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Sample records for burkholderia cenocepacia strain

  1. Host evasion by Burkholderia cenocepacia

    Directory of Open Access Journals (Sweden)

    Shyamala eGanesan

    2012-01-01

    Full Text Available Burkholderia cenocepacia is an opportunistic respiratory pathogen of individuals with cystic fibrosis (CF. It is one of the highly transmissible species of Burkholderia cepacia complex and very resistant to almost all the antibiotics. Approximately 1/3rd of B. cenocepacia infected CF patients go on to develop fatal ‘cepacia syndrome’. During the last two decades, substantial progress has been made with regards to evasion of host innate defense mechanisms by B. cenocepacia. Almost all strains of B. cenocepacia has capacity to survive and replicate intracellularly in both airway epithelial cells and macrophages, which are primary centennials of the lung and play a pivotal role in clearance of infecting bacteria. Some strains of B. cenocepaica, which express cable pili and the associated 22kDa adhesin are also capable of transmigrating across airway epithelium and persist in mouse models of infection. In this review, we will discuss how this type of interaction between B. cenocepacia and host may lead to persistence of bacteria and contribute to lung inflammation in CF patients.

  2. Identification of Burkholderia cenocepacia strain H111 virulence factors using nonmammalian infection hosts

    DEFF Research Database (Denmark)

    Schwager, Stephan; Agnoli, Kirsty; Köthe, Manuela

    2013-01-01

    Burkholderia cenocepacia H111, a strain isolated from a cystic fibrosis patient, has been shown to effectively kill the nematode Caenorhabditis elegans. We used the C. elegans model of infection to screen a mini-Tn5 mutant library of B. cenocepacia H111 for attenuated virulence....... Of the approximately 5,500 B. cenocepacia H111 random mini-Tn5 insertion mutants that were screened, 22 showed attenuated virulence in C. elegans. Except for the quorum-sensing regulator cepR, none of the mutated genes coded for the biosynthesis of classical virulence factors such as extracellular proteases...... or siderophores. Instead, the mutants contained insertions in metabolic and regulatory genes. Mutants attenuated in virulence in the C. elegans infection model were also tested in the Drosophila melanogaster pricking model, and those also attenuated in this model were further tested in Galleria mellonella. Six...

  3. Environmental Burkholderia cenocepacia Strain Enhances Fitness by Serial Passages during Long-Term Chronic Airways Infection in Mice

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    Alessandra Bragonzi

    2017-11-01

    Full Text Available Burkholderia cenocepacia is an important opportunistic pathogen in cystic fibrosis (CF patients, and has also been isolated from natural environments. In previous work, we explored the virulence and pathogenic potential of environmental B. cenocepacia strains and demonstrated that they do not differ from clinical strains in some pathogenic traits. Here, we investigated the ability of the environmental B. cenocepacia Mex1 strain, isolated from the maize rhizosphere, to persist and increase its virulence after serial passages in a mouse model of chronic infection. B. cenocepacia Mex1 strain, belonging to the recA lineage IIIA, was embedded in agar beads and challenged into the lung of C57Bl/6 mice. The mice were sacrificed after 28 days from infection and their lungs were tested for bacterial loads. Agar beads containing the pool of B. cenocepacia colonies from the four sequential passages were used to infect the mice. The environmental B. cenocepacia strain showed a low incidence of chronic infection after the first passage; after the second, third and fourth passages in mice, its ability to establish chronic infection increased significantly and progressively up to 100%. Colonial morphology analysis and genetic profiling of the Mex1-derived clones recovered after the fourth passage from infected mice revealed that they were indistinguishable from the challenged strain both at phenotypic and genetic level. By testing the virulence of single clones in the Galleria mellonella infection model, we found that two Mex1-derived clones significantly increased their pathogenicity compared to the parental Mex1 strain and behaved similarly to the clinical and epidemic B. cenocepacia LMG16656T. Our findings suggest that serial passages of the environmental B. cenocepacia Mex1 strain in mice resulted in an increased ability to determine chronic lung infection and the appearance of clonal variants with increased virulence in non-vertebrate hosts.

  4. Burkholderia cenocepacia Vaginal Infection in Patient with Smoldering Myeloma and Chronic Hepatitis C

    OpenAIRE

    Petrucca, Andrea; Cipriani, Paola; Sessa, Rosa; Teggi, Antonella; Pustorino, Rosalia; Santapaola, Daniela; Nicoletti, Mauro

    2004-01-01

    We report a case of a vaginal infection caused by a strain of Burkholderia cenocepacia. The strain was isolated from vaginal swab specimens from a 68-year-old woman with smoldering myeloma and chronic hepatitis C virus infection who was hospitalized for abdominal abscess. Treatment with piperacillin/tazobactam eliminated B. cenocepacia infection and vaginal symptoms.

  5. Deciphering the role of RND efflux transporters in Burkholderia cenocepacia.

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    Silvia Bazzini

    Full Text Available Burkholderia cenocepacia J2315 is representative of a highly problematic group of cystic fibrosis (CF pathogens. Eradication of B. cenocepacia is very difficult with the antimicrobial therapy being ineffective due to its high resistance to clinically relevant antimicrobial agents and disinfectants. RND (Resistance-Nodulation-Cell Division efflux pumps are known to be among the mediators of multidrug resistance in gram-negative bacteria. Since the significance of the 16 RND efflux systems present in B. cenocepacia (named RND-1 to -16 has been only partially determined, the aim of this work was to analyze mutants of B. cenocepacia strain J2315 impaired in RND-4 and RND-9 efflux systems, and assess their role in the efflux of toxic compounds. The transcriptomes of mutants deleted individually in RND-4 and RND-9 (named D4 and D9, and a double-mutant in both efflux pumps (named D4-D9, were compared to that of the wild-type B. cenocepacia using microarray analysis. Microarray data were confirmed by qRT-PCR, phenotypic experiments, and by Phenotype MicroArray analysis. The data revealed that RND-4 made a significant contribution to the antibiotic resistance of B. cenocepacia, whereas RND-9 was only marginally involved in this process. Moreover, the double mutant D4-D9 showed a phenotype and an expression profile similar to D4. The microarray data showed that motility and chemotaxis-related genes appeared to be up-regulated in both D4 and D4-D9 strains. In contrast, these gene sets were down-regulated or expressed at levels similar to J2315 in the D9 mutant. Biofilm production was enhanced in all mutants. Overall, these results indicate that in B. cenocepacia RND pumps play a wider role than just in drug resistance, influencing additional phenotypic traits important for pathogenesis.

  6. Novel diagnostic PCR assay for Burkholderia cenocepacia epidemic strain ST32 and its utility in monitoring infection in cystic fibrosis patients

    Czech Academy of Sciences Publication Activity Database

    Dědečková, K.; Kalferstová, L.; Strnad, Hynek; Vávrová, J.; Dřevínek, P.

    2013-01-01

    Roč. 12, č. 5 (2013), s. 475-481 ISSN 1569-1993 Institutional support: RVO:68378050 Keywords : Burkholderia cenocepacia * diagnostic PCR * B. cenocepacia ST32 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.820, year: 2013

  7. Antimicrobial Properties of an Oxidizer Produced by Burkholderia cenocepacia P525

    Science.gov (United States)

    A compound with both oxidizing properties and antibiotic properties was extracted and purified from broth cultures of Burkholderia cenocepacia strain P525. A four step purification procedure was used to increase its specific activity ~ 400 fold and to yield a HPLC- UV chromatogram containing a sing...

  8. Efflux-mediated resistance to a benzothiadiazol derivative effective against Burkholderia cenocepacia

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    Viola Camilla eScoffone

    2015-08-01

    Full Text Available Burkholderia cenocepacia is a major concern for people suffering from Cystic Fibrosis as it contributes to serious respiratory tract infections. The lack of drugs effective against this opportunistic pathogen, along with the high level of resistance to multiple antibiotics, render the treatment of these infections particularly difficult.Here a new compound, belonging to the 2,1,3-benzothiadiazol-5-yl family (10126109, with a bactericidal effect and a MIC of 8 µg/ml against B. cenocepacia, is described. The compound is not cytotoxic and effective against B. cenocepacia clinical isolates and members of all the known Burkholderia cepacia complex species.Spontaneous mutants resistant to 10126109 were isolated and mutations in the MerR transcriptional regulator BCAM1948 were identified. In this way, a mechanism of resistance to this new molecule was described, which relies on the overexpression of the RND-9 efflux pump. Indeed, rnd-9 overexpression was confirmed by qRT-PCR, and RND-9 was identified in the membrane fractions of the mutant strains. Moreover, the increase in the MIC values of different drugs in the mutant strains, together with complementation experiments, suggested the involvement of RND-9 in the efflux of 10126109, thus indicating again the central role of efflux transporters in B. cenocepacia drug resistance.

  9. Construction of a large-scale Burkholderia cenocepacia J2315 transposon mutant library

    Science.gov (United States)

    Wong, Yee-Chin; Pain, Arnab; Nathan, Sheila

    2014-09-01

    Burkholderia cenocepacia, a pathogenic member of the Burkholderia cepacia complex (Bcc), has emerged as a significant threat towards cystic fibrosis patients, where infection often leads to the fatal clinical manifestation known as cepacia syndrome. Many studies have investigated the pathogenicity of B. cenocepacia as well as its ability to become highly resistant towards many of the antibiotics currently in use. In addition, studies have also been undertaken to understand the pathogen's capacity to adapt and survive in a broad range of environments. Transposon based mutagenesis has been widely used in creating insertional knock-out mutants and coupled with recent advances in sequencing technology, robust tools to study gene function in a genome-wide manner have been developed based on the assembly of saturated transposon mutant libraries. In this study, we describe the construction of a large-scale library of B. cenocepacia transposon mutants. To create transposon mutants of B. cenocepacia strain J2315, electrocompetent bacteria were electrotransformed with the EZ-Tn5 transposome. Tetracyline resistant colonies were harvested off selective agar and pooled. Mutants were generated in multiple batches with each batch consisting of ˜20,000 to 40,000 mutants. Transposon insertion was validated by PCR amplification of the transposon region. In conclusion, a saturated B. cenocepacia J2315 transposon mutant library with an estimated total number of 500,000 mutants was successfully constructed. This mutant library can now be further exploited as a genetic tool to assess the function of every gene in the genome, facilitating the discovery of genes important for bacterial survival and adaptation, as well as virulence.

  10. Efflux pump genes of the resistance-nodulation-division family in Burkholderia cenocepacia genome

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    Manina Giulia

    2006-07-01

    Full Text Available Abstract Background Burkholderia cenocepacia is recognized as opportunistic pathogen that can cause lung infections in cystic fibrosis patients. A hallmark of B. cenocepacia infections is the inability to eradicate the organism because of multiple intrinsic antibiotic resistance. As Resistance-Nodulation-Division (RND efflux systems are responsible for much of the intrinsic multidrug resistance in Gram-negative bacteria, this study aims to identify RND genes in the B. cenocepacia genome and start to investigate their involvement into antimicrobial resistance. Results Genome analysis and homology searches revealed 14 open reading frames encoding putative drug efflux pumps belonging to RND family in B. cenocepacia J2315 strain. By reverse transcription (RT-PCR analysis, it was found that orf3, orf9, orf11, and orf13 were expressed at detectable levels, while orf10 appeared to be weakly expressed in B. cenocepacia. Futhermore, orf3 was strongly induced by chloramphenicol. The orf2 conferred resistance to fluoroquinolones, tetraphenylphosphonium, streptomycin, and ethidium bromide when cloned and expressed in Escherichia coli KAM3, a strain lacking the multidrug efflux pump AcrAB. The orf2-overexpressing E. coli also accumulate low concentrations of ethidium bromide, which was restored to wild type level in the presence of CCCP, an energy uncoupler altering the energy of the drug efflux pump. Conclusion The 14 RND pumps gene we have identified in the genome of B. cenocepacia suggest that active efflux could be a major mechanism underlying antimicrobial resistance in this microorganism. We have characterized the ORF2 pump, one of these 14 potential RND efflux systems. Its overexpression in E. coli conferred resistance to several antibiotics and to ethidium bromide but it remains to be determined if this pump play a significant role in the antimicrobial intrinsic resistance of B. cenocepacia. The characterization of antibiotic efflux pumps in B

  11. NtrC-dependent control of exopolysaccharide synthesis and motility in Burkholderia cenocepacia H111.

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    Yilei Liu

    Full Text Available Burkholderia cenocepacia is a versatile opportunistic pathogen that survives in a wide variety of environments, which can be limited in nutrients such as nitrogen. We have previously shown that the sigma factor σ54 is involved in the control of nitrogen assimilation and virulence in B. cenocepacia H111. In this work, we investigated the role of the σ54 enhancer binding protein NtrC in response to nitrogen limitation and in the pathogenicity of H111. Of 95 alternative nitrogen sources tested the ntrC showed defects in the utilisation of nitrate, urea, L-citrulline, acetamide, DL-lactamide, allantoin and parabanic acid. RNA-Seq and phenotypic analyses of an ntrC mutant strain showed that NtrC positively regulates two important phenotypic traits: exopolysaccharide (EPS production and motility. However, the ntrC mutant was not attenuated in C. elegans virulence.

  12. Differential roles of RND efflux pumps in antimicrobial drug resistance of sessile and planktonic Burkholderia cenocepacia cells.

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    Buroni, Silvia; Matthijs, Nele; Spadaro, Francesca; Van Acker, Heleen; Scoffone, Viola C; Pasca, Maria Rosalia; Riccardi, Giovanna; Coenye, Tom

    2014-12-01

    Burkholderia cenocepacia is notorious for causing respiratory tract infections in people with cystic fibrosis. Infections with this organism are particularly difficult to treat due to its high level of intrinsic resistance to most antibiotics. Multidrug resistance in B. cenocepacia can be ascribed to different mechanisms, including the activity of efflux pumps and biofilm formation. In the present study, the effects of deletion of the 16 operons encoding resistance-nodulation-cell division (RND)-type efflux pumps in B. cenocepacia strain J2315 were investigated by determining the MICs of various antibiotics and by investigating the antibiofilm effect of these antibiotics. Finally, the expression levels of selected RND genes in treated and untreated cultures were investigated using reverse transcriptase quantitative PCR (RT-qPCR). Our data indicate that the RND-3 and RND-4 efflux pumps are important for resistance to various antimicrobial drugs (including tobramycin and ciprofloxacin) in planktonic B. cenocepacia J2315 populations, while the RND-3, RND-8, and RND-9 efflux systems protect biofilm-grown cells against tobramycin. The RND-8 and RND-9 efflux pumps are not involved in ciprofloxacin resistance. Results from the RT-qPCR experiments on the wild-type strain B. cenocepacia J2315 suggest that there is little regulation at the level of mRNA expression for these efflux pumps under the conditions tested. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  13. Genetic Determinants Associated With in Vivo Survival of Burkholderia cenocepacia in the Caenorhabditis elegans Model

    KAUST Repository

    Wong, Yee-Chin

    2018-05-29

    A Burkholderia cenocepacia infection usually leads to reduced survival and fatal cepacia syndrome in cystic fibrosis patients. The identification of B. cenocepacia essential genes for in vivo survival is key to designing new anti-infectives therapies. We used the Transposon-Directed Insertion Sequencing (TraDIS) approach to identify genes required for B. cenocepacia survival in the model infection host, Caenorhabditis elegans. A B. cenocepacia J2315 transposon pool of ∼500,000 mutants was used to infect C. elegans. We identified 178 genes as crucial for B. cenocepacia survival in the infected nematode. The majority of these genes code for proteins of unknown function, many of which are encoded by the genomic island BcenGI13, while other gene products are involved in nutrient acquisition, general stress responses and LPS O-antigen biosynthesis. Deletion of the glycosyltransferase gene wbxB and a histone-like nucleoid structuring (H-NS) protein-encoding gene (BCAL0154) reduced bacterial accumulation and attenuated virulence in C. elegans. Further analysis using quantitative RT-PCR indicated that BCAL0154 modulates B. cenocepacia pathogenesis via transcriptional regulation of motility-associated genes including fliC, fliG, flhD, and cheB1. This screen has successfully identified genes required for B. cenocepacia survival within the host-associated environment, many of which are potential targets for developing new antimicrobials.

  14. Genetic Determinants Associated With in Vivo Survival of Burkholderia cenocepacia in the Caenorhabditis elegans Model

    KAUST Repository

    Wong, Yee-Chin; Abd El Ghany, Moataz; Ghazzali, Raeece N. M.; Yap, Soon-Joo; Hoh, Chee-Choong; Pain, Arnab; Nathan, Sheila

    2018-01-01

    A Burkholderia cenocepacia infection usually leads to reduced survival and fatal cepacia syndrome in cystic fibrosis patients. The identification of B. cenocepacia essential genes for in vivo survival is key to designing new anti-infectives therapies. We used the Transposon-Directed Insertion Sequencing (TraDIS) approach to identify genes required for B. cenocepacia survival in the model infection host, Caenorhabditis elegans. A B. cenocepacia J2315 transposon pool of ∼500,000 mutants was used to infect C. elegans. We identified 178 genes as crucial for B. cenocepacia survival in the infected nematode. The majority of these genes code for proteins of unknown function, many of which are encoded by the genomic island BcenGI13, while other gene products are involved in nutrient acquisition, general stress responses and LPS O-antigen biosynthesis. Deletion of the glycosyltransferase gene wbxB and a histone-like nucleoid structuring (H-NS) protein-encoding gene (BCAL0154) reduced bacterial accumulation and attenuated virulence in C. elegans. Further analysis using quantitative RT-PCR indicated that BCAL0154 modulates B. cenocepacia pathogenesis via transcriptional regulation of motility-associated genes including fliC, fliG, flhD, and cheB1. This screen has successfully identified genes required for B. cenocepacia survival within the host-associated environment, many of which are potential targets for developing new antimicrobials.

  15. Identification of putative noncoding RNA genes in the Burkholderia cenocepacia J2315 genome

    DEFF Research Database (Denmark)

    Coenye, T.; Drevinek, P.; Mahenthiralingam, E.

    2007-01-01

    Noncoding RNA (ncRNA) genes are not involved in the production of mRNA and proteins, but produce transcripts that function directly as structural or regulatory RNAs. In the present study, the presence of ncRNA genes in the genome of Burkholderia cenocepacia J2315 was evaluated by combining...

  16. The temperate Burkholderia phage AP3 of the Peduovirinae shows efficient antimicrobial activity against B. cenocepacia of the IIIA lineage.

    Science.gov (United States)

    Roszniowski, Bartosz; Latka, Agnieszka; Maciejewska, Barbara; Vandenheuvel, Dieter; Olszak, Tomasz; Briers, Yves; Holt, Giles S; Valvano, Miguel A; Lavigne, Rob; Smith, Darren L; Drulis-Kawa, Zuzanna

    2017-02-01

    Burkholderia phage AP3 (vB_BceM_AP3) is a temperate virus of the Myoviridae and the Peduovirinae subfamily (P2likevirus genus). This phage specifically infects multidrug-resistant clinical Burkholderia cenocepacia lineage IIIA strains commonly isolated from cystic fibrosis patients. AP3 exhibits high pairwise nucleotide identity (61.7 %) to Burkholderia phage KS5, specific to the same B. cenocepacia host, and has 46.7-49.5 % identity to phages infecting other species of Burkholderia. The lysis cassette of these related phages has a similar organization (putative antiholin, putative holin, endolysin, and spanins) and shows 29-98 % homology between specific lysis genes, in contrast to Enterobacteria phage P2, the hallmark phage of this genus. The AP3 and KS5 lysis genes have conserved locations and high amino acid sequence similarity. The AP3 bacteriophage particles remain infective up to 5 h at pH 4-10 and are stable at 60 °C for 30 min, but are sensitive to chloroform, with no remaining infective particles after 24 h of treatment. AP3 lysogeny can occur by stable genomic integration and by pseudo-lysogeny. The lysogenic bacterial mutants did not exhibit any significant changes in virulence compared to wild-type host strain when tested in the Galleria mellonella moth wax model. Moreover, AP3 treatment of larvae infected with B. cenocepacia revealed a significant increase (P < 0.0001) in larvae survival in comparison to AP3-untreated infected larvae. AP3 showed robust lytic activity, as evidenced by its broad host range, the absence of increased virulence in lysogenic isolates, the lack of bacterial gene disruption conditioned by bacterial tRNA downstream integration site, and the absence of detected toxin sequences. These data suggest that the AP3 phage is a promising potent agent against bacteria belonging to the most common B. cenocepacia IIIA lineage strains.

  17. The genome of Burkholderia cenocepacia J2315, an epidemic pathogen of cystic fibrosis patients

    DEFF Research Database (Denmark)

    Holden, Matthew T G; Seth-Smith, Helena M B; Crossman, Lisa C

    2009-01-01

    ; the resulting chronic infections are associated with severe declines in lung function and increased mortality rates. B. cenocepacia strain J2315 was isolated from a CF patient and is a member of the epidemic ET12 lineage that originated in Canada or the United Kingdom and spread to Europe. The 8.06-Mb genome...... be pathogenic to both plants and man, J2315 is representative of a lineage of B. cenocepacia rarely isolated from the environment and which spreads between CF patients. Comparative analysis revealed that ca. 21% of the genome is unique in comparison to other strains of B. cenocepacia, highlighting the genomic...... success as an epidemic CF pathogen....

  18. Evaluation of the electron transfer flavoprotein as an antibacterial target in Burkholderia cenocepacia.

    Science.gov (United States)

    Stietz, Maria S; Lopez, Christina; Osifo, Osasumwen; Tolmasky, Marcelo E; Cardona, Silvia T

    2017-10-01

    There are hundreds of essential genes in multidrug-resistant bacterial genomes, but only a few of their products are exploited as antibacterial targets. An example is the electron transfer flavoprotein (ETF), which is required for growth and viability in Burkholderia cenocepacia. Here, we evaluated ETF as an antibiotic target for Burkholderia cepacia complex (Bcc). Depletion of the bacterial ETF during infection of Caenorhabditis elegans significantly extended survival of the nematodes, proving that ETF is essential for survival of B. cenocepacia in this host model. In spite of the arrest in respiration in ETF mutants, the inhibition of etf expression did not increase the formation of persister cells, when treated with high doses of ciprofloxacin or meropenem. To test if etf translation could be inhibited by RNA interference, antisense oligonucleotides that target the etfBA operon were synthesized. One antisense oligonucleotide was effective in inhibiting etfB translation in vitro but not in vivo, highlighting the challenge of reduced membrane permeability for the design of drugs against B. cenocepacia. This work contributes to the validation of ETF of B. cenocepacia as a target for antibacterial therapy and demonstrates the utility of a C. elegans liquid killing assay to validate gene essentiality in an in vivo infection model.

  19. Candidate Essential Genes in Burkholderia cenocepacia J2315 Identified by Genome-Wide TraDIS

    KAUST Repository

    Wong, Yee-Chin

    2016-08-22

    Burkholderia cenocepacia infection often leads to fatal cepacia syndrome in cystic fibrosis patients. However, antibiotic therapy rarely results in complete eradication of the pathogen due to its intrinsic resistance to many clinically available antibiotics. Recent attention has turned to the identification of essential genes as the proteins encoded by these genes may serve as potential targets for development of novel antimicrobials. In this study, we utilized TraDIS (Transposon Directed Insertion-site Sequencing) as a genome-wide screening tool to facilitate the identification of B. cenocepacia genes essential for its growth and viability. A transposon mutant pool consisting of approximately 500,000 mutants was successfully constructed, with more than 400,000 unique transposon insertion sites identified by computational analysis of TraDIS datasets. The saturated library allowed for the identification of 383 genes that were predicted to be essential in B. cenocepacia. We extended the application of TraDIS to identify conditionally essential genes required for in vitro growth and revealed an additional repertoire of 439 genes to be crucial for B. cenocepacia growth under nutrient-depleted conditions. The library of B. cenocepacia mutants can subsequently be subjected to various biologically related conditions to facilitate the discovery of genes involved in niche adaptation as well as pathogenicity and virulence.

  20. Candidate Essential Genes in Burkholderia cenocepacia J2315 Identified by Genome-Wide TraDIS

    KAUST Repository

    Wong, Yee-Chin; Abd El Ghany, Moataz; Naeem, Raeece; Lee, Kok-Wei; Tan, Yung-Chie; Pain, Arnab; Nathan, Sheila

    2016-01-01

    Burkholderia cenocepacia infection often leads to fatal cepacia syndrome in cystic fibrosis patients. However, antibiotic therapy rarely results in complete eradication of the pathogen due to its intrinsic resistance to many clinically available antibiotics. Recent attention has turned to the identification of essential genes as the proteins encoded by these genes may serve as potential targets for development of novel antimicrobials. In this study, we utilized TraDIS (Transposon Directed Insertion-site Sequencing) as a genome-wide screening tool to facilitate the identification of B. cenocepacia genes essential for its growth and viability. A transposon mutant pool consisting of approximately 500,000 mutants was successfully constructed, with more than 400,000 unique transposon insertion sites identified by computational analysis of TraDIS datasets. The saturated library allowed for the identification of 383 genes that were predicted to be essential in B. cenocepacia. We extended the application of TraDIS to identify conditionally essential genes required for in vitro growth and revealed an additional repertoire of 439 genes to be crucial for B. cenocepacia growth under nutrient-depleted conditions. The library of B. cenocepacia mutants can subsequently be subjected to various biologically related conditions to facilitate the discovery of genes involved in niche adaptation as well as pathogenicity and virulence.

  1. Candidate essential genes in Burkholderia cenocepacia J2315 identified by genome-wide TraDIS

    Directory of Open Access Journals (Sweden)

    Yee-Chin Wong

    2016-08-01

    Full Text Available Burkholderia cenocepacia infection often leads to fatal cepacia syndrome in cystic fibrosis patients. However, antibiotic therapy rarely results in complete eradication of the pathogen due to its intrinsic resistance to many clinically available antibiotics. Recent attention has turned to the identification of essential genes as the proteins encoded by these genes may serve as potential targets for development of novel antimicrobials. In this study, we utilized TraDIS (Transposon Directed Insertion-site Sequencing as a genome-wide screening tool to facilitate the identification of B. cenocepacia genes essential for its growth and viability. A transposon mutant pool consisting of approximately 500,000 mutants was successfully constructed, with more than 400,000 unique transposon insertion sites identified by computational analysis of TraDIS datasets. The saturated library allowed for the identification of 383 genes that were predicted to be essential in B. cenocepacia. We extended the application of TraDIS to identify conditionally essential genes required for in vitro growth and revealed an additional repertoire of 439 genes to be crucial for B. cenocepacia growth under nutrient-depleted conditions. The library of B. cenocepacia mutants can subsequently be subjected to various biologically related conditions to facilitate the discovery of genes involved in niche adaptation as well as pathogenicity and virulence.

  2. Burkholderia cenocepacia K56-2 trimeric autotransporter adhesin BcaA binds TNFR1 and contributes to induce airway inflammation.

    Science.gov (United States)

    Mil-Homens, Dalila; Pinto, Sandra N; Matos, Rute G; Arraiano, Cecília; Fialho, Arsenio M

    2017-04-01

    Chronic lung disease caused by persistent bacterial infections is a major cause of morbidity and mortality in patients with cystic fibrosis (CF). CF pathogens acquire antibiotic resistance, overcome host defenses, and impose uncontrolled inflammation that ultimately may cause permanent damage of lungs' airways. Among the multiple CF-associated pathogens, Burkholderia cenocepacia and other Burkholderia cepacia complex bacteria have become prominent contributors of disease progression. Here, we demonstrate that BcaA, a trimeric autotransporter adhesin (TAA) from the epidemic strain B. cenocepacia K56-2, is a tumor necrosis factor receptor 1-interacting protein able to regulate components of the tumor necrosis factor signaling pathway and ultimately leading to a significant production of the proinflammatory cytokine IL-8. Notably, this study is the first to demonstrate that a protein belonging to the TAA family is involved in the induction of the inflammatory response during B. cenocepacia infections, contributing to the success of the pathogen. Moreover, our results reinforce the relevance of the TAA BcaA as a multifunctional protein with a major role in B. cenocepacia virulence. © 2016 John Wiley & Sons Ltd.

  3. Cyanide toxicity to Burkholderia cenocepacia is modulated by polymicrobial communities and environmental factors

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    Steve P. Bernier

    2016-05-01

    Full Text Available Microbes within polymicrobial communities can establish positive and negative interactions that have the potential to influence the overall behaviour of the community. Pseudomonas aeruginosa and species of the Burkholderia cepacia complex (Bcc can co-exist in the lower airways, however several studies have shown that P. aeruginosa can effectively kill the Bcc in vitro, for which hydrogen cyanide was recently proposed to play a critical role. Here we show that modification of the environment (i.e. culture medium, long-term genetic adaptation of P. aeruginosa to the cystic fibrosis (CF lung, or the addition of another bacterial species to the community can alter the sensitivity of Burkholderia cenocepacia to P. aeruginosa toxins. We specifically demonstrate that undefined rich media leads to higher susceptibility of B. cenocepacia to P. aeruginosa toxins like cyanide as compared to a synthetic medium (SCFM, that mimics the CF lung nutritional content. Overall, our study shows that the polymicrobial environment can have profound effects on negative interactions mediated by P. aeruginosa against B. cenocepacia. In fact, evolved P. aeruginosa or the presence of other species such as Staphylococcus aureus can directly abolish the direct competition mediated by cyanide and consequently maintaining a higher level of species diversity within the community.

  4. Effect of nitrofurans and NO generators on biofilm formation by Pseudomonas aeruginosa PAO1 and Burkholderia cenocepacia 370.

    Science.gov (United States)

    Zaitseva, Julia; Granik, Vladimir; Belik, Alexandr; Koksharova, Olga; Khmel, Inessa

    2009-06-01

    Antibacterial drugs in the nitrofuran series, such as nitrofurazone, furazidin, nitrofurantoin and nifuroxazide, as well as the nitric oxide generators sodium nitroprusside and isosorbide mononitrate in concentrations that do not suppress bacterial growth, were shown to increase the capacity of pathogenic bacteria Pseudomonas aeruginosa PAO1 and Burkholderia cenocepacia 370 to form biofilms. At 25-100microg/ml, nitrofurans 2-2.5-fold enhanced biofilm formation of P. aeruginosa PAO1, and NO donors 3-6-fold. For B. cenocepacia 370, the enhancement was 2-5-fold (nitrofurans) and 4.5-fold (sodium nitroprusside), respectively.

  5. Biochemical Characterization of Glutamate Racemase-A New Candidate Drug Target against Burkholderia cenocepacia Infections.

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    Aygun Israyilova

    Full Text Available The greatest obstacle for the treatment of cystic fibrosis patients infected with the Burkholderia species is their intrinsic antibiotic resistance. For this reason, there is a need to develop new effective compounds. Glutamate racemase, an essential enzyme for the biosynthesis of the bacterial cell wall, is an excellent candidate target for the design of new antibacterial drugs. To this aim, we recombinantly produced and characterized glutamate racemase from Burkholderia cenocepacia J2315. From the screening of an in-house library of compounds, two Zn (II and Mn (III 1,3,5-triazapentadienate complexes were found to efficiently inhibit the glutamate racemase activity with IC50 values of 35.3 and 10.0 μM, respectively. Using multiple biochemical approaches, the metal complexes have been shown to affect the enzyme activity by binding to the enzyme-substrate complex and promoting the formation of an inhibited dimeric form of the enzyme. Our results corroborate the value of glutamate racemase as a good target for the development of novel inhibitors against Burkholderia.

  6. Evaluation of combination therapy for Burkholderia cenocepacia lung infection in different in vitro and in vivo models.

    Directory of Open Access Journals (Sweden)

    Freija Van den Driessche

    Full Text Available Burkholderia cenocepacia is an opportunistic pathogen responsible for life-threatening infections in cystic fibrosis patients. B. cenocepacia is extremely resistant towards antibiotics and therapy is complicated by its ability to form biofilms. We investigated the efficacy of an alternative antimicrobial strategy for B. cenocepacia lung infections using in vitro and in vivo models. A screening of the NIH Clinical Collection 1&2 was performed against B. cenocepacia biofilms formed in 96-well microtiter plates in the presence of tobramycin to identify repurposing candidates with potentiator activity. The efficacy of selected hits was evaluated in a three-dimensional (3D organotypic human lung epithelial cell culture model. The in vivo effect was evaluated in the invertebrate Galleria mellonella and in a murine B. cenocepacia lung infection model. The screening resulted in 60 hits that potentiated the activity of tobramycin against B. cenocepacia biofilms, including four imidazoles of which econazole and miconazole were selected for further investigation. However, a potentiator effect was not observed in the 3D organotypic human lung epithelial cell culture model. Combination treatment was also not able to increase survival of infected G. mellonella. Also in mice, there was no added value for the combination treatment. Although potentiators of tobramycin with activity against biofilms of B. cenocepacia were identified in a repurposing screen, the in vitro activity could not be confirmed nor in a more sophisticated in vitro model, neither in vivo. This stresses the importance of validating hits resulting from in vitro studies in physiologically relevant model systems.

  7. The CRP/FNR family protein Bcam1349 is a c-di-GMP effector that regulates biofilm formation in the respiratory pathogen Burkholderia cenocepacia

    DEFF Research Database (Denmark)

    Fazli, Mustafa; O'Connell, Aileen; Nilsson, Martin

    2011-01-01

    Burkholderia cenocepacia is an opportunistic respiratory pathogen that can cause severe infections in immune-compromised individuals and is associated with poor prognosis for patients suffering from cystic fibrosis. The second messenger cyclic diguanosine monophosphate (c-di-GMP) has been shown...... to control a wide range of functions in bacteria, but little is known about these regulatory mechanisms in B. cenocepacia. Here we investigated the role that c-di-GMP plays in the regulation of biofilm formation and virulence in B. cenocepacia. Elevated intracellular levels of c-di-GMP promoted wrinkly...... colony, pellicle and biofilm formation in B. cenocepacia. A screen for transposon mutants unable to respond to elevated levels of c-di-GMP led to the identification of the mutant bcam1349 that did not display increased biofilm and pellicle formation with excessive c-di-GMP levels, and displayed a biofilm...

  8. In-Frame and Unmarked Gene Deletions in Burkholderia cenocepacia via an Allelic Exchange System Compatible with Gateway Technology.

    Science.gov (United States)

    Fazli, Mustafa; Harrison, Joe J; Gambino, Michela; Givskov, Michael; Tolker-Nielsen, Tim

    2015-06-01

    Burkholderia cenocepacia is an emerging opportunistic pathogen causing life-threatening infections in immunocompromised individuals and in patients with cystic fibrosis, which are often difficult, if not impossible, to treat. Understanding the genetic basis of virulence in this emerging pathogen is important for the development of novel treatment regimes. Generation of deletion mutations in genes predicted to encode virulence determinants is fundamental to investigating the mechanisms of pathogenesis. However, there is a lack of appropriate selectable and counterselectable markers for use in B. cenocepacia, making its genetic manipulation problematic. Here we describe a Gateway-compatible allelic exchange system based on the counterselectable pheS gene and the I-SceI homing endonuclease. This system provides efficiency in cloning homology regions of target genes and allows the generation of precise and unmarked gene deletions in B. cenocepacia. As a proof of concept, we demonstrate its utility by deleting the Bcam1349 gene, encoding a cyclic di-GMP (c-di-GMP)-responsive regulator protein important for biofilm formation. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  9. Saturation mutagenesis of a CepR binding site as a means to identify new quorum-regulated promoters in Burkholderia cenocepacia

    Science.gov (United States)

    Burkholderia cenocepacia, an opportunistic pathogen of humans, encodes the CepI and CepR proteins, which resemble the LuxI and LuxR quorum sensing proteins of Vibrio fischeri. CepI directs the synthesis of octanoylhomoserine lactone (OHL), while CepR is an OHL dependent transcription factor. In pr...

  10. Burkholderia cenocepacia type VI secretion system mediates escape of type II secreted proteins into the cytoplasm of infected macrophages.

    Directory of Open Access Journals (Sweden)

    Roberto Rosales-Reyes

    Full Text Available Burkholderia cenocepacia is an opportunistic pathogen that survives intracellularly in macrophages and causes serious respiratory infections in patients with cystic fibrosis. We have previously shown that bacterial survival occurs in bacteria-containing membrane vacuoles (BcCVs resembling arrested autophagosomes. Intracellular bacteria stimulate IL-1β secretion in a caspase-1-dependent manner and induce dramatic changes to the actin cytoskeleton and the assembly of the NADPH oxidase complex onto the BcCV membrane. A Type 6 secretion system (T6SS is required for these phenotypes but surprisingly it is not required for the maturation arrest of the BcCV. Here, we show that macrophages infected with B. cenocepacia employ the NLRP3 inflammasome to induce IL-1β secretion and pyroptosis. Moreover, IL-1β secretion by B. cenocepacia-infected macrophages is suppressed in deletion mutants unable to produce functional Type VI, Type IV, and Type 2 secretion systems (SS. We provide evidence that the T6SS mediates the disruption of the BcCV membrane, which allows the escape of proteins secreted by the T2SS into the macrophage cytoplasm. This was demonstrated by the activity of fusion derivatives of the T2SS-secreted metalloproteases ZmpA and ZmpB with adenylcyclase. Supporting this notion, ZmpA and ZmpB are required for efficient IL-1β secretion in a T6SS dependent manner. ZmpA and ZmpB are also required for the maturation arrest of the BcCVs and bacterial intra-macrophage survival in a T6SS-independent fashion. Our results uncover a novel mechanism for inflammasome activation that involves cooperation between two bacterial secretory pathways, and an unanticipated role for T2SS-secreted proteins in intracellular bacterial survival.

  11. The exopolysaccharide gene cluster Bcam1330-Bcam1341 is involved in Burkholderia cenocepacia biofilm formation, and its expression is regulated by c-di-GMP and Bcam1349

    DEFF Research Database (Denmark)

    Fazli, Mustafa; McCarthy, Yvonne; Givskov, Michael

    2013-01-01

    In Burkholderia cenocepacia, the second messenger cyclic diguanosine monophosphate (c-di-GMP) has previously been shown to positively regulate biofilm formation and the expression of cellulose and type-I fimbriae genes through binding to the transcriptional regulator Bcam1349. Here, we provide...... evidence that cellulose and type-I fimbriae are not involved in B. cenocepacia biofilm formation in flow chambers, and we identify a novel Bcam1349/c-di-GMP-regulated exopolysaccharide gene cluster which is essential for B. cenocepacia biofilm formation. Overproduction of Bcam1349 in trans promotes wrinkly...... matrix exopolysaccharide and to be essential for flow-chamber biofilm formation. We demonstrate that Bcam1349 binds to the promoter region of genes in the Bcam1330-Bcam1341 cluster and that this binding is enhanced by the presence of c-di-GMP. Furthermore, we demonstrate that overproduction of both c-di-GMP...

  12. High-resolution structure of the M14-type cytosolic carboxypeptidase from Burkholderia cenocepacia refined exploiting PDB-REDO strategies

    Energy Technology Data Exchange (ETDEWEB)

    Rimsa, Vadim; Eadsforth, Thomas C. [University of Dundee, Dundee DD1 5EH, Scotland (United Kingdom); Joosten, Robbie P. [Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam (Netherlands); Hunter, William N., E-mail: w.n.hunter@dundee.ac.uk [University of Dundee, Dundee DD1 5EH, Scotland (United Kingdom)

    2014-02-01

    The structure of a bacterial M14-family carboxypeptidase determined exploiting microfocus synchrotron radiation and highly automated refinement protocols reveals its potential to act as a polyglutamylase. A potential cytosolic metallocarboxypeptidase from Burkholderia cenocepacia has been crystallized and a synchrotron-radiation microfocus beamline allowed the acquisition of diffraction data to 1.9 Å resolution. The asymmetric unit comprises a tetramer containing over 1500 amino acids, and the high-throughput automated protocols embedded in PDB-REDO were coupled with model–map inspections in refinement. This approach has highlighted the value of such protocols for efficient analyses. The subunit is constructed from two domains. The N-terminal domain has previously only been observed in cytosolic carboxypeptidase (CCP) proteins. The C-terminal domain, which carries the Zn{sup 2+}-containing active site, serves to classify this protein as a member of the M14D subfamily of carboxypeptidases. Although eukaryotic CCPs possess deglutamylase activity and are implicated in processing modified tubulin, the function and substrates of the bacterial family members remain unknown. The B. cenocepacia protein did not display deglutamylase activity towards a furylacryloyl glutamate derivative, a potential substrate. Residues previously shown to coordinate the divalent cation and that contribute to peptide-bond cleavage in related enzymes such as bovine carboxypeptidase are conserved. The location of a conserved basic patch in the active site adjacent to the catalytic Zn{sup 2+}, where an acetate ion is identified, suggests recognition of the carboxy-terminus in a similar fashion to other carboxypeptidases. However, there are significant differences that indicate the recognition of substrates with different properties. Of note is the presence of a lysine in the S1′ recognition subsite that suggests specificity towards an acidic substrate.

  13. High-resolution structure of the M14-type cytosolic carboxypeptidase from Burkholderia cenocepacia refined exploiting PDB-REDO strategies

    International Nuclear Information System (INIS)

    Rimsa, Vadim; Eadsforth, Thomas C.; Joosten, Robbie P.; Hunter, William N.

    2014-01-01

    The structure of a bacterial M14-family carboxypeptidase determined exploiting microfocus synchrotron radiation and highly automated refinement protocols reveals its potential to act as a polyglutamylase. A potential cytosolic metallocarboxypeptidase from Burkholderia cenocepacia has been crystallized and a synchrotron-radiation microfocus beamline allowed the acquisition of diffraction data to 1.9 Å resolution. The asymmetric unit comprises a tetramer containing over 1500 amino acids, and the high-throughput automated protocols embedded in PDB-REDO were coupled with model–map inspections in refinement. This approach has highlighted the value of such protocols for efficient analyses. The subunit is constructed from two domains. The N-terminal domain has previously only been observed in cytosolic carboxypeptidase (CCP) proteins. The C-terminal domain, which carries the Zn 2+ -containing active site, serves to classify this protein as a member of the M14D subfamily of carboxypeptidases. Although eukaryotic CCPs possess deglutamylase activity and are implicated in processing modified tubulin, the function and substrates of the bacterial family members remain unknown. The B. cenocepacia protein did not display deglutamylase activity towards a furylacryloyl glutamate derivative, a potential substrate. Residues previously shown to coordinate the divalent cation and that contribute to peptide-bond cleavage in related enzymes such as bovine carboxypeptidase are conserved. The location of a conserved basic patch in the active site adjacent to the catalytic Zn 2+ , where an acetate ion is identified, suggests recognition of the carboxy-terminus in a similar fashion to other carboxypeptidases. However, there are significant differences that indicate the recognition of substrates with different properties. Of note is the presence of a lysine in the S1′ recognition subsite that suggests specificity towards an acidic substrate

  14. Lack of MyD88 protects the immunodeficient host against fatal lung inflammation triggered by the opportunistic bacteria Burkholderia cenocepacia.

    Science.gov (United States)

    Ventura, Grasiella M de C; Balloy, Viviane; Ramphal, Reuben; Khun, Huot; Huerre, Michel; Ryffel, Bernhard; Plotkowski, Maria-Cristina M; Chignard, Michel; Si-Tahar, Mustapha

    2009-07-01

    Burkholderia cenocepacia is an opportunistic pathogen of major concern for cystic fibrosis patients as well as immunocompromised cancer patients and transplant recipients. The mechanisms by which B. cenocepacia triggers a rapid health deterioration of the susceptible host have yet to be characterized. TLR and their key signaling intermediate MyD88 play a central role in the detection of microbial molecular patterns and in the initiation of an effective immune response. We performed a study to better understand the role of TLR-MyD88 signaling in B. cenocepacia-induced pathogenesis in the immunocompromised host, using an experimental murine model. The time-course of several dynamic parameters, including animal survival, bacterial load, and secretion of critical inflammatory mediators, was compared in infected and immunosuppressed wild-type and MyD88(-/-) mice. Notably, when compared with wild-type mice, infected MyD88(-/-) animals displayed significantly reduced levels of inflammatory mediators (including KC, TNF-alpha, IL-6, MIP-2, and G-CSF) in blood and lung airspaces. Moreover, despite a higher transient bacterial load in the lungs, immunosuppressed mice deficient in MyD88 had an unexpected survival advantage. Finally, we showed that this B. cenocepacia-induced life-threatening infection of wild-type mice involved the proinflammatory cytokine TNF-alpha and could be prevented by corticosteroids. Altogether, our findings demonstrate that a MyD88-dependent pathway can critically contribute to a detrimental host inflammatory response that leads to fatal pneumonia.

  15. [The Effect of Introduction of the Heterologous Gene Encoding the N-acyl-homoserine Lactonase (aiiA) on the Properties of Burkholderia cenocepacia 370].

    Science.gov (United States)

    Plyuta, V A; Lipasova, V A; Koksharova, O A; Veselova, M A; Kuznetsov, A E; Khmel, I A

    2015-08-01

    To study the role of Quorum Sensing (QS) regulation in the control of the cellular processes of Burkholderia cenocepacia 370, plasmid pME6863 was transferred into its cells. The plasmid contains a heterologous gene encoding for AiiA N-acyl-homoserine lactonase, which degrades the signaling molecules of the QS system of N-acyl-homoserine lactones (AHL). An absence or reduction of AHL in the culture was revealed with the biosensors Chromobacterium violaceum CV026 and Agrobacterium tumifaciens NT1/pZLR4, respectively. The presence of the aiiA gene, which was cloned from Bacillus sp. A24 in the cells of B. cenocepacia 370, resulted in a lack of hemolytic activity, which reduced the extracellular proteolytic activity and decreased the cells' ability to migration in swarms on the surface of the agar medium. The introduction of the aiiA gene did not affect lipase activity, fatty acids synthesis, HCN synthesis, or biofilm formation. Hydrogen peroxide was shown to stimulate biofilm formation by B. cenocepacia 370 in concentrations that inhibited or weakly suppressed bacterial growth. The introduction of the aiiA gene into the cells did not eliminate this effect but it did reduce it.

  16. ParABS Systems of the Four Replicons of Burkholderia cenocepacia: New Chromosome Centromeres Confer Partition Specificity†

    Science.gov (United States)

    Dubarry, Nelly; Pasta, Franck; Lane, David

    2006-01-01

    Most bacterial chromosomes carry an analogue of the parABS systems that govern plasmid partition, but their role in chromosome partition is ambiguous. parABS systems might be particularly important for orderly segregation of multipartite genomes, where their role may thus be easier to evaluate. We have characterized parABS systems in Burkholderia cenocepacia, whose genome comprises three chromosomes and one low-copy-number plasmid. A single parAB locus and a set of ParB-binding (parS) centromere sites are located near the origin of each replicon. ParA and ParB of the longest chromosome are phylogenetically similar to analogues in other multichromosome and monochromosome bacteria but are distinct from those of smaller chromosomes. The latter form subgroups that correspond to the taxa of their hosts, indicating evolution from plasmids. The parS sites on the smaller chromosomes and the plasmid are similar to the “universal” parS of the main chromosome but with a sequence specific to their replicon. In an Escherichia coli plasmid stabilization test, each parAB exhibits partition activity only with the parS of its own replicon. Hence, parABS function is based on the independent partition of individual chromosomes rather than on a single communal system or network of interacting systems. Stabilization by the smaller chromosome and plasmid systems was enhanced by mutation of parS sites and a promoter internal to their parAB operons, suggesting autoregulatory mechanisms. The small chromosome ParBs were found to silence transcription, a property relevant to autoregulation. PMID:16452432

  17. Burkholderia species infections in patients with cystic fibrosis in British Columbia, Canada. 30 years' experience.

    Science.gov (United States)

    Zlosnik, James E A; Zhou, Guohai; Brant, Rollin; Henry, Deborah A; Hird, Trevor J; Mahenthiralingam, Eshwar; Chilvers, Mark A; Wilcox, Pearce; Speert, David P

    2015-01-01

    We have been collecting Burkholderia species bacteria from patients with cystic fibrosis (CF) for the last 30 years. During this time, our understanding of their multispecies taxonomy and infection control has evolved substantially. To evaluate the long-term (30 year) epidemiology and clinical outcome of Burkholderia infection in CF, and fully define the risks associated with infection by each species. Isolates from Burkholderia-positive patients (n=107) were speciated and typed annually for each infected patient. Microbiological and clinical data were evaluated by thorough review of patient charts, and statistical analyses performed to define significant epidemiological factors. Before 1995, the majority of new Burkholderia infections were caused by epidemic clones of Burkholderia cenocepacia. After implementation of new infection control measures in 1995, Burkholderia multivorans became the most prevalent species. Survival analysis showed that patients with CF infected with B. cenocepacia had a significantly worse outcome than those with B. multivorans, and a novel finding was that, after Burkholderia infection, the prognosis for females was significantly worse than for males. B. multivorans and B. cenocepacia have been the predominant Burkholderia species infecting people with CF in Vancouver. The implementation of infection control measures were successful in preventing new acquisition of epidemic strains of B. cenocepacia, leaving nonclonal B. multivorans as the most prevalent species. Historically, survival after infection with B. cenocepacia has been significantly worse than B. multivorans infection, and, of new significance, we show that females tend toward worse clinical outcomes.

  18. Regulation of Burkholderia cenocepacia biofilm formation by RpoN and the c-di-GMP effector BerB

    DEFF Research Database (Denmark)

    Fazli, Mustafa; Rybtke, Morten Levin; Steiner, Elisabeth

    2017-01-01

    Knowledge about the molecular mechanisms that are involved in the regulation of biofilm formation is essential for the development of biofilm-control measures. It is well established that the nucleotide second messenger cyclic diguanosine monophosphate (c-di-GMP) is a positive regulator of biofilm...... formation in many bacteria, but more knowledge about c-di-GMP effectors is needed. We provide evidence that c-di-GMP, the alternative sigma factor RpoN (σ54), and the enhancer-binding protein BerB play a role in biofilm formation of Burkholderia cenocepacia by regulating the production of a biofilm......-stabilizing exopolysaccharide. Our findings suggest that BerB binds c-di-GMP, and activates RpoN-dependent transcription of the berA gene coding for a c-di-GMP-responsive transcriptional regulator. An increased level of the BerA protein in turn induces the production of biofilm-stabilizing exopolysaccharide in response to high...

  19. The AHL- and BDSF-dependent quorum sensing systems control specific and overlapping sets of genes in Burkholderia cenocepacia H111.

    Directory of Open Access Journals (Sweden)

    Nadine Schmid

    Full Text Available Quorum sensing in Burkholderia cenocepacia H111 involves two signalling systems that depend on different signal molecules, namely N-acyl homoserine lactones (AHLs and the diffusible signal factor cis-2-dodecenoic acid (BDSF. Previous studies have shown that AHLs and BDSF control similar phenotypic traits, including biofilm formation, proteolytic activity and pathogenicity. In this study we mapped the BDSF stimulon by RNA-Seq and shotgun proteomics analysis. We demonstrate that a set of the identified BDSF-regulated genes or proteins are also controlled by AHLs, suggesting that the two regulons partially overlap. The detailed analysis of two mutually regulated operons, one encoding three lectins and the other one encoding the large surface protein BapA and its type I secretion machinery, revealed that both AHLs and BDSF are required for full expression, suggesting that the two signalling systems operate in parallel. In accordance with this, we show that both AHLs and BDSF are required for biofilm formation and protease production.

  20. A sensor kinase recognizing the cell-cell signal BDSF (cis-2-dodecenoic acid) regulates virulence in Burkholderia cenocepacia

    DEFF Research Database (Denmark)

    McCarthy, Y.; Yang, Liang; Twomey, K.B.

    2010-01-01

    Xanthomonas campestris. The mechanism of perception of this signal and the range of functions regulated in B. cenocepacia are, however, unknown. A screen for transposon mutants unable to respond to exogenous signal identified BCAM0227 as a potential BDSF sensor. BCAM0227 is a histidine sensor kinase...... with an input domain unrelated to that of RpfC, the DSF sensor found in xanthomonads. Transcriptome profiling established the scope of the BDSF regulon and demonstrated that the sensor controls expression of a subset of these genes. A chimeric sensor kinase in which the input domain of BCAM0227 replaced...... the input domain of RpfC was active in BDSF signal perception when expressed in X. campestris. Mutation of BCAM0227 gave rise to reduced cytotoxicity to Chinese hamster ovary cells and reduced virulence to Wax moth larvae and in the agar-bead mouse model of pulmonary infection. The findings identify BCAM...

  1. 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.

  2. Occidiofungin is an important component responsible for the antifungal activity of Burkholderia pyrrocinia strain Lyc2.

    Science.gov (United States)

    Wang, X Q; Liu, A X; Guerrero, A; Liu, J; Yu, X Q; Deng, P; Ma, L; Baird, S M; Smith, L; Li, X D; Lu, S E

    2016-03-01

    To identify the taxonomy of tobacco rhizosphere-isolated strain Lyc2 and investigate the mechanisms of the antifungal activities, focusing on antimicrobials gene clusters identification and function analysis. Multilocus sequence typing and 16S rRNA analyses indicated that strain Lyc2 belongs to Burkholderia pyrrocinia. Bioassay results indicated strain Lyc2 showed significant antifungal activities against a broad range of plant and animal fungal pathogens and control efficacy on seedling damping off disease of cotton. A 55·2-kb gene cluster which was homologous to ocf gene clusters in Burkholderia contaminans MS14 was confirmed to be responsible for antifungal activities by random mutagenesis; HPLC was used to verify the production of antifungal compounds. Multiple antibiotic and secondary metabolized biosynthesis gene clusters predicated by antiSMASH revealed the broad spectrum of antimicrobials activities of the strain. Our results revealed the mechanisms of antifungal activities of strain Lyc2 and expand our knowledge about production of occidiofungin in the bacteria Burkholderia. Understanding the mechanisms of antifungal activities of strain Lyc2 has contributed to discovery of new antibiotics and expand our knowledge of production of occidiofungin in the bacteria Burkholderia. © 2015 The Society for Applied Microbiology.

  3. Proof that Burkholderia Strains Form Effective Symbioses with Legumes: a Study of Novel Mimosa-Nodulating Strains from South America

    Science.gov (United States)

    Chen, Wen-Ming; de Faria, Sergio M.; Straliotto, Rosângela; Pitard, Rosa M.; Simões-Araùjo, Jean L.; Chou, Jui-Hsing; Chou, Yi-Ju; Barrios, Edmundo; Prescott, Alan R.; Elliott, Geoffrey N.; Sprent, Janet I.; Young, J. Peter W.; James, Euan K.

    2005-01-01

    Twenty Mimosa-nodulating bacterial strains from Brazil and Venezuela, together with eight reference Mimosa-nodulating rhizobial strains and two other β-rhizobial strains, were examined by amplified rRNA gene restriction analysis. They fell into 16 patterns and formed a single cluster together with the known β-rhizobia, Burkholderia caribensis, Burkholderia phymatum, and Burkholderia tuberum. The 16S rRNA gene sequences of 15 of the 20 strains were determined, and all were shown to belong to the genus Burkholderia; four distinct clusters could be discerned, with strains isolated from the same host species usually clustering very closely. Five of the strains (MAP3-5, Br3407, Br3454, Br3461, and Br3469) were selected for further studies of the symbiosis-related genes nodA, the NodD-dependent regulatory consensus sequences (nod box), and nifH. The nodA and nifH sequences were very close to each other and to those of B. phymatum STM815, B. caribensis TJ182, and Cupriavidus taiwanensis LMG19424 but were relatively distant from those of B. tuberum STM678. In addition to nodulating their original hosts, all five strains could also nodulate other Mimosa spp., and all produced nodules on Mimosa pudica that had nitrogenase (acetylene reduction) activities and structures typical of effective N2-fixing symbioses. Finally, both wild-type and green fluorescent protein-expressing transconjugant strains of Br3461 and MAP3-5 produced N2-fixing nodules on their original hosts, Mimosa bimucronata (Br3461) and Mimosa pigra (MAP3-5), and hence this confirms strongly that Burkholderia strains can form effective symbioses with legumes. PMID:16269788

  4. Efflux Pump-mediated Drug Resistance in Burkholderia

    Directory of Open Access Journals (Sweden)

    Nicole L Podnecky

    2015-04-01

    Full Text Available Several members of the genus Burkholderia are prominent pathogens. Infections caused by these bacteria are difficult to treat because of significant antibiotic resistance. Virtually all Burkholderia species are also resistant to polymyxin, prohibiting use of drugs like colistin that are available for treatment of infections caused by most other drug resistant Gram-negative bacteria. Despite clinical significance and antibiotic resistance of Burkholderia species, characterization of efflux pumps lags behind other non-enteric Gram-negative pathogens such as Acinetobacter baumannii and Pseudomonas aeruginosa. Although efflux pumps have been described in several Burkholderia species, they have been best studied in B. cenocepacia and B. pseudomallei. As in other non-enteric Gram-negatives, efflux pumps of the resistance nodulation cell division (RND family are the clinically most significant efflux systems in these two species. Several efflux pumps were described in B. cenocepacia, which when expressed confer resistance to clinically significant antibiotics, including aminoglycosides, chloramphenicol, fluoroquinolones, and tetracyclines. Three RND pumps have been characterized in B. pseudomallei, two of which confer either intrinsic or acquired resistance to aminoglycosides, macrolides, chloramphenicol, fluoroquinolones, tetracyclines, trimethoprim, and in some instances trimethoprim+sulfamethoxazole. Several strains of the host-adapted B. mallei, a clone of B. pseudomallei, lack AmrAB-OprA and are therefore aminoglycoside and macrolide susceptible. B. thailandensis is closely related to B. pseudomallei, but non-pathogenic to humans. Its pump repertoire and ensuing drug resistance profile parallels that of B. pseudomallei. An efflux pump in B. vietnamiensis plays a significant role in acquired aminoglycoside resistance. Summarily, efflux pumps are significant players in Burkholderia drug resistance.

  5. 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.

  6. Complete Genome Sequence of a Burkholderia pseudomallei Strain Isolated from a Pet Green Iguana in Prague, Czech Republic

    Science.gov (United States)

    Thomas, Prasad; El-Adawy, Hosny; Mertens, Katja; Melzer, Falk; Hnizdo, Jan; Stamm, Ivonne

    2017-01-01

    ABSTRACT Burkholderia pseudomallei was isolated from pus from an abscess of a pet iguana living in a private household in Prague, Czech Republic. This paper presents the complete genome sequence of B. pseudomallei strain VB976100. PMID:28280033

  7. CD4+ T cell epitopes of FliC conserved between strains of Burkholderia: implications for vaccines against melioidosis and cepacia complex in cystic fibrosis.

    Science.gov (United States)

    Musson, Julie A; Reynolds, Catherine J; Rinchai, Darawan; Nithichanon, Arnone; Khaenam, Prasong; Favry, Emmanuel; Spink, Natasha; Chu, Karen K Y; De Soyza, Anthony; Bancroft, Gregory J; Lertmemongkolchai, Ganjana; Maillere, Bernard; Boyton, Rosemary J; Altmann, Daniel M; Robinson, John H

    2014-12-15

    Burkholderia pseudomallei is the causative agent of melioidosis characterized by pneumonia and fatal septicemia and prevalent in Southeast Asia. Related Burkholderia species are strong risk factors of mortality in cystic fibrosis (CF). The B. pseudomallei flagellar protein FliC is strongly seroreactive and vaccination protects challenged mice. We assessed B. pseudomallei FliC peptide binding affinity to multiple HLA class II alleles and then assessed CD4 T cell immunity in HLA class II transgenic mice and in seropositive individuals in Thailand. T cell hybridomas were generated to investigate cross-reactivity between B. pseudomallei and the related Burkholderia species associated with Cepacia Complex CF. B. pseudomallei FliC contained several peptide sequences with ability to bind multiple HLA class II alleles. Several peptides were shown to encompass strong CD4 T cell epitopes in B. pseudomallei-exposed individuals and in HLA transgenic mice. In particular, the p38 epitope is robustly recognized by CD4 T cells of seropositive donors across diverse HLA haplotypes. T cell hybridomas against an immunogenic B. pseudomallei FliC epitope also cross-reacted with orthologous FliC sequences from Burkholderia multivorans and Burkholderia cenocepacia, important pathogens in CF. Epitopes within FliC were accessible for processing and presentation from live or heat-killed bacteria, demonstrating that flagellin enters the HLA class II Ag presentation pathway during infection of macrophages with B. cenocepacia. Collectively, the data support the possibility of incorporating FliC T cell epitopes into vaccination programs targeting both at-risk individuals in B. pseudomallei endemic regions as well as CF patients. Copyright © 2014 by The American Association of Immunologists, Inc.

  8. Involvement of the efflux pumps in chloramphenicol selected strains of Burkholderia thailandensis: proteomic and mechanistic evidence.

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    Fabrice V Biot

    Full Text Available Burkholderia is a bacterial genus comprising several pathogenic species, including two species highly pathogenic for humans, B. pseudomallei and B. mallei. B. thailandensis is a weakly pathogenic species closely related to both B. pseudomallei and B. mallei. It is used as a study model. These bacteria are able to exhibit multiple resistance mechanisms towards various families of antibiotics. By sequentially plating B. thailandensis wild type strains on chloramphenicol we obtained several resistant variants. This chloramphenicol-induced resistance was associated with resistance against structurally unrelated antibiotics including quinolones and tetracyclines. We functionally and proteomically demonstrate that this multidrug resistance phenotype, identified in chloramphenicol-resistant variants, is associated with the overexpression of two different efflux pumps. These efflux pumps are able to expel antibiotics from several families, including chloramphenicol, quinolones, tetracyclines, trimethoprim and some β-lactams, and present a partial susceptibility to efflux pump inhibitors. It is thus possible that Burkholderia species can develop such adaptive resistance mechanisms in response to antibiotic pressure resulting in emergence of multidrug resistant strains. Antibiotics known to easily induce overexpression of these efflux pumps should be used with discernment in the treatment of Burkholderia infections.

  9. Morphological characterization of several strains of the rice-pathogenic bacterium Burkholderia glumae in North Sumatra

    Science.gov (United States)

    Hasibuan, M.; Safni, I.; Lisnawita; Lubis, K.

    2018-02-01

    Burkholderia glumae is a quarantine seed-borne bacterial pathogen causing panicle blight disease on rice. This pathogen has been detected in some locations in Java, and recently, farmers in North Sumatra have reported rice yield loss with symptoms similar with those on rice infeced by the rice-pathogenic bacterium B. glumae. This research was aimed to isolate several bacterial strains from several rice varieties in various locations in North Sumatra and characterize the morphology of the strains to detect and identify the unknown bacterial strains presumably B. glumae. Several rice seed varieties were collected from Medan and Deli Serdang Districts. The seed samples were extracted, isolated and purified, then grown in semi-selective media PPGA. The morphological characteristics of the bacterial strains were determined including Gram staining, bacterial colony’s and bacterial cell’s morphology. The results showed that of eleven strains isolated, two strains were Gram negative and nine strains were Gram positive. On the basis of colony morphology, all strains had circular form, flat elevation and cream colour while the colony margin varied, i.e. entire and undulate. Most strains had bacillus/rod shape (8 strains) and only 3 strains were coccus.

  10. Identification and cloning of four riboswitches from Burkholderia pseudomallei strain K96243

    Science.gov (United States)

    Munyati-Othman, Noor; Fatah, Ahmad Luqman Abdul; Piji, Mohd Al Akmarul Fizree Bin Md; Ramlan, Effirul Ikhwan; Raih, Mohd Firdaus

    2015-09-01

    Structured RNAs referred as riboswitches have been predicted to be present in the genome sequence of Burkholderia pseudomallei strain K96243. Four of the riboswitches were identified and analyzed through BLASTN, Rfam search and multiple sequence alignment. The RNA aptamers belong to the following riboswitch classifications: glycine riboswitch, cobalamin riboswitch, S-adenosyl-(L)-homocysteine (SAH) riboswitch and flavin mononucleotide (FMN) riboswitch. The conserved nucleotides for each aptamer were identified and were marked on the secondary structure generated by RNAfold. These riboswitches were successfully amplified and cloned for further study.

  11. X-ray crystal structures of the pheromone-binding domains of two quorum-hindered transcription factors, YenR of Yersinia enterocolitica and CepR2 of Burkholderia cenocepacia: KIM et al.

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    Kim, Youngchang [Midwest Center for Structural Genomics, Biosciences, Argonne National Laboratory, Argonne Illinois 60439; Structural Biology Center, Biosciences, Argonne National Laboratory, Argonne Illinois 60439; Chhor, Gekleng [Midwest Center for Structural Genomics, Biosciences, Argonne National Laboratory, Argonne Illinois 60439; Tsai, Ching-Sung [Department of Microbiology, Cornell University, Ithaca New York 14853; Fox, Gabriel [Department of Microbiology, Cornell University, Ithaca New York 14853; Chen, Chia-Sui [Department of Microbiology, Cornell University, Ithaca New York 14853; Winans, Nathan J. [Department of Microbiology, Cornell University, Ithaca New York 14853; Jedrzejczak, Robert [Structural Biology Center, Biosciences, Argonne National Laboratory, Argonne Illinois 60439; Joachimiak, Andrzej [Midwest Center for Structural Genomics, Biosciences, Argonne National Laboratory, Argonne Illinois 60439; Structural Biology Center, Biosciences, Argonne National Laboratory, Argonne Illinois 60439; Department of Biochemistry and Molecular Biology, University of Chicago, Chicago Illinois 60637; Winans, Stephen C. [Department of Microbiology, Cornell University, Ithaca New York 14853

    2017-07-24

    The ability of LuxR-type proteins to regulate transcription is controlled by bacterial pheromones, N-acylhomoserine lactones (AHLs). Most LuxR-family proteins require their cognate AHLs for activity, and some of them require AHLs for folding and stability, and for protease-resistance. However, a few members of this family are able to fold, dimerize, bind DNA, and regulate transcription in the absence of AHLs; moreover, these proteins are antagonized by their cognate AHLs. One such protein is YenR of Yersinia enterocolitica, which is antagonized by N-3-oxohexanoyl-l-homoserine lactone (OHHL). This pheromone is produced by the OHHL synthase, a product of the adjacent yenI gene. Another example is CepR2 of Burkholderia cenocepacia, which is antagonized by N-octanoyl-l-homoserine lactone (OHL), whose synthesis is directed by the cepI gene of the same bacterium. Here, we describe the high-resolution crystal structures of the AHL binding domains of YenR and CepR2. YenR was crystallized in the presence and absence of OHHL. While this ligand does not cause large scale changes in the YenR structure, it does alter the orientation of several highly conserved YenR residues within and near the pheromone-binding pocket, which in turn caused a significant movement of a surface-exposed loop.

  12. Diversities in virulence, antifungal activity, pigmentation and DNA fingerprint among strains of Burkholderia glumae.

    Science.gov (United States)

    Karki, Hari S; Shrestha, Bishnu K; Han, Jae Woo; Groth, Donald E; Barphagha, Inderjit K; Rush, Milton C; Melanson, Rebecca A; Kim, Beom Seok; Ham, Jong Hyun

    2012-01-01

    Burkholderia glumae is the primary causal agent of bacterial panicle blight of rice. In this study, 11 naturally avirulent and nine virulent strains of B. glumae native to the southern United States were characterized in terms of virulence in rice and onion, toxofalvin production, antifungal activity, pigmentation and genomic structure. Virulence of B. glumae strains on rice panicles was highly correlated to virulence on onion bulb scales, suggesting that onion bulb can be a convenient alternative host system to efficiently determine the virulence of B. glumae strains. Production of toxoflavin, the phytotoxin that functions as a major virulence factor, was closely associated with the virulence phenotypes of B. glumae strains in rice. Some strains of B. glumae showed various levels of antifungal activity against Rhizoctonia solani, the causal agent of sheath blight, and pigmentation phenotypes on casamino acid-peptone-glucose (CPG) agar plates regardless of their virulence traits. Purple and yellow-green pigments were partially purified from a pigmenting strain of B. glumae, 411gr-6, and the purple pigment fraction showed a strong antifungal activity against Collectotrichum orbiculare. Genetic variations were detected among the B. glumae strains from DNA fingerprinting analyses by repetitive element sequence-based PCR (rep-PCR) for BOX-A1R-based repetitive extragenic palindromic (BOX) or enterobacterial repetitive intergenic consensus (ERIC) sequences of bacteria; and close genetic relatedness among virulent but pigment-deficient strains were revealed by clustering analyses of DNA fingerprints from BOX-and ERIC-PCR.

  13. An objective approach for Burkholderia pseudomallei strain selection as challenge material for medical countermeasures efficacy testing

    Directory of Open Access Journals (Sweden)

    Kristopher E. Van Zandt

    2012-09-01

    Full Text Available Burkholderia pseudomallei is the causative agent of melioidosis, a rare disease of biodefense concern with high mortality and extreme difficulty in treatment. No human vaccines are available that protect against B. pseudomallei infection, and with the current limitations of antibiotic treatment, the development of new preventative and therapeutic interventions is crucial. Although clinical trials could be used to test the efficacy of new medical countermeasures (MCMs, the high mortality rates associated with melioidosis raises significant ethical issues concerning treating individuals with new compounds with unknown efficacies. The US Food and Drug Administration (FDA has formulated a set of guidelines for the licensure of new MCMs to treat diseases in which it would be unethical to test the efficacy of these drugs in humans. The FDA Animal Rule 21 CFR 314 calls for consistent, well-characterized B. pseudomallei strains to be used as challenge material in animal models. In order to facilitate the efficacy testing of new MCMs for melioidosis using animal models, we intend to develop a well-characterized panel of strains for use. This panel will comprise of strains that were isolated from human cases, have a low passage history, are virulent in animal models, and are well characterized phenotypically and genotypically. We have reviewed published and unpublished data on various B. pseudomallei strains to establish an objective method for selecting the strains to be included in the panel of B. pseudomallei strains with attention to five categories: animal infection models, genetic characterization, clinical and passage history, and availability of the strain to the research community. We identified 109 strains with data in at least one of the five categories, scored each strain based on the gathered data and identified 6 strains as candidate for a B. pseudomallei strain panel.

  14. An objective approach for Burkholderia pseudomallei strain selection as challenge material for medical countermeasures efficacy testing.

    Science.gov (United States)

    Van Zandt, Kristopher E; Tuanyok, Apichai; Keim, Paul S; Warren, Richard L; Gelhaus, H Carl

    2012-01-01

    Burkholderia pseudomallei is the causative agent of melioidosis, a rare disease of biodefense concern with high mortality and extreme difficulty in treatment. No human vaccines are available that protect against B. pseudomallei infection, and with the current limitations of antibiotic treatment, the development of new preventative and therapeutic interventions is crucial. Although clinical trials could be used to test the efficacy of new medical countermeasures (MCMs), the high mortality rates associated with melioidosis raises significant ethical issues concerning treating individuals with new compounds with unknown efficacies. The US Food and Drug Administration (FDA) has formulated a set of guidelines for the licensure of new MCMs to treat diseases in which it would be unethical to test the efficacy of these drugs in humans. The FDA "Animal Rule" 21 CFR 314 calls for consistent, well-characterized B. pseudomallei strains to be used as challenge material in animal models. In order to facilitate the efficacy testing of new MCMs for melioidosis using animal models, we intend to develop a well-characterized panel of strains for use. This panel will comprise of strains that were isolated from human cases, have a low passage history, are virulent in animal models, and are well-characterized phenotypically and genotypically. We have reviewed published and unpublished data on various B. pseudomallei strains to establish an objective method for selecting the strains to be included in the panel of B. pseudomallei strains with attention to five categories: animal infection models, genetic characterization, clinical and passage history, and availability of the strain to the research community. We identified 109 strains with data in at least one of the five categories, scored each strain based on the gathered data and identified six strains as candidate for a B. pseudomallei strain panel.

  15. Immune Recognition of the Epidemic Cystic Fibrosis Pathogen Burkholderia dolosa.

    Science.gov (United States)

    Roux, Damien; Weatherholt, Molly; Clark, Bradley; Gadjeva, Mihaela; Renaud, Diane; Scott, David; Skurnik, David; Priebe, Gregory P; Pier, Gerald; Gerard, Craig; Yoder-Himes, Deborah R

    2017-06-01

    Burkholderia dolosa caused an outbreak in the cystic fibrosis (CF) clinic at Boston Children's Hospital from 1998 to 2005 and led to the infection of over 40 patients, many of whom died due to complications from infection by this organism. To assess whether B. dolosa significantly contributes to disease or is recognized by the host immune response, mice were infected with a sequenced outbreak B. dolosa strain, AU0158, and responses were compared to those to the well-studied CF pathogen Pseudomonas aeruginosa In parallel, mice were also infected with a polar flagellin mutant of B. dolosa to examine the role of flagella in B. dolosa lung colonization. The results showed a higher persistence in the host by B. dolosa strains, and yet, neutrophil recruitment and cytokine production were lower than those with P. aeruginosa The ability of host immune cells to recognize B. dolosa was then assessed, B. dolosa induced a robust cytokine response in cultured cells, and this effect was dependent on the flagella only when bacteria were dead. Together, these results suggest that B. dolosa can be recognized by host cells in vitro but may avoid or suppress the host immune response in vivo through unknown mechanisms. B. dolosa was then compared to other Burkholderia species and found to induce similar levels of cytokine production despite being internalized by macrophages more than Burkholderia cenocepacia strains. These data suggest that B. dolosa AU0158 may act differently with host cells and is recognized differently by immune systems than are other Burkholderia strains or species. Copyright © 2017 American Society for Microbiology.

  16. Rapid emergence of a ceftazidime-resistant Burkholderia multivorans strain in a cystic fibrosis patient.

    Science.gov (United States)

    Stokell, Joshua R; Gharaibeh, Raad Z; Steck, Todd R

    2013-12-01

    Burkholderia multivorans poses a serious health threat to cystic fibrosis patients due to innate resistance to multiple antibiotics and acquisition of resistance to a range of antibiotics due to the frequent use of antibiotics to treat chronic infections. Monitoring antibiotic susceptibility is crucial to managing patient care. We identified the rapid emergence of a ceftazidime-resistant strain in a single patient within four days during a hospitalization for treatment of an exacerbation. B. multivorans was isolated from expectorated sputum samples using Burkholderia cepacia selective agar. A macrodilution assay was performed on all isolates to determine the minimum inhibitory concentration of ceftazidime. Approximately 4000 colonies were scored to identify the percent of ceftazidime-resistant colonies. Extracted DNA was used to determine the total bacterial counts and abundance of B. multivorans using quantitative PCR. An increase from no detectable B. multivorans ceftazidime-resistant colonies to over 75% of all colonies tested occurred within a four-day period. The resistant population remained dominant in 6 of the 8 samples in the following 17 months of the study. qPCR revealed an association between change in the percent of resistant colonies and abundance of B. multivorans, but not of total bacteria. No association was found between the acquisition of resistance to ceftazidime and other antibiotics commonly used to treat B. multivorans infections. The rapid emergence of a ceftazidime-resistant by B. multivorans strain occurred during a hospitalization while under selective pressure of antibiotics. The resistant strain maintained dominance in the B. multivorans population which resulted in an overall decline in a patient health and treatment efficacy. Copyright © 2013 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

  17. Bacteria of the Burkholderia cepacia complex are cyanogenic under biofilm and colonial growth conditions

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    Hoshino Saiko

    2008-06-01

    Full Text Available Abstract Background The Burkholderia cepacia complex (Bcc is a collection of nine genotypically distinct but phenotypically similar species. They show wide ecological diversity and include species that are used for promoting plant growth and bio-control as well species that are opportunistic pathogens of vulnerable patients. Over recent years the Bcc have emerged as problematic pathogens of the CF lung. Pseudomonas aeruginosa is another important CF pathogen. It is able to synthesise hydrogen cyanide (HCN, a potent inhibitor of cellular respiration. We have recently shown that HCN production by P. aeruginosa may have a role in CF pathogenesis. This paper describes an investigation of the ability of bacteria of the Bcc to make HCN. Results The genome of Burkholderia cenocepacia has 3 putative HCN synthase encoding (hcnABC gene clusters. B. cenocepacia and all 9 species of the Bcc complex tested were able to make cyanide at comparable levels to P. aeruginosa, but only when grown surface attached as colonies or during biofilm growth on glass beads. In contrast to P. aeruginosa and other cyanogenic bacteria, cyanide was not detected during planktonic growth of Bcc strains. Conclusion All species in the Bcc are cyanogenic when grown as surface attached colonies or as biofilms.

