de risco associados à infecção. Foram investigadas 283 propriedades criadoras de ovinos, das quais foram colhidas 498 amostras de soro sanguíneo de carneiros, a partir de oito meses de idade, nas mesorregiões do Sertão Paraibano e Borborema. Todos os soros foram examinados pela IDGA (teste de triagem e RFC (teste confirmatório. De acordo com as análises, 8,59% (I.C.95% = 5,83%-12,48% das propriedades apresentaram evidência sorológica de infecção por B. ovis, com uma prevalência de 5,57% (I.C.95% = 3,86%-7,97% de reprodutores sororreagentes. Nas propriedades que higienizavam suas instalações com maior freqüência, a soropositividade foi estatisticamente inferior (p A sero-epidemiological survey was conducted to determine the prevalence and distribution of Brucella ovis infection in rams in the state of Paraíba, northeastern Brazil. The risk factors for the infection were also verified. Serum samples from 498 rams, 8 months of age or older, of 283 sheep herds in the Sertão Paraibano and Borborema mesoregions were investigated. All sera were examined by AGID test (screening test and CFT (confirmatory test. From the total of examined herds, 8.59% (95% CI = 5.83%-12.48% were seropositive for Brucella ovis. The prevalence of seropositive rams was 5.57% (95% CI = 3.86%-7.97%. The seropositivity was lower in herds where cleanliness was frequently made (p< 0.05.

  2. 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.

  3. U-Omp19 from Brucella abortus Is a Useful Adjuvant for Vaccine Formulations against Salmonella Infection in Mice

    Science.gov (United States)

    Risso, Gabriela S.; Carabajal, Marianela V.; Bruno, Laura A.; Ibañez, Andrés E.; Coria, Lorena M.; Pasquevich, Karina A.; Lee, Seung-Joo; McSorley, Stephen J.; Briones, Gabriel; Cassataro, Juliana

    2017-01-01

    Most pathogens infect through mucosal surfaces, and parenteral immunization typically fails to induce effective immune responses at these sites. Development of oral-administered vaccines capable of inducing mucosal as well as systemic immunity while bypassing the issues of antigen degradation and immune tolerance could be crucial for the control of enteropathogens. This study demonstrates that U-Omp19, a bacterial protease inhibitor with immunostimulatory features, coadministered with Salmonella antigens by the oral route, enhances mucosal and systemic immune responses in mice. U-Omp19 was able to increase antigen-specific production of IFN-γ and IL-17 and mucosal (IgA) antibody response. Finally, oral vaccination with U-Omp19 plus Salmonella antigens conferred protection against virulent challenge with Salmonella Typhimurium, with a significant reduction in bacterial loads. These findings prove the efficacy of this novel adjuvant in the Salmonella infection model and support the potential of U-Omp19 as a suitable adjuvant in oral vaccine formulations against mucosal pathogens requiring T helper (Th)1–Th17 protective immune responses.

  4. Recent advances in Brucella abortus vaccines

    OpenAIRE

    Dorneles, Elaine Maria S.; Sriranganathan, Nammalwar; Andrey P. Lage

    2015-01-01

    Abstract Brucella abortus vaccines play a central role in bovine brucellosis control/eradication programs and have been successfully used worldwide for decades. Strain 19 and RB51 are the approved B. abortus vaccines strains most commonly used to protect cattle against infection and abortion. However, due to some drawbacks shown by these vaccines much effort has been undertaken for the development of new vaccines, safer and more effective, that could also be used in other susceptible species ...

  5. Recent advances in Brucella abortus vaccines

    OpenAIRE

    2015-01-01

    International audience; Brucella abortus vaccines play a central role in bovine brucellosis control/eradication programs and have been successfully used worldwide for decades. Strain 19 and RB51 are the approved B. abortus vaccines strains most commonly used to protect cattle against infection and abortion. However, due to some drawbacks shown by these vaccines much effort has been undertaken for the development of new vaccines, safer and more effective, that could also be used in other susce...

  6. Identification of Brucella by MALDI-TOF mass spectrometry. Fast and reliable identification from agar plates and blood cultures.

    Directory of Open Access Journals (Sweden)

    Laura Ferreira

    Full Text Available BACKGROUND: MALDI-TOF mass spectrometry (MS is a reliable method for bacteria identification. Some databases used for this purpose lack reference profiles for Brucella species, which is still an important pathogen in wide areas around the world. We report the creation of profiles for MALDI-TOF Biotyper 2.0 database (Bruker Daltonics, Germany and their usefulness for identifying brucellae from culture plates and blood cultures. METHODOLOGY/PRINCIPAL FINDINGS: We created MALDI Biotyper 2.0 profiles for type strains belonging to B. melitensis biotypes 1, 2 and 3; B. abortus biotypes 1, 2, 5 and 9; B. suis, B. canis, B ceti and B. pinnipedialis. Then, 131 clinical isolates grown on plate cultures were used in triplicate to check identification. Identification at genus level was always correct, although in most cases the three replicates reported different identification at species level. Simulated blood cultures were performed with type strains belonging to the main human pathogenic species (B. melitensis, B. abortus, B. suis and B. canis, and studied by MALDI-TOF MS in triplicate. Identification at genus level was always correct. CONCLUSIONS/SIGNIFICANCE: MALDI-TOF MS is reliable for Brucella identification to the genus level from culture plates and directly from blood culture bottles.

  7. Identification of Brucella by MALDI-TOF Mass Spectrometry. Fast and Reliable Identification from Agar Plates and Blood Cultures

    Science.gov (United States)

    Ferreira, Laura; Vega Castaño, Silvia; Sánchez-Juanes, Fernando; González-Cabrero, Sandra; Menegotto, Fabiola; Orduña-Domingo, Antonio

    2010-01-01

    Background MALDI-TOF mass spectrometry (MS) is a reliable method for bacteria identification. Some databases used for this purpose lack reference profiles for Brucella species, which is still an important pathogen in wide areas around the world. We report the creation of profiles for MALDI-TOF Biotyper 2.0 database (Bruker Daltonics, Germany) and their usefulness for identifying brucellae from culture plates and blood cultures. Methodology/Principal Findings We created MALDI Biotyper 2.0 profiles for type strains belonging to B. melitensis biotypes 1, 2 and 3; B. abortus biotypes 1, 2, 5 and 9; B. suis, B. canis, B ceti and B. pinnipedialis. Then, 131 clinical isolates grown on plate cultures were used in triplicate to check identification. Identification at genus level was always correct, although in most cases the three replicates reported different identification at species level. Simulated blood cultures were performed with type strains belonging to the main human pathogenic species (B. melitensis, B. abortus, B. suis and B. canis), and studied by MALDI-TOF MS in triplicate. Identification at genus level was always correct. Conclusions/Significance MALDI-TOF MS is reliable for Brucella identification to the genus level from culture plates and directly from blood culture bottles. PMID:21151913

  8. Small GTPases and Brucella entry into the endoplasmic reticulum.

    Science.gov (United States)

    de Bolle, Xavier; Letesson, Jean-Jacques; Gorvel, Jean-Pierre

    2012-12-01

    A key determinant for intracellular pathogenic bacteria to ensure their virulence within host cells is their ability to bypass the endocytic pathway and to reach a safe niche of replication. In the case of Brucella, the bacterium targets the ER (endoplasmic reticulum) to create a replicating niche called the BCV (Brucella-containing vacuole). The ER is a suitable strategic place for pathogenic Brucella. Indeed, bacteria can be hidden from host cell defences to persist within the host, and they can take advantage of the membrane reservoir delivered by the ER to replicate. Interaction with the ER leads to the presence on the BCV of the GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and the small GTPase Rab2 known to be located on secretory vesicles that traffic between the ER and the Golgi apparatus. GAPDH and the small GTPase Rab2 controls Brucella replication at late times post-infection. A specific interaction between the human small GTPase Rab2 and a Brucella spp. protein named RicA was identified. Altered kinetics of intracellular trafficking and faster proliferation of the Brucella abortus ΔricA mutant was observed compared with the wild-type strain. RicA is the first reported effector with a proposed function for B. abortus.

  9. Abortion in cattle due to infection with Staphylococcus lugdunensis.

    Science.gov (United States)

    Ardigò, Paolo; D'Incau, Mario; Pongolini, Stefano

    2014-11-01

    An aborted fetus of 7 months gestation, the associated placenta, and a single blood sample from the dam were submitted for diagnostic investigation to the diagnostic laboratory of the Lombardy and Emilia-Romagna Experimental Zooprophylactic Institute in Parma, Italy. The serum was negative for Neospora caninum, Coxiella burnetii, Chlamydophila abortus, Bovine herpesvirus 1 (BHV-1), Bovine viral diarrhea virus (BVDV), Brucella abortus, and Brucella melitensis. Fetal tissues and placental cotyledons were pooled and tested by polymerase chain reaction (PCR) for the presence of BHV-1, Bovine herpesvirus 4, BVDV, N. caninum, C. burnetii, Chlamydophila spp., Schmallemberg virus, and Leptospira interrogans. All PCR assays were negative. Bacteriological examinations performed on the fetal organs revealed a pure growth of Staphylococcus lugdunensis in all organs cultured. In human beings, S. lugdunensis is responsible for community-acquired and nosocomial infections, in both immunocompetent and immunocompromised patients. In veterinary medicine, the pathogenic potential of S. lugdunensis has not been fully investigated. The incidence of S. lugdunensis is regarded as being underreported because it could be easily misidentified as Staphylococcus aureus. The current report documents the ability of S. lugdunensis to cause abortion in cattle, indicating the need for accurate diagnostic procedures to identify this emerging and zoonotic pathogen whose incidence is likely underestimated in both human and veterinary medicine.

  10. "EPIDEMIOLOGY OF BRUCELLOSIS IN ISFAHAN (SEASONAL VARIATION IN MILK CONTAMINATION WITH BRUCELLA"

    Directory of Open Access Journals (Sweden)

    A. Owrang

    1972-06-01

    Full Text Available Milk samples were collected once every two weeks from 69 milk distribution centers for a period of one year. The samples were examined for Brucella infection by means of a ring test. It was shown that the infection of milk has no relation to any special season. Cattle, sheep and goat milk were all found infected. Some of the positive milk specimens were cultured and Brucella was isolated.

  11. Application of multiplex polymerase chain reaction to identify Brucella%应用多重聚合酶链式反应鉴别布鲁杆菌

    Institute of Scientific and Technical Information of China (English)

    韩丽红; 刘志国; 王妙; 刘日宏; 崔步云

    2013-01-01

    目的 探讨应用多重聚合酶链式反应(Muhiple-PCR)进行布鲁杆菌株种型的鉴别.方法 以6株布鲁杆菌标准菌株(牛种、羊种、猪种、犬种、绵羊附睾种、沙林鼠种布鲁杆菌标准菌株)作为阳性对照,以大肠埃希菌O∶157和小肠结肠炎耶尔森菌O∶9作为阴性对照,待测布鲁杆菌株29株.先用布鲁杆菌属特异性聚合酶链式反应(BCSP31-PCR)扩增上述待测布鲁杆菌株,扩增结果为阳性的菌株再用Muhiple-PCR方法进行布鲁杆菌种型鉴别.结果 29株待测布鲁杆菌株BCSP31-PCR方法扩增均为阳性,进一步Multiple-PCR方法扩增均为阳性,其中有20株鉴定为羊种菌,5株鉴定为猪种菌、3株鉴定为牛种菌、1株鉴定为犬种菌.结论 Multiple-PCR方法是一种快速、特异、简便、低风险的布鲁杆菌种型鉴定方法.%Objective To evaluate the effect of multiplex polymerase chain reaction(Multiple-PCR) in identification of Brucella strains.Methods Six standard Brucella strains (Brucella abortus,Brucella melitensis,Brucella suis,Brucella canis,Brucella ovis,Brucella neotomae) were used as positive controls and Escherichia coli O∶157 and Yersinia enterocolitica O∶9 were used as negative controls.A total of 29 Brucella strains were tested.Brucella strains were amplified by BCSP31-PCR and the species of Brucella with positive results were identified with Multiple-PCR method.Results The results of all 29 amplified Brucella strains were positive with BCSP31-PCR.The identified results of all Brucella strains were positive with Multiple-PCR,including 20 strains of Brucella melitensis,5 isolates of Brucella suis,3 strains of Brucella abortus and 1 strain of Brucella canis.Conclusion Multiple-PCR method is a rapid,specific,simple and low risk method for identification of Brucella species.

  12. A comparison of titers of anti-Brucella antibodies of naturally infected and healthy vaccinated cattle by standard tube agglutination test, microtiter plate agglutination test, indirect hemagglutination assay, and indirect enzyme-linked immunosorbent assay

    Directory of Open Access Journals (Sweden)

    Anju Mohan

    2016-07-01

    Full Text Available Aim: We determined the antibody response in cattle naturally infected with brucellosis and normal healthy adult cattle vaccinated during calf hood with strain 19. Materials and Methods: The antibody titers were measured by standard tube agglutination test (STAT, microtiter plate agglutination test (MAT, indirect hemagglutination assay (IHA, and indirect enzyme-linked immunosorbent assay (iELISA as per standard protocols. Results: The mean STAT titers were 1.963±0.345 in infected cattle and 1.200±0.155 in healthy vaccinated cattle. The difference was extremely significant (p<0.0001. The mean MAT titers were 2.244±0.727 in infected cattle and 1.200±0.155 in healthy vaccinated cattle. The difference was very significant (p<0.005. The mean IHA titers in infected cattle were 2.284±0.574, and those in healthy vaccinated cattle were 1.200±0.155. The difference was extremely significant (p=0.0002. However, the difference in mean iELISA titers of infected cattle (1.3678±0.014 and healthy vaccinated cattle (1.367±0.014 was non-significant. The infected animals showed very high titers of agglutinating antibodies compared to the vaccinated animals. However, it cannot be ascertained whether these antibodies are due to vaccine or response to infection. Since the infected animals had been vaccinated earlier, the current infection may suggest that vaccination was unable to induce protective levels of antibody. The heightened antibody response after infection may also indicate a secondary immune response to the antigens common to the vaccine strain and wild Brucella organisms. Conclusion: The brucellosis infected animals showed very high titers of agglutinating antibodies compared to the vaccinated animals.

  13. 河北省布鲁菌流行菌株生物学特征分析%Analysis on biological characteristics of epidemic Brucella strain in Hebei province

    Institute of Scientific and Technical Information of China (English)

    钱振宇; 姜霞; 贾肇一; 何宝花; 王颖童; 李月平; 孙印旗; 刘晓丽

    2013-01-01

    目的 了解河北省布鲁菌分离株的生物学特征.方法 应用传统分型鉴定方法和分子生物学方法确定布鲁菌属与种型鉴定.结果 8株分离菌株经传统分型方法鉴定均为羊种菌,羊1型1株,羊3型6株,粗糙型羊种菌1株;进一步采用布鲁菌属特异性BCSP31-PCR均扩增出223 bp的特异性条带;经种/型特异性AMOS-PCR均扩增出731bp的特异性条带,鉴定为羊种布鲁菌.结论 河北省近两年人感染布鲁氏菌的流行菌株为羊种菌3型,AMOS-PCR方法可以作为该省布鲁菌分离株鉴定的辅助手段.%OBJECTIVE To understand the biological characteristics of Brucella isolated in Hebei Province.METHODS 8 strains of Brucella from 2010 to 2011 were identified by traditional methods and molecular techniques.RESULTS The results showed that 8 strains were identified as B.melitensis by traditional methods.Among them,1 strain was classified as B.melitensis biotype 1,6 strains were B.melitensis biotype 3,1 strain was rough B.melitensis.And all these 8 strains were further identified as Brucella spp.by genus specific BCSP31-PCR.Specific fragments of the eight Bacteria spp.from Hebei province were identified by AMOS PCR as B.melitensis.CONCLUSION These data indicate that B.melitensis biotype 3 is the predominant species of Brucella in Hebei province,and AMOS assays may be used as the assistant tools in the identification of Brucella strains.

  14. Characterization of the Urease Operon of Brucella abortus and Assessment of Its Role in Virulence of the Bacterium▿

    OpenAIRE

    Sangari, Félix J; Seoane, Asunción; Rodríguez, María Cruz; Agüero, Jesús; García Lobo, Juan M.

    2006-01-01

    Most members of the genus Brucella show strong urease activity. However, the role of this enzyme in the pathogenesis of Brucella infections is poorly understood. We isolated several Tn5 insertion mutants deficient in urease activity from Brucella abortus strain 2308. The mutations of most of these mutants mapped to a 5.7-kbp DNA region essential for urease activity. Sequencing of this region, designated ure1, revealed the presence of seven open reading frames corresponding to the urease struc...

  15. 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.

  16. 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

    2014-05-01

    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.

  17. 福建省布鲁氏菌分离株的分子生物学鉴定%Molecular identification of the Brucella strains isolated in Fujian province

    Institute of Scientific and Technical Information of China (English)

    邓艳琴; 王加熊; 林代华; 陈亮; 王灵岚

    2009-01-01

    AMOS-PCR and MLVA were carried out to identify the Brucella strains isolated in Fujian province, which were classified as B. melitensis biovar 2 or 3 by conventional microbiological tests. All these 3 isolates were identified to be B. melitensis by AMOS-PCR. The genetic patterns obtained by MLVA were queried in the Brucella 2007 database and clustered with B. melitensis strains. It is evident that these two molecular assays may be used as the assistant tools in the identification of Brucella strains.%目的 利用分子生物学方法鉴定福建省布鲁氏菌分离株.方法 对布鲁氏菌分离株进行AMOS-PCR和MLVA分型.结果 3株福建省分离株经AMOS-PCR鉴定为羊种,与传统生物学分型一致;MLVA分型将其分为3个基因型,聚类分析鉴定为羊种.结论 AMOS-PCR、MLVA分型可以作为我省布鲁氏菌分离株鉴定的辅助手段.

  18. 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. PMID:26959235

  19. 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.

  20. Caracterização epidemiológica e fatores de risco associados à infecção por Brucella ovis em ovinos deslanados do semiárido paraibano Epidemiological characterization and risk factors associated with Brucella ovis infection in sheep in the Brazilian semiarid

    Directory of Open Access Journals (Sweden)

    Fabrine A. Santos

    2013-04-01

    Full Text Available Este trabalho teve como objetivo determinar a prevalência de rebanhos ovinos positivos (focos e de animais soropositivos para Brucella ovis na mesorregião do Sertão, Estado da Paraíba, Nordeste do Brasil, bem como identificar fatores de risco. Foram colhidas amostras de sangue de 1.134 animais procedentes de 103 rebanhos em 17 municípios. Para o diagnóstico sorológico da infecção por B. ovis foi utilizado o teste de imunodifusão em gel de ágar (IDGA. Um rebanho foi considerado positivo quando apresentou pelo menos um animal soropositivo. Das 103 propriedades utilizadas 21 (20,39% apresentaram pelo menos um animal soropositivo e dos 1.134 animais, 59 (5,20% foram soropositivos. Realizar higiene nas instalações com periodicidade anual (odds ratio = 7,13; IC 95% = 1,56-32,47; p=0,011 e aquisição de animais (odds ratio = 6,06; IC 95% = 1,39-26,48; p=0,017 foram identificados como fatores de risco. Com base na análise de fatores de risco, recomenda-se a realização de diagnóstico da infecção por B. ovis previamente à aquisição de animais e realização periódica de higienização das instalações.The aim of this investigation was to determine the seroprevalence of Brucella ovis in sheep flocks and individual sheep in the Sertão mesorregion, Paraíba state, Northeastern Brazil, as well as to identify risk factors. Blood samples were collected from 1,134 sheep from 103 flocks in 17 counties. For the serological diagnosis of B. ovis infection the agar gel immunodiffusion test (AGID was carried out. A flock was considered positive when there was at least one seropositive animal. Of the 103 flocks used, 21 (20.39% presented at least one seropositive sheep, and of the 1,134 sheep examined 59 (5.20% seropositive animals were diagnosed. Cleaning of facilities (odds ratio = 7.13; 95% CI=1.56-32.47; p=0.011 and purchase of animals (odds ratio = 6.06; 95% CI=1.39-26.48; p=0.017 were identified as risk factors. Based on the risk factor

  1. BrucellaBase: Genome information resource.

    Science.gov (United States)

    Sankarasubramanian, Jagadesan; Vishnu, Udayakumar S; Khader, L K M Abdul; Sridhar, Jayavel; Gunasekaran, Paramasamy; Rajendhran, Jeyaprakash

    2016-09-01

    Brucella sp. causes a major zoonotic disease, brucellosis. Brucella belongs to the family Brucellaceae under the order Rhizobiales of Alphaproteobacteria. We present BrucellaBase, a web-based platform, providing features of a genome database together with unique analysis tools. We have developed a web version of the multilocus sequence typing (MLST) (Whatmore et al., 2007) and phylogenetic analysis of Brucella spp. BrucellaBase currently contains genome data of 510 Brucella strains along with the user interfaces for BLAST, VFDB, CARD, pairwise genome alignment and MLST typing. Availability of these tools will enable the researchers interested in Brucella to get meaningful information from Brucella genome sequences. BrucellaBase will regularly be updated with new genome sequences, new features along with improvements in genome annotations. BrucellaBase is available online at http://www.dbtbrucellosis.in/brucellabase.html or http://59.99.226.203/brucellabase/homepage.html.

  2. 2010-2013年贵州省12株布鲁菌分离株分子分型研究%Molecular typing of 12 Brucella strains isolated in Guizhou province in 2010-2013

    Institute of Scientific and Technical Information of China (English)

    王月; 陈红; 刘英; 周敬祝; 李世军; 黄艳; 唐光鹏; 王定明; 陈贵春

    2015-01-01

    31-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). Results 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. Conclusion 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.

  3. 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.

  4. 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.

  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. 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

    2017-03-03

    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.

  7. Species-specific PCR for the Diagnosis and Determination of Antibiotic Susceptibilities of Brucella Strains Isolated from Tehran, Iran

    Science.gov (United States)

    Irajian, Gholam Reza; Masjedian Jazi, Faramarz; Mirnejad, Reza; Piranfar, Vahhab; Zahraei salehi, Taghi; Amir Mozafari, Noor; Ghaznavi-rad, Ehsanollah; Khormali, Mahmoud

    2016-01-01

    Background: Brucellosis is an endemic zoonotic disease in the Middle East. This study intended to design a uniplex PCR assay for the detection and differentiation of Brucella at the species level and determining the antibiotic susceptibility pattern of Brucella in Iran. Methods: Sixty-eight Brucella specimens (38 animal and 30 human specimens) were analyzed using PCR (using one pair of primers). Antibiotic susceptibility patterns were evaluated and compared using the E-Test and disk diffusion susceptibility test. Tigecycline susceptibility pattern was compared with other antibiotics. Results: Thirty six isolates of B. melitensis, 2 isolates of B. abortus and 1 isolate of B. suis from the 38 animal specimens, 24 isolates of B. melitensis and 6 isolates of B. abortus from the 30 human specimens were differentiated. The MIC50 values of doxycycline for human and animal specimens were 125 and 10 μg/ml, respectively, tigecycline 0.064 μg/ml for human specimens and 0.125μg/ml for animal specimens, and trimethoprim/ sulfamethoxazole and ciprofloxacin 0.065 and 0.125μg/ml, respectively, for both human and animal specimens. The highest MIC50 value of streptomycin in the human specimens was 0.5μg/ml and 1μg/ml for the animal specimens. The greatest resistance shown was to tetracycline and gentamicin, respectively. Conclusion: Uniplex PCR for the detection and differentiation of Brucella at the strain level is faster and less expensive than multiplex PCR, and the antibiotics doxycycline, rifampin, trimethoprim-sulfamethoxazole, ciprofloxacin, and ofloxacin are the most effective antibiotics for treating brucellosis. Resistance to tigecycline is increasing, and we recommend that it be used in a combination regimen. PMID:27799972

  8. 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.

  9. Seroprevalence and potential risk factors for Brucella spp. infection in traditional cattle, sheep and goats reared in urban, periurban and rural areas of Niger.

