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1

Melioidosis Caused by Burkholderia pseudomallei in Drinking Water, Thailand, 2012  

PubMed Central

We identified 10 patients in Thailand with culture-confirmed melioidosis who had Burkholderia pseudomallei isolated from their drinking water. The multilocus sequence type of B. pseudomallei from clinical specimens and water samples were identical for 2 patients. This finding suggests that drinking water is a preventable source of B. pseudomallei infection. PMID:24447771

Wongsuvan, Gumphol; Aanensen, David; Ngamwilai, Sujittra; Saiprom, Natnaree; Rongkard, Patpong; Thaipadungpanit, Janjira; Kanoksil, Manas; Chantratita, Narisara; Day, Nicholas P.J.; Peacock, Sharon J.

2014-01-01

2

Burkholderia pseudomallei musculoskeletal infections (melioidosis) in India.  

PubMed

Melioidosis, an infection due to gram negative Burkholderia pseudomallei, is an important cause of sepsis in east Asia especially Thailand and northern Australia. It usually causes abscesses in lung, liver, spleen, skeletal muscle and parotids especially in patients with diabetes, chronic renal failure and thalassemia. Musculoskeletal melioidosis is not common in India even though sporadic cases have been reported mostly involving soft tissues. During a two-year-period, we had five patients with musculoskeletal melioidosis. All patients presented with multifocal osteomyelitis, recurrent osteomyelitis or septic arthritis. One patient died early because of septicemia and multi-organ failure. All patients were diagnosed on the basis of positive pus culture. All patients were treated by surgical debridement followed by a combination of antibiotics; (ceftazidime, amoxy-clavulanic acid, co-trimoxazole and doxycycline) for six months except for one who died due to fulminant septicemia. All other patients recovered completely with no recurrences. With increasing awareness and better diagnostic facilities, probably musculoskeletal melioidosis will be increasingly diagnosed in future. PMID:20419012

Pandey, Vivek; Rao, Sripathi P; Rao, Sugandhi; Acharya, Kiran Kv; Chhabra, Sarabjeet Singh

2010-04-01

3

Burkholderia pseudomallei musculoskeletal infections (melioidosis) in India  

PubMed Central

Melioidosis, an infection due to gram negative Burkholderia pseudomallei, is an important cause of sepsis in east Asia especially Thailand and northern Australia. It usually causes abscesses in lung, liver, spleen, skeletal muscle and parotids especially in patients with diabetes, chronic renal failure and thalassemia. Musculoskeletal melioidosis is not common in India even though sporadic cases have been reported mostly involving soft tissues. During a two-year-period, we had five patients with musculoskeletal melioidosis. All patients presented with multifocal osteomyelitis, recurrent osteomyelitis or septic arthritis. One patient died early because of septicemia and multi-organ failure. All patients were diagnosed on the basis of positive pus culture. All patients were treated by surgical debridement followed by a combination of antibiotics; (ceftazidime, amoxy-clavulanic acid, co-trimoxazole and doxycycline) for six months except for one who died due to fulminant septicemia. All other patients recovered completely with no recurrences. With increasing awareness and better diagnostic facilities, probably musculoskeletal melioidosis will be increasingly diagnosed in future. PMID:20419012

Pandey, Vivek; Rao, Sripathi P; Rao, Sugandhi; Acharya, Kiran KV; Chhabra, Sarabjeet Singh

2010-01-01

4

Burkholderia pseudomallei capsular polysaccharide conjugates provide protection against acute melioidosis.  

PubMed

Burkholderia pseudomallei, the etiologic agent of melioidosis, is a CDC tier 1 select agent that causes severe disease in both humans and animals. Diagnosis and treatment of melioidosis can be challenging, and in the absence of optimal chemotherapeutic intervention, acute disease is frequently fatal. Melioidosis is an emerging infectious disease for which there are currently no licensed vaccines. Due to the potential malicious use of B. pseudomallei as well as its impact on public health in regions where the disease is endemic, there is significant interest in developing vaccines for immunization against this disease. In the present study, type A O-polysaccharide (OPS) and manno-heptose capsular polysaccharide (CPS) antigens were isolated from nonpathogenic, select-agent-excluded strains of B. pseudomallei and covalently linked to carrier proteins. By using these conjugates (OPS2B1 and CPS2B1, respectively), it was shown that although high-titer IgG responses against the OPS or CPS component of the glycoconjugates could be raised in BALB/c mice, only those animals immunized with CPS2B1 were protected against intraperitoneal challenge with B. pseudomallei. Extending upon these studies, it was also demonstrated that when the mice were immunized with a combination of CPS2B1 and recombinant B. pseudomallei LolC, rather than with CPS2B1 or LolC individually, they exhibited higher survival rates when challenged with a lethal dose of B. pseudomallei. Collectively, these results suggest that CPS-based glycoconjugates are promising candidates for the development of subunit vaccines for immunization against melioidosis. PMID:24866807

Scott, Andrew E; Burtnick, Mary N; Stokes, Margaret G M; Whelan, Adam O; Williamson, E Diane; Atkins, Timothy P; Prior, Joann L; Brett, Paul J

2014-08-01

5

Less Is More: Burkholderia pseudomallei and Chronic Melioidosis  

PubMed Central

ABSTRACT The Gram-negative bacterium Burkholderia pseudomallei is the causative agent of melioidosis, a serious infectious disease of humans and animals. Once considered an esoteric tropical disease confined to Southeast Asia and northern Australia, research on B. pseudomallei has recently gained global prominence due to its classification as a potential bioterrorism agent by countries such as the United States and also by increasing numbers of case reports from regions where it is not endemic. An environmental bacterium typically found in soil and water, assessing the true global prevalence of melioidosis is challenged by the fact that clinical symptoms associated with B. pseudomallei infection are extremely varied and may be confused with diverse conditions such as lung cancer, tuberculosis, or Staphyloccocus aureus infection. These diagnostic challenges, coupled with lack of awareness among clinicians, have likely contributed to underdiagnosis and the high mortality rate of melioidosis, as initial treatment is often either inappropriate or delayed. Even after antibiotic treatment, relapses are frequent, and after resolution of acute symptoms, chronic melioidosis can also occur, and the symptoms can persist for months to years. In a recent article, Price et al. [mBio 4(4):e00388-13, 2013, doi:10.1128/mBio.00388-13] demonstrate how comparative genomic sequencing can reveal the repertoire of genetic changes incurred by B. pseudomallei during chronic human infection. Their results have significant clinical ramifications and highlight B. pseudomallei’s ability to survive in a wide range of potential niches within hosts, through the acquisition of genetic adaptations that optimize fitness and resource utilization. PMID:24065633

Nandi, Tannistha; Tan, Patrick

2013-01-01

6

Less is more: Burkholderia pseudomallei and chronic melioidosis.  

PubMed

The Gram-negative bacterium Burkholderia pseudomallei is the causative agent of melioidosis, a serious infectious disease of humans and animals. Once considered an esoteric tropical disease confined to Southeast Asia and northern Australia, research on B. pseudomallei has recently gained global prominence due to its classification as a potential bioterrorism agent by countries such as the United States and also by increasing numbers of case reports from regions where it is not endemic. An environmental bacterium typically found in soil and water, assessing the true global prevalence of melioidosis is challenged by the fact that clinical symptoms associated with B. pseudomallei infection are extremely varied and may be confused with diverse conditions such as lung cancer, tuberculosis, or Staphyloccocus aureus infection. These diagnostic challenges, coupled with lack of awareness among clinicians, have likely contributed to underdiagnosis and the high mortality rate of melioidosis, as initial treatment is often either inappropriate or delayed. Even after antibiotic treatment, relapses are frequent, and after resolution of acute symptoms, chronic melioidosis can also occur, and the symptoms can persist for months to years. In a recent article, Price et al. [mBio 4(4):e00388-13, 2013, doi:10.1128/mBio.00388-13] demonstrate how comparative genomic sequencing can reveal the repertoire of genetic changes incurred by B. pseudomallei during chronic human infection. Their results have significant clinical ramifications and highlight B. pseudomallei's ability to survive in a wide range of potential niches within hosts, through the acquisition of genetic adaptations that optimize fitness and resource utilization. PMID:24065633

Nandi, Tannistha; Tan, Patrick

2013-01-01

7

Genomic plasticity of the causative agent of melioidosis, Burkholderia pseudomallei  

PubMed Central

Burkholderia pseudomallei is a recognized biothreat agent and the causative agent of melioidosis. This Gram-negative bacterium exists as a soil saprophyte in melioidosis-endemic areas of the world and accounts for 20% of community-acquired septicaemias in northeastern Thailand where half of those affected die. Here we report the complete genome of B. pseudomallei, which is composed of two chromosomes of 4.07 megabase pairs and 3.17 megabase pairs, showing significant functional partitioning of genes between them. The large chromosome encodes many of the core functions associated with central metabolism and cell growth, whereas the small chromosome carries more accessory functions associated with adaptation and survival in different niches. Genomic comparisons with closely and more distantly related bacteria revealed a greater level of gene order conservation and a greater number of orthologous genes on the large chromosome, suggesting that the two replicons have distinct evolutionary origins. A striking feature of the genome was the presence of 16 genomic islands (GIs) that together made up 6.1% of the genome. Further analysis revealed these islands to be variably present in a collection of invasive and soil isolates but entirely absent from the clonally related organism B. mallei. We propose that variable horizontal gene acquisition by B. pseudomallei is an important feature of recent genetic evolution and that this has resulted in a genetically diverse pathogenic species. PMID:15377794

Holden, Matthew T. G.; Titball, Richard W.; Peacock, Sharon J.; Cerdeno-Tarraga, Ana M.; Atkins, Timothy; Crossman, Lisa C.; Pitt, Tyrone; Churcher, Carol; Mungall, Karen; Bentley, Stephen D.; Sebaihia, Mohammed; Thomson, Nicholas R.; Bason, Nathalie; Beacham, Ifor R.; Brooks, Karen; Brown, Katherine A.; Brown, Nat F.; Challis, Greg L.; Cherevach, Inna; Chillingworth, Tracy; Cronin, Ann; Crossett, Ben; Davis, Paul; DeShazer, David; Feltwell, Theresa; Fraser, Audrey; Hance, Zahra; Hauser, Heidi; Holroyd, Simon; Jagels, Kay; Keith, Karen E.; Maddison, Mark; Moule, Sharon; Price, Claire; Quail, Michael A.; Rabbinowitsch, Ester; Rutherford, Kim; Sanders, Mandy; Simmonds, Mark; Songsivilai, Sirirurg; Stevens, Kim; Tumapa, Sarinna; Vesaratchavest, Monkgol; Whitehead, Sally; Yeats, Corin; Barrell, Bart G.; Oyston, Petra C. F.; Parkhill, Julian

2004-01-01

8

Genomic patterns of pathogen evolution revealed by comparison of Burkholderia pseudomallei, the causative agent of melioidosis, to avirulent Burkholderia thailandensis  

E-print Network

Background: The Gram-negative bacterium Burkholderia pseudomallei (Bp) is the causative agent of the human disease melioidosis. To understand the evolutionary mechanisms contributing to Bp virulence, we performed a comparative ...

Yu, Yiting

9

Multilocus Sequence Typing and Evolutionary Relationships among the Causative Agents of Melioidosis and Glanders, Burkholderia pseudomallei and Burkholderia mallei  

Microsoft Academic Search

A collection of 147 isolates of Burkholderia pseudomallei, B. mallei, and B. thailandensis was characterized by multilocus sequence typing (MLST). The 128 isolates of B. pseudomallei, the causative agent of melioidosis, were obtained from diverse geographic locations, from humans and animals with disease, and from the environment and were resolved into 71 sequence types. The utility of the MLST scheme

Daniel Godoy; Gaynor Randle; Andrew J. Simpson; David M. Aanensen; Tyrone L. Pitt; Reimi Kinoshita; Brian G. Spratt

2003-01-01

10

Mechanisms of antibiotic resistance in Burkholderia pseudomallei: implications for treatment of melioidosis  

PubMed Central

Burkholderia pseudomallei is the etiologic agent of melioidosis. This multifaceted disease is difficult to treat, resulting in high morbidity and mortality. Treatment of B. pseudomallei infections is lengthy and necessitates an intensive phase (parenteral ceftazidime, amoxicillin–clavulanic acid or meropenem) and an eradication phase (oral trimethoprim–sulfamethoxazole). The main resistance mechanisms affecting these antibiotics include enzymatic inactivation, target deletion and efflux from the cell, and are mediated by chromosomally encoded genes. Overproduction and mutations in the class A PenA ?-lactamase cause ceftazidime and amoxicillin–clavulanic acid resistance. Deletion of the penicillin binding protein 3 results in ceftazidime resistance. BpeEF–OprC efflux pump expression causes trimethoprim and trimethoprim–sulfamethoxazole resistance. Although resistance is still relatively rare, therapeutic efficacies may be compromised by resistance emergence due to increased use of antibiotics in endemic regions. Novel agents and therapeutic strategies are being tested and, in some instances, show promise as anti-B. pseudomallei infectives. PMID:23231488

Schweizer, Herbert P

2013-01-01

11

Within-Host Evolution of Burkholderia pseudomallei in Four Cases of Acute Melioidosis  

PubMed Central

Little is currently known about bacterial pathogen evolution and adaptation within the host during acute infection. Previous studies of Burkholderia pseudomallei, the etiologic agent of melioidosis, have shown that this opportunistic pathogen mutates rapidly both in vitro and in vivo at tandemly repeated loci, making this organism a relevant model for studying short-term evolution. In the current study, B. pseudomallei isolates cultured from multiple body sites from four Thai patients with disseminated melioidosis were subjected to fine-scale genotyping using multilocus variable-number tandem repeat analysis (MLVA). In order to understand and model the in vivo variable-number tandem repeat (VNTR) mutational process, we characterized the patterns and rates of mutations in vitro through parallel serial passage experiments of B. pseudomallei. Despite the short period of infection, substantial divergence from the putative founder genotype was observed in all four melioidosis cases. This study presents a paradigm for examining bacterial evolution over the short timescale of an acute infection. Further studies are required to determine whether the mutational process leads to phenotypic alterations that impact upon bacterial fitness in vivo. Our findings have important implications for future sampling strategies, since colonies in a single clinical sample may be genetically heterogeneous, and organisms in a culture taken late in the infective process may have undergone considerable genetic change compared with the founder inoculum. PMID:20090837

Limmathurotsakul, Direk; Max, Tamara L.; Sarovich, Derek S.; Vogler, Amy J.; Dale, Julia L.; Ginther, Jennifer L.; Leadem, Benjamin; Colman, Rebecca E.; Foster, Jeffrey T.; Tuanyok, Apichai; Wagner, David M.; Peacock, Sharon J.; Pearson, Talima; Keim, Paul

2010-01-01

12

Genome-Wide Expression Analysis of Burkholderia pseudomallei Infection in a Hamster Model of Acute Melioidosis  

PubMed Central

Burkholderia pseudomallei is the causative agent of melioidosis and represents a potential bioterrorism threat. In the current studies we have examined gene expression in B. pseudomallei in an animal model of acute melioidosis using whole-genome microarrays. Gene expression profiles were generated by comparing transcriptional levels of B. pseudomallei-expressed genes in infected hamster organs including liver, lung, and spleen following intraperitoneal and intranasal routes of infection to those from bacteria grown in vitro. Differentially expressed genes were similar in infected livers irrespective of the route of infection. Reduced expression of a number of housekeeping genes suggested a lower bacterial growth rate during infection. Energy production during growth in vivo involved specific biochemical pathways such as isomerization of 3-phosphoglycerate, catabolism of d-glucosamine and inositol, and biosynthesis of particular amino acids. In addition, the induction of genes known to be involved in oxidative phosphorylation including ubiquinol oxidase, ferredoxin oxidoreductase, and formate dehydrogenase enzymes suggested the use of alternative pathways for energy production, while the expression of genes coding for ATP-synthase and NADH-dehydrogenase enzymes was reduced. Our studies have identified differentially expressed genes which include potential virulence genes such as those for a putative phospholipase C and a putative two-component regulatory system, and they have also provided a better understanding of bacterial metabolism in response to the host environment during acute melioidosis. PMID:16988221

Tuanyok, Apichai; Tom, Marina; Dunbar, John; Woods, Donald E.

2006-01-01

13

Burkholderia pseudomallei Known Siderophores and Hemin Uptake Are Dispensable for Lethal Murine Melioidosis  

PubMed Central

Burkholderia pseudomallei is a mostly saprophytic bacterium, but can infect humans where it causes the difficult-to-manage disease melioidosis. Even with proper diagnosis and prompt therapeutic interventions mortality rates still range from >20% in Northern Australia to over 40% in Thailand. Surprisingly little is yet known about how B. pseudomallei infects, invades and survives within its hosts, and virtually nothing is known about the contribution of critical nutrients such as iron to the bacterium's pathogenesis. It was previously assumed that B. pseudomallei used iron-acquisition systems commonly found in other bacteria, for example siderophores. However, our previous discovery of a clinical isolate carrying a large chromosomal deletion missing the entire malleobactin gene cluster encoding the bacterium's major high-affinity siderophore while still being fully virulent in a murine melioidosis model suggested that other iron-acquisition systems might make contributions to virulence. Here, we deleted the major siderophore malleobactin (mba) and pyochelin (pch) gene clusters in strain 1710b and revealed a residual siderophore activity which was unrelated to other known Burkholderia siderophores such as cepabactin and cepaciachelin, and not due to increased secretion of chelators such as citrate. Deletion of the two hemin uptake loci, hmu and hem, showed that Hmu is required for utilization of hemin and hemoglobin and that Hem cannot complement a Hmu deficiency. Prolonged incubation of a hmu hem mutant in hemoglobin-containing minimal medium yielded variants able to utilize hemoglobin and hemin suggesting alternate pathways for utilization of these two host iron sources. Lactoferrin utilization was dependent on malleobactin, but not pyochelin synthesis and/or uptake. A mba pch hmu hem quadruple mutant could use ferritin as an iron source and upon intranasal infection was lethal in an acute murine melioidosis model. These data suggest that B. pseudomallei may employ a novel ferritin-iron acquisition pathway as a means to sustain in vivo growth. PMID:22745846

Kvitko, Brian H.; Goodyear, Andrew; Propst, Katie L.; Dow, Steven W.; Schweizer, Herbert P.

2012-01-01

14

Antimicrobial susceptibility patterns of Burkholderia pseudomallei among melioidosis cases in Kedah, Malaysia.  

PubMed

Burkholderia pseudomallei, the causative agent of melioidosis is an important cause of morbidity and mortality particularly among diabetics. We evaluated 228 isolates of B. pseudomallei for antimicrobial sensitivity during 2005-2010 using the disc diffusion technique, of which 144 were obtained from blood culture. More than 90% of the strains were susceptible to cefoperazone, ceftazidime, chloramphenicol and imipenem. Eighty-two percent of the isolates were susceptible to tetracycline and amoxicillin/clavulanate. The susceptibilities to ciprofloxacin was 78% and to trimethoprim-sulfamethoxezole was 47%. The susceptibilities to aminoglycoside antibiotics were low (21% to gentamicin and 6% to amikacin). The susceptibilities were similar between isolates from females and males, bacteremic and abacteremic cases, diabetics and non-diabetics, pneumonia and non-pneumonia cases and between those who died and those who survived. Our findings show antibiotic susceptibility patterns are not a major factor in determining outcomes of B. pseudomallei infection. Monitoring the drug susceptibilities among B. pseudomallei isolates needs to be conducted regularly to guide empiric therapy for melioidosis, as it causes high mortality, especially among diabetic cases. PMID:24974653

Hassan, Muhammad R A; Vijayalakshmi, Natesan; Pani, Subhada Prasad; Peng, Ng P; Mehenderkar, Ranjith; Voralu, Kirtanaa; Michael, Edwin

2014-05-01

15

Loop-Mediated Isothermal Amplification Method Targeting the TTS1 Gene Cluster for Detection of Burkholderia pseudomallei and Diagnosis of Melioidosis  

Microsoft Academic Search

Melioidosis is a severe infection caused by Burkholderia pseudomallei. The timely implementation of effective antimicrobial treatment requires rapid diagnosis. Loop-mediated isothermal amplification (LAMP) targeting the TTS1 gene cluster was developed for the detection of B. pseudomallei. LAMP was sensitive and specific for the laboratory detection of this organism. The lower limit of detection was 38 genomic copies per reaction, and

Narisara Chantratita; Ella Meumann; Direk Limmathurotsakul; Vanaporn Wuthiekanun; Saran Wannapasni; Nicholas P. J. Day; Sharon J. Peacock

16

The Burkholderia pseudomallei ?asd Mutant Exhibits Attenuated Intracellular Infectivity and Imparts Protection against Acute Inhalation Melioidosis in Mice ?  

PubMed Central

Burkholderia pseudomallei, the cause of serious and life-threatening diseases in humans, is of national biodefense concern because of its potential use as a bioterrorism agent. This microbe is listed as a select agent by the CDC; therefore, development of vaccines is of significant importance. Here, we further investigated the growth characteristics of a recently created B. pseudomallei 1026b ?asd mutant in vitro, in a cell model, and in an animal model of infection. The mutant was typified by an inability to grow in the absence of exogenous diaminopimelate (DAP); upon single-copy complementation with a wild-type copy of the asd gene, growth was restored to wild-type levels. Further characterization of the B. pseudomallei ?asd mutant revealed a marked decrease in RAW264.7 murine macrophage cytotoxicity compared to the wild type and the complemented ?asd mutant. RAW264.7 cells infected by the ?asd mutant did not exhibit signs of cytopathology or multinucleated giant cell (MNGC) formation, which were observed in wild-type B. pseudomallei cell infections. The ?asd mutant was found to be avirulent in BALB/c mice, and mice vaccinated with the mutant were protected against acute inhalation melioidosis. Thus, the B. pseudomallei ?asd mutant may be a promising live attenuated vaccine strain and a biosafe strain for consideration of exclusion from the select agent list. PMID:21807903

Norris, Michael H.; Propst, Katie L.; Kang, Yun; Dow, Steven W.; Schweizer, Herbert P.; Hoang, Tung T.

2011-01-01

17

Growth, motility and resistance to oxidative stress of the melioidosis pathogen Burkholderia pseudomallei are enhanced by epinephrine.  

PubMed

Burkholderia pseudomallei causes melioidosis, a severe invasive disease endemic in South-East Asia and Northern Australia. Bacterial pathogens of several genera have been reported to be able to sense and respond to the stress-related catecholamine hormone epinephrine. Here, we report that epinephrine induces growth of B. pseudomallei in minimal serum-rich medium and heat-inactivated whole human serum and enhances bacterial motility, transcription of flagellar genes and flagellin synthesis. The effect of epinephrine on motility, but not bacterial growth, could be partially reversed by the alpha-adrenergic receptor antagonist phentolamine. Epinephrine also altered the transcription of iron-regulated genes encoding superoxide dismutase (sodB) and the malleobactin receptor (fmtA). Consistent with induction of sodB expression, epinephrine-treated B. pseudomallei exhibited increased resistance to superoxide. Epinephrine treatment did not stimulate Type III secretion via the virulence-associated Bsa apparatus or the ability of B. pseudomallei to invade epithelial cells in culture. This study provides the first evidence that epinephrine, a hormone released from the host under stress and upon therapy, can affect B. pseudomallei virulence-associated properties. PMID:24753312

Intarak, Narin; Muangsombut, Veerachat; Vattanaviboon, Paiboon; Stevens, Mark P; Korbsrisate, Sunee

2014-10-01

18

Burkholderia pseudomallei Penetrates the Brain via Destruction of the Olfactory and Trigeminal Nerves: Implications for the Pathogenesis of Neurological Melioidosis  

PubMed Central

ABSTRACT Melioidosis is a potentially fatal disease that is endemic to tropical northern Australia and Southeast Asia, with a mortality rate of 14 to 50%. The bacterium Burkholderia pseudomallei is the causative agent which infects numerous parts of the human body, including the brain, which results in the neurological manifestation of melioidosis. The olfactory nerve constitutes a direct conduit from the nasal cavity into the brain, and we have previously reported that B. pseudomallei can colonize this nerve in mice. We have now investigated in detail the mechanism by which the bacteria penetrate the olfactory and trigeminal nerves within the nasal cavity and infect the brain. We found that the olfactory epithelium responded to intranasal B. pseudomallei infection by widespread crenellation followed by disintegration of the neuronal layer to expose the underlying basal layer, which the bacteria then colonized. With the loss of the neuronal cell bodies, olfactory axons also degenerated, and the bacteria then migrated through the now-open conduit of the olfactory nerves. Using immunohistochemistry, we demonstrated that B. pseudomallei migrated through the cribriform plate via the olfactory nerves to enter the outer layer of the olfactory bulb in the brain within 24 h. We also found that the bacteria colonized the thin respiratory epithelium in the nasal cavity and then rapidly migrated along the underlying trigeminal nerve to penetrate the cranial cavity. These results demonstrate that B. pseudomallei invasion of the nerves of the nasal cavity leads to direct infection of the brain and bypasses the blood-brain barrier. PMID:24736221

St. John, James A.; Ekberg, Jenny A. K.; Dando, Samantha J.; Meedeniya, Adrian C. B.; Horton, Rachel E.; Batzloff, Michael; Owen, Suzzanne J.; Holt, Stephanie; Peak, Ian R.; Ulett, Glen C.; Mackay-Sim, Alan; Beacham, Ifor R.

2014-01-01

19

Genomic patterns of pathogen evolution revealed by comparison of Burkholderia pseudomallei, the causative agent of melioidosis, to avirulent Burkholderia thailandensis  

PubMed Central

Background The Gram-negative bacterium Burkholderia pseudomallei (Bp) is the causative agent of the human disease melioidosis. To understand the evolutionary mechanisms contributing to Bp virulence, we performed a comparative genomic analysis of Bp K96243 and B. thailandensis (Bt) E264, a closely related but avirulent relative. Results We found the Bp and Bt genomes to be broadly similar, comprising two highly syntenic chromosomes with comparable numbers of coding regions (CDs), protein family distributions, and horizontally acquired genomic islands, which we experimentally validated to be differentially present in multiple Bt isolates. By examining species-specific genomic regions, we derived molecular explanations for previously-known metabolic differences, discovered potentially new ones, and found that the acquisition of a capsular polysaccharide gene cluster in Bp, a key virulence component, is likely to have occurred non-randomly via replacement of an ancestral polysaccharide cluster. Virulence related genes, in particular members of the Type III secretion needle complex, were collectively more divergent between Bp and Bt compared to the rest of the genome, possibly contributing towards the ability of Bp to infect mammalian hosts. An analysis of pseudogenes between the two species revealed that protein inactivation events were significantly biased towards membrane-associated proteins in Bt and transcription factors in Bp. Conclusion Our results suggest that a limited number of horizontal-acquisition events, coupled with the fine-scale functional modulation of existing proteins, are likely to be the major drivers underlying Bp virulence. The extensive genomic similarity between Bp and Bt suggests that, in some cases, Bt could be used as a possible model system for studying certain aspects of Bp behavior. PMID:16725056

Yu, Yiting; Kim, H Stanley; Chua, Hui Hoon; Lin, Chi Ho; Sim, Siew Hoon; Lin, Daoxun; Derr, Alan; Engels, Reinhard; DeShazer, David; Birren, Bruce; Nierman, William C; Tan, Patrick

2006-01-01

20

Loop-Mediated Isothermal Amplification Method Targeting the TTS1 Gene Cluster for Detection of Burkholderia pseudomallei and Diagnosis of Melioidosis?  

PubMed Central

Melioidosis is a severe infection caused by Burkholderia pseudomallei. The timely implementation of effective antimicrobial treatment requires rapid diagnosis. Loop-mediated isothermal amplification (LAMP) targeting the TTS1 gene cluster was developed for the detection of B. pseudomallei. LAMP was sensitive and specific for the laboratory detection of this organism. The lower limit of detection was 38 genomic copies per reaction, and LAMP was positive for 10 clinical B. pseudomallei isolates but negative for 5 B. thailandensis and 5 B. mallei isolates. A clinical evaluation was conducted in northeast Thailand to compare LAMP to an established real-time PCR assay targeting the same TTS1 gene cluster. A total of 846 samples were obtained from 383 patients with suspected melioidosis, 77 of whom were subsequently diagnosed with culture-confirmed melioidosis. Of these 77 patients, a positive result was obtained from one or more specimens by PCR in 26 cases (sensitivity, 34%; 95% confidence interval [CI], 23.4 to 45.4%) and by LAMP in 34 cases (sensitivity, 44%; 95% CI, 32.8 to 55.9%) (P = 0.02). All samples from 306 patients that were culture negative for B. pseudomallei were negative by PCR (specificity, 100%; 95% CI, 98.8 to 100%), but 5 of 306 patients (1.6%) were positive by LAMP (specificity, 98.4%; 95% CI, 96.2 to 99.5%) (P = 0.03). The diagnostic accuracies of PCR and LAMP were 86.7% (95% CI, 82.9 to 89.9%) and 87.5% (95% CI, 83.7 to 90.6%), respectively (P = 0.47). Both assays were very insensitive when applied to blood samples; PCR and LAMP were positive for 0 and 1 of 44 positive blood cultures, respectively. The PCR and LAMP assays evaluated here are not sufficiently sensitive to replace culture in our clinical setting. PMID:18039797

Chantratita, Narisara; Meumann, Ella; Thanwisai, Aunchalee; Limmathurotsakul, Direk; Wuthiekanun, Vanaporn; Wannapasni, Saran; Tumapa, Sarinna; Day, Nicholas P. J.; Peacock, Sharon J.

2008-01-01

21

Analysis of the Prevalence, Secretion and Function of a Cell Cycle-Inhibiting Factor in the Melioidosis Pathogen Burkholderia pseudomallei  

PubMed Central

Enteropathogenic and enterohaemorrhagic Escherichia coli express a cell cycle-inhibiting factor (Cif), that is injected into host cells via a Type III secretion system (T3SS) leading to arrest of cell division, delayed apoptosis and cytoskeletal rearrangements. A homologue of Cif has been identified in Burkholderia pseudomallei (CHBP; Cif homologue in B. pseudomallei; BPSS1385), which shares catalytic activity, but its prevalence, secretion and function are ill-defined. Among 43 available B. pseudomallei genome sequences, 33 genomes (76.7%) harbor the gene encoding CHBP. Western blot analysis using antiserum raised to a synthetic CHBP peptide detected CHBP in 46.6% (7/15) of clinical B. pseudomallei isolates from the endemic area. Secretion of CHBP into bacterial culture supernatant could not be detected under conditions where a known effector (BopE) was secreted in a manner dependent on the Bsa T3SS. In contrast, CHBP could be detected in U937 cells infected with B. pseudomallei by immunofluorescence microscopy and Western blotting in a manner dependent on bsaQ. Unlike E. coli Cif, CHBP was localized within the cytoplasm of B. pseudomallei-infected cells. A B. pseudomallei chbP insertion mutant showed a significant reduction in cytotoxicity and plaque formation compared to the wild-type strain that could be restored by plasmid-mediated trans-complementation. However, there was no defect in actin-based motility or multinucleated giant cell formation by the chbP mutant. The data suggest that the level or timing of CHBP secretion differs from a known Bsa-secreted effector and that CHBP is required for selected virulence-associated phenotypes in vitro. PMID:24809950

Pumirat, Pornpan; Broek, Charles Vander; Juntawieng, Niramol; Muangsombut, Veerachat; Kiratisin, Pattarachai; Pattanapanyasat, Kovit; Stevens, Joanne M.; Stevens, Mark P.; Korbsrisate, Sunee

2014-01-01

22

Selection of Burkholderia pseudomallei protease-binding peptides by phage display  

Microsoft Academic Search

Burkholderia pseudomallei is a causative agent of melioidosis, a fatal community acquired septicemia in Southeast Asia and Northern Australia. A protease has been proposed to be one of the major pathogenic factors to play a significant role in melioidosis. We have used phage display technology to identify peptides binding to B. pseudomallei protease. By screening a constrained cyclic heptapeptide library, five

Wimol Petkanchanapong; Sarah Fredriksson; Virapong Prachayasittikul; Leif Bülow

2000-01-01

23

Evaluation of a Latex Agglutination Assay for the Identification of Burkholderia pseudomallei and Burkholderia mallei  

PubMed Central

Cases of melioidosis and glanders are rare in the United States, but the etiologic agents of each disease (Burkholderia pseudomallei and Burkholderia mallei, respectively) are classified as Tier 1 select agents because of concerns about their potential use as bioterrorism agents. A rapid, highly sensitive, and portable assay for clinical laboratories and field use is required. Our laboratory has further evaluated a latex agglutination assay for its ability to identify B. pseudomallei and B. mallei isolates. This assay uses a monoclonal antibody that specifically recognizes the capsular polysaccharide produced by B. pseudomallei and B. mallei, but is absent in closely related Burkholderia species. A total of 110 B. pseudomallei and B. mallei were tested, and 36 closely related Burkholderia species. The latex agglutination assay was positive for 109 of 110 (99.1% sensitivity) B. pseudomallei and B. mallei isolates tested. PMID:24710616

Duval, Brea D.; Elrod, Mindy G.; Gee, Jay E.; Chantratita, Narisara; Tandhavanant, Sarunporn; Limmathurotsakul, Direk; Hoffmaster, Alex R.

2014-01-01

24

Evaluation of a latex agglutination assay for the identification of Burkholderia pseudomallei and Burkholderia mallei.  

PubMed

Cases of melioidosis and glanders are rare in the United States, but the etiologic agents of each disease (Burkholderia pseudomallei and Burkholderia mallei, respectively) are classified as Tier 1 select agents because of concerns about their potential use as bioterrorism agents. A rapid, highly sensitive, and portable assay for clinical laboratories and field use is required. Our laboratory has further evaluated a latex agglutination assay for its ability to identify B. pseudomallei and B. mallei isolates. This assay uses a monoclonal antibody that specifically recognizes the capsular polysaccharide produced by B. pseudomallei and B. mallei, but is absent in closely related Burkholderia species. A total of 110 B. pseudomallei and B. mallei were tested, and 36 closely related Burkholderia species. The latex agglutination assay was positive for 109 of 110 (99.1% sensitivity) B. pseudomallei and B. mallei isolates tested. PMID:24710616

Duval, Brea D; Elrod, Mindy G; Gee, Jay E; Chantratita, Narisara; Tandhavanant, Sarunporn; Limmathurotsakul, Direk; Hoffmaster, Alex R

2014-06-01

25

Burkholderia pseudomallei traced to water treatment plant in Australia.  

PubMed Central

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. PMID:10653571

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

2000-01-01

26

Burkholderia pseudomallei Isolates in 2 Pet Iguanas, California, USA  

PubMed Central

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. PMID:24447394

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

2014-01-01

27

Biochemical characteristics of clinical and environmental isolates of Burkholderia pseudomallei  

Microsoft Academic Search

The biochemical characteristics of 213 isolates of Burkholderia pseudomallei from patients with melioidosis and 140 isolates from the soil in central and northeastern Thailand were compared. Whereas the biochemical profiles of all the clinical isolates were similar, all soil isolates from the central area and 25% of isolates from northeastern Thailand comprised a different phenotype. This was characterised by the

VANAPORN WUTHIEKANUN; M. D. Smith; D. A. B. Dance; AMANDA L. WALSH; T. L. PITTT; N. J. White

1996-01-01

28

Misidentification of Burkholderia pseudomallei as Burkholderia cepacia by the VITEK 2 system.  

PubMed

A previously healthy Chinese male returned from working in the Malaysian jungle with a fever. A blood culture grew Gram-negative bacilli that were initially identified as Burkholderia cepacia by the VITEK 2 system but were subsequently found to be Burkholderia pseudomallei by partial sequencing of the 16S rRNA gene. The identification of B. pseudomallei using commercially available automated systems is problematic and clinicians in non-endemic areas should be aware of the possibility of melioidosis in patients with a relevant travel history and blood cultures growing Burkholderia spp. PMID:22820689

Zong, Zhiyong; Wang, Xiaohui; Deng, Yiyun; Zhou, Taoyou

2012-10-01

29

Ecology of Burkholderia pseudomallei and the interactions between environmental Burkholderia spp. and human–animal hosts  

Microsoft Academic Search

Early workers thought that melioidosis was a zoonosis with a reservoir in rodents, but we now know that Burkholderia pseudomallei is a widely distributed environmental saprophyte. In northeast Thailand, two thirds of paddy fields yield the organism, and 80% of children have antibodies by the time they are 4 years old. However, interpretation of these results has been complicated by

David A. B Dance

2000-01-01

30

Distribution of type III secretion gene clusters in Burkholderia pseudomallei, B. thailandensis and B. mallei  

Microsoft Academic Search

Burkholderia pseudomallei, the causative agent of melioidosis, carries a cluster of genes closely related in organisation to the type III secretion (TTS) system gene clusters of the plant pathogens Ralstonia solanacearum and Xanthomonas spp. The TTS gene cluster (TTS1) is present only in B. pseudomallei and not in avirulent B. thailandensis. Adjacent to the gene cluster encoding putative secreton structural

LUCILLE RAINBOW; C. ANTHONY HART; CRAIG WINSTANLEY

31

Strategies toward vaccines against Burkholderia mallei and Burkholderia pseudomallei.  

PubMed

Burkholderia mallei and Burkholderia pseudomallei are Gram-negative, rod-shaped bacteria, and are the causative agents of the diseases glanders and melioidosis, respectively. These bacteria have been recognized as important pathogens for over 100 years, yet a relative dearth of available information exists regarding their virulence determinants and immunopathology. Infection with either of these bacteria presents with nonspecific symptoms and can be either acute or chronic, impeding rapid diagnosis. The lack of a vaccine for either bacterium also makes them potential candidates for bioweaponization. Together with their high rate of infectivity via aerosols and resistance to many common antibiotics, both bacteria have been classified as category B priority pathogens by the US NIH and US CDC, which has spurred a dramatic increase in interest in these microorganisms. Attempts have been made to develop vaccines for these infections, which would not only benefit military personnel, a group most likely to be targeted in an intentional release, but also individuals who may come in contact with glanders-infected animals or live in areas where melioidosis is endemic. This review highlights some recent attempts of vaccine development for these infections and the strategies used to improve the efficacy of vaccine approaches. PMID:18980539

Bondi, Sara K; Goldberg, Joanna B

2008-11-01

32

Development of Burkholderia mallei and pseudomallei vaccines  

PubMed Central

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-? and TNF-? 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 vaccines have typically provided less robust immunity, but are safer to administer to a wider variety of people, including immune compromised individuals because they do not reactivate or cause disease. The challenges facing B. mallei and B. pseudomalllei vaccine development include identification of broadly protective antigens, design of efficient vaccine delivery and adjuvant systems, and a better understanding of the correlates of protection from both acute and chronic infection. PMID:23508691

Silva, Ediane B.; Dow, Steven W.

2013-01-01

33

Autotransporters and Their Role in the Virulence of Burkholderia pseudomallei and Burkholderia mallei  

PubMed Central

Burkholderia pseudomallei and Burkholderia mallei are closely related Gram-negative bacteria responsible for the infectious diseases melioidosis and glanders, respectively. Autotransporters (ATs) comprise a large and diverse family of secreted and outer membrane proteins that includes virulence-associated invasins, adhesins, proteases, and actin-nucleating factors. The B. pseudomallei K96243 genome contains 11 predicted ATs, eight of which share homologs in the B. mallei ATCC 23344 genome. This review distils key findings from in silico, in vitro, and in vivo studies on the ATs of B. pseudomallei and B. mallei. To date, the best characterized of the predicted ATs of B. pseudomallei and B. mallei is BimA, a predicted trimeric AT mediating actin-based motility which varies in sequence and mode of action between Burkholderia species. Of the remaining eight predicted B. pseudomallei trimeric autotransporters, five of which are also present in B. mallei, two (BoaA and BoaB), have been implicated in bacterial adhesion to epithelial cells. Several predicted Burkholderia ATs are recognized by human humoral and cell-mediated immunity, indicating that they are expressed during infection and may be useful for diagnosis and vaccine-mediated protection. Further studies on the mode of secretion and functions of Burkholderia ATs will facilitate the rational design of control strategies. PMID:21811486

Adler, Natalie R. Lazar; Stevens, Joanne M.; Stevens, Mark P.; Galyov, Edouard E.

2011-01-01

34

Comparison of Diagnostic Laboratory Methods for Identification of Burkholderia pseudomallei  

PubMed Central

Limited experience and a lack of validated diagnostic reagents make Burkholderia pseudomallei, the cause of melioidosis, difficult to recognize in the diagnostic microbiology laboratory. We compared three methods of confirming the identity of presumptive B. pseudomallei strains using a collection of Burkholderia species drawn from diverse geographic, clinical, and environmental sources. The 95 isolates studied included 71 B. pseudomallei and 3 B. thailandensis isolates. The API 20NE method identified only 37% of the B. pseudomallei isolates. The agglutinating antibody test identified 82% at first the attempt and 90% including results of a repeat test with previously negative isolates. Gas-liquid chromatography analysis of bacterial fatty acid methyl esters (GLC-FAME) identified 98% of the B. pseudomallei isolates. The agglutination test produced four false positive results, one B. cepacia, one B. multivorans, and two B. thailandensis. API produced three false positive results, one positive B. cepacia and two positive B. thailandensis. GLC-FAME analysis was positive for one B. cepacia isolate. On the basis of these results, the most robust B. pseudomallei discovery pathway combines the previously recommended isolate screening tests (Gram stain, oxidase test, gentamicin and polymyxin susceptibility) with monoclonal antibody agglutination on primary culture, followed by a repeat after 24 h incubation on agglutination-negative isolates and GLC-FAME analysis. Incorporation of PCR-based identification within this schema may improve percentages of recognition further but requires more detailed evaluation. PMID:15872242

Inglis, Timothy J. J.; Merritt, Adam; Chidlow, Glenys; Aravena-Roman, Max; Harnett, Gerry

2005-01-01

35

In vitro antimicrobial activity of natural toxins and animal venoms tested against Burkholderia pseudomallei  

Microsoft Academic Search

BACKGROUND: Burkholderia pseudomallei are the causative agent of melioidosis. Increasing resistance of the disease to antibiotics is a severe problem in treatment regime and has led to intensification of the search for new drugs. Antimicrobial peptides are the most ubiquitous in nature as part of the innate immune system and host defense mechanism. METHODS: Here, we investigated a group of

R Perumal Samy; A Pachiappan; P Gopalakrishnakone; Maung M Thwin; Yap E Hian; Vincent TK Chow; Ho Bow; Joseph T Weng

2006-01-01

36

Adaptation and Antibiotic Tolerance of Anaerobic Burkholderia pseudomallei ? †  

PubMed Central

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

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

2011-01-01

37

Clinical Features and Laboratory Diagnosis of Infection with the Potential Bioterrorism Agents Burkholderia Mallei and Burkholderia Pseudomallei  

PubMed Central

Burkholderia mallei and Burkholderia pseudomallei are the causative organisms of Glanders and Melioidosis, respectively. Although now rare in Western countries, both organisms have recently gained much interest because of their unique potential as bioterrorism agents. These organisms are less familiar to medical and laboratory personnel than other select bioterrorism bacterial agents and thus heightened awareness of Glanders and Melioidosis is crucial in order to enable adequate emergency preparedness and response to deliberate release of B. mallei and B. pseudomallei. The microbiological diagnosis of both species in the clinical laboratory is complicated. This paper reviews the various challenges and pitfalls associated with the diagnosis of Melioidosis and Glanders in the clinical setting, with emphasis on the role of sentinel laboratories. PMID:23675037

Gilad, Jacob; Schwartz, David; Amsalem, Yoram

2007-01-01

38

Systematic Review and Consensus Guidelines for Environmental Sampling of Burkholderia pseudomallei  

PubMed Central

Background 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. Methods/Principal Findings 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. Conclusions/Significance The proposed consensus guidelines provide the basis for the development of an accurate and comprehensive global map of environmental B. pseudomallei. PMID:23556010

Limmathurotsakul, Direk; Dance, David A. B.; Wuthiekanun, Vanaporn; Kaestli, Mirjam; Mayo, Mark; Warner, Jeffrey; Wagner, David M.; Tuanyok, Apichai; Wertheim, Heiman; Yoke Cheng, Tan; Mukhopadhyay, Chiranjay; Puthucheary, Savithiri; Day, Nicholas P. J.; Steinmetz, Ivo; Currie, Bart J.; Peacock, Sharon J.

2013-01-01

39

Polar Lipids of Burkholderia pseudomallei Induce Different Host Immune Responses  

PubMed Central

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-?, 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-? 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

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

2013-01-01

40

Potential misidentification of Burkholderia pseudomallei by API 20NE.  

PubMed

Biochemical confirmation of the identity of Burkholderia pseudomallei in Singapore previously relied on the API 20NE panel of tests. After introducing an alternative proprietary biochemical panel, the Microbact 24E (MedVet, Adelaide, Australia), we noted that the API panel identified some presumptive B. pseudomallei isolates as other species. We therefore compared the performance of the API 20NE against the Microbact 24E with 50 distinct clinical isolates of B. pseudomallei, after 24 hours and after five days incubation of primary cultures. The API panel correctly identified 40 isolates. Four results were unacceptable or uninterpretable. Six isolates were misidentified as other species; the commonest being Chromobacterium violaceum. One of these was again identified as C. violaceum by the repeat API panel. Fourteen isolates, including the six misidentified isolates and four isolate pairs from separate sources in four separate patients, were typed using PCR amplification of repetitive extragenic palindromic sequences (REPS). The isolates identified as C. violaceum appeared to have identical REPS patterns, suggesting that some of the errant API results may be due to a single locally prevalent strain of B. pseudomallei. A previous suggestion that C. violaceum may produce a melioidosis-like illness may therefore be due to laboratory misidentification of B. pseudomallei with the API 20NE biochemical test panel. PMID:9534210

Inglis, T J; Chiang, D; Lee, G S; Chor-Kiang, L

1998-02-01

41

Functional reconstitution, gene isolation and topology modelling of porins from Burkholderia pseudomallei and Burkholderia thailandensis.  

PubMed Central

The sequences for Omp38 from Burkholderia pseudomallei and Burkholderia thailandensis have been deposited in the DDBJ, EMBL, GenBank(R) and GSDB Nucleotide Sequence Databases under the accession numbers AY312416 and AY312417 respectively. The intracellular pathogen Burkholderia pseudomallei is the causative agent of tropical melioidosis, and Burkholderia thailandensis is a closely-related Gram-negative bacterium that does not cause serious disease. Like other bacteria, the major outer membrane (OM) porins of Burkholderia strains, Bps Omp38 and Bth Omp38 may have roles in antibiotic resistance and immunity. We purified both proteins and found them to be immunologically related, SDS-resistant, heat-sensitive trimers with M (r) of approx. 110000. In functional liposome-swelling assays, both proteins showed similar permeabilities for small sugar molecules, compatible with a pore diameter of between 1.2 and 1.6 nm. Secondary structure analysis by FTIR (Fourier-transform infrared) spectroscopy revealed almost identical spectra with predominantly beta-sheet structures, typical of bacterial porins. MALDI-TOF (matrix-assisted laser-desorption ionization-time of flight) MS and ESI/MS (electrospray ionization MS) analysis of each protein showed extensive sequence similarities to the OpcP1 porin from Burkholderia cepacia (later found to be 76.5% identical). Based on information from the incomplete B. pseudomallei genome-sequencing project, the genes encoding Omp38 were identified and amplified by PCR from B. pseudomallei and B. thailandensis genomic DNA. The nucleotide sequences are 99.7% identical, and the predicted processed proteins are 100% identical. Topology prediction and molecular modelling suggest that this newly-isolated and cloned porin is a 16-stranded beta-barrel and the external loops of the protein could be important determinants of the immune response to infection. PMID:14567756

Siritapetawee, Jaruwan; Prinz, Heino; Samosornsuk, Worada; Ashley, Richard H; Suginta, Wipa

2004-01-01

42

Post-Exposure Therapeutic Efficacy of COX-2 Inhibition against Burkholderia pseudomallei  

PubMed Central

Burkholderia pseudomallei is a Gram-negative, facultative intracellular bacillus and the etiologic agent of melioidosis, a severe disease in Southeast Asia and Northern Australia. Like other multidrug-resistant pathogens, the inherent antibiotic resistance of B. pseudomallei impedes treatment and highlights the need for alternative therapeutic strategies that can circumvent antimicrobial resistance mechanisms. In this work, we demonstrate that host prostaglandin E2 (PGE2) production plays a regulatory role in the pathogenesis of B. pseudomallei. PGE2 promotes B. pseudomallei intracellular survival within macrophages and bacterial virulence in a mouse model of pneumonic melioidosis. PGE2-mediated immunosuppression of macrophage bactericidal effector functions is associated with increased arginase 2 (Arg2) expression and decreased nitric oxide (NO) production. Treatment with a commercially-available COX-2 inhibitor suppresses the growth of B. pseudomallei in macrophages and affords significant protection against rapidly lethal pneumonic melioidosis when administered post-exposure to B. pseudomallei-infected mice. COX-2 inhibition may represent a novel immunotherapeutic strategy to control infection with B. pseudomallei and other intracellular pathogens. PMID:23675544

Asakrah, Saja; Nieves, Wildaliz; Mahdi, Zaid; Agard, Mallory; Zea, Arnold H.; Roy, Chad J.; Morici, Lisa A.

2013-01-01

43

Molecular Characterization of Putative Virulence Determinants in Burkholderia pseudomallei  

PubMed Central

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. PMID:25215325

Puah, Suat Moi; Puthucheary, S. D.; Wang, Jin Town; Pan, Yi Jiun; Chua, Kek Heng

2014-01-01

44

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

PubMed

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. PMID:24145517

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

45

Effect of colony morphology variation of Burkholderia pseudomallei on intracellular survival and resistance to antimicrobial environments in human macrophages in vitro  

Microsoft Academic Search

BACKGROUND: Primary diagnostic cultures from patients with melioidosis demonstrate variation in colony morphology of the causative organism, Burkholderia pseudomallei. Variable morphology is associated with changes in the expression of a range of putative virulence factors. This study investigated the effect of B. pseudomallei colony variation on survival in the human macrophage cell line U937 and under laboratory conditions simulating conditions

Sarunporn Tandhavanant; Aunchalee Thanwisai; Direk Limmathurotsakul; Sunee Korbsrisate; Nicholas PJ Day; Sharon J Peacock; Narisara Chantratita

2010-01-01

46

Burkholderia pseudomallei Misidentified by Automated System  

PubMed Central

After returning from Thailand, a 35-year-old man from Switzerland was hospitalized with an abscess of the head. Material cultured from the abscess and adjacent bone grew a gram-negative rod, which was misidentified by an automated microbiology system as Burkholderia cepacia. The organism was eventually identified by molecular methods as B. pseudomallei. PMID:19891868

Weissert, Christoph; Dollenmaier, Gunter; Rafeiner, Philippe; Riehm, Julia

2009-01-01

47

Burkholderia pseudomallei-Induced Expression of Suppressor of Cytokine Signaling 3 and Cytokine-Inducible Src Homology 2-Containing Protein in Mouse Macrophages: a Possible Mechanism for Suppression of the Response to Gamma Interferon Stimulation  

Microsoft Academic Search

Burkholderia pseudomallei, the causative agent of melioidosis, is a facultative intracellular gram-negative bacterium that is able to survive and multiply in macrophages. Previously, we reported that B. pseudomallei was able to escape macrophage killing by interfering with the expression of inducible nitric oxide synthase (iNOS). In the present study, we extended this finding and demonstrated that B. pseudomallei was able

P. Ekchariyawat; S. Pudla; K. Limposuwan; S. Arjcharoen; S. Sirisinha; P. Utaisincharoen

2005-01-01

48

Accurate and Rapid Identification of the Burkholderia pseudomallei Near-Neighbour, Burkholderia ubonensis, Using Real-Time PCR  

PubMed Central

Burkholderia ubonensis is an environmental bacterium belonging to the Burkholderia cepacia complex (Bcc), a group of genetically related organisms that are associated with opportunistic but generally nonfatal infections in healthy individuals. In contrast, the near-neighbour species Burkholderia pseudomallei causes melioidosis, a disease that can be fatal in up to 95% of cases if left untreated. B. ubonensis is frequently misidentified as B. pseudomallei from soil samples using selective culturing on Ashdown’s medium, reflecting both the shared environmental niche and morphological similarities of these species. Additionally, B. ubonensis shows potential as an important biocontrol agent in B. pseudomallei-endemic regions as certain strains possess antagonistic properties towards B. pseudomallei. Current methods for characterising B. ubonensis are laborious, time-consuming and costly, and as such this bacterium remains poorly studied. The aim of our study was to develop a rapid and inexpensive real-time PCR-based assay specific for B. ubonensis. We demonstrate that a novel B. ubonensis-specific assay, Bu550, accurately differentiates B. ubonensis from B. pseudomallei and other species that grow on selective Ashdown’s agar. We anticipate that Bu550 will catalyse research on B. ubonensis by enabling rapid identification of this organism from Ashdown’s-positive colonies that are not B. pseudomallei. PMID:23967229

Price, Erin P.; Sarovich, Derek S.; Webb, Jessica R.; Ginther, Jennifer L.; Mayo, Mark; Cook, James M.; Seymour, Meagan L.; Kaestli, Mirjam; Theobald, Vanessa; Hall, Carina M.; Busch, Joseph D.; Foster, Jeffrey T.; Keim, Paul; Wagner, David M.; Tuanyok, Apichai; Pearson, Talima; Currie, Bart J.

2013-01-01

49

Workshop on Treatment of and Postexposure Prophylaxis for Burkholderia pseudomallei and B. mallei Infection, 2010  

PubMed Central

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 melioidosis and glanders, respectively. Drugs recommended by consensus of the participants are ceftazidime or meropenem for initial intensive therapy, and trimethoprim/sulfamethoxazole or amoxicillin/clavulanic acid for eradication therapy. For postexposure prophylaxis, recommended drugs are trimethoprim/sulfamethoxazole or co-amoxiclav. To improve the timely diagnosis of melioidosis and glanders, further development and wide distribution of rapid diagnostic assays were also recommended. Standardized animal models and B. pseudomallei strains are needed for further development of therapeutic options. Training for laboratory technicians and physicians would facilitate better diagnosis and treatment options. PMID:23171644

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

50

Obligatory Role of Gamma Interferon for Host Survival in a Murine Model of Infection with Burkholderia pseudomallei  

Microsoft Academic Search

Burkholderia pseudomallei, the causative agent of melioidosis, is a gram-negative bacterium capable of causing either acute lethal sepsis or chronic but eventually fatal disease in infected individuals. However, despite the clinical importance of this infection in areas where it is endemic, there is essentially no information on the mechanisms of protective immunity to the bacterium. We describe here a murine

P. SANTANIRAND; V. S. HARLEY; D. A. B. DANCE; B. S. DRASAR; G. J. BANCROFT

1999-01-01

51

Growing Burkholderia pseudomallei in Biofilm Stimulating Conditions Significantly Induces Antimicrobial Resistance  

Microsoft Academic Search

BackgroundBurkholderia pseudomallei, a Gram-negative bacterium that causes melioidosis, was reported to produce biofilm. As the disease causes high relapse rate when compared to other bacterial infections, it therefore might be due to the reactivation of the biofilm forming bacteria which also provided resistance to antimicrobial agents. However, the mechanism on how biofilm can provide tolerance to antimicrobials is still unclear.Methodology\\/Principal

Chakrit Sawasdidoln; Suwimol Taweechaisupapong; Rasana W. Sermswan; Unchalee Tattawasart; Sumalee Tungpradabkul; Surasakdi Wongratanacheewin; Stefan Bereswill

2010-01-01

52

Identification of tomato plant as a novel host model for Burkholderia pseudomallei  

Microsoft Academic Search

BACKGROUND: Burkholderia pseudomallei is the causative agent for melioidosis, a disease with significant mortality and morbidity in endemic regions. Its versatility as a pathogen is reflected in its relatively huge 7.24 Mb genome and the presence of many virulence factors including three Type Three Secretion Systems known as T3SS1, T3SS2 and T3SS3. Besides being a human pathogen, it is able

Yian Hoon Lee; Yahua Chen; Xuezhi Ouyang; Yunn-Hwen Gan

2010-01-01

53

The chemical arsenal of Burkholderia pseudomallei is essential for pathogenicity.  

PubMed

Increasing evidence has shown that small-molecule chemistry in microbes (i.e., secondary metabolism) can modulate the microbe-host response in infection and pathogenicity. The bacterial disease melioidosis is conferred by the highly virulent, antibiotic-resistant pathogen Burkholderia pseudomallei (BP). Whereas some macromolecular structures have been shown to influence BP virulence (e.g., secretion systems, cellular capsule, pili), the role of the large cryptic secondary metabolome encoded within its genome has been largely unexplored for its importance to virulence. Herein we demonstrate that BP-encoded small-molecule biosynthesis is indispensible for in vivo BP pathogenicity. Promoter exchange experiments were used to induce high-level molecule production from two gene clusters (MPN and SYR) found to be essential for in vivo virulence. NMR structural characterization of these metabolites identified a new class of lipopeptide biosurfactants/biofilm modulators (the malleipeptins) and syrbactin-type proteasome inhibitors, both of which represent overlooked small-molecule virulence factors for BP. Disruption of Burkholderia virulence by inhibiting the biosynthesis of these small-molecule biosynthetic pathways may prove to be an effective strategy for developing novel melioidosis-specific therapeutics. PMID:24884988

Biggins, John B; Kang, Hahk-Soo; Ternei, Melinda A; DeShazer, David; Brady, Sean F

2014-07-01

54

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

PubMed Central

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 thrive inside J774A.1 murine macrophages, suggesting a possible role for these proteins in survival within professional phagocytic cells. Conclusions The boaA and boaB genes specify adhesins that mediate adherence to epithelial cells of the human respiratory tract. The boaA gene product is shared by B. pseudomallei and B. mallei whereas BoaB appears to be a B. pseudomallei-specific adherence factor. PMID:20920184

2010-01-01

55

Burkholderia pseudomallei transcriptional adaptation in macrophages  

PubMed Central

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. PMID:22823543

2012-01-01

56

Role of RelA and SpoT in Burkholderia pseudomallei Virulence and Immunity  

PubMed Central

Burkholderia pseudomallei is a Gram-negative soil bacterium and the causative agent of melioidosis, a disease of humans and animals. It is also listed as a category B bioterrorism threat agent by the U.S. Centers for Disease Control and Prevention, and there is currently no melioidosis vaccine available. Small modified nucleotides such as the hyperphosphorylated guanosine molecules ppGpp and pppGpp play an important role as signaling molecules in prokaryotes. They mediate a global stress response under starvation conditions and have been implicated in the regulation of virulence and survival factors in many bacterial species. In this study, we created a relA spoT double mutant in B. pseudomallei strain K96243, which lacks (p)ppGpp-synthesizing enzymes, and investigated its phenotype in vitro and in vivo. The B. pseudomallei ?relA ?spoT mutant displayed a defect in stationary-phase survival and intracellular replication in murine macrophages. Moreover, the mutant was attenuated in the Galleria mellonella insect model and in both acute and chronic mouse models of melioidosis. Vaccination of mice with the ?relA ?spoT mutant resulted in partial protection against infection with wild-type B. pseudomallei. In summary, (p)ppGpp signaling appears to represent an essential component of the regulatory network governing virulence gene expression and stress adaptation in B. pseudomallei, and the ?relA ?spoT mutant may be a promising live-attenuated vaccine candidate. PMID:22778096

Conejero, Laura; Spink, Natasha; Wand, Matthew E.; Bancroft, Gregory J.; Titball, Richard W.

2012-01-01

57

Comparative Burkholderia pseudomallei natural history virulence studies using an aerosol murine model of infection.  

PubMed

Melioidosis is an endemic disease caused by the bacterium Burkholderia pseudomallei. Concerns exist regarding B. pseudomallei use as a potential bio-threat agent causing persistent infections and typically manifesting as severe pneumonia capable of causing fatal bacteremia. Development of suitable therapeutics against melioidosis is complicated due to high degree of genetic and phenotypic variability among B. pseudomallei isolates and lack of data establishing commonly accepted strains for comparative studies. Further, the impact of strain variation on virulence, disease presentation, and mortality is not well understood. Therefore, this study evaluate and compare the virulence and disease progression of B. pseudomallei strains K96243 and HBPUB10303a, following aerosol challenge in a standardized BALB/c mouse model of infection. The natural history analysis of disease progression monitored conditions such as weight, body temperature, appearance, activity, bacteremia, organ and tissue colonization (pathological and histological analysis) and immunological responses. This study provides a detailed, direct comparison of infection with different B. pseudomallei strains and set up the basis for a standardized model useful to test different medical countermeasures against Burkholderia species. Further, this protocol serves as a guideline to standardize other bacterial aerosol models of infection or to define biomarkers of infectious processes caused by other intracellular pathogens. PMID:24603493

Massey, Shane; Yeager, Linsey A; Blumentritt, Carla A; Vijayakumar, Sudhamathi; Sbrana, Elena; Peterson, Johnny W; Brasel, Trevor; LeDuc, James W; Endsley, Janice J; Torres, Alfredo G

2014-01-01

58

Human Melioidosis, Malawi, 2011  

PubMed Central

A case of human melioidosis caused by a novel sequence type of Burkholderia pseudomallei occurred in a child in Malawi, southern Africa. A literature review showed that human cases reported from the continent have been increasing. PMID:23735189

Katangwe, Thembi; Purcell, Janet; Bar-Zeev, Naor; Denis, Brigitte; Montgomery, Jacqui; Alaerts, Maaike; Heyderman, Robert Simon; Dance, David A.B.; Kennedy, Neil; Feasey, Nicholas

2013-01-01

59

Characterization of the Burkholderia pseudomallei K96243 Capsular Polysaccharide I Coding Region  

PubMed Central

Burkholderia pseudomallei is the causative agent of melioidosis, a disease endemic to regions of Southeast Asia and Northern Australia. Both humans and a range of other animal species are susceptible to melioidosis, and the production of a group 3 polysaccharide capsule in B. pseudomallei is essential for virulence. B. pseudomallei capsular polysaccharide (CPS) I comprises unbranched manno-heptopyranose residues and is encoded by a 34.5-kb locus on chromosome 1. Despite the importance of this locus, the role of all of the genes within this region is unclear. We inactivated 18 of these genes and analyzed their phenotype using Western blotting and immunofluorescence staining. Furthermore, by combining this approach with bioinformatic analysis, we were able to develop a model for CPS I biosynthesis and export. We report that inactivating gmhA, wcbJ, and wcbN in B. pseudomallei K96243 retains the immunogenic integrity of the polysaccharide despite causing attenuation in the BALB/c murine infection model. Mice immunized with the B. pseudomallei K96243 mutants lacking a functional copy of either gmhA or wcbJ were afforded significant levels of protection against a wild-type B. pseudomallei K96243 challenge. PMID:22252864

Cuccui, Jon; Milne, Timothy S.; Harmer, Nicholas; George, Alison J.; Harding, Sarah V.; Dean, Rachel E.; Scott, Andrew E.; Sarkar-Tyson, Mitali; Wren, Brendan W.; Prior, Joann L.

2012-01-01

60

A Burkholderia pseudomallei Outer Membrane Vesicle Vaccine Provides Protection against Lethal Sepsis  

PubMed Central

The environmental Gram-negative encapsulated bacillus Burkholderia pseudomallei is the causative agent of melioidosis, a disease associated with high morbidity and mortality rates in areas of Southeast Asia and northern Australia in which the disease is endemic. B. pseudomallei is also classified as a tier I select agent due to the high level of lethality of the bacterium and its innate resistance to antibiotics, as well as the lack of an effective vaccine. Gram-negative bacteria, including B. pseudomallei, secrete outer membrane vesicles (OMVs) which are enriched with multiple protein, lipid, and polysaccharide antigens. Previously, we demonstrated that immunization with multivalent B. pseudomallei-derived OMVs protects highly susceptible BALB/c mice against an otherwise lethal aerosol challenge. In this work, we evaluated the protective efficacy of OMV immunization against intraperitoneal challenge with a heterologous strain because systemic infection with phenotypically diverse environmental B. pseudomallei strains poses another hazard and a challenge to vaccine development. We demonstrated that B. pseudomallei OMVs derived from strain 1026b afforded significant protection against septicemic infection with B. pseudomallei strain K96243. OMV immunization induced robust OMV-, lipopolysaccharide-, and capsular polysaccharide-specific serum IgG (IgG1, IgG2a, and IgG3) and IgM antibody responses. OMV-immune serum promoted bacterial killing in vitro, and passive transfer of B. pseudomallei OMV immune sera protected naive mice against a subsequent challenge. These results indicate that OMV immunization provides antibody-mediated protection against acute, rapidly lethal sepsis in mice. B. pseudomallei-derived OMVs may represent an efficacious multivalent vaccine strategy against melioidosis. PMID:24671550

Nieves, Wildaliz; Petersen, Hailey; Judy, Barbara M.; Blumentritt, Carla A.; Russell-Lodrigue, Kasi; Roy, Chad J.; Torres, Alfredo G.

2014-01-01

61

Alteration of the phenotypic and pathogenic patterns of Burkholderia pseudomallei that persist in a soil environment.  

PubMed

Melioidosis is caused by the soil-borne pathogen Burkholderia pseudomallei. To investigate whether the distinct phenotypic and virulent characteristics result from environmental adaptations in the soil or from the host body, two pairs of isogenic strains were generated by passages in soil or mice. After cultivation in soil, the levels of 3-hydroxytetradecanoic acid, biofilm formation, flagellar expression, and ultrastructure were altered in the bacteria. Uniformly fatal melioidosis developed as a result of infection with mouse-derived strains; however, the survival rates of mice infected with soil-derived strains prolonged. After primary infection or reinfection with soil-derived strains, the mice developed a low degree of bacterial hepatitis and bacterial colonization in the liver and bone marrow compared with mice that were infected with isogenic or heterogenic mouse-derived strains. We suggest that specific phenotypic and pathogenic patterns can be induced through infection with B. pseudomallei that has been cultured in different (soil versus mouse) environments. PMID:24445207

Chen, Yao-Shen; Shieh, Wun-Ju; Goldsmith, Cynthia S; Metcalfe, Maureen G; Greer, Patricia W; Zaki, Sherif R; Chang, Hsin-Hou; Chan, Hao; Chen, Ya-Lei

2014-03-01

62

Fatal Burkholderia pseudomallei Infection Initially Reported as a Bacillus Species, Ohio, 2013.  

PubMed

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 samples for culturing and real-time polymerase chain reaction testing. Family members and pets tested were seronegative for B. pseudomallei. Environmental samples were negative by real-time polymerase chain reaction and culture. Although the patient never traveled internationally, the isolate genotype was consistent with an isolate that originated in Southeast Asia. This investigation identified the fifth reported locally acquired non-laboratory melioidosis case in the contiguous United States. Physicians and laboratories should be aware of this potentially emerging disease and refer positive cultures to a Laboratory Response Network laboratory. PMID:25092821

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

2014-10-01

63

Enzymatic and molecular characterisation of leucine aminopeptidase of Burkholderia pseudomallei  

PubMed Central

Background Leucine aminopeptidase (LAP) has been known to be a housekeeping protease, DNA-binding protein and repressor or activator in the operon regulation of virulence-associated genes in several bacterial species. LAP activity was consistently detected in overnight cultures of Burkholderia pseudomallei, the causative agent of melioidosis and this enzyme was partially purified and characterised in this study. The intra- and inter-species nucleotide and deduced amino acid sequence variation of LAP encoding gene (pepA) was determined. A pepA/PCR-RFLP assay was designed to facilitate the identification of major LAP sequence types amongst clinical and environmental isolates of B. pseudomallei. Results LAP activity was detected in B. pseudomallei culture supernantants by zymographic analysis. Optimum activity was at pH 9 and stable at 50°C. Enhanced enzymatic activity was observed in the presence of metallic ions Mg2+, Ca2+, Na+ and K+. LAP activity was inhibited by EDTA, 1,10-phenanthroline, amastatin, Mn2+ and Zn2+. Sequence analysis of the complete nucleotide and deduced amino acid sequences of LAP-encoding (pepA) gene showed close genetic relatedness to B. mallei (similarity 99.7%/99.6%), but not with B. thailandensis (96.4%/96.4%). Eight pepA sequence types were identified by comparison with a 596 bp DNA fragment encompassing central regions of the pepA gene. A pepA/PCR-RFLP was designed to differentiate pepA sequence types. Based on restriction analysis with StuI and HincII enzymes of the amplified pepA gene, clinical and environmental isolates showed different predominant RFLP types. Type I was the most predominant type amongst 73.6% (67/91) of the clinical isolates, while Type II was predominant in 55.6% (5/9) of the environmental isolates. Conclusions This study showed that LAP is a secretory product of B. pseudomallei with features similar to LAP of other organisms. Identification of major LAP sequence types of B. pseudomallei was made possible based on RFLP analysis of the pepA gene. The high LAP activity detected in both B. pseudomallei and B. thailandensis, suggests that LAP is probably a housekeeping enzyme rather than a virulence determinant. PMID:23682954

2013-01-01

64

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

PubMed Central

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. PMID:23057010

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

2012-01-01

65

Genomic Diversity of Burkholderia pseudomallei Clinical Isolates: Subtractive Hybridization Reveals a Burkholderia mallei-Specific Prophage in B. pseudomallei 1026b  

PubMed Central

Burkholderia pseudomallei is the etiologic agent of the disease melioidosis and is a category B biological threat agent. The genomic sequence of B. pseudomallei K96243 was recently determined, but little is known about the overall genetic diversity of this species. Suppression subtractive hybridization was employed to assess the genetic variability between two distinct clinical isolates of B. pseudomallei, 1026b and K96243. Numerous mobile genetic elements, including a temperate bacteriophage designated ?1026b, were identified among the 1026b-specific suppression subtractive hybridization products. Bacteriophage ?1026b was spontaneously produced by 1026b, and it had a restricted host range, infecting only Burkholderia mallei. It possessed a noncontractile tail, an isometric head, and a linear 54,865-bp genome. The mosaic nature of the ?1026b genome was revealed by comparison with bacteriophage ?E125, a B. mallei-specific bacteriophage produced by Burkholderia thailandensis. The ?1026b genes for DNA packaging, tail morphogenesis, host lysis, integration, and DNA replication were nearly identical to the corresponding genes in ?E125. On the other hand, ?1026b genes involved in head morphogenesis were similar to head morphogenesis genes encoded by Pseudomonas putida and Pseudomonas aeruginosa bacteriophages. Consistent with this observation, immunogold electron microscopy demonstrated that polyclonal antiserum against ?E125 reacted with the tail of ?1026b but not with the head. The results presented here suggest that B. pseudomallei strains are genetically heterogeneous and that bacteriophages are major contributors to the genomic diversity of this species. The bacteriophage characterized in this study may be a useful diagnostic tool for differentiating B. pseudomallei and B. mallei, two closely related biological threat agents. PMID:15175308

DeShazer, David

2004-01-01

66

Liver abscess caused by Burkholderia pseudomallei in a young man: A case report and review of literature.  

PubMed

Pyogenic liver abscess is a common entity in Indian subcontinent and is mostly caused by gram negative bacteria. Melioidosis is not commonly seen in India and only a few cases are reported. It can give rise to multiple abscesses at different sites including liver. We report a case of isolated liver abscess caused by Burkholderia pseudomallei (B. pseudomallei) in a 29-year-old recently diagnosed diabetic, immunocompetent male. Diagnosis was made by imaging and culture of pus aspirated from the abscess and he was treated with percutaneous pigtail catheter drainage followed by antibiotics (meropenem and trimethoprim-sulphmethoxazole). Melioidosis is an emerging infection in India and has high mortality rate, so early diagnosis and prompt management is warranted which requires clinical vigilance and an intensive microbiological workup. Clinicians should be aware of isolated liver abscess caused by B. pseudomallei in appropriate clinical settings. PMID:25325075

Pal, Partha; Ray, Sayantan; Moulick, Avijit; Dey, Subhasis; Jana, Anirban; Banerjee, Kokila

2014-10-16

67

Liver abscess caused by Burkholderia pseudomallei in a young man: A case report and review of literature  

PubMed Central

Pyogenic liver abscess is a common entity in Indian subcontinent and is mostly caused by gram negative bacteria. Melioidosis is not commonly seen in India and only a few cases are reported. It can give rise to multiple abscesses at different sites including liver. We report a case of isolated liver abscess caused by Burkholderia pseudomallei (B. pseudomallei) in a 29-year-old recently diagnosed diabetic, immunocompetent male. Diagnosis was made by imaging and culture of pus aspirated from the abscess and he was treated with percutaneous pigtail catheter drainage followed by antibiotics (meropenem and trimethoprim-sulphmethoxazole). Melioidosis is an emerging infection in India and has high mortality rate, so early diagnosis and prompt management is warranted which requires clinical vigilance and an intensive microbiological workup. Clinicians should be aware of isolated liver abscess caused by B. pseudomallei in appropriate clinical settings. PMID:25325075

Pal, Partha; Ray, Sayantan; Moulick, Avijit; Dey, Subhasis; Jana, Anirban; Banerjee, Kokila

2014-01-01

68

Antimicrobial Susceptibility and Genetic Characterisation of Burkholderia pseudomallei Isolated from Malaysian Patients  

PubMed Central

Burkholderia pseudomallei, the causative agent of melioidosis, is intrinsically resistant to many antibiotics. Ceftazidime (CAZ), the synthetic ?-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). PMID:25379514

Khosravi, Yalda; Mariappan, Vanitha; Ng, Shet-Lee

2014-01-01

69

Within-Host Evolution of Burkholderia pseudomallei over a Twelve-Year Chronic Carriage Infection  

PubMed Central

ABSTRACT Burkholderia pseudomallei causes the potentially fatal disease melioidosis. It is generally accepted that B. pseudomallei is a noncommensal bacterium and that any culture-positive clinical specimen denotes disease requiring treatment. Over a 23-year study of melioidosis cases in Darwin, Australia, just one patient from 707 survivors has developed persistent asymptomatic B. pseudomallei carriage. To better understand the mechanisms behind this unique scenario, we performed whole-genome analysis of two strains isolated 139 months apart. During this period, B. pseudomallei underwent several adaptive changes. Of 23 point mutations, 78% were nonsynonymous and 43% were predicted to be deleterious to gene function, demonstrating a strong propensity for positive selection. Notably, a nonsense mutation inactivated the universal stress response sigma factor RpoS, with pleiotropic implications. The genome underwent substantial reduction, with four deletions in chromosome 2 resulting in the loss of 221 genes. The deleted loci included genes involved in secondary metabolism, environmental survival, and pathogenesis. Of 14 indels, 11 occurred in coding regions and 9 resulted in frameshift mutations that dramatically affected predicted gene products. Disproportionately, four indels affected lipopolysaccharide biosynthesis and modification. Finally, we identified a frameshift mutation in both P314 isolates within wcbR, an important component of the capsular polysaccharide I locus, suggesting virulence attenuation early in infection. Our study illustrates a unique clinical case that contrasts a high-consequence infectious agent with a long-term commensal infection and provides further insights into bacterial evolution within the human host. PMID:23860767

Price, Erin P.; Sarovich, Derek S.; Mayo, Mark; Tuanyok, Apichai; Drees, Kevin P.; Kaestli, Mirjam; Beckstrom-Sternberg, Stephen M.; Babic-Sternberg, James S.; Kidd, Timothy J.; Bell, Scott C.; Keim, Paul; Pearson, Talima; Currie, Bart J.

2013-01-01

70

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

SciTech Connect

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.

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

71

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

PubMed Central

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. PMID:23126230

2012-01-01

72

The Condition-Dependent Transcriptional Landscape of Burkholderia pseudomallei  

PubMed Central

Burkholderia pseudomallei (Bp), the causative agent of the often-deadly infectious disease melioidosis, contains one of the largest prokaryotic genomes sequenced to date, at 7.2 Mb with two large circular chromosomes (1 and 2). To comprehensively delineate the Bp transcriptome, we integrated whole-genome tiling array expression data of Bp exposed to >80 diverse physical, chemical, and biological conditions. Our results provide direct experimental support for the strand-specific expression of 5,467 Sanger protein-coding genes, 1,041 operons, and 766 non-coding RNAs. A large proportion of these transcripts displayed condition-dependent expression, consistent with them playing functional roles. The two Bp chromosomes exhibited dramatically different transcriptional landscapes — Chr 1 genes were highly and constitutively expressed, while Chr 2 genes exhibited mosaic expression where distinct subsets were expressed in a strongly condition-dependent manner. We identified dozens of cis-regulatory motifs associated with specific condition-dependent expression programs, and used the condition compendium to elucidate key biological processes associated with two complex pathogen phenotypes — quorum sensing and in vivo infection. Our results demonstrate the utility of a Bp condition-compendium as a community resource for biological discovery. Moreover, the observation that significant portions of the Bp virulence machinery can be activated by specific in vitro cues provides insights into Bp's capacity as an “accidental pathogen”, where genetic pathways used by the bacterium to survive in environmental niches may have also facilitated its ability to colonize human hosts. PMID:24068961

Nandi, Tannistha; Kreisberg, Jason F.; Chua, Hui Hoon; Sun, Guangwen; Chen, Yahua; Mueller, Claudia; Conejero, Laura; Eshaghi, Majid; Ang, Roy Moh Lik; Liu, Jianhua; Sobral, Bruno W.; Korbsrisate, Sunee; Gan, Yunn Hwen; Titball, Richard W.; Bancroft, Gregory J.; Valade, Eric; Tan, Patrick

2013-01-01

73

Evaluation of a Burkholderia pseudomallei Outer Membrane Vesicle Vaccine in Nonhuman Primates  

PubMed Central

Burkholderia pseudomallei (Bps)is the causative agent of melioidosis and is endemic in regions of northern Australia and Southeast Asia. Bps is inherently resistant to multiple antibiotics and is considered a potential biological warfare agent by the U.S. DHHS. Therefore, effective vaccines are necessary to prevent natural infection and to safeguard against biological attack with this organism. In our previous work we have shown that immunization with naturally derived outer membrane vesicles (OMVs) from Bps provides significant protection against lethal aerosol and systemic infection in BALB/c mice. In this work, we evaluated the safety and immunogenicity of escalating doses of OMV vaccine in rhesus macaques. We show that immunization of rhesus macaques with Bps OMVs generates humoral immuneresponses to protective protein and polysaccharide antigens without any associated toxicity or reactogenicity. These results lay the groundwork for evaluation of protective efficacy of the OMV vaccine in the nonhuman primate model of melioidosis. PMID:25165491

Petersen, Hailey; Nieves, Wildaliz; Russell-Lodrigue, Kasi; Roy, Chad J.; Morici, Lisa A.

2014-01-01

74

Comparison of Rapid, Automated Ribotyping and DNA Macrorestriction Analysis of Burkholderia pseudomallei  

PubMed Central

An automated ribotyping device (RiboPrinter) was used to determine the ribotypes of a collection of Burkholderia pseudomallei isolates. In a preliminary evaluation with the restriction enzymes BamHI and EcoRI, the protocol with EcoRI was more discriminating. The reproducibilities of the ribotypes obtained with EcoRI (EcoRI ribotypes) were determined by testing three levels of bacterial loads. The performance of the manufacturer's software was assessed by comparing the machine-optimized ribotypes with the type determined from the original gel image analyzed with Bionumerics software. The library of B. pseudomallei EcoRI ribotypes was then compared with the ribotypes obtained by DNA macrorestriction analysis of XbaI digests by pulsed-field gel electrophoresis. The typeability of B. pseudomallei by EcoRI ribotyping was 100%, and the discrimination index was 0.94. The slightly greater discrimination provided by DNA macrorestriction analysis (0.96) was achieved at the expense of a significantly longer processing time of 6 days, although the method was only half the cost of automated ribotyping. Typeability by macrorestriction analysis was lower (97%) unless a thiourea step was added to neutralize the action of Tris-dependent endonucleases. The digital record of B. pseudomallei isolates analyzed thus far provides a useful resource for future epidemiological studies and will help shorten the response time in the event of a further melioidosis outbreak or the deliberate release of B. pseudomallei as a biohazard. PMID:12202553

Inglis, Timothy J. J.; O'Reilly, Lyn; Foster, Niki; Clair, Adele; Sampson, Judy

2002-01-01

75

Effects of soil pH, temperature and water content on the growth of Burkholderia pseudomallei.  

PubMed

Optimum conditions were determined for the growth of Burkholderia pseudomallei in natural soils or waters. It grows better in paddy soil, crop-covered and fallow field than in fresh and salty water. Although the optimal temperature and pH for the growth were 37 or 42 degrees C, and 6.5 or 7.5 in an environmental-mimicking soil medium, this bacterium can still grow at 4 degrees C, which was suggested to be related with the occurrence of melioidosis in some cold areas. In soil media with water content < 15. B. pseudomallei did not grow until 60 d of incubation, suggesting that water contents of soils in which it dwelled would be one important factor in determining the growth rate. PMID:12800512

Chen, Y S; Chen, S C; Kao, C M; Chen, Y L

2003-01-01

76

Immunogenic recombinant Burkholderia pseudomallei MprA serine protease elicits protective immunity in mice  

PubMed Central

Burkholderia pseudomallei is resistant to a diverse group of antimicrobials including third generation cephalosporins whilst quinolones and aminoglycosides have no reliable effect. As therapeutic options are limited, development of more effective forms of immunotherapy is vital to avoid a fatal outcome. In an earlier study, we reported on the B. pseudomallei serine MprA protease, which is relatively stable over a wide pH and temperature range and digests physiological proteins. The present study was carried out to evaluate the immunogenicity and protective efficacy of the MprA as a potential vaccine candidate. In BALB/c mice immunized with recombinant MprA protease (smBpF4), a significantly high IgG titer was detectable. Isotyping studies revealed that the smBpF4-specific antibodies produced were predominantly IgG1, proposing that immunization with smBpF4 triggered a Th2 immune response. Mice were immunized with smBpF4 and subsequently challenged with B. pseudomallei via the intraperitoneal route. Whilst control mice succumbed to the infection by day 9, smBpF4-immunized mice were protected against the lethal challenge and survived beyond 25 days post-infection. In conclusion, MprA is immunogenic in melioidosis patients whilst also eliciting a strong immune response upon bacterial challenge in mice and presents itself as a potential vaccine candidate for the treatment of melioidosis. PMID:22919676

Chin, Chui-Yoke; Tan, Swee-Chen; Nathan, Sheila

2012-01-01

77

In vitro antimicrobial activity of natural toxins and animal venoms tested against Burkholderia pseudomallei  

PubMed Central

Background Burkholderia pseudomallei are the causative agent of melioidosis. Increasing resistance of the disease to antibiotics is a severe problem in treatment regime and has led to intensification of the search for new drugs. Antimicrobial peptides are the most ubiquitous in nature as part of the innate immune system and host defense mechanism. Methods Here, we investigated a group of venoms (snakes, scorpions and honey bee venoms) for antimicrobial properties against two strains of Gram-negative bacteria Burkholderia pseudomallei by using disc-diffusion assay for in vitro susceptibility testing. The antibacterial activities of the venoms were compared with that of the isolated L-amino acid oxidase (LAAO) and phospholipase A2 (PLA2s) enzymes. MICs were determined using broth dilution method. Bacterial growth was assessed by measurement of optical density at the lowest dilutions (MIC 0.25 mg/ml). The cell viability was measured using tetrazolium salts (XTT) based cytotoxic assay. Results The studied venoms showed high antimicrobial activity. The venoms of C. adamanteus, Daboia russelli russelli, A. halys, P. australis, B. candidus and P. guttata were equally as effective as Chloramphenicol and Ceftazidime (30 ?g/disc). Among those tested, phospholipase A2 enzymes (crotoxin B and daboiatoxin) showed the most potent antibacterial activity against Gram-negative (TES) bacteria. Naturally occurring venom peptides and phospholipase A2 proved to possess highly potent antimicrobial activity against Burkholderia pseudomallei. The XTT-assay results showed that the cell survival decreased with increasing concentrations (0.05–10 mg/mL) of Crotalus adamanteus venom, with no effect on the cell viability evident at 0.5 mg/mL. Conclusion This antibacterial profile of snake venoms reported herein will be useful in the search for potential antibacterial agents against drug resistant microorganisms like B. pseudomallei. PMID:16784542

Perumal Samy, R; Pachiappan, A; Gopalakrishnakone, P; Thwin, Maung M; Hian, Yap E; Chow, Vincent TK; Bow, Ho; Weng, Joseph T

2006-01-01

78

Screening for potential anti-infective agents towards Burkholderia pseudomallei infection  

NASA Astrophysics Data System (ADS)

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.

Eng, Su Anne; Nathan, Sheila

2014-09-01

79

In Vitro Activity of Doripenem against Burkholderia pseudomallei  

Microsoft Academic Search

The MIC50 and MIC90 values of doripenem, determined by Etest, for 110 isolates of Burkholderia pseudoma- llei were 0.5 and 0.75 g\\/ml, respectively. There were significant correlations between MICs determined by Etest and MICs determined by agar dilution, MICs determined by Etest and inhibition zone size, and MICs determined by agar dilution and inhibition zone size. Burkholderia pseudomallei, a gram-negative

Visanu Thamlikitkul; Suwanna Trakulsomboon

2009-01-01

80

Identification of a Predicted Trimeric Autotransporter Adhesin Required for Biofilm Formation of Burkholderia pseudomallei  

PubMed Central

The autotransporters are a large and diverse family of bacterial secreted and outer membrane proteins, which are present in many Gram-negative bacterial pathogens and play a role in numerous environmental and virulence-associated interactions. As part of a larger systematic study on the autotransporters of Burkholderia pseudomallei, the causative agent of the severe tropical disease melioidosis, we have constructed an insertion mutant in the bpss1439 gene encoding an unstudied predicted trimeric autotransporter adhesin. The bpss1439 mutant demonstrated a significant reduction in biofilm formation at 48 hours in comparison to its parent 10276 wild-type strain. This phenotype was complemented to wild-type levels by the introduction of a full-length copy of the bpss1439 gene in trans. Examination of the wild-type and bpss1439 mutant strains under biofilm-inducing conditions by microscopy after 48 hours confirmed that the bpss1439 mutant produced less biofilm compared to wild-type. Additionally, it was observed that this phenotype was due to low levels of bacterial adhesion to the abiotic surface as well as reduced microcolony formation. In a murine melioidosis model, the bpss1439 mutant strain demonstrated a moderate attenuation for virulence compared to the wild-type strain. This attenuation was abrogated by in trans complementation, suggesting that bpss1439 plays a subtle role in the pathogenesis of B. pseudomallei. Taken together, these studies indicate that BPSS1439 is a novel predicted autotransporter involved in biofilm formation of B. pseudomallei; hence, this factor was named BbfA, Burkholderia biofilm factor A. PMID:24223950

Lazar Adler, Natalie R.; Dean, Rachel E.; Saint, Richard J.; Stevens, Mark P.; Prior, Joann L.; Atkins, Timothy P.; Galyov, Edouard E.

2013-01-01

81

Persistent Gastric Colonization with Burkholderia pseudomallei and Dissemination from the Gastrointestinal Tract following Mucosal Inoculation of Mice  

PubMed Central

Melioidosis is a disease of humans caused by opportunistic infection with the soil and water bacterium Burkholderia pseudomallei. Melioidosis can manifest as an acute, overwhelming infection or as a chronic, recurrent infection. At present, it is not clear where B. pseudomallei resides in the mammalian host during the chronic, recurrent phase of infection. To address this question, we developed a mouse low-dose mucosal challenge model of chronic B. pseudomallei infection and investigated sites of bacterial persistence over 60 days. Sensitive culture techniques and selective media were used to quantitate bacterial burden in major organs, including the gastrointestinal (GI) tract. We found that the GI tract was the primary site of bacterial persistence during the chronic infection phase, and was the only site from which the organism could be consistently cultured during a 60-day infection period. The organism could be repeatedly recovered from all levels of the GI tract, and chronic infection was accompanied by sustained low-level fecal shedding. The stomach was identified as the primary site of GI colonization as determined by fluorescent in situ hybridization. Organisms in the stomach were associated with the gastric mucosal surface, and the propensity to colonize the gastric mucosa was observed with 4 different B. pseudomallei isolates. In contrast, B. pseudomallei organisms were present at low numbers within luminal contents in the small and large intestine and cecum relative to the stomach. Notably, inflammatory lesions were not detected in any GI tissue examined in chronically-infected mice. Only low-dose oral or intranasal inoculation led to GI colonization and development of chronic infection of the spleen and liver. Thus, we concluded that in a mouse model of melioidosis B. pseudomallei preferentially colonizes the stomach following oral inoculation, and that the chronically colonized GI tract likely serves as a reservoir for dissemination of infection to extra-intestinal sites. PMID:22624016

Goodyear, Andrew; Bielefeldt-Ohmann, Helle; Schweizer, Herbert; Dow, Steven

2012-01-01

82

Comparative in vitro susceptibility of Burkholderia pseudomallei to doripenem, ertapenem, tigecycline and moxifloxacin.  

PubMed

Burkholderia pseudomallei, the causative agent of melioidosis, continues to present therapeutic challenges in endemic areas. A number of clinical issues have prompted consideration of alternative antimicrobial therapies. These include stability in 24-h infusion pumps, broad-spectrum coverage in the empirical treatment of community-acquired pneumonia, cost, the need for effective oral agents and rare reports of emerging resistance. This study aimed to examine the in vitro susceptibility of B. pseudomallei to four new antimicrobial agents, namely moxifloxacin, tigecycline, ertapenem and doripenem. A total of 100 clinical isolates were tested by Etest and disk diffusion. As there are no interpretative standards for these antimicrobials, MIC(90) values (minimum inhibitory concentrations for 90% of the isolates) were compared with those for meropenem. MIC values for each agent were correlated with zone of inhibition diameters. MICs for doripenem were broadly similar to those for meropenem, with a MIC(90) of 1.5 ?g/mL (range 0.38-4 ?g/mL). There was good correlation (r=-0.71; P<0.001) between the MIC and disk diffusion for doripenem. Ertapenem, tigecycline and moxifloxacin had limited in vitro activity in this study, although no interpretative criteria exist for these agents. Further in vitro, animal and clinical studies are suggested to validate the efficacy of doripenem in the management of melioidosis. PMID:21481571

Harris, Patrick; Engler, Cathy; Norton, Robert

2011-06-01

83

Evolutionary Analysis of Burkholderia pseudomallei Identifies Putative Novel Virulence Genes, Including a Microbial Regulator of Host Cell Autophagy  

PubMed Central

Burkholderia pseudomallei, the causative agent of melioidosis, contains a large pathogen genome (7.2 Mb) with ?2,000 genes of putative or unknown function. Interactions with potential hosts and environmental factors may induce rapid adaptations in these B. pseudomallei genes, which can be discerned through evolutionary analysis of multiple B. pseudomallei genomes. Here we show that several previously uncharacterized B. pseudomallei genes bearing genetic signatures of rapid adaptation (positive selection) can induce diverse cellular phenotypes when expressed in mammalian cells. Notably, several of these phenotypes are plausibly related to virulence, including multinuclear giant cell formation, apoptosis, and autophagy induction. Specifically, we show that BPSS0180, a type VI cluster-associated gene, is capable of inducing autophagy in both phagocytic and nonphagocytic mammalian cells. Following infection of macrophages, a B. pseudomallei mutant disrupted in BPSS0180 exhibited significantly decreased colocalization with LC3 and impaired intracellular survival; these phenotypes were rescued by introduction of an intact BPSS0180 gene. The results suggest that BPSS0180 may be a novel inducer of host cell autophagy that contributes to B. pseudomallei intracellular growth. More generally, our study highlights the utility of applying evolutionary principles to microbial genomes to identify novel virulence genes. PMID:24097950

Singh, Arvind Pratap; Lai, Shu-chin; Nandi, Tannistha; Chua, Hui Hoon; Ooi, Wen Fong; Ong, Catherine; Boyce, John D.; Adler, Ben

2013-01-01

84

Identification and Discrimination of Burkholderia pseudomallei, B. mallei, and B. thailandensis by Real-Time PCR Targeting Type III Secretion System Genes  

PubMed Central

Burkholderia pseudomallei and B. mallei are two highly pathogenic bacteria, responsible for melioidosis and glanders, respectively. The two are closely related and can also be mistaken for B. thailandensis, a nonpathogenic species. To improve their differential identification, we describe a hydrolysis probe-based real-time PCR method using the uneven distribution of type III secretion system genes among these three species. PMID:15583328

Thibault, F. M.; Valade, E.; Vidal, D. R.

2004-01-01

85

Distribution of type III secretion gene clusters in Burkholderia pseudomallei, B. thailandensis and B. mallei.  

PubMed

Burkholderia pseudomallei, the causative agent of melioidosis, carries a cluster of genes closely related in organisation to the type III secretion (TTS) system gene clusters of the plant pathogens Ralstonia solanacearum and Xanthomonas spp. The TTS gene cluster (TTS1) is present only in B. pseudomallei and not in avirulent B. thailandensis. Adjacent to the gene cluster encoding putative secreton structural proteins lie a number of open reading frames (ORFs) encoding putative proteins with little or no homology to known proteins, with the exception of one predicted protein with homology to Pseudomonas syringae HrpK. In both R. solanacearum and Xanthomonas spp., genes in this location encode secreted effector proteins. RT-PCR analysis indicated that TTS genes, including two of these ORFs, are expressed in broth at 37 degrees C. Analysis of genome sequence data identified a second cluster of TTS genes (TTS2) present in both B. pseudomallei and B. mallei (99% identity). However, B. mallei appears to lack the TTS1 gene cluster. PCR assays indicated that TTS2 was also present in B. thailandensis. TTS1 and TTS2 are similar in gene organisation, but nucleotide sequences are sufficiently divergent to suggest that the two TTS systems may have different roles. PMID:11990489

Rainbow, Lucille; Hart, C Anthony; Winstanley, Craig

2002-05-01

86

Burkholderia pseudomallei in soil samples from an oceanarium in Hong Kong detected using a sensitive PCR assay  

PubMed Central

Melioidosis, caused by Burkholderia pseudomallei, is an emerging infectious disease with an expanding geographical distribution. Although assessment of the environmental load of B. pseudomallei is important for risk assessment in humans or animals in endemic areas, traditional methods of bacterial culture for isolation have low sensitivities and are labor-intensive. Using a specific polymerase chain reaction (PCR) assay targeting a Tat domain protein in comparison with a bacterial culture method, we examined the prevalence of B. pseudomallei in soil samples from an oceanarium in Hong Kong where captive marine mammals and birds have contracted melioidosis. Among 1420 soil samples collected from various sites in the oceanarium over a 15-month period, B. pseudomallei was detected in nine (0.6%) soil samples using bacterial culture, whereas it was detected in 96 (6.8%) soil samples using the specific PCR assay confirmed by sequencing. The PCR-positive samples were detected during various months, with higher detection rates observed during summer months. Positive PCR detection was significantly correlated with ambient temperature (P<0.0001) and relative humidity (P=0.011) but not with daily rainfall (P=0.241) or a recent typhoon (P=0.787). PCR-positive samples were obtained from all sampling locations, with the highest detection rate in the valley. Our results suggest that B. pseudomallei is prevalent and endemic in the oceanarium. The present PCR assay is more sensitive than the bacterial culture method, and it may be used to help better assess the transmission of melioidosis and to design infection control measures for captive animals in this unique and understudied environment.

Lau, Susanna KP; Chan, San-Yuen; Curreem, Shirly OT; Hui, Suk-Wai; Lau, Candy CY; Lee, Paul; Ho, Chi-Chun; Martelli, Paolo; Woo, Patrick CY

2014-01-01

87

Identification of the conserved hypothetical protein BPSL0317 in Burkholderia pseudomallei K96243  

NASA Astrophysics Data System (ADS)

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.

Yusoff, Nur Syamimi; Damiri, Nadzirah; Firdaus-Raih, Mohd

2014-09-01

88

Characterization of BcaA, a Putative Classical Autotransporter Protein in Burkholderia pseudomallei  

PubMed Central

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 Bp340?bcaA and Bp340?bcaB 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 Bp340?bcaA, Bp340?bcaB, and wild-type bacteria. However, significantly fewer bacteria were recovered from the spleen of Bp340?bcaA-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. PMID:23340315

Campos, Cristine G.; Borst, Luke

2013-01-01

89

Quorum Sensing Negatively Regulates Multinucleate Cell Formation during Intracellular Growth of Burkholderia pseudomallei in Macrophage-Like Cells  

PubMed Central

Burkholderia pseudomallei is a Gram-negative environmental bacterium and the causative agent of melioidosis, a potentially fatal, acute or chronic disease endemic in the tropics. Acyl homoserine lactone (AHL)-mediated quorum sensing and signalling have been associated with virulence and biofilm formation in numerous bacterial pathogens. In the canonical acyl-homoserine lactone signalling paradigm, AHLs are detected by a response regulator. B. pseudomallei encodes three AHL synthases, encoded by bpsI1, bpsI2 and bpsI3, and five regulator genes. In this study, we mutated the B. pseudomallei AHL synthases individually and in double and triple combination. Five AHLs were detected and quantified by tandem liquid chromatography-mass spectroscopy. The major AHLs produced were N-octanoylhomoserine lactone and N-(3-hydroxy-decanoyl)homoserine lactone, the expression of which depended on bpsI1 and bpsI2, respectively. B. pseudomallei infection of macrophage cells causes cell fusion, leading to multinucleated cells (3 or more nuclei per cell). A triple mutant defective in production of all three AHL synthases was associated with a striking phenotype of massively enhanced host cellular fusion in macrophages. However, neither abrogation of host cell fusion, achieved by mutation of bimA or hcp1, nor enhancement of fusion altered intracellular replication of B. pseudomallei. Furthermore, when tested in murine models of acute melioidosis the AHL synthase mutants were not attenuated for virulence. Collectively, this study identifies important new aspects of the genetic basis of AHL synthesis in B. pseudomallei and the roles of these AHLs in systemic infection and in cell fusion in macrophages for this important human pathogen. PMID:23704903

Horton, Rachel E.; Grant, Gary D.; Matthews, Ben; Batzloff, Michael; Owen, Suzzanne J.; Kyan, Stephanie; Flegg, Cameron P.; Clark, Amanda M.; Ulett, Glen C.; Morrison, Nigel; Peak, Ian R.; Beacham, Ifor R.

2013-01-01

90

Rapid Identification of Burkholderia pseudomallei in Blood Cultures by a Monoclonal Antibody Assay  

PubMed Central

Burkholderia pseudomallei is the causative agent of melioidosis. In northeast Thailand, this gram-negative bacterium is a major cause of mortality from septicemia. The definitive diagnosis of this disease is made by bacterial culture. In this study, we produced a monoclonal antibody (MAb) specific to the 30-kDa protein of B. pseudomallei by in vivo and in vitro immunization of BALB/c mice with a crude culture filtrate antigen. The MAb could directly agglutinate with all 243 clinical isolates of B. pseudomallei but not with other gram-negative bacteria, except for one strain of Burkholderia mallei. However, the MAb cross-reacted with the gram-positive Bacillus sp. and Streptococcus pyogenes. B. pseudomallei in brain heart infusion broth (BHIB) subcultured from a BacT/Alert automated blood culture system could be identified by simple agglutination with this MAb assay. The sensitivity and specificity of direct agglutination compared to the “gold standard,” the culture method, were 94.12 and 98.25%, respectively. However, the MAb adsorbed to polystyrene beads or latex particles directly identified the bacterium in blood culture specimens and in BHIB subcultured from a BacT/Alert automated blood culture system. The sensitivity of the latex agglutination test was 100% for both blood culture and BHIB specimens. The specificity was 85.96 and 96.49% for the blood culture and BHIB specimens, respectively. The specificity could be increased if the nonspecific materials in the blood culture broths were eradicated by centrifugation at low speeds. Thus, a combination of blood culture and the agglutination method could be used for the rapid diagnosis of melioidosis in the routine bacteriological laboratory. This method could speed up detection of the bacterium in blood culture by at least 2 days, compared to the conventional bacterial culture method. In addition, the MAb is stable at room temperature for 2 weeks and at 4, ?20, and ?70°C for at least 1 year. The latex reagent was stable for at least 6 months at 4°C. PMID:10523570

Pongsunk, Supinya; Thirawattanasuk, Nittaya; Piyasangthong, Nuanchan; Ekpo, Pattama

1999-01-01

91

The Madagascar hissing cockroach as a novel surrogate host for Burkholderia pseudomallei, B. mallei and B. thailandensis  

PubMed Central

Background Burkholderia pseudomallei and Burkholderia mallei are gram-negative pathogens responsible for the diseases melioidosis and glanders, respectively. Both species cause disease in humans and animals and have been designated as category B select agents by the Centers for Disease Control and Prevention (CDC). Burkholderia thailandensis is a closely related bacterium that is generally considered avirulent for humans. While it can cause disease in rodents, the B. thailandensis 50% lethal dose (LD50) is typically???104-fold higher than the B. pseudomallei and B. mallei LD50 in mammalian models of infection. Here we describe an alternative to mammalian hosts in the study of virulence and host-pathogen interactions of these Burkholderia species. Results Madagascar hissing cockroaches (MH cockroaches) possess a number of qualities that make them desirable for use as a surrogate host, including ease of breeding, ease of handling, a competent innate immune system, and the ability to survive at 37°C. MH cockroaches were highly susceptible to infection with B. pseudomallei, B. mallei and B. thailandensis and the LD50 was <10 colony-forming units (cfu) for all three species. In comparison, the LD50 for Escherichia coli in MH cockroaches was >105?cfu. B. pseudomallei, B. mallei, and B. thailandensis cluster 1 type VI secretion system (T6SS-1) mutants were all attenuated in MH cockroaches, which is consistent with previous virulence studies conducted in rodents. B. pseudomallei mutants deficient in the other five T6SS gene clusters, T6SS-2 through T6SS-6, were virulent in both MH cockroaches and hamsters. Hemocytes obtained from MH cockroaches infected with B. pseudomallei harbored numerous intracellular bacteria, suggesting that this facultative intracellular pathogen can survive and replicate inside of MH cockroach phagocytic cells. The hemolymph extracted from these MH cockroaches also contained multinuclear giant cells (MNGCs) with intracellular B. pseudomallei, which indicates that infected hemocytes can fuse while flowing through the insect’s open circulatory system in vivo. Conclusions The results demonstrate that MH cockroaches are an attractive alternative to mammals to study host-pathogen interactions and may allow the identification of new Burkholderia virulence determinants. The importance of T6SS-1 as a virulence factor in MH cockroaches and rodents suggests that the primary role of this secretion system is to target evasion of the innate immune system. PMID:22892068

2012-01-01

92

The Multiple Roles of Hypothetical Gene BPSS1356 in Burkholderia pseudomallei  

PubMed Central

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 ?? 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 ?BPSS1356 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 ?BPSS1356 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. PMID:24927285

Yam, Hokchai; Abdul Rahim, Ainihayati; Mohamad, Suriani; Mahadi, Nor Muhammad; Abdul Manaf, Uyub; Shu-Chien, Alexander Chong; Najimudin, Nazalan

2014-01-01

93

ATP-binding cassette systems in Burkholderia pseudomallei and Burkholderia mallei  

Microsoft Academic Search

BACKGROUND: ATP binding cassette (ABC) systems are responsible for the import and export of a wide variety of molecules across cell membranes and comprise one of largest protein superfamilies found in prokarya, eukarya and archea. ABC systems play important roles in bacterial lifestyle, virulence and survival. In this study, an inventory of the ABC systems of Burkholderia pseudomallei strain K96243

David N Harland; Elie Dassa; Richard W Titball; Katherine A Brown; Helen S Atkins

2007-01-01

94

Multiplex qPCR for reliable detection and differentiation of Burkholderia mallei and Burkholderia pseudomallei  

PubMed Central

Background Burkholderia mallei and B. pseudomallei are two closely related species of highly virulent bacteria that can be difficult to detect. Pathogenic Burkholderia are endemic in many regions worldwide and cases of infection, sometimes brought by travelers from unsuspected regions, also occur elsewhere. Rapid, sensitive methods for identification of B. mallei and B. pseudomallei are urgently needed in the interests of patient treatment and epidemiological surveillance. Methods Signature sequences for sensitive, specific detection of pathogenic Burkholderia based on published genomes were identified and a qPCR assay was designed and validated. Results A single-reaction quadruplex qPCR assay for the detection of pathogenic Burkholderia, which includes a marker for internal control of DNA extraction and amplification, was developed. The assay permits differentiation of B. mallei and B. pseudomallei strains, and probit analysis showed a very low detection limit. Use of a multicopy signature sequence permits detection of less than 1 genome equivalent per reaction. Conclusions The new assay permits rapid detection of pathogenic Burkholderia and combines enhanced sensitivity, species differentiation, and inclusion of an internal control for both DNA extraction and PCR amplification. PMID:23409683

2013-01-01

95

Comparison of DNA Extraction Kits for Detection of Burkholderia pseudomallei in Spiked Human Whole Blood Using Real-Time PCR  

PubMed Central

Burkholderia pseudomallei, the etiologic agent of melioidosis, is endemic in northern Australia and Southeast Asia and can cause severe septicemia that may lead to death in 20% to 50% of cases. Rapid detection of B. pseudomallei infection is crucial for timely treatment of septic patients. This study evaluated seven commercially available DNA extraction kits to determine the relative recovery of B. pseudomallei DNA from spiked EDTA-containing human whole blood. The evaluation included three manual kits: the QIAamp DNA Mini kit, the QIAamp DNA Blood Mini kit, and the High Pure PCR Template Preparation kit; and four automated systems: the MagNAPure LC using the DNA Isolation Kit I, the MagNAPure Compact using the Nucleic Acid Isolation Kit I, and the QIAcube using the QIAamp DNA Mini kit and the QIAamp DNA Blood Mini kit. Detection of B. pseudomallei DNA extracted by each kit was performed using the B. pseudomallei specific type III secretion real-time PCR (TTS1) assay. Crossing threshold (CT) values were used to compare the limit of detection and reproducibility of each kit. This study also compared the DNA concentrations and DNA purity yielded for each kit. The following kits consistently yielded DNA that produced a detectable signal from blood spiked with 5.5×104 colony forming units per mL: the High Pure PCR Template Preparation, QIAamp DNA Mini, MagNA Pure Compact, and the QIAcube running the QIAamp DNA Mini and QIAamp DNA Blood Mini kits. The High Pure PCR Template Preparation kit yielded the lowest limit of detection with spiked blood, but when this kit was used with blood from patients with confirmed cases of melioidosis, the bacteria was not reliably detected indicating blood may not be an optimal specimen. PMID:23460920

Podnecky, Nicole L.; Elrod, Mindy G.; Newton, Bruce R.; Dauphin, Leslie A.; Shi, Jianrong; Chawalchitiporn, Sutthinan; Baggett, Henry C.; Hoffmaster, Alex R.; Gee, Jay E.

2013-01-01

96

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

PubMed

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

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

2014-11-15

97

Characterization and analysis of the Burkholderia pseudomallei BsaN virulence regulon  

PubMed Central

Background Burkholderia pseudomallei is a facultative intracellular pathogen and the causative agent of melioidosis. A conserved type III secretion system (T3SS3) and type VI secretion system (T6SS1) are critical for intracellular survival and growth. The T3SS3 and T6SS1 genes are coordinately and hierarchically regulated by a TetR-type regulator, BspR. A central transcriptional regulator of the BspR regulatory cascade, BsaN, activates a subset of T3SS3 and T6SS1 loci. Results To elucidate the scope of the BsaN regulon, we used RNAseq analysis to compare the transcriptomes of wild-type B. pseudomallei KHW and a bsaN deletion mutant. The 60 genes positively-regulated by BsaN include those that we had previously identified in addition to a polyketide biosynthesis locus and genes involved in amino acid biosynthesis. BsaN was also found to repress the transcription of 51 genes including flagellar motility loci and those encoding components of the T3SS3 apparatus. Using a promoter-lacZ fusion assay in E. coli, we show that BsaN together with the chaperone BicA directly control the expression of the T3SS3 translocon, effector and associated regulatory genes that are organized into at least five operons (BPSS1516-BPSS1552). Using a mutagenesis approach, a consensus regulatory motif in the promoter regions of BsaN-regulated genes was shown to be essential for transcriptional activation. Conclusions BsaN/BicA functions as a central regulator of key virulence clusters in B. pseudomallei within a more extensive network of genetic regulation. We propose that BsaN/BicA controls a gene expression program that facilitates the adaption and intracellular survival of the pathogen within eukaryotic hosts. PMID:25085508

2014-01-01

98

Antimicrobial Action of the Cyclic Peptide Bactenecin on Burkholderia pseudomallei Correlates with Efficient Membrane Permeabilization  

PubMed Central

Burkholderia pseudomallei is a category B agent that causes Melioidosis, an acute and chronic disease with septicemia. The current treatment regimen is a heavy dose of antibiotics such as ceftazidime (CAZ); however, the risk of a relapse is possible. Peptide antibiotics are an alternative to classical antibiotics as they exhibit rapid action and are less likely to result in the development of resistance. The aim of this study was to determine the bactericidal activity against B. pseudomallei and examine the membrane disrupting abilities of the potent antimicrobial peptides: bactenecin, RTA3, BMAP-18 and CA-MA. All peptides exhibited >97% bactericidal activity at 20 µM, with bactenecin having slightly higher activity. Long term time-kill assays revealed a complete inhibition of cell growth at 50 µM bactenecin and CA-MA. All peptides inhibited biofilm formation comparable to CAZ, but exhibited faster kinetics (within 1 h). Bactenecin exhibited stronger binding to LPS and induced perturbation of the inner membrane of live cells. Interaction of bactenecin with model membranes resulted in changes in membrane fluidity and permeability, leading to leakage of dye across the membrane at levels two-fold greater than that of other peptides. Modeling of peptide binding on the membrane showed stable and deep insertion of bactenecin into the membrane (up to 9 Å). We propose that bactenecin is able to form dimers or large ?-sheet structures in a concentration dependent manner and subsequently rapidly permeabilize the membrane, leading to cytosolic leakage and cell death in a shorter period of time compared to CAZ. Bactenecin might be considered as a potent antimicrobial agent for use against B. pseudomallei. PMID:23785532

Phophetleb, Onanong; Nasompag, Sawinee; Thammasirirak, Sompong; Daduang, Sakda; Taweechaisupapong, Suwimol; Lomize, Andrei L.; Patramanon, Rina

2013-01-01

99

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

PubMed

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. PMID:24961697

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

100

Proteomic analysis of the Burkholderia pseudomallei type II secretome reveals hydrolytic enzymes, novel proteins, and the deubiquitinase TssM.  

PubMed

Burkholderia pseudomallei, the etiologic agent of melioidosis, is an opportunistic pathogen that harbors a wide array of secretion systems, including a type II secretion system (T2SS), three type III secretion systems (T3SS), and six type VI secretion systems (T6SS). The proteins exported by these systems provide B. pseudomallei with a growth advantage in vitro and in vivo, but relatively little is known about the full repertoire of exoproducts associated with each system. In this study, we constructed deletion mutations in gspD and gspE, T2SS genes encoding an outer membrane secretin and a cytoplasmic ATPase, respectively. The secretion profiles of B. pseudomallei MSHR668 and its T2SS mutants were noticeably different when analyzed by SDS-PAGE. We utilized liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify proteins present in the supernatants of B. pseudomallei MSHR668 and B. pseudomallei ?gspD grown in rich and minimal media. The MSHR668 supernatants contained 48 proteins that were either absent or substantially reduced in the supernatants of ?gspD strains. Many of these proteins were putative hydrolytic enzymes, including 12 proteases, two phospholipases, and a chitinase. Biochemical assays validated the LC-MS/MS results and demonstrated that the export of protease, phospholipase C, and chitinase activities is T2SS dependent. Previous studies had failed to identify the mechanism of secretion of TssM, a deubiquitinase that plays an integral role in regulating the innate immune response. Here we present evidence that TssM harbors an atypical signal sequence and that its secretion is mediated by the T2SS. This study provides the first in-depth characterization of the B. pseudomallei T2SS secretome. PMID:24866793

Burtnick, Mary N; Brett, Paul J; DeShazer, David

2014-08-01

101

BPSS1504, a Cluster 1 Type VI Secretion Gene, Is Involved in Intracellular Survival and Virulence of Burkholderia pseudomallei  

PubMed Central

Burkholderia pseudomallei is a Gram-negative rod and the causative agent of melioidosis, an emerging infectious disease of tropical and subtropical areas worldwide. B. pseudomallei harbors a remarkable number of virulence factors, including six type VI secretion systems (T6SS). Using our previously described plaque assay screening system, we identified a B. pseudomallei transposon mutant defective in the BPSS1504 gene that showed reduced plaque formation. The BPSS1504 locus is encoded within T6SS cluster 1 (T6SS1), which is known to be involved in the pathogenesis of B. pseudomallei in mammalian hosts. For further analysis, a B. pseudomallei BPSS1504 deletion (Bp?BPSS1504) mutant and complemented mutant strain were constructed. B. pseudomallei lacking the BPSS1504 gene was highly attenuated in BALB/c mice, whereas the in vivo virulence of the complemented mutant strain was fully restored to the wild-type level. The Bp?BPSS1504 mutant showed impaired intracellular replication and formation of multinucleated giant cells in macrophages compared with wild-type bacteria, whereas the induction of actin tail formation within host cells was not affected. These observations resembled the phenotype of a mutant lacking hcp1, which is an integral component of the T6SS1 apparatus and is associated with full functionality of the T6SS1. Transcriptional expression of the T6SS components vgrG, tssA, and hcp1, as well as the T6SS regulators virAG, bprC, and bsaN, was not dependent on BPSS1504 expression. However, secretion of Hcp1 was not detectable in the absence of BPSS1504. Thus, BPSS1504 seems to serve as a T6SS component that affects Hcp1 secretion and is therefore involved in the integrity of the T6SS1 apparatus. PMID:24595140

Hopf, Verena; Gohler, Andre; Eske-Pogodda, Kristin; Bast, Antje; Steinmetz, Ivo

2014-01-01

102

Comparison of the protective effects of killed Burkholderia pseudomallei and CpG oligodeoxynucleotide against live challenge.  

PubMed

Melioidosis is a fatal disease caused by Burkholderia pseudomallei. Currently there is no vaccine available. Synthetic oligodeoxynucleotides with unmethylated CpG dinucleotide motifs (CpG ODN) can stimulate vertebrate immune cells and clear certain pathogens that are susceptible to a strong Th1 response. In our previous study, pretreatment with CpG ODN alone or CpG-ODN with cationic liposomes for 2-10 or 30 days before B. pseudomallei infection in mice conferred 80-100% protection. In the present study we investigated the protective effect of CpG-ODN together with heat-killed (HK) or paraformaldehyde-killed B. pseudomallei (PP). HK or PP were used to immunize BALB/c mice twice at 15-day intervals before intra-peritoneal challenge with 5LD50 of B. pseudomallei and observed for 30 days. We found that PP could significantly protect mice (60%) with an increased survival time (24.8±11.63 days) while in the HK and PBS groups, all infected mice died within 6 days. Although either CpG ODN or PP conferred significant protection, giving them in combination did not enhance it further. Serum IFN-? levels on day-5 (before challenge) of the PP and PP+CpG ODN groups were significantly higher than those of the PBS control group. The results further support the importance of IFN-? in host protection against B. pseudomallei and suggest further study on paraformaldehyde-killed bacteria as a component of a future B. pseudomallei vaccine. PMID:25223269

Puangpetch, Apichaya; Anderson, Robert; Huang, Yan Y; Saengsot, Rojana; Sermswan, Rasana W; Wongratanacheewin, Surasakdi

2014-10-14

103

Caspase-1-Dependent and -Independent Cell Death Pathways in Burkholderia pseudomallei Infection of Macrophages  

PubMed Central

The cytosolic pathogen Burkholderia pseudomallei and causative agent of melioidosis has been shown to regulate IL-1? 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-1? 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-1?, 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-dependent and -independent cell death mechanisms in the pathogenesis of B. pseudomallei infection. PMID:24626296

Schmidt, Imke H. E.; Pudla, Matsayapan; Brakopp, Stefanie; Hopf, Verena; Breitbach, Katrin; Steinmetz, Ivo

2014-01-01

104

Autochthonous Melioidosis in Humans, Madagascar, 2012 and 2013  

PubMed Central

Melioidosis is an often fatal infectious disease affecting humans and animals in the tropics. Only sporadic cases have been reported from Africa and the Indian Ocean region. We describe 2 confirmed autochthonous cases of human melioidosis in Madagascar, both from novel genotypes of Burkholderia pseudomallei.

Djaomazala, Innocente; Dubois-Cauwelaert, Natasha; Raharimanga, Vaomalala; Ralison, Fidiarivony; Herindrainy, Perlinot; Andriamalala, Nivosoa C.; Sarovich, Derek S.; Mayo, Mark; Kaestli, Mirjam; Currie, Bart J.

2014-01-01

105

Functional Characterization and Evaluation of In Vitro Protective Efficacy of Murine Monoclonal Antibodies BURK24 and BURK37 against Burkholderia pseudomallei  

PubMed Central

Burkholderia pseudomallei, the causative agent of melioidosis has been recognized by CDC as a category B select agent. Although substantial efforts have been made for development of vaccine molecules against the pathogen, significant hurdles still remain. With no licensed vaccines available and high relapse rate of the disease, there is a pressing need for development of alternate protection strategies. Antibody-mediated passive protection is promising in this regard and our primary interest was to unravel this frontier of specific mAbs against Burkholderia pseudomallei infections, as functional characterization of antibodies is a pre-requisite to demonstrate them as protective molecules. To achieve this, we designed our study on in vitro-based approach and assessed two mAbs, namely BURK24 and BURK37, reactive with outer membrane proteins and lipopolysaccharide of the pathogen respectively, for their ability to manifest inhibitory effects on the pathogenesis mechanisms of B. pseudomallei including biofilm formation, invasion and induction of apoptosis. The experiments were performed using B. pseudomallei standard strain NCTC 10274 and a clinical isolate, B. pseudomallei 621 recovered from a septicemia patient with diabetic ailment. The growth kinetic studies of the pathogen in presence of various concentrations of each individual mAb revealed their anti-bacterial properties. Minimal inhibitory concentration and minimal bactericidal concentration of both the mAbs were determined by using standards of Clinical and Laboratory Standards Institute (CLSI) and experiments were performed using individual mAbs at their respective bacteriostatic concentration. As an outcome, both mAbs exhibited significant anti-Burkholderia pseudomallei properties. They limited the formation of biofilm by the bacterium and completely crippled its invasion into human alveolar adenocarcinoma epithelial cells. Also, the mAbs were appreciably successful in preventing the bacterium to induce apoptosis in A549 cells. The present study design revealed the protection attributes possessed by BURK24 and BURK37 that has to be further substantiated by additional in vivo studies. PMID:24614539

Peddayelachagiri, Bhavani V.; Paul, Soumya; Makam, Shivakiran S.; Urs, Radhika M.; Kingston, Joseph J.; Tuteja, Urmil; Sripathy, Murali H.; Batra, Harsh V.

2014-01-01

106

Quorum sensing regulates dpsA and the oxidative stress response in Burkholderia pseudomallei.  

PubMed

Burkholderia pseudomallei is the causative agent of melioidosis, a fatal human tropical disease. The non-specific DNA-binding protein DpsA plays a key role in protecting B. pseudomallei from oxidative stress mediated, for example, by organic hydroperoxides. The regulation of dpsA expression is poorly understood but one possibility is that it is regulated in a cell population density-dependent manner via N-acylhomoserine lactone (AHL)-dependent quorum sensing (QS) since a lux-box motif has been located within the dpsA promoter region. Using liquid chromatography and tandem mass spectrometry, it was first established that B. pseudomallei strain PP844 synthesizes AHLs. These were identified as N-octanoylhomoserine lactone (C8-HSL), N-(3-oxooctanoyl)homoserine lactone (3-oxo-C8-HSL), N-(3-hydroxyoctanoyl)-homoserine lactone (3-hydroxy-C8-HSL), N-decanoylhomoserine lactone (C10-HSL), N-(3-hydroxydecanoyl) homoserine lactone (3-hydroxy-C10-HSL) and N-(3-hydroxydodecanoyl)homoserine lactone (3-hydroxy-C12-HSL). Mutation of the genes encoding the LuxI homologue BpsI or the LuxR homologue BpsR resulted in the loss of C8-HSL and 3-oxo-C8-HSL synthesis, demonstrating that BpsI was responsible for directing the synthesis of these AHLs only and that bpsI expression and hence C8-HSL and 3-oxo-C8-HSL production depends on BpsR. In bpsI, bpsR and bpsIR mutants, dpsA expression was substantially down-regulated. Furthermore, dpsA expression in Escherichia coli required both BpsR and C8-HSL. bpsIR-deficient mutants exhibited hypersensitivity to the organic hydroperoxide tert-butyl hydroperoxide by displaying a reduction in cell viability which was restored by provision of exogenous C8-HSL (bpsI mutant only), by complementation with the bpsIR genes or by overexpression of dpsA. These data indicate that in B. pseudomallei, QS regulates the response to oxidative stress at least in part via the BpsR/C8-HSL-dependent regulation of DpsA. PMID:17159218

Lumjiaktase, Putthapoom; Diggle, Stephen P; Loprasert, Suvit; Tungpradabkul, Sumalee; Daykin, Mavis; Cámara, Miguel; Williams, Paul; Kunakorn, Mongkol

2006-12-01

107

Structural characterization of Burkholderia pseudomallei adenylate kinase (Adk): Profound asymmetry in the crystal structure of the 'open' state  

SciTech Connect

In all organisms adenylate kinases (Adks) play a vital role in cellular energy metabolism and nucleic acid synthesis. Due to differences in catalytic properties between the Adks found in prokaryotes and in the cytoplasm of eukaryotes, there is interest in targeting this enzyme for new drug therapies against infectious bacterial agents. Here we report the 2.1 {angstrom} resolution crystal structure for the 220-residue Adk from Burkholderia pseudomallei (BpAdk), the etiological agent responsible for the infectious disease melioidosis. The general structure of apo BpAdk is similar to other Adk structures, composed of a CORE subdomain with peripheral ATP-binding (ATP{sub bd}) and LID subdomains. The two molecules in the asymmetric unit have significantly different conformations, with a backbone RMSD of 1.46 {angstrom}. These two BpAdk conformations may represent 'open' Adk sub-states along the preferential pathway to the 'closed' substrate-bound state.

Buchko, G.W.; Robinson, H.; Abendroth, J.; Staker, B. L.; Myler, P. J.

2010-04-16

108

A genetic programming approach for Burkholderia Pseudomallei diagnostic pattern discovery  

PubMed Central

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

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

2009-01-01

109

Leveraging structure determination with fragment screening for infectious disease drug targets: MECP synthase from Burkholderia pseudomallei  

SciTech Connect

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.

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

110

Role of Inducible Nitric Oxide Synthase and NADPH Oxidase in Early Control of Burkholderia pseudomallei Infection in Mice  

Microsoft Academic Search

Infection with the soil bacterium Burkholderia pseudomallei can result in a variety of clinical outcomes, including asymptomatic infection. The initial immune defense mechanisms which might contribute to the various outcomes after environmental contact with B. pseudomallei are largely unknown. We have previously shown that relatively resistant C57BL\\/6 mice can restrict bacterial B. pseudomallei growth more efficiently within 1 day after

Katrin Breitbach; Sonja Klocke; Thomas Tschernig; Nico van Rooijen; Ulrich Baumann; Ivo Steinmetz

2006-01-01

111

Evaluation of the Remel RapID NF Plus Rapid Biochemical Method for Identification of Burkholderia pseudomallei  

PubMed Central

Typical methods for the identification of Burkholderia pseudomallei colonies produce results in 18 h. The Remel RapID NF Plus kit produces results in 4 h. We used the kit for 190 stored B. pseudomallei isolates and correctly identified 189 of them. This kit produces consistent results for known B. pseudomallei isolates. PMID:24671779

Engler, Cathy; Norton, Robert

2014-01-01

112

Characterization of an autotransporter adhesin protein shared by Burkholderia mallei and Burkholderia pseudomallei  

PubMed Central

Background Autotransporters form a large family of outer membrane proteins specifying diverse biological traits of Gram-negative bacteria. In this study, we report the identification and characterization of a novel autotransporter gene product of Burkholderia mallei (locus tag BMA1027 in strain ATCC 23344). Results Database searches identified the gene in at least seven B. mallei isolates and the encoded proteins were found to be 84% identical. Inactivation of the gene encoding the autotransporter in the genome of strain ATCC 23344 substantially reduces adherence to monolayers of HEp-2 laryngeal cells and A549 type II pneumocytes, as well as to cultures of normal human bronchial epithelium (NHBE). Consistent with these findings, expression of the autotransporter on the surface of recombinant E. coli bacteria increases adherence to these cell types by 5–7 fold. The gene specifying the autotransporter was identified in the genome of 29 B. pseudomallei isolates and disruption of the gene in strain DD503 reduced adherence to NHBE cultures by 61%. Unlike B. mallei, the mutation did not impair binding of B. pseudomallei to A549 or HEp-2 cells. Analysis of sera from mice infected via the aerosol route with B. mallei and B. pseudomallei revealed that animals inoculated with as few as 10 organisms produce antibodies against the autotransporter, therefore indicating expression in vivo. Conclusions Our data demonstrate that we have identified an autotransporter protein common to the pathogenic species B. mallei and B. pseudomallei which mediates adherence to respiratory epithelial cells and is expressed in vivo during the course of aerosol infection. PMID:24731253

2014-01-01

113

Melioidosis as a Consequence of Sporting Activity  

PubMed Central

In the tropical city of Darwin, Northern Territory, Australia, dry season soil sampling cultured Burkholderia pseudomallei from 7 (70%) of 10 sports fields. However, during the 23 years of the Darwin Prospective Melioidosis Study, only 5 (0.6%) of 785 melioidosis cases have been attributed to infection from sports fields. In one soccer player with cutaneous melioidosis, B. pseudomallei cultured from the player was identical by multilocus sequence typing and multilocus variable-number tandem repeat analysis with an isolate recovered from soil at the location on the sports field where he was injured. Melioidosis is uncommon in otherwise healthy sports persons in melioidosis-endemic regions but still needs consideration in persons with abrasion injuries that involve contact with soil. PMID:23732257

Hill, Audrey A.; Mayo, Mark; Kaestli, Mirjam; Price, Erin P.; Richardson, Leisha J.; Godoy, Daniel; Spratt, Brian G.; Currie, Bart J.

2013-01-01

114

Immunostimulatory CpG Oligodeoxynucleotide Confers Protection in a Murine Model of Infection with Burkholderia pseudomallei  

Microsoft Academic Search

Although CpG oligodeoxynucleotides (CpG ODNs) are known to enhance resistance against infection in a number of animal models, little is known about the CpG-induced protection against acute fatal sepsis such as that associated with the highly virulent bacterium Burkholderia pseudomallei. We previously demonstrated in an in vitro study that immunostimulatory CpG ODN 1826 enhances phagocytosis of B. pseudomallei and induces

Surasakdi Wongratanacheewin; Wannapa Kespichayawattana; Pakamas Intachote; Sathit Pichyangkul; Rasana W. Sermswan; Arthur M. Krieg; Stitaya Sirisinha

2004-01-01

115

Near-atomic resolution analysis of BipD, a component of the type III secretion system of Burkholderia pseudomallei  

PubMed Central

Burkholderia pseudomallei, the causative agent of melioidosis, possesses a type III protein secretion apparatus that is similar to those found in Salmonella and Shigella. A major function of these secretion systems is to inject virulence-associated proteins into target cells of the host organism. The bipD gene of B. pseudomallei encodes a secreted virulence factor that is similar in sequence and is most likely to be functionally analogous to IpaD from Shigella and SipD from Salmonella. Proteins in this family are thought to act as extracellular chaperones at the tip of the secretion needle to help the hydrophobic translocator proteins enter the target cell membrane, where they form a pore and may also link the translocon pore with the secretion needle. BipD has been crystallized in a monoclinic crystal form that diffracted X-rays to 1.5?Å resolution and the structure was refined to an R factor of 16.1% and an R free of 19.8% at this resolution. The putative dimer interface that was observed in previous crystal structures was retained and a larger surface area was buried in the new crystal form. PMID:20823511

Pal, M.; Erskine, P. T.; Gill, R. S.; Wood, S. P.; Cooper, J. B.

2010-01-01

116

Two fatal cases of melioidosis on the Thai-Myanmar border  

PubMed Central

Melioidosis is endemic in areas of Southeast Asia, however, there are no published reports from the Thai-Myanmar border. We report the first two documented cases of fatal melioidosis in this region. This is of great public health importance and highlights the need to both increase clinical awareness of melioidosis on the Thai-Myanmar border, and to assess the true burden of disease in the area through improved case detection and Burkholderia pseudomallei prevalence studies. PMID:24715973

Nosten, Francois

2014-01-01

117

Nasal Acai Polysaccharides Potentiate Innate Immunity to Protect against Pulmonary Francisella tularensis and Burkholderia pseudomallei Infections  

Microsoft Academic Search

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

Jerod A. Skyberg; MaryClare F. Rollins; Jeff S. Holderness; Nicole L. Marlenee; Igor A. Schepetkin; Andrew Goodyear; Steven W. Dow; Mark A. Jutila; David W. Pascual

2012-01-01

118

Comparison of the Susceptibilities of Burkholderia pseudomallei to Meropenem and Ceftazidime by Conventional and Intracellular Methods  

Microsoft Academic Search

The effect of the two antibiotics ceftazidime and meropenem on a collection of 46 Burkholderia pseudomallei isolates representing clinical and environmental sources across northern Australia was investigated by using a series of in vitro test methods. The susceptibility testing methods used included Kirby-Bauer disk diffusion, Etest MIC, broth microdilution MIC, and a modification of the microdilution method in which Acanthamoeba

T. J. J. Inglis; F. Rodrigues; P. Rigby; R. Norton; B. J. Currie

2004-01-01

119

Incidence, risk factors and clinical epidemiology of melioidosis: a complex socio-ecological emerging infectious disease in the Alor Setar region of Kedah, Malaysia  

Microsoft Academic Search

BACKGROUND: Melioidosis, a severe and fatal infectious disease caused by Burkholderia pseudomallei, is believed to an emerging global threat. However, data on the natural history, risk factors, and geographic epidemiology of the disease are still limited. METHODS: We undertook a retrospective analysis of 145 confirmed cases extracted from a hospital-based Melioidosis Registry set up from 2005 in Hospital Sultanah Bahiyah,

Muhammad RA Hassan; Subhada P Pani; Ng P Peng; Kirtanaa Voralu; Natesan Vijayalakshmi; Ranjith Mehanderkar; Norasmidar A Aziz; Edwin Michael

2010-01-01

120

Burkholderia pseudomallei kills Caenorhabditis elegans through virulence mechanisms distinct from intestinal lumen colonization  

PubMed Central

The nematode Caenorhabditis elegans is hypersusceptible to Burkholderia pseudomallei infection. However, the virulence mechanisms underlying rapid lethality of C. elegans upon B. pseudomallei infection remain poorly defined. To probe the host-pathogen interaction, we constructed GFP-tagged B. pseudomallei and followed bacterial accumulation within the C. elegans intestinal lumen. Contrary to slow-killing by most bacterial pathogens, B. pseudomallei caused fairly limited intestinal lumen colonization throughout the period of observation. Using grinder-defective mutant worms that allow the entry of intact bacteria also did not result in full intestinal lumen colonization. In addition, we observed a significant decline in C. elegans defecation and pharyngeal pumping rates upon B. pseudomallei infection. The decline in defecation rates ruled out the contribution of defecation to the limited B. pseudomallei colonization. We also demonstrated that the limited intestinal lumen colonization was not attributed to slowed host feeding as bacterial loads did not change significantly when feeding was stimulated by exogenous serotonin. Both these observations confirm that B. pseudomallei is a poor colonizer of the C. elegans intestine. To explore the possibility of toxin-mediated killing, we examined the transcription of the C. elegans ABC transporter gene, pgp-5, upon B. pseudomallei infection of the ppgp-5::gfp reporter strain. Expression of pgp-5 was highly induced, notably in the pharynx and intestine, compared with Escherichia coli-fed worms, suggesting that the host actively thwarted the pathogenic assaults during infection. Collectively, our findings propose that B. pseudomallei specifically and continuously secretes toxins to overcome C. elegans immune responses. PMID:23076282

Ooi, Soon-Keat; Lim, Tian-Yeh; Lee, Song-Hua; Nathan, Sheila

2012-01-01

121

Discrimination of Burkholderia mallei/pseudomallei from Burkholderia thailandensis by sequence comparison of a fragment of the ribosomal protein S21 (rpsU) gene  

PubMed Central

Discrimination of Burkholderia (B.) pseudomallei and B. mallei from environmental B. thailandensis is challenging. We describe a discrimination method based on sequence comparison of the ribosomal protein S21 (rpsU) gene. The rpsU gene was sequenced in ten B. pseudomallei, six B. mallei, one B. thailandensis reference strains, six isolates of B. pseudomallei, and 37 of B. thailandensis. Further rpsU sequences of six B. pseudomallei, three B. mallei, and one B. thailandensis were identified via NCBI GenBank. Three to four variable base-positions were identified within a 120-base-pair fragment, allowing discrimination of the B. pseudomallei/mallei-cluster from B. thailandensis, whose sequences clustered identically. All B. mallei and three B. pseudomallei sequences were identical, while 17/22 B. pseudomallei strains differed in one nucleotide (78A>C). Sequences of the rpsU fragment of ‘out-stander’ reference strains of B. cepacia, B. gladioli, B. plantarii, and B. vietnamensis clustered differently. Sequence comparison of the described rpsU gene fragment can be used as a supplementary diagnostic procedure for the discrimination of B. mallei/pseudomallei from B. thailandensis as well as from other species of the genus Burkholderia, keeping in mind that it does not allow for a differentiation between B. mallei and B. pseudomallei. PMID:23227305

Frickmann, H.; Chantratita, N.; Gauthier, Y. P.; Neubauer, H.; Hagen, R. M.

2012-01-01

122

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

SciTech Connect

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.

Gardner, S; Jaing, C

2012-03-27

123

Survival, Sublethal Injury, and Recovery of Environmental Burkholderia pseudomallei in Soil Subjected to Desiccation  

PubMed Central

Environmental Burkholderia pseudomallei isolated from sandy soil at Castle Hill, Townsville, in the dry tropic region of Queensland, Australia, was inoculated into sterile-soil laboratory microcosms subjected to variable soil moisture. Survival and sublethal injury of the B. pseudomallei strain were monitored by recovery using culture-based methods. Soil extraction buffer yielded higher recoveries as an extraction agent than sterile distilled water. B. pseudomallei was not recoverable when inoculated into desiccated soil but remained recoverable from moist soil subjected to 91 days' desiccation and showed a growth response to increased soil moisture over at least 113 days. Results indicate that endemic dry tropic soil may act as a reservoir during the dry season, with an increase in cell number and potential for mobilization from soil into water in the wet season. PMID:23377947

Larsen, Eloise; Smith, James J.; Norton, Robert

2013-01-01

124

The BpsIR Quorum-Sensing System of Burkholderia pseudomallei  

PubMed Central

BpsIR, a LuxIR quorum-sensing homolog, is required for optimal expression of virulence and secretion of exoproducts in Burkholderia pseudomallei. Cell density-dependent expression of bpsI and bpsR, the positive regulation of bpsIR expression by BpsR, and the synthesis of N-octanoyl-homoserine lactone (C8HSL) by BpsI are described in this report. PMID:15629951

Song, Yan; Xie, Chao; Ong, Yong-Mei; Gan, Yunn-Hwen; Chua, Kim-Lee

2005-01-01

125

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

PubMed Central

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 ?flgK 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. PMID:21720554

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

2011-01-01

126

Burkholderia pseudomallei suppresses Caenorhabditis elegans immunity by specific degradation of a GATA transcription factor.  

PubMed

Burkholderia pseudomallei is a Gram-negative soil bacterium that infects both humans and animals. Although cell culture studies have revealed significant insights into factors contributing to virulence and host defense, the interactions between this pathogen and its intact host remain to be elucidated. To gain insights into the host defense responses to B. pseudomallei infection within an intact host, we analyzed the genome-wide transcriptome of infected Caenorhabditis elegans and identified ?6% of the nematode genes that were significantly altered over a 12-h course of infection. An unexpected feature of the transcriptional response to B. pseudomallei was a progressive increase in the proportion of down-regulated genes, of which ELT-2 transcriptional targets were significantly enriched. ELT-2 is an intestinal GATA transcription factor with a conserved role in immune responses. We demonstrate that B. pseudomallei down-regulation of ELT-2 targets is associated with degradation of ELT-2 protein by the host ubiquitin-proteasome system. Degradation of ELT-2 requires the B. pseudomallei type III secretion system. Together, our studies using an intact host provide evidence for pathogen-mediated host immune suppression through the destruction of a host transcription factor. PMID:23980181

Lee, Song-Hua; Wong, Rui-Rui; Chin, Chui-Yoke; Lim, Tian-Yeh; Eng, Su-Anne; Kong, Cin; Ijap, Nur Afifah; Lau, Ming-Seong; Lim, Mei-Perng; Gan, Yunn-Hwen; He, Fang-Lian; Tan, Man-Wah; Nathan, Sheila

2013-09-10

127

Melioidosis: molecular aspects of pathogenesis.  

PubMed

Burkholderia pseudomallei is a gram-negative bacterium that causes melioidosis, a multifaceted disease that is highly endemic in southeast Asia and northern Australia. This facultative intracellular pathogen possesses a large genome that encodes a wide array of virulence factors that promote survival in vivo by manipulating host cell processes and disarming elements of the host immune system. Antigens and systems that play key roles in B. pseudomallei virulence include capsular polysaccharide, lipopolysaccharide, adhesins, specialized secretion systems, actin-based motility and various secreted factors. This review provides an overview of the current and steadily expanding knowledge regarding the molecular mechanisms used by this organism to survive within a host and their contribution to the pathogenesis of melioidosis. PMID:25312349

Stone, Joshua K; DeShazer, David; Brett, Paul J; Burtnick, Mary N

2014-12-01

128

Nasal Acai Polysaccharides Potentiate Innate Immunity to Protect against Pulmonary Francisella tularensis and Burkholderia pseudomallei Infections  

PubMed Central

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 abilities as an immunotherapeutic to treat F. tularensis and B. pseudomallei infections. In vitro, Acai PS impaired replication of Francisella in primary human macrophages co-cultured with autologous NK cells via augmentation of NK cell IFN-?. Furthermore, Acai PS administered nasally before or after infection protected mice against type A F. tularensis aerosol challenge with survival rates up to 80%, and protection was still observed, albeit reduced, when mice were treated two days post-infection. Nasal Acai PS administration augmented intracellular expression of IFN-? by NK cells in the lungs of F. tularensis-infected mice, and neutralization of IFN-? ablated the protective effect of Acai PS. Likewise, nasal Acai PS treatment conferred protection against pulmonary infection with B. pseudomallei strain 1026b. Acai PS dramatically reduced the replication of B. pseudomallei in the lung and blocked bacterial dissemination to the spleen and liver. Nasal administration of Acai PS enhanced IFN-? responses by NK and ?? T cells in the lungs, while neutralization of IFN-? totally abrogated the protective effect of Acai PS against pulmonary B. pseudomallei infection. Collectively, these results demonstrate Acai PS is a potent innate immune agonist that can resolve F. tularensis and B. pseudomallei infections, suggesting this innate immune agonist has broad-spectrum activity against virulent intracellular pathogens. PMID:22438809

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

129

Nasal Acai polysaccharides potentiate innate immunity to protect against pulmonary Francisella tularensis and Burkholderia pseudomallei Infections.  

PubMed

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 abilities as an immunotherapeutic to treat F. tularensis and B. pseudomallei infections. In vitro, Acai PS impaired replication of Francisella in primary human macrophages co-cultured with autologous NK cells via augmentation of NK cell IFN-?. Furthermore, Acai PS administered nasally before or after infection protected mice against type A F. tularensis aerosol challenge with survival rates up to 80%, and protection was still observed, albeit reduced, when mice were treated two days post-infection. Nasal Acai PS administration augmented intracellular expression of IFN-? by NK cells in the lungs of F. tularensis-infected mice, and neutralization of IFN-? ablated the protective effect of Acai PS. Likewise, nasal Acai PS treatment conferred protection against pulmonary infection with B. pseudomallei strain 1026b. Acai PS dramatically reduced the replication of B. pseudomallei in the lung and blocked bacterial dissemination to the spleen and liver. Nasal administration of Acai PS enhanced IFN-? responses by NK and ?? T cells in the lungs, while neutralization of IFN-? totally abrogated the protective effect of Acai PS against pulmonary B. pseudomallei infection. Collectively, these results demonstrate Acai PS is a potent innate immune agonist that can resolve F. tularensis and B. pseudomallei infections, suggesting this innate immune agonist has broad-spectrum activity against virulent intracellular pathogens. PMID:22438809

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

130

Type three secretion system-mediated escape of Burkholderia pseudomallei into the host cytosol is critical for the activation of NF?B  

PubMed Central

Background Burkholderia pseudomallei is the causative agent of melioidosis, a potentially fatal disease endemic in Southeast Asia and Northern Australia. This Gram-negative pathogen possesses numerous virulence factors including three “injection type” type three secretion systems (T3SSs). B. pseudomallei has been shown to activate NF?B in HEK293T cells in a Toll-like receptor and MyD88 independent manner that requires T3SS gene cluster 3 (T3SS3 or T3SSBsa). However, the mechanism of how T3SS3 contributes to NF?B activation is unknown. Results Known T3SS3 effectors are not responsible for NF?B activation. Furthermore, T3SS3-null mutants are able to activate NF?B almost to the same extent as wildtype bacteria at late time points of infection, corresponding to delayed escape into the cytosol. NF?B activation also occurs when bacteria are delivered directly into the cytosol by photothermal nanoblade injection. Conclusions T3SS3 does not directly activate NF?B but facilitates bacterial escape into the cytosol where the host is able to sense the presence of the pathogen through cytosolic sensors leading to NF?B activation. PMID:24884837

2014-01-01

131

Burkholderia pseudomallei activates complement and is ingested but not killed by polymorphonuclear leukocytes.  

PubMed Central

The mechanism by which Burkholderia pseudomallei is resistant to lysis by human serum is unknown but may include interference with complement activation, effective opsonization, or complement-mediated lysis. We investigated the interaction of B. pseudomallei with complement in the presence and absence of specific antibody to determine potential mechanisms of serum resistance. We demonstrated rapid activation and consumption of complement by B. pseudomallei which, in the absence of specific antibody, occurred predominantly via the alternative pathway. Complement activation was associated with deposition of the opsonically active C3b and iC3b fragments on the bacterial surface. C5b-9, detected on the bacterial surface after opsonic periods of 1 to 60 min, was susceptible to elution by 1 M NaCl, indicating that resistance to complement-mediated lysis may result from deposition of the membrane attack complex in a nonmicrobicidal location. To define the role of opsonins, we investigated the ability of polymorphonuclear leukocytes (PMNL) to phagocytose B. pseudomallei. Phagocytosis of bacteria by PMNL, and the observed oxidative response, was significantly increased by opsonization of organisms with complement and/or specific antibody. Despite opsonophagocytosis by PMNL and the production of an oxidative response, no significant bacterial killing was observed. PMID:8945532

Egan, A M; Gordon, D L

1996-01-01

132

Isolation and characterization of the outer-membrane proteins of Burkholderia (Pseudomonas) pseudomallei.  

PubMed

Membranes obtained from whole-cell lysates of Burkholderia (Pseudomonas) pseudomallei (strain 319a) were separated into four fractions by sucrose density gradient centrifugation. Membranes were characterized by enzymic and chemical analyses, and by SDS-PAGE. Cytoplasmic membranes and two forms of outer membranes (OM-1, OM-2) were detected. The major outer-membrane proteins had M(r) values of 70,000, 38,000, 31,000, 24,000 and 17,000. To determine which outer-membrane proteins were common to B. pseudomallei strains, OM-1 fractions from 12 different strains were prepared. SDS-PAGE analysis of these fractions demonstrated that the five major outer-membrane proteins were common to the strains tested. Further studies have shown that an M(r) 110,000 protein, which is oligomeric in that it migrates as an M(r) 38,000 protein upon heating at 95 degrees C and which is peptidoglycan-associated, serves as a porin in B. pseudomallei. Using proteoliposomes reconstituted from this protein and phospholipid, it was demonstrated by the liposome-swelling assay that this protein acts as a porin through which small saccharides may diffuse. Further characterization of this M(r) 38,000 protein will be important in delineating the role of this molecule in the permeability of the B. pseudomallei outer membrane. PMID:7516793

Gotoh, N; White, N J; Chaowagul, W; Woods, D E

1994-04-01

133

Expression and refolding of Omp38 from Burkholderia pseudomallei and Burkholderia thailandensis, and its function as a diffusion porin  

PubMed Central

In the present paper, we describe cloning and expression of two outer membrane proteins, BpsOmp38 (from Burkholderia pseudomallei) and BthOmp38 (from Burkholderia thailandensis) lacking signal peptide sequences, using the pET23d(+) expression vector and Escherichia coli host strain Origami(DE3). The 38 kDa proteins, expressed as insoluble inclusion bodies, were purified, solubilized in 8 M urea, and then subjected to refolding experiments. As seen on SDS/PAGE, the 38 kDa band completely migrated to ?110 kDa when the purified monomeric proteins were refolded in a buffer system containing 10% (w/v) Zwittergent® 3-14, together with a subsequent heating to 95 °C for 5 min. CD spectroscopy revealed that the 110 kDa proteins contained a predominant ?-sheet structure, which corresponded completely to the structure of the Omp38 proteins isolated from B. pseudomallei and B. thailandensis. Immunoblot analysis using anti-BpsOmp38 polyclonal antibodies and peptide mass analysis by MALDI–TOF (matrix-assisted laser-desorption ionization–time-of-flight) MS confirmed that the expressed proteins were BpsOmp38 and BthOmp38. The anti-BpsOmp38 antibodies considerably exhibited the inhibitory effects on the permeation of small sugars through the Omp38-reconstituted liposomes. A linear relation between relative permeability rates and Mr of neutral sugars and charged antibiotics suggested strongly that the in vitro re-assembled Omp38 functioned fully as a diffusion porin. PMID:15329048

2004-01-01

134

Rapid Burkholderia pseudomallei identification and antibiotic resistance determination by bacteriophage amplification and MALDI-TOF MS  

PubMed Central

Phage amplification detected by MALDI-TOF MS was investigated for rapid and simultaneous Burkholderia pseudomallei identification and ceftazidime resistance determination. B. pseudomallei ceftazidime susceptible and resistant ?purM mutant strains Bp82 and Bp82.3 were infected with broadly targeting B. pseudomallei phage ?X216 and production of the m/z 37.6 kDa phage capsid protein observed by MALDI-TOF MS over the course of 3 h infections. This allowed for repoducible phage-based bacterial ID within 2 h of the onset of infection. MALDI-TOF MS-measured time to detection correlated with in silico modeling, which predicted an approximate 2 h detection time. Ceftazidime susceptible strain Bp82, while detectable in the absence of the drug, owing to the reliance of phage amplification on a viable host, was not detectable when 10 ?g/mL ceftazidime was added at the onset of infection. In contrast, resistant strain Bp82.3 was detected in the same 2 h timeframe both with and without the addition of ceftazidime. PMID:25050191

Cox, Christopher R; Saichek, Nicholas R; Schweizer, Herbert P; Voorhees, Kent J

2014-01-01

135

The role of NOD2 in murine and human melioidosis  

PubMed Central

NOD2 is a cytosolic pathogen recognition receptor that regulates susceptibility to a variety of infections and chronic diseases. Burkholderia pseudomallei, a facultative intracellular bacterium, causes the tropical infection melioidosis. We hypothesized that NOD2 may participate in host defense in melioidosis. We performed a series of in vitro assays and in vivo experiments, and analyzed the association of human genetic variation with infection to delineate the contribution of NOD2 to the host response to B. pseudomallei. We found that transfection with NOD2 mediated NF-?B activation induced by B. pseudomallei stimulation of HEK293 cells. After low dose inoculation with aerosolized B. pseudomallei, Nod2-deficient mice showed impaired clinical responses and permitted greater bacterial replication in the lung and dissemination to the spleen compared to wild type mice. IL-6 and KC levels were higher in the lungs of Nod2-deficient mice. In a cohort of 1,562 Thai subjects, a common genetic polymorphism in the NOD2 region, rs7194886, was associated with melioidosis and this effect was most pronounced in women. rs7194886 was not associated with differences in cytokine production induced by whole blood stimulation with the NOD2 ligand, MDP, or B. pseudomallei. These findings are the first to characterize the role of NOD2 in host defense in mammalian melioidosis. PMID:24298015

Myers, Nicolle D.; Chantratita, Narisara; Berrington, William R.; Chierakul, Wirongrong; Limmathurotsakul, Direk; Wuthiekanun, Vanaporn; Robertson, Johanna D.; Liggitt, H. Denny; Peacock, Sharon J.; Skerrett, Shawn J.; West, T. Eoin

2013-01-01

136

Cellular Reporter Screens for Inhibitors of Burkholderia pseudomallei Targets in Pseudomonas aeruginosa  

PubMed Central

Summary To facilitate the discovery of new therapeutics for Burkholderia pseudomallei infections, we have developed cellular reporter screens for inhibitors of B. pseudomallei targets in the surrogate host Pseudomonas aeruginosa. P. aeruginosa strains carrying deletions of essential genes were engineered to be dependent on the IPTG-regulated expression of their B. pseudomallei orthologs on a broad-host-range plasmid. P. aeruginosa genes which are upregulated in response to depletion of each target gene product were fused to the Photorhabdus luminescens luxCDABE operon via pGSV3-lux-SpR to generate reporter strains with increased bioluminescence upon target inhibition. A total of 11 of 19 B. pseudomallei genes complemented deletions of their orthologs in P. aeruginosa. The dependence of growth on IPTG levels varied from complete dependence (ftsQ, gyrA, glmU, secA), to slower growth in the absence of IPTG (coaD, efp, mesJ), to apparently normal growth in the absence of IPTG (ligA, lpxA, folA, ipk). Reporter screening strains have been constructed for three gene targets (gyrA, glmU, secA), and one (gyrA) has been applied to 68,000 compounds resulting in a primary hit rate of 0.5% and a confirmed hit rate of 0.06% including several fluoroquinolones. These results provide proof of principle for surrogate cellular reporter screens as a useful approach to identify inhibitors of essential gene products. PMID:19121678

Moir, D. T.; Di, M.; Moore, R. A.; Schweizer, H. P.; Woods, D. E.

2009-01-01

137

Delineating the Importance of Serum Opsonins and the Bacterial Capsule in Affecting the Uptake and Killing of Burkholderia pseudomallei by Murine Neutrophils and Macrophages  

PubMed Central

Infection of susceptible hosts by the encapsulated Gram-negative bacterium Burkholderia pseudomallei (Bp) causes melioidosis, with septic patients attaining mortality rates ?40%. Due to its high infectivity through inhalation and limited effective therapies, Bp is considered a potential bioweapon. Thus, there is great interest in identifying immune effectors that effectively kill Bp. Our goal is to compare the relative abilities of murine macrophages and neutrophils to clear Bp, as well as determine the importance of serum opsonins and bacterial capsule. Our findings indicate that murine macrophages and neutrophils are inherently unable to clear either unopsonized Bp or the relatively-avirulent acapsular bacterium B. thailandensis (Bt). Opsonization of Bp and Bt with complement or pathogen-specific antibodies increases macrophage-uptake, but does not promote clearance, although antibody-binding enhances complement deposition. In contrast, complement opsonization of Bp and Bt causes enhanced uptake and killing by neutrophils, which is linked with rapid ROS induction against bacteria exhibiting a threshold level of complement deposition. Addition of bacteria-specific antibodies enhances complement deposition, but antibody-binding alone cannot elicit neutrophil clearance. Bp capsule provides some resistance to complement deposition, but is not anti-phagocytic or protective against reactive oxygen species (ROS)-killing. Macrophages were observed to efficiently clear Bp only after pre-activation with IFN?, which is independent of serum- and/or antibody-opsonization. These studies indicate that antibody-enhanced complement activation is sufficient for neutrophil-clearance of Bp, whereas macrophages are ineffective at clearing serum-opsonized Bp unless pre-activated with IFN?. This suggests that effective immune therapies would need to elicit both antibodies and Th1-adaptive responses for successful prevention/eradication of melioidosis. PMID:25144195

Mulye, Minal; Bechill, Michael P.; Grose, William; Ferreira, Viviana P.; Lafontaine, Eric R.; Wooten, R. Mark

2014-01-01

138

DpsA protects the human pathogen Burkholderia pseudomallei against organic hydroperoxide.  

PubMed

The human pathogen, Burkholderia pseudomalle, is able to survive and multiply in hostile environments such as within macrophages. In an attempt to understand its strategy to cope with oxidative stress, the physiological role and gene regulation of a nonspecific DNA-binding protein (DpsA) was investigated. Expression of dpsA increases in response to oxidative stress through increased transcription from the upstream katG (catalase-peroxidase) promoter, which is OxyR dependent. dpsA is also transcribed from its own promoter, which is activated by osmotic stress in an OxyR-independent manner. DpsA-deficient mutants are hypersensitive to tert-butyl hydroperoxide, while overexpression of DpsA leads to increased resistance to organic oxidants. B. pseudomallei DpsA can also protect Escherichia coli against organic hydroperoxide toxicity. The mechanism of DpsA-mediated resistance to organic hydroperoxides was shown to differ from that of alkyl hydroperoxide reductase. PMID:15241582

Loprasert, Suvit; Whangsuk, Wirongrong; Sallabhan, Ratiboot; Mongkolsuk, Skorn

2004-09-01

139

Reactivation of latent melioidosis presenting with acute pyelonephritis and bacteremia.  

PubMed

We report an interesting case involving an 81-year-old male patient who presented to our hospital with sepsis secondary to pyelonephritis and bacteremia. Blood cultures were positive for a Gram-negative bacillus, Burkholderia pseudomallei. His risk factors were diabetes, chronic obstructive pulmonary disease including travel to an endemic area. The patient received antibiotic therapy in accordance with the national guidelines. This is the first report of reactivation of latent melioidosis in the literature manifesting as pyelonephritis and bacteremia. PMID:24678468

Shaaban, Hamid; Hallit, Rabih; Slim, Jihad; Sree, Aruna; Sensakovic, John W

2014-01-01

140

Reactivation of latent melioidosis presenting with acute pyelonephritis and bacteremia  

PubMed Central

We report an interesting case involving an 81-year-old male patient who presented to our hospital with sepsis secondary to pyelonephritis and bacteremia. Blood cultures were positive for a Gram-negative bacillus, Burkholderia pseudomallei. His risk factors were diabetes, chronic obstructive pulmonary disease including travel to an endemic area. The patient received antibiotic therapy in accordance with the national guidelines. This is the first report of reactivation of latent melioidosis in the literature manifesting as pyelonephritis and bacteremia. PMID:24678468

Shaaban, Hamid; Hallit, Rabih; Slim, Jihad; Sree, Aruna; Sensakovic, John W.

2014-01-01

141

Burkholderia pseudomallei RpoS regulates OxyR and the katG-dpsA operon under conditions of oxidative stress.  

PubMed

Burkholderia pseudomallei, the causative agent of the potentially fatal tropical disease melioidosis, is known to be highly resistant to oxidative stress although the mechanism of this resistance remains to be fully elucidated. Previous studies have shown that an OxyR is involved in the regulation of oxidative stress via the katG and dpsA genes encoding KatG and DpsA and that the alternative sigma factor, RpoS, plays a critical role in resistance to oxidative stress by regulating katG and katE genes. Here it is shown that RpoS is essential for expression of the oxidative stress regulator OxyR, since a mutant strain lacking RpoS failed to induce oxyR expression both during normal growth and under conditions of oxidative stress. It is further demonstrated that the RpoS acts as a positive transcriptional regulator of oxyR and dpsA expression, while OxyR acts as a negative transcriptional regulator of the katG-dpsA operon via OxyR repressor under normal growth conditions, and as a positive transcriptional regulator via OxyR under conditions of oxidative stress. Therefore both RpoS and OxyR are required to promote expression of both the katG-dpsA operon and dpsA gene. PMID:20618685

Jangiam, Witawat; Loprasert, Suvit; Smith, Duncan R; Tungpradabkul, Sumalee

2010-07-01

142

The HicA toxin from Burkholderia pseudomallei has a role in persister cell formation  

PubMed Central

TA (toxin–antitoxin) systems are widely distributed amongst bacteria and are associated with the formation of antibiotic tolerant (persister) cells that may have involvement in chronic and recurrent disease. We show that overexpression of the Burkholderia pseudomallei HicA toxin causes growth arrest and increases the number of persister cells tolerant to ciprofloxacin or ceftazidime. Furthermore, our data show that persistence towards ciprofloxacin or ceftazidime can be differentially modulated depending on the level of induction of HicA expression. Deleting the hicAB locus from B. pseudomallei K96243 significantly reduced persister cell frequencies following exposure to ciprofloxacin, but not ceftazidime. The structure of HicA(H24A) was solved by NMR and forms a dsRBD-like (dsRNA-binding domain-like) fold, composed of a triple-stranded ?-sheet, with two helices packed against one face. The surface of the protein is highly positively charged indicative of an RNA-binding protein and His24 and Gly22 were functionality important residues. This is the first study demonstrating a role for the HicAB system in bacterial persistence and the first structure of a HicA protein that has been experimentally characterized. PMID:24502667

Butt, Aaron; Higman, Victoria A.; Williams, Christopher; Crump, Matthew P.; Hemsley, Claudia M.; Harmer, Nicholas; Titball, Richard W.

2014-01-01

143

Development of a Prototype Lateral Flow Immunoassay (LFI) for the Rapid Diagnosis of Melioidosis  

PubMed Central

Burkholderia pseudomallei is a soil-dwelling bacterium and the causative agent of melioidosis. Isolation of B. pseudomallei from clinical samples is the “gold standard” for the diagnosis of melioidosis; results can take 3–7 days to produce. Alternatively, antibody-based tests have low specificity due to a high percentage of seropositive individuals in endemic areas. There is a clear need to develop a rapid point-of-care antigen detection assay for the diagnosis of melioidosis. Previously, we employed In vivo Microbial Antigen Discovery (InMAD) to identify potential B. pseudomallei diagnostic biomarkers. The B. pseudomallei capsular polysaccharide (CPS) and numerous protein antigens were identified as potential candidates. Here, we describe the development of a diagnostic immunoassay based on the detection of CPS. Following production of a CPS-specific monoclonal antibody (mAb), an antigen-capture immunoassay was developed to determine the concentration of CPS within a panel of melioidosis patient serum and urine samples. The same mAb was used to produce a prototype Active Melioidosis Detect Lateral Flow Immunoassay (AMD LFI); the limit of detection of the LFI for CPS is comparable to the antigen-capture immunoassay (?0.2 ng/ml). The analytical reactivity (inclusivity) of the AMD LFI was 98.7% (76/77) when tested against a large panel of B. pseudomallei isolates. Analytical specificity (cross-reactivity) testing determined that 97.2% of B. pseudomallei near neighbor species (35/36) were not reactive. The non-reactive B. pseudomallei strain and the reactive near neighbor strain can be explained through genetic sequence analysis. Importantly, we show the AMD LFI is capable of detecting CPS in a variety of patient samples. The LFI is currently being evaluated in Thailand and Australia; the focus is to optimize and validate testing procedures on melioidosis patient samples prior to initiation of a large, multisite pre-clinical evaluation. PMID:24651568

Houghton, Raymond L.; Reed, Dana E.; Hubbard, Mark A.; Dillon, Michael J.; Chen, Hongjing; Currie, Bart J.; Mayo, Mark; Sarovich, Derek S.; Theobald, Vanessa; Limmathurotsakul, Direk; Wongsuvan, Gumphol; Chantratita, Narisara; Peacock, Sharon J.; Hoffmaster, Alex R.; Duval, Brea; Brett, Paul J.; Burtnick, Mary N.; AuCoin, David P.

2014-01-01

144

Urokinase receptor is necessary for bacterial defense against pneumonia-derived septic melioidosis by facilitating phagocytosis.  

PubMed

Urokinase receptor (urokinase-type plasminogen activator receptor [uPAR], CD87), a GPI-anchored protein, is considered to play an important role in inflammation and fibrinolysis. The Gram-negative bacterium Burkholderia pseudomallei is able to survive and replicate within leukocytes and causes melioidosis, an important cause of pneumonia-derived community-acquired sepsis in Southeast Asia. In this study, we investigated the expression and function of uPAR both in patients with septic melioidosis and in a murine model of experimental melioidosis. uPAR mRNA and surface expression was increased in patients with septic melioidosis in/on both peripheral blood monocytes and granulocytes as well as in the pulmonary compartment during experimental pneumonia-derived melioidosis in mice. uPAR-deficient mice intranasally infected with B. pseudomallei showed an enhanced growth and dissemination of B. pseudomallei when compared with wild-type mice, corresponding with increased pulmonary and hepatic inflammation. uPAR knockout mice demonstrated significantly reduced neutrophil migration toward the pulmonary compartment after inoculation with B. pseudomallei. Further in vitro experiments showed that uPAR-deficient macrophages and granulocytes display a markedly impaired phagocytosis of B. pseudomallei. Additional studies showed that uPAR deficiency did not influence hemostatic and fibrinolytic responses during severe melioidosis. These data suggest that uPAR is crucially involved in the host defense against sepsis caused by B. pseudomallei by facilitating the migration of neutrophils toward the primary site of infection and subsequently facilitating the phagocytosis of B. pseudomallei. PMID:20142364

Wiersinga, W Joost; Kager, Liesbeth M; Hovius, Joppe W R; van der Windt, Gerritje J W; de Vos, Alex F; Meijers, Joost C M; Roelofs, Joris J; Dondorp, Arjen; Levi, Marcel; Day, Nicholas P; Peacock, Sharon J; van der Poll, Tom

2010-03-15

145

The aftermath of the Western Australian melioidosis outbreak.  

PubMed

Melioidosis became a notifiable disease in Western Australia (WA) 2 years after the West Kimberley melioidosis outbreak. Two cases of melioidosis caused by the outbreak genotype of Burkholderia pseudomallei (National Collection of Type Cultures [NCTC] 13177) occurred in 1998 and 1999 in persons who visited the outbreak location at the time. No other infections caused by the outbreak strain have been recorded in WA since that time, despite an average of four culture-positive cases per year. Sporadic cases of melioidosis often follow tropical storms and cyclones during summer, and they have been detected outside the endemic area when cyclones travel far inland. In 2007, environmental isolates resembling NCTC 13177 were found 500 km east of the outbreak location after unusually severe weather. Recent whole-genome analysis places NCTC 13177 genetically close to other Australian isolates. Additional biogeographic and ecological studies are needed to establish the relative importance of environmental cofactors in disease pathogenesis. PMID:21633018

Inglis, Timothy J J; O'Reilly, Lyn; Merritt, Adam J; Levy, Avram; Heath, Christopher H; Heath, Christopher

2011-06-01

146

The Aftermath of the Western Australian Melioidosis Outbreak  

PubMed Central

Melioidosis became a notifiable disease in Western Australia (WA) 2 years after the West Kimberley melioidosis outbreak. Two cases of melioidosis caused by the outbreak genotype of Burkholderia pseudomallei (National Collection of Type Cultures [NCTC] 13177) occurred in 1998 and 1999 in persons who visited the outbreak location at the time. No other infections caused by the outbreak strain have been recorded in WA since that time, despite an average of four culture-positive cases per year. Sporadic cases of melioidosis often follow tropical storms and cyclones during summer, and they have been detected outside the endemic area when cyclones travel far inland. In 2007, environmental isolates resembling NCTC 13177 were found 500 km east of the outbreak location after unusually severe weather. Recent whole-genome analysis places NCTC 13177 genetically close to other Australian isolates. Additional biogeographic and ecological studies are needed to establish the relative importance of environmental cofactors in disease pathogenesis. PMID:21633018

Inglis, Timothy J. J.; O'Reilly, Lyn; Merritt, Adam J.; Levy, Avram; Heath, Christopher

2011-01-01

147

Burkholderia vaccines: are we moving forward?  

PubMed Central

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

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

148

Burkholderia vaccines: are we moving forward?  

PubMed

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

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

149

Multiple-antigen ELISA for melioidosis - a novel approach to the improved serodiagnosis of melioidosis  

PubMed Central

Background Burkholderia pseudomallei, the causative agent of melioidosis, is endemic to Southeast Asia and northern Australia. Clinical manifestations of disease are diverse, ranging from chronic infection to acute septicaemia. The current gold standard of diagnosis involves bacterial culture and identification which is time consuming and often too late for early medical intervention. Hence, rapid diagnosis of melioidosis is crucial for the successful management of melioidosis. Methods The study evaluated 4 purified B. pseudomallei recombinant proteins (TssD-5, Omp3, smBpF4 and Omp85) as potential diagnostic agents for melioidosis. A total of 68 sera samples from Malaysian melioidosis patients were screened for the presence of specific antibodies towards these proteins using enzyme-linked immunosorbent assay (ELISA). Sera from patients with various bacterial and viral infections but negative for B. pseudomallei, as well as sera from healthy individuals, were also included as non-melioidosis controls. The Mann Whitney test was performed to compare the statistical differences between melioidosis patients and the non-melioidosis controls. Results TssD-5 demonstrated the highest sensitivity of 71% followed by Omp3 (59%), smBpF4 (41%) and Omp85 (19%). All 4 antigens showed equally high specificity (89-96%). A cocktail of all 4 antigens resulted in slightly reduced sensitivity of 65% but improved specificity (99%). Multiple-antigen ELISA provided improved sensitivity of 88.2% whilst retaining good specificity (96%). There was minimal reactivity with sera from healthy individuals proposing the utility of these antigens to demarcate diseased from non-symptomatic individuals in an endemic country. Conclusions TssD-5 demonstrated high detection sensitivity and specificity and the results were obtained within a few hours compared to time consuming culture and IFAT methods commonly used in a clinical setting. The use of multiple-antigens resulted in improved sensitivity (88.2%) whilst maintaining superior specificity. These data highlight the use of TssD-5 and other recombinant antigens tested in this study as potential serodiagnostic agents for melioidosis. PMID:23556548

2013-01-01

150

Natural Infection of Burkholderia pseudomallei in an Imported Pigtail Macaque (Macaca nemestrina) and Management of the Exposed Colony  

PubMed Central

Identification of the select agent Burkholderia pseudomallei in macaques imported into the United States is rare. A purpose-bred, 4.5-y-old pigtail macaque (Macaca nemestrina) imported from Southeast Asia was received from a commercial vendor at our facility in March 2012. After the initial acclimation period of 5 to 7 d, physical examination of the macaque revealed a subcutaneous abscess that surrounded the right stifle joint. The wound was treated and resolved over 3 mo. In August 2012, 2 mo after the stifle joint wound resolved, the macaque exhibited neurologic clinical signs. Postmortem microbiologic analysis revealed that the macaque was infected with B. pseudomallei. This case report describes the clinical evaluation of a B. pseudomallei-infected macaque, management and care of the potentially exposed colony of animals, and protocols established for the animal care staff that worked with the infected macaque and potentially exposed colony. This article also provides relevant information on addressing matters related to regulatory issues and risk management of potentially exposed animals and animal care staff. PMID:24326230

Johnson, Crystal H; Skinner, Brianna L; Dietz, Sharon M; Blaney, David; Engel, Robyn M; Lathrop, George W; Hoffmaster, Alex R; Gee, Jay E; Elrod, Mindy G; Powell, Nathaniel; Walke, Henry

2013-01-01

151

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.  

PubMed

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. PMID:25044501

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

2014-10-01

152

The coagulation system in melioidosis: from pathogenesis to new treatment strategies.  

PubMed

Melioidosis, caused by the Gram-negative bacterium Burkholderia pseudomallei, is a dreadful disease common in South-East Asia and Northern Australia and is characterized by chronic suppurative lesions and pneumonia. Melioidosis may evolve into severe sepsis with multi-organ failure with high mortalities, despite proper antibiotic therapy. Besides activation of a strong pro-inflammatory host response, the coagulation system plays an important role during melioidosis, which is thought to be host-protective. In particular, a procoagulant state together with downregulation of anticoagulant pathways and activation of fibrinolysis are present, all closely interrelated with parameters of inflammation. This review presents an overview of recent studies in which the role of coagulation, anti-coagulation and fibrinolysis during melioidosis was investigated both in patients and in experimental settings. PMID:24962103

Kager, Liesbeth Martine; van der Poll, Tom; Wiersinga, Willem Joost

2014-08-01

153

Evaluation of a New Commercially Available Immunoglobulin M and Immunoglobulin G Immunochromatographic Test for Diagnosis of Melioidosis Infection  

Microsoft Academic Search

An immunochromatographic test for the rapid determination of immunoglobulin M (IgM) and IgG anti- bodies to Burkholderia pseudomallei was evaluated by using sera from bacteriologically confirmed melioidosis patients and high-risk and clinically suspected patients, along with disease control groups. The sensitivities were 100 and 93% for the IgG and IgM tests, respectively, while the specificity was 95% for both assays.

ANDREA J. CUZZUBBO; V. CHENTHAMARAKSHAN; JAMUNA VADIVELU; SAVITHIRI D. PUTHUCHEARY; DENISE ROWLAND; PETER L. DEVINE

2000-01-01

154

Deficiency of protease-activated receptor-1 limits bacterial dissemination during severe Gram-negative sepsis (melioidosis).  

PubMed

Protease-activated receptor-1 (PAR-1) is a G-coupled transmembrane receptor expressed by multiple cell types present in the lungs that can be activated by various proteases generated during acute inflammation. In this study we aimed to investigate the role of PAR-1 during melioidosis, a common cause of (pneumo)sepsis in Southeast Asia in a murine model of intranasal inoculation of the causative pathogen Burkholderia (B.) pseudomallei. Our results show that endogenous PAR-1 facilitates bacterial growth and dissemination during murine melioidosis, which is associated with increased cell influxes. However, these observations have no impact on survival. PMID:24239704

Kager, Liesbeth M; Wiersinga, W Joost; Roelofs, Joris J T H; van 't Veer, Cornelis; van der Poll, Tom

2014-02-01

155

Melioidosis: Epidemiology, Pathophysiology, and Management  

PubMed Central

Melioidosis, caused by the gram-negative saprophyte Burkholderia pseudomallei, is a disease of public health importance in southeast Asia and northern Australia that is associated with high case-fatality rates in animals and humans. It has the potential for epidemic spread to areas where it is not endemic, and sporadic case reports elsewhere in the world suggest that as-yet-unrecognized foci of infection may exist. Environmental determinants of this infection, apart from a close association with rainfall, are yet to be elucidated. The sequencing of the genome of a strain of B. pseudomallei has recently been completed and will help in the further identification of virulence factors. The presence of specific risk factors for infection, such as diabetes, suggests that functional neutrophil defects are important in the pathogenesis of melioidosis; other studies have defined virulence factors (including a type III secretion system) that allow evasion of killing mechanisms by phagocytes. There is a possible role for cell-mediated immunity, but repeated environmental exposure does not elicit protective humoral or cellular immunity. A vaccine is under development, but economic constraints may make vaccination an unrealistic option for many regions of endemicity. Disease manifestations are protean, and no inexpensive, practical, and accurate rapid diagnostic tests are commercially available; diagnosis relies on culture of the organism. Despite the introduction of ceftazidime- and carbapenem-based intravenous treatments, melioidosis is still associated with a significant mortality attributable to severe sepsis and its complications. A long course of oral eradication therapy is required to prevent relapse. Studies exploring the role of preventative measures, earlier clinical identification, and better management of severe sepsis are required to reduce the burden of this disease. PMID:15831829

Cheng, Allen C.; Currie, Bart J.

2005-01-01

156

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

NASA Astrophysics Data System (ADS)

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.

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

2010-04-01

157

Glyburide Reduces Bacterial Dissemination in a Mouse Model of Melioidosis  

PubMed Central

Background Burkholderia pseudomallei infection (melioidosis) is an important cause of community-acquired Gram-negative sepsis in Northeast Thailand, where it is associated with a ?40% mortality rate despite antimicrobial chemotherapy. We showed in a previous cohort study that patients taking glyburide (?=?glibenclamide) prior to admission have lower mortality and attenuated inflammatory responses compared to patients not taking glyburide. We sought to define the mechanism underlying this observation in a murine model of melioidosis. Methods Mice (C57BL/6) with streptozocin-induced diabetes were inoculated with ?6×102 cfu B. pseudomallei intranasally, then treated with therapeutic ceftazidime (600 mg/kg intraperitoneally twice daily starting 24 h after inoculation) in order to mimic the clinical scenario. Glyburide (50 mg/kg) or vehicle was started 7 d before inoculation and continued until sacrifice. The minimum inhibitory concentration of glyburide for B. pseudomallei was determined by broth microdilution. We also examined the effect of glyburide on interleukin (IL) 1? by bone-marrow-derived macrophages (BMDM). Results Diabetic mice had increased susceptibility to melioidosis, with increased bacterial dissemination but no effect was seen of diabetes on inflammation compared to non-diabetic controls. Glyburide treatment did not affect glucose levels but was associated with reduced pulmonary cellular influx, reduced bacterial dissemination to both liver and spleen and reduced IL1? production when compared to untreated controls. Other cytokines were not different in glyburide-treated animals. There was no direct effect of glyburide on B. pseudomallei growth in vitro or in vivo. Glyburide directly reduced the secretion of IL1? by BMDMs in a dose-dependent fashion. Conclusions Diabetes increases the susceptibility to melioidosis. We further show, for the first time in any model of sepsis, that glyburide acts as an anti-inflammatory agent by reducing IL1? secretion accompanied by diminished cellular influx and reduced bacterial dissemination to distant organs. We found no evidence for a direct effect of glyburide on the bacterium. PMID:24147174

Koh, Gavin C. K. W.; Weehuizen, Tassili A.; Breitbach, Katrin; Krause, Kathrin; de Jong, Hanna K.; Kager, Liesbeth M.; Hoogendijk, Arjan J.; Bast, Antje; Peacock, Sharon J.; van der Poll, Tom; Steinmetz, Ivo; Wiersinga, W. Joost

2013-01-01

158

In Vitro Activities of Amoxicillin-Clavulanate, Doxycycline, Ceftazidime, Imipenem, and Trimethoprim-Sulfamethoxazole against Biofilm of Brazilian Strains of Burkholderia pseudomallei  

PubMed Central

This study aimed at investigating the in vitro activities of amoxicillin-clavulanate, doxycycline, ceftazidime, imipenem, and trimethoprim-sulfamethoxazole against Burkholderia pseudomallei in planktonic and biofilm forms, through broth microdilution and resazurin-based viability staining, respectively. In planktonic growth, the strains were susceptible to the drugs, while in biofilm growth, significantly higher antimicrobial concentrations were required, especially for ceftazidime and imipenem, surpassing the resistance breakpoints. These results highlight the importance of the routine evaluation of biofilm antimicrobial susceptibility. PMID:24002089

Bandeira, Tereza de Jesus Pinheiro Gomes; Moreira, Camila Alencar; Castelo-Branco, Debora de Souza Collares Maia; Neto, Manoel Paiva de Araujo; Cordeiro, Rossana de Aguiar; Rodrigues, Terezinha de Jesus Santos; Rocha, Marcos Fabio Gadelha; Sidrim, Jose Julio Costa

2013-01-01

159

In vitro activities of amoxicillin-clavulanate, doxycycline, ceftazidime, imipenem, and trimethoprim-sulfamethoxazole against biofilm of Brazilian strains of Burkholderia pseudomallei.  

PubMed

This study aimed at investigating the in vitro activities of amoxicillin-clavulanate, doxycycline, ceftazidime, imipenem, and trimethoprim-sulfamethoxazole against Burkholderia pseudomallei in planktonic and biofilm forms, through broth microdilution and resazurin-based viability staining, respectively. In planktonic growth, the strains were susceptible to the drugs, while in biofilm growth, significantly higher antimicrobial concentrations were required, especially for ceftazidime and imipenem, surpassing the resistance breakpoints. These results highlight the importance of the routine evaluation of biofilm antimicrobial susceptibility. PMID:24002089

Bandeira, Tereza de Jesus Pinheiro Gomes; Moreira, Camila Alencar; Brilhante, Raimunda Sâmia Nogueira; Castelo-Branco, Débora de Souza Collares Maia; Neto, Manoel Paiva de Araújo; Cordeiro, Rossana de Aguiar; Rodrigues, Terezinha de Jesus Santos; Rocha, Marcos Fábio Gadelha; Sidrim, José Júlio Costa

2013-11-01

160

The Type VI Secretion System Spike Protein VgrG5 Mediates Membrane Fusion during Intercellular Spread by Pseudomallei Group Burkholderia Species  

PubMed Central

Pseudomallei group Burkholderia species are facultative intracellular parasites that spread efficiently from cell to cell by a mechanism involving the fusion of adjacent cell membranes. Intercellular fusion requires the function of the cluster 5 type VI secretion system (T6SS-5) and its associated valine-glycine repeat protein, VgrG5. Here we show that VgrG5 alleles are conserved and functionally interchangeable between Burkholderia pseudomallei and its relatives B. mallei, B. oklahomensis, and B. thailandensis. We also demonstrate that the integrity of the VgrG5 C-terminal domain is required for fusogenic activity, and we identify sequence motifs, including two hydrophobic segments, that are important for fusion. Mutagenesis and secretion experiments using B. pseudomallei strains engineered to express T6SS-5 in vitro show that the VgrG5 C-terminal domain is dispensable for T6SS-mediated secretion of Hcp5, demonstrating that the ability of VgrG5 to mediate membrane fusion can be uncoupled from its essential role in type VI secretion. We propose a model in which a unique fusogenic activity at the C terminus of VgrG5 facilitates intercellular spread by B. pseudomallei and related species following injection across the plasma membranes of infected cells. PMID:24421040

Toesca, Isabelle J.; French, Christopher T.

2014-01-01

161

Development and Characterization of a Caprine Aerosol Infection Model of Melioidosis  

PubMed Central

Infection with Burkholderia pseudomallei causes the disease melioidosis, which often presents as a serious suppurative infection that is typically fatal without intensive treatment and is a significant emerging infectious disease in Southeast Asia. Despite intensive research there is still much that remains unknown about melioidosis pathogenesis. New animal models of melioidosis are needed to examine novel aspects of pathogenesis as well as for the evaluation of novel therapeutics. The objective of the work presented here was to develop a subacute to chronic caprine model of melioidosis and to characterize the progression of disease with respect to clinical presentation, hematology, clinical microbiology, thoracic radiography, and gross and microscopic pathology. Disease was produced in all animals following an intratracheal aerosol of 104 CFU delivered, with variable clinical manifestations indicative of subacute and chronic disease. Bronchointerstitial pneumonia was apparent microscopically by day 2 and radiographically and grossly apparent by day 7 post infection (PI). Early lesions of bronchopneumonia soon progressed to more severe bronchointerstitial pneumonia with pyogranuloma formation. Extrapulmonary dissemination appeared to be a function of pyogranuloma invasion of pulmonary vasculature, which peaked around day 7 PI. Histopathology indicated that leukocytoclastic vasculitis was the central step in dissemination of B. pseudomallei from the lungs as well as in the establishment of new lesions. While higher doses of organism in goats can produce acute fatal disease, the dose investigated and resulting disease had many similarities to human melioidosis and may warrant further development to provide a model for the study of both natural and bioterrorism associated disease. PMID:22916225

Soffler, Carl; Bosco-Lauth, Angela M.; Aboellail, Tawfik A.; Marolf, Angela J.; Bowen, Richard A.

2012-01-01

162

Overexpression of the Endothelial Protein C Receptor Is Detrimental during Pneumonia-Derived Gram-negative Sepsis (Melioidosis)  

PubMed Central

Background The endothelial protein C receptor (EPCR) enhances anticoagulation by accelerating activation of protein C to activated protein C (APC) and mediates anti-inflammatory effects by facilitating APC-mediated signaling via protease activated receptor-1. We studied the role of EPCR in the host response during pneumonia-derived sepsis instigated by Burkholderia (B.) pseudomallei, the causative agent of melioidosis, a common form of community-acquired Gram-negative (pneumo)sepsis in South-East Asia. Methodology/Principal Findings Soluble EPCR was measured in plasma of patients with septic culture-proven melioidosis and healthy controls. Experimental melioidosis was induced by intranasal inoculation of B. pseudomallei in wild-type (WT) mice and mice with either EPCR-overexpression (Tie2-EPCR) or EPCR-deficiency (EPCR?/?). Mice were sacrificed after 24, 48 or 72 hours. Organs and plasma were harvested to measure colony forming units, cellular influxes, cytokine levels and coagulation parameters. Plasma EPCR-levels were higher in melioidosis patients than in healthy controls and associated with an increased mortality. Tie2-EPCR mice demonstrated enhanced bacterial growth and dissemination to distant organs during experimental melioidosis, accompanied by increased lung damage, neutrophil influx and cytokine production, and attenuated coagulation activation. EPCR?/? mice had an unremarkable response to B. pseudomallei infection as compared to WT mice, except for a difference in coagulation activation in plasma. Conclusion/Significance Increased EPCR-levels correlate with accelerated mortality in patients with melioidosis. In mice, transgenic overexpression of EPCR aggravates outcome during Gram-negative pneumonia-derived sepsis caused by B. pseudomallei, while endogenous EPCR does not impact on the host response. These results add to a better understanding of the regulation of coagulation during severe (pneumo)sepsis. PMID:23875041

Kager, Liesbeth M.; Schouten, Marcel; Wiersinga, W. Joost; de Boer, J. Daan; Lattenist, Lionel C. W.; Roelofs, Joris J. T. H.; Meijers, Joost C. M.; Levi, Marcel; Dondorp, Arjen M.; Esmon, Charles T.; van 't Veer, Cornelis; van der Poll, Tom

2013-01-01

163

Treatment and prophylaxis of melioidosis  

PubMed Central

Melioidosis, infection with Burkholderia pseudomallei, is being recognised with increasing frequency and is probably more common than currently appreciated. Treatment recommendations are based on a series of clinical trials conducted in Thailand over the past 25 years. Treatment is usually divided into two phases: in the first, or acute phase, parenteral drugs are given for ?10 days with the aim of preventing death from overwhelming sepsis; in the second, or eradication phase, oral drugs are given, usually to complete a total of 20 weeks, with the aim of preventing relapse. Specific treatment for individual patients needs to be tailored according to clinical manifestations and response, and there remain many unanswered questions. Some patients with very mild infections can probably be cured by oral agents alone. Ceftazidime is the mainstay of acute-phase treatment, with carbapenems reserved for severe infections or treatment failures and amoxicillin/clavulanic acid (co-amoxiclav) as second-line therapy. Trimethoprim/sulfamethoxazole (co-trimoxazole) is preferred for the eradication phase, with the alternative of co-amoxiclav. In addition, the best available supportive care is needed, along with drainage of abscesses whenever possible. Treatment for melioidosis is unaffordable for many in endemic areas of the developing world, but the relative costs have reduced over the past decade. Unfortunately there is no likelihood of any new or cheaper options becoming available in the immediate future. Recommendations for prophylaxis following exposure to B. pseudomallei have been made, but the evidence suggests that they would probably only delay rather than prevent the development of infection. PMID:24613038

Dance, David

2014-01-01

164

Treatment and prophylaxis of melioidosis.  

PubMed

Melioidosis, infection with Burkholderia pseudomallei, is being recognised with increasing frequency and is probably more common than currently appreciated. Treatment recommendations are based on a series of clinical trials conducted in Thailand over the past 25 years. Treatment is usually divided into two phases: in the first, or acute phase, parenteral drugs are given for ?10 days with the aim of preventing death from overwhelming sepsis; in the second, or eradication phase, oral drugs are given, usually to complete a total of 20 weeks, with the aim of preventing relapse. Specific treatment for individual patients needs to be tailored according to clinical manifestations and response, and there remain many unanswered questions. Some patients with very mild infections can probably be cured by oral agents alone. Ceftazidime is the mainstay of acute-phase treatment, with carbapenems reserved for severe infections or treatment failures and amoxicillin/clavulanic acid (co-amoxiclav) as second-line therapy. Trimethoprim/sulfamethoxazole (co-trimoxazole) is preferred for the eradication phase, with the alternative of co-amoxiclav. In addition, the best available supportive care is needed, along with drainage of abscesses whenever possible. Treatment for melioidosis is unaffordable for many in endemic areas of the developing world, but the relative costs have reduced over the past decade. Unfortunately there is no likelihood of any new or cheaper options becoming available in the immediate future. Recommendations for prophylaxis following exposure to B. pseudomallei have been made, but the evidence suggests that they would probably only delay rather than prevent the development of infection. PMID:24613038

Dance, David

2014-04-01

165

Redefining the PF06864 Pfam Family Based on Burkholderia pseudomallei PilO2Bp S-SAD Crystal Structure  

PubMed Central

Type IV pili are surface-exposed filaments and bacterial virulence factors, represented by the Tfpa and Tfpb types, which assemble via specific machineries. The Tfpb group is further divided into seven variants, linked to heterogeneity in the assembly machineries. Here we focus on PilO2Bp, a protein component of the Tfpb R64 thin pilus variant assembly machinery from the pathogen Burkholderia pseudomallei. PilO2Bp belongs to the PF06864 Pfam family, for which an improved definition is presented based on newly derived Hidden Markov Model (HMM) profiles. The 3D structure of the N-terminal domain of PilO2Bp (N-PilO2Bp), here reported, is the first structural representative of the PF06864 family. N-PilO2Bp presents an actin-like ATPase fold that is shown to be present in BfpC, a different variant assembly protein; the new HMM profiles classify BfpC as a PF06864 member. Our results provide structural insight into the PF06864 family and on the Type IV pili assembly machinery. PMID:24728008

Manjasetty, Babu A.; Yero, Daniel; Perletti, Lucia; Belrhali, Hassan; Daura, Xavier; Gourlay, Louise J.; Bolognesi, Martino

2014-01-01

166

Nitric Oxide from IFN?-Primed Macrophages Modulates the Antimicrobial Activity of ?-Lactams against the Intracellular Pathogens Burkholderia pseudomallei and Nontyphoidal Salmonella  

PubMed Central

Our investigations show that nonlethal concentrations of nitric oxide (NO) abrogate the antibiotic activity of ?-lactam antibiotics against Burkholderia pseudomallei, Escherichia coli and nontyphoidal Salmonella enterica serovar Typhimurium. NO protects B. pseudomallei already exposed to ?-lactams, suggesting that this diatomic radical tolerizes bacteria against the antimicrobial activity of this important class of antibiotics. The concentrations of NO that elicit antibiotic tolerance repress consumption of oxygen (O2), while stimulating hydrogen peroxide (H2O2) synthesis. Transposon insertions in genes encoding cytochrome c oxidase-related functions and molybdenum assimilation confer B. pseudomallei a selective advantage against the antimicrobial activity of the ?-lactam antibiotic imipenem. Cumulatively, these data support a model by which NO induces antibiotic tolerance through the inhibition of the electron transport chain, rather than by potentiating antioxidant defenses as previously proposed. Accordingly, pharmacological inhibition of terminal oxidases and nitrate reductases tolerizes aerobic and anaerobic bacteria to ?-lactams. The degree of NO-induced ?-lactam antibiotic tolerance seems to be inversely proportional to the proton motive force (PMF), and thus the dissipation of ?H+ and ?? electrochemical gradients of the PMF prevents ?-lactam-mediated killing. According to this model, NO generated by IFN?-primed macrophages protects intracellular Salmonella against imipenem. On the other hand, sublethal concentrations of imipenem potentiate the killing of B. pseudomallei by NO generated enzymatically from IFN?-primed macrophages. Our investigations indicate that NO modulates the antimicrobial activity of ?-lactam antibiotics. PMID:25121731

Jones-Carson, Jessica; Zweifel, Adrienne E.; Tapscott, Timothy; Austin, Chad; Brown, Joseph M.; Jones, Kenneth L.; Voskuil, Martin I.; Vazquez-Torres, Andres

2014-01-01

167

Melioidosis presenting as lymphadenitis: a case report  

PubMed Central

Background Melioidosis is an infection caused by the facultative intracellular gram-negative bacterium; Burkholderia pseudomallei. It gives rise to protean clinical manifestations and has a varied prognosis. Although it was rare in Sri Lanka increasing numbers of cases are being reported with high morbidity and mortality. Here we report a case of melioidosis presenting with lymphadenitis which was diagnosed early and treated promptly with a good outcome. Case presentation A 53-year-old Sinhalese woman with diabetes presented with fever and left sided painful inguinal lymphadenitis for one month. She had undergone incision and drainage of a thigh abscess three months previously and had been treated with a short course of antibiotics. There was no record that abscess material was tested microbiologically. She had neutrophil leukocytosis and elevated inflammatory markers. Initial pus culture revealed a scanty growth of “Pseudomonas sp.” and Escherichia coli which were sensitive to ceftazidime and resistant to gentamicin. Due to the history of diabetes, recurrent abscess formation and the suggestive sensitivity pattern of the bacterial isolates, we actively investigated for melioidosis. The bacterial isolate was subsequently identified as B. pseudomallei by polymerase chain reaction and antibodies to melioidin antigen were found to be raised at a titre of 1:160. The patient was treated with high dose intravenous ceftazidime for four weeks followed by eradication therapy with cotrimoxazole and doxycycline. As the patient was intolerant to cotrimoxazole, the antibiotics were changed to a combination of co-amoxyclav and doxycycline and continued for 12 weeks. The patient was well after 6 months without any relapse. Conclusions Melioidosis is an emerging infection in South Asia. It may present with recurrent abscesses. Therefore it is very important to send pus for culture whenever an abscess is drained. However, it should be noted that the reporting laboratory may be unfamiliar with this bacterium and the isolate may be misidentified as Pseudomonas or even E. coli. Melioidosis should be suspected when an isolate with the typical antibiotic sensitivity pattern of ceftazidime sensitivity and gentamicin resistance is cultured, especially in a patient with diabetes. This will expedite diagnosis and prompt treatment leading to an excellent prognosis. PMID:24927768

2014-01-01

168

Regulation of the katG-dpsA operon and the importance of KatG in survival of Burkholderia pseudomallei exposed to oxidative stress.  

PubMed

Homologues of the catalase-peroxidase gene katG and the gene for the non-specific DNA binding protein dpsA were identified downstream of oxyR in Burkholderia pseudomallei. Northern experiments revealed that both katG and dpsA are co-transcribed during oxidative stress. Under conditions where the katG promoter is not highly induced, dpsA is transcribed from a second promoter located within the katG-dpsA intergenic region. A katG insertion mutant was found to be hypersensitive to various oxidants. Analysis of katG expression in the oxyR mutant indicates that OxyR is a dual function regulator that represses the expression of katG during normal growth and activates katG during exposure to oxidative stress. Both reduced and oxidized OxyR were shown to bind to the katG promoter. PMID:12729890

Loprasert, Suvit; Whangsuk, Wirongrong; Sallabhan, Ratiboot; Mongkolsuk, Skorn

2003-05-01

169

A non-invasive intratracheal inoculation method for the study of pulmonary melioidosis  

PubMed Central

Pulmonary melioidosis, a disease manifestation caused by the bacterium Burkholderia pseudomallei, has been studied using aerosols or intranasal (IN) inoculation in small animal models. Both have inherent disadvantages which may not accurately model primary pulmonary melioidosis in humans. Intratracheal inoculation (IT) by direct visualization of the tracheal opening offers an alternative technique for infection that overcomes the disadvantages of aerosol and IN challenge. In this study, we describe a method which requires relatively inexpensive equipment, little training, and is compliant with the operational constraints of a BSL3 laboratory. Results obtained using trypan blue demonstrated that an inoculum can be accurately delivered into the lungs of mice within a biosafety cabinet (BSC). Whole body imaging and histopathology confirmed that mice inoculated intratracheally with B. pseudomallei develop the primary focus of infection in the lungs, and not the nasal passages which can lead to invasion of the central nervous system and potential neurologic complications. Further, based on colony counts and bioluminescent imaging, dissemination to secondary organs occurred as expected. Taken together, this intratracheal method of inoculation fulfills four goals: (1) to accurately deliver B. pseudomallei into the lungs of the animal model, (2) to avoid potentially confounding complications due to primary infections at sites other than the lung, (3) to maintain normal organ dissemination, and (4) to be BSL3 compliant. PMID:23267442

Revelli, David A.; Boylan, Julie A.; Gherardini, Frank C.

2012-01-01

170

Neutrophil Elastase Causes Tissue Damage That Decreases Host Tolerance to Lung Infection with Burkholderia Species  

PubMed Central

Two distinct defense strategies can protect the host from infection: resistance is the ability to destroy the infectious agent, and tolerance is the ability to withstand infection by minimizing the negative impact it has on the host's health without directly affecting pathogen burden. Burkholderia pseudomallei is a Gram-negative bacterium that infects macrophages and causes melioidosis. We have recently shown that inflammasome-triggered pyroptosis and IL-18 are equally important for resistance to B. pseudomallei, whereas IL-1? is deleterious. Here we show that the detrimental role of IL-1? during infection with B. pseudomallei (and closely related B. thailandensis) is due to excessive recruitment of neutrophils to the lung and consequent tissue damage. Mice deficient in the potentially damaging enzyme neutrophil elastase were less susceptible than the wild type C57BL/6J mice to infection, although the bacterial burdens in organs and the extent of inflammation were comparable between C57BL/6J and elastase-deficient mice. In contrast, lung tissue damage and vascular leakage were drastically reduced in elastase-deficient mice compared to controls. Bradykinin levels were higher in C57BL/6 than in elastase-deficient mice; administration of a bradykinin antagonist protected mice from infection, suggesting that increased vascular permeability mediated by bradykinin is one of the mechanisms through which elastase decreases host tolerance to melioidosis. Collectively, these results demonstrate that absence of neutrophil elastase increases host tolerance, rather than resistance, to infection by minimizing host tissue damage. PMID:25166912

Miller, Mark A.; Re, Fabio

2014-01-01

171

In Vivo Manipulation of ?9+ T Cells in the Common Marmoset (Callithrix Jacchus) with Phosphoantigen and Effect on the Progression of Respiratory Melioidosis  

PubMed Central

Burkholderia pseudomallei is a dangerous human pathogen. Phosphoantigens specifically the target primate specific ?9+?2+ T cells subset and some have been developed as potential immunotherapeutics. Previously, we demonstrated that, when stimulated with the phosphoantigen CHDMAPP, ?9+?2+ T cells aid in the killing of intracellular B. pseudomallei bacteria. Moreover, we found that common marmoset (Callithrix Jacchus) ?9+ T cells increase in frequency and respond to the phosphoantigen CHDMAPP and/or B. pseudomallei, in combination with IL-2, in a similar manner to human ?9+?2+ T cells. Here we evaluate the efficacy of the phosphoantigen CHDMAPP, in combination with IL-2, as a therapy against B. pseudomallei infection, in vivo. We found that the previous studies predicted the in vivo responsiveness of ?9+ T cells to the CHDMAPP+IL-2 treatment and significant expansion of the numbers of peripheral and splenic ?9+ T cells were observed. This effect was similar to those reported in other primate species treated with phosphoantigen. Furthermore, splenocytes were retrieved 7 days post onset of treatment, restimulated with CHDMAPP or heat-killed B. pseudomallei and the cultured ?9+ T cells demonstrated no reduction in IFN-? response when CHDMAPP+IL-2 animals were compared to IL-2 only treated animals. Using an established model of B. pseudomallei infection in the marmoset, we assessed the potential for using phosphoantigen as a novel immunotherapy. The CHDMAPP treatment regime had no effect on the progression of respiratory melioidosis and this was despite the presence of elevated numbers of ?9+ T cells in the spleen, liver and lung and an increased proportion of IFN-?+ cells in response to infection. We therefore report that the common marmoset has proven a good model for studying the effect in vivo of ?9+ T cell stimulation; however, ?9+ T cells have little or no effect on the progression of lethal, respiratory B. pseudomallei infection. PMID:24098670

Laws, Thomas R.; Nelson, Michelle; Bonnafous, Cecile; Sicard, Helene; Taylor, Christopher; Salguero, Francisco Javier; Atkins, Timothy P.; Oyston, Petra C. F.; Rowland, Caroline A.

2013-01-01

172

Genomic acquisition of a capsular polysaccharide virulence cluster by non-pathogenic Burkholderia isolates  

PubMed Central

Background Burkholderia thailandensis is a non-pathogenic environmental saprophyte closely related to Burkholderia pseudomallei, the causative agent of the often fatal animal and human disease melioidosis. To study B. thailandensis genomic variation, we profiled 50 isolates using a pan-genome microarray comprising genomic elements from 28 Burkholderia strains and species. Results Of 39 genomic regions variably present across the B. thailandensis strains, 13 regions corresponded to known genomic islands, while 26 regions were novel. Variant B. thailandensis isolates exhibited isolated acquisition of a capsular polysaccharide biosynthesis gene cluster (B. pseudomallei-like capsular polysaccharide) closely resembling a similar cluster in B. pseudomallei that is essential for virulence in mammals; presence of this cluster was confirmed by whole genome sequencing of a representative variant strain (B. thailandensis E555). Both whole-genome microarray and multi-locus sequence typing analysis revealed that the variant strains formed part of a phylogenetic subgroup distinct from the ancestral B. thailandensis population and were associated with atypical isolation sources when compared to the majority of previously described B. thailandensis strains. In functional assays, B. thailandensis E555 exhibited several B. pseudomallei-like phenotypes, including colony wrinkling, resistance to human complement binding, and intracellular macrophage survival. However, in murine infection assays, B. thailandensis E555 did not exhibit enhanced virulence relative to other B. thailandensis strains, suggesting that additional factors are required to successfully colonize and infect mammals. Conclusions The discovery of such novel variant strains demonstrates how unbiased genomic surveys of non-pathogenic isolates can reveal insights into the development and emergence of new pathogenic species. PMID:20799932

2010-01-01

173

Activities of Daily Living Associated with Acquisition of Melioidosis in Northeast Thailand: A Matched Case-Control Study  

PubMed Central

Background Melioidosis is a serious infectious disease caused by the Category B select agent and environmental saprophyte, Burkholderia pseudomallei. Most cases of naturally acquired infection are assumed to result from skin inoculation after exposure to soil or water. The aim of this study was to provide evidence for inoculation, inhalation and ingestion as routes of infection, and develop preventive guidelines based on this evidence. Methods/Principal Findings A prospective hospital-based 1?2 matched case-control study was conducted in Northeast Thailand. Cases were patients with culture-confirmed melioidosis, and controls were patients admitted with non-infectious conditions during the same period, matched for gender, age, and diabetes mellitus. Activities of daily living were recorded for the 30-day period before onset of symptoms, and home visits were performed to obtain drinking water and culture this for B. pseudomallei. Multivariable conditional logistic regression analysis based on 286 cases and 512 controls showed that activities associated with a risk of melioidosis included working in a rice field (conditional odds ratio [cOR]?=?2.1; 95% confidence interval [CI] 1.4–3.3), other activities associated with exposure to soil or water (cOR?=?1.4; 95%CI 0.8–2.6), an open wound (cOR?=?2.0; 95%CI 1.2–3.3), eating food contaminated with soil or dust (cOR?=?1.5; 95%CI 1.0–2.2), drinking untreated water (cOR?=?1.7; 95%CI 1.1–2.6), outdoor exposure to rain (cOR?=?2.1; 95%CI 1.4–3.2), water inhalation (cOR?=?2.4; 95%CI 1.5–3.9), current smoking (cOR?=?1.5; 95%CI 1.0–2.3) and steroid intake (cOR?=?3.1; 95%CI 1.4–6.9). B. pseudomallei was detected in water source(s) consumed by 7% of cases and 3% of controls (cOR?=?2.2; 95%CI 0.8–5.8). Conclusions/Significance We used these findings to develop the first evidence-based guidelines for the prevention of melioidosis. These are suitable for people in melioidosis-endemic areas, travelers and military personnel. Public health campaigns based on our recommendations are under development in Thailand. PMID:23437412

Limmathurotsakul, Direk; Kanoksil, Manas; Wuthiekanun, Vanaporn; Kitphati, Rungrueng; deStavola, Bianca; Day, Nicholas P. J.; Peacock, Sharon J.

2013-01-01

174

Burkholderia thailandensis Is Virulent in Drosophila melanogaster  

PubMed Central

Melioidosis is a serious infectious disease endemic to Southeast Asia and Northern Australia. This disease is caused by the Gram-negative bacterium Burkholderia pseudomallei; Burkholderia thailandensis is a closely-related organism known to be avirulent in humans. B. thailandensis has not previously been used to infect Drosophila melanogaster. We examined the effect of B. thailandensis infection on fly survival, on antimicrobial peptide expression, and on phagocytic cells. In the fruit fly, which possesses only an innate immune system, B. thailandensis is highly virulent, causing rapid death when injected or fed. One intriguing aspect of this infection is its temperature dependence: infected flies maintained at 25°C exhibit rapid bacterial proliferation and death in a few days, while infected animals maintained at 18°C exhibit very slow bacterial proliferation and take weeks to die; this effect is due in part to differences in immune activity of the host. Death in this infection is likely due at least in part to a secreted toxin, as injection of flies with sterile B. thailandensis-conditioned medium is able to kill. B. thailandensis infection strongly induces the expression of antimicrobial peptides, but this is insufficient to inhibit bacterial proliferation in infected flies. Finally, the function of fly phagocytes is not affected by B. thailandensis infection. The high virulence of B. thailandensis in the fly suggests the possibility that this organism is a natural pathogen of one or more invertebrates. PMID:23209596

Pilatova, Martina; Dionne, Marc S.

2012-01-01

175

The effect of glibenclamide on the pathogenesis of melioidosis  

E-print Network

organism) and when aerosolised, might see utility as a bioterror agent with a high mortality rate. 1.1 Burkholderia pseudomallei 1.1.1 Taxonomy B. pseudomallei was for a long time placed in the genus Pseudomonas on the basis of its Gram-stain appearance... Burkholderia cepacia, Pseudomonas aeruginosa, Chromobacterium violaceum and other non-lactose fermenting Gram-negative bacilli. Variation in the API profile occurs primarily in citrate utilisation, and in sucrose and amygdalin assimilation. The antibiotic...

Koh, Gavin Christian Kia Wee

2012-03-06

176

Whole-Genome Assemblies of 56 Burkholderia Species  

PubMed Central

Burkholderia is a genus of betaproteobacteria that includes three notable human pathogens: B. cepacia, B. pseudomallei, and B. mallei. While B. pseudomallei and B. mallei are considered potential biowarfare agents, B. cepacia infections are largely limited to cystic fibrosis patients. Here, we present 56 Burkholderia genomes from 8 distinct species. PMID:25414490

Daligault, H. E.; Davenport, K. W.; Minogue, T. D.; Bishop-Lilly, K. A.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Frey, K. G.; Gibbons, H. S.; Jaissle, J.; Koroleva, G. I.; Ladner, J. T.; Lo, C.-C.; Munk, C.; Palacios, G. F.; Redden, C. L.; Rosenzweig, C. N.; Scholz, M. B.

2014-01-01

177

Effect of colony morphology variation of Burkholderia pseudomallei on intracellular survival and resistance to antimicrobial environments in human macrophages in vitro  

E-print Network

. pseudomallei, 1 × 105 U937 cells per well were trans- ferred to a 24 well-tissue culture plate (BD Falcon) and activated by the addition of 50 ng/ml of phorbol 12- myristate 13-acetate (PMA) (Sigma) over 2 days [20]. The medium was then replaced with 1 ml...

Tandhavanant, Sarunporn; Thanwisai, Aunchalee; Limmathurotsakul, Direk; Korbsrisate, Sunee; Day, Nicholas P J; Peacock, Sharon J; Chantratita, Narisara

2010-11-30

178

[Glanders, melioidosis and biowarfare].  

PubMed

MANY COMMON FACTORS: Glanders and melioidosis are infectious diseases that are caused by the bacteria of the Burkholderia species. These infections are endemic in tropical regions and can lead to la broad spectrum of common clinical manifestations. TWO PRINCIPLE CLINICAL FORMS: The most frequent clinical presentation is the pulmonary form, which can mimic pulmonary tuberculosis. The septicemic form is the most severe form, and lethal in nearly 50% of cases. WEAPONS FOR BIOTERRORISM AND WAR: Very few organisms are required to cause disease by aerosolisation, which could be the main route of contamination for humans after a deliberate release. This property has permitted yet the use of these bacteria as biological warfare weapon during the past century. We have to consider these agents as possible biological warfare agents. Europeans guidelines for treatment and post-exposure prophylaxis are detailed. PMID:15687969

Guilhot, Amélie; Bricaire, François; Bossi, Philippe

2005-01-29

179

Melioidosis: the tip of the iceberg?  

PubMed Central

For nearly 80 years clinical melioidosis has been considered a rare disease. This bacterial infection is caused by Pseudomonas pseudomallei, a saprophyte found in soil and surface water of endemic areas. Consequently, those who have most contact with soil, the rural poor, are likely to be at greatest risk of infection. Since the diversity of clinical manifestations necessitates the isolation and identification of the causative organism for a definitive diagnosis of melioidosis and the population at greatest risk within endemic areas rarely have access to an appropriate level of health care, the disease has probably been underrecognized. Melioidosis is now known to be an important cause of human morbidity and mortality in Thailand, and this may be true throughout Southeast Asia, which is usually regarded as the main endemic area for the disease. In Australia, melioidosis causes a smaller number of human infections, while disease among livestock has important economic and possible public health implications. Sporadic reports of the infection indicate its presence in several other tropical regions: in the Indian subcontinent, Africa, and Central and South America. Clinical melioidosis may be highly prevalent in these areas, but underdiagnosed as a result of a lack of awareness of the clinical and microbiological features of the disease, or simply because of a lack of health care facilities. Furthermore, during the last two decades the importation and transmission of melioidosis within nontropical zones have been documented. The causative organism is not difficult to grow, and modern antibiotics have improved disease prognosis. Further studies are needed to determine the true worldwide distribution and prevalence of melioidosis so that improved therapeutic and preventive measures can be developed and applied. PMID:2004347

Dance, D A

1991-01-01

180

Stable, Site-Specific Fluorescent Tagging Constructs Optimized for Burkholderia Species?  

PubMed Central

Several vectors that facilitate stable fluorescent labeling of Burkholderia pseudomallei and Burkholderia thailandensis were constructed. These vectors combined the effectiveness of the mini-Tn7 site-specific transposition system with fluorescent proteins optimized for Burkholderia spp., enabling bacterial tracking during cellular infection. PMID:20851961

Norris, Michael H.; Kang, Yun; Wilcox, Bruce; Hoang, Tung T.

2010-01-01

181

Isolation of Pseudomonas pseudomallei from soil in north-eastern Thailand  

Microsoft Academic Search

In order to optimize the recovery from soil of Pseudomonas pseudomallei, the cause of melioidosis, 3 selective broths were compared. A basal salt solution containing l-threonine (TBSS) performed significantly better than trypticase soy broth containing crystal violet and colistin 50 mg\\/L (CVC50), both in isolation rate and suppression of overgrowth of other organisms, but the addition of colistin to TBSS

Vanaporn Wuthiekanun; Michael D. Smith; David A. B. Dance; Nicholas J. White

1995-01-01

182

Melioidosis in a returned traveller  

PubMed Central

Melioidosis is an infection endemic to Southeast Asia and Northern Australia, and is associated with significant morbidity and mortality. The present report describes a case of chronic melioidosis in a returning traveller from the Philippines. Clinical suspicion of this illness is warranted in individuals with a history of travel to endemic regions. Safety in handling clinical specimens is paramount because laboratory transmission has been described.

Chagla, Zain; Aleksova, Natasha; Quirt, Jaclyn; Emery, Joel; Kraeker, Christian; Haider, Shariq

2014-01-01

183

Fatal Melioidosis in Goats in Bangkok, Thailand  

PubMed Central

Bangkok, Thailand, is a city considered to be at low risk for melioidosis. We describe 10 goats that died of melioidosis in Bangkok. Half of them were born and reared in the city. Multilocus sequence typing ruled out an outbreak. This finding challenges the assumption that melioidosis is rarely acquired in central Thailand. PMID:24891468

Tonpitak, Walaiporn; Sornklien, Chulabha; Chawanit, Mongkol; Pavasutthipaisit, Suvarin; Wuthiekanun, Vanaporn; Hantrakun, Viriya; Amornchai, Premjit; Thaipadungpanit, Janjira; Day, Nicholas P. J.; Yingst, Samuel; Peacock, Sharon J.; Limmathurotsakul, Direk

2014-01-01

184

Melioidosis as an emerging global problem  

Microsoft Academic Search

There is remarkably little known about the incidence of melioidosis outside a few countries (Thailand, Australia, Singapore and Malaysia). Presumably it is widespread in tropical south east Asia. Elsewhere there are tantalising glimpses of the tip of what may be a large iceberg. Since a specific diagnosis of melioidosis requires awareness on the part of clinicians, and the existence of

David A. B Dance

2000-01-01

185

Human melioidosis: an emerging medical problem  

Microsoft Academic Search

Summary Human melioidosis is endemic in South East Asia and tropical Australia. However, increasing numbers of case reports are coming from other parts of the world. Increasing world travel and the potential for person-to-person infection in non-endemic areas make the likelihood of physicians and medical microbiologists encountering the disease far greater than heretofore. Both the disease, melioidosis, and the causative

C. J. Smith; J. C. Allen; M. Noor Embi; O. Othmant; N. Razakt; G. Ismail

1987-01-01

186

Genus-wide acid tolerance accounts for the biogeographical distribution of soil Burkholderia populations.  

PubMed

Bacteria belonging to the genus Burkholderia are highly versatile with respect to their ecological niches and lifestyles, ranging from nodulating tropical plants to causing melioidosis and fatal infections in cystic fibrosis patients. Despite the clinical importance and agronomical relevance of Burkholderia species, information about the factors influencing their occurrence, abundance and diversity in the environment is scarce. Recent findings have demonstrated that pH is the main predictor of soil bacterial diversity and community structure, with the highest diversity observed in neutral pH soils. As many Burkholderia species have been isolated from low pH environments, we hypothesized that acid tolerance may be a general feature of this genus, and pH a good predictor of their occurrence in soils. Using a combination of environmental surveys at trans-continental and local scales, as well as in vitro assays, we show that, unlike most bacteria, Burkholderia species have a competitive advantage in acidic soils, but are outcompeted in alkaline soils. Physiological assays and diversity analysis based on 16S rRNA clone libraries demonstrate that pH tolerance is a general phenotypic trait of the genus Burkholderia. Our results provide a basis for building a predictive understanding of the biogeographical patterns exhibited by Burkholderia sp. PMID:23945027

Stopnisek, Nejc; Bodenhausen, Natacha; Frey, Beat; Fierer, Noah; Eberl, Leo; Weisskopf, Laure

2014-06-01

187

Immunogenic Consensus Sequence T helper Epitopes for a Pan-Burkholderia Biodefense Vaccine  

PubMed Central

Background Biodefense vaccines against Category B bioterror agents Burkholderia pseudomallei (BPM) and Burkholderia mallei (BM) are needed, as they are both easily accessible to terrorists and have strong weaponization potential. Burkholderia cepaciae (BC), a related pathogen, causes chronic lung infections in cystic fibrosis patients. Since BPM, BM and BC are all intracellular bacteria, they are excellent targets for T cell-based vaccines. However, the sheer volume of available genomic data requires the aid of immunoinformatics for vaccine design. Using EpiMatrix, ClustiMer and EpiAssembler, a set of immunoinformatic vaccine design tools, we screened the 31 available Burkholderia genomes and performed initial tests of our selections that are candidates for an epitope-based multi-pathogen vaccine against Burkholderia species. Results Immunoinformatics analysis of 31 Burkholderia genomes yielded 350,004 9-mer candidate vaccine peptides of which 133,469 had perfect conservation across the 10 BM genomes, 175,722 had perfect conservation across the 11 BPM genomes and 40,813 had perfect conservation across the 10 BC genomes. Further screening with EpiMatrix yielded 54,010 high-scoring Class II epitopes; these were assembled into 2,880 longer highly conserved ‘immunogenic consensus sequence’ T helper epitopes. 100% of the peptides bound to at least one HLA class II allele in vitro, 92.7% bound to at least two alleles, 82.9% to three, and 75.6% of the binding results were consistent with the immunoinformatics analysis. Conclusions Our results show it is possible to rapidly identify promiscuous T helper epitopes conserved across multiple Burkholderia species and test their binding to HLA ligands in vitro. The next step in our process will be to test the epitopes ex vivo using peripheral leukocytes from BC, BPM infected humans and for immunogenicity in human HLA transgenic mice. We expect that this approach will lead to development of a licensable, pan-Burkholderia biodefense vaccine.

De Groot, Anne S.; Ardito, Matthew; Moise, Leonard; Gustafson, Eric A.; Spero, Denice; Tejada, Gloria; Martin, William

2014-01-01

188

In vitro susceptibilities of Pseudomonas pseudomallei to 27 antimicrobial agents.  

PubMed Central

Clinical isolates of Pseudomonas pseudomallei isolated in Thailand from 1981 to 1989 were tested for their in vitro susceptibilities to 27 antimicrobial agents, including older and newer quinolones, broad-spectrum cephems, carbapenems, monobactams, penicillins, tetracyclines, chloramphenicol, rifamycin, sulfamethoxazole, trimethoprim, and fosfomycin. Tosufloxacin, meropenem, CS-533, and minocycline were active against P. pseudomallei at levels comparable to or even greater than those of antimicrobial agents tested in previous studies, such as ciprofloxacin, ceftazidime, imipenem, carumonam, and piperacillin. Drug-resistant P. pseudomallei was found in only 1% of the isolates. The drug-resistant P. pseudomallei isolates displayed a unique pattern of susceptibility to the above-listed drugs. PMID:2291671

Yamamoto, T; Naigowit, P; Dejsirilert, S; Chiewsilp, D; Kondo, E; Yokota, T; Kanai, K

1990-01-01

189

Global Analysis of the Burkholderia thailandensis Quorum Sensing-Controlled Regulon  

PubMed Central

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

Majerczyk, Charlotte; Brittnacher, Mitchell; Jacobs, Michael; Armour, Christopher D.; Radey, Mathew; Schneider, Emily; Phattarasokul, Somsak; Bunt, Richard

2014-01-01

190

Clinical-Epidemiological Features of 13 Cases of Melioidosis in Brazil  

PubMed Central

The aim of this work was to catalog the clinical and ecoepidemiological characteristics of melioidosis in Brazil. The clinical-epidemiological features of melioidosis in Ceará are similar to those in other regions where the disease is endemic. These findings support the inclusion of this Brazilian state as part of the zone of endemicity for melioidosis. PMID:22814457

Bandeira, Tereza J. P. G.; Cordeiro, Rossana A.; Grangeiro, Thalles B.; Lima, Rita A. C.; Ribeiro, Joyce F.; Castelo-Branco, Debora S. C. M.; Rodrigues, Jorge L. N.; Coelho, Ivo C. B.; Magalhaes, Francisco G.; Rocha, Marcos F. G.; Sidrim, Jose J. C.

2012-01-01

191

Isolation of Burkholderia cenocepacia and Burkholderia vietnamiensis from human sewage  

Microsoft Academic Search

Fresh human sewage was examined from a sewage treatment plant for the presence of members of the Burkholderia cepacia complex (BCC) of bacterial organisms and confirmed the presence of viable B. cenocepacia and B. vietnamiensis, by a combination of cultural, phenotypic and genotypic techniques. Both these organisms are important respiratory pathogens for patients with cystic fibrosis (CF). Presently, the survival

Damian McNeely; John E. Moore; J. Stuart Elborn; B. Cherie Millar; Jackie Rendall; James S. G. Dooley

2009-01-01

192

CHROMOSOMAL MULTIPLICITY IN BURKHOLDERIA CEPACIA  

EPA Science Inventory

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 ...

193

Burkholderia thailandensis harbors two identical rhl gene clusters responsible for the biosynthesis of rhamnolipids  

PubMed Central

Background Rhamnolipids are surface active molecules composed of rhamnose and ?-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-hydroxyalkanoyloxy)alkanoic 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 ?rhlA 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 results add another Burkholderia species to the list of bacteria able to produce rhamnolipids and this, by the means of two identical functional gene clusters. Our results also demonstrate the very impressive tensio-active properties these long-chain rhamnolipids possess in comparison to the well-studied short-chain ones from P. aeruginosa. PMID:20017946

2009-01-01

194

Occurrence of Multiple Genomovars of Burkholderia cepacia in Cystic Fibrosis Patients and Proposal of Burkholderia multivorans sp. nov  

Microsoft Academic Search

We performed an integrated genotypic and phenotypic analysis of 128 strains of the genera Burkholderia, Rabtonia, and Pseudomonas in order to study the taxonomic structure of Burkholderia cepacia and its relation- ships with other Burkholderia species. Our data show that presumed B. cepacia strains isolated from cystic fibrosis patients belong to at least five distinct genomic species, one of which

P. VANDAMME; B. HOLMES; M. VANCANNEYT; T. COENYE; B. HOSTE; R. COOPMAN; H. REVETS; S. LAUWERS; M. GILLIS; K. KERSTERS; J. R. W. GOVAN

195

The Twin Arginine Translocation System Is Essential for Aerobic Growth and Full Virulence of Burkholderia thailandensis  

PubMed Central

The twin arginine translocation (Tat) system in bacteria is responsible for transporting folded proteins across the cytoplasmic membrane, and in some bacteria, Tat-exported substrates have been linked to virulence. We report here that the Tat machinery is present in Burkholderia pseudomallei, B. mallei, and B. thailandensis, and we show that the system is essential for aerobic but not anaerobic growth. Switching off of the Tat system in B. thailandensis grown anaerobically resulted in filamentous bacteria, and bacteria showed increased sensitivity to some ?-lactam antibiotics. In Galleria mellonella and zebrafish infection models, the Tat conditional mutant was attenuated. The aerobic growth-restricted phenotype indicates that Tat substrates may play a functional role in oxygen-dependent energy conservation. In other bacteria, aerobic growth restriction in Tat mutants has been attributed to the inability to translocate PetA, the Rieske iron-sulfur protein which forms part of the quinol-cytochrome c oxidoreductase complex. Here, we show that PetA is not responsible for aerobic growth restriction in B. thailandensis. However, we have identified an operon encoding 2 proteins of unknown function (BTH_I2176 and BTH_I2175) that play a role in aerobic growth restriction, and we present evidence that BTH_I2176 is Tat translocated. PMID:24214943

Wagley, Sariqa; Hemsley, Claudia; Thomas, Rachael; Moule, Madeleine G.; Vanaporn, Muthita; Andreae, Clio; Robinson, Matthew; Goldman, Stan; Wren, Brendan W.; Butler, Clive S.

2014-01-01

196

The multifarious, multireplicon Burkholderia cepacia complex  

Microsoft Academic Search

The Burkholderia cepacia complex (Bcc) is a collection of genetically distinct but phenotypically similar bacteria that are divided into at least nine species. Bcc bacteria are found throughout the environment, where they can have both beneficial and detrimental effects on plants and some members can also degrade natural and man-made pollutants. Bcc bacteria are now recognized as important opportunistic pathogens

Eshwar Mahenthiralingam; Teresa A. Urban; Joanna B. Goldberg

2005-01-01

197

Quorum sensing systems influence Burkholderia cenocepacia virulence.  

PubMed

Burkholderia cepacia complex strains communicate using N-acyl homoserine lactones and BDSF-dependent quorum sensing (QS) systems. Burkholderia cenocepacia QS systems include CepIR, CciIR, CepR2 and BDSF. Analysis of CepR, CciIR, CepR2 and RpfF (BDSF synthase) QS regulons revealed that these QS systems both independently regulate and coregulate many target genes, often in an opposing manner. The role of QS and several QS-regulated genes in virulence has been determined using vertebrate, invertebrate and plant infection models. Virulence phenotypes are strain and model dependent, suggesting that different QS-regulated genes are important depending on the strain and type of infection. QS inhibitors in combination with antibiotics can reduce biofilm formation and virulence in infection models. PMID:23231487

Subramoni, Sujatha; Sokol, Pamela A

2012-12-01

198

An ancient but promiscuous host–symbiont association between Burkholderia gut symbionts and their heteropteran hosts  

Microsoft Academic Search

Here, we investigated 124 stinkbug species representing 20 families and 5 superfamilies for their Burkholderia gut symbionts, of which 39 species representing 6 families of the superfamilies Lygaeoidea and Coreoidea were Burkholderia-positive. Diagnostic PCR surveys revealed high frequencies of Burkholderia infection in natural populations of the stinkbugs, and substantial absence of vertical transmission of Burkholderia infection to their eggs. In

Yoshitomo Kikuchi; Takahiro Hosokawa; Takema Fukatsu

2011-01-01

199

Glibenclamide reduces pro-inflammatory cytokine production by neutrophils of diabetes patients in response to bacterial infection  

NASA Astrophysics Data System (ADS)

Type 2 diabetes mellitus is a major risk factor for melioidosis, which is caused by Burkholderia pseudomallei. Our previous study has shown that polymorphonuclear neutrophils (PMNs) from diabetic subjects exhibited decreased functions in response to B. pseudomallei. Here we investigated the mechanisms regulating cytokine secretion of PMNs from diabetic patients which might contribute to patient susceptibility to bacterial infections. Purified PMNs from diabetic patients who had been treated with glibenclamide (an ATP-sensitive potassium channel blocker for anti-diabetes therapy), showed reduction of interleukin (IL)-1? and IL-8 secretion when exposed to B. pseudomallei. Additionally, reduction of these pro-inflammatory cytokines occurred when PMNs from diabetic patients were treated in vitro with glibenclamide. These findings suggest that glibenclamide might be responsible for the increased susceptibility of diabetic patients, with poor glycemic control, to bacterial infections as a result of its effect on reducing IL-1? production by PMNs.

Kewcharoenwong, Chidchamai; Rinchai, Darawan; Utispan, Kusumawadee; Suwannasaen, Duangchan; Bancroft, Gregory J.; Ato, Manabu; Lertmemongkolchai, Ganjana

2013-11-01

200

40 CFR 725.1075 - Burkholderia cepacia complex.  

Code of Federal Regulations, 2011 CFR

...ACT REPORTING REQUIREMENTS AND REVIEW PROCESSES FOR MICROORGANISMS Significant New Uses for Specific Microorganisms § 725.1075 Burkholderia cepacia complex. (a) Microorganism and significant new uses subject to reporting....

2011-07-01

201

40 CFR 725.1075 - Burkholderia cepacia complex.  

Code of Federal Regulations, 2010 CFR

...ACT REPORTING REQUIREMENTS AND REVIEW PROCESSES FOR MICROORGANISMS Significant New Uses for Specific Microorganisms § 725.1075 Burkholderia cepacia complex. (a) Microorganism and significant new uses subject to reporting....

2010-07-01

202

40 CFR 725.1075 - Burkholderia cepacia complex.  

Code of Federal Regulations, 2012 CFR

...ACT REPORTING REQUIREMENTS AND REVIEW PROCESSES FOR MICROORGANISMS Significant New Uses for Specific Microorganisms § 725.1075 Burkholderia cepacia complex. (a) Microorganism and significant new uses subject to reporting....

2012-07-01

203

40 CFR 725.1075 - Burkholderia cepacia complex.  

Code of Federal Regulations, 2013 CFR

...ACT REPORTING REQUIREMENTS AND REVIEW PROCESSES FOR MICROORGANISMS Significant New Uses for Specific Microorganisms § 725.1075 Burkholderia cepacia complex. (a) Microorganism and significant new uses subject to reporting....

2013-07-01

204

TNT and nitroaromatic compounds are chemoattractants for Burkholderia cepacia R34 and Burkholderia sp. strain DNT.  

PubMed

Nitroaromatic compounds are toxic and potential carcinogens. In this study, a drop assay was used to detect chemotaxis toward nitroaromatic compounds for wild-type Burkholderia cepacia R34, wild-type Burkholderia sp. strain DNT, and a 2,4-dinitrotoluene (2,4-DNT) dioxygenase mutant strain (S5). The three strains are chemotactic toward 2,4,6-trinitrotoluene (TNT), 2,3-DNT, 2,4-DNT, 2,5-DNT, 2-nitrotoluene (NT), 4NT, and 4-methyl-5-nitrocatechol (4M5NC), but not toward 2,6-DNT. Of these, only 2,4-DNT is a carbon and energy source for B. cepacia R34 and Burkholderia sp. strain DNT, and 4M5NC is an intermediate in the 2,4-DNT degradation pathway. It was determined that the 2,4-DNT dioxygenase genes are not required for the chemotaxis for these nitroaromatic compounds because the DNT DDO mutant S5 has a chemotactic response toward 2,4-DNT although 2,4-DNT is not metabolized by S5; hence, 2,4-DNT itself is the chemoattractant. This is the first report of chemotaxis toward TNT, 2,3-DNT, 2,4-DNT, 2,5-DNT, 2NT, 4NT, and 4M5NC. PMID:15856226

Leungsakul, Thammajun; Keenan, Brendan G; Smets, Barth F; Wood, Thomas K

2005-12-01

205

Burkholderia cepacia: current clinical issues, environmental controversies and ethical dilemmas  

Microsoft Academic Search

Burkholderia cepacia: current clinical issues, environmental controversies and ethical dilemmas. A.M. Jones, M.E. Dodd, A.K. Webb. #ERS Journals Ltd 2001. ABSTRACT: Burkholderia cepacia is a plant phytogen and is known as a hardy and versatile organism. Over the past two decades it has emerged as a pathogen in the cystic fibrosis (CF) community, with devastating effects. Pulmonary colonisation can lead

A. M. Jones; M. E. Dodd; A. K. Webb

2001-01-01

206

Burkholderia multivorans survival and trafficking within macrophages.  

PubMed

Cystic fibrosis (CF) patients are at great risk of opportunistic lung infection, particularly by members of the Burkholderia cepacia complex (Bcc). This group of bacteria can cause damage to the lung tissue of infected patients and are difficult to eradicate due to their high levels of antibiotic resistance. Although the highly virulent Burkholderia cenocepacia has been the focus of virulence research for the past decade, Burkholderia multivorans is emerging as the most prevalent Bcc species infecting CF patients in North America. Despite several studies detailing the intramacrophage trafficking and survival of B. cenocepacia, no such data exist for B. multivorans. The results of this study demonstrated that the clinical CF isolates C5568 and C0514 and an environmental B. multivorans isolate, ATCC 17616, were able to replicate and survive within murine macrophages in a manner similar to that of B. cenocepacia strain K56-2. These strains were also able to survive but were unable to replicate within human THP-1 macrophages. Differences in macrophage uptake were observed among all three B. multivorans strains; these variances were attributed to major differences in O-antigen production. Unlike B. cenocepacia-containing vacuoles, which delay phagosomal maturation in murine macrophages by 6 h, all B. multivorans-containing vacuoles co-localized with lysosome-associated membrane protein-1, a late endosome/lysosomal marker, and the lysosomal marker dextran within 2 h of uptake. Together, these results indicated that, whilst both Bcc species were able to survive and replicate within macrophages, they utilized different intramacrophage survival strategies. To observe differences in virulence, the strains were compared using the Galleria mellonella (wax worm) model. When compared with the B. multivorans strains tested, B. cenocepacia K56-2 was highly virulent in this model and killed all worms within 24 h when injected at 10(7) c.f.u. B. multivorans clinical isolates C5568 and C0514 were significantly more virulent than the soil isolate ATCC 17616, which was avirulent even when worms were injected with 10(7) c.f.u. These results suggest strain differences in the virulence of B. multivorans isolates. PMID:23105020

Schmerk, Crystal L; Valvano, Miguel A

2013-02-01

207

Burkholderia Species Are Major Inhabitants of White Lupin Cluster Roots?†  

PubMed Central

The formation of cluster roots by plants represents a highly efficient strategy for acquisition of sparingly available phosphate. This particular root type is characterized by a densely branched structure and high exudation of organic acids and protons, which are likely to influence the resident bacterial community. Until now, the identity of the bacterial populations living in cluster roots has not been investigated. We applied cultivation-dependent and cultivation-independent methods to characterize the dominant bacterial genera inhabiting the growing cluster roots of white lupin. We observed a high relative abundance of Burkholderia species (up to 58% of all isolated strains and 44% of all retrieved 16S rRNA sequences) and a significant enrichment with increasing cluster root age. Most of the sequences retrieved clustered together with known plant- or fungus-associated Burkholderia species, while only one of 98 sequences was affiliated with the Burkholderia cepacia complex. In vitro assays revealed that Burkholderia strains were much more tolerant to low pH than non-Burkholderia strains. Moreover, many strains produced large amounts of siderophores and were able to utilize citrate and oxalate as carbon sources. These features seem to represent important traits for the successful colonization and maintenance of Burkholderia species in white lupin cluster roots. PMID:21908626

Weisskopf, Laure; Heller, Stefanie; Eberl, Leo

2011-01-01

208

Identification of Pseudomonas pseudomallei in clinical practice: use of simple screening tests and API 20NE  

Microsoft Academic Search

The API 20NE kit and a simple screening system involving Gram's stain, the oxidase reaction, colistin and gentamicin resistance, and colonial characteristics on a differential agar medium, were used to test 400 strains of Pseudomonas pseudomallei. The API kit identified 390 (97.5%) strains correctly on first testing and all but one of the remainder on second testing. Only one strain

D A Dance; V Wuthiekanun; P Naigowit; N J White

1989-01-01

209

VgrG-5 is a Burkholderia type VI secretion system-exported protein required for multinucleated giant cell formation and virulence.  

PubMed

The type VI secretion system (T6SS) has emerged as a critical virulence factor for the group of closely related Burkholderia spp. that includes Burkholderia pseudomallei, B. mallei, and B. thailandensis. While the genomes of these bacteria, referred to as the Bptm group, appear to encode several T6SSs, we and others have shown that one of these, type VI secretion system 5 (T6SS-5), is required for virulence in mammalian infection models. Despite its pivotal role in the pathogenesis of the Bptm group, the effector repertoire of T6SS-5 has remained elusive. Here we used quantitative mass spectrometry to compare the secretome of wild-type B. thailandensis to that of a mutant harboring a nonfunctional T6SS-5. This analysis identified VgrG-5 as a novel secreted protein whose export depends on T6SS-5 function. Bioinformatics analysis revealed that VgrG-5 is a specialized VgrG protein that harbors a C-terminal domain (CTD) conserved among Bptm group species. We found that a vgrG-5 ?CTD mutant is avirulent in mice and is unable to stimulate the fusion of host cells, a hallmark of the Bptm group previously shown to require T6SS-5 function. The singularity of VgrG-5 as a detected T6SS-5 substrate, taken together with the essentiality of its CTD for virulence, suggests that the protein is critical for the effector activity of T6SS-5. Intriguingly, we show that unlike the bacterial-cell-targeting T6SSs characterized so far, T6SS-5 localizes to the bacterial cell pole. We propose a model whereby the CTD of VgrG-5-, propelled by T6SS-5-, plays a key role in inducing membrane fusion, either by the recruitment of other factors or by direct participation. PMID:24452686

Schwarz, Sandra; Singh, Pragya; Robertson, Johanna D; LeRoux, Michele; Skerrett, Shawn J; Goodlett, David R; West, T Eoin; Mougous, Joseph D

2014-04-01

210

VgrG-5 Is a Burkholderia Type VI Secretion System-Exported Protein Required for Multinucleated Giant Cell Formation and Virulence  

PubMed Central

The type VI secretion system (T6SS) has emerged as a critical virulence factor for the group of closely related Burkholderia spp. that includes Burkholderia pseudomallei, B. mallei, and B. thailandensis. While the genomes of these bacteria, referred to as the Bptm group, appear to encode several T6SSs, we and others have shown that one of these, type VI secretion system 5 (T6SS-5), is required for virulence in mammalian infection models. Despite its pivotal role in the pathogenesis of the Bptm group, the effector repertoire of T6SS-5 has remained elusive. Here we used quantitative mass spectrometry to compare the secretome of wild-type B. thailandensis to that of a mutant harboring a nonfunctional T6SS-5. This analysis identified VgrG-5 as a novel secreted protein whose export depends on T6SS-5 function. Bioinformatics analysis revealed that VgrG-5 is a specialized VgrG protein that harbors a C-terminal domain (CTD) conserved among Bptm group species. We found that a vgrG-5 ?CTD mutant is avirulent in mice and is unable to stimulate the fusion of host cells, a hallmark of the Bptm group previously shown to require T6SS-5 function. The singularity of VgrG-5 as a detected T6SS-5 substrate, taken together with the essentiality of its CTD for virulence, suggests that the protein is critical for the effector activity of T6SS-5. Intriguingly, we show that unlike the bacterial-cell-targeting T6SSs characterized so far, T6SS-5 localizes to the bacterial cell pole. We propose a model whereby the CTD of VgrG-5—, propelled by T6SS-5—, plays a key role in inducing membrane fusion, either by the recruitment of other factors or by direct participation. PMID:24452686

Singh, Pragya; Robertson, Johanna D.; LeRoux, Michele; Skerrett, Shawn J.; Goodlett, David R.; West, T. Eoin; Mougous, Joseph D.

2014-01-01

211

42 CFR 73.9 - Responsible Official.  

Code of Federal Regulations, 2013 CFR

...neurotoxin producing species of Clostridium, Burkholderia mallei, Burkholderia pseudomallei, Francisella tularensis, Ebola viruses, Marburg virus, Variola major virus (Smallpox virus), Variola minor (Alastrim), or Yersinia pestis....

2013-10-01

212

77 FR 66850 - Public Workshop on Burkholderia: Exploring Current Issues and Identifying Regulatory Science Gaps  

Federal Register 2010, 2011, 2012, 2013

...of the bacteria to several classes of antibiotics. For example, a 20-year prospective...percent (Ref. 2). Prolonged courses of antibiotics are required to treat melioidosis...Ref. 5). Because of the lengthy antibiotic therapy required to treat...

2012-11-07

213

Burkholderia thailandensis oacA mutants facilitate the expression of Burkholderia mallei-like O polysaccharides.  

PubMed

Previous studies have shown that the O polysaccharides (OPS) expressed by Burkholderia mallei are similar to those produced by Burkholderia thailandensis except that they lack the 4-O-acetyl modifications on their 6-deoxy-?-l-talopyranosyl residues. In the present study, we describe the identification and characterization of an open reading frame, designated oacA, expressed by B. thailandensis that accounts for this phenomenon. Utilizing the B. thailandensis and B. mallei lipopolysaccharide (LPS)-specific monoclonal antibodies Pp-PS-W and 3D11, Western immunoblot analyses demonstrated that the LPS antigens expressed by the oacA mutant, B. thailandensis ZT0715, were antigenically similar to those produced by B. mallei ATCC 23344. In addition, immunoblot analyses demonstrated that when B. mallei ATCC 23344 was complemented in trans with oacA, it synthesized B. thailandensis-like LPS antigens. To elucidate the structure of the OPS moieties expressed by ZT0715, purified samples were analyzed via nuclear magnetic resonance spectroscopy. As predicted, these studies demonstrated that the loss of OacA activity influenced the O acetylation phenotype of the OPS moieties. Unexpectedly, however, the results indicated that the O methylation status of the OPS antigens was also affected by the loss of OacA activity. Nonetheless, it was revealed that the LPS moieties expressed by the oacA mutant reacted strongly with the B. mallei LPS-specific protective monoclonal antibody 9C1-2. Based on these findings, it appears that OacA is required for the 4-O acetylation and 2-O methylation of B. thailandensis OPS antigens and that ZT0715 may provide a safe and cost-effective source of B. mallei-like OPS to facilitate the synthesis of glanders subunit vaccine candidates. PMID:21115721

Brett, Paul J; Burtnick, Mary N; Heiss, Christian; Azadi, Parastoo; DeShazer, David; Woods, Donald E; Gherardini, Frank C

2011-02-01

214

Degradation of polyaromatic hydrocarbons by Burkholderia cepacia 2A-12  

Microsoft Academic Search

A new strain of bacterium degrading polyaromatic hydrocarbons (PAHs), Burkholderia cepacia 2A-12, was isolated from oil-contaminated soil. Of three PAHs, the isolated strain could utilize naphthalene (Nap) and phenanthrene (Phe) as a sole carbon source but not pyrene (Pyr). However, the strain could degrade Pyr when a cosubstrate such as yeast extract (YE) was supplemented. The PAH degradation rate of

Tae Jung Kim; Eun Young Lee; Youn Jung Kim; Kyung-Suk Cho; Hee Wook Ryu

2003-01-01

215

Nitrogen Fixation Genes in an Endosymbiotic Burkholderia Strain  

Microsoft Academic Search

In this paper we report the identification and characterization of a DNA region containing putative nif genes and belonging to a Burkholderia endosymbiont of the arbuscular mycorrhizal fungus Gigaspora margarita .A genomic library of total DNA extracted from the fungal spores was also representative of the bacterial genome and was used to investigate the prokaryotic genome. Screening of the library

DANIELA MINERDI; RENATO FANI; ROMINA GALLO; ALESSANDRA BOARINO; PAOLA BONFANTE

2001-01-01

216

Burkholderia bryophila sp. nov. and Burkholderia megapolitana sp. nov., moss-associated species with antifungal and plant-growth-promoting properties.  

PubMed

A polyphasic taxonomic study including DNA-DNA reassociation experiments and an extensive biochemical characterization was performed on 14 Burkholderia isolates from moss gametophytes of nutrient-poor plant communities on the southern Baltic Sea coast in northern Germany. The strains were classified within two novel species, for which the names Burkholderia bryophila sp. nov. and Burkholderia megapolitana sp. nov. are proposed. The former species also includes isolates from grassland and agricultural soil collected in previous studies. Strains Burkholderia bryophila 1S18(T) (=LMG 23644(T) =CCUG 52993(T)) and Burkholderia megapolitana A3(T) (=LMG 23650(T) =CCUG 53006(T)) are the proposed type strains. They were isolated from Sphagnum rubellum and Aulacomnium palustre, respectively, growing in the 'Ribnitzer Grosses Moor' nature reserve (Mecklenburg-Pommern, Germany). All moss isolates of both novel species showed antifungal activity against phytopathogens as well as plant-growth-promoting properties. PMID:17911288

Vandamme, Peter; Opelt, Katja; Knöchel, Nadine; Berg, Christian; Schönmann, Susan; De Brandt, Evie; Eberl, Leo; Falsen, Enevold; Berg, Gabriele

2007-10-01

217

Removal of Burkholderia cepacia biofilms with oxidants  

NASA Technical Reports Server (NTRS)

Iodine is used to disinfect the water system aboard US space shuttles and is the anticipated biocide for the international space station. Water quality on spacecraft must be maintained at the highest possible levels for the safety of the crew. Furthermore, the treatment process used to maintain the quality of water on research must be robust and operate for long periods with minimal crew intervention. Biofilms are recalcitrant and pose a major threat with regard to chronic contamination of spacecraft water systems. We measured the effectiveness of oxidizing biocides on the removal and regrowth of Burkholderia (Pseudomonas) cepacia biofilms. B. cepacia, isolated from the water distribution system of the space shuttle Discovery, was grown in continuous culture to produce a bacterial contamination source for biofilm formation and removal studies. A 10(7) CFU ml-1 B. cepacia suspension, in distilled water, was used to form biofilms on 3000 micrometers2 glass surfaces. Rates of attachment were measured directly with image analysis and were found to be 7.8, 15.2, and 22.8 attachment events h-1 for flow rates of 20.7, 15.2, and 9.8 ml min-1, respectively. After 18 h of formation, the B. cepacia biofilms were challenged with oxidants (ozone, chlorine, and iodine) and the rates of biofilm removal determined by image analysis. Fifty percent of the biofilm material was removed in the first hour of continous treatment with 24 mg l-1 chlorine or 2 mg l-1 ozone. Iodine (48 mg l-1) did not remove any measurable cellular material after 6 h continuous contact. After this first removal of biofilms by the oxidants, the surface was allowed to refoul and was again treated with the biocide. Iodine was the only compound that was unable to remove cellular debris from either primary or secondary biofilms. Moreover, treating primary biofilms with iodine increased the rate of formation of secondary biofilms, from 4.4 to 5.8 attachment events h-1. All the oxidants tested inactivated the B. cepacia associated with both primary and secondary biofilms. The amount of biocide needed to inactivate 50% of planktonic B. cepacia in 10 min at 25 degrees C was 8.4, 0.5, and 0.2 mg l-1 for iodine, chlorine, and ozone, respectively. The data suggest that iodine maynot be the best chemical for treating of biofilms when removal of cellular material is required.

Koenig, D. W.; Mishra, S. K.; Pierson, D. L.

1995-01-01

218

An ancient but promiscuous host-symbiont association between Burkholderia gut symbionts and their heteropteran hosts  

PubMed Central

Here, we investigated 124 stinkbug species representing 20 families and 5 superfamilies for their Burkholderia gut symbionts, of which 39 species representing 6 families of the superfamilies Lygaeoidea and Coreoidea were Burkholderia-positive. Diagnostic PCR surveys revealed high frequencies of Burkholderia infection in natural populations of the stinkbugs, and substantial absence of vertical transmission of Burkholderia infection to their eggs. In situ hybridization confirmed localization of the Burkholderia in their midgut crypts. In the lygaeoid and coreoid stinkbugs, development of midgut crypts in their alimentary tract was coincident with the Burkholderia infection, suggesting that the specialized morphological configuration is pivotal for establishment and maintenance of the symbiotic association. The Burkholderia symbionts were easily isolated as pure culture on standard microbiological media, indicating the ability of the gut symbionts to survive outside the host insects. Molecular phylogenetic analysis showed that the gut symbionts of the lygaeoid and coreoid stinkbugs belong to a ?-proteobacterial clade together with Burkholderia isolates from soil environments and Burkholderia species that induce plant galls. On the phylogeny, the stinkbug-associated, environmental and gall-forming Burkholderia strains did not form coherent groups, indicating host–symbiont promiscuity among these stinkbugs. Symbiont culturing revealed that slightly different Burkholderia genotypes often coexist in the same insects, which is also suggestive of host–symbiont promiscuity. All these results strongly suggest an ancient but promiscuous host–symbiont relationship between the lygaeoid/coreoid stinkbugs and the Burkholderia gut symbionts. Possible mechanisms as to how the environmentally transmitted promiscuous symbiotic association has been stably maintained in the evolutionary course are discussed. PMID:20882057

Kikuchi, Yoshitomo; Hosokawa, Takahiro; Fukatsu, Takema

2011-01-01

219

The role of cystic fibrosis conductance regulator in the clearance of Burkholderia cenocepacia by macrophages.  

E-print Network

??Burkholderia cenocepacia is an opportunistic Gram-negative bacterium that causes chronic respiratory infection in cystic fibrosis (CF) patients, leading to rapid decline of lung function. CF… (more)

Ostapska, Hanna

2012-01-01

220

Nodulation of Cyclopia spp. (Leguminosae, Papilionoideae) by Burkholderia tuberum  

PubMed Central

Background and Aims Species of the genus Burkholderia, from the Betaproteobacteria, have been isolated from legume nodules, but so far they have only been shown to form symbioses with species of Mimosa, sub-family Mimosoideae. This work investigates whether Burkholderia tuberum strains STM678 (isolated from Aspalathus carnosa) and DUS833 (from Aspalathus callosa) can nodulate species of the South African endemic papilionoid genera Cyclopia (tribe Podalyrieae) and Aspalathus (Crotalarieae) as well as the promiscuous legume Macroptilium atropurpureum (Phaseoleae). Method Bacterial strains and the phylogeny of their symbiosis-related (nod) genes were examined via 16S rRNA gene sequencing. Seedlings were grown in liquid culture and inoculated with one of the two strains of B. tuberum or with Sinorhizobium strain NGR 234 (from Lablab purpureus), Mesorhizobium strain DUS835 (from Aspalathus linearis) or Methylobacterium nodulans (from Crotalaria podocarpa). Some nodules, inoculated with green fluorescence protein (GFP)-tagged strains, were examined by light and electron microscopy coupled with immunogold labelling with a Burkholderia-specific antibody. The presence of active nitrogenase was checked by immunolabelling of nitrogenase and by the acetylene reduction assay. B. tuberum STM678 was also tested on a wide range of legumes from all three sub-families. Key Results Nodules were not formed on any of the Aspalathus spp. Only B. tuberum nodulated Cyclopia falcata, C. galioides, C. genistoides, C. intermedia and C. pubescens. It also effectively nodulated M. atropurpureum but no other species tested. GFP-expressing inoculant strains were located inside infected cells of C. genistoides, and bacteroids in both Cyclopia spp. and M. atropurpureum were immunogold-labelled with antibodies against Burkholderia and nitrogenase. Nitrogenase activity was also shown using the acetylene reduction assay. This is the first demonstration that a ?-rhizobial strain can effectively nodulate papilioinoid legumes. Conclusions Papilionoid legumes from widely different tribes can be nodulated by ?-rhizobia, forming both indeterminate (Cyclopia) and determinate (Macroptilium) nodules. PMID:17881339

Elliott, Geoffrey N.; Chen, Wen-Ming; Bontemps, Cyril; Chou, Jui-Hsing; Young, J. Peter W.; Sprent, Janet I.; James, Euan K.

2007-01-01

221

Draft Genome Sequence of the Lignin-Degrading Burkholderia sp. Strain LIG30, Isolated from Wet Tropical Forest Soil.  

PubMed

Burkholderia species are common soil Betaproteobacteria capable of degrading recalcitrant aromatic compounds and xenobiotics. Burkholderia sp. strain LIG30 was isolated from wet tropical forest soil and is capable of utilizing lignin as a sole carbon source. Here we report the draft genome sequence of Burkholderia sp. strain LIG30. PMID:24948777

Woo, Hannah L; Utturkar, Sagar; Klingeman, Dawn; Simmons, Blake A; DeAngelis, Kristen M; Brown, Steven D; Hazen, Terry C

2014-01-01

222

Draft Genome Sequence of the Lignin-Degrading Burkholderia sp. Strain LIG30, Isolated from Wet Tropical Forest Soil  

PubMed Central

Burkholderia species are common soil Betaproteobacteria capable of degrading recalcitrant aromatic compounds and xenobiotics. Burkholderia sp. strain LIG30 was isolated from wet tropical forest soil and is capable of utilizing lignin as a sole carbon source. Here we report the draft genome sequence of Burkholderia sp. strain LIG30. PMID:24948777

Woo, Hannah L.; Utturkar, Sagar; Klingeman, Dawn; Simmons, Blake A.; DeAngelis, Kristen M.; Brown, Steven D.

2014-01-01

223

Passive protection of diabetic rats with antisera specific for the polysaccharide portion of the lipopolysaccharide isolated from Pseudomonas pseudomallei  

PubMed Central

Polyclonal and monoclonal antisera raised to tetanus toxoid-conjugated polysaccharide of lipopolysaccharide (lps) and purified lps of Pseudomonas pseudomallei that reacted with a collection of 41 strains of this bacterium from 23 patients are described. The common antigen recognized by these sera was within the polysaccharide component of the lps of the cells. The sera were specific for P pseudomallei in that none of 37 strains of other bacteria, including 20 Gram-negative and three Gram-positive species, were recognized, although cross-reaction occurred using the anticonjugate serum with some strains of Pseudomonas cepacia serotype A, a closely related bacterium. Passive protection studies using a diabetic rat model of P pseudomallei infection showed that partially purified rabbit polyclonal and mouse monoclonal antisera were protective when the median lethal dose was raised by four to five orders of magnitude. The wide distribution of the polysaccharide antigen among isolates of P pseudomallei used in this study and the protective role of antibody to the conjugated polysaccharide antigen suggest potential as a vaccine. PMID:22346496

Bryan, Larry E; Wong, Sallene; Woods, Don E; Dance, David AB; Chaowagul, W

1994-01-01

224

Immunoproteomic Analysis of Proteins Expressed by Two Related Pathogens, Burkholderia multivorans and Burkholderia cenocepacia, during Human Infection  

PubMed Central

Burkholderia cepacia complex (Bcc) is an opportunistic bacterial pathogen that causes chronic infections in people with cystic fibrosis (CF). It is a highly antibiotic resistant organism and Bcc infections are rarely cleared from patients, once they are colonized. The two most clinically relevant species within Bcc are Burkholderia cenocepacia and Burkholderia multivorans. The virulence of these pathogens has not been fully elucidated and the virulence proteins expressed during human infection have not been identified to date. Furthermore, given its antibiotic resistance, prevention of infection with a prophylactic vaccine may represent a better alternative than eradication of an existing infection. We have compared the immunoproteome of two strains each from these two species of Bcc, with the aim of identifying immunogenic proteins which are common to both species. Fourteen immunoreactive proteins were exclusive to both B. cenocepacia strains, while 15 were exclusive to B. multivorans. A total of 15 proteins were immunogenic across both species. DNA-directed RNA polymerase, GroEL, 38kDa porin and elongation factor-Tu were immunoreactive proteins expressed by all four strains examined. Many proteins which were immunoreactive in both species, warrant further investigations in order to aid in the elucidation of the mechanisms of pathogenesis of this difficult organism. In addition, identification of some of these could also allow the development of protective vaccines which may prevent colonisation. PMID:24260482

Shinoy, Minu; Dennehy, Ruth; Coleman, Lorraine; Carberry, Stephen; Schaffer, Kirsten; Callaghan, Maire; Doyle, Sean; McClean, Siobhan

2013-01-01

225

Genome characterization of a novel Burkholderia cepacia complex genomovar isolated from dieback affected mango orchards.  

PubMed

We characterized the genome of the antibiotic resistant, caseinolytic and non-hemolytic Burkholderia sp. strain TJI49, isolated from mango trees (Mangifera indica L.) with dieback disease. This isolate produced severe disease symptoms on the indicator plants. Next generation DNA sequencing and short-read assembly generated the 60X deep 7,631,934 nucleotide draft genome of Burkholderia sp. TJI49 which comprised three chromosomes and at least one mega plasmid. Genome annotation studies revealed a total 8,992 genes, out of which 8,940 were protein coding genes. Comparative genomics and phylogenetics identified Burkholderia sp. TJI49 as a distinct species of Burkholderia cepacia complex (BCC), closely related to B. multivorans ATCC17616. Genome-wide sequence alignment of this isolate with replicons of BCC members showed conservation of core function genes but considerable variations in accessory genes. Subsystem-based gene annotation identified the active presence of wide spread colonization island and type VI secretion system in Burkholderia sp. TJI49. Sequence comparisons revealed (a) 28 novel ORFs that have no database matches and (b) 23 ORFs with orthologues in species other than Burkholderia, indicating horizontal gene transfer events. Fold recognition of novel ORFs identified genes encoding pertactin autotransporter-like proteins (a constituent of type V secretion system) and Hap adhesion-like proteins (involved in cell-cell adhesion) in the genome of Burkholderia sp. TJI49. The genomic characterization of this isolate provided additional information related to the 'pan-genome' of Burkholderia species. PMID:23653265

Khan, Asifullah; Asif, Huma; Studholme, David J; Khan, Ishtiaq A; Azim, M Kamran

2013-11-01

226

Exploring the HME and HAE1 efflux systems in the genus Burkholderia  

PubMed Central

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 HAE1 systems in the Burkholderia genus. Conclusion A complete knowledge of the presence and distribution of RND proteins in Burkholderia species was obtained and an evolutionary model was depicted. Data presented in this work may serve as a basis for future experimental tests, focused especially on HAE1 proteins, aimed at the identification of novel targets in antimicrobial therapy against Burkholderia species. PMID:20525265

2010-01-01

227

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

EPA Science Inventory

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 ...

228

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

EPA Science Inventory

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...

229

Conserved and Hybrid meta-Cleavage Operons from PAH-degrading Burkholderia RP007  

Microsoft Academic Search

We have compared the sequence and gene order of meta-cleavage pathway operons from ?- and ?-subgroups of the Proteobacteria with operons from Burkholderia sp. strain RP007 which belongs to the ?-subgroup of the Proteobacteria. Burkholderia RP007 was isolated for its ability to degrade phenanthrene and contains two meta-cleavage operons. One exhibits a comparable gene order to previously characterised ?-subgroup Proteobacterial

Andrew D. Laurie; Gareth Lloyd-Jones

1999-01-01

230

Occurrence of Multiple Genomovars of Burkholderia cepacia in Cystic Fibrosis Patients and Proposal of  

E-print Network

We performed an integrated genotypic and phenotypic analysis of 128 strains of the genera Burkholderia, Rabtonia, and Pseudomonas in order to study the taxonomic structure of Burkholderia cepacia and its relationships with other Burkholderia species. Our data show that presumed B. cepacia strains isolated from cystic fibrosis patients belong to at least five distinct genomic species, one of which was identified as Burkholderia vietnamiensis. This group of five phenotypically similar species is referred to as the B. cepacia complex. The name Burkholderia multivorans is proposed for one of these genomic species, which was formerly referred to as B. cepacia genomovar 11; the remaining B. cepacia groups are referred to as genomovars I, 111, and lV, pending additional differential phenotypic tests. The role and pathogenic potential of each of these taxa, particularly in view of the potentially fatal infections in cystic fibrosis patients, need further evaluation. The data presented also demonstrate that Pseudomonas glathei and Pseudomonas pyrrocinia should be reclassified as Burkholderia species. The taxonomy and the description of the genus Pseudomonus have been changing for more than a century (16). The discovery of five major phylogenetic lineages has played a major role (23) and was at the origin of the description of

Burkholderia Multivorans Sp. Nov; H. Revets; S. Lauwers; M. Gillis; L K. Kersters; J. R. W. Govan

231

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

PubMed Central

Burkholderia legume symbionts (also called ?-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, ?-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 ? to ?-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

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

2013-01-01

232

Coexistence of Burkholderia, Cupriavidus, and Rhizobium sp. Nodule Bacteria on two Mimosa spp. in Costa Rica  

PubMed Central

rRNA gene sequencing and PCR assays indicated that 215 isolates of root nodule bacteria from two Mimosa species at three sites in Costa Rica belonged to the genera Burkholderia, Cupriavidus, and Rhizobium. This is the first report of Cupriavidus sp. nodule symbionts for Mimosa populations within their native geographic range in the neotropics. Burkholderia spp. predominated among samples from Mimosa pigra (86% of isolates), while there was a more even distribution of Cupriavidus, Burkholderia, and Rhizobium spp. on Mimosa pudica (38, 37, and 25% of isolates, respectively). All Cupriavidus and Burkholderia genotypes tested formed root nodules and fixed nitrogen on both M. pigra and M. pudica, and sequencing of rRNA genes in strains reisolated from nodules verified identity with inoculant strains. Inoculation tests further indicated that both Cupriavidus and Burkholderia spp. resulted in significantly higher plant growth and nodule nitrogenase activity (as measured by acetylene reduction assays) relative to plant performance with strains of Rhizobium. Given the prevalence of Burkholderia and Cupriavidus spp. on these Mimosa legumes and the widespread distribution of these plants both within and outside the neotropics, it is likely that both ?-proteobacterial genera are more ubiquitous as root nodule symbionts than previously believed. PMID:16461667

Barrett, Craig F.; Parker, Matthew A.

2006-01-01

233

Diazotrophic Burkholderia Species Associated with Field-Grown Maize and Sugarcane  

PubMed Central

Until recently, diazotrophy was known in only one of the 30 formally described species of Burkholderia. Novel N2-fixing plant-associated Burkholderia species such as B. unamae, B. tropica, and B. xenovorans have been described, but their environmental distribution is scarcely known. In the present study, the occurrence of N2-fixing Burkholderia species associated with different varieties of sugarcane and maize growing in regions of Mexico and Brazil was analyzed. Only 111 out of more than 900 isolates recovered had N2-fixing ability as demonstrated by the acetylene reduction assay. All 111 isolates also yielded a PCR product with primers targeting the nifH gene, which encodes a key enzyme in the process of nitrogen fixation. These 111 isolates were confirmed as belonging to the genus Burkholderia by using a new 16S rRNA-specific primer pair for diazotrophic species (except B. vietnamiensis) and closely related nondiazotrophic Burkholderia. In Mexico, many isolates of B. unamae (predominantly associated with sugarcane) and B. tropica (more often associated with maize) were recovered. However, in Brazil B. tropica was not identified among the isolates analyzed, and only a few B. unamae isolates were recovered from one sugarcane variety. Most Brazilian diazotrophic Burkholderia isolates (associated with both sugarcane and maize plants) belonged to a novel species, as revealed by amplified 16S rRNA gene restriction profiles, 16S rRNA gene sequencing, and protein electrophoresis. In addition, transmissibility factors such as the cblA and esmR genes, identified among clinical and environmental isolates of opportunistic pathogens of B. cenocepacia and other species of the B. cepacia complex, were not detected in any of the plant-associated diazotrophic Burkholderia isolates analyzed. PMID:16672447

Perin, L.; Martínez-Aguilar, L.; Castro-González, R.; Estrada-de los Santos, P.; Cabellos-Avelar, T.; Guedes, H. V.; Reis, V. M.; Caballero-Mellado, J.

2006-01-01

234

Screening a mushroom extract library for activity against Acinetobacter baumannii and Burkholderia cepacia and the identification of a compound with anti-Burkholderia activity  

Microsoft Academic Search

BACKGROUND: Acinetobacter baumannii and species within the Burkholderia cepacia complex (BCC) are significant opportunistic bacterial pathogens of humans. These species exhibit a high degree of antibiotic resistance, and some clinical isolates are resistant to all currently available antimicrobial drugs used for treatment. Thus, new drugs are needed to treat infections by these species. Mushrooms could be a potential source for

William R Schwan; Craig Dunek; Michael Gebhardt; Kathleen Engelbrecht; Tiffany Klett; Aaron Monte; Joseph Toce; Marc Rott; Thomas J Volk; John J LiPuma; Xue-Ting Liu; Ronald McKelvey

2010-01-01

235

Oxalotrophy, a widespread trait of plant-associated Burkholderia species, is involved in successful root colonization of lupin and maize by Burkholderia phytofirmans  

PubMed Central

Plant roots and shoots harbor complex bacterial communities. Early seed and plantlet colonization plays a key role in determining which bacterial populations will successfully invade plant tissues, yet the mechanisms enabling plants to select for beneficial rather than harmful populations are largely unknown. In this study, we demonstrate a role of oxalate as a determinant in this selection process, using members of the genus Burkholderia as model organisms. Oxalotrophy, i.e., the ability to use oxalate as a carbon source, was found to be a property strictly associated with plant-beneficial species of the Burkholderia genus, while plant pathogenic (B. glumae, B. plantarii) or human opportunistic pathogens (Burkholderia cepacia complex strains) were unable to degrade oxalate. We further show that oxalotrophy is required for successful plant colonization by the broad host endophyte Burkholderia phytofirmans PsJN: an engineered ?oxc mutant, which lost the ability to grow on oxalate, was significantly impaired in early colonization of both lupin and maize compared with the wild-type. This work suggests that in addition to the role of oxalate in heavy metal tolerance of plants and in virulence of phytopathogenic fungi, it is also involved in specifically recruiting plant-beneficial members from complex bacterial communities. PMID:24409174

Kost, Thomas; Stopnisek, Nejc; Agnoli, Kirsty; Eberl, Leo

2014-01-01

236

Draft Genome Sequence of Burkholderia pyrrocinia Lyc2, a Biological Control Strain That Can Suppress Multiple Plant Microbial Pathogens.  

PubMed

Burkholderia pyrrocinia strain Lyc2 was isolated from the tobacco rhizosphere in China. This bacterium exhibits a remarkable capacity to inhibit the growth of multiple pathogens and shows strong suppression of cotton seedling damping-off. Here, we present the draft genome sequence of Burkholderia pyrrocinia strain Lyc2. PMID:25278535

Wang, Xiao-Qiang; Showmaker, Kurt C; Yu, Xiao-Qing; Bi, Tao; Hsu, Chuan-Yu; Baird, Sonya M; Peterson, Daniel G; Li, Xiang-Dong; Lu, Shi-En

2014-01-01

237

Draft Genome Sequence of Burkholderia pyrrocinia Lyc2, a Biological Control Strain That Can Suppress Multiple Plant Microbial Pathogens  

PubMed Central

Burkholderia pyrrocinia strain Lyc2 was isolated from the tobacco rhizosphere in China. This bacterium exhibits a remarkable capacity to inhibit the growth of multiple pathogens and shows strong suppression of cotton seedling damping-off. Here, we present the draft genome sequence of Burkholderia pyrrocinia strain Lyc2. PMID:25278535

Wang, Xiao-Qiang; Showmaker, Kurt C.; Yu, Xiao-Qing; Bi, Tao; Hsu, Chuan-Yu; Baird, Sonya M.; Peterson, Daniel G.

2014-01-01

238

Genome Annotation of Burkholderia sp. SJ98 with Special Focus on Chemotaxis Genes  

PubMed Central

Burkholderia sp. strain SJ98 has the chemotactic activity towards nitroaromatic and chloronitroaromatic compounds. Recently our group published draft genome of strain SJ98. In this study, we further sequence and annotate the genome of stain SJ98 to exploit the potential of this bacterium. We specifically annotate its chemotaxis genes and methyl accepting chemotaxis proteins. Genome of Burkholderia sp. SJ98 was annotated using PGAAP pipeline that predicts 7,268 CDSs, 52 tRNAs and 3 rRNAs. Our analysis based on phylogenetic and comparative genomics suggest that Burkholderia sp. YI23 is closest neighbor of the strain SJ98. The genes involved in the chemotaxis of strain SJ98 were compared with genes of closely related Burkholderia strains (i.e. YI23, CCGE 1001, CCGE 1002, CCGE 1003) and with well characterized bacterium E. coli K12. It was found that strain SJ98 has 37 che genes including 19 methyl accepting chemotaxis proteins that involved in sensing of different attractants. Chemotaxis genes have been found in a cluster along with the flagellar motor proteins. We also developed a web resource that provides comprehensive information on strain SJ98 that includes all analysis data (http://crdd.osdd.net/raghava/genomesrs/burkholderia/). PMID:23940608

Kumar, Shailesh; Vikram, Surendra; Raghava, Gajendra Pal Singh

2013-01-01

239

Genetic Diversity of Burkholderia contaminans Isolates from Cystic Fibrosis Patients in Argentina  

PubMed Central

A total of 120 Burkholderia cepacia complex isolates collected during 2004–2010 from 66 patients in two cystic fibrosis reference centers in Argentina were analyzed. Burkholderia contaminans was the species most frequently recovered (57.6%), followed by Burkholderia cenocepacia (15%), a species distribution not reported so far. The recA-PCR-based techniques applied to the B. contaminans isolates revealed that 85% of the population carried the recA-ST-71 allele. Our results showed the utility of BOX-PCR genotyping in analyzing B. contaminans diversity. This approach allowed us to address clonal transmission during an outbreak and the genetic changes occurring in infecting bacteria over the course of chronic infection. PMID:23135937

Martina, Pablo; Bettiol, Marisa; Vescina, Cecilia; Montanaro, Patricia; Mannino, M. Constanza; Prieto, Claudia I.; Vay, Carlos; Naumann, Dieter; Schmitt, Juergen; Yantorno, Osvaldo; Lagares, Antonio

2013-01-01

240

Regulation of biofilm formation in Pseudomonas and Burkholderia species.  

PubMed

In the present review, we describe and compare the molecular mechanisms that are involved in the regulation of biofilm formation by Pseudomonas putida, Pseudomonas fluorescens, Pseudomonas aeruginosa and Burkholderia cenocepacia. Our current knowledge suggests that biofilm formation is regulated by cyclic diguanosine-5'-monophosphate (c-di-GMP), small RNAs (sRNA) and quorum sensing (QS) in all these bacterial species. The systems that employ c-di-GMP as a second messenger regulate the production of exopolysaccharides and surface proteins which function as extracellular matrix components in the biofilms formed by the bacteria. The systems that make use of sRNAs appear to regulate the production of exopolysaccharide biofilm matrix material in all these species. In the pseudomonads, QS regulates the production of extracellular DNA, lectins and biosurfactants which all play a role in biofilm formation. In B.cenocepacia QS regulates the expression of a large surface protein, lectins and extracellular DNA that all function as biofilm matrix components. Although the three regulatory systems all regulate the production of factors used for biofilm formation, the molecular mechanisms involved in transducing the signals into expression of the biofilm matrix components differ between the species. Under the conditions tested, exopolysaccharides appears to be the most important biofilm matrix components for P.aeruginosa, whereas large surface proteins appear to be the most important biofilm matrix components for P.putida, P.fluorescens, and B.cenocepacia. PMID:24592823

Fazli, Mustafa; Almblad, Henrik; Rybtke, Morten Levin; Givskov, Michael; Eberl, Leo; Tolker-Nielsen, Tim

2014-07-01

241

Binding of protegrin-1 to Pseudomonas aeruginosa and Burkholderia cepacia  

PubMed Central

Background Pseudomonas aeruginosa and Burkholderia cepacia infections of cystic fibrosis patients' lungs are often resistant to conventional antibiotic therapy. Protegrins are antimicrobial peptides with potent activity against many bacteria, including P. aeruginosa. The present study evaluates the correlation between protegrin-1 (PG-1) sensitivity/resistance and protegrin binding in P. aeruginosa and B. cepacia. Methods The PG-1 sensitivity/resistance and PG-1 binding properties of P. aeruginosa and B. cepacia were assessed using radial diffusion assays, radioiodinated PG-1, and surface plasmon resonance (BiaCore). Results The six P. aeruginosa strains examined were very sensitive to PG-1, exhibiting minimal active concentrations from 0.0625–0.5 ?g/ml in radial diffusion assays. In contrast, all five B. cepacia strains examined were greater than 10-fold to 100-fold more resistant, with minimal active concentrations ranging from 6–10 ?g/ml. When incubated with a radioiodinated variant of PG-1, a sensitive P. aeruginosa strain bound considerably more protegrin molecules per cell than a resistant B. cepacia strain. Binding/diffusion and surface plasmon resonance assays revealed that isolated lipopolysaccharide (LPS) and lipid A from the sensitive P. aeruginosa strains bound PG-1 more effectively than LPS and lipid A from resistant B. cepacia strains. Conclusion These findings support the hypothesis that the relative resistance of B. cepacia to protegrin is due to a reduced number of PG-1 binding sites on the lipid A moiety of its LPS. PMID:11980587

Albrecht, Mark T; Wang, Wei; Shamova, Olga; Lehrer, Robert I; Schiller, Neal L

2002-01-01

242

RNAseq-based Transcriptome Analysis of Burkholderia glumae Quorum Sensing  

PubMed Central

Burkholderia glumae causes rice grain rot and sheath rot by producing toxoflavin, the expression of which is regulated by quorum sensing (QS). The QS systems of B. glumae rely on N-octanoyl homoserine lactone, synthesized by TofI and its cognate receptor TofR, to activate the genes for toxoflavin biosynthesis and an IclR-type transcriptional regulator gene, qsmR. To understand genome-wide transcriptional profiling of QS signaling, we employed RNAseq of the wild-type B. glumae BGR1 with QS-defective mutant, BGS2 (BGR1 tofI::?) and QS-dependent transcriptional regulator mutant, BGS9 (BGR1 qsmR::?). A comparison of gene expression profiling among the wild-type BGR1 and the two mutants before and after QS onset as well as gene ontology (GO) enrichment analysis from differential expressed genes (DEGs) revealed that genes involved in motility were highly enriched in TofI-dependent DEGs, whereas genes for transport and DNA polymerase were highly enriched in QsmR-dependent DEGs. Further, a combination of pathways with these DEGs and phenotype analysis of mutants pointed to a couple of metabolic processes, which are dependent on QS in B. glumae, that were directly or indirectly related with bacterial motility. The consistency of observed bacterial phenotypes with GOs or metabolic pathways in QS-regulated genes implied that integration RNAseq with GO enrichment or pathways would be useful to study bacterial physiology and phenotypes.

Kim, Sunyoung; Park, Jungwook; Kim, Ji Hyeon; Lee, Jongyun; Bang, Bongjun; Hwang, Ingyu; Seo, Young-Su

2013-01-01

243

Monitoring Therapeutic Treatments against Burkholderia Infections Using Imaging Techniques  

PubMed Central

Burkholderia mallei, the etiologic agent of glanders, are Category B select agents with biothreat potential, and yet effective therapeutic treatments are lacking. In this study, we showed that CpG administration increased survival, demonstrating protection in the murine glanders model. Bacterial recovery from infected lungs, liver and spleen was significantly reduced in CpG-treated animals as compared with non-treated mice. Reciprocally, lungs of CpG-treated infected animals were infiltrated with higher levels of neutrophils and inflammatory monocytes, as compared to control animals. Employing the B. mallei bioluminescent strain CSM001 and the Neutrophil-Specific Fluorescent Imaging Agent, bacterial dissemination and neutrophil trafficking were monitored in real-time using multimodal in vivo whole body imaging techniques. CpG-treatment increased recruitment of neutrophils to the lungs and reduced bioluminescent bacteria, correlating with decreased bacterial burden and increased protection against acute murine glanders. Our results indicate that protection of CpG-treated animals was associated with recruitment of neutrophils prior to infection and demonstrated, for the first time, simultaneous real time in vivo imaging of neutrophils and bacteria. This study provides experimental evidence supporting the importance of incorporating optimized in vivo imaging methods to monitor disease progression and to evaluate the efficacy of therapeutic treatment during bacterial infections. PMID:24349761

Mott, Tiffany M.; Johnston, R. Katie; Vijayakumar, Sudhamathi; Estes, D. Mark; Motamedi, Massoud; Sbrana, Elena; Endsley, Janice J.; Torres, Alfredo G.

2013-01-01

244

Identification of quorum sensing-controlled genes in Burkholderia ambifaria  

PubMed Central

The Burkholderia cepacia complex (Bcc) comprises strains with a virulence potential toward immunocompromised patients as well as plant growth–promoting rhizobacteria (PGPR). Owing to the link between quorum sensing (QS) and virulence, most studies among Bcc species have been directed toward QS of pathogenic bacteria. We have investigated the QS of B. ambifaria, a PGPR only infrequently recovered from patients. The cepI gene, responsible for the synthesis of the main signaling molecule N-octanoylhomoserine lactone (C8-HSL), was inactivated. Phenotypes of the B. ambifaria cepI mutant we observed, such as increased production of siderophores and decreased proteolytic and antifungal activities, are in agreement with those of other Bcc cepI mutants. The cepI mutant was then used as background strain for a whole-genome transposon-insertion mutagenesis strategy, allowing the identification of 20 QS-controlled genes, corresponding to 17 loci. The main functions identified are linked to antifungal and antimicrobial properties, as we have identified QS-controlled genes implicated in the production of pyrrolnitrin, burkholdines (occidiofungin-like molecules), and enacyloxins. This study provides insights in the QS-regulated functions of a PGPR, which could lead to beneficial potential biotechnological applications. PMID:23382083

Chapalain, Annelise; Vial, Ludovic; Laprade, Natacha; Dekimpe, Valerie; Perreault, Jonathan; Deziel, Eric

2013-01-01

245

Burkholderia cenocepacia zinc metalloproteases influence resistance to antimicrobial peptides.  

PubMed

Burkholderia cenocepacia secretes two zinc-dependent metalloproteases, designated ZmpA and ZmpB. Previously, ZmpA and ZmpB have been shown to cleave several proteins important in host defence. In this study, the ability of ZmpA and ZmpB to digest and inactivate antimicrobial peptides involved in innate immunity was examined. ZmpB but not ZmpA cleaved beta-defensin-1. ZmpA but not ZmpB cleaved the cathelicidin LL-37. Both enzymes cleaved elafin and secretory leukocyte inhibitor, which are antimicrobial peptides as well as neutrophil elastase inhibitors. Both ZmpA and ZmpB cleaved protamine, a fish antimicrobial peptide, and a zmpA zmpB mutant was more sensitive to protamine killing than the parental strain. ZmpA or ZmpB cleavage of elafin inactivated its anti-protease activity. The effect of ZmpA and ZmpB on the neutrophil proteases elastase and cathepsin G was also examined but neither enzyme was active against these host proteases. These studies suggest that ZmpA and ZmpB may influence the resistance of B. cenocepacia to host antimicrobial peptides as well as alter the host protease/anti-protease balance in chronic respiratory infections. PMID:19542010

Kooi, Cora; Sokol, Pamela A

2009-09-01

246

Burkholderia, a Genus Rich in Plant-Associated Nitrogen Fixers with Wide Environmental and Geographic Distribution  

PubMed Central

The genus Burkholderia comprises 19 species, including Burkholderia vietnamiensis which is the only known N2-fixing species of this bacterial genus. The first isolates of B. vietnamiensis were recovered from the rhizosphere of rice plants grown in a phytotron, but its existence in natural environments and its geographic distribution were not reported. In the present study, most N2-fixing isolates recovered from the environment of field-grown maize and coffee plants cultivated in widely separated regions of Mexico were phenotypically identified as B. cepacia using the API 20NE system. Nevertheless, a number of these isolates recovered from inside of maize roots, as well as from the rhizosphere and rhizoplane of maize and coffee plants, showed similar or identical features to those of B. vietnamiensis TVV75T. These features include nitrogenase activity with 10 different carbon sources, identical or very similar nifHDK hybridization patterns, very similar protein electrophoregrams, identical amplified 16S rDNA restriction (ARDRA) profiles, and levels of DNA-DNA reassociation higher than 70% with total DNA from strain TVV75T. Although the ability to fix N2 is not reported to be a common feature among the known species of the genus Burkholderia, the results obtained show that many diazotrophic Burkholderia isolates analyzed showed phenotypic and genotypic features different from those of the known N2-fixing species B. vietnamiensis as well as from those of B. kururiensis, a bacterium identified in the present study as a diazotrophic species. DNA-DNA reassociation assays confirmed the existence of N2-fixing Burkholderia species different from B. vietnamiensis. In addition, this study shows the wide geographic distribution and substantial capability of N2-fixing Burkholderia spp. for colonizing diverse host plants in distantly separated environments. PMID:11375196

Estrada-De Los Santos, Paulina; Bustillos-Cristales, Rocio; Caballero-Mellado, Jesus

2001-01-01

247

Expression of Caenorhabditis elegans antimicrobial peptide NLP-31 in Escherichia coli  

NASA Astrophysics Data System (ADS)

Burkholderia pseudomallei is the causative agent of melioidosis, a fulminant disease endemic in Southeast Asia and Northern Australia. The standardized form of therapy is antibiotics treatment; however, the bacterium has become increasingly resistant to these antibiotics. This has spurred the need to search for alternative therapeutic agents. Antimicrobial peptides (AMPs) are small proteins that possess broad-spectrum antimicrobial activity. In a previous study, the nematode Caenorhabditis elegans was infected by B. pseudomallei and a whole animal transcriptome analysis identified a number of AMP-encoded genes which were induced significantly in the infected worms. One of the AMPs identified is NLP-31 and to date, there are no reports of anti-B. pseudomallei activity demonstrated by NLP-31. To produce NLP-31 protein for future studies, the gene encoding for NLP-31 was cloned into the pET32b expression vector and transformed into Escherichia coli BL21(DE3). Protein expression was induced with 1 mM IPTG for 20 hours at 20°C and recombinant NLP-31 was detected in the soluble fraction. Taken together, a simple optimized heterologous production of AMPs in an E. coli expression system has been successfully developed.

Lim, Mei-Perng; Nathan, Sheila

2014-09-01

248

Mixotrophic metabolism in Burkholderia kururiensis subsp. thiooxydans subsp. nov., a facultative chemolithoautotrophic thiosulfate oxidizing bacterium isolated from rhizosphere soil and proposal for classification of the type strain of Burkholderia kururiensis as Burkholderia kururiensis subsp. kururiensis subsp. nov  

Microsoft Academic Search

A thiosulfate-oxidizing facultative chemolithoautotrophic Burkholderia sp. strain ATSB13T was previously isolated from rhizosphere soil of tobacco plant. Strain ATSB13T was aerobic, Gram-staining-negative, rod shaped and motile by means of sub-terminal flagellum. Strain ATSB13T exhibited mixotrophic growth in a medium containing thiosulfate plus acetate. A phylogenetic study based on 16S rRNA gene\\u000a sequence analysis indicated that strain ATSB13T was most closely

Rangasamy Anandham; Pandiyan Indira Gandhi; Soon Wo Kwon; Tong Min Sa; Yong Ki Kim; Hyeong Jin Jee

2009-01-01

249

A prospective comparison of co-amoxiclav and the combination of chloramphenicol, doxycycline, and co-trimoxazole for the oral maintenance treatment of melioidosis  

Microsoft Academic Search

An open randomized comparison of the oral ‘conventional’ regimen (combination of chloramphenicol, cotrimoxazole and doxycycline) and co-amoxiclav for the maintenance treatment of melioidosis was conducted in Ubon Ratchatani, north-eastern Thailand, between 1989 and 1992. The total antibiotic treatment duration was 20 weeks. Of 101 patients followed, 10 (10%; 95% confidence interval [CI] 4·9–17·5%) subsequently relapsed: 2 of 52 patients (4%)

A. Rajchanuvong; W. Chaowagul; Y. Suputtamongkol; M. D. Smith; D. A. B. Dance; N. J. White

1995-01-01

250

Immune Cell Activation in Melioidosis: Increased Serum Levels of Interferon-? and Soluble Interleukin2 Receptors without Change in Soluble CD8 Protein  

Microsoft Academic Search

To evaluate immune cell activation in patients with melioidosis, serum samples were assayed for interferon-? (IFN-?), soluble interleukin-2 receptors (sIL-2R), and soluble CD8 protein (sCD8). Forty patients with sepsis (23 fatal cases, 17 survivors) and 13 with localized disease were studied during acute illness; 12 additional patients were studied after discharge while on maintenance antimicrobial therapy. Serum concentrations of IFN-?

Arthur E. Brown; David A. B. Dance; Yupin Suputtamongkol; Wipada Chaowagul; Somchai Kongchareon; H. Kyle Webster; Nicholas J. White

1991-01-01

251

Toluene 2-Monooxygenase-Dependent Growth of Burkholderia cepacia G4/PR1 on Diethyl Ether  

PubMed Central

Aerobic bacterial growth on aromatic hydrocarbons typically requires oxygenase enzymes, which are known to fortuitously oxidize nongrowth substrates. In this study, we found that oxidation of diethyl ether by toluene 2-monooxygenase supported more rapid growth of Burkholderia cepacia G4/PR1 than did the aromatic substrates n-propylbenzene and o-xylene. The wild-type Burkholderia cepacia G4 failed to grow on diethyl ether. Purified toluene 2-monooxygenase protein components oxidized diethyl ether stoichiometrically to ethanol and acetaldehyde. Butyl methyl ether, diethyl sulfide, and 2-chloroethyl ethyl ether were oxidized by B. cepacia G4/PR1. PMID:16535583

Hur, H.; Newman, L. M.; Wackett, L. P.; Sadowsky, M. J.

1997-01-01

252

Pathogens penetrating the central nervous system: infection pathways and the cellular and molecular mechanisms of invasion.  

PubMed

The brain is well protected against microbial invasion by cellular barriers, such as the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB). In addition, cells within the central nervous system (CNS) are capable of producing an immune response against invading pathogens. Nonetheless, a range of pathogenic microbes make their way to the CNS, and the resulting infections can cause significant morbidity and mortality. Bacteria, amoebae, fungi, and viruses are capable of CNS invasion, with the latter using axonal transport as a common route of infection. In this review, we compare the mechanisms by which bacterial pathogens reach the CNS and infect the brain. In particular, we focus on recent data regarding mechanisms of bacterial translocation from the nasal mucosa to the brain, which represents a little explored pathway of bacterial invasion but has been proposed as being particularly important in explaining how infection with Burkholderia pseudomallei can result in melioidosis encephalomyelitis. PMID:25278572

Dando, Samantha J; Mackay-Sim, Alan; Norton, Robert; Currie, Bart J; St John, James A; Ekberg, Jenny A K; Batzloff, Michael; Ulett, Glen C; Beacham, Ifor R

2014-10-01

253

42 CFR 73.6 - Exemptions for overlap select agents and toxins.  

Code of Federal Regulations, 2013 CFR

...of any of the following overlap select agents or toxins must be immediately reported by telephone, facsimile, or e-mail: Bacillus anthracis, Burkholderia mallei and Burkholderia pseudomallei . This report must be followed by submission of...

2013-10-01

254

Investigating early stages of biocorrosion with XPS: AISI 304 stainless steel exposed to Burkholderia species  

Microsoft Academic Search

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

Leena-Sisko Johansson; Tuomas Saastamoinen

1999-01-01

255

Drosophila melanogaster as a Model Host for the Burkholderia cepacia Complex  

E-print Network

Drosophila melanogaster as a Model Host for the Burkholderia cepacia Complex Jose´e Castonguay: Castonguay-Vanier J, Vial L, Tremblay J, De´ziel E (2010) Drosophila melanogaster as a Model Host with vertebrate organisms. Methodology/Principal Findings: The aim of this study was to establish Drosophila

Paris-Sud XI, Université de

256

Emergence of Burkholderia cepacia in Honolulu: A Case of Nursing Home-acquired B. cepacia sepsis  

PubMed Central

Burkholderia cepacia has rarely been reported in Honolulu. Its emergence as a nursing home-acquired pathogen with high mortality rate is concerning. This case report describes a local nursing home patient who was diagnosed with B. cepacia sepsis in 2012. PMID:24069571

Hua, Charles NC

2013-01-01

257

Role of type IV secretion systems in trafficking of virulence determinants of Burkholderia cenocepacia  

E-print Network

in CF patients is often associated with a decline in respiratory function, and can result in acute systemic infection. Burkholderia cenocepacia strain K 56-2 is part of the epidemic and clinically problematic ET12 lineage. Two type IV secretion systems...

Engledow, Amanda Suzanne

2009-06-02

258

Title : Liquid C. elegans culture for the study of Burkholderia pathogenesis Authors : Vaughn Cooper1  

E-print Network

Title : Liquid C. elegans culture for the study of Burkholderia pathogenesis Authors : Vaughn Email: vaughn.cooper@unh.edu Abstract : The nematode C. elegans has been adopted as a key model system for the study of bacterial pathogenesis. The practical benefits of this system are notable: ease of culture

259

Analysis of the endophytic lifestyle and plant growth promotion of Burkholderia terricola ZR2-12  

Microsoft Academic Search

Members of the genus Burkholderia are highly versatile bacteria that can be beneficial as well as pathogenic for their eukaryotic hosts. Furthermore, many\\u000a strains exhibit a remarkable biotechnological potential. To study the ecosystem function and lifestyle of B. terricola, we analysed the interactions with plants and survival in soil as well as the mechanisms behind it. We used a combination

Ilona Gasser; Massimiliano Cardinale; Henry Müller; Stefanie Heller; Leo Eberl; Nicole Lindenkamp; Chlud Kaddor; Alexander Steinbüchel; Gabriele Berg

260

Perturbation of maize rhizosphere microflora following seed bacterization with Burkholderia cepacia MCI 7  

Microsoft Academic Search

Introduction of a large quantity of exogenous microorganisms may disrupt a local ecosystem and affect the natural microflora. In this work we investigated the effects of the introduction of a plant growth promoting strain of Burkholderia cepacia into the rhizosphere of maize on both indigenous B. cepacia populations and microbial community structure of total culturable bacteria using the concept of

Carlo Nacamulli; Annamaria Bevivino; Claudia Dalmastri; Silvia Tabacchioni; Luigi Chiarini

1997-01-01

261

Pilus-mediated epithelial cell death in response to infection with Burkholderia cenocepacia  

Microsoft Academic Search

Burkholderia cenocepacia is an opportunistic pathogen that can cause serious infections in cystic fibrosis (CF) patients. The ET12 lineage appears particularly virulent in CF; however, its pathogenesis is poorly understood and may be associated with host response. To help characterize this response, the ability of B. cenocepacia to induce cytotoxicity and apoptosis in an epithelial cell model was examined. Upon

K-John Cheung Jr.; Gang Li; Teresa A. Urban; Joanna B. Goldberg; Adam Griffith; Fuqu Lu; Jane L. Burns

2007-01-01

262

Structural, evolutionary and genetic analysis of the histidine biosynthetic “core” in the genus Burkholderia  

Microsoft Academic Search

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

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

2009-01-01

263

Burkholderia cepacia Complex Infection in Italian Patients with Cystic Fibrosis: Prevalence, Epidemiology, and Genomovar Status  

Microsoft Academic Search

The prevalence, epidemiology, and genomovar status of Burkholderia cepacia complex strains recovered from Italian cystic fibrosis (CF) patients were investigated using genetic typing and species identification methods. Four CF treatment centers were examined: two in Sicily, one in central Italy, and one in northern Italy. B. cepacia complex bacteria were isolated from 59 out of 683 CF patients attending these

ANTONELLA AGODI; ESHWAR MAHENTHIRALINGAM; MARTINA BARCHITTA; VIVIANA GIANNINO; AGATA SCIACCA; STEFANIA STEFANI

2001-01-01

264

Burkholderia and Cupriavidus spp. are the preferred symbionts of Mimosa spp. in southern China.  

PubMed

Rhizobia were isolated from invasive Mimosa spp. (M. diplotricha and M. pudica) in Dehong district of the province of Yunnan in subtropical southern China. Almost all of the 98 isolates were ?-rhizobia in the genera Burkholderia and Cupriavidus. These strains were analysed for their distribution characteristics together with strains from a previous study from Sishuangbanna. The proportion of nodules containing each ?-rhizobial genus varied between Mimosa species, with Cupriavidus being predominant in M. diplotricha nodules (63.3% compared to 36.7% occupation with Burkholderia), but with M. pudica showing a slight preference for Burkholderia over Cupriavidus, with them occupying 56.5% and 43.5% of nodules, respectively. The symbiosis-essential genes nodA and nifH were present in all the Burkholderia and Cupriavidus strains tested, and their phylogenies indicated that these Mimosa symbionts share symbiotic genes with native South American rhizobia. The evolutionary discrepancies among 16S rRNA genes, nodA and nifH of Mimosa spp. symbionts, suggests that the nod and nif genes of ?-rhizobia evolved independently. PMID:22268711

Liu, XiaoYun; Wei, Shuang; Wang, Fang; James, Euan K; Guo, XiaoYe; Zagar, Catherine; Xia, Liu Gui; Dong, Xin; Wang, Yi Peng

2012-05-01

265

Transmission and prevalence of Burkholderia cepacia in welsh cystic fibrosis patients  

Microsoft Academic Search

From 1987 to 1994, 16 of 162 cystic fibrosis (CF) patients attending CF clinics at three different hospitals in South Wales, U.K. were found to have respiratory secretions colonized with Burkholderia cepacia (B. cepacia). Bacteriological typing by polymerase chain reaction (PCR) ribotyping demonstrated seven strains of B. cepacia among these 16 CF patients. This typing confirmed that cross-infection was the

L. Millar-Jones; H. C. Ryley; A. Paull; M. C. Goodchild

1998-01-01

266

Transmission of Burkholderia cepacia Complex: Evidence for New Epidemic Clones Infecting Cystic Fibrosis Patients in Italy  

Microsoft Academic Search

To analyze national prevalence, genomovar distribution, and epidemiology of the Burkholderia cepacia complex in Italy, 225 putative B. cepacia complex isolates were obtained from 225 cystic fibrosis (CF) patients attending 18 CF centers. The genomovar status of these isolates was determined by a polyphasic approach, which included whole-cell protein electrophoresis and recA restriction fragment length polymorphism (RFLP) anal- ysis. Two

S. Campana; G. Taccetti; N. Ravenni; F. Favari; L. Cariani; A. Sciacca; D. Savoia; A. Collura; E. Fiscarelli; G. De Intinis; M. Busetti; A. Cipolloni; A. d'Aprile; E. Provenzano; I. Collebrusco; P. Frontini; G. Stassi; M. Trancassini; D. Tovagliari; A. Lavitola; C. J. Doherty; T. Coenye; J. R. W. Govan; P. Vandamme

2005-01-01

267

Quantification of Burkholderia coxL Genes in Hawaiian Volcanic Deposits?  

PubMed Central

Isolation of multiple carbon monoxide (CO)-oxidizing Burkholderia strains and detection by culture-independent approaches suggest that Burkholderia may be an important component of CO-oxidizing communities in Hawaiian volcanic deposits. The absolute and relative abundance of the bacteria in these communities remains unknown, however. In this study, a quantitative PCR (Q-PCR) approach has been developed to enumerate Burkholderia coxL genes (large subunit of carbon monoxide dehydrogenase). This represents the first attempt to enumerate coxL genes from CO oxidizers in environmental samples. coxL copy numbers have been determined for samples from three sites representing a vegetation gradient on a 1959 volcanic deposit that included unvegetated cinders (bare), edges of vegetated sites (edge), and sites within tree stands (canopy). Q-PCR has also been used to estimate copy numbers of Betaproteobacteria 16S rRNA gene copy numbers and total Bacteria 16S rRNA. coxL genes could not be detected in the bare site (detection limit, ?4.7 × 103 copies per reaction) but average 1.0 × 108 ± 2.4 × 107 and 8.6 × 108 ± 7.6 ×107 copies g?1 (dry weight) in edge and canopy sites, respectively, which differ statistically (P = 0.0007). Average Burkholderia coxL gene copy numbers, expressed as a percentage of total Bacteria 16S rRNA gene copy numbers, are 6.2 and 0.7% for the edge and canopy sites, respectively. Although the percentage of Burkholderia coxL is lower in the canopy site, significantly greater gene copy numbers demonstrate that absolute abundance of coxL increases in vegetated sites and contributes to the expansion of CO oxidizer communities during biological succession on volcanic deposits. PMID:20139318

Weber, C. F.; King, G. M.

2010-01-01

268

Evidence of environmental and vertical transmission of Burkholderia symbionts in the oriental chinch bug, Cavelerius saccharivorus (Heteroptera: Blissidae).  

PubMed

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. PMID:25038101

Itoh, Hideomi; Aita, Manabu; Nagayama, Atsushi; Meng, Xian-Ying; Kamagata, Yoichi; Navarro, Ronald; Hori, Tomoyuki; Ohgiya, Satoru; Kikuchi, Yoshitomo

2014-10-01

269

Spontaneous and evolutionary changes in the antibiotic resistance of Burkholderia cenocepacia observed by global gene expression analysis  

Microsoft Academic Search

Background  \\u000a Burkholderia cenocepacia is a member of the Burkholderia cepacia complex group of bacteria that cause infections in individuals with cystic fibrosis. B. cenocepacia isolate J2315 has been genome sequenced and is representative of a virulent, epidemic CF strain (ET12). Its genome encodes\\u000a multiple antimicrobial resistance pathways and it is not known which of these is important for intrinsic or

Andrea Sass; Angela Marchbank; Elizabeth Tullis; John J LiPuma; Eshwar Mahenthiralingam

2011-01-01

270

Improved molecular detection of Burkholderia cepacia genomovar III and Burkholderia multivorans directly from sputum of patients with cystic fibrosis.  

PubMed

Optimum detection of the Burkholderia cepacia complex (BCC) from sputum of patients with cystic fibrosis (CF) is essential in preventing patient-to-patient transmission of this organism. The aim of this study was to develop an improved PCR assay with reference to sensitivity for the direct detection of BCC organisms from CF sputum employing the recA locus. The sensitivity results of three recA PCR assays were compared using various combinations of previously published primers. These included (i) a single-round approach using the primer set BCR1/BCR2, yielding a 1036-bp product, (ii) a single-round approach using the primer set BCR1/Mr, yielding a 465-bp product, and (iii) a semi-nested PCR (SN-PCR) approach using the primer set BCR1/BCR2 followed by BCR1/Mr. The sensitivity of these assays were determined by spiking B. cepacia-free sputum with known numbers of four strains of BCC, namely, genomovar II [B. multivorans] (C1576), genomovar IIIa (C5424, C6433) and genomovar IIIb (C1394). Following optimization, the chosen assay was performed on 14 patients. Employment of the single-round assay with BCR1/BCR2 was the least sensitive with a detection threshold of 10(7) cfu/g sputum for GIIIa and GIIIb, and 10(8) cfu/g sputum for GII. Sensitivity was improved by targeting the smaller amplification region of the recA locus (465 bp) employing the BCR1/Mr primer pair, in combination with a single-round approach, whereby the detection threshold was improved by 1 log for each genomovar. Employment of the semi-nested assay demonstrated optimum sensitivity, whereby the detection threshold increased to 10(1) and 10(2) cfu/g sputum for genomovar IIIa/IIIb and genomovar II, respectively. Subsequent genomovar characterisation can be performed by sequencing of the PCR amplicon without the need for culture which may be beneficial in patients in the initial stages of colonisation or who are transiently colonised and who may be culture-negative for BCC. PMID:11830304

Moore, John E; Xu, Jiru; Millar, Beverley C; Crowe, Mary; Elborn, J Stuart

2002-04-01

271

Genome sequence of the Lebeckia ambigua-nodulating "Burkholderia sprentiae” strain WSM5005T  

PubMed Central

Burkholderia sprentiae” strain WSM5005T is an aerobic, motile, Gram-negative, non-spore-forming rod that was isolated in Australia from an effective N2-fixing root nodule of Lebeckia ambigua collected in Klawer, Western Cape of South Africa, in October 2007. Here we describe the features of “Burkholderia sprentiae” strain WSM5005T, together with the genome sequence and its annotation. The 7,761,063 bp high-quality-draft genome is arranged in 8 scaffolds of 236 contigs, contains 7,147 protein-coding genes and 76 RNA-only encoding genes, and is one of 20 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Community Sequencing Program. PMID:24976894

Reeve, Wayne; De Meyer, Sofie; Terpolilli, Jason; Melino, Vanessa; Ardley, Julie; Rui, Tian; Tiwari, Ravi; Howieson, John; Yates, Ron; O'Hara, Graham; Lu, Megan; Bruce, David; Detter, Chris; Tapia, Roxanne; Han, Cliff; Wei, Chia-Lin; Huntemann, Marcel; Han, James; Chen, I-Min; Mavromatis, Konstantinos; Markowitz, Victor; Szeto, Ernest; Ivanova, Natalia; Mikhailova, Natalia; Ovchinnikova, Galina; Pagani, Ioanna; Pati, Amrita; Goodwin, Lynne; Peters, Lin; Pitluck, Sam; Woyke, Tanja; Kyrpides, Nikos

2013-01-01

272

Bioassay-guided isolation of a low molecular weight PHB from Burkholderia sp. with phytotoxic activity.  

PubMed

This work reports on the bioassay-guided isolation and identification of the macrocyclic pentolide 1, a cyclic polyhydroxybutyrate (PHB) with low molecular weight. This metabolite is produced by Burkholderia sp. and it exhibited phytotoxic activity in a Lemna minor bioassay. Its structure was determined by (1)H and (13)C NMR, heteronuclear multiple quantum correlation, heteronuclear multiple bond correlation, IR, and electrospray ionization tandem mass spectrometry analyses. The period for maximum production of the pentolide was optimized and determined on the basis of multiple reaction monitoring experiments at 15 days. The potential of Burkholderia sp. as a producer of higher biopolymers of PHB was also investigated. The methodology employed here accelerated the isolation and characterization of a phytotoxic metabolite whose structure can serve as a model for the synthesis of new classes of herbicides. PMID:23722946

Petta, Tânia; