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Sample records for glanders

  1. Humoral immune responses in a human case of glanders.

    Science.gov (United States)

    Waag, David M; England, Marilyn J; DeShazer, David

    2012-05-01

    Within 2 months of acquiring glanders, a patient developed 8-, 16-, and 4-fold increases, respectively, in specific IgA, IgG, and IgM serological titers against Burkholderia mallei. Within 14 months of infection, the titers decreased to the baseline. Serum from this patient was also highly reactive against Burkholderia pseudomallei whole cells. Burkholderia mallei whole cells did not react with sera from patients with other diseases. Therefore, an assay using a B. mallei cellular diagnostic antigen may be useful for the serodiagnosis of glanders.

  2. Efficacy of postexposure therapy against glanders in mice.

    Science.gov (United States)

    Waag, David M

    2015-04-01

    Burkholderia mallei, the causative agent of glanders, is a CDC Tier 1 Select Agent for which there is no preventive vaccine and antibiotic therapy is difficult. In this study, we show that a combination of vaccination using killed cellular vaccine and therapy using moxifloxacin, azithromycin, or sulfamethoxazole-trimethoprim can protect BALB/c mice from lethal infection even when given 5 days after infectious challenge. Vaccination only, or antibiotic therapy only, was not efficacious. Although antibiotics evaluated experimentally can protect when given before or 1 day after challenge, this time course is not realistic in the cases of natural infection or biological attack, when the patient seeks treatment after symptoms develop or after a biological attack has been confirmed and the agent has been identified. Antibiotics can be efficacious after a prolonged interval between exposure and treatment, but only if the animals were previously vaccinated. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  3. Outer Membrane Vesicle Vaccines from Biosafe Surrogates Prevent Acute Lethal Glanders in Mice

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    Michael H. Norris

    2018-01-01

    Full Text Available Burkholderia mallei is a host-adapted Gram-negative mammalian pathogen that causes the severe disease glanders. Glanders can manifest as a rapid acute progression or a chronic debilitating syndrome primarily affecting solipeds and humans in close association with infected animals. In USA, B. mallei is classified as one of the most important bacterial biothreat agents. Presently, there is no licensed glanders vaccine available for humans or animals. In this work, outer membrane vesicles (OMVs were isolated from three attenuated biosafe bacterial strains, Burkholderia pseudomallei Bp82, B. thailandensis E555, and B. thailandensis TxDOH and used to vaccinate mice. B. thailandensis OMVs induced significantly higher antibody responses that were investigated. B. mallei specific serum antibody responses were of higher magnitude in mice vaccinated with B. thailandensis OMVs compared to levels in mice vaccinated with B. pseudomallei OMVs. OMVs derived from biosafe strains protected mice from acute lethal glanders with vesicles from the two B. thailandensis strains affording significant protection (>90% up to 35 days post-infection with some up to 60 days. Organ loads from 35-day survivors indicated bacteria colonization of the lungs, liver, and spleen while those from 60 days had high CFUs in the spleens. The highest antibody producing vaccine (B. thailandensis E555 OMVs also protected C57BL/6 mice from acute inhalational glanders with evidence of full protection.

  4. Glanders in donkeys (Equus asinus in the state of Pernambuco, Brazil: a case report

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    Rinaldo Aparecido Mota

    2010-03-01

    Full Text Available The clinical, anatomical and histopatological findings of glanders diagnosis in donkeys in the state of Pernambuco-Brazil are reported. The animals were euthanized and necropsied, and evaluated for lesions in respiratory and lymphatic systems, confirming the disease by isolation of Burkholderia mallei and Strauss test.

  5. Use of a Western blot technique for the serodiagnosis of glanders

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    de Souza Marcilia MA

    2011-01-01

    Full Text Available Abstract Background The in vivo diagnosis of glanders relies on the highly sensitive complement fixation test (CFT. Frequently observed false positive results are troublesome for veterinary authorities and cause financial losses to animal owners. Consequently, there is an urgent need to develop a test with high specificity. Hence, a Western blot assay making use of a partly purified lipopolysaccaride (LPS containing antigen of three Burkholderia mallei strains was developed. The test was validated investigating a comprehensive set of positive and negative sera obtained from horses and mules from endemic and non endemic areas. Results The developed Western blot assay showed a markedly higher diagnostic specificity when compared to the prescribed CFT and therefore can be used as a confirmatory test. However, the CFT remains the test of choice for routine testing of glanders due to its high sensitivity, its feasibility using standard laboratory equipment and its worldwide distribution in diagnostic laboratories. Conclusions The CFT should be amended by the newly validated Western blot to increase the positive likelihood ratio of glanders serodiagnosis in non endemic areas or areas with low glanders prevalence. Its use for international trade of horses and mules should be implemented by the OIE.

  6. Development of novel O-polysaccharide based glycoconjugates for immunization against glanders

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    Mary N Burtnick

    2012-11-01

    Full Text Available Burkholderia mallei, the etiologic agent of glanders, causes severe disease in humans and animals and is a potential agent of biological warfare and terrorism. Diagnosis and treatment of glanders can be challenging, and in the absence of chemotherapeutic intervention, acute human disease is invariably fatal. At present, there are no human or veterinary vaccines available for immunization against disease. One of the goals of our research, therefore, is to identify and characterize protective antigens expressed by B. mallei and use them to develop efficacious glanders vaccine candidates. Previous studies have demonstrated that the O-polysaccharide (OPS expressed by B. mallei is both a virulence factor and a protective antigen. Recently, we demonstrated that Burkholderia thailandensis, a closely related but non-pathogenic species, can be genetically manipulated to express OPS antigens that are recognized by B. mallei OPS-specific monoclonal antibodies. As a result, these antigens have become important components of the various OPS-based subunit vaccines that we are currently developing in our laboratory. In this study, we describe a method for isolating B. mallei-like OPS antigens from B. thailandensis oacA mutants. Utilizing these purified OPS antigens, we also describe a simple procedure for coupling the polysaccharides to protein carriers such as cationized bovine serum albumin, diphtheria toxin mutant CRM197 and cholera toxin B subunit. Additionally, we demonstrate that high titer IgG responses against purified B. mallei LPS can be generated by immunizing mice with the resulting constructs. Collectively, these approaches provide a rational starting point for the development of novel OPS-based glycoconjugates for immunization against glanders.

  7. Effectiveness of an antimicrobial treatment scheme in a confined glanders outbreak

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    Saqib Muhammad

    2012-11-01

    Full Text Available Abstract Background Glanders is a contagious and fatal zoonotic disease of solipeds caused by the Gram-negative bacterium Burkholderia (B. mallei. Although regulations call for culling of diseased animals, certain situations e.g. wild life conservation, highly valuable breeding stock, could benefit from effective treatment schemes and post-exposure prophylaxis. Results Twenty three culture positive glanderous horses were successfully treated during a confined outbreak by applying a treatment protocol of 12 weeks duration based on the parenteral administration of enrofloxacin and trimethoprim plus sulfadiazine, followed by the oral administration of doxycycline. Induction of immunosupression in six randomly chosen horses after completion of treatment did not lead to recrudescence of disease. Conclusion This study demonstrates that long term treatment of glanderous horses with a combination of various antibiotics seems to eliminate the agent from the organism. However, more studies are needed to test the effectiveness of this treatment regime on B. mallei strains from different endemic regions. Due to its cost and duration, this treatment can only be an option in certain situations and should not replace the current “testing and culling” policy, in conjunction with adequate compensation to prevent spreading of disease.

  8. Genotyping of Burkholderia mallei from an outbreak of glanders in Bahrain suggests multiple introduction events.

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    Holger C Scholz

    2014-09-01

    Full Text Available Glanders, caused by the gram-negative bacterium Burkholderia mallei, is a highly infectious zoonotic disease of solipeds causing severe disease in animals and men. Although eradicated from many Western countries, it recently emerged in Asia, the Middle-East, Africa, and South America. Due to its rareness, little is known about outbreak dynamics of the disease and its epidemiology.We investigated a recent outbreak of glanders in Bahrain by applying high resolution genotyping (multiple locus variable number of tandem repeats, MLVA and comparative whole genome sequencing to B. mallei isolated from infected horses and a camel. These results were compared to samples obtained from an outbreak in the United Arab Emirates in 2004, and further placed into a broader phylogeographic context based on previously published B. mallei data. The samples from the outbreak in Bahrain separated into two distinct clusters, suggesting a complex epidemiological background and evidence for the involvement of multiple B. mallei strains. Additionally, the samples from Bahrain were more closely related to B. mallei isolated from horses in the United Arab Emirates in 2004 than other B. mallei which is suggestive of repeated importation to the region from similar geographic sources.High-resolution genotyping and comparative whole genome analysis revealed the same phylogenetic patterns among our samples. The close relationship of the Dubai/UAE B. mallei populations to each other may be indicative of a similar geographic origin that has yet to be identified for the infecting strains. The recent emergence of glanders in combination with worldwide horse trading might pose a new risk for human infections.