    Directory of Open Access Journals (Sweden)

    Abdou Razac Boukary

    Full Text Available INTRODUCTION: In Niamey, Niger, interactions within the interface between animals, humans and the environment induce a potential risk of brucellosis transmission between animals and from animals to humans. Currently, little is known about the transmission of Brucella in this context. RESULTS: 5,192 animals from 681 herds were included in the study. Serum samples and hygroma fluids were collected. A household survey enabled to identify the risk factors for transmission of brucellosis. The true adjusted herd-level prevalence of brucellosis ranged between 11.2% and 17.2% and the true adjusted animal-population level prevalence was 1.3% (95% CI: 0.9-1.8% based on indirect ELISA test for Brucella antibodies. Animals aged of 1-4 years were found to be more susceptible than animals less than 1 year old (Odds ratio [OR] of 2.7; 95% CI: 1.43-5.28. For cattle, the odds of brucellosis seropositivity were higher in rural compared to the periurban areas (OR of 2.8; 95% CI: 1.48-5.17 whereas for small ruminants the risk of seropositivity appeared to be higher in urban compared to periurban areas (OR of 5.5; 95% CI: 1.48-20.38. At herd level, the risk of transmission was increased by transhumance (OR of 5.4; 95% CI: 2.84-10.41, the occurrence of abortions (OR of 3.0; 95% CI: 1.40-6.41, and for herds having more than 50 animals (OR of 11.0; 95% CI: 3.75-32.46. Brucella abortus biovar 3 was isolated from the hygromas. CONCLUSION: brucellosis in Niger is a serious problem among cattle especially in the rural areas around Niamey and among sheep in the urban areas of Niamey. The seroprevalence varies across strata and animal species with important risk factors including herd size, abortion and transhumance at herd level and age at animal population level. For effective control of brucellosis, an integrated approach seems appropriate involving all stakeholders working in public and animal health.

  10. 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.

  11. The role of TREM-2 in internalization and intracellular survival of Brucella abortus in murine macrophages.

    Science.gov (United States)

    Wei, Pan; Lu, Qiang; Cui, Guimei; Guan, Zhenhong; Yang, Li; Sun, Changjiang; Sun, Wanchun; Peng, Qisheng

    2015-02-15

    Triggering receptor expressed on myeloid cells-2 (TREM-2) is a cell surface receptor primarily expressed on macrophages and dendritic cells. TREM-2 functions as a phagocytic receptor for bacteria as well as an inhibitor of Toll like receptors (TLR) induced inflammatory cytokines. However, the role of TREM-2 in Brucella intracellular growth remains unknown. To investigate whether TREM-2 is involved in Brucella intracellular survival, we chose bone marrow derived macrophages (BMDMs), in which TREM-2 is stably expressed, as cell model. Colony formation Units (CFUs) assay suggests that TREM-2 is involved in the internalization of Brucella abortus (B. abortus) by macrophages, while silencing of TREM-2 decreases intracellular survival of B. abortus. To further study the underlying mechanisms of TREM-2-mediated bacterial intracellular survival, we examined the activation of B. abortus-infected macrophages through determining the kinetics of activation of the three MAPKs, including ERK, JNK and p38, and measuring TNFα production in response to lipopolysaccharide (LPS) of Brucella (BrLPS) or B. abortus stimulation. Our data show that TREM-2 deficiency promotes activation of Brucella-infected macrophages. Moreover, our data also demonstrate that macrophage activation promotes killing of Brucella by enhancing nitric oxygen (NO), but not reactive oxygen species (ROS) production, macrophage apoptosis or cellular death. Taken together, these findings provide a novel interpretation of Brucella intracellular growth through inhibition of NO production produced by TREM-2-mediated activated macrophages.

  12. 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.

  13. 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. PMID:28036367

  14. In vitro antimicrobial susceptibility testing of human Brucella melitensis isolates from Qatar between 2014 - 2015

    NARCIS (Netherlands)

    Deshmukh, A.; Hagen, F.; Sharabasi, O.A.; Abraham, M.; Wilson, G.; Doiphode, S.; Maslamani, M.A.; Meis, J.F.G.M.

    2015-01-01

    BACKGROUND: Brucellosis is one of the most common zoonotic disease affecting humans and animals and is endemic in many parts of the world including the Gulf Cooperation Council region (GCC). The aim of this study was to identify the species and determine the antimicrobial susceptibility pattern of B

  15. Analysis of the brucella pathogen and its molecular genotype in Guangdong province%广东省布鲁杆菌的病原学及分子分型分析

    Institute of Scientific and Technical Information of China (English)

    陈经雕; 刘美真; 柯碧霞; 谭海玲; 李柏生; 张万里; 柯昌文

    2012-01-01

    目的 分析广东省布鲁杆菌病原学及分子分型特征.方法 对2010年广东省分离出的19株布鲁杆菌菌株进行传统生物学表型验证,同时进行PCR试验和脉冲凝胶电泳(PFGE)分子分型.结果 4株菌为羊种1型,2株菌为猪种3型,其余均为羊种3型,属特异性B基因阳性,菌株间的PFGE分型相似值介于67.9%~ 100.0%.除了来自珠海的4株菌为爆发,同源性为100.0%,其余菌株均为散发病例.结论 广东省布鲁杆菌病例存在高度散发和分散爆发的流行特征,与前几年比较呈现出种型的多样化,羊种3型仍是目前广东省布鲁杆菌流行株的优势菌型,PFGE方法可在种的水平区分布鲁杆菌的3个种,但不能有效区分同种异型.%Objective To analysis the etiology and molecular classification of brucella strains isolated in Guangdong province in 2010.Methods The strains of 19 brucella were verified and identified by some methods including traditional biology phenotype confirmation,PCR amplification and pulsed field gel electrophoresis (PFGE).Results On phenotype level,4 strains were brucella melitensis biovar 1,2 strains were brucella suis biovar 3,and the rest were brucella melitensis biovar 3,which were specific B genes positive strains,and the PFGE typing similar values ranging from 67.9% to 100%.In addition to the four strains from Zhuhai for the outbreak,the homology was 100%,and the rest were sporadic cases.Conclusions Brucella cases,in Guangdong province,are highly sporadic and dispersed outbreaks.Compared with a few years ago,it shows species diversification,and brucella melitensis biovar 3 is still the dominant serotype.PFGE can be used to distinguish the three species of brucella,but it can't effectively distinguish the allotypes.

  16. Pyruvate kinase is necessary for Brucella abortus full virulence in BALB/c mouse

    OpenAIRE

    2016-01-01

    International audience; AbstractBrucellosis, caused by a facultative intracellular pathogen Brucella, is one of the most prevalent zoonosis worldwide. Host infection relies on several uncanonical virulence factors. A recent research hotpot is the links between carbon metabolism and bacterial virulence. In this study, we found that a carbon metabolism-related pyruvate kinase (Pyk) encoded by pyk gene (locus tag BAB_RS24320) was associated with Brucella virulence. Determination of bacterial gro...

  17. Spontaneous excision of the O-polysaccharide wbkA glycosyltranferase gene is a cause of dissociation of smooth to rough Brucella colonies.

    Science.gov (United States)

    Mancilla, Marcos; Marín, Clara M; Blasco, José M; Zárraga, Ana María; López-Goñi, Ignacio; Moriyón, Ignacio

    2012-04-01

    The brucellae are Gram-negative pathogens that cause brucellosis, a zoonosis of worldwide importance. The genus Brucella includes smooth and rough species that differ in that they carry smooth and rough lipopolysaccharides, respectively. Brucella abortus, B. melitensis, and B. suis are typical smooth species. However, these smooth brucellae dissociate into rough mutants devoid of the lipopolysaccharide O-polysaccharide, a major antigen and a virulence determinant encoded in regions wbo (included in genomic island-2) and wbk. We demonstrate here the occurrence of spontaneous recombination events in those three Brucella species leading to the deletion of a 5.5-kb fragment carrying the wbkA glycosyltranferase gene and to the appearance of rough mutants. Analysis of the recombination intermediates suggested homologous recombination between the ISBm1 insertion sequences flanking wbkA as the mechanism generating the deletion. Excision of wbkA was reduced but not abrogated in a recA-deficient mutant, showing the existence of both RecA-dependent and -independent processes. Although the involvement of the ISBm1 copies flanking wbkA suggested a transpositional event, the predicted transpositional joint could not be detected. This absence of detectable transposition was consistent with the presence of polymorphism in the inverted repeats of one of the ISBm1 copies. The spontaneous excision of wbkA represents a novel dissociation mechanism of smooth brucellae that adds to the previously described excision of genomic island-2. This ISBm1-mediated wbkA excision and the different %GC levels of the excised fragment and of other wbk genes suggest that the Brucella wbk locus is the result of at least two horizontal acquisition events.

  18. Effectiveness of Rose Bengal test and fluorescence polarization assay in the diagnosis of Brucella spp. infections in free range cattle reared in endemic areas in Zambia.

    Science.gov (United States)

    Muma, J B; Lund, A; Nielsen, K; Matope, G; Munyeme, M; Mwacalimba, K; Skjerve, E

    2009-06-01

    The effectiveness of Rose Bengal test (RBT) and fluorescence polarization assay (FPA) in diagnosing cattle brucellosis in endemic areas was assessed and RBT and FPA test agreement was compared (n = 319). The sensitivity of RBT and FPA in detecting low Brucella titres were evaluated in paired sera (n = 34). A logistic regression model was constructed to predict cattle test result in FPA using RBT as the main predictor and incorporating bio-data and animal history. There was 79.3% agreement between the RBT and FPA (Kappa = 0.59; Std error = 0.05; p = 0.000) and a high correspondence between high RBT scores and positive FPA results suggesting that sera with high RBT score may not require confirmation with tests such as competitive-ELISA or CFT. High FPA cut-off points were more likely to miss animals with low antibody titres. The RBT had a reduced ability in detecting low antibody titres compared to the FPA. FPA test interpretation was improved if a priori information, such as sex and age was used. Under the challenging disease surveillance conditions prevailing in rural Africa, field-testing methods that are sensitive and specific; allow single animal contact, low technical skills in data interpretation are suitable.

  19. Investigation and genetic characteristic analysis of Brucella strain isolated from high-risk population in a human Brucellosis epidemic in Guizhou province%贵州省一起人间布鲁氏菌病疫情高危人群调查及分离菌遗传特征分析

    Institute of Scientific and Technical Information of China (English)

    王月; 王定明; 李世军; 陈红; 李沛丽; 刘英; 马青; 周敬祝; 余春; 黄艳; 唐光鹏

    2016-01-01

    Objective In order to provide scientific basis for Brucellosis control and prevention, sero⁃epidemiological survey, Brucella isolation and genetic characteristic analysis were conducted for the high⁃risk population in a human Brucellosis epidemic in Weining county of Guizhou province. Methods Tube agglutination test was used to detect the anti⁃Brucella antibody for the high⁃risk human population. Rose Bengal Plate Test was applied to detect the antibody in goat blood samples. The blood of antibody positive human population was collected for Brucella isolation. Conventional methods, genus specific BCSP31-PCR and species⁃specific AMOS⁃PCR were used to identify the bacteria strain, and the genetic characteristic was analyzed using MLVA and MLST techniques. Results Six people of 43 high⁃risk populations were confirmed as anti⁃Brucella antibody positive, 64 out of 302 goat blood samples were anti⁃Brucella antibody positive, with positive rate of 21.19%. A suspected Brucella strain were isolated from one of the high⁃risk human populations and wasidentified as B. melitensis biovar 3. MLVA-16 analysis indicated that the bacteria strain was most closely clustered with B. melitensis biovar 3. Multilocus sequence typing analysis indicated the strain was ST8. Conclusion Genetic characteristic of Brucella strain isolated from the Brucelosis epidemic was consistent with that of M. melitensis biovar 3. Although antibody and Brucella were detected, the high⁃risk populations did not displayed symptoms, so all of them were asymptomatic infections. The epidemic was seemingly imported due to goats trading, suggested that health and animal disease control and prevention departments and doctors should pay great attention to it.%目的:对贵州省威宁县一起人间布鲁氏菌病(布病)疫情中的高危人群进行血清学调查、菌株分离鉴定及遗传特征分析,为布病疫情的防控提供科学依据。方法采用试管

  20. 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...

  1. 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...