  18. Characterization of Burkholderia pseudomallei Strains Using a Murine Intraperitoneal Infection Model and In Vitro Macrophage Assays.

    Directory of Open Access Journals (Sweden)

    Susan L Welkos

    Full Text Available Burkholderia pseudomallei, the etiologic agent of melioidosis, is a gram-negative facultative intracellular bacterium. This bacterium is endemic in Southeast Asia and Northern Australia and can infect humans and animals by several routes. It has also been estimated to present a considerable risk as a potential biothreat agent. There are currently no effective vaccines for B. pseudomallei, and antibiotic treatment can be hampered by nonspecific symptomology, the high incidence of naturally occurring antibiotic resistant strains, and disease chronicity. Accordingly, there is a concerted effort to better characterize B. pseudomallei and its associated disease. Before novel vaccines and therapeutics can be tested in vivo, a well characterized animal model is essential. Previous work has indicated that mice may be a useful animal model. In order to develop standardized animal models of melioidosis, different strains of bacteria must be isolated, propagated, and characterized. Using a murine intraperitoneal (IP infection model, we tested the virulence of 11 B. pseudomallei strains. The IP route offers a reproducible way to rank virulence that can be readily reproduced by other laboratories. This infection route is also useful in distinguishing significant differences in strain virulence that may be masked by the exquisite susceptibility associated with other routes of infection (e.g., inhalational. Additionally, there were several pathologic lesions observed in mice following IP infection. These included varisized abscesses in the spleen, liver, and haired skin. This model indicated that commonly used laboratory strains of B. pseudomallei (i.e., K96243 and 1026b were significantly less virulent as compared to more recently acquired clinical isolates. Additionally, we characterized in vitro strain-associated differences in virulence for macrophages and described a potential inverse relationship between virulence in the IP mouse model of some strains

  19. Characterization of Burkholderia pseudomallei Strains Using a Murine Intraperitoneal Infection Model and In Vitro Macrophage Assays.

    Science.gov (United States)

    Welkos, Susan L; Klimko, Christopher P; Kern, Steven J; Bearss, Jeremy J; Bozue, Joel A; Bernhards, Robert C; Trevino, Sylvia R; Waag, David M; Amemiya, Kei; Worsham, Patricia L; Cote, Christopher K

    2015-01-01

    Burkholderia pseudomallei, the etiologic agent of melioidosis, is a gram-negative facultative intracellular bacterium. This bacterium is endemic in Southeast Asia and Northern Australia and can infect humans and animals by several routes. It has also been estimated to present a considerable risk as a potential biothreat agent. There are currently no effective vaccines for B. pseudomallei, and antibiotic treatment can be hampered by nonspecific symptomology, the high incidence of naturally occurring antibiotic resistant strains, and disease chronicity. Accordingly, there is a concerted effort to better characterize B. pseudomallei and its associated disease. Before novel vaccines and therapeutics can be tested in vivo, a well characterized animal model is essential. Previous work has indicated that mice may be a useful animal model. In order to develop standardized animal models of melioidosis, different strains of bacteria must be isolated, propagated, and characterized. Using a murine intraperitoneal (IP) infection model, we tested the virulence of 11 B. pseudomallei strains. The IP route offers a reproducible way to rank virulence that can be readily reproduced by other laboratories. This infection route is also useful in distinguishing significant differences in strain virulence that may be masked by the exquisite susceptibility associated with other routes of infection (e.g., inhalational). Additionally, there were several pathologic lesions observed in mice following IP infection. These included varisized abscesses in the spleen, liver, and haired skin. This model indicated that commonly used laboratory strains of B. pseudomallei (i.e., K96243 and 1026b) were significantly less virulent as compared to more recently acquired clinical isolates. Additionally, we characterized in vitro strain-associated differences in virulence for macrophages and described a potential inverse relationship between virulence in the IP mouse model of some strains and in the

  20. Phylogenomic Study of Burkholderia glathei-like Organisms, Proposal of 13 Novel Burkholderia Species and Emended Descriptions of Burkholderia sordidicola, Burkholderia zhejiangensis, and Burkholderia grimmiae

    Science.gov (United States)

    Peeters, Charlotte; Meier-Kolthoff, Jan P.; Verheyde, Bart; De Brandt, Evie; Cooper, Vaughn S.; Vandamme, Peter

    2016-01-01

    Partial gyrB gene sequence analysis of 17 isolates from human and environmental sources revealed 13 clusters of strains and identified them as Burkholderia glathei clade (BGC) bacteria. The taxonomic status of these clusters was examined by whole-genome sequence analysis, determination of the G+C content, whole-cell fatty acid analysis and biochemical characterization. The whole-genome sequence-based phylogeny was assessed using the Genome Blast Distance Phylogeny (GBDP) method and an extended multilocus sequence analysis (MLSA) approach. The results demonstrated that these 17 BGC isolates represented 13 novel Burkholderia species that could be distinguished by both genotypic and phenotypic characteristics. BGC strains exhibited a broad metabolic versatility and developed beneficial, symbiotic, and pathogenic interactions with different hosts. Our data also confirmed that there is no phylogenetic subdivision in the genus Burkholderia that distinguishes beneficial from pathogenic strains. We therefore propose to formally classify the 13 novel BGC Burkholderia species as Burkholderia arvi sp. nov. (type strain LMG 29317T = CCUG 68412T), Burkholderia hypogeia sp. nov. (type strain LMG 29322T = CCUG 68407T), Burkholderia ptereochthonis sp. nov. (type strain LMG 29326T = CCUG 68403T), Burkholderia glebae sp. nov. (type strain LMG 29325T = CCUG 68404T), Burkholderia pedi sp. nov. (type strain LMG 29323T = CCUG 68406T), Burkholderia arationis sp. nov. (type strain LMG 29324T = CCUG 68405T), Burkholderia fortuita sp. nov. (type strain LMG 29320T = CCUG 68409T), Burkholderia temeraria sp. nov. (type strain LMG 29319T = CCUG 68410T), Burkholderia calidae sp. nov. (type strain LMG 29321T = CCUG 68408T), Burkholderia concitans sp. nov. (type strain LMG 29315T = CCUG 68414T), Burkholderia turbans sp. nov. (type strain LMG 29316T = CCUG 68413T), Burkholderia catudaia sp. nov. (type strain LMG 29318T = CCUG 68411T) and Burkholderia peredens sp. nov. (type strain LMG 29314T = CCUG

  1. Molecular typing of Burkholderia cepacia complex isolated from patients attending an Italian Cystic Fibrosis Centre.

    Science.gov (United States)

    Teri, Antonio; Sottotetti, Samantha; Biffi, Arianna; Girelli, Daniela; D'Accico, Monica; Arghittu, Milena; Colombo, Carla; Corti, Fabiola; Pizzamiglio, Giovanna; Cariani, Lisa

    2018-04-01

    Bacteria from the Burkholderia cepacia complex (Bcc) are capable of causing severe infections in patients with cystic fibrosis (CF). Bcc infection is often extremely difficult to treat due to its intrinsic resistance to multiple antibiotics. In addition, it seems to speed up the decline of lung function and is considered a contraindication for lung transplantation in CF. This study investigates the species of the Bcc strains recovered from chronically infected CF subjects by means of: isolation, identification methods and complete recA nucleotide sequences of 151 samples. Molecular typing showed that B. cenocepacia III is the dominant strain found in the group of subjects being treated at the Milan CF Centre (Italy) and that the infection is chronically maintained by the same species. Defining species by means of molecular analysis yields important information for the clinician in order to establish the most appropriate therapy and implement correct measures for prevention of transmission among CF subjects.

  2. Purification, biochemical characterization, and genetic cloning of the phytase produced by Burkholderia sp. strain a13.

    Science.gov (United States)

    Graminho, Eduardo Rezende; Takaya, Naoki; Nakamura, Akira; Hoshino, Takayuki

    2015-01-01

    A phytase-producing bacterium, Burkholderia sp. a13 (JCM 30421), was isolated from Lake Kasumigaura by enrichment cultivation using minimum medium containing phytic acid as the sole phosphorus source. The phytase production by strain a13 was induced by the presence of phytic acid and repressed by the addition of glucose. The purified enzyme had a molecular weight of 44 kDa and a phytase activity of 174 μmol min(-1) mg(-1). The enzyme showed broad substrate specificity, but the highest activity was observed with phytic acid. The enzyme activity was strongly inhibited by Cu(2+), Zn(2+), Hg(2+), and iodoacetic acid, indicating the requirement of a thiol group for the activity. Genetic cloning reveals that the mature portion of this enzyme consists of 428 amino acids with a calculated molecular weight of 46 kDa. The amino acid sequence showed the highest similarity to the phytase produced by Hafnia alvei with 48% identity; it also contained histidine acid phosphatase (HAP) motifs (RHGXRXP and HD), indicating the classification of this enzyme in the HAP phytase family. We have successfully expressed the cloned gene in Escherichia coli from its putative initiation codon, showing that the gene actually encodes the phytase.

  3. Burkholderia cepacia complex infection in an Adult Cystic Fibrosis unit in Madrid.

    Science.gov (United States)

    Correa-Ruiz, Ana; Girón, Rosa; Buendía, Buenaventura; Medina-Pascual, M José; Valenzuela, Claudia; López-Brea, Manuel; Sáez-Nieto, Juan Antonio

    2013-12-01

    Burkholderia cepacia complex have emerged as significant pathogens in cystic fibrosis (CF) patients due to the risk of cepacia syndrome and the innate multi-resistance of the microorganisms to antibiotics. The aim of this study was to describe the antimicrobial susceptibility profiles, the genotypes and subtypes of BCC, and the clinical evolution of CF patients with BCC. The lung function and Brasfield and Shwachman score were assessed in 12 patients. BCC were identified and susceptibility was studied by MicroScan (Siemens). Species and genospecies of BCC were confirmed by molecular methods in a Reference Centre (Majadahonda). BCC were identified in 12 of 70 patients (17.1%) over a ten year period. The mean age to colonization by BCC was 24.4 years (SD: 7.71). B. cenocepacia was isolated in 4 patients (33.3%), B. contaminans was isolated in 3 patients (25%), both B. vietnamiensis and B. stabilis were isolated in 2 patients (16.7%), and B. cepacia, B. multivorans and B. late were isolated in one patient (8.3%). Among the B. cenocepacia, subtype IIIa was identified in two strains, and subtype IIIb was identified in the other two strains. There was susceptibility to meropenem in 90% of BCC, 80% to cotrimoxazole, 60% to minocycline, 50% to ceftazidime, and 40% to levofloxacin. B. cenocepacia was the most prevalent species among the BCC isolated in CF adult patients, and subtypes IIIa and IIIb were identified in the 50% of the strains. Meropenem and cotrimoxazole showed the best activity. Copyright © 2012 Elsevier España, S.L. All rights reserved.

  4. Designing Probes for Immunodiagnostics: Structural Insights into an Epitope Targeting Burkholderia Infections.

    Science.gov (United States)

    Capelli, Riccardo; Matterazzo, Elena; Amabili, Marco; Peri, Claudio; Gori, Alessandro; Gagni, Paola; Chiari, Marcella; Lertmemongkolchai, Ganjana; Cretich, Marina; Bolognesi, Martino; Colombo, Giorgio; Gourlay, Louise J

    2017-10-13

    Structure-based epitope prediction drives the design of diagnostic peptidic probes to reveal specific antibodies elicited in response to infections. We previously identified a highly immunoreactive epitope from the peptidoglycan-associated lipoprotein (Pal) antigen from Burkholderia pseudomallei, which could also diagnose Burkholderia cepacia infections. Here, considering the high phylogenetic conservation within Burkholderia species, we ask whether cross-reactivity can be reciprocally displayed by the synthetic epitope from B. cenocepacia. We perform comparative analyses of the conformational preferences and diagnostic performances of the corresponding epitopes from the two Burkholderia species when presented in the context of the full-length proteins or as isolated peptides. The effects of conformation on the diagnostic potential and cross-reactivity of Pal peptide epitopes are rationalized on the basis of the 1.8 Å crystal structure of B. cenocepacia Pal and through computational analyses. Our results are discussed in the context of designing new diagnostic molecules for the early detection of infectious diseases.

  5. Study of class I integron in a Burkholderia cepacia complex strain isolated from blood colture

    Directory of Open Access Journals (Sweden)

    Linda Furlanis

    2011-06-01

    Full Text Available The Burkholderia cepacia complex (Bcc consists of several species that cause lung infections in patients with cystic fibrosis but are also capable to colonize immunocompromised patients. Once established, the infection is usually difficult to eradicate, as Bcc is intrinsically resistant to many antibiotics. Besides, the acquisition of additional resistance determinants by horizontal gene transfer makes very difficult the therapeutic approach to these infections. Among horizontally acquired DNAs, integrons have been frequently reported in many Gramnegative bacteria that affect human health, but they have not been found frequently in Burkholderia isolates until now. In the present work we report on a Bcc isolate, recovered from the blood of an immunocompromised patient, that carries a 2.3 kb class I integron already described in a Salmonella enterica isolate eight years ago, coding for aacA4, aadA1 and catB2 in its cassette array.

  6. An Objective Approach for Burkholderia pseudomallei Strain Selection as Challenge Material for Medical Countermeasures Efficacy Testing

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    Van Zandt, Kristopher E.; Tuanyok, Apichai; Keim, Paul S.; Warren, Richard L.; Gelhaus, H. Carl

    2012-01-01

    Burkholderia pseudomallei is the causative agent of melioidosis, a rare disease of biodefense concern with high mortality and extreme difficulty in treatment. No human vaccines are available that protect against B. pseudomallei infection, and with the current limitations of antibiotic treatment, the development of new preventative and therapeutic interventions is crucial. Although clinical trials could be used to test the efficacy of new medical countermeasures (MCMs), the high mortality rate...

  7. Trimeric autotransporter adhesins in members of the Burkholderia cepacia complex: a multifunctional family of proteins implicated in virulence

    Directory of Open Access Journals (Sweden)

    Arsénio Mendes Fialho

    2011-12-01

    Full Text Available Trimeric autotransporter adhesins (TAAs are multimeric surface proteins, involved in various biological traits of pathogenic Gram-negative bacteria including adherence, biofilm formation, invasion, survival within eukaryotic cells, serum resistance and cytotoxicity. TAAs have a modular architecture composed by a conserved membrane-anchored C-terminal domain and a variable number of stalk and head domains. In this study, a bioinformatic approach has been used to analyze the distribution and architecture of TAAs among Burkholderia cepacia complex (Bcc genomes. Fifteen genomes were probed revealing a total of 74 encoding sequences. Compared with other bacterial species, the Bcc genomes contain a disproportionately large number of TAAs (two genes to up to 8 genes, such as in B.cenocepacia. Phylogenetic analysis showed that the TAAs grouped into at least eight distinct clusters. TAAs with serine-rich repeats are clearly well separated from others, thereby representing a different evolutionary lineage. Comparative gene mapping across Bcc genomes reveals that TAA genes are inserted within conserved synteny blocks. We further focused our analysis on the epidemic strain B. cenocepacia J2315 in which 7 TAAs were annotated. Among these, 3 TAA-encoding genes (BCAM019, BCAM0223 and BCAM0224 are organized into a cluster and are candidates for multifunctional virulence factors. Here we review the current insights into the functional role of BCAM0224 as a model locus.

  8. Characterization of methyl parathion degradation by a Burkholderia zhejiangensis strain, CEIB S4-3, isolated from agricultural soils.

    Science.gov (United States)

    Popoca-Ursino, Elida C; Martínez-Ocampo, Fernando; Dantán-González, Edgar; Sánchez-Salinas, Enrique; Ortiz-Hernández, Ma Laura

    2017-12-01

    Through the use of an enrichment technique, we isolated from the agricultural soils of Morelos in central México a strain of Burkholderia zhejiangensis identified as CEIB S4-3, it's could use the pesticide methyl parathion (MP) as the only source of carbon and degrade completely p-nitrophenol (PNP). For more efficient MP and PNP degradation by the CEIB S4-3 strain, the absence of an extra carbon source, a large inoculum and an MP concentration up to 50 mg/l are required. Sequence and annotation analysis of the draft genome, showed presence of mpd functional gene, which was expressed and its activity on the MP was confirmed. Additionally, the genes coding for enzymes in the benzoquinone pathway (conducted by Gram-negative bacteria) and the benzenotriol pathway (conducted by Gram-positive bacteria) were found, which was corroborated by identification of intermediary metabolites by HPLC. Thus, we propose that B. zhejiangensis CEIB S4-3 uses both degradation pathways.

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

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    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.

  10. Expression, purification, crystallization and preliminary X-ray analysis of maleylacetate reductase from Burkholderia sp. strain SJ98

    International Nuclear Information System (INIS)

    Chauhan, Archana; Islam, Zeyaul; Jain, Rakesh Kumar; Karthikeyan, Subramanian

    2009-01-01

    Purification and preliminary X-ray crystallographic analysis of maleylacetate reductase encoded by the pnpD gene is reported. Maleylacetate reductase (EC 1.3.1.32) is an important enzyme that is involved in the degradation pathway of aromatic compounds and catalyzes the reduction of maleylacetate to 3-oxoadipate. The gene pnpD encoding maleylacetate reductase in Burkholderia sp. strain SJ98 was cloned, expressed in Escherichia coli and purified by affinity chromatography. The enzyme was crystallized in both native and SeMet-derivative forms by the sitting-drop vapour-diffusion method using PEG 3350 as a precipitant at 293 K. The crystals belonged to space group P2 1 2 1 2, with unit-cell parameters a = 72.91, b = 85.94, c = 53.07 Å. X-ray diffraction data for the native and SeMet-derivative crystal were collected to 2.7 and 2.9 Å resolution, respectively

  11. Complete genome sequence of Burkholderia sp. strain PAMC28687, a potential octopine-utilizing bacterium isolated from Antarctica lichen.

    Science.gov (United States)

    Han, So-Ra; Yu, Sang-Cheol; Ahn, Do-Hwan; Park, Hyun; Oh, Tae-Jin

    2016-05-20

    We report the complete genome sequence of Burkholderia sp. PAMC28687, which was isolated from the Antarctica lichen Useea sp., for better understanding of its catabolic traits in utilizing octopine as a source of carbon/nitrogen between Burkholderia and lichen. The genome consists of three circular chromosomes with five circular plasmids for the total 6,881,273bp sized genome with a G+C content of 58.14%. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Literature Review of DNA-Based Subspecies Analysis of Bacillus Anthracis Burkholderia Pseudomallel Burkholderia Mallei, and Yersinia Pestis

    National Research Council Canada - National Science Library

    Harvey, Steven

    1999-01-01

    ...; Bacillus anthracis, Burkholderia pseudomallei, Burkholderia mallei, and Yersinia pestis. Considerable research has been accomplished for the identification of polymorphisms from the strains B. anthracis and B. pseudomallei. The B...

  13. Draft Genome Sequence of the Soil Bacterium Burkholderia terrae Strain BS001, Which Interacts with Fungal Surface Structures

    DEFF Research Database (Denmark)

    Nazir, Rashid; Hansen, Martin A.; Sorensen, Soren

    2012-01-01

    Burkholderia terrae BS001 is a soil bacterium which was originally isolated from the mycosphere of the ectomycorrhizal fungus Laccaria proxima. It exhibits a range of fungus-interacting traits which reveal its propensity to actively interact at fungal interfaces. Here, we present the approximately...

  14. Burkholderia monticola sp. nov., isolated from mountain soil.

    Science.gov (United States)

    Baek, Inwoo; Seo, Boram; Lee, Imchang; Yi, Hana; Chun, Jongsik

    2015-02-01

    An ivory/yellow, Gram-stain-negative, short-rod-shaped, aerobic bacterial strain, designated JC2948(T), was isolated from a soil sample taken from Gwanak Mountain, Republic of Korea. 16S rRNA gene sequence analysis indicated that strain JC2948(T) belongs to the genus Burkholderia. The test strain showed highest sequence similarities to Burkholderia tropica LMG 22274(T) (97.6 %), Burkholderia acidipaludis NBRC 101816(T) (97.5 %), Burkholderia tuberum LMG 21444(T) (97.5 %), Burkholderia sprentiae LMG 27175(T) (97.4 %), Burkholderia terricola LMG 20594(T) (97.3 %) and Burkholderia diazotrophica LMG 26031(T) (97.1 %). Based on average nucleotide identity (ANI) values, the new isolate represents a novel genomic species as it shows less than 90 % ANI values with other closely related species. Also, other phylosiological and biochemical comparisons allowed the phenotypic differentiation of strain JC2948(T) from other members of the genus Burkholderia. Therefore, we suggest that this strain should be classified as the type strain of a novel species of the genus Burkholderia. The name Burkholderia monticola sp. nov. (type strain, JC2948(T) = JCM 19904(T) = KACC 17924(T)) is proposed. © 2015 IUMS.

  15. Unusual Multiple Production of N-Acylhomoserine Lactones a by Burkholderia sp. Strain C10B Isolated from Dentine Caries

    Directory of Open Access Journals (Sweden)

    Share Yuan Goh

    2014-05-01

    Full Text Available Bacteria realize the ability to communicate by production of quorum sensing (QS molecules called autoinducers, which regulate the physiological activities in their ecological niches. The oral cavity could be a potential area for the presence of QS bacteria. In this study, we report the isolation of a QS bacterial isolate C10B from dentine caries. Preliminary screening using Chromobacterium violaceum CV026 biosensor showed that isolate C10B was able to produce N-acylhomoserine lactones (AHLs. This bacterium was further identified as a member of Burkholderia, an opportunistic pathogen. The isolated Burkholderia sp. was confirmed to produce N-hexanoyl-L-homoserine lactone (C6-HSL, N-octanoyl-L-homoserine lactone (C8-HSL, N-decanoyl-L-homoserine lactone (C10-HSL and N-dodecanoyl-L-homoserine lactone (C12-HSL.

  16. Key role for efflux in the preservative susceptibility and adaptive resistance of Burkholderia cepacia complex bacteria.

    Science.gov (United States)

    Rushton, Laura; Sass, Andrea; Baldwin, Adam; Dowson, Christopher G; Donoghue, Denise; Mahenthiralingam, Eshwar

    2013-07-01

    Bacteria from the Burkholderia cepacia complex (Bcc) are encountered as industrial contaminants, and little is known about the species involved or their mechanisms of preservative resistance. Multilocus sequence typing (MLST) revealed that multiple Bcc species may cause contamination, with B. lata (n = 17) and B. cenocepacia (n = 11) dominant within the collection examined. At the strain level, 11 of the 31 industrial sequence types identified had also been recovered from either natural environments or clinical infections. Minimal inhibitory (MIC) and minimum bactericidal (MBC) preservative concentrations varied across 83 selected Bcc strains, with industrial strains demonstrating increased tolerance for dimethylol dimethyl hydantoin (DMDMH). Benzisothiazolinone (BIT), DMDMH, methylisothiazolinone (MIT), a blend of 3:1 methylisothiazolinone-chloromethylisothiazolinone (M-CMIT), methyl paraben (MP), and phenoxyethanol (PH), were all effective anti-Bcc preservatives; benzethonium chloride (BC) and sodium benzoate (SB) were least effective. Since B. lata was the dominant industrial Bcc species, the type strain, 383(T) (LMG 22485(T)), was used to study preservative tolerance. Strain 383 developed stable preservative tolerance for M-CMIT, MIT, BIT, and BC, which resulted in preservative cross-resistance and altered antibiotic susceptibility, motility, and biofilm formation. Transcriptomic analysis of the B. lata 383 M-CMIT-adapted strain demonstrated that efflux played a key role in its M-CMIT tolerance and elevated fluoroquinolone resistance. The role of efflux was corroborated using the inhibitor l-Phe-Arg-β-napthylamide, which reduced the MICs of M-CMIT and ciprofloxacin. In summary, intrinsic preservative tolerance and stable adaptive changes, such as enhanced efflux, play a role in the ability of Bcc bacteria to cause industrial contamination.

  17. Preliminary data on antibacterial activity of Echinacea purpurea-associated bacterial communities against Burkholderia cepacia complex strains, opportunistic pathogens of Cystic Fibrosis patients.

    Science.gov (United States)

    Chiellini, Carolina; Maida, Isabel; Maggini, Valentina; Bosi, Emanuele; Mocali, Stefano; Emiliani, Giovanni; Perrin, Elena; Firenzuoli, Fabio; Mengoni, Alessio; Fani, Renato

    2017-03-01

    Burkholderia cepacia complex bacteria (Bcc) represent a serious threat for immune-compromised patient affected by Cystic Fibrosis (CF) since they are resistant to many substances and to most antibiotics. For this reason, the research of new natural compounds able to inhibit the growth of Bcc strains has raised new interest during the last years. A source of such natural compounds is represented by medicinal plants and, in particular, by bacterial communities associated with these plants able to produce molecules with antimicrobial activity. In this work, a panel of 151 (endophytic) bacteria isolated from three different compartments (rhizospheric soil, roots, and stem/leaves) of the medicinal plant Echinacea purpurea were tested (using the cross-streak method) for their ability to inhibit the growth of 10 Bcc strains. Data obtained revealed that bacteria isolated from the roots of E. purpurea are the most active in the inhibition of Bcc strains, followed by bacteria isolated from the rhizospheric soil, and endophytes from stem/leaf compartment. At the same time, Bcc strains of environmental origin showed a higher resistance toward inhibition than the Bcc strains with clinical (i.e. CF patients) origin. Differences in the inhibition activity of E. purpurea-associated bacteria are mainly linked to the environment -the plant compartment- rather than to their taxonomical position. Copyright © 2016 Elsevier GmbH. All rights reserved.

  18. Burkholderia humisilvae sp. nov., Burkholderia solisilvae sp. nov. and Burkholderia rhizosphaerae sp. nov., isolated from forest soil and rhizosphere soil.

    Science.gov (United States)

    Lee, Jae-Chan; Whang, Kyung-Sook

    2015-09-01

    Strains Y-12(T) and Y-47(T) were isolated from mountain forest soil and strain WR43(T) was isolated from rhizosphere soil, at Daejeon, Korea. The three strains grew at 10-55 °C (optimal growth at 28-30 °C), at pH 3.0-8.0 (optimal growth at pH 6.0) and in the presence of 0-4.0% (w/v) NaCl, growing optimally in the absence of added NaCl. On the basis of 16S rRNA gene sequence analysis, the three strains were found to belong to the genus Burkholderia, showing the closest phylogenetic similarity to Burkholderia diazotrophica JPY461(T) (97.2-97.7%); the similarity between the three sequences ranged from 98.3 to 98.7%. Additionally, the three strains formed a distinct group in phylogenetic trees based on the housekeeping genes recA and gyrB. The predominant ubiquinone was Q-8, the major fatty acids were C16 : 0 and C17  : 0 cyclo and the DNA G+C content of the novel isolates was 61.6-64.4 mol%. DNA-DNA relatedness among the three strains and the type strains of the closest species of the genus Burkholderia was less than 50%. On the basis of 16S rRNA, recA and gyrB gene sequence similarities, chemotaxonomic and phenotypic data, the three strains represent three novel species within the genus Burkholderia, for which the names Burkholderia humisilvae sp. nov. (type strain Y-12(T)= KACC 17601(T) = NBRC 109933(T) = NCAIM B 02543(T)), Burkholderia solisilvae sp. nov. (type strain Y-47(T) = KACC 17602(T)= NBRC 109934(T) = NCAIM B 02539(T)) and Burkholderia rhizosphaerae sp. nov. (type strain WR43(T) = KACC 17603(T) = NBRC 109935(T) = NCAIM B 02541(T)) are proposed.

  19. Members of the genus Burkholderia: good and bad guys

    Science.gov (United States)

    Eberl, Leo; Vandamme, Peter

    2016-01-01

    In the 1990s several biocontrol agents on that contained Burkholderia strains were registered by the United States Environmental Protection Agency (EPA). After risk assessment these products were withdrawn from the market and a moratorium was placed on the registration of Burkholderia-containing products, as these strains may pose a risk to human health. However, over the past few years the number of novel Burkholderia species that exhibit plant-beneficial properties and are normally not isolated from infected patients has increased tremendously. In this commentary we wish to summarize recent efforts that aim at discerning pathogenic from beneficial Burkholderia strains. PMID:27303639

  20. CHROMOSOMAL MULTIPLICITY IN BURKHOLDERIA CEPACIA

    Science.gov (United States)

    We have used CHEF gel electrophoresis to screen preparations of large DNA from different Burkholderia cepacia isolates for the presence of DNA species corresponding to the linearized forms of the three chromosomes of 3.4,2.5, and 0.9 Mb identified in B. cepacia strain 17616. DNA ...

  1. Burkholderia megalochromosomata sp. nov., isolated from grassland soil.

    Science.gov (United States)

    Baek, Inwoo; Seo, Boram; Lee, Imchang; Lee, Kihyun; Park, Sang-Cheol; Yi, Hana; Chun, Jongsik

    2015-03-01

    A Gram-stain negative, rod-shaped, non-spore-forming, obligate aerobic bacterial strain, JC2949(T), was isolated from grassland soil in Gwanak Mountain, Seoul, Republic of Korea. Phylogenetic analysis, based on 16S rRNA sequences, indicated that strain JC2949(T) belongs to the genus Burkholderia, showing highest sequence similarities with Burkholderia grimmiae R27(T) (98.8 %), Burkholderia cordobensis LMG 27620(T) (98.6 %), Burkholderia jiangsuensis MP-1T(T) (98.6 %), Burkholderia zhejiangensis OP-1(T) (98.5 %), Burkholderia humi LMG 22934(T) (97.5 %), Burkholderia terrestris LMG 22937(T) (97.3 %), Burkholderia telluris LMG 22936(T) (97.2 %) and Burkholderia glathei ATCC 29195(T) (97.0 %). The major fatty acids of strain JC2949(T) were C18 : 1ω7c, summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) and C16 : 0. Its predominant polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol and an unknown amino phospholipid. The dominant isoprenoid quinone was ubiquinone Q-8. The pairwise average nucleotide identity values between strain JC2949(T) and the genomes of 30 other species of the genus Burkholderia ranged from 73.4-90.4 %, indicating that the isolate is a novel genomic species within this genus. Based on phenotypic and chemotaxonomic comparisons, it is clear that strain JC2949(T) represents a novel species of the genus Burkholderia. We propose the name for this novel species to be Burkholderia megalochromosomata sp. nov. The type strain is JC2949(T) ( = KACC 17925(T) = JCM 19905(T)). © 2015 IUMS.

  2. The Role of Hydrophobicity and Surface Receptors at Hyphae of Lyophyllum sp. Strain Karsten in the Interaction with Burkholderia terrae BS001 – Implications for Interactions in Soil

    Science.gov (United States)

    Vila, Taissa; Nazir, Rashid; Rozental, Sonia; dos Santos, Giulia M. P.; Calixto, Renata O. R.; Barreto-Bergter, Eliana; Wick, Lukas Y.; van Elsas, Jan Dirk

    2016-01-01

    The soil bacterium Burkholderia terrae strain BS001 can interact with varying soil fungi, using mechanisms that range from the utilization of carbon/energy sources such as glycerol to the ability to reach novel territories in soil via co-migration with growing fungal mycelia. Here, we investigate the intrinsic properties of the B. terrae BS001 interaction with the basidiomycetous soil fungus Lyophyllum sp. strain Karsten. In some experiments, the ascomycetous Trichoderma asperellum 302 was also used. The hyphae of Lyophyllum sp. strain Karsten were largely hydrophilic on water-containing media versus hydrophobic when aerial, as evidenced by contact angle analyses (CA). Co-migration of B. terrae strain BS001 cells with the hyphae of the two fungi occurred preferentially along the - presumably hydrophilic - soil-dwelling hyphae, whereas aerial hyphae did not allow efficient migration, due to reduced thickness of their surrounding mucous films. Moreover, the cell numbers over the length of the hyphae in soil showed an uneven distribution, i.e., the CFU numbers increased from minima at the inoculation point to maximal numbers in the middle of the extended hyphae, then decreasing toward the terminal side. Microscopic analyses of the strain BS001 associations with the Lyophyllum sp. strain Karsten hyphae in the microcosms confirmed the presence of B. terrae BS001 cells on the mucous matter that was present at the hyphal surfaces of the fungi used. Cell agglomerates were found to accumulate at defined sites on the hyphal surfaces, which were coined ‘fungal-interactive’ hot spots. Evidence was further obtained for the contention that receptors for a physical bacterium-fungus interaction occur at the Lyophyllum sp. strain Karsten hyphal surface, in which the specific glycosphingolipid ceramide monohexoside (CMH) plays an important role. Thus, bacterial adherence may be mediated by heterogeneously distributed fungal-specific receptors, implying the CMH moieties. This

  3. 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

  4. Environmental Transmission of the Gut Symbiont Burkholderia to Phloem-Feeding Blissus insularis.

    Science.gov (United States)

    Xu, Yao; Buss, Eileen A; Boucias, Drion G

    2016-01-01

    The plant-phloem-feeding Blissus insularis possesses specialized midgut crypts, which harbor a dense population of the exocellular bacterial symbiont Burkholderia. Most individual B. insularis harbor a single Burkholderia ribotype in their midgut crypts; however, a diverse Burkholderia community exists within a host population. To understand the mechanism underlying the consistent occurrence of various Burkholderia in B. insularis and their specific association, we investigated potential gut symbiont transmission routes. PCR amplification detected a low titer of Burkholderia in adult reproductive tracts; however, fluorescence in situ hybridization assays failed to produce detectable signals in these tracts. Furthermore, no Burkholderia-specific PCR signals were detected in eggs and neonates, suggesting that it is unlikely that B. insularis prenatally transmits gut symbionts via ovarioles. In rearing experiments, most nymphs reared on St. Augustinegrass treated with cultured Burkholderia harbored the cultured Burkholderia strains. Burkholderia was detected in the untreated host grass of B. insularis, and most nymphs reared on untreated grass harbored a Burkholderia ribotype that was closely related to a plant-associated Burkholderia strain. These findings revealed that B. insularis neonates acquired Burkholderia primarily from the environment (i.e., plants and soils), even though the possibility of acquisition via egg surface cannot be excluded. In addition, our study explains how the diverse Burkholderia symbiont community in B. insularis populations can be maintained.

  5. Genes involved in degradation of para-nitrophenol are differentially arranged in form of non-contiguous gene clusters in Burkholderia sp. strain SJ98.

    Directory of Open Access Journals (Sweden)

    Surendra Vikram

    Full Text Available Biodegradation of para-Nitrophenol (PNP proceeds via two distinct pathways, having 1,2,3-benzenetriol (BT and hydroquinone (HQ as their respective terminal aromatic intermediates. Genes involved in these pathways have already been studied in different PNP degrading bacteria. Burkholderia sp. strain SJ98 degrades PNP via both the pathways. Earlier, we have sequenced and analyzed a ~41 kb fragment from the genomic library of strain SJ98. This DNA fragment was found to harbor all the lower pathway genes; however, genes responsible for the initial transformation of PNP could not be identified within this fragment. Now, we have sequenced and annotated the whole genome of strain SJ98 and found two ORFs (viz., pnpA and pnpB showing maximum identity at amino acid level with p-nitrophenol 4-monooxygenase (PnpM and p-benzoquinone reductase (BqR. Unlike the other PNP gene clusters reported earlier in different bacteria, these two ORFs in SJ98 genome are physically separated from the other genes of PNP degradation pathway. In order to ascertain the identity of ORFs pnpA and pnpB, we have performed in-vitro assays using recombinant proteins heterologously expressed and purified to homogeneity. Purified PnpA was found to be a functional PnpM and transformed PNP into benzoquinone (BQ, while PnpB was found to be a functional BqR which catalyzed the transformation of BQ into hydroquinone (HQ. Noticeably, PnpM from strain SJ98 could also transform a number of PNP analogues. Based on the above observations, we propose that the genes for PNP degradation in strain SJ98 are arranged differentially in form of non-contiguous gene clusters. This is the first report for such arrangement for gene clusters involved in PNP degradation. Therefore, we propose that PNP degradation in strain SJ98 could be an important model system for further studies on differential evolution of PNP degradation functions.

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

    Directory of Open Access Journals (Sweden)

    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.

  7. Genome-centric evaluation of Burkholderia sp. strain SRS-W-2-2016 resistant to high concentrations of uranium and nickel isolated from the Savannah River Site (SRS, USA

    Directory of Open Access Journals (Sweden)

    Ashish Pathak

    2017-06-01

    Full Text Available Savannah River Site (SRS, an approximately 800-km2 former nuclear weapons production facility located near Aiken, SC remains co-contaminated by heavy metals and radionuclides. To gain a better understanding on microbially-mediated bioremediation mechanisms, several bacterial strains resistant to high concentrations of Uranium (U and Nickel (Ni were isolated from the Steeds Pond soils located within the SRS site. One of the isolated strains, designated as strain SRS-W-2-2016, grew robustly on both U and Ni. To fully understand the arsenal of metabolic functions possessed by this strain, a draft whole genome sequence (WGS was obtained, assembled, annotated and analyzed. Genome-centric evaluation revealed the isolate to belong to the Burkholderia genus with close affiliation to B. xenovorans LB400, an aggressive polychlorinated biphenyl-degrader. At a coverage of 90×, the genome of strain SRS-W-2-2016 consisted of 8,035,584 bases with a total number of 7071 putative genes assembling into 191 contigs with an N50 contig length of 134,675 bases. Several gene homologues coding for resistance to heavy metals/radionuclides were identified in strain SRS-W-2-2016, such as a suite of outer membrane efflux pump proteins similar to nickel/cobalt transporter regulators, peptide/nickel transport substrate and ATP-binding proteins, permease proteins, and a high-affinity nickel-transport protein. Also noteworthy were two separate gene fragments in strain SRS-W-2-2016 homologous to the spoT gene; recently correlated with bacterial tolerance to U. Additionally, a plethora of oxygenase genes were also identified in the isolate, potentially involved in the breakdown of organic compounds facilitating the strain's successful colonization and survival in the SRS co-contaminated soils. The WGS project of Burkholderia sp. strain SRS-W-2-2016 is available at DDBJ/ENA/GenBank under the accession #MSDV00000000.

  8. Burkholderia cordobensis sp. nov., from agricultural soils.

    Science.gov (United States)

    Draghi, Walter O; Peeters, Charlotte; Cnockaert, Margo; Snauwaert, Cindy; Wall, Luis G; Zorreguieta, Angeles; Vandamme, Peter

    2014-06-01

    Two Gram-negative, rod-shaped bacteria were isolated from agricultural soils in Córdoba province in central Argentina. Their 16S rRNA gene sequences demonstrated that they belong to the genus Burkholderia, with Burkholderia zhejiangensis as most closely related formally named species; this relationship was confirmed through comparative gyrB sequence analysis. Whole-cell fatty acid analysis supported their assignment to the genus Burkholderia. Burkholderia sp. strain YI23, for which a whole-genome sequence is available, represents the same taxon, as demonstrated by its highly similar 16S rRNA (100% similarity) and gyrB (99.1-99.7%) gene sequences. The results of DNA-DNA hybridization experiments and physiological and biochemical characterization further substantiated the genotypic and phenotypic distinctiveness of the Argentinian soil isolates, for which the name Burkholderia cordobensis sp. nov. is proposed, with strain MMP81(T) ( = LMG 27620(T) = CCUG 64368(T)) as the type strain. © 2014 IUMS.

  9. Characterization of Clinically Attenuated Burkholderia mallei by Whole-Genome Sequencing: Candidate Strain for Exclusion from Select Agent Lists

    Science.gov (United States)

    2008-04-01

    subjected to WGS and the resulting data compared those from the virulent ATCC 23344 strain[22] which was know to have caused recent near- death in a...Human Services ASfPaR (2007) http://www.hhs. gov/aspr/barda/documents/phemce_implplan_041607final.pdf. 11. Koenig RL (2006) The Fourth Horseman : One Man’s

  10. Phylogenomic Study of Burkholderia glathei-like Organisms, Proposal of 13 Novel Burkholderia Species and Emended Descriptions of Burkholderia sordidicola, Burkholderia zhejiangensis, and Burkholderia grimmiae

    OpenAIRE

    Peeters, Charlotte; Meier-Kolthoff, Jan P.; Verheyde, Bart; De Brandt, Evie; Cooper, Vaughn S.; Vandamme, Peter

    2016-01-01

    Partial gyrB gene sequence analysis of 17 isolates from human and environmental sources revealed 13 clusters of strains and identified them as Burkholderia glathei Glade (BGC) bacteria. The taxonomic status of these clusters was examined by whole-genome sequence analysis, determination of the G+C content, whole-cell fatty acid analysis and biochemical characterization. The whole-genome sequence-based phylogeny was assessed using the Genome Blast Distance Phylogeny (GBDP) method and an extende...

  11. Activity of Tobramycin against Cystic Fibrosis Isolates of Burkholderia cepacia Complex Grown as Biofilms.

    Science.gov (United States)

    Kennedy, Sarah; Beaudoin, Trevor; Yau, Yvonne C W; Caraher, Emma; Zlosnik, James E A; Speert, David P; LiPuma, John J; Tullis, Elizabeth; Waters, Valerie

    2016-01-01

    Pulmonary infection with Burkholderia cepacia complex in cystic fibrosis (CF) patients is associated with more-rapid lung function decline and earlier death than in CF patients without this infection. In this study, we used confocal microscopy to visualize the effects of various concentrations of tobramycin, achievable with systemic and aerosolized drug administration, on mature B. cepacia complex biofilms, both in the presence and absence of CF sputum. After 24 h of growth, biofilm thickness was significantly reduced by exposure to 2,000 μg/ml of tobramycin for Burkholderia cepacia, Burkholderia multivorans, and Burkholderia vietnamiensis; 200 μg/ml of tobramycin was sufficient to reduce the thickness of Burkholderia dolosa biofilm. With a more mature 48-h biofilm, significant reductions in thickness were seen with tobramycin at concentrations of ≥100 μg/ml for all Burkholderia species. In addition, an increased ratio of dead to live cells was observed in comparison to control with tobramycin concentrations of ≥200 μg/ml for B. cepacia and B. dolosa (24 h) and ≥100 μg/ml for Burkholderia cenocepacia and B. dolosa (48 h). Although sputum significantly increased biofilm thickness, tobramycin concentrations of 1,000 μg/ml were still able to significantly reduce biofilm thickness of all B. cepacia complex species with the exception of B. vietnamiensis. In the presence of sputum, 1,000 μg/ml of tobramycin significantly increased the dead-to-live ratio only for B. multivorans compared to control. In summary, although killing is attenuated, high-dose tobramycin can effectively decrease the thickness of B. cepacia complex biofilms, even in the presence of sputum, suggesting a possible role as a suppressive therapy in CF. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  12. Test of aerobic TCE degradation by willows (Salix viminalis) and willows inoculated with TCE-cometabolizing strains of Burkholderia cepacia.

    Science.gov (United States)

    Clausen, Lauge Peter Westergaard; Broholm, Mette Martina; Gosewinkel, Ulrich; Trapp, Stefan

    2017-08-01

    Trichloroethylene (TCE) is a widespread soil and groundwater pollutant and clean-up is often problematic and expensive. Phytoremediation may be a cost-effective solution at some sites. This study investigates TCE degradation by willows (S. viminalis) and willows inoculated with three strains of B. cepacia (301C, PR1-31 and VM1330-pTOM), using chloride formation as an indicator of dehalogenation. Willows were grown in non-sterile, hydroponic conditions for 3 weeks in chloride-free nutrient solution spiked with TCE. TCE was added weekly due to rapid loss by volatilization. Chloride and TCE in solution were measured every 2-3 days and chloride and metabolite concentrations in plants were measured at test termination. Based on transpiration, no tree toxicity of TCE exposure was observed. However, trees grown in chloride-free solution showed severely inhibited transpiration. No or very little chloride was formed during the test, and levels of chloride in TCE-exposed trees were not elevated. Chloride concentrations in chloride containing TCE-free nutrient solution doubled within 23 days, indicating active exclusion of chloride by root cell membranes. Only traces of TCE-metabolites were detected in plant tissue. We conclude that TCE is not, or to a limited extent (less than 3%), aerobically degraded by the willow trees. The three strains of B. cepacia did not enhance TCE mineralization. Future successful application of rhizo- and phytodegradation of TCE requires measures to be taken to improve the degradation rates.

  13. Use of a Burkholderia cenocepacia ABTS Oxidizer in a Microbial Fuel Cell

    Science.gov (United States)

    Microbial fuel cells (MFCs) often use biological processes to generate electrons from organic material contained in the anode chamber and abiotic processes employing atmospheric oxygen as the oxidant in the cathode chamber. This study investigated the accumulation of an oxidant in bacterial cultures...

  14. Draft genome sequence of Burkholderia sordidicola S170, a potential plant growth promoter isolated from coniferous forest soil in the Czech Republic

    DEFF Research Database (Denmark)

    Lladó, Salvador; Xu, Zhuofei; Sørensen, Søren Johannes

    2014-01-01

    Burkholderia species are key players in the accumulation of carbon from cellulose decomposition in coniferous forest ecosystems. We report here the draft genome of Burkholderia sordidicola strain S170, containing features associated with known genes involved in plant growth promotion...

  15. Burkholderia humptydooensis sp. nov., a New Species Related to Burkholderia thailandensis and the Fifth Member of the Burkholderia pseudomallei Complex.

    Science.gov (United States)

    Tuanyok, Apichai; Mayo, Mark; Scholz, Holger; Hall, Carina M; Allender, Christopher J; Kaestli, Mirjam; Ginther, Jennifer; Spring-Pearson, Senanu; Bollig, Molly C; Stone, Joshua K; Settles, Erik W; Busch, Joseph D; Sidak-Loftis, Lindsay; Sahl, Jason W; Thomas, Astrid; Kreutzer, Lisa; Georgi, Enrico; Gee, Jay E; Bowen, Richard A; Ladner, Jason T; Lovett, Sean; Koroleva, Galina; Palacios, Gustavo; Wagner, David M; Currie, Bart J; Keim, Paul

    2017-03-01

    During routine screening for Burkholderia pseudomallei from water wells in northern Australia in areas where it is endemic, Gram-negative bacteria (strains MSMB43 T , MSMB121, and MSMB122) with a similar morphology and biochemical pattern to B. pseudomallei and B. thailandensis were coisolated with B. pseudomallei on Ashdown's selective agar. To determine the exact taxonomic position of these strains and to distinguish them from B. pseudomallei and B. thailandensis , they were subjected to a series of phenotypic and molecular analyses. Biochemical and fatty acid methyl ester analysis was unable to distinguish B. humptydooensis sp. nov. from closely related species. With matrix-assisted laser desorption ionization-time of flight analysis, all isolates grouped together in a cluster separate from other Burkholderia spp. 16S rRNA and recA sequence analyses demonstrated phylogenetic placement for B. humptydooensis sp. nov. in a novel clade within the B. pseudomallei group. Multilocus sequence typing (MLST) analysis of the three isolates in comparison with MLST data from 3,340 B. pseudomallei strains and related taxa revealed a new sequence type (ST318). Genome-to-genome distance calculations and the average nucleotide identity of all isolates to both B. thailandensis and B. pseudomallei , based on whole-genome sequences, also confirmed B. humptydooensis sp. nov. as a novel Burkholderia species within the B. pseudomallei complex. Molecular analyses clearly demonstrated that strains MSMB43 T , MSMB121, and MSMB122 belong to a novel Burkholderia species for which the name Burkholderia humptydooensis sp. nov. is proposed, with the type strain MSMB43 T (American Type Culture Collection BAA-2767; Belgian Co-ordinated Collections of Microorganisms LMG 29471; DDBJ accession numbers CP013380 to CP013382). IMPORTANCE Burkholderia pseudomallei is a soil-dwelling bacterium and the causative agent of melioidosis. The genus Burkholderia consists of a diverse group of species, with

  16. Burkholderia thailandensis: Genetic Manipulation.

    Science.gov (United States)

    Garcia, Erin C

    2017-05-16

    Burkholderia thailandensis is a Gram-negative bacterium endemic to Southeast Asian and northern Australian soils. It is non-pathogenic; therefore, it is commonly used as a model organism for the related human pathogens Burkholderia mallei and Burkholderia pseudomallei. B. thailandensis is relatively easily genetically manipulated and a variety of robust genetic tools can be used in this organism. This unit describes protocols for conjugation, natural transformation, mini-Tn7 insertion, and allelic exchange in B. thailandensis. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

  17. Plant growth-promoting Burkholderia species isolated from annual ryegrass in Portuguese soils.

    Science.gov (United States)

    Castanheira, N; Dourado, A C; Kruz, S; Alves, P I L; Delgado-Rodríguez, A I; Pais, I; Semedo, J; Scotti-Campos, P; Sánchez, C; Borges, N; Carvalho, G; Barreto Crespo, M T; Fareleira, P

    2016-03-01

    To search for culturable Burkholderia species associated with annual ryegrass in soils from natural pastures in Portugal, with plant growth-promoting effects. Annual ryegrass seedlings were used to trap Burkholderia from two different soils in laboratory conditions. A combined approach using genomic fingerprinting and sequencing of 16S rRNA and recA genes resulted in the identification of Burkholderia strains belonging to the species Burkholderia graminis, Burkholderia fungorum and the Burkholderia cepacia complex. Most strains were able to solubilize mineral phosphate and to synthesize indole acetic acid; some of them could produce siderophores and antagonize the phytopathogenic oomycete, Phytophthora cinnamomi. A strain (G2Bd5) of B. graminis was selected for gnotobiotic plant inoculation experiments. The main effects were the stimulation of root growth and enhancement of leaf lipid synthesis and turnover. Fluorescence in situ hybridization and confocal laser microscopy evidenced that strain G2Bd5 is a rhizospheric and endophytic colonizer of annual ryegrass. This work revealed that annual ryegrass can naturally associate with members of the genus Burkholderia. A novel plant growth promoting strain of B. graminis was obtained. The novel strain belongs to the plant-associated Burkholderia cluster and is a promising candidate for exploitation as plant inoculant in field conditions. © 2015 The Society for Applied Microbiology.

  18. Selection and evaluation of reference genes for RT-qPCR expression studies on Burkholderia tropica strain Ppe8, a sugarcane-associated diazotrophic bacterium grown with different carbon sources or sugarcane juice.

    Science.gov (United States)

    da Silva, Paula Renata Alves; Vidal, Marcia Soares; de Paula Soares, Cleiton; Polese, Valéria; Simões-Araújo, Jean Luís; Baldani, José Ivo

    2016-11-01

    Among the members of the genus Burkholderia, Burkholderia tropica has the ability to fix nitrogen and promote sugarcane plant growth as well as act as a biological control agent. There is little information about how this bacterium metabolizes carbohydrates as well as those carbon sources found in the sugarcane juice that accumulates in stems during plant growth. Reverse transcription quantitative PCR (RT-qPCR) can be used to evaluate changes in gene expression during bacterial growth on different carbon sources. Here we tested the expression of six reference genes, lpxC, gyrB, recA, rpoA, rpoB, and rpoD, when cells were grown with glucose, fructose, sucrose, mannitol, aconitic acid, and sugarcane juice as carbon sources. The lpxC, gyrB, and recA were selected as the most stable reference genes based on geNorm and NormFinder software analyses. Validation of these three reference genes during strain Ppe8 growth on the same carbon sources showed that genes involved in glycogen biosynthesis (glgA, glgB, glgC) and trehalose biosynthesis (treY and treZ) were highly expressed when Ppe8 was grown in aconitic acid relative to other carbon sources, while otsA expression (trehalose biosynthesis) was reduced with all carbon sources. In addition, the expression level of the ORF_6066 (gluconolactonase) gene was reduced on sugarcane juice. The results confirmed the stability of the three selected reference genes (lpxC, gyrB, and recA) during the RT-qPCR and also their robustness by evaluating the relative expression of genes involved in glycogen and trehalose biosynthesis when strain Ppe8 was grown on different carbon sources and sugarcane juice.

  19. Diverse Burkholderia Species Isolated from Soils in the Southern United States with No Evidence of B. pseudomallei.

    Directory of Open Access Journals (Sweden)

    Carina M Hall

    Full Text Available The global distribution of the soil-dwelling bacterium Burkholderia pseudomallei, causative agent of melioidosis, is poorly understood. We used established culturing methods developed for B. pseudomallei to isolate Burkholderia species from soil collected at 18 sampling sites in three states in the southern United States (Arizona (n = 4, Florida (n = 7, and Louisiana (n = 7. Using multi-locus sequence typing (MLST of seven genes, we identified 35 Burkholderia isolates from these soil samples. All species belonged to the B. cepacia complex (Bcc, including B. cenocepacia, B. cepacia, B. contaminans, B. diffusa, B. metallica, B. seminalis, B. vietnamiensis and two unnamed members of the Bcc. The MLST analysis provided a high level of resolution among and within these species. Despite previous clinical cases within the U.S. involving B. pseudomallei and its close phylogenetic relatives, we did not isolate any of these taxa. The Bcc contains a number of opportunistic pathogens that cause infections in cystic fibrosis patients. Interestingly, we found that B. vietnamiensis was present in soil from all three states, suggesting it may be a common component in southern U.S. soils. Most of the Burkholderia isolates collected in this study were from Florida (30/35; 86%, which may be due to the combination of relatively moist, sandy, and acidic soils found there compared to the other two states. We also investigated one MLST gene, recA, for its ability to identify species within Burkholderia. A 365bp fragment of recA recovered nearly the same species-level identification as MLST, thus demonstrating its cost effective utility when conducting environmental surveys for Burkholderia. Although we did not find B. pseudomallei, our findings document that other diverse Burkholderia species are present in soils in the southern United States.

  20. Diverse Burkholderia Species Isolated from Soils in the Southern United States with No Evidence of B. pseudomallei.

    Science.gov (United States)

    Hall, Carina M; Busch, Joseph D; Shippy, Kenzie; Allender, Christopher J; Kaestli, Mirjam; Mayo, Mark; Sahl, Jason W; Schupp, James M; Colman, Rebecca E; Keim, Paul; Currie, Bart J; Wagner, David M

    2015-01-01

    The global distribution of the soil-dwelling bacterium Burkholderia pseudomallei, causative agent of melioidosis, is poorly understood. We used established culturing methods developed for B. pseudomallei to isolate Burkholderia species from soil collected at 18 sampling sites in three states in the southern United States (Arizona (n = 4), Florida (n = 7), and Louisiana (n = 7)). Using multi-locus sequence typing (MLST) of seven genes, we identified 35 Burkholderia isolates from these soil samples. All species belonged to the B. cepacia complex (Bcc), including B. cenocepacia, B. cepacia, B. contaminans, B. diffusa, B. metallica, B. seminalis, B. vietnamiensis and two unnamed members of the Bcc. The MLST analysis provided a high level of resolution among and within these species. Despite previous clinical cases within the U.S. involving B. pseudomallei and its close phylogenetic relatives, we did not isolate any of these taxa. The Bcc contains a number of opportunistic pathogens that cause infections in cystic fibrosis patients. Interestingly, we found that B. vietnamiensis was present in soil from all three states, suggesting it may be a common component in southern U.S. soils. Most of the Burkholderia isolates collected in this study were from Florida (30/35; 86%), which may be due to the combination of relatively moist, sandy, and acidic soils found there compared to the other two states. We also investigated one MLST gene, recA, for its ability to identify species within Burkholderia. A 365bp fragment of recA recovered nearly the same species-level identification as MLST, thus demonstrating its cost effective utility when conducting environmental surveys for Burkholderia. Although we did not find B. pseudomallei, our findings document that other diverse Burkholderia species are present in soils in the southern United States.

  1. Isolation and Characterization of Burkholderia rinojensis sp. nov., a Non-Burkholderia cepacia Complex Soil Bacterium with Insecticidal and Miticidal Activities

    Science.gov (United States)

    Fernandez, Lorena E.; Koivunen, Marja; Yang, April; Flor-Weiler, Lina; Marrone, Pamela G.

    2013-01-01

    Isolate A396, a bacterium isolated from a Japanese soil sample demonstrated strong insecticidal and miticidal activities in laboratory bioassays. The isolate was characterized through biochemical methods, fatty acid methyl ester (FAME) analysis, sequencing of 16S rRNA, multilocus sequence typing and analysis, and DNA-DNA hybridization. FAME analysis matched A396 to Burkholderia cenocepacia, but this result was not confirmed by 16S rRNA or DNA-DNA hybridization. 16S rRNA sequencing indicated closest matches with B. glumae and B. plantarii. DNA-DNA hybridization experiments with B. plantarii, B. glumae, B. multivorans, and B. cenocepacia confirmed the low genetic similarity (11.5 to 37.4%) with known members of the genus. PCR-based screening showed that A396 lacks markers associated with members of the B. cepacia complex. Bioassay results indicated two mechanisms of action: through ingestion and contact. The isolate effectively controlled beet armyworms (Spodoptera exigua; BAW) and two-spotted spider mites (Tetranychus urticae; TSSM). In diet overlay bioassays with BAW, 1% to 4% (vol/vol) dilution of the whole-cell broth caused 97% to 100% mortality 4 days postexposure, and leaf disc treatment bioassays attained 75% ± 22% mortality 3 days postexposure. Contact bioassays led to 50% larval mortality, as well as discoloration, stunting, and failure to molt. TSSM mortality reached 93% in treated leaf discs. Activity was maintained in cell-free supernatants and after heat treatment (60°C for 2 h), indicating that a secondary metabolite or excreted thermostable enzyme might be responsible for the activity. Based on these results, we describe the novel species Burkholderia rinojensis, a good candidate for the development of a biocontrol product against insect and mite pests. PMID:24096416

  2. An efficient system for the generation of marked genetic mutants in members of the genus Burkholderia.

    Science.gov (United States)

    Shastri, Sravanthi; Spiewak, Helena L; Sofoluwe, Aderonke; Eidsvaag, Vigdis A; Asghar, Atif H; Pereira, Tyrone; Bull, Edward H; Butt, Aaron T; Thomas, Mark S

    2017-01-01

    To elucidate the function of a gene in bacteria it is vital that targeted gene inactivation (allelic replacement) can be achieved. Allelic replacement is often carried out by disruption of the gene of interest by insertion of an antibiotic-resistance marker followed by subsequent transfer of the mutant allele to the genome of the host organism in place of the wild-type gene. However, due to their intrinsic resistance to many antibiotics only selected antibiotic-resistance markers can be used in members of the genus Burkholderia, including the Burkholderia cepacia complex (Bcc). Here we describe the construction of improved antibiotic-resistance cassettes that specify resistance to kanamycin, chloramphenicol or trimethoprim effectively in the Bcc and related species. These were then used in combination with and/or to construct a series enhanced suicide vectors, pSHAFT2, pSHAFT3 and pSHAFT-GFP to facilitate effective allelic replacement in the Bcc. Validation of these improved suicide vectors was demonstrated by the genetic inactivation of selected genes in the Bcc species Burkholderia cenocepacia and B. lata, and in the non-Bcc species, B. thailandensis. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  3. Understanding the Pathogenicity of Burkholderia contaminans, an Emerging Pathogen in Cystic Fibrosis

    Czech Academy of Sciences Publication Activity Database

    Nunvář, J.; Kalferstová, L.; Bloodworth, R.A.M.; Kolář, Michal; Degrossi, J.; Lubovich, S.; Cardona, S.T.; Dřevínek, P.

    2016-01-01

    Roč. 11, č. 8 (2016), č. článku e0160975. E-ISSN 1932-6203 R&D Projects: GA MZd(CZ) NT12405; GA MZd(CZ) NV15-28017A Institutional support: RVO:68378050 Keywords : quorum sensing systems * cepacia complex * pseudomonas-aeruginosa * strain ms14 * spontaneous mutations * molecular-spectrum * escherichia-coli * structural basis * sequence data * cenocepacia Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.806, year: 2016

  4. Common duckweed (Lemna minor is a versatile high-throughput infection model for the Burkholderia cepacia complex and other pathogenic bacteria.

    Directory of Open Access Journals (Sweden)

    Euan L S Thomson

    Full Text Available Members of the Burkholderia cepacia complex (Bcc have emerged in recent decades as problematic pulmonary pathogens of cystic fibrosis (CF patients, with severe infections progressing to acute necrotizing pneumonia and sepsis. This study presents evidence that Lemna minor (Common duckweed is useful as a plant model for the Bcc infectious process, and has potential as a model system for bacterial pathogenesis in general. To investigate the relationship between Bcc virulence in duckweed and Galleria mellonella (Greater wax moth larvae, a previously established Bcc infection model, a duckweed survival assay was developed and used to determine LD50 values. A strong correlation (R(2 = 0.81 was found between the strains' virulence ranks in the two infection models, suggesting conserved pathways in these vastly different hosts. To broaden the application of the duckweed model, enteropathogenic Escherichia coli (EPEC and five isogenic mutants with previously established LD50 values in the larval model were tested against duckweed, and a strong correlation (R(2 = 0.93 was found between their raw LD50 values. Potential virulence factors in B. cenocepacia K56-2 were identified using a high-throughput screen against single duckweed plants. In addition to the previously characterized antifungal compound (AFC cluster genes, several uncharacterized genes were discovered including a novel lysR regulator, a histidine biosynthesis gene hisG, and a gene located near the gene encoding the recently characterized virulence factor SuhB(Bc. Finally, to demonstrate the utility of this model in therapeutic applications, duckweed was rescued from Bcc infection by treating with bacteriophage at 6-h intervals. It was observed that phage application became ineffective at a timepoint that coincided with a sharp increase in bacterial invasion of plant tissue. These results indicate that common duckweed can serve as an effective infection model for the investigation of bacterial

  5. BIOAUGMENTATION WITH BURKHOLDERIA CEPACIA PR1301 FOR IN SITU BIOREMEDIATION OF TRICHLOROETHYLENE CONTAMINATED GROUNDWATER (RESEARCH BRIEF)

    Science.gov (United States)

    A pilot field study was conducted at the Moffett Federal Airfield, Mountain View, California, to determine whether effective in-situ aerobic cometabolic biodegradation of TCE could be accomplished through bioaugmentation with a genetically modified strain of Burkholderia cepacia ...

  6. Burkholderia caballeronis sp. nov., a nitrogen fixing species isolated from tomato (Lycopersicon esculentum) with the ability to effectively nodulate Phaseolus vulgaris.

    Science.gov (United States)

    Martínez-Aguilar, Lourdes; Salazar-Salazar, Corelly; Méndez, Rafael Díaz; Caballero-Mellado, Jesús; Hirsch, Ann M; Vásquez-Murrieta, María Soledad; Estrada-de los Santos, Paulina

    2013-12-01

    During a survey of Burkholderia species with potential use in agrobiotechnology, a group of 12 strains was isolated from the rhizosphere and rhizoplane of tomato plants growing in Mexico (Nepantla, Mexico State). A phylogenetic analysis of 16S rRNA gene sequences showed that the strains are related to Burkholderia kururiensis and Burkholderia mimosarum (97.4 and 97.1 %, respectively). However, they induced effective nitrogen-fixing nodules on roots of Phaseolus vulgaris. Based on polyphasic taxonomy, the group of strains represents a novel species for which the name Burkholderia caballeronis sp. nov. is proposed. The type species is TNe-841(T) (= LMG 26416(T) = CIP 110324(T)).

  7. Characterisation of the simultaneous molybdenum reduction and glyphosate degradation by Burkholderia vietnamiensis AQ5-12 and Burkholderia sp. AQ5-13.

    Science.gov (United States)

    Manogaran, Motharasan; Ahmad, Siti Aqlima; Yasid, Nur Adeela; Yakasai, Hafeez Muhammad; Shukor, Mohd Yunus

    2018-02-01

    In this novel study, we report on the use of two molybdenum-reducing bacteria with the ability to utilise the herbicide glyphosate as the phosphorus source. The bacteria reduced sodium molybdate to molybdenum blue (Mo-blue), a colloidal and insoluble product, which is less toxic. The characterisation of the molybdenum-reducing bacteria was carried out using resting cells immersed in low-phosphate molybdenum media. Two glyphosate-degrading bacteria, namely Burkholderia vietnamiensis AQ5-12 and Burkholderia sp. AQ5-13, were able to use glyphosate as a phosphorous source to support molybdenum reduction to Mo-blue. The bacteria optimally reduced molybdenum between the pHs of 6.25 and 8. The optimum concentrations of molybdate for strain Burkholderia vietnamiensis strain AQ5-12 was observed to be between 40 and 60 mM, while for Burkholderia sp. AQ5-13, the optimum molybdate concentration occurred between 40 and 50 mM. Furthermore, 5 mM of phosphate was seen as the optimum concentration supporting molybdenum reduction for both bacteria. The optimum temperature aiding Mo-blue formation ranged from 30 to 40 °C for Burkholderia vietnamiensis strain AQ5-12, whereas for Burkholderia sp. AQ5-13, the range was from 35 to 40 °C. Glucose was the best electron donor for supporting molybdate reduction, followed by sucrose, fructose and galactose for both strains. Ammonium sulphate was the best nitrogen source in supporting molybdenum reduction. Interestingly, increasing the glyphosate concentrations beyond 100 and 300 ppm for Burkholderia vietnamiensis strain AQ5-12 and Burkholderia sp. AQ5-13, respectively, significantly inhibited molybdenum reduction. The ability of these bacteria to reduce molybdenum while degrading glyphosate is a useful process for the bioremediation of both toxicants.

  8. Burkholderia species associated with legumes of Chiapas, Mexico, exhibit stress tolerance and growth in aromatic compounds.

    Science.gov (United States)

    de León-Martínez, José A; Yañez-Ocampo, Gustavo; Wong-Villarreal, Arnoldo

    Leguminous plants have received special interest for the diversity of β-proteobacteria in their nodules and are promising candidates for biotechnological applications. In this study, 15 bacterial strains were isolated from the nodules of the following legumes: Indigofera thibaudiana, Mimosa diplotricha, Mimosa albida, Mimosa pigra, and Mimosa pudica, collected in 9 areas of Chiapas, Mexico. The strains were grouped into four profiles of genomic fingerprints through BOX-PCR and identified based on their morphology, API 20NE biochemical tests, sequencing of the 16S rRNA, nifH and nodC genes as bacteria of the Burkholderia genus, genetically related to Burkholderia phenoliruptrix, Burkholderia phymatum, Burkholderia sabiae, and Burkholderia tuberum. The Burkholderia strains were grown under stress conditions with 4% NaCl, 45°C, and benzene presence at 0.1% as the sole carbon source. This is the first report on the isolation of these nodulating species of the Burkholderia genus in legumes in Mexico. Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

  9. Burkholderia pseudomallei traced to water treatment plant in Australia.

    Science.gov (United States)

    Inglis, T J; Garrow, S C; Henderson, M; Clair, A; Sampson, J; O'Reilly, L; Cameron, B

    2000-01-01

    Burkholderia pseudomallei was isolated from environmental specimens 1 year after an outbreak of acute melioidosis in a remote coastal community in northwestern Australia. B. pseudomallei was isolated from a water storage tank and from spray formed in a pH-raising aerator unit. Pulsed-field gel electrophoresis confirmed the aerator and storage tank isolates were identical to the outbreak strain, WKo97.

  10. Burkholderia susongensis sp. nov., a mineral-weathering bacterium isolated from weathered rock surface.

    Science.gov (United States)

    Gu, Jia-Yu; Zang, Sheng-Gang; Sheng, Xia-Fang; He, Lin-Yan; Huang, Zhi; Wang, Qi

    2015-03-01

    A novel type of mineral-weathering bacterium was isolated from the weathered surface of rock (mica schist) collected from Susong (Anhui, China). Cells of strain L226(T) were Gram-stain-negative. The strain grew optimally at 30 °C, with 1 % (w/v) NaCl and at pH 7.0 in trypticase soy broth. On the basis of 16S rRNA gene phylogeny, strain L226(T) was shown to belong to the genus Burkholderia and the closest phylogenetic relatives were Burkholderia sprentiae WSM5005(T) (98.3 %), Burkholderia acidipaludis NBRC 101816(T) (98.2 %), Burkholderia tuberum STM678(T) (97.2 %) and Burkholderia diazotrophica JPY461(T) (97.1 %). The DNA G+C content was 63.5 mol% and the respiratory quinone was Q-8. The major fatty acids were C16 : 0, C17 : 0 cyclo and C19 : 0 cyclo ω8c. The polar lipid profile of strain L226(T) consisted of a mixture of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, unknown lipids and unidentified aminophospholipids. Based on the low level of DNA-DNA relatedness (ranging from 25.8 % to 34.4 %) to the tested type strains of species of the genus Burkholderia and unique phenotypic characteristics, it is suggested that strain L226(T) represents a novel species of the genus Burkholderia, for which the name Burkholderia susongensis sp. nov., is proposed. The type strain is L226(T) ( = CCTCC AB2014142(T) = JCM 30231(T)). © 2015 IUMS.

  11. Unusual distribution of Burkholderia cepacia complex species in Danish cystic fibrosis clinics may stem from restricted transmission between patients

    DEFF Research Database (Denmark)

    Nørskov-Lauritsen, Niels; Johansen, Helle Krogh; Fenger, Mette G

    2010-01-01

    Forty-four of 48 Burkholderia cepacia complex strains cultured from Danish cystic fibrosis patients were Burkholderia multivorans, a distribution of species that has not been reported before. Although cases of cross infections were demonstrated, no major epidemic clone was found. The species...

  12. 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

  13. Exploring the HME and HAE1 efflux systems in the genus Burkholderia

    Directory of Open Access Journals (Sweden)

    Pasca Maria

    2010-06-01

    Full Text Available Abstract Background The genus Burkholderia includes a variety of species with opportunistic human pathogenic strains, whose increasing global resistance to antibiotics has become a public health problem. In this context a major role could be played by multidrug efflux pumps belonging to Resistance Nodulation Cell-Division (RND family, which allow bacterial cells to extrude a wide range of different substrates, including antibiotics. This study aims to i identify rnd genes in the 21 available completely sequenced Burkholderia genomes, ii analyze their phylogenetic distribution, iii define the putative function(s that RND proteins perform within the Burkholderia genus and iv try tracing the evolutionary history of some of these genes in Burkholderia. Results BLAST analysis of the 21 Burkholderia sequenced genomes, using experimentally characterized ceoB sequence (one of the RND family counterpart in the genus Burkholderia as probe, allowed the assembly of a dataset comprising 254 putative RND proteins. An extensive phylogenetic analysis revealed the occurrence of several independent events of gene loss and duplication across the different lineages of the genus Burkholderia, leading to notable differences in the number of paralogs between different genomes. A putative substrate [antibiotics (HAE1 proteins/heavy-metal (HME proteins] was also assigned to the majority of these proteins. No correlation was found between the ecological niche and the lifestyle of Burkholderia strains and the number/type of efflux pumps they possessed, while a relation can be found with genome size and taxonomy. Remarkably, we observed that only HAE1 proteins are mainly responsible for the different number of proteins observed in strains of the same species. Data concerning both the distribution and the phylogenetic analysis of the HAE1 and HME in the Burkholderia genus allowed depicting a likely evolutionary model accounting for the evolution and spreading of HME and HAE

  14. Burkholderia in gladiool lastige bacterie

    NARCIS (Netherlands)

    Kok, B.J.; Aanholt, van J.T.M.

    2009-01-01

    In de bollen- en bloementeelt van gladiolen komt de laatste jaren de bacterieziekte Burkholderia gladiola voor die onder vochtige warme omstandigheden veel uitval veroorzaken. PPO onderzocht een aantal maatregelen om de ziekte in kralen, pitten en knollen te bestrijden

  15. 16S rRNA gene-based phylogenetic microarray for simultaneous identification of members of the genus Burkholderia.

    Science.gov (United States)

    Schönmann, Susan; Loy, Alexander; Wimmersberger, Céline; Sobek, Jens; Aquino, Catharine; Vandamme, Peter; Frey, Beat; Rehrauer, Hubert; Eberl, Leo

    2009-04-01

    For cultivation-independent and highly parallel analysis of members of the genus Burkholderia, an oligonucleotide microarray (phylochip) consisting of 131 hierarchically nested 16S rRNA gene-targeted oligonucleotide probes was developed. A novel primer pair was designed for selective amplification of a 1.3 kb 16S rRNA gene fragment of Burkholderia species prior to microarray analysis. The diagnostic performance of the microarray for identification and differentiation of Burkholderia species was tested with 44 reference strains of the genera Burkholderia, Pandoraea, Ralstonia and Limnobacter. Hybridization patterns based on presence/absence of probe signals were interpreted semi-automatically using the novel likelihood-based strategy of the web-tool Phylo- Detect. Eighty-eight per cent of the reference strains were correctly identified at the species level. The evaluated microarray was applied to investigate shifts in the Burkholderia community structure in acidic forest soil upon addition of cadmium, a condition that selected for Burkholderia species. The microarray results were in agreement with those obtained from phylogenetic analysis of Burkholderia 16S rRNA gene sequences recovered from the same cadmiumcontaminated soil, demonstrating the value of the Burkholderia phylochip for determinative and environmental studies.

  16. Comparative genome analysis of rice-pathogenic Burkholderia provides insight into capacity to adapt to different environments and hosts.

    Science.gov (United States)

    Seo, Young-Su; Lim, Jae Yun; Park, Jungwook; Kim, Sunyoung; Lee, Hyun-Hee; Cheong, Hoon; Kim, Sang-Mok; Moon, Jae Sun; Hwang, Ingyu

    2015-05-06

    In addition to human and animal diseases, bacteria of the genus Burkholderia can cause plant diseases. The representative species of rice-pathogenic Burkholderia are Burkholderia glumae, B. gladioli, and B. plantarii, which primarily cause grain rot, sheath rot, and seedling blight, respectively, resulting in severe reductions in rice production. Though Burkholderia rice pathogens cause problems in rice-growing countries, comprehensive studies of these rice-pathogenic species aiming to control Burkholderia-mediated diseases are only in the early stages. We first sequenced the complete genome of B. plantarii ATCC 43733T. Second, we conducted comparative analysis of the newly sequenced B. plantarii ATCC 43733T genome with eleven complete or draft genomes of B. glumae and B. gladioli strains. Furthermore, we compared the genome of three rice Burkholderia pathogens with those of other Burkholderia species such as those found in environmental habitats and those known as animal/human pathogens. These B. glumae, B. gladioli, and B. plantarii strains have unique genes involved in toxoflavin or tropolone toxin production and the clustered regularly interspaced short palindromic repeats (CRISPR)-mediated bacterial immune system. Although the genome of B. plantarii ATCC 43733T has many common features with those of B. glumae and B. gladioli, this B. plantarii strain has several unique features, including quorum sensing and CRISPR/CRISPR-associated protein (Cas) systems. The complete genome sequence of B. plantarii ATCC 43733T and publicly available genomes of B. glumae BGR1 and B. gladioli BSR3 enabled comprehensive comparative genome analyses among three rice-pathogenic Burkholderia species responsible for tissue rotting and seedling blight. Our results suggest that B. glumae has evolved rapidly, or has undergone rapid genome rearrangements or deletions, in response to the hosts. It also, clarifies the unique features of rice pathogenic Burkholderia species relative to other

  17. Plant-Associated Symbiotic Burkholderia Species Lack Hallmark Strategies Required in Mammalian Pathogenesis

    Science.gov (United States)

    Fong, Stephanie; Yerrapragada, Shailaja; Estrada-de los Santos, Paulina; Yang, Paul; Song, Nannie; Kano, Stephanie; de Faria, Sergio M.; Dakora, Felix D.; Weinstock, George; Hirsch, Ann M.

