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Sample records for brucella abortus rb51

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

    Science.gov (United States)

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

    2016-08-01

    The aim of this study was to evaluate the shedding of Brucella abortus in the milk of cows vaccinated with a full dose of RB51 during lactation. Eighteen cows, nine previously vaccinated with S19 as calves and nine non-vaccinated, were immunized subcutaneously with 1.3×10(10)CFU of B. abortus RB51, 30-60days after parturition. Milk samples from all animals were collected daily until day 7, and at weekly interval for the next 9 weeks after vaccination. To evaluate the shedding of B. abortus, milk samples were submitted for culture and PCR. No B. abortus was isolated from any sample tested. Only one sample, collected on first day after vaccination from a cow previously vaccinated, was faintly positive in the PCR. In conclusion, the public health hazard associated with milk consumption from cows vaccinated with RB51 in post-partum is very low, despite vaccination with the full dose and regardless of previous S19 vaccination. PMID:27143220

  2. Immune Response of Calves Vaccinated with Brucella abortus S19 or RB51 and Revaccinated with RB51

    OpenAIRE

    Elaine M S Dorneles; Lima, Graciela K.; Andréa Teixeira-Carvalho; Araújo, Márcio S S; Martins-Filho, Olindo A; Nammalwar Sriranganathan; Hamzeh Al Qublan; Heinemann, Marcos B; Andrey P. Lage

    2015-01-01

    Brucella abortus S19 and RB51 strains have been successfully used to control bovine brucellosis worldwide; however, currently, most of our understanding of the protective immune response induced by vaccination comes from studies in mice. The aim of this study was to characterize and compare the immune responses induced in cattle prime-immunized with B. abortus S19 or RB51 and revaccinated with RB51. Female calves, aged 4 to 8 months, were vaccinated with either vaccine S19 (0.6-1.2 x 1011 CFU...

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

    OpenAIRE

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

    2010-01-01

    Approximately 250 000 water buffalo (Bubalus bubalis) live in the Campania region of southern Italy where the breeding of this species is very popular. Of these animals, almost 150 000 are concentrated in the Caserta province where the prevalence of Brucella abortus in this species represents approximately 20% at herd level. The Italian brucellosis eradication programme provides a slaughter and vaccination strategy for this province. B. abortus strain RB51 (RB51) has become the official vacci...

  4. Comparison of Biological and Immunological Characterization of Lipopolysaccharides From Brucella abortus RB51 and S19

    OpenAIRE

    Kianmehr, Zahra; Kaboudanian Ardestani, Sussan; Soleimanjahi, Hoorieh; Fotouhi, Fatemeh; Alamian, Saeed; Ahmadian, Shahin

    2015-01-01

    Background: Brucella abortus RB51 is a rough stable mutant strain, which has been widely used as a live vaccine for prevention of brucellosis in cattle instead of B. abortus strain S19. B. abortus lipopolysaccharide (LPS) has unique properties in comparison to other bacterial LPS. Objectives: In the current study, two types of LPS, smooth (S-LPS) and rough (R-LPS) were purified from B. abortus S19 and RB51, respectively. The aim of this study was to evaluate biological and immunological prope...

  5. Immune Response of Calves Vaccinated with Brucella abortus S19 or RB51 and Revaccinated with RB51

    Science.gov (United States)

    Dorneles, Elaine M. S.; Lima, Graciela K.; Teixeira-Carvalho, Andréa; Araújo, Márcio S. S.; Martins-Filho, Olindo A.; Sriranganathan, Nammalwar; Al Qublan, Hamzeh; Heinemann, Marcos B.; Lage, Andrey P.

    2015-01-01

    Brucella abortus S19 and RB51 strains have been successfully used to control bovine brucellosis worldwide; however, currently, most of our understanding of the protective immune response induced by vaccination comes from studies in mice. The aim of this study was to characterize and compare the immune responses induced in cattle prime-immunized with B. abortus S19 or RB51 and revaccinated with RB51. Female calves, aged 4 to 8 months, were vaccinated with either vaccine S19 (0.6–1.2 x 1011 CFU) or RB51 (1.3 x 1010 CFU) on day 0, and revaccinated with RB51 (1.3 x 1010 CFU) on day 365 of the experiment. Characterization of the immune response was performed using serum and peripheral blood mononuclear cells. Blood samples were collected on days 0, 28, 210, 365, 393 and 575 post-immunization. Results showed that S19 and RB51 vaccination induced an immune response characterized by proliferation of CD4+ and CD8+ T-cells; IFN-ɣ and IL-17A production by CD4+ T-cells; cytotoxic CD8+ T-cells; IL-6 secretion; CD4+ and CD8+ memory cells; antibodies of IgG1 class; and expression of the phenotypes of activation in T-cells. However, the immune response stimulated by S19 compared to RB51 showed higher persistency of IFN-ɣ and CD4+ memory cells, induction of CD21+ memory cells and higher secretion of IL-6. After RB51 revaccination, the immune response was chiefly characterized by increase in IFN-ɣ expression, proliferation of antigen-specific CD4+ and CD8+ T-cells, cytotoxic CD8+ T-cells and decrease of IL-6 production in both groups. Nevertheless, a different polarization of the immune response, CD4+- or CD8+-dominant, was observed after the booster with RB51 for S19 and RB51 prime-vaccinated animals, respectively. Our results indicate that after prime vaccination both vaccine strains induce a strong and complex Th1 immune response, although after RB51 revaccination the differences between immune profiles induced by prime-vaccination become accentuated. PMID:26352261

  6. Immune Response of Calves Vaccinated with Brucella abortus S19 or RB51 and Revaccinated with RB51.

    Directory of Open Access Journals (Sweden)

    Elaine M S Dorneles

    Full Text Available Brucella abortus S19 and RB51 strains have been successfully used to control bovine brucellosis worldwide; however, currently, most of our understanding of the protective immune response induced by vaccination comes from studies in mice. The aim of this study was to characterize and compare the immune responses induced in cattle prime-immunized with B. abortus S19 or RB51 and revaccinated with RB51. Female calves, aged 4 to 8 months, were vaccinated with either vaccine S19 (0.6-1.2 x 1011 CFU or RB51 (1.3 x 1010 CFU on day 0, and revaccinated with RB51 (1.3 x 1010 CFU on day 365 of the experiment. Characterization of the immune response was performed using serum and peripheral blood mononuclear cells. Blood samples were collected on days 0, 28, 210, 365, 393 and 575 post-immunization. Results showed that S19 and RB51 vaccination induced an immune response characterized by proliferation of CD4+ and CD8+ T-cells; IFN-ɣ and IL-17A production by CD4+ T-cells; cytotoxic CD8+ T-cells; IL-6 secretion; CD4+ and CD8+ memory cells; antibodies of IgG1 class; and expression of the phenotypes of activation in T-cells. However, the immune response stimulated by S19 compared to RB51 showed higher persistency of IFN-ɣ and CD4+ memory cells, induction of CD21+ memory cells and higher secretion of IL-6. After RB51 revaccination, the immune response was chiefly characterized by increase in IFN-ɣ expression, proliferation of antigen-specific CD4+ and CD8+ T-cells, cytotoxic CD8+ T-cells and decrease of IL-6 production in both groups. Nevertheless, a different polarization of the immune response, CD4+- or CD8+-dominant, was observed after the booster with RB51 for S19 and RB51 prime-vaccinated animals, respectively. Our results indicate that after prime vaccination both vaccine strains induce a strong and complex Th1 immune response, although after RB51 revaccination the differences between immune profiles induced by prime-vaccination become accentuated.

  7. Immune Response of Calves Vaccinated with Brucella abortus S19 or RB51 and Revaccinated with RB51.

    Science.gov (United States)

    Dorneles, Elaine M S; Lima, Graciela K; Teixeira-Carvalho, Andréa; Araújo, Márcio S S; Martins-Filho, Olindo A; Sriranganathan, Nammalwar; Al Qublan, Hamzeh; Heinemann, Marcos B; Lage, Andrey P

    2015-01-01

    Brucella abortus S19 and RB51 strains have been successfully used to control bovine brucellosis worldwide; however, currently, most of our understanding of the protective immune response induced by vaccination comes from studies in mice. The aim of this study was to characterize and compare the immune responses induced in cattle prime-immunized with B. abortus S19 or RB51 and revaccinated with RB51. Female calves, aged 4 to 8 months, were vaccinated with either vaccine S19 (0.6-1.2 x 1011 CFU) or RB51 (1.3 x 1010 CFU) on day 0, and revaccinated with RB51 (1.3 x 1010 CFU) on day 365 of the experiment. Characterization of the immune response was performed using serum and peripheral blood mononuclear cells. Blood samples were collected on days 0, 28, 210, 365, 393 and 575 post-immunization. Results showed that S19 and RB51 vaccination induced an immune response characterized by proliferation of CD4+ and CD8+ T-cells; IFN-ɣ and IL-17A production by CD4+ T-cells; cytotoxic CD8+ T-cells; IL-6 secretion; CD4+ and CD8+ memory cells; antibodies of IgG1 class; and expression of the phenotypes of activation in T-cells. However, the immune response stimulated by S19 compared to RB51 showed higher persistency of IFN-ɣ and CD4+ memory cells, induction of CD21+ memory cells and higher secretion of IL-6. After RB51 revaccination, the immune response was chiefly characterized by increase in IFN-ɣ expression, proliferation of antigen-specific CD4+ and CD8+ T-cells, cytotoxic CD8+ T-cells and decrease of IL-6 production in both groups. Nevertheless, a different polarization of the immune response, CD4+- or CD8+-dominant, was observed after the booster with RB51 for S19 and RB51 prime-vaccinated animals, respectively. Our results indicate that after prime vaccination both vaccine strains induce a strong and complex Th1 immune response, although after RB51 revaccination the differences between immune profiles induced by prime-vaccination become accentuated. PMID:26352261

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

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    Vincenzo Caporale

    2010-03-01

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

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

    Science.gov (United States)

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

    2010-01-01

    Approximately 250,000 water buffalo (Bubalus bubalis) live in the Campania region of southern Italy where the breeding of this species is very popular. Of these animals, almost 150,000 are concentrated in the Caserta province where the prevalence of Brucella abortus in this species represents approximately 20% at herd level. The Italian brucellosis eradication programme provides a slaughter and vaccination strategy for this province. B. abortus strain RB51 (RB51) has become the official vaccine for the prevention of brucellosis in cattle in several countries. The aim of this study was to evaluate the efficacy of RB51 in water buffalo compared to the B. abortus S19 vaccine (S19). The study was performed in accordance with a protocol described in mice. Female buffalo aged five months were inoculated. Five received a RB51 dosage on two occasions that was three times greater than that approved for use in cattle and a booster after one month, five received B. abortus S19 vaccine at the standard dosage and three controls received a phosphate buffer solution. Buffalo were then challenged with a virulent B. abortus strain 544 thirty days post vaccination. Antibodies that developed in the five animals vaccinated with RB51 were not detected by the Rose Bengal test or complement fixation test (CFT) and were also tested by CFT prepared with RB51 antigen. After culling, B. abortus was cultured from the spleen, retropharyngeal and supra-mammary lymph nodes. A statistical evaluation was performed to assess the immunogenicity values obtained in buffalo vaccinated with S19, compared to those obtained in buffalo vaccinated with the RB51 vaccine and in the unvaccinated control group. PMID:20391363

  10. Efficacy of dart or booster vaccination with strain RB51 in protecting bison against experimental Brucella abortus challenge

    Science.gov (United States)

    Vaccination is an effective tool for reducing the prevalence of brucellosis in natural hosts. In this study, we characterized the efficacy of the Brucella abortus strain RB51 (RB51) vaccine in bison when delivered by single intramuscular vaccination (Hand RB51), single pneumatic dart delivery (Dart ...

  11. Biosafety of parenteral Brucella abortus RB51 vaccine in bison calves

    Science.gov (United States)

    Roffe, T.J.; Olsen, S.C.; Gidlewski, T.; Jensen, A.E.; Palmer, M.V.; Huber, R.

    1999-01-01

    Vaccination is considered among the primary management tools for reducing brucellosis prevalence in Greater Yellowstone Area (GYA) ungulates. Before their use, however, vaccine safety and efficacy must be demonstrated. Twenty-seven female bison (Bison bison) calves (approx 5 months old) were vaccinated with Brucella abortus Strain RB51 (1.5 x 1010 colony forming units [CFU], subcutaneously) as part of routine management. We assessed the persistence, pathology, shedding, and transmission associated with RB51 by serial necropsy, bacteriology, histopathology, and serology of 20 of these 27 vaccinated calves, and RB51 serology of 10 nonvaccinated, commingling adult females. With the exception of 1 calf, RB51 dot-blot titers at necropsy were cows were negative at a 1:20 dilution at 26 weeks PV. Our results suggest that RB51 persists longer in bison calves than in domestic cattle and is systemically distributed within lymphatic tissues. However, bison apparently clear the RB51 vaccine strain without shedding, transmission, or significant adverse reactions.

  12. Comparison of Biological and Immunological Characterization of Lipopolysaccharides From Brucella abortus RB51 and S19

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    Kianmehr, Zahra; Kaboudanian Ardestani, Sussan; Soleimanjahi, Hoorieh; Fotouhi, Fatemeh; Alamian, Saeed; Ahmadian, Shahin

    2015-01-01

    Background: Brucella abortus RB51 is a rough stable mutant strain, which has been widely used as a live vaccine for prevention of brucellosis in cattle instead of B. abortus strain S19. B. abortus lipopolysaccharide (LPS) has unique properties in comparison to other bacterial LPS. Objectives: In the current study, two types of LPS, smooth (S-LPS) and rough (R-LPS) were purified from B. abortus S19 and RB51, respectively. The aim of this study was to evaluate biological and immunological properties of purified LPS as an immunogenical determinant. Materials and Methods: Primarily, S19 and RB51 LPS were extracted and purified by two different modifications of the phenol water method. The final purity of LPS was determined by chemical analysis (2-keto-3-deoxyoctonate (KDO), glycan, phosphate and protein content) and different staining methods, following sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). C57BL/6 mice were immunized subcutaneously three times at biweekly intervals with the same amount of purified LPSs. The humoral immunity was evaluated by measuring specific IgG levels and also different cytokine levels, such as IFN-γ, TNF-α, IL-4 and IL-10, were determined for assessing T-cell immune response. Results: Biochemical analysis data and SDS-PAGE profile showed that the chemical nature of S19 LPS is different from RB51 LPS. Both S and R-LPS induce an immune response. T-cell immune response induced by both S and R-LPS had almost the same pattern whereas S19 LPS elicited humoral immunity, which was higher than RB51 LPS. Conclusions: Purified LPS can be considered as a safe adjuvant and can be used as a component in prophylactic and therapeutic vaccines targeting infectious disease, cancer and allergies. PMID:26862376

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

    Science.gov (United States)

    Thirty-one bison heifers were randomly assigned to saline (control; n=7) or single vaccination (n=24) with 1010 CFU of B. abortus strain RB51 (RB51). Some vaccinated bison were randomly selected for booster vaccination with 10**10 CFU of RB51 at 11 months after initial vaccination (n=16). When comp...

  14. Enhancement of the Brucella AMOS PCR assay for differentiation of Brucella abortus vaccine strains S19 and RB51.

    OpenAIRE

    Bricker, B J; Halling, S. M.

    1995-01-01

    Because the brucellosis eradication program uses slaughter and quarantine as control measures, it would benefit from faster methods of bacterial identification. Distinguishing vaccine strains from strains that cause infections among vaccinated herds in the field is essential. To accomplish this, our PCR-based, species-specific assay (B. J. Bricker and S. M. Halling, J. Clin. Microbiol. 32:2660-2666, 1994) was updated to identify Brucella abortus vaccine strains S19 and RB51. Three new oligonu...

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

    Science.gov (United States)

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

    2002-01-01

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

  16. Brucella abortus Strain RB51 Vaccine: Immune Response after Calfhood Vaccination and Field Investigation in Italian Cattle Population

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    Manuela Tittarelli

    2008-01-01

    Full Text Available Immune response to Brucella abortus strain RB51 vaccine was measured in cattle vaccinated at calfhood. After an increase at day 6 post-vaccination (pv, the antibody level recorded in the 10 vaccinated animals remained constant for two months, and then progressively decreased. All vaccinated animals remained negative from day 162 pv to the end of the study (day 300 pv. Only at days 13 and 14 pv the RB51-CFT showed 100% sensitivity (credibility interval (CI 76.2%–100%. The results indicate that the possibility to use RB51-CFT for the identification of cattle vaccinated at calfhood with RB51 is limited in time. A field investigation was carried out on 26,975 sera collected on regional basis from the Italian cattle population. The study outcomes indicate that in case of RB51-CFT positive results observed in officially Brucellosis-free (OBF areas and, in any case, when an illegal use of RB51 vaccine is suspected, the use of the RB51-CFT alone is not sufficient to identify all the vaccinated animals. The design of a more sophisticated diagnostic protocol including an epidemiological investigation, the use of RB51-CFT, and the use of the skin test with RB51 as antigen is deemed more appropriate for the identification of RB51 vaccinated animals.

  17. Reporte de primer caso humano de aislamiento y tipificación de Brucella abortus RB 51: first report in Chile Isolation and identification of Brucella abortus RB 51 in human

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    M. Villarroel

    2000-01-01

    Full Text Available En el Laboratorio de Microbiología del Hospital Base de Osorno, se aisló una cepa de Brucella sp, que posteriormente se tipificó en el Laboratorio Regional de Diagnósticos del Servicio Agrícola y Ganadero, SAG Osorno. Esta cepa fue aislada de un hemocultivo realizado a un paciente con sintomatología clínica compatible con Brucelosis. El resultado de la tipificación fue Brucella abortus RB 51, cepa vaccinal que se presumía apatógena para el hombre. Este es el primer reporte a nivel mundial de un aislamiento de B. abortus RB 51 en humanosA Brucella strain was isolated from a haemoculture of a clinically ill veterinarian practitioner, at the Laboratory of Microbiology, Hospital Base Osorno, Chile. This strain was identified as Brucella abortus RB 51 at Laboratorio de Diagnósticos from the Servicio Agrícola y Ganadero (SAG, Osorno, based mainly in the following characteristics: resistence to Rifampicin, aerobic growth and the production of only rough colonies. The strain RB 51 was confirmed by PCR analysis

  18. Validation of the Abbreviated Brucella AMOS PCR as a Rapid Screening Method for Differentiation of Brucella abortus Field Strain Isolates and the Vaccine Strains, 19 and RB51

    OpenAIRE

    Ewalt, Darla R; Bricker, Betsy J.

    2000-01-01

    The Brucella AMOS PCR assay was previously developed to identify and differentiate specific Brucella species. In this study, an abbreviated Brucella AMOS PCR test was evaluated to determine its accuracy in differentiating Brucella abortus into three categories: field strains, vaccine strain 19 (S19), and vaccine strain RB51/parent strain 2308 (S2308). Two hundred thirty-one isolates were identified and tested by the conventional biochemical tests and Brucella AMOS PCR. This included 120 isola...

  19. Overexpression of Brucella putative glycosyltransferase WbkA in B. abortus RB51 leads to production of exopolysaccharide

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    Neha eDabral

    2015-06-01

    Full Text Available Brucella spp. are Gram-negative, facultative intracellular bacteria that cause brucellosis in mammals. Brucella strains containing the O-polysaccharide in their cell wall structure exhibit a smooth phenotype whereas the strains devoid of the polysaccharide show rough phenotype. B. abortus strain RB51 is a stable rough attenuated mutant which is used as a licensed live vaccine for bovine brucellosis. Previous studies have shown that the wboA gene, which encodes a glycosyltransferase required for the synthesis of O-polysaccharide, is disrupted in B. abortus RB51 by an IS711 element. Although complementation of strain RB51 with a functional wboA gene results in O-polysaccharide synthesis in the cytoplasm, it does not result in smooth phenotype. The aim of this study was to determine if overexpression of Brucella WbkA or WbkE, two additional putative glycosyltransferases essential for O-polysaccharide synthesis, in strain RB51 would result in the O-polysaccharide synthesis and smooth phenotype. Our results demonstrate that overexpression of wbkA or wbkE gene in RB51 does not result in O-polysaccharide expression as shown by Western blotting with specific antibodies. However, wbkA, but not wbkE, overexpression leads to the development of a clumping phenotype and the production of exopolysaccharide(s containing mannose, galactose, N-acetylglucosamine and N-acetylgalactosamine. Moreover, we found that the clumping recombinant strain displays increased adhesion to polystyrene plates. The recombinant strain was similar to strain RB51 in its attenuation characteristic and in its ability to induce protective immunity against virulent B. abortus challenge in mice.

  20. Immune Responses of Elk to Initial and Booster Vaccinations with Brucella abortus Strain RB51 or 19

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    S. C. Olsen; Fach, S. J.; Palmer, M. V.; Sacco, R. E.; Stoffregen, W. C.; Waters, W.R.

    2006-01-01

    Previous studies have suggested that currently available brucellosis vaccines induce poor or no protection in elk (Cervus elaphus nelsoni). In this study, we characterized the immunologic responses of elk after initial or booster vaccination with Brucella abortus strains RB51 (SRB51) and 19 (S19). Elk were vaccinated with saline or 1010 CFU of SRB51 or S19 (n = seven animals/treatment) and booster vaccinated with a similar dosage of the autologous vaccine at 65 weeks. Compared to nonvaccinate...

  1. Comparison of immune responses and resistance to brucellosis in mice vaccinated with Brucella abortus 19 or RB51.

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    Stevens, M G; S. C. Olsen; Pugh, G W; Brees, D

    1995-01-01

    Immune responses and resistance to infection with Brucella abortus 2308 (S2308) were measured in mice following vaccination with B. abortus 19 (S19) or the lipopolysaccharide (LPS) O-antigen-deficient mutant, strain RB51 (SRB51). Live bacteria persisted for 8 weeks in spleens of mice vaccinated with 5 x 10(6) or 5 x 10(8) CFU of SRB51, whereas bacteria persisted for 12 weeks in mice vaccinated with 5 x 10(6) CFU of S19. Mice vaccinated with 5 x 10(6) or 5 x 10(8) CFU of SRB51 had increased re...

  2. 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

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    Pauline eNol

    2016-02-01

    Full Text Available In recent years, elk (Cervus canadensis have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area. In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the development of effective disease management strategies for wild elk herds is of utmost importance, not only for the prevention of reintroduction of brucellosis to cattle, but also for the overall health of the Greater Yellowstone Area elk populations. In two studies, we evaluated the efficacy of B. abortus strain RB51 over-expressing superoxide dismutase and glycosytransferase for protecting elk from infection and disease caused by B. abortus after experimental infection with a virulent B. abortus strain. Our data indicate that the recombinant vaccine does not protect elk against brucellosis. Further work is needed for development of an effective brucellosis vaccine for use in elk

  3. Brucella abortus S19 and RB51 vaccine immunogenicity test: Evaluation of three mice (BALB/c, Swiss and CD-1) and two challenge strains (544 and 2308).

    Science.gov (United States)

    Miranda, Karina Leite; Dorneles, Elaine Maria Seles; Pauletti, Rebeca Barbosa; Poester, Fernando Padilla; Lage, Andrey Pereira

    2015-01-15

    The aim of the present study was to evaluate the use of different mouse strains (BALB/c, Swiss and CD-1) and different challenge strains (Brucella abortus 544 and 2308) in the study of B. abortus vaccine (S19 and RB51) immunogenicity test in the murine model. No significant difference in B. abortus vaccine potency assay was found with the use of B. abortus 544 or B. abortus 2308 as challenge strain. Results of variance analysis showed an interaction between treatment and mouse strain; therefore these parameters could not be compared separately. When CD-1 groups were compared, those vaccinated showed significantly lower counts than non-vaccinated ones (PS19 or RB51). Similar results were observed on BALB/c groups. However, in Swiss mouse groups, S19 was more protective than RB51 (Pabortus strains 544 and 2308, can be used in immunogenicity tests of S19 and RB51 vaccines. PMID:25498211

  4. Valutazione dell’efficacia del vaccino Brucella abortus ceppo RB51 rispetto al vaccino di referenza Brucella abortus ceppo 19 nel bufalo

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    Massimo Scacchia

    2010-03-01

    Full Text Available Il patrimonio zootecnico della specie bufalina (Bubalus bubalis della regione Campania, è di 250 000 capi, di questi 150 000 allevati in aziende zootecniche della provincia di Caserta. In queste aziende, nel 2007, l’infezione da Brucella abortus ha avuto la prevalenza media, per allevamento, del 20%. Complessivamente, i 2/3 degli allevamenti positivi hanno evidenziato una prevalenza superiore al 10% e, di questi, i 3/4 una prevalenza superiore al 20%. Prendendo il 20% come valore di riferimento, la metà degli allevamenti infetti (22% degli allevamenti casertani ha evidenziato prevalenze inferiori o uguali al 20%, la restante metà (un altro 22% del totale prevalenze comprese tra il 20 e il 56%. In questo contesto epidemiologico è stato adottato un piano di eradicazione della brucellosi che prevedeva l’abbattimento dei capi infetti e la vaccinazione del restante patrimonio bufalino delle zone con più alta incidenza. Per la profilassi vaccinale della brucellosi, il Manual of diagnostic tests and vaccines for terrestrial animals (OIE prevede l’utilizzo del vaccino B. abortus S19 (S19. Purtroppo, l’utilizzo del vaccino negli animali adulti non è privo di possibili effetti indesiderati. Per superare questo aspetto negativo è stato ipotizzato l’impiego del vaccino di B. abortus RB51 (RB51 anche se in letteratura scientifica, sono risultati disponibili pochi dati relativi alla corretta dose vaccinale, all’efficacia e all’innocuità del vaccino nel bufalo. A tale scopo è stato condotto uno studio comparativo tra i due vaccini. Sono state utilizzate 13 femmine di bufalo di 5 mesi di età provenienti da un allevamento ufficialmente indenne da brucellosi. Un gruppo di 5 animali è stato vaccinato due volte, a distanza di un mese, con una dose di RB51 tre volte superiore a quella prevista per i bovini; un secondo gruppo di 5 bufale con S19 rispettando il dosaggio raccomandato per i bovini e un terzo gruppo, di 3 animali di controllo

  5. Medición de respuesta inmune humoral y celular frente a antígenos de Brucella abortus RB51 en bovinos (Evaluation of Humoral and cellular immune response evaluation against Brucella abortus strain RB51 antigens in bovine

    Directory of Open Access Journals (Sweden)

    N.I. Montaña S.

    1998-01-01

    Full Text Available Con el objeto de evaluar comparativamente el tipo de respuesta inmune inducido por los antígenos estructurales purificados y vacunas vivas de Brucella abortus, 14 bovinos criollos hembras de 19 meses de edad fueron distribuidos al azar, en cinco grupos experimentales e inmunizados vía subcutánea de la siguiente manera: OMP-II (proteínas de membrana externa, OMP-II-Cadena-O, OMP II acoplado a polisacárido-O, B. abortus cepa RB51, B. abortus cepa 19 (C19 y solución salina para el grupo control. Dos meses posinmunización, los animales fueron desafiados, vía intramuscular, con cepa patógena de referencia B. abortus 2308 y evaluados en su nivel de protección 30 días posdesafío. La respuesta inmune humoral específica fue determinada mediante las pruebas convencionales, de aglutinación Rosa de Bengala, fijación de complemento, inmunodifusión radial, ELISA indirecta, a los 8, 15, 30, 60 y 90 días posinmunización. Adicionalmente se determinó la subclase de IgG por ELISA tipo doble sandwich. La evaluación de respuesta celular fue determinada mediante linfoproliferación, expresada en índices de estimulación (IS, obtenidos por los niveles de incorporación de 3H timidina, medición de Interferón gamma (IFN-g, igualmente expresado como IS y cuantificación de subpoblaciones linfoides CD4+ y CD8+ por citofluorometría. Animales vacunados con cepas vivas demostraron protección total al reto, mientras se observó un 66% de protección en los vacunados con antígenos purificados. Se observó ausencia de anticuerpos a lipopolisacárido, sLPS, por las pruebas convencionales para B. abortus en todos los grupos inmunizados, excepto el vacunado con cepa 19, confirmando la ventaja diagnóstica de la utilización de la cepa RB51 o sus antígenos purificados. La linfoproliferación como una de las medidas de respuesta celular no demostró IS significativos (p0.05 frente a los antígenos estructurales purificados, OMP-II, Cadena-O y sLPS en

  6. EXPRESSION OF BACILLUS ANTHRACIS PROTECTIVE ANTIGEN IN VACCINE STRAIN BRUCELLA ABORTUS RB51

    OpenAIRE

    Poff, Sherry Ann

    1997-01-01

    Bacillus anthracis is a facultative intracellular bacterial pathogen that can cause cutaneous, gastrointestinal or respiratory disease in many vertebrates, including humans. Commercially available anthrax vaccines for immunization of humans are of limited duration and do not protect against the respiratory form of the disease. Brucella abortus is a facultative intracellular bacterium that causes chronic infection in animals and humans. As with other intracellular pathogens, cell mediated im...

  7. Genetic stability of Brucella abortus S19 and RB51 vaccine strains by multiple locus variable number tandem repeat analysis (MLVA16).

    Science.gov (United States)

    Dorneles, Elaine Maria Seles; de Faria, Ana Paula Paiva; Pauletti, Rebeca Barbosa; Santana, Jordana Almeida; Caldeira, George Afonso Vítor; Heinemann, Marcos Bryan; Titze-de-Almeida, Ricardo; Lage, Andrey Pereira

    2013-10-01

    The aims of the present study were (i) to assess the in vitro genetic stability of S19 and RB51 Brucella abortus vaccines strains and (ii) to evaluate the ability of multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) as a tool to be used in the quality control of live vaccines against brucellosis. Sixty-three batches of commercial S19 (n=53) and RB51 (n=10) vaccines, produced between 2006 and 2009, were used in this study. S19 and RB51 vaccines were obtained from, respectively, seven and two different manufacturers. Ten in vitro serial passages were performed on reference strains and on selected batches of commercial vaccines. All batches, reference strains and strains of serial passages were typed by the MLVA16. The results demonstrated that B. abortus S19 and RB51 vaccine strains are genetically stable and very homogeneous in their respective groups. Anyway, batches of S19 from one manufacturer and batches of RB51 from another presented genotypes distincts from the reference vaccine strains. In both cases, differences were found on locus Bruce07, which had addition of one repeat unit in the case of S19 batches and the deletion of one repeat unit in the case of RB51 batches. In summary, MLVA16 proved to be a molecular tool capable of discriminating small genomic variations and should be included in in vitro official tests. PMID:23933375

  8. Immune Responses of Elk to Initial and Booster Vaccinations with Brucella abortus Strain RB51 or 19

    Science.gov (United States)

    Olsen, S. C.; Fach, S. J.; Palmer, M. V.; Sacco, R. E.; Stoffregen, W. C.; Waters, W. R.

    2006-01-01

    Previous studies have suggested that currently available brucellosis vaccines induce poor or no protection in elk (Cervus elaphus nelsoni). In this study, we characterized the immunologic responses of elk after initial or booster vaccination with Brucella abortus strains RB51 (SRB51) and 19 (S19). Elk were vaccinated with saline or 1010 CFU of SRB51 or S19 (n = seven animals/treatment) and booster vaccinated with a similar dosage of the autologous vaccine at 65 weeks. Compared to nonvaccinates, elk vaccinated with SRB51 or S19 had greater (P 0.05) from the responses of nonvaccinated elk. Gamma interferon production in response to autologous or nonautologous Brucella antigens did not differ (P > 0.05) between controls and vaccinates after booster vaccination. Flow cytometric techniques suggested that proliferation occurred more frequently in immunoglobulin M-positive cells, with differences between vaccination and control treatments in CD4+ and CD8+ subset proliferation detected only at 22 weeks after initial vaccination. After booster vaccination, one technique ([3H]thymidine incorporation) suggested that proliferative responses to SRB51 antigen, but not S19 antigen, were greater (P < 0.05) in vaccinates compared to the responses of nonvaccinates. However, in general, flow cytometric and other techniques failed to detect significant anamnestic responses to autologous or nonautologous Brucella antigens in S19 or SRB51 vaccinates after booster vaccination. Although some cellular immune responses were detected after initial or booster vaccination of elk with SRB51 or S19, our data suggest that responses tend to be transient and much less robust than previously reported in SRB51-vaccinated cattle (Bos taurus) or bison (Bison bison). These data may explain why the vaccination of elk with S19 and SRB51 induces poor protection against brucellosis. PMID:17028213

  9. Immune responses and protection against experimental challenge after vaccination of bison with Brucella abortus strains RB51 or RB51 overexpressing superoxide dismutase and Glycosyltransferase genes

    Science.gov (United States)

    Vaccination is a tool that could be beneficial in managing the high prevalence of brucellosis in free-ranging bison in Yellowstone National Park. In this study, we characterized immunologic responses and protection against experimental challenge after vaccination of bison with Brucella abortus stra...

  10. Comparison of spleen cell proliferation in response to Brucella abortus 2308 lipopolysaccharide or proteins in mice vaccinated with strain 19 or RB51.

    OpenAIRE

    Stevens, M G; S. C. Olsen; Pugh, G W

    1995-01-01

    Mice vaccinated with Brucella abortus 19 (S19) or RB51 (SRB51) had spleen cells which proliferated in response to proteins of 32, 27, 18, and < 18 kDa but not in response to proteins of 106, 80, and 49 kDa from B. abortus 2308 (S2308) following vaccination and challenge infection with S2308. Spleen cells from mice vaccinated with S19 but not with SRB51 had increased proliferation in response to S2308 lipopolysaccharide (LPS) following challenge infection with S2308. We previously reported tha...

  11. Immune responses of elk to initial and booster vaccinations with Brucella abortus strain RB51 or 19.

    Science.gov (United States)

    Olsen, S C; Fach, S J; Palmer, M V; Sacco, R E; Stoffregen, W C; Waters, W R

    2006-10-01

    Previous studies have suggested that currently available brucellosis vaccines induce poor or no protection in elk (Cervus elaphus nelsoni). In this study, we characterized the immunologic responses of elk after initial or booster vaccination with Brucella abortus strains RB51 (SRB51) and 19 (S19). Elk were vaccinated with saline or 10(10) CFU of SRB51 or S19 (n=seven animals/treatment) and booster vaccinated with a similar dosage of the autologous vaccine at 65 weeks. Compared to nonvaccinates, elk vaccinated with SRB51 or S19 had greater (PS19 after initial vaccination and after booster vaccination. Compared to nonvaccinated elk, greater (PS19 (22 weeks) treatment groups. In general, proliferative responses of vaccinates to nonautologous antigens did not differ (P>0.05) from the responses of nonvaccinated elk. Gamma interferon production in response to autologous or nonautologous Brucella antigens did not differ (P>0.05) between controls and vaccinates after booster vaccination. Flow cytometric techniques suggested that proliferation occurred more frequently in immunoglobulin M-positive cells, with differences between vaccination and control treatments in CD4+ and CD8+ subset proliferation detected only at 22 weeks after initial vaccination. After booster vaccination, one technique ([3H]thymidine incorporation) suggested that proliferative responses to SRB51 antigen, but not S19 antigen, were greater (PBrucella antigens in S19 or SRB51 vaccinates after booster vaccination. Although some cellular immune responses were detected after initial or booster vaccination of elk with SRB51 or S19, our data suggest that responses tend to be transient and much less robust than previously reported in SRB51-vaccinated cattle (Bos taurus) or bison (Bison bison). These data may explain why the vaccination of elk with S19 and SRB51 induces poor protection against brucellosis. PMID:17028213

  12. 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...

  13. Efecto de una dieta con bajo aporte de selenio sobre la respuesta inmune a la vacuna Brucella abortus Cepa RB51 en vacas lecheras Effect of a low selenium diet on the immune response to Brucella abortus strain RB51 vaccine in dairy cows

    Directory of Open Access Journals (Sweden)

    V Leyán

    2006-01-01

    Full Text Available Se estudió el efecto de una dieta con bajo aporte de selenio (Se sobre la respuesta inmune a la vacuna Brucella abortus Cepa RB51 y la concentración de inmunoglobulinas séricas en vacas. Se utilizaron 12 vacas Friesian, estabuladas desde aproximadamente dos meses preparto y hasta el cuarto mes de lactancia mantenidas con una dieta basada en heno de pradera con bajo contenido de Se (0,02 ppm de MS y balanceada según requerimientos para el resto de nutrientes. Seis vacas conformaron el grupo de animales con bajo aporte de Se (Se-D y otras seis el grupo de animales suplementados (Se-S con selenato de bario (1 mg de Se/kg , 45 días previos al parto. Los animales fueron inmunizados con la vacuna RB51 al cuarto mes del experimento. Muestras de sangre fueron obtenidas previo a la suplementación con Se y cada 15 días hasta el término del experimento. El balance de Se fue medido mediante la actividad sanguínea de GSH-Px. Las concentraciones séricas de IgG, IgM e IgA se determinaron por inmunodifusión y los anticuerpos específicos contra Brucella abortus mediante ELISA y la respuesta inmune celular mediante pruebas de intradermorreacción a antígenos de Brucella abortus y estudio histológico de la reacción. La dieta con bajo contenido de Se provocó una disminución lenta y progresiva de la actividad de GSH-Px (The effect of a diet with a low selenium (Se content on the immune response to Brucella abortus Strain RB51 vaccine in dairy cows and in their serum inmunoglobulin concentrations was studied. Twelve pregnant Friesian cows (7 to 8 months were randomly allocated into two homogeneous groups of six animals each. Animals were maintained during 6 months in individual cubicles with water ad libitum and a diet based on grass hay with a low Se content (0.02 ppm base on dry matter and nutritionally balanced for other nutrients. One group was maintained only with the low Se diet (Se-D and the other group (Se-S was treated with barium selenate

  14. Suplementación con selenio en vaquillas: Efecto sobre la respuesta inmune a las vacunas Brucella abortus cepa RB51 y toxoide tetánico Selenium suplementation in heifers: Effect on the immune response to Brucella abortus strain RB51 and tetanus toxoid vaccines

    Directory of Open Access Journals (Sweden)

    V Leyán

    2005-01-01

    Full Text Available El trabajo tiene por objetivo evaluar el efecto de una suplementación con selenio (Se sobre la respuesta inmune a las vacunas Toxoide tetánico y Brucella abortus cepa RB51 en vaquillas con un adecuado balance metabólico de selenio (GSH-Px >130 U/g Hb. Para ello se empelaron 32 vaquillas Friesian de 18 a 24 meses de edad, asignadas al azar a dos grupos de 16 animales; uno suplementado (Se-S el día 0 con una dosis de selenato de bario (1 mg Se/kg, s.c., permaneciendo el otro como control no suplementado (No-S. Todas las vaquillas fueron mantenidas durante 9 meses (abril a enero a pastoreo sobre una pradera naturalizada con un contenido de Se de 0,04 ppm/MS. Los animales fueron inmunizados con vacuna RB51 el día 60 y posteriormente con Toxoide tetánico los días 120 y 150. Muestras de sangre fueron obtenidas previo a la suplementación y cada 15 días hasta el término del experimento. El balance metabólico de selenio fue evaluado mediante la actividad sanguínea de Glutatión peroxidasa (GSH-Px. La respuesta inmune humoral se evaluó determinando los anticuerpos séricos específicos para ambos antígenos mediante ELISA y la respuesta inmune celular mediante pruebas de intradermorreacción a antígenos de Brucella abortus. La administración de Se aumentó (P 0,05 en ambos grupos experimentales, mientras que la respuesta celular a la vacuna RB51 fue menor (P The effect of selenium (Se supplementation on the immune response to tetanus toxoid and Brucella abortus strain RB51 vaccines was studied in heifers with a normal Se status (GSH-Px activity > 130 U/g Hb. Frisian heifers (n-32, 18 to 24 months old were randomly allocated into two groups of 16 animals each. Animals from one group were supplemented (Se-S with one dose of barium selenate (1 mg/Se/kg. s.c. on day 0; animals from the other group remained as a control without supplementation (No-S. The heifers grazed during 9 months (April to January a pasture that contained 0.04 ppm/DM of Se

  15. Valutazione dell’efficacia del vaccino Brucella abortus ceppo RB51 rispetto al vaccino di referenza Brucella abortus ceppo 19 nel bufalo

    OpenAIRE

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

    2010-01-01

    Il patrimonio zootecnico della specie bufalina (Bubalus bubalis) della regione Campania, è di 250 000 capi, di questi 150 000 allevati in aziende zootecniche della provincia di Caserta. In queste aziende, nel 2007, l’infezione da Brucella abortus ha avuto la prevalenza media, per allevamento, del 20%. Complessivamente, i 2/3 degli allevamenti positivi hanno evidenziato una prevalenza superiore al 10% e, di questi, i 3/4 una prevalenza superiore al 20%. Prendendo il 20% come valore di riferime...

  16. Suplementación con selenio en vaquillas: Efecto sobre la respuesta inmune a las vacunas Brucella abortus cepa RB51 y toxoide tetánico Selenium suplementation in heifers: Effect on the immune response to Brucella abortus strain RB51 and tetanus toxoid vaccines

    OpenAIRE

    V Leyán; D Pesutic; G Schurig; F. WITTWER; P.A. CONTRERAS; J. KRUZE

    2005-01-01

    El trabajo tiene por objetivo evaluar el efecto de una suplementación con selenio (Se) sobre la respuesta inmune a las vacunas Toxoide tetánico y Brucella abortus cepa RB51 en vaquillas con un adecuado balance metabólico de selenio (GSH-Px >130 U/g Hb). Para ello se empelaron 32 vaquillas Friesian de 18 a 24 meses de edad, asignadas al azar a dos grupos de 16 animales; uno suplementado (Se-S) el día 0 con una dosis de selenato de bario (1 mg Se/kg, s.c.), permaneciendo el otro como control no...

  17. Characterization of recombinant B. abortus strain RB51SOD towards understanding the uncorrelated innate and adaptive immune responses induced by RB51SOD compared to its parent vaccine strain RB51

    Directory of Open Access Journals (Sweden)

    Jianguo eZhu

    2011-11-01

    Full Text Available Brucella abortus is a Gram-negative, facultative intracellular pathogen for several mammals, including humans. Live attenuated B. abortus strain RB51 is currently the official vaccine used against bovine brucellosis in the United States and several other countries. Overexpression of protective B. abortus antigen Cu/Zn superoxide dismutase (SOD in a recombinant strain of RB51 (strain RB51SOD significantly increases its vaccine efficacy against virulent B. abortus challenge in a mouse model. An attempt has been made to better understand the mechanism of the enhanced protective immunity of RB51SOD compared to its parent strain RB51. We previously reported that RB51SOD stimulated enhanced Th1 immune response. In this study, we further found that T effector cells derived from RB51SOD-immunized mice exhibited significantly higher cytotoxic T lymphocyte (CTL activity than T effector cells derived from RB51-immunized mice against virulent B. abortus-infected target cells. Meanwhile, the macrophage responses to these two strains were also studied. Compared to RB51, RB51SOD cells had a lower survival rate in macrophages and induced lower levels of macrophage apoptosis and necrosis. The decreased survival of RB51SOD cells correlates with the higher sensitivity of RB51SOD, compared to RB51, to the bactericidal action of either Polymyxin B or sodium dodecyl sulfate (SDS. Furthermore, a physical damage to the outer membrane of RB51SOD was observed by electron microscopy. Possibly due to the physical damage, overexpressed Cu/Zn SOD in RB51SOD was found to be released into the bacterial cell culture medium. Therefore, the stronger adaptive immunity induced by RB51SOD did not correlate with the low level of innate immunity induced by RB51SOD compared to RB51. This unique and apparently contradictory profile is likely associated with the differences in outer membrane integrity and Cu/Zn SOD release.

  18. Immune Responses and Protection against Experimental Brucella suis biovar 1 Challenge in Non-vaccinated or RB51-Vaccinated Cattle

    Science.gov (United States)

    Twenty Hereford heifers, approximately 9 months of age, were vaccinated with saline (control) or 2 x 10**10 CFU of Brucella abortus strain RB51 (RB51) vaccine. Immunologic responses after inoculation demonstrated significantly greater (P<0.05) antibody and proliferative responses to RB51 antigens i...

  19. Immune responses and safety after dart or booster vaccination of bison with Brucella abortus strain RB51

    Science.gov (United States)

    One alternative in the Bison remote vaccination environmental impact statement (EIS) for Yellowstone National Park includes inoculation of both calves and yearlings. Although RB51 vaccination of bison does protect against experimental challenge, it was unknown whether booster vaccination might enhan...

  20. Protective role of antibodies induced by Brucella melitensis B115 against B. melitensis and Brucella abortus infections in mice.

    Science.gov (United States)

    Adone, Rosanna; Francia, Massimiliano; Pistoia, Claudia; Petrucci, Paola; Pesciaroli, Michele; Pasquali, Paolo

    2012-06-01

    It has been demonstrated that antibodies specific for O-PS antigen of Brucella smooth strains are involved in the protective immunity of brucellosis. Since the rough strain Brucella melitensis B115 was able to protect mice against wild Brucella strains brucellosis despite the lack of anti-OPS antibodies, in this study we evaluated the biological significance of antibodies induced by this strain, directed to antigens other than O-PS, passively tranferred to untreated mice prior to infection with Brucella abortus 2308 and B. melitensis 16M virulent strains. The protective ability of specific antisera collected from mice vaccinated with B. melitensis B115, B. abortus RB51 and B. abortus S19 strains was compared. The results indicated that antibodies induced by B115 were able to confer a satisfactory protection, especially against B. abortus 2308, similar to that conferred by the antiserum S19, while the RB51 antiserum was ineffective. These findings suggest that antibodies induced by B115 could act as opsonins as well as antibodies anti-O-PS, thus triggering more efficient internalization and degradation of bacteria within phagocytes. This is the first study assessing the efficacy of antibodies directed to antigens other than O-PS in the course of brucellosis infection. PMID:22521283

  1. 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...

  2. Virulence Criteria for Brucella abortus Strains as Determined by Interferon Regulatory Factor 1-Deficient Mice

    OpenAIRE

    Ko, Jinkyung; Gendron-Fitzpatrick, Annette; Thomas A Ficht; Gary A Splitter

    2002-01-01

    Interferon regulatory factor 1-deficient (IRF-1−/−) mice infected with virulent Brucella abortus 2308 at 5 × 105 CFU developed acute hepatitis similar to many natural hosts but, unlike natural hosts, IRF-1−/− mice were unable to resolve infection and died. In contrast, IRF-1−/− mice survived when infected at 5 × 105 CFU with several attenuated Brucella strains (S19, RB51, cbp, and cyd). The survival of infected IRF-1−/− mice is likely a function of the level of virulence of each Brucella stra...

  3. Experimental infection of Richardson's ground squirrels (Spermophilus richardsonii) with attenuated and virulent strains of Brucella abortus.

    Science.gov (United States)

    Nol, Pauline; Olsen, Steven C; Rhyan, Jack C

    2009-01-01

    A previous investigation of the safety of Brucella abortus strain RB51 (sRB51) in various nontarget species suggested that Richardson's ground squirrels (Spermophilus richardsonii) may develop persistent infections when orally inoculated with the vaccine. In the present study, sRB51, B. abortus strain 19 (s19), and virulent B. abortus strain 9941 (s9941) were administered orally to Richardson's ground squirrels to further characterize B. abortus infection in this species. Six groups of nongravid ground squirrels were orally inoculated with 6 x 10(8) colony forming units (cfu) sRB51 (n = 10), 2.5 x 10(4) cfu s19 (n = 10), 2.5 x 10(7) cfu s19 (n = 6), 1.3 x 10(6) cfu s9941 (n = 5), 2.1 x 10(8) cfu s9941 (n = 5), or vaccine diluent (control; n = 4). One of five animals in the lower-dose s19 group and two of three animals in the higher-dose s19 group showed persistence of bacteria in various tissues at 14 wk postinoculation (PI). At 18 wk PI, one of five animals in the sRB51 group and one of five animals in the high-dose s9941 group were culture positive. Although we did detect some persistence of B. abortus strains at 18 wk, we found no evidence of pathology caused by B. abortus strains in nonpregnant Richardson's ground squirrels based on clinical signs, gross lesions, and microscopic lesions. PMID:19204348

  4. Novel vector vaccine against Brucella abortus based on influenza A viruses expressing Brucella L7/L12 or Omp16 proteins: evaluation of protection in pregnant heifers.

    Science.gov (United States)

    Tabynov, Kaissar; Yespembetov, Bolat; Sansyzbay, Abylai

    2014-10-14

    The present study provides the first information about the protection of a novel influenza viral vector vaccine expressing the Brucella proteins ribosomal L7/L12 or Omp16 containing the adjuvant Montanide Gel01 in pregnant heifers. Immunization of pregnant heifers was conducted via the conjunctival (n=10) or subcutaneous (n=10) route using cross prime and booster vaccination schedules at an interval of 28 days. The vector vaccine was evaluated in comparison with positive control groups vaccinated with Brucella abortus S19 (n=10) or B. abortus RB51 (n=10) and a negative (PBS+Montanide Gel01; n=10) control group. Via both the conjunctival or subcutaneous route, evaluation of protectiveness against abortion, effectiveness of vaccination and index of infection (in heifers and their fetuses or calves) demonstrated the vector vaccine provided good protection against B. abortus 544 infection compared to the negative control group (PBS+Montanide Gel01) and comparable protection to commercial vaccines B. abortus S19 or B. abortus RB51. PMID:25218295

  5. Safety of the novel influenza viral vector Brucella abortus vaccine in pregnant heifers

    Directory of Open Access Journals (Sweden)

    Kaissar Tabynov

    2016-01-01

    Full Text Available ABSTRACT: The present study provides the first information about the safety of a new influenza viral vector vaccine expressing the Brucella ribosomal protein L7/L12 or Omp16 containing the adjuvant Montanide Gel01 in pregnant heifers. Immunization of pregnant heifers was conducted via the conjunctival (n=10 or subcutaneous (n=10 route using cross prime and booster vaccination schedules at an interval of 28 days. The vector vaccine was evaluated in comparison with positive control groups vaccinated with B. abortus S19 (n=10 or B. abortus RB51 (n=10 and a negative (PBS+Montanide Gel01; n=10 control group. Clinical studies, thermometry, assessment of local reactogenicity and observation of abortion showed that the vector vaccine via the conjunctival or subcutaneous route was completely safe for pregnant heifers compared to the commercial vaccines B. abortus S19 or B. abortus RB51. The only single adverse event was the formation of infiltration at the site of subcutaneous injection; this reaction was not observed for the conjunctival route.

  6. 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

    Full Text Available Brucelosis, enfermedad causada por la bacteria intracelular facultativa Brucella abortus, es una zoonosis ampliamente distribuida a nivel mundial que afecta principalmente al ganado bovino, causando esterilidad en machos y abortos en hembras en gestación. La resistencia depende del desarrollo de una inmunidad mediada por células, con la participación de células T CD4+ de tipo Th1, que secreten interferón gama (-INF, citosina que estimula la actividad bactericida por macrófagos y la actividad citotóxica de linfocitos T CD8+, que son capaces de matar células infectadas con Brucella. Brucella posee como componentes antigénicos importantes el lipopolisacárido (LPS y las proteínas, entre las que se destaca por su demostrada capacidad inmune la superóxido dismutasa (SOD. La prevención de la diseminación de la brucelosis se fundamenta en el desarrollo de vacunas eficientes contra B. abortus, utilizándose cepas atenuadas de Brucella abortus como la cepa 19, cepa 45/20 y la cepa RB51; vacunas subcelulares en base a antígenos que forman parte de la estructura de la bacteria y vacunas basadas en moléculas de ácidos nucleicos, como las vacunas ADN y las vacunas ARN. En la presente revisión se pretende dar una visión actualizada sobre la brucelosis, su patogenia y cuadro clínico. Se hace un análisis de las características genéticas, antigénicas e inmunológicas de Brucella. Luego, una exposición de las vacunas actualmente en uso para su prevención y los estudios con vacunas subcelulares para finalizar con las nuevas tendencias en la generación de vacunas, como las vacunas ADN y ARN para Brucella.Brucellosis, a disease caused by the intracellular facultative bacteria Brucella abortus, is a widely distributed zoonosis throughout the world, affecting mainly cattle and causing sterility in males and abortion in pregnant females. Protection against B. abortus relies on the activation of cellular immunity involving lymphocytes T CD4

  7. 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. PMID:25644010

  8. Mutant Brucella abortus Membrane Fusogenic Protein Induces Protection against Challenge Infection in Mice

    Science.gov (United States)

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

    2015-01-01

    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. PMID:25644010

  9. Proteomic analysis of Brucella abortus cell envelope and identification of immunogenic candidate proteins for vaccine development.

    Science.gov (United States)

    Connolly, Joseph P; Comerci, Diego; Alefantis, Timothy G; Walz, Alexander; Quan, Marian; Chafin, Ryan; Grewal, Paul; Mujer, Cesar V; Ugalde, Rodolfo A; DelVecchio, Vito G

    2006-07-01

    Brucella abortus is the etiologic agent of bovine brucellosis and causes a chronic disease in humans known as undulant fever. In livestock the disease is characterized by abortion and sterility. Live, attenuated vaccines such as S19 and RB51 have been used to control the spread of the disease in animals; however, they are considered unsafe for human use and they induce abortion in pregnant cattle. For the development of a safer and equally efficacious vaccine, immunoproteomics was utilized to identify novel candidate proteins from B. abortus cell envelope (CE). A total of 163 proteins were identified using 2-DE with MALDI-TOF MS and LC-MS/MS. Some of the major protein components include outer-membrane protein (OMP) 25, OMP31, Omp2b porin, and 60 kDa chaperonin GroEL. 2-DE Western blot analyses probed with antiserum from bovine and a human patient infected with Brucella identified several new immunogenic proteins such as fumarate reductase flavoprotein subunit, F0F1-type ATP synthase alpha subunit, and cysteine synthase A. The elucidation of the immunome of B. abortus CE identified a number of candidate proteins for developing vaccines against Brucella infection in bovine and humans. PMID:16739129

  10. Deletion of znuA virulence factor attenuates Brucella abortus and confers protection against wild-type challenge.

    Science.gov (United States)

    Yang, Xinghong; Becker, Todd; Walters, Nancy; Pascual, David W

    2006-07-01

    znuA is known to be an important factor for survival and normal growth under low Zn(2+) concentrations for Escherichia coli, Haemophilus spp., Neisseria gonorrhoeae, and Pasteurella multocida. We hypothesized that the znuA gene present in Brucella melitensis 16 M would be similar to znuA in B. abortus and questioned whether it may also be an important factor for growth and virulence of Brucella abortus. Using the B. melitensis 16 M genome sequence, primers were designed to construct a B. abortus deletion mutant. A znuA knockout mutation in B. abortus 2308 (DeltaznuA) was constructed and found to be lethal in low-Zn(2+) medium. When used to infect macrophages, DeltaznuA B. abortus showed minimal growth. Further study with DeltaznuA B. abortus showed that its virulence in BALB/c mice was attenuated, and most of the bacteria were cleared from the spleen within 8 weeks. Protection studies confirmed the DeltaznuA mutant as a potential live vaccine, since protection against wild-type B. abortus 2308 challenge was as effective as that obtained with the RB51 or S19 vaccine strain. PMID:16790759

  11. Increases of efficacy as vaccine against Brucella abortus infection in mice by simultaneous inoculation with avirulent smooth bvrS/bvrR and rough wbkA mutants.

    Science.gov (United States)

    Grilló, María Jesús; Manterola, Lorea; de Miguel, María Jesús; Muñoz, Pilar María; Blasco, José María; Moriyón, Ignacio; López-Goñi, Ignacio

    2006-04-01

    The Brucella abortus S19 and RB51 strains are the most widely used live vaccines against bovine brucellosis. However, both can induce abortion and milk excretion, S19 vaccination interferes in serological tests, and RB51 is less effective. We have shown previously that a rough wbkAB. abortus mutant is attenuated and a better vaccine than RB51 in BALB/c mice, and that mutants in the two-component regulatory system bvrS/bvrR are markedly attenuated while keeping a smooth lipopolysaccharide (S-LPS). In this work, we tested whether simultaneous inoculation with live bvrS increases wbkA vaccine efficacy in mice. Even at high doses, the bvrS mutant was cleared much faster from spleens than the wbkA mutant. The splenic persistence of the wbkA mutant increased when inoculated along with the bvrS mutant, but also with inactivated bvrS cells or with purified B. abortus S-LPS, strongly suggesting that S-LPS in the bvrS mutant played a determinant role in the wbkA persistence. When inoculated alone, both mutants protected against virulent B. abortus but less than when inoculated simultaneously, and the protection afforded by the combination was better than that obtained with B. abortus S19. Increased protection was also obtained after simultaneous inoculation of the wbkA mutant and inactivated bvrS cells or purified S-LPS, showing again the role played by the S-LPS in the bvrS cells. In mice, the bvrS-wbkA combination induced an antibody response reduced with respect to B. abortus S19 vaccination. Thus, the simultaneous use of live bvrS and wbkA B. abortus mutants seems a promising approach to overcome the problems of the S19 andRB51 vaccines. PMID:16439039

  12. A potent Brucella abortus 2308 Δery live vaccine allows for the differentiation between natural and vaccinated infection.

    Science.gov (United States)

    Zhang, Junbo; Yin, Shuanghong; Guo, Fei; Meng, Ren; Chen, Chuangfu; Zhang, Hui; Li, Zhiqiang; Fu, Qiang; Shi, Huijun; Hu, Shengwei; Ni, Wei; Li, Tiansen; Zhang, Ke

    2014-08-01

    Brucellosis is a globally distributed zoonotic disease that causes animal and human diseases. However, the current Brucella abortus vaccines (S19 and RB51) are deficient; they can cause abortion in pregnant animals. Moreover, when the vaccine S19 is used, tests cannot differentiate natural from vaccinated infection. Therefore, a safer and more potent vaccine is needed. A Brucella abortus 2308 ery promoter mutant (Δery) was constructed to overcome these drawbacks. The growth of the Δery mutant was significantly attenuated in macrophages and mice and induced high protective immunity in mice. Moreover, Δery induced an anti-Brucella-specific IgG (immunoglobulin G) response and stimulated the expression of interferon-gamma (INF-γ) and interleukin-4 (IL-4). Furthermore, the expression of EryA antigen allowed for the serological differentiation between natural and vaccinated infection in mice. These results indicate that the Δery mutant is a potential attenuated live vaccine candidate against virulent Brucella abortus 2308 (S2308) infection. PMID:24994009

  13. Brucella abortus S19 genome sequenced, points toward virulence genes

    OpenAIRE

    Whyte, Barry James

    2008-01-01

    Researchers at the Virginia Bioinformatics Institute at Virginia Tech; the National Animal Disease Center in Ames, Iowa; and collaborators at 454 Life Sciences, Branford, Conn., have sequenced the genome of Brucella abortus strain S19.

  14. Adverse events in humans associated with accidental exposure to the livestock brucellosis vaccine RB51.

    Science.gov (United States)

    Ashford, David A; di Pietra, Jennifer; Lingappa, Jairam; Woods, Christopher; Noll, Heather; Neville, Bridget; Weyant, Robbin; Bragg, Sandra L; Spiegel, Richard A; Tappero, Jordan; Perkins, Bradley A

    2004-09-01

    Brucella abortus strain RB51 vaccine, is an attenuated live bacterial vaccine that was licensed conditionally by the Center for Veterinary Biologics, Veterinary Services, Animal and Plant Health Inspection Service, USDA, on 23 February 1996, for vaccination of cattle in the United States. Accidental human inoculations can occur during vaccination of cattle, and previous live Brucella vaccines designed for cattle have been known to cause brucellosis in humans. The Centers for Disease Control and Prevention (CDC) established passive surveillance for accidental inoculation with the RB51 vaccine in the United States to determine if this veterinary vaccine is associated with human disease, to describe the circumstances of accidental inoculation, to evaluate the potential efficacy of post-exposure chemoprophylaxis, and to develop recommendations for post-exposure management following exposure to RB51. Reports were received from 26 individuals. Accidental exposure to RB51 occurred by needle stick injury in 21 people (81%), conjunctival spray exposure in four (15%), and spray exposure of an open wound in one (4%) individual. At least one systemic symptom was reported in 19 (73%) people, including three (12%) who reported persistent local reactions with systemic involvement. One case required surgery, and B. abortus strain RB51 was isolated from the wound of that individual. Seven cases reported no adverse event associated with accidental exposure. Nine cases reported previous exposure to Brucella vaccines, including one case who also reported a previous diagnosis of brucellosis following exposure to S19 vaccine. Accidental needle stick injuries and conjunctival or open wound exposures of humans with the RB51 vaccine are associated with both local and systemic adverse events in the United States that are consistent with brucellosis; however, it remains undetermined if strain RB51 vaccine can cause systemic brucellosis in humans. Early culture attempts on those exposed and

  15. A combined vaccine against Brucella abortus and infectious bovine rhinotracheitis

    OpenAIRE

    Kamaraj, Govindasamy; Chinchkar, Shankar R.; Rajendra, Lingala; Srinivasan, Villuppanoor Alwar

    2009-01-01

    The present study was undertaken to study the immune response in calves vaccinated with Brucella abortus strain 19, infectious bovine rhinotracheitis (IBR) vaccines in monovalent form and combined vaccine containing both antigen. The seroconversion of monovalent and combined vaccines was tested in seronegative cattle calves. IBR vaccine alone and combination with live Brucella abortus S19 vaccine elicited an anamnestic response on day 60 post booster but started declining from day 90 onwards ...

  16. Genotipos de aislados de campo de Brucella abortus de distintas regiones geográficas de Chile Genotypes of Brucella abortus field isolates from different geographical regions of Chile

    Directory of Open Access Journals (Sweden)

    M Mancilla

    2008-01-01

    Full Text Available La brucelosis bovina es una enfermedad zoonótica, endémica de alto impacto económico. La identificación genética de las cepas prevalentes de Brucella abortus, el patógeno, es clave para establecer estrategias epidemiológicas de control de la enfermedad. La secuencia de inserción IS711 ha sido utilizada como un marcador genético para diferenciar entre especies de Brucella, miembros de una misma especie y dentro de un mismo biovar. Hemos analizado los perfiles de IS711-RFLP de 46 aislados de B. abortus, recolectados durante el periodo 1997-2005, provenientes de 16 áreas geográficas diferentes de Chile. Todos los aislados fueron previamente identificados como B. abortus biovar 1, utilizando las técnicas convencionales. De estos, el 87% compartieron el mismo perfil de IS711-RFLP, mientras que el 8,7% correspondió al patrón de la cepa vacuna RB51. En este trabajo se informa el hallazgo de dos cepas indistinguibles por PCR AMOS con perfiles nuevos de IS711-RFLP, no reportados previamente.Bovine brucellosis is an endemic, zoonotic disease of high economic impact. The genetic identification of the prevalent Brucella abortus strains, the pathogen, is key to pursue further epidemiological strategies for disease control. The insertion sequence IS711 has been used as genetic marker to differentiate among Brucella species, members of the same specie and within the same biovar. We have analyzed the IS711-RFLP pattern for 46 B. abortus isolates, collected during the period of 1997-2005 from 16 different geographical areas of Chile. All isolates were previously identified by conventional techniques as B. abortus biovar 1. Of these, 87% sharedthesame IS711 DNA profile, while an 8.7 % corresponded to the pattern of RB51 vaccine strain. We report the finding of two new strains, not differentiated by AMOS PCR, which showed unreported patterns of IS711-RFLP.

  17. Recombinant Brucella abortus gene expressing immunogenic protein

    Energy Technology Data Exchange (ETDEWEB)

    Mayfield, J.E.; Tabatabai, L.B.

    1991-06-11

    This patent describes a synthetic recombinant DNA molecule containing a DNA sequence. It comprises a gene of Brucella abortus encoding an immunogenic protein having a molecular weight of approximately 31,000 daltons as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis under denaturing conditions, the protein having an isoelectric point around 4.9, and containing a twenty-five amino acid sequence from its amino terminal end consisting of Gln-Ala-Pro-Thr-Phe-Phe-Arg-Ile-Gly-Thr-Gly-Gly-Thr-Ala-Gly-Thr-Tyr-Tyr-Pro-Ile-Gly-Gly-Leu-Ile-Ala, wherein Gln, Ala, Pro, Thr, Phe, Arg, Ile, Gly, Tyr, and Leu, respectively, represent glutamine, alanine, proline, threonine, phenylalanine, arginine, isolecuine, glycine, tyrosine, and leucine.

  18. 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.

  19. 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. PMID:24171573

  20. Inactivation of formyltransferase (wbkC) gene generates a Brucella abortus rough strain that is attenuated in macrophages and in mice.

    Science.gov (United States)

    Lacerda, Thaís Lourdes Santos; Cardoso, Patrícia Gomes; Augusto de Almeida, Leonardo; Camargo, Ilana Lopes Baratella da Cunha; Afonso, Daniela Almeida Freitas; Trant, Cyntia Cardoso; Macedo, Gilson Costa; Campos, Eleonora; Cravero, Silvio L; Salcedo, Suzana P; Gorvel, Jean-Pierre; Oliveira, Sérgio Costa

    2010-08-01

    Rough mutants of Brucella abortus were generated by disruption of wbkC gene which encodes the formyltransferase enzyme involved in LPS biosynthesis. In bone marrow-derived macrophages the B. abortusDeltawbkC mutants were attenuated, could not reach a replicative niche and induced higher levels of IL-12 and TNF-alpha when compared to parental smooth strains. Additionally, mutants exhibited attenuation in vivo in C57BL/6 and interferon regulatory factor-1 knockout mice. DeltawbkC mutant strains induced lower protective immunity in C56BL/6 than smooth vaccine S19 but similar to rough vaccine RB51. Finally, we demonstrated that Brucella wbkC is critical for LPS biosynthesis and full bacterial virulence. PMID:20580469

  1. 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.

  2. [Multiplication of Brucella abortus and production of nitric oxide in two macrophage cell lines of different origin].

    Science.gov (United States)

    Serafino, J; Conde, S; Zabal, O; Samartino, L

    2007-01-01

    Brucella abortus is a bacterium which causes abortions and infertility in cattle and undulant fever in humans. It multiplies intracellularly, evading the mechanisms of cellular death. Nitric oxide (NO) is important in the regulation of the immune response. In the present work, we studied the ability of three B. abortus strains to survive intracellularly in two macrophage cell lines. The bacterial multiplication in both cell lines was determined at two different times in UFC/ ml units. Moreover the inoculated cells were also observed under light-field and fluorescence microscopy stained with Giemsa and acridine orange, respectively. The stain of both cellular lines showed similar results with respect to the UFC/ml determination. The presence of B. abortus was confirmed by electronic microscopy. In both macrophage cell lines inoculated with the rough strain RB51, the multiplication diminished and the level of NO was higher, compared with cells inoculated with smooth strains (S19 and 2308). These results suggest that the absence of O-chain of LPS probably affects the intracellular growth of B. abortus. PMID:18390151

  3. Endocarditis por Brucella abortus: Reporte del primer caso en C.R Brucella abortus Endocarditis

    Directory of Open Access Journals (Sweden)

    Manuel Antonio Villalobos-Zúñiga

    2011-09-01

    Full Text Available Paciente masculino de 36 años de edad, proveniente de la zona rural de Costa Rica, con un cuadro clínico de 8 meses de evolución de fiebre, mialgias, artralgias, pérdida de peso y lumbalgia; referido por la detección de un soplo de insuficiencia aórtica. El ecocardiograma reveló endocarditis de la válvula aórtica, y se obtuvieron 4 hemocultivos positivos por Brucella abortus biotipo 3, con serologías negativas por brucelosis. Se inició tratamiento con antibióticos y luego se le realizó un reemplazo valvular aórtico; 4 meses después ingresó con dolor torácico que se atribuyó a una oclusión de la arteria descendente anterior, demostrada angiográficamente, por posible embolismo. En la actualidad cursa clínicamente estable con manejo médico para su cardiopatía, sin recaída infecciosa.The case of a 36-year-old patient from a rural area is presented. He came with an 8 month history of fever, myalgias, arthralgias, weight loss and lower back pain; who also had an aortic insufficiency murmur detected. The diagnosis of aortic valve endocarditis was made by echocardiography, and had 4 positive blood cultures for Brucella abortus biotype 3, and negative serologic test for brucellosis. He was started on antibiotics and later on underwent aortic valve replacement, with a late coronary cardioembolism as a complication.

  4. Influence of Brucella abortus lipopolysaccharide as an adjuvant on the immunogenicity of HPV-16 L1VLP vaccine in mice.

    Science.gov (United States)

    Kianmehr, Zahra; Soleimanjahi, Hoorieh; Ardestani, Susan Kaboudanian; Fotouhi, Fatemeh; Abdoli, Asghar

    2015-04-01

    Brucella abortus lipopolysaccharide (LPS) has less toxicity and no pyrogenic properties in comparison with other bacterial LPS. It is a toll-like receptor 4 agonist and has been shown to have the potential use as a vaccine adjuvant. In this study, the immunostimulatory properties of LPS from smooth and rough strains of B. abortus (S19 and RB51) as adjuvants were investigated for the human papillomavirus type 16 (HPV16) L1 virus-like particles (L1VLPs) vaccines. C57BL/6 mice were immunized subcutaneously three times either with HPV-16 L1VLPs alone, or in combination with smooth LPS (S-LPS), rough LPS (R-LPS), aluminum hydroxide or a mixture of them as adjuvant. The humoral immunity was evaluated by measuring the specific and total IgG levels, and also the T-cell immune response of mice was evaluated by measuring different cytokines such as IFN-γ, TNF-α, IL-4, IL-10 and IL-17. Results showed that serum anti-HPV16 L1VLP IgG antibody titers was significantly higher in mice immunized with a combination of VLPs and R-LPS or S-LPS compared with other immunized groups. Co-administration of HPV-16 L1VLPs with R-LPS elicited the highest levels of splenocytes cytokines (IFN-γ, IL-4, IL-17 and TNF-α) and also effectively induced improvement of a Th1-type cytokine response characterized with a high ratio of IFN-γ/IL-10. The data indicate that B. abortus LPS particularly RB51-LPS enhances the immune responses to HPV-16 L1VLPs and suggests its potential as an adjuvant for the development of a potent prophylactic HPV vaccine and other candidate vaccines.

  5. Influence of Brucella abortus lipopolysaccharide as an adjuvant on the immunogenicity of HPV-16 L1VLP vaccine in mice.

    Science.gov (United States)

    Kianmehr, Zahra; Soleimanjahi, Hoorieh; Ardestani, Susan Kaboudanian; Fotouhi, Fatemeh; Abdoli, Asghar

    2015-04-01

    Brucella abortus lipopolysaccharide (LPS) has less toxicity and no pyrogenic properties in comparison with other bacterial LPS. It is a toll-like receptor 4 agonist and has been shown to have the potential use as a vaccine adjuvant. In this study, the immunostimulatory properties of LPS from smooth and rough strains of B. abortus (S19 and RB51) as adjuvants were investigated for the human papillomavirus type 16 (HPV16) L1 virus-like particles (L1VLPs) vaccines. C57BL/6 mice were immunized subcutaneously three times either with HPV-16 L1VLPs alone, or in combination with smooth LPS (S-LPS), rough LPS (R-LPS), aluminum hydroxide or a mixture of them as adjuvant. The humoral immunity was evaluated by measuring the specific and total IgG levels, and also the T-cell immune response of mice was evaluated by measuring different cytokines such as IFN-γ, TNF-α, IL-4, IL-10 and IL-17. Results showed that serum anti-HPV16 L1VLP IgG antibody titers was significantly higher in mice immunized with a combination of VLPs and R-LPS or S-LPS compared with other immunized groups. Co-administration of HPV-16 L1VLPs with R-LPS elicited the highest levels of splenocytes cytokines (IFN-γ, IL-4, IL-17 and TNF-α) and also effectively induced improvement of a Th1-type cytokine response characterized with a high ratio of IFN-γ/IL-10. The data indicate that B. abortus LPS particularly RB51-LPS enhances the immune responses to HPV-16 L1VLPs and suggests its potential as an adjuvant for the development of a potent prophylactic HPV vaccine and other candidate vaccines. PMID:25187406

  6. The Effect of Irradiation on the Immunogenity of Brucella Abortus

    International Nuclear Information System (INIS)

    An experiment was carried out to study the effect of irradiation on the immunogenity of B. abortus. The B. abortus were irradiated by Gamma Cells (60Co). An experiment were divided into four groups. The first group (V1) was inoculated by irradiated B. abortus with the dose of 0.25 kGy. The second group (V2) was inoculated by irradiated B. abortus with the dose of 0.50 kGy. The third group (V3) was inoculated by irradiated B. abortus with the dose of 0.75 kGy. The fourth group (V4) was inoculated by Brucella vaccine 8.19. The observation respectively were included purely test, safety test, RBT serological test, diffusion test, development the colony of B. abortus in lien, and pathology anatomic inspection. The results obtained showed that 0.25 kGy was the expectantly dose of irradiation which could not only decreasing the infectivity of B. abortus but also has the ability to become a good immunogen for stimulating the immune response in the experiment animals. (author)

  7. A combined vaccine against Brucella abortus and infectious bovine rhinotracheitis.

    Science.gov (United States)

    Kamaraj, Govindasamy; Chinchkar, Shankar R; Rajendra, Lingala; Srinivasan, Villuppanoor Alwar

    2009-06-01

    The present study was undertaken to study the immune response in calves vaccinated with Brucella abortus strain 19, infectious bovine rhinotracheitis (IBR) vaccines in monovalent form and combined vaccine containing both antigen. The seroconversion of monovalent and combined vaccines was tested in seronegative cattle calves. IBR vaccine alone and combination with live Brucella abortus S19 vaccine elicited an anamnestic response on day 60 post booster but started declining from day 90 onwards against IBR. B. abortus S19 alone and in combination with IBR vaccine gave more than 2 log protection in mice two weeks post challenge. Fluorescence polarization assay analysis with sera samples of calves vaccinated with B. abortus S19 monovalent vaccine alone and in combination with IBR vaccine revealed the presence of B. abortus antibodies. The components of the combined vaccine did not show any evidence of interference in the development of immunity. This combined vaccine may provide economical and affordable biological for the control of brucellosis and IBR. PMID:23100765

  8. 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.

  9. Immunological response to the Brucella abortus GroEL homolog.

    OpenAIRE

    Lin, J.; Adams, L G; Ficht, T A

    1996-01-01

    Western blot (immunoblot) analysis of sera from cattle vaccinated with Brucella abortus S19 exhibit an elevated serologic response to Hsp62, the GroEL homolog (BaGroEL). Serologic screening of individual cows vaccinated with B. abortus S19 revealed no correlation between the immune response to BaGroEL and protection against a challenge with virulent organisms. The humoral immune response to BaGroEL was restricted to a region of the mature protein which mapped to amino acids 317 to 355 and may...

  10. Oral immunization of mice with gamma-irradiated Brucella neotomae induces protection against intraperitoneal and intranasal challenge with virulent B. abortus 2308.

    Science.gov (United States)

    Dabral, Neha; Martha-Moreno-Lafont; Sriranganathan, Nammalwar; Vemulapalli, Ramesh

    2014-01-01

    Brucella spp. are Gram-negative, facultative intracellular coccobacilli that cause one of the most frequently encountered zoonosis worldwide. Humans naturally acquire infection through consumption of contaminated dairy and meat products and through direct exposure to aborted animal tissues and fluids. No vaccine against brucellosis is available for use in humans. In this study, we tested the ability of orally inoculated gamma-irradiated B. neotomae and B. abortus RB51 in a prime-boost immunization approach to induce antigen-specific humoral and cell mediated immunity and protection against challenge with virulent B. abortus 2308. Heterologous prime-boost vaccination with B. abortus RB51 and B. neotomae and homologous prime-boost vaccination of mice with B. neotomae led to the production of serum and mucosal antibodies specific to the smooth LPS. The elicited serum antibodies included the isotypes of IgM, IgG1, IgG2a, IgG2b and IgG3. All oral vaccination regimens induced antigen-specific CD4(+) and CD8(+) T cells capable of secreting IFN-γ and TNF-α. Upon intra-peritoneal challenge, mice vaccinated with B. neotomae showed the highest level of resistance against virulent B. abortus 2308 colonization in spleen and liver. Experiments with different doses of B. neotomae showed that all tested doses of 10(9), 10(10) and 10(11) CFU-equivalent conferred significant protection against the intra-peritoneal challenge. However, a dose of 10(11) CFU-equivalent of B. neotomae was required for affording protection against intranasal challenge as shown by the reduced bacterial colonization in spleens and lungs. Taken together, these results demonstrate the feasibility of using gamma-irradiated B. neotomae as an effective and safe oral vaccine to induce protection against respiratory and systemic infections with virulent Brucella.

  11. Studies on recombinant glucokinase (r-glk) protein of Brucella abortus as a candidate vaccine molecule for brucellosis.

    Science.gov (United States)

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

    2014-09-29

    Brucellosis is one of the most prevalent zoonotic diseases of worldwide distribution caused by the infection of genus Brucella. Live attenuated vaccines such as B. abortus S19, B. abortus RB51 and B. melitensis Rev1 are found most effective against brucellosis infection in animals, contriving a number of serious side effects and having chances to revert back into their active pathogenic form. In order to engineer a safe and effective vaccine candidate to be used in both animals and human, a recombinant subunit vaccine molecule comprising the truncated region of glucokinase (r-glk) gene from B. abortus S19 was cloned and expressed in Escherichia coli BL21DE3 host. Female BALB/c mice immunized with purified recombinant protein developed specific antibody titer of 1:64,000. The predominant IgG2a and IgG2b isotypes signified development of Th1 directed immune responses. In vitro cell cytotoxicity assay using anti-r-glk antibodies incubated with HeLa cells showed 81.20% and 78.5% cell viability against lethal challenge of B. abortus 544 and B. melitensis 16M, respectively. The lymphocyte proliferative assay indicated a higher splenic lymphocyte responses at 25μg/ml concentration of protein which implies the elevated development of memory immune responses. In contrast to control, the immunized group of mice intra-peritoneal (I.P.) challenged with B. abortus 544 were significantly protected with no signs of necrosis and vacuolization in their liver and spleen tissue. The elevated B-cell response associated with Th1 adopted immunity, significant in vitro cell viability as well as protection afforded in experimental animals after challenge, supplemented with histopathological analysis are suggestive of r-glk protein as a prospective candidate vaccine molecule against brucellosis. PMID:25131740

  12. Mass vaccination as a complementary tool in the control of a severe outbreak of bovine brucellosis due to Brucella abortus in Extremadura, Spain.

    Science.gov (United States)

    Sanz, Cristina; Sáez, José Luis; Alvarez, Julio; Cortés, María; Pereira, Gema; Reyes, Aurelia; Rubio, Félix; Martín, Javier; García, Nerea; Domínguez, Lucas; Hermoso-de-Mendoza, María; Hermoso-de-Mendoza, Javier

    2010-11-01

    We report the evolution of an outbreak of bovine brucellosis (Brucella abortus) in the region of Extremadura (Spain) involving more than 1000 herds and nearly 40,000 animals. S19 vaccination of young cattle combined with a test and slaughter strategy did not result in a rapid decrease in herd prevalence and animal incidence; these parameters showed a constant decreasing trend only when a combination of restriction of cattle movements, increased test frequency, S19 vaccination and mass RB51 vaccination (with yearly revaccinations) were applied to all susceptible populations. These measures were applied for 5 years; abortions following RB51 vaccination of pregnant cows were limited to the first inoculation and the involvement of the vaccine strain could only be demonstrated in 78 out of 897 abortions. Our results demonstrate the usefulness - and lack of significant side effects - of RB51 mass vaccination as a complementary tool to control bovine brucellosis outbreaks in areas where the disease cannot be contained using more conservative approaches. PMID:20833439

  13. Plasmid transfer and plasmid-mediated genetic exchange in Brucella abortus.

    OpenAIRE

    Rigby, C E; Fraser, A.D.

    1989-01-01

    Naturally-occurring plasmids and gene transfer mechanisms have not yet been reported in brucellae. Here we show that Brucella abortus is capable of maintaining and transferring the broad-host-range plasmids pTH10 (IncP), pSa (IncW) and R751 (IncP), and describe pTH10-mediated transfer of B. abortus chromosomal genes to Escherichia coli. All three plasmids transferred by conjugation from E. coli to B. abortus S19, and from B. abortus S19 to B. abortus 292 (biovar 4). They were stably maintaine...

  14. Immunization with recombinant Brucella species outer membrane protein Omp16 or Omp19 in adjuvant induces specific CD4+ and CD8+ T cells as well as systemic and oral protection against Brucella abortus infection.

    Science.gov (United States)

    Pasquevich, Karina A; Estein, Silvia M; García Samartino, Clara; Samartino, Clara García; Zwerdling, Astrid; Coria, Lorena M; Barrionuevo, Paula; Fossati, Carlos A; Giambartolomei, Guillermo H; Cassataro, Juliana

    2009-01-01

    Available vaccines against Brucella spp. are live attenuated Brucella strains. In order to engineer a better vaccine to be used in animals and humans, our laboratory aims to develop an innocuous subunit vaccine. Particularly, we are interested in the outer membrane proteins (OMPs) of B. abortus: Omp16 and Omp19. In this study, we assessed the use of these proteins as vaccines against Brucella in BALB/c mice. Immunization with lipidated Omp16 (L-Omp16) or L-Omp19 in incomplete Freund's adjuvant (IFA) conferred significant protection against B. abortus infection. Vaccination with unlipidated Omp16 (U-Omp16) or U-Omp19 in IFA induced a higher degree of protection than the respective lipidated versions. Moreover, the level of protection induced after U-Omp16 or U-Omp19 immunization in IFA was similar to that elicited by live B. abortus S19 immunization. Flow cytometric analysis showed that immunization with U-Omp16 or U-Omp19 induced antigen-specific CD4(+) as well as CD8(+) T cells producing gamma interferon. In vivo depletion of CD4(+) or CD8(+) T cells in mice immunized with U-Omp16 or U-Omp19 plus IFA resulted in a loss of the elicited protection, indicating that both cell types are mediating immune protection. U-Omp16 or U-Omp19 vaccination induced a T helper 1 response, systemic protection in aluminum hydroxide formulation, and oral protection with cholera toxin adjuvant against B. abortus infection. Both immunization routes exhibited a similar degree of protection to attenuated Brucella vaccines (S19 and RB51, respectively). Overall these results indicate that U-Omp16 or U-Omp19 would be a useful candidate for a subunit vaccine against human and animal brucellosis. PMID:18981242

  15. Immunization with Recombinant Brucella Species Outer Membrane Protein Omp16 or Omp19 in Adjuvant Induces Specific CD4+ and CD8+ T Cells as Well as Systemic and Oral Protection against Brucella abortus Infection▿

    Science.gov (United States)

    Pasquevich, Karina A.; Estein, Silvia M.; Samartino, Clara García; Zwerdling, Astrid; Coria, Lorena M.; Barrionuevo, Paula; Fossati, Carlos A.; Giambartolomei, Guillermo H.; Cassataro, Juliana

    2009-01-01

    Available vaccines against Brucella spp. are live attenuated Brucella strains. In order to engineer a better vaccine to be used in animals and humans, our laboratory aims to develop an innocuous subunit vaccine. Particularly, we are interested in the outer membrane proteins (OMPs) of B. abortus: Omp16 and Omp19. In this study, we assessed the use of these proteins as vaccines against Brucella in BALB/c mice. Immunization with lipidated Omp16 (L-Omp16) or L-Omp19 in incomplete Freund's adjuvant (IFA) conferred significant protection against B. abortus infection. Vaccination with unlipidated Omp16 (U-Omp16) or U-Omp19 in IFA induced a higher degree of protection than the respective lipidated versions. Moreover, the level of protection induced after U-Omp16 or U-Omp19 immunization in IFA was similar to that elicited by live B. abortus S19 immunization. Flow cytometric analysis showed that immunization with U-Omp16 or U-Omp19 induced antigen-specific CD4+ as well as CD8+ T cells producing gamma interferon. In vivo depletion of CD4+ or CD8+ T cells in mice immunized with U-Omp16 or U-Omp19 plus IFA resulted in a loss of the elicited protection, indicating that both cell types are mediating immune protection. U-Omp16 or U-Omp19 vaccination induced a T helper 1 response, systemic protection in aluminum hydroxide formulation, and oral protection with cholera toxin adjuvant against B. abortus infection. Both immunization routes exhibited a similar degree of protection to attenuated Brucella vaccines (S19 and RB51, respectively). Overall these results indicate that U-Omp16 or U-Omp19 would be a useful candidate for a subunit vaccine against human and animal brucellosis. PMID:18981242

  16. Seroprevalence of Brucella abortus and Leptospira hardjo in cattle

    Directory of Open Access Journals (Sweden)

    S. Jegaveera Pandian

    2015-02-01

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

  17. A repA-based ELISA for discriminating cattle vaccinated with Brucella suis 2 from those naturally infected with Brucella abortus and Brucella melitensis.

    Science.gov (United States)

    Wang, Jing-Yu; Wu, Ning; Liu, Wan-Hua; Ren, Juan-Juan; Tang, Pan; Qiu, Yuan-Hao; Wang, Chi-Young; Chang, Ching-Dong; Liu, Hung-Jen

    2014-01-01

    The commonest ways of diagnosing brucellosis in animals include the Rose-Bengal plate agglutination test, the buffered plate agglutination test (BPA), the slide agglutination test, the complement fixation test, and the indirect enzyme linked immunosorbent assay (I-ELISA). However, these methods cannot discriminate the Brucella vaccine strain (Brucella suis strain 2; B. suis S2) from naturally acquired virulent strains. Of the six common Brucella species, Brucella melitensis, Brucella abortus, and B. suis are the commonest species occurring in China. To develop an ELISA assay that can differentiate between cows inoculated with B. suis S2 and naturally infected with B. abortus and B. melitensis, genomic sequences from six Brucella spp. (B. melitensis, B. abortus, B. suis, Brucella canis, Brucella neotomae and Brucella ovis) were compared using Basic Local Alignment Search Tool software. One particular gene, the repA-related gene, was found to be a marker that can differentiate B. suis from B. abortus and B. melitensis. The repA-related gene of B. suis was PCR amplified and subcloned into the pET-32a vector. Expressed repA-related protein was purified and used as an antigen. The repA-based ELISA was optimized and used as specific tests. In the present study, serum from animals inoculated with the B. suis S2 vaccine strain had positive repA-based ELISA results. In contrast, the test-positive reference sera against B. abortus and B. melitensis had negative repA-based ELISA results. The concordance rate between B. abortus antibody-negative (based on the repA-based ELISA) and the Brucella gene-positive (based on the 'Bruce ladder' multiplex PCR) was 100%. Therefore, the findings suggest that the repA-based ELISA is a useful tool for differentiating cows vaccinated with the B. suis S2 and naturally infected with B. abortus and B. melitensis. PMID:24941369

  18. Desenvolvimento e avaliação de uma cepa knockout de Brucella abortus obtida pela deleção do gene virB10 Development and evaluation of a strain of Brucella abortus gotten by the knockout of the virB10 gene

    Directory of Open Access Journals (Sweden)

    Fabiane G. de Souza

    2009-11-01

    Full Text Available Brucella spp. são bactérias gram-negativas, intracelulares facultativas que são patogênicas para muitas espécies de mamíferos causando a brucelose, uma zoonose difundida mundialmente. Por isso a busca de alternativas de controle mais eficientes se faz necessário como o desenvolvimento de novas cepas que possam ser testadas como potenciais imunógenos. Neste estudo realizou-se a deleção do gene virB10 da cepa S2308 de Brucella abortus gerando uma cepa knockout provavelmente incapaz de produzir a proteína nativa correspondente. O gene virB10 faz parte de um operon que codifica para um sistema de secreção do tipo IV, essencial para a sobrevivência intracelular e multiplicação da bactéria em células hospedeiras. A deleção foi realizada pela construção do plasmídeo suicida pBlue:virB10:kan e eletroporação deste em células eletrocompetentes de B. abortus S2308, ocorrendo a troca do gene selvagem pelo gene interrompido, com o gene de resistência a canamicina, por recombinação homóloga dupla. Camundongos BALB/c foram inoculados com as cepas S19, RB-51, ΔvirB10 de B. abortus e B. abortus S2308 selvagem; os resultados demonstraram que camundongos BALB/c inoculados com S19 e camundongos BALB/c inoculados com S2308 apresentaram queda mais rápida de linha de tendência, quando comparadas aos demais grupos, para recuperação bacteriana (RB e peso esplênico (PE respectivamente. Os grupos que receberam ΔvirB10 S2308 de B. abortus e RB-51 demonstraram comportamento semelhante para ambas as características. Na sexta semana após a inoculação, os resultados para RB (log de UFC ± desvio padrão e PE (peso esplênico ± desvio padrão, respectivamente, mostraram: grupos inoculados com as cepas S2308 (4,44±1,97 e 0,44±0,11, S19 (1,83±2,54 e 0,31±0,04, RB-51 (0,00±0,00 e 0,20±0,01 e ΔvirB10 S2308 (1,43±1,25 e 0,19±0,03. Considerado o clearance bacteriano, todos os grupos diferiram estatisticamente do grupo que recebeu S

  19. Deletion of the BCSP31 gene of Brucella abortus by replacement.

    OpenAIRE

    Halling, S. M.; Detilleux, P G; Tatum, F M; Judge, B A; Mayfield, J E

    1991-01-01

    The 31-kDa salt-extractable immunogenic protein, BCSP31, was deleted from several Brucella abortus strains by replacement with a marker gene encoding resistance to the antibiotics kanamycin and neomycin. The BCSP31 gene replacement plasmids, constructed with ColE1-derived vectors, were introduced by electroporation into B. abortus strain 19 (S19), into a rough variant of B. abortus S19, and into B. abortus S2308, and antibiotic-resistant transformants were isolated. B. abortus S19 is an atten...

  20. Different resistance patterns of reference and field strains of Brucella abortus

    Directory of Open Access Journals (Sweden)

    Karina L. Miranda

    2015-03-01

    Full Text Available The aim of this study was to evaluate the growth of the B. abortus reference strains and field isolates on media containing different inhibitor agents. Reference strains were seeded on tryptose agar containing: i-erythritol (1.0 mg/mL, fuchsin (20 μg/mL and 80 μg/mL, thionin (2.5 μg/mL and 10 μg/mL, rifampicin (200 μg/mL and safranin O (200 μg/mL. Field isolates were tested only on media containing i-erythritol, rifampicin and thionin. Furthermore, each suspension was also inoculated on tryptose agar incubated in air, to test its ability to grow without CO2. Sensitivity to fuchsin was similar among reference strains evaluated. Growth of S19, 544 and 2308 but not RB51 were inhibited on media containing rifampicin. Medium with safranin O showed no inhibition for RB51, 544 and 2308, but it partially inhibited the S19 growth as well as medium containing i-erythritol. Treatment/control growth ratio for 2308 on tryptose agar containing thionin (2.5 μg/mL was approximatelly 1.0, whereas S19 and RB51 showed 0.85 and 0.89 ratios, respectively. Growth of 544, S19 and RB51 but not 2308 was completely inhibited on medium with thionin (10 μg/mL. All field strains grew on medium containing i-erythritol, but were completelly inhibited by rifampicin. With exception of A1 (B. abortus biovar 3 all field isolates grew on medium with thionin, although some strains showed a treatment/control growth ratio of 0.75–0.80 (10 μg/mL. These results showed that tryptose agar with thionin, i-erythritol or rifampicin could be useful for differentiating vaccine, challenge and field strains of B. abortus.

  1. Different resistance patterns of reference and field strains of Brucella abortus

    Science.gov (United States)

    Miranda, Karina L.; Dorneles, Elaine M. S.; Poester, Fernando P.; Martins, Paulo S.; Pauletti, Rebeca B.; Lage, Andrey P.

    2015-01-01

    The aim of this study was to evaluate the growth of the B. abortus reference strains and field isolates on media containing different inhibitor agents. Reference strains were seeded on tryptose agar containing: i-erythritol (1.0 mg/mL), fuchsin (20 μg/mL and 80 μg/mL), thionin (2.5 μg/mL and 10 μg/mL), rifampicin (200 μg/mL) and safranin O (200 μg/mL). Field isolates were tested only on media containing i-erythritol, rifampicin and thionin. Furthermore, each suspension was also inoculated on tryptose agar incubated in air, to test its ability to grow without CO 2 . Sensitivity to fuchsin was similar among reference strains evaluated. Growth of S19, 544 and 2308 but not RB51 were inhibited on media containing rifampicin. Medium with safranin O showed no inhibition for RB51, 544 and 2308, but it partially inhibited the S19 growth as well as medium containing i-erythritol. Treatment/control growth ratio for 2308 on tryptose agar containing thionin (2.5 μg/mL) was approximatelly 1.0, whereas S19 and RB51 showed 0.85 and 0.89 ratios, respectively. Growth of 544, S19 and RB51 but not 2308 was completely inhibited on medium with thionin (10 μg/mL). All field strains grew on medium containing i-erythritol, but were completelly inhibited by rifampicin. With exception of A1 ( B. abortus biovar 3) all field isolates grew on medium with thionin, although some strains showed a treatment/control growth ratio of 0.75–0.80 (10 μg/mL). These results showed that tryptose agar with thionin, i-erythritol or rifampicin could be useful for differentiating vaccine, challenge and field strains of B. abortus. PMID:26221116

  2. Different resistance patterns of reference and field strains of Brucella abortus.

    Science.gov (United States)

    Miranda, Karina L; Dorneles, Elaine M S; Poester, Fernando P; Martins Filho, Paulo S; Pauletti, Rebeca B; Lage, Andrey P

    2015-03-01

    The aim of this study was to evaluate the growth of the B. abortus reference strains and field isolates on media containing different inhibitor agents. Reference strains were seeded on tryptose agar containing: i-erythritol (1.0 mg/mL), fuchsin (20 μg/mL and 80 μg/mL), thionin (2.5 μg/mL and 10 μg/mL), rifampicin (200 μg/mL) and safranin O (200 μg/mL). Field isolates were tested only on media containing i-erythritol, rifampicin and thionin. Furthermore, each suspension was also inoculated on tryptose agar incubated in air, to test its ability to grow without CO 2 . Sensitivity to fuchsin was similar among reference strains evaluated. Growth of S19, 544 and 2308 but not RB51 were inhibited on media containing rifampicin. Medium with safranin O showed no inhibition for RB51, 544 and 2308, but it partially inhibited the S19 growth as well as medium containing i-erythritol. Treatment/control growth ratio for 2308 on tryptose agar containing thionin (2.5 μg/mL) was approximatelly 1.0, whereas S19 and RB51 showed 0.85 and 0.89 ratios, respectively. Growth of 544, S19 and RB51 but not 2308 was completely inhibited on medium with thionin (10 μg/mL). All field strains grew on medium containing i-erythritol, but were completelly inhibited by rifampicin. With exception of A1 ( B. abortus biovar 3) all field isolates grew on medium with thionin, although some strains showed a treatment/control growth ratio of 0.75-0.80 (10 μg/mL). These results showed that tryptose agar with thionin, i-erythritol or rifampicin could be useful for differentiating vaccine, challenge and field strains of B. abortus. PMID:26221116

  3. 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.

  4. 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.

  5. Soroepidemiologia da brucelose canina causada por Brucella canis e Brucella abortus na cidade de Alfenas, MG Seroepidemiology of canine brucellosis caused by Brucella canis and Brucella abortus in Alfenas, MG, Brazil

    Directory of Open Access Journals (Sweden)

    A.C. Almeida

    2004-04-01

    Full Text Available The prevalence of canine brucellosis was evaluated in the city of Alfenas, MG through the technique of agarose gel imunodifusion for Brucella canis and slow serum agglutination test with 2-mercaptoetanol for Brucella abortus. The prevalence was of 14.2% and 2.8%, respectively, for B. canis and B. abortus. The positives, characterized by animals above one year of age (77.8%, and mongrel dogs (56.2%, showed a prevalence of 50 and 48% for males and females, respectively. The canine brucellosis was prevalent in the city principally in dogs of outskirts.

  6. DeltaznuADeltapurE Brucella abortus 2308 mutant as a live vaccine candidate.

    Science.gov (United States)

    Yang, Xinghong; Thornburg, Theresa; Walters, Nancy; Pascual, David W

    2010-01-22

    To create a new, safe brucellosis live vaccine, a double mutant strain was constructed from Brucella abortus 2308. Using the DeltaznuA B. abortus 2308 mutant, a second mutation was introduced by deleting purE gene. The DeltaznuA DeltapurE B. abortus 2308 strain was less capable of surviving in macrophages. When evaluated in vivo, it was cleared within 8 weeks (wks) from mice, causing significantly less inflammation than spleens obtained from wild-type B. abortus 2308-infected mice. Furthermore, two doses of DeltaznuA DeltapurE B. abortus 2308 conferred 0.79 log protection, similar to S19 as did a single dose of DeltaznuA B. abortus 2308. Thus, this study shows the DeltaznuA DeltapurE B. abortus 2308 strain to be a potential livestock vaccine candidate. PMID:19914192

  7. Virulent Brucella abortus Prevents Lysosome Fusion and Is Distributed within Autophagosome-Like Compartments

    OpenAIRE

    Pizarro-Cerdá, Javier; Moreno, Edgardo; Sanguedolce, Veronique; Mege, Jean-Louis; Gorvel, Jean-Pierre

    1998-01-01

    Virulent and attenuated Brucella abortus strains attach to and penetrate nonprofessional phagocytic HeLa cells. Compared to pathogenic Brucella, the attenuated strain 19 hardly replicates within cells. The majority of the strain 19 bacteria colocalized with the lysosome marker cathepsin D, suggesting that Brucella-containing phagosomes had fused with lysosomes, in which they may have degraded. The virulent bacteria prevented lysosome-phagosome fusion and were found distributed in the perinucl...

  8. Epitope Mapping of the Brucella melitensis BP26 Immunogenic Protein: Usefulness for Diagnosis of Sheep Brucellosis

    OpenAIRE

    Seco-Mediavilla, Patricia; Verger, Jean-Michel; Grayon, Maggy; Cloeckaert, Axel; Marín, Clara M.; Zygmunt, Michel S; Fernández-Lago, Luis; Vizcaíno, Nieves

    2003-01-01

    Sequencing of bp26, the gene encoding the Brucella sp. immunogenic BP26 periplasmic protein, was performed in the reference strains of Brucella abortus, B. suis, and B. ovis. The three bp26 sequences were almost identical to that published for B. melitensis 16M bp26, and only minor nucleotide substitutions, without modifying the amino acid sequence, were observed between species. The bp26 genes of the seven B. abortus biovar reference strains and B. abortus S19 and RB51 vaccine strains were a...

  9. Diagnostic characterization of a feral swine herd enzootically infected with Brucella.

    Science.gov (United States)

    Stoffregen, William C; Olsen, Steven C; Jack Wheeler, C; Bricker, Betsy J; Palmer, Mitchell V; Jensen, Allen E; Halling, Shirley M; Alt, David P

    2007-05-01

    Eighty feral swine were trapped from a herd that had been documented to be seropositive for Brucella and which had been used for Brucella abortus RB51 vaccine trials on a 7,100-hectare tract of land in South Carolina. The animals were euthanized and complete necropsies were performed. Samples were taken for histopathology, Brucella culture, and Brucella serology. Brucella was cultured from 62 (77.5%) animals. Brucella suis was isolated from 55 animals (68.8%), and all isolates were biovar 1. Brucella abortus was isolated from 28 animals (35.0%), and isolates included field strain biovar 1 (21 animals; 26.3%), vaccine strain Brucella abortus S19 (8 animals, 10.0%), and vaccine strain Brucella abortus RB51 (6 animals, 7.5%). Males were significantly more likely to be culture positive than females (92.9% vs. 60.6%). Thirty-nine animals (48.8%) were seropositive. Males also had a significantly higher seropositivity rate than females (61.9% vs. 34.2%). The relative sensitivity rates were significantly higher for the standard tube test (44.6%) and fluorescence polarization assay (42.6%) than the card agglutination test (13.1%). Lesions consistent with Brucella infection were commonly found in the animals surveyed and included inflammatory lesions of the lymph nodes, liver, kidney, and male reproductive organs, which ranged from lymphoplasmacytic to pyogranulomatous with necrosis. This is the first report of an apparent enzootic Brucella abortus infection in a feral swine herd suggesting that feral swine may serve as a reservoir of infection for Brucella abortus as well as Brucella suis for domestic livestock. PMID:17459850

  10. 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. PMID:27001541

  11. Outer Membrane Proteins of Brucella abortus Vaccinal and Field Strains and their Immune Response in Buffaloes

    OpenAIRE

    Rukhshanda Munir*, M. Afzal1, M. Hussain2, S. M. S. Naqvi3 and A. Khanum3

    2010-01-01

    Outer membrane proteins (OMPs) of three strains of B. abortus i.e. S19, RB51 and a local field isolate of biotype 1 were isolated through disrupting cells to generate membranes by centrifugation and sodium lauryl sarcosinate solubilisation of inner membrane proteins. Distinct OMP profiles of each strain were seen on SDS-PAGE. SDS-PAGE analysis of S19 and field isolate revealed eight protein bands in each strain. The OMPs of S19 had molecular masses 89.0, 73.0, 53.7, 49.0, 38.0, 27.0, 22.3, a...

  12. 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

    OpenAIRE

    Hai Jiang; Heng Wang; Liqing Xu; Guiying Hu; Junying Ma; Pei Xiao; Weixing Fan; Dongdong Di; Guozhong Tian; Mengguang Fan; Jingchuan Mi; Ruiping Yu; Litao Song; Hongyan Zhao; Dongri Piao

    2013-01-01

    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 wer...

  13. Brucella abortus transits through the autophagic pathway and replicates in the endoplasmic reticulum of nonprofessional phagocytes

    OpenAIRE

    Pizarro-Cerda, J. (Javier); Meresse, S. (Stéphane); Parton, R G; van der Goot, G; Sola-Landa, A.; Lopez-Goñi, I. (Ignacio); Moreno, E.; Gorvel, J P

    1998-01-01

    Brucella abortus is an intracellular pathogen that replicates within a membrane-bounded compartment. In this study, we have examined the intracellular pathway of the virulent B. abortus strain 2308 (S2308) and the attenuated strain 19 (S19) in HeLa cells. At 10 min after inoculation, both bacterial strains are transiently detected in phagosomes characterized by the presence of early endosomal markers such as the early endosomal antigen 1. At approximately 1 h postinoculation, bacteria are loc...

  14. Brucella abortus 1119-3 O-chain polysaccharide to differentiate sera from B. abortus S-19-vaccinated and field-strain-infected cattle by agar gel immunodiffusion.

    OpenAIRE

    Cherwonogrodzky, J W; Nielsen, K H

    1988-01-01

    Purified Brucella abortus 1119-3 and Brucella melitensis 16M lipopolysaccharide O-chain polysaccharides were not precipitated in agar gel immunodiffusion by any of 24 sera from vaccinated cattle but were precipitated by 18 of 24 sera from infected cattle. This difference can be used to differentiate sera of cattle vaccinated with B. abortus S-19 from sera of some field-strain-infected cattle.

  15. Occupational infection due to Brucella abortus S19 among workers involved in vaccine production in Argentina.

    Science.gov (United States)

    Wallach, J C; Ferrero, M C; Victoria Delpino, M; Fossati, C A; Baldi, P C

    2008-08-01

    The pathological consequences of exposure to the vaccine strain Brucella abortus S19 were evaluated in 30 employees from vaccine-manufacturing plants. Active brucellosis was diagnosed in 21 subjects, of whom only five recalled an accidental exposure. Clinical manifestations were mild, and only one patient presented a complication. After antimicrobial therapy, initially symptomatic patients either experienced clinical remission or had mild persistent symptoms. This is the first study reporting infection by B. abortus S19 among workers from vaccine-manufacturing plants, which in many cases was acquired from unnoticed exposures. Measures to improve the safety of B. abortus S19 handling should be implemented. PMID:18727806

  16. Outer Membrane Proteins of Brucella abortus Vaccinal and Field Strains and their Immune Response in Buffaloes

    Directory of Open Access Journals (Sweden)

    Rukhshanda Munir*, M. Afzal1, M. Hussain2, S. M. S. Naqvi3 and A. Khanum3

    2010-04-01

    Full Text Available Outer membrane proteins (OMPs of three strains of B. abortus i.e. S19, RB51 and a local field isolate of biotype 1 were isolated through disrupting cells to generate membranes by centrifugation and sodium lauryl sarcosinate solubilisation of inner membrane proteins. Distinct OMP profiles of each strain were seen on SDS-PAGE. SDS-PAGE analysis of S19 and field isolate revealed eight protein bands in each strain. The OMPs of S19 had molecular masses 89.0, 73.0, 53.7, 49.0, 38.0, 27.0, 22.3, and 17.7 kDa, while field isolate had OMPs of 151.3, 89.0, 75.8, 67.6, 37.0, 27.0, 24.0 and 19.0 kDa. B. abortus RB51 yielded 11 OMP bands ranging from 12.5 to 107.1 kDa, with 34.2, 15.8 and 12.5 kDa as additional OMPs. Western immunoblot analysis using antisera raised against all three strains in buffaloes indicated an almost similar pattern of immuno-reactive OMPs in S19 and field strain. Two OMPs of molecular weight 37-38 and 19 kDa were immuno-reactive in all strains in buffaloes. There is possibility of use of these OMPs in a recombinant vaccine for B. abortus. A distinct protein of molecular weight of 151.3 kDa was identified in field strain but not in both vaccine strains of B. abortus. Use of this OMP in a diagnostic assay may differentiate between vaccinated and infected animals.

  17. [The application and research advances of Brucella vaccines].

    Science.gov (United States)

    Ding, Jia-Bo; Mao, Kai-Rong; Cheng, Jun-Sheng; Dai, Zhi-Hong; Jiang, Yu-Wen

    2006-10-01

    Brucellosis is a crucial zoonosis caused by Brucella, which has some traits of wide hosts, great infectivity and difficulty in cure. Brucellosis caused great losses to farming and people's health. Vaccination is the main measure used to control Brucellosis, and some attenuated Brucella strains were often used as vaccines. To find more effective vaccines, Scientists are now constructing recombinant strains, DNA vaccines and subunit vaccines, as well as inducing new attenuated strains from isolations. The present applications of B. abortus strain 19 (S19) , B. melitensis Rev. 1 (Rev. 1), B. suis strain 2 (S2), B. abortus strain 45/20 (45/20) and rough strain B. abortus 51 (RB51) were discussed. And some recent research work on Brucella vaccines, such as Brucella recombinant vaccines, DNA vaccines and so on, were reviewed in this paper. PMID:17172046

  18. 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. PMID:26093199

  19. A rapid cycleave PCR method for distinguishing the vaccine strain Brucella abortus A19 in China.

    Science.gov (United States)

    Nan, Wenlong; Zhang, Yueyong; Tan, Pengfei; Xu, Zouliang; Chen, Yuqi; Mao, Kairong; Chen, Yiping

    2016-05-01

    Brucellosis is a widespread zoonotic disease caused by Brucella spp. Immunization with attenuated vaccines has proved to be an effective method of prevention; however, it may also interfere with diagnosis. Brucella abortus strain A19, which is homologous to B. abortus strain S19, is widely used for the prevention of bovine brucellosis in China. For effective monitoring of the control of brucellosis, it is essential to distinguish A19 from field strains. Single-nucleotide polymorphism-based assays offer a new approach to such discrimination studies. In the current study, we developed a cycleave PCR assay that successfully distinguished attenuated vaccine strains A19 and S19 from 22 strains of B. abortus and 57 strains of 5 other Brucella species. The assay gave a negative reaction with 4 non-Brucella species. The minimum sensitivity of the assay, evaluated using 10-fold dilutions of chromosomal DNA, was 7.6 fg for the A19 strain and 220 fg for the single non-A19/non-S19 Brucella strain tested (B. abortus 104M). The assay was also reproducible (intra- and interassay coefficients of variation: 0.003-0.01 and 0.004-0.025, respectively). The cycleave assay gave an A19/S19-specific reaction in 3 out of 125 field serum samples, with the same 3 samples being positive in an alternative A19/S19-specific molecular assay. The cycleave assay gave a total of 102 Brucella-specific reactions (3 being the A19/S19-specific reactions), whereas an alternative Brucella-specific assay gave 92 positive reactions (all also positive in the cycleave assay). Therefore, this assay represents a simple, rapid, sensitive, and specific tool for use in brucellosis control. PMID:27075847

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

    OpenAIRE

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

    1998-01-01

    A vector for the expression of foreign antigens in the vaccine strain Brucella abortus S19 was developed by using a DNA fragment containing the regulatory sequences and the signal peptide of the Brucella bcsp31 gene. This fragment was cloned in broad-host-range plasmid pBBR4MCS, resulting in plasmid pBEV. As a reporter protein, a repetitive antigen of Trypanosoma cruzi was used. The recombinant fusion protein is stably expressed and secreted into the Brucella periplasmic space, inducing a goo...

  1. Identification of biovars of Brucella abortus in aborted cattle and buffaloes herd in Sri Lanka

    Directory of Open Access Journals (Sweden)

    M. A. R. Priyantha

    Full Text Available Bovine brucellosis is an endemic disease in Sri Lanka, caused by Brucella abortus and had been reported all part of the country for last six decades. Since available biovar is still unknown, the objective of the study was to identify the biovar of B.abortus from sporadically aborted cattle and buffaloes. Samples were collected from 18 aborted herds out of 19 herds of Cattle and Buffaloes in the year 2010. Rose Bengal plate test and Compliment Fixation test were carried out. Milk, vaginal swabs, placental contents and aborted fetus were collected and cultured by conventional bacteriological methods. The detection of biovars were based on growth on Thionin and Bacto fuschin,CO2 requirement, H2S production, serum agglutination with Brucella negative, A,M and R reference antiserum. Eighteen herds investigated out of 19 herds reported, 11 herds were serologically positive for brucellosis (61.11% and only Brucella abortus were isolated from 8 individuals from six herds. All were identified as Biovar 3 of Brucella abortus. [Vet. World 2011; 4(12.000: 542-545

  2. Identificación de la cepa vacunal Brucella abortus S19 en muestras de leche de vaca

    OpenAIRE

    Luary Carolina Martínez Chavarría; Antonio Verdugo Rodríguez; Rigoberto Hernández Castro

    2006-01-01

    La brucelosis es una enfermedad infectocontagiosa de origen bacteriano que afecta tanto al humano como a diferentes especies animales domésticas y silvestres. En la década pasada, la vacuna que se usaba ampliamente en los bovinos era la cepa vacunal B. abortus S19, que tiene una deleción en dos de los genes del operón ery que participa en el catabolismo del eritritol. En México, desde 1997 se aprobó la cepa B. abortus RB51 como vacuna para el ganado. Esta cepa presenta una secuencia de inserc...

  3. 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.

  4. 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...

  5. Antigens of Brucella abortus S19 immunodominant for bovine lymphocytes as identified by one- and two-dimensional cellular immunoblotting.

    OpenAIRE

    Brooks-Worrell, B M; Splitter, G A

    1992-01-01

    Cellular immune responses are influential for protection against intracellular bacteria such as brucellae. Therefore, identification of Brucella abortus antigens that activate primed bovine lymphocytes is fundamental for discerning the breadth of cellular response in bovine brucellosis. Potentially antigenic components of B. abortus S19 were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by nitrocellulose blotting. Specific one-dimensional blot segments induced...

  6. Genome sequence of Brucella abortus vaccine strain S19 compared to virulent strains yields candidate virulence genes

    OpenAIRE

    Crasta, Oswald R.; Folkerts, Otto; Fei, Zhangjun; Mane, Shrinivasrao P.; Evans, Clive; Martino-Catt, Susan; Bricker, Betsy; Yu, GongXin; Du, Lei; Sobral, Bruno W.

    2008-01-01

    The Brucella abortus strain S19, a spontaneously attenuated strain, has been used as a vaccine strain in vaccination of cattle against brucellosis for six decades. Despite many studies, the physiological and molecular mechanisms causing the attenuation are not known. We have applied pyrosequencing technology together with conventional sequencing to rapidly and comprehensively determine the complete genome sequence of the attenuated Brucella abortus vaccine strain S19. The main goal of this st...

  7. Cloning of Brucella abortus gene and characterization of expressed 26-kilodalton periplasmic protein: potential use for diagnosis.

    OpenAIRE

    Rossetti, O L; Arese, A I; Boschiroli, M L; Cravero, S L

    1996-01-01

    Brucella spp. are the causative agents of brucellosis in many different hosts, including humans. Most of the serological methods of diagnosis are based on the detection of antilipopolysaccharide antibodies, which makes the differentiation of vaccinated animals from infected animals difficult. By using molecular biology techniques, a gene that encodes a 26-kDa protein (BP26) was isolated from a Brucella abortus S19 genome lambda gt11 library. This protein is in the periplasm of B. abortus and ...

  8. The Brucella abortus Cyclic β-1,2-Glucan Virulence Factor Is Substituted with O-Ester-Linked Succinyl Residues

    OpenAIRE

    Roset, Mara S.; Ciocchini, Andrés E.; Ugalde, Rodolfo A.; Iñón de Iannino, Nora

    2006-01-01

    Brucella periplasmic cyclic β-1,2-glucan plays an important role during bacterium-host interaction. Nuclear magnetic resonance spectrometry analysis, thin-layer chromatography, and DEAE-Sephadex chromatography were used to characterize Brucella abortus cyclic glucan. In the present study, we report that a fraction of B. abortus cyclic β-1,2-glucan is substituted with succinyl residues, which confer anionic character on the cyclic β-1,2-glucan. The oligosaccharide backbone is substituted at C-...

  9. Genome Sequence of Brucella abortus Vaccine Strain S19 Compared to Virulent Strains Yields Candidate Virulence Genes

    OpenAIRE

    Crasta, Oswald R.; Otto Folkerts; Zhangjun Fei; Mane, Shrinivasrao P.; Clive Evans; Susan Martino-Catt; Betsy Bricker; GongXin Yu; Lei Du; Sobral, Bruno W.

    2008-01-01

    The Brucella abortus strain S19, a spontaneously attenuated strain, has been used as a vaccine strain in vaccination of cattle against brucellosis for six decades. Despite many studies, the physiological and molecular mechanisms causing the attenuation are not known. We have applied pyrosequencing technology together with conventional sequencing to rapidly and comprehensively determine the complete genome sequence of the attenuated Brucella abortus vaccine strain S19. The main goal of this st...

  10. Biotyping and genotyping (MLVA16 of Brucella abortus isolated from cattle in Brazil, 1977 to 2008.

    Directory of Open Access Journals (Sweden)

    Sílvia Minharro

    Full Text Available Brucellosis is a worldwide distributed zoonosis that causes important economic losses to animal production. In Brazil, information on the distribution of biovars and genotypes of Brucella spp. is scarce or unavailable. This study aimed (i to biotype and genotype 137 Brazilian cattle isolates (from 1977 to 2008 of B. abortus and (ii to analyze their distribution. B. abortus biovars 1, 2 and 3 (subgroup 3b were confirmed and biovars 4 and 6 were first described in Brazil. Genotyping by the panel 1 revealed two groups, one clustering around genotype 40 and another around genotype 28. Panels 2A and 2B disclosed a high diversity among Brazilian B. abortus strains. Eighty-nine genotypes were found by MLVA16. MLVA16 panel 1 and 2 showed geographic clustering of some genotypes. Biotyping and MLVA16 genotyping of Brazilian B. abortus isolates were useful to better understand the epidemiology of bovine brucellosis in the region.

  11. 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.

  12. 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

    2016-01-01

    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 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. PMID:27144565

  13. Literatuuronderzoek naar gegevens betreffende de betekenis van een aantal verwekkers van zoonosen in verband met de vleesconsumptie. VIII: Brucella abortus

    NARCIS (Netherlands)

    Bos; J.M.; Engel; H.W.B.; Groothuis; D.G.; Knapen; F. van; Weiss; J.W.

    1986-01-01

    Brucellose bij de mens heeft meestal een weinig specifiek, griepachtig beeld. In Nederland is de voornaamste verwekker Brucella abortus, waar toe het literatuuronderzoek zich dan ook beperkt. Bij dieren is het rund de voornaamste gastheer en leidt een infectie bij drachtige dieren tot abortus.

  14. Isolation of Brucella abortus ssb and uvrA genes from a genomic library by use of lymphocytes as probes.

    OpenAIRE

    Zhu, Y.; S.C. Oliveira; Splitter, G A

    1993-01-01

    Brucella abortus proteins from virulent S2308 expressed from a pBluescript II SK- genomic library stimulated peripheral blood mononuclear (PBM) cell proliferation from cattle vaccinated with B. abortus S19. The method described here permits a rapid and directed approach to isolate genes encoding antigens of B. abortus that interact with lymphocytes primed to the living bacterium. The supernatants from the bacterial host JM109 (DE3) were cultured with freshly isolated bovine PBM cells. A total...

  15. A dual-targeting approach to inhibit Brucella abortus replication in human cells

    Science.gov (United States)

    Czyż, Daniel M.; Jain-Gupta, Neeta; Shuman, Howard A.; Crosson, Sean

    2016-01-01

    Brucella abortus is an intracellular bacterial pathogen and an etiological agent of the zoonotic disease known as brucellosis. Brucellosis can be challenging to treat with conventional antibiotic therapies and, in some cases, may develop into a debilitating and life-threatening chronic illness. We used multiple independent assays of in vitro metabolism and intracellular replication to screen a library of 480 known bioactive compounds for novel B. abortus anti-infectives. Eighteen non-cytotoxic compounds specifically inhibited B. abortus replication in the intracellular niche, which suggests these molecules function by targeting host cell processes. Twenty-six compounds inhibited B. abortus metabolism in axenic culture, thirteen of which are non-cytotoxic to human host cells and attenuate B. abortus replication in the intracellular niche. The most potent non-cytotoxic inhibitors of intracellular replication reduce B. abortus metabolism in axenic culture and perturb features of mammalian cellular biology including mitochondrial function and receptor tyrosine kinase signaling. The efficacy of these molecules as inhibitors of B. abortus replication in the intracellular niche suggests “dual-target” compounds that coordinately perturb host and pathogen are promising candidates for development of improved therapeutics for intracellular infections. PMID:27767061

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

    Science.gov (United States)

    Gao, Jianpeng; Tian, Mingxing; Bao, Yanqing; Li, Peng; Liu, Jiameng; Ding, Chan; Wang, Shaohui; Li, Tao; Yu, Shengqing

    2016-01-01

    Brucellosis, 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 growth curves and resistance to environmental stress factors showed that Pyk plays an important role in B. abortus growth, especially under the conditions of nutrition deprivation, and resistance to oxidative stress. Additionally, cell infection assay showed that Pyk is necessary for B. abortus survival and evading fusion with lysosomes within RAW264.7 cells. Moreover, animal experiments exhibited that the Pyk deletion significantly reduced B. abortus virulence in a mouse infection model. Our results elucidated the role of the Pyk in B. abortus virulence and provided information for further investigation of Brucella virulence associated carbon metabolism. PMID:27561260

  17. Brucella abortus Induces the Secretion of Proinflammatory Mediators from Glial Cells Leading to Astrocyte Apoptosis

    Science.gov (United States)

    García Samartino, Clara; Delpino, M. Victoria; Pott Godoy, Clara; Di Genaro, María Silvia; Pasquevich, Karina A.; Zwerdling, Astrid; Barrionuevo, Paula; Mathieu, Patricia; Cassataro, Juliana; Pitossi, Fernando; Giambartolomei, Guillermo H.

    2010-01-01

    Central nervous system (CNS) invasion by bacteria of the genus Brucella results in an inflammatory disorder called neurobrucellosis. In this study we present in vivo and in vitro evidence that B. abortus and its lipoproteins activate the innate immunity of the CNS, eliciting an inflammatory response that leads to astrogliosis, a characteristic feature of neurobrucellosis. Intracranial injection of heat-killed B. abortus (HKBA) or outer membrane protein 19 (Omp19), a B. abortus lipoprotein model, induced astrogliosis in mouse striatum. Moreover, infection of astrocytes and microglia with B. abortus induced the secretion of interleukin (IL)−6, IL-1β, tumor necrosis factor (TNF)-α, macrophage chemoattractant protein−1, and KC (CXCL1). HKBA also induced these inflammatory mediators, suggesting the involvement of a structural component of the bacterium. Accordingly, Omp19 induced the same cytokine and chemokine secretion pattern. B. abortus infection induced astrocyte, but not microglia, apoptosis. Indeed, HKBA and Omp19 elicited not only astrocyte apoptosis but also proliferation, two features observed during astrogliosis. Apoptosis induced by HKBA and L-Omp19 was completely suppressed in cells of TNF receptor p55−/− mice or when the general caspase inhibitor Z-VAD-FMK was added to cultures. Hence, TNF-α signaling via TNF receptor (TNFR) 1 through the coupling of caspases determines apoptosis. Our results provide proof of the principle that Brucella lipoproteins could be key virulence factors in neurobrucellosis and that astrogliosis might contribute to neurobrucellosis pathogenesis. PMID:20093491

  18. T lymphocytes subsets and cytokine pattern induced by vaccination against bovine brucellosis employing S19 calfhood vaccination and adult RB51 revaccination.

    Science.gov (United States)

    Dorneles, Elaine M S; Teixeira-Carvalho, Andréa; Araújo, Márcio S S; Lima, Graciela Kunrath; Martins-Filho, Olindo A; Sriranganathan, Nammalwar; Lage, Andrey P

    2014-10-21

    The aims of this study were to address the protective immune response induced by S19 vaccination (n=10) and RB51 revaccination, in pregnant (n=9) and non-pregnant (n=10) S19 calfhood-vaccinated cattle as follows: evaluate the in vitro CD4(+) and CD8(+) T-lymphocytes specific proliferation, and in vitro expression of IFN-γ by CD4(+) and CD8(+) T-cells and IL-4 by CD4(+), CD8(+) and CD21(+) lymphocytes subset. Upon in vitro stimulation with γ-irradiated Brucella abortus 2308, blood mononuclear cells from S19 vaccinated and RB51 revaccinated cows exhibited significantly higher proliferation of CD4(+) and CD8(+) T-lymphocytes and CD4(+)IFN-γ(+) T-cells compared to non-vaccinated animals. RB51 revaccination, regardless of the pregnancy status, did not enhance the proliferation of CD4(+) or CD8(+) T-cells nor IFN-γ or IL-4 production. Data from the present study suggest that cattle's cellular immune response induced after brucellosis vaccination and revaccination is due to CD4(+) and CD8(+) T-lymphocytes, being CD4(+) T-cells the main source of IFN-γ. PMID:25218192

  19. 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.

  20. Immunogenic response induced by wzm and wzt gene deletion mutants from Brucella abortus S19.

    Science.gov (United States)

    Wang, Xiu-Ran; Yan, Guang-Mou; Zhang, Rui; Lang, Xu-Long; Yang, Yan-Ling; Li, Xiao-Yan; Chen, Si; Qian, Jing; Wang, Xing-Long

    2014-02-01

    Brucellosis is an infectious disease affecting humans and animals worldwide. Effective methods of control include inducing immunity in animals by vaccination and elimination. Brucella abortus S19 is one of the popular vaccines for control of cattle brucellosis, as it has low virulence. In this paper, allelic exchange plasmids of wzm and wzt genes were constructed and partially knocked out to evaluate the effects on the induction of immunity to Brucella abortus S19 mutants. Cytokine secretion in vitro, INF-γ induction in vivo and antibody dynamics were evaluated. These data suggested that the immunity-eliciting ability of the wzm and wzt gene deletion mutants was similar, although reduced compared with the S19 strain. The results demonstrated that the wzt gene may be more important in the regulation of the induction of immunity than the wzm gene. PMID:24247358

  1. Brucella abortus Induces the Premature Death of Human Neutrophils through the Action of Its Lipopolysaccharide.

    Science.gov (United States)

    Barquero-Calvo, Elías; Mora-Cartín, Ricardo; Arce-Gorvel, Vilma; de Diego, Juana L; Chacón-Díaz, Carlos; Chaves-Olarte, Esteban; Guzmán-Verri, Caterina; Buret, Andre G; Gorvel, Jean-Pierre; Moreno, Edgardo

    2015-05-01

    Most bacterial infections induce the activation of polymorphonuclear neutrophils (PMNs), enhance their microbicidal function, and promote the survival of these leukocytes for protracted periods of time. Brucella abortus is a stealthy pathogen that evades innate immunity, barely activates PMNs, and resists the killing mechanisms of these phagocytes. Intriguing clinical signs observed during brucellosis are the low numbers of Brucella infected PMNs in the target organs and neutropenia in a proportion of the patients; features that deserve further attention. Here we demonstrate that B. abortus prematurely kills human PMNs in a dose-dependent and cell-specific manner. Death of PMNs is concomitant with the intracellular Brucella lipopolysaccharide (Br-LPS) release within vacuoles. This molecule and its lipid A reproduce the premature cell death of PMNs, a phenomenon associated to the low production of proinflammatory cytokines. Blocking of CD14 but not TLR4 prevents the Br-LPS-induced cell death. The PMNs cell death departs from necrosis, NETosis and classical apoptosis. The mechanism of PMN cell death is linked to the activation of NADPH-oxidase and a modest but steadily increase of ROS mediators. These effectors generate DNA damage, recruitments of check point kinase 1, caspases 5 and to minor extent of caspase 4, RIP1 and Ca++ release. The production of IL-1β by PMNs was barely stimulated by B. abortus infection or Br-LPS treatment. Likewise, inhibition of caspase 1 did not hamper the Br-LPS induced PMN cell death, suggesting that the inflammasome pathway was not involved. Although activation of caspases 8 and 9 was observed, they did not seem to participate in the initial triggering mechanisms, since inhibition of these caspases scarcely blocked PMN cell death. These findings suggest a mechanism for neutropenia in chronic brucellosis and reveal a novel Brucella-host cross-talk through which B. abortus is able to hinder the innate function of PMN. PMID:25946018

  2. Brucella abortus Induces the Premature Death of Human Neutrophils through the Action of Its Lipopolysaccharide.

    Directory of Open Access Journals (Sweden)

    Elías Barquero-Calvo

    2015-05-01

    Full Text Available Most bacterial infections induce the activation of polymorphonuclear neutrophils (PMNs, enhance their microbicidal function, and promote the survival of these leukocytes for protracted periods of time. Brucella abortus is a stealthy pathogen that evades innate immunity, barely activates PMNs, and resists the killing mechanisms of these phagocytes. Intriguing clinical signs observed during brucellosis are the low numbers of Brucella infected PMNs in the target organs and neutropenia in a proportion of the patients; features that deserve further attention. Here we demonstrate that B. abortus prematurely kills human PMNs in a dose-dependent and cell-specific manner. Death of PMNs is concomitant with the intracellular Brucella lipopolysaccharide (Br-LPS release within vacuoles. This molecule and its lipid A reproduce the premature cell death of PMNs, a phenomenon associated to the low production of proinflammatory cytokines. Blocking of CD14 but not TLR4 prevents the Br-LPS-induced cell death. The PMNs cell death departs from necrosis, NETosis and classical apoptosis. The mechanism of PMN cell death is linked to the activation of NADPH-oxidase and a modest but steadily increase of ROS mediators. These effectors generate DNA damage, recruitments of check point kinase 1, caspases 5 and to minor extent of caspase 4, RIP1 and Ca++ release. The production of IL-1β by PMNs was barely stimulated by B. abortus infection or Br-LPS treatment. Likewise, inhibition of caspase 1 did not hamper the Br-LPS induced PMN cell death, suggesting that the inflammasome pathway was not involved. Although activation of caspases 8 and 9 was observed, they did not seem to participate in the initial triggering mechanisms, since inhibition of these caspases scarcely blocked PMN cell death. These findings suggest a mechanism for neutropenia in chronic brucellosis and reveal a novel Brucella-host cross-talk through which B. abortus is able to hinder the innate function of PMN.

  3. Protection against infection in mice vaccinated with a Brucella abortus mutant.

    OpenAIRE

    Boschiroli, M L; Cravero, S L; Arese, A I; Campos, E.; Rossetti, O L

    1997-01-01

    This study determines whether a genetically engineered mutant of Brucella abortus, strain M-1, possesses differences in protective properties compared to the parental strain, vaccine S19. M-1 is a mutant unable to express BP26, a periplasmic protein with potential use in diagnosis. Mice vaccinated with S19 developed antibodies against BP26, while those vaccinated with M-1 did not. However, mice vaccinated with S19 or M-1 were similarly protected against challenge with pathogenic strain 2308, ...

  4. Evaluation of Brucella abortus Phosphoglucomutase (pgm) Mutant as a New Live Rough-Phenotype Vaccine

    OpenAIRE

    Ugalde, Juan Esteban; Comerci, Diego José; Leguizamón, M. Susana; Ugalde, Rodolfo Augusto

    2003-01-01

    Brucella abortus S19 is the vaccine most frequently used against bovine brucellosis. Although it induces good protection levels, it cannot be administered to pregnant cattle, revaccination is not advised due to interference in the discrimination between infected and vaccinated animals during immune-screening procedures, and the vaccine is virulent for humans. Due to these reasons, there is a continuous search for new bovine vaccine candidates that may confer protection levels comparable to th...

  5. Proteomic Profile of Brucella abortus-Infected Bovine Chorioallantoic Membrane Explants

    Science.gov (United States)

    Mol, Juliana P. S.; Pires, Simone F.; Chapeaurouge, Alexander D.; Perales, Jonas; Santos, Renato L.; Andrade, Hélida M.; Lage, Andrey P.

    2016-01-01

    Brucella abortus is the etiological agent of bovine brucellosis, a zoonotic disease that causes significant economic losses worldwide. The differential proteomic profile of bovine chorioallantoic membrane (CAM) explants at early stages of infection with B. abortus (0.5, 2, 4, and 8 h) was determined. Analysis of CAM explants at 0.5 and 4 h showed the highest differences between uninfected and infected CAM explants, and therefore were used for the Differential Gel Electrophoresis (DIGE). A total of 103 spots were present in only one experimental group and were selected for identification by mass spectrometry (MALDI/ToF-ToF). Proteins only identified in extracts of CAM explants infected with B. abortus were related to recognition of PAMPs by TLR, production of reactive oxygen species, intracellular trafficking, and inflammation. PMID:27104343

  6. In vitro and in vivo characterization of smooth small colony variants of Brucella abortus S19.

    Science.gov (United States)

    Jacob, J; Hort, G M; Overhoff, P; Mielke, M E A

    2006-02-01

    Brucella abortus is known to produce chronic infections in both humans and a variety of animal species. However, the mechanisms underlying the persistence of the bacteria in the presence of an ongoing immune response are still unknown. In this respect we made use of the observation that in vitro grown B. abortus S19 exhibits heterogenicity in colony size when plated onto TS agar, while experimental infection of mice uniformly results in the in vivo selection of the small colony variant. We demonstrate that the spontaneous smooth small colony variant is characterized not only by a slower growth rate in vitro but also by an increased tolerance to hyperosmotic medium and, most importantly, a less effective clearance from spleens and livers of experimentally infected mice. On a molecular level, a gene with homology to a formerly described galactoside transport ATP binding protein (mglA) was differentially expressed in small versus large colonies of B. abortus S19. PMID:16239117

  7. Purification and properties of Cu-Zn superoxide dismutase extracted from Brucella abortus strain 19

    Energy Technology Data Exchange (ETDEWEB)

    Tabatabai, L.B. (ARS-USDA, Ames, IA (United States))

    1991-03-11

    Recent work showed that a recombinant 20 kDa protein from Brucella abortus expressed in E. coli is a Cu-Zn superoxide dismutase (SOD). Western blot and ELISA results indicated that cattle with brucellosis have antibody to SOD. Here the authors report the purification and properties of the native B. abortus Cu-Zn SOD. SOD was extracted from methanol-killed Brucella abortus strain 19 with 0.1 M sodium citrate-1.0 M sodium chloride solution. The extract was dialyzed and protein precipitated by ammonium sulfate at 70-100% saturation was collected. The SOD was purified by HPLC anion exchange chromatography. SOD activity was assayed with a coupled enzyme assay using xanthine oxidase-cytochrome C reduction assay. The authors determined that the Brucella SOD is present in two molecular forms both inhibitable with KCN with Ki's of 0.32 mM and 4.98 mM, respectively. No other form of SOD was identified in the extract. Polyclonal antibody to SOD and polyclonal antibody to SOD synthetic peptide residues 134-143 inhibited SOD activity by 50% and 13%, respectively. Both SOD and the synthetic peptide inhibited binding of anti-SOD antibody to SOD by 60% and 20%, respectively. Based on these results the SOD and its amphipathic peptide will be considered as candidates for the design of synthetic multiple peptide vaccines and diagnostic reagents for bovine brucellosis.

  8. Molecular Epidemiology of Brucella abortus in Northern Ireland-1991 to 2012.

    Directory of Open Access Journals (Sweden)

    Adrian Allen

    Full Text Available Brucellosis is the most common bacterial zoonoses worldwide. Bovine brucellosis caused by Brucella abortus has far reaching animal health and economic impacts at both the local and national levels. Alongside traditional veterinary epidemiology, the use of molecular typing has recently been applied to inform on bacterial population structure and identify epidemiologically-linked cases of infection. Multi-locus variable number tandem repeat VNTR analysis (MLVA was used to investigate the molecular epidemiology of a well-characterised Brucella abortus epidemic in Northern Ireland involving 387 herds between 1991 and 2012.MLVA identified 98 unique B. abortus genotypes from disclosing isolates in the 387 herds involved in the epidemic. Clustering algorithms revealed the relatedness of many of these genotypes. Combined with epidemiological information on chronology of infection and geographic location, these genotype data helped to identify 7 clonal complexes which underpinned the outbreak over the defined period. Hyper-variability of some VNTR loci both within herds and individual animals led to detection of multiple genotypes associated with single outbreaks. However with dense sampling, these genotypes could still be associated with specific clonal complexes thereby permitting inference of epidemiological links. MLVA- based epidemiological monitoring data were congruent with an independent classical veterinary epidemiology study carried out in the same territory.MLVA is a useful tool in ongoing disease surveillance of B. abortus outbreaks, especially when combined with accurate epidemiological information on disease tracings, geographical clustering of cases and chronology of infection.

  9. Cytokine responses in camels (Camelus bactrianus) vaccinated with Brucella abortus strain 19 vaccine.

    Science.gov (United States)

    Odbileg, Raadan; Purevtseren, Byambaa; Gantsetseg, Dorj; Boldbaatar, Bazartseren; Buyannemekh, Tumurjav; Galmandakh, Zagd; Erdenebaatar, Janchivdorj; Konnai, Satoru; Onuma, Misao; Ohashi, Kazuhiko

    2008-02-01

    In the present study, we determined the levels of cytokines produced by camel (Camelus bactrianus) peripheral blood mononuclear cells (PBMCs) in response to live attenuated Brucella abortus (B. abortus) S19 vaccine. Seven camels were vaccinated with commercial B. abortus S19 vaccine, and their cytokine responses were determined using a real-time PCR assay. Cytokine responses to B. abortus S19 were examined at 6 hr, 48 hr and 1, 2 and 3 weeks post-vaccination. Serological tests were performed to further confirm these immune responses. The results revealed that IFN-gamma and IL-6 were upregulated during the first week post-vaccination. Low level expressions of IL-1alpha, IL-1beta, TNFalpha and IL-10 and no expression of IL-2 and IL-4 were observed compared with the control camels. The findings showed that B. abortus stimulates cell-mediated immunity by directly activating camel Th1 cells to secrete IFN-gamma. This quantification of cytokine expression in camels is essential for understanding of Camelidae disease development and protective immune responses. This is the first report of in vivo camel cytokine quantification after vaccination. PMID:18319583

  10. Expression of Babesia bovis rhoptry-associated protein 1 (RAP1) in Brucella abortus S19.

    Science.gov (United States)

    Sabio y García, Julia V; Farber, Marisa; Carrica, Mariela; Cravero, Silvio; Macedo, Gilson C; Bigi, Fabiana; Oliveira, Sergio C; Rossetti, Osvaldo; Campos, Eleonora

    2008-05-01

    Brucella abortus strain 19 (live vaccine) induces a strong humoral and cellular immune response and therefore, it is an attractive vector for the delivery of heterologous antigens. The objective of the present study was to express the rhoptry-associated protein (RAP1) of Babesia bovis in B. abortus S19, as a model for heterologous expression of immunostimulatory antigens from veterinary pathogens. A plasmid for the expression of recombinant proteins fused to the aminoterminal of the outer membrane lipoprotein OMP19 was created, pursuing the objective of increasing the immunogenicity of the recombinant antigen being expressed by its association to a lipid moiety. Recombinant strains of B. abortus S19 expressing RAP1 as a fusion protein either with the first amino acids of beta-galactosidase (S19pBB-RAP1) or B. abortus OMP19 (S19pBB19-RAP1) were generated. Plasmid stability and the immunogenicity of the heterologous proteins were analyzed. Mice immunized with S19pBB-RAP1 or S19pBB19-RAP1 developed specific humoral immune response to RAP1, IgG2a being the predominant antibody isotype. Furthermore, a specific cellular immune response to recombinant RAP1 was elicited in vitro by lymphocytes from mice immunized with both strains. Therefore, we concluded that B. abortus S19 expressing RAP1 is immunostimulatory and may provide the basis for combined heterologous vaccines for babesiosis and brucellosis. PMID:18462974

  11. Brucella suis S2, brucella melitensis Rev. 1 and Brucella abortus S19 living vaccines: residual virulence and immunity induced against three Brucella species challenge strains in mice.

    Science.gov (United States)

    Bosseray, N; Plommet, M

    1990-10-01

    Live attenuated Brucella suis S2 vaccine was compared to living vaccines B. abortus S19 and B. melitensis Rev. 1 in mice. Residual virulence was estimated by ability to multiply and persist in spleen and lymph nodes. Immunogenicity was estimated by spleen counts of control and vaccinated mice challenged either with the reference B. abortus 544 strain or with virulent B. melitensis H38 and B. suis 1330 strains. S2 vaccine had lower residual virulence; expressed as 50% recovery time, persistence was 4.3 weeks, compared to 7.1 and 9.0 weeks for S19 and Rev. 1 vaccines. Immunity induced by the three vaccines was similar 45 days after vaccination. At 150 days, immunity by S19 and Rev.1 was still similar against the three challenge strains. In contrast, immunity induced by S2 had declined against the B. melitensis strain. Thus, a recall vaccination may be required for vaccination of sheep to confer a long-lasting immunity. PMID:2123586

  12. Characterization of Brucella abortus O-polysaccharide and core lipopolysaccharide mutants and demonstration that a complete core is required for rough vaccines to be efficient against Brucella abortus and Brucella ovis in the mouse model

    OpenAIRE

    Monreal, D.; Grillo, M.J. (María Jesús); Gonzalez-fernandez, D.; Marin, C.M.; de Miguel, M. J.; Lopez-Goñi, I. (Ignacio); Blasco, J.M. (José); Cloeckaert, A.; Moriyon, I. (Ignacio)

    2003-01-01

    Brucella abortus rough lipopolysaccharide (LPS) mutants were obtained by transposon insertion into two wbk genes (wbkA [putative glycosyltransferase; formerly rfbU] and per [perosamine synthetase]), into manB (pmm [phosphomannomutase; formerly rfbK]), and into an unassigned gene. Consistent with gene-predicted roles, electrophoretic analysis, 2-keto-3-manno-D-octulosonate measurements, and immunoblots with monoclonal antibodies to O-polysaccharide, outer and inner core epitopes showed no O-po...

  13. Humoral immune response against lipopolysaccharide and cytoplasmic proteins of Brucella abortus in cattle vaccinated with B. abortus S19 or experimentally infected with Yersinia enterocolitica serotype 0:9.

    OpenAIRE

    Baldi, P C; Giambartolomei, G H; Goldbaum, F A; Abdón, L F; C.A. Velikovsky; Kittelberger, R; Fossati, C A

    1996-01-01

    The humoral immune responses against three different antigens of Brucella abortus were monitored by enzyme-linked immunosorbent assay in cattle vaccinated with B. abortus S19 or experimentally infected with Yersinia enterocolitica serotype 0:9. Immunoglobulin G (IgG) and IgM responses against (i) B. abortus lipopolysaccharide (LPS), (ii) total cytoplasmic proteins depleted of LPS (LPS-free CYT), and (iii) B. abortus 18-kDa cytoplasmic protein were measured. Vaccinated animals and Yersinia-inf...

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

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    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.

  15. Identification and Characterization of a Brucella abortus ATP-Binding Cassette Transporter Homolog to Rhizobium meliloti ExsA and Its Role in Virulence and Protection in Mice

    OpenAIRE

    G.M.S. Rosinha; Freitas, Daniela A.; Miyoshi, Anderson; Azevedo, Vasco; Campos, Eleonora; Cravero, Silvio L; Rossetti, Osvaldo; Splitter, Gary; S.C. Oliveira

    2002-01-01

    Brucella abortus is a facultative intracellular bacterial pathogen that causes abortion in domestic animals and undulant fever in humans. The mechanism of virulence of Brucella spp. is not fully understood yet. Furthermore, genes that allow Brucella to reach the intracellular niche and to interact with host cells need to be identified. Using the genomic survey sequence (GSS) approach, we identified the gene encoding an ATP-binding cassette (ABC) transporter of B. abortus strain S2308. The ded...

  16. In Vitro Antibacterial Effects of Five Volatile Oil Extracts Against Intramacrophage Brucella Abortus 544

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    Ayman Al-Mariri

    2012-06-01

    Full Text Available Background: Brucella abortus is a gram-negative facultative intracellular bacterium that can cause a highly contagious disease in sheep, goats, cattle and one-humped camels. It is responsible for one of the most important zoonosis in human. The aim of this study was to evaluate the role of Mentha piperita, Origanum majorana, Citrus lemon, Cinnamomum verum and Myristica fragrans essential volatile oil extracts on human macrophages infected by B. abortus 544. Methods: Essential volatile oil extracts from M. piperita, O. majorana, C. lemon, C. verum and M. fragrans were extracted. Human macrophages were cultured at a density of 2×105 cells per well in sterile 96-well microtiter plates, and infected with B. abortus 544 at a ratio of 1:100 bacteria/cell. Then essential volatile oil extracts were added at a concentration of 1%. At specified times; cells were washed, lysed with 0.1% Triton, and plated on 2YT agar to determine the number of intracellular bacteria. Results: Cinnamomum verum volatile oil at a concentration of 1% had the highest antibacterial activity against B. abortus 544 inside human macrophages. Its inhibitory effect observed from 24 h and continued till 144 h after the infection. Moreover, C. verum (0.1% in combination with 1% concentration of M. piperita, O. majorana, C. lemon or M. fragrans volatile oil extracts produced a synergistic inhibitory effect against B. abortus 544. Conclusion: The results indicate that, among the five selected oil extracts, C. verum volatile oil applied either separately or in combination with other oil extracts had the most effective antimicrobial activity against Brucella.

  17. 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. PMID:15972510

  18. 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.

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    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.

  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.

    Science.gov (United States)

    Jiang, Hai; Wang, Heng; Xu, Liqing; Hu, Guiying; Ma, Junying; Xiao, Pei; Fan, Weixing; Di, Dongdong; Tian, Guozhong; Fan, Mengguang; Mi, Jingchuan; Yu, Ruiping; Song, Litao; Zhao, Hongyan; Piao, Dongri; Cui, Buyun

    2013-01-01

    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. PMID:24124546

  20. Expression and validation of D-erythrulose 1-phosphate dehydrogenase from Brucella abortus: a diagnostic reagent for bovine brucellosis.

    Science.gov (United States)

    Eoh, Hyungjin; Jeon, Bo-Young; Kim, Zhiyeol; Kim, Seung-Cheol; Cho, Sang-Nae

    2010-07-01

    Brucella abortus is a bacterium of brucellosis causing abortion in cattle. The diagnosis of bovine brucellosis mainly relies on serologic tests using smooth lipopolysaccharide (S-LPS) from B. abortus. However, the usefulness of this method is limited by false-positive reactions due to cross-reaction with other Gram-negative bacteria. In the present study, the eryC gene encoding B. abortus d-erythrulose 1-phosphate dehydrogenase, which is involved in the erythritol metabolism in virulent B. abortus strain but is absent from a B. abortus vaccine strain (S19), was cloned. Recombinant EryC was expressed and purified for the evaluation as a diagnostic reagent for bovine brucellosis. Other B. abortus proteins, Omp16, PP26, and CP39 were also purified and their seroreactivities were compared. Recombinant EryC, Omp16, PP26, and PP39 were all reactive to B. abortus-positive serum. The specificity of recombinant Omp16, PP26, CP39, and EryC, were shown to be approximately 98%, whereas that of B. abortus whole cell lysates was shown to be 95%. The sensitivity of Omp16, PP26, CP39, and EryC were 10%, 51%, 64%, and 43%, respectively, whereas that of B. abortus whole cell lysates was 53%. These results suggested that B. abortus EryC would be a potential reagent for diagnosis for bovine brucellosis as a single protein antigen. PMID:20622221

  1. Deletion of znuA Virulence Factor Attenuates Brucella abortus and Confers Protection against Wild-Type Challenge

    OpenAIRE

    Yang, Xinghong; Becker, Todd; Walters, Nancy; Pascual, David W.

    2006-01-01

    znuA is known to be an important factor for survival and normal growth under low Zn2+ concentrations for Escherichia coli, Haemophilus spp., Neisseria gonorrhoeae, and Pasteurella multocida. We hypothesized that the znuA gene present in Brucella melitensis 16 M would be similar to znuA in B. abortus and questioned whether it may also be an important factor for growth and virulence of Brucella abortus. Using the B. melitensis 16 M genome sequence, primers were designed to construct a B. abortu...

  2. 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.

  3. A combined DNA vaccine provides protective immunity against Mycobacterium bovis and Brucella abortus in cattle.

    Science.gov (United States)

    Hu, Xi-Dan; Yu, Da-Hai; Chen, Su-Ting; Li, Shu-Xia; Cai, Hong

    2009-04-01

    We evaluated the immunogenicity and protective efficacy of a combined DNA vaccine containing six genes encoding immunodominant antigens from Mycobacterium bovis and Brucella abortus. The number of lymph node and spleen cultures positive for M. bovis and B. abortus from calves immunized with the combined DNA vaccine was significantly reduced (p abortus 544. The combined DNA vaccine group displayed stronger antigen-specific interferon-gamma (IFN-gamma) responses and antigen-specific IFN-gamma ELISPOT activities 2 months after final immunization and after challenge. Antigen-specific CD4(+) and CD8(+) T cell responses in the combined DNA vaccine group were higher than either the Bacillus Calmette-Guerin (BCG)-positive or S19-positive control group. Likewise, more calves in the DNA vaccine group exhibited antigen-specific IgG titers and had higher IgG titers than those in the BCG- or S19-immunized groups 2 months after the final immunization. Moreover, two antigens in the combined DNA vaccine induced significant antigen-specific IFN-gamma responses 6 months after challenge (p S19 against B. abortus. This is the first report to demonstrate that a single combined DNA vaccine protects cattle against two infectious diseases. PMID:19364278

  4. Characterisation of Brucella abortus strain 19 cultures isolated from vaccinated cattle.

    Science.gov (United States)

    Thomas, E L; Bracewell, C D; Corbel, M J

    1981-01-31

    Thirty-four cultures recovered from material of bovine origin in England, Scotland and Wales were identified unequivocally as Brucella abortus strain 19 (S19). All had the properties of carbon dioxide-independent B abortus biotype 1 strains, were inhibited by penicillin G and thionin blue at standard concentrations and behaved in oxidative metabolism and guinea pig virulence tests as typical S19. Their sensitivity to i-erythritol varied somewhat between cultures as did reference subcultures of S19. Of the total number of isolates, 11 were recovered from abortion material or cyetic products, 10 were from calves which died from a hypersensitivity reaction within 24 hours of S19 vaccination and the remainder were from milk or internal organs. From the evidence available, there is little to suggest that calfhood vaccination with S19 has resulted in persistent systemic infection in other than a very small proportion of the animals inoculated. PMID:6789543

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

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    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

  6. Identification of new IS711 insertion sites in Brucella abortus field isolates

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    Moriyón Ignacio

    2011-08-01

    Full Text Available Abstract Background Brucellosis is a zoonosis caused by Brucella spp., a group of highly homogeneous bacteria. The insertion sequence IS711 is characteristic of these bacteria, and occurs in variable numbers and positions, but always constant within a given species. This species-associated polymorphism is used in molecular typing and identification. Field isolates of B. abortus, the most common species infecting cattle, typically carry seven IS711 copies (one truncated. Thus far, IS711 transposition has only been shown in vitro and only for B. ovis and B. pinnipedialis, two species carrying a high number of IS711 copies, but never in other Brucella species, neither in vitro nor in field strains. Results We found several B. abortus strains isolated from milk and aborted fetuses that carried additional IS711 copies in two hitherto undescribed insertion sites: one in an intergenic region near to the 3' end of a putative lactate permease gene and the other interrupting the sequence of a marR transcriptional regulator gene. Interestingly, the second type of insertion was identified in isolates obtained repeatedly from the same herd after successive brucellosis outbreaks, an observation that proves the stability and virulence of the new genotype under natural conditions. Sequence analyses revealed that the new copies probably resulted from the transposition of a single IS711 copy common to all Brucella species sequenced so far. Conclusions Our results show that the replicative transposition of IS711 can occur under field conditions. Therefore, it represents an active mechanism for the emergence of genetic diversity in B. abortus thus contributing to intra-species genetic polymorphism.

  7. The Brucella abortus cyclic beta-1,2-glucan virulence factor is substituted with O-ester-linked succinyl residues.

    Science.gov (United States)

    Roset, Mara S; Ciocchini, Andrés E; Ugalde, Rodolfo A; Iñón de Iannino, Nora

    2006-07-01

    Brucella periplasmic cyclic beta-1,2-glucan plays an important role during bacterium-host interaction. Nuclear magnetic resonance spectrometry analysis, thin-layer chromatography, and DEAE-Sephadex chromatography were used to characterize Brucella abortus cyclic glucan. In the present study, we report that a fraction of B. abortus cyclic beta-1,2-glucan is substituted with succinyl residues, which confer anionic character on the cyclic beta-1,2-glucan. The oligosaccharide backbone is substituted at C-6 positions with an average of two succinyl residues per glucan molecule. This O-ester-linked succinyl residue is the only substituent of Brucella cyclic glucan. A B. abortus open reading frame (BAB1_1718) homologous to Rhodobacter sphaeroides glucan succinyltransferase (OpgC) was identified as the gene encoding the enzyme responsible for cyclic glucan modification. This gene was named cgm for cyclic glucan modifier and is highly conserved in Brucella melitensis and Brucella suis. Nucleotide sequencing revealed that B. abortus cgm consists of a 1,182-bp open reading frame coding for a predicted membrane protein of 393 amino acid residues (42.7 kDa) 39% identical to Rhodobacter sphaeroides succinyltransferase. cgm null mutants in B. abortus strains 2308 and S19 produced neutral glucans without succinyl residues, confirming the identity of this protein as the cyclic-glucan succinyltransferase enzyme. In this study, we demonstrate that succinyl substituents of cyclic beta-1,2-glucan of B. abortus are necessary for hypo-osmotic adaptation. On the other hand, intracellular multiplication and mouse spleen colonization are not affected in cgm mutants, indicating that cyclic-beta-1,2-glucan succinylation is not required for virulence and suggesting that no low-osmotic stress conditions must be overcome during infection. PMID:16816173

  8. Brucella abortus Strain 2308 Wisconsin Genome: Importance of the Definition of Reference Strains

    Science.gov (United States)

    Suárez-Esquivel, Marcela; Ruiz-Villalobos, Nazareth; Castillo-Zeledón, Amanda; Jiménez-Rojas, César; Roop II, R. Martin; Comerci, Diego J.; Barquero-Calvo, Elías; Chacón-Díaz, Carlos; Caswell, Clayton C.; Baker, Kate S.; Chaves-Olarte, Esteban; Thomson, Nicholas R.; Moreno, Edgardo; Letesson, Jean J.; De Bolle, Xavier; Guzmán-Verri, Caterina

    2016-01-01

    Brucellosis is a bacterial infectious disease affecting a wide range of mammals and a neglected zoonosis caused by species of the genetically homogenous genus Brucella. As in most studies on bacterial diseases, research in brucellosis is carried out by using reference strains as canonical models to understand the mechanisms underlying host pathogen interactions. We performed whole genome sequencing analysis of the reference strain B. abortus 2308 routinely used in our laboratory, including manual curated annotation accessible as an editable version through a link at https://en.wikipedia.org/wiki/Brucella#Genomics. Comparison of this genome with two publically available 2308 genomes showed significant differences, particularly indels related to insertional elements, suggesting variability related to the transposition of these elements within the same strain. Considering the outcome of high resolution genomic techniques in the bacteriology field, the conventional concept of strain definition needs to be revised. PMID:27746773

  9. Use of an indirect elisa for Brucella abortus diagnosis in Cuba

    International Nuclear Information System (INIS)

    Introducing immunoassays in Brucella diagnosis requires a comparative study with reference techniques such as the complement fixation reaction (CFR). Sensitivity and relative specificity studies allowed us to observe the behaviour of this immunoassay, using samples from free of disease, free by vaccination and affected areas. Sensitivity results for a cut-off point of 40PP and a confidence interval of 95% ranged from 94.8 to 99.5% and the specificity between 94.1 and 97.5%. For free of disease areas a cut-off point of 22 PP was calculated that reached a 99% specificity. This immunoassay for the detection of antibodies against Brucella abortus must be used with two different cut-off points, depending on the epidemiologic conditions of the country, with CFR in affected or vaccinated areas as a confirmative method. (author)

  10. Identification of Brucella abortus virulence proteins that modulate the host immune response.

    Science.gov (United States)

    Wang, Yufei; Chen, Zeliang; Qiu, Yefeng; Ke, Yuehua; Xu, Jie; Yuan, Xitong; Li, Xianbo; Fu, Simei; Cui, Mingquan; Xie, Yongfei; Du, Xinying; Wang, Zhoujia; Huang, Liuyu

    2012-01-01

    Brucellosis is an important zoonotic disease of almost worldwide distribution. One significant immune phenomenon of this disease is the ability of the pathogen to hide and survive in the host, establishing long lasting chronic infections. Brucella was found to have the ability to actively modulate the host immune response in order to establish chronic infections, but the mechanism by which the pathogen achieves this remains largely unknown. In our screening for protective antigens of Brucella abortus, 3 proteins (BAB1_0597, BAB1_0917, and BAB2_0431) were found to induce significantly higher levels of gamma interferon (IFNγ) in splenocytes of PBS immunized mice than those immunized with S19. This finding strongly implied that these three proteins inhibit the production of IFNγ. Previous studies have shown that LPS, PrpA, and Btp1/TcpB are three important immunomodulatory molecules with the capacity to interfere with host immune response. They have been shown to have the ability to inhibit the secretion of IFNγ, or to increase the production of IL-10. Due to the role of these proteins in virulence and immunomodulation, they likely offer significant potential as live, attenuated Brucella vaccine candidates. Understanding the mechanisms by which these proteins modulate the host immune responses will deepen our knowledge of Brucella virulence and provide important information on the development of new vaccines against Brucellosis. PMID:22743689

  11. A Live Vaccine from Brucella abortus Strain 82 for Control of Cattle Brucellosis in the Russian Federation

    Science.gov (United States)

    During the first half of the 20th century, widespread regulatory efforts to control cattle brucellosis (Brucella abortus) in the Union of Soviet Socialist Republics were essentially nonexistent, and control was limited to selective test and slaughter of serologic agglutination reactors. By the 1950...

  12. 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...

  13. Draft Genome Sequence of the Intermediate Rough Vaccine Strain Brucella abortus S19Δper Mutant

    Science.gov (United States)

    Chaudhuri, Pallab; Goswami T, Tapas K.; Lalsiamthara, Jonathan; Kaur, Gurpreet; Vishnu, Udayakumar S.; Sankarasubramanian, Jagadesan; Gunasekaran, Paramasamy

    2015-01-01

    Here, we report the genome sequence of the intermediate rough vaccine strain mutant, Brucella abortus S19Δper. The length of the draft genome was 3,271,238 bp, with 57.2% G+C content. A total of 3,204 protein-coding genes and 56 RNA genes were predicted. PMID:26564050

  14. Genomic fingerprinting and development of a dendrogram for Brucella spp. isolated from seals, porpoises, and dolphins.

    Science.gov (United States)

    Jensen, A E; Cheville, N F; Thoen, C O; MacMillan, A P; Miller, W G

    1999-03-01

    Genomic DNA from reference strains and biovars of the genus Brucella was analyzed using pulsed-field gel electrophoresis (PFGE). Fingerprints were compared to estimate genetic relatedness among the strains and to obtain information on evolutionary relationships. Electrophoresis of DNA digested with the restriction endonuclease XbaI produced fragment profiles for the reference type strains that distinguished these strains to the level of species. Included in this study were strains isolated from marine mammals. The PFGE profiles from these strains were compared with those obtained from the reference strains and biovars. Isolates from dolphins had similar profiles that were distinct from profiles of Brucella isolates from seals and porpoises. Distance matrix analyses were used to produce a dendrogram. Biovars of B. abortus were clustered together in the dendrogram; similar clusters were shown for biovars of B. melitensis and for biovars of B. suis. Brucella ovis, B. canis, and B. neotomae differed from each other and from B. abortus, B. melitensis, and B. suis. The relationship between B. abortus strain RB51 and other Brucella biovars was compared because this strain has replaced B. abortus strain 19 for use as a live vaccine in cattle and possibly in bison and elk. These results support the current taxonomy of Brucella species and the designation of an additional genomic group(s) of Brucella. The PFGE analysis in conjunction with distance matrix analysis was a useful tool for calculating genetic relatedness among the Brucella species. PMID:10098687

  15. Evaluation of Immunogenicity of Divalent DNA Vaccine Encoding Brucella melitensis Omp31 and P39 Genes in Balb/c Mice

    OpenAIRE

    A Doosti; GR Javadi; Sardari, S.; MA Shokrgozar; P Ghassemi-Dehkordi

    2007-01-01

    Introduction & Objective: Brucella is a facultative intracellular pathogen and one of the etiologic agents of brucellosis that can infect humans and domestic animals. Attenuated strains such as B. melitensis Rve1 and B. abortus S19 and Rb51 are being used to control brucellosis in domestic animals. However, no safe and effective vaccine is available for human use. This study was designed to evaluate the immunogenicity and the protective efficacy of a divalent fusion DNA vaccine encoding both ...

  16. Immunoproteomics of Brucella abortus reveals differential antibody profiles between S19-vaccinated and naturally infected cattle.

    Science.gov (United States)

    Pajuaba, Ana C A M; Silva, Deise A O; Almeida, Karine C; Cunha-Junior, Jair P; Pirovani, Carlos P; Camillo, Luciana R; Mineo, José R

    2012-03-01

    Brucella abortus is a Gram-negative intracellular bacterium that causes infectious abortion in food-producing animals and chronic infection in humans. This study aimed to characterize a B. abortus S19 antigen preparation obtained by Triton X-114 (TX-114) extraction through immunoproteomics to differentiate infected from vaccinated cattle. Three groups of bovine sera were studied: GI, 30 naturally infected cows; GII, 30 S19-vaccinated heifers; and GIII, 30 nonvaccinated seronegative cows. One-dimensional (1D) and two-dimensional electrophoretic profiles of TX-114 hydrophilic phase antigen revealed a broad spectrum of polypeptides (10-79 kDa). 1D immunoblot showed widespread seroreactivity profile in GI compared with restricted profile in GII. Three antigenic components (10, 12, 17 kDa) were recognized exclusively by GI sera, representing potential markers of infection and excluding vaccinal response. The proteomic characterization revealed 56 protein spots, 27 of which were antigenic spots showing differential seroreactivity profile between GI and GII, especially polypeptides abortus S19 proteins (Invasion protein B, Sod, Dps, Ndk, and Bfr), which were related with antigenicity in naturally infected cattle. In conclusion, immunoproteomics of this new antigen preparation enabled the characterization of proteins that could be used as tools to develop sensitive and specific immunoassays for serodiagnosis of bovine brucellosis, with emphasis on differentiation between S19 vaccinated and infected cattle. PMID:22539433

  17. Second generation competitive enzyme immunoassay for detection of bovine antibody to Brucella abortus.

    Science.gov (United States)

    Nielsen, K; Smith, P; Yu, W L; Elmgren, C; Nicoletti, P; Perez, B; Bermudez, R; Renteria, T

    2007-09-20

    A second generation competitive enzyme immunoassay (CELISA) for detection of bovine antibody to Brucella abortus was developed. This assay was different from previously developed CELISAs in that the detection reagent used was a recombinant combination of the receptor portions of protein A and protein G, labelled with horseradish peroxidase. This eliminates the need for polyclonal anti-mouse-enzyme conjugate reagents for detection thus allowing for true standardization. The assay utilized a monoclonal antibody specific for a common epitope of the O-polysaccharide (OPS) of smooth lipopolysaccharide (SLPS) derived from B. abortus S1119.3 but which did not react with protein A/G. This monoclonal antibody was used to compete with antibody in the bovine test serum. Binding of bovine antibody to the smooth lipopolysaccharide antigen was then measured directly with the protein A/G enzyme conjugate. In this case, development of colour in the reaction was indicative of the presence of bovine antibody. The performance characteristics, sensitivity, specificity and exclusion of B. abortus S19 vaccinated animals, of the assay were very similar to those of the classical CELISA. PMID:17467200

  18. Transcriptome analysis of the Brucella abortus BvrR/BvrS two-component regulatory system.

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    Cristina Viadas

    Full Text Available BACKGROUND: The two-component BvrR/BvrS system is essential for Brucella abortus virulence. It was shown previously that its dysfunction alters the expression of some major outer membrane proteins and the pattern of lipid A acylation. To determine the genes regulated by BvrR/BvrS, we performed a whole-genome microarray analysis using B. abortus RNA obtained from wild type and bvrR mutant cells grown in the same conditions. METHODOLOGY/PRINCIPAL FINDINGS: A total of 127 differentially expressed genes were found: 83 were over expressed and 44 were less expressed in the bvrR mutant. Two operons, the phosphotransferase system and the maltose transport system, were down-regulated. Several genes involved in cell envelope or outer membrane biogenesis were differentially expressed: genes for outer membrane proteins (omp25a, omp25d, lipoproteins, LPS and fatty acid biosynthesis, stress response proteins, chaperones, flagellar genes, and twelve genes encoding ABC transport systems. Ten genes related with carbon metabolism (pckA and fumB among others were up-regulated in the bvrR mutant, and denitrification genes (nirK, norC and nosZ were also regulated. Notably, seven transcriptional regulators were affected, including VjbR, ExoR and OmpR that were less expressed in the bvrR mutant. Finally, the expression of eleven genes which have been previously related with Brucella virulence was also altered. CONCLUSIONS/SIGNIFICANCE: All these data corroborate the impact of BvrR/BvrS on cell envelope modulation, confirm that this system controls the carbon and nitrogen metabolism, and suggest a cross-talk among some regulators to adjust the Brucella physiology to the shift expected to occur during the transit from the extracellular to the intracellular niche.