  2. Evidence of exposure to Brucella sp. in harbor porpoises (Phocoena phocoena) from the Bay of Fundy, Canada.

    Science.gov (United States)

    Neimanis, A S; Koopman, H N; Westgate, A J; Nielsen, K; Leighton, F A

    2008-04-01

    Novel strains of Brucella recently have been discovered in marine mammals. To investigate Brucella exposure and infection in a general population of cetaceans, blood and tissue samples were collected and analyzed from wild harbor porpoises (Phocoena phocoena) incidentally caught in fishing gear in the Bay of Fundy, Canada. Two of 170 (1.2%) animals had detectable antibodies against Brucella, but no organisms were isolated from genital swabs or tissues from 22 and 8 porpoises, respectively. Genetic analysis of inflamed testes from 20 animals yielded no amplification of Brucella DNA. This is the first evidence of exposure to Brucella in porpoises from the western North Atlantic, and the prevalence is much lower than documented for conspecifics from the eastern North Atlantic.

  3. Research progress of laboratory diagnostic techniques for Brucella%布鲁杆菌实验室诊断技术研究进展

    Institute of Scientific and Technical Information of China (English)

    周梅

    2012-01-01

    布鲁菌病是一种人畜共患传染病,生物种型较多,感染主范围广泛,传播途径多样.对人类致病的主要有羊、牛、猪、犬等4种,呈急性或慢性感染,临床表现为败血、发热性疾病或骨组织、器官系统的局部感染.布鲁菌病的诊断主要以流行病学史、临床表现以及实验检查为依据.因布鲁菌病的临床表现复杂,必须通过实验室分析来确证,诊断可直接进行血培养或通过间接免疫反应对微生物抗原进行检测.布鲁菌病临床表现不一,无症状感染形成抗体常是其特点,因此,当不能直接进行血培养时,血清学检查常发挥重要作用.布鲁杆菌常规细菌培养分离及鉴定耗时,操作杂,存在操作人员面临被布鲁杆菌感染及病菌从实验室扩散的风险.近年来出现的新型免疫学方法和聚合酶链反应(PCR)具有较高的敏感性和特异性.作者将布鲁杆菌实验室诊断检测技术进展做一综述.%Brucellosis is an important zoonosis. There are numerous species of Brucella, which can infect the hosts in various routes of transmission. Brucellosis in humans can be caused by any of the 4 main species (B. abortus, B. melitensis, B. suis and B. ca-nis). This is a disease presenting as acute or chronic infection in humans, and manifesting as a septicaemia febrile illness or localized infection of bone, tissue, or organ systems. Brucellosis diagnosis is primarily based on epidemiological history, clinical manifestations and laboratory examinations. Since the disease presents with a great variety of clinical manifestations the diagnosis must be confirmed by laboratory analyses. The diagnosis can be done directly by isolation of Brucellae from blood cultures, or indirectly by the detection of the immune response against the microbe's antigens. Asymptomatic infection is frequently characterized by antibody formation in people with no history of symptoms consistent with brucellosis. Hence, serological

  4. Establishment of loop-mediated isothermal amplification (LAMP) for rapid detection of Brucella spp. and application to milk and blood samples.

    Science.gov (United States)

    Song, Liuyan; Li, Juntao; Hou, Shuiping; Li, Xunde; Chen, Shouyi

    2012-09-01

    Brucella spp. are facultative intracellular bacteria that infect humans and animals. In this study, the loop-mediated isothermal amplification (LAMP) was used to detect the Brucella-specific gene omp25. Reaction conditions were optimized as temperature 65°C, reaction time 60 min, Mg(2+) concentration 8.0 mmol/L, polymerase content Bst DNA, 0.5 μL, deoxyribonucleotide concentration 1.6 mmol/L, and inner/outer primer ratio 1:8. The LAMP method was evaluated with 4 Brucella species and 29 non-Brucella bacteria species. Positive reactions were observed on all the 4 Brucella species but not on any non-Brucella species. The limit of detection of the LAMP method was 3.81 CFU Brucella spp. Using the LAMP method, 7 of 110 raw milk samples and 5 of 59 sheep blood samples were detected positive of Brucella spp. Results indicated that LAMP is a fast, specific, sensitive, inexpensive, and suitable method for diagnosis of Brucella spp. infection.

  5. Isolation and identification of Brucella in raw milk%牛乳样品中布鲁氏菌的分离和鉴定

    Institute of Scientific and Technical Information of China (English)

    易新萍; 马晓菁; 闫晶华; 谷文喜; 刘丽娅; 叶锋; 吐尔洪·努尔; 钟旗

    2013-01-01

    To investigate the prevalence of Brvcella in Xinjiang and improve the isolation method, serum samples collected from 3 dairy farms were tested by serum agglutination test (SAT). The results indicated that 3 dairy farms were Brucellosis-positive between 2010 and 2011. Twenty-five raw milk samples were collected from Brucellosis-positive cows based on SAT test and 9 Brucella isolates were isolated by using a modified Brucella selective supplement in the culture, which were confirmed by VirB8-PCR. In addition to the AMOS-PCR assays, the 9 isolates of Brucella were identified as B.abortus (7 isolates) and B.melitensis (2 isolates), which were further classified into B.abortus biovars 3 and B.melitensis biovars 3 by biochemical test. The results showed that the method of rapid isolation and identification of Brucella was useful for detection of Brucella.%为进一步改良布鲁氏菌(Brucella)的分离和鉴定方法,本研究于2010年~2012年对新疆部分奶牛场进行Brucella病的监测,采用改良Brucella选择添加剂培养分离的方法,从试管凝集试验(SAT)检测阳性的25头奶牛的牛乳样中分离到9株分离菌,经VirB8-PCR方法扩增Brucella的VirB8基因对分离株进行鉴定,结果表明9个分离株均为Brucella.同时采用Brucella分子分型PCR及生物学分型方法进一步对分离株鉴定,结果显示其中7个分离株为牛3型,另外2个分离株为羊3型.本研究为奶牛Brucella病的检疫与防控提供了快速分离及鉴定方法.

  6. 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.

  7. 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.

  8. 膜基因芯片技术鉴定布鲁杆菌的初步研究%A preliminary study of membrane gene chip assay in screening and species identification of Brucella

    Institute of Scientific and Technical Information of China (English)

    李铁锋; 李坚; 王照; 滕昊林; 王赢; 陈雪娇; 马文丽; 张宝; 桑建国

    2015-01-01

    目的 建立鉴定布鲁杆菌属及种的膜基因芯片技术方法.方法 试验菌株羊种标准株16M,牛种标准株544A,猪种标准株1330S;临床标本羊种81190,牛种80176、104M,猪种80177、19971、19972,共9个样本,均为中国疾病预防控制中心鼠布基地菌种库保存.提取各菌种DNA,进行PCR扩增,并以耶尔森菌O∶9为对照.设计布鲁杆菌鞭毛蛋白基因flhA保守片段中两个位点(379、576)的单个核苷酸多态性探针,在探针中引物入生物素,并将探针固定在膜上,制成膜芯片,与目的片段进行杂交,用生物素显色技术检测目的片段.结果 9个样本的扩增产物长度均为357 bp,与预期结果一致.与布鲁杆菌有相同抗原的耶尔森菌O∶9无特异性PCR产物产生.在鞭毛蛋白基因序列379和576两个核苷酸位点上,猪种为379G和576C,牛种为379T和576T,羊种为379T和576C,与所设计探针序列完全相同.经膜基因芯片实验验证,猪种菌株在379G、576C及F位点(公共探针)显色;牛种菌株在379T、576T及F位点显色;羊种菌株在379T、576C及F位点显色.结论 膜基因芯片可用于布鲁杆菌属和种的鉴定,且是一种优越的鉴定方法.%Objective To establish an accurate testing method for detection of Brucella and Brucella Abortus,Brucella Melitensis,Brucella suis.Methods The strains include the standard Brucella melitensis 16M,Brucella abortus 544A,the standard Brucella suis 1330s,and the clinical specimens Brucella melitensis 81190,Brucella abortus 80176,104M,Brucella suis 80177,19971,19972.All 9 strains were saved at the Base for Prevention and Control of Plague and Brucellosis,the Chinese Center for Disease Prevention and Control.DNA was extracted and used to amplify by PCR,Yersinia pestis O ∶ 9 served as control.The single nucleotide polymorphisms probes were designed on the basis of twosites (379,576) in conservative segment from Blucella flhA,and made into gene chip.According to part sequence of flh

  9. 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 skinfol

  10. 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 tr

  11. Construction and Identification of omp31-Deleted Mutant of Brucella Standard Strain 16M%布鲁氏菌16M△omp31基因缺失株的构建与鉴定

    Institute of Scientific and Technical Information of China (English)

    王慧; 张亚丽; 王远志; 陈创夫; 任晓丽

    2013-01-01

    To construct the omp31 deletion mutant of Brucella melitensis 16M,the upstream and downstream of the omp31 gene and SacB gene were amplified by PCR from Brucella melitensis 16M and Bacillus subtilis. After constructing omp31-SacB re-combinant fragment into plasmid 18-T simple vector, the suicide plasmid pGEM-7zf+-△omp31-SacB was further obtained and transformed into Brucella melitensis 16M by electroporation. The Aomp31 mutant strain was screened out by homologous recombination and its stability was detected by continuous bacteria culture. The results showed that Brucella melitensis 16M △omp31 mutant strain was successfully generated and reversion was not observed in 15 generations. This research lays a foundation of further study on the anti-apoptosis mechanism and construction of new types of vaccines of Brucella.%为了构建布鲁氏菌16M△omp31基因缺失株,采用PCR方法分别从亲本株16 M上扩增omp31基因的侧翼看序列及枯草芽孢杆菌SacB基因,并将所得片段与pMD18-T载体相连并测序,利用双酶切的方法分别将其连入自杀载体pGEM-7zf+,获得亚克隆pGEM-7zf+-△omp31-SacB.将所构建好的自杀载体通过电转化入布鲁氏菌16M感受态细胞中,经2次同源重组后筛选出16M△omp31基因缺失株,并对获得缺失株进行遗传稳定性检测.结果显示:成功获得布鲁氏菌16 M△omp31基因缺失株,该缺失株在15代内未发生回复性突变.本研究为今后研究布鲁氏菌抗凋亡机制奠定基础.

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

    OpenAIRE

    Francesco Casalinuovo; Lucia Ciambrone; Antonio Cacia; Paola Rippa

    2016-01-01

    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 ly...

  13. Multiple-locus variable-number tandem-repeat analysis genotyping of human Brucella isolates from Turkey.

    Science.gov (United States)

    Kiliç, Selçuk; Ivanov, Ivan N; Durmaz, Riza; Bayraktar, Mehmet Refik; Ayaslioglu, Ergin; Uyanik, M Hamidullah; Aliskan, Hikmet; Yasar, Ekrem; Bayramoglu, Gülçin; Arslantürk, Ahmet; Vergnaud, Gilles; Kantardjiev, Todor V

    2011-09-01

    A multiple-locus variable-number tandem-repeat analysis (MLVA) was applied to investigate the epidemiological relationship and genetic diversity among 162 human Brucella isolates collected from all geographic regions of Turkey in an 8-year period (2001 to 2008). The isolates were genotyped by using an MLVA assay developed in Orsay, France (MLVA-16(Orsay)) including eight minisatellite (panel 1) and eight microsatellite (panel 2, subdivided into 2A and 2B) markers. Panels 1 and 2A distinguish 14 genotypes; two of these represented 85% of the strains. Panel 2B displayed a very high discriminatory power. Three loci from panel 2B had diversity index values higher than 0.74. MLVA-16(Orsay) yielded 105 genotypes; 73 were represented by a unique isolate, and 32 included two to eight isolates. The isolates from different patients within the same outbreak or from the same patient before first-line therapy and after relapse showed identical genotypes. A number of MLVA genotypes appeared to be partially restricted to some geographic areas and displayed no annual variation, possibly reflecting persistence of genotypes in certain areas for a time span of at least a decade. This study, representing the first molecular typing results of human Brucella isolates from Turkey, indicated that Turkish human Brucella melitensis isolates were most closely related to the neighboring countries' isolates included in the East Mediterranean group.

  14. Multiple-Locus Variable-Number Tandem-Repeat Analysis Genotyping of Human Brucella Isolates from Turkey▿†

    Science.gov (United States)

    Kılıç, Selçuk; Ivanov, Ivan N.; Durmaz, Rıza; Bayraktar, Mehmet Refik; Ayaşlıoğlu, Ergin; Uyanık, M. Hamidullah; Alışkan, Hikmet; Yaşar, Ekrem; Bayramoğlu, Gülçin; Arslantürk, Ahmet; Vergnaud, Gilles; Kantardjiev, Todor V.

    2011-01-01

    A multiple-locus variable-number tandem-repeat analysis (MLVA) was applied to investigate the epidemiological relationship and genetic diversity among 162 human Brucella isolates collected from all geographic regions of Turkey in an 8-year period (2001 to 2008). The isolates were genotyped by using an MLVA assay developed in Orsay, France (MLVA-16Orsay) including eight minisatellite (panel 1) and eight microsatellite (panel 2, subdivided into 2A and 2B) markers. Panels 1 and 2A distinguish 14 genotypes; two of these represented 85% of the strains. Panel 2B displayed a very high discriminatory power. Three loci from panel 2B had diversity index values higher than 0.74. MLVA-16Orsay yielded 105 genotypes; 73 were represented by a unique isolate, and 32 included two to eight isolates. The isolates from different patients within the same outbreak or from the same patient before first-line therapy and after relapse showed identical genotypes. A number of MLVA genotypes appeared to be partially restricted to some geographic areas and displayed no annual variation, possibly reflecting persistence of genotypes in certain areas for a time span of at least a decade. This study, representing the first molecular typing results of human Brucella isolates from Turkey, indicated that Turkish human Brucella melitensis isolates were most closely related to the neighboring countries' isolates included in the East Mediterranean group. PMID:21795514

  15. A case of unusual septic knee arthritis with Brucella abortus after arthroscopic meniscus surgery.

    Science.gov (United States)

    Lee, Keun Hwa; Kang, Hyunseong; Kim, Taejung; Choi, Sungwook

    2016-01-01

    We present a 51-year-old male patient with Brucella abortus septic arthritis in the right knee following arthroscopic meniscus surgery. He had eaten a traditional dish of raw minced cattle conceptus (bovine fetus) that was prepared after the cow was slaughtered. Despite treatment with empirical antibiotics and debridement of the postoperative surgical wound, the infection persisted without improvement. Polymerase chain reaction sequencing identified Brucella abortus from tissue samples obtained from the patient. After confirmation of the diagnosis of brucellar infection, antibiotics were replaced with doxycycline and rifampin, which were used for 4 months. In patients with a non-specific arthralgia who eat raw meat or live close to animals, it is important to consider the possibility of septic arthritis due to infection with Brucella spp.

  16. Biotype identification and epidemiological analysis of twenty-four Brucella strains%24株布鲁菌的种型鉴定和流行病学分析

    Institute of Scientific and Technical Information of China (English)

    张磊; 屈平华; 吴尚为; 陈经雕

    2016-01-01

    Objective To identify the biotype and analyze the epidemiological characteristics of twentyfour Brurella strains from the primary hospitals in Guangdong Province.Methods The twenty-four Brucella strains,collected from Oct.2009 to Oct.2015,were identified by routine biochemical methods,VITEK 2 COMPACT automatic microbial identification analyzer,16S rRNA gene sequencing and biology phenotype based on serological and bacteriophages lysis test.The etiology was analyzed based on clinical data,biotypes of the isolates and other clinical information.Results All of the twenty-four strains were Gram-negative coccobacilli,including two strains of Brucella suis biotype Ⅱ,four strains of Brucella melitensis biotype Ⅰ and eighteen strains of Brucella melitensis biotype Ⅲ.By GN card of VITEK 2 COMPACT automatic microbial identification analyzer,one strain was mistaken as Bordetella bronchiseptica and two strains were mistaken as Ochrobaetrum anthropi.The 16S rRNA gene sequencing showed they were high homology to Ochrobactrum intermedium and Ochrobaetrum anthropi,which completely excluded the possibility of Bordetella bronchiseptica.Tbe clinical data showed that all of the twenty-four patients were adults with an average age of 49.0 years old,men and women were twelve people respectively,with no significant gender differences and no occupational exposure,which presenting a wide and diverse range of nonspecific clinical signs and symptoms,but brucellosis was not aware of by the physician.Conclusion Brucella melitensis biotype Ⅲ is the main pathogen of brucellosis,with the characteristics of sporadic outbreak and occult infection in the primary hospitals in Guangdong Province.%目的 对广东省基层地区24株布鲁菌进行种型鉴定和流行病学分析.方法 对24株来自于2009年10月至2015年10月广东省基层医院血培养阳性的布鲁菌进行传统生化鉴定、VITEK 2仪器鉴定、16S rRNA基因测序、血清学试验和噬菌体试验,比较不

  17. Brucella suis-Impaired Specific Recognition of Phagosomes by Lysosomes due to Phagosomal Membrane Modifications

    Science.gov (United States)

    Naroeni, Aroem; Jouy, Nicolas; Ouahrani-Bettache, Safia; Liautard, Jean-Pierre; Porte, Françoise

    2001-01-01

    Brucella species are gram-negative, facultatively intracellular bacteria that infect humans and animals. These organisms can survive and replicate within a membrane-bound compartment in phagocytic and nonprofessional phagocytic cells. Inhibition of phagosome-lysosome fusion has been proposed as a mechanism for intracellular survival in both types of cells. However, the biochemical mechanisms and microbial factors implicated in Brucella maturation are still completely unknown. We developed two different approaches in an attempt to gain further insight into these mechanisms: (i) a fluorescence microscopy analysis of general intracellular trafficking on whole cells in the presence of Brucella and (ii) a flow cytometry analysis of in vitro reconstitution assays showing the interaction between Brucella suis-containing phagosomes and lysosomes. The fluorescence microscopy results revealed that fusion properties of latex bead-containing phagosomes with lysosomes were not modified in the presence of live Brucella suis in the cells. We concluded that fusion inhibition was restricted to the pathogen phagosome and that the host cell fusion machinery was not altered by the presence of live Brucella in the cell. By in vitro reconstitution experiments, we observed a specific association between killed B. suis-containing phagosomes and lysosomes, which was dependent on exogenously supplied cytosol, energy, and temperature. This association was observed with killed bacteria but not with live bacteria. Hence, this specific recognition inhibition seemed to be restricted to the pathogen phagosomal membrane, as noted in the in vivo experiments. PMID:11119541

  18. Regulatory effect of Astragalus Polysaccharides on immune function of mice infected by Brucella suis S2%黄芪多糖对布鲁菌S2株感染小鼠免疫功能的调节作用

    Institute of Scientific and Technical Information of China (English)

    王晓曼; 陈志磊; 王少毅; 许振国; 杨瑞强; 张树全; 曹志然

    2014-01-01

    To study the regulating effect of Astragalus Polysaccharides ( APS) to the mice infected by Brucella suis S2.Methods:120 BALB/c mice were randomly divided into 4 groups:experimental mice were injected APS 1 ml ( 0.4,1.2,3 mg/ml) via peritoneal cavity respectively once a day and the control group was injected with the same volume of saline for 3 days,then infected with Brucella suis S2 1 ml (1×107 L-1 ) by ip.Five mice of each group were killed through eye bloodletting at 1,6,12,24,48, 72 h respectively post-infection with Brucella suis S 2 and the peritoneal macrophage were obtained respectively to make smear.Phagocytic rate and phagocytic index were calculated by the Wright Giemsa staining after infected 1 h.TNF-α,IL-12 and IFN-γlevels of serum at different time points were measured by ELISA.The bacterial load of MΦand spleen were measured by coating method.Results:The phagocytic rate and phagocytic index of MΦin APS 3 dose groups were higher than those of the control group ( P<0.05 ).The microbial load of MΦin APS 3 dose groups at 1 h infected by Brucella suis S 2 were significantly higher than those of control,but significantly lower than those of control at 6,12,24,48,72 h after infected by Brucella suis S2.The microbial load of spleen in APS 3 dose groups at 6 h infected by Brucella suis S 2 were significantly higher than those of control ,but significantly lower than those of control at 12,24,48,72h after infected by Brucella suis S2.The concentrations of TNF-α,IL-12 and IFN-γin the serum of APS groups had significantly been improved ( P<0.05 ).Conclusion: APS can promote the activation of MΦin vivo and strengthen the activity of phagocytosis and killing to Brucella suis S 2.APS can promote the secretion of TNF-α,IL-12 and IFN-γof mice,strengthen the cellular immune response of mice to Brucella suis S 2.%目的:研究黄芪多糖( APS)对猪布鲁菌S2株感染小鼠免疫功能的调节作用。方法:120只BALB/c小鼠随机分为4组,

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

    Directory of Open Access Journals (Sweden)

    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

  20. Recent advances in Brucella abortus vaccines.

    Science.gov (United States)

    Dorneles, Elaine M S; Sriranganathan, Nammalwar; Lage, Andrey P

    2015-07-08

    Brucella abortus vaccines play a central role in bovine brucellosis control/eradication programs and have been successfully used worldwide for decades. Strain 19 and RB51 are the approved B. abortus vaccines strains most commonly used to protect cattle against infection and abortion. However, due to some drawbacks shown by these vaccines much effort has been undertaken for the development of new vaccines, safer and more effective, that could also be used in other susceptible species of animals. In this paper, we present a review of the main aspects of the vaccines that have been used in the brucellosis control over the years and the current research advances in the development of new B. abortus vaccines.

  1. Evaluación de un modelo de infección murino para la evaluación de vacunas homólogas y heterólogas contra Brucella canis Evaluation of a mouse infection model to evaluate homologue and heterologue vaccines against Brucella canis in mouse model

    Directory of Open Access Journals (Sweden)

    M Clausse

    2011-12-01

    Full Text Available La brucelosis canina es una enfermedad zoonótica que afecta a los caninos y provoca importantes pérdidas económicas en criaderos. En la actualidad, no existe una vacuna para la prevención de esta enfermedad. Una limitante en el desarrollo de una vacuna eficaz es la falta de un modelo experimental que permita evaluar la infección con Brucella canis. En este trabajo, se desarrolló un modelo ratón para evaluar la infección esplénica por B. canis empleando distintas dosis, tiempos de sacrificio y vías de inoculación. Además, se ensayaron inmunógenos homólogos (B. canis inactivada y heterólogos (B. ovis inactivada y viva como potenciales vacunas control. Las bacterinas fueron formuladas en Marcol 52, Montanide o Quil A. La inoculación de 106-107 UFC (unidades formadoras de colonias de B. canis por las vías intraperitoneal o intravenosa no mostró diferencias en la carga bacteriana esplénica. El recuento de UFC en bazo fue similar cuando los ratones se sacrificaron a 14, 21 o 30 días post-inoculación. B. ovis PA76250 viva confirió el mejor nivel de protección contra B. canis mientras que una protección menor, pero significativa, fue obtenida cuando se administró la bacterina B. canis M- formulada en Marcol 52 o Montanide. Los resultados obtenidos indican que el modelo ratón es adecuado para evaluar, en el futuro, la eficacia de potenciales inmunógenos contra la brucelosis canina.The development of an effective vaccine against brucellosis is a research area of much interest since there is no available vaccine against canine brucellosis. One limitation in testing any immunization strategy is the lack of a suitable laboratory animal that is permissive to the infection. In this work, a mouse model was developed to study splenic B. canis RM6/66 infection using different doses, times of sacrifice and inoculation routes. In addition, homologous (B. canis M- bacterin and heterologous (live or B. ovis bacterin rough whole cell

  2. Characterization and evaluation of an arbitrary primed Polymerase Chain Reaction (PCR) product for the specific detection of Brucella species.

    Science.gov (United States)

    Qasem, Jafar A; AlMomin, Sabah; Al-Mouqati, Salwa A; Kumar, Vinod

    2015-03-01

    Laboratory detection of Brucella is based largely on bacterial isolation and phenotypic characterization. These methods are lengthy and labor-intensive and have been associated with a heightened risk of laboratory-acquired infection. Antibody based indirect detection methods also suffer from limitations in proper diagnosis of the organism. To overcome these problems, nucleic acid amplification has been explored for rapid detection and confirmation of the presence of Brucella spp. PCR-based diagnostics is useful for screening large populations of livestock to identify infected individuals and confirms the presence of the pathogen. Random Amplification of Polymorphic DNA (RAPD) was performed and identified a 1.3 kb PCR fragment specifically amplifiable from DNA isolated from Brucella. A BLAST search revealed no significant homology with the reported sequences from species other than the members of Brucella. The isolated fragment seems to be a part of d-alanine-d-alanine ligase gene in Brucella sp. Translational BLAST revealed certain degree of homology of this sequence with orthologs of this gene reported from other microbial species at the deduced amino acid level. The sequence information was used to develop PCR based assays to detect Brucella sp. from various samples. The minimum detection limit of Brucella from blood and milk samples spiked with Brucella DNA was found to be 1 ng/ml and 10 ng/ml, respectively. In conclusion, we demonstrated that the PCR based detection protocol was successfully used for the detection of Brucella from various organs and spiked samples of diseased sheep. Diagnosis of Brucellosis by PCR based method reported in this study is relatively rapid, specific and simple.

  3. Aislamiento de Brucella canis en un humano conviviente con caninos infectados. Informe de un caso.

    Directory of Open Access Journals (Sweden)

    Martha Olivera

    2009-12-01

    Full Text Available Objective: To report a case of bacteremia in a woman living in close contact with dogs infected with Brucella canis. Methods: Previous written consent of the woman a blood sample was taken to perform rapid agglutination test, immunofluorescence, and hemoculture. Results: Brucella canis was isolated and the isolation was confirmed by the Instituto Nacional ANLIS «Carlos G. Malbran» from Argentina. The woman did not present any symptoms associated with brucellosis. Conclusion: There is a risk of contamination by B. canis for people keeping tight contact with infected dogs.

  4. ELISA for brucellosis detection based on three Brucella recombinant proteins.

    Science.gov (United States)

    Thepsuriyanont, Pikun; Intarapuk, Apiradee; Chanket, Panita; Tunyong, Wittawat; Kalambaheti, Thareerat

    2014-01-01

    Control of brucellosis among farm animals, wildlife and humans require reliable diagnosis. Rose Bengal serological test (RBT) is based on lipopolysaccharide antigen of Brucella, which may cross react with other gram-negative bacteria and produce false positive result. Immunoreactive proteins, such as outer-membrane protein BP26, ribosome recycling factor protein CP24 and Brucella lumazine synthase (BLS), previously reported to be recognized by infected sheep sera, were selected for production of recombinant proteins for use in an ELISA in order to investigate immune response among goats and cows, in comparison with commercial RBT. Cut-off value for ELISA was based on the immune response of in vitro fertilized goats and cows. Goats positive for Brucella culture or by RBT were ELISA positive for either IgG or IgM against at least one recombinant protein. For animals with negative RBT, animals with positive ELISA could be detected, and 61.6% possessed ELISA values as high as in infected animals. Thus, this ELISA procedure is proposed as an alternative to RBT for screening of brucellosis in farm animals.

  5. 3 misidentification cases of Brucella species by automated microbial identification system and literature review*%细菌自动鉴定仪3例布鲁氏菌误鉴定及文献复习

    Institute of Scientific and Technical Information of China (English)

    杨婧; 任晓庆; 褚美玲; 马均; 任微; 王璐; 孟冬娅

    2012-01-01

    have obvious limitations in identifying slowly growing bacteria such as Brucella melitensis. Misidentification could mislead clinical therapy and epidemiological investigation of such infectious disease as Brucellosis. It could be suggested that the microbial identification system manufacturers improve the expert system and notice to discriminate with Brucella species when the above-mentioned strains were identified so as to effectively prevent lab-acquired infection caused by Brucella species. Articles lacking molecular biological diagnostic methods to i-dentify the above-mentioned rare pathogens should be reported deliberately.

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

    Directory of Open Access Journals (Sweden)

    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.

  7. Pesquisa de anticorpos anti-Brucella abortus e anti-Brucella ovis em ovinos no município de Uberlândia, MG

    Directory of Open Access Journals (Sweden)

    S.R.S. Salaberry

    2011-08-01

    Full Text Available The first epidemiologic inquiry to Brucella abortus (B. abortus and Brucella ovis (B. ovis was carried out in sheep from Uberlândia county, MG. A total of 334 blood serum samples of sheep from both sexes and different ages and breeds were collected in 12 farms. An epidemiologic questionnaire was applied for each farm. Tests for B. abortus and B. ovis antibodies were Buffered Acidified Antigen and Complement Fixation, respectively. None of the sheep was reactive to B. abortus and B. ovis; however, the adoption of sanitary measures is important to avoid the introduction of infections caused by these bacteria.

  8. Evaluation of DNA extraction protocols for Brucella abortus pcr detection in aborted fetuses or calves born from cows experimentally infected with strain 2308 Avaliação de diferentes protocolos de extração de DNA para detecção de Brucella abortus a partir de abortos ou de bezerros nascidos de vacas experimentalmente infectadas com estirpe 2308

    Directory of Open Access Journals (Sweden)

    M. Matrone

    2009-09-01

    Full Text Available The objective of the present study was to improve the detection of B. abortus by PCR in organs of aborted fetuses from infected cows, an important mechanism to find infected herds on the eradication phase of the program. So, different DNA extraction protocols were compared, focusing the PCR detection of B. abortus in clinical samples collected from aborted fetuses or calves born from cows challenged with the 2308 B. abortus strain. Therefore, two gold standard groups were built based on classical bacteriology, formed from: 32 lungs (17 positives, 26 spleens (11 positives, 23 livers (8 positives and 22 bronchial lymph nodes (7 positives. All samples were submitted to three DNA extraction protocols, followed by the same amplification process with the primers B4 and B5. From the accumulated results for organ, the proportion of positives for the lungs was higher than the livers (p=0.04 or bronchial lymph nodes (p=0.004 and equal to the spleens (p=0.18. From the accumulated results for DNA extraction protocol, the proportion of positives for the Boom protocol was bigger than the PK (pO objetivo do presente estudo foi aperfeiçoar a detecção de Brucella abortus pela PCR em homogeneizados de órgãos de fetos abortados por vacas infectadas, importante mecanismo para descobrir focos da doença na fase de erradicação. Assim, foram comparados diferentes protocolos de extração de DNA, visando à detecção de B. abortus pela PCR em amostras clínicas colhidas de abortos ou de bezerros oriundos de vacas desafiadas com a estirpe 2308 de B. abortus. Para tanto, foram construídos dois grupos padrão ouro com base na bacteriologia clássica, constituídos por: 32 pulmões (17 positivos, 26 baços (11 positivos, 23 fígados (8 positivos e 22 linfonodos bronquiais (7 positivos. Todas essas amostras foram submetidas a três protocolos de extração de DNA, seguidos do mesmo processo de amplificação com os primers B4 e B5. Nos resultados acumulados por

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

    Directory of Open Access Journals (Sweden)

    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.

  10. Dextran sulfate sodium upregulates MAPK signaling for the uptake and subsequent intracellular survival of Brucella abortus in murine macrophages.

    Science.gov (United States)

    Reyes, Alisha Wehdnesday Bernardo; Arayan, Lauren Togonon; Simborio, Hannah Leah Tadeja; Hop, Huynh Tan; Min, WonGi; Lee, Hu Jang; Kim, Dong Hee; Chang, Hong Hee; Kim, Suk

    2016-02-01

    Brucellosis is one of the major zoonoses worldwide that inflicts important health problems in animal and human. Here, we demonstrated that dextran sulfate sodium (DSS) significantly increased adhesion of Brucella (B.) abortus in murine macrophages compared to untreated cells. Even without infection, Brucella uptake into macrophages increased and F-actin reorganization was induced compared with untreated cells. Furthermore, DSS increased the phosphorylation of MAPKs (ERK1/2 and p38α) in Brucella-infected, DSS-treated cells compared with the control cells. Lastly, DSS markedly increased the intracellular survival of Brucella abortus in macrophages by up to 48 h. These results suggest that DSS enhanced the adhesion and phagocytosis of B. abortus into murine macrophages by stimulating the MAPK signaling proteins phospho-ERK1/2 and p38α and that DSS increased the intracellular survival of B. abortus by inhibiting colocalization of Brucella-containing vacuoles (BCVs) with the late endosome marker LAMP-1. This study emphasizes the enhancement of the phagocytic and intracellular modulatory effects of DSS, which may suppress the innate immune system and contribute to prolonged Brucella survival and chronic infection.

  11. Evaluation of DNA extraction protocols for Brucella abortus pcr detection in aborted fetuses or calves born from cows experimentally infected with strain 2308.

    Science.gov (United States)

    Matrone, M; Keid, L B; Rocha, V C M; Vejarano, M P; Ikuta, C Y; Rodriguez, C A R; Ferreira, F; Dias, R A; Ferreira Neto, J S

    2009-07-01

    The objective of the present study was to improve the detection of B. abortus by PCR in organs of aborted fetuses from infected cows, an important mechanism to find infected herds on the eradication phase of the program. So, different DNA extraction protocols were compared, focusing the PCR detection of B. abortus in clinical samples collected from aborted fetuses or calves born from cows challenged with the 2308 B. abortus strain. Therefore, two gold standard groups were built based on classical bacteriology, formed from: 32 lungs (17 positives), 26 spleens (11 positives), 23 livers (8 positives) and 22 bronchial lymph nodes (7 positives). All samples were submitted to three DNA extraction protocols, followed by the same amplification process with the primers B4 and B5. From the accumulated results for organ, the proportion of positives for the lungs was higher than the livers (p=0.04) or bronchial lymph nodes (p=0.004) and equal to the spleens (p=0.18). From the accumulated results for DNA extraction protocol, the proportion of positives for the Boom protocol was bigger than the PK (pprotocols (p=0.5). Some positive samples from the classical bacteriology were negative to the PCR and vice-versa. Therefore, the best strategy for B. abortus detection in the organs of aborted fetuses or calves born from infected cows is the use, in parallel, of isolation by classical bacteriology and the PCR, with the DNA extraction performed by the Boom protocol.

  12. N-Formyl-Perosamine Surface Homopolysaccharides Hinder the Recognition of Brucella abortus by Mouse Neutrophils.

    Science.gov (United States)

    Mora-Cartín, Ricardo; Chacón-Díaz, Carlos; Gutiérrez-Jiménez, Cristina; Gurdián-Murillo, Stephany; Lomonte, Bruno; Chaves-Olarte, Esteban; Barquero-Calvo, Elías; Moreno, Edgardo

    2016-06-01

    Brucella abortus is an intracellular pathogen of monocytes, macrophages, dendritic cells, and placental trophoblasts. This bacterium causes a chronic disease in bovines and in humans. In these hosts, the bacterium also invades neutrophils; however, it fails to replicate and just resists the killing action of these leukocytes without inducing significant activation or neutrophilia. Moreover, B. abortus causes the premature cell death of human neutrophils. In the murine model, the bacterium is found within macrophages and dendritic cells at early times of infection but seldom in neutrophils. Based on this observation, we explored the interaction of mouse neutrophils with B. abortus In contrast to human, dog, and bovine neutrophils, naive mouse neutrophils fail to recognize smooth B. abortus bacteria at early stages of infection. Murine normal serum components do not opsonize smooth Brucella strains, and neutrophil phagocytosis is achieved only after the appearance of antibodies. Alternatively, mouse normal serum is capable of opsonizing rough Brucella mutants. Despite this, neutrophils still fail to kill Brucella, and the bacterium induces cell death of murine leukocytes. In addition, mouse serum does not opsonize Yersinia enterocolitica O:9, a bacterium displaying the same surface polysaccharide antigen as smooth B. abortus Therefore, the lack of murine serum opsonization and absence of murine neutrophil recognition are specific, and the molecules responsible for the Brucella camouflage are N-formyl-perosamine surface homopolysaccharides. Although the mouse is a valuable model for understanding the immunobiology of brucellosis, direct extrapolation from one animal system to another has to be undertaken with caution.

  13. 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...

  14. Identification and sequence analysis of IS6501, an insertion sequence in Brucella spp.: relationship between genomic structure and the number of IS6501 copies.

    Science.gov (United States)

    Ouahrani, S; Michaux, S; Sri Widada, J; Bourg, G; Tournebize, R; Ramuz, M; Liautard, J P

    1993-12-01

    An insertion sequence (IS) element of Brucella ovis, named IS6501, was isolated and its complete nucleotide sequence determined. IS6501 is 836 bp in length and occurs 20-35 times in the B. ovis genome and 5-15 times in other Brucella species. Analysis of the junctions at the sites of insertion revealed a small target site duplication of four bases and inverted repeats of 17 bp with one mismatch. IS6501 presents significant similarity (53.4%) with IS427 identified in Agrobacterium tumefaciens, suggesting a common ancestral sequence. A long ORF of 708 bp was identified encoding a protein with a predicted molecular mass of 26 kDa and sharing sequence identity with the hypothetical protein 1 of A. tumefaciens and with the transposase of Mycobacterium tuberculosis. IS6501 is present in all Brucella strains we have tested. Restriction fragment length polymorphism of reference and field strains of two species (B. melitensis and B. ovis) was studied using either pulsed field gel electrophoresis (PFGE) on XbaI-digested DNA or hybridization of EcoRI-digested DNA using IS6501 as a probe. The genome of B. melitensis biovar 3 contains about 10 IS copies per genome and field strains of the same species could not be distinguished either by IS hybridization or by XbaI (PFGE) restriction patterns. In contrast, the number of IS copies in the B. ovis genome is around 30 and the different field strains can be differentiated by both methods.(ABSTRACT TRUNCATED AT 250 WORDS)

  15. Molecular characterisation of Brucella species.

    Science.gov (United States)

    Scholz, H C; Vergnaud, G

    2013-04-01

    The genus Brucella (Mayer and Shaw, 1920) currently consists often species with validly published names. Within most species further differentiation into biovars exists. Genetically, all Brucella species are highly related to each other, exhibiting sequence similarity values of 98% to 100% in aligned regions (core genome). The population structure is clonal. Despite this close genetic relatedness, the various species can be clearly distinguished from each other by application of high-resolution molecular typing tools, in addition to assessment of phenotype and host preference. Accurate species delineation can be achieved by conventional multiplex polymerase chain reaction (PCR), single nucleotide polymorphism (SNP) analysis and multilocus sequence typing (MLST) or multilocus sequence analysis (MLSA). The last is also suitable for phylogenetic reconstructions, owing to the highly clonal evolution of the different species. Highly discriminatory multilocus variable number of tandem repeats (VNTR) analysis (MLVA) allows both species delineation and differentiation of individual isolates and thus represents a perfect first-line toolfor molecular epidemiological studies within outbreak investigations. More recently,whole genome sequencing (WGS)and the resulting global genome-wide SNP analysis have become available. These novel approaches should help in further understanding the evolution, host specificity and pathogenicity of the genus Brucella.

  16. Intraspecies biodiversity of the genetically homologous species Brucella microti.

    Science.gov (United States)

    Al Dahouk, Sascha; Hofer, Erwin; Tomaso, Herbert; Vergnaud, Gilles; Le Flèche, Philippe; Cloeckaert, Axel; Koylass, Mark S; Whatmore, Adrian M; Nöckler, Karsten; Scholz, Holger C

    2012-03-01

    Brucellosis is one of the major bacterial zoonoses worldwide. In the past decade, an increasing number of atypical Brucella strains and species have been described. Brucella microti in particular has attracted attention, because this species not only infects mammalian hosts but also persists in soil. An environmental reservoir may pose a new public health risk, leading to the reemergence of brucellosis. In a polyphasic approach, comprising conventional microbiological techniques and extensive biochemical and molecular techniques, all currently available Brucella microti strains were characterized. While differing in their natural habitats and host preferences, B. microti isolates were found to possess identical 16S rRNA, recA, omp2a, and omp2b gene sequences and identical multilocus sequence analysis (MLSA) profiles at 21 different genomic loci. Only highly variable microsatellite markers of multiple-locus variable-number tandem repeat (VNTR) analysis comprising 16 loci (MLVA-16) showed intraspecies discriminatory power. In contrast, biotyping demonstrated striking differences within the genetically homologous species. The majority of the mammalian isolates agglutinated only with monospecific anti-M serum, whereas soil isolates agglutinated with anti-A, anti-M, and anti-R sera. Bacteria isolated from animal sources were lysed by phages F1, F25, Tb, BK2, Iz, and Wb, whereas soil isolates usually were not. Rough strains of environmental origin were lysed only by phage R/C. B. microti exhibited high metabolic activities similar to those of closely related soil organisms, such as Ochrobactrum spp. Each strain was tested with 93 different substrates and showed an individual metabolic profile. In summary, the adaptation of Brucella microti to a specific habitat or host seems to be a matter of gene regulation rather than a matter of gene configuration.

  17. Public health consequences of a false-positive laboratory test result for Brucella--Florida, Georgia, and Michigan, 2005.

    Science.gov (United States)

    2008-06-01

    Human brucellosis, a nationally notifiable disease, is uncommon in the United States. Most human cases have occurred in returned travelers or immigrants from regions where brucellosis is endemic, or were acquired domestically from eating illegally imported, unpasteurized fresh cheeses. In January 2005, a woman aged 35 years who lived in Nassau County, Florida, received a diagnosis of brucellosis, based on results of a Brucella immunoglobulin M (IgM) enzyme immunoassay (EIA) performed in a commercial laboratory using analyte specific reagents (ASRs); this diagnosis prompted an investigation of dairy products in two other states. Subsequent confirmatory antibody testing by Brucella microagglutination test (BMAT) performed at CDC on the patient's serum was negative. The case did not meet the CDC/Council of State and Territorial Epidemiologists' (CSTE) definition for a probable or confirmed brucellosis case, and the initial EIA result was determined to be a false positive. This report summarizes the case history, laboratory findings, and public health investigations. CDC recommends that Brucella serology testing only be performed using tests cleared or approved by the Food and Drug Administration (FDA) or validated under the Clinical Laboratory Improvement Amendments (CLIA) and shown to reliably detect the presence of Brucella infection. Results from these tests should be considered supportive evidence for recent infection only and interpreted in the context of a clinically compatible illness and exposure history. EIA is not considered a confirmatory Brucella antibody test; positive screening test results should be confirmed by Brucella-specific agglutination (i.e., BMAT or standard tube agglutination test) methods.

  18. 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

    OpenAIRE

    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...

  19. 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 abortus strain 2308. The incidences of abortion and infection were 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.

  20. 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.

  1. 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.

  2. 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.

  3. A Familial Cluster of Human Brucellosis Attributable to Contact with Imported Infected Goats in Shuyang, Jiangsu Province, China, 2013.

    Science.gov (United States)

    Tan, Zhongming; Huang, Yong; Liu, Genyan; Zhou, Weizhong; Xu, Xilou; Zhang, Zibing; Shen, Qing; Tang, Fenyang; Zhu, Yefei

    2015-10-01

    Brucellosis remains a serious public health issue in developing countries, including China. On August 8, 2013, four cases of brucellosis from one extended family were reported at Shuyang County, Jiangsu Province, China. Active case finding was performed to identify the source and the risk factors of the infection and to prevent additional cases. Multiple-locus variable number tandem repeat analysis (MLVA) was used for molecular subtyping analysis. Six people from two extended families met the case definition for brucellosis infection; four were blood culture positive for Brucella melitensis biotype 3. Four additional family members were found seropositive by using a serological test. Isolates from the four patients were indistinguishable by MLVA profiling, displaying a unique type for Jiangsu Province. Field epidemiological data combined with MLVA genotyping supported a common source of the isolates from the different patients. We recommend stronger reinforcement measures for animal quarantine practices, enhanced cooperation with veterinary service organizations, and implementation of measures that strengthen public education on brucellosis to prevent further human outbreaks in Jiangsu Province.

  4. Sequencing of an atypical Brucella strain 16S rDNA%1株非典型布鲁杆菌的16S rDNA序列分析

    Institute of Scientific and Technical Information of China (English)

    刘志国; 王妙; 刘日宏; 崔步云

    2015-01-01

    目的 分析非典型布鲁杆菌的16S rDNA序列,评价16S rDNA序列测定方法鉴定布鲁杆菌的可行性.方法 用常规方法对l株非典型菌株进行初步鉴定;提取菌株的DNA,双向测定16S rDNA全序列,在NCBI对该序列进行Blast比对,用DNAMAN软件进行菌株的16S rDNA序列一致性比对;同时,从GenBank下载与布鲁杆菌有血清学交叉反应菌株的16S rDNA全序列,用MEGA 6.0构建16S rDNA系统发生树.结果 常规鉴定显示,试验菌株为非典型布鲁杆菌.试验菌株的16S rDNA序列与布鲁杆菌基因的相似性为99%,与牛种布鲁杆菌544A、羊种布鲁杆菌16M的16S rDNA序列的一致性为96.99%.系统发生树显示,试验菌株为布鲁杆菌,且与牛种布鲁杆菌544A、羊种布鲁杆菌16M有较为密切的亲缘关系.结论 试验菌株为非典型布鲁杆菌,16S rDNA序列测定是一种简便、快速、准确的非典型布鲁杆菌鉴定方法.%Objective To sequence an atypical Brucella strain 16S rDNA,and to evaluate the feasibility of 16S rDNA sequencing method for identification of Brucella.Methods Preliminary identification of atypical strains was carried out with conventional method.Strain DNA was extracted,and 16S rDNA complete sequence was bidirectional sequenced,and Blast in NCBI and DNAMAN software were used for comparison of the sequence identities of the 16S rDNA.Moreover,16S rDNA complete sequence of the stains those were known to cross-react serologically with Brucella was downloaded from GenBank,MEGA 6.0 was used to construct the phylogenetic tree.Results The conventional identification results revealed that it was an atypical Brucella,the gene similarity between the sequences of the test strain 16S rDNA and Brucella was 99%,between 16S rDNA sequence of Brucella abortus 544A and Brucella melitensis 16M was 96.99%.Phylogenetic tree revealed that the test stain was a Brucella,and closely related to Brucella abortus 544A and Brucella melitensis 16M.Conclusion The

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

    Directory of Open Access Journals (Sweden)

    R Rivers

    2006-01-01

    +, subset Th1, secreting gamma-interferon (γ-INF, a cytokine stimulatings macrophage bactericidal activity and cytotoxic activity of lymphocytes TCD8+, which are able to lyse Brucella infected cells. The main antigenic components of Brucella are lipopolysaccharide (LPS and proteins, especially superoxide dismutase (SOD with demonstrated immune potential. Brucellosis spreading is prevented with vaccines using attenuated or inactivated strains of B. abortus, such as strains 19, 45/20 and RB51. On the other hand, several investigators are making efforts to obtain immunity using antigenic structures of Brucella as subcellular vaccines and recently, genetic vaccines based on DNA and RNA molecules. The aim of this review is to give a current overview about brucellosis, its pathogenicity and the clinical syndrome. Firstly, an analysis of the genetic, antigenic and immune characteristics of Brucella is presented. Secondly, the vaccines presently used for prevention and the research on subcellular vaccines are discussed. Finally, the new approach in the vaccine investigation, genetic DNA and RNA vaccines, for Brucella is presented.

  6. [A case of Brucella spondylodiscitis after a visit to Lebanon].

    Science.gov (United States)

    Nielsen, Stig Lønberg; Johansen, Isik Somuncu

    2012-02-13

    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 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.

  7. 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.

  8. Evaluation of the ability of two multiplex PCR assays (Bruce-ladder and Suis-ladder) to distinguish six Brucella species and Brucella suis at the biovar level%应用多重PCR鉴别布鲁氏菌种及猪种5个生物型研究

    Institute of Scientific and Technical Information of China (English)

    陈军; 崔步云; 赵鸿雁; 朴东日; 姜海; 邰新平; 田国忠

    2013-01-01

    Objective To evaluate the ability of two types of multiplex PCR assays (Bruce-ladder and Suis-ladder) to distinguish six Brucella species and five biovars of Brucella suis. Methods A Bruce-ladder multiplex PCR assay was performed using 8 pairs of primers to distinguish six Brucella species. A Suis-ladder multiplex PCR assay was performed using 4 pairs of primers to distinguish five biovars of B. suis. The tested strains, which consisted of 27 reference strains of six Brucella species and 239 Brucella isolates, were examined using PCR assays and methods of biological identification. Results The Bruce-ladder multiplex PCR assay differentiated six Brucella species, including Brucella abortus, Brucella melitensis , Brucella suis , Brucella canis , Brucella ovis , and Brucella neotomae , in a single step. The Suis-ladder multiplex PCR assay differentiated five biovars of B. suis in a single step. The electrophoresis patterns of the vaccine strain S2 and biovar 1 of B. suis were identical. The total rate of concordance between methods of biological identification and multiplex PCR assays was 100%. Conclusion The Bruce-ladder and Suis-ladder multiplex PCR assays are rapid and specific methods of respectively identifying Brucella species and B. suis at the biovar level.%目的 应用布鲁氏菌种多重PCR和猪种多重PCR对布鲁氏菌进行鉴定. 方法 对两套多重PCR方法进行条件优化,应用布鲁氏菌参考菌株、疫苗株和临床分离菌株进行评价,并与生物学分型方法进行比较. 结果 布鲁氏菌种多重PCR扩增(包括牛种布鲁氏菌8个生物型,猪种5个生物型,羊种3个生物型,以及绵羊附睾、犬和沙漠森林野鼠种各1个生物型)可获得152~1 682 bp的8个DNA片段,但种间DNA片段数不一.猪种多重PCR扩增猪种5个生物型菌株得到197~774 bp的7个DNA片段,不同生物型的DNA扩增图谱不同;猪种疫苗株S2扩增图谱与猪种生物1型相同,多重PCR分型方法与生物学

  9. iNOS-producing inflammatory dendritic cells constitute the major infected cell type during the chronic Leishmania major infection phase of C57BL/6 resistant mice.

    Directory of Open Access Journals (Sweden)

    Carl De Trez

    2009-06-01

    Full Text Available Leishmania major parasites reside and multiply in late endosomal compartments of host phagocytic cells. Immune control of Leishmania growth absolutely requires expression of inducible Nitric Oxide Synthase (iNOS/NOS2 and subsequent production of NO. Here, we show that CD11b+ CD11c+ Ly-6C+ MHC-II+ cells are the main iNOS-producing cells in the footpad lesion and in the draining lymph node of Leishmania major-infected C57BL/6 mice. These cells are phenotypically similar to iNOS-producing inflammatory DC (iNOS-DC observed in the mouse models of Listeria monocytogenes and Brucella melitensis infection. The use of DsRed-expressing parasites demonstrated that these iNOS-producing cells are the major infected population in the lesions and the draining lymph nodes. Analysis of various genetically deficient mouse strains revealed the requirement of CCR2 expression for the recruitment of iNOS-DC in the draining lymph nodes, whereas their activation is strongly dependent on CD40, IL-12, IFN-gamma and MyD88 molecules with a partial contribution of TNF-alpha and TLR9. In contrast, STAT-6 deficiency enhanced iNOS-DC recruitment and activation in susceptible BALB/c mice, demonstrating a key role for IL-4 and IL-13 as negative regulators. Taken together, our results suggest that iNOS-DC represent a major class of Th1-regulated effector cell population and constitute the most frequent infected cell type during chronic Leishmania major infection phase of C57BL/6 resistant mice.

  10. Sequential real-time PCR assays applied to identification of genomic signatures in formalin-fixed paraffin-embedded tissues: a case report about brucella-induced osteomyelitis.

    Science.gov (United States)

    Zhang, Binxue; Wear, Douglas J; Stojadinovic, Alexander; Izadjoo, Mina

    2013-01-01

    Brucellosis is a zoonotic infection transmitted from animals to human by ingestion of infected food products, direct contact with an infected animal, or inhalation of aerosols. Brucella infection-induced osteomyelitis may present only with nonspecific clinical and radiographic findings, mild elevations in serum inflammatory markers, as well as nonspecific histological changes. We studied a case of an Iraqi war veteran with multifocal vertebral body and left iliac bone lesions on radio nucleotide scans and magnetic resonance imaging, clinically suspected osteomyelitis possibly because of Brucella. Although histomorphological findings were nonspecific, consisting of chronic inflammatory cell infiltrate and reactive fibrosis, tissue gram and silver impregnation stains of bone biopsies were informative, revealing gram-negative coccobacilli consistent in size with Brucella species. Total nucleic acids were extracted from formalin-fixed paraffin-embedded tissues and amplified by sequential real-time polymerase chain reaction, targeting genes coding (1) outer membrane protein (omp-31) of Brucella species and (2) insertion sequence (IS711) of Brucella abortus (b-abt). Polymerase chain reaction results confirmed B. abortus as the causative pathogens for presumed diagnosis of Brucella osteomyelitis.

  11. Brucella and Osteoarticular Cell Activation: Partners in Crime

    Science.gov (United States)

    Giambartolomei, Guillermo H.; Arriola Benitez, Paula C.; Delpino, M. Victoria

    2017-01-01

    Osteoarticular brucellosis is the most common presentation of human active disease although its prevalence varies widely. The three most common forms of osteoarticular involvement are sacroiliitis, spondylitis, and peripheral arthritis. The molecular mechanisms implicated in bone damage have been recently elucidated. B. abortus induces bone damage through diverse mechanisms in which TNF-α and the receptor activator of nuclear factor kappa-B ligand (RANKL)-the natural modulator of bone homeostasis are involved. These processes are driven by inflammatory cells, like monocytes/macrophages, neutrophils, Th17 CD4+ T, and B cells. In addition, Brucella abortus has a direct effect on osteoarticular cells and tilts homeostatic bone remodeling. These bacteria inhibit bone matrix deposition by osteoblasts (the only bone cells involved in bone deposition), and modify the phenotype of these cells to produce matrix metalloproteinases (MMPs) and cytokine secretion, contributing to bone matrix degradation. B. abortus also affects osteoclasts (cells naturally involved in bone resorption) by inducing an increase in osteoclastogenesis and osteoclast activation; thus, increasing mineral and organic bone matrix resorption, contributing to bone damage. Given that the pathology induced by Brucella species involved joint tissue, experiments conducted on synoviocytes revealed that besides inducing the activation of these cells to secrete chemokines, proinflammatory cytokines and MMPS, the infection also inhibits synoviocyte apoptosis. Brucella is an intracellular bacterium that replicates preferentially in the endoplasmic reticulum of macrophages. The analysis of B. abortus-infected synoviocytes indicated that bacteria also replicate in their reticulum suggesting that they could use this cell type for intracellular replication during the osteoarticular localization of the disease. Finally, the molecular mechanisms of osteoarticular brucellosis discovered recently shed light on how the

  12. Brucella and Osteoarticular Cell Activation: Partners in Crime.

    Science.gov (United States)

    Giambartolomei, Guillermo H; Arriola Benitez, Paula C; Delpino, M Victoria

    2017-01-01

    Osteoarticular brucellosis is the most common presentation of human active disease although its prevalence varies widely. The three most common forms of osteoarticular involvement are sacroiliitis, spondylitis, and peripheral arthritis. The molecular mechanisms implicated in bone damage have been recently elucidated. B. abortus induces bone damage through diverse mechanisms in which TNF-α and the receptor activator of nuclear factor kappa-B ligand (RANKL)-the natural modulator of bone homeostasis are involved. These processes are driven by inflammatory cells, like monocytes/macrophages, neutrophils, Th17 CD4(+) T, and B cells. In addition, Brucella abortus has a direct effect on osteoarticular cells and tilts homeostatic bone remodeling. These bacteria inhibit bone matrix deposition by osteoblasts (the only bone cells involved in bone deposition), and modify the phenotype of these cells to produce matrix metalloproteinases (MMPs) and cytokine secretion, contributing to bone matrix degradation. B. abortus also affects osteoclasts (cells naturally involved in bone resorption) by inducing an increase in osteoclastogenesis and osteoclast activation; thus, increasing mineral and organic bone matrix resorption, contributing to bone damage. Given that the pathology induced by Brucella species involved joint tissue, experiments conducted on synoviocytes revealed that besides inducing the activation of these cells to secrete chemokines, proinflammatory cytokines and MMPS, the infection also inhibits synoviocyte apoptosis. Brucella is an intracellular bacterium that replicates preferentially in the endoplasmic reticulum of macrophages. The analysis of B. abortus-infected synoviocytes indicated that bacteria also replicate in their reticulum suggesting that they could use this cell type for intracellular replication during the osteoarticular localization of the disease. Finally, the molecular mechanisms of osteoarticular brucellosis discovered recently shed light on how the

  13. Biological characteristics of five strains of Brucella isolated in frozen human blood samples%冻存人血中分离的5株布鲁杆菌的生物学特征分析

    Institute of Scientific and Technical Information of China (English)

    王锐泽; 皮鑫; 张翠; 刘新彬; 关超玲; 赵倩; 赵硕; 李萌; 甄清

    2015-01-01

    Objective To understand the survival situation of Brucella in frozen human blood samples.Methods Blood samples of outpatients with brucellosis were from the Songyuan Center for Disease Control and Prevention.Twenty-eight blood samples of patients who had a symptom of fever and epidemiological contact history were collected,which were kept in tubes containing sodium citrate anticoagulation,and stored in the fridge at-20 ℃.Isolation and culture of pathogen in frozen human blood samples were carried out.Suspected colonies were detected by Grams stain and microscope examination,then further tested by serum agglutination test with Brucella,A and M factors positive serum.The strains were detected with Brucella tautonym primers by multiplex-PCR amplification.Results In the twenty-eight blood samples frozen at different times,colonies were found in five samples after isolation and culture of pathogen for 4-5 d.The colonies were arranged in transparence,moist or milky lawn;the Grams staining was negative;serum agglutination test with Brucella,A and M factors positive serum were positive.Five strains of Brucella were preliminary considered as Brucella melitensis type 3.The freezing time of Brucella being isolated in five human blood samples was from 6 to 17 d.The results of multiplex-PCR showed that a band of 223 bp could be amplified from strains of Brucella abortus and Brucella melitensis,and that a band of 488 bp could be amplified from strain of Brucella abortus,and 310 bp from Brucella melitensis.The results of five tested strains were identical with those of Brucella melitensis.Conclusion Brucella could be survived in frozen human blood samples for a certain time,and multiplex-PCR amplification with Brucella tautonym primers could provide a basis for diagnose and treatment of Brucella.%目的 了解布鲁杆菌在冻存血液中的存活情况.方法 在松原市疾病预防控制中心布鲁杆菌病(简称布病)门诊就诊的人群中,收集有发热症状及

  14. Neurological complications of brucella spondylitis.

    Science.gov (United States)

    Mousa, A M; Bahar, R H; Araj, G F; Koshy, T S; Muhtaseb, S A; al-Mudallal, D S; Marafie, A A

    1990-01-01

    Twenty-two patients with brucella spondylitis and neurobrucellosis were studied during a 2-year period. The diagnosis was based on history of exposure, compatible signs and symptoms, high antibody titre and/or positive culture of a clinical specimen(s). Spondylitis was confirmed by plain radiographs, bone scan, CT and in some cases by histology. Neurobrucellosis was confirmed by CSF examination and culture, myelography, NCV, EMG and CT head. The spondylitis was early in 4 cases, chronic active in 12, smouldering "partially healed" in 3 and healed in 3 cases. Of these, 15 patients (68%) had neurological complications of various types. Plain radiographs were not a good index of activity of spondylitis. Tc99 bone scan was not specific and it remained positive long after the completion of therapy. CT was superior in revealing details of bone destruction, soft tissue swelling and entrapment of nerve roots and cord. The 3 modalities were complementary. Spondylitis is commonly associated with neurobrucellosis and symptoms of one may over shadow those of the other and in some cases neurobrucellosis may be subclinical. In all cases of spondylitis, a thorough search for neurobrucellosis should be made and vice versa. Prolonged treatment with a combination of 3 anti-brucella drugs is recommended and prolonged follow-up is necessary.

  15. Brucella ceti and brucellosis in cetaceans

    Directory of Open Access Journals (Sweden)

    Edgardo eMoreno

    2012-02-01

    Full Text Available 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.

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

    Directory of Open Access Journals (Sweden)

    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.