    2014-01-01

    Burkholderia is a diverse and dynamic genus, containing pathogenic species as well as species that form complex interactions with plants. Pathogenic strains, such as B. pseudomallei and B. mallei, can cause serious disease in mammals, while other Burkholderia strains are opportunistic pathogens, infecting humans or animals with a compromised immune system. Although some of the opportunistic Burkholderia pathogens are known to promote plant growth and even fix nitrogen, the risk of infection to infants, the elderly, and people who are immunocompromised has not only resulted in a restriction on their use, but has also limited the application of non-pathogenic, symbiotic species, several of which nodulate legume roots or have positive effects on plant growth. However, recent phylogenetic analyses have demonstrated that Burkholderia species separate into distinct lineages, suggesting the possibility for safe use of certain symbiotic species in agricultural contexts. A number of environmental strains that promote plant growth or degrade xenobiotics are also included in the symbiotic lineage. Many of these species have the potential to enhance agriculture in areas where fertilizers are not readily available and may serve in the future as inocula for crops growing in soils impacted by climate change. Here we address the pathogenic potential of several of the symbiotic Burkholderia strains using bioinformatics and functional tests. A series of infection experiments using Caenorhabditis elegans and HeLa cells, as well as genomic characterization of pathogenic loci, show that the risk of opportunistic infection by symbiotic strains such as B. tuberum is extremely low. PMID:24416172

  18. Competition between Burkholderia pseudomallei and B. thailandensis.

    Science.gov (United States)

    Ngamdee, Wikanda; Tandhavanant, Sarunporn; Wikraiphat, Chanthiwa; Reamtong, Onrapak; Wuthiekanun, Vanaporn; Salje, Jeanne; Low, David A; Peacock, Sharon J; Chantratita, Narisara

    2015-03-03

    Burkholderia pseudomallei is a Gram-negative bacterium that causes melioidosis, an often fatal disease in tropical countries. Burkholderia thailandensis is a non-virulent but closely related species. Both species are soil saprophytes but are almost never isolated together. We identified two mechanisms by which B. pseudomallei affects the growth of B. thailandensis. First, we found that six different isolates of B. pseudomallei inhibited the growth of B. thailandensis on LB agar plates. Second, our results indicated that 55% of isolated strains of B. pseudomallei produced a secreted compound that inhibited the motility but not the viability of B. thailandensis. Analysis showed that the active compound was a pH-sensitive and heat-labile compound, likely a protein, which may affect flagella processing or facilitate their degradation. Analysis of bacterial sequence types (STs) demonstrated an association between this and motility inhibition. The active compound was produced from B. pseudomallei during the stationary growth phase. Taken together, our results indicate that B. pseudomallei inhibits both the growth and motility of its close relative B. thailandensis. The latter phenomenon appears to occur via a previously unreported mechanism involving flagellar processing or degradation.

  19. Non-obligate predatory bacterium burkholderia casidaeand uses thereof

    OpenAIRE

    1998-01-01

    A novel predator bacterium Burkholderia casidae is disclosed. The invention is directed to the isolation and use of Burkholderia casidae to control microbial diseases of plants. The genetic, biochemical and physiological characteristics of Burkholderia casidae are described. Biocontrol compositions comprising Burkholderia casidae, and antimicrobial compounds and antimicrobial preparations prepared from Burkholderia casidae are also disclosed, as are methods for accomplishing all of the forego...

  20. Non-obligate predatory bacterium Burkholderia casidae and uses thereof

    OpenAIRE

    2001-01-01

    A novel predator bacterium Burkholderia casidae is disclosed. The invention is directed to the isolation and use of Burkholderia casidae to control microbial diseases of plants. The genetic, biochemical and physiological characteristics of Burkholderia casidae are described. Biocontrol compositions comprising Burkholderia casidae, and antimicrobial compounds and antimicrobial preparations prepared from Burkholderia casidae are also disclosed, as are methods for accomplishing all of the forego...

  1. The phylogenetic distribution and ecological role of carbon monoxide oxidation in the genus Burkholderia.

    Science.gov (United States)

    Weber, Carolyn F; King, Gary M

    2012-01-01

    Burkholderia is a physiologically and ecologically diverse genus that occurs commonly in assemblages of soil and rhizosphere bacteria. Although Burkholderia is known for its heterotrophic versatility, we demonstrate that 14 distinct environmental isolates oxidized carbon monoxide (CO) and possessed the gene encoding the catalytic subunit of form I CO dehydrogenase (coxL). DNA from a Burkholderia isolate obtained from a passalid beetle also contained coxL as do the genomic sequences of species H160 and Ch1-1. Isolates were able to consume CO at concentrations ranging from 100 ppm (vol/vol) to sub-ambient ( 2.5 mM), but mixotrophic consumption of CO and pyruvate occurred when initial pyruvate concentrations were lower (c. 400 lM). With the exception of an isolate most closely related to Burkholderia cepacia, all CO-oxidizing isolates examined were members of a nonpathogenic clade and were most closely related to Burkholderia species, B. caledonica, B. fungorum, B. oxiphila, B. mimosarum, B. nodosa, B. sacchari, B. bryophila, B. ferrariae, B. ginsengesoli, and B. unamae. However, none of these type strains oxidized CO or contained coxL based on results from PCR analyses. Collectively, these results demonstrate that the presence of CO oxidation within members of the Burkholderia genus is variable but it is most commonly found among rhizosphere inhabitants that are not closely related to B. cepacia.

  2. Crosstalk between sugarcane and a plant-growth promoting Burkholderia species

    Science.gov (United States)

    Paungfoo-Lonhienne, Chanyarat; Lonhienne, Thierry G. A.; Yeoh, Yun Kit; Donose, Bogdan C.; Webb, Richard I.; Parsons, Jeremy; Liao, Webber; Sagulenko, Evgeny; Lakshmanan, Prakash; Hugenholtz, Philip; Schmidt, Susanne; Ragan, Mark A.

    2016-01-01

    Bacterial species in the plant-beneficial-environmental clade of Burkholderia represent a substantial component of rhizosphere microbes in many plant species. To better understand the molecular mechanisms of the interaction, we combined functional studies with high-resolution dual transcriptome analysis of sugarcane and root-associated diazotrophic Burkholderia strain Q208. We show that Burkholderia Q208 forms a biofilm at the root surface and suppresses the virulence factors that typically trigger immune response in plants. Up-regulation of bd-type cytochromes in Burkholderia Q208 suggests an increased energy production and creates the microaerobic conditions suitable for BNF. In this environment, a series of metabolic pathways are activated in Burkholderia Q208 implicated in oxalotrophy, microaerobic respiration, and formation of PHB granules, enabling energy production under microaerobic conditions. In the plant, genes involved in hypoxia survival are up-regulated and through increased ethylene production, larger aerenchyma is produced in roots which in turn facilitates diffusion of oxygen within the cortex. The detected changes in gene expression, physiology and morphology in the partnership are evidence of a sophisticated interplay between sugarcane and a plant-growth promoting Burkholderia species that advance our understanding of the mutually beneficial processes occurring in the rhizosphere. PMID:27869215

  3. Burkholderia insulsa sp. nov., a facultatively chemolithotrophic bacterium isolated from an arsenic-rich shallow marine hydrothermal system.

    Science.gov (United States)

    Rusch, Antje; Islam, Shaer; Savalia, Pratixa; Amend, Jan P

    2015-01-01

    Enrichment cultures inoculated with hydrothermally influenced nearshore sediment from Papua New Guinea led to the isolation of an arsenic-tolerant, acidophilic, facultatively aerobic bacterial strain designated PNG-April(T). Cells of this strain were Gram-stain-negative, rod-shaped, motile and did not form spores. Strain PNG-April(T) grew at temperatures between 4 °C and 40 °C (optimum 30-37 °C), at pH 3.5 to 8.3 (optimum pH 5-6) and in the presence of up to 2.7% NaCl (optimum 0-1.0%). Both arsenate and arsenite were tolerated up to concentrations of at least 0.5 mM. Metabolism in strain PNG-April(T) was strictly respiratory. Heterotrophic growth occurred with O2 or nitrate as electron acceptors, and aerobic lithoautotrophic growth was observed with thiosulfate or nitrite as electron donors. The novel isolate was capable of N2-fixation. The respiratory quinones were Q-8 and Q-7. Phylogenetically, strain PNG-April(T) belongs to the genus Burkholderia and shares the highest 16S rRNA gene sequence similarity with the type strains of Burkholderia fungorum (99.8%), Burkholderia phytofirmans (98.8%), Burkholderia caledonica (98.4%) and Burkholderia sediminicola (98.4%). Differences from these related species in several physiological characteristics (lipid composition, carbohydrate utilization, enzyme profiles) and DNA-DNA hybridization suggested the isolate represents a novel species of the genus Burkholderia, for which we propose the name Burkholderia insulsa sp. nov. The type strain is PNG-April(T) ( = DSM 28142(T) = LMG 28183(T)). © 2015 IUMS.

  4. Biochemical and Functional Studies on the Burkholderia cepacia Complex bceN Gene, Encoding a GDP-D-Mannose 4,6-Dehydratase

    Science.gov (United States)

    Pinheiro, Pedro F.; Leitão, Jorge H.

    2013-01-01

    This work reports the biochemical and functional analysis of the Burkholderia cenocepacia J2315 bceN gene, encoding a protein with GDP-D-mannose 4,6-dehydratase enzyme activity (E.C.4.2.1.47). Data presented indicate that the protein is active when in the tetrameric form, catalyzing the conversion of GDP-D-mannose into GDP-4-keto-6-deoxy-D-mannose. This sugar nucleotide is the intermediary necessary for the biosynthesis of GDP-D-rhamnose, one of the sugar residues of cepacian, the major exopolysaccharide produced by environmental and human, animal and plant pathogenic isolates of the Burkholderia cepacia complex species. Vmax and Km values of 1.5±0.2 µmol.min−1.mg−1 and 1024±123 µM, respectively, were obtained from the kinetic characterization of the B. cenocepacia J2315 BceN protein by NMR spectroscopy, at 25°C and in the presence of 1 mol MgCl2 per mol of protein. The enzyme activity was strongly inhibited by the substrate, with an estimated Ki of 2913±350 µM. The lack of a functional bceN gene in a mutant derived from B. cepacia IST408 slightly reduced cepacian production. However, in the B. multivorans ATCC17616 with bceN as the single gene in its genome with predicted GMD activity, a bceN mutant did not produce cepacian, indicating that this gene product is required for cepacian biosynthesis. PMID:23460819

  5. Biochemical and functional studies on the Burkholderia cepacia complex bceN gene, encoding a GDP-D-mannose 4,6-dehydratase.

    Directory of Open Access Journals (Sweden)

    Sílvia A Sousa

    Full Text Available This work reports the biochemical and functional analysis of the Burkholderia cenocepacia J2315 bceN gene, encoding a protein with GDP-D-mannose 4,6-dehydratase enzyme activity (E.C.4.2.1.47. Data presented indicate that the protein is active when in the tetrameric form, catalyzing the conversion of GDP-D-mannose into GDP-4-keto-6-deoxy-D-mannose. This sugar nucleotide is the intermediary necessary for the biosynthesis of GDP-D-rhamnose, one of the sugar residues of cepacian, the major exopolysaccharide produced by environmental and human, animal and plant pathogenic isolates of the Burkholderia cepacia complex species. Vmax and Km values of 1.5±0.2 µmol.min(-1.mg(-1 and 1024±123 µM, respectively, were obtained from the kinetic characterization of the B. cenocepacia J2315 BceN protein by NMR spectroscopy, at 25°C and in the presence of 1 mol MgCl2 per mol of protein. The enzyme activity was strongly inhibited by the substrate, with an estimated Ki of 2913±350 µM. The lack of a functional bceN gene in a mutant derived from B. cepacia IST408 slightly reduced cepacian production. However, in the B. multivorans ATCC17616 with bceN as the single gene in its genome with predicted GMD activity, a bceN mutant did not produce cepacian, indicating that this gene product is required for cepacian biosynthesis.

  6. Burkholderia novacaledonica sp. nov. and B. ultramafica sp. nov. isolated from roots of Costularia spp. pioneer plants of ultramafic soils in New Caledonia.

    Science.gov (United States)

    Guentas, Linda; Gensous, Simon; Cavaloc, Yvon; Ducousso, Marc; Amir, Hamid; De Georges de Ledenon, Benjamin; Moulin, Lionel; Jourand, Philippe

    2016-05-01

    The taxonomic status of eleven rhizospheric bacterial strains belonging to the genus Burkholderia and isolated from roots of Costularia (Cyperaceae), tropical herbaceous pioneer plants growing on ultramafic soils in New Caledonia, was investigated using a polyphasic taxonomic approach. The genetic analyses (16S rRNA genes, gyrB, recA, nreB and cnr) confirmed that all strains are Burkholderia and cluster into two separated groups. The DNA hybridization results showed low relatedness values to the closest relatives Burkholderia species. The phenotypic analyses confirmed that the two groups of strains could be differentiated from each other and from other known Burkholderia species. This polyphasic study revealed that these two groups of strains represent each a novel species of Burkholderia, for which the names Burkholderia novacaledonica sp. nov. (type strain STM10272(T)=LMG28615(T)=CIP110887(T)) and B. ultramafica sp. nov. (type strain STM10279(T)=LMG28614(T)=CIP110886(T)) are proposed, respectively. These strains of Burkholderia presented specific ecological traits such as the tolerance to the extreme edaphic constraints of ultramafic soils: they grew at pH between 4 and 8 and tolerate the strong unbalanced Ca/Mg ratio (1/19) and the high concentrations of heavy metals i.e. Co, Cr, Mn and Ni. Noteworthy B. ultramafica tolerated nickel until 10mM and B. novacaledonica up to 5mM. The presence of the nickel (nreB) and cobalt/nickel (cnr) resistance determinants encoding for protein involved in metal tolerance was found in all strains of both groups. Moreover, most of the strains were able to produce plant growth promoting molecules (ACC, IAA, NH3 and siderophores). Such ecological traits suggest that these new species of Burkholderia might be environmentally adaptable plant-associated bacteria and beneficial to plants. Copyright © 2016 Elsevier GmbH. All rights reserved.

  7. Phylogeographic, genomic, and meropenem susceptibility analysis of Burkholderia ubonensis.

    Science.gov (United States)

    Price, Erin P; Sarovich, Derek S; Webb, Jessica R; Hall, Carina M; Jaramillo, Sierra A; Sahl, Jason W; Kaestli, Mirjam; Mayo, Mark; Harrington, Glenda; Baker, Anthony L; Sidak-Loftis, Lindsay C; Settles, Erik W; Lummis, Madeline; Schupp, James M; Gillece, John D; Tuanyok, Apichai; Warner, Jeffrey; Busch, Joseph D; Keim, Paul; Currie, Bart J; Wagner, David M

    2017-09-01

    The bacterium Burkholderia ubonensis is commonly co-isolated from environmental specimens harbouring the melioidosis pathogen, Burkholderia pseudomallei. B. ubonensis has been reported in northern Australia and Thailand but not North America, suggesting similar geographic distribution to B. pseudomallei. Unlike most other Burkholderia cepacia complex (Bcc) species, B. ubonensis is considered non-pathogenic, although its virulence potential has not been tested. Antibiotic resistance in B. ubonensis, particularly towards drugs used to treat the most severe B. pseudomallei infections, has also been poorly characterised. This study examined the population biology of B. ubonensis, and includes the first reported isolates from the Caribbean. Phylogenomic analysis of 264 B. ubonensis genomes identified distinct clades that corresponded with geographic origin, similar to B. pseudomallei. A small proportion (4%) of strains lacked the 920kb chromosome III replicon, with discordance of presence/absence amongst genetically highly related strains, demonstrating that the third chromosome of B. ubonensis, like other Bcc species, probably encodes for a nonessential pC3 megaplasmid. Multilocus sequence typing using the B. pseudomallei scheme revealed that one-third of strains lack the "housekeeping" narK locus. In comparison, all strains could be genotyped using the Bcc scheme. Several strains possessed high-level meropenem resistance (≥32 μg/mL), a concern due to potential transmission of this phenotype to B. pseudomallei. In silico analysis uncovered a high degree of heterogeneity among the lipopolysaccharide O-antigen cluster loci, with at least 35 different variants identified. Finally, we show that Asian B. ubonensis isolate RF23-BP41 is avirulent in the BALB/c mouse model via a subcutaneous route of infection. Our results provide several new insights into the biology of this understudied species.

  8. Burkholderia sp. induces functional nodules on the South African invasive legume Dipogon lignosus (Phaseoleae) in New Zealand soils.

    Science.gov (United States)

    Liu, Wendy Y Y; Ridgway, Hayley J; James, Trevor K; James, Euan K; Chen, Wen-Ming; Sprent, Janet I; Young, J Peter W; Andrews, Mitchell

    2014-10-01

    The South African invasive legume Dipogon lignosus (Phaseoleae) produces nodules with both determinate and indeterminate characteristics in New Zealand (NZ) soils. Ten bacterial isolates produced functional nodules on D. lignosus. The 16S ribosomal RNA (rRNA) gene sequences identified one isolate as Bradyrhizobium sp., one isolate as Rhizobium sp. and eight isolates as Burkholderia sp. The Bradyrhizobium sp. and Rhizobium sp. 16S rRNA sequences were identical to those of strains previously isolated from crop plants and may have originated from inocula used on crops. Both 16S rRNA and DNA recombinase A (recA) gene sequences placed the eight Burkholderia isolates separate from previously described Burkholderia rhizobial species. However, the isolates showed a very close relationship to Burkholderia rhizobial strains isolated from South African plants with respect to their nitrogenase iron protein (nifH), N-acyltransferase nodulation protein A (nodA) and N-acetylglucosaminyl transferase nodulation protein C (nodC) gene sequences. Gene sequences and enterobacterial repetitive intergenic consensus (ERIC) PCR and repetitive element palindromic PCR (rep-PCR) banding patterns indicated that the eight Burkholderia isolates separated into five clones of one strain and three of another. One strain was tested and shown to produce functional nodules on a range of South African plants previously reported to be nodulated by Burkholderia tuberum STM678(T) which was isolated from the Cape Region. Thus, evidence is strong that the Burkholderia strains isolated here originated in South Africa and were somehow transported with the plants from their native habitat to NZ. It is possible that the strains are of a new species capable of nodulating legumes.

  9. Comparative Genomics of Burkholderia singularis sp. nov., a Low G+C Content, Free-Living Bacterium That Defies Taxonomic Dissection of the Genus Burkholderia

    Directory of Open Access Journals (Sweden)

    Peter Vandamme

    2017-09-01

    Full Text Available Four Burkholderia pseudomallei-like isolates of human clinical origin were examined by a polyphasic taxonomic approach that included comparative whole genome analyses. The results demonstrated that these isolates represent a rare and unusual, novel Burkholderia species for which we propose the name B. singularis. The type strain is LMG 28154T (=CCUG 65685T. Its genome sequence has an average mol% G+C content of 64.34%, which is considerably lower than that of other Burkholderia species. The reduced G+C content of strain LMG 28154T was characterized by a genome wide AT bias that was not due to reduced GC-biased gene conversion or reductive genome evolution, but might have been caused by an altered DNA base excision repair pathway. B. singularis can be differentiated from other Burkholderia species by multilocus sequence analysis, MALDI-TOF mass spectrometry and a distinctive biochemical profile that includes the absence of nitrate reduction, a mucoid appearance on Columbia sheep blood agar, and a slowly positive oxidase reaction. Comparisons with publicly available whole genome sequences demonstrated that strain TSV85, an Australian water isolate, also represents the same species and therefore, to date, B. singularis has been recovered from human or environmental samples on three continents.

  10. Comparative Genomics of Burkholderia singularis sp. nov., a Low G+C Content, Free-Living Bacterium That Defies Taxonomic Dissection of the Genus Burkholderia

    Science.gov (United States)

    Vandamme, Peter; Peeters, Charlotte; De Smet, Birgit; Price, Erin P.; Sarovich, Derek S.; Henry, Deborah A.; Hird, Trevor J.; Zlosnik, James E. A.; Mayo, Mark; Warner, Jeffrey; Baker, Anthony; Currie, Bart J.; Carlier, Aurélien

    2017-01-01

    Four Burkholderia pseudomallei-like isolates of human clinical origin were examined by a polyphasic taxonomic approach that included comparative whole genome analyses. The results demonstrated that these isolates represent a rare and unusual, novel Burkholderia species for which we propose the name B. singularis. The type strain is LMG 28154T (=CCUG 65685T). Its genome sequence has an average mol% G+C content of 64.34%, which is considerably lower than that of other Burkholderia species. The reduced G+C content of strain LMG 28154T was characterized by a genome wide AT bias that was not due to reduced GC-biased gene conversion or reductive genome evolution, but might have been caused by an altered DNA base excision repair pathway. B. singularis can be differentiated from other Burkholderia species by multilocus sequence analysis, MALDI-TOF mass spectrometry and a distinctive biochemical profile that includes the absence of nitrate reduction, a mucoid appearance on Columbia sheep blood agar, and a slowly positive oxidase reaction. Comparisons with publicly available whole genome sequences demonstrated that strain TSV85, an Australian water isolate, also represents the same species and therefore, to date, B. singularis has been recovered from human or environmental samples on three continents. PMID:28932212

  11. Burkholderia ginsengiterrae sp. nov. and Burkholderia panaciterrae sp. nov., antagonistic bacteria against root rot pathogen Cylindrocarpon destructans, isolated from ginseng soil.

    Science.gov (United States)

    Farh, Mohamed El-Agamy; Kim, Yeon-Ju; Van An, Hoang; Sukweenadhi, Johan; Singh, Priyanka; Huq, Md Amdadul; Yang, Deok-Chun

    2015-04-01

    Strain DCY85(T) and DCY85-1(T), isolated from rhizosphere of ginseng, were rod-shaped, Gram-reaction-negative, strictly aerobic, catalase positive and oxidase negative. 16S rRNA gene sequence analysis revealed that strain DCY85(T) as well as DCY85-1(T) belonged to the genus Burkholderia and were closely related to Burkholderia fungorum KACC 12023(T) (98.1 and 98.0 % similarity, respectively). The major polar lipids of strain DCY85(T) and DCY85-1(T) were phosphatidylethanolamine, one unidentified aminolipid and two unidentified phospholipids. The major fatty acids of both strains are C16:0, C18:1 ω7c and summed feature 3 (C16:1 ω6c and/or C16:1 ω7c). The predominant isoprenoid quinone of each strain DCY85(T) and DCY85-1(T) was ubiquinone (Q-8) and the G+C content of their genomic DNA was 66.0 and 59.4 mol%, respectively, which fulfill the characteristic range of the genus Burkholderia. The polyamine content of both DCY85(T) and DCY85-1(T) was putrescine. Although both DCY85(T) and DCY85-1(T) have highly similar 16S rRNA and identical RecA and gyrB sequences, they show differences in phenotypic and chemotaxonomic characteristics. DNA-DNA hybridization results proved the consideration of both strains as two different species. Based on the results from our polyphasic characterization, strain DCY85(T) and DCY85-1(T) are considered novel Burkholderia species for which the name Burkholderia ginsengiterrae sp. nov and Burkholderia panaciterrae sp. nov are, respectively, proposed. An emended description of those strains is also proposed. DCY85(T) and DCY85-1(T) showed antagonistic activity against the common root rot pathogen of ginseng, Cylindrocarpon destructans. The proposed type strains are DCY85(T) (KCTC 42054(T) = JCM 19888(T)) and DCY85-1(T) (KCTC 42055(T) = JCM 19889(T)).

  12. NOVEL ORGANIZATION OF THE GENES FOR PHTHALATE DEGRADATION FROM BURKHOLDERIA CEPACIA DBO1

    Science.gov (United States)

    Burkholderia cepacia DBO1 is able to utilize phthalate as the sole source of carbon and energy for growth. Two overlapping cosmid clones containing the genes for phthalate degradation were isolated from this strain. Subcloning and activity analysis localized the genes for phthala...

  13. The role of siderophores in metal homeostasis of members of the genus Burkholderia.

    Science.gov (United States)

    Mathew, Anugraha; Jenul, Christian; Carlier, Aurelien L; Eberl, Leo

    2016-02-01

    Although members of the genus Burkholderia can utilize a high-affinity iron uptake system to sustain growth under iron-limiting conditions, many strains also produce siderophores, suggesting that they may serve alternative functions. Here we demonstrate that the two Burkholderia siderophores pyochelin and ornibactin can protect the cells from metal toxicity and thus play an alternative role in metal homeostasis. We also demonstrate that metals such as copper and zinc induce the production of ornibactin. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  14. Phylogenetically Diverse Burkholderia Associated with Midgut Crypts of Spurge Bugs, Dicranocephalus spp. (Heteroptera: Stenocephalidae).

    Science.gov (United States)

    Kuechler, Stefan Martin; Matsuura, Yu; Dettner, Konrad; Kikuchi, Yoshitomo

    2016-06-25

    Diverse phytophagous heteropteran insects, commonly known as stinkbugs, are associated with specific gut symbiotic bacteria, which have been found in midgut cryptic spaces. Recent studies have revealed that members of the stinkbug families Coreidae and Alydidae of the superfamily Coreoidea are consistently associated with a specific group of the betaproteobacterial genus Burkholderia, called the "stinkbug-associated beneficial and environmental (SBE)" group, and horizontally acquire specific symbionts from the environment every generation. However, the symbiotic system of another coreoid family, Stenocephalidae remains undetermined. We herein investigated four species of the stenocephalid genus Dicranocephalus. Examinations via fluorescence in situ hybridization (FISH) and transmission electron microscopy (TEM) revealed the typical arrangement and ultrastructures of midgut crypts and gut symbionts. Cloning and molecular phylogenetic analyses of bacterial genes showed that the midgut crypts of all species are colonized by Burkholderia strains, which were further assigned to different subgroups of the genus Burkholderia. In addition to the SBE-group Burkholderia, a number of stenocephalid symbionts belonged to a novel clade containing B. sordidicola and B. udeis, suggesting a specific symbiont clade for the Stenocephalidae. The symbiotic systems of stenocephalid bugs may provide a unique opportunity to study the ongoing evolution of symbiont associations in the stinkbug-Burkholderia interaction.

  15. In Vitro Activity of Ceftolozane-Tazobactam against Burkholderia pseudomallei.

    Science.gov (United States)

    Slack, Andrew; Parsonson, Fiona; Cronin, Katie; Engler, Kathy; Norton, Robert

    2018-06-25

    We investigated the in vitro activity of a novel fifth-generation cephalosporin-tazobactam combination, ceftolozane-tazobactam against Burkholderia pseudomallei , the etiological agent of melioidosis. Using both disc diffusion and minimum inhibitory concentration (MIC) strip techniques against 56 clinical isolates and an NCTC strain, the MIC to ceftolozane-tazobactam was found to be between 0.75 and 4 mcg/mL. The MIC50 was found to be 1.5 mcg/mL and MIC90 was 2.0 mcg/mL. This study provides initial evidence of ceftolozane-tazobactam as a novel agent in the management of melioidosis.

  16. Distinct colicin M-like bacteriocin-immunity pairs in Burkholderia.

    Science.gov (United States)

    Ghequire, Maarten G K; De Mot, René

    2015-11-27

    The Escherichia coli bacteriocin colicin M (ColM) acts via degradation of the cell wall precursor lipid II in target cells. ColM producers avoid self-inhibition by a periplasmic immunity protein anchored in the inner membrane. In this study, we identified colM-like bacteriocin genes in genomes of several β-proteobacterial strains belonging to the Burkholderia cepacia complex (Bcc) and the Burkholderia pseudomallei group. Two selected Burkholderia ambifaria proteins, designated burkhocins M1 and M2, were produced recombinantly and showed antagonistic activity against Bcc strains. In their considerably sequence-diverged catalytic domain, a conserved aspartate residue equally proved pivotal for cytotoxicity. Immunity to M-type burkhocins is conferred upon susceptible strains by heterologous expression of a cognate gene located either upstream or downstream of the toxin gene. These genes lack homology with currently known ColM immunity genes and encode inner membrane-associated proteins of two distinct types, differing in predicted transmembrane topology and moiety exposed to the periplasm. The addition of burkhocins to the bacteriocin complement of Burkholderia reveals a wider phylogenetic distribution of ColM-like bacteriotoxins, beyond the γ-proteobacterial genera Escherichia, Pectobacterium and Pseudomonas, and illuminates the diversified nature of immunity-providing proteins.

  17. Burkholderia jiangsuensis sp. nov., a methyl parathion degrading bacterium, isolated from methyl parathion contaminated soil.

    Science.gov (United States)

    Liu, Xu-Yun; Li, Chun-Xiu; Luo, Xiao-Jing; Lai, Qi-Liang; Xu, Jian-He

    2014-09-01

    A methyl parathion (MP) degrading bacterial strain, designated MP-1(T), was isolated from a waste land where pesticides were formerly manufactured in Jiangsu province, China. Polyphasic taxonomic studies showed that MP-1(T) is a Gram-stain-negative, non-spore-forming, rod-shaped and motile bacterium. The bacterium could grow at salinities of 0-1 % (w/v) and temperatures of 15-40 °C. Strain MP-1(T) could reduce nitrate to nitrite, utilize d-glucose and l-arabinose, but not produce indole, or hydrolyse gelatin. Phylogenetic analysis based on 16S rRNA gene sequences demonstrated that MP-1(T) belongs to the genus Burkholderia, showing highest sequence similarity to Burkholderia grimmiae DSM 25160(T) (98.5 %), and similar strains including Burkholderia zhejiangensis OP-1(T) (98.2 %), Burkholderia choica LMG 22940(T) (97.5 %), Burkholderia glathei DSM 50014(T) (97.4 %), Burkholderia terrestris LMG 22937(T) (97.2 %) and Burkholderia telluris LMG 22936(T) (97.0 %). In addition, the gyrB and recA gene segments of strain MP-1(T) exhibited less than 89.0 % and 95.1 % similarities with the most highly-related type strains indicated above. The G+C content of strain MP-1(T) was 62.6 mol%. The major isoprenoid quinone was ubiquinone Q-8. The predominant polar lipids comprised phosphatidyl ethanolamine, phosphatidyl glycerol, aminolipid and phospholipid. The principal fatty acids in strain MP-1(T) were C18 : 1ω7c/C18 : 1ω6c (23.3 %), C16 : 0 (16.8 %), cyclo-C17 : 0 (15.0 %), C16 : 1ω7c/C16 : 1ω6 (8.5 %), cyclo-C19 : 0ω8c (8.1 %), C16 : 1 iso I/C14 : 0 3-OH (5.7 %), C16 : 0 3-OH (5.6 %) and C16 : 02-OH (5.1 %). The DNA-DNA relatedness values between strain MP-1(T) and the three type strains (B. grimmiae DSM 25160(T), B. zhejiangensis OP-1(T) and B. glathei DSM 50014(T)) ranged from 24.6 % to 37.4 %. In accordance with phenotypic and genotypic characteristics, strain MP-1(T) represents a novel

  18. Insecticide applications to soil contribute to the development of Burkholderia mediating insecticide resistance in stinkbugs.

    Science.gov (United States)

    Tago, Kanako; Kikuchi, Yoshitomo; Nakaoka, Sinji; Katsuyama, Chie; Hayatsu, Masahito

    2015-07-01

    Some soil Burkholderia strains are capable of degrading the organophosphorus insecticide, fenitrothion, and establish symbiosis with stinkbugs, making the host insects fenitrothion-resistant. However, the ecology of the symbiotic degrading Burkholderia adapting to fenitrothion in the free-living environment is unknown. We hypothesized that fenitrothion applications affect the dynamics of fenitrothion-degrading Burkholderia, thereby controlling the transmission of symbiotic degrading Burkholderia from the soil to stinkbugs. We investigated changes in the density and diversity of culturable Burkholderia (i.e. symbiotic and nonsymbiotic fenitrothion degraders and nondegraders) in fenitrothion-treated soil using microcosms. During the incubation with five applications of pesticide, the density of the degraders increased from less than the detection limit to around 10(6)/g of soil. The number of dominant species among the degraders declined with the increasing density of degraders; eventually, one species predominated. This process can be explained according to the competitive exclusion principle using V(max) and K(m) values for fenitrothion metabolism by the degraders. We performed a phylogenetic analysis of representative strains isolated from the microcosms and evaluated their ability to establish symbiosis with the stinkbug Riptortus pedestris. The strains that established symbiosis with R. pedestris were assigned to a cluster including symbionts commonly isolated from stinkbugs. The strains outside the cluster could not necessarily associate with the host. The degraders in the cluster predominated during the initial phase of degrader dynamics in the soil. Therefore, only a few applications of fenitrothion could allow symbiotic degraders to associate with their hosts and may cause the emergence of symbiont-mediated insecticide resistance. © 2015 John Wiley & Sons Ltd.

  19. Detection of misidentifications of species from the Burkholderia cepacia complex and description of a new member, the soil bacterium Burkholderia catarinensis sp. nov.

    Science.gov (United States)

    Bach, Evelise; Sant'Anna, Fernando Hayashi; Magrich Dos Passos, João Frederico; Balsanelli, Eduardo; de Baura, Valter Antonio; Pedrosa, Fábio de Oliveira; de Souza, Emanuel Maltempi; Passaglia, Luciane Maria Pereira

    2017-08-31

    The correct identification of bacteria from the Burkholderia cepacia complex (Bcc) is crucial for epidemiological studies and treatment of cystic fibrosis infections. However, genome-based identification tools are revealing many controversial Bcc species assignments. The aim of this work is to re-examine the taxonomic position of the soil bacterium B. cepacia 89 through polyphasic and genomic approaches. recA and 16S rRNA gene sequence analysis positioned strain 89 inside the Bcc group. However, based on the divergence score of seven concatenated allele sequences, and values of average nucleotide identity, and digital DNA:DNA hybridization, our results suggest that strain 89 is different from other Bcc species formerly described. Thus, we propose to classify Burkholderia sp. 89 as the novel species Burkholderia catarinensis sp. nov. with strain 89T (=DSM 103188T = BR 10601T) as the type strain. Moreover, our results call the attention to some probable misidentifications of Bcc genomes at the National Center for Biotechnology Information database. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. 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

  1. Co-existence of Rhizobia and Diverse Non-rhizobial Bacteria in the Rhizosphere and Nodules of Dalbergia odorifera Seedlings Inoculated with Bradyrhizobium elkanii, Rhizobium multihospitium–Like and Burkholderia pyrrocinia–Like Strains

    Directory of Open Access Journals (Sweden)

    Junkun Lu

    2017-11-01

    Full Text Available Rhizobia induce root nodules and fix atmospheric N2 for most legume species in exchange for carbon. However, the diverse endophytic non-rhizobial bacteria in legume nodules that co-exist with rhizobia are often ignored because they are difficult to cultivate using routine cultivation approaches. To enhance our understanding of the incidence and diversity of legume–bacteria associations, a high-throughput sequencing analysis of bacterial 16S rRNA genes was used to examine the bacterial community in the rhizospheres and root nodules of Dalbergia odorifera seedlings that were uninoculated or inoculated with Bradyrhizobium elkanii H255, Rhizobium multihospitium–like HT221, or Burkholderia pyrrocinia–like H022238, in two growth media (nitrogen [N]-supplied soil or N-omitted potting mix. Seedlings inoculated with Bradyrhizobium had significantly more nodules than seedlings in the other inoculation conditions, regardless of growth media. Using the 15N natural abundance method, it was shown that the inoculated plants had significantly higher N2 fixation efficiency (48–57% and specific nodule activity [269–313 μg N mg−1 of dry weight (dwt nodule] compared to the uninoculated plants (203 μg N mg−1 dwt nodule. The 16S rRNA gene analysis showed that there was generally a higher bacterial diversity in the rhizosphere than in the nodules in the corresponding condition. Both rhizobial inoculation and media status significantly altered the bacterial communities in the rhizospheres and nodules (P < 0.05, with the exception of the inoculated soil rhizospheres. Regarding non-rhizobial bacteria, three genera, i.e., Lactococcus, Bacillus, and Pseudomonas, were consistently enriched in the rhizosphere and Bradyrhizobium, Chloroplast norank (which belongs to Cyanobacteria, and Lactococcus were commonly found in the nodules. In contrast, common rhizobial genera (including Rhizobium, Mesorhizobium, and Burkholderia were only present in the nodules at low

  2. Prevalence and Identification of Burkholderia pseudomallei and Near-Neighbor Species in the Malabar Coastal Region of India

    Science.gov (United States)

    Peddayelachagiri, Bhavani V.; Paul, Soumya; Nagaraj, Sowmya; Gogoi, Madhurjya; Sripathy, Murali H.; Batra, Harsh V.

    2016-01-01

    Accurate identification of pathogens with biowarfare importance requires detection tools that specifically differentiate them from near-neighbor species. Burkholderia pseudomallei, the causative agent of a fatal disease melioidosis, is one such biothreat agent whose differentiation from its near-neighbor species is always a challenge. This is because of its phenotypic similarity with other Burkholderia species which have a wide spread geographical distribution with shared environmental niches. Melioidosis is a major public health concern in endemic regions including Southeast Asia and northern Australia. In India, the disease is still considered to be emerging. Prevalence surveys of this saprophytic bacterium in environment are under-reported in the country. A major challenge in this case is the specific identification and differentiation of B. pseudomallei from the growing list of species of Burkholderia genus. The objectives of this study included examining the prevalence of B. pseudomallei and near-neighbor species in coastal region of South India and development of a novel detection tool for specific identification and differentiation of Burkholderia species. Briefly, we analyzed soil and water samples collected from Malabar coastal region of Kerala, South India for prevalence of B. pseudomallei. The presumptive Burkholderia isolates were identified using recA PCR assay. The recA PCR assay identified 22 of the total 40 presumptive isolates as Burkholderia strains (22.72% and 77.27% B. pseudomallei and non-pseudomallei Burkholderia respectively). In order to identify each isolate screened, we performed recA and 16S rDNA sequencing. This two genes sequencing revealed that the presumptive isolates included B. pseudomallei, non-pseudomallei Burkholderia as well as non-Burkholderia strains. Furthermore, a gene termed D-beta hydroxybutyrate dehydrogenase (bdha) was studied both in silico and in vitro for accurate detection of Burkholderia genus. The optimized bdha

  3. Burkholderia Species Are the Most Common and Preferred Nodulating Symbionts of the Piptadenia Group (Tribe Mimoseae)

    Science.gov (United States)

    Bournaud, Caroline; de Faria, Sergio Miana; dos Santos, José Miguel Ferreira; Tisseyre, Pierre; Silva, Michele; Chaintreuil, Clémence; Gross, Eduardo; James, Euan K.; Prin, Yves; Moulin, Lionel

    2013-01-01

    Burkholderia legume symbionts (also called α-rhizobia) are ancient in origin and are the main nitrogen-fixing symbionts of species belonging to the large genus Mimosa in Brazil. We investigated the extent of the affinity between Burkholderia and species in the tribe Mimoseae by studying symbionts of the genera Piptadenia (P.), Parapiptadenia (Pp.), Pseudopiptadenia (Ps.), Pityrocarpa (Py.), Anadenanthera (A.) and Microlobius (Mi.), all of which are native to Brazil and are phylogenetically close to Mimosa, and which together with Mimosa comprise the “Piptadenia group”. We characterized 196 strains sampled from 18 species from 17 locations in Brazil using two neutral markers and two symbiotic genes in order to assess their species affiliations and the evolution of their symbiosis genes. We found that Burkholderia are common and highly diversified symbionts of species in the Piptadenia group, comprising nine Burkholderia species, of which three are new ones and one was never reported as symbiotic (B. phenoliruptrix). However, α-rhizobia were also detected and were occasionally dominant on a few species. A strong sampling site effect on the rhizobial nature of symbionts was detected, with the symbiont pattern of the same legume species changing drastically from location to location, even switching from β to α-rhizobia. Coinoculation assays showed a strong affinity of all the Piptadenia group species towards Burkholderia genotypes, with the exception of Mi. foetidus. Phylogenetic analyses of neutral and symbiotic markers showed that symbiosis genes in Burkholderia from the Piptadenia group have evolved mainly through vertical transfer, but also by horizontal transfer in two species. PMID:23691052

  4. Polysaccharide microarray technology for the detection of Burkholderia pseudomallei and Burkholderia mallei antibodies.

    Science.gov (United States)

    Parthasarathy, Narayanan; DeShazer, David; England, Marilyn; Waag, David M

    2006-11-01

    A polysaccharide microarray platform was prepared by immobilizing Burkholderia pseudomallei and Burkholderia mallei polysaccharides. This polysaccharide array was tested with success for detecting B. pseudomallei and B. mallei serum (human and animal) antibodies. The advantages of this microarray technology over the current serodiagnosis of the above bacterial infections were discussed.

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

    Science.gov (United States)

    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

  6. Isolation and Identification of Burkholderia glumae from Symptomless Rice Seeds

    Directory of Open Access Journals (Sweden)

    Bo Zhu

    2008-06-01

    Full Text Available A survey on isolation and detection of the casual organism of bacterial grain rot of rice was conducted during 1997–2006. In 2006, six pathogenic bacterial strains were isolated from two symptomless seed samples of rice (Oryza sativa L. originally produced in Hainan Province and then planted in Zhejiang Province, China. They were identified as Burkholderia glumae which is the causal organism of bacterial grain rot of rice by physiological characteristics, colony morphology, pathogenicity test, Biolog, fatty acid methyl ester (FAME analysis and RAPD-PCR compared with the four standard reference strains. It is confirmed that there is the infection of B. glumae in so-called ‘health looking seeds’.

  7. Construction and characterization of stable, constitutively expressed, chromosomal green and red fluorescent transcriptional fusions in the select agents, Bacillus anthracis, Yersinia pestis, Burkholderia mallei, and Burkholderia pseudomallei.

    Science.gov (United States)

    Su, Shengchang; Bangar, Hansraj; Saldanha, Roland; Pemberton, Adin; Aronow, Bruce; Dean, Gary E; Lamkin, Thomas J; Hassett, Daniel J

    2014-10-01

    Here, we constructed stable, chromosomal, constitutively expressed, green and red fluorescent protein (GFP and RFP) as reporters in the select agents, Bacillus anthracis, Yersinia pestis, Burkholderia mallei, and Burkholderia pseudomallei. Using bioinformatic approaches and other experimental analyses, we identified P0253 and P1 as potent promoters that drive the optimal expression of fluorescent reporters in single copy in B. anthracis and Burkholderia spp. as well as their surrogate strains, respectively. In comparison, Y. pestis and its surrogate strain need two chromosomal copies of cysZK promoter (P2cysZK) for optimal fluorescence. The P0253-, P2cysZK-, and P1-driven GFP and RFP fusions were first cloned into the vectors pRP1028, pUC18R6KT-mini-Tn7T-Km, pmini-Tn7-gat, or their derivatives. The resultant constructs were delivered into the respective surrogates and subsequently into the select agent strains. The chromosomal GFP- and RFP-tagged strains exhibited bright fluorescence at an exposure time of less than 200 msec and displayed the same virulence traits as their wild-type parental strains. The utility of the tagged strains was proven by the macrophage infection assays and lactate dehydrogenase release analysis. Such strains will be extremely useful in high-throughput screens for novel compounds that could either kill these organisms, or interfere with critical virulence processes in these important bioweapon agents and during infection of alveolar macrophages. © 2014 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  8. RsaM: a transcriptional regulator of Burkholderia spp. with novel fold

    Energy Technology Data Exchange (ETDEWEB)

    Michalska, Karolina [Midwest Center for Structural Genomics, Argonne National Laboratory, IL USA; Structural Biology Center, Biosciences Division, Argonne National Laboratory, IL USA; Chhor, Gekleng [Midwest Center for Structural Genomics, Argonne National Laboratory, IL USA; Clancy, Shonda [Midwest Center for Structural Genomics, Argonne National Laboratory, IL USA; Jedrzejczak, Robert [Midwest Center for Structural Genomics, Argonne National Laboratory, IL USA; Babnigg, Gyorgy [Midwest Center for Structural Genomics, Argonne National Laboratory, IL USA; Winans, Stephen C. [Department of Microbiology, Cornell University, Ithaca NY USA; Joachimiak, Andrzej [Midwest Center for Structural Genomics, Argonne National Laboratory, IL USA; Structural Biology Center, Biosciences Division, Argonne National Laboratory, IL USA; Department of Biochemistry and Molecular Biology, University of Chicago, IL USA

    2014-07-04

    Burkholderia cepacia complex (Bcc) is a set of closely related bacterial species that are notorious pathogens of cystic fibrosis patients, responsible for life-threatening lung infections. Expression of several virulence factors of Bcc is controlled by a mechanism known as quorum sensing (QS). QS is a means of bacterial communication used to coordinate gene expression in a cell-density-dependent manner. The system involves the production of diffusible signaling molecules (N-acyl-L-homoserine lactones, AHLs), that bind to cognate transcriptional regulators and influence their ability to regulate gene expression. One such system that is highly conserved in Bcc consists of CepI and CepR. CepI is AHL synthase, while CepR is an AHL-dependent transcription factor. In most members of the Bcc group, the cepI and cepR genes are divergently transcribed and separated by additional genes. One of them, bcam1869, encodes the BcRsaM protein, which was recently postulated to modulate the abundance or activity of CepI or CepR. Here we show the crystal structure of BcRsaM from B. cenocepacia J2315. It is a single-domain protein with unique topology and presents a novel fold. The protein is a dimer in the crystal and in solution. This regulator has no known DNA binding motifs and direct binding of BcRsaM to the cepI promoter could not be detected in in vitro assays. Therefore, we propose that the modulatory action of RsaM might result from interactions with other components of the QS machinery rather than from direct association with the DNA promoter.

  9. 40 CFR 725.1075 - Burkholderia cepacia complex.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Burkholderia cepacia complex. 725.1075... Specific Microorganisms § 725.1075 Burkholderia cepacia complex. (a) Microorganism and significant new uses subject to reporting. (1) The microorganisms identified as the Burkholderia cepacia complex defined as...

  10. Phosphorus uptake of an arbuscular mycorrhizal fungus is not effected by the biocontrol bacterium ¤Burkholderia cepacia¤

    DEFF Research Database (Denmark)

    Ravnskov, S.; Larsen, J.; Jakobsen, I.

    2002-01-01

    The biocontrol bacterium Burkholderia cepacia is known to suppress a broad range of root pathogenic fungi, while its impact on other beneficial non-target organisms such as arbuscular mycorrhizal (AM) fungi is unknown. Direct interactions between five B. cepacia strains and the AM fungus, Glomus ...

  11. 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...

  12. Alanine racemase mutants of Burkholderia pseudomallei and Burkholderia mallei and use of alanine racemase as a non-antibiotic-based selectable marker.

    Directory of Open Access Journals (Sweden)

    Sheryl L W Zajdowicz

    Full Text Available Burkholderia pseudomallei and Burkholderia mallei are category B select agents and must be studied under BSL3 containment in the United States. They are typically resistant to multiple antibiotics, and the antibiotics used to treat B. pseudomallei or B. mallei infections may not be used as selective agents with the corresponding Burkholderia species. Here, we investigated alanine racemase deficient mutants of B. pseudomallei and B. mallei for development of non-antibiotic-based genetic selection methods and for attenuation of virulence. The genome of B. pseudomallei K96243 has two annotated alanine racemase genes (bpsl2179 and bpss0711, and B. mallei ATCC 23344 has one (bma1575. Each of these genes encodes a functional enzyme that can complement the alanine racemase deficiency of Escherichia coli strain ALA1. Herein, we show that B. pseudomallei with in-frame deletions in both bpsl2179 and bpss0711, or B. mallei with an in-frame deletion in bma1575, requires exogenous D-alanine for growth. Introduction of bpsl2179 on a multicopy plasmid into alanine racemase deficient variants of either Burkholderia species eliminated the requirement for D-alanine. During log phase growth without D-alanine, the viable counts of alanine racemase deficient mutants of B. pseudomallei and B. mallei decreased within 2 hours by about 1000-fold and 10-fold, respectively, and no viable bacteria were present at 24 hours. We constructed several genetic tools with bpsl2179 as a selectable genetic marker, and we used them without any antibiotic selection to construct an in-frame ΔflgK mutant in the alanine racemase deficient variant of B. pseudomallei K96243. In murine peritoneal macrophages, wild type B. mallei ATCC 23344 was killed much more rapidly than wild type B. pseudomallei K96243. In addition, the alanine racemase deficient mutant of B. pseudomallei K96243 exhibited attenuation versus its isogenic parental strain with respect to growth and survival in murine

  13. Alanine Racemase Mutants of Burkholderia pseudomallei and Burkholderia mallei and Use of Alanine Racemase as a Non-Antibiotic-Based Selectable Marker

    Science.gov (United States)

    Zajdowicz, Sheryl L. W.; Jones-Carson, Jessica; Vazquez-Torres, Andres; Jobling, Michael G.; Gill, Ronald E.; Holmes, Randall K.

    2011-01-01

    Burkholderia pseudomallei and Burkholderia mallei are category B select agents and must be studied under BSL3 containment in the United States. They are typically resistant to multiple antibiotics, and the antibiotics used to treat B. pseudomallei or B. mallei infections may not be used as selective agents with the corresponding Burkholderia species. Here, we investigated alanine racemase deficient mutants of B. pseudomallei and B. mallei for development of non-antibiotic-based genetic selection methods and for attenuation of virulence. The genome of B. pseudomallei K96243 has two annotated alanine racemase genes (bpsl2179 and bpss0711), and B. mallei ATCC 23344 has one (bma1575). Each of these genes encodes a functional enzyme that can complement the alanine racemase deficiency of Escherichia coli strain ALA1. Herein, we show that B. pseudomallei with in-frame deletions in both bpsl2179 and bpss0711, or B. mallei with an in-frame deletion in bma1575, requires exogenous d-alanine for growth. Introduction of bpsl2179 on a multicopy plasmid into alanine racemase deficient variants of either Burkholderia species eliminated the requirement for d-alanine. During log phase growth without d-alanine, the viable counts of alanine racemase deficient mutants of B. pseudomallei and B. mallei decreased within 2 hours by about 1000-fold and 10-fold, respectively, and no viable bacteria were present at 24 hours. We constructed several genetic tools with bpsl2179 as a selectable genetic marker, and we used them without any antibiotic selection to construct an in-frame ΔflgK mutant in the alanine racemase deficient variant of B. pseudomallei K96243. In murine peritoneal macrophages, wild type B. mallei ATCC 23344 was killed much more rapidly than wild type B. pseudomallei K96243. In addition, the alanine racemase deficient mutant of B. pseudomallei K96243 exhibited attenuation versus its isogenic parental strain with respect to growth and survival in murine peritoneal macrophages

  14. Genome sequencing and transposon mutagenesis of Burkholderia seminalis TC3.4.2R3 identify genes contributing to suppression of orchid necrosis caused by B. gladioli

    Science.gov (United States)

    Thirty six strains of Burkholderia spp. isolated from sugarcane were evaluated for biological control of leaf and pseudobulb necrosis of orchid caused by B. gladioli. Twenty nine of the sugarcane strains suppressed the disease in greenhouse assays. We generated a draft genomic sequence of one suppr...

  15. A reverse-phase protein microarray-based screen identifies host signaling dynamics upon Burkholderia spp. infection

    Directory of Open Access Journals (Sweden)

    Chih-Yuan eChiang

    2015-07-01

    Full Text Available Burkholderia is a diverse genus of Gram-negative bacteria that cause high mortality rate in humans and cattle. The lack of effective therapeutic treatments poses serious public health threats. Insights toward host-Burkholderia spp. interaction are critical in understanding the pathogenesis of the infection as well as identifying therapeutic targets for drug development. Reverse-phase protein microarray (RPMA technology was previously proven to characterize novel biomarkers and molecular signatures associated with infectious diseases and cancers. In the present study, this technology was utilized to interrogate changes in host protein expression and post-translational phosphorylation events in macrophages infected with a collection of geographically diverse strains of Burkholderia spp. The expression or phosphorylation state of 25 proteins was altered during Burkholderia spp. infections and of which eight proteins were selected for further validation by immunoblotting. Kinetic expression patterns of phosphorylated AMPK-α1, Src, and GSK3β suggested the importance of their roles in regulating Burkholderia spp. mediated innate immune responses. Modulating inflammatory responses by perturbing AMPK-α1, Src, and GSK3β activities may provide novel therapeutic targets for future treatments.

  16. The lipopolysaccharide core oligosaccharide of Burkholderia plays a critical role in maintaining a proper gut symbiosis with the bean bug Riptortus pedestris.

    Science.gov (United States)

    Kim, Jiyeun Kate; Jang, Ho Am; Kim, Min Seon; Cho, Jae Hyun; Lee, Junbeom; Di Lorenzo, Flaviana; Sturiale, Luisa; Silipo, Alba; Molinaro, Antonio; Lee, Bok Luel

    2017-11-24

    Lipopolysaccharide, the outer cell-wall component of Gram-negative bacteria, has been shown to be important for symbiotic associations. We recently reported that the lipopolysaccharide O-antigen of Burkholderia enhances the initial colonization of the midgut of the bean bug, Riptortus pedestris However, the midgut-colonizing Burkholderia symbionts lack the O-antigen but display the core oligosaccharide on the cell surface. In this study, we investigated the role of the core oligosaccharide, which directly interacts with the host midgut, in the Riptortus-Burkholderia symbiosis. To this end, we generated the core oligosaccharide mutant strains, Δ wabS , Δ wabO , Δ waaF, and Δ waaC, and determined the chemical structures of their oligosaccharides, which exhibited different compositions. The symbiotic properties of these mutant strains were compared with those of the wild-type and O-antigen-deficient Δ wbiG strains. Upon introduction into Riptortus via the oral route, the core oligosaccharide mutant strains exhibited different rates of colonization of the insect midgut. The symbiont titers in fifth-instar insects revealed significantly reduced population sizes of the inner core oligosaccharide mutant strains Δ waaF and Δ waaC These two strains also negatively affected host growth rate and fitness. Furthermore, R. pedestris individuals colonized with the Δ waaF and Δ waaC strains were vulnerable to septic bacterial challenge, similar to insects without a Burkholderia symbiont. Taken together, these results suggest that the core oligosaccharide from Burkholderia symbionts plays a critical role in maintaining a proper symbiont population and in supporting the beneficial effects of the symbiont on its host in the Riptortus-Burkholderia symbiosis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Burkholderia humi sp nov., Burkholderia choica sp nov., Burkholderia telluris sp nov., Burkholderia terrestris sp nov and Burkholderia udeis sp nov. : Burkholderia glathei-like bacteria from soil and rhizosphere soil

    NARCIS (Netherlands)

    Vandamme, Peter; De Brandt, Evie; Houf, Kurt; Salles, Joana Falcao; van Elsas, Jan Dirk; Spilker, Theodore; LiPuma, John J.

    2013-01-01

    Analysis of partial gyrB gene sequences revealed six taxa in a group of 17 Burkholderia glathei-like isolates which were further examined by (GTG)(5)-PCR fingerprinting, 16S rRNA gene sequence analysis, DNA-DNA hybridizations, determination of the DNA G+C content, whole-cell fatty acid analysis and

  18. Identification of Burkholderia spp. in the Clinical Microbiology Laboratory: Comparison of Conventional and Molecular Methods

    Science.gov (United States)

    van Pelt, Cindy; Verduin, Cees M.; Goessens, Wil H. F.; Vos, Margreet C.; Tümmler, Burkhard; Segonds, Christine; Reubsaet, Frans; Verbrugh, Henri; van Belkum, Alex

    1999-01-01

    Cystic fibrosis (CF) predisposes patients to bacterial colonization and infection of the lower airways. Several species belonging to the genus Burkholderia are potential CF-related pathogens, but microbiological identification may be complicated. This situation is not in the least due to the poorly defined taxonomic status of these bacteria, and further validation of the available diagnostic assays is required. A total of 114 geographically diverse bacterial isolates, previously identified in reference laboratories as Burkholderia cepacia (n = 51), B. gladioli (n = 14), Ralstonia pickettii (n = 6), B. multivorans (n = 2), Stenotrophomonas maltophilia (n = 3), and Pseudomonas aeruginosa (n = 11), were collected from environmental, clinical, and reference sources. In addition, 27 clinical isolates putatively identified as Burkholderia spp. were recovered from the sputum of Dutch CF patients. All isolates were used to evaluate the accuracy of two selective growth media, four systems for biochemical identification (API 20NE, Vitek GNI, Vitek NFC, and MicroScan), and three different PCR-based assays. The PCR assays amplify different parts of the ribosomal DNA operon, either alone or in combination with cleavage by various restriction enzymes (PCR-restriction fragment length polymorphism [RFLP] analysis). The best system for the biochemical identification of B. cepacia appeared to be the API 20NE test. None of the biochemical assays successfully grouped the B. gladioli strains. The PCR-RFLP method appeared to be the optimal method for accurate nucleic acid-mediated identification of the different Burkholderia spp. With this method, B. gladioli was also reliably classified in a separate group. For the laboratory diagnosis of B. cepacia, we recommend parallel cultures on blood agar medium and selective agar plates. Further identification of colonies with a Burkholderia phenotype should be performed with the API 20NE test. For final confirmation of species identities, PCR

  19. Retraction for Ramos et al., The second RNA chaperone, Hfq2, is also required for survival under stress and full virulence of Burkholderia cenocepacia J2315.

    Science.gov (United States)

    2014-11-01

    Volume 193, no. 7, p.1515–1526, 2011. Problems related to images published in this paper have been brought to our attention. Figure 8 contains duplicated images as well as images previously published in articles in Microbiology and Microbial Pathogenesis, i.e., the following: S. A. Sousa, C. G. Ramos, L. M. Moreira, and J. H. Leitão, Microbiology 156:896–908, 2010. http://dx.doi.org/10.1099/mic.0.035139-0. C. G. Ramos, S. A. Sousa, A. M. Grilo, L. Eberl, and J. H. Leitão, Microb. Pathog. 48:168–177, 2010. http://dx.doi.org/10.1016 /j.micpath.2010.02.006. Therefore, we retract the paper.Wedeeply regret this situation and apologize for any inconvenience to the editors and readers of Journal of Bacteriology, Microbial Pathogenesis, and Microbiology.

  20. Gene expression profiling of Burkholderia cenocepacia at the time of cepacia syndrome: loss of motility as a marker of poor prognosis?

    Czech Academy of Sciences Publication Activity Database

    Kalferstová, L.; Kolář, Michal; Fila, L.; Vávrová, J.; Dřevínek, P.

    2015-01-01

    Roč. 53, č. 5 (2015), s. 1515-1522 ISSN 0095-1137 Grant - others:GA MŠk(CZ) LD11029; GA MZd(CZ) NT12405 Institutional support: RVO:68378050 Keywords : CYSTIC-FIBROSIS PATIENTS * COMPLEX * VIRULENCE Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.631, year: 2015

  1. Burkholderia metalliresistens sp. nov., a multiple metal-resistant and phosphate-solubilising species isolated from heavy metal-polluted soil in Southeast China.

    Science.gov (United States)

    Guo, Jun Kang; Ding, Yong Zhen; Feng, Ren Wei; Wang, Rui Gang; Xu, Ying Ming; Chen, Chun; Wei, Xiu Li; Chen, Wei Min

    2015-06-01

    A metal-resistant and phosphate-solubilising bacterium, designated as strain D414(T), was isolated from heavy metal (Pb, Cd, Cu and Zn)-polluted paddy soils at the surrounding area of Dabao Mountain Mine in Southeast China. The minimum inhibitory concentrations of heavy metals for strain D414(T) were 2000 mg L(-1) (Cd), 800 mg L(-1) (Pb), 150 mg L(-1) (Cu) and 2500 mg L(-1) (Zn). The strain possessed plant growth-promoting properties, such as 1-aminocyclopropane-1-carboxylate assimilation, indole production and phosphate solubilisation. Analysis of 16S rRNA gene sequence indicated that the isolate is a member of the genus Burkholderia where strain D414(T) formed a distinct phyletic line with validly described Burkholderia species. Strain D414(T) is closely related to Burkholderia tropica DSM 15359(T), B. bannensis NBRC E25(T) and B. unamae DSM 17197(T), with 98.5, 98.3 and 98.3 % sequence similarities, respectively. Furthermore, less than 34 % DNA-DNA relatedness was detected between strain D414(T) and the type strains of the phylogenetically closest species of Burkholderia. The dominant fatty acids of strain D414(T) were C14:0, C16:0, C17:0 cyclo and C18:1 ω7c. The DNA G+C content was 62.3 ± 0.5 mol%. On the basis of genotypic, phenotypic and phylogenetic data, strain D414(T) represents a novel species, for which the name Burkholderia metalliresistens sp. nov. is proposed, with D414(T) (=CICC 10561(T) = DSM 26823(T)) as the type strain.

  2. Draft Genome Sequence of Pseudomonas sp. Strain Ep R1 Isolated from Echinacea purpurea Roots and Effective in the Growth Inhibition of Human Opportunistic Pathogens Belonging to the Burkholderia cepacia Complex.

    Science.gov (United States)

    Maggini, Valentina; Presta, Luana; Miceli, Elisangela; Fondi, Marco; Bosi, Emanuele; Chiellini, Carolina; Fagorzi, Camilla; Bogani, Patrizia; Di Pilato, Vincenzo; Rossolini, Gian Maria; Mengoni, Alessio; Firenzuoli, Fabio; Perrin, Elena; Fani, Renato

    2017-05-18

    In this announcement, we detail the draft genome sequence of the Pseudomonas sp. strain Ep R1, isolated from the roots of the medicinal plant Echinacea purpurea The elucidation of this genome sequence may allow the identification of genes associated with the production of antimicrobial compounds. Copyright © 2017 Maggini et al.

  3. Development of ceftazidime resistance in an acute Burkholderia pseudomallei infection

    Directory of Open Access Journals (Sweden)

    Sarovich DS

    2012-08-01

    Full Text Available Derek S Sarovich,1,2,* Erin P Price,1,2,* Direk Limmathurotsakul,3 James M Cook,1 Alex T Von Schulze,1 Spenser R Wolken,1 Paul Keim,1 Sharon J Peacock,3,4 Talima Pearson1 1Center for Microbial Genetics and Genomics, Northern Arizona University, Flagstaff, AZ, USA; 2Tropical and Emerging Infectious Diseases Division, Menzies School of Health Research, Darwin, Australia; 3Department of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand; 4Department of Medicine, University of Cambridge, Cambridge, United Kingdom*These authors contributed equally to this workAbstract: Burkholderia pseudomallei, a bacterium that causes the disease melioidosis, is intrinsically resistant to many antibiotics. First-line antibiotic therapy for treating melioidosis is usually the synthetic β-lactam, ceftazidime (CAZ, as almost all B. pseudomallei strains are susceptible to this drug. However, acquired CAZ resistance can develop in vivo during treatment with CAZ, which can lead to mortality if therapy is not switched to a different drug in a timely manner. Serial B. pseudomallei isolates obtained from an acute Thai melioidosis patient infected by a CAZ susceptible strain, who ultimately succumbed to infection despite being on CAZ therapy for the duration of their infection, were analyzed. Isolates that developed CAZ resistance due to a proline to serine change at position 167 in the β-lactamase PenA were identified. Importantly, these CAZ resistant isolates remained sensitive to the alternative melioidosis treatments; namely, amoxicillin-clavulanate, imipenem, and meropenem. Lastly, real-time polymerase chain reaction-based assays capable of rapidly identifying CAZ resistance in B. pseudomallei isolates at the position 167 mutation site were developed. The ability to rapidly identify the emergence of CAZ resistant B. pseudomallei populations in melioidosis patients will allow timely alterations in treatment strategies

  4. Use of a Real-Time PCR TaqMan Assay for Rapid Identification and Differentiation of Burkholderia pseudomallei and Burkholderia mallei

    OpenAIRE

    U'Ren, Jana M.; Van Ert, Matthew N.; Schupp, James M.; Easterday, W. Ryan; Simonson, Tatum S.; Okinaka, Richard T.; Pearson, Talima; Keim, Paul

    2005-01-01

    A TaqMan allelic-discrimination assay designed around a synonymous single-nucleotide polymorphism was used to genotype Burkholderia pseudomallei and Burkholderia mallei isolates. The assay rapidly identifies and discriminates between these two highly pathogenic bacteria and does not cross-react with genetic near neighbors, such as Burkholderia thailandensis and Burkholderia cepacia.

  5. 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.

  6. Burkholderia in gladiolen: voortgezet diagnostisch onderzoek 2007

    NARCIS (Netherlands)

    Vink, P.; Hollinger, T.C.

    2008-01-01

    In 2006 is middels een infectieproef bekend geworden dat de bacterie Burkholderia gladioli in staat is een ziekte bij gladiolen te veroorzaken waardoor de sier- en handelswaarde zeer negatief worden beïnvloed. In 2007 is in het kader van het voortgezet diagnostisch onderzoek nagegaan of de bacterie

  7. GENOME ANALYSIS OF BURKHOLDERIA CEPACIA AC1100

    Science.gov (United States)

    Burkholderia cepacia is an important organism in bioremediation of environmental pollutants and it is also of increasing interest as a human pathogen. The genomic organization of B. cepacia is being studied in order to better understand its unusual adaptive capacity and genome pl...

  8. 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

  9. Human Infection with Burkholderia thailandensis, China, 2013.

    Science.gov (United States)

    Chang, Kai; Luo, Jie; Xu, Huan; Li, Min; Zhang, Fengling; Li, Jin; Gu, Dayong; Deng, Shaoli; Chen, Ming; Lu, Weiping

    2017-08-01

    Burkholderia thailandensis infection in humans is uncommon. We describe a case of B. thailandensis infection in a person in China, a location heretofore unknown for B. thailandensis. We identified the specific virulence factors of B. thailandensis, which may indicate a transition to a new virulent form.

  10. Burkholderia pseudomallei septicaemia - A case report

    Directory of Open Access Journals (Sweden)

    Dias M

    2004-01-01

    Full Text Available Burkholderia pseudomallei, a natural saprophyte widely distributed in soil, stagnant waters of endemic areas, is said to infect humans through breaks in the skin or through inhalation causing protean clinical manifestations including fatal septicaemia. A case of septicaemia in a elderly female diabetic due to B. pseudomallei following a history of fall is being reported with complete details.

  11. Burkholderia pseudomallei Antibodies in Children, Cambodia

    Science.gov (United States)

    Pheaktra, Ngoun; Putchhat, Hor; Sin, Lina; Sen, Bun; Kumar, Varun; Langla, Sayan; Peacock, Sharon J.; Day, Nicholas P.

    2008-01-01

    Antibodies to Burkholderia pseudomallei were detected in 16% of children in Siem Reap, Cambodia. This organism was isolated from 30% of rice paddies in the surrounding vicinity. Despite the lack of reported indigenous cases, melioidosis is likely to occur in Cambodia. PMID:18258125

  12. Recovery of a Burkholderia thailandensis-like isolate from an Australian water source

    Directory of Open Access Journals (Sweden)

    Wilkins Patricia P

    2008-04-01

    Full Text Available Abstract Background Burkholderia thailandensis, a close relative of Burkholderia pseudomallei, has previously been reported only from Southeast Asia and North America. It is biochemically differentiated from B. pseudomallei by the ability to utilize arabinose. During the course of environmental sampling for B. pseudomallei in the Northern Territory of Australia, an isolate, MSMB 43, was recovered that is arabinose positive. Results Genetic analysis using 16S rDNA sequencing and DNA/DNA hybridization indicates that MSMB 43 is most similar to B. thailandensis although multi-locus sequence typing indicates that this isolate is divergent from both B. pseudomallei and other described B. thailandensis. Conclusion We report the isolation and initial characterization of strain MSMB 43, which is a B. thailandensis-like isolate recovered in Australia.

  13. AN IMPORTED CASE OF ACUTE MELIOIDOSIS CAUSED BY ST881 BURKHOLDERIA PSEUDOMALLEI.

    Science.gov (United States)

    Zong, Zhiyong; Wang, Xiaohui; Deng, Yiyun

    2016-03-01

    A previously healthy Chinese male working in Malaysia returned to China with high fever. A blood culture showed Burkholderia pseudomallei strain WCBP1. This isolate was sequenced, showing type, ST881, which appears to be present in Malaysia. WCP1 had unusual susceptibility to aminoglycosides and habored the Yersinia-like fimbrial gene cluster for virulence. The patient's condition deteriorated rapidly but he recovered after receiving meropenem and intensive care support. Melioidosis is a potential problem among Chinese imigrant workers with strains new to China being identified.

  14. Symbiotic factors in Burkholderia essential for establishing an association with the bean bug, Riptortus pedestris.

    Science.gov (United States)

    Kim, Jiyeun Kate; Lee, Bok Luel

    2015-01-01

    Symbiotic bacteria are common in insects and intimately affect the various aspects of insect host biology. In a number of insect symbiosis models, it has been possible to elucidate the effects of the symbiont on host biology, whereas there is a limited understanding of the impact of the association on the bacterial symbiont, mainly due to the difficulty of cultivating insect symbionts in vitro. Furthermore, the molecular features that determine the establishment and persistence of the symbionts in their host (i.e., symbiotic factors) have remained elusive. However, the recently established model, the bean bug Riptortus pedestris, provides a good opportunity to study bacterial symbiotic factors at a molecular level through their cultivable symbionts. Bean bugs acquire genus Burkholderia cells from the environment and harbor them as gut symbionts in the specialized posterior midgut. The genome of the Burkholderia symbiont was sequenced, and the genomic information was used to generate genetically manipulated Burkholderia symbiont strains. Using mutant symbionts, we identified several novel symbiotic factors necessary for establishing a successful association with the host gut. In this review, these symbiotic factors are classified into three categories based on the colonization dynamics of the mutant symbiont strains: initiation, accommodation, and persistence factors. In addition, the molecular characteristics of the symbiotic factors are described. These newly identified symbiotic factors and on-going studies of the Riptortus-Burkholderia symbiosis are expected to contribute to the understanding of the molecular cross-talk between insects and bacterial symbionts that are of ecological and evolutionary importance. © 2014 Wiley Periodicals, Inc.

  15. Enhanced degradation of haloacid by heterologous expression in related Burkholderia species.

    Science.gov (United States)

    Su, Xianbin; Deng, Liyu; Kong, Ka Fai; Tsang, Jimmy S H

    2013-10-01

    Haloacids are environmental pollutant and can be transformed to non-toxic alkanoic acids by microbial dehalogenase. Bacterium Burkholderia species MBA4 was enriched from soil for its ability to bioremediate haloacids such as mono-chloroacetate (MCA), mono-bromoacetate (MBA), 2-mono-chloropropionate, and 2-mono-bromopropionate. MBA4 produces an inducible dehalogenase Deh4a that catalyzes the dehalogenation process. The growth of MBA4 on haloacid also relies on the presence of a haloacid-uptake system. Similar dehalogenase genes can be found in the genome of many related species. However, wildtype Burkholderia caribensis MWAP64, Burkholderia phymatum STM815, and Burkholderia xenovorans LB400 were not able to grow on MCA. When a plasmid containing the regulatory and structural gene of Deh4a was transformed to these species, they were able to grow on haloacid. The specific enzyme activities in these recombinants ranges from 2- to 30-fold that of MBA4 in similar condition. Reverse transcription-quantitative real-time PCR showed that the relative transcript levels in these recombinant strains ranges from 9 to over 1,600 times that of MBA4 in similar condition. A recombinant has produced nearly five times of dehalogenase that MBA4 could ever achieve. While the expressions of Deh4a were more relaxed in these phylogenetically related species, an MCA-uptake activity was found to be inducible. These metabolically engineered strains are better degraders than the haloacid-enriched MBA4. Copyright © 2013 Wiley Periodicals, Inc.

  16. 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

  17. Burkholderia contaminans Biofilm Regulating Operon and Its Distribution in Bacterial Genomes.

    Science.gov (United States)

    Voronina, Olga L; Kunda, Marina S; Ryzhova, Natalia N; Aksenova, Ekaterina I; Semenov, Andrey N; Romanova, Yulia M; Gintsburg, Alexandr L

    2016-01-01

    Biofilm formation by Burkholderia spp. is a principal cause of lung chronic infections in cystic fibrosis patients. A "lacking biofilm production" (LBP) strain B. contaminans GIMC4587:Bct370-19 has been obtained by insertion modification of clinical strain with plasposon mutagenesis. It has an interrupted transcriptional response regulator (RR) gene. The focus of our investigation was a two-component signal transduction system determination, including this RR. B. contaminans clinical and LBP strains were analyzed by whole genome sequencing and bioinformatics resources. A four-component operon (BiofilmReg) has a key role in biofilm formation. The relative location (i.e., by being separated by another gene) of RR and histidine kinase genes is unique in BiofilmReg. Orthologs were found in other members of the Burkholderiales order. Phylogenetic analysis of strains containing BiofilmReg operons demonstrated evidence for earlier inheritance of a three-component operon. During further evolution one lineage acquired a fourth gene, whereas others lost the third component of the operon. Mutations in sensor domains have created biodiversity which is advantageous for adaptation to various ecological niches. Different species Burkholderia and Achromobacter strains all demonstrated similar BiofilmReg operon structure. Therefore, there may be an opportunity to develop a common drug which is effective for treating all these causative agents.

  18. Cross-species comparison of the Burkholderia pseudomallei, Burkholderia thailandensis, and Burkholderia mallei quorum-sensing regulons.