  19. Heterologous expression of Brucella abortus GroEL heat-shock protein in Lactococcus lactis

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    Langella Philippe

    2006-03-01

    Full Text Available Abstract Background Brucella abortus is a facultative intracellular pathogen that mainly infects cattle and humans. Current vaccines rely on live attenuated strains of B. abortus, which can revert to their pathogenic status and thus are not totally safe for use in humans. Therefore, the development of mucosal live vaccines using the food-grade lactic acid bacterium, Lactococcus lactis, as an antigen delivery vector, is an attractive alternative and a safer vaccination strategy against B. abortus. Here, we report the construction of L. lactis strains genetically modified to produce B. abortus GroEL heat-shock protein, a candidate antigen, in two cellular locations, intracellular or secreted. Results Only the secreted form of GroEL was stably produced in L. lactis, suggesting a detrimental effect of GroEL protein when intracellularly produced in this bacterium. Only trace amounts of mature GroEL were detected in the supernatant fraction of induced lactococcal cultures, and the GroEL precursor remained stacked in the cell fraction. Attempts to raise the secretion yields were made, but even when GroEL was fused to a synthetic propeptide, secretion of this antigen was not improved. Conclusion We found that L. lactis is able to produce, and to secrete, a stable form of GroEL into the extracellular medium. Despite the low secretion efficiency of GroEL, which suggest that this antigen interacts with the cell envelope of L. lactis, secretion seems to be the best way to achieve both production and protein yields, regardless of cellular location. The L. lactis strain secreting GroEL has potential for in vivo immunization.

  20. 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.

  1. Brucella abortus Choloylglycine Hydrolase Affects Cell Envelope Composition and Host Cell Internalization

    Science.gov (United States)

    Marchesini, María Inés; Connolly, Joseph; Delpino, María Victoria; Baldi, Pablo C.; Mujer, Cesar V.; DelVecchio, Vito G.; Comerci, Diego J.

    2011-01-01

    Choloylglycine hydrolase (CGH, E.C. 3.5.1.24) is a conjugated bile salt hydrolase that catalyses the hydrolysis of the amide bond in conjugated bile acids. Bile salt hydrolases are expressed by gastrointestinal bacteria, and they presumably decrease the toxicity of host's conjugated bile salts. Brucella species are the causative agents of brucellosis, a disease affecting livestock and humans. CGH confers Brucella the ability to deconjugate and resist the antimicrobial action of bile salts, contributing to the establishment of a successful infection through the oral route in mice. Additionally, cgh-deletion mutant was also attenuated in intraperitoneally inoculated mice, which suggests that CGH may play a role during systemic infection other than hydrolyzing conjugated bile acids. To understand the role CGH plays in B. abortus virulence, we infected phagocytic and epithelial cells with a cgh-deletion mutant (Δcgh) and found that it is defective in the internalization process. This defect along with the increased resistance of Δcgh to the antimicrobial action of polymyxin B, prompted an analysis of the cell envelope of this mutant. Two-dimensional electrophoretic profiles of Δcgh cell envelope-associated proteins showed an altered expression of Omp2b and different members of the Omp25/31 family. These results were confirmed by Western blot analysis with monoclonal antibodies. Altogether, the results indicate that Brucella CGH not only participates in deconjugation of bile salts but also affects overall membrane composition and host cell internalization. PMID:22174816

  2. Recombinant bovine interleukin 2 enhances immunity and protection induced by Brucella abortus vaccines in cattle.

    Science.gov (United States)

    Wyckoff, John H; Howland, Jeri L; Scott, Catherine M O'Connell; Smith, Robert A; Confer, Anthony W

    2005-11-30

    Augmentation of immunization of cattle Brucella abortus S19 or a B. abortus soluble protein extract (SPEBA) vaccine through administration of recombinant bovine IL 2 (rBoIL 2) was evaluated. Seventy-five heifers were divided among 6 groups that were treated with the following: Group 1, no treatment; Group 2, rBoIL 2 (1microg/kg) on day 0; Group 3, SPEBA (2 mg) on day 0 and week 9; Group 4, SPEBA + rBoIL 2 on day 0, SPEBA on week 9; Group 5, S19 (10(7) CFU) on day 0 and week 9; Group 6, S19 + rBoIL 2 on day 0, S19 only on week 9. Approximately, 6 months after vaccination, cattle were bred by natural service, and at mid-gestation pregnant cattle were challenged intraconjunctivally with 9.1 x 10(5) CFU of virulent B. abortus S2308. Pre- and post-challenge antibody responses were measured by an enzyme-linked immunosorbent assay, a particle concentration fluorescence assay, and the card test. Lymphoproliferation (LP) responses to gamma-irradiated B. abortus and SPEBA antigens were measured in peripheral blood mononuclear cells. After vaccination, antibody responses to B. abortus elevated rapidly in SPEBA- and S19-vaccinates with and without rBoIL 2, however, these responses were significantly (P S19 resulted in significant (P abortus antigens following challenge. Characterization of the cytokine response of bovine monocyte-derived macrophages by real-time polymerase chain reaction indicated that in vitro stimulation of these cells with rBoIL 2 resulted in a profound up-regulation of genes encoding tumor necrosis factor-alpha, IL 12p40, and interferon-gamma reflecting activation of the cells. Overall, rBoIL 2-treatment was associated with fewer infections, sero-conversions and a significant (P = 0.02) level of protection against abortion as compared to vaccination alone or no treatment. PMID:16242273

  3. Otpornost na antimikrobne tvari, fenotipska obilježja i fagotipovi sojeva vrste B. abortus izdvojenih iz goveda i azijskih bivola (Bubalus bubalis) u Trinidadu.

    OpenAIRE

    Adesiyun, Abiodun A.; Baird, Kadian; Stewart-Johnson, Alva

    2011-01-01

    Određivana su fenotipska obilježja, fagotipovi i otpornost na antimikrobna sredstva sojeva bakterije Brucella abortus izdvojenih iz goveda i azijskog bivola (Bubalus bubalis) u Trinidadu i Tobagu. Ukupno je 86 izolata bilo izdvojeno iz limfnih čvorova 14 klinički zdravih serološki pozitivnih goveda i 17 indijskih bivola, iz ozljeda kože devet bivola te tkiva pobačenih plodova 16 bivola. Analizirana su bila i dva cjepna soja bakterije B. abortus, soj 19 (S19) i RB51 (SRB51). Svi analizirani so...

  4. 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

  5. An ecological perspective on the changing face of Brucella abortus in the western United States

    Science.gov (United States)

    Cross, Paul C.; Maichak, Eric J.; Brennan, Angela; Scurlock, Brandon M.; Henningsen, John C.; Luikart, Gordon

    2013-01-01

    After a hiatus during the 1990s, outbreaks of Brucella abortus in cattle are occurring more frequently in some of the western states of the United States, namely, Montana, Wyoming and Idaho. This increase is coincident with increasing brucellosis seroprevalence in elk (Cervus elaphus), which is correlated with elk density. Vaccines are a seductive solution, but their use in wildlife systems remains limited by logistical, financial, and scientific constraints. Cattle vaccination is ongoing in the region. Livestock regulations, however, tend to be based on serological tests that test for previous exposure and available vaccines do not protect against seroconversion. The authors review recent ecological studies of brucellosis, with particular emphasis on the Greater Yellowstone Area, and highlight the management options and implications of this work, including the potential utility of habitat modifications and targeted hunts, as well as scavengers and predators. Finally, the authors discuss future research directions that will help us to understand and manage brucellosis in wildlife.

  6. 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.

  7. Crystal structure of cyclic nucleotide-binding-like protein from Brucella abortus.

    Science.gov (United States)

    He, Zheng; Gao, Yuan; Dong, Jing; Ke, Yuehua; Li, Xuemei; Chen, Zeliang; Zhang, Xuejun C

    2015-12-25

    The cyclic nucleotide-binding (CNB)-like protein (CNB-L) from Brucella abortus shares sequence homology with CNB domain-containing proteins. We determined the crystal structure of CNB-L at 2.0 Å resolution in the absence of its C-terminal helix and nucleotide. The 3D structure of CNB-L is in a two-fold symmetric form. Each protomer shows high structure similarity to that of cGMP-binding domain-containing proteins, and likely mimics their nucleotide-free conformation. A key residue, Glu17, mediates the dimerization and prevents binding of cNMP to the canonical ligand-pocket. The structurally observed dimer of CNB-L is stable in solution, and thus is likely to be biologically relevant.

  8. DETECCION DE Brucella abortus POR PCR EN MUESTRAS DE SANGRE Y LECHE DE VACUNOS

    Directory of Open Access Journals (Sweden)

    Xiomara Mosquera C

    2008-12-01

    Full Text Available Objetivo. Evaluar el uso de la Reacción en Cadena de la Polimerasa (PCR para la detección de Brucella abortus en muestras de sangre y leche de vacunos. Materiales y métodos. Este estudio de tipo descriptivo fue realizado durante los años 2004 y 2005. Se analizaron 136 animales de tres fincas localizadas en el municipio de Durania, Norte de Santander, Colombia. Se evaluó la presencia de anticuerpos en la leche mediante la prueba del anillo (PAL. Se amplificó el fragmento de 223pb del gen BCSP31. Se emplearon los cebadores B4 y B5 de la región interna de la secuencia del gen BCSP31 (GenBank, número M20404. Resultados. En aquellos animales positivos se obtuvo una muestra de sangre y leche para el análisis por PCR, la sangre no fue analizada por serología. Se evaluaron diferentes métodos de extracción de ADN. Se encontró que un 13.2% (18/136 de las muestras de leche fueron positivas a la PAL. Se analizaron 33 muestras de leche negativas por PAL de las cuales el 30.3% (10/33 resultaron positivas por PCR. Al analizar las muestras de sangre de los animales positivos por PAL el 94.1% (16/17 fueron positivas por PCR, mientras que el 47% (8/17 de las muestras de leche positivas por PAL, fueron positivas por PCR. Conclusiones. Se demostró la amplificación de un fragmento de ADN de Brucella abortus en muestras de sangre y leche de vacunos. Los resultados preliminares demostraron que es posible usar PCR como prueba diagnóstica de brucelosis en Colombia.

  9. DNA vaccine encoding L7/L12-P39 of Brucella abortus induces protective immunity in BALB/c mice

    Institute of Scientific and Technical Information of China (English)

    LUO De-yan; LI Peng; XING Li; ZHAO Guang-yu; SHI Wei; ZHANG Song-le; WANG Xi-liang

    2006-01-01

    @@ Brucella abortus is a gram-negative, facultative, intracellular bacterium that infects both cattle and humans, causing abortion and infertility in the former and undulant fever, endocarditis, arthritis, and osteomyelitis. Resistance to Brucella depends on acquired cell-mediated immunity (CMI).1 Live attenuated vaccines can stimulate strong CMI response, which are usually very effective against brucellosis and are used to control brucellosis in domestic animals. However, there is no safe and effective vaccine available for human because the vaccine strains used for animals are considered too virulent for humans. A vaccine that will be noninfectious to humans but effective in stimulating a broad protective immune response is needed.2

  10. Effects of partial deletion of the wzm and wzt genes on lipopolysaccharide synthesis and virulence of Brucella abortus S19.

    Science.gov (United States)

    Wang, Xiuran; Wang, Lin; Lu, Tiancheng; Yang, Yanling; Chen, Si; Zhang, Rui; Lang, Xulong; Yan, Guangmou; Qian, Jing; Wang, Xiaoxu; Meng, Lingyi; Wang, Xinglong

    2014-06-01

    Brucellosis is a worldwide human and animal infectious disease, and the effective methods of its control are immunisation of animals by vaccination and elimination. Brucella abortus S19 is one of the popular vaccines with virulence in the control of cattle Brucellosis. In the present study, allelic exchange plasmids of wzm and wzt genes and partial knockout mutants of wzm and wzt were constructed to evaluate the resulting difference in virulence of B. abortus S19. PCR analysis revealed that the target genes were knocked out. The mutants were rough mutants and they could be differentiated from natural infection by the Rose Bengal plate and standard agglutination tests. The molecular weights of lipopolysaccharides of the Δwzm and Δwzt mutants were clustered between 25 and 40 kDa, and 30 and 35 kDa separately, and were markedly different from those in B. abortus S19. The virulence of B. abortus Δwzm and Δwzt was decreased compared with that of B. abortus S19 in mice. All these results identified that there were several differences between the wzm and wzt genes on lipopolysaccharide synthesis and on the virulence of B. abortus. PMID:24718931

  11. Optimization of production of Brucella abortus S19 culture in bioreactor using soyabean casein digest medium.

    Science.gov (United States)

    Kamaraj, Govindasamy; Rajendra, Lingala; Shankar, Chinchkar Ramachandra; Srinivasan, Villuppanoor Alwar

    2010-10-01

    A method of cultivating Brucella abortus S19 culture in bioreactor was attempted using three different media. Culture conditions in bioreactor were optimized by varying agitation and aeration parameters. Varying the aeration ranging from 0.5 vvm to 0.8 vvm and agitation rate ranging from 250 rpm to 400 rpm during bacterial growth was found to yield highest viable count within 48 hours of culture period. A count of > 1 x 10(11) CFU per ml within 48 to 60 hours post seeding was obtained consistently in all five consecutive batches (P > 0.05) with 6 x 10(11) CFU per ml being the maximum yield when the organism is grown in soyabean casein digest medium. B. abortus S19 maintained its smooth characteristics throughout its growth in bioreactor. The vaccine prepared with soyabean casein digest medium was found to be potent and safe with a protective index of 3.33 in mice. The vaccine was tested in 10 cattle calves of 3 to 13 months age and all the vaccinated animals were seropositive on 28, 60, 90, 120 and 150 days post-vaccination when analyzed by fluorescence polarization assay (FPA). PMID:21213590

  12. Molecular structure of the Brucella abortus metalloprotein RicA, a Rab2-binding virulence effector.

    Science.gov (United States)

    Herrou, Julien; Crosson, Sean

    2013-12-17

    The Gram-negative intracellular pathogen Brucella abortus is the causative agent of brucellosis, which is among the most common zoonoses globally. The B. abortus RicA protein binds the host-expressed guanosine nucleotide-binding protein, Rab2, and modulates B. abortus infection biology. We have solved the first X-ray crystal structure of RicA to 2.7 Å resolution and have quantified the affinity of RicA binding to human Rab2 in its GDP-bound and nucleotide-free forms. RicA adopts a classic γ-carbonic anhydrase (γ-CA) fold containing a left-handed β-helix followed by a C-terminal α-helix. Two homotrimers of RicA occupy the crystallographic asymmetric unit. Though no zinc was included in the purification or crystallization buffers, zinc is contained within the RicA crystals, as demonstrated by X-ray fluorescence spectroscopy. Electron density for a Zn(2+) ion coordinated by three histidine residues is evident in the putative active site of RicA. However, purified RicA preparations do not exhibit carbonic anhydrase activity, suggesting that Zn(2+) may not be the physiologically relevant metal cofactor or that RicA is not a bona fide carbonic anhydrase enzyme. Isothermal titration calorimetry (ITC) measurements of purified RicA binding to purified human Rab2 and GDP-Rab2 revealed similar equilibrium affinities (Kd ≈ 35 and 40 μM, respectively). This study thus defines RicA as a Zn(2+)-binding γ-carbonic anhydrase-like protein that binds the human membrane fusion/trafficking protein Rab2 with low micromolar affinity in vitro. These results support a model in which γ-CA family proteins may evolve unique cellular functions while retaining many of the structural hallmarks of archetypal γ-CA enzymes.

  13. Seroprevalence of brucellosis in sheep and isolation of Brucella abortus biovar 6 in Kassala State, Eastern Sudan.

    Science.gov (United States)

    Gumaa, M M; Osman, H M; Omer, M M; El Sanousi, E M; Godfroid, J; Ahmed, A M

    2014-12-01

    Brucellosis is one of the important zoonotic diseases among livestock. This study was carried out to estimate the prevalence of brucellosis and isolate Brucella spp. in sheep in Kassala State in the east of Sudan. Two thousand and five serum samples were randomly collected from nine different localities. All serum samples were examined by the Rose Bengal plate test (RBPT) and the modified RBPT (mRBPT). Forty-three (2.15%, 95% confidence interval [CI]: 1.6,3.0) and 68 (3.4%, 95% CI: 2.6, 4.2) samples were positive with the RBPT and the mRBPT, respectively. According to a known diagnostic sensitivity of 86.6% and a known diagnostic specificity of 97.6% for the mRBPT, the true prevalence was estimated to be 1.2% (95% CI: 0.3, 2.2). Different tissue samples were collected from 41 mRBPT seropositive animals. Brucella abortus biovar 6 was isolated from a pyometra of a seropositive ewe. It is important to note that B. abortus biovar 6 cannot be differentiated from Brucella melitensis biovar 2 by routine bacteriology. Only phage typing performed in reference laboratories will allow accurate identification of the strain. The fact that B. abortus biovar 6 does not require CO2 for growth, combined with the fact that it has been isolated from a small ruminant in this study, could easily have led to misidentification (as B. melitensis biovar 2), to wrong epidemiological inferences and to the implementation of inappropriate control measures. The results presented here suggest that sheep are spillover hosts, as previously described for camels, and that the actual reservoir of B. abortus biovar 6 is cattle in Kassala State, Eastern Sudan. This study highlights the importance of isolating and identifying Brucella spp. in different livestock species in order to accurately decipher brucellosis epidemiology in sub-Saharan Africa. PMID:25812219

  14. Reaction of Native and Denatured Brucella abortus (S19) Proteins with Antibody Using Affinity Chromatography and Immunoblotting

    OpenAIRE

    Karimi, R.; A Mostafaie; B. Tabaraie; Y. Bahrami; J. Abdolalizadeh

    2005-01-01

    Western blotting or immunoblotting commonly use for study of reaction between antigens and antibodies. Denaturation of many proteins in immunoblotting can affect greatly the reactivity of antibodies and outcome of the procedure.In this study proteins of Brucella abortus (S19) was extracted by a mild method and reaction of the extracted proteins with serum of infected human and goat and immunized rabbit compared by affinity chromatography and immunoblotting. Gamma globulin (m...

  15. Brucella abortus ornithine lipids are dispensable outer membrane components devoid of a marked pathogen-associated molecular pattern.

    Directory of Open Access Journals (Sweden)

    Leyre Palacios-Chaves

    Full Text Available The brucellae are α-Proteobacteria facultative intracellular parasites that cause an important zoonosis. These bacteria escape early detection by innate immunity, an ability associated to the absence of marked pathogen-associated molecular patterns in the cell envelope lipopolysaccharide, lipoproteins and flagellin. We show here that, in contrast to the outer membrane ornithine lipids (OL of other Gram negative bacteria, Brucella abortus OL lack a marked pathogen-associated molecular pattern activity. We identified two OL genes (olsB and olsA and by generating the corresponding mutants found that olsB deficient B. abortus did not synthesize OL or their lyso-OL precursors. Liposomes constructed with B. abortus OL did not trigger IL-6 or TNF-α release by macrophages whereas those constructed with Bordetella pertussis OL and the olsB mutant lipids as carriers were highly active. The OL deficiency in the olsB mutant did not promote proinflammatory responses or generated attenuation in mice. In addition, OL deficiency did not increase sensitivity to polymyxins, normal serum or complement consumption, or alter the permeability to antibiotics and dyes. Taken together, these observations indicate that OL have become dispensable in the extant brucellae and are consistent within the trend observed in α-Proteobacteria animal pathogens to reduce and eventually eliminate the envelope components susceptible of recognition by innate immunity.

  16. 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

    2015-05-01

    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.

  17. Circulating strains of Brucella abortus in cattle in the province of Santo Domingo de los Tsáchilas - Ecuador.

    Directory of Open Access Journals (Sweden)

    Richar Ivan Rodríguez Hidalgo

    2015-03-01

    Full Text Available In Ecuador, the Province of Santo Domingo de los Tsáchilas represents the largest informal market of livestock due to its strategic position in the country; thus, given the high mobility of cattle in this region, the aim of this study was to determine the strain variation of Brucella sp.. Part of the study was the isolation, biotyping and genotyping of Brucella species isolated from milk and supra-mammary lymph nodes of sero-positive bovines. Protocols used for isolation, biotyping and genotyping of Brucella species were selective Farrell medium, biochemical assays and IS711-PCR, AMOS-PCR and HOOF-Prints techniques, respectively. In total, 656 animals from sero-positive dairy herds and from the slaughterhouse of the province were diagnosed by Rose Bengal and Wright’s Slow Agglutination test with EDTA. From these animals, 50 animals were found sero-positive for brucellosis. Twenty-five lymph nodes and 25 milk samples from each group of positive reactors were cultured and growth. Isolation was possible in 4 (16% and 9 (36%, respectively; and of these, 10 isolates were diagnosed as Brucella sp. All 4 isolates of lymphatic tissue corresponded to Brucella abortus biotype 1, confirmed as field strains by molecular analysis. Milk isolations, biochemically showed a more dispersed pattern in which B. abortus biotypes 1 and 4 were found; yet four samples gave a pattern similar to B. abortus biotype 2; however, only biotypes 1 and 4 were confirmed by molecular analysis. The concordance between biochemical and molecular diagnostic tests reached 76.9%.

  18. Immunization of mice with recombinant protein CobB or AsnC confers protection against Brucella abortus infection.

    Directory of Open Access Journals (Sweden)

    Simei Fu

    Full Text Available Due to drawbacks of live attenuated vaccines, much more attention has been focused on screening of Brucella protective antigens as subunit vaccine candidates. Brucella is a facultative intracellular bacterium and cell mediated immunity plays essential roles for protection against Brucella infection. Identification of Brucella antigens that present T-cell epitopes to the host could enable development of such vaccines. In this study, 45 proven or putative pathogenesis-associated factors of Brucella were selected according to currently available data. After expressed and purified, 35 proteins were qualified for analysis of their abilities to stimulate T-cell responses in vitro. Then, an in vitro gamma interferon (IFN-γ assay was used to identify potential T-cell antigens from B. abortus. In total, 7 individual proteins that stimulated strong IFN-γ responses in splenocytes from mice immunized with B. abortus live vaccine S19 were identified. The protective efficiencies of these 7 recombinant proteins were further evaluated. Mice given BAB1_1316 (CobB or BAB1_1688 (AsnC plus adjuvant could provide protection against virulent B. abortus infection, similarly with the known protective antigen Cu-Zn SOD and the license vaccine S19. In addition, CobB and AsnC could induce strong antibodies responses in BALB/c mice. Altogether, the present study showed that CobB or AsnC protein could be useful antigen candidates for the development of subunit vaccines against brucellosis with adequate immunogenicity and protection efficacy.

  19. Immunization of mice with recombinant protein CobB or AsnC confers protection against Brucella abortus infection.

    Science.gov (United States)

    Fu, Simei; Xu, Jie; Li, Xianbo; Xie, Yongfei; Qiu, Yefeng; Du, Xinying; Yu, Shuang; Bai, Yaoxia; Chen, Yanfen; Wang, Tongkun; Wang, Zhoujia; Yu, Yaqing; Peng, Guangneng; Huang, Kehe; Huang, Liuyu; Wang, Yufei; Chen, Zeliang

    2012-01-01

    Due to drawbacks of live attenuated vaccines, much more attention has been focused on screening of Brucella protective antigens as subunit vaccine candidates. Brucella is a facultative intracellular bacterium and cell mediated immunity plays essential roles for protection against Brucella infection. Identification of Brucella antigens that present T-cell epitopes to the host could enable development of such vaccines. In this study, 45 proven or putative pathogenesis-associated factors of Brucella were selected according to currently available data. After expressed and purified, 35 proteins were qualified for analysis of their abilities to stimulate T-cell responses in vitro. Then, an in vitro gamma interferon (IFN-γ) assay was used to identify potential T-cell antigens from B. abortus. In total, 7 individual proteins that stimulated strong IFN-γ responses in splenocytes from mice immunized with B. abortus live vaccine S19 were identified. The protective efficiencies of these 7 recombinant proteins were further evaluated. Mice given BAB1_1316 (CobB) or BAB1_1688 (AsnC) plus adjuvant could provide protection against virulent B. abortus infection, similarly with the known protective antigen Cu-Zn SOD and the license vaccine S19. In addition, CobB and AsnC could induce strong antibodies responses in BALB/c mice. Altogether, the present study showed that CobB or AsnC protein could be useful antigen candidates for the development of subunit vaccines against brucellosis with adequate immunogenicity and protection efficacy. PMID:22383953

  20. 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

  1. Mouse cytokine profile skewed towards Th2 in pregnancy during infection with Brucella abortus S19 strain.

    Science.gov (United States)

    Wamonje, Francis O; Waihenya, Rebecca K; Ng'ang'a, Zipporah; Ngeranwa, Joseph N

    2011-04-01

    The two classes of cytokines Th1 and Th2 determine the type of immune response elicited. The Th2 immune response is associated with successful pregnancy. Brucellosis is an intracellular bacterium that elicits the Th1 response and is known to cause spontaneous abortion in mammalian species. This study sought to determine if Brucella infection causes spontaneous abortion by causing the circulating cytokine profile be Th1 dominant during pregnancy. Forty-eight Swiss white mice were used in this murine model and the S19 strain of Brucella abortus was used in as the infective agent. Pregnant mice in the test group were injected intraperitoneally with 10(5-8) CFU of Brucella and cytokine profile evaluated over the three trimesters of pregnancy. Pregnant mice in the control group were left to go through normal pregnancy and their cytokine profile evaluated over the three trimesters of pregnancy. Cytokines in serum samples were analyzed by Cytometric Bead Array. The data was analyzed using the Paired T- test and p Brucella abortus cannot cause spontaneous abortion by altering the mouse cytokine profile towards Th1 in pregnancy. Elevated IL-4 levels with corresponding suppression of IFN-γ can be used as a marker for successful pregnancy in Brucellosis. PMID:25566611

  2. 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.

  3. Protection levels in vaccinated heifers with experimental vaccines Brucella abortus M1-luc and INTA 2.

    Science.gov (United States)

    Fiorentino, M A; Campos, E; Cravero, S; Arese, A; Paolicchi, F; Campero, C; Rossetti, O

    2008-12-10

    Brucella abortus M1-luc is a mutant strain derived from S19 vaccine strain in which most of bp26 sequence has been replaced by the luciferase coding gene. Strain I2 is a double mutant derived from M1-luc in which most of omp19 has been deleted without introduction of any genetic markers. In BALB/c mice, M1-luc presented equivalent performance to S19 regarding persistence, splenomegaly and protection against challenge. Interestingly, I2 was more attenuated than S19, with no reduction of protection against challenge. In order to evaluate the potential for vaccine use of these strains in the natural host, four groups of 15 heifers, 6-month old, were either non-vaccinated or vaccinated with S19, M1-luc or I2. To at reached 17-month old, heifers were synchronized with two doses of PGF2alpha and received natural service during 60 days with two bulls. Pregnant heifers were challenged at approximately six gestation months with virulent B. abortus S2308. Blood samples post-challenge of heifers were collected for serologic test as well as specimens of aborted fetuses and premature calves for bacterial isolation and histopathological analyses. Protection levels against abortion were 78.6% for S19, 81.8% for M1-luc and 45.5% for I2, compared to the 25% that did not abort from the non-vaccinated group. These results indicate that in bovines BP26 had no influence in protective capacity of S19, correlating with the results obtained in mice. However, contrarily to what was previously observed in mice, lack of expression of Omp19 rendered in less protection capacity of S19 in the natural host. PMID:18565697

  4. Inhibitory effect of red ginseng acidic polysaccharide from Korean red ginseng on phagocytic activity and intracellular replication of Brucella abortus in RAW 264.7 cells

    Science.gov (United States)

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

    2016-01-01

    Korean red ginseng (KRG) has long been used in traditional Korean and Oriental medicine. However, the anti-bacterial mechanism and therapeutic efficiency of KGR for intracellular Brucella infection are still unclear. In this study, the bactericidal activity of Korean red ginseng acidic polysaccharide (RGAP) on Brucella (B.) abortus and its cytotoxic effects on RAW 264.7 cells were evaluated. In addition, B. abortus internalization and intracellular replication in macrophages were investigated after RGAP treatment. RGAP-incubated cells displayed a marked reduction in the adherence, internalization and intracellular growth of B. abortus in macrophages. Furthermore, decreased F-actin fluorescence was observed relative to untreated B. abortus-infected cells. Western blot analysis of intracellular signaling proteins revealed reduced ERK, JNK and p38α phosphorylation levels in B. abortus-infected RGAP-treated cells compared to the control. Moreover, elevated co-localization of B. abortus-containing phagosomes with lysosome-associated membrane protein 1 (LAMP-1) were observed in RGAP-treated cells compared with the control. Overall, the results of this study suggest that RGAP can disrupt phagocytic activity of B. abortus via suppression of mitogen-activated protein kinases (MAPKs) signaling proteins ERK, JNK and p38 levels and inhibit intracellular replication of B. abortus by enhancing phagolysosome fusion, which may provide an alternative control of brucellosis. PMID:26726017

  5. Prediction of T cell epitopes of Brucella abortus and evaluation of their protective role in mice.

    Science.gov (United States)

    Afley, Prachiti; Dohre, Sudhir K; Prasad, G B K S; Kumar, Subodh

    2015-09-01

    Brucellae are Gram-negative intracellular bacteria that cause an important zoonotic disease called brucellosis. The animal vaccines are available but have disadvantage of causing abortions in a proportion of pregnant animals. The animal vaccines are also pathogenic to humans. Recent trend in vaccine design has shifted to epitope-based vaccines that are safe and specific. In this study, efforts were made to identify MHC-I- and MHC-II-restricted T cell epitopes of Brucella abortus and evaluate their vaccine potential in mice. The peptides were designed using online available immunoinformatics tools, and five MHC-I- and one MHC-II-restricted T cell peptides were selected on the basis of their ability to produce interferon gamma (IFN-γ) in in vivo studies. The selected peptides were co-administered with poly DL-lactide-co-glycolide (PLG) microparticles and evaluated for immunogenicity and protection in BALB/c mice. Mice immunized with peptides either entrapped in PLG microparticles (EPLG-Pep) or adsorbed on PLG particles (APLG-Pep) showed significantly higher splenocyte proliferation and IFN-γ generation to all selected peptides than the mice immunized with corresponding irrelevant peptides formulated PLG microparticles or phosphate-buffered saline (PBS). A significant protection compared to PBS control was also observed in EPLG-Pep and APLG-Pep groups. A plasmid DNA vaccine construct (pVaxPep) for peptides encoding DNA sequences was generated and injected to mice by in vivo electroporation. Significant protection was observed (1.66 protection units) when compared with PBS and empty vector control group animals. Overall, the MHC-I and MHC-II peptides identified in this study are immunogenic and protective in mouse model and support the feasibility of peptide-based vaccine for brucellosis. PMID:26150246

  6. Preparation by ultrafiltration and control by high-performance liquid chromatography of the native hapten of Brucella abortus for use in radial immunodiffusion diagnostic test.

    OpenAIRE

    Zygmunt, M S; Dubray, G

    1987-01-01

    An effective method was developed for the preparation of Brucella abortus native hapten by use of an ultrafiltration system. The membranes PM30 and YM5 were used to separate the lipopolysaccharide and the native hapten. The yield of the native hapten was higher than that obtained by a more complex procedure reported previously. This method is economical for the large-scale preparation of B. abortus native hapten. The effects of ultrafiltration were evaluated by a quick and sensitive high-perf...

  7. Brucella abortus Cyclic β-1,2-Glucan Mutants Have Reduced Virulence in Mice and Are Defective in Intracellular Replication in HeLa Cells

    OpenAIRE

    Briones, Gabriel; Iñón de Iannino, Nora; Roset, Mara; VIGLIOCCO, ANA; Paulo, Patricia Silva; Ugalde, Rodolfo A.

    2001-01-01

    Null cyclic β-1,2-glucan synthetase mutants (cgs mutants) were obtained from Brucella abortus virulent strain 2308 and from B. abortus attenuated vaccinal strain S19. Both mutants show greater sensitivity to surfactants like deoxycholic acid, sodium dodecyl sulfate, and Zwittergent than the parental strains, suggesting cell surface alterations. Although not to the same extent, both mutants display reduced virulence in mice and defective intracellular multiplication in HeLa cells. The B. abort...

  8. Molecular Cloning and Characterization of cgt, the Brucella abortus Cyclic β-1,2-Glucan Transporter Gene, and Its Role in Virulence

    OpenAIRE

    Roset, Mara S.; Ciocchini, Andrés E.; Ugalde, Rodolfo A.; Iñón de Iannino, Nora

    2004-01-01

    The animal pathogen Brucella abortus contains a gene cgt, which complemented Sinorhizobium meliloti nodule development (ndvA) and Agrobacterium tumefaciens chromosomal virulence (chvA) mutants. Complemented strains recovered the presence of anionic cyclic β-1,2-glucan, motility, tumor induction in A. tumefaciens, and nodule occupancy in S. meliloti, all traits strictly associated with the presence of cyclic β-1,2-glucan in the periplasm. Nucleotide sequencing revealed that B. abortus cgt cont...

  9. Endocarditis por Brucella abortus: Reporte del primer caso en C.R

    Directory of Open Access Journals (Sweden)

    Manuel Antonio Villalobos-Zúñiga

    2011-09-01

    Full Text Available Paciente masculino de 36 años de edad, proveniente de la zona rural de Costa Rica, con un cuadro clínico de 8 meses de evolución de fiebre, mialgias, artralgias, pérdida de peso y lumbalgia; referido por la detección de un soplo de insuficiencia aórtica. El ecocardiograma reveló endocarditis de la válvula aórtica, y se obtuvieron 4 hemocultivos positivos por Brucella abortus biotipo 3, con serologías negativas por brucelosis. Se inició tratamiento con antibióticos y luego se le realizó un reemplazo valvular aórtico; 4 meses después ingresó con dolor torácico que se atribuyó a una oclusión de la arteria descendente anterior, demostrada angiográficamente, por posible embolismo. En la actualidad cursa clínicamente estable con manejo médico para su cardiopatía, sin recaída infecciosa.

  10. Effectiveness of Brucella abortus Strain 19 single calfhood vaccination in elk (Cervus elaphus)

    Science.gov (United States)

    Roffe, Thomas J.; Jones, Lee C.; Coffin, Kenneth; Sweeney, Steven J.; Williams, Beth; Quist, Charlotte

    2002-01-01

    Brucellosis in Greater Yellowstone Area (GYA) bison and elk has been a source of controversy and focus of the Greater Yellowstone Interagency Brucellosis Committee (GYIBC) for years. Brucellosis has been eradicated from cattle in the 3 states of Wyoming, Montana, and Idaho and all three states currently are classified as “brucellosis free” with regard to livestock. Yet free-ranging elk that attend feedgrounds in the GYA, and bison in Yellowstone and Grand Teton National Parks, still have high seroprevalence to the disease and are viewed as a threat to the state-federal cooperative national brucellosis eradication program. Recently, cattle in eastern Idaho were found infected with brucellosis and transmission was apparently from fed elk. The GYIBC, formed of state and federal agencies involved in wildlife and livestock management in the 3 states, has committed to eventual elimination of the disease from wildlife. Management tools to control or eliminate the disease are limited; however, wildlife vaccination is one of the methods currently employed. Effective wildlife vaccination depends on dose efficacy, deliverability, and safety to non-targeted species. We commenced a single-dose efficacy study of vaccine Brucella abortus strain 19 (S19) in elk in 1999.

  11. Construction of Cu-Zn superoxide dismutase deletion mutants of Brucella abortus: analysis of survival in vitro in epithelial and phagocytic cells and in vivo in mice.

    OpenAIRE

    Tatum, F M; Detilleux, P G; Sacks, J M; Halling, S. M.

    1992-01-01

    Cu-Zn superoxide dismutase (SOD) deletion mutants of Brucella abortus S2308, a virulent strain, and S19, a vaccine strain, were generated by gene replacement. A deletion plasmid, pBA delta sodknr, was constructed by excising the Cu-Zn SOD gene (Cu-Zn sod) from a 2.3-kb B. abortus DNA fragment of plasmid pBA20-1527 and inserting a 1.4-kb DNA fragment encoding kanamycin resistance into the Cu-Zn sod excision site. The deletion plasmid was introduced into B. abortus by electroporation, and South...

  12. The Attenuated Brucella abortus Strain 19 Invades, Persists in, and Activates Human Dendritic Cells, and Induces the Secretion of IL-12p70 but Not IL-23

    OpenAIRE

    Mario Weinhold; Martin Eisenblätter; Edith Jasny; Michael Fehlings; Antje Finke; Hermine Gayum; Ursula Rüschendorf; Pablo Renner Viveros; Verena Moos; Kristina Allers; Thomas Schneider; Schaible, Ulrich E; Schumann, Ralf R.; Martin E Mielke; Ralf Ignatius

    2013-01-01

    Background: Bacterial vectors have been proposed as novel vaccine strategies to induce strong cellular immunity. Attenuated strains of Brucella abortus comprise promising vector candidates since they have the potential to induce strong CD4+ and CD8+ T-cell mediated immune responses in the absence of excessive inflammation as observed with other Gram-negative bacteria. However, some Brucella strains interfere with the maturation of dendritic cells (DCs), which is essential for antigen-speci...

  13. Control of Bovine Brucellosis from Persistently Infected Holdings Using RB51 Vaccination with Test-and-Slaughter: A Comparative Case Report from a High Incidence Area in Portugal.

    Science.gov (United States)

    Caetano, M C; Afonso, F; Ribeiro, R; Fonseca, A P; Abernethy, D A; Boinas, F

    2016-02-01

    Bovine brucellosis due to Brucella abortus infection causes significant reproductive and production losses in cattle and is a major zoonosis. Eradication of this disease has proved difficult to achieve in Portugal where it still occurs in some regions despite an ongoing national eradication programme. In 2004, the Alentejo region, a major cattle producing area, reported one of the highest levels of bovine brucellosis in the country, especially in one divisional area. In that area, bovine brucellosis was particularly problematic in a holding of ten herds, the largest extensive cattle unit in the country, which remained infected despite an extensive test-and-slaughter programme and depopulation of five herds. A 5-year programme of RB51 vaccination with biannual test-and-slaughter was thus implemented in 2004. The apparent animal seroprevalence decreased from 19% (646/3,400) to 3% (88/2930) on the third herd-level test and remained below 0.8% (27/3324) after the fourth test. After the tenth test, the holding had a prevalence of 0.1% (2/2332) and only one herd remained positive with a within-herd prevalence of 1.1% (2/177). The results were compared to all other herds (n = 10) in the divisional area that were also persistently infected but were subject only to test-and-slaughter before being depopulated. In these herds, the strategy of test-and-slaughter did not reduce the prevalence, which remained significantly higher than the vaccinated group (median = 0.48% and 8.5% in vaccinated versus non-vaccinated herds; Wilcoxon rank sum test; P Portugal provided a valuable case study to the official veterinary services by illustrating the value of RB51 vaccination with parallel testing and improved biosecurity as a comprehensive and sustainable strategy for bovine brucellosis control in persistently infected herds.

  14. Effect of ochratoxin and aflatoxin on serum proteins, complement activity, and antibody production to Brucella abortus in guinea pigs.

    Science.gov (United States)

    Richard, J L; Thurston, J R; Deyoe, B L; Booth, G D

    1975-01-01

    The effect of ochratoxin alone and in combination with aflatoxin and Brucella abortus antigen on complement activity, serum proteins, and antibody response in guinea pigs was investigated. Ochratoxin did not affect complement activity or antibody response and there was no interaction between ochratoxin and aflatoxin on any of the responses tested. Ochratoxin significantly lowered the level of beta-globulin in serum of guinea pigs. There was no significant interaction between aflatoxin and antigen on lowering of the serum albumin levels of guinea pigs. PMID:45955

  15. The Dutch Brucella abortus monitoring programme for cattle: the impact of false-positive serological reactions and comparison of serological tests

    NARCIS (Netherlands)

    Emmerzaal, A.; Wit, de J.J.; Dijkstra, T.; Bakker, D.; Ziiderveld, van F.G.

    2002-01-01

    The Dutch national Brucella abortus eradication programme for cattle started in 1959. Sporadic cases occurred yearly until 1995; the last infected herd was culled in 1996. In August 1999 the Netherlands was declared officially free of bovine brucellosis by the European Union. Before 1999, the progra

  16. Immunization with Recombinant Brucella Species Outer Membrane Protein Omp16 or Omp19 in Adjuvant Induces Specific CD4+ and CD8+ T Cells as Well as Systemic and Oral Protection against Brucella abortus Infection

    OpenAIRE

    Pasquevich, Karina A.; Estein, Silvia M.; Samartino, Clara García; Zwerdling, Astrid; Coria, Lorena M.; Barrionuevo, Paula; Fossati, Carlos A.; Giambartolomei, Guillermo H.; Cassataro, Juliana

    2009-01-01

    Available vaccines against Brucella spp. are live attenuated Brucella strains. In order to engineer a better vaccine to be used in animals and humans, our laboratory aims to develop an innocuous subunit vaccine. Particularly, we are interested in the outer membrane proteins (OMPs) of B. abortus: Omp16 and Omp19. In this study, we assessed the use of these proteins as vaccines against Brucella in BALB/c mice. Immunization with lipidated Omp16 (L-Omp16) or L-Omp19 in incomplete Freund's adjuvan...

  17. Isolation of two biologically active cell surface proteins from Brucella abortus by chromatofocusing

    Energy Technology Data Exchange (ETDEWEB)

    Tabatabai, L.B.; Deyoe, B.L.

    1983-01-01

    Brucella abortus contains a group of immunogenic cell surface proteins which have potential value as a vaccine or as a diagnostic reagent for the prevention and diagnosis of bovine brucellosis. Under nondenaturing conditions, these proteins range in molecular weight from 10,000-124,000, as determined by high performance liquid chromatography (HPLC) on TSK 3000sw. By analytical isoelectrofocusing, 6 major protein bands could be distinguished with pI's ranging from 4.0 to 6.0 and 3 additional major proteins with pI's of 7.5, 9.5, and 10. By chromatofocusing on Polybuffer Exchanger 94 with a pH gradient from 6-4, two of the six proteins from pI 4-6 were separated, a pI 4.9 and a pI 4.7 protein; a third fraction contained the high pI proteins. The former two proteins were homogeneous by analytical isoelectrofocusing, and a molecular weight of 54,000 daltons was found for both protein species by HPLC on TSK 3000sw. The pI 4-6 and not the pI 9.5 and 10 proteins, could be radiolabeled when intact cells were radioiodinated with diazotized (/sup 125/I)-iodosulfanilic acid. Biological activity of the proteins as assessed in lemmings indicated that immunization with the pI 4.7 and 4.9 proteins afforded better protection against experimental brucellosis than immunization with the high pI proteins. These results support our view that a single surface protein may be sufficient for the prevention of experimental brucellosis.

  18. Reduced Susceptibility to Rifampicin and Resistance to Multiple Antimicrobial Agents among Brucella abortus Isolates from Cattle in Brazil

    Science.gov (United States)

    Barbosa Pauletti, Rebeca; Reinato Stynen, Ana Paula; Pinto da Silva Mol, Juliana; Seles Dorneles, Elaine Maria; Alves, Telma Maria; de Sousa Moura Souto, Monalisa; Minharro, Silvia; Heinemann, Marcos Bryan; Lage, Andrey Pereira

    2015-01-01

    This study aimed to determine the susceptibility profile of Brazilian Brucella abortus isolates from cattle to eight antimicrobial agents that are recommended for the treatment of human brucellosis and to correlate the susceptibility patterns with origin, biotype and MLVA16-genotype of the strains. Screening of 147 B. abortus strains showed 100% sensitivity to doxycycline and ofloxacin, one (0.68%) strain resistant to ciprofloxacin, two strains (1.36%) resistant to streptomycin, two strains (1.36%) resistant to trimethoprim-sulfamethoxazole and five strains (3.40%) resistant to gentamicin. For rifampicin, three strains (2.04%) were resistant and 54 strains (36.73%) showed reduced sensitivity. Two strains were considered multidrug resistant. In conclusion, the majority of B. abortus strains isolated from cattle in Brazil were sensitive to the antimicrobials commonly used for the treatment of human brucellosis; however, a considerable proportion of strains showed reduced susceptibility to rifampicin and two strains were considered multidrug resistant. Moreover, there was no correlation among the drug susceptibility pattern, origin, biotype and MLVA16-genotypes of these strains. PMID:26181775

  19. Detection of antibodies against Brucella abortus, Leptospira spp., and Apicomplexa protozoa in water buffaloes in the Northeast of Argentina.

    Science.gov (United States)

    Konrad, José L; Campero, Lucía M; Caspe, Gastón S; Brihuega, Bibiana; Draghi, Graciela; Moore, Dadin P; Crudeli, Gustavo A; Venturini, María C; Campero, Carlos M

    2013-11-01

    Water buffalo industry has become a profitable activity worldwide, including the Northeast of Argentina (NEA). However, research on diseases affecting this species is scarce. The aim of the present study was to detect antibodies against Brucella abortus, Leptospira spp., Neospora caninum, Toxoplasma gondii, and Sarcocystis spp. in 500 water buffalo cows from five ranches (100 animals each) in the NEA. Serum samples were tested for B. abortus by fluorescence polarization assay, Leptospira spp. by microagglutination test, and N. caninum, T. gondii, and Sarcocystis spp. by indirect fluorescent antibody tests. Overall, the proportion of seropositive animals was 6.4, 22.2, 42.2, 25.4, and 50.8 % for brucellosis, leptospirosis, neosporosis, toxoplasmosis, and sarcocystosis, respectively. The proportion of seropositive animals for all diseases was statistically different among herds (p < 0.05). Statistical differences were also detected among age groups for brucellosis and neosporosis (p < 0.05). The detection of specific antibodies to B. abortus, Leptospira spp., and several Apicomplexa protozoans in water buffaloes in the NEA is reported in this study. PMID:23765549

  20. Genome sequence of Brucella abortus vaccine strain S19 compared to virulent strains yields candidate virulence genes.

    Science.gov (United States)

    Crasta, Oswald R; Folkerts, Otto; Fei, Zhangjun; Mane, Shrinivasrao P; Evans, Clive; Martino-Catt, Susan; Bricker, Betsy; Yu, GongXin; Du, Lei; Sobral, Bruno W

    2008-01-01

    The Brucella abortus strain S19, a spontaneously attenuated strain, has been used as a vaccine strain in vaccination of cattle against brucellosis for six decades. Despite many studies, the physiological and molecular mechanisms causing the attenuation are not known. We have applied pyrosequencing technology together with conventional sequencing to rapidly and comprehensively determine the complete genome sequence of the attenuated Brucella abortus vaccine strain S19. The main goal of this study is to identify candidate virulence genes by systematic comparative analysis of the attenuated strain with the published genome sequences of two virulent and closely related strains of B. abortus, 9-941 and 2308. The two S19 chromosomes are 2,122,487 and 1,161,449 bp in length. A total of 3062 genes were identified and annotated. Pairwise and reciprocal genome comparisons resulted in a total of 263 genes that were non-identical between the S19 genome and any of the two virulent strains. Amongst these, 45 genes were consistently different between the attenuated strain and the two virulent strains but were identical amongst the virulent strains, which included only two of the 236 genes that have been implicated as virulence factors in literature. The functional analyses of the differences have revealed a total of 24 genes that may be associated with the loss of virulence in S19. Of particular relevance are four genes with more than 60 bp consistent difference in S19 compared to both the virulent strains, which, in the virulent strains, encode an outer membrane protein and three proteins involved in erythritol uptake or metabolism. PMID:18478107

  1. Genome sequence of Brucella abortus vaccine strain S19 compared to virulent strains yields candidate virulence genes.

    Directory of Open Access Journals (Sweden)

    Oswald R Crasta

    Full Text Available The Brucella abortus strain S19, a spontaneously attenuated strain, has been used as a vaccine strain in vaccination of cattle against brucellosis for six decades. Despite many studies, the physiological and molecular mechanisms causing the attenuation are not known. We have applied pyrosequencing technology together with conventional sequencing to rapidly and comprehensively determine the complete genome sequence of the attenuated Brucella abortus vaccine strain S19. The main goal of this study is to identify candidate virulence genes by systematic comparative analysis of the attenuated strain with the published genome sequences of two virulent and closely related strains of B. abortus, 9-941 and 2308. The two S19 chromosomes are 2,122,487 and 1,161,449 bp in length. A total of 3062 genes were identified and annotated. Pairwise and reciprocal genome comparisons resulted in a total of 263 genes that were non-identical between the S19 genome and any of the two virulent strains. Amongst these, 45 genes were consistently different between the attenuated strain and the two virulent strains but were identical amongst the virulent strains, which included only two of the 236 genes that have been implicated as virulence factors in literature. The functional analyses of the differences have revealed a total of 24 genes that may be associated with the loss of virulence in S19. Of particular relevance are four genes with more than 60 bp consistent difference in S19 compared to both the virulent strains, which, in the virulent strains, encode an outer membrane protein and three proteins involved in erythritol uptake or metabolism.

  2. Serological relationship between cattle exposed to Brucella abortus, Yersinia enterocolitica O:9 and Escherichia coli O157:H7.

    Science.gov (United States)

    Nielsen, K; Smith, P; Widdison, J; Gall, D; Kelly, L; Kelly, W; Nicoletti, P

    2004-05-20

    Sera from cattle naturally infected with Brucella abortus (n = 160), vaccinated with B. abortus S19 (n = 88) or immunized with Yersinia enterocolitica O:9 (n = 25) or Escherichia coli O157:H7 (n = 80) were collected. The sera were compared for antibody content to the same bacteria by indirect enzyme immunoassay (IELISA), fluorescence polarization assay (FPA) and competitive enzyme immunoassay (CELISA). Cattle sera (n = 523) collected randomly from across Canada were tested in the same tests. Sera from the B. abortus infected group reacted positively in the brucellosis IELISA (IELISA(Br)), CELISA and FPA (FPA(Br)) and the Y. enterocolitica IELISA (IELISA(Ye)) while the Y. enterocolitica FPA (FPA(Ye)) detected antibody in 93.8% and the E. coli IELISA (IELISA(Ec)) 86.9% and the E. coli FPA (FPA(Ec)) 48.1%. About 70% of the sera from B. abortus S19 vaccinated animals reacted in the three IELISAs, 45% in the CELISA, and 37.7% in the FPA(Ec), 21.6% in the FPA(Br) and 5.7% in the FPA(Ye). Sera from E. coli O:157 exposed cattle reacted mainly in the IELISA(Ec) and FPA(Ec) although surprisingly 87.5% reacted in the IELISA(Ye) and only 3.8% in the IELISA(Br). No reactions were observed with these sera in the FPA(Br) and FPA(Ye) but one serum gave a low positive reaction in the CELISA. All sera from Y. enterocolitica O:9 exposed cattle reacted in the IELISA(Br) and IELISA(Ye) and 80% in the IELISA(Ec). In the CELISA, 44% gave a positive reaction and 64% were positive in the FPA(Br), 28% in the FPA(Ye) and 12% in the FPA(Ec). Of the 523 Canadian sera, about 50% reacted in the E. coli tests with only minor reactions in the Y. enterocolitica O:9 and B. abortus assays. From the data, the cross reaction between E. coli O157:H7, Y. enterocilitica O:9 and B. abortus is dependent on the test used. Thus, extensive cross reaction was observed with the IELISA with much less reactivity in the FPA and the CELISA. PMID:15135510

  3. Brucella alters endocytic pathway in J774 macrophages.

    Science.gov (United States)

    Arenas, Graciela N; Grilli, Diego J; Samartino, Luis E; Magadán, Javier; Mayorga, Luis S

    2010-01-01

    Brucella is a facultative intracellular bacterium which causes chronic infections in mammals by surviving and replicating within host cells. The putative role of the endoplasmic reticulum (ER) in the formation of the phagosome in non-professional phagocytes is supported by several research groups, but still leaves open the question of the fate of Brucella inside professional phagocytes and its resistance mechanisms therein. Macrophages are particularly important for the survival and spreading of Brucella during infection. The intracellular transport of Brucella in these cells has not been thoroughly characterized. To study the maturation process of Brucella-containing phagosomes in phagocytes, we comparatively monitored the intracellular transport of a virulent strain (2308) with two vaccine strains (S19 and RB51) in J 774 macrophages. Then, we compared the behavior of all three strains studied through transmission electron microscopy. The results indicate that the virulent strain not only occupies two different kinds of compartments but also alters the endocytic pathway of the cell it parasitizes, unlike what has been reported for non-professional phagocytes, like HeLa cell. Besides, differences are observed in the behavior of both Brucella abortus vaccine strains. PMID:21178473

  4. Serological response to an indirect and a competitive elisa in heifers vaccinated with Brucella abortus strain 19

    International Nuclear Information System (INIS)

    The different serologic techniques for bovine brucellosis diagnosis have different abilities to detect antibodies after vaccination with Brucella abortus strain 19. The humoral response in heifers vaccinated with Brucella abortus strain 19 was evaluated by using several serologic techniques. In the experimental field of INTA, Pilcaniyeu, Rio Negro province, sixteen 5 months old heifers were vaccinated subcutaneously with a standard dose (2ml, containing 20x109 to 10x109 living organisms) of Brucella abortus strain 19. Sera from all the heifers were obtained on 18 occasions (one 87 days before vaccination, one immediately before vaccination and on 16 occasions after vaccination, during 488 days) and analyzed by buffered plate antigen test, rose bengal test, standard tube agglutination test, 2-mercaptoetanol test, complement fixation test, indirect ELISA, and competitive ELISA. Prior vaccination, 100% of the heifers gave negative results in all the techniques used, while 100% of them gave positive reaction in the first sampling after vaccination to all the techniques, with the exception of standard tube agglutination test that showed agglutinating titters of 1/100 or higher (positive threshold) in only 71.4% of the heifers. The indirect ELISA technique showed a reducing percentage of positive animals up until 316 days after vaccination, after which positive results were obtained. The competitive ELISA gave positive results in a variable number of heifers up to 253 days after vaccination when 100% of the sera were negative to this technique. Buffered plate antigen test was the technique that gave positive results for a longest period, being 100% of the animals negative to this technique at 450 days after vaccination. The other serological techniques assayed gave positive results during variable periods of time, intermediate between standard tube agglutination test and buffered plate antigen test. Although the present results were obtained from a limited number of

  5. Vaccination with Brucella abortus Recombinant In Vivo-Induced Antigens Reduces Bacterial Load and Promotes Clearance in a Mouse Model for Infection

    OpenAIRE

    Jake E Lowry; Isaak, Dale D.; Leonhardt, Jack A.; Giulia Vernati; Jessie C Pate; Andrews, Gerard P.

    2011-01-01

    Current vaccines used for the prevention of brucellosis are ineffective in inducing protective immunity in animals that are chronically infected with Brucella abortus, such as elk. Using a gene discovery approach, in vivo-induced antigen technology (IVIAT) on B. abortus, we previously identified ten loci that encode products up-regulated during infection in elk and consequently may play a role in virulence. In our present study, five of the loci (D15, 0187, VirJ, Mdh, AfuA) were selected for ...

  6. 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.

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

    Science.gov (United States)

    Osman, Amira E. F.; Hassan, Abdullahi N.; Ali, Ali E.; Abdoel, Theresia H.; Smits, Henk L.

    2015-01-01

    Background 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. Methods 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. Results 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. Conclusions 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. PMID:25938483

  8. Diagnóstico sorológico da brucelose bovina em animais adultos vacinados com dose reduzida da cepa 19 de Brucella abortus Serological diagnosis of bovine brucellosis in adult herd vaccinated with Brucella abortus strain 19 reduced dose

    Directory of Open Access Journals (Sweden)

    Gustavo Coelho Jardim

    2006-09-01

    Full Text Available Com o presente trabalho avaliou-se o uso de dose reduzida da vacina produzida com a amostra 19 de Brucella abortus, em rebanho adulto negativo para a enfermidade, por meio de técnicas de diagnóstico sorológico preconizadas pelo Programa Nacional de Controle e Erradicação da Brucelose e Tuberculose Animal e por um ensaio indireto de imunoadsorção enzimática (ELISA ID. A prova de fixação de complemento detectou 46,77% de positivos, o antígeno acidificado tamponado 67,74%, o 2-mercaptoetanol com soroaglutinação lenta 87,09% e o ELISA ID 100%. A dose reduzida interferiu no diagnóstico sorológico. Nenhuma das técnicas apresentou especificidade adequada para uso em rebanho nestas condições, até 3 meses após a vacinação.The study evaluated the use of a reduced dose of the Brucella abortus strain 19 vaccine, in an adult herd negative for the disease, by serological diagnostic techniques, advocated by the Brazilian Program for Animal Brucellosis and Tuberculosis Control and Eradication, and by an indirect ELISA. The complement fixation test detecteed 46.77% positives, the rose bengal test 67.74%, the mercaptoethanol with standard agglutination test 87.09% and the ELISA ID 100%. The reduced dose influenced the serological diagnosis. None of the techniques reached a suitable specificity for use in the herd under those conditions, up to 3 months after vaccination.

  9. Evaluation of the effects of erythritol on gene expression in Brucella abortus

    OpenAIRE

    María Cruz Rodríguez; Cristina Viadas; Asunción Seoane; Félix Javier Sangari; Ignacio López-Goñi; Juan María García-Lobo

    2012-01-01

    Bacteria of the genus Brucella have the unusual capability to catabolize erythritol and this property has been associated with their virulence mainly because of the presence of erythritol in bovine foetal tissues and because the attenuated S19 vaccine strain is the only Brucella strain unable to oxydize erythritol. In this work we have analyzed the transcriptional changes produced in Brucella by erythritol by means of two high throughput approaches: RNA hybridization against a microarray cont...

  10. Mutant Brucella abortus Membrane Fusogenic Protein Induces Protection against Challenge Infection in Mice

    OpenAIRE

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

    2015-01-01

    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 t...

  11. Characterization of Brucella abortus and Brucella melitensis native haptens as outer membrane O-type polysaccharides independent from the smooth lipopolysaccharide.

    Science.gov (United States)

    Aragón, V; Díaz, R; Moreno, E; Moriyón, I

    1996-02-01

    Brucella native haptens (NHs) extracted with hot water from smooth (S)-type B. abortus and B. melitensis were purified to high levels of serological activity and compared with the polysaccharide obtained by acid hydrolysis (PS) of the S lipopolysaccharide (S-LPS). By 13C nuclear magnetic resonance analysis, NHs showed the spectrum of a homopolymer of alpha-1,2- or alpha-1,2- plus alpha-1,3-linked 4-formamido-4,6-dideoxy-D-mannose (N-formylperosamine) previously reported for the LPS O chain. However, while PS contained up to 0.6% 3-deoxy-D-manno-2-octulosonate, this LPS-core marker was absent from NH. High performance liquid chromatography and thin-layer chromatography showed heterogeneity in NH purified from whole cells but not in PS. By immunoprecipitation, polysaccharides indistinguishable from NH were demonstrated in extracts obtained with phenol-water, saline at 60 degrees C, and ether-water treatments, and none of these treatments caused S-LPS hydrolysis detectable with antibodies to the O chain and lipid A. Two lines of evidence showed that NH was in the cell surface. First, NH became biotinylated when B. abortus live cells were labelled with biotin-hydrazide, and the examination of cell fractions and electron microscopy sections with streptavidin-peroxidase and streptavidin-coloidal gold, respectively, showed that labelling was extrinsic. Moreover, whereas only traces of NH were found in cytosols, the amount of NH was enriched in cell envelopes and in the outer membrane blebs spontaneously released by brucellae during growth. Interactions between NH and S-LPS were observed in crude cell extracts, and such interactions could be reconstituted by using purified NH and LPS. The results demonstrate that NH is not a hydrolytic product of S-LPS and suggest a model in which LPS-independent O-type polysaccharides (NH) are intertwined with the O chain in the outer membrane of S-type brucellae. PMID:8576040

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

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    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.

  13. Summary of field trials using the direct and competitive enzyme immunoassays for detection of antibody to brucella abortus

    International Nuclear Information System (INIS)

    Two indirect and two competitive enzyme immunoassays for detection of antibody to Brucella abortus, validated elsewhere, were field tested in five different Latin American laboratories. Testing was performed according to standardised protocols using sera obtained in each area. Sera from B. abortus infected herds, from vaccinated (but serologically negative in a screening test) and non-vaccinated cattle were tested in each assay and compared to the results obtained with conventional diagnostic tests used for diagnosis of brucellosis in each country. Relative sensitivity and specificity values were calculated for each country as well as a weighted summary combining the data from all the participating laboratories. The result demonstrate that all ELISAs performed as well as, or better than, the conventional aerological tests. Given the inherent errors in the use of the latter in the diagnosis of brucellosis, it is recommended that the ELISAs described here be considered as replacements for the conventional tests. The CELISA using the lipopolysaccharide antigen with the competing monoclonal antibody M84, should be considered as the most useful because of cross-species and vaccination considerations. (author)

  14. 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. PMID:26302373

  15. 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.

  16. Brucella abortus Inhibits Major Histocompatibility Complex Class II Expression and Antigen Processing through Interleukin-6 Secretion via Toll-Like Receptor 2▿

    Science.gov (United States)

    Barrionuevo, Paula; Cassataro, Juliana; Delpino, M. Victoria; Zwerdling, Astrid; Pasquevich, Karina A.; Samartino, Clara García; Wallach, Jorge C.; Fossati, Carlos A.; Giambartolomei, Guillermo H.

    2008-01-01

    The strategies that allow Brucella abortus to survive inside macrophages for prolonged periods and to avoid the immunological surveillance of major histocompatibility complex class II (MHC-II)-restricted gamma interferon (IFN-γ)-producing CD4+ T lymphocytes are poorly understood. We report here that infection of THP-1 cells with B. abortus inhibited expression of MHC-II molecules and antigen (Ag) processing. Heat-killed B. abortus (HKBA) also induced both these phenomena, indicating the independence of bacterial viability and involvement of a structural component of the bacterium. Accordingly, outer membrane protein 19 (Omp19), a prototypical B. abortus lipoprotein, inhibited both MHC-II expression and Ag processing to the same extent as HKBA. Moreover, a synthetic lipohexapeptide that mimics the structure of the protein lipid moiety also inhibited MHC-II expression, indicating that any Brucella lipoprotein could down-modulate MHC-II expression and Ag processing. Inhibition of MHC-II expression and Ag processing by either HKBA or lipidated Omp19 (L-Omp19) depended on Toll-like receptor 2 and was mediated by interleukin-6. HKBA or L-Omp19 also inhibited MHC-II expression and Ag processing of human monocytes. In addition, exposure to the synthetic lipohexapeptide inhibited Ag-specific T-cell proliferation and IFN-γ production of peripheral blood mononuclear cells from Brucella-infected patients. Together, these results indicate that there is a mechanism by which B. abortus may prevent recognition by T cells to evade host immunity and establish a chronic infection. PMID:17984211

  17. Early Transcriptional Responses of Bovine Chorioallantoic Membrane Explants to Wild Type, ΔvirB2 or ΔbtpB Brucella abortus Infection

    Science.gov (United States)

    Mol, Juliana P. S.; Costa, Erica A.; Carvalho, Alex F.; Sun, Yao-Hui; Tsolis, Reneé M.; Paixão, Tatiane A.; Santos, Renato L.

    2014-01-01

    The pathogenesis of the Brucella-induced inflammatory response in the bovine placenta is not completely understood. In this study we evaluated the role of the B. abortus Type IV secretion system and the anti-inflammatory factor BtpB in early interactions with bovine placental tissues. Transcription profiles of chorioallantoic membrane (CAM) explants inoculated with wild type (strain 2308), ΔvirB2 or ΔbtpB Brucella abortus were compared by microarray analysis at 4 hours post infection. Transcripts with significant variation (>2 fold change; P<0.05) were functionally classified, and transcripts related to defense and inflammation were assessed by quantitative real time RT-PCR. Infection with wild type B. abortus resulted in slightly more genes with decreased than increased transcription levels. Conversely, infection of trophoblastic cells with the ΔvirB2 or the ΔbtpB mutant strains, that lack a functional T4SS or that has impaired inhibition of TLR signaling, respectively, induced more upregulated than downregulated genes. Wild type Brucella abortus impaired transcription of host genes related to immune response when compared to ΔvirB and ΔbtpB mutants. Our findings suggest that proinflammatory genes are negatively modulated in bovine trophoblastic cells at early stages of infection. The virB operon and btpB are directly or indirectly related to modulation of these host genes. These results shed light on the early interactions between B. abortus and placental tissue that ultimately culminate in inflammatory pathology and abortion. PMID:25259715

  18. Early transcriptional responses of bovine chorioallantoic membrane explants to wild type, ΔvirB2 or ΔbtpB Brucella abortus infection.

    Directory of Open Access Journals (Sweden)

    Juliana P S Mol

    Full Text Available The pathogenesis of the Brucella-induced inflammatory response in the bovine placenta is not completely understood. In this study we evaluated the role of the B. abortus Type IV secretion system and the anti-inflammatory factor BtpB in early interactions with bovine placental tissues. Transcription profiles of chorioallantoic membrane (CAM explants inoculated with wild type (strain 2308, ΔvirB2 or ΔbtpB Brucella abortus were compared by microarray analysis at 4 hours post infection. Transcripts with significant variation (>2 fold change; P<0.05 were functionally classified, and transcripts related to defense and inflammation were assessed by quantitative real time RT-PCR. Infection with wild type B. abortus resulted in slightly more genes with decreased than increased transcription levels. Conversely, infection of trophoblastic cells with the ΔvirB2 or the ΔbtpB mutant strains, that lack a functional T4SS or that has impaired inhibition of TLR signaling, respectively, induced more upregulated than downregulated genes. Wild type Brucella abortus impaired transcription of host genes related to immune response when compared to ΔvirB and ΔbtpB mutants. Our findings suggest that proinflammatory genes are negatively modulated in bovine trophoblastic cells at early stages of infection. The virB operon and btpB are directly or indirectly related to modulation of these host genes. These results shed light on the early interactions between B. abortus and placental tissue that ultimately culminate in inflammatory pathology and abortion.

  19. Characterization of the heat shock response in Brucella abortus and isolation of the genes encoding the GroE heat shock proteins.

    OpenAIRE

    Lin, J.; Adams, L G; Ficht, T A

    1992-01-01

    In an effort to define the heat shock response in the bovine intracellular pathogen Brucella abortus, a rough variant lacking extensive lipopolysaccharide was pulse-labeled with [35S]methionine following exposure to elevated temperatures. The major heat shock proteins observed following sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography migrate at 70, 62, 18, and 10 kDa. The maximum response was observed between 42 and 46 degrees C and within 2 to 3 h of the shif in...

  20. Reaction of Native and Denatured Brucella abortus (S19 Proteins with Antibody Using Affinity Chromatography and Immunoblotting

    Directory of Open Access Journals (Sweden)

    R. Karimi

    2005-01-01

    Full Text Available Western blotting or immunoblotting commonly use for study of reaction between antigens and antibodies. Denaturation of many proteins in immunoblotting can affect greatly the reactivity of antibodies and outcome of the procedure.In this study proteins of Brucella abortus (S19 was extracted by a mild method and reaction of the extracted proteins with serum of infected human and goat and immunized rabbit compared by affinity chromatography and immunoblotting. Gamma globulin (mostly IgG fraction of the sera was precipitated by half saturation of ammonium sulfate and linked to activated sepharose 4B. The extracted proteins were loaded on the affinity column. Attached proteins was eluted by low pH and analyzed by SDS-PAGE. Reaction of the total extract and eluted fractions with IgG fraction of sera was evaluated by Western blotting.Upon the results of affinity chromatography and immunoblotting, Brucella proteins can be classified in four groups: 1- The proteins that adsorbed to the affinity column and react with IgG in westernblotting. 2- Proteins that react with IgG in native state but no in denatured state. 3- Proteins that do not react with IgG in native state but react in denatured state. 4- Proteins that do not react with IgG in native and denatured state.

  1. DETECTION OF Leptospira spp. AND Brucella abortus ANTIBODIES IN FREE-LIVING JAGUARS (Panthera onca IN TWO PROTECTED AREAS OF NORTHERN PANTANAL, BRAZIL

    Directory of Open Access Journals (Sweden)

    Selma Samiko Miyazaki ONUMA

    2015-04-01

    Full Text Available This study aimed to assess the exposure of free-living jaguars (Panthera onca to Leptospira spp. and Brucella abortus in two conservation units in the Pantanal of Mato Grosso, Brazil. The presence of antibodies in blood samples of eleven jaguars was investigated using autochthonous antigens isolated in Brazil added to reference antigen collection applied to diagnosis of leptospirosis by Microscopic Agglutination Test (MAT. The Rose Bengal test was applied for B. abortus antibodies. Two (18.2% jaguars were seroreactive for the Leptospira spp. antigen and the serovar considered as most infective in both animals was a Brazilian isolate of serovar Canicola (L01. All jaguars were seronegative for B. abortus. These data indicate that the inclusion of autochthonous antigens in serological studies can significantly increase the number of reactive animals, as well as modify the epidemiological profile of Leptospira spp. infection.

  2. Protective immune-response of aluminium hydroxide gel adjuvanted phage lysate of Brucella abortus S19 in mice against direct virulent challenge with B. abortus 544.

    Science.gov (United States)

    Jain, Lata; Rawat, Mayank; Prajapati, Awadhesh; Tiwari, Ashok Kumar; Kumar, Bablu; Chaturvedi, V K; Saxena, H M; Ramakrishnan, Sarvanan; Kumar, Jatin; Kerketta, Priscilla

    2015-09-01

    The prophylactic efficacies of plain and alum adsorbed lysate were evaluated by direct virulent challenge in mice model. A recently isolated brucellaphage 'ϕLd' was used for generation of lysates. Twenty four h incubated Brucella abortus S19 broth cultures standardized to contain approximately 10(8) CFU/ml were found suitable for generation of lysates. Three lysate batches produced through separate cycles did not show any significant variation with respect to protein and polysaccharide contents, endotoxin level and phage counts, indicating that compositionally stable lysate preparations can be generated through an optimized production process. Three polypeptides of ∼16, 19 and 23 kDa could be identified as immuno-dominant antigens of the lysate which induced both humoral and cell-mediated immune responses in a dose dependent manner. Results of efficacy evaluation trial confirmed dose-dependent protective potencies of lysate preparation. The lysate with an antigenic dose of 0.52 μg protein and 60 μg CHO adsorbed on aluminium gel (0.1 percent aluminium concentration) exhibited the highest protective potency which was greater than that induced by standard S19 vaccine. Phage lysate methodology provides a very viable option through which an improved immunizing preparation with all desirable traits can be developed against brucellosis, and integrated with immunization programmes in a more efficient manner. PMID:26156404

  3. Identification of Brucella abortus virulence proteins that modulate the host immune response

    OpenAIRE

    Wang, Yufei; Chen, Zeliang; Qiu, Yefeng; Ke, Yuehua; Xu, Jie; Yuan, Xitong; Li, Xianbo; Fu, Simei; Cui, Mingquan; Xie, Yongfei; Du, Xinying; Wang, Zhoujia; Huang, Liuyu

    2012-01-01

    Brucellosis is an important zoonotic disease of almost worldwide distribution. One significant immune phenomenon of this disease is the ability of the pathogen to hide and survive in the host, establishing long lasting chronic infections. Brucella was found to have the ability to actively modulate the host immune response in order to establish chronic infections, but the mechanism by which the pathogen achieves this remains largely unknown. In our screening for protective antigens of Brucella...

  4. Evaluation of the effects of erythritol on gene expression in Brucella abortus.

    Directory of Open Access Journals (Sweden)

    María Cruz Rodríguez

    Full Text Available Bacteria of the genus Brucella have the unusual capability to catabolize erythritol and this property has been associated with their virulence mainly because of the presence of erythritol in bovine foetal tissues and because the attenuated S19 vaccine strain is the only Brucella strain unable to oxydize erythritol. In this work we have analyzed the transcriptional changes produced in Brucella by erythritol by means of two high throughput approaches: RNA hybridization against a microarray containing most of Brucella ORF's constructed from the Brucella ORFeome and next generation sequencing of Brucella mRNA in an Illumina GAIIx platform. The results obtained showed the overexpression of a group of genes, many of them in a single cluster around the ery operon, able to co-ordinately mediate the transport and degradation of erythritol into three carbon atoms intermediates that will be then converted into fructose-6P (F6P by gluconeogenesis. Other induced genes participating in the nonoxidative branch of the pentose phosphate shunt and the TCA may collaborate with the ery genes to conform an efficient degradation of sugars by this route. On the other hand, several routes of amino acid and nucleotide biosynthesis are up-regulated whilst amino acid transport and catabolism genes are down-regulated. These results corroborate previous descriptions indicating that in the presence of erythritol, this sugar was used preferentially over other compounds and provides a neat explanation of the the reported stimulation of growth induced by erythritol.

  5. Evaluation of the effects of erythritol on gene expression in Brucella abortus.

    Science.gov (United States)

    Rodríguez, María Cruz; Viadas, Cristina; Seoane, Asunción; Sangari, Félix Javier; López-Goñi, Ignacio; García-Lobo, Juan María

    2012-01-01

    Bacteria of the genus Brucella have the unusual capability to catabolize erythritol and this property has been associated with their virulence mainly because of the presence of erythritol in bovine foetal tissues and because the attenuated S19 vaccine strain is the only Brucella strain unable to oxydize erythritol. In this work we have analyzed the transcriptional changes produced in Brucella by erythritol by means of two high throughput approaches: RNA hybridization against a microarray containing most of Brucella ORF's constructed from the Brucella ORFeome and next generation sequencing of Brucella mRNA in an Illumina GAIIx platform. The results obtained showed the overexpression of a group of genes, many of them in a single cluster around the ery operon, able to co-ordinately mediate the transport and degradation of erythritol into three carbon atoms intermediates that will be then converted into fructose-6P (F6P) by gluconeogenesis. Other induced genes participating in the nonoxidative branch of the pentose phosphate shunt and the TCA may collaborate with the ery genes to conform an efficient degradation of sugars by this route. On the other hand, several routes of amino acid and nucleotide biosynthesis are up-regulated whilst amino acid transport and catabolism genes are down-regulated. These results corroborate previous descriptions indicating that in the presence of erythritol, this sugar was used preferentially over other compounds and provides a neat explanation of the the reported stimulation of growth induced by erythritol. PMID:23272076

  6. 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.

  7. Crystallographic and kinetic study of riboflavin synthase from Brucella abortus, a chemotherapeutic target with an enhanced intrinsic flexibility

    Energy Technology Data Exchange (ETDEWEB)

    Serer, María I.; Bonomi, Hernán R. [IIBBA–CONICET, Avenida Patricias Argentinas 435, C1405BWE Buenos Aires (Argentina); Guimarães, Beatriz G. [Synchrotron SOLEIL, L’Orme des Merisiers, Saint-Aubin BP 48, 91192 Gif-sur-Yvette CEDEX (France); Rossi, Rolando C. [Universidad de Buenos Aires, Junín 956, C1113AAD Buenos Aires (Argentina); Goldbaum, Fernando A.; Klinke, Sebastián, E-mail: sklinke@leloir.org.ar [IIBBA–CONICET, Avenida Patricias Argentinas 435, C1405BWE Buenos Aires (Argentina)

    2014-05-01

    This work reports crystal structures of trimeric riboflavin synthase from the pathogen B. abortus both as the apo protein and in complex with several ligands of interest. It is shown that ligand binding drives the assembly of the unique active site of the trimer, and these findings are complemented by a detailed kinetic study on this enzyme, in which marked inhibition by substrate and product was observed. Riboflavin synthase (RS) catalyzes the last step of riboflavin biosynthesis in microorganisms and plants, which corresponds to the dismutation of two molecules of 6,7-dimethyl-8-ribityllumazine to yield one molecule of riboflavin and one molecule of 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione. Owing to the absence of this enzyme in animals and the fact that most pathogenic bacteria show a strict dependence on riboflavin biosynthesis, RS has been proposed as a potential target for antimicrobial drug development. Eubacterial, fungal and plant RSs assemble as homotrimers lacking C{sub 3} symmetry. Each monomer can bind two substrate molecules, yet there is only one active site for the whole enzyme, which is located at the interface between two neighbouring chains. This work reports the crystallographic structure of RS from the pathogenic bacterium Brucella abortus (the aetiological agent of the disease brucellosis) in its apo form, in complex with riboflavin and in complex with two different product analogues, being the first time that the structure of an intact RS trimer with bound ligands has been solved. These crystal models support the hypothesis of enhanced flexibility in the particle and also highlight the role of the ligands in assembling the unique active site. Kinetic and binding studies were also performed to complement these findings. The structural and biochemical information generated may be useful for the rational design of novel RS inhibitors with antimicrobial activity.

  8. Desenvolvimento e avaliação de uma cepa knockout de Brucella abortus obtida pela deleção do gene virB10 Development and evaluation of a strain of Brucella abortus gotten by the knockout of the virB10 gene

    OpenAIRE

    Fabiane G. de Souza; Ana L.A.R. Osório; Bárbara G. Csordas; Rafael Q. Prado; Carina Elisei; Cleber O. Soares; Araújo, Flábio R; Stênio P Fragoso; Grácia M.S. Rosinha

    2009-01-01

    Brucella spp. são bactérias gram-negativas, intracelulares facultativas que são patogênicas para muitas espécies de mamíferos causando a brucelose, uma zoonose difundida mundialmente. Por isso a busca de alternativas de controle mais eficientes se faz necessário como o desenvolvimento de novas cepas que possam ser testadas como potenciais imunógenos. Neste estudo realizou-se a deleção do gene virB10 da cepa S2308 de Brucella abortus gerando uma cepa knockout provavelmente incapaz de produzir ...

  9. Evaluation of Brucella abortus S19 vaccines commercialized in Brazil: immunogenicity, residual virulence and MLVA15 genotyping.

    Science.gov (United States)

    Miranda, Karina Leite; Poester, Fernando Padilla; Minharro, Silvia; Dorneles, Elaine Maria Seles; Stynen, Ana Paula Reinato; Lage, Andrey Pereira

    2013-06-24

    Live attenuated Brucella abortus S19 is the most effective vaccine against brucellosis in cattle. The assessment of the immunological parameters is essential to guarantee the biological quality of live anti-bacteria vaccines. The evaluation of genetic stability of live bacterial vaccines is also important in quality control. The aims of the present study were to compare (i) the immunogenicity and residual virulence, and (ii) the genotypic profile (MLVA15) of the eight S19 vaccines commercialized in Brazil to the USDA S19 reference strain. Two batches of each of the eight S19 commercial vaccines used in Brazil (A-H) were tested. They were submitted to the potency and residual virulence in vivo tests recommended by OIE and typed by the multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) described for Brucella spp. Our results demonstrated that all S19 vaccines commercialized in Brazil would be approved by Brazilian and OIE recommendations for potency and residual virulence. Furthermore, the S19 vaccine is genetically very homogeneous, as all but two batches (from the same manufacturer) tested showed identical MLVA15 profile. The two batches with different profiles presented six repeat units in locus Bruce07, instead of the five found in all other strains, including the USDA S19 reference strain. Although presenting a slightly different profile, this vaccine was also protective, as demonstrated by the immunogenicity and residual virulence assays performed. Therefore, the commercial Brazilian S19 vaccines were in accordance to Brazilian and international standards for immunogenicity and residual virulence tests. Moreover, our results also show that MLVA could be a useful inclusion to the list of in vitro tests required by the official control authorities to be applied to the commercial S19 vaccines, as an efficient assay to guarantee the quality and stability of the vaccine strains. PMID:23664986

  10. Expression of MPB83 from Mycobacterium bovis in Brucella abortus S19 induces specific cellular immune response against the recombinant antigen in BALB/c mice.

    Science.gov (United States)

    Sabio y García, Julia V; Bigi, Fabiana; Rossetti, Osvaldo; Campos, Eleonora

    2010-12-01

    Immunodominant MPB83 antigen from Mycobacterium bovis was expressed as a chimeric protein fused to either β-galactosidase, outer membrane lipoprotein OMP19 or periplasmic protein BP26 in gram-negative Brucella abortus S19, in all cases driven by each gene's own promoter. All fusion proteins were successfully expressed and localized in the expected subcellular fraction. Moreover, OMP19-MPB83 was processed as a lipoprotein when expressed in B. abortus. Splenocytes from BALB/c mice immunized with the recombinant S19 strains carrying the genes coding for the heterologous antigens in replicative plasmids, showed equally specific INF-γ production in response to MPB83 stimulation. Association to the lipid moiety of OMP19 presented no advantage in terms of immunogenicity for MPB83. In contrast, fusion to BP26, which was encoded by an integrative plasmid, resulted in a weaker immune response. None of the constructions affected the survival rate or the infection pattern of Brucella. We concluded that B. abortus S19 is an appropriate candidate for the expression of M. bovis antigens both associated to the membrane or cytosolic fraction and may provide the basis for a future combined vaccine for bovine brucellosis and tuberculosis. PMID:20888425

  11. Overview of the activity of a Brucella abortus preparation, Bru-Pel.

    Science.gov (United States)

    Youngner, J S; Feingold, D S; Keleti, G

    1978-11-01

    The properties of a nonviable, aqueous ether-extracted Brucela abortus preparation, Bru-Pel, are described. In addition to inducing a "virus-type" interferon response and protecting mice against challenge with otherwise lethal doses of Semliki Forest virus, Bru-Pel is demonstrated to have potent antitumor properties in mice. These antitumor effects appear to be mediated by an increase in nonspecific resistance similar to that seen with other experimental antitumor agents.

  12. A Bile Salt Hydrolase of Brucella abortus Contributes to the Establishment of a Successful Infection through the Oral Route in Mice▿ †

    Science.gov (United States)

    Delpino, M. Victoria; Marchesini, María I.; Estein, Silvia M.; Comerci, Diego J.; Cassataro, Juliana; Fossati, Carlos A.; Baldi, Pablo C.

    2007-01-01

    Choloylglycine hydrolase (CGH), a bile salt hydrolase, has been annotated in all the available genomes of Brucella species. We obtained the Brucella CGH in recombinant form and demonstrated in vitro its capacity to cleave glycocholate into glycine and cholate. Brucella abortus 2308 (wild type) and its isogenic Δcgh deletion mutant exhibited similar growth rates in tryptic soy broth in the absence of bile. In contrast, the growth of the Δcgh mutant was notably impaired by both 5% and 10% bile. The bile resistance of the complemented mutant was similar to that of the wild-type strain. In mice infected through the intragastric or the intraperitoneal route, splenic infection was significantly lower at 10 and 20 days postinfection in animals infected with the Δcgh mutant than in those infected with the wild-type strain. For both routes, no differences in spleen CFU were found between animals infected with the wild-type strain and those infected with the complemented mutant. Mice immunized intragastrically with recombinant CGH mixed with cholera toxin (CGH+CT) developed a specific mucosal humoral (immunoglobulin G [IgG] and IgA) and cellular (interleukin-2) immune responses. Fifteen days after challenge by the same route with live B. abortus 2308 cells, splenic CFU counts were 10-fold lower in mice immunized with CGH+CT than in mice immunized with CT or phosphate-buffered saline. This study shows that CGH confers on Brucella the ability to resist the antimicrobial action of bile salts. The results also suggest that CGH may contribute to the ability of Brucella to infect the host through the oral route. PMID:17088355

  13. 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 (Panti-rO+rP polysera exhibited enhanced viability against challenge with B. abortus 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 (P<0.001) protected in the rO+rP group (log units of protection, spleen: 2.38, 2.12; liver: 1.04, 0.81, respectively) than in rO (spleen: 1.43, 1.21; liver: 0.7, 0.47) and rP (spleen: 1.24, 1.17; liver: 0.65, 0.34). Findings from this study depicted that rO+rP cocktail is highly immunogenic with the Th1 predominant serum antibody titers and T-cell mediated immune protection, would be a valuable intervention in the development of a safer and improved Brucella vaccine.

  14. Vaccination with Brucella abortus recombinant in vivo-induced antigens reduces bacterial load and promotes clearance in a mouse model for infection.

    Directory of Open Access Journals (Sweden)

    Jake E Lowry

    Full Text Available Current vaccines used for the prevention of brucellosis are ineffective in inducing protective immunity in animals that are chronically infected with Brucella abortus, such as elk. Using a gene discovery approach, in vivo-induced antigen technology (IVIAT on B. abortus, we previously identified ten loci that encode products up-regulated during infection in elk and consequently may play a role in virulence. In our present study, five of the loci (D15, 0187, VirJ, Mdh, AfuA were selected for further characterization and compared with three additional antigens with virulence potential (Hia, PrpA, MltA. All eight genes were PCR-amplified from B. abortus and cloned into E. coli. The recombinant products were then expressed, purified, adjuvanted, and delivered subcutaneously to BALB/c mice. After primary immunization and two boosts, mice were challenged i.p. with 5 x 10⁴ CFU of B. abortus strain 19. Spleens from challenged animals were harvested and bacterial loads determined by colony count at various time points. While vaccination with four of the eight individual proteins appeared to have some effect on clearance kinetics, mice vaccinated with recombinant Mdh displayed the most significant reduction in bacterial colonization. Furthermore, mice immunized with Mdh maintained higher levels of IFN-γ in spleens compared to other treatment groups. Collectively, our in vivo data gathered from the S19 murine colonization model suggest that vaccination with at least three of the IVIAT antigens conferred an enhanced ability of the host to respond to infection, reinforcing the utility of this methodology for the identification of potential vaccine candidates against brucellosis. Mechanisms for immunity to one protein, Mdh, require further in vitro exploration and evaluation against wild-type B. abortus challenge in mice, as well as other hosts. Additional studies are being undertaken to clarify the role of Mdh and other IVI antigens in B. abortus virulence

  15. Vaccination with Brucella abortus recombinant in vivo-induced antigens reduces bacterial load and promotes clearance in a mouse model for infection.

    Science.gov (United States)

    Lowry, Jake E; Isaak, Dale D; Leonhardt, Jack A; Vernati, Giulia; Pate, Jessie C; Andrews, Gerard P

    2011-01-01

    Current vaccines used for the prevention of brucellosis are ineffective in inducing protective immunity in animals that are chronically infected with Brucella abortus, such as elk. Using a gene discovery approach, in vivo-induced antigen technology (IVIAT) on B. abortus, we previously identified ten loci that encode products up-regulated during infection in elk and consequently may play a role in virulence. In our present study, five of the loci (D15, 0187, VirJ, Mdh, AfuA) were selected for further characterization and compared with three additional antigens with virulence potential (Hia, PrpA, MltA). All eight genes were PCR-amplified from B. abortus and cloned into E. coli. The recombinant products were then expressed, purified, adjuvanted, and delivered subcutaneously to BALB/c mice. After primary immunization and two boosts, mice were challenged i.p. with 5 x 10⁴ CFU of B. abortus strain 19. Spleens from challenged animals were harvested and bacterial loads determined by colony count at various time points. While vaccination with four of the eight individual proteins appeared to have some effect on clearance kinetics, mice vaccinated with recombinant Mdh displayed the most significant reduction in bacterial colonization. Furthermore, mice immunized with Mdh maintained higher levels of IFN-γ in spleens compared to other treatment groups. Collectively, our in vivo data gathered from the S19 murine colonization model suggest that vaccination with at least three of the IVIAT antigens conferred an enhanced ability of the host to respond to infection, reinforcing the utility of this methodology for the identification of potential vaccine candidates against brucellosis. Mechanisms for immunity to one protein, Mdh, require further in vitro exploration and evaluation against wild-type B. abortus challenge in mice, as well as other hosts. Additional studies are being undertaken to clarify the role of Mdh and other IVI antigens in B. abortus virulence and induction of

  16. Granulocyte-Macrophage Colony-Stimulating Factor- and Tumor Necrosis Factor Alpha-Mediated Matrix Metalloproteinase Production by Human Osteoblasts and Monocytes after Infection with Brucella abortus

    Science.gov (United States)

    Scian, Romina; Barrionuevo, Paula; Giambartolomei, Guillermo H.; Fossati, Carlos A.; Baldi, Pablo C.; Delpino, M. Victoria

    2011-01-01

    Osteoarticular complications are common in human brucellosis, but the pathogenic mechanisms involved are largely unknown. Since matrix metalloproteinases (MMPs) are involved in joint and bone damage in inflammatory and infectious diseases, we investigated the production of MMPs by human osteoblasts and monocytes, either upon Brucella abortus infection or upon reciprocal stimulation with factors produced by each infected cell type. B. abortus infection of the normal human osteoblastic cell line hFOB 1.19 triggered a significant release of MMP-2, which was mediated in part by granulocyte-macrophage colony-stimulating factor (GM-CSF) acting on these same cells. Supernatants from infected osteoblasts exhibited increased levels of monocyte chemoattractant protein 1 and induced the migration of human monocytes (THP-1 cell line). Infection with B. abortus induced a high MMP-9 secretion in monocytes, which was also induced by heat-killed B. abortus and by the Omp19 lipoprotein from B. abortus. These effects were mediated by Toll-like receptor 2 and by the action of tumor necrosis factor alpha (TNF-α) produced by these same cells. Supernatants from B. abortus-infected monocytes induced MMP-2 secretion in uninfected osteoblasts, and this effect was mediated by TNF-α. Similarly, supernatants from infected osteoblasts induced MMP-9 secretion in uninfected monocytes. This effect was mediated by GM-CSF, which induced TNF-α production by monocytes, which in turn induced MMP-9 in these cells. These results suggest that MMPs could be potentially involved in the tissue damage observed in osteoarticular brucellosis. PMID:20956574

  17. Molecular cloning and characterization of cgt, the Brucella abortus cyclic beta-1,2-glucan transporter gene, and its role in virulence.

    Science.gov (United States)

    Roset, Mara S; Ciocchini, Andrés E; Ugalde, Rodolfo A; Iñón de Iannino, Nora

    2004-04-01

    The animal pathogen Brucella abortus contains a gene cgt, which complemented Sinorhizobium meliloti nodule development (ndvA) and Agrobacterium tumefaciens chromosomal virulence (chvA) mutants. Complemented strains recovered the presence of anionic cyclic beta-1,2-glucan, motility, tumor induction in A. tumefaciens, and nodule occupancy in S. meliloti, all traits strictly associated with the presence of cyclic beta-1,2-glucan in the periplasm. Nucleotide sequencing revealed that B. abortus cgt contains a 1,797-bp open reading frame coding for a predicted membrane protein of 599 amino acids (65.9 kDa) that is 58.5 and 59.9% identical to S. meliloti NdvA and A. tumefaciens ChvA, respectively. Additionally, B. abortus cgt, like S. meliloti ndvA and A. tumefaciens chvA possesses ATP-binding motifs and the ABC signature domain features of a typical ABC transporter. Characterization of Cgt was carried out by the construction of null mutants in B. abortus 2308 and S19 backgrounds. Both mutants do not transport cyclic beta-1,2-glucan to the periplasm, as shown by the absence of anionic cyclic glucan, and they display reduced virulence in mice and defective intracellular multiplication in HeLa cells. These results suggest that cyclic beta-1,2-glucan must be transported into the periplasmatic space to exert its action as a virulence factor. PMID:15039351

  18. The Attenuated Brucella abortus Strain 19 Invades, Persists in, and Activates Human Dendritic Cells, and Induces the Secretion of IL-12p70 but Not IL-23.

    Directory of Open Access Journals (Sweden)

    Mario Weinhold

    Full Text Available Bacterial vectors have been proposed as novel vaccine strategies to induce strong cellular immunity. Attenuated strains of Brucella abortus comprise promising vector candidates since they have the potential to induce strong CD4(+ and CD8(+ T-cell mediated immune responses in the absence of excessive inflammation as observed with other Gram-negative bacteria. However, some Brucella strains interfere with the maturation of dendritic cells (DCs, which is essential for antigen-specific T-cell priming. In the present study, we investigated the interaction of human monocyte-derived DCs with the smooth attenuated B. abortus strain (S 19, which has previously been employed successfully to vaccinate cattle.We first looked into the potential of S19 to hamper the cytokine-induced maturation of DCs; however, infected cells expressed CD25, CD40, CD80, and CD86 to a comparable extent as uninfected, cytokine-matured DCs. Furthermore, S19 activated DCs in the absence of exogeneous stimuli, enhanced the expression of HLA-ABC and HLA-DR, and was able to persist intracellularly without causing cytotoxicity. Thus, DCs provide a cellular niche for persisting brucellae in vivo as a permanent source of antigen. S19-infected DCs produced IL-12/23p40, IL-12p70, and IL-10, but not IL-23. While heat-killed bacteria also activated DCs, soluble mediators were not involved in S19-induced activation of human DCs. HEK 293 transfectants revealed cellular activation by S19 primarily through engagement of Toll-like receptor (TLR2.Thus, as an immunological prerequisite for vaccine efficacy, B. abortus S19 potently infects and potently activates (most likely via TLR2 human DCs to produce Th1-promoting cytokines.

  19. The Attenuated Brucella abortus Strain 19 Invades, Persists in, and Activates Human Dendritic Cells, and Induces the Secretion of IL-12p70 but Not IL-23

    Science.gov (United States)

    Weinhold, Mario; Eisenblätter, Martin; Jasny, Edith; Fehlings, Michael; Finke, Antje; Gayum, Hermine; Rüschendorf, Ursula; Renner Viveros, Pablo; Moos, Verena; Allers, Kristina; Schneider, Thomas; Schaible, Ulrich E.; Schumann, Ralf R.; Mielke, Martin E.; Ignatius, Ralf

    2013-01-01

    Background Bacterial vectors have been proposed as novel vaccine strategies to induce strong cellular immunity. Attenuated strains of Brucella abortus comprise promising vector candidates since they have the potential to induce strong CD4+ and CD8+ T-cell mediated immune responses in the absence of excessive inflammation as observed with other Gram-negative bacteria. However, some Brucella strains interfere with the maturation of dendritic cells (DCs), which is essential for antigen-specific T-cell priming. In the present study, we investigated the interaction of human monocyte-derived DCs with the smooth attenuated B. abortus strain (S) 19, which has previously been employed successfully to vaccinate cattle. Methodology/Principal findings We first looked into the potential of S19 to hamper the cytokine-induced maturation of DCs; however, infected cells expressed CD25, CD40, CD80, and CD86 to a comparable extent as uninfected, cytokine-matured DCs. Furthermore, S19 activated DCs in the absence of exogeneous stimuli, enhanced the expression of HLA-ABC and HLA-DR, and was able to persist intracellularly without causing cytotoxicity. Thus, DCs provide a cellular niche for persisting brucellae in vivo as a permanent source of antigen. S19-infected DCs produced IL-12/23p40, IL-12p70, and IL-10, but not IL-23. While heat-killed bacteria also activated DCs, soluble mediators were not involved in S19-induced activation of human DCs. HEK 293 transfectants revealed cellular activation by S19 primarily through engagement of Toll-like receptor (TLR)2. Conclusions/Significance Thus, as an immunological prerequisite for vaccine efficacy, B. abortus S19 potently infects and potently activates (most likely via TLR2) human DCs to produce Th1-promoting cytokines. PMID:23805193

  20. Safety of the novel influenza viral vector Brucella abortus vaccine in pregnant heifers

    OpenAIRE

    Kaissar Tabynov; Sholpan Ryskeldinova; Zhailaubay Kydyrbayev; Abylai Sansyzbay

    2016-01-01

    ABSTRACT: The present study provides the first information about the safety of a new influenza viral vector vaccine expressing the Brucella ribosomal protein L7/L12 or Omp16 containing the adjuvant Montanide Gel01 in pregnant heifers. Immunization of pregnant heifers was conducted via the conjunctival (n=10) or subcutaneous (n=10) route using cross prime and booster vaccination schedules at an interval of 28 days. The vector vaccine was evaluated in comparison with positive control groups vac...

  1. Different resistance patterns of reference and field strains of Brucella abortus

    OpenAIRE

    Karina L. Miranda; Elaine M S Dorneles; Poester, Fernando P; Paulo S. Martins Filho; Pauletti, Rebeca B.; Andrey P. Lage

    2015-01-01

    The aim of this study was to evaluate the growth of the B. abortus reference strains and field isolates on media containing different inhibitor agents. Reference strains were seeded on tryptose agar containing: i-erythritol (1.0 mg/mL), fuchsin (20 μg/mL and 80 μg/mL), thionin (2.5 μg/mL and 10 μg/mL), rifampicin (200 μg/mL) and safranin O (200 μg/mL). Field isolates were tested only on media containing i-erythritol, rifampicin and thionin. Furthermore, each suspension was also inoculated on ...

  2. 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.

  3. The Dutch Brucella abortus monitoring programme for cattle: the impact of false-positive serological reactions and comparison of serological tests.

    Science.gov (United States)

    Emmerzaal, A; de Wit, J J; Dijkstra, Th; Bakker, D; van Zijderveld, F G

    2002-02-01

    The Dutch national Brucella abortus eradication programme for cattle started in 1959. Sporadic cases occurred yearly until 1995; the last infected herd was culled in 1996. In August 1999 the Netherlands was declared officially free of bovine brucellosis by the European Union. Before 1999, the programme to monitor the official Brucella-free status of bovine herds was primarily based on periodical testing of dairy herds with the milk ring test (MRT) and serological testing of all animals older than 1 year of age from non-dairy herds, using the micro-agglutination test (MAT) as screening test. In addition, serum samples of cattle that aborted were tested with the MAT. The high number of false positive reactions in both tests and the serum agglutination test (SAT) and complement fixation test (CFT) used for confirmation seemed to result in unnecessary blockade of herds, subsequent testing and slaughter of animals. For this reason, a validation study was performed in which three indirect enzyme-linked immunosorbent assays (ELISAs), the CFT and the SAT were compared using a panel of sera from brucellosis-free cattle, sera from experimentally infected cattle, and sera from cattle experimentally infected with bacteria which are known to induce cross-reactive antibodies (Pasteurella, Salmonella, Yersinia, and Escherichia). Moreover, four ELISAs and the MRT were compared using a panel of 1000 bulk milk samples from Brucella-free herds and 12 milk samples from Brucella abortus- infected cattle. It is concluded that the ELISA obtained from ID-Lelystad is the most suitable test to monitor the brucelosis free status of herds because it gives rise to fewer false-positive reactions than the SAT.

  4. Oral vaccination with microencapsuled strain 19 vaccine confers enhanced protection against Brucella abortus strain 2308 challenge in red deer (Cervus elaphus elaphus).

    Science.gov (United States)

    Arenas-Gamboa, Angela M; Ficht, Thomas A; Davis, Donald S; Elzer, Philip H; Kahl-McDonagh, Melissa; Wong-Gonzalez, Alfredo; Rice-Ficht, Allison C

    2009-10-01

    Bison (Bison bison) and elk (Cervus elaphus nelsoni) in the Greater Yellowstone Area (GYA), USA, are infected with Brucella abortus, the causative agent of bovine brucellosis, and they serve as a wildlife reservoir for the disease. Bovine brucellosis recently has been transmitted from infected elk to cattle in Montana, Wyoming, and Idaho and has resulted in their loss of brucellosis-free status. An efficacious Brucella vaccine with a delivery system suitable for wildlife would be a valuable tool in a disease prevention and control program. We evaluated Strain 19 (S19) in a sustained release vehicle consisting of alginate microspheres containing live vaccine. In a challenge study using red deer (Cervus elaphus elaphus) as a model for elk, alginate, a naturally occurring polymer combined with a protein of Fasciola hepatica vitelline protein B was used to microencapsulate S19. Red deer were orally or subcutaneously immunized with 1.5 x 10(10) colony-forming units (CFUs) using microencapsulated S19. Humoral and cellular profiles were analyzed bimonthly throughout the study. The vaccinated red deer and nonvaccinated controls were challenged 1 yr postimmunization conjunctivally with 1 x 10(9) CFUs of B. abortus strain 2308. Red deer vaccinated with oral microencapsulated S19 had a statistically significant lower bacterial tissue load compared with controls. These data indicate for the first time that protection against Brucella-challenge can be achieved by combining a commonly used vaccine with a novel oral delivery system such as alginate-vitelline protein B microencapsulation. This system is a potential improvement for efficacious Brucella-vaccine delivery to wildlife in the GYA. PMID:19901378

  5. Excretion of Brucella abortus vaccine B19 strain during a reproductive cycle in dairy cows

    Directory of Open Access Journals (Sweden)

    W. A. Pacheco

    2012-06-01

    Full Text Available This paper aimed to determine the excretion period of B19 vaccine strain during a complete reproductive cycle (from estrus synchronization, artificial insemination, pregnancy and until 30 days after parturition of dairy cows from 3 to 9 years old that were previously vaccinated from 3 to 8 months. Three groups were monitored with monthly milk and urine collection during 12 months: G1 with seven cows from 3 to 4 years old; G2 with three cows from 5 to 6 years old; and G3 with four cows from 7 to 9 years old. Urine and milk samples were submitted to bacteriological culture and urine and PCR reactions for detection of Brucella spp. and PCR-multiplex for B19 strain identification. Ring test (RT was also performed in the milk samples, and serum samples were tested by buffered acidified plate antigen test (BAPA. All animals were serologically negative at BAPA and Brucella spp. was not isolated from both urine and milk samples. RT revealed 13/210 (6.2% positive milk samples. PCR reactions detected DNA of Brucella spp. in 86/420 (20.5% samples. In urine it was found a significantly higher frequency (35.2%; 74/210 than in milk (5.7%; 12/210, more frequently from the estrus to 150 days of pregnancy and after parturition (6.7%; 10/150, and from 150 days of pregnancy to parturition (3.4%; 2/60, and they were all identified as B19 strain. In three groups, intermittent excretion of B19 strain was detected mainly in urine samples, which confirmed its multiplication and persistence in cows for until 9 years.

  6. 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.

  7. A combined DNA vaccine encoding BCSP31, SOD, and L7/L12 confers high protection against Brucella abortus 2308 by inducing specific CTL responses.

    Science.gov (United States)

    Yu, Da-Hai; Hu, Xi-Dan; Cai, Hong

    2007-06-01

    We constructed a combined DNA vaccine comprising genes encoding the antigens BCSP31, superoxide dismutase (SOD), and L7/L12 and evaluated its immunogenicity and protective efficacy. Immunization of mice with the combined DNA vaccine offered high protection against Brucella abortus (B. abortus) infection. The vaccine induced a vigorous specific immunoglobulin G (IgG) response, with higher IgG2a than IgG1 titers. Cytokine profiling performed at the same time showed a biased Th1-type immune response with significantly increased interferon-gamma and tumor necrosis factor-alpha stimulation. CD8(+), but not CD4(+), T cells accumulated at significantly higher levels after administration of the vaccine. Granzyme B-producing CD8(+) T cells were significantly higher in number in samples prepared from combined DNA-vaccinated mice compared with S19-vaccinated mice, demonstrating that the cytotoxicity lysis pathway is involved in the response to Brucella infection. The success of our combined DNA vaccine in a mouse model suggests its potential efficacy against brucellosis infection in large animals. PMID:17570767

  8. virB-Mediated Survival of Brucella abortus in Mice and Macrophages Is Independent of a Functional Inducible Nitric Oxide Synthase or NADPH Oxidase in Macrophages

    Science.gov (United States)

    Sun, Yao-Hui; den Hartigh, Andreas B.; de Lima Santos, Renato; Adams, L. Garry; Tsolis, Renée M.

    2002-01-01

    The Brucella abortus virB locus is required for establishing chronic infection in the mouse. Using in vitro and in vivo models, we investigated whether virB is involved in evasion of the bactericidal activity of NADPH oxidase and the inducible nitric oxide synthase (iNOS) in macrophages. Elimination of NADPH oxidase or iNOS activity in macrophages in vitro increased recovery of wild-type B. abortus but not recovery of a virB mutant. In mice lacking either NADPH oxidase or iNOS, however, B. abortus infected and persisted to the same extent as it did in congenic C57BL/6 mice up until 60 days postinfection, suggesting that these host defense mechanisms are not critical for limiting bacterial growth in the mouse. A virB mutant did not exhibit increased survival in either of the knockout mouse strains, indicating that this locus does not contribute to evasion of nitrosative or oxidative killing mechanisms in vivo. PMID:12183526

  9. Interferon-γ is crucial for surviving a Brucella abortus infection in both resistant C57BL/6 and susceptible BALB/c mice

    Science.gov (United States)

    Murphy, Erin A; Sathiyaseelan, Janaki; Parent, Michelle A; Zou, Baixiang; Baldwin, Cynthia L

    2001-01-01

    Brucella abortus is an intracellular bacterial pathogen that causes chronic infections in humans and a number of agriculturally important species of animals. It has been shown that BALB/c mice are more susceptible to infections with virulent strains of Brucella abortus than C57BL/6 or C57BL/10 strains. In experiments described here, gene knock-out mice were utilized to elucidate some of the salient components of resistance. Resistant C57BL/6 mice with gene deletions or disruptions in the interferon-γ (IFN-γ), perforin or β2-microglobulin genes had decreased abilities to control intracellular infections with B. abortus strain 2308 during the first week after infection. However, only the IFN-γ knock-out mice had a sustained inability to control infections and this resulted in death of the mice at approximately 6 weeks post-infection. These mice had a continual increase in the number of bacterial colony-forming units (CFU) in their spleens until death. When BALB/c mice with the disrupted IFN-γ gene were infected they had more splenic CFU at one week post-infection than control mice but the increase was not statistically significant and by 3 weeks they did not have more CFU than control mice. Moreover, the number of splenic bacteria did not increase in the BALB/c IFN-γ knock-out mice between 6 and 10·5 weeks, although they died at 10·5 weeks, the time by which normal BALB/c mice were clearing the infection. Death in both strains of IFN-γ gene disrupted mice coincided with symptoms of cachexia and macrophages comprised ≥75% of the splenic leucocytes. PMID:11529943

  10. Immunization of Mice with Recombinant Protein CobB or AsnC Confers Protection against Brucella abortus Infection

    OpenAIRE

    Simei Fu; Jie Xu; Xianbo Li; Yongfei Xie; Yefeng Qiu; Xinying Du; Shuang Yu; Yaoxia Bai; Yanfen Chen; Tongkun Wang; Zhoujia Wang; Yaqing Yu; Guangneng Peng; Kehe Huang; Liuyu Huang

    2012-01-01

    Due to drawbacks of live attenuated vaccines, much more attention has been focused on screening of Brucella protective antigens as subunit vaccine candidates. Brucella is a facultative intracellular bacterium and cell mediated immunity plays essential roles for protection against Brucella infection. Identification of Brucella antigens that present T-cell epitopes to the host could enable development of such vaccines. In this study, 45 proven or putative pathogenesis-associated factors of Bruc...

  11. Vaccination with recombinant flagellar proteins FlgJ and FliN induce protection against Brucella abortus 544 infection in BALB/c mice.

    Science.gov (United States)

    Li, Xianbo; Xu, Jie; Xie, Yongfei; Qiu, Yefeng; Fu, Simei; Yuan, Xitong; Ke, Yuehua; Yu, Shuang; Du, Xinying; Cui, Mingquan; Chen, Yanfen; Wang, Tongkun; Wang, Zhoujia; Yu, Yaqing; Huang, Kehe; Huang, Liuyu; Peng, Guangneng; Chen, Zeliang; Wang, Yufei

    2012-12-28

    Brucella has been considered as a non-motile, facultative intracellular pathogenic bacterium. However, the genome sequences of different Brucella species reveal the presence of the flagellar genes needed for the construction of a functional flagellum. Due to its roles in the interaction between pathogen and host, we hypothesized that some of the flagellar proteins might induce protective immune responses and these proteins will be good subunit vaccine candidates. This study was conducted to screening of protective antigens among these flagellar proteins. Firstly, according to the putative functional roles, a total of 30 flagellar genes of Brucella abortus were selected for in vitro expression. 15 of these flagellar genes were successfully expressed as his-tagged recombinant proteins in Escherichia coli ER2566. Then, these proteins were purified and used to analyze their T cell immunity induction activity by an in vitro gamma interferon (IFN-γ) assay. Five of the flagellar proteins could stimulate significantly higher levels of IFN-γ secretion in splenocytes from S19 immunized mice, indicating their T cell induction activity. Finally, immunogenicity and protection activity of these 5 flagellar proteins were evaluated in BALB/c mice. Results showed that immunization with FlgJ (BAB1_0260) or FliN (BAB2_0122) plus adjuvant could provide protection against B. abortus 544 infection. Furthermore, mice immunized with FlgJ and FliN developed a vigorous immunoglobulin G response, and in vitro stimulation of their splenocytes with immunizing proteins induced the secretion of IFN-γ. Altogether, these data suggest that flagellar proteins FlgJ and FliN are protective antigens that could produce humoral and cell-mediated responses in mice and candidates for use in future studies of vaccination against brucellosis. PMID:22854331

  12. 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...

  13. 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.

  14. Intermediate rough Brucella abortus S19Δper mutant is DIVA enable, safe to pregnant guinea pigs and confers protection to mice.

    Science.gov (United States)

    Lalsiamthara, Jonathan; Gogia, Neha; Goswami, Tapas K; Singh, R K; Chaudhuri, Pallab

    2015-05-21

    Brucella abortus S19 is a smooth strain used as live vaccine against bovine brucellosis. Smooth lipopolysaccharide (LPS) is responsible for its residual virulence and serological interference. Rough mutants defective of LPS are more attenuated but confers lower level of protection. We describe a modified B. abortus S19 strain, named as S19Δper, which exhibits intermediate rough phenotype with residual O-polysaccharide (OPS). Deletion of perosamine synthetase gene resulted in substantial attenuation of S19Δper mutant without affecting immunogenic properties. It mounted strong immune response in Swiss albino mice and conferred protection similar to S19 vaccine. Immunized mice produced higher levels of IFN-γ, IgG2a and thus has immune response inclined towards Th1 cell mediated immunity. Sera from immunized animals did not show agglutination reaction with RBPT antigen and thus could serve as DIVA (Differentiating Infected from Vaccinated Animals) vaccine. S19Δper mutant displayed more susceptibility to serum complement mediated killing and sensitivity to polymyxin B. Pregnant guinea pigs injected with S19Δper mutant completed full term of pregnancy and did not cause abortion, still birth or birth of weak offspring. S19Δper mutant with intermediate rough phenotype displayed remarkable resemblance to S19 vaccine strain with improved properties of safety, immunogenicity and DIVA capability for control of bovine brucellosis. PMID:25869887

  15. Vaccination of adult animals with a reduced dose of Brucella abortus S19 vaccine to control brucellosis on dairy farms in endemic areas of India.

    Science.gov (United States)

    Chand, Puran; Chhabra, Rajesh; Nagra, Juhi

    2015-01-01

    Bovine brucellosis is an economically important disease which seriously affects dairy farming by causing colossal losses. It can be controlled by practicing vaccination of animals with Brucella abortus S19 vaccine (S19 vaccine). In the present study, adult bovines were vaccinated on seven dairy farms with a reduced dose of S19 vaccine to control brucellosis. Serological screening of adult animals (N = 1,082) by Rose Bengal test (RBT) and ELISA prior to vaccination revealed the presence and absence of brucellosis on five and two farms, respectively. The positive animals (N = 171) were segregated and those which tested negative (N = 911) were vaccinated by conjunctival route with a booster after 4 months. The conjunctival vaccination induced weak antibody response in animals, which vanished within a period of 9 to 12 weeks. Abortion in 12 animals at various stages of pregnancy and post-vaccination was recorded, but none was attributed to S19 vaccine. However, virulent B. abortus was incriminated in six heifers, and the cause of abortion could not be established in six animals. The six aborted heifers perhaps acquired infection through in utero transmission or from the environment which remained undetected until abortion. These findings suggested that vaccination of adult animals with a reduced dose of S19 vaccine by conjunctival route did not produce adverse effects like abortion in pregnant animals and persistent vaccinal antibody titers, which are the major disadvantages of subcutaneous vaccination of adult animals. PMID:25274621

  16. Evaluation of Indirect Enzyme-Linked Immunosorbent Assays and IgG Avidity Assays Using a Protein A-Peroxidase Conjugate for Serological Distinction between Brucella abortus S19-Vaccinated and -Infected Cows ▿

    OpenAIRE

    Pajuaba, Ana C. A. M.; Deise A O Silva; Mineo, José R.

    2010-01-01

    This study aimed to evaluate the use of protein A-peroxidase (horseradish peroxidase [HRPO]) in indirect enzyme-linked immunosorbent assays (iELISAs) and IgG avidity assays for serological distinction between Brucella abortus S19-vaccinated and -infected cows. Four groups were analyzed: GI, 41 nonvaccinated seropositive cows; GII, 79 S19-vaccinated heifers analyzed at 3 months postvaccination; GIII, 105 S19-vaccinated cows analyzed after 24 months of age; and GIV, 278 nonvaccinated seronegati...

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

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    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.

  18. SNP Research on Brucella abortus Vaccine Strain A19%中国牛种布鲁氏菌疫苗株A19 SNP位点的研究

    Institute of Scientific and Technical Information of China (English)

    谭鹏飞; 南文龙; 彭大新; 毛开荣; 陈义平

    2014-01-01

    To distinguish Brucella abortus vaccine strain A19 from field strains, the single nucleotide polymorphism (SNP) signatures for B. abortus A19 were initially analyzed using bioinformatics method and gene sequencing. Then, the specificity of several SNPs was verified, by comparing the nucleotide sequences of these SNPs with common species and biovars of Brucella and three Brucella vaccines. The results showed 29 SNPs were successfully screened from genome of B. abortus A19. Furthermore, ClpX G825-C825,LysR A605-C605and Omp2b G503-A503were confirmed to be specific to B. abortus A19 (or B. abortus S19). Our study systematically revealed the SNP distribution of B. abortus A19, which provided a molecular basis for differentiating B. abortus A19 from field strains.%为鉴别我国牛种布鲁氏菌疫苗株A19与野生菌株,运用生物信息学方法结合基因测序,对疫苗株A19基因组单核苷酸多态性( SNP )位点分析筛选,选取其中部分SNP位点,通过与布鲁氏菌常见种、生物型标准参考菌株和疫苗株基因组SNP 位置核苷酸测序比较,验证SNP 位点的A19特异性。结果表明,共筛选获得A19基因组29个SNP 位点,验证ClpX G825-C825、LysR A605-C605、Omp2b G503-A503这3个SNP位点为A19(或S19)特异,揭示了A19基因组SNP位点分布情况,为疫苗株A19与野生菌株鉴别提供了分子依据。

  19. Novel influenza virus vectors expressing Brucella L7/L12 or Omp16 proteins in cattle induced a strong T-cell immune response, as well as high protectiveness against B. abortus infection.

    Science.gov (United States)

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

    2014-04-11

    This paper presents the results of a study of the immunogenicity and protectiveness of new candidate vector vaccine against Brucella abortus - a bivalent vaccine formulation consisting of a mixture of recombinant influenza A subtype H5N1 or H1N1 (viral constructs vaccine formulation) viruses expressing Brucella ribosomal protein L7/L12 and Omp16, in cattle. To increase the effectiveness of the candidate vaccine, adjuvants such as Montanide Gel01 or chitosan were included in its composition. Immunization of cattle (heifers aged 1-1.5 years, 5 animals per group) with the viral constructs vaccine formulation only, or its combination with adjuvants Montanide Gel01 or chitosan, was conducted via the conjunctival method using cross prime (influenza virus subtype H5N1) and booster (influenza virus subtype H1N1) vaccination schedules at an interval of 28 days. Vaccine candidates were evaluated in comparison with the positive (B. abortus S19) and negative (PBS) controls. The viral constructs vaccine formulations, particularly in combination with Montanide Gel01 adjuvant promoted formation of IgG antibodies (with a predominance of antibodies of isotype IgG2a) against Brucella L7/L12 and Omp16 proteins in ELISA. Moreover, these vaccines in cattle induced a strong antigen-specific T-cell immune response, as indicated by a high number of CD4(+) and CD8(+) cells, as well as the concentration of IFN-γ, and most importantly provided a high level of protectiveness comparable to the commercial B. abortus S19 vaccine and superior to the B. abortus S19 vaccine in combination with Montanide Gel01 adjuvant. Based on these findings, we recommended the bivalent vaccine formulation containing the adjuvant Montanide Gel01 for practical use in cattle. PMID:24598723

  20. Bovine SLC11A1 3' UTR SSCP genotype evaluated by a macrophage in vitro killing assay employing a Brucella abortus strain.

    Science.gov (United States)

    Martinez, R; Toro, R; Montoya, F; Burbano, M; Tobón, J; Gallego, J; Dunner, S; Cañón, J

    2008-08-01

    The 3' untranslated region (3' UTR) of the bovine natural resistance-associated macrophage gene (NRAMP1 or SLC11A1) was genotyped in Colombian Creole Blanco Orejinegro (BON) (Bos taurus) (n = 140) and Zebu Brahman (Bos indicus) (Z) (n = 20) cattle and their crosses (BON x Zebu Brahman [B x Z] [n = 10]; Zebu Brahman x BON [Z x B] [n = 10]), and in animals from a Holstein x BON (H x B) (n = 10) cross. Direct sequencing and single-strand conformation polymorphism analysis (SSCP) helped in detecting the polymorphic behaviour. The association between resistance to brucellosis infection and SSCP genotype was evaluated using a macrophage in vitro killing assay employing a virulent Brucella abortus strain. The 3' UTR (GT) repeated polymorphism was gentoyped and its association with resistance to brucellosis was evaluated. When all breeds were grouped, a high frequency in the homozygote GT(12) (AA genotype) (0.823) and a very low frequency in the homozygote GT(10) (BB genotype) (0.047) were detected. The BON (0.963), Z x B (0.60) and H x B (1.00) cattle showed high GT(12) allele frequencies, unlike that seen for the B x Z and Zebu cattle (0.3002 and 0.218, respectively). The GT(10) allele was only found in the Zebu cattle (0.391). A significant association (p < 0.001) was found between the B. abortus macrophage in vitro killing assay phenotypes and the bovine SLC11A1 3' UTR genotypes, which suggests that the A allele may be associated with resistance. Because only nine animals had the BB genotype, the results require some confirmation in more extensive populations. PMID:18717968

  1. FREQÜÊNCIA DE AGLUTININAS ANTI-Brucella abortus EM CAPRINOS E OVINOS DO SERTÃO DO ESTADO DE PERNAMBUCO, BRASIL FREQUENCY OF ANTI-Brucella abortus AGGLUTININS IN GOATS AND sheep OF THE “SERTÃO” (BACKLANDS OF THE STATE OF PERNAMBUCO, BRAZIL

    Directory of Open Access Journals (Sweden)

    Vânia Lúcia de Assis Santana

    2008-12-01

    Full Text Available Objetivou-se investigar a freqüência de aglutininas anti-Brucella abortus em caprinos e ovinos do Sertão do Estado de Pernambuco, Brasil. Foram processadas 700 amostras de soros sangüíneos, das quais 340 eram da espécie caprina (115 machos e 225 fêmeas e 360 (136 machos e 224 fêmeas ovina. Empregou-se a técnica do antígeno acidificado tamponado (AAT corado com rosa bengala (RB. Das 340 amostras de caprinos avaliadas, duas (0,6% foram reagentes ao AAT. Não se observaram associações significativas para as variáveis faixa etária (p= 0,430, raça (p= 0,936 e sexo (p= 0,562. Das 360 amostras de ovinos, nove (2,5% foram reagentes. Também não houve associação significativa entre as variáveis analisadas e a soropositividade para brucelose: faixa etária (p= 0,522; raça (p= 0,576 e sexo (p= 0,461. Verificou-se associação significativa (p= 0,042 entre as espécies estudadas e soropositividade para brucelose nos animais investigados. A soropositividade para Brucella abortus em caprinos e ovinos foi descrita pela primeira vez no Sertão de Pernambuco, fato que pode dificultar o sucesso do Programa Nacional de Controle e Erradicação da Brucelose, tendo em vista que nessa região é comum a criação consorciada de pequenos ruminantes com bovinos, além de representar riscos à Saúde Pública.

    PALAVRAS-CHAVES: Brucelose, ovinos, caprinos, pequenos ruminantes, sorodiagnóstico. The objective was to investigate the frequency of anti-Brucella abortus agglutinins in goats and sheep of the backlands of the State of Pernambuco, Brazil. 700 samples of sanguine serums were processed, of which 340 were of the goat (115 males and 225 females and 360 (136 males and 224 females sheep. The technique of the Tamponed Acidified Antigen (AAT dyed with Bengalese Rose (BR was used. Of the 340 samples of goat evaluated two (0.6% were reactive to AAT. Significant associations were not observed for the variable age group (p = 0.430; race (p = 0

  2. Imunodifusão em gel de ágar com polissacarídeos de membrana de Brucella abortus 1119-3 no diagnóstico da brucelose bovina Membrane polysaccharides of Brucella abortus 1119-3 in agar gel immunodifusion test in diagnosis of bovine brucellosis

    Directory of Open Access Journals (Sweden)

    J. Megid

    1999-10-01

    Full Text Available Comparou-se a prova de imunodifusão em gel de ágar (IDGA, utilizando extrato polissacarídico (POLI O, obtido da amostra de B. abortus 1119-3, com os testes de soroaglutinação rápida em placa, de soroaglutinação lenta em tubos, de antígeno acidificado e de 2-mercaptoetanol para o diagnóstico da brucelose bovina. O IDGA mostrou alta especificidade, porém sensibilidade inferior aos métodos convencionais.An agar gel immunodiffusion (AGID test using the polysaccharide (POLIO O extract obtained from Brucella abortus strain 1119-3, was compared to plate agglutination, tube agglutination, rose bengal plate and mercaptoethanol tests in the diagnosis of bovine brucellosis. The AGID assay with B. aborturs polysaccharide antigens, presented a greater specificity but lower sensitivity for detecting brucellosis infected animals as compared to the conventional methods of antibody detection.

  3. Confirmação de infecção por Brucella abortus em um rebanho bovino certificado livre em Minas Gerais: relato de caso

    Directory of Open Access Journals (Sweden)

    P.M. Soares Filho

    2012-10-01

    Full Text Available Relata-se a ocorrência de um surto de brucelose em um rebanho de aproximadamente 1000 animais, livre da doença há 18 anos, certificado pelo Ministério da Agricultura, Pecuária e Abastecimento desde 2006. Dois animais reagiram aos testes sorológicos de diagnóstico por ocasião dos procedimentos de recertificação em 2008. Após o sacrifício deles, Brucella abortus, biovariedade 1, amostra não vacinal, foi isolada e identificada por meio de provas bioquímicas e de biologia molecular (PCR AMOS. A origem do agente no rebanho é de difícil determinação. No entanto, a adoção de procedimentos preconizados pelo Programa Nacional de Controle e Erradicação da Brucelose permitiu evitar a disseminação da enfermidade. Ocorrências como essas, em que rebanhos livres foram infectados após anos sem a ocorrência de brucelose, nunca haviam sido relatadas no Brasil.

  4. Serological profile of buffalo (Bubalus bubalis) female calves vaccinated with standard Brucella abortus strain 19 vaccine using rose bengal, 2-mercaptoethanol and complement fixation tests.

    Science.gov (United States)

    Nardi, G Júnior; Ribeiro, M G; Jorge, A M; Megid, J; Silva, L M P

    2012-03-01

    The serological profiles of 21 female buffaloes vaccinated between 3 and 8 months of age using Brucella abortus strain 19 (S19) were evaluated by rose bengal (RBT), 2-mercaptoethanol (2ME) and complement fixation (CFT) tests. The serum strains were collected in day zero, 15, 30, 45, 60th days and subsequently to each 30 months, until 720th day after vaccination. No animal showed reaction in day zero. In 15th day above 95% of animals revealed reaction in all tests. All the animals presented absence of reactions in CFT, RBT and 2ME tests at 270, 300 and 360 days after vaccination, respectively. Our finding highlighted early response in CFT compared than other conventional agglutination tests. None of animals presented oscillation of titers or reactions in any test after 360 day of study, which enables the use of these tests after this period without interference of antibodies from S19 vaccine origin between 3 and 8 months in buffalo heifers. PMID:22284623

  5. Evaluation of Brucella abortus S19 vaccine strains by bacteriological tests, molecular analysis of ery loci and virulence in BALB/c mice.

    Science.gov (United States)

    Mukherjee, Falguni; Jain, Jainendra; Grilló, Maria Jesús; Blasco, José María; Nair, Mrinalini

    2005-09-01

    Two Brucella abortus S19 commercial vaccine strains used for vaccination against brucellosis in India and three S19 strains available as international reference were examined by microbiological assays and molecular analysis of the ery loci involved in erythritol metabolism, and tested for residual virulence in BALB/c mice. According to the sensitivity to penicillin and i-erythritol, the five strains tested had the phenotypic characteristics of strain S19. However, on culture medium containing i-erythritol, all strains developed spontaneous i-erythritol resistant colonies at mutation rates ranging from 1.42x10(-2) to 1.33x10(-6). The S19 characteristic 702 bp deletion in the erythrulose 1-phosphate dehydrogenase gene of the ery locus was present only in the three reference strains but not in the two commercial vaccines. Both commercial strains and one of the reference strains showed reduced virulence in BALB/c mice. The presence or absence in S19 strains of the 702 bp deletion in the ery locus had no correlation with either the rates of spontaneous mutation to erythritol resistance or the residual virulence in mice. PMID:16081301

  6. Simultaneous subcutaneous and conjunctival administration of the influenza viral vector based Brucella abortus vaccine to pregnant heifers provides better protection against B. abortus 544 infection than the commercial B. abortus S19 vaccine.

    Science.gov (United States)

    Tabynov, Kaissar; Orynbayev, Mukhit; Renukaradhya, Gourapura J; Sansyzbay, Abylai

    2016-09-30

    In this study, we explored possibility of increasing the protective efficacy of our novel influenza viral vector based B. abortus vaccine (Flu-BA) in pregnant heifers by adapting an innovative method of vaccine delivery. We administered the vaccine concurrently via the conjunctival and subcutaneous routes to pregnant heifers, and these routes were previously tested individually. The Flu-BA vaccination of pregnant heifers (n=9) against a challenge B. abortus 544 infection provided protection from abortion, infection of heifers and fetuses/calves by 88.8%, 100% and 100%, respectively (alpha=0.004-0.0007 vs. negative control; n=7). Our candidate vaccine using this delivery method provided slightly better protection than the commercial B. abortus S19 vaccine in pregnant heifers (n=8), which provided protection from abortion, infection of heifers and fetuses/calves by 87.5%, 75% and 87.5%, respectively. This improved method of the Flu-BA vaccine administration is highly recommended for the recovery of farms which has high prevalence of brucellosis. PMID:27595898

  7. 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

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    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

  8. Use of S-[2,3-Bispalmitoyiloxy-(2R)-Propyl]-R-Cysteinyl-Amido-Monomethoxy Polyethylene Glycol as an Adjuvant Improved Protective Immunity Associated with a DNA Vaccine Encoding Cu,Zn Superoxide Dismutase of Brucella abortus in Mice

    OpenAIRE

    Retamal-Díaz, Angello; Riquelme-Neira, Roberto; Sáez, Darwin; Rivera, Alejandra; Fernández, Pablo; Cabrera, Alex; Guzmán, Carlos A.; Oñate, Angel

    2014-01-01

    This study was conducted to evaluate the immunogenicity and protective efficacy of a DNA vaccine encoding Brucella abortus Cu,Zn superoxide dismutase (SOD) using the Toll-like receptor 2/6 agonist S-[2,3-bispalmitoyiloxy-(2R)-propyl]-R-cysteinyl-amido-monomethoxy polyethylene glycol (BPPcysMPEG) as an adjuvant. Intranasal coadministration of BPPcysMPEG with a plasmid carrying the SOD-encoding gene (pcDNA-SOD) into BALB/c mice elicited antigen-specific humoral and cellular immune responses. Hu...

  9. The immunological properties of Brucella ribosomal preparations.

    Science.gov (United States)

    Corbel, M J

    1976-01-01

    Ribosomes were isolated from Brucella abortus strains 19 and 45/20 by disruption of the cells followed by differential ultracentrifugation. The ribosome preparations contained 2-3 components reacting in immunodiffusion tests but were free of detectable lipopolysaccharide-protein agglutinogen. They crossreacted with antisera to Br. abortus, Br. melitensis, Br. suis and Br. ovis and elicited intradermal delayed hypersensitivity reactions in animals infected with Br. abortus, Br. melitensis or Br. suis. The ribosomes were antigenic in rabbits, guinea pigs and mice. Those from Br. abortus S19 induced agglutinins reaction with smooth brucella strains whereas those from Br. abortus 45/20 induced agglutinins reacting with rough brucella strains. Cattle vaccinated with S19 or 45/20 vaccines or infected with Br. abortus developed pricipitins to ribosomal components at an early stage in the immune response. PMID:816681

  10. The Brucella abortus S19 DeltavjbR live vaccine candidate is safer than S19 and confers protection against wild-type challenge in BALB/c mice when delivered in a sustained-release vehicle.

    Science.gov (United States)

    Arenas-Gamboa, A M; Ficht, T A; Kahl-McDonagh, M M; Gomez, G; Rice-Ficht, A C

    2009-02-01

    Brucellosis is an important zoonotic disease of nearly worldwide distribution. Despite the availability of live vaccine strains for bovine (S19, RB51) and small ruminants (Rev-1), these vaccines have several drawbacks, including residual virulence for animals and humans. Safe and efficacious immunization systems are therefore needed to overcome these disadvantages. A vjbR knockout was generated in the S19 vaccine and investigated for its potential use as an improved vaccine candidate. Vaccination with a sustained-release vehicle to enhance vaccination efficacy was evaluated utilizing the live S19 DeltavjbR::Kan in encapsulated alginate microspheres containing a nonimmunogenic eggshell precursor protein of the parasite Fasciola hepatica (vitelline protein B). BALB/c mice were immunized intraperitoneally with either encapsulated or nonencapsulated S19 DeltavjbR::Kan at a dose of 1 x 10(5) CFU per animal to evaluate immunogenicity, safety, and protective efficacy. Humoral responses postvaccination indicate that the vaccine candidate was able to elicit an anti-Brucella-specific immunoglobulin G response even when the vaccine was administered in an encapsulated format. The safety was revealed by the absence of splenomegaly in mice that were inoculated with the mutant. Finally, a single dose with the encapsulated mutant conferred higher levels of protection compared to the nonencapsulated vaccine. These results suggest that S19 DeltavjbR::Kan is safer than S19, induces protection in mice, and should be considered as a vaccine candidate when administered in a sustained-release manner. PMID:19047401

  11. Immunogenicity and Protective Response Induced by Recombinant Plasmids Based on the BAB1_0267 and BAB1_0270 Open Reading Frames of Brucella abortus 2308 in BALB/c Mice

    Science.gov (United States)

    Gómez, Leonardo A.; Alvarez, Francisco I.; Fernández, Pablo A.; Flores, Manuel R.; Molina, Raúl E.; Coloma, Roberto F.; Oñate, Angel A.

    2016-01-01

    Immunogenicity induced by recombinant plasmids based on the BAB1_0267 and BAB1_0270 open reading frames (ORFs) of Brucella abortus 2308 was evaluated. Bioinformatics analyses indicate that the BAB1_0267 and BAB1_0270 ORFs encode a protein with a SH3 domain and a Zn-dependent metalloproteinase, respectively. Both ORFs have important effects on intracellular survival and replication of B. abortus 2308, mediated via professional and non-professional phagocytic cells. Our results show that immunization with the recombinant plasmid based on the BAB1_0267 ORF significantly increases the production of IgG1, levels of IFN-γ and the lymphoproliferative response of splenocytes. However, BAB1_0267 did not provide significant levels of protection. The plasmid based on the BAB1_0270 significantly increased IgG2a production, levels of IFN-γ and TNF-α, and the lymphoproliferative response of splenocytes. These results demonstrate that immunization with the BAB1_0270 derived recombinant plasmid induce a Th1-type immune response, correlated with a heightened resistance to B. abortus 2308 infection in mice. It is concluded that the Th1-type immune response against bacterial Zn-dependent metalloproteinase induces a protective response in mice, and that pV270 recombinant plasmid is an effective candidate microbicide against brucellosis. PMID:27747197

  12. Respuesta serológica y tiempo de saneamiento en rebaños bovinos con brucelosis vacunados con Cepa 19 o Cepa RB-51; Xª Región, Chile Serologic response and time to eradication in herds with brucellosis vaccinated with strain 19 or strain RB-51; 10th Region, Chile

    Directory of Open Access Journals (Sweden)

    M RAMIREZ

    2002-01-01

    Full Text Available Se comparó la respuesta serológica y el tiempo de saneamiento en rebaños bovinos con brucelosis, vacunados con vacuna Cepa 19 o Cepa RB-51. Se estudiaron los registros serológicos de 79 rebaños de la provincia de Valdivia, Xª región de Chile. Los rebaños se habian incorporado al Programa de Erradicacion de Brucelosis Bovina entre 1996 y 1999, y al momento de este estudio se encontraban bajo la condicion de "rebaño saneado". Veintiséis rebaños, con 540 vacas y una seroprevalencia inicial de 14.1%, fueron vacunados con la vacuna Cepa 19 y 53 rebaños, con 1104 vacas y una seroprevalencia inicial de 7.6%, recibieron Cepa RB-51. Periódicamente se colectaron muestras de suero sanguíneo y se examinaron para anticuerpos de Brucella spp. usando las pruebas de Rosa de Bengala y Fijacion de Complemento. Se evaluaron las seroprevalencias, el tiempo de saneamiento y los intervalos de tiempo dentro de éste, el número de exámenes y el lapso de tiempo entre los exámenes. Sesenta y seis de 369, vacas, previamente negativas vacunadas con Cepa 19, seroconvirtieron, pero ninguna de las 917 vacas vacunadas con RB-51 seroconvirtió. El tiempo de saneamiento para los rebaños vacunados con Cepa 19 fluctuó desde 304 a 1025 días (mediana 481 días, y para los rebaños vacunados con Cepa RB-51 desde 140 a 753 días (mediana 401 días; p = 0.003. El tamaño del rebaño, las políticas de reemplazo, el tipo de asistencia veterinaria y la severidad de los signos clínicos de brucelosis no afectaron los lapsos de tiempo. Los rebaños con Cepa 19 fueron muestreados en promedio 4.4 veces y los rebaños con Cepa RB-51 fueron muestreados solo 3.4 veces (p The serologic response to brucellosis vaccination and the time to eradication of brucellosis from herds were compared in dairy cattle vaccinated with either vaccine strains 19 or RB-51. Serologic records from 79 herds from the Province of Valdivia, 10th region of Chile, were evaluated. Herds had been

  13. 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.

  14. TLR2 and TLR4 signaling pathways are required for recombinant Brucella abortus BCSP31-induced cytokine production, functional upregulation of mouse macrophages, and the Th1 immune response in vivo and in vitro.

    Science.gov (United States)

    Li, Jia-Yun; Liu, Yuan; Gao, Xiao-Xue; Gao, Xiang; Cai, Hong

    2014-09-01

    Brucella abortus is a zoonotic Gram-negative pathogen that causes brucelosis in ruminants and humans. Toll-like receptors (TLRs) recognize Brucella abortus and initiate antigen-presenting cell activities that affect both innate and adaptive immunity. In this study, we focused on recombinant Brucella cell-surface protein 31 (rBCSP31) to determine its effects on mouse macrophages. Our results demonstrated that rBCSP31 induced TNF-α, IL-6 and IL-12p40 production, which depended on the activation of mitogen-activated protein kinases (MAPKs) by stimulating the rapid phosphorylation of p38 and JNK and the activation of transcription factor NF-κB in macrophages. In addition, continuous exposure (>24 h) of RAW264.7 cells to rBCSP31 significantly enhanced IFN-γ-induced expression of MHC-II and the ability to present rBCSP31 peptide to CD4(+) T cells. Furthermore, we found that rBCSP31 could interact with both TLR2 and TLR4. The rBCSP31-induced cytokine production by macrophages from TLR2(-/-) and TLR4(-/-) mice was lower than that from C57BL/6 macrophages, and the activation of NF-κB and MAPKs was attenuated in macrophages from TLR2(-/-) and TLR4(-/-) mice. In addition, CD4(+) T cells from C57BL/6 mice immunized with rBCSP31 produced higher levels of IFN-γ and IL-2 compared with CD4(+) T cells from TLR2(-/-) and TLR4(-/-) mice. Macrophages from immunized C57BL/6 mice produced higher levels of IL-12p40 than those from TLR2(-/-) and TLR4(-/-) mice. Furthermore, immunization with rBCSP31 provided better protection in C57BL/6 mice than in TLR2(-/-) and TLR4(-/-) mice after B. abortus 2308 challenge. These results indicate that rBCSP31 is a TLR2 and TLR4 agonist that induces cytokine production, upregulates macrophage function and induces the Th1 immune response.

  15. Maintenance of Brucella abortus free herds; a review with emphasis on the epidemiology and the problems in diagnosing brucellosis in areas of low prevalence

    NARCIS (Netherlands)

    Bercovich, Z.

    1998-01-01

    This review covers some epidemiological aspects that allow Brucella to survive, spread, and maintain itself in the environment. Because the success of maintaining Brucella-free herds is determined by the efficiency of the serological tests to detect a single infected animal the limitations of the tr

  16. Activation of bovine neutrophils by Brucella spp.

    Science.gov (United States)

    Keleher, Lauren L; Skyberg, Jerod A

    2016-09-01

    Brucellosis is a globally important zoonotic infectious disease caused by gram negative bacteria of the genus Brucella. While many species of Brucella exist, Brucella melitensis, Brucella abortus, and Brucella suis are the most common pathogens of humans and livestock. The virulence of Brucella is largely influenced by its ability to evade host factors, including phagocytic killing mechanisms, which are critical for the host response to infection. The aim of this study was to characterize the bovine neutrophil response to virulent Brucella spp. Here, we found that virulent strains of smooth B. abortus, B. melitensis, B. suis, and virulent, rough, strains of Brucella canis possess similar abilities to resist killing by resting, or IFN-γ-activated, bovine neutrophils. Bovine neutrophils responded to infection with a time-dependent oxidative burst that varied little between Brucella spp. Inhibition of TAK1, or SYK kinase blunted the oxidative burst of neutrophils in response to Brucella infection. Interestingly, Brucella spp. did not induce robust death of bovine neutrophils. These results indicate that bovine neutrophils respond similarly to virulent Brucella spp. In addition, virulent Brucella spp., including naturally rough strains of B. canis, have a conserved ability to resist killing by bovine neutrophils. PMID:27436438

  17. Résistance de Brucella abortus isolée de produits alimentaires libanais à base de lait contre les antibiotiques utilisés couramment

    OpenAIRE

    Alwan, Nisreen; Saleh, Imane; Beydoun, Elias; Barbour, Elie; Ghosn, Nada; Harakeh, Steve

    2010-01-01

    International audience Considering the marked importance of Brucella organisms as food-borne pathogens and the lack of published literature on the evaluation of the microbiological quality of dairy-based food products in the Middle East, this study was performed to address this gap. The main aim of the present study was to assess the antimicrobial resistance patterns of Brucella isolates recovered from a total of 164 cultured samples of Lebanese dairy-based food products (Baladi cheese, Sh...

  18. 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.

  19. Evaluation of indirect enzyme-linked immunosorbent assays and IgG avidity assays using a protein A-peroxidase conjugate for serological distinction between Brucella abortus S19-vaccinated and -infected cows.

    Science.gov (United States)

    Pajuaba, Ana C A M; Silva, Deise A O; Mineo, José R

    2010-04-01

    This study aimed to evaluate the use of protein A-peroxidase (horseradish peroxidase [HRPO]) in indirect enzyme-linked immunosorbent assays (iELISAs) and IgG avidity assays for serological distinction between Brucella abortus S19-vaccinated and -infected cows. Four groups were analyzed: GI, 41 nonvaccinated seropositive cows; GII, 79 S19-vaccinated heifers analyzed at 3 months postvaccination; GIII, 105 S19-vaccinated cows analyzed after 24 months of age; and GIV, 278 nonvaccinated seronegative cows. IgG levels and avidity to B. abortus smooth lipopolysaccharide (S-LPS) were determined using anti-bovine IgG-HRPO or protein A-HRPO conjugates. Similar levels of IgG anti-S-LPS were found with GI using both conjugates. Lower IgG levels were detected with GII, GIII, and GIV using protein A-HRPO. Both conjugates showed high performance in discriminating GI from GIII, with high sensitivity (Se; 97.6%) and specificity (Sp; 97.1%). Protein A-HRPO was better in distinguishing GI from GIV (Se, 97.6%; Sp, 94.6%) and GI from GII (Se, 80.5%; Sp, 94.9%). Protein A-HRPO excluded a higher number of positive samples with GII and GIV. IgG avidity showed that protein A-HRPO, but not anti-IgG-HRPO, was able to distinguish nonvaccinated from vaccinated cattle, showing a higher avidity index (AI) with GI than with GII, with 78% of serum samples in GII showing an AI of abortus S-LPS antigen and protein A-HRPO conjugate for preferential detection of the IgG2 subclass was shown to be suitable for serological distinction between S19-vaccinated and -infected cows. Also, antibodies generated after vaccination showed lower avidity, suggesting a role for the IgG2 subclass as an antibody of higher-affinity maturation after infection, constituting an additional tool for differentiating vaccinated from infected cattle. PMID:20147498

  20. Generation and characterization of β1,2-gluco-oligosaccharide probes from Brucella abortus cyclic β-glucan and their recognition by C-type lectins of the immune system

    Science.gov (United States)

    Zhang, Hongtao; Palma, Angelina S; Zhang, Yibing; Childs, Robert A; Liu, Yan; Mitchell, Daniel A; Guidolin, Leticia S; Weigel, Wilfried; Mulloy, Barbara; Ciocchini, Andrés E; Feizi, Ten; Chai, Wengang

    2016-01-01

    The β1,2-glucans produced by bacteria are important in invasion, survival and immunomodulation in infected hosts be they mammals or plants. However, there has been a lack of information on proteins which recognize these molecules. This is partly due to the extremely limited availability of the sequence-defined oligosaccharides and derived probes for use in the study of their interactions. Here we have used the cyclic β1,2-glucan (CβG) of the bacterial pathogen Brucella abortus, after removal of succinyl side chains, to prepare linearized oligosaccharides which were used to generate microarrays. We describe optimized conditions for partial depolymerization of the cyclic glucan by acid hydrolysis and conversion of the β1,2-gluco-oligosaccharides, with degrees of polymerization 2–13, to neoglycolipids for the purpose of generating microarrays. By microarray analyses, we show that the C-type lectin receptor DC-SIGNR, like the closely related DC-SIGN we investigated earlier, binds to the β1,2-gluco-oligosaccharides, as does the soluble immune effector serum mannose-binding protein. Exploratory studies with DC-SIGN are suggestive of the recognition also of the intact CβG by this receptor. These findings open the way to unravelling mechanisms of immunomodulation mediated by β1,2-glucans in mammalian systems. PMID:27053576

  1. Comparison of five serological tests to detect Brucella abortus antibodies and a report on prevalence of the disease in livestock in the state of Yucatan, Mexico

    International Nuclear Information System (INIS)

    One hundred and seventy five sera samples from cattle were tested for reactions to 5 serological tests for Brucella antibodies. The tests used were Rose Bengal (RBT), 2-mercaptoethanol (2ME), rivanol (R), ELISA and complement fixation (CFT). The last of these was used as a standard against which the others were compared. Overall sensitivity and specificity for the 4 other tests by comparison were as follows: RBT, 100%, and 38%; 2ME, 90 and 99%; R, 86% and 100%, ELISA 97% and 99%. These figures varied within sub-groups of vaccinated and non-vaccinated animals. The ELISA test was able to differentiate vaccination and non-vaccination titres. Use of the RBT as a screening test followed by either CFT or ELISA for confirmation would yield the most reliable results. 2323 samples from cattle on 66 farms were tested with RBT as a screening test and 2ME for confirmation. 2.5% were positive for Brucella antigens. 1583 samples from pigs on 61 farms were tested by the same procedure and none was found to be positive for Brucella antibodies. (author). 13 refs, 1 fig., 5 tabs

  2. Prueba de inmunodifusión radial con hapteno nativo para diferenciar bovinos con revacunaciones repetidas con la cepa S19 de Brucella abortus

    OpenAIRE

    Esperanza González Miranda; Laura Hernández Andrade; Efrén Díaz Aparicio

    2006-01-01

    El objetivo del estudio fue establecer la capacidad de la inmunodifusión radial (IDR) con hapteno nativo, para diferenciar anticuerpos post-vacunales de anticuerpos por infección, en bovinos con revacunaciones repetidas de cepa S19. Se trabajó en un establo del estado de México, con presencia de abortos, retención de placenta, expulsiones y fetos momificados. Las becerras se vacunaban con dosis clásica de cepa S19 de B. abortus, y ya adultas revacunaciones anuales repetidas con dosis reducida...

  3. 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.

  4. Cloning and prokaryotic expression of virB8 gene of Brucella abortus%牛布氏菌virB8基因的克隆及原核表达

    Institute of Scientific and Technical Information of China (English)

    张瑞; 王秀然; 夏力亮; 李晓艳; 王兴龙; 王景龙; 冉会玲; 钱晶

    2012-01-01

    Objective To clone the virBS gene of Brucella abortus and express in E. coli . Methods The virBS gene was amplified by PCR from genomic DNA of B. abortus S19 and inserted into pEASY-El vector. The constructed recombinant plasmid pEASY-virB8 was transformed to E. coli BL21 (DE3) for expression under induction of IPTG. The expressed product was purified by HisTrap, FF chromatography and identified by SDS-PAGE and Western blot. Resultus The length of amplified virBS gene was 720 bp. Restriction analysis and sequencing proved that recombinant plasmid pEASY-virB8 was constructed correctly. The expressed recombinant protein,with a relative molecular mass of about 30 000,contained 17. 3% of total somatic protein and reached a purity of 85% after purification. At a protein concentration of 1. 52 mg/ml determined by Lowry method,the expressed product showed specific binding to mouse monoclonal antibody against His. Conclusion The virB8 gene of B. abortus was successfully cloned and expressed in E. coli,which provided an effective candidate antigen for development of novel vaccines.%目的 克隆牛布氏菌virB8基因并在大肠杆菌中进行表达.方法 从牛布氏菌S19基因组DNA中PCR扩增virB8基因片段,插入pEASY-E1载体中,构建重组表达质粒pEASY-virB8,转化E. coli BL.21(DE3),IPTG诱导表达.表达产物经HisTrapTM FF纯化后,进行SDS-PAGE分析和Western blot鉴定.结果 扩增的virB8基因大小为720 bp;重组表达质粒pEASY-virB8经双酶切及测序证实构建正确;表达的重组蛋白相对分子质量约为30000,表达量占菌体总蛋白的17.3%;纯化的重组蛋白纯度为85%,Lowry法测定蛋白浓度为1.52 mg/ml,可与鼠抗His单抗特异性结合.结论 成功克隆了牛布氏菌virB8基因,并在大肠杆菌中表达了重组蛋白,为新型疫苗的研发及布氏菌鉴别诊断方法的建立提供了有效的候选抗原.

  5. 地理信息系统在青海省布鲁杆菌病菌种空间分布中的应用研究%Application of a geographic information system on spatial distribution of Brucella abortus species in Qinghai Province

    Institute of Scientific and Technical Information of China (English)

    秦豫民; 徐立青; 马俊英; 杨旭欣; 田广; 胡桂英; 薛红梅; 杨宁海; 马丽

    2013-01-01

    Objective To establish a geographic information system for Brucella abortus species and study the spatial distribution of Brucella abortus species and strains in Qinghai Province.Methods We got the strains'information from Brucellosis Prevention and Control Department,Qinghai Institute for Endemic Diseases Prevention and Control.Through the geographical information technology software,we established the geographic information system database for Brucella abortus species in Qinghai Province and system fast query function.Time and space distribution of Brucella abortus species and strains was retrieved by the system fast query function and the layer set.Results Through the system fast query function,we got 113 strains' time distribution information.Through the layer set of time and space distribution for spatial distribution,we got 113 strains' and 3 species' spatial distribution information.One hundred and thirteen strains distributed in 21 counties(towns,cities) of Qinghai Province,most in Dulan County (21 strains),Qilian County (16 strains),Dachaidan Town (15 strains),Xinghai County(13 strains) and Gonghe County(10 strains).One hundred and three Brucella melitensis strains distributed in 17 counties(towns,cities) of Qinghai Province,most in Dulan County(20 strains),Qilian County(16 strains),Dachaidan Town (15 strains),Xinghai County (13 strains) and Gonghe County (10 strains); 8 Brucella abortus strains distributed in 6 counties(cities) ; 2 Brucella suis strains distributed in 2 counties.Conclusions We have successfully established a geographic information system for Brucella abortus species and strains in Qinghai Province.Through the layer set for spatial distribution,we could intuitively observe the spatial distribution of Brucella abortus species and strains in Qinghai Province.%目的 建立布鲁杆菌菌种地理信息系统,了解青海省布鲁杆菌病菌株与菌种的空间分布.方法 113株布鲁杆菌菌株资料来源于青海省地方病预防控

  6. The protein moiety of Brucella abortus outer membrane protein 16 is a new bacterial pathogen-associated molecular pattern that activates dendritic cells in vivo, induces a Th1 immune response, and is a promising self-adjuvanting vaccine against systemic and oral acquired brucellosis.

    Science.gov (United States)

    Pasquevich, Karina A; García Samartino, Clara; Coria, Lorena M; Estein, Silvia M; Zwerdling, Astrid; Ibañez, Andrés E; Barrionuevo, Paula; Oliveira, Fernanda Souza de; Carvalho, Natalia Barbosa; Borkowski, Julia; Oliveira, Sergio Costa; Warzecha, Heribert; Giambartolomei, Guillermo H; Cassataro, Juliana

    2010-05-01

    Knowing the inherent stimulatory properties of the lipid moiety of bacterial lipoproteins, we first hypothesized that Brucella abortus outer membrane protein (Omp)16 lipoprotein would be able to elicit a protective immune response without the need of external adjuvants. In this study, we demonstrate that Omp16 administered by the i.p. route confers significant protection against B. abortus infection and that the protective response evoked is independent of the protein lipidation. To date, Omp16 is the first Brucella protein that without the requirement of external adjuvants is able to induce similar protection levels to the control live vaccine S19. Moreover, the protein portion of Omp16 (unlipidated Omp16 [U-Omp16]) elicits a protective response when administered by the oral route. Either systemic or oral immunization with U-Omp16 elicits a Th1-specific response. These abilities of U-Omp16 indicate that it is endowed with self-adjuvanting properties. The adjuvanticity of U-Omp16 could be explained, at least in part, by its capacity to activate dendritic cells in vivo. U-Omp16 is also able to stimulate dendritic cells and macrophages in vitro. The latter property and its ability to induce a protective Th1 immune response against B. abortus infection have been found to be TLR4 dependent. The facts that U-Omp16 is an oral protective Ag and possesses a mucosal self-adjuvanting property led us to develop a plant-made vaccine expressing U-Omp16. Our results indicate that plant-expressed recombinant U-Omp16 is able to confer protective immunity, when given orally, indicating that a plant-based oral vaccine expressing U-Omp16 could be a valuable approach to controlling this disease. PMID:20351187

  7. The Brucella abortus S19 ΔvjbR Live Vaccine Candidate Is Safer than S19 and Confers Protection against Wild-Type Challenge in BALB/c Mice When Delivered in a Sustained-Release Vehicle▿

    OpenAIRE

    Arenas-Gamboa, A. M.; Ficht, T A; Kahl-McDonagh, M. M.; Gomez, G.; Rice-Ficht, A C

    2008-01-01

    Brucellosis is an important zoonotic disease of nearly worldwide distribution. Despite the availability of live vaccine strains for bovine (S19, RB51) and small ruminants (Rev-1), these vaccines have several drawbacks, including residual virulence for animals and humans. Safe and efficacious immunization systems are therefore needed to overcome these disadvantages. A vjbR knockout was generated in the S19 vaccine and investigated for its potential use as an improved vaccine candidate. Vaccina...

  8. Properties of Brucella-phages lytic for non-smooth Brucella strains.

    Science.gov (United States)

    Corbel, M J

    1984-01-01

    A series of host-range mutants has been selected for brucella-phage R. Two of these mutants designated R/O and R/C have been used for typing purposes. Phage R/O is lytic for non-smooth strains of Brucella abortus and for B. ovis. It is genetically unstable however and produces mutants lytic for smooth B. obortus and B. suis. Phage R/C is lytic for non-smooth B. abortus and for B. ovis and B. canis. It is much more stable than phages R or R/O and shows little or no lytic activity on smooth Brucella strains. It has been effective in differentiating B. canis from B. suis in tests on a limited number of strains. In their properties, all of the brucella-phages of the R series resemble their parent phage.

  9. 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.

  10. Brucella-Salmonella lipopolysaccharide chimeras are less permeable to hydrophobic probes and more sensitive to cationic peptides and EDTA than are their native Brucella sp. counterparts.

    OpenAIRE

    Freer, E; Moreno, E.; Moriyon, I. (Ignacio); Pizarro-Cerda, J. (Javier); Weintraub, A; Gorvel, J P

    1996-01-01

    A rough (R) Brucella abortus 45/20 mutant was more sensitive to the bactericidal activity of polymyxin B and lactoferricin B than was its smooth (S) counterpart but considerably more resistant than Salmonella montevideo. The outer membrane (OM) and isolated lipopolysaccharide (LPS) of S. montevideo showed a higher affinity for these cationic peptides than did the corresponding B. abortus OM and LPS. We took advantage of the moderate sensitivity of R B. abortus to cationic peptides to construc...

  11. 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

  12. 西藏职业技术学院实验农场牛布病与结核病的净化措施%Discussion on the Cleaning Measures of Tibet Vocational Technology College Test Farm Brucellosis Brucella Abortus and Tuberculosis

    Institute of Scientific and Technical Information of China (English)

    金红岩; 封家旺

    2011-01-01

    牛布病和结核病分别是由布鲁氏杆菌和结核分枝杆菌引起的人畜共患传染病,严重危害养牛业可持续发展和人民身体健康,危害公共卫生安全.一旦发生,如不能采取有效的净化措施,不仅给畜牧养殖业造成很大损失,而且也严重威胁人体健康.随着奶牛业在全国各地迅速发展,奶牛交易流通的增加,及奶产品在人们饮食结构中的比例越来越大,奶牛的布氏杆菌病、结核病发病率已在悄然上升,饮用未经消毒的牛奶易使人感染布氏杆菌病和牛型结核病[1].%Bovine brocellosis and tuberculosis are the anthropozoonosis caused by bmcellsis leucosis and myeobaeterium tuberculosis which seriously harms the sustainable development of cattle industry and people's health and the public health security. Once happened, if no proper effective cleaning measures are adopted, it will result in the big loss for livestock breeding industry as well as people's health. With the rapid development of dairy industry and the increase in number of cow trade and the the big proportion of milk at people's daily food, the brucellosis brucella abortus and tuberculosis have increased quietly. Drinking the unsterilized milk will make people infect the brucellosis brucella abortus and tuberculosis.

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

    NARCIS (Netherlands)

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

    1996-01-01

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

  14. 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.

  15. Evaluation of Immunogenicity of Divalent DNA Vaccine Encoding Brucella melitensis Omp31 and P39 Genes in Balb/c Mice

    Directory of Open Access Journals (Sweden)

    A Doosti

    2007-10-01

    Full Text Available Introduction & Objective: Brucella is a facultative intracellular pathogen and one of the etiologic agents of brucellosis that can infect humans and domestic animals. Attenuated strains such as B. melitensis Rve1 and B. abortus S19 and Rb51 are being used to control brucellosis in domestic animals. However, no safe and effective vaccine is available for human use. This study was designed to evaluate the immunogenicity and the protective efficacy of a divalent fusion DNA vaccine encoding both the B. melitensis Omp31 protein and P39 protein, designated pCDNA3 recombinant vector. Materials & Methods: This experimental study was performed in Biotechnology Research Center of Islamic Azad University, Shahrekord branch in summer, 1386. Construction of pCDNA3 recombinant vector containing Omp31 and P39 genes of B. melitensis was completed. Then, 12 Balb/c mice were immunized intramuscularly with 100 mg per 50 micro liters of this DNA vaccine. Control mice, 12 Balb/c mice, were simultaneously injected with PBS. During the 1st, 7th, 15th and 30th days the mice received the injections. Afterwards, the ELISA cytokine assay was performed and data were analyzed by SPSS software. Results: Intramuscular injection of the divalent DNA vaccine elicited cellular immune responses in Balb/c mice. The ELISA cytokine assay with serum of vaccinated mice showed high level of IFN-γ and low changes of IL-4 in compare with control mice. Conclusion: Use of divalent genetic vaccine based on the Omp31 and P39 genes can elicit a strong cellular immune response against Brucellosis.

  16. 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...

  17. An Aerosolized Brucella spp. Challenge Model for Laboratory Animals

    Science.gov (United States)

    To characterize the optimal aerosol dosage of Brucella abortus strain 2308 (S2308) and B. melitensis (S16M) in a laboratory animal model of brucellosis, dosages of 10**3 to 10**10 CFU were nebulized to mice. Although tissue weights were minimally influenced, total colony-forming units (CFU) per tis...

  18. Partial Protection against Brucella Infection in Mice by Immunization with Nonpathogenic Alphaproteobacteria▿

    Science.gov (United States)

    Delpino, M. Victoria; Estein, Silvia M.; Fossati, Carlos A.; Baldi, Pablo C.

    2007-01-01

    Previous findings indicate that Brucella antigens and those from nonpathogenic alphaproteobacteria (NPAP) are cross-recognized by the immune system. We hypothesized that immunization with NPAP would protect mice from Brucella infection. Mice were immunized subcutaneously with heat-killed Ochrobactrum anthropi, Sinorhizobium meliloti, Mesorhizobium loti, Agrobacterium tumefaciens, or Brucella melitensis H38 (standard positive control) before intravenous challenge with Brucella abortus 2308. Cross-reacting serum antibodies against Brucella antigens were detected at the moment of challenge in all NPAP-immunized mice. Thirty days after B. abortus challenge, splenic CFU counts were significantly lower in mice immunized with O. anthropi, M. loti, and B. melitensis H38 than in the phosphate-buffered saline controls (protection levels were 0.80, 0.66, and 1.99 log units, respectively). In mice immunized intraperitoneally with cytosoluble extracts from NPAP or Brucella abortus, protection levels were 1.58 for the latter, 0.63 for O. anthropi, and 0.40 for M. loti. To test whether the use of live NPAP would increase protection further, mice were both immunized and challenged by the oral route. Immunization with NPAP induced a significant increase in serum immunoglobulin G (IgG), but not serum or fecal IgA, against Brucella antigens. After challenge, anti-Brucella IgA increased significantly in the sera and feces of mice orally immunized with O. anthropi. For all NPAP, protection levels were higher than those obtained with systemic immunizations but were lower than those obtained by oral immunization with heat-killed B. abortus. These results show that immunization with NPAP, especially O. anthropi, confers partial protection against Brucella challenge. However, such protection is lower than that conferred by immunization with whole Brucella or its cytosoluble fraction. PMID:17715332

  19. Effect of P39 Gene Deletion in Live Brucella Vaccine Strains on Residual Virulence and Protective Activity in Mice

    OpenAIRE

    Tibor, Anne; Jacques, Isabelle; Guilloteau, Laurence; Verger, Jean-Michel; Grayon, Maggy; Wansard, Valerie; Letesson, Jean-Jacques

    1998-01-01

    The 39-kilodalton protein (P39) has previously been shown to be an immunodominant protein in Brucella infections. P39 gene deletion mutants of vaccine strains Brucella abortus S19 and Brucella melitensis Rev.1 were constructed by gene replacement. This deletion did not significantly modify the residual virulence of both vaccine strains in CD-1 mice. CD-1 mice vaccinated with the parent or mutant strains were protected against a virulent challenge. Mutant vaccine strains devoid of P39 could pr...

  20. 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.

  1. 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.

  2. 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.

  3. Construction of mutant Brucella Abortus S19 strain and evaluation of immunization in mice%流产布氏杆菌S19突变株构建及在小鼠感染模型中的免疫保护评估

    Institute of Scientific and Technical Information of China (English)

    郑孝辉; 胡森; 王加兰; 刘林涛; 高红霞; 步志高

    2009-01-01

    通过构建标记疫苗株来解决流产布氏杆菌(B.abortus)鉴别诊断方面的缺陷,本研究以bp26基因作为重组靶住点,S19为亲本,利用bp26基因ORF外侧序列作为同源重组序列,卡那霉素抗性基因(Kanr)为抗性筛选标记,通过双交叉重组筛选获得bp26基因缺失突变的重组S19株,命名为S19-△26.小鼠感染结果表明,突变株S19-△26的残留毒力与亲本株S19相比较没有发生明显改变,康复时间约为15周,突变株S19-△26、亲本株S19和B.abortus强毒株S544接种小鼠后的第3周能检测出"O"抗原的特异性抗体,而第6周开始S19和S544接种小鼠BP26特异性抗体明显升高,S19-△26接种的小鼠一直没检测到BP26特异性抗体.小鼠免疫保护试验显示,脾脏分离CFU数比空白对照要低310g10,S544攻击后脾脏细菌分离数表明突变株具有与亲本疫苗株免疫保护性无明显差异.结果表明,S19-△26免疫能够通过血清学方法与野生型B.abortus感染后的免疫反应相区别,具备作为标记疫苗的潜力.%Brucella abortus strain S19 played an important role in preventing and monitoring brucellosis with high protective rate. S19-△26, a Mutant bp26 gene of B. Abortus S19 strain was constructed by replacing the bp26 gene of attenuated S19 with kanamycin resistence gene. Compared with Si9, mutant S19-△26 possessed the same recover time (15 weeks) and residual virulence in mice, However it did not induce anti-bp26 antibody production in Mice, which would differentiate animals infected with S19-△26 from those infected by S19 and S544. The mutant S19-△26 possesses the same protective ability as S19, which could reduce CFU number of strain S544 infection in mice by 31og10. Altogether, these results indicate that B. Abortus S19-△26 could be a promising vaccine strain in preventing brucellosis.

  4. Protective Properties of Rifampin-Resistant Rough Mutants of Brucella melitensis

    OpenAIRE

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

    2005-01-01

    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, ...

  5. 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.

  6. 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. PMID:26626213

  7. Host Interferon-Gamma Inducible Protein Contributes to Brucella Survival

    Directory of Open Access Journals (Sweden)

    Jennifer eRitchie

    2012-04-01

    Full Text Available Brucella spp. are highly adapted intracellular pathogens of mammals that cause chronic infections while surving and replicating in host monocytes and macrophages. Although monocytes are normally susceptible to infection, pretreatment with pro-inflammatory cytokine interferon- (IFN- activates cellular defense mechanisms that increase intracellular killing of Brucella and prevents bacterial replication. We examined the contribution of the IFN- inducible GTPase, LRG-47, to B. abortus 2308 infection in in vitro and in vivo murine models. Infecting nonactivated macrophages from LRG-47-/- mice revealed that loss of this host protein negatively effected the intracellular survival and replication of IgG opsonized B. abortus. In contrast, survival and replication of non-opsonized B. abortus was the same in both C57/B6 and LRG-47-/- peritoneal macrophages. Following IFN-γ activation of LRG-47-/- monocytes, IgG opsonized B. abortus survived better than non-opsonized bacteria. Similar experiments using macrophages from BALB/c and C57/B6 mice found no difference in intracellular survival or replication between non-opsonized and IgG opsonized B. abortus in either resting or IFN-γ activated monocytes. The differential fate of opsonized and non-opsonized B. abortus was only observed in macrophages collected from LRG-47-/- mice. Given the specific nature of the relationship between this host protein and the mechanism of Brucella internalization, LRG-47-/- mice were infected with B. abortus to assess whether the loss of the lrg47 protein would affect the ability of the bacteria to colonize or persist within the host. B. abortus were able to establish and maintain similar numbers of bacteria in both C57/B6 mice and LRG-47-/- through 3 weeks post intraperitoneal infection. By nine weeks p.i. fewer B. abortus were recovered from LRG-47-/- mice than controls, suggesting that the host protein has a positive role in maintaining long term persistence of the

  8. 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

  9. 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.