  9. Aerogenic vaccination with a Burkholderia mallei auxotroph protects against aerosol-initiated glanders in mice.

    Science.gov (United States)

    Ulrich, Ricky L; Amemiya, Kei; Waag, David M; Roy, Chad J; DeShazer, David

    2005-03-14

    Burkholderia mallei is an obligate mammalian pathogen that causes the zoonotic disease glanders. Two live attenuated B. mallei strains, a capsule mutant and a branched-chain amino acid auxotroph, were evaluated for use as vaccines against aerosol-initiated glanders in mice. Animals were aerogenically vaccinated and serum samples were obtained before aerosol challenge with a high-dose (>300 times the LD50) of B. mallei ATCC 23344. Mice vaccinated with the capsule mutant developed a Th2-like Ig subclass antibody response and none survived beyond 5 days. In comparison, the auxotrophic mutant elicited a Th1-like Ig subclass antibody response and 25% of the animals survived for 1 month postchallenge. After a low-dose (5 times the LD50) aerosol challenge, the survival rates of auxotroph-vaccinated and unvaccinated animals were 50 and 0%, respectively. Thus, live attenuated strains that promote a Th1-like Ig response may serve as promising vaccine candidates against aerosol infection with B. mallei.

  10. Assessment of the effectiveness of the PPD-mallein produced in Brazil for diagnosing glanders in mules.

    Science.gov (United States)

    da Silva, Karla Patrícia Chaves; de Campos Takaki, Galba Maria; da Silva, Leonildo Bento Galiza; Saukas, Tomoe Noda; Santos, André Souza; Mota, Rinaldo Aparecido

    2013-01-01

    To assess the potency of the PPD-mallein produced in Brazil, five animals were from a property identified as a focus of glanders. These animals had suggestive clinical signs of the disease and the other five, from a property free from glanders, showed no clinical signs and were serology negative (control group). PPD-mallein from Burkholderia mallei was obtained by precipitation with trichloroacetic acid and ammonium sulfate. The animals were inoculated according to the criteria established by Department of Agriculture, Livestock and Supply (MAPA) for the diagnosis of glanders. After 48 h of application of PPD-mallein, there was swelling in the area of application, presence of ocular secretion and tears in sick animals. The control group showed no inflammatory reaction at the site of inoculation of PPD-mallein. This immunogen produced in Brazil and still being tested was effective for identifying the infection in true positive animals and excluding the truly negative ones, being a new possibility for diagnosis and control of glanders.

  11. Development of capsular polysaccharide-based glycoconjugates for immunization against melioidosis and glanders

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    Mary N Burtnick

    2012-08-01

    Full Text Available Burkholderia pseudomallei and Burkholderia mallei, the etiologic agents of melioidosis and glanders respectively, cause severe disease in humans and animals and are considered potential agents of biological warfare and terrorism. Diagnosis and treatment of infections caused by these pathogens can be challenging and, in the absence of chemotherapeutic intervention, acute disease is frequently fatal. At present, there are no human or veterinary vaccines available for immunization against these emerging/re-emerging infectious diseases. One of the long term objectives of our research, therefore, is to identify and characterize protective antigens expressed by B. pseudomallei and B. mallei and use them to develop efficacious vaccine candidates. Previous studies have demonstrated that the 6-deoxy-heptan capsular polysaccharide (CPS expressed by these bacterial pathogens is both a virulence determinant and a protective antigen. Consequently, this carbohydrate moiety has become an important component of the various subunit vaccines that we are currently developing in our laboratory. In the present study, we describe a reliable method for isolating CPS antigens from O-polysaccharide deficient strains of B. pseudomallei; including a derivative of the select agent excluded strain Bp82. Utilizing these purified CPS samples, we also describe a simple procedure for covalently linking these T-cell independent antigens to carrier proteins. In addition, we demonstrate that high titer IgG responses can be raised against the CPS component of such constructs. Collectively, these approaches provide a tangible starting point for the development of novel CPS-based glycoconjugates for immunization against melioidosis and glanders.

  12. Recombinant Salmonella Expressing Burkholderia mallei LPS O Antigen Provides Protection in a Murine Model of Melioidosis and Glanders.

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    Moustafa, Dina A; Scarff, Jennifer M; Garcia, Preston P; Cassidy, Sara K B; DiGiandomenico, Antonio; Waag, David M; Inzana, Thomas J; Goldberg, Joanna B

    2015-01-01

    Burkholderia pseudomallei and Burkholderia mallei are the etiologic agents of melioidosis and glanders, respectively. These bacteria are highly infectious via the respiratory route and can cause severe and often fatal diseases in humans and animals. Both species are considered potential agents of biological warfare; they are classified as category B priority pathogens. Currently there are no human or veterinary vaccines available against these pathogens. Consequently efforts are directed towards the development of an efficacious and safe vaccine. Lipopolysaccharide (LPS) is an immunodominant antigen and potent stimulator of host immune responses. B. mallei express LPS that is structurally similar to that expressed by B. pseudomallei, suggesting the possibility of constructing a single protective vaccine against melioidosis and glanders. Previous studies of others have shown that antibodies against B. mallei or B. pseudomallei LPS partially protect mice against subsequent lethal virulent Burkholderia challenge. In this study, we evaluated the protective efficacy of recombinant Salmonella enterica serovar Typhimurium SL3261 expressing B. mallei O antigen against lethal intranasal infection with Burkholderia thailandensis, a surrogate for biothreat Burkholderia spp. in a murine model that mimics melioidosis and glanders. All vaccine-immunized mice developed a specific antibody response to B. mallei and B. pseudomallei O antigen and to B. thailandensis and were significantly protected against challenge with a lethal dose of B. thailandensis. These results suggest that live-attenuated SL3261 expressing B. mallei O antigen is a promising platform for developing a safe and effective vaccine.

  13. Recombinant Salmonella Expressing Burkholderia mallei LPS O Antigen Provides Protection in a Murine Model of Melioidosis and Glanders.

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    Dina A Moustafa

    Full Text Available Burkholderia pseudomallei and Burkholderia mallei are the etiologic agents of melioidosis and glanders, respectively. These bacteria are highly infectious via the respiratory route and can cause severe and often fatal diseases in humans and animals. Both species are considered potential agents of biological warfare; they are classified as category B priority pathogens. Currently there are no human or veterinary vaccines available against these pathogens. Consequently efforts are directed towards the development of an efficacious and safe vaccine. Lipopolysaccharide (LPS is an immunodominant antigen and potent stimulator of host immune responses. B. mallei express LPS that is structurally similar to that expressed by B. pseudomallei, suggesting the possibility of constructing a single protective vaccine against melioidosis and glanders. Previous studies of others have shown that antibodies against B. mallei or B. pseudomallei LPS partially protect mice against subsequent lethal virulent Burkholderia challenge. In this study, we evaluated the protective efficacy of recombinant Salmonella enterica serovar Typhimurium SL3261 expressing B. mallei O antigen against lethal intranasal infection with Burkholderia thailandensis, a surrogate for biothreat Burkholderia spp. in a murine model that mimics melioidosis and glanders. All vaccine-immunized mice developed a specific antibody response to B. mallei and B. pseudomallei O antigen and to B. thailandensis and were significantly protected against challenge with a lethal dose of B. thailandensis. These results suggest that live-attenuated SL3261 expressing B. mallei O antigen is a promising platform for developing a safe and effective vaccine.

  14. Glanders (Burkholderia Mallei)

    Science.gov (United States)

    ... primarily a disease affecting horses. It also affects donkeys and mules and can be naturally contracted by ... file Microsoft Word file Microsoft Excel file Audio/Video file Apple Quicktime file RealPlayer file Text file ...

  15. Phenotypic characterization of a novel virulence-factor deletion strain of Burkholderia mallei that provides partial protection against inhalational glanders in mice

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    Joel A. Bozue

    2016-02-01

    Full Text Available Burkholderia mallei (Bm is a highly infectious intracellular pathogen classified as a category B biological agent by the Centers for Disease Control and Prevention. After respiratory exposure, Bm establishes itself within host macrophages before spreading into major organ systems, which can lead to chronic infection, sepsis, and death. Previously, we combined computational prediction of host-pathogen interactions with yeast two-hybrid experiments and identified novel virulence factor genes in Bm, including BMAA0553, BMAA0728 (tssN, and BMAA1865. In the present study, we used recombinant allelic exchange to construct deletion mutants of BMAA0553 and tssN (ΔBMAA0553 and ΔTssN, respectively and showed that both deletions completely abrogated virulence at doses of >100 times the LD50 of the wild-type Bm strain. Analysis of ΔBMAA0553- and ΔTssN-infected mice showed starkly reduced bacterial dissemination relative to wild-type Bm, and subsequent in vitro experiments characterized pathogenic phenotypes with respect to intracellular growth, macrophage uptake and phagosomal escape, actin-based motility, and multinucleated giant cell formation. Based on observed in vitro and in vivo phenotypes, we explored the use of ΔTssN as a candidate live-attenuated vaccine. Mice immunized with aerosolized ΔTssN showed a 21-day survival rate of 67% after a high-dose aerosol challenge with the wild-type Bm ATCC 23344 strain, compared to a 0% survival rate for unvaccinated mice. However, analysis of histopathology and bacterial burden showed that while the surviving vaccinated mice were protected from acute infection, Bm was still able to establish a chronic infection. Vaccinated mice showed a modest IgG response, suggesting a limited potential of ΔTssN as a vaccine candidate, but also showed prolonged elevation of pro-inflammatory cytokines, underscoring the role of cellular and innate immunity in mitigating acute infection in inhalational glanders.

  16. Disease: H00316 [KEGG MEDICUS

    Lifescience Database Archive (English)

    Full Text Available H00316 Glanders Glanders is an infectious disease of equines that can be transmitt...ent of glanders is Burkholderia mallei, a gram-negative aerobic bacterium. In equines and humans, B. mallei

  17. In Vitro Antibiotic Susceptibilities of Burkholderia mallei (Causative Agent of Glanders) Determined by Broth Microdilution and E-Test

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    Heine, Henry S.; England, Marilyn J.; Waag, David M.; Byrne, W. Russell

    2001-01-01

    In vitro susceptibilities to 28 antibiotics were determined for 11 strains of Burkholderia mallei by the broth microdilution method. The B. mallei strains demonstrated susceptibility to aminoglycosides, macrolides, quinolones, doxycycline, piperacillin, ceftazidime, and imipenem. For comparison and evaluation, 17 antibiotic susceptibilities were also determined by the E-test. E-test values were always lower than the broth dilution values. Establishing and comparing antibiotic susceptibilities of specific B. mallei strains will provide reference information for assessing new antibiotic agents. PMID:11408233

  18. NATO Planning Guide for the Estimation of Chemical, Biological, Radiological, and Nuclear (CBRN) Casualties (AMedP-8(C)) - Parameters for Estimation of Casualties from Exposure to Specified Biological Agents. Addenda to Allied Medical Publication 8

    Science.gov (United States)

    2011-01-01

    no. 2510 (1909): 319–25; I. Sobol , “A Case of Chronic Nasal Glanders,” Acta Oto-Laryngologica 18, no. 4 (1933): 500–9; J. F. Burgess, “Chronic...Observations on Human Glanders;” Sobol , “A Case of Chronic Nasal Glanders;” Burgess, “Chronic Glanders;” Herold and Erickson, "Human Glanders: Case...Human Subject;” Hunting, Glanders: A Clinical Treatise; Bernstein and Carling, “Observations on Human Glanders;” Sobol , “A Case of Chronic Nasal

  19. Phenotypic Characterization of a Novel Virulence-Factor Deletion Strain of Burkholderia mallei that Provides Partial Protection against Inhalational Glanders in Mice

    Science.gov (United States)

    2016-02-26

    Fort Detrick, MD, USA, 2 Telemedicine and Advanced Technology Research Center, Biotechnology HPC Software Applications Institute, United States Army...allelic exchange mutants, we grew the Bm cointegrate strain in yeast- extract tryptone (YT) broth medium and then serially diluted it onto YT agar...indicated. Cells were fixed with 4% paraformaldehyde and then blocked in PBS containing 0.25% saponin , 0.2% Bovine Serum Albumin (BSA) fraction V

  20. Development of a Polymerase Chain Reaction Assay for the Specific Identification of Burkholderia mallei and Differentiation from Burkholderia pseudomallei and Other Closely Related Burkholderiaceae

    National Research Council Canada - National Science Library

    Ulrich, Ricky L; Ulrich, Melanie P; Schell, Mark A; Kim, H. S; DeShazer, David

    2005-01-01

    Burkholderia mallei and Burkholderia pseudomallei, the etiologic agents responsible for glanders and melioidosis, respectively, are genetically and phenotypically similar and are category B biothreat agents...

  1. Monoclonal antibodies passively protect BALB/c mice against Burkholderia mallei aerosol challenge.

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    Treviño, Sylvia R; Permenter, Amy R; England, Marilyn J; Parthasarathy, Narayanan; Gibbs, Paul H; Waag, David M; Chanh, Tran C

    2006-03-01

    Glanders is a debilitating disease with no vaccine available. Murine monoclonal antibodies were produced against Burkholderia mallei, the etiologic agent of glanders, and were shown to be effective in passively protecting mice against a lethal aerosol challenge. The antibodies appeared to target lipopolysaccharide. Humoral antibodies may be important for immune protection against B. mallei infection.

  2. Distinct human antibody response to the biological warfare agent Burkholderia mallei.

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    Varga, John J; Vigil, Adam; DeShazer, David; Waag, David M; Felgner, Philip; Goldberg, Joanna B

    2012-10-01

    The genetic similarity between Burkholderia mallei (glanders) and Burkholderia pseudomallei (melioidosis) had led to the general assumption that pathogenesis of each bacterium would be similar. In 2000, the first human case of glanders in North America since 1945 was reported in a microbiology laboratory worker. Leveraging the availability of pre-exposure sera for this individual and employing the same well-characterized protein array platform that has been previously used to study a large cohort of melioidosis patients in southeast Asia, we describe the antibody response in a human with glanders. Analysis of 156 peptides present on the array revealed antibodies against 17 peptides with a > 2-fold increase in this infection. Unexpectedly, when the glanders data were compared with a previous data set from B. pseudomallei infections, there were only two highly increased antibodies shared between these two infections. These findings have implications in the diagnosis and treatment of B. mallei and B. pseudomallei infections.

  3. Innate Immune Response to Burkholderia mallei

    Science.gov (United States)

    2017-02-16

    highly contagious and often fatal disease, glanders. With its high rate of infectivity via aerosol and recalcitrance towards antibiotics , this...pathology (1-7). Glanders transmits amongst animals via respiratory secretions and exudates from skin lesions. In human infections, the primary modes of...resulted in the uniform production of cytokines interleukin 1 beta (IL-1β), tumor necrosis factor alpha (TNF-α), and murine keratinocyte-derived

  4. History of the Chemical Warfare Service in World War II. Biological Warfare Research in the United States, Volume 2

    Science.gov (United States)

    1947-11-01

    regarded as a pos- itive test. No individual, however, with a known history of brucell - osis or skin sensitivity was given either Brucellergen...investigation made of the organism and toxin. Brucell - osis caused the greatest tiaie loss per case, averaging 177 days; glanders was next with 121 days...Eeningopneumonitis viruses, brucel - losis, tularemia, end melioido3is. Studies were also made using this type chamber on the employment of a dye as a tracer in

  5. A CpG oligonucleotide can protect mice from a low aerosol challenge dose of Burkholderia mallei.

    Science.gov (United States)

    Waag, David M; McCluskie, Michael J; Zhang, Ningli; Krieg, Arthur M

    2006-03-01

    Treatment with an oligodeoxynucleotide (ODN) containing CPG motifs (CpG ODN 7909) was found to protect BALB/c mice from lung infection or death after aerosol challenge with Burkholderia mallei. Protection was associated with enhanced levels of gamma interferon (IFN-gamma)-inducible protein 10, interleukin-12 (IL-12), IFN-gamma, and IL-6. Preexposure therapy with CpG ODNs may protect victims of a biological attack from glanders.

  6. New England Bioterrorism Preparedness Workshop

    Science.gov (United States)

    2002-04-04

    Hypoxia • GI – Fever – Nausea/vomiting – Diarrhea (+/-bloody) • Rash and fever – Vesicular – Petechial • Neurologic – cranial nerve palsies, HA...plague) • variola major (smallpox) • Francisella tularensis (tularemia) • Viral hemorrhagic fever Agents of Concern: CDC Category B • Coxiella...burnetti (Q fever ) • Brucella species (brucellosis) • Burkholderia mallei (glanders) • ricin toxin from Ricinus communis (castor beans) • epsilon toxin of

  7. Use of the common marmoset to study Burkholderia mallei infection.

    Directory of Open Access Journals (Sweden)

    Tomislav Jelesijevic

    Full Text Available Burkholderia mallei is a host-adapted bacterium that does not persist outside of its equine reservoir. The organism causes the zoonosis glanders, which is endemic in Asia, Africa, the Middle East and South America. Infection by B. mallei typically occurs via the respiratory or percutaneous route, and the most common manifestations are life-threatening pneumonia and bacteremia. Glanders is difficult to diagnose and requires prolonged antibiotic therapy with low success rates. There is no vaccine to protect against B. mallei and there is concern regarding its use as a biothreat agent. Thus, experiments were performed to establish a non-human primate model of intranasal infection to study the organism and develop countermeasures. Groups of marmosets (Callithrix jacchus were inoculated intranasally with B. mallei strain ATCC 23344 and monitored for clinical signs of illness for up to 13 days. We discovered that 83% of marmosets inoculated with doses of 2.5 X 10(4 to 2.5 X 10(5 bacteria developed acute lethal infection within 3-4 days. Signs of disease were severe and included lethargy, inappetence, conjunctivitis, mucopurulent and hemorrhagic nasal discharges, and increased respiratory effort with abdominal lifts. Burkholderia mallei was cultured from the lungs, spleen and liver of these animals, and pathologic examination of tissues revealed lesions characteristic of glanders. Challenge experiments also revealed that 91% of animals infected with doses ranging from 25 to 2.5 X 10(3 bacteria exhibited mild non-specific signs of illness and were culture negative. One marmoset inoculated with 2.5 X 10(3 organisms developed moderate signs of disease and reached humane end-points 8 days post-infection. The liver and spleen of this animal were colonized with the agent and pathological analysis of tissues showed nasal, splenic and hepatic lesions. Taken together, these data indicate that the marmoset is a suitable model to study respiratory infection by B

  8. STUDIES OF METHOD FOR DETERMINING THE PROTEIN CONCENTRATION OF "MALEIN PPD" BY THE KJELDAHL METHOD

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    Ciuca, V

    2017-06-01

    Full Text Available Glanders is a contagious and fatal disease of horses, donkeys, and mules, caused by infection with the bacterium Burkholderia mallei. The pathogen causes nodules and ulcerations in the upper respiratory tract and lungs. Glanders is transmissible to humans by direct contact with diseased animals or with infected or contaminated material. In the untreated acute disease, the mortality rate can reach 95% within 3 weeks Malein PPD - the diagnostic product contain max 2mg/ml Burkholderia mallei. The amount of protein in the biological product "Malein PPD" is measured as nitrogen from protein molecule, applying the Kjeldahl (method determination of nitrogen by sulphuric acid digestion. The validation study aims to demonstrate the determination of the protein of the Malein PPD, by sulphuric acid digestion, it is an appropriate analytical method, reproducible and meets the quality requirements of diagnostic reagents. The paper establishes the performance characteristics of the method considered and identify the factors that influence these characteristics. The method for determining the concentration of protein, by the Kjeldahl method is considered valid if the results obtained for each validation parameter are within the admissibility criteria.The validation procedure includes details on protocol working to determine the protein of the Malein PPD, validation criteria, experimental results, mathematical calculations.

  9. Development of Burkholderia mallei and pseudomallei vaccines

    Science.gov (United States)

    Silva, Ediane B.; Dow, Steven W.

    2013-01-01

    Burkholderia mallei and Burkholderia pseudomallei are Gram-negative bacteria that cause glanders and melioidosis, respectively. Inhalational infection with either organism can result in severe and rapidly fatal pneumonia. Inoculation by the oral and cutaneous routes can also produce infection. Chronic infection may develop after recovery from acute infection with both agents, and control of infection with antibiotics requires prolonged treatment. Symptoms for both meliodosis and glanders are non-specific, making diagnosis difficult. B. pseudomallei can be located in the environment, but in the host, B. mallei and B. psedomallei are intracellular organisms, and infection results in similar immune responses to both agents. Effective early innate immune responses are critical to controlling the early phase of the infection. Innate immune signaling molecules such as TLR, NOD, MyD88, and pro-inflammatory cytokines such as IFN-γ and TNF-α play key roles in regulating control of infection. Neutrophils and monocytes are critical cells in the early infection for both microorganisms. Both monocytes and macrophages are necessary for limiting dissemination of B. pseudomallei. In contrast, the role of adaptive immune responses in controlling Burkholderia infection is less well understood. However, T cell responses are critical for vaccine protection from Burkholderia infection. At present, effective vaccines for prevention of glanders or meliodosis have not been developed, although recently development of Burkholderia vaccines has received renewed attention. This review will summarize current and past approaches to develop B. mallei and B. pseudomalllei vaccines, with emphasis on immune mechanisms of protection and the challenges facing the field. At present, immunization with live attenuated bacteria provides the most effective and durable immunity, and it is important therefore to understand the immune correlates of protection induced by live attenuated vaccines. Subunit

  10. Characterization of in vitro phenotypes of Burkholderia pseudomallei and Burkholderia mallei strains potentially associated with persistent infection in mice.

    Science.gov (United States)

    Bernhards, R C; Cote, C K; Amemiya, K; Waag, D M; Klimko, C P; Worsham, P L; Welkos, S L

    2017-03-01

    Burkholderia pseudomallei (Bp) and Burkholderia mallei (Bm), the agents of melioidosis and glanders, respectively, are Tier 1 biothreats. They infect humans and animals, causing disease ranging from acute and fatal to protracted and chronic. Chronic infections are especially challenging to treat, and the identification of in vitro phenotypic markers which signal progression from acute to persistent infection would be extremely valuable. First, a phenotyping strategy was developed employing colony morphotyping, chemical sensitivity testing, macrophage infection, and lipopolysaccharide fingerprint analyses to distinguish Burkholderia strains. Then mouse spleen isolates collected 3-180 days after infection were characterized phenotypically. Isolates from long-term infections often exhibited increased colony morphology differences and altered patterns of antimicrobial sensitivity and macrophage infection. Some of the Bp and Bm persistent infection isolates clearly displayed enhanced virulence in mice. Future studies will evaluate the potential role and significance of these phenotypic markers in signaling the establishment of a chronic infection.

  11. The type IV pilin of Burkholderia mallei is highly immunogenic but fails to protect against lethal aerosol challenge in a murine model.

    Science.gov (United States)

    Fernandes, Paula J; Guo, Qin; Waag, David M; Donnenberg, Michael S

    2007-06-01

    Burkholderia mallei is the cause of glanders and a proven biological weapon. We identified and purified the type IV pilin protein of this organism to study its potential as a subunit vaccine. We found that purified pilin was highly immunogenic. Furthermore, mice infected via sublethal aerosol challenge developed significant increases in titers of antibody against the pilin, suggesting that it is expressed in vivo. Nevertheless, we found no evidence that high-titer antipilin antisera provided passive protection against a sublethal or lethal aerosol challenge and no evidence of protection afforded by active immunization with purified pilin. These results contrast with the utility of type IV pilin subunit vaccines against other infectious diseases and highlight the need for further efforts to identify protective responses against this pathogen.

  12. Characterization of the Burkholderia mallei tonB Mutant and Its Potential as a Backbone Strain for Vaccine Development.

    Directory of Open Access Journals (Sweden)

    Tiffany M Mott

    Full Text Available In this study, a Burkholderia mallei tonB mutant (TMM001 deficient in iron acquisition was constructed, characterized, and evaluated for its protective properties in acute inhalational infection models of murine glanders and melioidosis.Compared to the wild-type, TMM001 exhibits slower growth kinetics, siderophore hyper-secretion and the inability to utilize heme-containing proteins as iron sources. A series of animal challenge studies showed an inverse correlation between the percentage of survival in BALB/c mice and iron-dependent TMM001 growth. Upon evaluation of TMM001 as a potential protective strain against infection, we found 100% survival following B. mallei CSM001 challenge of mice previously receiving 1.5 x 10(4 CFU of TMM001. At 21 days post-immunization, TMM001-treated animals showed significantly higher levels of B. mallei-specific IgG1, IgG2a and IgM when compared to PBS-treated controls. At 48 h post-challenge, PBS-treated controls exhibited higher levels of serum inflammatory cytokines and more severe pathological damage to target organs compared to animals receiving TMM001. In a cross-protection study of acute inhalational melioidosis with B. pseudomallei, TMM001-treated mice were significantly protected. While wild type was cleared in all B. mallei challenge studies, mice failed to clear TMM001.Although further work is needed to prevent chronic infection by TMM001 while maintaining immunogenicity, our attenuated strain demonstrates great potential as a backbone strain for future vaccine development against both glanders and melioidosis.

  13. Caracterização fenotípica e molecular de amostras de Burkholderia mallei isoladas na Região Nordeste do Brasil Phenotypic and molecular characterization of Burkholderia mallei isolated in northeastern Brazil

    Directory of Open Access Journals (Sweden)

    Karla P.C. Silva

    2009-05-01

    Full Text Available Objetivou-se com este trabalho realizar o estudo bioquímico e molecular de amostras de Burkholderia mallei isoladas de eqüídeos com diagnóstico clínico e sorológico para o mormo e provenientes da Região Metropolitana do Recife-PE e Zona da Mata dos Estados de Alagoas e Pernambuco. Foram realizadas as técnicas microbiológicas para o isolamento e identificação fenotípica de B. mallei e as técnicas moleculares de ribotipagem-PCR e RAPD-PCR. Das oito amostras estudadas, quatro apresentaram pequenas variações fenotípicas. Nas técnicas moleculares, as amostras formaram quatro grupos de diferentes perfis ribotípicos, demonstrando também quatro perfis genotípicos. Houve associação nos resultados da Ribotipagem-PCR e RAPD-PCR. As variações nos perfis ribotípicos e genotípicos foram associadas às diferentes regiões estudadas. De acordo com os resultados obtidos, conclui-se que as pequenas variações bioquímicas não estão associadas aos diferentes perfis moleculares e que essas diferenças demonstram uma heterogeneidade que está associada à procedência das amostras, indicando que a infecção nos animais ocorre por clones diferentes das amostras analisadas.The objective of this paper was to study the molecular performance and phenotypic characterization of Burkholderia mallei isolated from horses with clinical and serological diagnosis of glanders, originating from the Metropolitan District of Recife and Zona da Mata of Pernambuco and Alagoas. The isolation and biochemical identification of B. mallei was carried out by microbiological and molecular techniques of PCR-fingerprinting and RAPD-PCR. From the eight samples studied, four showed little phenotype variations. In the molecular tests, the samples formed 4 groups of different ribotype profiles and 4 genotype profiles. There was some association of PCR-fingerprinting with RAPD-PCR results. It was concluded that the slight biochemical variations were not associated with

  14. Burkholderia Vaccines: Are We Moving Forward?

    Directory of Open Access Journals (Sweden)

    Leang-Chung eChoh

    2013-02-01

    Full Text Available The genus Burkholderia consists of diverse species which includes both ‘friends’ and ‘foes’. Some of the ‘friendly’ Burkholderia spp. are extensively used in the biotechnological and agricultural industry for bioremediation and biocontrol. However, several members of the genus including B. pseudomallei, B. mallei and B. cepacia, are known to cause fatal disease in both humans and animals. B. pseudomallei and B. mallei are the causative agents of melioidosis and glanders, respectively, while B. cepacia infection is lethal to cystic fibrosis patients. Due to the high rate of infectivity and intrinsic resistance to many commonly used antibiotics, together with high mortality rate, B. mallei and B. pseudomallei are considered to be potential biological warfare agents. Treatments of the infections caused by these bacteria are often unsuccessful with frequent relapse of the infection. Thus, we are at a crucial stage of the need for Burkholderia vaccines. Although the search for a prophylactic therapy candidate continues, to date development of vaccines has not advanced beyond research to human clinical trials. In this article, we review the current research on development of safe vaccines with high efficacy against B. pseudomallei, B. mallei and B. cepacia. It can be concluded that further research will enable elucidation of the potential benefits and risks of Burkholderia vaccines.

  15. Population-Sequencing as a Biomarker of Burkholderia mallei and Burkholderia pseudomallei Evolution through Microbial Forensic Analysis

    Directory of Open Access Journals (Sweden)

    John P. Jakupciak

    2013-01-01

    Full Text Available Large-scale genomics projects are identifying biomarkers to detect human disease. B. pseudomallei and B. mallei are two closely related select agents that cause melioidosis and glanders. Accurate characterization of metagenomic samples is dependent on accurate measurements of genetic variation between isolates with resolution down to strain level. Often single biomarker sensitivity is augmented by use of multiple or panels of biomarkers. In parallel with single biomarker validation, advances in DNA sequencing enable analysis of entire genomes in a single run: population-sequencing. Potentially, direct sequencing could be used to analyze an entire genome to serve as the biomarker for genome identification. However, genome variation and population diversity complicate use of direct sequencing, as well as differences caused by sample preparation protocols including sequencing artifacts and mistakes. As part of a Department of Homeland Security program in bacterial forensics, we examined how to implement whole genome sequencing (WGS analysis as a judicially defensible forensic method for attributing microbial sample relatedness; and also to determine the strengths and limitations of whole genome sequence analysis in a forensics context. Herein, we demonstrate use of sequencing to provide genetic characterization of populations: direct sequencing of populations.

  16. Chronic suppurative joint effusion due to burkholderia pseudomallei: A case report

    Directory of Open Access Journals (Sweden)

    Madhavi Deshmukh

    2013-01-01

    Full Text Available Burkholderia pseudomallei, a Gram-negative bacillus is the causative agent of Melioidosis, a glanders-like disease, primarily a disease of animals. Melioidosis has been only a rare and sporadic disease in humans outside its endemic region. Currently, diagnosis of B. pseudomallei in the clinical laboratory is very difficult, owing to low awareness of physicians to the nonspecific clinical manifestations, lack of responsiveness among microbiologists outside endemic areas, identification systems in the average sentinel laboratory, and the biosafety conditions necessary to process these organisms. We report a case of chronic left hip joint effusion in a known case of diabetes mellitus. Gram stain of computed tomography (CT-guided aspirate from the joint revealed Gram-negative bacilli along with pus cells. Culture was confirmed as Burkholderia pseudomallei on Vitek2C, which was sensitive to ceftazidime and trimethoprim/sulfmethoxazole. Unfortunately, patient could not be started on appropriate antibiotics due to delay in detection and patient succumbed to severe septicemia. This case is reported to highlight importance of automated identification and sensitivity especially in nonendemic areas and unusual antibiogram of this organism for which disc diffusion method is not standardized.

  17. Burkholderia mallei tssM encodes a putative deubiquitinase that is secreted and expressed inside infected RAW 264.7 murine macrophages.

    Science.gov (United States)

    Shanks, John; Burtnick, Mary N; Brett, Paul J; Waag, David M; Spurgers, Kevin B; Ribot, Wilson J; Schell, Mark A; Panchal, Rekha G; Gherardini, Frank C; Wilkinson, Keith D; Deshazer, David

    2009-04-01

    Burkholderia mallei, a category B biothreat agent, is a facultative intracellular pathogen that causes the zoonotic disease glanders. The B. mallei VirAG two-component regulatory system activates the transcription of approximately 60 genes, including a large virulence gene cluster encoding a type VI secretion system (T6SS). The B. mallei tssM gene encodes a putative ubiquitin-specific protease that is physically linked to, and transcriptionally coregulated with, the T6SS gene cluster. Mass spectrometry and immunoblot analysis demonstrated that TssM was secreted in a virAG-dependent manner in vitro. Surprisingly, the T6SS was found to be dispensable for the secretion of TssM. The C-terminal half of TssM, which contains Cys and His box motifs conserved in eukaryotic deubiquitinases, was purified and biochemically characterized. Recombinant TssM hydrolyzed multiple ubiquitinated substrates and the cysteine at position 102 was critical for enzymatic activity. The tssM gene was expressed within 1 h after uptake of B. mallei into RAW 264.7 murine macrophages, suggesting that the TssM deubiquitinase is produced in this intracellular niche. Although the physiological substrate(s) is currently unknown, the TssM deubiquitinase may provide B. mallei a selective advantage in the intracellular environment during infection.