    Science.gov (United States)

    Majerczyk, Charlotte D; Brittnacher, Mitchell J; Jacobs, Michael A; Armour, Christopher D; Radey, Matthew C; Bunt, Richard; Hayden, Hillary S; Bydalek, Ryland; Greenberg, E Peter

    2014-11-01

    Burkholderia pseudomallei, Burkholderia thailandensis, and Burkholderia mallei (the Bptm group) are close relatives with very different lifestyles: B. pseudomallei is an opportunistic pathogen, B. thailandensis is a nonpathogenic saprophyte, and B. mallei is a host-restricted pathogen. The acyl-homoserine lactone quorum-sensing (QS) systems of these three species show a high level of conservation. We used transcriptome sequencing (RNA-seq) to define the quorum-sensing regulon in each species, and we performed a cross-species analysis of the QS-controlled orthologs. Our analysis revealed a core set of QS-regulated genes in all three species, as well as QS-controlled factors shared by only two species or unique to a given species. This global survey of the QS regulons of B. pseudomallei, B. thailandensis, and B. mallei serves as a platform for predicting which QS-controlled processes might be important in different bacterial niches and contribute to the pathogenesis of B. pseudomallei and B. mallei. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  19. Cross-Species Comparison of the Burkholderia pseudomallei, Burkholderia thailandensis, and Burkholderia mallei Quorum-Sensing Regulons

    Science.gov (United States)

    Majerczyk, Charlotte D.; Brittnacher, Mitchell J.; Jacobs, Michael A.; Armour, Christopher D.; Radey, Matthew C.; Bunt, Richard; Hayden, Hillary S.; Bydalek, Ryland

    2014-01-01

    Burkholderia pseudomallei, Burkholderia thailandensis, and Burkholderia mallei (the Bptm group) are close relatives with very different lifestyles: B. pseudomallei is an opportunistic pathogen, B. thailandensis is a nonpathogenic saprophyte, and B. mallei is a host-restricted pathogen. The acyl-homoserine lactone quorum-sensing (QS) systems of these three species show a high level of conservation. We used transcriptome sequencing (RNA-seq) to define the quorum-sensing regulon in each species, and we performed a cross-species analysis of the QS-controlled orthologs. Our analysis revealed a core set of QS-regulated genes in all three species, as well as QS-controlled factors shared by only two species or unique to a given species. This global survey of the QS regulons of B. pseudomallei, B. thailandensis, and B. mallei serves as a platform for predicting which QS-controlled processes might be important in different bacterial niches and contribute to the pathogenesis of B. pseudomallei and B. mallei. PMID:25182491

  20. Burkholderia dipogonis sp. nov., isolated from root nodules of Dipogon lignosus in New Zealand and Western Australia.

    Science.gov (United States)

    Sheu, Shih-Yi; Chen, Ming-Hui; Liu, Wendy Y Y; Andrews, Mitchell; James, Euan K; Ardley, Julie K; De Meyer, Sofie E; James, Trevor K; Howieson, John G; Coutinho, Bruna G; Chen, Wen-Ming

    2015-12-01

    Seven strains, ICMP 19430T, ICMP 19429, ICMP 19431, WSM4637, WSM4638, WSM4639 and WSM4640, were isolated from nitrogen-fixing nodules on roots of the invasive South African legume Dipogon lignosus (subfamily Papilionoideae, tribe Phaseoleae) in New Zealand and Western Australia, and their taxonomic positions were investigated by using a polyphasic approach. All seven strains grew at 10-37 °C (optimum, 25-30 °C), at pH 4.0-9.0 (optimum, pH 6.0-7.0) and with 0-2 % (w/v) NaCl (optimum growth in the absence of NaCl). On the basis of 16S rRNA gene sequence analysis, the strains showed 99.0-99.5 % sequence similarity to the closest type strain, Burkholderia phytofirmans PsJNT, and 98.4-99.7 % sequence similarity to Burkholderia caledonica LMG 19076T. The predominant fatty acids were C18 : 1ω7c (21.0 % of the total fatty acids in strain ICMP 19430T), C16 : 0 (19.1 %), C17 : 0 cyclo (18.9 %), summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c; 10.7 %) and C19 : 0 cyclov ω8c (7.5 %). The polar lipid profile consisted of a mixture of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and several uncharacterized aminophospholipids and phospholipids. The major isoprenoid quinone was Q-8 and the DNA G+C content of strain ICMP 19430T was 63.2 mol%. The DNA–DNA relatedness of the novel strains with respect to the closest neighbouring members of the genus Burkholderia was 55 % or less. On the basis of 16S rRNA and recA gene sequence similarities and chemotaxonomic and phenotypic data,these strains represent a novel symbiotic species in the genus Burkholderia, for which the name Burkholderia dipogonis sp. nov. is proposed, with the type strain ICMP 19430T (=LMG28415T=HAMBI 3637T).

  1. Detection of Burkholderia pseudomallei O-antigen serotypes in near-neighbor species

    Directory of Open Access Journals (Sweden)

    Stone Joshua K

    2012-11-01

    Full Text Available Abstract Background Burkholderia pseudomallei is the etiological agent of melioidosis and a CDC category B select agent with no available effective vaccine. Previous immunizations in mice have utilized the lipopolysaccharide (LPS as a potential vaccine target because it is known as one of the most important antigenic epitopes in B. pseudomallei. Complicating this strategy are the four different B. pseudomallei LPS O-antigen types: A, B, B2, and rough. Sero-crossreactivity is common among O-antigens of Burkholderia species. Here, we identified the presence of multiple B. pseudomallei O-antigen types and sero-crossreactivity in its near-neighbor species. Results PCR screening of O-antigen biosynthesis genes, phenotypic characterization using SDS-PAGE, and immunoblot analysis showed that majority of B. mallei and B. thailandensis strains contained the typical O-antigen type A. In contrast, most of B. ubonensis and B. thailandensis-like strains expressed the atypical O-antigen types B and B2, respectively. Most B. oklahomensis strains expressed a distinct and non-seroreactive O-antigen type, except strain E0147 which expressed O-antigen type A. O-antigen type B2 was also detected in B. thailandensis 82172, B. ubonensis MSMB108, and Burkholderia sp. MSMB175. Interestingly, B. thailandensis-like MSMB43 contained a novel serotype B positive O-antigen. Conclusions This study expands the number of species which express B. pseudomallei O-antigen types. Further work is required to elucidate the full structures and how closely these are to the B. pseudomallei O-antigens, which will ultimately determine the efficacy of the near-neighbor B serotypes for vaccine development.

  2. Microbiological assessment of Burkholderia cepacia complex (Bcc ...

    African Journals Online (AJOL)

    Nancy Omar

    2014-09-18

    Sep 18, 2014 ... tum 4/35 (11.4%) and urine 1/35 (2.9%). Other studies reported higher rates of isolation of B. cepa- cia complex from specimens other than those in our study. Gales et al. (2005)3 found that out of 176 NFGNB (83/176) belonging to Burkholderia spp.: 52/83 (62.7%) were from blood, 25/83 (30.1%) were from ...

  3. Burkholderia thailandensis: Growth and Laboratory Maintenance.

    Science.gov (United States)

    Garcia, Erin C; Cotter, Peggy A

    2016-08-12

    Burkholderia thailandensis is a nonpathogenic Gram-negative bacterium found in tropical soils. Closely related to several human pathogens, its ease of genetic manipulation, rapid growth in the laboratory, and low virulence make B. thailandensis a commonly used model organism. This unit describes the fundamental protocols for in vitro growth and maintenance of B. thailandensis in the laboratory. © 2016 by John Wiley & Sons, Inc. Copyright © 2016 John Wiley & Sons, Inc.

  4. Polymorphisms within the prnD and pltC genes from pyrrolnitrin and pyoluteorin-producing Pseudomonas and Burkholderia spp

    NARCIS (Netherlands)

    Souza, J.T.; Raaijmakers, J.M.

    2003-01-01

    Pyrrolnitrin (PRN) and pyoluteorin (PLT) are broad-spectrum antibiotics produced by several strains of Pseudomonas and Burkholderia species. Both antibiotics play an important role in the suppression of multiple plant pathogenic fungi. Primers were developed from conserved sequences and amplified

  5. Burkholderia kirstenboschensis sp. nov. nodulates papilionoid legumes indigenous to South Africa.

    Science.gov (United States)

    Steenkamp, Emma T; van Zyl, Elritha; Beukes, Chrizelle W; Avontuur, Juanita R; Chan, Wai Yin; Palmer, Marike; Mthombeni, Lunghile S; Phalane, Francina L; Sereme, T Karabo; Venter, Stephanus N

    2015-12-01

    Despite the diversity of Burkholderia species known to nodulate legumes in introduced and native regions, relatively few taxa have been formally described. For example, the Cape Floristic Region of South Africa is thought to represent one of the major centres of diversity for the rhizobial members of Burkholderia, yet only five species have been described from legumes occurring in this region and numerous are still awaiting taxonomic treatment. Here, we investigated the taxonomic status of 12 South African root-nodulating Burkholderia isolates from native papilionoid legumes (Hypocalyptus coluteoides, H. oxalidifolius, H. sophoroides and Virgilia oroboides). Analysis of four gene regions (16S rRNA, recA, atpD and rpoB) revealed that the isolates represent a genealogically unique and exclusive assemblage within the genus. Its distinctness was supported by all other aspects of the polyphasic approach utilized, including the genome-based criteria DNA-DNA hybridization (≥70.9%) and average nucleotide identities (≥96%). We accordingly propose the name B. kirstenboschensis sp. nov. for this taxon with isolate Kb15(T) (=LMG 28727(T); =SARC 695(T)) as its type strain. Our data showed that intraspecific genome size differences (≥0.81 Mb) and the occurrence of large DNA regions that are apparently unique to single individuals (16-23% of an isolate's genome) can significantly limit the value of data obtained from DNA-DNA hybridization experiments. Substitution of DNA-DNA hybridization with whole genome sequencing as a prerequisite for the description of Burkholderia species will undoubtedly speed up the pace at which their diversity are documented, especially in hyperdiverse regions such as the Cape Floristic Region. Copyright © 2015 Elsevier GmbH. All rights reserved.

  6. Competition Experiments for Legume Infection Identify Burkholderia phymatum as a Highly Competitive β-Rhizobium

    Directory of Open Access Journals (Sweden)

    Martina Lardi

    2017-08-01

    Full Text Available Members of the genus Burkholderia (β-proteobacteria have only recently been shown to be able to establish a nitrogen-fixing symbiosis with several legumes, which is why they are also referred to as β-rhizobia. Therefore, very little is known about the competitiveness of these species to nodulate different legume host plants. In this study, we tested the competitiveness of several Burkholderia type strains (B. diazotrophica, B. mimosarum, B. phymatum, B. sabiae, B. symbiotica and B. tuberum to nodulate four legumes (Phaseolus vulgaris, Macroptilium atropurpureum, Vigna unguiculata and Mimosa pudica under our closely defined growth conditions. The assessment of nodule occupancy of these species on different legume host plants revealed that B. phymatum was the most competitive strain in the three papilionoid legumes (bean, cowpea and siratro, while B. mimosarum outcompeted the other strains in mimosa. The analysis of phenotypes known to play a role in nodulation competitiveness (motility, exopolysaccharide production and additional in vitro competition assays among β-rhizobial strains suggested that B. phymatum has the potential to be a very competitive legume symbiont.

  7. 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

  8. Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei

    Directory of Open Access Journals (Sweden)

    Suat Moi Puah

    2014-01-01

    Full Text Available The Gram-negative saprophyte Burkholderia pseudomallei is the causative agent of melioidosis, an infectious disease which is endemic in Southeast Asia and northern Australia. This bacterium possesses many virulence factors which are thought to contribute to its survival and pathogenicity. Using a virulent clinical isolate of B. pseudomallei and an attenuated strain of the same B. pseudomallei isolate, 6 genes BPSL2033, BP1026B_I2784, BP1026B_I2780, BURPS1106A_A0094, BURPS1106A_1131, and BURPS1710A_1419 were identified earlier by PCR-based subtractive hybridization. These genes were extensively characterized at the molecular level, together with an additional gene BPSL3147 that had been identified by other investigators. Through a reverse genetic approach, single-gene knockout mutants were successfully constructed by using site-specific insertion mutagenesis and were confirmed by PCR. BPSL2033::Km and BURPS1710A_1419::Km mutants showed reduced rates of survival inside macrophage RAW 264.7 cells and also low levels of virulence in the nematode infection model. BPSL2033::Km demonstrated weak statistical significance (P=0.049 at 8 hours after infection in macrophage infection study but this was not seen in BURPS1710A_1419::Km. Nevertheless, complemented strains of both genes were able to partially restore the gene defects in both in vitro and in vivo studies, thus suggesting that they individually play a minor role in the virulence of B. pseudomallei.

  9. Identification and characterization of Burkholderia multivorans CCA53.

    Science.gov (United States)

    Akita, Hironaga; Kimura, Zen-Ichiro; Yusoff, Mohd Zulkhairi Mohd; Nakashima, Nobutaka; Hoshino, Tamotsu

    2017-07-06

    A lignin-degrading bacterium, Burkholderia sp. CCA53, was previously isolated from leaf soil. The purpose of this study was to determine phenotypic and biochemical features of Burkholderia sp. CCA53. Multilocus sequence typing (MLST) analysis based on fragments of the atpD, gltD, gyrB, lepA, recA and trpB gene sequences was performed to identify Burkholderia sp. CCA53. The MLST analysis revealed that Burkholderia sp. CCA53 was tightly clustered with B. multivorans ATCC BAA-247 T . The quinone and cellular fatty acid profiles, carbon source utilization, growth temperature and pH were consistent with the characteristics of B. multivorans species. Burkholderia sp. CCA53 was therefore identified as B. multivorans CCA53.

  10. Transcriptome analysis of Acidovorax avenae subsp. avenae cultivated in vivo and co-culture with Burkholderia seminalis

    OpenAIRE

    Bin Li; Muhammad Ibrahim; Mengyu Ge; Zhouqi Cui; Guochang Sun; Fei Xu; Michael Kube

    2014-01-01

    Response of bacterial pathogen to environmental bacteria and its host is critical for understanding of microbial adaption and pathogenesis. Here, we used RNA-Seq to comprehensively and quantitatively assess the transcriptional response of Acidovorax avenae subsp. avenae strain RS-1 cultivated in vitro, in vivo and in co-culture with rice rhizobacterium Burkholderia seminalis R456. Results revealed a slight response to other bacteria, but a strong response to host. In particular, a large numbe...

  11. Membrane-active mechanism of LFchimera against Burkholderia pseudomallei and Burkholderia thailandensis

    NARCIS (Netherlands)

    Kanthawong, S.; Puknun, A.; Bolscher, J.G.M.; Nazmi, K.; van Marle, J.; de Soet, J.J.; Veerman, E.C.I.; Wongratanacheewin, S.; Taweechaisupapong, S.

    2014-01-01

    LFchimera, a construct combining two antimicrobial domains of bovine lactoferrin, lactoferrampin265-284 and lactoferricin17-30, possesses strong bactericidal activity. As yet, no experimental evidence was presented to evaluate the mechanisms of LFchimera against Burkholderia isolates. In this study

  12. Relationships within the Proteobacteria of plant pathogenic Acidovorax species and subspecies, Burkholderia species, and Herbaspirillum rubrisubalbicans by sequence analysis of 16S rDNA, numerical analysis and determinative tests.

    Science.gov (United States)

    Hu, F P; Young, J M; Triggs, C M; Park, D C; Saul, D J

    2001-12-01

    Sequence data for 16S rDNA of the type strains of Acidovorax avenae subsp. avenae, A. avenae subsp. cattleyae, A. avenae subsp. citrulli, A. konjaci and Herbaspirillum rubrisubalbicans were compared with GenBank library accessions of Burkholderia spp., Comamonas sp., Ralstonia solanacearum and Variovorax sp. Maximum Parsimony analysis produced two clusters: 1. Acidovorax spp., Comamonas sp., and Variovorax sp. (all in the Comamonadaceae), and 2. Burkholderia spp., Ralstonia solanacearum, and Herbaspirillum rubrisubalbicans. Maximum Likelihood analysis produced only one cluster (of the Comamonadaceae). Using nutritional and laboratory tests, all Acidovorax spp., Burkholderia spp., and Herbaspirillum rubrisubalbicans were discriminated in distinct clusters at the species level, and could be identified by selected determinative tests. There were no phenotypic tests constituted as a circumscription of the genera and which permitted the allocation of strains to genera. Strain identification as species allowed allocation to genera only by inference. The nomenclatural implications of these data are discussed.

  13. Understanding the direction of evolution in Burkholderia glumae through comparative genomics.

    Science.gov (United States)

    Lee, Hyun-Hee; Park, Jungwook; Kim, Jinnyun; Park, Inmyoung; Seo, Young-Su

    2016-02-01

    Members of the genus Burkholderia occupy remarkably diverse niches, with genome sizes ranging from ~3.75 to 11.29 Mbp. The genome of Burkholderia glumae ranges in size from ~5.81 to 7.89 Mbp. Unlike other plant pathogenic bacteria, B. glumae can infect a wide range of monocot and dicot plants. Comparative genome analysis of B. glumae strains can provide insight into genome variation as well as differential features of whole metabolism or pathways between multiple strains of B. glumae infecting the same host. Comparative analysis of complete genomes among B. glumae BGR1, B. glumae LMG 2196, and B. glumae PG1 revealed the largest departmentalization of genes onto separate replicons in B. glumae BGR1 and considerable downsizing of the genome in B. glumae LMG 2196. In addition, the presence of large-scale evolutionary events such as rearrangement and inversion and the development of highly specialized systems were found to be related to virulence-associated features in the three B. glumae strains. This connection may explain why this bacterium broadens its host range and reinforces its interaction with hosts.

  14. Biochemical Characterization and Structural Basis of Reactivity and Regioselectivity Differences between Burkholderia thailandensis and Burkholderia glumae 1,6-Didesmethyltoxoflavin N-Methyltransferase.

    Science.gov (United States)

    Fenwick, Michael K; Almabruk, Khaled H; Ealick, Steven E; Begley, Tadhg P; Philmus, Benjamin

    2017-08-01

    Burkholderia glumae converts the guanine base of guanosine triphosphate into an azapteridine and methylates both the pyrimidine and triazine rings to make toxoflavin. Strains of Burkholderia thailandensis and Burkholderia pseudomallei have a gene cluster encoding seven putative biosynthetic enzymes that resembles the toxoflavin gene cluster. Four of the enzymes are similar in sequence to BgToxBCDE, which have been proposed to make 1,6-didesmethyltoxoflavin (1,6-DDMT). One of the remaining enzymes, BthII1283 in B. thailandensis E264, is a predicted S-adenosylmethionine (SAM)-dependent N-methyltransferase that shows a low level of sequence identity to BgToxA, which sequentially methylates N6 and N1 of 1,6-DDMT to form toxoflavin. Here we show that, unlike BgToxA, BthII1283 catalyzes a single methyl transfer to N1 of 1,6-DDMT in vitro. In addition, we investigated the differences in reactivity and regioselectivity by determining crystal structures of BthII1283 with bound S-adenosylhomocysteine (SAH) or 1,6-DDMT and SAH. BthII1283 contains a class I methyltransferase fold and three unique extensions used for 1,6-DDMT recognition. The active site structure suggests that 1,6-DDMT is bound in a reduced form. The plane of the azapteridine ring system is orthogonal to its orientation in BgToxA. In BthII1283, the modeled SAM methyl group is directed toward the p orbital of N1, whereas in BgToxA, it is first directed toward an sp 2 orbital of N6 and then toward an sp 2 orbital of N1 after planar rotation of the azapteridine ring system. Furthermore, in BthII1283, N1 is hydrogen bonded to a histidine residue whereas BgToxA does not supply an obvious basic residue for either N6 or N1 methylation.

  15. Genetic diversity and microevolution of Burkholderia pseudomallei in the environment.

    Directory of Open Access Journals (Sweden)

    Narisara Chantratita

    2008-02-01

    Full Text Available The soil dwelling Gram-negative pathogen Burkholderia pseudomallei is the cause of melioidosis. The diversity and population structure of this organism in the environment is poorly defined.We undertook a study of B. pseudomallei in soil sampled from 100 equally spaced points within 237.5 m(2 of disused land in northeast Thailand. B. pseudomallei was present on direct culture of 77/100 sampling points. Genotyping of 200 primary plate colonies from three independent sampling points was performed using a combination of pulsed field gel electrophoresis (PFGE and multilocus sequence typing (MLST. Twelve PFGE types and nine sequence types (STs were identified, the majority of which were present at only a single sampling point. Two sampling points contained four STs and the third point contained three STs. Although the distance between the three sampling points was low (7.6, 7.9, and 13.3 meters, respectively, only two STs were present in more than one sampling point. Each of the three samples was characterized by the localized expansion of a single B. pseudomallei clone (corresponding to STs 185, 163, and 93. Comparison of PFGE and MLST results demonstrated that two STs contained strains with variable PFGE banding pattern types, indicating geographic structuring even within a single MLST-defined clone.We discuss the implications of this extreme structuring of genotype and genotypic frequency in terms of micro-evolutionary dynamics and ecology, and how our results may inform future sampling strategies.

  16. Brain abscess caused by Burkholderia pseudomallei

    International Nuclear Information System (INIS)

    Padigione, A.; Spelman, D.; Ferris, N.

    1997-01-01

    Full text: Melioidosis, or infection with Burkholderia pseudomallei, is an important human disease in South East Asia and Northern Australia. Neurological manifestations are well recognized amongst its protean presentations, but direct focal central nervous system infection is infrequently described with only 9 adult and 5 paediatric cases reported in the English language literature. A case of brain abscess due to Burkholderia pseudomallei occurring in a 20 year old Dutch visitor to Australia which progressed despite antibiotic treatment is described. A review of the clinical manifestations, Magnetic Resonance (MR) appearance, diagnosis and treatment of melioidosis is presented, highlighting that: (i) physicians outside endernic areas should consider melioidosis in any patient with an appropriate travel history, (ii) MR imaging is more sensitive then CT in diagnosing early brain infection, especially of the brainstem; (iii) Bacterial culture, the mainstay of diagnosis, has many shortcomings; (iv)In vitro antibiotic sensitivity testing may not translate into clinical efficacy; and (v) Steroids appear to have little role, even in severe disease

  17. Development of a multiplex PCR assay for the detection and differentiation of Burkholderia pseudomallei, Burkholderia mallei, Burkholderia thailandensis, and Burkholderia cepacia complex.

    Science.gov (United States)

    Zakharova, Irina; Teteryatnikova, Natalya; Toporkov, Andrey; Viktorov, Dmitry

    2017-10-01

    Two species of Burkholderia pseudomallei complex (Bpc), B. pseudomallei and B. mallei, can cause severe life-threatening infections. Rapidly discerning individual species within the group and separating them from other opportunistic pathogens of the Burkholderia cepacia complex (Bcc) is essential to establish a correct diagnosis and for epidemiological surveillance. In this study, a multiplex PCR assay based on the detection of an individual set of chromosomal beta-lactamase genes for single-step identification and differentiation of B. pseudomallei, B. mallei, B. thailandensis, and Bcc was developed. Two pairs of primers specific to a distinct class of B metallo-beta-lactamase genes and a pair of primers specific to the oxacillin-hydrolyzing class D beta-lactamase gene were demonstrated to successfully discriminate species within Bpc and from Bcc. The assay sensitivity was 9561 genomic equivalents (GE) for B. pseudomallei, 7827 GE for B. mallei, 8749 GE for B. thailandensis and 6023 GE for B. cepacia. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Burkholderia glumae: next major pathogen of rice?

    Science.gov (United States)

    Ham, Jong Hyun; Melanson, Rebecca A; Rush, Milton C

    2011-05-01

    Burkholderia glumae causes bacterial panicle blight of rice, which is an increasingly important disease problem in global rice production. Toxoflavin and lipase are known to be major virulence factors of this pathogen, and their production is dependent on the TofI/TofR quorum-sensing system, which is mediated by N-octanoyl homoserine lactone. Flagellar biogenesis and a type III secretion system are also required for full virulence of B. glumae. Bacterial panicle blight is thought to be caused by seed-borne B. glumae; however, its disease cycle is not fully understood. In spite of its economic importance, neither effective control measures for bacterial panicle blight nor rice varieties showing complete resistance to the disease are currently available. A better understanding of the molecular mechanisms underlying B. glumae virulence and of the rice defence mechanisms against the pathogen would lead to the development of better methods of disease control for bacterial panicle blight. Bacteria; Proteobacteria; Betaproteobacteria; Burkholderiales; Burkholderiaceae; Burkholderia. Gram-negative, capsulated, motile, lophotrichous flagella, pectolytic. Aborted seed, empty grains as a result of failure of grain filling, brown spots on panicles, seedling rot. Seed sterilization, planting partially resistant lines (no completely resistant line is available). KNOWN VIRULENCE FACTORS: Toxoflavin, lipase, type III effectors. © 2010 LSU AGCENTER. MOLECULAR PLANT PATHOLOGY © 2010 BSPP AND BLACKWELL PUBLISHING LTD.

  19. Burkholderia xenovorans LB400 harbors a multi-replicon, 9.73-Mbp genome shaped for versatility.

    Science.gov (United States)

    Chain, Patrick S G; Denef, Vincent J; Konstantinidis, Konstantinos T; Vergez, Lisa M; Agulló, Loreine; Reyes, Valeria Latorre; Hauser, Loren; Córdova, Macarena; Gómez, Luis; González, Myriam; Land, Miriam; Lao, Victoria; Larimer, Frank; LiPuma, John J; Mahenthiralingam, Eshwar; Malfatti, Stephanie A; Marx, Christopher J; Parnell, J Jacob; Ramette, Alban; Richardson, Paul; Seeger, Michael; Smith, Daryl; Spilker, Theodore; Sul, Woo Jun; Tsoi, Tamara V; Ulrich, Luke E; Zhulin, Igor B; Tiedje, James M

    2006-10-17

    Burkholderia xenovorans LB400 (LB400), a well studied, effective polychlorinated biphenyl-degrader, has one of the two largest known bacterial genomes and is the first nonpathogenic Burkholderia isolate sequenced. From an evolutionary perspective, we find significant differences in functional specialization between the three replicons of LB400, as well as a more relaxed selective pressure for genes located on the two smaller vs. the largest replicon. High genomic plasticity, diversity, and specialization within the Burkholderia genus are exemplified by the conservation of only 44% of the genes between LB400 and Burkholderia cepacia complex strain 383. Even among four B. xenovorans strains, genome size varies from 7.4 to 9.73 Mbp. The latter is largely explained by our findings that >20% of the LB400 sequence was recently acquired by means of lateral gene transfer. Although a range of genetic factors associated with in vivo survival and intercellular interactions are present, these genetic factors are likely related to niche breadth rather than determinants of pathogenicity. The presence of at least eleven "central aromatic" and twenty "peripheral aromatic" pathways in LB400, among the highest in any sequenced bacterial genome, supports this hypothesis. Finally, in addition to the experimentally observed redundancy in benzoate degradation and formaldehyde oxidation pathways, the fact that 17.6% of proteins have a better LB400 paralog than an ortholog in a different genome highlights the importance of gene duplication and repeated acquirement, which, coupled with their divergence, raises questions regarding the role of paralogs and potential functional redundancies in large-genome microbes.

  20. Burkholderia xernovorans LB400 harbors a multi-replicon, 9.73-Mbp genome shaped for versatility

    Energy Technology Data Exchange (ETDEWEB)

    Chain, Patrick S. G. [Lawrence Livermore National Laboratory (LLNL); Denef, Vincent [University of California, Berkeley; Konstantinidis, Konstantinos T [Michigan State University, East Lansing; Vergez, Lisa [Lawrence Livermore National Laboratory (LLNL); Agullo, Loreine [Universidad Tecnica Federico Santa Maria, Casilla 110-V; Reyes, Valeria Latorre [Universidad Tecnica Federico Santa Maria, Casilla 110-V; Hauser, Loren John [ORNL; Cordova, Macarena [Universidad Tecnica Federico Santa Maria, Casilla 110-V; Gomez, Luis [Universidad Tecnica Federico Santa Maria, Casilla 110-V; Gonzalez, Myriam [Universidad Tecnica Federico Santa Maria, Casilla 110-V; Land, Miriam L [ORNL; Lao, Victoria [Lawrence Livermore National Laboratory (LLNL); Larimer, Frank W [ORNL; LiPuma, John J [University of Michigan; Mahenthiralingam, Eshwar [Cardiff University, Wales; Malfatti, Stephanie [Lawrence Livermore National Laboratory (LLNL); Marx, Christopher J [Harvard University; Parnell, J Jacob [Michigan State University, East Lansing; Ramette, Alban [Michigan State University, East Lansing; Richardson, P M [U.S. Department of Energy, Joint Genome Institute; Seeger, Michael [Universidad Tecnica Federico Santa Maria, Casilla 110-V; Smith, Daryl [University of British Columbia, Vancouver; Spilker, Theodore [University of Michigan; Sul, Woo Jun [Michigan State University, East Lansing; Tsoi, Tamara V [Michigan State University, East Lansing; Zhulin, Igor B [University of Tennessee, Knoxville (UTK) & Oak Ridge National Laboratory (ORNL); Tiedje, James M. [Michigan State University, East Lansing

    2006-01-01

    Burkholderia xenovorans LB400 (LB400), a well studied, effective polychlorinated biphenyl-degrader, has one of the two largest known bacterial genomes and is the first nonpathogenic Burkholderia isolate sequenced. From an evolutionary perspective, we find significant differences in functional specialization between the three replicons of LB400, as well as a more relaxed selective pressure for genes located on the two smaller vs. the largest replicon. High genomic plasticity, diversity, and specialization within the Burkholderia genus are exemplified by the conservation of only 44% of the genes between LB400 and Burkholderia cepacia complex strain 383. Even among four B. xenovorans strains, genome size varies from 7.4 to 9.73 Mbp. The latter is largely explained by our findings that >20% of the LB400 sequence was recently acquired by means of lateral gene transfer. Although a range of genetic factors associated with in vivo survival and intercellular interactions are present, these genetic factors are likely related to niche breadth rather than determinants of pathogenicity. The presence of at least eleven 'central aromatic' and twenty 'peripheral aromatic' pathways in LB400, among the highest in any sequenced bacterial genome, supports this hypothesis. Finally, in addition to the experimentally observed redundancy in benzoate degradation and formaldehyde oxidation pathways, the fact that 17.6% of proteins have a better LB400 paralog than an ortholog in a different genome highlights the importance of gene duplication and repeated acquirement, which, coupled with their divergence, raises questions regarding the role of paralogs and potential functional redundancies in large-genome microbes.

  1. Reclassification of the Specialized Metabolite Producer Pseudomonas mesoacidophila ATCC 31433 as a Member of the Burkholderia cepacia Complex.

    Science.gov (United States)

    Loveridge, E Joel; Jones, Cerith; Bull, Matthew J; Moody, Suzy C; Kahl, Małgorzata W; Khan, Zainab; Neilson, Louis; Tomeva, Marina; Adams, Sarah E; Wood, Andrew C; Rodriguez-Martin, Daniel; Pinel, Ingrid; Parkhill, Julian; Mahenthiralingam, Eshwar; Crosby, John

    2017-07-01

    Pseudomonas mesoacidophila ATCC 31433 is a Gram-negative bacterium, first isolated from Japanese soil samples, that produces the monobactam isosulfazecin and the β-lactam-potentiating bulgecins. To characterize the biosynthetic potential of P. mesoacidophila ATCC 31433, its complete genome was determined using single-molecule real-time DNA sequence analysis. The 7.8-Mb genome comprised four replicons, three chromosomal (each encoding rRNA) and one plasmid. Phylogenetic analysis demonstrated that P. mesoacidophila ATCC 31433 was misclassified at the time of its deposition and is a member of the Burkholderia cepacia complex, most closely related to Burkholderia ubonensis The sequenced genome shows considerable additional biosynthetic potential; known gene clusters for malleilactone, ornibactin, isosulfazecin, alkylhydroxyquinoline, and pyrrolnitrin biosynthesis and several uncharacterized biosynthetic gene clusters for polyketides, nonribosomal peptides, and other metabolites were identified. Furthermore, P. mesoacidophila ATCC 31433 harbors many genes associated with environmental resilience and antibiotic resistance and was resistant to a range of antibiotics and metal ions. In summary, this bioactive strain should be designated B. cepacia complex strain ATCC 31433, pending further detailed taxonomic characterization. IMPORTANCE This work reports the complete genome sequence of Pseudomonas mesoacidophila ATCC 31433, a known producer of bioactive compounds. Large numbers of both known and novel biosynthetic gene clusters were identified, indicating that P. mesoacidophila ATCC 31433 is an untapped resource for discovery of novel bioactive compounds. Phylogenetic analysis demonstrated that P. mesoacidophila ATCC 31433 is in fact a member of the Burkholderia cepacia complex, most closely related to the species Burkholderia ubonensis Further investigation of the classification and biosynthetic potential of P. mesoacidophila ATCC 31433 is warranted. Copyright © 2017

  2. A prophage tail-like protein is deployed by Burkholderia bacteria to feed on fungi.

    Science.gov (United States)

    Swain, Durga Madhab; Yadav, Sunil Kumar; Tyagi, Isha; Kumar, Rahul; Kumar, Rajeev; Ghosh, Srayan; Das, Joyati; Jha, Gopaljee

    2017-09-01

    Some bacteria can feed on fungi, a phenomenon known as mycophagy. Here we show that a prophage tail-like protein (Bg_9562) is essential for mycophagy in Burkholderia gladioli strain NGJ1. The purified protein causes hyphal disintegration and inhibits growth of several fungal species. Disruption of the Bg_9562 gene abolishes mycophagy. Bg_9562 is a potential effector secreted by a type III secretion system (T3SS) and is translocated into fungal mycelia during confrontation. Heterologous expression of Bg_9562 in another bacterial species, Ralstonia solanacearum, confers mycophagous ability in a T3SS-dependent manner. We propose that the ability to feed on fungi conferred by Bg_9562 may help the bacteria to survive in certain ecological niches. Furthermore, considering its broad-spectrum antifungal activity, the protein may be potentially useful in biotechnological applications to control fungal diseases.Some bacteria can feed on live fungi through unclear mechanisms. Here, the authors show that a T3SS-secreted protein, which is homologous to phage tail proteins, allows a Burkholderia gladioli strain to kill and feed on various fungal species.

  3. Membrane-active mechanism of LFchimera against Burkholderia pseudomallei and Burkholderia thailandensis.

    Science.gov (United States)

    Kanthawong, Sakawrat; Puknun, Aekkalak; Bolscher, Jan G M; Nazmi, Kamran; van Marle, Jan; de Soet, Johannes J; Veerman, Enno C I; Wongratanacheewin, Surasakdi; Taweechaisupapong, Suwimol

    2014-10-01

    LFchimera, a construct combining two antimicrobial domains of bovine lactoferrin, lactoferrampin265-284 and lactoferricin17-30, possesses strong bactericidal activity. As yet, no experimental evidence was presented to evaluate the mechanisms of LFchimera against Burkholderia isolates. In this study we analyzed the killing activity of LFchimera on the category B pathogen Burkholderia pseudomallei in comparison to the lesser virulent Burkholderia thailandensis often used as a model for the highly virulent B. pseudomallei. Killing kinetics showed that B. thailandensis E264 was more susceptible for LFchimera than B. pseudomallei 1026b. Interestingly the bactericidal activity of LFchimera appeared highly pH dependent; B. thailandensis killing was completely abolished at and below pH 6.4. FITC-labeled LFchimera caused a rapid accumulation within 15 min in the cytoplasm of both bacterial species. Moreover, freeze-fracture electron microscopy demonstrated extreme effects on the membrane morphology of both bacterial species within 1 h of incubation, accompanied by altered membrane permeability monitored as leakage of nucleotides. These data indicate that the mechanism of action of LFchimera is similar for both species and encompasses disruption of the plasma membrane and subsequently leakage of intracellular nucleotides leading to cell dead.

  4. Workshop on treatment of and postexposure prophylaxis for Burkholderia pseudomallei and B. mallei Infection, 2010

    NARCIS (Netherlands)

    Lipsitz, Rebecca; Garges, Susan; Aurigemma, Rosemarie; Baccam, Prasith; Blaney, David D.; Cheng, Allen C.; Currie, Bart J.; Dance, David; Gee, Jay E.; Larsen, Joseph; Limmathurotsakul, Direk; Morrow, Meredith G.; Norton, Robert; O'Mara, Elizabeth; Peacock, Sharon J.; Pesik, Nicki; Rogers, L. Paige; Schweizer, Herbert P.; Steinmetz, Ivo; Tan, Gladys; Tan, Patrick; Wiersinga, W. Joost; Wuthiekanun, Vanaporn; Smith, Theresa L.

    2012-01-01

    The US Public Health Emergency Medical Countermeasures Enterprise convened subject matter experts at the 2010 HHS Burkholderia Workshop to develop consensus recommendations for postexposure prophylaxis against and treatment for Burkholderia pseudomallei and B. mallei infections, which cause

  5. Alteration in cell surface properties of Burkholderia spp. during surfactant-aided biodegradation of petroleum hydrocarbons

    Energy Technology Data Exchange (ETDEWEB)

    Mohanty, Sagarika; Mukherji, Suparna [Indian Institute of Technology Bombay, Mumbai (India). Centre for Environmental Science and Engineering (CESE)

    2012-04-15

    Chemical surfactants may impact microbial cell surface properties, i.e., cell surface hydrophobicity (CSH) and cell surface charge, and may thus affect the uptake of components from non-aqueous phase liquids (NAPLs). This work explored the impact of Triton X-100, Igepal CA 630, and Tween 80 (at twice the critical micelle concentration, CMC) on the cell surface characteristics of Burkholderia cultures, Burkholderia cepacia (ES1, aliphatic degrader) and Burkholderia multivorans (NG1, aromatic degrader), when grown on a six-component model NAPL. In the presence of Triton X-100, NAPL biodegradation was enhanced from 21% to 60% in B. cepacia and from 18% to 53% in B. multivorans. CSH based on water contact angle (50-52 ) was in the same range for both strains while zeta potential at neutral pH was -38 and -31 mV for B. cepacia and B. multivorans, respectively. In the presence of Triton X-100, their CSH increased to greater than 75 and the zeta potential decreased. This induced a change in the mode of uptake and initiated aliphatic hydrocarbon degradation by B. multivorans and increased the rate of aliphatic hydrocarbon degradation in B. cepacia. Igepal CA 630 and Tween 80 also altered the cell surface properties. For B. cepacia grown in the presence of Triton X-100 at two and five times its CMC, CSH increased significantly in the log growth phase. Growth in the presence of the chemical surfactants also affected the abundance of chemical functional groups on the cell surface. Cell surface changes had maximum impact on NAPL degradation in the presence of emulsifying surfactants, Triton X-100 and Igepal CA630.

  6. Biosurfactant Production by Pseudomonas aeruginosa and Burkholderia gladioli Isolated from Mangrove Sediments Using Alternative Substrates

    Directory of Open Access Journals (Sweden)

    Karla Maria Catter

    2016-10-01

    Full Text Available Biosurfactants are surface-active agents produced by a variety of microorganisms. To make biosurfactant production economically feasible, several alternative carbon sources have been proposed. This study describes biosurfactant production by strains of Pseudomonas aeruginosa and Burkholderia gladioli isolated from mangrove sediments in Northeastern Brazil and cultured in mineral media enriched with waste cooking oil. The biosurfactants were tested for drop collapse, emulsion formation and stability and surface tension. P. aeruginosa performed better both at lowering the surface tension (from 69 to 28 mN/m and at forming stable emulsions (approximately 80% at 48 hours of culture. The strains tested in this study were found to be efficient biosurfactant producers when cultured on substrates enriched with vegetable oil. DOI: http://dx.doi.org/10.17807/orbital.v8i5.771

  7. Cometabolic degradation of trichloroethylene by Burkholderia cepacia G4 with poplar leaf homogenate.

    Science.gov (United States)

    Kang, Jun Won; Doty, Sharon Lafferty

    2014-07-01

    Trichloroethylene (TCE), a chlorinated organic solvent, is one of the most common and widespread groundwater contaminants worldwide. Among the group of TCE-degrading aerobic bacteria, Burkholderia cepacia G4 is the best-known representative. This strain requires the addition of specific substrates, including toluene, phenol, and benzene, to induce the enzymes to degrade TCE. However, the substrates are toxic and introducing them into the soil can result in secondary contamination. In this study, poplar leaf homogenate containing natural phenolic compounds was tested for the ability to induce the growth of and TCE degradation by B. cepacia G4. The results showed that the G4 strain could grow and degrade TCE well with the addition of phytochemicals. The poplar leaf homogenate also functioned as an inducer of the toluene-ortho-monooxygenase (TOM) gene in B. cepacia G4.

  8. Common Duckweed (Lemna minor) Is a Versatile High-Throughput Infection Model For the Burkholderia cepacia Complex and Other Pathogenic Bacteria

    Science.gov (United States)

    Thomson, Euan L. S.; Dennis, Jonathan J.

    2013-01-01

    Members of the Burkholderia cepacia complex (Bcc) have emerged in recent decades as problematic pulmonary pathogens of cystic fibrosis (CF) patients, with severe infections progressing to acute necrotizing pneumonia and sepsis. This study presents evidence that Lemna minor (Common duckweed) is useful as a plant model for the Bcc infectious process, and has potential as a model system for bacterial pathogenesis in general. To investigate the relationship between Bcc virulence in duckweed and Galleria mellonella (Greater wax moth) larvae, a previously established Bcc infection model, a duckweed survival assay was developed and used to determine LD50 values. A strong correlation (R2 = 0.81) was found between the strains’ virulence ranks in the two infection models, suggesting conserved pathways in these vastly different hosts. To broaden the application of the duckweed model, enteropathogenic Escherichia coli (EPEC) and five isogenic mutants with previously established LD50 values in the larval model were tested against duckweed, and a strong correlation (R2 = 0.93) was found between their raw LD50 values. Potential virulence factors in B. cenocepacia K56-2 were identified using a high-throughput screen against single duckweed plants. In addition to the previously characterized antifungal compound (AFC) cluster genes, several uncharacterized genes were discovered including a novel lysR regulator, a histidine biosynthesis gene hisG, and a gene located near the gene encoding the recently characterized virulence factor SuhBBc. Finally, to demonstrate the utility of this model in therapeutic applications, duckweed was rescued from Bcc infection by treating with bacteriophage at 6-h intervals. It was observed that phage application became ineffective at a timepoint that coincided with a sharp increase in bacterial invasion of plant tissue. These results indicate that common duckweed can serve as an effective infection model for the investigation of bacterial virulence

  9. Characterization of the papilionoid-Burkholderia interaction in the Fynbos biome: The diversity and distribution of beta-rhizobia nodulating Podalyria calyptrata (Fabaceae, Podalyrieae).

    Science.gov (United States)

    Lemaire, Benny; Van Cauwenberghe, Jannick; Verstraete, Brecht; Chimphango, Samson; Stirton, Charles; Honnay, Olivier; Smets, Erik; Sprent, Janet; James, Euan K; Muasya, A Muthama

    2016-02-01

    The South African Fynbos soils are renowned for nitrogen-fixing Burkholderia associated with diverse papilionoid legumes of the tribes Crotalarieae, Hypocalypteae, Indigofereae, Phaseoleae and Podalyrieae. However, despite numerous rhizobial studies in the region, the symbiotic diversity of Burkholderia has not been investigated in relation to a specific host legume and its geographical provenance. This study analyzed the diversity of nodulating strains of Burkholderia from the legume species Podalyria calyptrata. Diverse lineages were detected that proved to be closely related to Burkholderia taxa, originating from hosts in other legume tribes. By analyzing the genetic variation of chromosomal (recA) and nodulation (nodA) sequence data in relation to the sampling sites we assessed the geographical distribution patterns of the P. calyptrata symbionts. Although we found a degree of genetically differentiated rhizobial populations, a correlation between genetic (recA and nodA) and geographic distances among populations was not observed, suggesting high rates of dispersal and rhizobial colonization within Fynbos soils. Copyright © 2015 Elsevier GmbH. All rights reserved.

  10. Live imaging of symbiosis: spatiotemporal infection dynamics of a GFP-labelled Burkholderia symbiont in the bean bug Riptortus pedestris

    Science.gov (United States)

    Kikuchi, Yoshitomo; Fukatsu, Takema

    2014-01-01

    Many insects possess endosymbiotic bacteria inside their body, wherein intimate interactions occur between the partners. While recent technological advancements have deepened our understanding of metabolic and evolutionary features of the symbiont genomes, molecular mechanisms underpinning the intimate interactions remain difficult to approach because the insect symbionts are generally uncultivable. The bean bug Riptortus pedestris is associated with the betaproteobacterial Burkholderia symbiont in a posterior region of the midgut, which develops numerous crypts harbouring the symbiont extracellularly. Distinct from other insect symbiotic systems, R. pedestris acquires the Burkholderia symbiont not by vertical transmission but from the environment every generation. By making use of the cultivability and the genetic tractability of the symbiont, we constructed a transgenic Burkholderia strain labelled with green fluorescent protein (GFP), which enabled detailed observation of spatiotemporal dynamics and the colonization process of the symbiont in freshly prepared specimens. The symbiont live imaging revealed that, at the second instar, colonization of the symbiotic midgut M4 region started around 6 h after inoculation (hai). By 24 hai, the symbiont cells appeared in the main tract and also in several crypts of the M4. By 48 hai, most of the crypts were colonized by the symbiont cells. By 72 hai, all the crypts were filled up with the symbiont cells and the symbiont localization pattern continued during the subsequent nymphal development. Quantitative PCR of the symbiont confirmed the infection dynamics quantitatively. These results highlight the stinkbug-Burkholderia gut symbiosis as an unprecedented model for comprehensive understanding of molecular mechanisms underpinning insect symbiosis. PMID:24103110

  11. Mutation of the cyclic di-GMP phosphodiesterase gene in Burkholderia lata SK875 attenuates virulence and enhances biofilm formation.

    Science.gov (United States)

    Jung, Hae-In; Kim, Yun-Jung; Lee, Yun-Jung; Lee, Hee-Soo; Lee, Jung-Kee; Kim, Soo-Ki

    2017-10-01

    Burkholderia sp. is a gram-negative bacterium that commonly exists in the environment, and can cause diseases in plants, animals, and humans. Here, a transposon mutant library of a Burkholderia lata isolate from a pig with swine respiratory disease in Korea was screened for strains showing attenuated virulence in Caenorhabditis elegans. One such mutant was obtained, and the Tn5 insertion junction was mapped to rpfR, a gene encoding a cyclic di-GMP phosphodiesterase that functions as a receptor. Mutation of rpfR caused a reduction in growth on CPG agar and swimming motility as well as a rough colony morphology on Congo red agar. TLC analysis showed reduced AHL secretion, which was in agreement with the results from plate-based and bioluminescence assays. The mutant strain produced significantly more biofilm detected by crystal violet staining than the parent strain. SEM of the mutant strain clearly showed that the overproduced biofilm contained a filamentous structure. These results suggest that the cyclic di-GMP phosphodiesterase RpfR plays an important role in quorum sensing modulation of the bacterial virulence and biofilm formation.

  12. Multilocus Sequence Typing of Historical Burkholderia pseudomallei Isolates Collected in Southeast Asia from 1964 to 1967 Provides Insight into the Epidemiology of Melioidosis

    OpenAIRE

    McCombie, Roberta L.; Finkelstein, Richard A.; Woods, Donald E.

    2006-01-01

    A collection of 207 historically relevant Burkholderia pseudomallei isolates was analyzed by multilocus sequence typing (MLST). The strain collection contains environmental isolates obtained from a geographical distribution survey of B. pseudomallei isolates in Thailand (1964 to 1967), as well as stock cultures and colony variants from the U.S. Army Medical Research Unit (Malaysia), the Walter Reed Army Institute for Research, and the Pasteur Institute (Vietnam). The 207 isolates of the colle...

  13. Transfer of 13 species of the genus Burkholderia to the genus Caballeronia and reclassification of Burkholderia jirisanensis as Paraburkholderia jirisanensis comb. nov.

    Science.gov (United States)

    Dobritsa, Anatoly P; Linardopoulou, Elena V; Samadpour, Mansour

    2017-10-01

    A recent study of a group of Burkholderia glathei-like bacteria resulted in the description of 13 novel species of the genus Burkholderia. However, our analysis of phylogenetic positions of these species and their molecular signatures (conserved protein sequence indels) showed that they belong to the genus Caballeronia, and we propose to transfer them to this genus. The reclassified species names are proposed as Caballeroniaarationis comb. nov., Caballeroniaarvi comb. nov., Caballeroniacalidae comb. nov., Caballeroniacatudaia comb. nov., Caballeroniaconcitans comb. nov., Caballeroniafortuita comb. nov., Caballeroniaglebae comb. nov., Caballeroniahypogeia comb. nov., Caballeroniapedi comb. nov., Caballeroniaperedens comb. nov., Caballeroniaptereochthonis comb. nov., Caballeroniatemeraria comb. nov. and Caballeronia turbans comb. nov. It is also proposed to reclassify Burkholderia jirisanensis as Paraburkholderiajirisanensis comb. nov. Based on the results of the polyphasic study, B. jirisanensis had been described as a member of the A-group of the genus Burkholderiaand the most closely related to Burkholderia rhizosphaerae, Burkholderia humisilvae and Burkholderia solisilvae currently classified as belonging to the genus Paraburkholderia.

  14. The Capsular Polysaccharide of Burkholderia pseudomallei Contributes to Survival in Serum by Reducing Complement Factor C3b Deposition

    OpenAIRE

    Reckseidler-Zenteno, Shauna L.; DeVinney, Rebekah; Woods, Donald E.

    2005-01-01

    Burkholderia pseudomallei produces an extracellular polysaccharide capsule -3)-2-O-acetyl-6-deoxy-β-d-manno-heptopyranose-(1- which has been shown to be an essential virulence determinant. The addition of purified capsule was shown to increase the virulence of a capsule mutant strain in the Syrian hamster model of acute melioidosis. An increase in the number of wild-type B. pseudomallei cells in the blood was seen by 48 h, while the number of capsule mutant cells in the blood declined by 48 h...

  15. Prevalence of Burkholderia pseudomallei in Guangxi, China.

    Science.gov (United States)

    Ma, G; Zheng, D; Cai, Q; Yuan, Z

    2010-01-01

    Melioidosis, an infectious disease caused by the Gram-negative bacterium Burkholderia pseudomallei, is now recognized as an important public health problem in Southeast Asia and tropical northern Australia. Although B. pseudomallei has been detected in various water and soil samples in southeast China, the enviromental distribution of B. pseudomallei in China is unclear. In the winter months of 2007, 154 and 130 soil and water samples, respectively, were collected from several locations in Guangxi, China. The samples were screened for B. pseudomallei by bacterial culture and identification and confirmed by PCR for species-specific 16S rDNA and flagellin genes. B. pseudomallei was detected in 8.4% of the soil samples but in none of the water samples. All positive samples were confined to a single low-lying region from rice paddy fields. Counts of B. pseudomallei ranged from 23 to 521 c.f.u./g soil. This is the first geographical distribution survey of B. pseudomallei in soil in Guangxi, China, and the data are of importance for further evaluating the impact of this pathogen on melioidosis in this region.

  16. Burkholderia pseudomallei transcriptional adaptation in macrophages

    Directory of Open Access Journals (Sweden)

    Chieng Sylvia

    2012-07-01

    Full Text Available Abstract Background Burkholderia pseudomallei is a facultative intracellular pathogen of phagocytic and non-phagocytic cells. How the bacterium interacts with host macrophage cells is still not well understood and is critical to appreciate the strategies used by this bacterium to survive and how intracellular survival leads to disease manifestation. Results Here we report the expression profile of intracellular B. pseudomallei following infection of human macrophage-like U937 cells. During intracellular growth over the 6 h infection period, approximately 22 % of the B. pseudomallei genome showed significant transcriptional adaptation. B. pseudomallei adapted rapidly to the intracellular environment by down-regulating numerous genes involved in metabolism, cell envelope, motility, replication, amino acid and ion transport system and regulatory function pathways. Reduced expression in catabolic and housekeeping genes suggested lower energy requirement and growth arrest during macrophage infection, while expression of genes encoding anaerobic metabolism functions were up regulated. However, whilst the type VI secretion system was up regulated, expression of many known virulence factors was not significantly modulated over the 6hours of infection. Conclusions The transcriptome profile described here provides the first comprehensive view of how B. pseudomallei survives within host cells and will help identify potential virulence factors and proteins that are important for the survival and growth of B. pseudomallei within human cells.

  17. Biological Control of White Rot in Garlic Using Burkholderia pyrrocinia CAB08106-4

    Directory of Open Access Journals (Sweden)

    Kwang Seop Han

    2013-03-01

    Full Text Available White rot caused by Sclerotium cepivorum was reported to be severe soil-born disease on garlic. Disease progress of white rot of garlic (Allium sativum L. was investigated during the growing season of 2009 to 2011 at Taean and Seosan areas. The white rot disease on bulb began to occur from late April and peaked in late May. The antifungal bacteria, Burkholderia pyrrocinia CAB08106-4 was tested in field bioassay for suppression of white rot disease. As a result of the nucleotide sequence of the gene 16S rRNA, CAB008106-4 strain used in this study has been identified as B. pyrrocinia. B. pyrrocinia CAB080106-4 isolate suppressed the white rot with 69.6% control efficacy in field test. These results suggested that B. pyrrocinia CAB08106-4 isolate could be an effective biological control agent against white rot of garlic.

  18. The multiple roles of hypothetical gene BPSS1356 in Burkholderia pseudomallei.

    Directory of Open Access Journals (Sweden)

    Hokchai Yam

    Full Text Available Burkholderia pseudomallei is an opportunistic pathogen and the causative agent of melioidosis. It is able to adapt to harsh environments and can live intracellularly in its infected hosts. In this study, identification of transcriptional factors that associate with the β' subunit (RpoC of RNA polymerase was performed. The N-terminal region of this subunit is known to trigger promoter melting when associated with a sigma factor. A pull-down assay using histidine-tagged B. pseudomallei RpoC N-terminal region as bait showed that a hypothetical protein BPSS1356 was one of the proteins bound. This hypothetical protein is conserved in all B. pseudomallei strains and present only in the Burkholderia genus. A BPSS1356 deletion mutant was generated to investigate its biological function. The mutant strain exhibited reduced biofilm formation and a lower cell density during the stationary phase of growth in LB medium. Electron microscopic analysis revealed that the ΔBPSS1356 mutant cells had a shrunken cytoplasm indicative of cell plasmolysis and a rougher surface when compared to the wild type. An RNA microarray result showed that a total of 63 genes were transcriptionally affected by the BPSS1356 deletion with fold change values of higher than 4. The expression of a group of genes encoding membrane located transporters was concurrently down-regulated in ΔBPSS1356 mutant. Amongst the affected genes, the putative ion transportation genes were the most severely suppressed. Deprivation of BPSS1356 also down-regulated the transcriptions of genes for the arginine deiminase system, glycerol metabolism, type III secretion system cluster 2, cytochrome bd oxidase and arsenic resistance. It is therefore obvious that BPSS1356 plays a multiple regulatory roles on many genes.

  19. Burkholderia pseudomallei isolates in 2 pet iguanas, California, USA.

    Science.gov (United States)

    Zehnder, Ashley M; Hawkins, Michelle G; Koski, Marilyn A; Lifland, Barry; Byrne, Barbara A; Swanson, Alexandra A; Rood, Michael P; Gee, Jay E; Elrod, Mindy Glass; Beesley, Cari A; Blaney, David D; Ventura, Jean; Hoffmaster, Alex R; Beeler, Emily S

    2014-02-01

    Burkholderia pseudomallei, the causative agent of melioidosis, was isolated from abscesses of 2 pet green iguanas in California, USA. The international trade in iguanas may contribute to importation of this pathogen into countries where it is not endemic and put persons exposed to these animals at risk for infection.

  20. Burkholderia pseudomallei Isolates in 2 Pet Iguanas, California, USA

    OpenAIRE

    Zehnder, Ashley M.; Hawkins, Michelle G.; Koski, Marilyn A.; Lifland, Barry; Byrne, Barbara A.; Swanson, Alexandra A.; Rood, Michael P.; Gee, Jay E.; Elrod, Mindy Glass; Beesley, Cari A.; Blaney, David D.; Ventura, Jean; Hoffmaster, Alex R.; Beeler, Emily S.

    2014-01-01

    Burkholderia pseudomallei, the causative agent of melioidosis, was isolated from abscesses of 2 pet green iguanas in California, USA. The international trade in iguanas may contribute to importation of this pathogen into countries where it is not endemic and put persons exposed to these animals at risk for infection.

  1. Dicty_cDB: Contig-U16219-1 [Dicty_cDB

    Lifescience Database Archive (English)

    Full Text Available CP000094 |pid:none) Pseudomonas fluorescens Pf0-1, ... 235 6e-60 AB241242_1( AB241242 |pid:none) Symbiotic p...59 CP000378_251( CP000378 |pid:none) Burkholderia cenocepacia AU 1054... 233 2e-59 AB241241_1( AB241241 |pid:none) Symbiotic...243 |pid:none) Trichomonas vaginalis strain G3 sm... 220 1e-55 AB241243_1( AB241243 |pid:none) Symbiotic pro

  2. Role of phosphate solubilizing Burkholderia spp. for successful colonization and growth promotion of Lycopodium cernuum L. (Lycopodiaceae) in lateritic belt of Birbhum district of West Bengal, India.

    Science.gov (United States)

    Ghosh, Ranjan; Barman, Soma; Mukherjee, Rajib; Mandal, Narayan C

    2016-02-01

    Profuse growth of Lycpodium cernuum L. was found in phosphate deficient red lateritic soil of West Bengal, India. Interaction of vesicular-arbuscular mycorrhiza (VAM) with Lycopodium rhizoids were described earlier but association of PGPR with their rhizoids were not studied. Three potent phosphate solubilizing bacterial strains (P4, P9 and P10) associated with L. cernuum rhizoids were isolated and identified by 16S rDNA homologies on Ez-Taxon database as Burkholderia tropica, Burkholderia unamae and Burkholderia cepacia respectively. Day wise kinetics of phosphate solubilization against Ca3(PO4)2 suggested P4 (580.56±13.38 μg ml(-1)) as maximum mineral phosphate solubilizer followed by P9 (517.12±17.15 μg ml(-1)) and P10 (485.18±14.23 μg ml(-1)) at 28 °C. Release of bound phosphates by isolated strains from ferric phosphate (FePO4), aluminum phosphate (AlPO4) and four different complex rock phosphates indicated their very good phosphate solubilizng efficacy. Nitrogen independent solubilizition also supports their nitrogen fixing capabilities. Inhibition of P solubilization by calcium salts and induction by EDTA suggested pH dependent chelation of metal cations by all of the isolates. Rhizoidal colonization potentials of Burkholderia spp. were confirmed by in planta experiment and also using scanning electron microscope (SEM). Increases of total phosphate content in Lycopodium plants upon soil treatment with these isolates were also recorded. In addition siderophore production on CAS agar medium, tryptophan dependent IAA production and antifungal activities against pathogenic fungi by rhizospheric isolates deep-rooted that they have definite role in nutrient mobilization for successful colonization of L. cernuum in nutrient deficient lateritic soil. Copyright © 2015 Elsevier GmbH. All rights reserved.

  3. Burkholderia: an update on taxonomy and biotechnological potential as antibiotic producers.

    Science.gov (United States)

    Depoorter, Eliza; Bull, Matt J; Peeters, Charlotte; Coenye, Tom; Vandamme, Peter; Mahenthiralingam, Eshwar

    2016-06-01

    Burkholderia is an incredibly diverse and versatile Gram-negative genus, within which over 80 species have been formally named and multiple other genotypic groups likely represent new species. Phylogenetic analysis based on the 16S rRNA gene sequence and core genome ribosomal multilocus sequence typing analysis indicates the presence of at least three major clades within the genus. Biotechnologically, Burkholderia are well-known for their bioremediation and biopesticidal properties. Within this review, we explore the ability of Burkholderia to synthesise a wide range of antimicrobial compounds ranging from historically characterised antifungals to recently described antibacterial antibiotics with activity against multiresistant clinical pathogens. The production of multiple Burkholderia antibiotics is controlled by quorum sensing and examples of quorum sensing pathways found across the genus are discussed. The capacity for antibiotic biosynthesis and secondary metabolism encoded within Burkholderia genomes is also evaluated. Overall, Burkholderia demonstrate significant biotechnological potential as a source of novel antibiotics and bioactive secondary metabolites.

  4. Bioleaching remediation of heavy metal-contaminated soils using Burkholderia sp. Z-90.

    Science.gov (United States)

    Yang, Zhihui; Zhang, Zhi; Chai, Liyuan; Wang, Yong; Liu, Yi; Xiao, Ruiyang

    2016-01-15

    Bioleaching is an environment-friendly and economical technology to remove heavy metals from contaminated soils. In this study, a biosurfactant-producing strain with capacity of alkaline production was isolated from cafeteria sewer sludge and its capability for removing Zn, Pb, Mn, Cd, Cu, and As was investigated. Phylogenetic analysis using 16S rDNA gene sequences confirmed that the strain belonged to Burkholderia sp. and named as Z-90. The biosurfactant was glycolipid confirmed by thin layer chromatography and Fourier-transform infrared spectroscopy. Z-90 broth was then used for bioleaching remediation of heavy metal-contaminated soils. The removal efficiency was 44.0% for Zn, 32.5% for Pb, 52.2% for Mn, 37.7% for Cd, 24.1% for Cu and 31.6% for As, respectively. Mn, Zn and Cd were more easily removed from soil than Cu, Pb and As, which was attributed to the presence of high acid-soluble fraction of Mn, Zn and Cd and high residual fraction of Cu, Pb and As. The heavy metal removal in soils was contributed to the adhesion of heavy metal-contaminated soil minerals with strain Z-90 and the formation of a metal complex with biosurfactant. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Exploring the Anti-Burkholderia cepacia Complex Activity of Essential Oils: A Preliminary Analysis

    Directory of Open Access Journals (Sweden)

    Isabel Maida

    2014-01-01

    Full Text Available In this work we have checked the ability of the essential oils extracted from six different medicinal plants (Eugenia caryophyllata, Origanum vulgare, Rosmarinus officinalis, Lavandula officinalis, Melaleuca alternifolia, and Thymus vulgaris to inhibit the growth of 18 bacterial type strains belonging to the 18 known species of the Burkholderia cepacia complex (Bcc. These bacteria are opportunistic human pathogens that can cause severe infection in immunocompromised patients, especially those affected by cystic fibrosis (CF, and are often resistant to multiple antibiotics. The analysis of the aromatograms produced by the six oils revealed that, in spite of their different chemical composition, all of them were able to contrast the growth of Bcc members. However, three of them (i.e., Eugenia caryophyllata, Origanum vulgare, and Thymus vulgaris were particularly active versus the Bcc strains, including those exhibiting a high degree or resistance to ciprofloxacin, one of the most used antibiotics to treat Bcc infections. These three oils are also active toward both environmental and clinical strains (isolated from CF patients, suggesting that they might be used in the future to fight B. cepacia complex infections.

  6. Secondary metabolites from Bacillus amyloliquefaciens isolated from soil can kill Burkholderia pseudomallei.

    Science.gov (United States)

    Boottanun, Patcharaporn; Potisap, Chotima; Hurdle, Julian G; Sermswan, Rasana W

    2017-12-01

    Bacillus species are Gram-positive bacteria found in abundance in nature and their secondary metabolites were found to possess various potential activities, notably antimicrobial. In this study, Bacillus amyloliquefaciens N2-4 and N3-8 were isolated from soil and their metabolites could kill Burkholderia pseudomallei, a Gram-negative pathogenic bacterium also found in soil in its endemic areas. Moreover, the metabolites were able to kill drug resistant isolates of B. pseudomallei and also inhibit other pathogenic bacteria such as Staphylococcus aureus, Escherichia coli and Acinetobacter baumannii but not the non-pathogenic Burkholderia thailandensis, which is closely related to B. pseudomallei. Since the antimicrobial activity of N3-8 was not partially decreased or abolished when treated with proteolytic enzymes or autoclaved, but N2-4 was, these two strains should have produced different compounds. The N3-8 metabolites with antimicrobial activity consisted of both protein and non-protein compounds. The inhibition spectrum of the precipitated proteins compared to the culture supernatant indicated a possible synergistic effect of the non-protein and peptide compounds of N3-8 isolates against other pathogens. When either N2-4 or N3-8 isolates was co-cultured with B. pseudomallei the numbers of the bacteria decreased by 5 log 10 within 72 h. Further purification and characterization of the metabolites is required for future use of the bacteria or their metabolites as biological controls of B. pseudomallei in the environment or for development as new drugs for problematic pathogenic bacteria.

  7. Characterization of the Burkholderia thailandensis SOS response by using whole-transcriptome shotgun sequencing.

    Science.gov (United States)

    Ulrich, Ricky L; Deshazer, David; Kenny, Tara A; Ulrich, Melanie P; Moravusova, Anna; Opperman, Timothy; Bavari, Sina; Bowlin, Terry L; Moir, Donald T; Panchal, Rekha G

    2013-10-01

    The bacterial SOS response is a well-characterized regulatory network encoded by most prokaryotic bacterial species and is involved in DNA repair. In addition to nucleic acid repair, the SOS response is involved in pathogenicity, stress-induced mutagenesis, and the emergence and dissemination of antibiotic resistance. Using high-throughput sequencing technology (SOLiD RNA-Seq), we analyzed the Burkholderia thailandensis global SOS response to the fluoroquinolone antibiotic, ciprofloxacin (CIP), and the DNA-damaging chemical, mitomycin C (MMC). We demonstrate that a B. thailandensis recA mutant (RU0643) is ∼4-fold more sensitive to CIP in contrast to the parental strain B. thailandensis DW503. Our RNA-Seq results show that CIP and MMC treatment (P SOS response were induced and include lexA, uvrA, dnaE, dinB, recX, and recA. At the genome-wide level, we found an overall decrease in gene expression, especially for genes involved in amino acid and carbohydrate transport and metabolism, following both CIP and MMC exposure. Interestingly, we observed the upregulation of several genes involved in bacterial motility and enhanced transcription of a B. thailandensis genomic island encoding a Siphoviridae bacteriophage designated E264. Using B. thailandensis plaque assays and PCR with B. mallei ATCC 23344 as the host, we demonstrate that CIP and MMC exposure in B. thailandensis DW503 induces the transcription and translation of viable bacteriophage in a RecA-dependent manner. This is the first report of the SOS response in Burkholderia spp. to DNA-damaging agents. We have identified both common and unique adaptive responses of B. thailandensis to chemical stress and DNA damage.

  8. Fluorescence and NMR spectroscopy together with molecular simulations reveal amphiphilic characteristics of a Burkholderia biofilm exopolysaccharide.

    Science.gov (United States)

    Kuttel, Michelle M; Cescutti, Paola; Distefano, Marco; Rizzo, Roberto

    2017-06-30

    Biofilms are a collective mode of bacterial life in which a self-produced matrix confines cells in close proximity to each other. Biofilms confer many advantages, including protection from chemicals (including antibiotics), entrapment of useful extracellular enzymes and nutrients, as well as opportunities for efficient recycling of molecules from dead cells. Biofilm matrices are aqueous gel-like structures composed of polysaccharides, proteins, and DNA stabilized by intermolecular interactions that may include non-polar connections. Recently, polysaccharides extracted from biofilms produced by species of the Burkholderia cepacia complex were shown to possess clusters of rhamnose, a 6-deoxy sugar with non-polar characteristics. Molecular dynamics simulations are well suited to characterizing the structure and dynamics of polysaccharides, but only relatively few such studies exist of their interaction with non-polar molecules. Here we report an investigation into the hydrophobic properties of the exopolysaccharide produced by Burkholderia multivorans strain C1576. Fluorescence experiments with two hydrophobic fluorescent probes established that this polysaccharide complexes hydrophobic species, and NMR experiments confirmed these interactions. Molecular simulations to model the hydrodynamics of the polysaccharide and the interaction with guest species revealed a very flexible, amphiphilic carbohydrate chain that has frequent dynamic interactions with apolar molecules; both hexane and a long-chain fatty acid belonging to the quorum-sensing system of B. multivorans were tested. A possible role of the non-polar domains of the exopolysaccharide in facilitating the diffusion of aliphatic species toward specific targets within the biofilm aqueous matrix is proposed. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. Molecular Signatures and Phylogenomic Analysis of the Genus Burkholderia: Proposal for Division of this Genus into the Emended Genus Burkholderia Containing Pathogenic Organisms and a New Genus Paraburkholderia gen. nov. Harboring Environmental Species

    Directory of Open Access Journals (Sweden)

    Aman eSawana

    2014-12-01

    Full Text Available The genus Burkholderia contains large number of diverse species which are not reliably distinguished by the available biochemical or molecular characteristics. We report here results of detailed phylogenetic and comparative genomic analyses of 45 sequenced species of the genus Burkholderia. In phylogenetic trees based upon concatenated sequences for 21 conserved proteins as well as 16S rRNA gene sequences, Burkholderia species grouped into two major clades. Within these main clades a number of smaller clades were also clearly distinguished. Our comparative analysis of protein sequences from Burkholderia spp. has identified 42 highly specific molecular markers in the form of conserved sequence indels (CSIs that are uniquely found in different clades of Burkholderia spp. Six of these CSIs are specific for a group of Burkholderia spp. (referred to as Clade I which contains all clinically relevant members of the genus as well as the phytopathogenic Burkholderia species. The second main clade (Clade II composed of the environmental Burkholderia species, is also distinguished by 2 of the identified CSIs. Additionally, our work has also identified 3 CSIs that are specific for the Burkholderia cepacia complex, 4 CSIs that are uniquely found in the Burkholderia pseudomallei group, 5 CSIs that are specific for the phytopathogenic Burkholderia spp. and 22 other CSI that distinguish two groups within Clade II. The described molecular markers provide highly specific means for the demarcation of different groups of Burkholderia spp. and for development of novel diagnostic assays for the clinically important members of the group. Based upon the results from different lines of studies, a division of the genus Burkholderia into two genera is proposed. In this new proposal, the emended genus Burkholderia will contain only the clinically relevant and phytopathogenic Burkholderia species, whereas all other Burkholderia spp. are transferred to a new genus

  10. Diversidade de bactérias diazotróficas endofíticas dos gêneros Herbaspirillum e Burkholderia na cultura do arroz inundado Diversity of endophytic diazotrophic bacteria of the genus Herbaspirillum and Burkholderia in wetland rice

    Directory of Open Access Journals (Sweden)

    Luciana da Silva Rodrigues

    2006-02-01

    Full Text Available O objetivo deste trabalho foi avaliar a diversidade de bactérias diazotróficas endofíticas, dos gêneros Herbaspirillum e Burkholderia, em duas variedades de arroz, consideradas de alta (IR 42 e baixa (IAC 4440 eficiência de fixação biológica de nitrogênio. Foram realizados dois experimentos em casa de vegetação, em vasos com dois tipos de solos, provenientes dos Estados de Goiás e do Rio de Janeiro. Foi feita a contagem do número de bactérias e o isolamento em diferentes partes e estágios de desenvolvimento das plantas, mediante o uso de meios de cultivo JNFb e JMV. Os isolados bacterianos foram caracterizados a partir de aspectos morfológicos das colônias, com o crescimento em meios de cultivo, e de testes fisiológicos (uso de fontes de carbono e atividade de redução de acetileno. A contagem revelou grande número de bactérias diazotróficas (10(6 células g-1 matéria fresca, presentes em ambas as variedades de arroz, principalmente nas amostras radiculares. Os dados, obtidos na matriz de similaridade, mostram a presença de representantes da espécie Herbaspirillum seropedicae, bem como a diversidade entre isolados pertencentes ao gênero Burkholderia.The objective of this work was to evaluate the diversity of endophytic diazotrophic bacteria of the genera Herbaspirillum and Burkholderia, in two rice varieties, considered of high (IR 42 and low (IAC 4440 contribution on BNF. Two experiments were conducted in greenhouse conditions, in order to study the association of endophytic diazotrophic bacteria with wetland rice varieties, which were planted in two types of soil: one from Rio de Janeiro State and another from Goiás State, Brazil. Bacterial population (in different parts and physiological stages of the plants were evaluated, followed by the both genera strains isolation using culture media. The isolated bacteria were characterized based on morphological and physiological aspects. High bacterial counts were detected

  11. Stress conditions triggering mucoid morphotype variation in Burkholderia species and effect on virulence in Galleria mellonella and biofilm formation in vitro.

    Directory of Open Access Journals (Sweden)

    Inês N Silva

    Full Text Available Burkholderia cepacia complex (Bcc bacteria are opportunistic pathogens causing chronic respiratory infections particularly among cystic fibrosis patients. During these chronic infections, mucoid-to-nonmucoid morphotype variation occurs, with the two morphotypes exhibiting different phenotypic properties. Here we show that in vitro, the mucoid clinical isolate Burkholderia multivorans D2095 gives rise to stable nonmucoid variants in response to prolonged stationary phase, presence of antibiotics, and osmotic and oxidative stresses. Furthermore, in vitro colony morphotype variation within other members of the Burkholderia genus occurred in Bcc and non-Bcc strains, irrespectively of their clinical or environmental origin. Survival to starvation and iron limitation was comparable for the mucoid parental isolate and the respective nonmucoid variant, while susceptibility to antibiotics and to oxidative stress was increased in the nonmucoid variants. Acute infection of Galleria mellonella larvae showed that, in general, the nonmucoid variants were less virulent than the respective parental mucoid isolate, suggesting a role for the exopolysaccharide in virulence. In addition, most of the tested nonmucoid variants produced more biofilm biomass than their respective mucoid parental isolate. As biofilms are often associated with increased persistence of pathogens in the CF lungs and are an indicative of different cell-to-cell interactions, it is possible that the nonmucoid variants are better adapted to persist in this host environment.

  12. More than skin deep: moisturizing body milk and Burkholderia cepacia.

    Science.gov (United States)

    Irwin, Amy E; Price, Connie Savor

    2008-01-01

    Alvarez-Lerma and colleagues observed over an 18-day period that five critically ill patients admitted to a multidisciplinary 18-bed intensive care unit contracted Burkholderia cepacia from unopened containers of moisturizing body milk, calling into question the use in critical care settings of cosmetic products that do not guarantee sterilization during the manufacturing process. Is this the answer to the problem, however, or should the use of lotions in such settings be re-examined?

  13. More than skin deep: moisturizing body milk and Burkholderia cepacia

    OpenAIRE

    Irwin, Amy E; Price, Connie Savor

    2008-01-01

    Alvarez-Lerma and colleagues observed over an 18-day period that five critically ill patients admitted to a multidisciplinary 18-bed intensive care unit contracted Burkholderia cepacia from unopened containers of moisturizing body milk, calling into question the use in critical care settings of cosmetic products that do not guarantee sterilization during the manufacturing process. Is this the answer to the problem, however, or should the use of lotions in such settings be re-examined?

  14. Characterization of integrons in Burkholderia cepacia clinical isolates

    Directory of Open Access Journals (Sweden)

    Linda Furlanis

    2010-03-01

    Full Text Available Burkholderia cepacia is an opportunistic pathogen able to colonize the airways of Cystic Fibrosis (CF patients, frequently developing chronic infections. In 20% of cases these infections cause severe and poorly controlled pathological situations because of the intrinsic antibiotic resistance expressed by the microorganism. CF patients are often subjected to antibiotic therapy: this facilitates the acquisition of antibiotic resistance determinants by the infecting bacteria. Integrons are mobile genetic elements that are widespread in bacterial populations and favor the acquisition of gene cassettes coding for these determinants.The presence of class 1 integrons was investigated by PCR with primers specific for the 5’ and 3’ ends in Burkholderia isolates recovered from patients in treatment at the CF center of Friuli Venezia Giulia. The same integron, carrying an uncommon allelic form (Ib of the aacA4 gene in its cassette array and conferring resistance to some aminoglycosides, was found in two independent isolates (different RAPD profiles infecting two different patients. In both isolates the integron was carried by plasmids and was still present 3 and 6 years later the first finding. Despite the exchange of integrons between bacterial pathogens is fully described, these items were not frequently found in Burkholderia isolates. Although the clinical relevance of the integron we identified is low (a single gene cassette encoding a widespread resistance,we feel concerned that these genetic elements begin to circulate in this bacterial species, as this could make more and more troublesome the treatment of infections notoriously difficult to eradicate.

  15. Symbiotic Burkholderia Species Show Diverse Arrangements of nif/fix and nod Genes and Lack Typical High-Affinity Cytochrome cbb3 Oxidase Genes.

    Science.gov (United States)

    De Meyer, Sofie E; Briscoe, Leah; Martínez-Hidalgo, Pilar; Agapakis, Christina M; de-Los Santos, Paulina Estrada; Seshadri, Rekha; Reeve, Wayne; Weinstock, George; O'Hara, Graham; Howieson, John G; Hirsch, Ann M

    2016-08-01

    Genome analysis of fourteen mimosoid and four papilionoid beta-rhizobia together with fourteen reference alpha-rhizobia for both nodulation (nod) and nitrogen-fixing (nif/fix) genes has shown phylogenetic congruence between 16S rRNA/MLSA (combined 16S rRNA gene sequencing and multilocus sequence analysis) and nif/fix genes, indicating a free-living diazotrophic ancestry of the beta-rhizobia. However, deeper genomic analysis revealed a complex symbiosis acquisition history in the beta-rhizobia that clearly separates the mimosoid and papilionoid nodulating groups. Mimosoid-nodulating beta-rhizobia have nod genes tightly clustered in the nodBCIJHASU operon, whereas papilionoid-nodulating Burkholderia have nodUSDABC and nodIJ genes, although their arrangement is not canonical because the nod genes are subdivided by the insertion of nif and other genes. Furthermore, the papilionoid Burkholderia spp. contain duplications of several nod and nif genes. The Burkholderia nifHDKEN and fixABC genes are very closely related to those found in free-living diazotrophs. In contrast, nifA is highly divergent between both groups, but the papilionoid species nifA is more similar to alpha-rhizobia nifA than to other groups. Surprisingly, for all Burkholderia, the fixNOQP and fixGHIS genes required for cbb3 cytochrome oxidase production and assembly are missing. In contrast, symbiotic Cupriavidus strains have fixNOQPGHIS genes, revealing a divergence in the evolution of two distinct electron transport chains required for nitrogen fixation within the beta-rhizobia.

  16. Insecticide-degrading Burkholderia symbionts of the stinkbug naturally occupy various environments of sugarcane fields in a Southeast island of Japan.

    Science.gov (United States)

    Tago, Kanako; Okubo, Takashi; Itoh, Hideomi; Kikuchi, Yoshitomo; Hori, Tomoyuki; Sato, Yuya; Nagayama, Atsushi; Hayashi, Kentaro; Ikeda, Seishi; Hayatsu, Masahito

    2015-01-01

    The stinkbug Cavelerius saccharivorus, which harbors Burkholderia species capable of degrading the organophosphorus insecticide, fenitrothion, has been identified on a Japanese island in farmers' sugarcane fields that have been exposed to fenitrothion. A clearer understanding of the ecology of the symbiotic fenitrothion degraders of Burkholderia species in a free-living environment is vital for advancing our knowledge on the establishment of degrader-stinkbug symbiosis. In the present study, we analyzed the composition and abundance of degraders in sugarcane fields on the island. Degraders were recovered from field samples without an enrichment culture procedure. Degrader densities in the furrow soil in fields varied due to differences in insecticide treatment histories. Over 99% of the 659 isolated degraders belonged to the genus Burkholderia. The strains related to the stinkbug symbiotic group predominated among the degraders, indicating a selection for this group in response to fenitrothion. Degraders were also isolated from sugarcane stems, leaves, and rhizosphere in fields that were continuously exposed to fenitrothion. Their density was lower in the plant sections than in the rhizosphere. A phylogenetic analysis of 16S rRNA gene sequences demonstrated that most of the degraders from the plants and rhizosphere clustered with the stinkbug symbiotic group, and some were identical to the midgut symbionts of C. saccharivorus collected from the same field. Our results confirmed that plants and the rhizosphere constituted environmental reservoirs for stinkbug symbiotic degraders. To the best of our knowledge, this is the first study to investigate the composition and abundance of the symbiotic fenitrothion degraders of Burkholderia species in farmers' fields.

  17. A putative lateral flagella of the cystic fibrosis pathogen Burkholderia dolosa regulates swimming motility and host cytokine production

    Science.gov (United States)

    Clark, Bradley S.; Weatherholt, Molly; Renaud, Diane; Scott, David; LiPuma, John J.; Priebe, Gregory; Gerard, Craig

    2018-01-01

    Burkholderia dolosa caused an outbreak in the cystic fibrosis clinic at Boston Children’s Hospital and was associated with high mortality in these patients. This species is part of a larger complex of opportunistic pathogens known as the Burkholderia cepacia complex (Bcc). Compared to other species in the Bcc, B. dolosa is highly transmissible; thus understanding its virulence mechanisms is important for preventing future outbreaks. The genome of one of the outbreak strains, AU0158, revealed a homolog of the lafA gene encoding a putative lateral flagellin, which, in other non-Bcc species, is used for movement on solid surfaces, attachment to host cells, or movement inside host cells. Here, we analyzed the conservation of the lafA gene and protein sequences, which are distinct from those of the polar flagella, and found lafA homologs to be present in numerous β-proteobacteria but notably absent from most other Bcc species. A lafA deletion mutant in B. dolosa showed a greater swimming motility than wild-type due to an increase in the number of polar flagella, but did not appear to contribute to biofilm formation, host cell invasion, or murine lung colonization or persistence over time. However, the lafA gene was important for cytokine production in human peripheral blood mononuclear cells, suggesting it may have a role in recognition by the human immune response. PMID:29346379

  18. Burkholderia bacteria infectiously induce the proto-farming symbiosis of Dictyostelium amoebae and food bacteria.

    Science.gov (United States)

    DiSalvo, Susanne; Haselkorn, Tamara S; Bashir, Usman; Jimenez, Daniela; Brock, Debra A; Queller, David C; Strassmann, Joan E

    2015-09-08

    Symbiotic associations can allow an organism to acquire novel traits by accessing the genetic repertoire of its partner. In the Dictyostelium discoideum farming symbiosis, certain amoebas (termed "farmers") stably associate with bacterial partners. Farmers can suffer a reproductive cost but also gain beneficial capabilities, such as carriage of bacterial food (proto-farming) and defense against competitors. Farming status previously has been attributed to amoeba genotype, but the role of bacterial partners in its induction has not been examined. Here, we explore the role of bacterial associates in the initiation, maintenance, and phenotypic effects of the farming symbiosis. We demonstrate that two clades of farmer-associated Burkholderia isolates colonize D. discoideum nonfarmers and infectiously endow them with farmer-like characteristics, indicating that Burkholderia symbionts are a major driver of the farming phenomenon. Under food-rich conditions, Burkholderia-colonized amoebas produce fewer spores than uncolonized counterparts, with the severity of this reduction being dependent on the Burkholderia colonizer. However, the induction of food carriage by Burkholderia colonization may be considered a conditionally adaptive trait because it can confer an advantage to the amoeba host when grown in food-limiting conditions. We observed Burkholderia inside and outside colonized D. discoideum spores after fruiting body formation; this observation, together with the ability of Burkholderia to colonize new amoebas, suggests a mixed mode of symbiont transmission. These results change our understanding of the D. discoideum farming symbiosis by establishing that the bacterial partner, Burkholderia, is an important causative agent of the farming phenomenon.

  19. 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.

  20. Cloning, expression and mutation of a triazophos hydrolase gene from Burkholderia sp. SZL-1.

    Science.gov (United States)

    Zhang, Hao; Li, Qiang; Guo, Su-Hui; Cheng, Ming-Gen; Zhao, Meng-Jun; Hong, Qing; Huang, Xing

    2016-06-01

    Triazophos is a broad-spectrum and highly effective insecticide, and the residues of triazophos have been frequently detected in the environment. A triazophos-degrading bacterium, Burkholderia sp. SZL-1, was isolated from a long-term triazophos-polluted soil. Strain SZL-1 could hydrolyze triazophos to 1-phenyl-3-hydroxy-1,2,4-triazole, which was further utilized as the carbon sources for growth. The triazophos hydrolase gene trhA, cloned from strain SZL-1, was expressed and homogenously purified using Ni-nitrilotriacetic acid affinity chromatography. TrhA is 55 kDa and displays maximum activity at 25°C, pH 8.0. This enzyme still has nearly 60% activity at the range of 15°C-50°C for 30 min. TrhA was mutated by sequential error prone PCR and screened for improved activity for triazophos degradation. One purified variant protein (Val89-Gly89) named TrhA-M1 showed up to 3-fold improvement in specific activity against triazophos, and the specificity constants of Kcat and Kcat/Km for TrhA-M1 were improved up to 2.3- and 8.28-fold, respectively, compared to the wild-type enzyme. The results in this paper provided potential material for the contaminated soil remediation and hydrolase genetic structure research. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  1. Mechanisms of Disease: Host-Pathogen Interactions between Burkholderia Species and Lung Epithelial Cells

    Science.gov (United States)

    David, Jonathan; Bell, Rachel E.; Clark, Graeme C.

    2015-01-01

    Members of the Burkholderia species can cause a range of severe, often fatal, respiratory diseases. A variety of in vitro models of infection have been developed in an attempt to elucidate the mechanism by which Burkholderia spp. gain entry to and interact with the body. The majority of studies have tended to focus on the interaction of bacteria with phagocytic cells with a paucity of information available with regard to the lung epithelium. However, the lung epithelium is becoming more widely recognized as an important player in innate immunity and the early response to infections. Here we review the complex relationship between Burkholderia species and epithelial cells with an emphasis on the most pathogenic species, Burkholderia pseudomallei and Burkholderia mallei. The current gaps in knowledge in our understanding are highlighted along with the epithelial host-pathogen interactions that offer potential opportunities for therapeutic intervention. PMID:26636042

  2. A conserved two-component regulatory system, PidS/PidR, globally regulates pigmentation and virulence-related phenotypes of Burkholderia glumae.

    Science.gov (United States)

    Karki, Hari Sharan; Barphagha, Inderjit Kaur; Ham, Jong Hyun

    2012-09-01

    Burkholderia glumae is a rice pathogenic bacterium that causes bacterial panicle blight. Some strains of this pathogen produce dark brown pigments when grown on casamino-acid peptone glucose (CPG) agar medium. A pigment-positive and highly virulent strain of B. glumae, 411gr-6, was randomly mutagenized with mini-Tn5gus, and the resulting mini-Tn5gus derivatives showing altered pigmentation phenotypes were screened on CPG agar plates to identify the genetic elements governing the pigmentation of B. glumae. In this study, a novel two-component regulatory system (TCRS) composed of the PidS sensor histidine kinase and the PidR response regulator was identified as an essential regulatory factor for pigmentation. Notably, the PidS/PidR TCRS was also required for the elicitation of the hypersensitive response on tobacco leaves, indicating the dependence of the hypersensitive response and pathogenicity (Hrp) type III secretion system of B. glumae on this regulatory factor. In addition, B. glumae mutants defective in the PidS/PidR TCRS showed less production of the phytotoxin, toxoflavin, and less virulence on rice panicles and onion bulbs relative to the parental strain, 411gr-6. The presence of highly homologous PidS and PidR orthologues in other Burkholderia species suggests that PidS/PidR-family TCRSs may exert the same or similar functions in different Burkholderia species, including both plant and animal pathogens. © 2012 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

  3. PhaR, a Negative Regulator of PhaP, Modulates the Colonization of a Burkholderia Gut Symbiont in the Midgut of the Host Insect, Riptortus pedestris.

    Science.gov (United States)

    Jang, Seong Han; Jang, Ho Am; Lee, Junbeom; Kim, Jong Uk; Lee, Seung Ah; Park, Kyoung-Eun; Kim, Byung Hyun; Jo, Yong Hun; Lee, Bok Luel

    2017-06-01

    Five genes encoding PhaP family proteins and one phaR gene have been identified in the genome of Burkholderia symbiont strain RPE75. PhaP proteins function as the surface proteins of polyhydroxyalkanoate (PHA) granules, and the PhaR protein acts as a negative regulator of PhaP biosynthesis. Recently, we characterized one phaP gene to understand the molecular cross talk between Riptortus insects and Burkholderia gut symbionts. In this study, we constructed four other phaP gene-depleted mutants (Δ phaP1 , Δ phaP2 , Δ phaP3 , and Δ phaP4 mutants), one phaR gene-depleted mutant, and a phaR -complemented mutant (Δ phaR/phaR mutant). To address the biological roles of four phaP family genes and the phaR gene during insect-gut symbiont interaction, these Burkholderia mutants were fed to the second-instar nymphs, and colonization ability and fitness parameters were examined. In vitro , the Δ phaP3 and Δ phaR mutants cannot make a PHA granule normally in a stressful environment. Furthermore, the Δ phaR mutation decreased the colonization ability in the host midgut and negatively affected the host insect's fitness compared with wild-type Burkholderia -infected insects. However, other phaP family gene-depleted mutants colonized well in the midgut of the fifth-instar nymph insects. However, in the case of females, the colonization rate of the Δ phaP3 mutant was decreased and the host's fitness parameters were decreased compared with the wild-type-infected host, suggesting that the environment of the female midgut may be more hostile than that of the male midgut. These results demonstrate that PhaR plays an important role in the biosynthesis of PHA granules and that it is significantly related to the colonization of the Burkholderia gut symbiont in the host insects' midgut. IMPORTANCE Bacterial polyhydroxyalkanoate (PHA) biosynthesis is a complex process requiring several enzymes. The biological roles of PHA granule synthesis enzymes and the surface proteins of PHA

  4. Molecular signatures and phylogenomic analysis of the genus Burkholderia: proposal for division of this genus into the emended genus Burkholderia containing pathogenic organisms and a new genus Paraburkholderia gen. nov. harboring environmental species.

    Science.gov (United States)

    Sawana, Amandeep; Adeolu, Mobolaji; Gupta, Radhey S

    2014-01-01

    The genus Burkholderia contains large number of diverse species which include many clinically important organisms, phytopathogens, as well as environmental species. However, currently, there is a paucity of biochemical or molecular characteristics which can reliably distinguish different groups of Burkholderia species. We report here the results of detailed phylogenetic and comparative genomic analyses of 45 sequenced species of the genus Burkholderia. In phylogenetic trees based upon concatenated sequences for 21 conserved proteins as well as 16S rRNA gene sequence based trees, members of the genus Burkholderia grouped into two major clades. Within these main clades a number of smaller clades including those corresponding to the clinically important Burkholderia cepacia complex (BCC) and the Burkholderia pseudomallei groups were also clearly distinguished. Our comparative analysis of protein sequences from Burkholderia spp. has identified 42 highly specific molecular markers in the form of conserved sequence indels (CSIs) that are uniquely found in a number of well-defined groups of Burkholderia spp. Six of these CSIs are specific for a group of Burkholderia spp. (referred to as Clade I in this work) which contains all clinically relevant members of the genus (viz. the BCC and the B. pseudomallei group) as well as the phytopathogenic Burkholderia spp. The second main clade (Clade II), which is composed of environmental Burkholderia species, is also distinguished by 2 identified CSIs that are specific for this group. Additionally, our work has also identified multiple CSIs that serve to clearly demarcate a number of smaller groups of Burkholderia spp. including 3 CSIs that are specific for the B. cepacia complex, 4 CSIs that are uniquely found in the B. pseudomallei group, 5 CSIs that are specific for the phytopathogenic Burkholderia spp. and 22 other CSI that distinguish two groups within Clade II. The described molecular markers provide highly specific means for

  5. Molecular signatures and phylogenomic analysis of the genus Burkholderia: proposal for division of this genus into the emended genus Burkholderia containing pathogenic organisms and a new genus Paraburkholderia gen. nov. harboring environmental species

    Science.gov (United States)

    Sawana, Amandeep; Adeolu, Mobolaji; Gupta, Radhey S.

    2014-01-01

    The genus Burkholderia contains large number of diverse species which include many clinically important organisms, phytopathogens, as well as environmental species. However, currently, there is a paucity of biochemical or molecular characteristics which can reliably distinguish different groups of Burkholderia species. We report here the results of detailed phylogenetic and comparative genomic analyses of 45 sequenced species of the genus Burkholderia. In phylogenetic trees based upon concatenated sequences for 21 conserved proteins as well as 16S rRNA gene sequence based trees, members of the genus Burkholderia grouped into two major clades. Within these main clades a number of smaller clades including those corresponding to the clinically important Burkholderia cepacia complex (BCC) and the Burkholderia pseudomallei groups were also clearly distinguished. Our comparative analysis of protein sequences from Burkholderia spp. has identified 42 highly specific molecular markers in the form of conserved sequence indels (CSIs) that are uniquely found in a number of well-defined groups of Burkholderia spp. Six of these CSIs are specific for a group of Burkholderia spp. (referred to as Clade I in this work) which contains all clinically relevant members of the genus (viz. the BCC and the B. pseudomallei group) as well as the phytopathogenic Burkholderia spp. The second main clade (Clade II), which is composed of environmental Burkholderia species, is also distinguished by 2 identified CSIs that are specific for this group. Additionally, our work has also identified multiple CSIs that serve to clearly demarcate a number of smaller groups of Burkholderia spp. including 3 CSIs that are specific for the B. cepacia complex, 4 CSIs that are uniquely found in the B. pseudomallei group, 5 CSIs that are specific for the phytopathogenic Burkholderia spp. and 22 other CSI that distinguish two groups within Clade II. The described molecular markers provide highly specific means for

  6. Purification and characterization of vanillin dehydrogenases from alkaliphile Micrococcus sp. TA1 and neutrophile Burkholderia cepacia TM1.

    Science.gov (United States)

    Mitsui, Ryoji; Hirota, Mizuho; Tsuno, Takuo; Tanaka, Mitsuo

    2010-02-01

    Vanillin dehydrogenases (VDHs) were purified and characterized from two bacterial strains that have different pH dependencies for growth. The alkaliphile Micrococcus sp. TA1, isolated from an alkaline spa, can grow on several aromatic compounds such as ferulic acid, vanillin, vanillic acid, and protocatechuic acid under alkaline conditions. The neutrophile Burkholderia cepacia TM1, which was isolated previously, also grew on the above-mentioned compounds because they functioned as the sole carbon source under neutral conditions. Purified VDHs showed activities toward some aromatic aldehydes. These enzymes have the same subunit molecular mass of about 57 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but differed in some of their observed properties. Native molecular masses also differed between the purified enzymes. These were 250 kDa for the enzyme from alkaliphilic strain TA1 and 110 kDa for that from neutrophilic strain TM1, as determined by gel filtration. The enzyme from strain TA1 required NADP(+) as a coenzyme for its activity, but that from strain TM1 required NAD(+). These results are important because this is the first report of an alkaliphilic bacterium consuming lignin monomers.

  7. Burkholderia glumae EN EL CULTIVO DE ARROZ EN COSTA RICA

    Directory of Open Access Journals (Sweden)

    Andrea Quesada-Gonz\\u00E1lez

    2014-01-01

    Full Text Available Burkholderia glumae en el cultivo de arroz en Costa Rica. El objetivo de este trabajo fue determinar la presencia de Burkholderia glumae en arroz en Costa Rica. La bacteria Burkholderia glumae está asociada al cultivo del arroz en el que provoca la enfermedad llamada añublo bacterial. Bajo condiciones ambientales favorables, la densidad bacteriana aumenta, lo que provoca que, bajo un sistema de regulación denominado quorum sensing, se expresen sus mecanismos de virulencia mediante la activación de genes responsables para la síntesis de la toxoflavina, que bloquea el flujo de nutrientes, para la biogénesis de flagelos y la respuesta quimiotáctica, y la producción de la enzima catalasa. Las plantas desarrollan la sintomatología que finalmente conlleva a un vaneamiento del grano provocando pérdidas económicas importantes. Se investigó la situación referente a la contaminación del grano de arroz causado por esta bacteria en Costa Rica durante los años 2009 y 2010, mediante un convenio entre la Corporación Nacional Arrocera y el Laboratorio de Fitopatología del Centro de Investigación en Protección de Cultivos de la Universidad de Costa Rica. Se usó la metodología de PCR de punto final recomendada por investigadores del Centro Internacional de Agricultura Tropical en Colombia y se reforzó la identificación, por medio de técnicas de microbiología convencional. Se obtuvieron resultados que indican la presencia de la bacteria en Costa Rica, la primera información sobre la prevalencia de un fitopatógeno bacteriano de gran importancia para el sector arrocero.

  8. Development of a Selective Medium for the Fungal Pathogen Fusarium graminearum Using Toxoflavin Produced by the Bacterial Pathogen Burkholderia glumae

    Directory of Open Access Journals (Sweden)

    Boknam Jung

    2013-12-01

    Full Text Available The ascomycete fungus Fusarium graminearum is a major causal agent for Fusarium head blight in cereals and produces mycotoxins such as trichothecenes and zearalenone. Isolation of the fungal strains from air or cereals can be hampered by various other airborne fungal pathogens and saprophytic fungi. In this study, we developed a selective medium specific to F. graminearum using toxoflavin produced by the bacterial pathogen Burkholderia glumae. F. graminearum was resistant to toxoflavin, while other fungi were sensitive to this toxin. Supplementing toxoflavin into medium enhanced the isolation of F. graminearum from rice grains by suppressing the growth of saprophytic fungal species. In addition, a medium with or without toxoflavin exposed to wheat fields for 1 h had 84% or 25%, respectively, of colonies identified as F. graminearum. This selection medium provides an efficient tool for isolating F. graminearum, and can be adopted by research groups working on genetics and disease forecasting.

  9. CpG oligodeoxyribonucleotides protect mice from Burkholderia pseudomallei but not Francisella tularensis Schu S4 aerosols.

    Science.gov (United States)

    Rozak, David A; Gelhaus, Herbert C; Smith, Mark; Zadeh, Mojgan; Huzella, Louis; Waag, David; Adamovicz, Jeffrey J

    2010-02-05

    Studies have shown that CpG oligodeoxyribonucleotides (ODN) protect mice from various bacterial pathogens, including Burkholderia pseudomallei and Francisella tularensis live vaccine strain (LVS), when administered before parenteral challenge. Given the potential to develop CpG ODN as a pre-treatment for multiple bacterial biological warfare agents, we examined survival, histopathology, and cytokine data from CpG ODN-treated C57BL/6 mice to determine whether previously-reported protection extended to aerosolized B. pseudomallei 1026b and highly virulent F. tularensis Schu S4 infections. We found that, although CpG ODN protected mice from aerosolized B. pseudomallei challenges, the immunostimulant failed to benefit the animals exposed to F. tularensis Schu S4 aerosols. Our results, which contrast with earlier F. tularensis LVS studies, highlight potential differences in Francisella species pathogenesis and underscore the need to evaluate immunotherapies against human pathogenic species.

  10. Cloning, purification, crystallization and preliminary X-ray analysis of the Burkholderia pseudomallei L1 ribosomal protein

    International Nuclear Information System (INIS)

    Abd Aziz, Abd Ghani; Ruzheinikov, Sergey N.; Sedelnikova, Svetlana E.; Mohamed, Rahmah; Nathan, Sheila; Baker, Patrick J.; Rice, David W.

    2012-01-01

    The L1 ribosomal protein from B. pseudomallei has been overexpressed, purified and crystallized in a form suitable for X-ray analysis. The gene encoding the L1 ribosomal protein from Burkholderia pseudomallei strain D286 has been cloned into the pETBLUE-1 vector system, overexpressed in Escherichia coli and purified. Crystals of the native protein were grown by the hanging-drop vapour-diffusion technique using PEG 3350 as a precipitant and diffracted to beyond 1.65 Å resolution. The crystals belonged to space group P2 1 2 1 2, with unit-cell parameters a = 53.6, b = 127.1, c = 31.8 Å and with a single molecule in the asymmetric unit

  11. Characterization of BcaA, a putative classical autotransporter protein in Burkholderia pseudomallei.

    Science.gov (United States)

    Campos, Cristine G; Borst, Luke; Cotter, Peggy A

    2013-04-01

    Burkholderia pseudomallei is a tier 1 select agent, and the causative agent of melioidosis, a disease with effects ranging from chronic abscesses to fulminant pneumonia and septic shock, which can be rapidly fatal. Autotransporters (ATs) are outer membrane proteins belonging to the type V secretion system family, and many have been shown to play crucial roles in pathogenesis. The open reading frame Bp1026b_II1054 (bcaA) in B. pseudomallei strain 1026b is predicted to encode a classical autotransporter protein with an approximately 80-kDa passenger domain that contains a subtilisin-related domain. Immediately 3' to bcaA is Bp11026_II1055 (bcaB), which encodes a putative prolyl 4-hydroxylase. To investigate the role of these genes in pathogenesis, large in-frame deletion mutations of bcaA and bcaB were constructed in strain Bp340, an efflux pump mutant derivative of the melioidosis clinical isolate 1026b. Comparison of Bp340ΔbcaA and Bp340ΔbcaB mutants to wild-type B. pseudomallei in vitro demonstrated similar levels of adherence to A549 lung epithelial cells, but the mutant strains were defective in their ability to invade these cells and to form plaques. In a BALB/c mouse model of intranasal infection, similar bacterial burdens were observed after 48 h in the lungs and liver of mice infected with Bp340ΔbcaA, Bp340ΔbcaB, and wild-type bacteria. However, significantly fewer bacteria were recovered from the spleen of Bp340ΔbcaA-infected mice, supporting the idea of a role for this AT in dissemination or in survival in the passage from the site of infection to the spleen.

  12. A case of native valve endocarditis caused by Burkholderia cepacia without predisposing factors

    Directory of Open Access Journals (Sweden)

    Han Seong

    2011-05-01

    Full Text Available Abstract Background Infective endocarditis is rarely caused by Burkholderia cepacia. This infection is known to occur particularly in immunocompromised hosts, intravenous heroin users, and in patients with prosthetic valve replacement. Most patients with Burkholderia cepacia endocarditis usually need surgical treatment in addition to antimicrobial treatment. Case Presentation Here, we report the case of a patient who developed Burkholderia cepacia-induced native valve endocarditis with consequent cerebral involvement without any predisposing factors; she was successfully treated by antimicrobial agents only. Conclusion In this report, we also present literature review of relevant cases.

  13. Recurrent urinary tract infection by burkholderia cepacia in a live related renal transplant recipient

    International Nuclear Information System (INIS)

    Zeshan, M.

    2012-01-01

    Burkholderia cepacia is high virulent organism usually causing lower respiratory tract infections especially in Cystic fibrosis (CF) patients and post lung transplant. Urinary tract infections with Burkholderia cepacia have been associated after bladder irrigation or use of contaminated hospital objects. Post renal transplant urinary tract infection (UTI) is the most common infectious complications. Recurrent urinary tract infection with Burkholderia cepacia is a rare finding. Complete anatomical evaluation is essential in case recurrent urinary tract infections (UTI) after renal transplant. Vesico-ureteric reflux (VUR) and neurogenic urinary bladder was found to be important risk factors. (author)

  14. Complete Genome sequence of Burkholderia phymatum STM815, a broad host range and efficient nitrogen-fixing symbiont of Mimosa species

    Energy Technology Data Exchange (ETDEWEB)

    Moulin, Lionel [UMR, France; Klonowska, Agnieszka [UMR, France; Caroline, Bournaud [UMR, France; Booth, Kristina [University of Massachusetts; Vriezen, Jan A.C. [University of Massachusetts; Melkonian, Remy [UMR, France; James, Euan [James Hutton Institute, Dundee, United Kingdom; Young, Peter W. [University of York, United Kingdom; Bena, Gilles [UMR, France; Hauser, Loren John [ORNL; Land, Miriam L [ORNL; Kyrpides, Nikos C [U.S. Department of Energy, Joint Genome Institute; Bruce, David [Los Alamos National Laboratory (LANL); Chain, Patrick S. G. [Lawrence Livermore National Laboratory (LLNL); Copeland, A [U.S. Department of Energy, Joint Genome Institute; Pitluck, Sam [U.S. Department of Energy, Joint Genome Institute; Woyke, Tanja [U.S. Department of Energy, Joint Genome Institute; Lizotte-Waniewski, Michelle [University of Massachusetts; Bristow, James [U.S. Department of Energy, Joint Genome Institute; Riley, Monica [Woods Hole Oceanographic Institution (WHOI), Woods Hole

    2014-01-01

    Burkholderia phymatum is a soil bacterium able to develop a nitrogen-fixing symbiosis with species of the legume genus Mimosa, and is frequently found associated specifically with Mimosa pudica. The type strain of the species, STM 815T, was isolated from a root nodule in French Guiana in 2000. The strain is an aerobic, motile, non-spore forming, Gram-negative rod, and is a highly competitive strain for nodulation compared to other Mimosa symbionts, as it also nodulates a broad range of other legume genera and species. The 8,676,562 bp genome is composed of two chromosomes (3,479,187 and 2,697,374 bp), a megaplasmid (1,904,893 bp) and a plasmid hosting the symbiotic functions (595,108 bp).

  15. [Screening and optimization of cholesterol conversion strain].

    Science.gov (United States)

    Fan, Dan; Xiong, Bingjian; Pang, Cuiping; Zhu, Xiangdong

    2014-10-04

    Bacterial strain SE-1 capable of transforming cholesterol was isolated from soil and characterized. The transformation products were identified. Fermentation conditions were optimized for conversion. Cholesterol was used as sole carbon source to isolate strain SE-1. Morphology, physiological and biochemical characteristics of strain SE-1 were studied. 16S rRNA gene was sequenced and subjected to phylogenetic analysis. Fermentation supernatants were extracted with chloroform, the transformation products were analyzed by silica gel thin layer chromatography and Sephadex LH20. Their structures were identified by 1H-NMR and 13C-NMR. Fermentation medium including carbon and nitrogen, methods of adding substrates and fermentation conditions for Strain SE-1 were optimized. Strain SE-1 was a Gram-negative bacterium, exhibiting the highest homologs to Burkholderia cepacia based on the physiological analysis. The sequence analysis of 16S rRNA gene of SE-1 strain and comparison with related Burkholderia show that SE-1 strain was very close to B. cepacia (Genbank No. U96927). The similarity was 99%. The result of silica gel thin layer chromatography shows that strain SE-1 transformed cholesterol to two products, 7beta-hydroxycholesterol and the minor product was 7-oxocholesterol. The optimum culture conditions were: molasses 5%, (NH4 )2SO4 0.3%, 4% of inoculation, pH 7.5 and 36 degrees C. Under the optimum culture condition, the conversion rate reached 34.4% when concentration of cholesterol-Tween 80 was 1 g/L. Cholesterol 7beta-hydroxylation conversion rate under optimal conditions was improved by 20.8%. Strain SE-1 isolated from soil is capable of converting cholesterol at lab-scale.

  16. A heterodimer comprised of two bovine lactoferrin antimicrobial peptides exhibits powerful bactericidal activity against Burkholderia pseudomallei.

    Science.gov (United States)

    Puknun, Aekkalak; Bolscher, Jan G M; Nazmi, Kamran; Veerman, Enno C I; Tungpradabkul, Sumalee; Wongratanacheewin, Surasakdi; Kanthawong, Sakawrat; Taweechaisupapong, Suwimol

    2013-07-01

    Melioidosis is a severe infectious disease that is endemic in Southeast Asia and Northern Australia. Burkholderia pseudomallei, the causative agent of this disease, has developed resistance to an increasing list of antibiotics, demanding a search for novel agents. Lactoferricin and lactoferrampin are two antimicrobial domains of lactoferrin with a broad spectrum of antimicrobial activity. A hybrid peptide (LFchimera) containing lactoferrampin (LFampin265-284) and a part of lactoferricin (LFcin17-30) has strikingly higher antimicrobial activities compared to the individual peptides. In this study, the antimicrobial activities of this chimeric construct (LFchimera1), as well as of another one containing LFcin17-30 and LFampin268-284, a shorter fragment of LFampin265-284 (LFchimera2), and the constituent peptides were tested against 7 isolates of B. pseudomallei and compared to the preferential antibiotic ceftazidime (CAZ). All isolates including B. pseudomallei 979b shown to be resistant to CAZ, at a density of 10(5) CFU/ml, could be killed by 5-10 μM of LFchimera1 within 2 h, while the other peptides as well as the antibiotic CAZ only inhibited the B. pseudomallei strains resulting in an overgrowth in 24 h. These data indicate that LFchimera1 could be considered for development of therapeutic agents against B. pseudomallei.

  17. Structural, evolutionary and genetic analysis of the histidine biosynthetic "core" in the genus Burkholderia.

    Science.gov (United States)

    Papaleo, Maria Cristiana; Russo, Edda; Fondi, Marco; Emiliani, Giovanni; Frandi, Antonio; Brilli, Matteo; Pastorelli, Roberta; Fani, Renato

    2009-12-01

    In this work a detailed analysis of the structure, the expression and the organization of his genes belonging to the core of histidine biosynthesis (hisBHAF) in 40 newly determined and 13 available sequences of Burkholderia strains was carried out. Data obtained revealed a strong conservation of the structure and organization of these genes through the entire genus. The phylogenetic analysis showed the monophyletic origin of this gene cluster and indicated that it did not undergo horizontal gene transfer events. The analysis of the intergenic regions, based on the substitution rate, entropy plot and bendability suggested the existence of a putative transcription promoter upstream of hisB, that was supported by the genetic analysis that showed that this cluster was able to complement Escherichia colihisA, hisB, and hisF mutations. Moreover, a preliminary transcriptional analysis and the analysis of microarray data revealed that the expression of the his core was constitutive. These findings are in agreement with the fact that the entire Burkholderiahis operon is heterogeneous, in that it contains "alien" genes apparently not involved in histidine biosynthesis. Besides, they also support the idea that the proteobacterial his operon was piece-wisely assembled, i.e. through accretion of smaller units containing only some of the genes (eventually together with their own promoters) involved in this biosynthetic route. The correlation existing between the structure, organization and regulation of his "core" genes and the function(s) they perform in cellular metabolism is discussed.

  18. Glutamate uptake is important for osmoregulation and survival in the rice pathogen Burkholderia glumae.

    Directory of Open Access Journals (Sweden)

    Yongsung Kang

    Full Text Available Bacteria exhibit an optimal growth rate in culture media with sufficient nutrients at an optimal temperature and pH. In addition, the concentration of solutes plays a critical role in bacterial growth and survival. Glutamate is known to be a major anionic solute involved in osmoregulation and the bacterial cell's response to changes in solute concentration. To determine how glutamate uptake is involved in osmoregulation in the rice bacterial pathogen Burkholderia glumae BGR1, we mutated the gltI gene encoding a periplasmic substrate binding protein of a glutamate transport system to abolish glutamate uptake, and monitored the growth of the gltI null mutant in Luria-Bertani medium. We found that the gltI null mutant showed a slower growth rate than the wild-type strain and experienced hyperosmotic stress resulting in water loss from the cytoplasm in stationary phase. When the incubation time was extended, the mutant population collapsed due to the hyperosmotic stress. The gltI null mutant exhibited loss of adaptability under both hypoosmotic and hyperosmotic stresses. The growth rate of the gltI null mutant was restored to the level of wild-type growth by exogenous addition of glycine betaine to the culture medium, indicating that glycine betaine is a compatible solute in B. glumae. These results indicate that glutamate uptake from the environment plays a key role in osmoregulation in B. glumae.

  19. Investigation into the susceptibility of Burkholderia cepacia complex isolates to photodynamic antimicrobial chemotherapy (PACT)

    Science.gov (United States)

    Cassidy, C. M.; Watters, A. L.; Donnelly, R. F.; Tunney, M. M.

    2009-06-01

    The main cause of morbidity and mortality in cystic fibrosis (CF) sufferers is progressive pulmonary damage caused by recurrent and often unremitting respiratory tract infection. Causative organisms include Pseudomonas aeruginosa and Haemophilus influenzae, but in recent years the Burkholderia cepacia complex has come to the fore. This group of highly drug-resistant Gram-negative bacteria are associated with a rapid decline in lung function and the often fatal cepacia syndrome, with treatment limited to patient segregation and marginally effective antibacterial regimens. Thus, development of an effective treatment is of the upmost importance. PACT, a non-target specific therapy, has proven successful in killing both Gram-positive and Gram-negative bacteria. In this study, planktonic cultures of six strains of the B. cepacia complex were irradiated (635 nm, 200 J cm-2,10 minutes irradiation) following 30 seconds incubation with methylene blue (MB) or meso-tetra (N-methyl-4-pyridyl) porphine tetra tosylate (TMP). Rates of kill of > 99 % were achieved with MB- and TMP-PACT. A MB concentration of 50 μg ml-1 and TMP concentration of 500 μg ml-1 were associated with highest percentage kills for each photosensitizer. PACT is an attractive option for treatment of B.cepacia complex infection. Further study, involving biofilm culture susceptibility, delivery of light to the target and in vivo testing will be necessary before it PACT becomes a viable treatment option for CF patients who are colonised or infected with B. cepacia complex.

  20. An outbreak of Burkholderia stabilis colonization in a nasal ward.

    Science.gov (United States)

    Wang, Lijun; Wang, Mei; Zhang, Junyi; Wu, Wei; Lu, Yuan; Fan, Yanyan

    2015-04-01

    The aim of this study was to describe an outbreak of Burkholderia stabilis colonization among patients in a nasal ward. Multilocus sequence typing (MLST) was used for the molecular typing of B. stabilis isolates. Microbiological records were reviewed to delineate the colonization outbreak period. One hundred seventy-one cultures of environment and equipment samples from the nasal ward were performed to trace the source of contamination. Infection control measures were taken in order to end the outbreak. All B. stabilis isolates were identified as a new MLST type, ST821. A total of 53 patients carried this B. stabilis in the nasal ward between March and September 2013, which was defined as the outbreak period. The source of the colonization was not determined because all environment cultures were negative for Burkholderia cepacia complex. No further B. stabilis carriers have been found in the ward since the implementation of interventions. Attention must be paid to asymptomatic colonization in order to identify outbreaks early. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  1. PCR detection of Burkholderia multivorans in water and soil samples.

    Science.gov (United States)

    Peeters, Charlotte; Daenekindt, Stijn; Vandamme, Peter

    2016-08-12

    Although semi-selective growth media have been developed for the isolation of Burkholderia cepacia complex bacteria from the environment, thus far Burkholderia multivorans has rarely been isolated from such samples. Because environmental B. multivorans isolates mainly originate from water samples, we hypothesized that water rather than soil is its most likely environmental niche. The aim of the present study was to assess the occurrence of B. multivorans in water samples from Flanders (Belgium) using a fast, culture-independent PCR assay. A nested PCR approach was used to achieve high sensitivity, and specificity was confirmed by sequencing the resulting amplicons. B. multivorans was detected in 11 % of the water samples (n = 112) and 92 % of the soil samples (n = 25) tested. The percentage of false positives was higher for water samples compared to soil samples, showing that the presently available B. multivorans recA primers lack specificity when applied to the analysis of water samples. The results of the present study demonstrate that B. multivorans DNA is commonly present in soil samples and to a lesser extent in water samples in Flanders (Belgium).

  2. Type VI Secretion is a Major Virulence Determinant in Burkholderia Mallei

    National Research Council Canada - National Science Library

    Schell, Mark A; Ulrich, Ricky L; Ribot, Wilson J; Brueggemann, Ernst E; Hines, Harry B; Chen, Dan; Lipscomb, Lyla; Kim, H. S; Mrazek, Jan; Nierman, William C; DeShazer, David

    2007-01-01

    Burkholderia mallei is a host-adapted pathogen and a category B biothreat agent. Although the B. mallei VirAG two-component regulatory system is required for virulence in hamsters, the virulence genes it regulates are unknown...

  3. AQUIFER PROTIST RESPONSE AND THE POTENTIAL FOR TCE BIOREMEDIATION WITH BURKHOLDERIA CEPACIA G4 PR1

    Science.gov (United States)

    The introduction of bacteria into the environment for bioremediation purposes (bioaugmentation) requires analysis and monitoring of the persistence and activity of microbial population for efficacy and risk assessment purposes. Burkholderia cepacia G4 PR123 and PR131 constitutive...

  4. Changes in agricultural management drive the diversity of Burkholderia species isolated from soil on PLAT medium

    NARCIS (Netherlands)

    Salles, JF; Samyn, E; Vandamme, P; van Veen, JA; van Elsas, JD

    In order to assess the diversity of culturable Burkholderia populations in rhizosphere and bulk soil and to evaluate how different agricultural management regimes and land use history affect this diversity, four treatments were evaluated: permanent grassland; grassland converted into maize

  5. Changes in agricultural management drive the diversity of Burkholderia species isolated from soil on PCAT medium

    NARCIS (Netherlands)

    Salles, J.F.; Samyn, E.; Vandamme, P.A.; Veen, van J.A.; Elsas, van J.D.

    2006-01-01

    In order to assess the diversity of culturable Burkholderia populations in rhizosphere and bulk soil and to evaluate how different agricultural management regimes and land use history affect this diversity, four treatments were evaluated: permanent grassland; grassland converted into maize

  6. Interim report on updated microarray probes for the LLNL Burkholderia pseudomallei SNP array

    Energy Technology Data Exchange (ETDEWEB)

    Gardner, S; Jaing, C

    2012-03-27

    The overall goal of this project is to forensically characterize 100 unknown Burkholderia isolates in the US-Australia collaboration. We will identify genome-wide single nucleotide polymorphisms (SNPs) from B. pseudomallei and near neighbor species including B. mallei, B. thailandensis and B. oklahomensis. We will design microarray probes to detect these SNP markers and analyze 100 Burkholderia genomic DNAs extracted from environmental, clinical and near neighbor isolates from Australian collaborators on the Burkholderia SNP microarray. We will analyze the microarray genotyping results to characterize the genetic diversity of these new isolates and triage the samples for whole genome sequencing. In this interim report, we described the SNP analysis and the microarray probe design for the Burkholderia SNP microarray.

  7. Potential of Burkholderia seminalis TC3.4.2R3 as Biocontrol Agent Against Fusarium oxysporum Evaluated by Mass Spectrometry Imaging

    Science.gov (United States)

    Araújo, Francisca Diana da Silva; Araújo, Welington Luiz; Eberlin, Marcos Nogueira

    2017-05-01

    Species of genus Burkholderia display different interaction profiles in the environment, causing either several diseases in plants and animals or being beneficial to some plants, promoting their growth, and suppressing phytopathogens. Burkholderia spp. also produce many types of biomolecules with antimicrobial activity, which may be commercially used to protect crops of economic interest, mainly against fungal diseases. Herein we have applied matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to investigate secondary metabolites produced by B. seminalis TC3.4.2R3 in monoculture and coculture with plant pathogen Fusarium oxysporum. The siderophore pyochelin and the rhamnolipid Rha-Rha-C15-C14 were detected in wild-type B. seminalis strain, and their productions were found to vary in mutant strains carrying disruptions in gene clusters associated with antimicrobial compounds. Two mycotoxins were detected in F. oxysporum. During coculture with B. seminalis, metabolites probably related to defense mechanisms of these microorganisms were observed in the interspecies interaction zone. Our findings demonstrate the effective application of MALDI-MSI in the detection of bioactive molecules involved in the defense mechanism of B. seminalis, and these findings suggest the potential use of this bacterium in the biocontrol of plant diseases caused by F. oxysporum.

  8. Case-crossover study of Burkholderia cepacia complex bloodstream infection associated with contaminated intravenous bromopride.

    Science.gov (United States)

    Martins, Ianick Souto; Pellegrino, Flávia Lúcia Piffano Costa; Freitas, Andrea d'Avila; Santos, Marisa da Silva; Ferraiuoli, Giovanna Ianini d'Alemeida; Vasques, Márcia Regina Guimarães; Amorim, Efigenia Lourdes Teixeira; Oliveira, Sandra; Nouér, Simone Aranha; Cardoso, Fernando Luiz Lopes; Mascarenhas, Luiz Affonso; Magalhães, Ana Cristina Gouveia; Cleinman, Isabella Barbosa; Figueiredo, Agnes Marie Sá; Moreira, Beatriz Meurer

    2010-05-01

    To investigate an outbreak of healthcare-associated Burkholderia cepacia complex (BCC) primary bloodstream infections (BCC-BSI). Case-crossover study in a public hospital, a university hospital and a private hospital in Rio de Janeiro, Brazil, from March 2006 to May 2006. Twenty-five patients with BCC-BSI. After determining the date BCC-BSI symptoms started for each patient, 3 time intervals of data collection were defined, each one with a duration of 3 days: the case period, starting just before BCC-BSI symptoms onset; the control period, starting 6 days before BCC-BSI symptoms onset; and the washout period, comprising the 3 days between the case period and the control period. Exposures evaluated were intravascular solutions and invasive devices and procedures. Potential risk factors were identified by using the McNemar chi(2) adjusted test. Cultures of samples of potentially contaminated solutions were performed. BCC strain typing was performed by pulsed-field gel electrophoresis using SpeI. The statistical analysis revealed that the use of bromopride and dipyrone was associated with BCC-BSI. A total of 21 clinical isolates from 17 (68%) of the 25 patients and an isolate obtained from the bromopride vial were available for strain typing. Six pulsotypes were detected. A predominant pulsotype (A) accounted for 11 isolates obtained from 11 patients (65%) in the 3 study hospitals. Our investigation, using a case-crossover design, of an outbreak of BCC-BSI infections concluded it was polyclonal but likely caused by infusion of contaminated bromopride. The epidemiological finding was validated by microbiological analysis. After recall of contaminated bromopride vials by the manufacturer, the outbreak was controlled.

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

    Directory of Open Access Journals (Sweden)

    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.

  10. The effect of environmental conditions on biofilm formation of Burkholderia pseudomallei clinical isolates.

    Directory of Open Access Journals (Sweden)

    Nur Siti K Ramli

    Full Text Available Burkholderia pseudomallei, a Gram-negative saprophytic bacterium, is the causative agent of the potentially fatal melioidosis disease in humans. In this study, environmental parameters including temperature, nutrient content, pH and the presence of glucose were shown to play a role in in vitro biofilm formation by 28 B. pseudomallei clinical isolates, including four isolates with large colony variants (LCVs and small colony variants (SCVs morphotypes. Enhanced biofilm formation was observed when the isolates were tested in LB medium, at 30 °C, at pH 7.2, and in the presence of as little as 2 mM glucose respectively. It was also shown that all SVCs displayed significantly greater capacity to form biofilms than the corresponding LCVs when cultured in LB at 37 °C. In addition, octanoyl-homoserine lactone (C(8-HSL, a quorum sensing molecule, was identified by mass spectrometry analysis in bacterial isolates referred to as LCV CTH, LCV VIT, SCV TOM, SCV CTH, 1 and 3, and the presence of other AHL's with higher masses; decanoyl-homoserine lactone (C(10-HSL and dodecanoyl-homoserine lactone (C(12-HSL were also found in all tested strain in this study. Last but not least, we had successfully acquired two Bacillus sp. soil isolates, termed KW and SA respectively, which possessed strong AHLs degradation activity. Biofilm formation of B. pseudomallei isolates was significantly decreased after treated with culture supernatants of KW and SA strains, demonstrating that AHLs may play a role in B. pseudomallei biofilm formation.

  11. Molecular mechanisms underlying the close association between soil Burkholderia and fungi

    Science.gov (United States)

    Stopnisek, Nejc; Zühlke, Daniela; Carlier, Aurélien; Barberán, Albert; Fierer, Noah; Becher, Dörte; Riedel, Katharina; Eberl, Leo; Weisskopf, Laure

    2016-01-01

    Bacterial species belonging to the genus Burkholderia have been repeatedly reported to be associated with fungi but the extent and specificity of these associations in soils remain undetermined. To assess whether associations between Burkholderia and fungi are widespread in soils, we performed a co-occurrence analysis in an intercontinental soil sample collection. This revealed that Burkholderia significantly co-occurred with a wide range of fungi. To analyse the molecular basis of the interaction, we selected two model fungi frequently co-occurring with Burkholderia, Alternaria alternata and Fusarium solani, and analysed the proteome changes caused by cultivation with either fungus in the widespread soil inhabitant B. glathei, whose genome we sequenced. Co-cultivation with both fungi led to very similar changes in the B. glathei proteome. Our results indicate that B. glathei significantly benefits from the interaction, which is exemplified by a lower abundance of several starvation factors that were highly expressed in pure culture. However, co-cultivation also gave rise to stress factors, as indicated by the increased expression of multidrug efflux pumps and proteins involved in oxidative stress response. Our data suggest that the ability of Burkholderia to establish a close association with fungi mainly lies in the capacities to utilize fungal-secreted metabolites and to overcome fungal defense mechanisms. This work indicates that beneficial interactions with fungi might contribute to the survival strategy of Burkholderia species in environments with sub-optimal conditions, including acidic soils. PMID:25989372

  12. Molecular mechanisms underlying the close association between soil Burkholderia and fungi.

    Science.gov (United States)

    Stopnisek, Nejc; Zühlke, Daniela; Carlier, Aurélien; Barberán, Albert; Fierer, Noah; Becher, Dörte; Riedel, Katharina; Eberl, Leo; Weisskopf, Laure

    2016-01-01

    Bacterial species belonging to the genus Burkholderia have been repeatedly reported to be associated with fungi but the extent and specificity of these associations in soils remain undetermined. To assess whether associations between Burkholderia and fungi are widespread in soils, we performed a co-occurrence analysis in an intercontinental soil sample collection. This revealed that Burkholderia significantly co-occurred with a wide range of fungi. To analyse the molecular basis of the interaction, we selected two model fungi frequently co-occurring with Burkholderia, Alternaria alternata and Fusarium solani, and analysed the proteome changes caused by cultivation with either fungus in the widespread soil inhabitant B. glathei, whose genome we sequenced. Co-cultivation with both fungi led to very similar changes in the B. glathei proteome. Our results indicate that B. glathei significantly benefits from the interaction, which is exemplified by a lower abundance of several starvation factors that were highly expressed in pure culture. However, co-cultivation also gave rise to stress factors, as indicated by the increased expression of multidrug efflux pumps and proteins involved in oxidative stress response. Our data suggest that the ability of Burkholderia to establish a close association with fungi mainly lies in the capacities to utilize fungal-secreted metabolites and to overcome fungal defense mechanisms. This work indicates that beneficial interactions with fungi might contribute to the survival strategy of Burkholderia species in environments with sub-optimal conditions, including acidic soils.

  13. Effects of the inoculation of Burkholderia vietnamensis and related endophytic diazotrophic bacteria on grain yield of rice.

    Science.gov (United States)

    Govindarajan, Munusamy; Balandreau, Jacques; Kwon, Soon-Wo; Weon, Hang-Yeon; Lakshminarasimhan, Cunthipuram

    2008-01-01

    During a survey of endophytic diazotrophic bacteria associated with different rice varieties in Tamilnadu, some "endophytes" were obtained. Thirteen bacterial isolates from surface-sterilized roots and shoots were obtained in pure culture, which produced indole acetic acid (IAA) and reduced acetylene to ethylene. Polymerase chain reaction (PCR) amplification confirmed the presence of nif-H gene in all the isolates. Morphological, biochemical, and molecular characteristics indicated that all of them belonged to the genus Burkholderia One of them, MGK3, was consistently more active in reducing acetylene, and 16S rDNA sequences of isolate MGK3 confirmed its identification as Burkholderia vietnamiensis. Colonization of rice root was confirmed by strain MGK3 marked with gusA gene. The inoculated roots showed a blue color, which was most intense at the points of lateral root emergence and at the root tip. Transverse sections of roots, 15 days after inoculation, revealed beta-glucuronidase (GUS) activity within many of the cortical intercellular spaces next to the stele and within the aerenchyma. Nitrogen fixation was quantified by using (15)N isotope dilution method with two different cultivars grown in pot and field experiments. Higher nitrogen fixation was observed in variety Ponni than in ADT-43, where nearly 42% (field) and 40% (pot) of the nitrogen was derived from the atmosphere (% Ndfa). Isolate MGK3 was used to inoculate rice seedlings in a comparison with four other diazotrophs, viz., Gluconacetobacter diazotrophicus LMG7603, Herbaspirillum seropedicae LMG6513, Azospirillum lipoferum 4B LMG4348, and B. vietnamiensis LMG10929. They were used to conduct two pot and four field inoculation experiments. MGK3 alone, and combined with other diazotrophs, performed best under both pot and field conditions: combined inoculation produced yield increases between 9.5 and 23.6%, while MGK3 alone increased yield by 5.6 to 12.16% over the uninoculated control treatment.

  14. Global and regional dissemination and evolution of Burkholderia pseudomallei

    Science.gov (United States)

    Chewapreecha, Claire; Holden, Matthew T. G.; Vehkala, Minna; Välimäki, Niko; Yang, Zhirong; Harris, Simon R; Mather, Alison E.; Tuanyok, Apichai; De Smet, Birgit; Le Hello, Simon; Bizet, Chantal; Mayo, Mark; Wuthiekanun, Vanaporn; Limmathurotsakul, Direk; Phetsouvanh, Rattanaphone; Spratt, Brian G; Corander, Jukka; Keim, Paul; Dougan, Gordon; Dance, David A. B.; Currie, Bart J; Parkhill, Julian; Peacock, Sharon J.

    2017-01-01

    The environmental bacterium Burkholderia pseudomallei causes an estimated 165,000 cases of human melioidosis per year worldwide, and is also classified as a biothreat agent. We used whole genome sequences of 469 B. pseudomallei isolates from 30 countries collected over 79 years to explore its geographic transmission. Our data point to Australia as an early reservoir, with transmission to Southeast Asia followed by onward transmission to South Asia, and East Asia. Repeated reintroduction was observed within the Malay Peninsula, and between countries bordered by the Mekong river. Our data support an African origin of the Central and South American isolates with introduction of B. pseudomallei into the Americas between 1650 and 1850, providing a temporal link with the slave trade. We also identified geographically distinct genes/variants in Australasian or Southeast Asian isolates alone, with virulence-associated genes being among those overrepresented. This provides a potential explanation for clinical manifestations of melioidosis that are geographically restricted. PMID:28112723

  15. A genetic programming approach for Burkholderia Pseudomallei diagnostic pattern discovery

    Science.gov (United States)

    Yang, Zheng Rong; Lertmemongkolchai, Ganjana; Tan, Gladys; Felgner, Philip L.; Titball, Richard

    2009-01-01

    Motivation: Finding diagnostic patterns for fighting diseases like Burkholderia pseudomallei using biomarkers involves two key issues. First, exhausting all subsets of testable biomarkers (antigens in this context) to find a best one is computationally infeasible. Therefore, a proper optimization approach like evolutionary computation should be investigated. Second, a properly selected function of the antigens as the diagnostic pattern which is commonly unknown is a key to the diagnostic accuracy and the diagnostic effectiveness in clinical use. Results: A conversion function is proposed to convert serum tests of antigens on patients to binary values based on which Boolean functions as the diagnostic patterns are developed. A genetic programming approach is designed for optimizing the diagnostic patterns in terms of their accuracy and effectiveness. During optimization, it is aimed to maximize the coverage (the rate of positive response to antigens) in the infected patients and minimize the coverage in the non-infected patients while maintaining the fewest number of testable antigens used in the Boolean functions as possible. The final coverage in the infected patients is 96.55% using 17 of 215 (7.4%) antigens with zero coverage in the non-infected patients. Among these 17 antigens, BPSL2697 is the most frequently selected one for the diagnosis of Burkholderia Pseudomallei. The approach has been evaluated using both the cross-validation and the Jack–knife simulation methods with the prediction accuracy as 93% and 92%, respectively. A novel approach is also proposed in this study to evaluate a model with binary data using ROC analysis. Contact: z.r.yang@ex.ac.uk PMID:19561021

  16. PKC-η-MARCKS Signaling Promotes Intracellular Survival of Unopsonized Burkholderia thailandensis.

    Science.gov (United States)

    Micheva-Viteva, Sofiya N; Shou, Yulin; Ganguly, Kumkum; Wu, Terry H; Hong-Geller, Elizabeth

    2017-01-01

    Pathogenic Burkholderia rely on host factors for efficient intracellular replication and are highly refractory to antibiotic treatment. To identify host genes that are required by Burkholderia spp. during infection, we performed a RNA interference (RNAi) screen of the human kinome and identified 35 host kinases that facilitated Burkholderia thailandensis intracellular survival in human monocytic THP-1 cells. We validated a selection of host kinases using imaging flow cytometry to assess efficiency of B. thailandensis survival in the host upon siRNA-mediated knockdown. We focused on the role of the novel protein kinase C isoform, PKC-η, in Burkholderia infection and characterized PKC-η/MARCKS signaling as a key event that promotes the survival of unopsonized B. thailandensis CDC2721121 within host cells. While infection of lung epithelial cells with unopsonized Gram-negative bacteria stimulated phosphorylation of Ser175/160 in the MARCKS effector domain, siRNA-mediated knockdown of PKC-η expression reduced the levels of phosphorylated MARCKS by >3-fold in response to infection with Bt CDC2721121. We compared the effect of the conventional PKC-α and novel PKC-η isoforms on the growth of B. thailandensis CDC2721121 within monocytic THP-1 cells and found that ≥75% knock-down of PRKCH transcript levels reduced intracellular bacterial load 100% more efficiently when compared to growth in cells siRNA-depleted of the classical PKC-α, suggesting that the PKC-η isoform can specifically mediate Burkholderia intracellular survival. Based on imaging studies of intracellular B. thailandensis , we found that PKC-η function stimulates phagocytic pathways that promote B. thailandensis escape into the cytoplasm leading to activation of autophagosome flux. Identification of host kinases that are targeted by Burkholderia during infection provides valuable molecular insights in understanding Burkholderia pathogenesis, and ultimately, in designing effective host

  17. Antimicrobial susceptibility pattern of clinical isolates of Burkholderia pseudomallei in Bangladesh.

    Science.gov (United States)

    Dutta, Subarna; Haq, Sabah; Hasan, Mohammad Rokibul; Haq, Jalaluddin Ashraful

    2017-07-20

    Melioidosis an infectious disease, caused by a Gram negative bacterium called Burkholderia pseudomallei, is endemic in Bangladesh. This organism is sensitive to limited number of antimicrobial agents and need prolonged treatment. There is no comprehensive data on the antimicrobial susceptibility profile of B. pseudomallei isolated in Bangladesh over last several years. The present study aimed to determine the antimicrobial susceptibility pattern of B. pseudomallei isolated in a tertiary care hospital of Dhaka city from 2009 to 2015. All B. pseudomallei isolated from melioidosis patients over a period of 7 years (2009-2015) in the Department of Microbiology of a 725-bed tertiary care referral hospital in Dhaka city, Bangladesh were included in the study. B. pseudomallei was identified by Gram stain, culture, specific biochemical tests, serology and PCR using specific primers constructed from 16s rRNA region of B. pseudomallei. Antimicrobial susceptibility to specific agents was determined by disk diffusion and minimum inhibitory concentration methods. A total of 20 isolates of B. pseudomallei which were isolated from patients coming from different geographic locations of Bangladesh were included in the study. All the isolates were uniformly sensitive (100%) to ceftazidime, imipenem, piperacillin-tazobactam, amoxicillin-clavulanic acid and tetracycline by both disk diffusion and MIC methods. Two strains were resistant to trimethoprim-sulfamethoxazole by disk diffusion method but were sensitive by MIC method. The MIC 50 and MIC 90 values of the above antimicrobial agents were almost similar. All the isolates were resistant to amikacin by both MIC and disk diffusion methods. The results of the study suggest that B. pseudomallei prevalent in Bangladesh were still susceptible to all recommended antimicrobial agents used for the treatment of melioidosis. However, regular monitoring is needed to detect any emergence of resistance and shifting of MIC 50 and MIC 90 values.

  18. Burkholderia pseudomallei isolates from Sarawak, Malaysian Borneo, are predominantly susceptible to aminoglycosides and macrolides.

    Science.gov (United States)

    Podin, Yuwana; Sarovich, Derek S; Price, Erin P; Kaestli, Mirjam; Mayo, Mark; Hii, KingChing; Ngian, Hieung; Wong, SeeChang; Wong, IngTien; Wong, JinShyan; Mohan, Anand; Ooi, MongHow; Fam, TemLom; Wong, Jack; Tuanyok, Apichai; Keim, Paul; Giffard, Philip M; Currie, Bart J

    2014-01-01

    Melioidosis is a potentially fatal disease caused by the saprophytic bacterium Burkholderia pseudomallei. Resistance to gentamicin is generally a hallmark of B. pseudomallei, and gentamicin is a selective agent in media used for diagnosis of melioidosis. In this study, we determined the prevalence and mechanism of gentamicin susceptibility found in B. pseudomallei isolates from Sarawak, Malaysian Borneo. We performed multilocus sequence typing and antibiotic susceptibility testing on 44 B. pseudomallei clinical isolates from melioidosis patients in Sarawak district hospitals. Whole-genome sequencing was used to identify the mechanism of gentamicin susceptibility. A novel allelic-specific PCR was designed to differentiate gentamicin-sensitive isolates from wild-type B. pseudomallei. A reversion assay was performed to confirm the involvement of this mechanism in gentamicin susceptibility. A substantial proportion (86%) of B. pseudomallei clinical isolates in Sarawak, Malaysian Borneo, were found to be susceptible to the aminoglycoside gentamicin, a rare occurrence in other regions where B. pseudomallei is endemic. Gentamicin sensitivity was restricted to genetically related strains belonging to sequence type 881 or its single-locus variant, sequence type 997. Whole-genome sequencing identified a novel nonsynonymous mutation within amrB, encoding an essential component of the AmrAB-OprA multidrug efflux pump. We confirmed the role of this mutation in conferring aminoglycoside and macrolide sensitivity by reversion of this mutation to the wild-type sequence. Our study demonstrates that alternative B. pseudomallei selective media without gentamicin are needed for accurate melioidosis laboratory diagnosis in Sarawak. This finding may also have implications for environmental sampling of other locations to test for B. pseudomallei endemicity.

  19. Extraction of lipase from Burkholderia cepacia by PEG/Phosphate ATPS and its biochemical characterization

    Directory of Open Access Journals (Sweden)

    Giovana da Silva Padilha

    2012-02-01

    Full Text Available This work aimed to study the partitioning of a lipase produced by Burkholderia cepacia in PEG/Phosphate aqueous two phase system (ATPS and its characterization. Lipase was produced by B. cepacia strains in a fermenter. Enzyme partitioning occurred at pH 6.0 and 8.0, using PEG 1500 and 6000 on two tie lines. Metal ions, pH and temperature effects on enzyme activity were evaluated. Five milliliter of 7.5% olive oil emulsion with 2.5% gumarabic in 0.1M sodium phosphate buffer at pH 8.0 and 37ºC were used for the activity determinations. Results showed that crude stratum from B. cepacia was partitioned by PEG1500/phosphate ATPS at pH 6.0 or 8.0 for, which the partitioning coefficients were 108-and 209-folds. Lipase presented optimal activity conditions at 37ºC and pH 8.0; it showed pH-stability for 4 h of incubation at different pH values at 37ºC. Metal ions such as Mn2+ , Co2+, I-and Ca2+ sustained enzymatic activities; however, it was inhibited by the presence of Fe2+, Hg2+ and Al3+ . Km and Vmax values were 0.258 U/mg and 43.90 g/L, respectively. A molecular weight of 33 kDa and an isoelectric point at pH 5.0 were determined by SDS-PAGE and IFS electrophoresis, respectively.

  20. Burkholderia phytofirmans PsJN reduces damages to freezing temperature in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Fan eSU

    2015-10-01

    Full Text Available Several plant growth-promoting rhizobacteria (PGPR are known to improve plant tolerance to multiple stresses, including low temperatures. However, mechanisms underlying this protection are still poorly understood. The aim of this study was to evaluate the role of the endophytic PGPR, Burkholderia phytofirmans strain PsJN (Bp PsJN, on Arabidopsis thaliana cold tolerance using photosynthesis parameters as physiological markers.Under standard conditions, our results indicated that Bp PsJN inoculation led to growth promotion of Arabidopsis plants without significant modification on photosynthesis parameters and chloroplast organization. However, bacterial colonization induced a cell wall strengthening in the mesophyllImpact of inoculation modes (either on seeds or by soil irrigation and their effects overnight at 0, -1 or -3°C, were investigated by following photosystem II (PSII activity and gas exchanges. Following low temperatures stress, a decrease of photosynthesis parameters was observed. In addition, during three consecutive nights or days at -1°C, PSII activity was monitored. Pigment contents, RuBisCO protein abundance, expression of several genes including RbcS, RbcL, CBF1, CBF2, CBF3, ICE1, COR15a, and COR78 were evaluated at the end of exposure. To assess the impact of the bacteria on cell ultrastructure under low temperatures, microscopic observations were achieved. Results indicated that freezing treatment induced significant changes in PSII activity as early as the first cold day, whereas the same impact on PSII activity was observed only during the third cold night. The significant effects conferred by PsJN were differential accumulation of pigments, and reduced expression of RbcL and COR78. Microscopical observations showed an alteration/disorganization in A. thaliana leaf mesophyll cells independently of the freezing treatments. The presence of bacteria during the three successive nights or days did not significantly improved A

  1. The Organization of the Quorum Sensing luxI/R Family Genes in Burkholderia

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    Sándor Pongor

    2013-07-01

    Full Text Available Members of the Burkholderia genus of Proteobacteria are capable of living freely in the environment and can also colonize human, animal and plant hosts. Certain members are considered to be clinically important from both medical and veterinary perspectives and furthermore may be important modulators of the rhizosphere. Quorum sensing via N-acyl homoserine lactone signals (AHL QS is present in almost all Burkholderia species and is thought to play important roles in lifestyle changes such as colonization and niche invasion. Here we present a census of AHL QS genes retrieved from public databases and indicate that the local arrangement (topology of QS genes, their location within chromosomes and their gene neighborhoods show characteristic patterns that differ between the known Burkholderia clades. In sequence phylogenies, AHL QS genes seem to cluster according to the local gene topology rather than according to the species, which suggests that the basic topology types were present prior to the appearance of current Burkholderia species. The data are available at http://net.icgeb.org/burkholderia/.

  2. Burkholderia sp. KCTC 11096BP modulates pepper growth and resistance against Phytophthora capsici

    International Nuclear Information System (INIS)

    Kang, S.M.; Hamayun, M.; Shinwari, Z.K.

    2016-01-01

    Biological control of crop diseases is desirable for sustainable agriculture as it minimizes chemical inputs in the agricultural system and promotes eco-friendly environment. We analyzed the favorable role of Burkholderia sp. KCTC 11096BP against the pathogen Phytophthora capsici in pepper. We screen thirty rhizobateria for their anti-pathogen activity, and found that Burkholderia sp. KCTC 11096BP exhibits maximum growth inhibition of the pathogen P. capsici. The bacterium inoculation to pepper plants significantly enhanced growth attributes of pepper in infected and control treatments. The total proteins (10.9%), and the amino acids viz. glycine (4.08 ug/g), leucine (3.3 ug/g), and alanine (3.26 ug/g) were preset in considerably higher quantities in Burkholderia sp. applied treatments as compare to control. The systemic acquired resistance (SAR) of the host plant was up-regulated by Burkholderia sp. KCTC, as endogenous salicylic acid (235.5 ng/g) and jasmonic acid (22.8 ng/g) levels were found higher in such treatments. It was concluded that Burkholderia sp. KCTC 11096BP mitigates the adverse effects of P. capsici on pepper crop and can improve crop productivity at the field level. (author)

  3. A Possible Link between Infection with Burkholderia Bacteria and Systemic Lupus Erythematosus Based on Epitope Mimicry

    Directory of Open Access Journals (Sweden)

    Wei Zhang

    2008-01-01

    Full Text Available We previously demonstrated that purified polyclonal and monoclonal anti-dsDNA antibodies bind a 15-mer peptide ASPVTARVLWKASHV in ELISA and Dot blot. This 15-mer peptide partial sequence ARVLWKASH shares similarity with burkholderia bacterial cytochrome B 561 partial sequence ARVLWRATH. In this study, we show that purified anti-dsDNA antibodies react with burkholderia fungorum bacterial cell lysates in Western blot. We used anti-dsDNA antibodies to make an anti-dsDNA antibodies affinity column and used this column to purify the burkholderia fungorum bacterial protein. Purified anti-dsDNA antibodies bind specifically to purified bacterial antigen and purified bacterial antigen blocked the anti-dsDNA antibodies binding to dsDNA antigen. Sera with anti-dsDNA antibodies bind specifically to purified bacterial antigen. We obtained protein partial sequence of RAGTDEGFG which is shared with burkholderia bacterial transcription regulator protein sequence. Sera with anti-dsDNA antibodies bind to RAGTDEGFG peptide better than control groups. These data support our hypothesis that the origin of anti-dsDNA antibodies in SLE may be associated with burkholderia bacterial infection.

  4. The Organization of the Quorum Sensing luxI/R Family Genes in Burkholderia

    Science.gov (United States)

    Choudhary, Kumari Sonal; Hudaiberdiev, Sanjarbek; Gelencsér, Zsolt; Coutinho, Bruna Gonçalves; Venturi, Vittorio; Pongor, Sándor

    2013-01-01

    Members of the Burkholderia genus of Proteobacteria are capable of living freely in the environment and can also colonize human, animal and plant hosts. Certain members are considered to be clinically important from both medical and veterinary perspectives and furthermore may be important modulators of the rhizosphere. Quorum sensing via N-acyl homoserine lactone signals (AHL QS) is present in almost all Burkholderia species and is thought to play important roles in lifestyle changes such as colonization and niche invasion. Here we present a census of AHL QS genes retrieved from public databases and indicate that the local arrangement (topology) of QS genes, their location within chromosomes and their gene neighborhoods show characteristic patterns that differ between the known Burkholderia clades. In sequence phylogenies, AHL QS genes seem to cluster according to the local gene topology rather than according to the species, which suggests that the basic topology types were present prior to the appearance of current Burkholderia species. The data are available at http://net.icgeb.org/burkholderia/. PMID:23820583

  5. Divergent homologs of the predicted small RNA BpCand697 in Burkholderia spp.

    Science.gov (United States)

    Damiri, Nadzirah; Mohd-Padil, Hirzahida; Firdaus-Raih, Mohd

    2015-09-01

    The small RNA (sRNA) gene candidate, BpCand697 was previously reported to be unique to Burkholderia spp. and is encoded at 3' non-coding region of a putative AraC family transcription regulator gene. This study demonstrates the conservation of BpCand697 sequence across 32 Burkholderia spp. including B. pseudomallei, B. mallei, B. thailandensis and Burkholderia sp. by integrating both sequence homology and secondary structural analyses of BpCand697 within the dataset. The divergent sequence of BpCand697 was also used as a discriminatory power in clustering the dataset according to the potential virulence of Burkholderia spp., showing that B. thailandensis was clearly secluded from the virulent cluster of B. pseudomallei and B. mallei. Finally, the differential co-transcript expression of BpCand697 and its flanking gene, bpsl2391 was detected in Burkholderia pseudomallei D286 after grown under two different culture conditions using nutrient-rich and minimal media. It is hypothesized that the differential expression of BpCand697-bpsl2391 co-transcript between the two standard prepared media might correlate with nutrient availability in the culture media, suggesting that the physical co-localization of BpCand697 in B. pseudomallei D286 might be directly or indirectly involved with the transcript regulation of bpsl2391 under the selected in vitro culture conditions.

  6. Burkholderia of Plant-Beneficial Group are Symbiotically Associated with Bordered Plant Bugs (Heteroptera: Pyrrhocoroidea: Largidae).

    Science.gov (United States)

    Takeshita, Kazutaka; Matsuura, Yu; Itoh, Hideomi; Navarro, Ronald; Hori, Tomoyuki; Sone, Teruo; Kamagata, Yoichi; Mergaert, Peter; Kikuchi, Yoshitomo

    2015-01-01

    A number of phytophagous stinkbugs (order Heteroptera: infraorder Pentatomomorpha) harbor symbiotic bacteria in a specific midgut region composed of numerous crypts. Among the five superfamilies of the infraorder Pentatomomorpha, most members of the Coreoidea and Lygaeoidea are associated with a specific group of the genus Burkholderia, called the "stinkbug-associated beneficial and environmental (SBE)" group, which is not vertically transmitted, but acquired from the environment every host generation. A recent study reported that, in addition to these two stinkbug groups, the family Largidae of the superfamily Pyrrhocoroidea also possesses a Burkholderia symbiont. Despite this recent finding, the phylogenetic position and biological nature of Burkholderia associated with Largidae remains unclear. Based on the combined results of fluorescence in situ hybridization, cloning analysis, Illumina deep sequencing, and egg inspections by diagnostic PCR, we herein demonstrate that the largid species are consistently associated with the "plant-associated beneficial and environmental (PBE)" group of Burkholderia, which are phylogenetically distinct from the SBE group, and that they maintain symbiosis through the environmental acquisition of the bacteria. Since the superfamilies Coreoidea, Lygaeoidea, and Pyrrhocoroidea are monophyletic in the infraorder Pentatomomorpha, it is plausible that the symbiotic association with Burkholderia evolved at the common ancestor of the three superfamilies. However, the results of this study strongly suggest that a dynamic transition from the PBE to SBE group, or vice versa, occurred in the course of stinkbug evolution.

  7. A Burkholderia pseudomallei colony variant necessary for gastric colonization.

    Science.gov (United States)

    Austin, C R; Goodyear, A W; Bartek, I L; Stewart, A; Sutherland, M D; Silva, E B; Zweifel, A; Vitko, N P; Tuanyok, A; Highnam, G; Mittelman, D; Keim, P; Schweizer, H P; Vázquez-Torres, A; Dow, S W C; Voskuil, M I

    2015-02-03

    Diverse colony morphologies are a hallmark of Burkholderia pseudomallei recovered from infected patients. We observed that stresses that inhibit aerobic respiration shifted populations of B. pseudomallei from the canonical white colony morphotype toward two distinct, reversible, yet relatively stable yellow colony variants (YA and YB). As accumulating evidence supports the importance of B. pseudomallei enteric infection and gastric colonization, we tested the response of yellow variants to hypoxia, acidity, and stomach colonization. Yellow variants exhibited a competitive advantage under hypoxic and acidic conditions and alkalized culture media. The YB variant, although highly attenuated in acute virulence, was the only form capable of colonization and persistence in the murine stomach. The accumulation of extracellular DNA (eDNA) was a characteristic of YB as observed by 4',6-diamidino-2-phenylindole (DAPI) staining of gastric tissues, as well as in an in vitro stomach model where large amounts of eDNA were produced without cell lysis. Transposon mutagenesis identified a transcriptional regulator (BPSL1887, designated YelR) that when overexpressed produced the yellow phenotype. Deletion of yelR blocked a shift from white to the yellow forms. These data demonstrate that YB is a unique B. pseudomallei pathovariant controlled by YelR that is specifically adapted to the harsh gastric environment and necessary for persistent stomach colonization. Seemingly uniform populations of bacteria often contain subpopulations that are genetically identical but display unique characteristics which offer advantages when the population is faced with infrequent but predictable stresses. The pathogen Burkholderia pseudomallei is capable of forming several reversible colony types, and it interconverted between one white type and two yellow types under certain environmental stresses. The two yellow forms exhibited distinct advantages in low-oxygen and acidic environments. One yellow

  8. 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

  9. Transcriptomic profiling of Burkholderia phymatum STM815, Cupriavidus taiwanensis LMG19424 and Rhizobium mesoamericanum STM3625 in response to Mimosa pudica root exudates illuminates the molecular basis of their nodulation competitiveness and symbiotic evolutionary history.

    Science.gov (United States)

    Klonowska, Agnieszka; Melkonian, Rémy; Miché, Lucie; Tisseyre, Pierre; Moulin, Lionel

    2018-01-30

    Rhizobial symbionts belong to the classes Alphaproteobacteria and Betaproteobacteria (called "alpha" and "beta"-rhizobia). Most knowledge on the genetic basis of symbiosis is based on model strains belonging to alpha-rhizobia. Mimosa pudica is a legume that offers an excellent opportunity to study the adaptation toward symbiotic nitrogen fixation in beta-rhizobia compared to alpha-rhizobia. In a previous study (Melkonian et al., Environ Microbiol 16:2099-111, 2014) we described the symbiotic competitiveness of M. pudica symbionts belonging to Burkholderia, Cupriavidus and Rhizobium species. In this article we present a comparative analysis of the transcriptomes (by RNAseq) of B. phymatum STM815 (BP), C. taiwanensis LMG19424 (CT) and R. mesoamericanum STM3625 (RM) in conditions mimicking the early steps of symbiosis (i.e. perception of root exudates). BP exhibited the strongest transcriptome shift both quantitatively and qualitatively, which mirrors its high competitiveness in the early steps of symbiosis and its ancient evolutionary history as a symbiont, while CT had a minimal response which correlates with its status as a younger symbiont (probably via acquisition of symbiotic genes from a Burkholderia ancestor) and RM had a typical response of Alphaproteobacterial rhizospheric bacteria. Interestingly, the upregulation of nodulation genes was the only common response among the three strains; the exception was an up-regulated gene encoding a putative fatty acid hydroxylase, which appears to be a novel symbiotic gene specific to Mimosa symbionts. The transcriptional response to root exudates was correlated to each strain nodulation competitiveness, with Burkholderia phymatum appearing as the best specialised symbiont of Mimosa pudica.

  10. Bacterial cell motility of Burkholderia gut symbiont is required to colonize the insect gut.

    Science.gov (United States)

    Lee, Jun Beom; Byeon, Jin Hee; Jang, Ho Am; Kim, Jiyeun Kate; Yoo, Jin Wook; Kikuchi, Yoshitomo; Lee, Bok Luel

    2015-09-14

    We generated a Burkholderia mutant, which is deficient of an N-acetylmuramyl-l-alanine amidase, AmiC, involved in peptidoglycan degradation. When non-motile ΔamiC mutant Burkholderia cells harboring chain form were orally administered to Riptortus insects, ΔamiC mutant cells were unable to establish symbiotic association. But, ΔamiC mutant complemented with amiC gene restored in vivo symbiotic association. ΔamiC mutant cultured in minimal medium restored their motility with single-celled morphology. When ΔamiC mutant cells harboring single-celled morphology were administered to the host insect, this mutant established normal symbiotic association, suggesting that bacterial motility is essential for the successful symbiosis between host insect and Burkholderia symbiont. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  11. Riptortus pedestris and Burkholderia symbiont: an ideal model system for insect-microbe symbiotic associations.

    Science.gov (United States)

    Takeshita, Kazutaka; Kikuchi, Yoshitomo

    2017-04-01

    A number of insects establish symbiotic associations with beneficial microorganisms in various manners. The bean bug Riptortus pedestris and allied stink bugs possess an environmentally acquired Burkholderia symbiont in their midgut crypts. Unlike other insect endosymbionts, the Burkholderia symbiont is easily culturable and genetically manipulatable outside the host. In conjunction with the experimental advantages of the host insect, the Riptortus-Burkholderia symbiosis is an ideal model system for elucidating the molecular bases underpinning insect-microbe symbioses, which opens a new window in the research field of insect symbiosis. This review summarizes current knowledge of this system and discusses future perspectives. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  12. An ensemble of structures of Burkholderia pseudomallei 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase

    Energy Technology Data Exchange (ETDEWEB)

    Davies, Douglas R.; Staker, Bart L.; Abendroth, Jan A.; Edwards, Thomas E.; Hartley, Robert; Leonard, Jess; Kim, Hidong; Rychel, Amanda L.; Hewitt, Stephen N.; Myler, Peter J.; Stewart, Lance J. (UWASH); (Emerald)

    2011-12-07

    Burkholderia pseudomallei is a soil-dwelling bacterium endemic to Southeast Asia and Northern Australia. Burkholderia is responsible for melioidosis, a serious infection of the skin. The enzyme 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase (PGAM) catalyzes the interconversion of 3-phosphoglycerate and 2-phosphoglycerate, a key step in the glycolytic pathway. As such it is an extensively studied enzyme and X-ray crystal structures of PGAM enzymes from multiple species have been elucidated. Vanadate is a phosphate mimic that is a powerful tool for studying enzymatic mechanisms in phosphoryl-transfer enzymes such as phosphoglycerate mutase. However, to date no X-ray crystal structures of phosphoglycerate mutase have been solved with vanadate acting as a substrate mimic. Here, two vanadate complexes together with an ensemble of substrate and fragment-bound structures that provide a comprehensive picture of the function of the Burkholderia enzyme are reported.

  13. The Small Protein HemP Is a Transcriptional Activator for the Hemin Uptake Operon in Burkholderia multivorans ATCC 17616.

    Science.gov (United States)

    Sato, Takuya; Nonoyama, Shouta; Kimura, Akane; Nagata, Yuji; Ohtsubo, Yoshiyuki; Tsuda, Masataka

    2017-08-15

    Iron and heme play very important roles in various metabolic functions in bacteria, and their intracellular homeostasis is maintained because high concentrations of free forms of these molecules greatly facilitate the Fenton reaction-mediated production of large amounts of reactive oxygen species that severely damage various biomolecules. The ferric uptake regulator (Fur) from Burkholderia multivorans ATCC 17616 is an iron-responsive global transcriptional regulator, and its fur deletant exhibits pleiotropic phenotypes. In this study, we found that the phenotypes of the fur deletant were suppressed by an additional mutation in hemP The transcription of hemP was negatively regulated by Fur under iron-replete conditions and was constitutive in the fur deletant. Growth of a hemP deletant was severely impaired in a medium containing hemin as the sole iron source, demonstrating the important role of HemP in hemin utilization. HemP was required as a transcriptional activator that specifically binds the promoter-containing region upstream of a Fur-repressive hmuRSTUV operon, which encodes the proteins for hemin uptake. A hmuR deletant was still able to grow using hemin as the sole iron source, albeit at a rate clearly lower than that of the wild-type strain. These results strongly suggested (i) the involvement of HmuR in hemin uptake and (ii) the presence in ATCC 17616 of at least part of other unknown hemin uptake systems whose expression depends on the HemP function. Our in vitro analysis also indicated high-affinity binding of HemP to hemin, and such a property might modulate transcriptional activation of the hmu operon. IMPORTANCE Although the hmuRSTUV genes for the utilization of hemin as a sole iron source have been identified in a few Burkholderia strains, the regulatory expression of these genes has remained unknown. Our analysis in this study using B. multivorans ATCC 17616 showed that its HemP protein is required for expression of the hmuRSTUV operon, and the

  14. Identification of Burkholderia pseudomallei Near-Neighbor Species in the Northern Territory of Australia.

    Directory of Open Access Journals (Sweden)

    Jennifer L Ginther

    Full Text Available Identification and characterization of near-neighbor species are critical to the development of robust molecular diagnostic tools for biothreat agents. One such agent, Burkholderia pseudomallei, a soil bacterium and the causative agent of melioidosis, is lacking in this area because of its genomic diversity and widespread geographic distribution. The Burkholderia genus contains over 60 species and occupies a large range of environments including soil, plants, rhizospheres, water, animals and humans. The identification of novel species in new locations necessitates the need to identify the true global distribution of Burkholderia species, especially the members that are closely related to B. pseudomallei. In our current study, we used the Burkholderia-specific recA sequencing assay to analyze environmental samples from the Darwin region in the Northern Territory of Australia where melioidosis is endemic. Burkholderia recA PCR negative samples were further characterized using 16s rRNA sequencing for species identification. Phylogenetic analysis demonstrated that over 70% of the bacterial isolates were identified as B. ubonensis indicating that this species is common in the soil where B. pseudomallei is endemic. Bayesian phylogenetic analysis reveals many novel branches within the B. cepacia complex, one novel B. oklahomensis-like species, and one novel branch containing one isolate that is distinct from all other samples on the phylogenetic tree. During the analysis with recA sequencing, we discovered 2 single nucleotide polymorphisms in the reverse priming region of B. oklahomensis. A degenerate primer was developed and is proposed for future use. We conclude that the recA sequencing technique is an effective tool to classify Burkholderia and identify soil organisms in a melioidosis endemic area.

  15. Identification of Burkholderia pseudomallei Near-Neighbor Species in the Northern Territory of Australia

    Science.gov (United States)

    Ginther, Jennifer L.; Mayo, Mark; Warrington, Stephanie D.; Kaestli, Mirjam; Mullins, Travis; Wagner, David M.; Currie, Bart J.; Tuanyok, Apichai; Keim, Paul

    2015-01-01

    Identification and characterization of near-neighbor species are critical to the development of robust molecular diagnostic tools for biothreat agents. One such agent, Burkholderia pseudomallei, a soil bacterium and the causative agent of melioidosis, is lacking in this area because of its genomic diversity and widespread geographic distribution. The Burkholderia genus contains over 60 species and occupies a large range of environments including soil, plants, rhizospheres, water, animals and humans. The identification of novel species in new locations necessitates the need to identify the true global distribution of Burkholderia species, especially the members that are closely related to B. pseudomallei. In our current study, we used the Burkholderia-specific recA sequencing assay to analyze environmental samples from the Darwin region in the Northern Territory of Australia where melioidosis is endemic. Burkholderia recA PCR negative samples were further characterized using 16s rRNA sequencing for species identification. Phylogenetic analysis demonstrated that over 70% of the bacterial isolates were identified as B. ubonensis indicating that this species is common in the soil where B. pseudomallei is endemic. Bayesian phylogenetic analysis reveals many novel branches within the B. cepacia complex, one novel B. oklahomensis-like species, and one novel branch containing one isolate that is distinct from all other samples on the phylogenetic tree. During the analysis with recA sequencing, we discovered 2 single nucleotide polymorphisms in the reverse priming region of B. oklahomensis. A degenerate primer was developed and is proposed for future use. We conclude that the recA sequencing technique is an effective tool to classify Burkholderia and identify soil organisms in a melioidosis endemic area. PMID:26121041

  16. Burkholderia pseudomallei: Challenges for the Clinical Microbiology Laboratory.

    Science.gov (United States)

    Hemarajata, Peera; Baghdadi, Jonathan D; Hoffman, Risa; Humphries, Romney M

    2016-12-01

    Melioidosis is a potentially fatal infection caused by the bacterium Burkholderia pseudomallei Clinical diagnosis of melioidosis can be challenging since there is no pathognomonic clinical syndrome, and the organism is often misidentified by methods used routinely in clinical laboratories. Although the disease is more prevalent in Thailand and northern Australia, sporadic cases may be encountered in areas where it is not endemic, including the United States. Since the organism is considered a tier 1 select agent according to the Centers for Disease Control and Prevention and the U.S. Department of Agriculture Animal and Plant Health Inspection Service, clinical laboratories must be proficient at rapidly recognizing isolates suspicious for B. pseudomallei, be able to safely perform necessary rule-out tests, and to refer suspect isolates to Laboratory Response Network reference laboratories. In this minireview, we report a case of melioidosis encountered at our institution and discuss the laboratory challenges encountered when dealing with clinical isolates suspicious for B. pseudomallei or clinical specimens from suspected melioidosis cases. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  17. Characterization of Burkholderia cepacia genomovar I as a potential ...

    African Journals Online (AJOL)

    From the phylogenetic tree, UPM B3 is a specific strain within B. cepacia complex species that belong to genovomar I which is associated with strains nonpathogenic to humans. Thus, B. cepacia strain UPM B3 has the potential to be used against G. boninense, the causal pathogen of basal stem rot (BSR) in oil palm.

  18. Isolation and characterization of Burkholderia fungorum Gan-35 with the outstanding ammonia nitrogen-degrading ability from the tailings of rare-earth-element mines in southern Jiangxi, China.

    Science.gov (United States)

    Feng, Ai-Juan; Xiao, Xi; Ye, Cong-Cong; Xu, Xiao-Ming; Zhu, Qing; Yuan, Jian-Ping; Hong, Yue-Hui; Wang, Jiang-Hai

    2017-12-01

    The exploitation of rare-earth-element (REE) mines has resulted in severe ammonia nitrogen pollution and induced hazards to environments and human health. Screening microorganisms with the ammonia nitrogen-degrading ability provides a basis for bioremediation of ammonia nitrogen-polluted environments. In this study, a bacterium with the outstanding ammonia nitrogen-degrading capability was isolated from the tailings of REE mines in southern Jiangxi Province, China. This strain was identified as Burkholderia fungorum Gan-35 according to phenotypic and phylogenetic analyses. The optimal conditions for ammonia-nitrogen degradation by strain Gan-35 were determined as follows: pH value, 7.5; inoculum dose, 10%; incubation time, 44 h; temperature, 30 °C; and C/N ratio, 15:1. Strain Gan-35 degraded 68.6% of ammonia nitrogen under the optimized conditions. Nepeta cataria grew obviously better in the ammonia nitrogen-polluted soil with strain Gan-35 than that without inoculation, and the decrease in ammonia-nitrogen contents of the former was also more obvious than the latter. Besides, strain Gan-35 exhibited the tolerance to high salinities. In summary, strain Gan-35 harbors the ability of both ammonia-nitrogen degradation at high concentrations and promoting plant growth. This work has reported a Burkholderia strain with the ammonia nitrogen-degrading capability for the first time and is also the first study on the isolation of a bacterium with the ammonia nitrogen-degrading ability from the tailings of REE mines. The results are useful for developing an effective method for microbial remediation of the ammonia nitrogen-polluted tailings of REE mines.

  19. Crystallization and preliminary X-ray analysis of the receiver domain of a putative response regulator, BPSL0128, from Burkholderia pseudomallei

    International Nuclear Information System (INIS)

    Abd Aziz, Abd Ghani; Sedelnikova, Svetlana E.; Ruzheinikov, Sergey N.; Thorpe, Simon; Mohamed, Rahmah; Nathan, Sheila; Rafferty, John B.; Baker, Patrick J.; Rice, David W.

    2012-01-01

    The receiver domain of a putative response regulator from B. pseudomallei, BPSL0128, has been crystallized in a form suitable for X-ray analysis. bpsl0128, a gene encoding a putative response regulator from Burkholderia pseudomallei strain D286, has been cloned into a pETBLUE-1 vector system, overexpressed in Escherichia coli and purified. The full-length protein is degraded during purification to leave a fragment corresponding to the putative receiver domain, and crystals of this protein that diffracted to beyond 1.75 Å resolution have been grown by the hanging-drop vapour-diffusion technique using PEG 6000 as the precipitant. The crystals belonged to one of the enantiomorphic pair of space groups P3 1 21 and P3 2 21, with unit-cell parameters a = b = 65.69, c = 105.01 Å and either one or two molecules in the asymmetric unit

  20. Effect of gamma irradiation on Burkholderia thailandensis (Burkholderia pseudomallei surrogate) survival under combinations of pH and NaCl

    International Nuclear Information System (INIS)

    Yoon, Yohan; Kim, Jae-Hun; Byun, Myung-Woo; Choi, Kyoung-Hee; Lee, Ju-Woon

    2010-01-01

    This study evaluated the effect of gamma irradiation on Burkholderia thailandensis (Burkholderia pseudomallei surrogate; potential bioterrorism agent) survival under different levels of NaCl and pH. B. thailandensis in Luria Bertani broth supplemented with NaCl (0-3%), and pH-adjusted to 4-7 was treated with gamma irradiation (0-0.5 kGy). Surviving cell counts of bacteria were then enumerated on tryptic soy agar. Data for the cell counts were also used to calculate D 10 values (the dose required to reduce 1 log CFU/mL of B. thailandensis). Cell counts of B. thailandensis were decreased (P 10 values ranged from 0.04 to 0.07 kGy, regardless of NaCl and pH level. These results indicate that low doses of gamma irradiation should be a useful treatment in decreasing the potential bioterrorism bacteria, which may possibly infect humans through foods.

  1. Polar lipids of Burkholderia pseudomallei induce different host immune responses.

    Directory of Open Access Journals (Sweden)

    Mercedes Gonzalez-Juarrero

    Full Text Available Melioidosis is a disease in tropical and subtropical regions of the world that is caused by Burkholderia pseudomallei. In endemic regions the disease occurs primarily in humans and goats. In the present study, we used the goat as a model to dissect the polar lipids of B. pseudomallei to identify lipid molecules that could be used for adjuvants/vaccines or as diagnostic tools. We showed that the lipidome of B. pseudomallei and its fractions contain several polar lipids with the capacity to elicit different immune responses in goats, namely rhamnolipids and ornithine lipids which induced IFN-γ, whereas phospholipids and an undefined polar lipid induced strong IL-10 secretion in CD4(+ T cells. Autologous T cells co-cultured with caprine dendritic cells (cDCs and polar lipids of B. pseudomallei proliferated and up-regulated the expression of CD25 (IL-2 receptor molecules. Furthermore, we demonstrated that polar lipids were able to up-regulate CD1w2 antigen expression in cDCs derived from peripheral blood monocytes. Interestingly, the same polar lipids had only little effect on the expression of MHC class II DR antigens in the same caprine dendritic cells. Finally, antibody blocking of the CD1w2 molecules on cDCs resulted in decreased expression for IFN-γ by CD4(+ T cells. Altogether, these results showed that polar lipids of B. pseudomallei are recognized by the caprine immune system and that their recognition is primarily mediated by the CD1 antigen cluster.

  2. Adaptation and Antibiotic Tolerance of Anaerobic Burkholderia pseudomallei ▿ †

    Science.gov (United States)

    Hamad, Mohamad A.; Austin, Chad R.; Stewart, Amanda L.; Higgins, Mike; Vázquez-Torres, Andrés; Voskuil, Martin I.

    2011-01-01

    The Gram-negative bacterium Burkholderia pseudomallei is the etiological agent of melioidosis and is remarkably resistant to most classes of antibacterials. Even after months of treatment with antibacterials that are relatively effective in vitro, there is a high rate of treatment failure, indicating that this pathogen alters its patterns of antibacterial susceptibility in response to cues encountered in the host. The pathology of melioidosis indicates that B. pseudomallei encounters host microenvironments that limit aerobic respiration, including the lack of oxygen found in abscesses and in the presence of nitric oxide produced by macrophages. We investigated whether B. pseudomallei could survive in a nonreplicating, oxygen-deprived state and determined if this physiological state was tolerant of conventional antibacterials. B. pseudomallei survived initial anaerobiosis, especially under moderately acidic conditions similar to those found in abscesses. Microarray expression profiling indicated a major shift in the physiological state of hypoxic B. pseudomallei, including induction of a variety of typical anaerobic-environment-responsive genes and genes that appear specific to anaerobic B. pseudomallei. Interestingly, anaerobic B. pseudomallei was unaffected by antibacterials typically used in therapy. However, it was exquisitely sensitive to drugs used against anaerobic pathogens. After several weeks of anaerobic culture, a significant loss of viability was observed. However, a stable subpopulation that maintained complete viability for at least 1 year was established. Thus, during the course of human infection, if a minor subpopulation of bacteria inhabited an oxygen-restricted environment, it might be indifferent to traditional therapy but susceptible to antibiotics frequently used to treat anaerobic infections. PMID:21537012

  3. Polar Lipids of Burkholderia pseudomallei Induce Different Host Immune Responses

    Science.gov (United States)

    Gonzalez-Juarrero, Mercedes; Mima, Naoko; Trunck, Lily A.; Schweizer, Herbert P.; Bowen, Richard A.; Dascher, Kyle; Mwangi, Waithaka; Eckstein, Torsten M.

    2013-01-01

    Melioidosis is a disease in tropical and subtropical regions of the world that is caused by Burkholderia pseudomallei. In endemic regions the disease occurs primarily in humans and goats. In the present study, we used the goat as a model to dissect the polar lipids of B. pseudomallei to identify lipid molecules that could be used for adjuvants/vaccines or as diagnostic tools. We showed that the lipidome of B. pseudomallei and its fractions contain several polar lipids with the capacity to elicit different immune responses in goats, namely rhamnolipids and ornithine lipids which induced IFN-γ, whereas phospholipids and an undefined polar lipid induced strong IL-10 secretion in CD4+ T cells. Autologous T cells co-cultured with caprine dendritic cells (cDCs) and polar lipids of B. pseudomallei proliferated and up-regulated the expression of CD25 (IL-2 receptor) molecules. Furthermore, we demonstrated that polar lipids were able to up-regulate CD1w2 antigen expression in cDCs derived from peripheral blood monocytes. Interestingly, the same polar lipids had only little effect on the expression of MHC class II DR antigens in the same caprine dendritic cells. Finally, antibody blocking of the CD1w2 molecules on cDCs resulted in decreased expression for IFN-γ by CD4+ T cells. Altogether, these results showed that polar lipids of B. pseudomallei are recognized by the caprine immune system and that their recognition is primarily mediated by the CD1 antigen cluster. PMID:24260378

  4. Recovery efficiencies for Burkholderia thailandensis from various aerosol sampling media

    Directory of Open Access Journals (Sweden)

    Paul eDabisch

    2012-06-01

    Full Text Available Burkholderia thailandensis is used in the laboratory as a surrogate of the more virulent B. pseudomallei. Since inhalation is believed to be a natural route of infection for B. pseudomallei, many animal studies with B. pseudomallei and B. thailandensis utilize the inhalation route of exposure. The aim of the present study was to quantify the recovery efficiency of culturable B. thailandensis from several common aerosol sampling devices to ensure that collected microorganisms could be reliably recovered post-collection. The sampling devices tested included 25-mm gelatin filters, 25-mm stainless steel disks used in Mercer cascade impactors, and two types of glass impingers. The results demonstrate that while several processing methods tested resulted in significantly lower physical recovery efficiencies than other methods, it was possible to obtain culturable recovery efficiencies for B. thailandensis and physical recovery efficiencies for 1 μm fluorescent spheres of at least 0.95 from all of the sampling media tested given an appropriate sample processing procedure. The results of the present study also demonstrated that the bubbling action of liquid media in all-glass impingers (AGIs can result in physical loss of material from the collection medium, although additional studies are needed to verify the exact mechanisms involved. Overall, the results of this study demonstrate that the collection mechanism as well as the post-collection processing method can significantly affect the recovery from and retention of culturable microorganisms in sampling media, potentially affecting the calculated airborne concentration and any subsequent estimations of risk or dose derived from such data.

  5. Influence of neutrophil defects on Burkholderia cepacia complex pathogenesis

    Directory of Open Access Journals (Sweden)

    Laura A. Porter

    2011-11-01

    Full Text Available The Burkholderia cepacia complex (Bcc is a group of Gram-negative bacteria that are ubiquitous in the environment and have emerged as opportunistic pathogens in immunocompromised patients. The primary patient populations infected with Bcc include individuals with cystic fibrosis (CF, as well as those with chronic granulomatous disease (CGD. While Bcc infection in CF is better characterized than in CGD, these two genetic diseases are not obviously similar and it is currently unknown if there is any commonality in host immune defects that is responsible for the susceptibility to Bcc. CF is caused by mutations in the CF transmembrane conductance regulator, resulting in manifestations in various organ systems, however the major cause of morbidity and mortality is currently due to bacterial respiratory infections. CGD, on the other hand, is a genetic disorder that is caused by defects in phagocyte NADPH oxidase. Because of the defect in CGD, phagocytes in these patients are unable to produce reactive oxygen species, which results in increased susceptibility to bacterial and fungal infections. Despite this significant defect in microbial clearance, the spectrum of pathogens frequently implicated in infections in CGD is relatively narrow and includes some bacterial species that are considered almost pathognomonic for this disorder. Very little is known about the cause of the specific susceptibility to Bcc over other potential pathogens more prevalent in the environment, and a better understanding of specific mechanisms required for bacterial virulence has become a high priority. This review will summarize both the current knowledge and future directions related to Bcc virulence in immunocompromised individuals with a focus on the roles of bacterial factors and neutrophil defects in pathogenesis.

  6. Diversity of 16S-23S rDNA internal transcribed spacer (ITS reveals phylogenetic relationships in Burkholderia pseudomallei and its near-neighbors.

    Directory of Open Access Journals (Sweden)

    Andrew P Liguori

    Full Text Available Length polymorphisms within the 16S-23S ribosomal DNA internal transcribed spacer (ITS have been described as stable genetic markers for studying bacterial phylogenetics. In this study, we used these genetic markers to investigate phylogenetic relationships in Burkholderia pseudomallei and its near-relative species. B. pseudomallei is known as one of the most genetically recombined bacterial species. In silico analysis of multiple B. pseudomallei genomes revealed approximately four homologous rRNA operons and ITS length polymorphisms therein. We characterized ITS distribution using PCR and analyzed via a high-throughput capillary electrophoresis in 1,191 B. pseudomallei strains. Three major ITS types were identified, two of which were commonly found in most B. pseudomallei strains from the endemic areas, whereas the third one was significantly correlated with worldwide sporadic strains. Interestingly, mixtures of the two common ITS types were observed within the same strains, and at a greater incidence in Thailand than Australia suggesting that genetic recombination causes the ITS variation within species, with greater recombination frequency in Thailand. In addition, the B. mallei ITS type was common to B. pseudomallei, providing further support that B. mallei is a clone of B. pseudomallei. Other B. pseudomallei near-neighbors possessed unique and monomorphic ITS types. Our data shed light on evolutionary patterns of B. pseudomallei and its near relative species.

  7. Novel engineered cationic antimicrobial peptides display broad-spectrum activity against Francisella tularensis, Yersinia pestis and Burkholderia pseudomallei.

    Science.gov (United States)

    Abdelbaqi, Suha; Deslouches, Berthony; Steckbeck, Jonathan; Montelaro, Ronald; Reed, Douglas S

    2016-02-01

    Broad-spectrum antimicrobials are needed to effectively treat patients infected in the event of a pandemic or intentional release of a pathogen prior to confirmation of the pathogen's identity. Engineered cationic antimicrobial peptides (eCAPs) display activity against a number of bacterial pathogens including multi-drug-resistant strains. Two lead eCAPs, WLBU2 and WR12, were compared with human cathelicidin (LL-37) against three highly pathogenic bacteria: Francisella tularensis, Yersinia pestis and Burkholderia pseudomallei. Both WLBU2 and WR12 demonstrated bactericidal activity greater than that of LL-37, particularly against F. tularensis and Y. pestis. Only WLBU2 had bactericidal activity against B. pseudomallei. WLBU2, WR12 and LL-37 were all able to inhibit the growth of the three bacteria in vitro. Because these bacteria can be facultative intracellular pathogens, preferentially infecting macrophages and dendritic cells, we evaluated the activity of WLBU2 against F. tularensis in an ex vivo infection model with J774 cells, a mouse macrophage cell line. In that model WLBU2 was able to achieve greater than 50% killing of F. tularensis at a concentration of 12.5 μM. These data show the therapeutic potential of eCAPs, particularly WLBU2, as a broad-spectrum antimicrobial for treating highly pathogenic bacterial infections.

  8. Transcriptome analysis of Acidovorax avenae subsp. avenae cultivated in vivo and co-culture with Burkholderia seminalis.

    Science.gov (United States)

    Li, Bin; Ibrahim, Muhammad; Ge, Mengyu; Cui, Zhouqi; Sun, Guochang; Xu, Fei; Kube, Michael

    2014-07-16

    Response of bacterial pathogen to environmental bacteria and its host is critical for understanding of microbial adaption and pathogenesis. Here, we used RNA-Seq to comprehensively and quantitatively assess the transcriptional response of Acidovorax avenae subsp. avenae strain RS-1 cultivated in vitro, in vivo and in co-culture with rice rhizobacterium Burkholderia seminalis R456. Results revealed a slight response to other bacteria, but a strong response to host. In particular, a large number of virulence associated genes encoding Type I to VI secretion systems, 118 putative non-coding RNAs, and 7 genomic islands (GIs) were differentially expressed in vivo based on comparative genomic and transcriptomic analyses. Furthermore, the loss of virulence for knockout mutants of 11 differentially expressed T6SS genes emphasized the importance of these genes in bacterial pathogenicity. In addition, the reliability of expression data obtained by RNA-Seq was supported by quantitative real-time PCR of the 25 selected T6SS genes. Overall, this study highlighted the role of differentially expressed genes in elucidating bacterial pathogenesis based on combined analysis of RNA-Seq data and knockout of T6SS genes.

  9. Genome-wide analysis reveals loci encoding anti-macrophage factors in the human pathogen Burkholderia pseudomallei K96243.

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    Andrea J Dowling

    2010-12-01

    Full Text Available Burkholderia pseudomallei is an important human pathogen whose infection biology is still poorly understood. The bacterium is endemic to tropical regions, including South East Asia and Northern Australia, where it causes melioidosis, a serious disease associated with both high mortality and antibiotic resistance. B. pseudomallei is a Gram-negative facultative intracellular pathogen that is able to replicate in macrophages. However despite the critical nature of its interaction with macrophages, few anti-macrophage factors have been characterized to date. Here we perform a genome-wide gain of function screen of B. pseudomallei strain K96243 to identify loci encoding factors with anti-macrophage activity. We identify a total of 113 such loci scattered across both chromosomes, with positive gene clusters encoding transporters and secretion systems, enzymes/toxins, secondary metabolite, biofilm, adhesion and signal response related factors. Further phenotypic analysis of four of these regions shows that the encoded factors cause striking cellular phenotypes relevant to infection biology, including apoptosis, formation of actin 'tails' and multi-nucleation within treated macrophages. The detailed analysis of the remaining host of loci will facilitate genetic dissection of the interaction of this important pathogen with host macrophages and thus further elucidate this critical part of its infection cycle.

  10. Biocontrol of Late Blight (Phytophthora capsici Disease and Growth Promotion of Pepper by Burkholderia cepacia MPC-7

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    Mao Sopheareth

    2013-03-01

    Full Text Available A chitinolytic bacterial strain having strong antifungal activity was isolated and identified as Burkholderia cepacia MPC-7 based on 16S rRNA gene analysis. MPC-7 solubilized insoluble phosphorous in hydroxyapatite agar media. It produced gluconic acid and 2-ketogluconic acid related to the decrease in pH of broth culture. The antagonist produced benzoic acid (BA and phenylacetic acid (PA. The authentic compounds, BA and PA, showed a broad spectrum of antimicrobial activity against yeast, several bacterial and fungal pathogens in vitro. To demonstrate the biocontrol efficiency of MPC-7 on late blight disease caused by Phytophthora capsici, pepper plants in pot trials were treated with modified medium only (M, M plus zoospore inoculation (MP, MPC-7 cultured broth (B and B plus zoospore inoculation (BP. With the sudden increase in root mortality, plants in MP wilted as early as five days after pathogen inoculation. However, plant in BP did not show any symptom of wilting until five days. Root mortality in BP was markedly reduced for as much as 50%. Plants in B had higher dry weight, P concentration in root, and larger leaf area compared to those in M and MP. These results suggested that B. cepacia MPC-7 should be considered as a candidate for the biological fertilizer as well as antimicrobial agent for pepper plants.

  11. N-acylhomoserine-lactone-mediated communication between Pseudomonas aeruginosa and Burkholderia cepacia in mixed biofilms

    DEFF Research Database (Denmark)

    Riedel, K.; Hentzer, Morten; Geisenberger, O.

    2001-01-01

    Pseudomonas aeruginosa and Burkholderia cepacia are capable of forming mixed biofilms in the lungs of cystic fibrosis patients. Both bacteria employ quorum-sensing systems, which rely on N-acylhomoserine lactone (AHL) signal molecules, to co- ordinate expression of virulence factors with the form...

  12. A heterodimer comprised of two bovine lactoferrin antimicrobial peptides exhibits powerful bactericidal activity against Burkholderia pseudomallei

    NARCIS (Netherlands)

    Puknun, A.; Bolscher, J.G.M.; Nazmi, K.; Veerman, E.C.I.; Tungpradabkul, S.; Wongratanacheewin, S.; Kanthawong, S.; Taweechaisupapong, S.

    2013-01-01

    Melioidosis is a severe infectious disease that is endemic in Southeast Asia and Northern Australia. Burkholderia pseudomallei, the causative agent of this disease, has developed resistance to an increasing list of antibiotics, demanding a search for novel agents. Lactoferricin and lactoferrampin

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

    Science.gov (United States)

    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.

  14. Rapid DNA vaccination against Burkholderia pseudomallei flagellin by tattoo or intranasal application

    NARCIS (Netherlands)

    Lankelma, Jacqueline M.; Wagemakers, Alex; Birnie, Emma; Haak, Bastiaan W.; Trentelman, Jos J. A.; Weehuizen, Tassili A. F.; Ersöz, Jasmin; Roelofs, Joris J. T. H.; Hovius, Joppe W.; Wiersinga, W. Joost; Bins, Adriaan D.

    2017-01-01

    Melioidosis is a severe infectious disease with a high mortality that is endemic in South-East Asia and Northern Australia. The causative pathogen, Burkholderia pseudomallei, is listed as potential bioterror weapon due to its high virulence and potential for easy dissemination. Currently, there is

  15. Study of the mode of action of a polygalacturonase from the phytopathogen Burkholderia cepacia

    DEFF Research Database (Denmark)

    Massa, C.; Clausen, Mads Hartvig; Stojan, J.

    2007-01-01

    We have recently isolated and heterologously expressed BcPeh28A, an endopolygalacturonase from the phytopathogenic Gram-negative bacterium Burkholderia cepacia. Endopolygalacturonases belong to glycoside hydrolase family 28 and are responsible for the hydrolysis of the non-esterified regions...

  16. Burkholderia pseudomallei Infection in a Cystic Fibrosis Patient from the Caribbean: A Case Report

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    Dimas Mateos Corral

    2008-01-01

    Full Text Available Burkholderia pseudomallei is a pathogen identified with increasing frequency in the respiratory tracts of cystic fibrosis (CF patients from endemic areas such as Southeast Asia and northern Australia. The following report describes the first known reported case in a CF patient from the Caribbean attending a North American CF clinic.

  17. Burkholderia pseudomallei infection in a cystic fibrosis patient from the Caribbean: A case report

    Science.gov (United States)

    Corral, Dimas Mateos; Coates, Allan L; Yau, Yvonne CW; Tellier, Raymond; Glass, Mindy; Jones, Steven M; Waters, Valerie J

    2008-01-01

    Burkholderia pseudomallei is a pathogen identified with increasing frequency in the respiratory tracts of cystic fibrosis (CF) patients from endemic areas such as Southeast Asia and northern Australia. The following report describes the first known reported case in a CF patient from the Caribbean attending a North American CF clinic. PMID:18716683

  18. Multivariate analyses of Burkholderia species in soil: effect of crop and land use history

    NARCIS (Netherlands)

    Salles, Joanna; Van Veen, J.A.; van Elsas, J.D.

    2004-01-01

    The assessment of Burkholderia diversity in agricultural areas is important considering the potential use of this genus for agronomic and environmental applications. Therefore, the aim of this work was to ascertain how plant species and land use management drive the diversity of the genus

  19. Multivariate analyses of Burkholderia species in soil : Effect of crop and land use history

    NARCIS (Netherlands)

    Salles, JF; van Veen, JA; van Elsas, JD

    The assessment of Burkholderia diversity in agricultural areas is important considering the potential use of this genus for agronomic and environmental applications. Therefore, the aim of this work was to ascertain how plant species and land use management drive the diversity of the genus

  20. Molecular method to assess the diversity of Burkholderia species in environmental samples

    NARCIS (Netherlands)

    Salles, J.; Souza, de F.A.; Elsas, van J.D.

    2002-01-01

    In spite of the importance of many members of the genus Burkholderia in the soil microbial community, no direct method to assess the diversity of this genus has been developed so far. The aim of this work was the development of soil DNA-based PCR-denaturing gradient gel electrophoresis (DGGE), a

  1. The relationship of biofilm production to biocontrol activity of Burkholderia pyrrocinia FP62

    Science.gov (United States)

    Foliar biocontrol agent (BCA) efficacy is often inconsistent due to poor colonization and survival on plant surfaces. Burkholderia pyrrocinia FP62, a superior leaf colonist and BCA of Botrytis cinerea, forms unsaturated biofilms on plant surfaces. To determine the relationship between biocontrol act...

  2. Determining the biochemical properties of the Oxalate Biosynthetic Component (Obc)1 from Burkholderia mallei

    Science.gov (United States)

    Oxalic acid is produced by a variety of organisms ranging from simple microbes to complex animals. This acid has been proposed to fulfill various physiological and pathological functions which vary between organisms. In bacteria from the Burkholderia genus, oxalate secretion has been shown to be quo...

  3. Multivariate Analyses of Burkholderia species in soil: effect of crop and land use history.

    NARCIS (Netherlands)

    Salles, J.F.; Veen, van J.A.; Elsas, van J.D.

    2004-01-01

    The assessment of Burkholderia diversity in agricultural areas is important considering the potential use of this genus for agronomic and environmental applications. Therefore, the aim of this work was to ascertain how plant species and land use management drive the diversity of the genus

  4. Effect of agricultural management regimes on Burkholderia community structure in soil

    NARCIS (Netherlands)

    Salles, Joanna; van Elsas, J.D.; Van Veen, J.A.

    2006-01-01

    The main objective of this study was to determine the Burkholderia community structure associated with areas under different agricultural management and to evaluate to which extent this community structure is affected by changes in agricultural management. Two fields with distinct soil history

  5. Assessing the potential for Burkholderia pseudomallei in the southeastern United States

    Science.gov (United States)

    Burkholderia pseudomallei, the causative agent of melioidosis, is an underreported zoonosis in many countries where environmental conditions may be favorable for B. pseudomallei. This soil saprophyte is most often detected in tropical areas such as Southeast Asia and Northern Australia where the cas...

  6. Effect of agricultural management regime on Burkholderia community structure in soil

    NARCIS (Netherlands)

    Salles, J.F.; Elsas, van J.D.; Veen, van J.A.

    2006-01-01

    The main objective of this study was to determine the Burkholderia community structure associated with areas under different agricultural management and to evaluate to which extent this community structure is affected by changes in agricultural management. Two fields with distinct soil history

  7. Effect of agricultural management regime on Burkholderia community structure in soil

    NARCIS (Netherlands)

    Salles, J. F.; van Elsas, J. D.; van Veen, J. A.

    The main objective of this study was to determine the Burkholderia community structure associated with areas under different agricultural management and to evaluate to which extent this community structure is affected by changes in agricultural management. Two fields with distinct soil history

  8. Changes in agricultural management drive the diversity of Burkholderia species isolated from soil on PCAT medium

    NARCIS (Netherlands)

    Salles, Joanna; Samyn, E.; Vandamme, P.; Van Veen, J.A.; van Elsas, J.D.

    2006-01-01

    Abstract In order to assess the diversity of culturable Burkholderia populations in rhizosphere and bulk soil and to evaluate how different agricultural management regimes and land use history affect this diversity, four treatments were evaluated: permanent grassland; grassland converted into maize

  9. Molecular method to assess the diversity of Burkholderia species in environmental samples

    NARCIS (Netherlands)

    Salles, Joanna; De Souza, F.A.; Van Elsas, J.D.

    2002-01-01

    In spite of the importance of many members of the genus Burkholderia in the soil microbial community, no direct method to assess the diversity of this genus has been developed so far. The aim of this work was the development of soil DNA-based PCR-denaturing gradient get electrophoresis (DGGE), a

  10. Neutrophil extracellular traps in the host defense against sepsis induced by Burkholderia pseudomallei (melioidosis)

    NARCIS (Netherlands)

    de Jong, Hanna K.; Koh, Gavin C. K. W.; Achouiti, Ahmed; van der Meer, Anne J.; Bulder, Ingrid; Stephan, Femke; Roelofs, Joris J. T. H.; Day, Nick P. J.; Peacock, Sharon J.; Zeerleder, Sacha; Wiersinga, W. Joost

    2014-01-01

    Neutrophil extracellular traps (NETs) are a central player in the host response to bacteria: neutrophils release extracellular DNA (nucleosomes) and neutrophil elastase to entrap and kill bacteria. We studied the role of NETs in Burkholderia pseudomallei infection (melioidosis), an important cause

  11. Two-Phase Bactericidal Mechanism of Silver Nanoparticles against Burkholderia pseudomallei.

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    Pawinee Siritongsuk

    Full Text Available Silver nanoparticles (AgNPs have a strong antimicrobial activity against a variety of pathogenic bacteria. The killing mechanism of AgNPs involves direct physical membrane destruction and subsequent molecular damage from both AgNPs and released Ag+. Burkholderia pseudomallei is the causative agent of melioidosis, an endemic infectious disease primarily found in northern Australia and Southeast Asia. B. pseudomallei is intrinsically resistant to most common antibiotics. In this study, the antimicrobial activity and mechanism of AgNPs (10-20 nm against B. pseudomallei were investigated. The MIC and MBC for nine B. pseudomallei strains ranged from 32-48 μg/mL and 96-128 μg/mL, respectively. Concentrations of AgNPs less than 256 μg/mL were not toxic to human red blood cells. AgNPs exhibited a two-phase mechanism: cell death induction and ROS induction. The first phase was a rapid killing step within 5 min, causing the direct damage of the cytoplasmic membrane of the bacterial cells, as observed by a time-kill assay and fluorescence microscopy. During the period of 5-30 min, the cell surface charge was rapidly neutralized from -8.73 and -7.74 to 2.85 and 2.94 mV in two isolates of B. pseudomallei, as revealed by zeta potential measurement. Energy-dispersive X-ray (EDX spectroscopy showed the silver element deposited on the bacterial membrane, and TEM micrographs of the AgNP-treated B. pseudomallei cells showed severe membrane damage and cytosolic leakage at 1/5 MIC and cell bursting at MBC. During the killing effect the released Ag+ from AgNPs was only 3.9% from the starting AgNPs concentration as observed with ICP-OES experiment. In the second phase, the ROS induction occurred 1-4 hr after the AgNP treatment. Altogether, we provide direct kinetic evidence of the AgNPs killing mechanism, by which cell death is separable from the ROS induction and AgNPs mainly contributes in the killing action. AgNPs may be considered a potential candidate to

  12. Tandem repeat regions within the Burkholderia pseudomallei genome and their application for high resolution genotyping

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    Harvey Steven P

    2007-03-01

    Full Text Available Abstract Background The facultative, intracellular bacterium Burkholderia pseudomallei is the causative agent of melioidosis, a serious infectious disease of humans and animals. We identified and categorized tandem repeat arrays and their distribution throughout the genome of B. pseudomallei strain K96243 in order to develop a genetic typing method for B. pseudomallei. We then screened 104 of the potentially polymorphic loci across a diverse panel of 31 isolates including B. pseudomallei, B. mallei and B. thailandensis in order to identify loci with varying degrees of polymorphism. A subset of these tandem repeat arrays were subsequently developed into a multiple-locus VNTR analysis to examine 66 B. pseudomallei and 21 B. mallei isolates from around the world, as well as 95 lineages from a serial transfer experiment encompassing ~18,000 generations. Results B. pseudomallei contains a preponderance of tandem repeat loci throughout its genome, many of which are duplicated elsewhere in the genome. The majority of these loci are composed of repeat motif lengths of 6 to 9 bp with 4 to 10 repeat units and are predominately located in intergenic regions of the genome. Across geographically diverse B. pseudomallei and B.mallei isolates, the 32 VNTR loci displayed between 7 and 28 alleles, with Nei's diversity values ranging from 0.47 and 0.94. Mutation rates for these loci are comparable (>10-5 per locus per generation to that of the most diverse tandemly repeated regions found in other less diverse bacteria. Conclusion The frequency, location and duplicate nature of tandemly repeated regions within the B. pseudomallei genome indicate that these tandem repeat regions may play a role in generating and maintaining adaptive genomic variation. Multiple-locus VNTR analysis revealed extensive diversity within the global isolate set containing B. pseudomallei and B. mallei, and it detected genotypic differences within clonal lineages of both species that were

  13. A midgut lysate of the Riptortus pedestris has antibacterial activity against LPS O-antigen-deficient Burkholderia mutants.

    Science.gov (United States)

    Jang, Ho Am; Seo, Eun Sil; Seong, Min Young; Lee, Bok Luel

    2017-02-01

    Riptortus pedestris, a common pest in soybean fields, harbors a symbiont Burkholderia in a specialized posterior midgut region of insects. Every generation of second nymphs acquires new Burkholderia cells from the environment. We compared in vitro cultured Burkholderia with newly in vivo colonized Burkholderia in the host midgut using biochemical approaches. The bacterial cell envelope of in vitro cultured and in vivo Burkholderia differed in structure, as in vivo bacteria lacked lipopolysaccharide (LPS) O-antigen. The LPS O-antigen deficient bacteria had a reduced colonization rate in the host midgut compared with that of the wild-type Burkholderia. To determine why LPS O-antigen-deficient bacteria are less able to colonize the host midgut, we examined in vitro survival rates of three LPS O-antigen-deficient Burkholderia mutants and lysates of five different midgut regions. The LPS O-antigen-deficient mutants were highly susceptible when cultured with the lysate of a specific first midgut region (M1), indicating that the M1 lysate contains unidentified substance(s) capable of killing LPS O-antigen-deficient mutants. We identified a 17 kDa protein from the M1 lysate, which was enriched in the active fractions. The N-terminal sequence of the protein was determined to be a soybean Kunitz-type trypsin inhibitor. These data suggest that the 17 kDa protein, which was originated from a main soybean source of the R. pedestris host, has antibacterial activity against the LPS O-antigen deficient (rough-type) Burkholderia. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Evidence of environmental and vertical transmission of Burkholderia symbionts in the oriental chinch bug, Cavelerius saccharivorus (Heteroptera: Blissidae).

    Science.gov (United States)

    Itoh, Hideomi; Aita, Manabu; Nagayama, Atsushi; Meng, Xian-Ying; Kamagata, Yoichi; Navarro, Ronald; Hori, Tomoyuki; Ohgiya, Satoru; Kikuchi, Yoshitomo

    2014-10-01

    The vertical transmission of symbiotic microorganisms is omnipresent in insects, while the evolutionary process remains totally unclear. The oriental chinch bug, Cavelerius saccharivorus (Heteroptera: Blissidae), is a serious sugarcane pest, in which symbiotic bacteria densely populate the lumen of the numerous tubule-like midgut crypts that the chinch bug develops. Cloning and sequence analyses of the 16S rRNA genes revealed that the crypts were dominated by a specific group of bacteria belonging to the genus Burkholderia of the Betaproteobacteria. The Burkholderia sequences were distributed into three distinct clades: the Burkholderia cepacia complex (BCC), the plant-associated beneficial and environmental (PBE) group, and the stinkbug-associated beneficial and environmental group (SBE). Diagnostic PCR revealed that only one of the three groups of Burkholderia was present in ∼89% of the chinch bug field populations tested, while infections with multiple Burkholderia groups within one insect were observed in only ∼10%. Deep sequencing of the 16S rRNA gene confirmed that the Burkholderia bacteria specifically colonized the crypts and were dominated by one of three Burkholderia groups. The lack of phylogenetic congruence between the symbiont and the host population strongly suggested host-symbiont promiscuity, which is probably caused by environmental acquisition of the symbionts by some hosts. Meanwhile, inspections of eggs and hatchlings by diagnostic PCR and egg surface sterilization demonstrated that almost 30% of the hatchlings vertically acquire symbiotic Burkholderia via symbiont-contaminated egg surfaces. The mixed strategy of symbiont transmission found in the oriental chinch bug might be an intermediate stage in evolution from environmental acquisition to strict vertical transmission in insects. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  15. Burkholderia pseudomallei Evades Nramp1 (Slc11a1- and NADPH Oxidase-Mediated Killing in Macrophages and Exhibits Nramp1-Dependent Virulence Gene Expression

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    Veerachat Muangsombut

    2017-08-01

    Full Text Available Bacterial survival in macrophages can be affected by the natural resistance-associated macrophage protein 1 (Nramp1; also known as solute carrier family 11 member a1 or Slc11a1 which localizes to phagosome membranes and transports divalent cations, including iron. Little is known about the role of Nramp1 in Burkholderia infection, in particular whether this differs for pathogenic species like Burkholderia pseudomallei causing melioidosis or non-pathogenic species like Burkholderia thailandensis. Here we show that transfected macrophages stably expressing wild-type Nramp1 (Nramp1+ control the net replication of B. thailandensis, but not B. pseudomallei. Control of B. thailandensis was associated with increased cytokine responses, and could be abrogated by blocking NADPH oxidase-mediated production of reactive oxygen species but not by blocking generation of reactive nitrogen species. The inability of Nramp1+ macrophages to control B. pseudomallei was associated with rapid escape of bacteria from phagosomes, as indicated by decreased co-localization with LAMP1 compared to B. thailandensis. A B. pseudomallei bipB mutant impaired in escape from phagosomes was controlled to a greater extent than the parent strain in Nramp1+ macrophages, but was also attenuated in Nramp1− cells. Consistent with reduced escape from phagosomes, B. thailandensis formed fewer multinucleated giant cells in Nramp1+ macrophages at later time points compared to B. pseudomallei. B. pseudomallei exhibited elevated transcription of virulence-associated genes of Type VI Secretion System cluster 1 (T6SS-1, the Bsa Type III Secretion System (T3SS-3 and the bimA gene required for actin-based motility in Nramp1+ macrophages. Nramp1+ macrophages were found to contain decreased iron levels that may impact on expression of such genes. Our data show that B. pseudomallei is able to evade Nramp1- and NADPH oxidase-mediated killing in macrophages and that expression of virulence

  16. Phylogenomic Analysis Reveals an Asian Origin for African Burkholderia pseudomallei and Further Supports Melioidosis Endemicity in Africa.

    Science.gov (United States)

    Sarovich, Derek S; Garin, Benoit; De Smet, Birgit; Kaestli, Mirjam; Mayo, Mark; Vandamme, Peter; Jacobs, Jan; Lompo, Palpouguini; Tahita, Marc C; Tinto, Halidou; Djaomalaza, Innocente; Currie, Bart J; Price, Erin P

    2016-01-01

    Burkholderia pseudomallei, an environmental bacterium that causes the deadly disease melioidosis, is endemic in northern Australia and Southeast Asia. An increasing number of melioidosis cases are being reported in other tropical regions, including Africa and the Indian Ocean islands. B. pseudomallei first emerged in Australia, with subsequent rare dissemination event(s) to Southeast Asia; however, its dispersal to other regions is not yet well understood. We used large-scale comparative genomics to investigate the origins of three B. pseudomallei isolates from Madagascar and two from Burkina Faso. Phylogenomic reconstruction demonstrates that these African B. pseudomallei isolates group into a single novel clade that resides within the more ancestral Asian clade. Intriguingly, South American strains reside within the African clade, suggesting more recent dissemination from West Africa to the Americas. Anthropogenic factors likely assisted in B. pseudomallei dissemination to Africa, possibly during migration of the Austronesian peoples from Indonesian Borneo to Madagascar ~2,000 years ago, with subsequent genetic diversity driven by mutation and recombination. Our study provides new insights into global patterns of B. pseudomallei dissemination and adds to the growing body of evidence of melioidosis endemicity in Africa. Our findings have important implications for melioidosis diagnosis and management in Africa. IMPORTANCE Sporadic melioidosis cases have been reported in the African mainland and Indian Ocean islands, but until recently, these regions were not considered areas where B. pseudomallei is endemic. Given the high mortality rate of melioidosis, it is crucial that this disease be recognized and suspected in all regions of endemicity. Previous work has shown that B. pseudomallei originated in Australia, with subsequent introduction into Asia; however, the precise origin of B. pseudomallei in other tropical regions remains poorly understood. Using

  17. Isolation of the highly pathogenic and zoonotic agent Burkholderia pseudomallei from a pet green Iguana in Prague, Czech Republic.

    Science.gov (United States)

    Elschner, Mandy C; Hnizdo, Jan; Stamm, Ivonne; El-Adawy, Hosny; Mertens, Katja; Melzer, Falk

    2014-11-28

    Melioidosis caused by Burkholderia (B.) pseudomallei is an endemic zoonotic disease mainly reported from northern Australia and Southeast Asia. In Europe, cases of human melioidosis have been reported only from patients travelling to endemic regions. Besides humans, B. pseudomallei has a very broad host range in domestic and wild animals. There are some reports about importation of B. pseudomallei-infected animals from endemic areas into Europe. The present report describes the first case of B. pseudomallei infection of a pet iguana in Europe. In a 5-year-old pet Iguana iguana living in a private household in Prague, Czech Republic, B. pseudomallei was isolated from pus of an abscess. The isolate VB976100 was identified by Vitek®2, MALDI-TOF mass spectrometry and polymerase chain reaction as B. pseudomallei. The molecular typing resulted in multi-locus sequence type 436 hitherto, which has been found only once worldwide in a B. pseudomallei strain isolated in the USA and originating from Guatemala. The identification as internal transcribed spacer type G indicates a close relatedness to strains mainly isolated in the Western Hemisphere. These findings support the hypothesis that the iguana became infected in this region or in a breeding facility through contact to other infected animals. The present case highlights the risk of importation of the highly pathogenic and zoonotic B. pseudomallei into non-endemic regions through animal trade. Therefore, veterinarians treating animals from these areas and physicians examining patients owning such animals should include melioidosis in differential diagnosis whenever specific symptoms appear. Furthermore, veterinary authorities responsible for supervision of traders and pet shops should be aware of this risk of zoonotic transmission.

  18. In Vitro Activity of Fusidic Acid (CEM-102, Sodium Fusidate) against Staphylococcus aureus Isolates from Cystic Fibrosis Patients and Its Effect on the Activities of Tobramycin and Amikacin against Pseudomonas aeruginosa and Burkholderia cepacia▿

    Science.gov (United States)

    McGhee, Pamela; Clark, Catherine; Credito, Kim; Beachel, Linda; Pankuch, Glenn A.; Appelbaum, Peter C.; Kosowska-Shick, Klaudia

    2011-01-01

    We tested the MICs of fusidic acid (CEM-102) plus other agents against 40 methicillin-resistant Staphylococcus aureus (MRSA) isolates from cystic fibrosis patients and the activities of fusidic acid with or without tobramycin or amikacin against Pseudomonas aeruginosa, MRSA, and Burkholderia cepacia isolates from cystic fibrosis patients in a 24-h time-kill study. Fusidic acid was potent (MICs, 0.125 to 0.5 μg/ml; a single 500-mg dose of fusidic acid at 8 h averaged 8 to 12. 5 μg/ml with 91 to 97% protein binding) against all MRSA strains. No antagonism was observed; synergy occurred for one MRSA strain treated with fusidic acid plus tobramycin. PMID:21343445

  19. In vitro activity of fusidic acid (CEM-102, sodium fusidate) against Staphylococcus aureus isolates from cystic fibrosis patients and its effect on the activities of tobramycin and amikacin against Pseudomonas aeruginosa and Burkholderia cepacia.

    Science.gov (United States)

    McGhee, Pamela; Clark, Catherine; Credito, Kim; Beachel, Linda; Pankuch, Glenn A; Appelbaum, Peter C; Kosowska-Shick, Klaudia

    2011-05-01

    We tested the MICs of fusidic acid (CEM-102) plus other agents against 40 methicillin-resistant Staphylococcus aureus (MRSA) isolates from cystic fibrosis patients and the activities of fusidic acid with or without tobramycin or amikacin against Pseudomonas aeruginosa, MRSA, and Burkholderia cepacia isolates from cystic fibrosis patients in a 24-h time-kill study. Fusidic acid was potent (MICs, 0.125 to 0.5 μg/ml; a single 500-mg dose of fusidic acid at 8 h averaged 8 to 12. 5 μg/ml with 91 to 97% protein binding) against all MRSA strains. No antagonism was observed; synergy occurred for one MRSA strain treated with fusidic acid plus tobramycin.

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

    Science.gov (United States)

    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.

  1. Genome-Wide Analysis of Type VI System Clusters and Effectors in Burkholderia Species

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    Thao Thi Nguyen

    2018-02-01

    Full Text Available Type VI secretion system (T6SS has been discovered in a variety of gram-negative bacteria as a versatile weapon to stimulate the killing of eukaryotic cells or prokaryotic competitors. Type VI secretion effectors (T6SEs are well known as key virulence factors for important pathogenic bacteria. In many Burkholderia species, T6SS has evolved as the most complicated secretion pathway with distinguished types to translocate diverse T6SEs, suggesting their essential roles in this genus. Here we attempted to detect and characterize T6SSs and potential T6SEs in target genomes of plant-associated and environmental Burkholderia species based on computational analyses. In total, 66 potential functional T6SS clusters were found in 30 target Burkholderia bacterial genomes, of which 33% possess three or four clusters. The core proteins in each cluster were specified and phylogenetic trees of three components (i.e., TssC, TssD, TssL were constructed to elucidate the relationship among the identified T6SS clusters. Next, we identified 322 potential T6SEs in the target genomes based on homology searches and explored the important domains conserved in effector candidates. In addition, using the screening approach based on the profile hidden Markov model (pHMM of T6SEs that possess markers for type VI effectors (MIX motif (MIX T6SEs, 57 revealed proteins that were not included in training datasets were recognized as novel MIX T6SE candidates from the Burkholderia species. This approach could be useful to identify potential T6SEs from other bacterial genomes.

  2. 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

  3. Fatal Burkholderia pseudomallei Infection Initially Reported as a Bacillus Species, Ohio, 2013

    OpenAIRE

    Doker, Thomas J.; Quinn, Celia L.; Salehi, Ellen D.; Sherwood, Joshua J.; Benoit, Tina J.; Elrod, Mindy Glass; Gee, Jay E.; Shadomy, Sean V.; Bower, William A.; Hoffmaster, Alex R.; Walke, Henry T.; Blaney, David D.; DiOrio, Mary S.

    2014-01-01

    A fatal case of melioidosis was diagnosed in Ohio one month after culture results were initially reported as a Bacillus species. To identify a source of infection and assess risk in patient contacts, we abstracted patient charts; interviewed physicians and contacts; genetically characterized the isolate; performed a Burkholderia pseudomallei antibody indirect hemagglutination assay on household contacts and pets to assess seropositivity; and collected household plant, soil, liquid, and insect...

  4. Nasal Acai Polysaccharides Potentiate Innate Immunity to Protect against Pulmonary Francisella tularensis and Burkholderia pseudomallei Infections

    OpenAIRE

    Skyberg, Jerod A.; Rollins, MaryClare F.; Holderness, Jeff S.; Marlenee, Nicole L.; Schepetkin, Igor A.; Goodyear, Andrew; Dow, Steven W.; Jutila, Mark A.; Pascual, David W.

    2012-01-01

    Pulmonary Francisella tularensis and Burkholderia pseudomallei infections are highly lethal in untreated patients, and current antibiotic regimens are not always effective. Activating the innate immune system provides an alternative means of treating infection and can also complement antibiotic therapies. Several natural agonists were screened for their ability to enhance host resistance to infection, and polysaccharides derived from the Acai berry (Acai PS) were found to have potent abilitie...

  5. Mining Host-Pathogen Protein Interactions to Characterize Burkholderia mallei Infectivity Mechanisms

    Science.gov (United States)

    2015-03-04

    the cytoskeleton, in lysosomes , and in the nuclear lumen. These results were consistent with the experimentally observed pathogen interference with...RESEARCH ARTICLE Mining Host- Pathogen Protein Interactions to Characterize Burkholderia mallei Infectivity Mechanisms Vesna Memišević1, Nela...Bacteriology Division, U.S. Army Medical Research Institute of Infectious Diseases , Fort Detrick, Maryland, United States of America * jaques.reifman.civ

  6. 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

  7. Characterization of the Phthalate Permease OphD from Burkholderia cepacia ATCC 17616†

    OpenAIRE

    Chang, Hung-Kuang; Zylstra, Gerben J.

    1999-01-01

    The ophD gene, encoding a permease for phthalate transport, was cloned from Burkholderia cepacia ATCC 17616. Expression of the gene in Escherichia coli results in the ability to transport phthalate rapidly into the cell. Uptake inhibition experiments show that 4-hydroxyphthalate, 4-chlorophthalate, 4-methylphthalate, and cinchomeronate compete for the phthalate permease. An ophD knockout mutant of 17616 grows slightly more slowly on phthalate but is still able to take up phthalate at rates eq...

  8. 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

  9. Global Analysis of the Burkholderia thailandensis Quorum Sensing-Controlled Regulon

    OpenAIRE

    Majerczyk, Charlotte; Brittnacher, Mitchell; Jacobs, Michael; Armour, Christopher D.; Radey, Mathew; Schneider, Emily; Phattarasokul, Somsak; Bunt, Richard; Greenberg, E. Peter

    2014-01-01

    Burkholderia thailandensis contains three acyl-homoserine lactone quorum sensing circuits and has two additional LuxR homologs. To identify B. thailandensis quorum sensing-controlled genes, we carried out transcriptome sequencing (RNA-seq) analyses of quorum sensing mutants and their parent. The analyses were grounded in the fact that we identified genes coding for factors shown previously to be regulated by quorum sensing among a larger set of quorum-controlled genes. We also found that gene...

  10. The In Vitro Antibiotic Susceptibility of Malaysian Isolates of Burkholderia pseudomallei

    Directory of Open Access Journals (Sweden)

    Norazah Ahmad

    2013-01-01

    Full Text Available Acute melioidosis may present as localised or septicaemic infections and can be fatal if left untreated. Burkholderia pseudomallei resistant to antibiotics used for the treatment of melioidosis had been reported. The aim of this study was to determine the in vitro antibiotic susceptibility patterns of Burkholderia pseudomallei isolated in Malaysia to a panel of antibiotics used for the treatment of melioidosis and also to potential alternative antibiotics such as tigecycline, ampicillin/sulbactam, and piperacillin/tazobactam. A total of 170 Burkholderia pseudomallei isolates were subjected to minimum inhibitory concentration determination using E-test method to eleven antibiotics. All isolates were sensitive to meropenem and piperacillin/tazobactam. For ceftazidime, imipenem, amoxicillin/clavulanic acid, and doxycycline resistance was observed in 1 isolate (0.6% for each of the antibiotics. Trimethoprim/sulfamethoxazole resistance was observed in 17 (10% isolates. For other antibiotics, ampicillin/sulbactam, chloramphenicol, tigecycline, and ciprofloxacin resistance were observed in 1 (0.6%, 6 (3.5%, 60 (35.3% and 98 (57.7% isolates respectively. One isolate B170/06 exhibited resistance to 4 antibiotics, namely, ciprofloxacin, chloramphenicol, trimethoprim/sulfamethoxazole, and tigecycline. In conclusion, the Malaysian isolates were highly susceptible to the current antibiotics used in the treatment of melioidosis in Malaysia. Multiple resistances to the antibiotics used in the maintenance therapy are the cause for a concern.

  11. Global Analysis of the Burkholderia thailandensis Quorum Sensing-Controlled Regulon

    Science.gov (United States)

    Majerczyk, Charlotte; Brittnacher, Mitchell; Jacobs, Michael; Armour, Christopher D.; Radey, Mathew; Schneider, Emily; Phattarasokul, Somsak; Bunt, Richard

    2014-01-01

    Burkholderia thailandensis contains three acyl-homoserine lactone quorum sensing circuits and has two additional LuxR homologs. To identify B. thailandensis quorum sensing-controlled genes, we carried out transcriptome sequencing (RNA-seq) analyses of quorum sensing mutants and their parent. The analyses were grounded in the fact that we identified genes coding for factors shown previously to be regulated by quorum sensing among a larger set of quorum-controlled genes. We also found that genes coding for contact-dependent inhibition were induced by quorum sensing and confirmed that specific quorum sensing mutants had a contact-dependent inhibition defect. Additional quorum-controlled genes included those for the production of numerous secondary metabolites, an uncharacterized exopolysaccharide, and a predicted chitin-binding protein. This study provides insights into the roles of the three quorum sensing circuits in the saprophytic lifestyle of B. thailandensis, and it provides a foundation on which to build an understanding of the roles of quorum sensing in the biology of B. thailandensis and the closely related pathogenic Burkholderia pseudomallei and Burkholderia mallei. PMID:24464461

  12. Global analysis of the Burkholderia thailandensis quorum sensing-controlled regulon.

    Science.gov (United States)

    Majerczyk, Charlotte; Brittnacher, Mitchell; Jacobs, Michael; Armour, Christopher D; Radey, Mathew; Schneider, Emily; Phattarasokul, Somsak; Bunt, Richard; Greenberg, E Peter

    2014-04-01

    Burkholderia thailandensis contains three acyl-homoserine lactone quorum sensing circuits and has two additional LuxR homologs. To identify B. thailandensis quorum sensing-controlled genes, we carried out transcriptome sequencing (RNA-seq) analyses of quorum sensing mutants and their parent. The analyses were grounded in the fact that we identified genes coding for factors shown previously to be regulated by quorum sensing among a larger set of quorum-controlled genes. We also found that genes coding for contact-dependent inhibition were induced by quorum sensing and confirmed that specific quorum sensing mutants had a contact-dependent inhibition defect. Additional quorum-controlled genes included those for the production of numerous secondary metabolites, an uncharacterized exopolysaccharide, and a predicted chitin-binding protein. This study provides insights into the roles of the three quorum sensing circuits in the saprophytic lifestyle of B. thailandensis, and it provides a foundation on which to build an understanding of the roles of quorum sensing in the biology of B. thailandensis and the closely related pathogenic Burkholderia pseudomallei and Burkholderia mallei.

  13. Burkholderia tropica una bacteria con gran potencial parasu uso en la agricultura

    Directory of Open Access Journals (Sweden)

    Hernando José Bolívar-Anillo

    2016-01-01

    Full Text Available El género Burkholderia con más de 90 especies reportadas hasta la fecha, se encuentra dividido en dos grupos mayores filogenéticamente distantes. El primer grupo se encuentra constituido por especies patógenas donde destacan los patógenos oportunistas referidos como el complejo Burkholderia cepacia (Bcc;el otro grupo está conformado por especies no patógenas con habilidades para la promoción del crecimiento vegetal y la rizoremediación. Burkholderia tropica es unabacteria con capacidad de fijar nitrógeno; aislada de la rizósfera, rizoplano, tallo y la raíz de plantas de maíz y caña de azúcar. Además de su capacidad diazotrofa, B. tropica presenta características que permiten catalogarla como una bacteria promotora del crecimiento vegetal, por su capacidad de producir sideróforos, solubilizar fosfatos, producir exo-heteropolisacáridos, además de utilizarse como biocontrol para algunos fitoparásitos, lo que la convierte en una bacteria prometedora para su aplicación en el sector agrícola.

  14. Investigating the Role of the Host Multidrug Resistance Associated Protein Transporter Family in Burkholderia cepacia Complex Pathogenicity Using a Caenorhabditis elegans Infection Model.

    Science.gov (United States)

    Tedesco, Pietro; Visone, Marco; Parrilli, Ermenegilda; Tutino, Maria Luisa; Perrin, Elena; Maida, Isabel; Fani, Renato; Ballestriero, Francesco; Santos, Radleigh; Pinilla, Clemencia; Di Schiavi, Elia; Tegos, George; de Pascale, Donatella

    2015-01-01

    This study investigated the relationship between host efflux system of the non-vertebrate nematode Caenorhabditis elegans and Burkholderia cepacia complex (Bcc) strain virulence. This is the first comprehensive effort to profile host-transporters within the context of Bcc infection. With this aim, two different toxicity tests were performed: a slow killing assay that monitors mortality of the host by intestinal colonization and a fast killing assay that assesses production of toxins. A Virulence Ranking scheme was defined, that expressed the toxicity of the Bcc panel members, based on the percentage of surviving worms. According to this ranking the 18 Bcc strains were divided in 4 distinct groups. Only the Cystic Fibrosis isolated strains possessed profound nematode killing ability to accumulate in worms' intestines. For the transporter analysis a complete set of isogenic nematode single Multidrug Resistance associated Protein (MRP) efflux mutants and a number of efflux inhibitors were interrogated in the host toxicity assays. The Bcc pathogenicity profile of the 7 isogenic C. elegans MRP knock-out strains functionality was classified in two distinct groups. Disabling host transporters enhanced nematode mortality more than 50% in 5 out of 7 mutants when compared to wild type. In particular mrp-2 was the most susceptible phenotype with increased mortality for 13 out 18 Bcc strains, whereas mrp-3 and mrp-4 knock-outs had lower mortality rates, suggesting a different role in toxin-substrate recognition. The use of MRP efflux inhibitors in the assays resulted in substantially increased (>40% on average) mortality of wild-type worms.

  15. Effect of gamma irradiation on Burkholderia thailandensis (Burkholderia pseudomallei surrogate) survival under combinations of pH and NaCl

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Yohan; Kim, Jae-Hun; Byun, Myung-Woo [Team for Radiation Food Science and Biotechnology, Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup, Jeollabuk 580-185 (Korea, Republic of); Choi, Kyoung-Hee [Department of Oral Microbiology, College of Dentistry, Wonkwang University, Iksan, Jeollabuk 570-749 (Korea, Republic of); Lee, Ju-Woon, E-mail: sjwlee@kaeri.re.k [Team for Radiation Food Science and Biotechnology, Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup, Jeollabuk 580-185 (Korea, Republic of)

    2010-04-15

    This study evaluated the effect of gamma irradiation on Burkholderia thailandensis (Burkholderia pseudomallei surrogate; potential bioterrorism agent) survival under different levels of NaCl and pH. B. thailandensis in Luria Bertani broth supplemented with NaCl (0-3%), and pH-adjusted to 4-7 was treated with gamma irradiation (0-0.5 kGy). Surviving cell counts of bacteria were then enumerated on tryptic soy agar. Data for the cell counts were also used to calculate D{sub 10} values (the dose required to reduce 1 log CFU/mL of B. thailandensis). Cell counts of B. thailandensis were decreased (P<0.05) as irradiation dose increased, and no differences (P>=0.05) in cell counts of the bacteria were observed among different levels of NaCl and pH. D{sub 10} values ranged from 0.04 to 0.07 kGy, regardless of NaCl and pH level. These results indicate that low doses of gamma irradiation should be a useful treatment in decreasing the potential bioterrorism bacteria, which may possibly infect humans through foods.

  16. Transfer of eleven species of the genus Burkholderia to the genus Paraburkholderia and proposal of Caballeronia gen. nov. to accommodate twelve species of the genera Burkholderia and Paraburkholderia.

    Science.gov (United States)

    Dobritsa, Anatoly P; Samadpour, Mansour

    2016-08-01

    It has been proposed to split the genus Burkholderia into two genera according to phylogenetic clustering: (1) a genus retaining this name and consisting mainly of animal and plant pathogens and (2) the genus Paraburkholderia including so-called environmental bacteria. The latter genus name has been validly published recently. During the period between the effective and valid publications of the genus name Paraburkholderia, 16 novel species of the genus Burkholderiawere described, but only two of them can be classified as members of this genus based on the emended genus description. Analysis of traits and phylogenetic positions of the other 11 species shows that they belong to the genus Paraburkholderia, and we propose to transfer them to this genus. The reclassified species names are proposed as Paraburkholderia dipogonis comb. nov., Paraburkholderia ginsengiterrae comb. nov., Paraburkholderia humisilvae comb. nov., Paraburkholderia insulsa comb. nov., Paraburkholderia kirstenboschensis comb. nov., Paraburkholderia metalliresistens comb. nov., Paraburkholderia monticola comb. nov., Paraburkholderia panaciterrae comb. nov., Paraburkholderia rhizosphaerae comb. nov., Paraburkholderia solisilvae comb. nov. and Paraburkholderia susongensis comb. nov. The remaining three species are transferred to the new genus Caballeronia gen. nov. proposed to accommodate twelve species of the genera Burkholderia and Paraburkholderia forming a distinctive clade in phylogenetic trees. The new genus members are Caballeronia choica comb. nov., Caballeronia cordobensis comb. nov., Caballeronia glathei comb. nov., Caballeronia grimmiae comb. nov., Caballeronia humi comb. nov., Caballeronia megalochromosomata comb. nov., Caballeronia jiangsuensis comb. nov., Caballeronia sordidicola comb. nov., Caballeronia telluris comb. nov., Caballeronia terrestris comb. nov., Caballeronia udeis comb. nov., and Caballeronia zhejiangensis comb. nov.

  17. Effect of gamma irradiation on Burkholderia thailandensis ( Burkholderia pseudomallei surrogate) survival under combinations of pH and NaCl

    Science.gov (United States)

    Yoon, Yohan; Kim, Jae-Hun; Byun, Myung-Woo; Choi, Kyoung-Hee; Lee, Ju-Woon

    2010-04-01

    This study evaluated the effect of gamma irradiation on Burkholderia thailandensis ( Burkholderia pseudomallei surrogate; potential bioterrorism agent) survival under different levels of NaCl and pH. B. thailandensis in Luria Bertani broth supplemented with NaCl (0-3%), and pH-adjusted to 4-7 was treated with gamma irradiation (0-0.5 kGy). Surviving cell counts of bacteria were then enumerated on tryptic soy agar. Data for the cell counts were also used to calculate D10 values (the dose required to reduce 1 log CFU/mL of B. thailandensis). Cell counts of B. thailandensis were decreased ( P<0.05) as irradiation dose increased, and no differences ( P≥0.05) in cell counts of the bacteria were observed among different levels of NaCl and pH. D10 values ranged from 0.04 to 0.07 kGy, regardless of NaCl and pH level. These results indicate that low doses of gamma irradiation should be a useful treatment in decreasing the potential bioterrorism bacteria, which may possibly infect humans through foods.

  18. Spatio-temporal responses of Arabidopsis leaves in photosynthetic performance and metabolite contents to Burkholderia phytofirmans PsJN

    Directory of Open Access Journals (Sweden)

    Fan eSu

    2016-03-01

    Full Text Available A valuable strategy to improve crop yield consists in the use of plant growth-promoting rhizobacteria (PGPRs. However, the influence of PGPR colonization on plant physiology is largely unknown. PGPR Burkholderia phytofirmans strain PsJN (Bp PsJN colonized only Arabidopsis thaliana roots after seed or soil inoculation. Foliar bacteria were detected only after leaf infiltration. Since different bacterial times of presence and/or locations in host plant could lead to different plant physiological responses, photosynthesis and metabolite profiles in A. thaliana leaves were thus investigated following leaf, root or seed inoculation with Bp PsJN. Only Bp PsJN leaf colonization transiently decreased cyclic electron transport and effective quantum yield of photosystem I (PSI, and prevented a decrease in net photosynthesis and stomatal opening compared to the corresponding control. Metabolomic analysis revealed that soluble sugars, amino acids or their derivatives accumulated differently in all Bp PsJN-inoculated plants. Octanoic acid accumulated only in case of inoculated plants. Modifications in vitamin, organic acid such as tricarboxylic acid intermediates, and hormone amounts were dependent on bacterial time of presence and location. Additionally, a larger array of amino acids and hormones (auxin, cytokinin, abscisic acid were modified by seed inoculation with Bp PsJN. Our work thereby provides evidence that relative short-term inoculation with Bp PsJN altered physiological status of A.thaliana leaves, whereas long-term bacterization triggered modifications on a larger set of metabolites. Our data highlighted the changes displayed during this plant-microbe interaction to trigger physiological and metabolic responses that could explain the increase in plant growth or stress tolerance conferred by the presence of Bp PsJN.

  19. Genomic characterization of Burkholderia pseudomallei isolates selected for medical countermeasures testing: comparative genomics associated with differential virulence.

    Directory of Open Access Journals (Sweden)

    Jason W Sahl

    Full Text Available Burkholderia pseudomallei is the causative agent of melioidosis and a potential bioterrorism agent. In the development of medical countermeasures against B. pseudomallei infection, the US Food and Drug Administration (FDA animal Rule recommends using well-characterized strains in animal challenge studies. In this study, whole genome sequence data were generated for 6 B. pseudomallei isolates previously identified as candidates for animal challenge studies; an additional 5 isolates were sequenced that were associated with human inhalational melioidosis. A core genome single nucleotide polymorphism (SNP phylogeny inferred from a concatenated SNP alignment from the 11 isolates sequenced in this study and a diverse global collection of isolates demonstrated the diversity of the proposed Animal Rule isolates. To understand the genomic composition of each isolate, a large-scale blast score ratio (LS-BSR analysis was performed on the entire pan-genome; this demonstrated the variable composition of genes across the panel and also helped to identify genes unique to individual isolates. In addition, a set of ~550 genes associated with pathogenesis in B. pseudomallei were screened against the 11 sequenced genomes with LS-BSR. Differential gene distribution for 54 virulence-associated genes was observed between genomes and three of these genes were correlated with differential virulence observed in animal challenge studies using BALB/c mice. Differentially conserved genes and SNPs associated with disease severity were identified and could be the basis for future studies investigating the pathogenesis of B. pseudomallei. Overall, the genetic characterization of the 11 proposed Animal Rule isolates provides context for future studies involving B. pseudomallei pathogenesis, differential virulence, and efficacy to therapeutics.

  20. Burkholderia thailandensis harbors two identical rhl gene clusters responsible for the biosynthesis of rhamnolipids

    Directory of Open Access Journals (Sweden)

    Woods Donald E

    2009-12-01

    Full Text Available Abstract Background Rhamnolipids are surface active molecules composed of rhamnose and β-hydroxydecanoic acid. These biosurfactants are produced mainly by Pseudomonas aeruginosa and have been thoroughly investigated since their early discovery. Recently, they have attracted renewed attention because of their involvement in various multicellular behaviors. Despite this high interest, only very few studies have focused on the production of rhamnolipids by Burkholderia species. Results Orthologs of rhlA, rhlB and rhlC, which are responsible for the biosynthesis of rhamnolipids in P. aeruginosa, have been found in the non-infectious Burkholderia thailandensis, as well as in the genetically similar important pathogen B. pseudomallei. In contrast to P. aeruginosa, both Burkholderia species contain these three genes necessary for rhamnolipid production within a single gene cluster. Furthermore, two identical, paralogous copies of this gene cluster are found on the second chromosome of these bacteria. Both Burkholderia spp. produce rhamnolipids containing 3-hydroxy fatty acid moieties with longer side chains than those described for P. aeruginosa. Additionally, the rhamnolipids produced by B. thailandensis contain a much larger proportion of dirhamnolipids versus monorhamnolipids when compared to P. aeruginosa. The rhamnolipids produced by B. thailandensis reduce the surface tension of water to 42 mN/m while displaying a critical micelle concentration value of 225 mg/L. Separate mutations in both rhlA alleles, which are responsible for the synthesis of the rhamnolipid precursor 3-(3-hydroxyalkanoyloxyalkanoic acid, prove that both copies of the rhl gene cluster are functional, but one contributes more to the total production than the other. Finally, a double ΔrhlA mutant that is completely devoid of rhamnolipid production is incapable of swarming motility, showing that both gene clusters contribute to this phenotype. Conclusions Collectively, these

  1. Insights into selenite reduction and biogenesis of elemental selenium nanoparticles by two environmental isolates of Burkholderia fungorum.

    Science.gov (United States)

    Khoei, Nazanin Seyed; Lampis, Silvia; Zonaro, Emanuele; Yrjälä, Kim; Bernardi, Paolo; Vallini, Giovanni

    2017-01-25

    Microorganisms capable of transforming toxic selenium oxyanions into non-toxic elemental selenium (Se°) may be considered as biocatalysts for the production of selenium nanoparticles (SeNPs), eventually exploitable in different biotechnological applications. Two Burkholderia fungorum strains (B. fungorum DBT1 and B. fungorum 95) were monitored during their growth for both capacity and efficiency of selenite (SeO 3 2- ) reduction and elemental selenium formation. Both strains are environmental isolates in origin: B. fungorum DBT1 was previously isolated from an oil refinery drainage, while B. fungorum 95 has been enriched from inner tissues of hybrid poplars grown in a soil contaminated by polycyclic aromatic hydrocarbons. Our results showed that B. fungorum DBT1 is able to reduce 0.5mM SeO 3 2- to Se° when cultured aerobically in liquid medium at 27°C, while B. fungorum 95 can reduce more than 1mM SeO 3 2- to Se° within 96h under the same growth conditions, with the appearance of SeNPs in cultures of both bacterial strains. Biogenic SeNPs were spherical, with pH-dependent charge and an average hydrodynamic diameter of 170nm and 200nm depending on whether they were produced by B. fungorum 95 or B. fungorum DBT1, respectively. Electron microscopy analyses evidenced that Se nanoparticles occurred intracellularly and extracellularly. The mechanism of SeNPs formation can be tentatively attributed to cytoplasmic enzymatic activation mediated by electron donors. Biogenic nanoparticles were then probably released outside the bacterial cells as a consequence of a secretory process or cell lysis. Nevertheless, formation of elemental selenium nanoparticles under aerobic conditions by B. fungorum DBT1 and B. fungorum 95 is likely due to intracellular reduction mechanisms. Biomedical and other high tech sectors might exploit these biogenic nanoparticles in the near future, once fully characterized and tested for their multiple properties. Copyright © 2016 Elsevier B.V. All

  2. Isolation of Burkholderia cepacia JB12 from lead- and cadmium-contaminated soil and its potential in promoting phytoremediation with tall fescue and red clover.

    Science.gov (United States)

    Jin, Zhong Min; Sha, Wei; Zhang, Yan Fu; Zhao, Jing; Ji, Hongyang

    2013-07-01

    Phytoremediation combined with suitable microorganisms and biodegradable chelating agents can be a means of reclaiming lands contaminated by toxic heavy metals. We investigated the ability of a lead- and cadmium-resistant bacterial strain (JB12) and the biodegradable chelator ethylenediamine-N,N'-disuccinic acid (EDDS) to improve absorption of these metals from soil by tall fescue and red clover. Strain JB12 was isolated from contaminated soil samples, analysed for lead and cadmium resistance, and identified as Burkholderia cepacia. Tall fescue and red clover were grown in pots to which we added JB12, (S,S)-EDDS, combined JB12 and EDDS, or water only. Compared with untreated plants, the biomass of plants treated with JB12 was significantly increased. Concentrations of lead and cadmium in JB12-treated plants increased significantly, with few exceptions. Plants treated with EDDS responded variably, but in those treated with combined EDDS and JB12, heavy metal concentrations increased significantly in tall fescue and in the aboveground parts of red clover. We conclude that JB12 is resistant to lead and cadmium. Its application to the soil improved the net uptake of these heavy metals by experimental plants. The potential for viable phytoremediation of lead- and cadmium-polluted soils with tall fescue and red clover combined with JB12 was further enhanced by the addition of EDDS.

  3. xylA and xylB overexpression as a successful strategy for improving xylose utilization and poly-3-hydroxybutyrate production in Burkholderia sacchari.

    Science.gov (United States)

    Guamán, Linda P; Oliveira-Filho, Edmar R; Barba-Ostria, Carlos; Gomez, José G C; Taciro, Marilda K; da Silva, Luiziana Ferreira

    2018-03-01

    Despite the versatility and many advantages of polyhydroxyalkanoates as petroleum-based plastic substitutes, their higher production cost compared to petroleum-based polymers has historically limited their large-scale production. One appealing approach to reducing production costs is to employ less expensive, renewable feedstocks. Xylose, for example is an abundant and inexpensive carbon source derived from hemicellulosic residues abundant in agro-industrial waste (sugarcane bagasse hemicellulosic hydrolysates). In this work, the production of poly-3-hydroxybutyrate P(3HB) from xylose was studied to develop technologies for conversion of agro-industrial waste into high-value chemicals and biopolymers. Specifically, this work elucidates the organization of the xylose assimilation operon of Burkholderia sacchari, a non-model bacterium with high capacity for P(3HB) accumulation. Overexpression of endogenous xylose isomerase and xylulokinase genes was successfully assessed, improving both specific growth rate and P(3HB) production. Compared to control strain (harboring pBBR1MCS-2), xylose utilization in the engineered strain was substantially improved with 25% increase in specific growth rate, 34% increase in P(3HB) production, and the highest P(3HB) yield from xylose reported to date for B. sacchari (Y P3HB/Xil  = 0.35 g/g). This study highlights that xylA and xylB overexpression is an effective strategy to improve xylose utilization and P(3HB) production in B. sacchari.

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

    Science.gov (United States)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  6. Nitrous oxide emission potentials of Burkholderia species isolated from the leaves of a boreal peat moss Sphagnum fuscum.

    Science.gov (United States)

    Nie, Yanxia; Li, Li; Wang, Mengcen; Tahvanainen, Teemu; Hashidoko, Yasuyuki

    2015-01-01

    Using a culture-based nitrous oxide (N2O) emission assay, three active N2O emitters were isolated from Sphagnum fuscum leaves and all identified as members of Burkholderia. These isolates showed N2O emission in the medium supplemented with [Formula: see text] but not with [Formula: see text], and Burkholderia sp. SF-E2 showed the most efficient N2O emission (0.20 μg·vial(-1)·day(-1)) at 1.0 mM KNO3. In Burkholderia sp. SF-E2, the optimum pH for N2O production was 5.0, close to that of the phyllosphere of Sphagnum mosses, while the optimum temperature was uniquely over 30 °C. The stimulating effect of additional 1.5 mM sucrose on N2O emission was ignorable, but Burkholderia sp. SF-E2 upon exposure to 100 mg·L(-1) E-caffeic acid showed uniquely 67-fold higher N2O emission. All of the three N2O emitters were negative in both acetylene inhibition assay and PCR assay for nosZ-detection, suggesting that N2O reductase or the gene itself is missing in the N2O-emitting Burkholderia.

  7. Rhizonin A from Burkholderia sp. KCTC11096 and Its Growth Promoting Role in Lettuce Seed Germination

    Directory of Open Access Journals (Sweden)

    Sang-Mo Kang

    2012-07-01

    Full Text Available We isolated and identified a gibberellin-producing Burkholderia sp. KCTC 11096 from agricultural field soils. The culture filtrate of plant growth promoting rhizobacteria (PGPR significantly increased the germination and growth of lettuce and Chinese cabbage seeds. The ethyl acetate extract of the PGPR culture showed significantly higher rate of lettuce seed germination and growth as compared to the distilled water treated control. The ethyl acetate fraction of the Burkholderia sp. was subjected to bioassay-guided isolation and we obtained for the first time from a Burkholderia sp. the plant growth promoting compound rhizonin A (1, which was characterized through NMR and MS techniques. Application of various concentrations of 1 significantly promoted the lettuce seed germination as compared to control.

  8. Chemotaxis and adherence to fungal surfaces are key components of the behavioral response of Burkholderia terrae BS001 to two selected soil fungi.

    Science.gov (United States)

    Haq, Irshad Ul; Calixto, Renata Oliveira da Rocha; Yang, Pu; Dos Santos, Giulia Maria Pires; Barreto-Bergter, Eliana; van Elsas, Jan Dirk

    2016-11-01

    Burkholderia terrae BS001 has previously been proposed to be a 'generalist' associate of soil fungi, but its strategies of interaction have been largely ignored. Here, we studied the chemotactic behavior of B. terrae BS001 towards Lyophyllum sp. strain Karsten and Trichoderma asperellum 302 and the role of fungal surface molecules in their physical interaction with the bacteria. To assess the involvement of the type 3 secretion system (T3SS), wild-type strain BS001 and T3SS mutant strain BS001-ΔsctD were used in the experiments. First, the two fungi showed divergent behavior when confronted with B. terrae BS001 on soil extract agar medium. Lyophyllum sp. strain Karsten revealed slow growth towards the bacterium, whereas T. asperellum 302 grew avidly over it. Both on soil extract and M9 agar, B. terrae BS001 and BS001-ΔsctD moved chemotactically towards the hyphae of both fungi, with a stronger response to Lyophyllum sp. strain Karsten than to T. asperellum 302. The presence of a progressively increasing glycerol level in the M9 agar enhanced the level of movement. Different oxalic acid concentrations exerted varied effects, with a significantly raised chemotactic response at lower, and a subdued response at higher concentrations. Testing of the adherence of B. terrae BS001 and BS001-ΔsctD to Lyophyllum sp. strain Karsten and to cell envelope-extracted ceramide monohexosides (CMHs) revealed that CMHs in both conidia and hyphae could bind strain BS001 cells. As BS001-ΔsctD adhered significantly less to the CMHs than BS001, the T3SS was presumed to have a role in the interaction. In contrast, such avid adherence was not detected with T. asperellum 302. Thus, B. terrae BS001 shows a behavior characterized by swimming towards Lyophyllum sp. strain Karsten and T. asperellum 302 and attachment to the CMH moiety in the cell envelope, in particular of the former. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  9. The cep quorum-sensing system of Burkholderia cepacia H111 controls biofilm formation and swarming motility

    DEFF Research Database (Denmark)

    Huber, B.; Riedel, K.; Hentzer, Morten

    2001-01-01

    Burkholderia cepacia and Pseudomonas aeruginosa often co-exist as mixed biofilms in the lungs of patients suffering from cystic fibrosis (CF). Here, the isolation of random mini-Tn5 insertion mutants of B. cepacia H111 defective in biofilm formation on an abiotic surface is reported. It is demons......Burkholderia cepacia and Pseudomonas aeruginosa often co-exist as mixed biofilms in the lungs of patients suffering from cystic fibrosis (CF). Here, the isolation of random mini-Tn5 insertion mutants of B. cepacia H111 defective in biofilm formation on an abiotic surface is reported...

  10. Burkholderia humptydooensis sp. nov., A Burkholderia thailandensis-Like Species and the Fifth Member of the pseudomallei Complex

    Science.gov (United States)

    2016-06-02

    biochemical pattern to B. pseudomallei and B. thailandensis were co-64 isolated with B. pseudomallei on Ashdown’s selective agar. To determine the...three B. ubonensis strains) (see SI doc and Fig. S2). 157 Minimum inhibitory concentrations (MICs) were determined by broth microdilution 158...rifampicin (0.0625–8 mg/L), chloramphenicol (0.5–64 mg/L), 162 trimethoprim /sulfamethoxazole (0.25–32/4.75–608 mg/L), streptomycin (0.25–32 mg/L

  11. Characterization of Burkholderia rhizoxinica and B. endofungorum isolated from clinical specimens.

    Directory of Open Access Journals (Sweden)

    Jay E Gee

    Full Text Available Eight isolates submitted to CDC from 1989 to 2006 from clinical specimens were initially identified as members of the genus Burkholderia based on preliminary cellular fatty acid analysis and/or 16S rRNA gene sequencing. With the recent descriptions of the new species B. rhizoxinica and B. endofungorum, which are considered endosymbiotic bacteria in Rhizopus microsporus fungi, we now identify seven of these clinical isolates as B. rhizoxinica and one as B. endofungorum based on biochemical testing, 16s rRNA, and DNA-DNA hybridization results. We also further characterize these isolates by assessing toxin production and/or by multiple locus sequence typing.

  12. The promise of bacteriophage therapy for Burkholderia cepacia complex respiratory infections.

    Directory of Open Access Journals (Sweden)

    Diana Dawn Semler

    2012-01-01

    Full Text Available In recent times, increased attention has been given to evaluating the efficacy of phage therapy, especially in scenarios where the bacterial infectious agent of interest is highly antibiotic resistant. In this regard, phage therapy is especially applicable to infections caused by the Burkholderia cepacia complex (BCC since members of the BCC are antibiotic pan-resistant. Current studies in BCC phage therapy are unique from many other avenues of phage therapy research in that the research is not only comprised of phage isolation, in vitro phage characterization and in vivo infection model efficacy, but also adapting aerosol drug delivery techniques to aerosol phage formulation delivery and storage.

  13. Investigating early stages of biocorrosion with XPS: AISI 304 stainless steel exposed to Burkholderia species

    Science.gov (United States)

    Johansson, Leena-Sisko; Saastamoinen, Tuomas

    1999-04-01

    We have investigated the interactions of an exopolymer-producing bacteria, Burkholderia sp. with polished AISI 304 stainless steel substrates using X-ray photoelectron spectroscopy (XPS). Steel coupons were exposed to the pure bacteria culture in a specially designed flowcell for 6 h during which the experiment was monitored in situ with an optical microscope. XPS results verified the formation of biofilm containing extracellular polymer on all the samples exposed to bacteria. Sputter results indicated that some ions needed for metabolic processes were trapped within the biofilm. Changes in the relative Fe concentration and Fe 2p peak shape indicated that also iron had accumulated into the biofilm.

  14. Eradication of Burkholderia cepacia Using Inhaled Aztreonam Lysine in Two Patients with Bronchiectasis

    Directory of Open Access Journals (Sweden)

    A. Iglesias

    2014-01-01

    Full Text Available There are not many articles about the chronic bronchial infection/colonization in patients with underlying lung disease other than cystic fibrosis (CF, especially with non-CF bronchiectasis (NCFBQ. The prevalence of B. cepacia complex is not well known in NCFBQ. The vast majority of published clinical data on Burkholderia infection in individuals with CF is comprised of uncontrolled, anecdotal, and/or single center experiences, and no consensus has emerged regarding treatment. We present two cases diagnosed with bronchiectasis (BQ of different etiology, with early pulmonary infection by B. cepacia complex, which was eradicated with inhaled aztreonam lysine.

  15. Molecular characterization of a novel family VIII esterase from burkholderia multivorans UWC10

    CSIR Research Space (South Africa)

    Rashamuse, KJ

    2007-02-01

    Full Text Available et al., 1992). Here we report the cloning, purification, and 3D model of a novel family VIII esterase from Burkholderia multivorans UWC10. To our knowledge no report of esterolytic activity from B. multivorans is currently available. METHODOLOGY... stream_source_info Rashamuse1_2007_d.pdf.txt stream_content_type text/plain stream_size 9884 Content-Encoding UTF-8 stream_name Rashamuse1_2007_d.pdf.txt Content-Type text/plain; charset=UTF-8 Molecular Characterization...

  16. Expression of a mineral phosphate solubilizing gene from Erwinia herbicola in two rhizobacterial strains.

    Science.gov (United States)

    Rodríguez, H; Gonzalez, T; Selman, G

    2001-11-30

    A genetic construction was carried out using the broad host range vector pKT230 and plasmid pMCG898, which encodes the Erwinia herbicola pyrroloquinoline quinone (PQQ) synthase, a gene involved in mineral phosphate solubilization (mps). The final construction was transformed and expressed in Escherichia coli MC1061, and the recombinant plasmids were transferred to Burkholderia cepacia IS-16 and Pseudomonas sp. PSS recipient cells by conjugation. Clones containing recombinant plasmids produced higher clearing halos in plates with insoluble phosphate as the unique (P) source, in comparison with those of strains without plasmids, demonstrating the heterologous expression of the E. herbicola gene in the recipient strains. This genetic manipulation allowed the increase in mps ability of both strains, enhancing their potentialities as growth promoters of agricultural crops. These results represent the first report on the application of the recombinant DNA methodology for the obtaining of improved phosphate solubilizing ability from rhizobacterial strains for biofertilization purposes.

  17. Degradation of toluene and trichloroethylene by Burkholderia cepacia G4 in growth-limited fed-batch culture

    NARCIS (Netherlands)

    Mars, Astrid E.; Houwing, Joukje; Dolfing, Jan; Janssen, Dick B.

    Burkholderia (Pseudomonas) cepacia G4 was cultivated in a fed-batch bioreactor on either toluene or toluene plus trichloroethylene (TCE), The culture was allowed to reach a constant cell density under conditions in which the amount of toluene supplied equals the maintenance energy demand of the

  18. South African Papilionoid Legumes Are Nodulated by Diverse Burkholderia with Unique Nodulation and Nitrogen-Fixation Loci

    Science.gov (United States)

    Beukes, Chrizelle W.; Venter, Stephanus N.; Law, Ian J.; Phalane, Francina L.; Steenkamp, Emma T.

    2013-01-01

    The root-nodule bacteria of legumes endemic to the Cape Floristic Region are largely understudied, even though recent reports suggest the occurrence of nodulating Burkholderia species unique to the region. In this study, we considered the diversity and evolution of nodulating Burkholderia associated with the endemic papilionoid tribes Hypocalypteae and Podalyrieae. We identified distinct groups from verified rhizobial isolates by phylogenetic analyses of the 16S rRNA and recA housekeeping gene regions. In order to gain insight into the evolution of the nodulation and diazotrophy of these rhizobia we analysed the genes encoding NifH and NodA. The majority of these 69 isolates appeared to be unique, potentially representing novel species. Evidence of horizontal gene transfer determining the symbiotic ability of these Cape Floristic Region isolates indicate evolutionary origins distinct from those of nodulating Burkholderia from elsewhere in the world. Overall, our findings suggest that Burkholderia species associated with fynbos legumes are highly diverse and their symbiotic abilities have unique ancestries. It is therefore possible that the evolution of these bacteria is closely linked to the diversification and establishment of legumes characteristic of the Cape Floristic Region. PMID:23874611

  19. Reassessment of the taxonomic position of Burkholderia andropogonis and description of Robbsia andropogonis gen. nov., comb. nov.

    Science.gov (United States)

    Lopes-Santos, Lucilene; Castro, Daniel Bedo Assumpção; Ferreira-Tonin, Mariana; Corrêa, Daniele Bussioli Alves; Weir, Bevan Simon; Park, Duckchul; Ottoboni, Laura Maria Mariscal; Neto, Júlio Rodrigues; Destéfano, Suzete Aparecida Lanza

    2017-06-01

    The phylogenetic classification of the species Burkholderia andropogonis within the Burkholderia genus was reassessed using 16S rRNA gene phylogenetic analysis and multilocus sequence analysis (MLSA). Both phylogenetic trees revealed two main groups, named A and B, strongly supported by high bootstrap values (100%). Group A encompassed all of the Burkholderia species complex, whi.le Group B only comprised B. andropogonis species, with low percentage similarities with other species of the genus, from 92 to 95% for 16S rRNA gene sequences and 83% for conserved gene sequences. Average nucleotide identity (ANI), tetranucleotide signature frequency, and percentage of conserved proteins POCP analyses were also carried out, and in the three analyses B. andropogonis showed lower values when compared to the other Burkholderia species complex, near 71% for ANI, from 0.484 to 0.724 for tetranucleotide signature frequency, and around 50% for POCP, reinforcing the distance observed in the phylogenetic analyses. Our findings provide an important insight into the taxonomy of B. andropogonis. It is clear from the results that this bacterial species exhibits genotypic differences and represents a new genus described herein as Robbsia andropogonis gen. nov., comb. nov.

  20. 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.

  1. Systematic review and consensus guidelines for environmental sampling of Burkholderia pseudomallei.

    Directory of Open Access Journals (Sweden)

    Direk Limmathurotsakul

    Full Text Available Burkholderia pseudomallei, a Tier 1 Select Agent and the cause of melioidosis, is a Gram-negative bacillus present in the environment in many tropical countries. Defining the global pattern of B. pseudomallei distribution underpins efforts to prevent infection, and is dependent upon robust environmental sampling methodology. Our objective was to review the literature on the detection of environmental B. pseudomallei, update the risk map for melioidosis, and propose international consensus guidelines for soil sampling.An international working party (Detection of Environmental Burkholderia pseudomallei Working Party (DEBWorP was formed during the VIth World Melioidosis Congress in 2010. PubMed (January 1912 to December 2011 was searched using the following MeSH terms: pseudomallei or melioidosis. Bibliographies were hand-searched for secondary references. The reported geographical distribution of B. pseudomallei in the environment was mapped and categorized as definite, probable, or possible. The methodology used for detecting environmental B. pseudomallei was extracted and collated. We found that global coverage was patchy, with a lack of studies in many areas where melioidosis is suspected to occur. The sampling strategies and bacterial identification methods used were highly variable, and not all were robust. We developed consensus guidelines with the goals of reducing the probability of false-negative results, and the provision of affordable and 'low-tech' methodology that is applicable in both developed and developing countries.The proposed consensus guidelines provide the basis for the development of an accurate and comprehensive global map of environmental B. pseudomallei.

  2. The Burkholderia cepacia rpoE gene is not involved in exopolysaccharide production and onion pathogenicity.

    Science.gov (United States)

    Devescovi, Giulia; Venturi, Vittorio

    2006-03-01

    Burkholderia cepacia was originally described as the causative agent of bacterial rot of onions, and it has now emerged as an important opportunistic pathogen causing severe chronic lung infections in patients having cystic fibrosis. Burkholderia cepacia is now classified into nine very closely related species (previously designated as genomovars), all of which have been isolated from both environmental and clinical sources and are collectively known as the B. cepacia complex. The alternative extracytoplasmic function sigma factor, sigmaE, has been determined in several bacterial species as making substantial contributions to bacterial survival under stress conditions. Here, we report the identification and characterization of the rpoE gene, encoding sigmaE, of B. cepacia. It is highly similar to sigmaE of other bacteria, including Escherichia coli and Pseudomonas aeruginosa. Studies using an rpoE knockout mutant of B. cepacia revealed that many stress adaptations, including osmotic, oxidative, desiccation, carbon, and nitrogen stress, were independent of sigmaE. Similarly, biofilm formation; production of exopolysaccharides, N-acyl homoserine lactones, and several exoenzymes; and onion pathogenicity were not affected by the absence of sigmaE. In contrast, sigmaE contributed to the adaptation to heat stress and phosphate starvation.

  3. Inactivation of Burkholderia pseudomallei on environmental surfaces using spray-applied, common liquid disinfectants.

    Science.gov (United States)

    Calfee, M W; Wendling, M

    2015-11-01

    Five commercially available liquid antimicrobials were evaluated for their ability to decontaminate common environmental surface materials, contaminated with Burkholderia pseudomallei, using a spray-based disinfectant delivery procedure. Tests were conducted at both an ambient temperature (c. 20°C) and a lower temperature (c. 12°C) condition. Nonporous materials (glass and aluminium) were more easily decontaminated than porous materials (wood, concrete and carpet). Citric acid (1%) demonstrated poor efficacy in all test conditions. Bleach (pH-adjusted), ethanol (70%), quaternary ammonium and PineSol®, demonstrated high (>6 log10 reduction) efficacies on glass and aluminium at both temperatures, but achieved varying results for wood, carpet and concrete. Temperature had minimal effect on decontamination efficacy during these tests. Much of the antimicrobial efficacy data for pathogenic micro-organisms are generated with testing that utilizes hard nonporous surface materials. These data are not directly translatable for decontaminant selection following an incident whereby complex and porous environmental surfaces are contaminated. This study presents efficacy data for spray-applied antimicrobial liquids, when used to decontaminate common environmental surfaces contaminated with Burkholderia pseudomallei. These data can help responders develop effective remediation strategies following an environmental contamination incident involving B. pseudomallei. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.

  4. Functional characterisation of Burkholderia pseudomallei biotin protein ligase: A toolkit for anti-melioidosis drug development.

    Science.gov (United States)

    Bond, Thomas E H; Sorenson, Alanna E; Schaeffer, Patrick M

    2017-06-01

    Burkholderia pseudomallei (Bp) is the causative agent of melioidosis. The bacterium is responsible for 20% of community-acquired sepsis cases and 40% of sepsis-related mortalities in northeast Thailand, and is intrinsically resistant to aminoglycosides, macrolides, rifamycins, cephalosporins, and nonureidopenicillins. There is no vaccine and its diagnosis is problematic. Biotin protein ligase (BirA) which is essential for fatty acid synthesis has been proposed as a drug target in bacteria. Very few bacterial BirA have been characterized, and a better understanding of these enzymes is necessary to further assess their value as drug targets. BirA within the Burkholderia genus have not yet been investigated. We present for the first time the cloning, expression, purification and functional characterisation of the putative Bp BirA and orthologous B. thailandensis (Bt) biotin carboxyl carrier protein (BCCP) substrate. A GFP-tagged Bp BirA was produced and applied for the development of a high-throughput (HT) assay based on our differential scanning fluorimetry of GFP-tagged proteins (DSF-GTP) principle as well as an electrophoretic mobility shift assay. Our biochemical data in combination with the new HT DSF-GTP and biotinylation activity assay could facilitate future drug screening efforts against this drug-resistant organism. Copyright © 2017 Elsevier GmbH. All rights reserved.

  5. Comparative and bioinformatics analyses of pathogenic bacterial secretomes identified by mass spectrometry in Burkholderia species.

    Science.gov (United States)

    Nguyen, Thao Thi; Chon, Tae-Soo; Kim, Jaehan; Seo, Young-Su; Heo, Muyoung

    2017-07-01

    Secreted proteins (secretomes) play crucial roles during bacterial pathogenesis in both plant and human hosts. The identification and characterization of secretomes in the two plant pathogens Burkholderia glumae BGR1 and B. gladioli BSR3, which cause diseases in rice such as seedling blight, panicle blight, and grain rot, are important steps to not only understand the disease-causing mechanisms but also find remedies for the diseases. Here, we identified two datasets of secretomes in B. glumae BGR1 and B. gladioli BSR3, which consist of 118 and 111 proteins, respectively, using mass spectrometry approach and literature curation. Next, we characterized the functional properties, potential secretion pathways and sequence information properties of secretomes of two plant pathogens in a comparative analysis by various computational approaches. The ratio of potential non-classically secreted proteins (NCSPs) to classically secreted proteins (CSPs) in B. glumae BGR1 was greater than that in B. gladioli BSR3. For CSPs, the putative hydrophobic regions (PHRs) which are essential for secretion process of CSPs were screened in detail at their N-terminal sequences using hidden Markov model (HMM)-based method. Total 31 pairs of homologous proteins in two bacterial secretomes were indicated based on the global alignment (identity ≥ 70%). Our results may facilitate the understanding of the species-specific features of secretomes in two plant pathogenic Burkholderia species.

  6. The use of nanoscale visible light-responsive photocatalyst TiO2-Pt for the elimination of soil-borne pathogens.

    Directory of Open Access Journals (Sweden)

    Ya-Lei Chen

    Full Text Available Exposure to the soil-borne pathogens Burkholderia pseudomallei and Burkholderia cenocepacia can lead to severe infections and even mortality. These pathogens exhibit a high resistance to antibiotic treatments. In addition, no licensed vaccine is currently available. A nanoscale platinum-containing titania photocatalyst (TiO(2-Pt has been shown to have a superior visible light-responsive photocatalytic ability to degrade chemical contaminants like nitrogen oxides. The antibacterial activity of the catalyst and its potential use in soil pathogen control were evaluated. Using the plating method, we found that TiO(2-Pt exerts superior antibacterial performance against Escherichia coli compared to other commercially available and laboratory prepared ultraviolet/visible light-responsive titania photocatalysts. TiO(2-Pt-mediated photocatalysis also affectively eliminates the soil-borne bacteria B. pseudomallei and B. cenocepacia. An air pouch infection mouse model further revealed that TiO(2-Pt-mediated photocatalysis could reduce the pathogenicity of both strains of bacteria. Unexpectedly, water containing up to 10% w/v dissolved soil particles did not reduce the antibacterial potency of TiO(2-Pt, suggesting that the TiO(2-Pt photocatalyst is suitable for use in soil-contaminated environments. The TiO(2-Pt photocatalyst exerted superior antibacterial activity against a broad spectrum of human pathogens, including B. pseudomallei and B. cenocepacia. Soil particles (<10% w/v did not significantly reduce the antibacterial activity of TiO(2-Pt in water. These findings suggest that the TiO(2-Pt photocatalyst may have potential applications in the development of bactericides for soil-borne pathogens.

  7. Bukholderia strains promote Mimosa spp. growth but not Macroptilium atropurpureum

    Directory of Open Access Journals (Sweden)

    Kaliane Sírio Araújo

    Full Text Available ABSTRACT The aim of this study was to evaluate the relationship and symbiotic efficiency of 14 strains of Burkholderia isolated from rupestrian grasslands, using M. atropurpureum and Mimosa tenuiflora as trap plants, with the species M. atropurpureum, Mimosa bimucronata and M. foliolosa. For the nodulation and symbiotic efficiency test in M. atropurpureum, long-neck bottles containing nutrient solution were used. The experiments with Mimosa spp. were carried out in tubes containing vermiculite (160 cm3 and sand (80 cm3 (2:1. The parameters under evaluation were number of nodules, nodules dry matter production, shoots dry matter, roots dry matter, and total dry matter production for all the species analyzed; and plant height, diameter, and the Dickson quality index for Mimosa species. Of the 14 tested strains, two nodulated M. atropurpureum; however, they were ineffective in promoting plant growth. All the tested strains established symbiosis with M. bimucronata, and 12 strains nodulated M. foliolosa. Of these, six promoted growth in M. bimucronata, and seven presented symbiotic efficiency in M. foliolosa. The strains UFLA 01-739, UFLA 01-748 and UFLA 01-751, isolated from M. tenuiflora, and UFLA 04-260 and UFLA 04-405, isolated from M. atropurpureum, stood out as potential inoculants for the Mimosa species evaluated in this study.

  8. 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.

  9. Octanoyl-Homoserine Lactone Is the Cognate Signal for Burkholderia mallei BmaR1-BmaI1 Quorum Sensing

    National Research Council Canada - National Science Library

    Duerkop, Breck A; Ulrich, Ricky L; Greenberg, E. P

    2007-01-01

    .... The obligate animal pathogen Burkholderia mallei produces several acyl-HSLs, and the B. mallei genome has four luxR and two luxI homologs, each of which has been established as a virulence factor...

  10. IN SITU BIOREMEDIATION OF TRICHLOROETHYLENE USING BURKHOLDERIA CEPACIA G4 PR1: ANALYSIS OF MICROBIAL ECOLOGY PARAMETERS FOR RISK ASSESSMENT (RESEARCH BRIEF)

    Science.gov (United States)

    The introduction of bacteria into aquifers for bioremediation purposes requires monitoring of the persistence and activity of microbial populations for efficacy and risk assessment purposes. Burkholderia cepacia G4 PR1 constitutively expresses a toluene ortho-monooxygenase (tom) ...

  11. A Unique Set of the Burkholderia Collagen-Like Proteins Provides Insight into Pathogenesis, Genome Evolution and Niche Adaptation, and Infection Detection.

    Science.gov (United States)

    Bachert, Beth A; Choi, Soo J; Snyder, Anna K; Rio, Rita V M; Durney, Brandon C; Holland, Lisa A; Amemiya, Kei; Welkos, Susan L; Bozue, Joel A; Cote, Christopher K; Berisio, Rita; Lukomski, Slawomir

    2015-01-01

    Burkholderia pseudomallei and Burkholderia mallei, classified as category B priority pathogens, are significant human and animal pathogens that are highly infectious and broad-spectrum antibiotic resistant. Currently, the pathogenicity mechanisms utilized by Burkholderia are not fully understood, and correct diagnosis of B. pseudomallei and B. mallei infection remains a challenge due to limited detection methods. Here, we provide a comprehensive analysis of a set of 13 novel Burkholderia collagen-like proteins (Bucl) that were identified among B. pseudomallei and B. mallei select agents. We infer that several Bucl proteins participate in pathogenesis based on their noncollagenous domains that are associated with the components of a type III secretion apparatus and membrane transport systems. Homology modeling of the outer membrane efflux domain of Bucl8 points to a role in multi-drug resistance. We determined that bucl genes are widespread in B. pseudomallei and B. mallei; Fischer's exact test and Cramer's V2 values indicate that the majority of bucl genes are highly associated with these pathogenic species versus nonpathogenic B. thailandensis. We designed a bucl-based quantitative PCR assay which was able to detect B. pseudomallei infection in a mouse with a detection limit of 50 CFU. Finally, chromosomal mapping and phylogenetic analysis of bucl loci revealed considerable genomic plasticity and adaptation of Burkholderia spp. to host and environmental niches. In this study, we identified a large set of phylogenetically unrelated bucl genes commonly found in Burkholderia select agents, encoding predicted pathogenicity factors, detection targets, and vaccine candidates.

  12. Test of aerobic TCE degradation by willows (Salix viminalis) and willows inoculated with TCE-cometabolizing strains of Burkholderia cepacia

    DEFF Research Database (Denmark)

    Clausen, Lauge Peter Westergaard; Broholm, Mette Martina; Gosewinkel, Ulrich Bay

    2017-01-01

    . cepacia (301C, PR1-31 and VM1330-pTOM), using chloride formation as an indicator of dehalogenation. Willows were grown in non-sterile, hydroponic conditions for 3 weeks in chloride-free nutrient solution spiked with TCE. TCE was added weekly due to rapid loss by volatilization. Chloride and TCE...

  13. Effect of two types of biosurfactants on phenanthrene availability to the bacterial bioreporter Burkholderia sartisoli strain RP037

    NARCIS (Netherlands)

    Tecon, R.; Van der Meer, J.R.

    2010-01-01

    Biosurfactants are tensio-active agents that have often been proposed as a means to enhance the aqueous solubility of hydrophobic organic contaminants, such as polycyclic aromatic hydrocarbons (PAHs). Biosurfactant-producing bacteria such as those belonging to the genus Pseudomonas might therefore

  14. Two Stable Variants of Burkholderia pseudomallei Strain MSHR5848 Express Broadly Divergent in vitro Phenotypes Associated with their Virulence Differences

    Science.gov (United States)

    2016-11-21

    initial growth advantage when exposed to nutrient- rich niches such as the intestinal tract or infected lungs. For instance, type 1 preferentially...possible initial growth advantage of type 1 in nutrient- rich niches , the evolving bacterial adaptation and deregulation of the utilization of...conditions, eg., the macrophage environment (Fig. 10) and inhibitory chemicals (PM substrates, Table 7A, B and Table S- 2). Survival in a defined niche such

  15. Hierarchical ZIF-8 toward Immobilizing Burkholderia cepacia Lipase for Application in Biodiesel Preparation

    Directory of Open Access Journals (Sweden)

    Miaad Adnan

    2018-05-01

    Full Text Available A hierarchical mesoporous zeolitic imidazolate framework (ZIF-8 was processed based on cetyltrimethylammonium bromide (CTAB as a morphological regulating agent and amino acid (l-histidine as assisting template agent. Burkholderia cepacia lipase (BCL was successfully immobilized by ZIF-8 as the carrier via an adsorption method (BCL-ZIF-8. The immobilized lipase (BCL showed utmost activity recovery up to 1279%, a 12-fold boost in its free counterpart. BCL-ZIF-8 was used as a biocatalyst in the transesterification reaction for the production of biodiesel with 93.4% yield. There was no significant lowering of conversion yield relative to original activity for BCL-ZIF-8 when continuously reused for eight cycles. This work provides a new outlook for biotechnological importance by immobilizing lipase on the hybrid catalyst (ZIF-8 and opens the door for its uses in the industrial field.

  16. Identification of the conserved hypothetical protein BPSL0317 in Burkholderia pseudomallei K96243

    Science.gov (United States)

    Yusoff, Nur Syamimi; Damiri, Nadzirah; Firdaus-Raih, Mohd

    2014-09-01

    Burkholderia pseudomallei K96243 is the causative agent of melioidosis, a disease which is endemic in Northern Australia and Southeastern Asia. The genome encodes several essential proteins including those currently annotated as hypothetical proteins. We studied the conservation and the essentiality of expressed hypothetical proteins in normal and different stress conditions. Based on the comparative genomics, we identified a hypothetical protein, BPSL0317, a potential essential gene that is being expressed in all normal and stress conditions. BPSL0317 is also phylogenetically conserved in the Burkholderiales order suggesting that this protein is crucial for survival among the order's members. BPSL0317 therefore has a potential to be a candidate antimicrobial drug target for this group of bacteria.

  17. Direct detection of the plant pathogens Burkholderia glumae, Burkholderia gladioli pv. gladioli, and Erwinia chrysanthemi pv. zeae in infected rice seedlings using matrix assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Kajiwara, Hideyuki

    2016-01-01

    The plant pathogens Burkholderia glumae, Burkholderia gladioli pv. gladioli, and Erwinia chrysanthemi pv. zeae were directly detected in extracts from infected rice seedlings by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). This method did not require culturing of the pathogens on artificial medium. In the MALDI-TOF MS analysis, peaks originating from bacteria were found in extracts from infected rice seedlings. The spectral peaks showed significantly high scores, in spite of minor differences in spectra. The spectral peaks originating from host plant tissues did not affect this direct MALDI-TOF MS analysis for the rapid identification of plant pathogens. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. Transports of acetate and haloacetate in Burkholderia species MBA4 are operated by distinct systems

    Directory of Open Access Journals (Sweden)

    Su Xianbin

    2012-11-01

    Full Text Available Abstract Background Acetate is a commonly used substrate for biosynthesis while monochloroacetate is a structurally similar compound but toxic and inhibits cell metabolism by blocking the citric acid cycle. In Burkholderia species MBA4 haloacetate was utilized as a carbon and energy source for growth. The degradation of haloacid was mediated by the production of an inducible dehalogenase. Recent studies have identified the presence of a concomitantly induced haloacetate-uptake activity in MBA4. This uptake activity has also been found to transport acetate. Since acetate transporters are commonly found in bacteria it is likely that haloacetate was transported by such a system in MBA4. Results The haloacetate-uptake activity of MBA4 was found to be induced by monochloroacetate (MCA and monobromoacetate (MBA. While the acetate-uptake activity was also induced by MCA and MBA, other alkanoates: acetate, propionate and 2-monochloropropionate (2MCPA were also inducers. Competing solute analysis showed that acetate and propionate interrupted the acetate- and MCA- induced acetate-uptake activities. While MCA, MBA, 2MCPA, and butyrate have no effect on acetate uptake they could significantly quenched the MCA-induced MCA-uptake activity. Transmembrane electrochemical potential was shown to be a driving force for both acetate- and MCA- transport systems. Conclusions Here we showed that acetate- and MCA- uptake in Burkholderia species MBA4 are two transport systems that have different induction patterns and substrate specificities. It is envisaged that the shapes and the three dimensional structures of the solutes determine their recognition or exclusion by the two transport systems.

  19. Understanding the Pathogenicity of Burkholderia contaminans, an Emerging Pathogen in Cystic Fibrosis.

    Science.gov (United States)

    Nunvar, Jaroslav; Kalferstova, Lucie; Bloodworth, Ruhi A M; Kolar, Michal; Degrossi, Jose; Lubovich, Silvina; Cardona, Silvia T; Drevinek, Pavel

    2016-01-01

    Several bacterial species from the Burkholderia cepacia complex (Bcc) are feared opportunistic pathogens that lead to debilitating lung infections with a high risk of developing fatal septicemia in cystic fibrosis (CF) patients. However, the pathogenic potential of other Bcc species is yet unknown. To elucidate clinical relevance of Burkholderia contaminans, a species frequently isolated from CF respiratory samples in Ibero-American countries, we aimed to identify its key virulence factors possibly linked with an unfavorable clinical outcome. We performed a genome-wide comparative analysis of two isolates of B. contaminans ST872 from sputum and blood culture of a female CF patient in Argentina. RNA-seq data showed significant changes in expression for quorum sensing-regulated virulence factors and motility and chemotaxis. Furthermore, we detected expression changes in a recently described low-oxygen-activated (lxa) locus which encodes stress-related proteins, and for two clusters responsible for the biosynthesis of antifungal and hemolytic compounds pyrrolnitrin and occidiofungin. Based on phenotypic assays that confirmed changes in motility and in proteolytic, hemolytic and antifungal activities, we were able to distinguish two phenotypes of B. contaminans that coexisted in the host and entered her bloodstream. Whole genome sequencing revealed that the sputum and bloodstream isolates (each representing a distinct phenotype) differed by over 1,400 mutations as a result of a mismatch repair-deficient hypermutable state of the sputum isolate. The inferred lack of purifying selection against nonsynonymous mutations and the high rate of pseudogenization in the derived isolate indicated limited evolutionary pressure during evolution in the nutrient-rich, stable CF sputum environment. The present study is the first to examine the genomic and transcriptomic differences between longitudinal isolates of B. contaminans. Detected activity of a number of putative virulence

  20. 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.

  1. Cell-bound lipases from Burkholderia sp. ZYB002: gene sequence analysis, expression, enzymatic characterization, and 3D structural model.

    Science.gov (United States)

    Shu, Zhengyu; Lin, Hong; Shi, Shaolei; Mu, Xiangduo; Liu, Yanru; Huang, Jianzhong

    2016-05-03

    The whole-cell lipase from Burkholderia cepacia has been used as a biocatalyst in organic synthesis. However, there is no report in the literature on the component or the gene sequence of the cell-bound lipase from this species. Qualitative analysis of the cell-bound lipase would help to illuminate the regulation mechanism of gene expression and further improve the yield of the cell-bound lipase by gene engineering. Three predictive cell-bound lipases, lipA, lipC21 and lipC24, from Burkholderia sp. ZYB002 were cloned and expressed in E. coli. Both LipA and LipC24 displayed the lipase activity. LipC24 was a novel mesophilic enzyme and displayed preference for medium-chain-length acyl groups (C10-C14). The 3D structural model of LipC24 revealed the open Y-type active site. LipA displayed 96 % amino acid sequence identity with the known extracellular lipase. lipA-inactivation and lipC24-inactivation decreased the total cell-bound lipase activity of Burkholderia sp. ZYB002 by 42 % and 14 %, respectively. The cell-bound lipase activity from Burkholderia sp. ZYB002 originated from a multi-enzyme mixture with LipA as the main component. LipC24 was a novel lipase and displayed different enzymatic characteristics and structural model with LipA. Besides LipA and LipC24, other type of the cell-bound lipases (or esterases) should exist.

  2. Brief communication genotyping of Burkholderia pseudomallei revealed high genetic variability among isolates from a single population group

    OpenAIRE

    Zueter, Abdelrahman Mohammad; Rahman, Zaidah Abdul; Yean, Chan Yean; Harun, Azian

    2015-01-01

    Burkholderia pseudomallei is a soil dwelling Gram-negative bacteria predominates in Southeast Asia zone and the tropical part of Australia. Genetic diversity has been explored among various populations and environments worldwide. To date, little data is available on MLST profiling of clinical B. pseudomallei isolates in peninsular Malaysia. In this brief report, thirteen culture positive B. pseudomallei cases collected from a single population of Terengganu state in the Western Peninsular Mal...

  3. Extensive cultivation of soil and water samples yields various pathogens in patients with cystic fibrosis but not Burkholderia multivorans.

    Science.gov (United States)

    Peeters, Charlotte; Depoorter, Eliza; Praet, Jessy; Vandamme, Peter

    2016-11-01

    While the epidemiology of Burkholderia cepacia complex (Bcc) bacteria in cystic fibrosis (CF) patients suggests that Burkholderia multivorans is acquired from environmental sources, this species has rarely been isolated from soil and water samples. Multiple isolation strategies were applied to water and soil samples that were previously shown to be B. multivorans PCR positive. These included direct plating and liquid enrichment procedures and the use of selective media, acclimatizing recovery and co-cultivation with CF sputum. MALDI-TOF mass spectrometry and sequence analysis of 16S rRNA and housekeeping genes were used to identify all isolates. None of the approaches yielded B. multivorans isolates. Other Burkholderia species, several Gram-negative non-fermenting bacteria (including Cupriavidus, Inquilinus, Pandoraea, Pseudomonas and Stenotrophomonas) and rapidly growing mycobacteria (including Mycobacterium chelonae) were all isolated from water and soil samples. The use of Bcc isolation media yielded a surprisingly wide array of rare but often clinically relevant CF pathogens, confirming that soil and water are reservoirs of these infectious agents. Copyright © 2016 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

  4. Rapid solubilization of insoluble phosphate by a novel environmental stress-tolerant Burkholderia vietnamiensis M6 isolated from ginseng rhizospheric soil.

    Science.gov (United States)

    Park, Ki-Hyun; Lee, O-Mi; Jung, Ho-Il; Jeong, Jin-Ha; Jeon, Young-Dong; Hwang, Dae-Youn; Lee, Chung-Yeol; Son, Hong-Joo

    2010-04-01

    We isolated and characterized novel insoluble phosphate (P)-solubilizing bacteria tolerant to environmental factors like high salt, low and high pHs, and low temperature. A bacterium M6 was isolated from a ginseng rhizospheric soil and confirmed to belong to Burkholderia vietnamiensis by BIOLOG system and 16S rRNA gene analysis. The optimal cultural conditions for the solubilization of P were 2.5% (w/v) glucose, 0.015% (w/v) urea, and 0.4% (w/v) MgCl(2).6H(2)O along with initial pH 7.0 at 35 degrees C. High-performance liquid chromatography analysis showed that B. vietnamiensis M6 produced gluconic and 2-ketogluconic acids. During the culture, the pH was reduced with increase in gluconic acid concentration and was inversely correlated with P solubilization. Insoluble P solubilization in the optimal medium was about 902 mg l(-1), which was approximately 1.6-fold higher than the yield in NBRIP medium (580 mg l(-1)). B. vietnamiensis M6 showed resistance against different environmental stresses like 10-45 degrees C, 1-5% (w/v) salt, and 2-11 pH range. The maximal concentration of soluble P produced by B. vietnamiensis M6 from Ca(3)(PO(4))(2), CaHPO(4), and hydroxyapatite was 1,039, 2,132, and 1,754 mg l(-1), respectively. However, the strain M6 produced soluble P with 20 mg l(-1) from FePO(4) after 2 days and 100 mg l(-1) from AlPO(4) after 6 days, respectively. Our results indicate that B. vietnamiensis M6 could be a potential candidate for the development of biofertilizer applicable to environmentally stressed soil.

  5. Choline Catabolism in Burkholderia thailandensis Is Regulated by Multiple Glutamine Amidotransferase 1-Containing AraC Family Transcriptional Regulators.

    Science.gov (United States)

    Nock, Adam M; Wargo, Matthew J

    2016-09-15

    Burkholderia thailandensis is a soil-dwelling bacterium that shares many metabolic pathways with the ecologically similar, but evolutionarily distant, Pseudomonas aeruginosa Among the diverse nutrients it can utilize is choline, metabolizable to the osmoprotectant glycine betaine and subsequently catabolized as a source of carbon and nitrogen, similar to P. aeruginosa Orthologs of genes in the choline catabolic pathway in these two bacteria showed distinct differences in gene arrangement as well as an additional orthologous transcriptional regulator in B. thailandensis In this study, we showed that multiple glutamine amidotransferase 1 (GATase 1)-containing AraC family transcription regulators (GATRs) are involved in regulation of the B. thailandensis choline catabolic pathway (gbdR1, gbdR2, and souR). Using genetic analyses and sequencing the transcriptome in the presence and absence of choline, we identified the likely regulons of gbdR1 (BTH_II1869) and gbdR2 (BTH_II0968). We also identified a functional ortholog for P. aeruginosa souR, a GATR that regulates the metabolism of sarcosine to glycine. GbdR1 is absolutely required for expression of the choline catabolic locus, similar to P. aeruginosa GbdR, while GbdR2 is important to increase expression of the catabolic locus. Additionally, the B. thailandensis SouR ortholog (BTH_II0994) is required for catabolism of choline and its metabolites as carbon sources, whereas in P. aeruginosa, SouR function can by bypassed by GbdR. The strategy employed by B. thailandensis represents a distinct regulatory solution to control choline catabolism and thus provides both an evolutionary counterpoint and an experimental system to analyze the acquisition and regulation of this pathway during environmental growth and infection. Many proteobacteria that occupy similar environmental niches have horizontally acquired orthologous genes for metabolism of compounds useful in their shared environment. The arrangement and differential

  6. Understanding regulation of the host-mediated gut symbiont population and the symbiont-mediated host immunity in the Riptortus-Burkholderia symbiosis system.

    Science.gov (United States)

    Kim, Jiyeun Kate; Lee, Jun Beom; Jang, Ho Am; Han, Yeon Soo; Fukatsu, Takema; Lee, Bok Luel

    2016-11-01

    Valuable insect models have tremendously contributed to our understanding of innate immunity and symbiosis. Bean bug, Riptortus pedestris, is a useful insect symbiosis model due to harboring cultivable monospecific gut symbiont, genus Burkholderia. Bean bug is a hemimetabolous insect whose immunity is not well-understood. However, we recently identified three major antimicrobial peptides of Riptortus and examined the relationship between gut symbiosis and host immunity. We found that the presence of Burkholderia gut symbiont positively affects Riptortus immunity. From studying host regulation mechanisms of symbiont population, we revealed that the symbiotic Burkholderia cells are much more susceptible to Riptortus immune responses than the cultured cells. We further elucidated that the immune-susceptibility of the Burkholderia gut symbionts is due to the drastic change of bacterial cell envelope. Finally, we show that the immune-susceptible Burkholderia symbionts are able to prosper in host owing to the suppression of immune responses of the symbiotic midgut. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Draft genome sequence of Paraburkholderia tropica Ppe8 strain, a sugarcane endophytic diazotrophic bacterium.

    Science.gov (United States)

    Silva, Paula Renata Alves da; Simões-Araújo, Jean Luiz; Vidal, Márcia Soares; Cruz, Leonardo Magalhães; Souza, Emanuel Maltempi de; Baldani, José Ivo

    Paraburkholderia tropica (syn Burkholderia tropica) are nitrogen-fixing bacteria commonly found in sugarcane. The Paraburkholderia tropica strain Ppe8 is part of the sugarcane inoculant consortium that has a beneficial effect on yield. Here, we report a draft genome sequence of this strain elucidating the mechanisms involved in its interaction mainly with Poaceae. A genome size of approximately 8.75Mb containing 7844 protein coding genes distributed in 526 subsystems was de novo assembled with ABySS and annotated by RAST. Genes related to the nitrogen fixation process, the secretion systems (I, II, III, IV, and VI), and related to a variety of metabolic traits, such as metabolism of carbohydrates, amino acids, vitamins, and proteins, were detected, suggesting a broad metabolic capacity and possible adaptation to plant association. Copyright © 2017 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  8. Degradación de Fenantreno por bacterias del género Burkholderia y Rhizobium aisladas de nódulos de mimosas

    Directory of Open Access Journals (Sweden)

    Arnoldo Wong-Villarreal

    2017-01-01

    Full Text Available El presente trabajo tuvo como objetivo identificar y evaluar la capacidad de degradación de microorganismos aislados de nódulos de mimosas, que puedan ser utilizados en procesos de biorremediación de suelos contaminados con fenantreno . Método . Se realizó el aislamiento de 122 cepas bacterianas de nódulos de mimosas; fueron crecidas en el medio de cultivo Maconkey para descartar enterobacterias. L as cepas bacterianas que dieron resultado negativo a esta prueba, fueron inoculadas en el medio de cultivo que contenía como úni ca fuente de carbono fenantreno; tres aislados tuvieron la capacidad de crecer en este medio. Las tres cepas fueron identificadas por secuencia del gen 1 6s ribosomal, se evaluó su capacidad de crecimiento en presencia de fenantreno mediante curvas de crecimiento microbiano; la capacidad para degradar fenantreno de las tres cepas fue cuantificada por cromatografía de gases acoplado a masas. Resultados . La s secuencias obtenidas del gen 16s ribosomal tienen relación genética con las especies de Burkholderia phenoliruptrix , Burkholderia phymatum y Rhizobium paknamense. El crecimiento microbiano de las tres cepas, suministradas con fenantreno, tuvieron un comp ortamiento similar al control , el cual contenía succinato como fuente de carbono. La cepa de Burkholderia sp. BB26 degradó 78.5 % , Burkholderia sp. BB24 68.5 % y Rhizobium sp. BY8 99%. Discusión . Los resultados de degradación de fenantreno por las cepas de Burkholderia sp. BB26 , Burkholderia sp. BB24 y Rhizobium sp. BY8 sugieren que las tres cepas tienen p otencial para utilizarse en procesos de biorremediación de suelos contaminados con fenantreno.

  9. An outbreak of Burkholderia cepacia complex in the paediatric unit of a tertiary care hospital

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    Swapna Mali

    2017-01-01

    Full Text Available Introduction: Burkholderia cepacia complex (Bcc has emerged as a serious nosocomial pathogen worldwide especially in patients with indwelling catheters and cystic fibrosis. Bcc is a common contaminant of pharmaceutical products. We describe an outbreak of Bcc bacteraemia amongst children admitted in Paediatric Intensive Care Unit (PICU and paediatric ward at a tertiary care hospital, Mumbai, in Western India. Materials and Methods: Blood culture samples from paediatric patients yielded growth of non-fermenting, oxidase positive, motile, Gram negative bacilli (NFGNB (76/909 over a period of 8 months. Based on conventional biochemical tests and antimicrobial susceptibility testing, these isolates were provisionally identified as Bcc. The increased, repeated and continued isolation of Bcc alerted the possibility of an outbreak confined to PICU and paediatric ward. Active surveillance was undertaken to trace the source and contain the outbreak. Isolates were subjected to recA polymerase chain reaction (PCR and Expanded multilocus sequence typing (EMLST. Results: Surveillance revealed the presence of Bcc on the upper surface of rubber stopper of sealed multidose amikacin vials. Isolates from blood culture and rubber stoppers were confirmed as Bcc by recA PCR. EMLST revealed that these isolates shared an identical novel sequence type 824 proving clonality. Timely interventions instituted led to control of the outbreak. Conclusion: This study highlights the importance of identification and molecular characterization of Bcc to establish its role in infection and outbreak.

  10. MOLECULAR DYNAMICS SIMULATION OF KINETIC RESOLUTION OF RACEMIC ALCOHOL USING BURKHOLDERIA CEPACIA LIPASE IN ORGANIC SOLVENTS

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    A. C. Mathpati

    2018-03-01

    Full Text Available Lipases, a subclass of hydrolases, have gained a lot of importance as they can catalyze esterification, transesterification and hydrolysis reaction in non-aqueous media. Lipases are also widely used for kinetic resolution of racemic alcohols into enantiopure compounds. The lipase activity is affected by organic solvents due to changes in the conformational rigidity of enzymes, the active site, or altering the solvation of the transition state. The activity of lipases strongly depends on the logP value of solvents. Molecular dynamics (MD can help to understand the effect of solvents on lipase conformation as well as protein-ligand complex. In this work, MD simulations of Burkholderia cepacia lipase (BCL and complex between R and S conformation of acetylated form of 1-phenylethanol with BCL using gromacs have been carried in various organic solvents. The RMSD values were within the range of 0.15 to 0.20 nm and radius of gyration was found to be with 1.65 to 1.9 nm. Major changes in the B factor compared to reference structure were observed between residues 60 to 80, 120 to 150 and 240 to 260. Higher unfolding was observed in toluene and diethyl ether compared to hexane and acetonitrile. R acetylated complex was found to favorably bind BCL compared to S form. The predicted enantioselectivity were in good agreement with the experimental data.

  11. An outbreak of Burkholderia cepacia complex in the paediatric unit of a tertiary care hospital.

    Science.gov (United States)

    Mali, Swapna; Dash, Lona; Gautam, Vikas; Shastri, Jayanthi; Kumar, Sunil

    2017-01-01

    Burkholderia cepacia complex (Bcc) has emerged as a serious nosocomial pathogen worldwide especially in patients with indwelling catheters and cystic fibrosis. Bcc is a common contaminant of pharmaceutical products. We describe an outbreak of Bcc bacteraemia amongst children admitted in Paediatric Intensive Care Unit (PICU) and paediatric ward at a tertiary care hospital, Mumbai, in Western India. Blood culture samples from paediatric patients yielded growth of non-fermenting, oxidase positive, motile, Gram negative bacilli (NFGNB) (76/909) over a period of 8 months. Based on conventional biochemical tests and antimicrobial susceptibility testing, these isolates were provisionally identified as Bcc. The increased, repeated and continued isolation of Bcc alerted the possibility of an outbreak confined to PICU and paediatric ward. Active surveillance was undertaken to trace the source and contain the outbreak. Isolates were subjected to recA polymerase chain reaction (PCR) and Expanded multilocus sequence typing (EMLST). Surveillance revealed the presence of Bcc on the upper surface of rubber stopper of sealed multidose amikacin vials. Isolates from blood culture and rubber stoppers were confirmed as Bcc by recA PCR. EMLST revealed that these isolates shared an identical novel sequence type 824 proving clonality. Timely interventions instituted led to control of the outbreak. This study highlights the importance of identification and molecular characterization of Bcc to establish its role in infection and outbreak.

  12. Antimicrobial Susceptibility and Genetic Characterisation of Burkholderia pseudomallei Isolated from Malaysian Patients

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    Yalda Khosravi

    2014-01-01

    Full Text Available Burkholderia pseudomallei, the causative agent of melioidosis, is intrinsically resistant to many antibiotics. Ceftazidime (CAZ, the synthetic β-lactam, is normally used as the first-line antibiotic therapy for treatment of melioidosis. However, acquired CAZ resistance can develop in vivo during treatment with CAZ, leading to mortality if therapy is not switched to a different antibiotic(s in a timely manner. In this study, susceptibilities of 81 B. pseudomallei isolates to nine different antimicrobial agents were determined using the disk diffusion method, broth microdilution test and Etest. Highest percentage of susceptibility was demonstrated to CAZ, amoxicillin/clavulanic acid, meropenem, imipenem, and trimethoprim/sulfamethoxazole. Although these drugs demonstrated the highest percentage of susceptibility in B. pseudomallei, the overall results underline the importance of the emergence of resistance in this organism. PCR results showed that, of the 81 B. pseudomallei, six multidrug resistant (MDR isolates carried bpeB, amrB, and BPSS1119 and penA genes. Genotyping of the isolates using random amplified polymorphic DNA analysis showed six different PCR fingerprinting patterns generated from the six MDR isolates clusters (A and eight PCR fingerprinting patterns generated for the remaining 75 non-MDR isolates clusters (B.

  13. Integration process of fermentation and liquid biphasic flotation for lipase separation from Burkholderia cepacia.

    Science.gov (United States)

    Sankaran, Revathy; Show, Pau Loke; Lee, Sze Ying; Yap, Yee Jiun; Ling, Tau Chuan

    2018-02-01

    Liquid Biphasic Flotation (LBF) is an advanced recovery method that has been effectively applied for biomolecules extraction. The objective of this investigation is to incorporate the fermentation and extraction process of lipase from Burkholderia cepacia using flotation system. Initial study was conducted to compare the performance of bacteria growth and lipase production using flotation and shaker system. From the results obtained, bacteria shows quicker growth and high lipase yield via flotation system. Integration process for lipase separation was investigated and the result showed high efficiency reaching 92.29% and yield of 95.73%. Upscaling of the flotation system exhibited consistent result with the lab-scale which are 89.53% efficiency and 93.82% yield. The combination of upstream and downstream processes in a single system enables the acceleration of product formation, improves the product yield and facilitates downstream processing. This integration system demonstrated its potential for biomolecules fermentation and separation that possibly open new opportunities for industrial production. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Toward modern inhalational bacteriophage therapy: nebulization of bacteriophages of Burkholderia cepacia complex.

    Science.gov (United States)

    Golshahi, Laleh; Seed, Kimberley D; Dennis, Jonathan J; Finlay, Warren H

    2008-12-01

    Antibiotic-resistant bacterial infections have renewed interest in finding substitute methods of treatment. The purpose of the present in vitro study was to investigate the possibility of respiratory delivery of a Burkholderia cepacia complex (BCC) bacteriophage by nebulized aerosol administration. Bacteriophages in isotonic saline were aerosolized with Pari LC star and eFlow nebulizers, at titers with mean value (standard deviation) of 2.15 x 10(8) (1.63 x 10(8)) plaque-forming unit (PFU)/mL in 2.5-mL nebulizer fills. The breathing pattern of an adult was simulated using a pulmonary waveform generator. During breath simulation, the size distributions of the nebulized aerosol were measured using phase doppler anemometry (PDA). Efficiency of nebulizer delivery was subsequently determined by collection of aerosol on low resistance filters and measurement of bacteriophage titers. These filter titers were used as input data to a mathematical lung deposition model to predict regional deposition of bacteriophages in the lung and initial bacteriophage titers in the liquid surface layer of each conducting airway generation. The results suggest that BCC bacteriophages can be nebulized successfully within a reasonable delivery time and predicted titers in the lung indicate that this method may hold potential for treatment of bacterial lung infections common among cystic fibrosis patients.

  15. Molecular architecture of the N-type ATPase rotor ring from Burkholderia pseudomallei.

    Science.gov (United States)

    Schulz, Sarah; Wilkes, Martin; Mills, Deryck J; Kühlbrandt, Werner; Meier, Thomas

    2017-04-01

    The genome of the highly infectious bacterium Burkholderia pseudomallei harbors an atp operon that encodes an N-type rotary ATPase, in addition to an operon for a regular F-type rotary ATPase. The molecular architecture of N-type ATPases is unknown and their biochemical properties and cellular functions are largely unexplored. We studied the B. pseudomallei N 1 N o -type ATPase and investigated the structure and ion specificity of its membrane-embedded c-ring rotor by single-particle electron cryo-microscopy. Of several amphiphilic compounds tested for solubilizing the complex, the choice of the low-density, low-CMC detergent LDAO was optimal in terms of map quality and resolution. The cryoEM map of the c-ring at 6.1 Å resolution reveals a heptadecameric oligomer with a molecular mass of ~141 kDa. Biochemical measurements indicate that the c 17 ring is H + specific, demonstrating that the ATPase is proton-coupled. The c 17 ring stoichiometry results in a very high ion-to-ATP ratio of 5.7. We propose that this N-ATPase is a highly efficient proton pump that helps these melioidosis-causing bacteria to survive in the hostile, acidic environment of phagosomes. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

  16. Survival and Intra-Nuclear Trafficking of Burkholderia pseudomallei: Strategies of Evasion from Immune Surveillance?

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    Jamuna Vadivelu

    2017-01-01

    Full Text Available During infection, successful bacterial clearance is achieved via the host immune system acting in conjunction with appropriate antibiotic therapy. However, it still remains a tip of the iceberg as to where persistent pathogens namely, Burkholderia pseudomallei (B. pseudomallei reside/hide to escape from host immune sensors and antimicrobial pressure.We used transmission electron microscopy (TEM to investigate post-mortem tissue sections of patients with clinical melioidosis to identify the localisation of a recently identified gut microbiome, B. pseudomallei within host cells. The intranuclear presence of B. pseudomallei was confirmed using transmission electron microscopy (TEM of experimentally infected guinea pig spleen tissues and Live Z-stack, and ImageJ analysis of fluorescence microscopy analysis of in vitro infection of A549 human lung epithelial cells.TEM investigations revealed intranuclear localization of B. pseudomallei in cells of infected human lung and guinea pig spleen tissues. We also found that B. pseudomallei induced actin polymerization following infection of A549 human lung epithelial cells. Infected A549 lung epithelial cells using 3D-Laser scanning confocal microscopy (LSCM and immunofluorescence microscopy confirmed the intranuclear localization of B. pseudomallei.B. pseudomallei was found within the nuclear compartment of host cells. The nucleus may play a role as an occult or transient niche for persistence of intracellular pathogens, potentially leading to recurrrent episodes or recrudescence of infection.

  17. What drives the occurrence of the melioidosis bacterium Burkholderia pseudomallei in domestic gardens?

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    Mirjam Kaestli

    2015-03-01

    Full Text Available Melioidosis is an often fatal infectious disease affecting humans and animals in tropical regions and is caused by the saprophytic environmental bacterium Burkholderia pseudomallei. Domestic gardens are not only a common source of exposure to soil and thus to B. pseudomallei, but they also have been found to contain more B. pseudomallei than other environments. In this study we addressed whether anthropogenic manipulations common to gardens such as irrigation or fertilizers change the occurrence of B. pseudomallei. We conducted a soil microcosm experiment with a range of fertilizers and soil types as well as a longitudinal interventional study over three years on an experimental fertilized field site in an area naturally positive for B. pseudomallei. Irrigation was the only consistent treatment to increase B. pseudomallei occurrence over time. The effects of fertilizers upon these bacteria depended on soil texture, physicochemical soil properties and biotic factors. Nitrates and urea increased B. pseudomallei load in sand while phosphates had a positive effect in clay. The high buffering and cation exchange capacities of organic material found in a commercial potting mix led to a marked increase in soil salinity with no survival of B. pseudomallei after four weeks in the potting mix sampled. Imported grasses were also associated with B. pseudomallei occurrence in a multivariate model. With increasing population density in endemic areas these findings inform the identification of areas in the anthropogenic environment with increased risk of exposure to B. pseudomallei.

  18. Leveraging structure determination with fragment screening for infectious disease drug targets: MECP synthase from Burkholderia pseudomallei

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    Begley, Darren W.; Hartley, Robert C.; Davies, Douglas R.; Edwards, Thomas E.; Leonard, Jess T.; Abendroth, Jan; Burris, Courtney A.; Bhandari, Janhavi; Myler, Peter J.; Staker, Bart L.; Stewart, Lance J. (UWASH); (Emerald)

    2011-09-28

    As part of the Seattle Structural Genomics Center for Infectious Disease, we seek to enhance structural genomics with ligand-bound structure data which can serve as a blueprint for structure-based drug design. We have adapted fragment-based screening methods to our structural genomics pipeline to generate multiple ligand-bound structures of high priority drug targets from pathogenic organisms. In this study, we report fragment screening methods and structure determination results for 2C-methyl-D-erythritol-2,4-cyclo-diphosphate (MECP) synthase from Burkholderia pseudomallei, the gram-negative bacterium which causes melioidosis. Screening by nuclear magnetic resonance spectroscopy as well as crystal soaking followed by X-ray diffraction led to the identification of several small molecules which bind this enzyme in a critical metabolic pathway. A series of complex structures obtained with screening hits reveal distinct binding pockets and a range of small molecules which form complexes with the target. Additional soaks with these compounds further demonstrate a subset of fragments to only bind the protein when present in specific combinations. This ensemble of fragment-bound complexes illuminates several characteristics of MECP synthase, including a previously unknown binding surface external to the catalytic active site. These ligand-bound structures now serve to guide medicinal chemists and structural biologists in rational design of novel inhibitors for this enzyme.

  19. A preliminary X-ray study of sedoheptulose-7-phosphate isomerase from Burkholderia pseudomallei

    International Nuclear Information System (INIS)

    Kim, Mi-Sun; Shin, Dong Hae

    2009-01-01

    Sedoheptulose-7-phosphate isomerase (GmhA) from B. pseudomallei is one of the targets of antibiotic adjuvants for melioidosis. In this study, GmhA has been cloned, expressed, purified and crystallized. Sedoheptulose-7-phosphate isomerase (GmhA) converts d-sedoheptulose 7-phosphate to d,d-heptose 7-phosphate. This is the first step in the biosynthesis pathway of NDP-heptose, which is responsible for the pleiotropic phenotype. This biosynthesis pathway is the target of inhibitors to increase the membrane permeability of Gram-negative pathogens or of adjuvants working synergistically with known antibiotics. Burkholderia pseudomallei is the causative agent of melioidosis, a seriously invasive disease in animals and humans in tropical and subtropical areas. GmhA from B. pseudomallei is one of the targets of antibiotic adjuvants for melioidosis. In this study, GmhA has been cloned, expressed, purified and crystallized. Synchrotron X-ray data were also collected to 1.9 Å resolution. The crystal belonged to the primitive orthorhombic space group P2 1 2 1 2 1 , with unit-cell parameters a = 61.3, b = 84.2, c = 142.3 Å. A full structural determination is under way in order to provide insights into the structure–function relationships of this protein

  20. A preliminary X-ray study of transketolase from Burkholderia pseudomallei

    International Nuclear Information System (INIS)

    Kim, Mi-Sun; Lim, Areum; Yang, Seung Won; Lee, Daeun; Park, Jimin; Shin, Dong Hae

    2012-01-01

    The transketolase TktA from B. pseudomallei has been cloned, expressed, purified and crystallized. Synchrotron X-ray data were collected to 2.0 Å resolution. TktA is the most critical enzyme in the nonoxidative pentose phosphate pathway. It catalyzes the conversion of xylulose 5-phosphate and ribose 5-phosphate into sedoheptulose 7-phosphate and glyceraldehyde 3-phosphate, and its products are used in the biosynthesis of acetyl-CoA, aromatic amino acids, nucleic acids and ADP-l-glycero-β-d-manno-heptose. TktA also has an unexpected role in chromosome structure that is independent of its metabolic responsibilities. Therefore, it is a new potent antibiotic target. In this study, TktA from Burkholderia pseudomallei has been cloned, expressed, purified and crystallized. Synchrotron X-ray data were also collected to 2.0 Å resolution. The crystal belonged to the monoclinic space group C2, with unit-cell parameters a = 146.2, b = 74.6, c = 61.6 Å, β = 113.0°. A full structural determination is under way in order to provide insight into the structure–function relationship of this protein

  1. Screening for potential anti-infective agents towards Burkholderia pseudomallei infection

    Science.gov (United States)

    Eng, Su Anne; Nathan, Sheila

    2014-09-01

    The established treatment for melioidosis is antibiotic therapy. However, a constant threat to this form of treatment is resistance development of the causative agent, Burkholderia pseudomallei, towards antibiotics. One option to circumvent this threat of antibiotic resistance is to search for new alternative anti-infectives which target the host innate immune system and/or bacterial virulence. In this study, 29 synthetic compounds were evaluated for their potential to increase the lifespan of an infected host. The nematode Caenorhabditis elegans was adopted as the infection model as its innate immune pathways are homologous to humans. Screens were performed in a liquid-based survival assay containing infected worms exposed to individual compounds and survival of untreated and compound-treated worms were compared. A primary screen identified nine synthetic compounds that extended the lifespan of B. pseudomallei-infected worms. Subsequently, a disc diffusion test was performed on these selected compounds to delineate compounds into those that enhanced the survival of worms via antimicrobial activity i.e. reducing the number of infecting bacteria, or into those that did not target pathogen viability. Out of the nine hits selected, two demonstrated antimicrobial effects on B. pseudomallei. Therefore, the findings from this study suggest that the other seven identified compounds are potential anti-infectives which could protect a host against B. pseudomallei infection without developing the risk of drug resistance.

  2. A new species of Burkholderia isolated from sugarcane roots promotes plant growth

    Science.gov (United States)

    Paungfoo-Lonhienne, Chanyarat; Lonhienne, Thierry G A; Yeoh, Yun Kit; Webb, Richard I; Lakshmanan, Prakash; Chan, Cheong Xin; Lim, Phaik-Eem; Ragan, Mark A; Schmidt, Susanne; Hugenholtz, Philip

    2014-01-01

    Sugarcane is a globally important food, biofuel and biomaterials crop. High nitrogen (N) fertilizer rates aimed at increasing yield often result in environmental damage because of excess and inefficient application. Inoculation with diazotrophic bacteria is an attractive option for reducing N fertilizer needs. However, the efficacy of bacterial inoculants is variable, and their effective formulation remains a knowledge frontier. Here, we take a new approach to investigating diazotrophic bacteria associated with roots using culture-independent microbial community profiling of a commercial sugarcane variety (Q208A) in a field setting. We first identified bacteria that were markedly enriched in the rhizosphere to guide isolation and then tested putative diazotrophs for the ability to colonize axenic sugarcane plantlets (Q208A) and promote growth in suboptimal N supply. One isolate readily colonized roots, fixed N2 and stimulated growth of plantlets, and was classified as a new species, Burkholderia australis sp. nov. Draft genome sequencing of the isolate confirmed the presence of nitrogen fixation. We propose that culture-independent identification and isolation of bacteria that are enriched in rhizosphere and roots, followed by systematic testing and confirming their growth-promoting capacity, is a necessary step towards designing effective microbial inoculants. PMID:24350979

  3. Molecular phylogeny of Burkholderia pseudomallei from a remote region of Papua New Guinea.

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    Anthony Baker

    Full Text Available BACKGROUND: The island of New Guinea is located midway between the world's two major melioidosis endemic regions of Australia and Southeast Asia. Previous studies in Papua New Guinea have demonstrated autochthonous melioidosis in Balimo, Western province. In contrast to other regions of endemicity, isolates recovered from both environmental and clinical sources demonstrate narrow genetic diversity over large spatial and temporal scales. METHODOLOGY/PRINCIPAL FINDINGS: We employed molecular typing techniques to determine the phylogenetic relationships of these isolates to each other and to others worldwide to aid in understanding the origins of the Papua New Guinean isolates. Multi-locus sequence typing of the 39 isolates resolved three unique sequence types. Phylogenetic reconstruction and Structure analysis determined that all isolates were genetically closer to those from Australia than those from Southeast Asia. Gene cluster analysis however, identified a Yersinia-like fimbrial gene cluster predominantly found among Burkholderia pseudomallei derived from Southeast Asia. Higher resolution VNTR typing and phylogenetic reconstruction of the Balimo isolates resolved 24 genotypes with long branch lengths. These findings are congruent with long term persistence in the region and a high level of environmental stability. CONCLUSIONS/SIGNIFICANCE: Given that anthropogenic influence has been hypothesized as a mechanism for the dispersal of B. pseudomallei, these findings correlate with limited movement of the indigenous people in the region. The palaeogeographical and anthropogenic history of Australasia and the results from this study indicate that New Guinea is an important region for the further study of B. pseudomallei origins and dissemination.

  4. Optimization of olive oil hydrolysis process using immobilized Lipase from Burkholderia cepacia sp. in Polyurethane

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    Nádia Ligianara Dewes Nyari

    2017-09-01

    Full Text Available The aim of this study was to achieve the best conditions for the  olive oil hydrolysis process at optimal pH and temperature using Burkholderia cepacia lipase immobilized in situ in rigid polyurethane support. The influences of the temperature (13.85 to 56.5ºC and pH (4.18 to 9.82 were evaluated by a central composite rotational experimental design 22. The operational stability and storage conditions were also studied. The olive oil hydrolysis process was optimized in pH 7.0, at 40°C and 15 min of reaction, with 66 and 93 U g-1 of hydrolysis activity in free and immobilized lipase, respectively, with > 700% yield. The immobilized remained stable for up to 40 days of storage at temperatures of 60oC, and for 100 days from 4 to 25°C. The operational stability of the immobilized was 6 continuous cycles. In this way, immobilization showed to be a promising alternative for its application in olive oil hydrolysis, having storage stability and reuse capability.

  5. Crystal structure of a β-aminopeptidase from an Australian Burkholderia sp.

    Science.gov (United States)

    John-White, Marietta; Dumsday, Geoff J; Johanesen, Priscilla; Lyras, Dena; Drinkwater, Nyssa; McGowan, Sheena

    2017-07-01

    β-Aminopeptidases are a unique group of enzymes that have the unusual capability to hydrolyze N-terminal β-amino acids from synthetic β-peptides. β-Peptides can form secondary structures mimicking α-peptide-like structures that are resistant to degradation by most known proteases and peptidases. These characteristics of β-peptides give them great potential as peptidomimetics. Here, the X-ray crystal structure of BcA5-BapA, a β-aminopeptidase from a Gram-negative Burkholderia sp. that was isolated from activated sludge from a wastewater-treatment plant in Australia, is reported. The crystal structure of BcA5-BapA was determined to a resolution of 2.0 Å and showed a tetrameric assembly typical of the β-aminopeptidases. Each monomer consists of an α-subunit (residues 1-238) and a β-subunit (residues 239-367). Comparison of the structure of BcA5-BapA with those of other known β-aminopeptidases shows a highly conserved structure and suggests a similar proteolytic mechanism of action.

  6. Host immunity in the protective response to vaccination with heat-killed Burkholderia mallei

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    Paessler Slobodan

    2008-09-01

    Full Text Available Abstract Background We performed initial cell, cytokine and complement depletion studies to investigate the possible role of these effectors in response to vaccination with heat-killed Burkholderia mallei in a susceptible BALB/c mouse model of infection. Results While protection with heat-killed bacilli did not result in sterilizing immunity, limited protection was afforded against an otherwise lethal infection and provided insight into potential host protective mechanisms. Our results demonstrated that mice depleted of either B cells, TNF-α or IFN-γ exhibited decreased survival rates, indicating a role for these effectors in obtaining partial protection from a lethal challenge by the intraperitoneal route. Additionally, complement depletion had no effect on immunoglobulin production when compared to non-complement depleted controls infected intranasally. Conclusion The data provide a basis for future studies of protection via vaccination using either subunit or whole-organism vaccine preparations from lethal infection in the experimental BALB/c mouse model. The results of this study demonstrate participation of B220+ cells and pro-inflammatory cytokines IFN-γ and TNF-α in protection following HK vaccination.

  7. Survival and Intra-Nuclear Trafficking of Burkholderia pseudomallei: Strategies of Evasion from Immune Surveillance?

    Science.gov (United States)

    Vadivelu, Jamuna; Vellasamy, Kumutha Malar; Thimma, Jaikumar; Mariappan, Vanitha; Kang, Wen-Tyng; Choh, Leang-Chung; Wong, Kum Thong

    2017-01-01

    Background During infection, successful bacterial clearance is achieved via the host immune system acting in conjunction with appropriate antibiotic therapy. However, it still remains a tip of the iceberg as to where persistent pathogens namely, Burkholderia pseudomallei (B. pseudomallei) reside/hide to escape from host immune sensors and antimicrobial pressure. Methods We used transmission electron microscopy (TEM) to investigate post-mortem tissue sections of patients with clinical melioidosis to identify the localisation of a recently identified gut microbiome, B. pseudomallei within host cells. The intranuclear presence of B. pseudomallei was confirmed using transmission electron microscopy (TEM) of experimentally infected guinea pig spleen tissues and Live Z-stack, and ImageJ analysis of fluorescence microscopy analysis of in vitro infection of A549 human lung epithelial cells. Results TEM investigations revealed intranuclear localization of B. pseudomallei in cells of infected human lung and guinea pig spleen tissues. We also found that B. pseudomallei induced actin polymerization following infection of A549 human lung epithelial cells. Infected A549 lung epithelial cells using 3D-Laser scanning confocal microscopy (LSCM) and immunofluorescence microscopy confirmed the intranuclear localization of B. pseudomallei. Conclusion B. pseudomallei was found within the nuclear compartment of host cells. The nucleus may play a role as an occult or transient niche for persistence of intracellular pathogens, potentially leading to recurrrent episodes or recrudescence of infection. PMID:28045926

  8. Development and validation of Burkholderia pseudomallei-specific real-time PCR assays for clinical, environmental or forensic detection applications.

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    Erin P Price

    Full Text Available The bacterium Burkholderia pseudomallei causes melioidosis, a rare but serious illness that can be fatal if untreated or misdiagnosed. Species-specific PCR assays provide a technically simple method for differentiating B. pseudomallei from near-neighbor species. However, substantial genetic diversity and high levels of recombination within this species reduce the likelihood that molecular signatures will differentiate all B. pseudomallei from other Burkholderiaceae. Currently available molecular assays for B. pseudomallei detection lack rigorous validation across large in silico datasets and isolate collections to test for specificity, and none have been subjected to stringent quality control criteria (accuracy, precision, selectivity, limit of quantitation (LoQ, limit of detection (LoD, linearity, ruggedness and robustness to determine their suitability for environmental, clinical or forensic investigations. In this study, we developed two novel B. pseudomallei specific assays, 122018 and 266152, using a dual-probe approach to differentiate B. pseudomallei from B. thailandensis, B. oklahomensis and B. thailandensis-like species; other species failed to amplify. Species specificity was validated across a large DNA panel (>2,300 samples comprising Burkholderia spp. and non-Burkholderia bacterial and fungal species of clinical and environmental relevance. Comparison of assay specificity to two previously published B. pseudomallei-specific assays, BurkDiff and TTS1, demonstrated comparable performance of all assays, providing between 99.7 and 100% specificity against our isolate panel. Last, we subjected 122018 and 266152 to rigorous quality control analyses, thus providing quantitative limits of assay performance. Using B. pseudomallei as a model, our study provides a framework for comprehensive quantitative validation of molecular assays and provides additional, highly validated B. pseudomallei assays for the scientific research community.

  9. Characterization of N-acylhomoserine lactone-degrading bacteria associated with the Zingiber officinale (ginger rhizosphere: Co-existence of quorum quenching and quorum sensing in Acinetobacter and Burkholderia

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    Chhabra Siri

    2011-03-01

    Full Text Available Abstract Background Cell-to-cell communication (quorum sensing (QS co-ordinates bacterial behaviour at a population level. Consequently the behaviour of a natural multi-species community is likely to depend at least in part on co-existing QS and quorum quenching (QQ activities. Here we sought to discover novel N-acylhomoserine lactone (AHL-dependent QS and QQ strains by investigating a bacterial community associated with the rhizosphere of ginger (Zingiber officinale growing in the Malaysian rainforest. Results By using a basal growth medium containing N-(3-oxohexanoylhomoserine lactone (3-oxo-C6-HSL as the sole source of carbon and nitrogen, the ginger rhizosphere associated bacteria were enriched for strains with AHL-degrading capabilities. Three isolates belonging to the genera Acinetobacter (GG2, Burkholderia (GG4 and Klebsiella (Se14 were identified and selected for further study. Strains GG2 and Se14 exhibited the broadest spectrum of AHL-degrading activities via lactonolysis while GG4 reduced 3-oxo-AHLs to the corresponding 3-hydroxy compounds. In GG2 and GG4, QQ was found to co-exist with AHL-dependent QS and GG2 was shown to inactivate both self-generated and exogenously supplied AHLs. GG2, GG4 and Se14 were each able to attenuate virulence factor production in both human and plant pathogens. Conclusions Collectively our data show that ginger rhizosphere bacteria which make and degrade a wide range of AHLs are likely to play a collective role in determining the QS-dependent phenotype of a polymicrobial community.

  10. Colonization of Tomato Root by Antagonistic Bacterial Strains to Fusarium Wilt of Tomato

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    Arif Wibowo

    2005-12-01

    Full Text Available Fusarium wilt of tomato caused by Fusarium oxysporum f.sp. lycopersici (Fol is an important disease in tomato which cause a significant loss of yield in major growing regions of the world. This study examined the ability of bacterial strains antagonistic to F. oxysporum f.sp. lycopersici (H5, H22, H63, H71, Burkholderia cepacia strain 65 and 526 to colonize tomato seedlings and the effect of plant growth. The effect of bacterial population size and air temperature on the bacterial colonization and their spread along the root systems was also assessed.The results of this study showed that the bacterial population at 28°/23° C day/night temperature 14 days after planting was significantly greater than 23°/18° C for 4 of 6 strains tested. Although there was no significant effect of temperature on bacterial population observed in this study, the ability of the baacterial strains to colonize the rhizosphere was significantly different. Three strains (H5, B. cepacia strain 65 and 526 survived well in the rhizosphere and at 4 weeks after planting rhizosphere populations per gram fresh root were not significantly different from those recovered 2 weeks after planting. The largest population of the bacterial inoculants developed in the basal region of the roots and this differed between strains by log10 2.7 cfu/cm root. The bacterial populations in other parts of the root were also strain dependent. Strain H71, for example, was able to colonize the root segments at a high population level. However strain H63 was recovered only in small number in all root segments.

  11. Rapid DNA vaccination against Burkholderia pseudomallei flagellin by tattoo or intranasal application.

    Science.gov (United States)

    Lankelma, Jacqueline M; Wagemakers, Alex; Birnie, Emma; Haak, Bastiaan W; Trentelman, Jos J A; Weehuizen, Tassili A F; Ersöz, Jasmin; Roelofs, Joris J T H; Hovius, Joppe W; Wiersinga, W Joost; Bins, Adriaan D

    2017-11-17

    Melioidosis is a severe infectious disease with a high mortality that is endemic in South-East Asia and Northern Australia. The causative pathogen, Burkholderia pseudomallei, is listed as potential bioterror weapon due to its high virulence and potential for easy dissemination. Currently, there is no licensed vaccine for prevention of melioidosis. Here, we explore the use of rapid plasmid DNA vaccination against B. pseudomallei flagellin for protection against respiratory challenge. We tested three flagellin DNA vaccines with different subcellular targeting designs. C57BL/6 mice were vaccinated via skin tattoo on day 0, 3 and 6 before intranasal challenge with B. pseudomallei on day 21. Next, the most effective construct was used as single vaccination on day 0 by tattoo or intranasal formulation. Mice were sacrificed 72 hours post-challenge to assess bacterial loads, cytokine responses, inflammation and microscopic lesions. A construct encoding a cellular secretion signal resulted in the most effective protection against melioidosis via tattooing, with a 10-fold reduction in bacterial loads in lungs and distant organs compared to the empty vector. Strikingly, a single intranasal administration of the same vaccine resulted in >1000-fold lower bacterial loads and increased survival. Pro-inflammatory cytokine responses were significantly diminished and strong reductions in markers for distant organ damage were observed. A rapid vaccination scheme using flagellin DNA tattoo provides significant protection against intranasal challenge with B. pseudomallei, markedly improved by a single administration via airway mucosa. Hence intranasal vaccination with flagellin-encoding DNA may be applicable when acute mass vaccination is indicated and warrants further testing.

  12. The concentrations of ambient Burkholderia pseudomallei during typhoon season in endemic area of melioidosis in Taiwan.

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    Ya-Lei Chen

    Full Text Available BACKGROUND: Melioidosis is a severe bacterial infection caused by Burkholderia pseudomallei with a high case-fatality rate. Epidemiological and animal studies show the possibility of inhalation transmission. However, no B. pseudomallei concentrations in ambient air have been researched. Here, we developed a method to quantify ambient B. pseudomallei and then measured concentrations of ambient B. pseudomallei during the typhoon season and the non-typhoon season to determine the factors influencing ambient B. pseudomallei levels. METHODS: We quantified ambient B. pseudomallei by using a filter/real-time qPCR method in the Zoynan Region in Kaohsiung, southern Taiwan. Twenty-four hour samples were collected at a sampling rate of 20 L/min every day from June 11 to December 21, 2012 including during the typhoon season (June to September and reference season (October to December. RESULTS: We successfully developed a filtration/real-time qPCR method to quantify ambient B. pseudomallei. To our knowledge, this is the first report describing concentrations of ambient B. pseudomallei. Ambient B. pseudomallei were only detected during the typhoon season when compared to the reference season. For the typhoons affecting the Zoynan Region, the positive rates of ambient B. pseudomallei were very high at 80% to 100%. During June to December, rainfall was positively correlated with ambient B. pseudomallei with a statistical significance. Sediment at a nearby pond significantly influenced the concentration of ambient B. pseudomallei. During the typhoon month, the typhoon was positively correlated with ambient B. pseudomallei whereas wind speed was reversely correlated with ambient B. pseudomallei. CONCLUSIONS: Our data suggest the possibility of transmission of B. pseudomallei via inhalation during the typhoon season.

  13. The concentrations of ambient Burkholderia pseudomallei during typhoon season in endemic area of melioidosis in Taiwan.

    Science.gov (United States)

    Chen, Ya-Lei; Yen, Yu-Chuan; Yang, Chun-Yuh; Lee, Min Sheng; Ho, Chi-Kung; Mena, Kristina D; Wang, Peng-Yau; Chen, Pei-Shih

    2014-01-01

    Melioidosis is a severe bacterial infection caused by Burkholderia pseudomallei with a high case-fatality rate. Epidemiological and animal studies show the possibility of inhalation transmission. However, no B. pseudomallei concentrations in ambient air have been researched. Here, we developed a method to quantify ambient B. pseudomallei and then measured concentrations of ambient B. pseudomallei during the typhoon season and the non-typhoon season to determine the factors influencing ambient B. pseudomallei levels. We quantified ambient B. pseudomallei by using a filter/real-time qPCR method in the Zoynan Region in Kaohsiung, southern Taiwan. Twenty-four hour samples were collected at a sampling rate of 20 L/min every day from June 11 to December 21, 2012 including during the typhoon season (June to September) and reference season (October to December). We successfully developed a filtration/real-time qPCR method to quantify ambient B. pseudomallei. To our knowledge, this is the first report describing concentrations of ambient B. pseudomallei. Ambient B. pseudomallei were only detected during the typhoon season when compared to the reference season. For the typhoons affecting the Zoynan Region, the positive rates of ambient B. pseudomallei were very high at 80% to 100%. During June to December, rainfall was positively correlated with ambient B. pseudomallei with a statistical significance. Sediment at a nearby pond significantly influenced the concentration of ambient B. pseudomallei. During the typhoon month, the typhoon was positively correlated with ambient B. pseudomallei whereas wind speed was reversely correlated with ambient B. pseudomallei. Our data suggest the possibility of transmission of B. pseudomallei via inhalation during the typhoon season.

  14. Epidemiological tracking and population assignment of the non-clonal bacterium, Burkholderia pseudomallei.

    Science.gov (United States)

    Dale, Julia; Price, Erin P; Hornstra, Heidie; Busch, Joseph D; Mayo, Mark; Godoy, Daniel; Wuthiekanun, Vanaporn; Baker, Anthony; Foster, Jeffrey T; Wagner, David M; Tuanyok, Apichai; Warner, Jeffrey; Spratt, Brian G; Peacock, Sharon J; Currie, Bart J; Keim, Paul; Pearson, Talima

    2011-12-01

    Rapid assignment of bacterial pathogens into predefined populations is an important first step for epidemiological tracking. For clonal species, a single allele can theoretically define a population. For non-clonal species such as Burkholderia pseudomallei, however, shared allelic states between distantly related isolates make it more difficult to identify population defining characteristics. Two distinct B. pseudomallei populations have been previously identified using multilocus sequence typing (MLST). These populations correlate with the major foci of endemicity (Australia and Southeast Asia). Here, we use multiple Bayesian approaches to evaluate the compositional robustness of these populations, and provide assignment results for MLST sequence types (STs). Our goal was to provide a reference for assigning STs to an established population without the need for further computational analyses. We also provide allele frequency results for each population to enable estimation of population assignment even when novel STs are discovered. The ability for humans and potentially contaminated goods to move rapidly across the globe complicates the task of identifying the source of an infection or outbreak. Population genetic dynamics of B. pseudomallei are particularly complicated relative to other bacterial pathogens, but the work here provides the ability for broad scale population assignment. As there is currently no independent empirical measure of successful population assignment, we provide comprehensive analytical details of our comparisons to enable the reader to evaluate the robustness of population designations and assignments as they pertain to individual research questions. Finer scale subdivision and verification of current population compositions will likely be possible with genotyping data that more comprehensively samples the genome. The approach used here may be valuable for other non-clonal pathogens that lack simple group-defining genetic characteristics

  15. Epidemiological tracking and population assignment of the non-clonal bacterium, Burkholderia pseudomallei.

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    Julia Dale

    2011-12-01

    Full Text Available Rapid assignment of bacterial pathogens into predefined populations is an important first step for epidemiological tracking. For clonal species, a single allele can theoretically define a population. For non-clonal species such as Burkholderia pseudomallei, however, shared allelic states between distantly related isolates make it more difficult to identify population defining characteristics. Two distinct B. pseudomallei populations have been previously identified using multilocus sequence typing (MLST. These populations correlate with the major foci of endemicity (Australia and Southeast Asia. Here, we use multiple Bayesian approaches to evaluate the compositional robustness of these populations, and provide assignment results for MLST sequence types (STs. Our goal was to provide a reference for assigning STs to an established population without the need for further computational analyses. We also provide allele frequency results for each population to enable estimation of population assignment even when novel STs are discovered. The ability for humans and potentially contaminated goods to move rapidly across the globe complicates the task of identifying the source of an infection or outbreak. Population genetic dynamics of B. pseudomallei are particularly complicated relative to other bacterial pathogens, but the work here provides the ability for broad scale population assignment. As there is currently no independent empirical measure of successful population assignment, we provide comprehensive analytical details of our comparisons to enable the reader to evaluate the robustness of population designations and assignments as they pertain to individual research questions. Finer scale subdivision and verification of current population compositions will likely be possible with genotyping data that more comprehensively samples the genome. The approach used here may be valuable for other non-clonal pathogens that lack simple group-defining genetic

  16. Inactivation of Toluene 2-Monooxygenase in Burkholderia cepacia G4 by Alkynes

    Science.gov (United States)

    Yeager, Chris M.; Bottomley, Peter J.; Arp, Daniel J.; Hyman, Michael R.

    1999-01-01

    High concentrations of acetylene (10 to 50% [vol/vol] gas phase) were required to inhibit the growth of Burkholderia cepacia G4 on toluene, while 1% (vol/vol) (gas phase) propyne or 1-butyne completely inhibited growth. Low concentrations of longer-chain alkynes (C5 to C10) were also effective inhibitors of toluene-dependent growth, and 2- and 3-alkynes were more potent inhibitors than their 1-alkyne counterparts. Exposure of toluene-grown B. cepacia G4 to alkynes resulted in the irreversible loss of toluene- and o-cresol-dependent O2 uptake activities, while acetate- and 3-methylcatechol-dependent O2 uptake activities were unaffected. Toluene-dependent O2 uptake decreased upon the addition of 1-butyne in a concentration- and time-dependent manner. The loss of activity followed first-order kinetics, with apparent rate constants ranging from 0.25 min−1 to 2.45 min−1. Increasing concentrations of toluene afforded protection from the inhibitory effects of 1-butyne. Furthermore, oxygen, supplied as H2O2, was required for inhibition by 1-butyne. These results suggest that alkynes are specific, mechanism-based inactivators of toluene 2-monooxygenase in B. cepacia G4, although the simplest alkyne, acetylene, was relatively ineffective compared to longer alkynes. Alkene analogs of acetylene and propyne—ethylene and propylene—were not inactivators of toluene 2-monooxygenase activity in B. cepacia G4 but were oxidized to their respective epoxides, with apparent Ks and Vmax values of 39.7 μM and 112.3 nmol min−1 mg of protein−1 for ethylene and 32.3 μM and 89.2 nmol min−1 mg of protein−1 for propylene. PMID:9925593

  17. Homology modeling and protein engineering of alkane monooxygenase in Burkholderia thailandensis MSMB121: in silico insights.

    Science.gov (United States)

    Jain, Chakresh Kumar; Gupta, Money; Prasad, Yamuna; Wadhwa, Gulshan; Sharma, Sanjeev Kumar

    2014-07-01

    The degradation of hydrocarbons plays an important role in the eco-balancing of petroleum products, pesticides and other toxic products in the environment. The degradation of hydrocarbons by microbes such as Geobacillus thermodenitrificans, Burkhulderia, Gordonia sp. and Acinetobacter sp. has been studied intensively in the literature. The present study focused on the in silico protein engineering of alkane monooxygenase (ladA)-a protein involved in the alkane degradation pathway. We demonstrated the improvement in substrate binding energy with engineered ladA in Burkholderia thailandensis MSMB121. We identified an ortholog of ladA monooxygenase found in B. thailandensis MSMB121, and showed it to be an enzyme involved in an alkane degradation pathway studied extensively in Geobacillus thermodenitrificans. Homology modeling of the three-dimensional structure of ladA was performed with a crystal structure (protein databank ID: 3B9N) as a template in MODELLER 9v11, and further validated using PROCHECK, VERIFY-3D and WHATIF tools. Specific amino acids were substituted in the region corresponding to amino acids 305-370 of ladA protein, resulting in an enhancement of binding energy in different alkane chain molecules as compared to wild protein structures in the docking experiments. The substrate binding energy with the protein was calculated using Vina (Implemented in VEGAZZ). Molecular dynamics simulations were performed to study the dynamics of different alkane chain molecules inside the binding pockets of wild and mutated ladA. Here, we hypothesize an improvement in binding energies and accessibility of substrates towards engineered ladA enzyme, which could be further facilitated for wet laboratory-based experiments for validation of the alkane degradation pathway in this organism.

  18. Activation of MAPK/ERK signaling by Burkholderia pseudomallei cycle inhibiting factor (Cif.

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    Mei Ying Ng

    Full Text Available Cycle inhibiting factors (Cifs are virulence proteins secreted by the type III secretion system of some Gram-negative pathogenic bacteria including Burkholderia pseudomallei. Cif is known to function to deamidate Nedd8, leading to inhibition of Cullin E3 ubiquitin ligases (CRL and consequently induction of cell cycle arrest. Here we show that Cif can function as a potent activator of MAPK/ERK signaling without significant activation of other signaling pathways downstream of receptor tyrosine kinases. Importantly, we found that the ability of Cif to activate ERK is dependent on its deamidase activity, but independent of Cullin E3 ligase inhibition. This suggests that apart from Nedd8, other cellular targets of Cif-dependent deamidation exist. We provide evidence that the mechanism involved in Cif-mediated ERK activation is dependent on recruitment of the Grb2-SOS1 complex to the plasma membrane. Further investigation revealed that Cif appears to modify the phosphorylation status of SOS1 in a region containing the CDC25-H and proline-rich domains. It is known that prolonged Cullin E3 ligase inhibition leads to cellular apoptosis. Therefore, we hypothesize that ERK activation is an important mechanism to counter the pro-apoptotic effects of Cif. Indeed, we show that Cif dependent ERK activation promotes phosphorylation of the proapoptotic protein Bim, thereby potentially conferring a pro-survival signal. In summary, we identified a novel deamidation-dependent mechanism of action of the B. pseudomallei virulence factor Cif/CHBP to activate MAPK/ERK signaling. Our study demonstrates that bacterial proteins such as Cif can serve as useful molecular tools to uncover novel aspects of mammalian signaling pathways.

  19. Antimicrobial activity of Tachyplesin 1 against Burkholderia pseudomallei: an in vitro and in silico approach

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    Lyn-Fay Lee

    2016-10-01

    Full Text Available Burkholderia pseudomallei, the causative agent of melioidosis, is intrinsically resistant to many conventional antibiotics. Therefore, alternative antimicrobial agents such as antimicrobial peptides (AMPs are extensively studied to combat this issue. Our study aims to identify and understand the mode of action of the potential AMP(s that are effective against B. pseudomallei in both planktonic and biofilm state as well as to predict the possible binding targets on using in vitro and in silico approaches. In the in vitro study, 11 AMPs were tested against 100 B. pseudomallei isolates for planktonic cell susceptibility, where LL-37, and PG1, demonstrated 100.0% susceptibility and TP1 demonstrated 83% susceptibility. Since the B. pseudomallei activity was reported on LL-37 and PG1, TP1 was selected for further investigation. TP1 inhibited B. pseudomallei cells at 61.69 μM, and membrane blebbing was observed using scanning electron microscopy. Moreover, TP1 inhibited B. pseudomallei cell growth, reaching bactericidal endpoint within 2 h post exposure as compared to ceftazidime (CAZ (8 h. Furthermore, TP1 was shown to suppress the growth of B. pseudomallei cells in biofilm state at concentrations above 221 μM. However, TP1 was cytotoxic to the mammalian cell lines tested. In the in silico study, molecular docking revealed that TP1 demonstrated a strong interaction to the common peptide or inhibitor binding targets for lipopolysaccharide of Escherichia coli, as well as autolysin, pneumolysin, and pneumococcal surface protein A (PspA of Streptococcus pneumoniae. Homology modelled B. pseudomallei PspA protein (YDP also showed a favourable binding with a strong electrostatic contribution and nine hydrogen bonds. In conclusion, TP1 demonstrated a good potential as an anti-B. pseudomallei agent.

  20. Phenotypic and functional characterization of human memory T cell responses to Burkholderia pseudomallei.

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    Patcharaporn Tippayawat

    Full Text Available Infection with the Gram-negative bacterium Burkholderia pseudomallei is an important cause of community-acquired lethal sepsis in endemic regions in southeast Asia and northern Australia and is increasingly reported in other tropical areas. In animal models, production of interferon-gamma (IFN-gamma is critical for resistance, but in humans the characteristics of IFN-gamma production and the bacterial antigens that are recognized by the cell-mediated immune response have not been defined.Peripheral blood from 133 healthy individuals who lived in the endemic area and had no history of melioidosis, 60 patients who had recovered from melioidosis, and 31 other patient control subjects were stimulated by whole bacteria or purified bacterial proteins in vitro, and IFN-gamma responses were analyzed by ELISPOT and flow cytometry.B. pseudomallei was a potent activator of human peripheral blood NK cells for innate production of IFN-gamma. In addition, healthy individuals with serological evidence of exposure to B. pseudomallei and patients recovered from active melioidosis developed CD4(+ (and CD8(+ T cells that recognized whole bacteria and purified proteins LolC, OppA, and PotF, members of the B. pseudomallei ABC transporter family. This response was primarily mediated by terminally differentiated T cells of the effector-memory (T(EMRA phenotype and correlated with the titer of anti-B. pseudomallei antibodies in the serum.Individuals living in a melioidosis-endemic region show clear evidence of T cell priming for the ability to make IFN-gamma that correlates with their serological status. The ability to detect T cell responses to defined B. pseudomallei proteins in large numbers of individuals now provides the opportunity to screen candidate antigens for inclusion in protein or polysaccharide-conjugate subunit vaccines against this important but neglected disease.

  1. Manganese scavenging and oxidative stress response mediated by type VI secretion system in Burkholderia thailandensis.

    Science.gov (United States)

    Si, Meiru; Zhao, Chao; Burkinshaw, Brianne; Zhang, Bing; Wei, Dawei; Wang, Yao; Dong, Tao G; Shen, Xihui

    2017-03-14

    Type VI secretion system (T6SS) is a versatile protein export machinery widely distributed in Gram-negative bacteria. Known to translocate protein substrates to eukaryotic and prokaryotic target cells to cause cellular damage, the T6SS has been primarily recognized as a contact-dependent bacterial weapon for microbe-host and microbial interspecies competition. Here we report contact-independent functions of the T6SS for metal acquisition, bacteria competition, and resistance to oxidative stress. We demonstrate that the T6SS-4 in Burkholderia thailandensis is critical for survival under oxidative stress and is regulated by OxyR, a conserved oxidative stress regulator. The T6SS-4 is important for intracellular accumulation of manganese (Mn 2+ ) under oxidative stress. Next, we identified a T6SS-4-dependent Mn 2+ -binding effector TseM, and its interacting partner MnoT, a Mn 2+ -specific TonB-dependent outer membrane transporter. Similar to the T6SS-4 genes, expression of mnoT is regulated by OxyR and is induced under oxidative stress and low Mn 2+ conditions. Both TseM and MnoT are required for efficient uptake of Mn 2+ across the outer membrane under Mn 2+ -limited and -oxidative stress conditions. The TseM-MnoT-mediated active Mn 2+ transport system is also involved in contact-independent bacteria-bacteria competition and bacterial virulence. This finding provides a perspective for understanding the mechanisms of metal ion uptake and the roles of T6SS in bacteria-bacteria competition.

  2. Environmental Free-Living Amoebae Isolated from Soil in Khon Kaen, Thailand, Antagonize Burkholderia pseudomallei.

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    Parumon Noinarin

    Full Text Available Presence of Burkholderia pseudomallei in soil and water is correlated with endemicity of melioidosis in Southeast Asia and northern Australia. Several biological and physico-chemical factors have been shown to influence persistence of B. pseudomallei in the environment of endemic areas. This study was the first to evaluate the interaction of B. pseudomallei with soil amoebae isolated from B. pseudomallei-positive soil site in Khon Kaen, Thailand. Four species of amoebae, Paravahlkampfia ustiana, Acanthamoeba sp., Naegleria pagei, and isolate A-ST39-E1, were isolated, cultured and identified based on morphology, movement and 18S rRNA gene sequence. Co-cultivation combined with a kanamycin-protection assay of B. pseudomallei with these amoebae at MOI 20 at 30°C were evaluated during 0-6 h using the plate count technique on Ashdown's agar. The fate of intracellular B. pseudomallei in these amoebae was also monitored by confocal laser scanning microscopy (CLSM observation of the CellTracker™ Orange-B. pseudomallei stained cells. The results demonstrated the ability of P. ustiana, Acanthamoeba sp. and isolate A-ST39-E1 to graze B. pseudomallei. However, the number of internalized B. pseudomallei substantially decreased and the bacterial cells disappeared during the observation period, suggesting they had been digested. We found that B. pseudomallei promoted the growth of Acanthamoeba sp. and isolate A-ST39-E1 in co-cultures at MOI 100 at 30°C, 24 h. These findings indicated that P. ustiana, Acanthamoeba sp. and isolate A-ST39-E1 may prey upon B. pseudomallei rather than representing potential environmental reservoirs in which the bacteria can persist.

  3. Solubilisation of inorganic phosphates by inoculant strains from tropical legumes

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    Leandro Marciano Marra

    2011-10-01

    Full Text Available Microbial solubilisation of low soluble inorganic phosphates is an important process contributing for the phosphorus available to plants in tropical soils. This study evaluates the ability of inoculant strains for tropical legumes to solubilise inorganic phosphates of low solubility that are found in tropical soils. Seven strains of Leguminosae nodulating bacteria (LNB were compared with one another and with a non-nodulating positive control, Burkholderia cepacia (LMG 1222T. Four of the strains are used as inoculants for cowpeas (Vigna unguiculata (Bradyrhizobium sp. UFLA 03-84; Bradyrhizobium elkani INPA 03-11B and Bradyrhizobium japonicum BR3267 or for common beans (Phaseolus vulgaris (Rhizobium tropici CIAT 899T. Rhizobium etli UFLA 02-100 and Rhizobium leguminosarum 316C10a are also efficient nodulators of beans and Cupriavidus taiwanensis LMG 19424T nodulates on Mimosa pudica. Two experiments, with solid and liquid media, were performed to determine whether the strains were able to solubilise CaHPO4, Al(H2PO43 or FePO4.2H2O. On solid GELP medium none of the strains dissolved FePO4.2H2O, but LMG 1222, UFLA 03-84 and CIAT 899 solubilised CaHPO4 particularly well. These strains, along with LMG 19424 and BR 3267, were also able to increase the solubility of Al(H2PO43. In liquid GELP medium, LMG 1222 solubilised all phosphate sources, but no legume nodulating strain could increase the solubility of Al(H2PO43. The strains CIAT 899 and UFLA 02-100 were the only legume nodulating bacteria able to solubilise the other phosphate sources in liquid media, dissolving both CaHPO4 and FePO4.2H2O. There was a negative correlation between the pH of the culture medium and the concentration of soluble phosphate when the phosphorus source was CaHPO4 or FePO4.2H2O. The contribution of these strains to increasing the phosphorus nutrition of legumes and non-legume plant species should be investigated further by in vivo experiments.

  4. The Animal Pathogen-Like Type III Secretion System is Required for the Intracellular Survival of Burkholderia mallei within J774.2 Macrophages

    Science.gov (United States)

    2006-03-30

    for B. mallei are horses, donkeys, and mules (solipeds), but other animals, including mice, hamsters, guinea pigs, monkeys , lions, and dogs, are...Spa/Prg and Shigella Ipa/Mxi/Spa TTS networks, are important for in vitro and in vivo survival of these pathogenic Burkholderia species (9–12, 14, 15

  5. Burkholderia sacchari DSM 17165: A source of compositionally-tunable block-copolymeric short-chain poly(hydroxyalkanoates) from xylose and levulinic acid

    Science.gov (United States)

    Burkholderia sacchari DSM 17165 was used as a biocatalyst for the production of poly-3-hydroxybutyrate-co-3-hydroxyvalerate block copolymers (Poly-3HB-block-3HV) from xylose and levulinic acid. Among the carbon source mixtures, levulinic acid was preferred and was consumed early in the fermentations...

  6. HemX is required for production of 2-ketogluconate, the predominant organic anion required for inorganic phosphate solubilization by Burkholderia sp. Ha185.

    Science.gov (United States)

    Hsu, Pei-Chun Lisa; Condron, Leo; O'Callaghan, Maureen; Hurst, Mark R H

    2015-12-01

    The bacterium Burkholderia sp. Ha185 readily solubilizes inorganic phosphate by releasing the low molecular weight organic anion, 2-ketogluconate. Using random transposon mutagenesis and in silico analysis, a mutation that caused almost complete abolition of phosphate solubilization was located within hemX, which is part of the hem operon. Burkholderia sp. Ha185 HemX is a multidomain protein, predicted to encode a bifunctional uroporphyrinogen-III synthetase/uroporphyrin-III C-methyltransferase, which has not previously been implicated in phosphate solubilization. Complementation of hemX restored the ability of the mutant to solubilize phosphate in both plate and liquid cultures. Based on a combination of organic-anion profiling, quantitative polymerase chain reaction and in silico analyses, hemX was confirmed to be solely responsible for hydroxyapatite solubilization in Burkholderia sp. Ha185. It is proposed that the biosynthesis of a yet to be determined redox cofactor by HemX is the main pathway for generating 2-ketogluconate via a haem-dependent gluconate 2-dehydrogenase in Burkholderia sp. Ha185. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  7. Differentiation of pulmonary bacterial pathogens in cystic fibrosis by volatile metabolites emitted by their in vitro cultures: Pseudomonas aeruginosa, Staphylococcus aureus, Stenotrophomonas maltophilia and the Burkholderia cepacia complex

    Czech Academy of Sciences Publication Activity Database

    Dryahina, Kseniya; Sovová, Kristýna; Nemec, A.; Španěl, Patrik

    2016-01-01

    Roč. 10, AUG 2016 (2016), s. 037102 ISSN 1752-7155 R&D Projects: GA ČR(CZ) GA14-14534S Institutional support: RVO:61388955 Keywords : Burkholderia cepacia complex * Pseudomonas aeruginosa * cystic fibrosis Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 4.318, year: 2016

  8. Proteogenomic Characterization of Monocyclic Aromatic Hydrocarbon Degradation Pathways in the Aniline-Degrading Bacterium Burkholderia sp. K24.

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    Sang-Yeop Lee

    Full Text Available Burkholderia sp. K24, formerly known as Acinetobacter lwoffii K24, is a soil bacterium capable of utilizing aniline as its sole carbon and nitrogen source. Genomic sequence analysis revealed that this bacterium possesses putative gene clusters for biodegradation of various monocyclic aromatic hydrocarbons (MAHs, including benzene, toluene, and xylene (BTX, as well as aniline. We verified the proposed MAH biodegradation pathways by dioxygenase activity assays, RT-PCR, and LC/MS-based quantitative proteomic analyses. This proteogenomic approach revealed four independent degradation pathways, all converging into the citric acid cycle. Aniline and p-hydroxybenzoate degradation pathways converged into the β-ketoadipate pathway. Benzoate and toluene were degraded through the benzoyl-CoA degradation pathway. The xylene isomers, i.e., o-, m-, and p-xylene, were degraded via the extradiol cleavage pathways. Salicylate was degraded through the gentisate degradation pathway. Our results show that Burkholderia sp. K24 possesses versatile biodegradation pathways, which may be employed for efficient bioremediation of aniline and BTX.

  9. Proteogenomic Characterization of Monocyclic Aromatic Hydrocarbon Degradation Pathways in the Aniline-Degrading Bacterium Burkholderia sp. K24

    Science.gov (United States)

    Yun, Sung Ho; Choi, Chi-Won; Yi, Yoon-Sun; Kim, Jonghyun; Chung, Young-Ho; Park, Edmond Changkyun; Kim, Seung Il

    2016-01-01

    Burkholderia sp. K24, formerly known as Acinetobacter lwoffii K24, is a soil bacterium capable of utilizing aniline as its sole carbon and nitrogen source. Genomic sequence analysis revealed that this bacterium possesses putative gene clusters for biodegradation of various monocyclic aromatic hydrocarbons (MAHs), including benzene, toluene, and xylene (BTX), as well as aniline. We verified the proposed MAH biodegradation pathways by dioxygenase activity assays, RT-PCR, and LC/MS-based quantitative proteomic analyses. This proteogenomic approach revealed four independent degradation pathways, all converging into the citric acid cycle. Aniline and p-hydroxybenzoate degradation pathways converged into the β-ketoadipate pathway. Benzoate and toluene were degraded through the benzoyl-CoA degradation pathway. The xylene isomers, i.e., o-, m-, and p-xylene, were degraded via the extradiol cleavage pathways. Salicylate was degraded through the gentisate degradation pathway. Our results show that Burkholderia sp. K24 possesses versatile biodegradation pathways, which may be employed for efficient bioremediation of aniline and BTX. PMID:27124467

  10. Increase of apatite dissolution rate by Scots pine roots associated or not with Burkholderia glathei PML1(12)Rp in open-system flow microcosms

    Science.gov (United States)

    Calvaruso, Christophe; Turpault, Marie-Pierre; Frey-Klett, Pascale; Uroz, Stéphane; Pierret, Marie-Claire; Tosheva, Zornitza; Kies, Antoine

    2013-04-01

    The release of nutritive elements through apatite dissolution represents the main source of phosphorus, calcium, and several micronutrients (e.g., Zn, Cu) for organisms in non-fertilized forest ecosystems. The aim of this study was to quantify, for the first time, the dissolution rate of apatite grains by tree roots that were or were not associated with a mineral weathering bacterial strain, and by various acids known to be produced by tree roots and soil bacterial strains in open-system flow microcosms. In addition, we explored whether the mobilization of trace elements (including rare earth elements) upon apatite dissolution was affected by the presence of trees and associated microorganisms. The dissolution rate of apatite by Scots pine plants that were or were not inoculated with the strain Burkholderia glathei PML1(12)Rp, and by inorganic (nitric) and organic (citric, oxalic and gluconic) acids at pH 5.5, 4.8, 3.8, 3.5, 3.0, and 2.0 was monitored in two controlled experiments: "plant-bacteria interaction" and "inorganic and organic acids". Analyses of the outlet solutions in the "plant-bacteria interaction" experiment showed that Scots pine roots and B. glathei PML1(12)Rp produced protons and organic acids such as gluconate, oxalate, acetate, and lactate. The weathering budget calculation revealed that Scots pines (with or without PML1(12)Rp) significantly increased (factor > 10) the release of Ca, P, As, Sr, Zn, U, Y, and rare earth elements such as Ce, La, Nd from apatite, compared to control abiotic treatment. Scanning electron microscopy observation confirmed traces of apatite dissolution in contact of roots. Most dissolved elements were taken up by Scots pine roots, i.e., approximately 50% of Ca, 70% of P, 30% of As, 70% of Sr, 90% of Zn, and 100% of U, Y, and rare earth elements. Interestingly, no significant additional effect due to the bacterial strain PML1(12)Rp on apatite dissolution and Scots pine nutrition and growth was observed. The "inorganic

  11. Interactions of Burkholderia terrae with soil fungi : Mechanisms, gene expression patterns and ecological behavior of Burkholderia terrae BS001 during its interaction with Lyophyllum sp. strain Karsten and Trichoderma asperellum 302 in soil

    NARCIS (Netherlands)

    Haq, Irshad U.

    2016-01-01

    Even though soil is a nutrient-limited environment, there are zones of high microbial activity. Mainly zones that are influenced by plants, fungi, or a combination of both, are of interest. The mycosphere (niche under the influence of fungi) is the zone where, in particular, bacterial-fungal

  12. Systematic mutagenesis of genes encoding predicted autotransported proteins of Burkholderia pseudomallei identifies factors mediating virulence in mice, net intracellular replication and a novel protein conferring serum resistance.

    Directory of Open Access Journals (Sweden)

    Natalie R Lazar Adler

    Full Text Available Burkholderia pseudomallei is the causative agent of the severe tropical disease melioidosis, which commonly presents as sepsis. The B. pseudomallei K96243 genome encodes eleven predicted autotransporters, a diverse family of secreted and outer membrane proteins often associated with virulence. In a systematic study of these autotransporters, we constructed insertion mutants in each gene predicted to encode an autotransporter and assessed them for three pathogenesis-associated phenotypes: virulence in the BALB/c intra-peritoneal mouse melioidosis model, net intracellular replication in J774.2 murine macrophage-like cells and survival in 45% (v/v normal human serum. From the complete repertoire of eleven autotransporter mutants, we identified eight mutants which exhibited an increase in median lethal dose of 1 to 2-log10 compared to the isogenic parent strain (bcaA, boaA, boaB, bpaA, bpaC, bpaE, bpaF and bimA. Four mutants, all demonstrating attenuation for virulence, exhibited reduced net intracellular replication in J774.2 macrophage-like cells (bimA, boaB, bpaC and bpaE. A single mutant (bpaC was identified that exhibited significantly reduced serum survival compared to wild-type. The bpaC mutant, which demonstrated attenuation for virulence and net intracellular replication, was sensitive to complement-mediated killing via the classical and/or lectin pathway. Serum resistance was rescued by in trans complementation. Subsequently, we expressed recombinant proteins of the passenger domain of four predicted autotransporters representing each of the phenotypic groups identified: those attenuated for virulence (BcaA, those attenuated for virulence and net intracellular replication (BpaE, the BpaC mutant with defects in virulence, net intracellular replication and serum resistance and those displaying wild-type phenotypes (BatA. Only BcaA and BpaE elicited a strong IFN-γ response in a restimulation assay using whole blood from seropositive donors

  13. Proteinograma sérico em muares naturalmente infectados pela Burkholderia mallei

    Directory of Open Access Journals (Sweden)

    Rinaldo Aparecido Mota

    2007-04-01

    Full Text Available A Burkholderia mallei é a bactéria causadora do mormo, doença de alta morbidade e letalidade para os eqüídeos, e também uma zoonose. Recentemente diagnosticada nos estados de Pernambuco e Alagoas, vem dizimando populações de eqüídeos na Região da Zona da Mata destes estados, causando grandes prejuízos à atividade canavieira que utiliza tais animais como tração. Objetivou-se com este trabalho investigar as alterações protéicas causadas pelo mormo. Foram estudados 90 muares adultos, de diferentes raças, destinados ao trabalho, provenientes da região canavieira, Zona da Mata, do Estado de Pernambuco. Estes foram divididos em três grupos: G1: composto por trinta animais sorologicamente negativos para o mormo; G2: composto por trinta animais sorologicamente positivos e sem sintomatologia clínica aparente e G3: composto por trinta animais sorologicamente positivos e com sintomatologia clínica aparente. Os resultados obtidos, referentes à média dos parâmetros estudados para G1, G2 e G3 foram respectivamente: proteína sérica total 7,33; 7,73 e 7,46g/dl; albumina 2,57; 2,43 e 1,81g/dl; globulinas 4,37; 4,86 e 5,64g/dl; relação albumina/globulinas 0,55; 0,47 e 0,34g/dl; alfa-globulina 1.06; 1.33 e 1,33g/dl; beta-globulina 1,10; 1,21 e 1,80g/dl e gama-globulina 2,21; 2,32 e 2,51g/dl. Conclui-se que as variações para os parâmetros estudados foram significativas, o aumento das globulinas caracteriza um estímulo antigênico nos animais positivos, bem como uma inversão na relação albumina/globulinas para os animais com clínica aparente em relação aos demais animais. Estes achados poderão ser considerados no diagnóstico, prognóstico e em pesquisas futuras que visem estudar formas de imunização contra esta importante enfermidade.

  14. Caspase-1-dependent and -independent cell death pathways in Burkholderia pseudomallei infection of macrophages.

    Directory of Open Access Journals (Sweden)

    Antje Bast

    2014-03-01

    Full Text Available The cytosolic pathogen Burkholderia pseudomallei and causative agent of melioidosis has been shown to regulate IL-1β and IL-18 production through NOD-like receptor NLRP3 and pyroptosis via NLRC4. Downstream signalling pathways of those receptors and other cell death mechanisms induced during B. pseudomallei infection have not been addressed so far in detail. Furthermore, the role of B. pseudomallei factors in inflammasome activation is still ill defined. In the present study we show that caspase-1 processing and pyroptosis is exclusively dependent on NLRC4, but not on NLRP3 in the early phase of macrophage infection, whereas at later time points caspase-1 activation and cell death is NLRC4- independent. In the early phase we identified an activation pathway involving caspases-9, -7 and PARP downstream of NLRC4 and caspase-1. Analyses of caspase-1/11-deficient infected macrophages revealed a strong induction of apoptosis, which is dependent on activation of apoptotic initiator and effector caspases. The early activation pathway of caspase-1 in macrophages was markedly reduced or completely abolished after infection with a B. pseudomallei flagellin FliC or a T3SS3 BsaU mutant. Studies using cells transfected with the wild-type and mutated T3SS3 effector protein BopE indicated also a role of this protein in caspase-1 processing. A T3SS3 inner rod protein BsaK mutant failed to activate caspase-1, revealed higher intracellular counts, reduced cell death and IL-1β secretion during early but not during late macrophage infection compared to the wild-type. Intranasal infection of BALB/c mice with the BsaK mutant displayed a strongly decreased mortality, lower bacterial loads in organs, and reduced levels of IL-1β, myeloperoxidase and neutrophils in bronchoalveolar lavage fluid. In conclusion, our results indicate a major role for a functional T3SS3 in early NLRC4-mediated caspase-1 activation and pyroptosis and a contribution of late caspase-1

  15. Quorum Sensing: A Transcriptional Regulatory System Involved in the Pathogenicity of Burkholderia mallei

    National Research Council Canada - National Science Library

    Ulrich, Ricky L; DeShazer, David; Hines, Harry B; Jeddeloh, Jeffrey A

    2004-01-01

    .... To determine if QS is involved in the virulence of B. mallei, we generated mutations in each putative luxIR homologue and tested the pathogenicities of the derivative strains in aerosol BALB/c mouse and intraperitoneal hamster models...

  16. Quorum Sensing: A transcriptional Regulatory System Involved in the Pathogenicity of Burkholderia mallei

    National Research Council Canada - National Science Library

    Ulrich, Ricky

    2004-01-01

    .... To determine if QS is involved in the virulence of B. mallei, we generated mutations in each putative luxIR homologue and tested the pathogenicities of the derivative strains in aerosol BALB/c mouse and intraperitoneal hamster models...

  17. Tandem Repeat Regions within the Burkholderia pseudomallei Genome and their Application for High-Resolution Genotyping

    National Research Council Canada - National Science Library

    U'Ren, Jana M; Schupp, James M; Pearson, Talima; Hornstra, Heidie; Friedman, Christine L; Smith, Kimothy L; Daugherty, Rebecca R; Rhoton, Shane D; Leadem, Ben; Georgia, Shalamar

    2007-01-01

    .... pseudomallei strain K96243 in order to develop a genetic typing method for B. pseudomallei. We then screened 104 of the potentially polymorphic loci across a diverse panel of isolates including B. pseudomallei, B. mallei and B. thailandensis...

  18. Strain-Modulated Epitaxy

    National Research Council Canada - National Science Library

    Brown, April

    1999-01-01

    Strain-Modulated Epitaxy (SME) is a novel approach, invented at Georgia Tech, to utilize subsurface stressors to control strain and therefore material properties and growth kinetics in the material above the stressors...

  19. Hamstring strain - aftercare

    Science.gov (United States)

    Pulled hamstring muscle; Sprain - hamstring ... There are 3 levels of hamstring strains: Grade 1 -- mild muscle strain or pull Grade 2 -- partial muscle tear Grade 3 -- complete muscle tear Recovery time depends ...

  20. Interleukin-12 induces a Th1-like response to Burkholderia mallei and limited protection in BALB/c mice.

    Science.gov (United States)

    Amemiya, Kei; Meyers, Jennifer L; Trevino, Sylvia R; Chanh, Tran C; Norris, Sarah L; Waag, David M

    2006-02-27

    We evaluated the effect of interleukin (IL)-12 on the immune response to Burkholderia mallei in BALB/c mice. Mice were vaccinated with non-viable B. mallei cells with or without IL-12. There was a seven- to nine-fold increase in IgG2a levels, and a significant increase in the proliferative response and interferon (IFN)-gamma production by splenocytes from mice that received B. mallei and IL-12. We saw an increase in survivors in the groups of mice that received B. mallei and IL-12 when challenged, compared to mice that received only B. mallei or IL-12. The results suggest that IL-12 can enhance the Th1-like immune response to B. mallei and mediate limited protection from a lethal challenge.

  1. Activity of a melimine derived peptide Mel4 against Stenotrophomonas, Delftia, Elizabethkingia, Burkholderia and biocompatibility as a contact lens coating

    DEFF Research Database (Denmark)

    Dutta, Debarun; Zhao, Timothy; Cheah, Kai Bing

    2017-01-01

    Purpose To determine the antimicrobial activity of the melimine derived peptide Mel4 against Delftia, Stenotrophomonas, Elizabethkingia, Burkholderia and to investigate biocompatibility of Mel4 as an antimicrobial coating on contact lenses in animals and humans. Methods In vitro antimicrobial...... activity of Mel4 was determined against the four Gram negative bacteria by investigating growth curves for 24 h followed by viable counts to determine the minimum inhibitory concentration (MIC). Contact lenses were coated by covalently binding Mel4, characterized by amino acid analysis, and were...... was active against all the bacteria tested (MIC50 ranged from 31–1000 μg ml−1) and produced an antimicrobial surface on contact lenses. Mel4-coating resulted hydrophilic surface without any significant change in contact lens parameters, and showed no signs of cytotoxicity or ocular irritation during rabbit...

  2. Effect of Azospirillum brasilense and Burkholderia unamae Bacteria on Maize Photosynthetic Activity Evaluated Using the Photoacoustic Technique

    Science.gov (United States)

    Gordillo-Delgado, F.; Marín, E.; Calderón, A.

    2016-09-01

    In this work, the photosynthetic process of maize plants ( Zea mays), which were grown using seeds inoculated with plant growth promoting bacteria Azospirillum brasilense and Burkholderia unamae, was monitored. Photothermal and photobaric signals obtained by a time-resolved photoacoustic measurement configuration were used for measuring the oxygen evolution rate in situ. A frequency-resolved configuration of the method was utilized to determine the oxygen diffusion coefficient and the thermal diffusivity of the maize leaves. The latter parameters, which can be used as indicators of the photosynthetic activity of maize, are found to vary according to the plant-microbe interaction. Treatment with plant growth promoting bacteria induced a decrease in the oxygen diffusion coefficient of about 20 %.

  3. Rapid Antimicrobial Susceptibility Testing of Bacillus anthracis, Yersinia pestis, and Burkholderia pseudomallei by Use of Laser Light Scattering Technology.

    Science.gov (United States)

    Bugrysheva, Julia V; Lascols, Christine; Sue, David; Weigel, Linda M

    2016-06-01

    Rapid methods to determine antimicrobial susceptibility would assist in the timely distribution of effective treatment or postexposure prophylaxis in the aftermath of the release of bacterial biothreat agents such as Bacillus anthracis, Yersinia pestis, or Burkholderia pseudomallei Conventional susceptibility tests require 16 to 48 h of incubation, depending on the bacterial species. We evaluated a method that is based on laser light scattering technology that measures cell density in real time. We determined that it has the ability to rapidly differentiate between growth (resistant) and no growth (susceptible) of several bacterial threat agents in the presence of clinically relevant antimicrobials. Results were available in 10 h of incubation. Use of laser scattering technology decreased the time required to determine antimicrobial susceptibility by 50% to 75% for B. anthracis, Y. pestis, and B. pseudomallei compared to conventional methods. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  4. Nanolipoprotein Particles (NLPs) as Versatile Vaccine Platforms for Co-delivery of Multiple Adjuvants with Subunit Antigens from Burkholderia spp. and F. tularensis - Technical Report

    Energy Technology Data Exchange (ETDEWEB)

    Fischer, N. O. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2015-01-13

    The goal of this proposal is to demonstrate that colocalization of protein subunit antigens and adjuvants on nanolipoprotein particles (NLPs) can increase the protective efficacy of subunit antigens from Burkholderia spp. and Francisella tularensis against an aerosol challenge. In the third quarter of the third year, F344 rats vaccinated with adjuvanted NLP formulations were challenged with F. tularensis SCHU S4 at Battelle. Preliminary data indicate that up to 65% of females vaccinated intranasally with an NLP-based formulation survived this challenge, compared to only 20% survival of naïve animals. In addition, NLPs were successfully formulated with Burkholderia protein antigens. IACUC approval for immunological assessments in BALB/c mice was received and we anticipate that these assessments will begin by March 2015, pending ACURO approval.

  5. A strain gauge

    DEFF Research Database (Denmark)

    2016-01-01

    The invention relates to a strain gauge of a carrier layer and a meandering measurement grid positioned on the carrier layer, wherein the strain gauge comprises two reinforcement members positioned on the carrier layer at opposite ends of the measurement grid in the axial direction....... The reinforcement members are each placed within a certain axial distance to the measurement grid with the axial distance being equal to or smaller than a factor times the grid spacing. The invention further relates to a multi-axial strain gauge such as a bi-axial strain gauge or a strain gauge rosette where each...... of the strain gauges comprises reinforcement members. The invention further relates to a method for manufacturing a strain gauge as mentioned above....

  6. Biorremediation of soil polluted by 75000 ppm of waste motor oil applying biostimulation and phytoremediation with Sorghum vulgare and Bacillus cereus or Burkholderia cepacia

    OpenAIRE

    Balderas-León Iván; Sánchez-Yáñez Juan Manuel

    2015-01-01

    Waste motor oil (WMO) pollutes soil and causing lost soil fertility. An alternative to solve this problem its bioremediation (BR) by double and following biostimulation (BS) with mineral solution (MS) and a legume as green manure (GM) then using phytoremediation (PR) with growth promoting vegetal bacteria (GPVB) like Bacillus cereus and Burkholderia cepacia to minimize remaining WMO. The aims of this research were: a) bioremediation of polluted soil by 75000 ppm of WMO by biostimulation and t...

  7. Nanolipoprotein Particles (NLPs) as Versatile Vaccine Platforms for Co-delivery of Multiple Adjuvants with Subunit Antigens from Burkholderia spp. and F. tularensis - Annual Technical Report

    Energy Technology Data Exchange (ETDEWEB)

    Fischer, N. O. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2015-04-16

    The goal of this proposal is to demonstrate that co-localization of protein subunit antigens and adjuvants on nanolipoprotein particles (NLPs) can increase the protective efficacy of recombinant subunit antigens from Burkholderia spp. and Francisella tularensis against an aerosol challenge. NLPs are are biocompatible, high-density lipoprotein mimetics that are amenable to the incorporation of multiple, chemically-disparate adjuvant and antigen molecules. We hypothesize that the ability to co-localize optimized adjuvant formulations with subunit antigens within a single particle will enhance the stimulation and activation of key immune effector cells, increasing the protective efficacy of subunit antigen-based vaccines. While Burkholderia spp. and F. tularensis subunit antigens are the focus of this proposal, we anticipate that this approach is applicable to a wide range of DOD-relevant biothreat agents. The F344 rat aerosol challenge model for F. tularensis has been successfully established at Battelle under this contract, and Year 3 efficacy studies performed at Battelle demonstrated that an NLP vaccine formulation was able to enhance survival of female F344 rats relative to naïve animals. In addition, Year 3 focused on the incorporation of multiple Burkholderia antigens (both polysaccharides and proteins) onto adjuvanted NLPs, with immunological analysis poised to begin in the next quarter.

  8. Three dimensional strained semiconductors

    Science.gov (United States)

    Voss, Lars; Conway, Adam; Nikolic, Rebecca J.; Leao, Cedric Rocha; Shao, Qinghui

    2016-11-08

    In one embodiment, an apparatus includes a three dimensional structure comprising a semiconductor material, and at least one thin film in contact with at least one exterior surface of the three dimensional structure for inducing a strain in the structure, the thin film being characterized as providing at least one of: an induced strain of at least 0.05%, and an induced strain in at least 5% of a volume of the three dimensional structure. In another embodiment, a method includes forming a three dimensional structure comprising a semiconductor material, and depositing at least one thin film on at least one surface of the three dimensional structure for inducing a strain in the structure, the thin film being characterized as providing at least one of: an induced strain of at least 0.05%, and an induced strain in at least 5% of a volume of the structure.

  9. Strain measurement technique

    International Nuclear Information System (INIS)

    1987-01-01

    The 10 contributions are concerned with selected areas of application, such as strain measurements in wood, rubber/metal compounds, sets of strain measurements on buildings, reinforced concrete structures without gaps, pipes buried in the ground and measurements of pressure fluctuations. To increase the availability and safety of plant, stress analyses were made on gas turbine rotors with HT-DMS or capacitive HT-DMS (high temperature strain measurements). (DG) [de

  10. Strained Silicon Photonics

    Directory of Open Access Journals (Sweden)

    Ralf B. Wehrspohn

    2012-05-01

    Full Text Available A review of recent progress in the field of strained silicon photonics is presented. The application of strain to waveguide and photonic crystal structures can be used to alter the linear and nonlinear optical properties of these devices. Here, methods for the fabrication of strained devices are summarized and recent examples of linear and nonlinear optical devices are discussed. Furthermore, the relation between strain and the enhancement of the second order nonlinear susceptibility is investigated, which may enable the construction of optically active photonic devices made of silicon.

  11. Burkholderia pseudomallei-derived miR-3473 enhances NF-κB via targeting TRAF3 and is associated with different inflammatory responses compared to Burkholderia thailandensis in murine macrophages.

    Science.gov (United States)

    Fang, Yao; Chen, Hai; Hu, Yi; Li, Qian; Hu, Zhiqiang; Ma, Tengfei; Mao, Xuhu

    2016-11-28

    Burkholderia pseudomallei (Bp) is the causative agent of melioidosis, a kind of tropical disease. Burkholderia thailandensis (Bt), with a high sequence similarity to Bp, is thought to be an avirulent organism. Since there are numerous similarities between Bp and Bt, their differences in pathogenesis of host response and related mechanism are still undermined. In recent years, microRNAs have been researched in many diseases, but seldom involved in bacterial infection, bacteria-host interaction or explaining the differences between virulent and avirulent species. We found that Bp and Bt had similar phenotypes in terms of intracellular replication, dissemination (reflected by multinucleated giant cell formation), TNF-α release and apoptosis in RAW264.7 macrophages or TC-1 pulmonary cell but in different level. Especially, at the late infection phases (after 12 h post infection), Bp showed faster intracellular growth, stronger cytotoxicity, and higher TNF-α release. After microRNA array analysis, we found some microRNAs were significantly expressed in macrophages treated by Bp. miR-3473 was one of them specifically induced, but not significantly changed in Bt-treated macrophages. In addition, TargetScan suggested that miR-3473 possibly target TRAF3 (TNF receptor-associated factor 3), a well-known negative regulator of the NF-κB pathway, which was probably involved in the TNF-α induction and apoptosis in cells with Bp infection. In vivo, it was found that miR-3473 expression of total lungs cells from Bp-treated was higher than that from Bt-treated mice. And miR-3473 inhibitor was able to decrease the TNF-α release of mice and prolong the survival of mice with Bp infection. In sum, miR-3473 plays an important role in the differential pathogenicity of Bp and Bt via miR-3473-TRAF3-TNF-α network, and regulates TNF-α release, cell apoptosis and animal survival after Bp treatment. In this study, we have found a specific microRNA is related to bacterial virulence and

  12. Regulator LdhR and d-Lactate Dehydrogenase LdhA of Burkholderia multivorans Play Roles in Carbon Overflow and in Planktonic Cellular Aggregate Formation.

    Science.gov (United States)

    Silva, Inês N; Ramires, Marcelo J; Azevedo, Lisa A; Guerreiro, Ana R; Tavares, Andreia C; Becker, Jörg D; Moreira, Leonilde M

    2017-10-01

    LysR-type transcriptional regulators (LTTRs) are the most commonly found regulators in Burkholderia cepacia complex, comprising opportunistic pathogens causing chronic respiratory infections in cystic fibrosis (CF) patients. Despite LTTRs being global regulators of pathogenicity in several types of bacteria, few have been characterized in Burkholderia Here, we show that gene ldhR of B. multivorans encoding an LTTR is cotranscribed with ldhA encoding a d-lactate dehydrogenase and evaluate their implication in virulence traits such as exopolysaccharide (EPS) synthesis and biofilm formation. A comparison of the wild type (WT) and its isogenic Δ ldhR mutant grown in medium with 2% d-glucose revealed a negative impact on EPS biosynthesis and on cell viability in the presence of LdhR. The loss of viability in WT cells was caused by intracellular acidification as a consequence of the cumulative secretion of organic acids, including d-lactate, which was absent from the Δ ldhR mutant supernatant. Furthermore, LdhR is implicated in the formation of planktonic cellular aggregates. WT cell aggregates reached 1,000 μm in size after 24 h in liquid cultures, in contrast to Δ ldhR mutant aggregates that never grew more than 60 μm. The overexpression of d-lactate dehydrogenase LdhA in the Δ ldhR mutant partially restored the formed aggregate size, suggesting a role for fermentation inside aggregates. Similar results were obtained for surface-attached biofilms, with WT cells producing more biofilm. A systematic evaluation of planktonic aggregates in Burkholderia CF clinical isolates showed aggregates in 40 of 74. As CF patients' lung environments are microaerophilic and bacteria are found as free aggregates/biofilms, LdhR and LdhA might have central roles in adapting to this environment. IMPORTANCE Cystic fibrosis patients often suffer from chronic respiratory infections caused by several types of microorganisms. Among them are the Burkholderia cepacia complex bacteria, which

  13. A strain gauge

    DEFF Research Database (Denmark)

    2017-01-01

    The invention relates to a strain gauge of a carrier layer and a meandering measurement grid (101) positioned on the carrier layer, wherein the measurement grid comprises a number of measurement grid sections placed side by side with gaps in between, and a number of end loops (106) interconnecting...... relates to a method for manufacturing a strain gauge as mentioned above....

  14. Evaluation of indigenous rhizobacterial strains with reduced dose of chemical fertilizer towards growth and yield of mustard (Brassica campestris under old alluvial soil zone of West Bengal, India

    Directory of Open Access Journals (Sweden)

    Shampa Dutta

    2017-12-01

    Full Text Available A field experiment had been carried out in the Crop Research and Seed Multiplication Farm of The University of Burdwan, West Bengal, India during the two consecutive winter seasons of 2011-2012 and 2012-2013 to study the effect of indigenous rhizospheric bacterial strains on growth, physiology and yield of mustard variety. Pseudomonas putida, Burkholderia cepacia, Burkholderia sp. and their mixture were used as seed inoculants for mustard cultivation. The experiment was laid down in a randomized complete block design (RCBD with three replications. Results revealed that indigenous inoculation (with reduced dose of chemical fertilizer significantly increased (p < 0.05 the yield of mustard as compared to uninoculated control (full recommended dose of NPK fertilizers. A combination treatment of biofertilizer and chemical fertilizer also increased plant height, plant biomass and other yield components compared to control. The comprehensive approach of plant growth promoting rhizobacteria (PGPR in agriculturally important crops should be carried out to explore the hidden potential of PGPR and to promote the quality and yield of crop under field conditions. Keywords: Indigenous rhizobacteria, Mustard, PGPR, Yield

  15. Differential sensitivity of polyhydroxyalkanoate producing bacteria to fermentation inhibitors and comparison of polyhydroxybutyrate production from Burkholderia cepacia and Pseudomonas pseudoflava.

    Science.gov (United States)

    Dietrich, Diane; Illman, Barbara; Crooks, Casey

    2013-06-04

    The aim of this study is determine the relative sensitivity of a panel of seven polyhydroxyalkanoate producing bacteria to a panel of seven lignocellulosic-derived fermentation inhibitors representing aliphatic acids, furans and phenolics. A further aim was to measure the polyhydroxybutyrate production of select organisms on lignocellulosic-derived monosaccharides arabinose, xylose, glucose and mannose. We examined the sensitivity of seven polyhydroxyalkanoate producing bacteria: Azohydromonas lata, Bacillus megaterium, Bacillus cereus, Burkholderia cepacia, Pseudomonas olevorans, Pseudomonas pseudoflava and Ralstonia eutropha, against seven fermentation inhibitors produced by the saccharification of lignocellulose: acetic acid, levulinic acid, coumaric acid, ferulic acid, syringaldehyde, furfural, and hyroxymethyfurfural. There was significant variation in the sensitivity of these microbes to representative phenolics ranging from 0.25-1.5 g/L coumaric and ferulic acid and between 0.5-6.0 g/L syringaldehyde. Inhibition ranged from 0.37-4 g/L and 0.75-6 g/L with acetic acid and levulinic acid, respectively. B. cepacia and P. pseudoflava were selected for further analysis of polyhydroxyalkanoate production. We find significant differences in sensitivity to the fermentation inhibitors tested and find these variations to be over a relevant concentration range given the concentrations of inhibitors typically found in lignocellulosic hydrolysates. Of the seven bacteria tested, B. cepacia demonstrated the greatest inhibitor tolerance. Similarly, of two organisms examined for polyhydroxybutyrate production, B. cepacia was notably more efficient when fermenting pentose substrates.

  16. Induction of Biofilm Formation in the Betaproteobacterium Burkholderia unamae CK43B Exposed to Exogenous Indole and Gallic Acid

    Science.gov (United States)

    Kim, Dongyeop; Sitepu, Irnayuli R.

    2013-01-01

    Burkholderia unamae CK43B, a member of the Betaproteobacteria that was isolated from the rhizosphere of a Shorea balangeran sapling in a tropical peat swamp forest, produces neither indole nor extracellular polymeric substances associated with biofilm formation. When cultured in a modified Winogradsky's medium supplemented with up to 1.7 mM indole, B. unamae CK43B maintains its planktonic state by cell swelling and effectively degrades exogenous indole. However, in medium supplemented with 1.7 mM exogenous indole and 1.0 mM gallic acid, B. unamae CK43B produced extracellular polymeric substances and formed a biofilm. The concentration indicated above of gallic acid alone had no effect on either the growth or the differentiation of B. unamae CK43B cells above a certain concentration threshold, whereas it inhibited indole degradation by B. unamae CK43B to 3-hydroxyindoxyl. In addition, coculture of B. unamae CK43B with indole-producing Escherichia coli in nutrient-rich Luria-Bertani medium supplemented with 1.0 mM gallic acid led to the formation of mixed cell aggregates. The viability and active growth of B. unamae CK43B cells in a coculture system with Escherichia coli were evidenced by fluorescence in situ hybridization. Our data thus suggest that indole facilitates intergenus communication between indole-producing gammaproteobacteria and some indole-degrading bacteria, particularly in gallic acid-rich environments. PMID:23747701

  17. Outbreak of Burkholderia cepacia complex bacteremia in a chemotherapy day care unit due to intrinsic contamination of an antiemetic drug

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    T Singhal

    2015-01-01

    Full Text Available Background: In the end of 2009, a large number of patients with cancer undergoing chemotherapy at the day care unit of a private hospital in Mumbai, India developed Burkholderia cepacia complex (BCC blood stream infection (BSI. Objective: The objectives were to identify the source of the outbreak and terminate the outbreak as rapidly as possible. Materials and Methods: All infection control protocols and processes were reviewed. Intensive training was started for all nursing staff involved in patient care. Cultures were sent from the environment (surfaces, water, air, intravenous fluids, disinfectants and antiseptics and opened/unopened medication. Results: A total of 13 patients with cancer with tunneled catheters were affected with BCC BSI. The isolates were of similar antimicrobial sensitivity. No significant breach of infection control protocols could be identified. Cultures from the prepared intravenous medication bags grew BCC. Subsequently, culture from unused vials of the antiemetic granisetron grew BCC, whereas those from the unopened IV fluid bag and chemotherapy medication were negative. On review, it was discovered that the outbreak started when a new brand of granisetron was introduced. The result was communicated to the manufacturer and the brand was withdrawn. There were no further cases. Conclusions: This outbreak was thus linked to intrinsic contamination of medication vials. We acknowledge a delay in identifying the source as we were concentrating more on human errors in medication preparation and less on intrinsic contamination. We recommend that in an event of an outbreak, unopened vials be cultured at the outset.

  18. Programmable DNA-binding proteins from Burkholderia provide a fresh perspective on the TALE-like repeat domain.

    Science.gov (United States)

    de Lange, Orlando; Wolf, Christina; Dietze, Jörn; Elsaesser, Janett; Morbitzer, Robert; Lahaye, Thomas

    2014-06-01

    The tandem repeats of transcription activator like effectors (TALEs) mediate sequence-specific DNA binding using a simple code. Naturally, TALEs are injected by Xanthomonas bacteria into plant cells to manipulate the host transcriptome. In the laboratory TALE DNA binding domains are reprogrammed and used to target a fused functional domain to a genomic locus of choice. Research into the natural diversity of TALE-like proteins may provide resources for the further improvement of current TALE technology. Here we describe TALE-like proteins from the endosymbiotic bacterium Burkholderia rhizoxinica, termed Bat proteins. Bat repeat domains mediate sequence-specific DNA binding with the same code as TALEs, despite less than 40% sequence identity. We show that Bat proteins can be adapted for use as transcription factors and nucleases and that sequence preferences can be reprogrammed. Unlike TALEs, the core repeats of each Bat protein are highly polymorphic. This feature allowed us to explore alternative strategies for the design of custom Bat repeat arrays, providing novel insights into the functional relevance of non-RVD residues. The Bat proteins offer fertile grounds for research into the creation of improved programmable DNA-binding proteins and comparative insights into TALE-like evolution. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  19. A genomic survey of positive selection in Burkholderia pseudomallei provides insights into the evolution of accidental virulence.

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    Tannistha Nandi

    2010-04-01

    Full Text Available Certain environmental microorganisms can cause severe human infections, even in the absence of an obvious requirement for transition through an animal host for replication ("accidental virulence". To understand this process, we compared eleven isolate genomes of Burkholderia pseudomallei (Bp, a tropical soil microbe and causative agent of the human and animal disease melioidosis. We found evidence for the existence of several new genes in the Bp reference genome, identifying 282 novel genes supported by at least two independent lines of supporting evidence (mRNA transcripts, database homologs, and presence of ribosomal binding sites and 81 novel genes supported by all three lines. Within the Bp core genome, 211 genes exhibited significant levels of positive selection (4.5%, distributed across many cellular pathways including carbohydrate and secondary metabolism. Functional experiments revealed that certain positively selected genes might enhance mammalian virulence by interacting with host cellular pathways or utilizing host nutrients. Evolutionary modifications improving Bp environmental fitness may thus have indirectly facilitated the ability of Bp to colonize and survive in mammalian hosts. These findings improve our understanding of the pathogenesis of melioidosis, and establish Bp as a model system for studying the genetics of accidental virulence.

  20. Biostimulation of soil polluted by 40000 ppm of waste motor oil and phytoremediation with Cicer arietinum and Burkholderia cepacia

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    Meza-Ramírez Janitzi Yunuén

    2016-08-01

    Full Text Available Soil polluted by 40000 ppm of waste residual oil (WRO, is a relative high hydrocarbons mix concentration according to Mexican regula