  10. Minimal requirements for growth of Brucella suis and other Brucella species.

    Science.gov (United States)

    Plommet, M

    1991-10-01

    Minimal nutritional requirements and temperature limits of growth were studied in Brucella suis and, comparatively, in a few other Brucella species. In a saline basic medium including thiosulphate, ammonium sulphate and glucose with addition of 2 or 4 vitamins (nicotinic acid, thiamin and panthotenic acid, biotin), 24 out of 25 B. suis, 4/6 B. melitensis and 1/6 B. abortus strains were able to grow. Some strains, however, needed to be initially induced to grow by other ingredients, CO2, other vitamins, or amino acids, or by a prolonged incubation. In the saline basic medium without ammonium, glutamic acid and/or alanine and arginine, with or without glucose, supported the growth of all the B. suis and B. melitensis strains, except 2 which required a sulphur amino acid. Five out of 6 B. abortus strains did not grow in either medium without addition of one or several aromatic amino acids or, for one strain, aspartic acid, or valine. One strain could also be induced to grow in ammonium medium by other amino acids. In a rich medium with yeast extract, all Brucella species grew at 18 degrees C and 42.5 degrees (except one) while most B. suis (14/17) grew also at 15 degrees C and 44 degrees C, in contrast to other brucellae of which a few strains only grew at these temperatures. In saline ammonium glucose medium, yeast extract at 0.1 g/l provided all the required vitamins and amino acids for all brucellae and at 1 g/l, it even provided enough nitrogen to support growth without ammonium. Such basic saline medium with yeast extract may be advantageously used in routine Brucella culture, instead of the classic undefined peptone mediums. B. suis biovar 1 strains did not differ significantly in their minimal nutritional requirements, precluding the use of these requirements to differentiate the strains, in particular the Chinese vaccine strain S2 from the reference strain 1330 or from other strains from different parts of the world. Finally, B. suis which is endowed with a

  11. In vivo differences in the virulence, pathogenicity, and induced protective immunity of wboA mutants from genetically different parent Brucella spp.

    Science.gov (United States)

    Wang, Zhen; Niu, Jianrui; Wang, Shuangshan; Lv, Yanli; Wu, Qingmin

    2013-02-01

    To explore the effects of the genetic background on the characteristics of wboA gene deletion rough mutants generated from different parent Brucella sp. strains, we constructed the rough-mutant strains Brucella melitensis 16 M-MB6, B. abortus 2308-SB6, B. abortus S19-RB6, and B. melitensis NI-NB6 and evaluated their survival, pathogenicity, and induced protective immunity in mice and sheep. In mice, the survival times of the four mutants were very different in the virulence assay, from less than 6 weeks for B. abortus S19-RB6 to 11 weeks for B. abortus 2308-SB6 and B. melitensis NI-NB6. However, B. abortus S19-RB6 and B. melitensis 16 M-MB6, with a shorter survival time in mice, offered better protection against challenges with B. abortus 2308 in protection tests than B. abortus 2308-SB6 and B. melitensis NI-NB6. It seems that the induced protective immunity of each mutant might not be associated with its survival time in vivo. In the cross-protection assay, both B. melitensis 16 M-MB6 and B. abortus S19-RB6 induced greater protection against homologous challenges than heterologous challenges. When pregnant sheep were inoculated with B. abortus S19-RB6 and B. melitensis 16 M-MB6, B. abortus S19-RB6 did not induce abortion, whereas B. melitensis 16 M-MB6 did. These results demonstrated the differences in virulence, pathogenicity, and protective immunity in vivo in the wboA deletion mutants from genetically different parent Brucella spp. and also indicated that future rough vaccine strain development could be promising if suitable parent Brucella strains and/or genes were selected. PMID:23239800

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

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    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. Genotyping of Brucella species using clade specific SNPs

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    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.

  14. Development of indirect ELISA for Brucella abortus coating with BP26%牛布鲁氏菌BP26间接ELISA检测方法的建立

    Institute of Scientific and Technical Information of China (English)

    左玉柱; 王增利; 路广计; 王振来; 郑丽丽

    2014-01-01

    为建立检测牛布鲁氏菌(Brucella)血清抗体的间接ELISA方法,本研究将原核表达并纯化的Brucella外膜蛋白BP26作为包被抗原,采用方阵法确定包被抗原和待检血清的最佳工作浓度,经过对各种反应条件进行优化,建立了牛Brucella间接ELISA抗体检测方法.该方法对其他相关的牛病病原无交叉反应,其组内和组间的变异系数均小于10%,具有良好的重复性.对100份临床血清样品进行检测,同时将建立的间接ELISA方法与IDEXX公司的同类试剂盒进行比较,符合率为93%.本研究为布鲁氏菌病提供了一种简便的血清学诊断方法.

  15. 流产布鲁氏菌bp26和omp10基因的同源性分析%Identity analysis of bp26 and omp10 genes of Brucella abortus

    Institute of Scientific and Technical Information of China (English)

    王军; 王瑞; 申之义; 杨莲茹

    2011-01-01

    The aim of the present study was to analyze the identity of genes coding for Brucella outer membrane proteins BP26 and OMP10 among Brucella of testing or reference strains and GenBank strains.Two pairs of primers specific to Brucella bp26 and omp10 genes were designed and synthesized according to GenBank sequences, respectively. bp26 and omp10 genes were amplified by PCR from the genomic DNAs extracted from testing or 2 reference Brucella strains. The PCR products were cloned into the pMD18-T vector and sequenced, respectively. The constructed plasmids were transformed into Escherichia coli DH5a. The extracted plasmid DNA were confirmed by PCR and sequencing. Sequencing results showed that the bp26 gene was 995 bp in length containing a complete 753 bp open reading frame(ORF). Sequence comparison revealed that the identity of bp26 genes in the testing strains was 100% ,and the testing and the reference M5 or S2 strains was 100% or 99.9% ,and the testing strains and the reference S19 or 870 sequences was 99.9% or 100%. The omp10 gene was 531 bp in length containing a complete 396 bp ORF.The identity of omp10 gene among the testing strains was 100%, the testing and the reference M5 or S2 strains were 100%,and the testing and the reference 544 sequence was 99.7%. The identity of bp26 gene and omp10 gene among the various types was more than 99 %,indicating that they were highly conserved.%利用布鲁氏菌外膜蛋白bp26基因和omp10基因,对分离的布鲁氏菌和参考布鲁氏菌菌株进行克隆和序列分析,以明确分离的布鲁氏菌和参考布鲁氏菌菌株与基因库中的布鲁氏菌外膜蛋白bp26基因和omp10基因间的同源性.根据GenBank中的布鲁氏菌bp26基因和omp10基因,分别设计合成了1对特异性引物,以提取的分离株布鲁氏菌和2株参考布鲁氏菌的总DNA为模板,通过PCR技术扩增得到了bp26基因和omp10基因,回收纯化后将这2个基因分别连接到pMD18-T载体上,热激

  16. The Cloning of bp26 and omplO Genes of Brucella Abortus in Huhhot%呼和浩特市区牛种布鲁菌bp26和omp10的克隆

    Institute of Scientific and Technical Information of China (English)

    王军; 王瑞; 杨莲茹; 郭志亮; 于翠玲; 申之义

    2011-01-01

    To clear genes homology on gene of the Brucella outer membrane protein BP26 and OMP10 genes among Brucella of testing or reference and Genbank strains, clone and Comparison for their DNA sequences . We designed and synthesized a specific primers with Brucella bp26 and omplO in Genbank. The genomic DNA of bp26 and omplO of testing or reference Brucella strains was extracted from each sample,PCR amplification was then carried out and thus obtaining bp26 and omplO. The PCR product was inserted into pMD18-T vector and sequenced. The plasmids were transformed into E. coli DH5α. Extracted E. coli plasmid DNA, PCR detected and sequenced. Sequencing showed that bp26 was 995 bp in length,including a 753 bp open reading of the complete framework . Sequence comparison reyealed that homology of bp26 among the testing strains was 100% ,homology of bp26 of the testing and the reference M5 or S2 strains were 100% or 99.9% , homology of bp26 of the testing strains and the reference S19 or 870 sequences was 99.9% or 100%. omplO was 531 bp containing a 396 bp open reading of the complete framework, homology of omplO among the testing strains was 100%. homology of omplO of the testing and the reference M5 or S2 strains were 100%. homology of omplO of the testing and the reference 544 sequence were 99.7%. There are bp26 and omplO gene homology between the various types in more than 99%. They is highly remain stable.%为明确试验布鲁菌和参考布鲁菌菌株与基因库中布鲁菌外膜蛋白BP26和OMP10基因间的同源性.利用布鲁菌外膜蛋白bp26和omp10基因,针对试验布鲁菌和参考布鲁菌菌株进行克隆和序列分析.根据GenBank发表的布鲁菌bp26基因和omp10基因,分别设计合成一对特异性引物,以提取的试验布鲁菌和2株参考布鲁菌的总DNA为模板,通过聚合酶链式反应(PCR)技术扩增得到bp26和omp10基因,回收纯化后将这两个基因分别连接到pMD18-T载体上,热激发

  17. 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.

  18. 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.

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

    Directory of Open Access Journals (Sweden)

    Yongqun eHe

    2012-02-01

    Full Text Available Brucella is a Gram-negative, facultative intracellular bacterium that causes zoonotic brucellosis in humans and various animals. Out of ten 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.

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

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    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.

  1. Respuesta serológica a la vacunación contra brucelosis en bovinos provenientes de un rebaño libre vacunados con dos dosis de vacuna Cepa RB-51 Serological response to brucellosis vaccination in bovines from a free herd vaccinated with two doses of RB-51

    Directory of Open Access Journals (Sweden)

    C Ramírez

    2009-01-01

    Full Text Available Se realizó una encuesta prospectiva con la finalidad de determinar la respuesta serológica a las pruebas oficiales aplicadas en Chile para la detección de brucelosis bovina que son Rosa de Bengala (RB, Fijación de Complemento (FC y Elisa de Competencia (C-Elisa, en hembras adultas vacunadas con dos dosis de vacuna Cepa RB-51 provenientes de un rebaño libre de la enfermedad con certificación vigente otorgada por el Servicio Agrícola y Ganadero (SAG del Ministerio de Agricultura de Chile. Se seleccionaron 100 hembras bovinas mayores de 18 meses con la finalidad de tener una confianza de un 90% de que la proporción de positivos no será mayor a un 2,95% si todos los animales muestreados resultan negativos. Los bovinos fueron vacunados vía subcutánes con vacuna Cepa RB-51 en dosis de 1-3.4 x 10(10 UFC con una diferencia de 30 días entre la primera y segunda aplicación; la fecha de la primera vacunación correspondió al día cero del estudio. Los días 0, 30 y 60 se tomaron muestras de 10 ml de sangre por venopunción coccígea de cada animal seleccionado. Las muestras se procesaron en el Laboratorio de Referencia de Brucelosis Bovina del SAG, Osorno, X Región, Chile. Ningún animal presentó reacción serológica atribuible a la vacunación con RB-51 que puedan interferir con las pruebas diagnósticas actualmente utilizadas para la detección de la brucelosis bovina.A prospective study was carried out in order to determine the serological response to the official tests used in Chile for the detection of bovine brucellosis which are Rose Bengal (RB, Complement Fixation (CF and Competing Elisa (C-ELISA, in adult cows vaccinated with two doses of vaccine strain RB-51 from a brucellosis free herd as certified by the Servicio Agrícola y Ganadero (SAG of the Ministry of Agriculture of Chile. A total of 100 female bovines older than 18 months were selected in order to be 90% confident that the proportion of positives in cows vaccinated twice

  2. Parallel gene loss and acquisition among strains of different Brucella species and biovars.

    Science.gov (United States)

    Zhong, Zhijun; Wang, Yufei; Xu, Jie; Chen, Yanfen; Ke, Yuehua; Zhou, Xiaoyan; Yuan, Xitong; Zhou, Dongsheng; Yang, Yi; Yang, Ruifu; Peng, Guangneng; Jiang, Hai; Yuan, Jing; Song, Hongbin; Cui, Buyun; Huang, Liuyu; Chen, Zeliang

    2012-08-01

    The genus Brucella is divided into six species; of these, B. melitensis and B. abortus are pathogenic to humans, and B. ovis and B. neotomae are nonpathogenic to humans. The definition of gene loss and acquisition is essential for understanding Brucella's ecology, evolutionary history, and host relationships. A DNA microarray containing unique genes of B. melitensis Type strain 16MT and B. abortus 9-941 was constructed and used to determine the gene contents of the representative strains of Brucella. Phylogenetic relationships were inferred from sequences of housekeeping genes. Gene loss and acquisition of different Brucella species were inferred. A total of 214 genes were found to be differentially distributed, and 173 of them were clustered into 15 genomic islands (GIs). Evidence of horizontal gene transfer was observed for 10 GIs. Phylogenetic analysis indicated that the 19 strains formed five clades, and some of the GIs had been lost or acquired independently among the different lineages. The derivation of Brucella lineages is concomitant with the parallel loss or acquisition of GIs, indicating a complex interaction between various Brucella species and hosts. PMID:22923103

  3. [Evaluation of 2 hemoculture media for the isolation of Brucella spp.

    Science.gov (United States)

    Fiorentino, M A; Cipolla, A L; Malena, C R; Paolicchi, F A

    2003-01-01

    The diagnostic efficiency of two hemoculture media for the detection of different species of Brucella strains was evaluated. Strains of Brucella melitensis, Brucella suis, Brucella abortus, Brucella ovis, and Brucella abortus S19 were used. Each strain was diluted in phosphate buffer saline (PBS) to obtain a concentration of 10(5) colony forming units/ml (CFU/ml). Blood from goats, pigs, cattle, and sheep was mixed with the bacterial suspension to obtain a final concentration minor or equal to 10(3) CFU/ml. These blood samples were inoculated into the following media: (i) Hemobrucella (HB), (ii) Tryptose citrated broth 2% (CTB), and (iii) Controls without blood for B. melitensis and B.suis. Subculture in dishes and CFU/ml counts were made at the 1st, 3rd, 8th, 10th, 20th, and 30th post-inoculation (PI) day. Best results were obtained in the HB medium for all strains, except for B. suis, which due to the presence of a contaminant did not reach its maximum development in this medium. All strains were recovered from both media at 24 h PI, except B. ovis that was isolated from HB at 72 h PI and was not recovered from CTB. All strains remained viable for a shorter period in CTB. Under the proposed experimental conditions the HB medium was more sensitive than CTB. Future experiments should evaluate the utility of this commercial medium in clinical cases of animal brucellosis. PMID:14587372

  4. 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 ...

  5. Genome sequences of three live attenuated vaccine strains of Brucella species and implications for pathogenesis and differential diagnosis.

    Science.gov (United States)

    Wang, Yufei; Ke, Yuehua; Wang, Zhoujia; Yuan, Xitong; Qiu, Yefeng; Zhen, Qing; Xu, Jie; Li, Tiefeng; Wang, Dali; Huang, Liuyu; Chen, Zeliang

    2012-11-01

    Live attenuated vaccines play essential roles in the prevention of brucellosis. Here, we report the draft genome sequences of three vaccine strains, Brucella melitensis M5-10, B. suis S2-30, and B. abortus 104M. Primary genome sequence analysis identified mutations, deletions, and insertions which have implications for attenuation and signatures for differential diagnosis. PMID:23045513

  6. Genome Sequences of Three Live Attenuated Vaccine Strains of Brucella Species and Implications for Pathogenesis and Differential Diagnosis

    OpenAIRE

    Wang, Yufei; Ke, Yuehua; Wang, Zhoujia; Yuan, Xitong; Qiu, Yefeng; Zhen, Qing; Xu, Jie; Li, Tiefeng; Wang, Dali; Huang, Liuyu; Chen, Zeliang

    2012-01-01

    Live attenuated vaccines play essential roles in the prevention of brucellosis. Here, we report the draft genome sequences of three vaccine strains, Brucella melitensis M5-10, B. suis S2-30, and B. abortus 104M. Primary genome sequence analysis identified mutations, deletions, and insertions which have implications for attenuation and signatures for differential diagnosis.

  7. In Vivo Differences in the Virulence, Pathogenicity, and Induced Protective Immunity of wboA Mutants from Genetically Different Parent Brucella spp.

    OpenAIRE

    Wang, Zhen; Niu, Jianrui; Wang, Shuangshan; Lv, Yanli; Wu, Qingmin

    2013-01-01

    To explore the effects of the genetic background on the characteristics of wboA gene deletion rough mutants generated from different parent Brucella sp. strains, we constructed the rough-mutant strains Brucella melitensis 16 M-MB6, B. abortus 2308-SB6, B. abortus S19-RB6, and B. melitensis NI-NB6 and evaluated their survival, pathogenicity, and induced protective immunity in mice and sheep. In mice, the survival times of the four mutants were very different in the virulence assay, from less t...

  8. Kinetic study of cytokines production by human peripheral blood mononuclear cells in response to Brucella DNA.

    Science.gov (United States)

    Lashkarbolouki, Taghi; Ardestani, Sussan K; Kariminia, Amina; Ziaee, Abed-Ali; Torkabadi, Ebrahim; Ebrahimi, Mohammad

    2008-01-01

    In spite of reports on cytokines induction by the Brucella DNA in murine model, there is no comparison between pathogenic and appropriate vaccine strains in human. We investigated the cytokines profile of human peripheral blood mononuclear cells (PBMCs) induced by DNA extracted from pathogenic isolates of Brucella melitensis and B. abortus as well as Rev1 and S19; the appropriate vaccine strains. It was observed that despite differential induction of Interleukin(IL)-12 and IL-10 production, identical IL-12/IL-10 concentration ratio was obtained by all Brucella strains DNAs that was 2 after 24 h and 4 after 5 days of incubation. In addition, IL-2 and Interferon(IFN)-gamma production were profoundly increased compared to the medium at day 3 and 5 respectively but IFN-alpha was not induced. Therefore, Brucella strains DNAs are Th1 inducing component with similar pattern in human PBMCs. PMID:17008080

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

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    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.

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

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    MacMillan Alastair P

    2007-04-01

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

  11. Interferon-γ promotes abortion due to Brucella infection in pregnant mice

    OpenAIRE

    Suzuki Hiroshi; Furuoka Hidefumi; Watanabe Kenta; Lee Dong; Kim Suk; Watarai Masahisa

    2005-01-01

    Abstract Background The mechanisms of abortion induced by bacterial infection are largely unknown. In the present study, we investigated abortion induced by Brucella abortus, a causative agent of brucellosis and facultative intracellular pathogen, in a mouse model. Results High rates of abortion were observed for bacterial infection on day 4.5 of gestation, but not for other days. Regardless of whether fetuses were aborted or stayed alive, the transmission of bacteria into the fetus and bacte...

  12. Killing of Brucella antigen-sensitized macrophages by T lymphocytes in bovine brucellosis.

    Science.gov (United States)

    Wyckoff, John H; Potts, Richard D

    2007-12-15

    The present study was an investigation into the role of T lymphocytes in the killing of antigen-sensitized macrophages (M Phi) in bovine brucellosis. Following confirmation of bovine T lymphocyte cell lines derived from Brucella abortus Strain 19 vaccinated steers as antigen-specific in proliferation studies using various antigens, we adapted an apoptosis assay for evaluation of cytotoxicity by these bovine T cells against autologous monocyte-derived macrophages (MDM Phi) as target cells. Various B. abortus antigen preparations were tested including whole gamma-irradiated B. abortus bacteria (gamma BA), a soluble cytosolic protein fraction and a membrane-associated protein fraction. Both polyclonal and cloned T lymphocyte cell lines exhibited cytotoxicity against MDM Phi targets in an antigen-specific fashion. Polyclonal and cloned T lymphocyte cell lines demonstrated cytotoxic responses to varying degrees against B. abortus antigens regardless of whether the antigen used was whole nonviable bacteria, a soluble protein extract or a membrane-associated fraction of extracted bacteria. To further develop correlation of these responses to an in vivo host defense mechanism, cytotoxicity was evaluated using target cells that had been infected with live B. abortus S19 or B. abortus Strain 2308. Cytotoxic responses were also demonstrated consistently against infected targets with either strain of B. abortus although in most cases, cytotoxicity was higher against target cells sensitized with gamma BA compared to those infected with live bacteria. Cloned T lymphocyte cell lines were all CD4+, CD8(-) cells indicating that the observed cytotoxic responses were most likely due to an inflammatory Th1 response and may represent an important host defense mechanism induced by vaccination with live attenuated strains of B. abortus in cattle. PMID:17904229

  13. The attenuation effect of UVc radiation doses in gram-negative bacteria (Brucella, Yersinia, Escherichia coli)

    International Nuclear Information System (INIS)

    The gram-negative bacteria Yersinia enterocolitica sero group O:3 and O:9, and Brucella (Melitensis and abortus) together with Escherichia coli (O:157, DH5α-pEt15b), were investigated to evaluate their susceptibility to UV radiation at 254 nm. If the dose of UVc was 18.7 mW/cm2, the time required for inactivation of Y. enterocolitica and E. coli DH5α-pEt15b and O:157 was 240s and 360s in the dark and light respectively; where if the dose was 19.5 mW/cm2, the time required was 60s in the dark and 120s in light respectively. The time required for inactivation of Brucella strains (melitensis and abortus) if the dose was 18.7 mW/cm2 was 240s in both dark and light, whereas it was 120s(dark) and 240s (light) respectively, when the dose was 19.5 mW/cm2. Using E. coli O:157 as control, it appears that Y. enterocolitica sero group O:3 and O:9 and vaccinal strains of Brucella (Rev. 1 and S19) are more sensitive to UV than wild Brucella strains. No relation was found between the sensitivity of Y. enterocolitica to UV and the presence or absence of a pYV+ virulence plasmid. (author)

  14. 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.

  15. 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.

  16. Proinflammatory Response of Human Osteoblastic Cell Lines and Osteoblast-Monocyte Interaction upon Infection with Brucella spp.▿

    Science.gov (United States)

    Delpino, M. Victoria; Fossati, Carlos A.; Baldi, Pablo C.

    2009-01-01

    The ability of Brucella spp. to infect human osteoblasts and the cytokine response of these cells to infection were investigated in vitro. Brucella abortus, B. suis, B. melitensis, and B. canis were able to infect the SaOS-2 and MG-63 osteoblastic cell lines, and the first three species exhibited intracellular replication. B. abortus internalization was not significantly affected by pretreatment of cells with cytochalasin D but was inhibited up to 92% by colchicine. A virB10 mutant of B. abortus could infect but not replicate within osteoblasts, suggesting a role for the type IV secretion system in intracellular survival. Infected osteoblasts produced low levels of chemokines (interleukin-8 [IL-8] and macrophage chemoattractant protein 1 [MCP-1]) and did not produce proinflammatory cytokines (IL-1β, IL-6, and tumor necrosis factor alpha [TNF-α]). However, osteoblasts stimulated with culture supernatants from Brucella-infected human monocytes (THP-1 cell line) produced chemokines at levels 12-fold (MCP-1) to 17-fold (IL-8) higher than those of infected osteoblasts and also produced IL-6. In the inverse experiment, culture supernatants from Brucella-infected osteoblasts induced the production of IL-8, IL-1β, IL-6, and TNF-α by THP-1 cells. The induction of TNF-α and IL-1β was largely due to granulocyte-macrophage colony-stimulating factor produced by infected osteoblasts, as demonstrated by inhibition with a specific neutralizing antibody. This study shows that Brucella can invade and replicate within human osteoblastic cell lines, which can directly and indirectly mount a proinflammatory response. Both phenomena may have a role in the chronic inflammation and bone and joint destruction observed in osteoarticular brucellosis. PMID:19103778

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

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    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.

  18. Immunoassays for the diagnosis of Brucella suis infection in pigs. Preliminary study

    International Nuclear Information System (INIS)

    Serological procedures to detect antibody against Brucella were developed for diagnosing bovine brucellosis and have been adapted for testing swine sera. The original tube and plate agglutination test and additional, more specific tests such as the buffered plate antigen (BPAT) or 2-mercaptoethanol (ME) utilize Brucella abortus whole cells as antigen. Regardless of the serological test used for swine diagnosis, detection of 80-90% of individual infected swine must be regarded as the best result that can be achieved at present. A primary binding assay such as the enzyme linked immunosorbent assay (ELISA) improved sensitivity and specificity of the measurement of bovine antibody to B. abortus. Limited investigation has been reported about the use of immunoassays for the diagnosis of swine brucellosis. The purpose of this preliminary study is to evaluate the immunoassays developed by K. Nielsen and collaborators for the serological diagnosis of bovine brucellosis using swine sera by comparing test results of BPAT and ME assays now available in diagnostic laboratories. Sera from brucellosis free herds (from Canada) and from swine infected groups (from Argentina) where Brucella suis was isolated were tested

  19. The use of green fluorescent protein as a marker for Brucella vaccines.

    Science.gov (United States)

    Chacón-Díaz, Carlos; Muñoz-Rodríguez, Melissa; Barquero-Calvo, Elías; Guzmán-Verri, Caterina; Chaves-Olarte, Esteban; Grilló, María Jesús; Moreno, Edgardo

    2011-01-10

    Brucellosis is an important malady of productive and wildlife animals and a worldwide zoonosis. The use of effective vaccines and the corresponding diagnostic tests that allow differentiating infected from vaccinated animals are essential tools to control the disease. For this, a prototype of Brucella abortus S19 vaccine expressing green fluorescent protein (S19-GFP) was constructed. The S19-GFP was readily identified under ultraviolet light by macroscopic and microscopic examination and maintained all the biochemical characteristics of the parental S19 vaccine. S19-GFP replicated ex vivo and in vivo, and protected mice against challenge with virulent B. abortus to the same extent as the isogenic S19. An immunoenzymatic assay designed to measure anti-GFP antibodies allowed the discrimination between mice vaccinated with S19-GFP and those immunized with S19. Both vaccines raised antibodies against lipopolysaccharide molecule to similar levels. This experimental model constitutes a "proof of concept" for the use of Brucella-GFP vaccines and associated diagnostic tests to distinguish vaccinated from naturally Brucella infected animals. PMID:21056079

  20. Assessment of Genetic Diversity of Zoonotic Brucella spp. Recovered from Livestock in Egypt Using Multiple Locus VNTR Analysis

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    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.

  1. EPIDEMIOLOGY AND MOLECULAR TYPING OF BRUCELLA STRAINS CIRCULATING IN GEORGIA.

    Science.gov (United States)

    Sidamonidze, K; Ramishvili, M; Kalandadze, I; Tsereteli, D; Nikolich, M P

    2015-10-01

    In 2009-2013, 851 cases of brucellosis were registered in Georgia. Most cases of brucellosis were found in eastern Georgia (91.3% of cases). Mainly men were infected with brucellosis (81.0%).The age group with the most frequent cases of brucellosis is 30-59 years (48.5%). Brucellosis is rarely found among children(0-4 years - 2.0%, 5-14 years - 8.0%). Brucellosis cases were linked to professional activity; mainly by farmers (33.0% of those infected) and shepherds (27.0%). Biotyping Brucella by microbiological methods alone has limitations, so molecular typing was implemented in this study to confirm species. Isolates from human blood and ruminant milk or blood were identified by a bacteriological algorithm and confirmed by real-time PCR (Brucella T1, Idaho Technology). Species identity was confirmed using the AMOS conventional PCR assay, which differentiates four human pathogenic species but cannot recognize certain biovars within them. This gap was addressed by using more universal species-specific Single Nucleotide Polymorphism (SNP) assays. Real-time PCR was used to confirm 86 Brucella strains (48 human, 38 animal isolates) obtained 2009-2011. AMOS PCR supported the biochemical test results for 53 B. melitensis and four B. abortus strains, but not for 29 suspected B. abortus human and animal isolates. SNP typing of all 86 isolates supported the AMOS PCR results but also confirmed the species of the 29 strains not amplified by AMOS PCR. In 2009-2013 years the prevalence of brucellosis was still high. Nowadays cases of brucellosis are higher in the western part of Georgia than in the 1991-2005 period by a factor of 2.62. Brucellosis continues to be mainly an infection in males, because men are mostly engaged in sheep and cattle care. Combined AMOS PCR and SNP typing in this study provided the first genetic confirmation that both B. abortus and B. melitensis are actively circulating in humans and animals in Georgia. PMID:26483376

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

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    Kavoosi G

    2001-07-01

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

  3. 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.

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

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    Joldoshbek Kasymbekov

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

  5. Brucella spp. in equines slaughtered in the south region of Brazil

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    R.F. Santos

    2016-08-01

    Full Text Available ABSTRACT Bacteria of the genus Brucella are widespread in many countries. These microorganisms can infect humans and many wild and domestic animal species. These bacteria have zoonotic potential, and can cause economic and public health problems since they can be transmitted by direct contact with sick animals, through consumption of contaminated milk, raw meat and its derivatives (Soares et al., 2015. Brucellosis is considered a chronic evolving disease, unusual in horses, predominantly caused by Brucella abortus. However, it is not characterized by reproductive disorders in horses, but primarily by abscess in the cervical region, bursa, tendons, and joints. Transmission is likely to occur via ingestion of contaminated water and pastures, especially in areas endemic for bovine brucellosis (Ribeiro et al., 2008. The slaughterhouse is a strategic point for obtaining information about the animal and animal products, edible or not. This study investigated the presence of anti-Brucella spp. immunoglobulins in the serum samples from horses slaughtered in a slaughterhouse in southern Brazil, to estimate the frequency of Brucella spp. antibodies and determine the spatial distribution of the cases.

  6. 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.

  7. The mglA gene and its flanking regions in Brucella: The role of mglA in tolerance to hostile environments, Fe-metabolism and in vivo persistence

    OpenAIRE

    Jacob, Jens; Finke, Antje; Mielke, Martin

    2012-01-01

    We previously demonstrated that a spontaneous smooth small-colony variant of Brucella abortus S19 is characterized by increased in vivo persistence and the differential expression of a gene predicted to encode a galactoside transport ATP binding protein (mglA). In order to further investigate the role of this gene in the context of its flanking regions, we analyzed the respective DNA sequences from the formerly described B. abortus S19 as well as from avirulent B. neotomae 5K33 and compared t...

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

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    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.

  9. 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

  10. 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. PMID:27164438

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

    Science.gov (United States)

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

    2005-01-01

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

  12. 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.

  13. An Oral Vaccine Based on U-Omp19 Induces Protection against B. abortus Mucosal Challenge by Inducing an Adaptive IL-17 Immune Response in Mice

    Science.gov (United States)

    Pasquevich, Karina A.; Ibañez, Andrés E.; Coria, Lorena M.; García Samartino, Clara; Estein, Silvia M.; Zwerdling, Astrid; Barrionuevo, Paula; Oliveira, Fernanda S.; Seither, Christine; Warzecha, Heribert; Oliveira, Sergio C.; Giambartolomei, Guillermo H.; Cassataro, Juliana

    2011-01-01

    As Brucella infections occur mainly through mucosal surfaces, the development of mucosal administered vaccines could be radical for the control of brucellosis. In this work we evaluated the potential of Brucella abortus 19 kDa outer membrane protein (U-Omp19) as an edible subunit vaccine against brucellosis. We investigated the protective immune response elicited against oral B. abortus infection after vaccination of mice with leaves from transgenic plants expressing U-Omp19; or with plant-made or E. coli-made purified U-Omp19. All tested U-Omp19 formulations induced protection against Brucella when orally administered without the need of adjuvants. U-Omp19 also induced protection against a systemic challenge when parenterally administered. This built-in adjuvant ability of U-Omp19 was independent of TLR4 and could be explained at least in part by its capability to activate dendritic cells in vivo. While unadjuvanted U-Omp19 intraperitoneally administered induced a specific Th1 response, following U-Omp19 oral delivery a mixed specific Th1-Th17 response was induced. Depletion of CD4+ T cells in mice orally vaccinated with U-Omp19 resulted in a loss of the elicited protection, indicating that this cell type mediates immune protection. The role of IL-17 against Brucella infection has never been explored. In this study, we determined that if IL-17A was neutralized in vivo during the challenge period, the mucosal U-Omp19 vaccine did not confer mucosal protection. On the contrary, IL-17A neutralization during the infection did not influence at all the subsistence and growth of this bacterium in PBS-immunized mice. All together, our results indicate that an oral unadjuvanted vaccine based on U-Omp19 induces protection against a mucosal challenge with Brucella abortus by inducing an adaptive IL-17 immune response. They also indicate different and important new aspects i) IL-17 does not contribute to reduce the bacterial burden in non vaccinated mice and ii) IL-17 plays a

  14. Interferon-γ promotes abortion due to Brucella infection in pregnant mice

    Directory of Open Access Journals (Sweden)

    Suzuki Hiroshi

    2005-05-01

    Full Text Available Abstract Background The mechanisms of abortion induced by bacterial infection are largely unknown. In the present study, we investigated abortion induced by Brucella abortus, a causative agent of brucellosis and facultative intracellular pathogen, in a mouse model. Results High rates of abortion were observed for bacterial infection on day 4.5 of gestation, but not for other days. Regardless of whether fetuses were aborted or stayed alive, the transmission of bacteria into the fetus and bacterial replication in the placenta were observed. There was a higher degree of bacterial colonization in the placenta than in other organs and many bacteria were detected in trophoblast giant cells in the placenta. Intracellular growth-defective virB4 mutant and attenuated vaccine strain S19 did not induce abortion. In the case of abortion, around day 7.5 of gestation (period of placental development, transient induction of IFN-γ production was observed for infection by the wild type strain, but not by the virB4 mutant and S19. Neutralization of IFN-γ, whose production was induced by infection with B. abortus, served to prevent abortion. Conclusion These results indicate that abortion induced by B. abortus infection is a result of transient IFN-γ production during the period of placental development.

  15. 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.

  16. 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.

  17. [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.

  18. Brucella Endocarditis in Prosthetic Valves

    Science.gov (United States)

    Mehanic, Snjezana; Mulabdic, Velida; Baljic, Rusmir; Hadzovic-Cengic, Meliha; Pinjo, Fikret; Hadziosmanovic, Vesna; Topalovic, Jasna

    2012-01-01

    SUMMARY CONFLICT OF INTEREST: none declared. Introduction Brucella endocarditis (BE) is a rare but severe and potentially lethal manifestation of brucellosis. Pre-existing valves lesions and prosthetic valves (PV) are favorable for BE. Case report We represent the case of a 46-year-old man who was treated at the Clinic for Infectious Diseases, Clinical Center of Sarajevo University, as blood culture positive (Brucella melitensis) mitral and aortic PV endocarditis. He was treated with combined anti-brucella and cardiac therapy. Surgical intervention was postponed due to cardiac instability. Four months later he passed away. Surgery was not performed. PMID:24493988

  19. 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.

  20. 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.

  1. 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

  2. Visual Detection of Brucella spp. in Spiked Bovine Semen Using Loop-Mediated Isothermal Amplification (LAMP) Assay.

    Science.gov (United States)

    Prusty, Bikash R; Chaudhuri, Pallab; Chaturvedi, V K; Saini, Mohini; Mishra, B P; Gupta, Praveen K

    2016-06-01

    Several pathogens including Brucella spp. are shed in semen of infected bulls and can be transmitted to cows through contaminated semen during artificial insemination. The present study reports omp2a and bcsp31 gene based loop-mediated isothermal amplification (LAMP) assays for detection of Brucella genomic DNA in semen from infected bulls. The positive results could be interpreted visually by change in colour of reaction mixture containing hydroxyl naphthol blue (HNB) dye from violet to sky blue. LAMP assays based on omp2a and bcsp31 could detect as little as 10 and 100 fg of B. abortus S19 genomic DNA, respectively. Sensitivity of omp2a and bcsp31 LAMP assays for direct detection of organisms in bovine semen was 2.28 × 10(1) CFU and 2.28 × 10(2) CFU of B. abortus S19 in spiked bovine semen, respectively. The omp2a LAMP assay was found equally sensitive to TaqMan probe based real-time PCR and 100 times more sensitive than conventional PCR in identifying Brucella in spiked semen. The diagnostic applicability of the omp2a LAMP assay was evaluated with seventy-nine bovine semen samples and results were re-evaluated through TaqMan probe based real-time PCR and conventional PCR. Taken together, the omp2a LAMP assay is easy to perform, rapid and sensitive in diagnosis of Brucella spp. in bovine semen. PMID:27570305

  3. A serological diagnostic survey for Brucella canis infection in Turkish patients with Brucellosis-like symptoms.

    Science.gov (United States)

    Sayan, Murat; Erdenlig, Sevil; Stack, Judy; Kilic, Selcuk; Guducuoglu, Huseyin; Aksoy, Yavuz; Baklan, Ayhan; Etiler, Nilay

    2011-01-01

    The incidence of Brucella canis infection in humans is unknown in Turkey. In this study, we investigated the prevalence of B. canis infection in human sera obtained from six regions in Turkey and comparatively evaluated the results obtained by agglutination-based techniques using standardized antigens made from B. canis. The patients (n = 1,746) presented with clinical symptoms that were similar to those of brucellosis. All patients who tested negative in the Rose Bengal test for the smooth Brucella strains (abortus, melitensis, and suis) were screened for evidence of B. canis infection using the rapid slide agglutination test (RSAT), the microagglutination test (MAT), and the 2-mercaptoethanol RSAT test (2ME-RSAT). Of the samples tested, 157 (8.9%), 68 (3.8%), and 66 (3.7%) were positive for B. canis, as determined by RSAT, MAT, and 2ME-RSAT, respectively. The diagnostic sensitivity, specificity, positive predictive value, and negative predictive value of RSAT were 100%, 94.6%, 42%, and 100%, respectively, and of MAT were 100%, 99.9%, 97%, and 100%, respectively. We recommend the routine use of MAT and 2ME-RSAT to check the sera of all patients with symptoms of brucellosis who are negative for brucellosis using a smooth Brucella antigen. PMID:22116333

  4. Laboratory exposure to Brucella melitensis in Denmark

    DEFF Research Database (Denmark)

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

    2013-01-01

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

  5. Chlamydophila abortus em animais de produção Chlamydophila abortus in production animals

    Directory of Open Access Journals (Sweden)

    Francielle Gibson da Silva

    2006-02-01

    Full Text Available A Chlamydophila abortus (anteriormente classificada como Chlamydia psittaci sorotipo 1 tem sido descrita em muitos países, associada principalmente com distúrbios reprodutivos em ovinos, bovinos e caprinos. O aborto enzoótico dos ovinos e caprinos e o aborto epizoótico dos bovinos são as doenças mais importantes causadas por esta bactéria. No Brasil, as pesquisas com C. abortus são praticamente inexistentes. O objetivo desta revisão é apresentar informações sobre modificações taxonômicas, ciclo de vida, epidemiologia, patogenia, sinais clínicos e diagnóstico da infecção por C. abortus principalmente em ovinos, bovinos e caprinos.Chlamydophila abortus (previously known as Chlamydia psittaci serovar 1 has been reported in many countries, associated with reproductive disorders in sheep, cattle, and goats. The enzootic abortion of sheep and goats and the epizootic bovine abortion are the most important diseases produced by this bacterium. In Brazil, there is scarce information about C. abortus. The objective of this review is to show information about taxonomic changes, life cycle, epidemiology, pathogenesis, clinical signs and diagnosis of C. abortus in sheep, cattle and goats.

  6. 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.

  7. DNAs from Brucella Strains Activate Efficiently Murine Immune System with Production of Cytokines, Reactive Oxygen and Nitrogen Species

    Directory of Open Access Journals (Sweden)

    Zahra Tavakoli

    2009-09-01

    Full Text Available Brucellosis is an infectious disease with high impact on innate immune responses which is induced partly by its DNA. In the present study the potential differences of wild type and patients isolates versus attenuated vaccine strains in terms of cytokines, ROS and NO induction on murine splenocytes and peritoneal macrophages were investigated.This panel varied in base composition and included DNA from B. abortus, B. melitensis, B.abortus strain S19 and melitensis strain Rev1, as attenuated live vaccine. Also we included Escherichia coli DNA, calf thymus DNA (a mammalian DNA, as controls. These DNA were evaluated for their ability to stimulate IL-12, TNF-α, IL-10, IFN-γ and ROS production from spleenocytes as well as NO production from peritoneal macrophages. Spleen cells were cultured in 24 well at a concentration of 106 cells/ ml with subsequent addition of 10 μg/ml of Brucella or Ecoli DNAs. These cultures were incubated at 37ºC with 5% CO2 for 5 days. Supernatants were harvested and cytokines, ROS and NOx were evaluated. It was observed that TNF-α was induced in days 1,3,5 by all Brucella strains DNAs and E. coli DNA, IL-10 only was induced in day 1, IFN-γ was induced only in day 5 and IL-12 not induced. ROS and NOx were produced by all strains; however, we observed higher production of NOx which were stimulated by DNA of B. melitensis.

  8. DNAs from Brucella strains activate efficiently murine immune system with production of cytokines, reactive oxygen and nitrogen species.

    Science.gov (United States)

    Tavakoli, Zahra; Ardestani, Sussan K; Lashkarbolouki, Taghi; Kariminia, Amina; Zahraei Salehi, Taghi; Tavassoli, Nasser

    2009-09-01

    Brucellosis is an infectious disease with high impact on innate immune responses which is induced partly by its DNA. In the present study the potential differences of wild type and patients isolates versus attenuated vaccine strains in terms of cytokines, ROS and NO induction on murine splenocytes and peritoneal macrophages were investigated. This panel varied in base composition and included DNA from B. abortus, B. melitensis, B.abortus strain S19 and melitensis strain Rev1, as attenuated live vaccine. Also we included Escherichia coli DNA, calf thymus DNA (a mammalian DNA), as controls. These DNA were evaluated for their ability to stimulate IL-12, TNF-alpha, IL-10, IFN-gamma and ROS production from spleenocytes as well as NO production from peritoneal macrophages. Spleen cells were cultured in 24 well at a concentration of 106 cells/ ml with subsequent addition of 10 microg/ml of Brucella or Ecoli DNAs. These cultures were incubated at 37 degrees C with 5% CO2 for 5 days. Supernatants were harvested and cytokines, ROS and NOx were evaluated. It was observed that TNF-alpha was induced in days 1,3,5 by all Brucella strains DNAs and E. coli DNA, IL-10 only was induced in day 1, IFN- gamma was induced only in day 5 and IL-12 not induced. ROS and NOx were produced by all strains; however, we observed higher production of NOx which were stimulated by DNA of B. melitensis. PMID:20124603

  9. Induction of Immune Response in BALB/c Mice with a DNA Vaccine Encoding Bacterioferritin or P39 of Brucella spp.

    OpenAIRE

    Al-Mariri, Ayman; Tibor, Anne; Mertens, Pascal; De Bolle, Xavier; Michel, Patrick; Godfroid, Jacques; Walravens, Karl; Letesson, Jean-Jacques

    2001-01-01

    In this study, we evaluated the ability of DNA vaccines encoding the bacterioferritin (BFR) or P39 proteins of Brucella spp. to induce cellular and humoral immune responses and to protect BALB/c mice against a challenge with B. abortus 544. We constructed eukaryotic expression vectors called pCIBFR and pCIP39, encoding BFR or P39 antigens, respectively, and we verified that these proteins were produced after transfection of COS-7 cells. PCIBFR or pCIP39 was injected intramuscularly three time...

  10. 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.

  11. 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

  12. Screening Brucella spp. in bovine raw milk by real-time quantitative PCR and conventional methods in a pilot region of vaccination, Edirne, Turkey.

    Science.gov (United States)

    Kaynak-Onurdag, F; Okten, S; Sen, B

    2016-05-01

    Brucellosis is a worldwide zoonotic disease transmitted to humans by consumption of contaminated milk and milk products. Brucellosis is endemic in Turkey, and Edirne has a high Brucella prevalence. Brucellosis is prevented by live-attenuated vaccines for animals and the vaccination program has been in place since 1984 in Turkey. Thrace is the pilot region for this vaccination program. The gold standard diagnostic technique for brucellosis is still the isolation of suspicious bacterial colonies followed by bacteriological identification, but it is very time consuming and laborious. In many studies, Brucella has been investigated by PCR techniques. However, PCR-based methods cannot differentiate between the vaccine strain and the virulent strain; thus, the vaccine strain may interfere with the virulent strain and causes false-positive reactions. To monitor brucellosis control programs effectively, it is important to distinguish vaccine and field strains of Brucella spp. In this study, raw milk samples were collected from 99 cows at 12 different barns in 5 villages of Edirne (Turkey). Bacteriological analyses and real-time quantitative (q)PCR experiments were applied to all samples. The DNA was isolated using Biospeedy DNA-Tricky Purification Kit (Bioeksen, Istanbul, Turkey). For all reactions, Roche Light Cycler Nano (Roche Diagnostics, Mannheim, Germany) instrument and Biospeedy EvaGreen qPCR Pre-Mix (Bioeksen) were used. The data were analyzed using Roche LightCycler NanoSoftware 1.0. For samples that were negative by bacteriological analyses and positive by qPCR, we developed a novel qPCR-based method to differentiate the virulent B. abortus strains and B. abortus S19 vaccine strain. We designed qPCR primers targeting the outer membrane protein of B. abortus. The qPCR products were sequenced using the ABI Prism Big Dye Terminator Cycle Sequencing Ready Reaction Kit on an ABI Prism 377 DNA sequencer (Applied Biosystems, Foster City, CA). In total, 2.02% of the

  13. Immunization with Brucella VirB proteins reduces organ colonization in mice through a Th1-type immune response and elicits a similar immune response in dogs.

    Science.gov (United States)

    Pollak, Cora N; Wanke, María Magdalena; Estein, Silvia M; Delpino, M Victoria; Monachesi, Norma E; Comercio, Elida A; Fossati, Carlos A; Baldi, Pablo C

    2015-03-01

    VirB proteins from Brucella spp. constitute the type IV secretion system, a key virulence factor mediating the intracellular survival of these bacteria. Here, we assessed whether a Th1-type immune response against VirB proteins may protect mice from Brucella infection and whether this response can be induced in the dog, a natural host for Brucella. Splenocytes from mice immunized with VirB7 or VirB9 responded to their respective antigens with significant and specific production of gamma interferon (IFN-γ), whereas interleukin-4 (IL-4) was not detected. Thirty days after an intraperitoneal challenge with live Brucella abortus, the spleen load of bacteria was almost 1 log lower in mice immunized with VirB proteins than in unvaccinated animals. As colonization reduction seemed to correlate with a Th1-type immune response against VirB proteins, we decided to assess whether such a response could be elicited in the dog. Peripheral blood mononuclear cells (PBMCs) from dogs immunized with VirB proteins (three subcutaneous doses in QuilA adjuvant) produced significantly higher levels of IFN-γ than cells from control animals upon in vitro stimulation with VirB proteins. A skin test to assess specific delayed-type hypersensitivity was positive in 4 out of 5 dogs immunized with either VirB7 or VirB9. As both proteins are predicted to locate in the outer membrane of Brucella organisms, the ability of anti-VirB antibodies to mediate complement-dependent bacteriolysis of B. canis was assessed in vitro. Sera from dogs immunized with either VirB7 or VirB9, but not from those receiving phosphate-buffered saline (PBS), produced significant bacteriolysis. These results suggest that VirB-specific responses that reduce organ colonization by Brucella in mice can be also elicited in dogs.

  14. Immunization with Brucella VirB proteins reduces organ colonization in mice through a Th1-type immune response and elicits a similar immune response in dogs.

    Science.gov (United States)

    Pollak, Cora N; Wanke, María Magdalena; Estein, Silvia M; Delpino, M Victoria; Monachesi, Norma E; Comercio, Elida A; Fossati, Carlos A; Baldi, Pablo C

    2015-03-01

    VirB proteins from Brucella spp. constitute the type IV secretion system, a key virulence factor mediating the intracellular survival of these bacteria. Here, we assessed whether a Th1-type immune response against VirB proteins may protect mice from Brucella infection and whether this response can be induced in the dog, a natural host for Brucella. Splenocytes from mice immunized with VirB7 or VirB9 responded to their respective antigens with significant and specific production of gamma interferon (IFN-γ), whereas interleukin-4 (IL-4) was not detected. Thirty days after an intraperitoneal challenge with live Brucella abortus, the spleen load of bacteria was almost 1 log lower in mice immunized with VirB proteins than in unvaccinated animals. As colonization reduction seemed to correlate with a Th1-type immune response against VirB proteins, we decided to assess whether such a response could be elicited in the dog. Peripheral blood mononuclear cells (PBMCs) from dogs immunized with VirB proteins (three subcutaneous doses in QuilA adjuvant) produced significantly higher levels of IFN-γ than cells from control animals upon in vitro stimulation with VirB proteins. A skin test to assess specific delayed-type hypersensitivity was positive in 4 out of 5 dogs immunized with either VirB7 or VirB9. As both proteins are predicted to locate in the outer membrane of Brucella organisms, the ability of anti-VirB antibodies to mediate complement-dependent bacteriolysis of B. canis was assessed in vitro. Sera from dogs immunized with either VirB7 or VirB9, but not from those receiving phosphate-buffered saline (PBS), produced significant bacteriolysis. These results suggest that VirB-specific responses that reduce organ colonization by Brucella in mice can be also elicited in dogs. PMID:25540276

  15. Development of immunochromatographic strip test using fluorescent, micellar silica nanosensors for rapid detection of B. abortus antibodies in milk samples.

    Science.gov (United States)

    Vyas, Swati S; Jadhav, Sushma V; Majee, Sharmila B; Shastri, Jayanthi S; Patravale, Vandana B

    2015-08-15

    Presence of bacteria such as Brucella spp. in dairy products is an immense risk to public health. Point of care immunoassays are rapid in that they can quickly screen various samples in a relatively short amount of time, are sensitive, specific and offer a great advantage in accurate and fast diagnosis of infectious diseases. We have fabricated a point of care rapid diagnostic assay that employs fluorescent, micellar silica nanosensors capable of specifically detecting Brucella IgG antibodies in milk samples of afflicted animals. Currently, point of care detection assays are not commercially available for field testing of farm animals using milk samples. The nanosensing allows precise detection of antibodies with low sample volumes (50 μl). We demonstrate recognition of B. abortus antibodies through capture by fluorescent silica nanosensors using spiked and raw milk samples validated by ELISA and PCR. The test results are accurate and repeatable with high sensitivity and specificity, and a short assay time of 10 min for antigenic recognition and do not require any sample processing procedures such as isolation and separation. Additionally, well defined antigenic components and surface biomarkers of various disease causing microbes can be broadly incorporated within the purview of this technology for accurate and rapid detection of suspected bovine pathological conditions, and can largely enable rapid field testing that can be implemented in farms and food industry. PMID:25829223

  16. The mglA gene and its flanking regions in Brucella: the role of mglA in tolerance to hostile environments, Fe-metabolism and in vivo persistence.

    Science.gov (United States)

    Jacob, Jens; Finke, Antje; Mielke, Martin

    2012-07-01

    We previously demonstrated that a spontaneous smooth small-colony variant of Brucella abortus S19 is characterized by increased in vivo persistence and the differential expression of a gene predicted to encode a galactoside transport ATP binding protein (mglA). In order to further investigate the role of this gene in the context of its flanking regions, we analyzed the respective DNA sequences from the formerly described B. abortus S19 as well as from avirulent B. neotomae 5K33 and compared these with published data from other Brucella species. Deletion mutagenesis of mglA in the large-colony variant of B. abortus S19 resulted in increased tolerance of the deletion mutant to a hyperosmotic (toxic), galactose-containing medium as well as to oxidative stress (H(2)O(2)). Whilst the deletion mutant is characterized by reduced growth on solid Fe(3+)-containing minimal medium (small-colony morphology), in vivo studies in mice demonstrated statistical significant differences in the bacterial load of spleens in the pre-immune, but not in the late phase of the infection. PMID:22534190

  17. 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.

  18. 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

  19. [Construction and characterization of a brucella suis S2 strain with a chloramphenicol resistance marker].

    Science.gov (United States)

    Mao, Kai-Rong; Ding, Jia-Bo; Cheng, Jun-Sheng; Jiang, Yu-Wen; Yao, Wen-Sheng

    2007-12-01

    Vaccination has not been used widely because of the interference in the discrimination between infected and vaccinated animals in immune-screening procedures. In the present study, chloramphenicol resistance gene (Cm(r)) was cloned into the genomic DNA of brucella suis S2 strain by homologous recombination with knocking out the WbkC gene, and obtained the recombinant rS2-WbkC. Further study confirmed that rS2-WbkC was conversed into rough-phenotype form smooth-phenotype. The recombinant keeps the ability to chloramphenicol resistance after 25 passages in tryptic soy agar (TSA). Mice tests showed rS2-WbkC offered similar protection to S2 strain, but more safe than S2. Serum collected form rS2-WbkC immunized mice could be easily distinguished from antiserum produced by smooth-phenotype brucella abortus. In view of these result, rS2-WbkC is a promising candidate for vaccine strain. PMID:18271249

  20. Elisa for the diagnosis and epidemiology of Brucella abortus infection in cattle in Chile

    International Nuclear Information System (INIS)

    A serum bank of 1251 adult cows sera was prepared. The sera originated from animals of three different epidemiological groups: 1) 244 from infected cows, strain 19 vaccinated when calves; 2) 507 from herds free of infection but all cows were strain 19 vaccinated when calves and 3) the last group, 500 sera from cows free of infection and non-vaccinated. All the sera where tested with the routine Rose Bengal (RB) Rivanol (RIV) and Complement Fixation (CF) tests and additionally three enzyme immunoassays were performed. They included two indirect Elisa both using the kit from the Joint FAO/IAEA Division, Vienna, Austria. One assay used a polyclonal conjugated antibody (I-ELISAp) and the other a monoclonal conjugated antibody (I-ELISAm). The third assay was a competitive ELISA (C-ELISA) performed with sLPS, plus monoclonal antibody, M84, and goat anti-mouse antibody-HRPO. Using the CFT as 'gold standard' the sensitivities of all the methods were: RB 87.1%, RIV 87.1%, I-ELISAp 100% I-ELISAm 100%. The calculated specificity was: RB 100%, RIV 100%, I-ELISAp 96.4% and I-ELISAm 100%. In the group of infected animals (244) the following results were obtained: RB 13.5%, RIV 11.9%, CF 12.7%, I-ELISAp 50.8% and I-ELISAm 22.9%. Results for the non-vaccinated group were: RB 0.2%, RIV 0%, CFT 0.2%, I-ELISAp 6.9% and I-ELISAm 2.9%. The C-ELISA was performed on samples from the positive group or with positivity values close to the cut-off value in the I-ELISAm. In the infected group 28 out of 63 animals were detected as infected and from the non-vaccinated herds none of 15 I-ELISAm positive samples were detected as infected in the C-ELISA. (author)

  1. Prevalence of brucella abortus antibodies in bovine serum from gusau modern abattoir, Zamfara state, Nigeria

    Directory of Open Access Journals (Sweden)

    N. Lawal

    2012-10-01

    Full Text Available A study for bovine brucellosis was conducted using serology to determine the status of the disease in slaughtered cattle. Three hundred and twenty (320 sera were collected from Gusau Modern Abattoir, Zamfara State. The sera were tested using Enzyme Linked Immunosorbent Assay obtained from Veterinary Laboratory Agencies, Weybridge, UK. An overall prevalence of 20% (64 positive was obtained with sex prevalence for males and females being 10.62% (34 positive and 9.37% (30 positive respectively out of 180 males and 140 females tested without significant association (P 24 months while; lower prevalence of 3.13% was recorded in age group

  2. Whole genome sequence comparison of ten diagnostic brucellaphages propagated on two Brucella abortus hosts

    OpenAIRE

    Tevdoradze, Ekaterine; Farlow, Jason; Kotorashvili, Adam; Skhirtladze, Natia; Antadze, Irina; Gunia, Sophio; Balarjishvili, Nana; Kvachadze, Leila; Kutateladze, Mzia

    2015-01-01

    Background Recently the genome sequences of two brucellaphages, isolated in Georgia (Tb) and Mexico (Pr) were reported revealing pronounced sequence homogeneity and the presence of two major indels discriminating the two phages. Subsequent genome sequencing of six diagnostic brucellaphages: Tbilisi (Tb), Firenze (Fz), Weybridge (Wb), S708, Berkeley (Bk) and R/C phages identified three major genetic groups. However, the propensity for fine-scale genetic variability of diverse brucellaphages gr...

  3. 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。

  4. 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

  5. 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.

  6. Brucella endocarditis on double valvular prosthesis.

    OpenAIRE

    Lezaun, R; Teruel, J.; Maître, M. J.; De Artaza, M

    1980-01-01

    The case is reported of a 48-year-old man suffering from Brucella endocarditis on a double prosthesis. The successful medical and surgical treatment is described. So far as the authors know, this is the first report of Brucella endocarditis from a heart valve prosthesis.

  7. Whole genome sequences of four Brucella strains.

    Science.gov (United States)

    Ding, Jiabo; Pan, Yuanlong; Jiang, Hai; Cheng, Junsheng; Liu, Taotao; Qin, Nan; Yang, Yi; Cui, Buyun; Chen, Chen; Liu, Cuihua; Mao, Kairong; Zhu, Baoli

    2011-07-01

    Brucella melitensis and Brucella suis are intracellular pathogens of livestock and humans. Here we report four genome sequences, those of the virulent strain B. melitensis M28-12 and vaccine strains B. melitensis M5 and M111 and B. suis S2, which show different virulences and pathogenicities, which will help to design a more effective brucellosis vaccine. PMID:21602346

  8. 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.

  9. 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.

  10. 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

  11. [Detection of anti-Brucella spp. antibodies in swine by agglutination techniques and indirect ELISA in the Buenos Aires and La Pampa provinces, Argentina].

    Science.gov (United States)

    Castro, H A; González, S R; Prat, M I; Baldi, P C

    2006-01-01

    Porcine brucellosis is one of the most important zoonoses in this country. Currently, there is no control program for porcine brucellosis in Argentina and the epidemiological situation is still unknown. The purpose of our study was to detect anti-Brucella spp. antibodies in swine in the southwest of the Buenos Aires province and the east of the La Pampa province. Blood samples were obtained when animals were slaughtered. The presence of anti-brucella antibodies was studied by the buffered plate agglutination test (BPA), the tube agglutination test (SAT), the 2-mercaptoethanol (2-ME) agglutination test and indirect ELISA tests, using the cytosolic fraction from Brucella abortus S19 (CYT), and lipopolysaccharide (LPS)-free cytosolic proteins (CP). Out of a total of 325 samples analyzed, 17.8% reacted positively to BPA, 13.8% to SAT, 8.0% to 2-ME, 21.0% to ELISA-CYT and 10.0% to ELISA-CP. These results agree with the few data available in our country and suggest that brucellosis screening should be extended to other regions. PMID:17037254

  12. 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

  13. BBP: Brucella genome annotation with literature mining and curation

    Directory of Open Access Journals (Sweden)

    He Yongqun

    2006-07-01

    Full Text Available Abstract Background Brucella species are Gram-negative, facultative intracellular bacteria that cause brucellosis in humans and animals. Sequences of four Brucella genomes have been published, and various Brucella gene and genome data and analysis resources exist. A web gateway to integrate these resources will greatly facilitate Brucella research. Brucella genome data in current databases is largely derived from computational analysis without experimental validation typically found in peer-reviewed publications. It is partially due to the lack of a literature mining and curation system able to efficiently incorporate the large amount of literature data into genome annotation. It is further hypothesized that literature-based Brucella gene annotation would increase understanding of complicated Brucella pathogenesis mechanisms. Results The Brucella Bioinformatics Portal (BBP is developed to integrate existing Brucella genome data and analysis tools with literature mining and curation. The BBP InterBru database and Brucella Genome Browser allow users to search and analyze genes of 4 currently available Brucella genomes and link to more than 20 existing databases and analysis programs. Brucella literature publications in PubMed are extracted and can be searched by a TextPresso-powered natural language processing method, a MeSH browser, a keywords search, and an automatic literature update service. To efficiently annotate Brucella genes using the large amount of literature publications, a literature mining and curation system coined Limix is developed to integrate computational literature mining methods with a PubSearch-powered manual curation and management system. The Limix system is used to quickly find and confirm 107 Brucella gene mutations including 75 genes shown to be essential for Brucella virulence. The 75 genes are further clustered using COG. In addition, 62 Brucella genetic interactions are extracted from literature publications. These

  14. 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.

  15. 9 CFR 113.32 - Detection of Brucella contamination.

    Science.gov (United States)

    2010-01-01

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

  16. 21 CFR 866.3085 - Brucella spp. serological reagents.

    Science.gov (United States)

    2010-04-01

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

  17. Human brucellosis in northwest Ecuador: typifying Brucella spp., seroprevalence, and associated risk factors.

    Science.gov (United States)

    Ron-Román, Jorge; Ron-Garrido, Lenin; Abatih, Emmanuel; Celi-Erazo, Maritza; Vizcaíno-Ordóñez, Laura; Calva-Pacheco, Jaime; González-Andrade, Pablo; Berkvens, Dirk; Benítez-Ortíz, Washington; Brandt, Jef; Fretin, David; Saegerman, Claude

    2014-02-01

    Human brucellosis in Ecuador is underreported and based only on passive surveillance. Since 2008, brucellosis was removed from the list of communicable diseases in the country. Until now, the true human brucellosis picture has not yet been determined. The aim of this study was to determine the seroprevalence of the disease, identify risk factors associated with brucellosis seropositivity in humans, and isolate circulating strains of Brucella spp. in the northwestern part of Ecuador. Between 2006 and 2008, a large transect survey was conducted, based on blood sampling of people from the northwestern part of Ecuador (n=3733) together with an epidemiological inquiry. On the basis of three diagnostic tests used in parallel, the overall seroprevalence was estimated as 1.88% (95% confidence interval [CI] 1.48-2.38). Based on a multivariable random effects logistic regression analysis, the main risk factors associated with human brucellosis seropositivity were contact with livestock (odds ratio [OR]=3.0; CI 1.25-7.08), consumption of fetus and placenta (OR=2.5; CI 1.18-5.22), and involvement in activities at risk for brucellosis infection (OR=1.8; CI 1.00-3.35). Noticeable variation in brucellosis seropositivity among humans within cantons was observed. The circulating strain was Brucella abortus biotype 4. This study emphasized that contact with livestock, consumption of fetus and placenta, and occupational hazard group were all significant risk factors for the transmission of brucellosis among individuals in the northwestern part of Ecuador. Alongside encouraging the launching of educational campaigns against brucellosis, especially in rural areas where 36% of the population lives, controlling this zoonotic disease in animals will directly benefit its prevention in humans, especially because there is no safe and efficacious vaccine against brucellosis in humans.

  18. 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.

  19. 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.

  20. 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.

  1. Detection of Chlamydophila abortus in Sheep (Ovis aries in Mexico

    Directory of Open Access Journals (Sweden)

    Juan M. Jiménez-Estrada

    2008-01-01

    Full Text Available Chlamydophila abortus is one of the pathogens which induce abortion in small ruminants; this pathogen has a tropism for ruminant placenta and causes the disease commonly referred to as Ovine Enzootic Abortion (OEA. In Europe are estimated economic losses of around 20 million pounds a year by OEA. In the American Continent the disease has been reported only in Canada, the United States, Colombia and Chile while in Mexico it is unknown whether OEA is common and it is causing abortions in flocks of sheep from “Estado de Mexico”. The objective of this study was investigating the prevalence of anti-Chlamydophila abortus IgG antibodies and detection of C. abortus DNA in sheep with clinical abort history by mean of ELISA assay (C. abortus ELISA, Institute Pourquier, Montpellier, France and molecular identification of the principal outer membrane protein (POMP 90-91B gene by PCR, respectively. A cross-sectional study was carried out to enroll and random sample of ewes from november 2003 until march 2005. A total of 349 sera and vaginal swabs samples were collected from 35 flocks of sheep from Xalatlaco. The results showed that the seropositive rate was 31.1% (14/45 for healthy and 21.3% (65/304 for sheep with history clinical of abort. In vaginal swabs, the PCR showed 0% (0/45 for healthy animals and 0.65% (2/304 for aborted sheep. Samples from the lungs and liver of the fetus of one of these animals were also positive for C. abortus. In conclusion, these results confirmed that infection with C. abortus is common and is affecting sheep flocks in the Mexican highlands. Therefore, is necessary that the authorities responsible for animal welfare in Mexico (SAGARPA to set up appropriate epidemiological surveillance and control programs to eradicate this disease.

  2. 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.

  3. Can Chlamydia abortus be transmitted by embryo transfer in goats?

    Science.gov (United States)

    Oseikria, M; Pellerin, J L; Rodolakis, A; Vorimore, F; Laroucau, K; Bruyas, J F; Roux, C; Michaud, S; Larrat, M; Fieni, F

    2016-10-01

    The objectives of this study were to determine (i) whether Chlamydia abortus would adhere to or penetrate the intact zona pellucida (ZP-intact) of early in vivo-derived caprine embryos, after in vitro infection; and (ii) the efficacy of the International Embryo Transfer Society (IETS) washing protocol for bovine embryos. Fifty-two ZP-intact embryos (8-16 cells), obtained from 14 donors were used in this experiment. The embryos were randomly divided into 12 batches. Nine batches (ZP-intact) of five embryos were incubated in a medium containing 4 × 10(7)Chlamydia/mL of AB7 strain. After incubation for 18 hours at 37 °C in an atmosphere of 5% CO2, the embryos were washed in batches in 10 successive baths of a phosphate buffer saline and 5% fetal calf serum solution in accordance with IETS guidelines. In parallel, three batches of ZP-intact embryos were used as controls by being subjected to similar procedures but without exposure to C. abortus. The 10 wash baths were collected separately and centrifuged for 1 hour at 13,000 × g. The washed embryos and the pellets of the 10 centrifuged wash baths were frozen at -20 °C before examination for evidence of C. abortus using polymerase chain reaction. C. abortus DNA was found in all of the infected batches of ZP-intact embryos (9/9) after 10 successive washes. It was also detected in the 10th wash fluid for seven batches of embryos, whereas for the two other batches, the last positive wash bath was the eighth and the ninth, respectively. In contrast, none of the embryos or their washing fluids in the control batches were DNA positive. These results report that C. abortus adheres to and/or penetrates the ZP of in vivo caprine embryos after in vitro infection, and that the standard washing protocol recommended by the IETS for bovine embryos, failed to remove it. The persistence of these bacteria after washing makes the embryo a potential means of transmission of the bacterium during embryo transfer from

  4. Detection of Brucella by loop-mediated isothermal amplification%布鲁杆菌环介导等温扩增快速检测方法的建立

    Institute of Scientific and Technical Information of China (English)

    刘锴

    2013-01-01

    Objective A loop-mediated isothermal amplification (LAMP)assay was developed for rapid detection of Brucella.Methods Using Primer 4.0,we designed four specific primers at brookings coli outer membrane protein (OMP31) gene conservative area,under the action of Bst polymerase larger pieces to realize the step-like isothermal amplification of DNA.In this method,on the basis of optimal amplification conditions,the specificity and sensitivity of this method was compared with conventional PCR,and LAMP visualization experiment was conducted.Results The detection results of Brucella abortus (B.abortus) strain 544,B.abortus strain 104M,Brucella melitensis (B.melitensis) strain Rev-1,B.melitensis strain 16M,Brucella suis (B.suis) strain S2,B.suis strainand 1330S,Brucella canis (B.canis) strain RM6/66,Brucellá ovis (B.ovis) strain 63/290,and Brucella neotomae.(B.neotomae) strain 5K33 were positive,and Yersinia enterocolitica (Y.enterocolitica) O ∶ 9,Escherichia coli (E.coli) O157 ∶H7,and (Salmonella typhimurium,S.typhimurium) 47729 were negative.The minimum detection limit was 8.5 × 10-8 mg/L.LAMP was more sensitive than PCR.The results can be determined by electrophoresis or through visual judgment.Conclusions LAMP is a simple,specific and sensitive method.LAMP assay is a useful tool for rapid detection of Brucella.%目的 建立一种环介导等温扩增(LAMP)快速检测动物布鲁杆菌的方法.方法 使用Primer 4.0,针对布鲁杆菌外膜蛋白(OMP31)基因保守区设计4条特异引物,在Bst大片段聚合酶的作用下,实现DNA的梯状等温扩增,并在此方法最适扩增条件优化的基础上,对其特异性、灵敏度进行实验,同时与普通PCR灵敏度进行比较,以及进行LAMP可视化实验.结果 本研究建立的检测方法对牛种布鲁杆菌(Brucella abortus,B.abortus)544A和104M、羊种布鲁杆菌(Brucella melitensis,B.melitensis)Rev-1和16M、猪种布鲁杆菌(Brucella suis,B.suis)S2和1330S

  5. Optimization of pulsed-field gel electrophoresis for molecular typing of Brucella using Xba Ⅰ%布鲁杆菌XbaⅠ酶切脉冲场凝胶电泳基因分型方法的条件优化

    Institute of Scientific and Technical Information of China (English)

    赵鸿雁; 田国忠; 王衡; 姜海; 李兰玉; 杜小莉; 崔晶花; 朴东日

    2013-01-01

    Objective To optimize the protocol of pulsed-field gel electrophoresis(PFGE) to evaluate the applicability for molecular typing of Brucella using Xba].Methods Factors that influence PFGE including selecting restriction endonuclease from different manufacturers and pulse time were optimized.The bands were marked with their molecular weight and then analyzed with BioNumerics (Version 4.0,Applied Maths BVBA,Belgium) software.Results The Xba I from Promega Ltd was selected due to its better digestion effect.The best pulsed-field gel electrophoresis was performed in two phases for 21 h which included 16 h for pulse time of 0.5-8.0 s and 5 h for pulse time of 10.0-56.0 s; the angle of 120°,gradient of 6.0 V/cm,temperature of 14 ℃.The 6 Brucella species and 19 reference biovar strains were tested by optimized conditions of PFGE using Xba Ⅰ.The strains from four Brucella species which included Brucella abortus,Brucella melitensis,Brucella suis and Brucella ovis were clustered into four groups.But Brucella canis was classified in Brucella suis group.The Brucella neotomae was classified in Brucella abortus group.The results of clustering analysis by PFGE were similar to that of biotyping methods.Conclusions PFGE genotyping method using Xba I has good discriminatory power for molecular typing for Brucella.The optimized PFGE protocol is worthy of application.%目的 优化布鲁杆菌XbaⅠ酶切脉冲场凝胶电泳基因分型方法.方法 通过测试不同厂家生产的Xba I限制性内切酶,优化脉冲场凝胶电泳条件和脉冲时间,所得凝胶电泳图谱用BioNumerics V 4.0软件和UPGMA方法进行聚类分析,获得脉冲场凝胶电泳分型结果.结果 优化后的条件为:XbaI限制性内切酶为美国Promega公司产品;脉冲时间为21 h,包括第一脉冲时间0.5~8.0 s,16h,第二脉冲时间10.0~56.0 s,5h;电压为6.0 V/cm;电场夹角120°;电泳温度为14℃.利用优化后的脉冲场凝胶电泳条件对布鲁杆菌6个菌种19

  6. Erythritol triggers expression of virulence traits in Brucella melitensis

    OpenAIRE

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

    2013-01-01

    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 B. 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. Immu...

  7. Generation of the Brucella melitensis ORFeome Version 1.1

    OpenAIRE

    Dricot, A. (Amélie); Rual, J. J. (Jean François); Lamesch, P; BERTIN, N.; Dupuy, D. (Denis); Hao, T.; Lambert, C; Hallez, R.; Delroisse, J.M. (Jean Marc); Vandenhaute, J; Lopez-Goñi, I. (Ignacio); Moriyon, I. (Ignacio); Garcia-Lobo, J M; Sangari, F.J. (Félix Javier); MacMillan, A.P.

    2004-01-01

    The bacteria of the Brucella genus are responsible for a worldwide zoonosis called brucellosis. They belong to the alpha-proteobacteria group, as many other bacteria that live in close association with a eukaryotic host. Importantly, the Brucellae are mainly intracellular pathogens, and the molecular mechanisms of their virulence are still poorly understood. Using the complete genome sequence of Brucella melitensis, we generated a database of protein-coding open reading frames (ORFs) and cons...

  8. 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.

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

    OpenAIRE

    Osman, Amira E. F.; Abdullahi N Hassan; Ali E Ali; Abdoel, Theresia H.; Smits, Henk L.

    2015-01-01

    Background 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. Methods 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 cu...

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

    Directory of Open Access Journals (Sweden)

    Jacques Isabelle

    2009-07-01

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

  11. 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.

  12. 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.

  13. 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

  14. 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

  15. Seroprevalence of Chlamydia abortus in camel in the western region of Libya

    Directory of Open Access Journals (Sweden)

    Rabia Elzlitne

    2016-06-01

    Conclusion: The present findings signify the C. abortus as a potential agent to cause abortion in Libyan camel (C. dromedarius. Besides, the persons who handle camels in Libya are at risk of infecting with C. abortus. [J Adv Vet Anim Res 2016; 3(2.000: 178-183

  16. 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...

  17. Whole Genome Sequences of Four Brucella Strains ▿

    OpenAIRE

    Ding, Jiabo; Pan, Yuanlong; Jiang, Hai; Cheng, Junsheng; Liu, Taotao; Qin, Nan; Yi YANG; Cui, Buyun; Chen, Chen; Liu, Cuihua; Mao, Kairong; Zhu, Baoli

    2011-01-01

    Brucella melitensis and Brucella suis are intracellular pathogens of livestock and humans. Here we report four genome sequences, those of the virulent strain B. melitensis M28-12 and vaccine strains B. melitensis M5 and M111 and B. suis S2, which show different virulences and pathogenicities, which will help to design a more effective brucellosis vaccine.

  18. Use of an indirect enzyme linked immunosorbent assay for the diagnosis and epidemiological studies of Brucella abortus in Chile

    International Nuclear Information System (INIS)

    It is well known that traditional brucellosis diagnostic tests have a lack of sensitivity and specificity depending on the area or country where they have been used. Seeking for better alternatives, we undertook an evaluation of an ELISA test for bovine brucellosis diagnosis, for its possible use in Chile. We collected 2 groups of sera from negative animals: one from an area free of S19 vaccination (n=491) and a second from vaccinated herds (n=349). These groups were used for determination of cut-offs and diagnostic specificity calculations. A positive group of sera was collected from animals (n=186) from infected herds which were all reactors in the Rose Bengal (RB) and Rivanol (RIV) tests; some were also bacteriological culture positive and positive to an immunodiffusion test. This group was used for sensitivity calculation. A final test group of 385 sera from herds with brucellosis, either negative or positive in the RB or RIV tests, were used for relative sensitivity and specificity calculations. The calculated cut-off values were 26% and 61% of Percentage Positivity (PP) for the free area and vaccinated negative groups, respectively. Real sensitivity was 100% and real specificity was 98.6% when a 26% cut-off was used. Using the 61% cut-off value, the relative sensitivity respect of the RB and RIV test were 95.7% and 100%, respectively. In addition, relative specificity with respect to the RB and RIV tests were 82.4% and 81.8%, respectively. (author)

  19. Infection Outcome and Cytokine Gene Expression in Brugia pahangi- Infected Gerbils (Meriones unguiculatus) Sensitized with Brucella abortus

    OpenAIRE

    Chirgwin, Sharon R.; Elzer, Philip H.; Coleman, Sharon U.; Nowling, Jena M.; Hagius, Sue D.; Edmonds, Matthew D.; Klei, Thomas R

    2002-01-01

    Filarial infections have been associated with the development of a strongly polarized Th2 host immune response and a severe impairment of mitogen-driven proliferation and type 1 cytokine production in mice and humans. The role of this polarization in the development of the broad spectra of clinical manifestations of lymphatic filariasis is still unknown. Recently, data gathered from humans as well as from immunocompromised mouse models suggest that filariasis elicits a complex host immune res...

  20. 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

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

    Science.gov (United States)

    McCready, Paula M.; Radnedge, Lyndsay; Andersen, Gary L.; Ott, Linda L.; Slezak, Thomas R.; Kuczmarski, Thomas A.

    2009-02-24

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

  2. Efficacy of Brucella suis strain 2 vaccine against Brucella ovis in rams.

    Science.gov (United States)

    Blasco, J M; Marín, C; Jiménez de Bagüés, M P; Barberán, M

    1993-10-01

    The protective efficacy against Brucella ovis of live vaccine Brucella suis strain 2 (S2) and Brucella melitensis strain Rev 1 has been evaluated in rams. Fourteen 4-month-old Brucella-free Aragonesa rams were vaccinated conjunctivally with 2 x 10(9) c.f.u. S2. Sixteen rams of the same breed, condition and age were conjunctivally vaccinated the same day with 1.6 x 10(9) Rev 1. Thirteen rams were unvaccinated controls. Eight months after vaccination all rams were challenged with 6 x 10(9) c.f.u. B. ovis and slaughtered 2 months thereafter for bacteriological and pathological studies. The percentage of infection in the group vaccinated with Rev 1 (43.7%) was significantly lower (p S2-vaccinated animals (78.6%) and unvaccinated controls (84.6%). No significant differences were found when comparing the percentages of infection corresponding to S2-vaccinated and control groups. The degree of infection (percentage of necropsy samples infected) was significantly lower in Rev 1-vaccinated (13%) than in S2-vaccinated (36.9%) or control groups (47.4%) (p S2-vaccinated and control groups. PMID:8296481

  3. 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.

  4. Two Unusual Presentations of Childhood Brucella Cases

    Directory of Open Access Journals (Sweden)

    Tolga Altuğ Şen

    2010-05-01

    Full Text Available Introduction: Brucellozis is still a common infectious disease in our country and sometimes it may be presented with uncommon clinical manifestations.Case 1: A ten years old male was presented to our clinic with complaints of malaise, weight loss, petechia, and bleeding of gums. On physical examination cervical lymphadenopathy and hepatosplenomegaly had been detected and in complete blood count pancytopenia was found.admitted to our clinic. In bone marrow aspiration hypocellular bone marrow was seen. His Brucella agglutination test was positive at 1:1280 titer and the blood culture was positive for Brucella mellitensis. The pancytopenia was resolved after the antibiotherapy. Case 2: A nine-year-old female was referred to our clinic with tachycardia, who had the cardiac rate of 136/min. The electrocardiography showed sinusal tachycardia and echocardiography was normal, no endocarditis or pericarditis was present. She had complaints of fatigue and lassitude for the last month. Her brucella agglutination test was positive at 1:1280 titer and blood culture was negative. After antibiotherapy her symptoms regressed, cardiac rate decreased to 80-100/min. Isolated tachycardia may be the early manifestaion of brucellosis in children which has not been reported previously. Conclusion: Brucellosis is a rare cause of pancytopenia, it should be considered in differential diagnosis with pancytopenia of children. Brucellosis was known to be involved cardiovascular system, but tachycardia which was not due to fever as the only sign of disease has not been reported previously made our case very interesting. (Journal of Current Pediatrics 2010; 8: 39-43

  5. Brucella meningoencephalitis with hydrocephalus masquerading as tuberculosis

    Institute of Scientific and Technical Information of China (English)

    Vishwanath Sathyanarayanan; Bekur Ragini; Abdul Razak; M Mukhya prana Prabhu

    2010-01-01

    Neurobrucellosis is a rare form of localized brucellosis usually with no systemic manifestations. We report a rare case of brucellosis presenting as meningoencephalitis associated with hydrocephalus. This patient had a lymphocytic predominantCSF and was initially treated with empirical anti tubercular therapy and steroids.A week later, when hisCSF culture grew Brucella species, the treatment was changed to a combination of streptomycin, doxycycline and rifampicin and the patient improved with this therapy. This case illustrates the need to consider neurobrucellosis as a close differential diagnosis of neurotuberculosis in endemic areas when the patient presents with meningo encephalitis with lymphocyticCSF.

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

    Science.gov (United States)

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

    2014-12-01

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

  7. Research progress in live attenuated Brucella vaccine development.

    Science.gov (United States)

    Wang, Zhen; Wu, Qingmin

    2013-01-01

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

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

    Science.gov (United States)

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

    2016-01-01

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

  9. 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.

  10. Brucella T4SS: the VIP pass inside host cells.

    Science.gov (United States)

    Lacerda, Thais Lourdes Santos; Salcedo, Suzana Pinto; Gorvel, Jean-Pierre

    2013-02-01

    For many Gram-negative bacteria, like Brucella, the type IV secretion system (T4SS) has a critical role in bacterial virulence. In Brucella, the VirB T4SS permits the injection of bacterial effectors inside host cells, leading to subversion of signaling pathways and favoring bacterial growth and pathogenesis. The virB operon promoter is tightly regulated by a combination of transcriptional activators and repressors that are expressed according to the environmental conditions encountered by Brucella. Recent advances have shed light on the Brucella T4SS regulatory mechanisms and also its substrates. Characterization of the targets and functions of these translocated effectors is underway and will help understand the role of the T4SS in the establishment of a replication niche inside host cells.

  11. Chlamydia abortus in Cows Oviducts, Occasional Event or Causal Connection?

    Science.gov (United States)

    Appino, S; Vincenti, L; Rota, A; Pellegrini, S; Chieppa, M N; Cadoni, V; Pregel, P

    2015-06-01

    Fifty-seven genital tracts of regularly slaughtered culled Piedmontese cows, aged 7.4 ± 4.3 years (mean ± SD), range: 2.6-15.6 years, were grossly and microscopically examined. DNA extracted from oviducts was subjected to PCR to evaluate the presence of Chlamydia spp. The 15 PCR-positive oviducts were subjected to Sanger sequencing and showed the presence of Chamydia abortus, with an identity range between 99 and 100%. Nine of the PCR-positive samples belonged to the 24 animals with a normal macroscopic appearance of the whole genital tract (percentage of positive oviducts in normal genital tracts 9/24 = 37.5%), while six belonged to the 33 genital tracts with lesions in one or more organs (percentage of positive oviducts in pathological genital tracts 6/33 = 18.1%); of these, a single animal had salpingitis. The detection of C. abortus in bovine oviducts is of particular interest because it has never been previously investigated or reported. PMID:25704292

  12. A History of the Development of Brucella Vaccines

    Directory of Open Access Journals (Sweden)

    Eric Daniel Avila-Calderón

    2013-01-01

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

  13. Confirmation of Chlamydophila abortus in infected cell culture using Indirect Immunofluorescence technique

    Directory of Open Access Journals (Sweden)

    Krishnan Nair G

    Full Text Available Chlamydophila abortus (C. abortus is an important abortifacient agent in bovines and ovines. Clinical diagnosis of the disease is often difficult. An early diagnosis can be achieved based on direct demonstration of the organism in clinical material and through the cultural recovery of the organism in embryonated chicken egg. For confirmatory diagnosis antigen detection methods or serological techniques can be adopted. The present study is aimed at the confirmatory diagnosis of C. abortus infection by indirect immunofluorescence technique following the isolation of the organism in cell culture. Specific apple green fluorescing inclusions of C. abortus in McCoy cell lines was detected from 72 h to 96 h post infection employing anti-chlamydial group specific monoclonal antibodies. Thus, a confirmatory diagnosis of the infection was possible with this study. [Vet. World 2011; 4(10.000: 473-474

  14. 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.

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

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    Nymo Ingebjørg H

    2011-08-01

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

  16. 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.

  17. Extended safety and efficacy studies of the attenuated Brucella vaccine candidates 16 M(Delta)vjbR and S19(Delta)vjbR in the immunocompromised IRF-1-/- mouse model.

    Science.gov (United States)

    Arenas-Gamboa, A M; Rice-Ficht, A C; Fan, Y; Kahl-McDonagh, M M; Ficht, T A

    2012-02-01

    The global distribution of brucellosis and high incidence in certain areas of the world warrant the development of a safer and efficacious vaccine. For the past 10 years, we have focused our attention on the development of a safer, but still highly protective, live attenuated vaccine for human and animal use. We have demonstrated the safety and protective efficacy of the vaccine candidates 16 MΔvjbR and S19ΔvjbR against homologous and heterologous challenge in multiple immunocompetent animal models, including mice and deer. In the present study, we conducted a series of experiments to determine the safety of the vaccine candidates in interferon regulatory factor-1-knockout (IRF-1(-/-)) mice. IRF-1(-/-) mice infected with either wild-type Brucella melitensis 16 M or the vaccine strain Brucella abortus S19 succumb to the disease within the first 3 weeks of infection, which is characterized by a marked granulomatous and neutrophilic inflammatory response that principally targets the spleen and liver. In contrast, IRF-1(-/-) mice inoculated with either the 16 MΔvjbR or S19ΔvjbR vaccine do not show any clinical or major pathological changes associated with vaccination. Additionally, when 16 MΔvjbR- or S19ΔvjbR-vaccinated mice are challenged with wild-type Brucella melitensis 16M, the degree of colonization in multiple organs, along with associated pathological changes, is significantly reduced. These findings not only demonstrate the safety and protective efficacy of the vjbR mutant in an immunocompromised mouse model but also suggest the participation of lesser-known mechanisms in protective immunity against brucellosis. PMID:22169089

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

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    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.

  19. 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. PMID:23421980

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

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    Loukaides Feidias

    2006-10-01

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

  1. Study on species identification of brucella with small fragments of PCR%小片段PCR产物对布鲁菌种属鉴定的研究

    Institute of Scientific and Technical Information of China (English)

    孙丽媛; 刘彬; 吕雪峰; 李凡

    2010-01-01

    目的 应用小片段PCR产物对布鲁菌进行快速种属鉴定.方法 收集2008年1月至2009年6月在吉林省松原地区从92例布鲁菌病患者血液中分离出的布鲁菌13株,其中羊布鲁菌9株,牛布鲁菌2株,猪布鲁菌2株.根据布鲁菌属细菌重复插入序列IS711及羊布鲁菌、牛布鲁菌及猪布鲁菌种间特异性基因片段设计引物,对3株布鲁菌标准菌株及13株临床分离布鲁菌分别进行PCR反应,获得相应PCR产物.将PCR产物T-A克隆并进行测序分析.对布鲁菌标准菌株DNA模板进行稀释,对每一稀释度的DNA模板分别进行10次PCR扩增,进行灵敏度和稳定性检测.结果 4种PCR反应均可获得特异的小片段PCR产物,片段长度分别为63、67、81和83 bp,T-A克隆测序结果 与目的 基因片段比较,符合率为100%,检测模板的最低DNA浓度为1 μg/L,分别用不同的引物对相应布鲁菌DNA模板进行10次PCR扩增,均得到预期结果 .结论 本研究建立的PCR反应能鉴定布鲁菌种属,具有良好的特异性、灵敏度和稳定性.%Objective To identify the species of brucella rapidly with small fragments of PCR products. Methods The primers were designed according to Brucella spp. repeated insertion sequence IS711 in addition to the specific gene sequences of Brucella melitensis, Brucella abortus and Brucella suis. The small fragments of 3 standard strains and 13 clinical isolates of Brucella were amplified by PCR. Then PCR products were T-A cloned and sequenced. The DNA of standard strains were diluted for sensitivity and stability testing. Results Four specific PCR products were obtained by PCR (63, 67, 81 and 83 bp). The results of T-A cloning and sequencing were in accord with the target gene fragment test. The detection limit of DNA was 1 μg/L. The expected results were achieved with different primers. Conclusion Species identification of Brucella with small fragments of PCR production is a method to identify Brucella strains

  2. 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.

  3. 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

    2014-11-01

    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.

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

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    Andreas F. Haag

    2010-01-01

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

  5. 全自动微生物分析系统对布鲁杆菌属和种鉴定效果的研究%Identification effects of automatic microbial analysis system on brucella genus and species

    Institute of Scientific and Technical Information of China (English)

    肖春霞; 赵鸿雁; 侯临平; 荣蓉; 刘熹; 赵赤鸿; 朴东日; 赵娜; 姜海

    2015-01-01

    Objective To identify and analyse the biochemical characterization of brucella and to evaluate its clinical application by VITEK2 COMPACT automatic microbial identification analyzer.Methods Seventeen strains of standard strains and 121 strains of experimental strains were from bacteria storehouse of brucella disease,Institute of Infectious Diseases Prevention and Control,China Center for Disease Control and Prevention.Experimental strains were from 26 provinces (municipalities and autonomous regions) from 1957 to 2014,including all previous strains from patients and goats,antelope,sheep,cattle,and pig.Reference standard strains and experimental strains were analyzed using the GN identification card on VITEK2 COMPACT automatic microbial identification analyzer,and biochemical identification of brucella strains was done.Identified abnormal strains were rechecked by traditional test methods,including oxidase experiment,urease experiment,semisolid experiment,determination of hydrogen sulfide experiment,basic fuchsin susceptibility experiment,phage lysis experiment,and A/M single-phase specific serum agglutination experiment.Results Of the 138 strains of brucella analyzed by the automatic microbial identification system,the results showed that the main identification indicators of brucella genus were:L-proline arylamidase (ProA),tyrosine arylamidase (TyrA),urease (URE),glycine arylamidase (GlyA),L-lactate alkalinisation (1LATK),and ELLMAN (ELLM).Compared with the system values,all strains biochemical function similar rate was 97.99% (135.23/138),including standard strains was 96.71% (16.44/17),experimental strains was 98.17% (118.79/ 121);time required for strains identification was 6.1-7.7 h,including standard strains was 7.3 h,experimental strains was 6.9 h.Identification indicators for distinguish brucella species were:ProA,TyrA,URE,and GlyA;for distinguish brucella melitensis was ELLM;for distinguish brucella abortus was 1LATK;for distinguish brucella suis was

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

    Science.gov (United States)

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

    2014-09-01

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

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

    Science.gov (United States)

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

    2014-09-01

    Immunisation with attenuated Brucella spp. vaccines prevents brucellosis, but may also interfere with diagnosis. In this study, a duplex PCR was developed to distinguish Brucella suis vaccine strain S2 from field strains of B. suis biovar 1 and other Brucella spp. The PCR detected 60 fg genomic DNA of B. suis S2 or biovar 1 field strains and was able to distinguish B. suis S2 and wild-type strains of B. suis biovar 1 among 76 field isolates representing all the common species and biovars, as well as four vaccine strains, of Brucella. PMID:25011712

  8. Neglected case of osteoarticular Brucella infection of the knee.

    Science.gov (United States)

    Yorgancigil, Huseyin; Yayli, Guler; Oyar, Orhan

    2003-12-01

    A 49-year-old farmer had a history of recurrent knee effusion for 20 years. He did not report undergoing any diagnostic or therapeutic procedures apart from repeated aspirations of the joint fluid. After the isolation of Brucella melitensis from the joint fluid, computed tomography-guided bone biopsy was performed and histopathologic examination of the biopsy sample confirmed the diagnosis of chronic Brucella osteomyelitis. Arthroscopic synovectomy combined with antimicrobial therapy with doxycyclin, rifampicin, and ciprofloxacin for six months resulted in clinical recovery. This case indicates that brucellosis should be suspected in patients with non-specific and chronic osteoarticular symptoms, especially in endemic regions. PMID:14652892

  9. Brucella ceti and Brucellosis in Cetaceans

    Science.gov (United States)

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

    2012-01-01

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

  10. A Study of Brucella Infection in Humans

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    Hasanjani Roushan Mohammad Reza

    2014-07-01

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

  11. Identification of Brucella spp. isolated from human brucellosis in Malaysia using high-resolution melt (HRM) analysis.

    Science.gov (United States)

    Mohamed Zahidi, Jama'ayah; Bee Yong, Tay; Hashim, Rohaidah; Mohd Noor, Azura; Hamzah, Siti Hawa; Ahmad, Norazah

    2015-04-01

    Molecular approaches have been investigated to overcome difficulties in identification and differentiation of Brucella spp. using conventional phenotypic methods. In this study, high-resolution melt (HRM) analysis was used for rapid identification and differentiation of members of Brucella genus. A total of 41 Brucella spp. isolates from human brucellosis were subjected to HRM analysis using 4 sets of primers, which identified 40 isolates as Brucella melitensis and 1 as Brucella canis. The technique utilized low DNA concentration and was highly reproducible. The assay is shown to be a useful diagnostic tool, which can rapidly differentiate Brucella up to species level. PMID:25641125

  12. A rare cause of native tricuspid valve endocarditis: Abortus

    Directory of Open Access Journals (Sweden)

    M.Sıddık Evsen

    2011-03-01

    Full Text Available A 28-year-old female patient, who at her 7 weeks ofpregnancy, admitted to hospital with abdominal painand vaginal bleeding. She had been hospitalized in anothercenter with the diagnosis of spontaneous completeabortion. After discharge, her clinical process, deteriorateddue to fever, chills and fatigue therefore she hadbeen admitted to emergency unit of that hospital onceagain, and received non-specific antibiotics. She was referredto our clinic because of persistant complaints.Transthoracic echocardiography showed vegetations onthe tricuspid valve leading to diagnosis of infective endocarditis,so treatment was started at our clinic. No microorganismisolated in blood cultures. Following 15-days antibiotic therapy no reduction was seen in the diameterof the vegetation, therefore surgical operationwas planned and a bioprosthetic tricuspid valve was putinto place. In this article we aimed to report the developmentof spontaneous abortus at 7 weeks of pregnancy,in order to emphasize that tricuspid valve endocarditiscan be developed secondary to very rare causes.J Clin Exp Invest 2011; 2(1: 102-105

  13. 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

  14. Comparison of the efficacy of Brucella suis strain 2 and Brucella melitensis Rev. 1 live vaccines against a Brucella melitensis experimental infection in pregnant ewes.

    Science.gov (United States)

    Verger, J M; Grayon, M; Zundel, E; Lechopier, P; Olivier-Bernardin, V

    1995-02-01

    The comparative efficacy of Brucella suis strain 2 (S2) and Brucella melitensis strain Rev. 1 (Rev. 1) live vaccines in protecting sheep against B. melitensis infection was evaluated by clinical and bacteriological examination of ewes vaccinated conjunctivally with a dose of 1 x 10(9) c.f.u. when 4 months old and then challenged with 5 x 10(7) c.f.u. of the B. melitensis virulent strain 53H38 (H38) at the middle of the first or second pregnancy following vaccination. Animals were considered to be protected when no abortion, no excretion of the challenge strain and no infection at slaughter occurred. The percentages of protection in Rev. 1-vaccinated groups challenged during either first (80%) or second (62%) pregnancy were significantly different (p S2-vaccinated and control groups. PMID:7625115

  15. [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

  16. Acute Brucella Hepatitis in an Urban Patient

    Directory of Open Access Journals (Sweden)

    Seyed Moayed Alavian

    2009-11-01

    Full Text Available A 35-year-old man was referred to our center because of low grade fever, vomiting, yellow sclera, and tenderness in the upper-right quadrant of his abdomen. Laboratory tests showed a white blood cell (WBC of 7100/µL, a platelet of 184,000/µL, an erythrocyte sedimentation rate (ESR of 4 mm/h, an alanine aminotransferase (ALT of 525 U/L, an aspartate aminotransferase AST of 142 U/L, a total bilirubin level of 4.23 mg/dL, and a direct bilirubin level of 3.16 mg/dL. Viral hepatitis markers, immunoglobuline M antibody to cytomegalovirus (anti-CMV IgM, Ebstein-Barr virus (EBV IgM, and immunologic markers of autoimmune hepatitis were negative. The patient was diagnosed with acute hepatitis. Alkaline phosphatase was in the normal range throughout the course of the disease. Because of the patient's occupation as a butcher and his history of eating semi-cooked sheep testes, serologic tests of brucellosis were conducted; the tests came out positive. We were concerned about the hepatotoxicity of standard regimens; therefore, we started treatment with streptomycin and ciprofloxacin regimens. Although liver enzyme had fallen and fever discontinued, the total and direct bilirubin concentrations in the patient's serum both increased in spite of using 2 weeks of the abovementioned drug regimen. The elevation of bilirubin could have been due to drug hepatotoxicity. Alternatively, a regimen containing ciprofloxacin may have not have been efficient enough and may have had effects on the direct bilirubin concentration. Fortunately, within 8 weeks, progressive recovery was noticed. Brucellosis should be considered in the differential diagnosis of fever and hepatitis for those who live in endemic areas, especially if his/her job was at high risk for acquiring brucellosis. We recommend taking a careful occupational and behavioral history for patients with acute hepatitis syndrome. We assumed that ciprofloxacin was not suitable for brucella hepatitis treatment and

  17. Brucella suis vaccine strain 2 induces endoplasmic reticulum stress that affects intracellular replication in goat trophoblast cells in vitro

    OpenAIRE

    Xiangguo eWang; pengfei elin; yang eli; caixia exiang; yanlong eyin; zhi echen; yue edu; dong ezhou; yaping ejin; Aihua eWang

    2016-01-01

    Brucella has been reported to impair placental trophoblasts, a cellular target where Brucella efficiently replicates in association with the endoplasmic reticulum (ER), and ultimately trigger abortion in pregnant animals. However, the precise effects of Brucella on trophoblast cells remain unclear. Here, we describe the infection and replication of Brucella suis vaccine strain 2 (B.suis.S2) in goat trophoblast cells (GTCs) and the cellular and molecular responses induced in vitro. Our studies...

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

    OpenAIRE

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

    2015-01-01

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

  19. Brucella suis Vaccine Strain 2 Induces Endoplasmic Reticulum Stress that Affects Intracellular Replication in Goat Trophoblast Cells In vitro

    OpenAIRE

    Wang, Xiangguo; Lin, PengFei; Li, Yang; Xiang, Caixia; Yin, Yanlong; Chen, Zhi; Du, Yue; Zhou, dong; Jin, Yaping; Wang, Aihua

    2016-01-01

    Brucella has been reported to impair placental trophoblasts, a cellular target where Brucella efficiently replicates in association with the endoplasmic reticulum (ER), and ultimately trigger abortion in pregnant animals. However, the precise effects of Brucella on trophoblast cells remain unclear. Here, we describe the infection and replication of Brucella suis vaccine strain 2 (B.suis.S2) in goat trophoblast cells (GTCs) and the cellular and molecular responses induced in vitro. Our studies...

  20. 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.

  1. 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.

  2. In Vitro Activities of Six New Fluoroquinolones against Brucella melitensis

    OpenAIRE

    Trujillano-Martín, Ignacio; García-Sánchez, Enrique; Martínez, Isaías Montes; Fresnadillo, María José; García-Sánchez, José Elías; García-Rodríguez, JoséÁngel

    1999-01-01

    We have tested the in vitro activities of eight fluoroquinolones against 160 Brucella melitensis strains. The most active was sitafloxacin (MIC at which 90% of the isolates are inhibited [MIC90], 0.12 μg/ml). In decreasing order, the activities (MIC90s) of the rest of the tested fluoroquinolones were as follows: levofloxacin, 0.5 μg/ml; ciprofloxacin, trovafloxacin, and moxifloxacin, 1 μg/ml; and ofloxacin, grepafloxacin, and gatifloxacin, 2 μg/ml.

  3. A potential novel Brucella species isolated from mandibular lymph nodes of red foxes in Austria.

    Science.gov (United States)

    Hofer, Erwin; Revilla-Fernández, Sandra; Al Dahouk, Sascha; Riehm, Julia M; Nöckler, Karsten; Zygmunt, Michel S; Cloeckaert, Axel; Tomaso, Herbert; Scholz, Holger C

    2012-02-24

    The wild red fox (Vulpes vulpes) is a known indicator species for natural foci of brucellosis. Here, we describe phenotypic and molecular characteristics of two atypical Brucella strains isolated from two foxes hunted 2008 in Eastern Austria. Both strains agglutinated with monospecific anti-Brucella A serum and were positive in ELISA with monoclonal antibodies directed against various Brucella lipopolysaccharide epitopes. However, negative nitrate reductase- and negative oxidase-reaction were atypical traits. Affiliation to the genus Brucella was confirmed by 16S rRNA gene sequencing and by detection of the Brucella specific insertion element IS711 and gene bcsp31 using real-time PCR. Both fox strains showed identical IS711 Southern blot profiles but were distinct from known brucellae. The number of IS711 copies detected was as high as found in B. ovis or marine mammal Brucella strains. Molecular analyses of the recA and omp2a/b genes suggest that both strains possibly represent a novel Brucella species.

  4. 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.

  5. 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.

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

    DEFF Research Database (Denmark)

    Bohlin, Jon; Snipen, Lars; Cloeckaert, Axel;

    2010-01-01

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

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  8. Effect of composition and method of preparation of liposomes on their stability and interaction with murine monocytes infected with Brucella abortus.

    OpenAIRE

    Vitas, A I; Diaz, R.; Gamazo, C

    1996-01-01

    The success of the use of liposomes as drug carriers depends on both their formulation and the method of preparation. We have carried out a series of in vitro studies using different formulations and preparation methods, with the aim of obtaining a type of liposome which is efficient in the treatment of brucellosis. On the basis of results obtained in studies of stability at 37 degrees C in the presence of serum lipoproteins and of the activation of phagocytic cells and antibiotic transport t...

  9. Distinct roles for hepcidin and interleukin-6 in the recovery from anemia in mice injected with heat-killed Brucella abortus

    NARCIS (Netherlands)

    Gardenghi, Sara; Renaud, Tom M; Meloni, Alessandra; Casu, Carla; Crielaard, Bart J; Bystrom, Laura M; Greenberg-Kushnir, Noa; Sasu, Barbra J; Cooke, Keegan S; Rivella, Stefano

    2014-01-01

    Anemia of inflammation (AI) is commonly observed in chronic inflammatory states and may hinder patient recovery and survival. Induction of hepcidin, mediated by interleukin 6, leads to iron-restricted erythropoiesis and anemia. Several translational studies have been directed at neutralizing hepcidi

  10. Occurrence of anti-Brucella abortus and anti-Leptospira spp. antibodies in buffaloes from Paraíba state, Northeastern Brazil

    Directory of Open Access Journals (Sweden)

    Arthur Willian de Lima Brasil

    2015-07-01

    Full Text Available The objective of this study was to determine the frequency of animals that tested positive for brucellosis and leptospirosis and the risk factors for these diseases in the State of Paraíba. A total of 136 buffaloes from 14 herds were examined. For brucellosis, we used the buffered acidified plate antigen (BAPA test as screening method and 2-mercaptoethanol as confirmatory test. For leptospirosis, we conducted a microscopic agglutination test (MAT, with a cut-off point of 1:100. Of the animals examined, two [1.5%; 95% CI = 0.4%–5.2%] were positive for brucellosis, and 38 (27.9%; 95% CI = 21.1%–36.0% were positive for leptospirosis. The brucellosis-positive animals were from two (14.3% herds, while nine (64.3% herds had leptospirosis-seropositive animals. The more frequent Leptospira spp. serotypes were Bratislava, Pomona, and Canicola. We concluded that leptospirosis was widespread in buffaloes in the state of Paraíba and suggested that breeding alongside horses and pigs might be na important factor in the spread of leptospirosis-positive animals. The presence of brucellosispositive animals indicated the possibility of negative-impacting measures on disease control in bovines, and it is therefore recommended that greater attention be given to these animals for brucellosis control.

  11. 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

    2013-09-01

    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.

  12. 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.

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

    Science.gov (United States)

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

    2013-01-01

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

  14. 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. PMID:24803576

  15. 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.

  16. 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.

  17. 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

  18. Intranasal infection with Chlamydia abortus induces dose-dependent latency and abortion in sheep.

    Directory of Open Access Journals (Sweden)

    David Longbottom

    Full Text Available BACKGROUND: Latency is a key feature of the animal pathogen Chlamydia abortus, where infection remains inapparent in the non-pregnant animal and only becomes evident during a subsequent pregnancy. Often the first sign that an animal is infected is abortion occurring late in gestation. Despite this, little is understood of the underlying mechanisms that control latency or the recrudescence of infection that occurs during subsequent pregnancy. The aim of this study was to develop an experimental model of latency by mimicking the natural route of infection through the intranasal inoculation of non-pregnant sheep with C. abortus. METHODOLOGY/PRINCIPAL FINDINGS: Three groups of sheep (groups 1, 2 and 3 were experimentally infected with different doses of C. abortus (5×10(3, 5×10(5 and 5×10(7 inclusion forming units (IFU, respectively prior to mating and monitored over 2 breeding cycles for clinical, microbiological, pathological, immunological and serological outcomes. Two further groups received either negative control inoculum (group 4a,b or were inoculated subcutaneously on day 70 of gestation with 2×10(6 IFU C. abortus (group 5. Animals in groups 1, 2 and 5 experienced an abortion rate of 50-67%, while only one animal aborted in group 3 and none in group 4a,b. Pathological, microbiological, immunological and serological analyses support the view that the maternal protective immune response is influenced by initial exposure to the bacterium. CONCLUSIONS/SIGNIFICANCE: The results show that intranasal administration of non-pregnant sheep with a low/medium dose of C. abortus results in a latent infection that leads in a subsequent pregnancy to infection of the placenta and abortion. In contrast a high dose stimulates protective immunity, resulting in a much lower abortion rate. This model will be useful in understanding the mechanisms of infection underlying latency and onset of disease, as well as in the development of novel therapeutics and

  19. Radiology changes in brucella spondylitis, experience with 26 cases

    International Nuclear Information System (INIS)

    Background/objective: brucellosis is an endemic zoonosis in the Middle East and despite all public health efforts it has not yet been eradicated in Iran. We aimed to highlight and categorize the imaging features of Brucella spondylitis. Material and method: twenty six cases of Brucella spondylitis were treated by the authors from 1982 up to 2003. The available imaging studies of all the cases are reviewed and include X-ray films, conventional myelography, computerized tomographic (CT Scan) and magnetic resonance imaging. Results: there were 21 male and 5 female patients with an age range of 5 to 62 years and the majority (60 %) in the 4 Th and 5 Th decades of life. Wright hemagglutination tests were positive in all cases. Plain x-ray films typically showed lysis of the end plates with osteophyte formation involving affected vertebrae, followed by narrowing of the inter spaces and destruction or collapse of the vertebral bodies in 7 cases. Myelography demonstrated various types of epidural filling defects and obstruction to the flow of contrast material in 10 cases. CT scan, available in 3 cases, showed erosion and cauliflower-like proliferation at the bony edges of the vertebral bodies and end plates. MRI findings varied depending upon the acute or chronic stages of the illness with hypo- or hyper-intense changes on T1 sequences, and primarily hyper-intense changes of T2 sequences in 8 cases. Conclusion: The findings in this series of patients suggest that imaging findings of Brucella spondylitis are scarcely specific. However when considering the medical history, place of origin of the patients, clinical presentation and laboratory findings, the early diagnosis of the illness may be possible before proceeding to surgical intervention

  20. 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.

  1. Prevalence of antibodies to Chlamydophila abortus in ovines in the Londrina area of Paraná state, Brazil

    Directory of Open Access Journals (Sweden)

    Bruna Azevedo de Carvalho Lima

    2014-10-01

    Full Text Available The aim of this paper was to estimate the prevalence of antibodies to Chlamydophila abortus in ewes from the Londrina region of Paraná state. Blood samples from 267 adult ewes from eight herds were submitted to complement fixation testing. Tests were considered positive when the titers ? 32. Nineteen animals (7.11% in 7 (87.50% herds tested positive for antibodies against C. abortus. The titers were 32 in 17 animals and were 64 and 128 in the other two animals. The clinical signs recorded were abortion, stillbirth, weak newborns and heat repetition. The high frequency of herds with at least one positive animal and a seroprevalence of 7.11% suggest the presence of Chlamydophila abortus in the animals of studied herds.

  2. Purification and characterization of an immunogenic aminopeptidase of Brucella melitensis.

    Science.gov (United States)

    Contreras-Rodriguez, Araceli; Ramirez-Zavala, Bernardo; Contreras, Andrea; Schurig, Gerhardt G; Sriranganathan, Nammalwar; Lopez-Merino, Ahide

    2003-09-01

    An immunogenic aminopeptidase was purified from Brucella melitensis strain VTRM1. The purification procedure consisted of ammonium sulfate fractionation and three chromatographic steps. This procedure resulted in a yield of 29% and a 144-fold increase in specific activity. The aminopeptidase appeared to be a monomeric enzyme with a molecular mass of 96 kDa and an isoelectric point of 4.8. Its activity was optimal at pH 7.0 at 40 degrees C. The enzyme was strongly inhibited by EDTA, 1,10-phenathroline, and divalent cations (Zn(2+) and Hg(2+)), suggesting that this protein was a metalloaminopeptidase. The enzyme showed preference for alanine at the N termini of aminoacyl derivatives. The K(m) values for L-alanine-p-nitroanilide (Ala-pNA) and Lys-pNA were 0.35 and 0.18 mM, respectively. The N-terminal sequence of aminopeptidase was used for a homologous search in the genomes of B. melitensis 16M and Brucella suis 1330. The analysis revealed an exact match of the probe sequence (36 bp) with an open reading frame of 2,652 bp encoding a protein predicted to be alanyl aminopeptidase (aminopeptidase N). Collectively, these data suggest designation of the B. melitensis enzyme as an aminopeptidase N. The aminopeptidase was recognized by sera from patients with acute and chronic brucellosis, suggesting that the enzyme may have important diagnostic implications. PMID:12933870

  3. 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.

  4. Prevalence of antibodies to Chlamydophila abortus in ovines in the Londrina area of Paraná state, Brazil

    OpenAIRE

    Bruna Azevedo de Carvalho Lima; Kledir Anderson Hofstaetter Spohr; Francielle Gibson da Silva Zacarias; Sergio Mangano de Almeida Santos; Mario de Luca Neto; Carlo Turilli; Julio Cesar de Freitas

    2014-01-01

    The aim of this paper was to estimate the prevalence of antibodies to Chlamydophila abortus in ewes from the Londrina region of Paraná state. Blood samples from 267 adult ewes from eight herds were submitted to complement fixation testing. Tests were considered positive when the titers ? 32. Nineteen animals (7.11%) in 7 (87.50%) herds tested positive for antibodies against C. abortus. The titers were 32 in 17 animals and were 64 and 128 in the other two animals. The clinical signs recorded w...

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

    Directory of Open Access Journals (Sweden)

    Maria Helena Matiko Akao Larsson

    1984-02-01

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

  6. 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