  18. Burkholderia mallei tssM Encodes a Putative Deubiquitinase That Is Secreted and Expressed inside Infected RAW 264.7 Murine Macrophages▿ †

    Science.gov (United States)

    Shanks, John; Burtnick, Mary N.; Brett, Paul J.; Waag, David M.; Spurgers, Kevin B.; Ribot, Wilson J.; Schell, Mark A.; Panchal, Rekha G.; Gherardini, Frank C.; Wilkinson, Keith D.; DeShazer, David

    2009-01-01

    Burkholderia mallei, a category B biothreat agent, is a facultative intracellular pathogen that causes the zoonotic disease glanders. The B. mallei VirAG two-component regulatory system activates the transcription of ∼60 genes, including a large virulence gene cluster encoding a type VI secretion system (T6SS). The B. mallei tssM gene encodes a putative ubiquitin-specific protease that is physically linked to, and transcriptionally coregulated with, the T6SS gene cluster. Mass spectrometry and immunoblot analysis demonstrated that TssM was secreted in a virAG-dependent manner in vitro. Surprisingly, the T6SS was found to be dispensable for the secretion of TssM. The C-terminal half of TssM, which contains Cys and His box motifs conserved in eukaryotic deubiquitinases, was purified and biochemically characterized. Recombinant TssM hydrolyzed multiple ubiquitinated substrates and the cysteine at position 102 was critical for enzymatic activity. The tssM gene was expressed within 1 h after uptake of B. mallei into RAW 264.7 murine macrophages, suggesting that the TssM deubiquitinase is produced in this intracellular niche. Although the physiological substrate(s) is currently unknown, the TssM deubiquitinase may provide B. mallei a selective advantage in the intracellular environment during infection. PMID:19168747

  19. Burkholderia vaccines: are we moving forward?

    Science.gov (United States)

    Choh, Leang-Chung; Ong, Guang-Han; Vellasamy, Kumutha M.; Kalaiselvam, Kaveena; Kang, Wen-Tyng; Al-Maleki, Anis R.; Mariappan, Vanitha; Vadivelu, Jamuna

    2013-01-01

    The genus Burkholderia consists of diverse species which includes both “friends” and “foes.” Some of the “friendly” Burkholderia spp. are extensively used in the biotechnological and agricultural industry for bioremediation and biocontrol. However, several members of the genus including B. pseudomallei, B. mallei, and B. cepacia, are known to cause fatal disease in both humans and animals. B. pseudomallei and B. mallei are the causative agents of melioidosis and glanders, respectively, while B. cepacia infection is lethal to cystic fibrosis (CF) patients. Due to the high rate of infectivity and intrinsic resistance to many commonly used antibiotics, together with high mortality rate, B. mallei and B. pseudomallei are considered to be potential biological warfare agents. Treatments of the infections caused by these bacteria are often unsuccessful with frequent relapse of the infection. Thus, we are at a crucial stage of the need for Burkholderia vaccines. Although the search for a prophylactic therapy candidate continues, to date development of vaccines has not advanced beyond research to human clinical trials. In this article, we review the current research on development of safe vaccines with high efficacy against B. pseudomallei, B. mallei, and B. cepacia. It can be concluded that further research will enable elucidation of the potential benefits and risks of Burkholderia vaccines. PMID:23386999

  20. Antibiotic resistance in Burkholderia species.

    Science.gov (United States)

    Rhodes, Katherine A; Schweizer, Herbert P

    2016-09-01

    The genus Burkholderia comprises metabolically diverse and adaptable Gram-negative bacteria, which thrive in often adversarial environments. A few members of the genus are prominent opportunistic pathogens. These include Burkholderia mallei and Burkholderia pseudomallei of the B. pseudomallei complex, which cause glanders and melioidosis, respectively. Burkholderia cenocepacia, Burkholderia multivorans, and Burkholderia vietnamiensis belong to the Burkholderia cepacia complex and affect mostly cystic fibrosis patients. Infections caused by these bacteria are difficult to treat because of significant antibiotic resistance. The first line of defense against antimicrobials in Burkholderia species is the outer membrane penetration barrier. Most Burkholderia contain a modified lipopolysaccharide that causes intrinsic polymyxin resistance. Contributing to reduced drug penetration are restrictive porin proteins. Efflux pumps of the resistance nodulation cell division family are major players in Burkholderia multidrug resistance. Third and fourth generation β-lactam antibiotics are seminal for treatment of Burkholderia infections, but therapeutic efficacy is compromised by expression of several β-lactamases and ceftazidime target mutations. Altered DNA gyrase and dihydrofolate reductase targets cause fluoroquinolone and trimethoprim resistance, respectively. Although antibiotic resistance hampers therapy of Burkholderia infections, the characterization of resistance mechanisms lags behind other non-enteric Gram-negative pathogens, especially ESKAPE bacteria such as Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Biosafety level 3 facility: essential infrastructure in biodefense strategy in the Republic of Croatia

    International Nuclear Information System (INIS)

    Cvetko Krajinovic, L.; Markotic, A.

    2009-01-01

    Wide spectrum of microorganisms nowadays present serious health risks to humans and animals and their potential for use as biological weapons has become an important concern for governments and responsible authorities. This has resulted in the implementation of measures (known as biodefense) directed toward containment of potentially harmful biological agents with the purpose to reduce or eliminate hazards to laboratory workers, other persons, and the outside environment. Many of such pathogens are dangerous pathogens which request biosafety level 3 (BSL-3) facility for research and management. Biosafety level 3 comprises the combinations of standard and special microbiological laboratory practices and techniques, safety equipment, and laboratory facilities recommended for work with indigenous or exotic agents that may cause serious or potentially lethal disease through inhalation route exposure. Croatia is endemic for many of these threatening pathogens/diseases (e.g. tularemia, pulmonary and non-pulmonary tuberculosis, brucellosis, Q fever, glanders, melioidosis, typhoid fever, viral hemorrhagic fevers, hepatitis B and C, HIV etc.). Its strategic geographic position and the overall world rise of international trade and travel unlocks the possibility for importing some new microorganisms or even occurrence of an outbreak of totally unknown infectious origin. We, also, cannot exclude the possibility of the so called deliberately emerging microbes used in intentional bioterrorist purposes. However, it is obvious that Croatia needs infrastructure and well trained human capacities on biosafety level 3 to cope with incoming public health challenges and threats. The fundamental objective of the laboratory under which dangerous agents can safely be handled, is surveillance and quick response, as a key elements in controlling of scenarios referred to above. For that purpose, the first BSL-3 facility in Croatia is in the final phase of its reconstruction at the University

  2. Biosafety level 3 facility: essential infrastructure in biodefense strategy in the Republic of Croatia

    Energy Technology Data Exchange (ETDEWEB)

    Cvetko Krajinovic, L; Markotic, A [University Hospital for Infectious Diseases Dr Fran Mihaljevic, Zagreb (Croatia)

    2009-07-01

    Wide spectrum of microorganisms nowadays present serious health risks to humans and animals and their potential for use as biological weapons has become an important concern for governments and responsible authorities. This has resulted in the implementation of measures (known as biodefense) directed toward containment of potentially harmful biological agents with the purpose to reduce or eliminate hazards to laboratory workers, other persons, and the outside environment. Many of such pathogens are dangerous pathogens which request biosafety level 3 (BSL-3) facility for research and management. Biosafety level 3 comprises the combinations of standard and special microbiological laboratory practices and techniques, safety equipment, and laboratory facilities recommended for work with indigenous or exotic agents that may cause serious or potentially lethal disease through inhalation route exposure. Croatia is endemic for many of these threatening pathogens/diseases (e.g. tularemia, pulmonary and non-pulmonary tuberculosis, brucellosis, Q fever, glanders, melioidosis, typhoid fever, viral hemorrhagic fevers, hepatitis B and C, HIV etc.). Its strategic geographic position and the overall world rise of international trade and travel unlocks the possibility for importing some new microorganisms or even occurrence of an outbreak of totally unknown infectious origin. We, also, cannot exclude the possibility of the so called deliberately emerging microbes used in intentional bioterrorist purposes. However, it is obvious that Croatia needs infrastructure and well trained human capacities on biosafety level 3 to cope with incoming public health challenges and threats. The fundamental objective of the laboratory under which dangerous agents can safely be handled, is surveillance and quick response, as a key elements in controlling of scenarios referred to above. For that purpose, the first BSL-3 facility in Croatia is in the final phase of its reconstruction at the University

  3. Combining Functional and Structural Genomics to Sample the Essential Burkholderia Structome

    Science.gov (United States)

    Baugh, Loren; Gallagher, Larry A.; Patrapuvich, Rapatbhorn; Clifton, Matthew C.; Gardberg, Anna S.; Edwards, Thomas E.; Armour, Brianna; Begley, Darren W.; Dieterich, Shellie H.; Dranow, David M.; Abendroth, Jan; Fairman, James W.; Fox, David; Staker, Bart L.; Phan, Isabelle; Gillespie, Angela; Choi, Ryan; Nakazawa-Hewitt, Steve; Nguyen, Mary Trang; Napuli, Alberto; Barrett, Lynn; Buchko, Garry W.; Stacy, Robin; Myler, Peter J.; Stewart, Lance J.; Manoil, Colin; Van Voorhis, Wesley C.

    2013-01-01

    Background The genus Burkholderia includes pathogenic gram-negative bacteria that cause melioidosis, glanders, and pulmonary infections of patients with cancer and cystic fibrosis. Drug resistance has made development of new antimicrobials critical. Many approaches to discovering new antimicrobials, such as structure-based drug design and whole cell phenotypic screens followed by lead refinement, require high-resolution structures of proteins essential to the parasite. Methodology/Principal Findings We experimentally identified 406 putative essential genes in B. thailandensis, a low-virulence species phylogenetically similar to B. pseudomallei, the causative agent of melioidosis, using saturation-level transposon mutagenesis and next-generation sequencing (Tn-seq). We selected 315 protein products of these genes based on structure-determination criteria, such as excluding very large and/or integral membrane proteins, and entered them into the Seattle Structural Genomics Center for Infection Disease (SSGCID) structure determination pipeline. To maximize structural coverage of these targets, we applied an “ortholog rescue” strategy for those producing insoluble or difficult to crystallize proteins, resulting in the addition of 387 orthologs (or paralogs) from seven other Burkholderia species into the SSGCID pipeline. This structural genomics approach yielded structures from 31 putative essential targets from B. thailandensis, and 25 orthologs from other Burkholderia species, yielding an overall structural coverage for 49 of the 406 essential gene families, with a total of 88 depositions into the Protein Data Bank. Of these, 25 proteins have properties of a potential antimicrobial drug target i.e., no close human homolog, part of an essential metabolic pathway, and a deep binding pocket. We describe the structures of several potential drug targets in detail. Conclusions/Significance This collection of structures, solubility and experimental essentiality data

  4. Combining functional and structural genomics to sample the essential Burkholderia structome.

    Directory of Open Access Journals (Sweden)

    Loren Baugh

    Full Text Available The genus Burkholderia includes pathogenic gram-negative bacteria that cause melioidosis, glanders, and pulmonary infections of patients with cancer and cystic fibrosis. Drug resistance has made development of new antimicrobials critical. Many approaches to discovering new antimicrobials, such as structure-based drug design and whole cell phenotypic screens followed by lead refinement, require high-resolution structures of proteins essential to the parasite.We experimentally identified 406 putative essential genes in B. thailandensis, a low-virulence species phylogenetically similar to B. pseudomallei, the causative agent of melioidosis, using saturation-level transposon mutagenesis and next-generation sequencing (Tn-seq. We selected 315 protein products of these genes based on structure-determination criteria, such as excluding very large and/or integral membrane proteins, and entered them into the Seattle Structural Genomics Center for Infection Disease (SSGCID structure determination pipeline. To maximize structural coverage of these targets, we applied an "ortholog rescue" strategy for those producing insoluble or difficult to crystallize proteins, resulting in the addition of 387 orthologs (or paralogs from seven other Burkholderia species into the SSGCID pipeline. This structural genomics approach yielded structures from 31 putative essential targets from B. thailandensis, and 25 orthologs from other Burkholderia species, yielding an overall structural coverage for 49 of the 406 essential gene families, with a total of 88 depositions into the Protein Data Bank. Of these, 25 proteins have properties of a potential antimicrobial drug target i.e., no close human homolog, part of an essential metabolic pathway, and a deep binding pocket. We describe the structures of several potential drug targets in detail.This collection of structures, solubility and experimental essentiality data provides a resource for development of drugs against

  5. Combining functional and structural genomics to sample the essential Burkholderia structome.

    Science.gov (United States)

    Baugh, Loren; Gallagher, Larry A; Patrapuvich, Rapatbhorn; Clifton, Matthew C; Gardberg, Anna S; Edwards, Thomas E; Armour, Brianna; Begley, Darren W; Dieterich, Shellie H; Dranow, David M; Abendroth, Jan; Fairman, James W; Fox, David; Staker, Bart L; Phan, Isabelle; Gillespie, Angela; Choi, Ryan; Nakazawa-Hewitt, Steve; Nguyen, Mary Trang; Napuli, Alberto; Barrett, Lynn; Buchko, Garry W; Stacy, Robin; Myler, Peter J; Stewart, Lance J; Manoil, Colin; Van Voorhis, Wesley C

    2013-01-01

    The genus Burkholderia includes pathogenic gram-negative bacteria that cause melioidosis, glanders, and pulmonary infections of patients with cancer and cystic fibrosis. Drug resistance has made development of new antimicrobials critical. Many approaches to discovering new antimicrobials, such as structure-based drug design and whole cell phenotypic screens followed by lead refinement, require high-resolution structures of proteins essential to the parasite. We experimentally identified 406 putative essential genes in B. thailandensis, a low-virulence species phylogenetically similar to B. pseudomallei, the causative agent of melioidosis, using saturation-level transposon mutagenesis and next-generation sequencing (Tn-seq). We selected 315 protein products of these genes based on structure-determination criteria, such as excluding very large and/or integral membrane proteins, and entered them into the Seattle Structural Genomics Center for Infection Disease (SSGCID) structure determination pipeline. To maximize structural coverage of these targets, we applied an "ortholog rescue" strategy for those producing insoluble or difficult to crystallize proteins, resulting in the addition of 387 orthologs (or paralogs) from seven other Burkholderia species into the SSGCID pipeline. This structural genomics approach yielded structures from 31 putative essential targets from B. thailandensis, and 25 orthologs from other Burkholderia species, yielding an overall structural coverage for 49 of the 406 essential gene families, with a total of 88 depositions into the Protein Data Bank. Of these, 25 proteins have properties of a potential antimicrobial drug target i.e., no close human homolog, part of an essential metabolic pathway, and a deep binding pocket. We describe the structures of several potential drug targets in detail. This collection of structures, solubility and experimental essentiality data provides a resource for development of drugs against infections and diseases

  6. Iron Acquisition Mechanisms and Their Role in the Virulence of Burkholderia Species

    Science.gov (United States)

    Butt, Aaron T.; Thomas, Mark S.

    2017-01-01

    Burkholderia is a genus within the β-Proteobacteriaceae that contains at least 90 validly named species which can be found in a diverse range of environments. A number of pathogenic species occur within the genus. These include Burkholderia cenocepacia and Burkholderia multivorans, opportunistic pathogens that can infect the lungs of patients with cystic fibrosis, and are members of the Burkholderia cepacia complex (Bcc). Burkholderia pseudomallei is also an opportunistic pathogen, but in contrast to Bcc species it causes the tropical human disease melioidosis, while its close relative Burkholderia mallei is the causative agent of glanders in horses. For these pathogens to survive within a host and cause disease they must be able to acquire iron. This chemical element is essential for nearly all living organisms due to its important role in many enzymes and metabolic processes. In the mammalian host, the amount of accessible free iron is negligible due to the low solubility of the metal ion in its higher oxidation state and the tight binding of this element by host proteins such as ferritin and lactoferrin. As with other pathogenic bacteria, Burkholderia species have evolved an array of iron acquisition mechanisms with which to capture iron from the host environment. These mechanisms include the production and utilization of siderophores and the possession of a haem uptake system. Here, we summarize the known mechanisms of iron acquisition in pathogenic Burkholderia species and discuss the evidence for their importance in the context of virulence and the establishment of infection in the host. We have also carried out an extensive bioinformatic analysis to identify which siderophores are produced by each Burkholderia species that is pathogenic to humans. PMID:29164069

  7. Identification of Burkholderia mallei and Burkholderia pseudomallei adhesins for human respiratory epithelial cells

    Directory of Open Access Journals (Sweden)

    Hogan Robert J

    2010-09-01

    Full Text Available Abstract Background Burkholderia pseudomallei and Burkholderia mallei cause the diseases melioidosis and glanders, respectively. A well-studied aspect of pathogenesis by these closely-related bacteria is their ability to invade and multiply within eukaryotic cells. In contrast, the means by which B. pseudomallei and B. mallei adhere to cells are poorly defined. The purpose of this study was to identify adherence factors expressed by these organisms. Results Comparative sequence analyses identified a gene product in the published genome of B. mallei strain ATCC23344 (locus # BMAA0649 that resembles the well-characterized Yersinia enterocolitica autotransporter adhesin YadA. The gene encoding this B. mallei protein, designated boaA, was expressed in Escherichia coli and shown to significantly increase adherence to human epithelial cell lines, specifically HEp2 (laryngeal cells and A549 (type II pneumocytes, as well as to cultures of normal human bronchial epithelium (NHBE. Consistent with these findings, disruption of the boaA gene in B. mallei ATCC23344 reduced adherence to all three cell types by ~50%. The genomes of the B. pseudomallei strains K96243 and DD503 were also found to contain boaA and inactivation of the gene in DD503 considerably decreased binding to monolayers of HEp2 and A549 cells and to NHBE cultures. A second YadA-like gene product highly similar to BoaA (65% identity was identified in the published genomic sequence of B. pseudomallei strain K96243 (locus # BPSL1705. The gene specifying this protein, termed boaB, appears to be B. pseudomallei-specific. Quantitative attachment assays demonstrated that recombinant E. coli expressing BoaB displayed greater binding to A549 pneumocytes, HEp2 cells and NHBE cultures. Moreover, a boaB mutant of B. pseudomallei DD503 showed decreased adherence to these respiratory cells. Additionally, a B. pseudomallei strain lacking expression of both boaA and boaB was impaired in its ability to

  8. Equine infectious anemia prevalence in feral donkeys from Northeast Brazil.

    Science.gov (United States)

    Oliveira, Fernanda G; Cook, R Frank; Naves, João H F; Oliveira, Cairo H S; Diniz, Rejane S; Freitas, Francisco J C; Lima, Joseney M; Sakamoto, Sidnei M; Leite, Rômulo C; Issel, Charles J; Reis, Jenner K P

    2017-05-01

    Equine infectious anemia virus (EIAV) is an important cause of morbidity and mortality throughout the world. Although the virus infects all members of the Equidae the vast majority of studies have been conducted in horses (Equus caballus) with comparatively little information available for other equid species. Brazil has one of the most abundant donkey (E. asinus) populations of any nation although the economic importance of these animals is declining as transportation becomes increasingly mechanized. As a result, considerable numbers of donkeys especially in the Northeast of the country have been released and allowed pursue an almost feral existence. Consequently, this large and growing population constitutes a significant risk as a reservoir for the maintenance and transmission of important equine infectious diseases such as glanders and equine arteritis virus in addition to EIAV. This study examines the prevalence of EIA in a semi-wild donkey population from Mossoró city, in Northeast Brazil, using AGID followed by cELISA, rgp90 ELISA and immunoblot (IB). Serum samples were collected from 367 donkeys without obvious EIA clinical signs. Subsequent testing revealed seropositive rates of 1.6% (6/367) in officially approved AGID tests, 3.3% (12/367) in cELISA and 14.4% (53/367) in the rgp90 ELISA. However, 88.7% (47/53) of the rgp90 ELISA positive samples were almost certainly false reactions because they failed to react with two or more antigens in IB. Consequently, the rpg90 ELISA has a similar sensitivity to AGID with donkey serum samples. Such high false positive rates have not been observed previously with serum samples from horses. Another highly significant finding is that 56.9% (33/58) of the donkey serum samples tested in IB had reactivity to EIAV p26 only. Although this could result from recent infection with the virus, it has been found that in some equids p26 only reactivity persists for extensive periods of time suggesting exposure to antigens

  9. Selected Abstracts of the 2nd Congress of joint European Neonatal Societies (jENS 2017; Venice (Italy; October 31-November 4, 2017; Session "Neonatal Infectious Diseases/Immunology"

    Directory of Open Access Journals (Sweden)

    --- Various Authors

    2017-10-01

    . Esaiassen, L. Juvet, J. van den Anker, C. KlingenbergABS 7. EFFECT OF CATHETER DWELL TIME ON RISK OF CENTRAL LINE ASSOCIATED BLOOD STREAM INFECTION IN NEONATES IN A DEVELOPING HEALTHCARE SYSTEM • S.B. Chaaya, R. Durmush, C.P.H. IbrahimABS 8. 20 YEARS OF EARLY NEONATAL SEPSIS DUE TO E. COLI: SHOULD WE CHANGE EMPIRICAL ANTIBIOTIC THERAPY? • M.C. Céspedes, N. Mendoza, M. Balasch, S. Gonzalez, A. Andreu, M.A. Frick, M.A. Linde, J.J. Gonzalez, P. SolerABS 9. COMPARISON OF THE INCIDENCE, CLINICAL FEATURES AND OUTCOMES OF INVASIVE CANDIDIASIS IN CHILDREN AND NEONATES • J.F. Hsu, S.M. Chu, M.H. TsaiABS 10. HAND HYGIENE COMPLIANCE IN NICU; AN OBSERVATIONAL STUDY • A. Gürol, K.Ş. Tekgündüz, S. Polat, S. Ejder Tekgündüz, S. Ejder ApayABS 11. NEONATAL SEPTIC ARTHRITIS IN NICU • F. Krasniqi, I. Krasniqi, X. Gojnovci, H. JashariABS 12. CURRENT PRACTICE AND NEONATAL STAFF’S KNOWLEDGE ON ANTIBIOTIC USE AND ANTIBIOTIC STEWARDSHIP PROGRAMMES ACROSS TWO NEONATAL NETWORKS • M. Scott, M. Carpenter, P. Satodia, P. NathABS 13. S100-ALARMIN-INDUCED INNATE IMMUNE PROGRAMMING PROTECTS NEWBORN INFANTS FROM SEPSIS • S. Pirr, T. Ulas, B. Fehlhaber, M.S. Bickes, T.G. Loof, T. Vogl, L. Mellinger, A.S. Heinemann, J. Burgmann, J. Schöning, S. Schreek, S. Pfeifer, F. Reuner, L. Völlger, M. Stanulla, M. von Köckritz-Blickwede, S. Glander, K. Barczyk-Kahlert, C.S. von Kaisenberg, J. Friesenhagen, L. Fischer-Riepe, S. Zenker, J.L. Schultze, J. Roth, D. ViemannABS 14. HUMAN BREAST MILK SUPPLIES NEWBORN INFANTS WITH HIGH AMOUNTS OF S100-ALARMINS PROTECTING FROM SEPSIS • S. Pirr, M. Richter, J. Pagel, C. Härtel, B. Fehlhaber, J. Roth, T. Vogl, D. ViemannABS 15. PREDICTORS OF MORTALITY IN NEONATAL SEPSIS • G.I. Sarosa, R. RahimABS 16. DIAGNOSTIC VALUE LYMPHOCYTE TO NEUTROPHIL RATIO IN PRETERM INFANTS WITH LATE ONSET SEPSIS • S.A. Ozdemir, E.A. Ozer, O. Ilhan, S. SutcuogluABS 17. C-REACTIVE PROTEIN: AN ADJUNCT OR DILEMMA FOR DIAGNOSIS OF NEONATAL MENINGITIS • F. Yasin, S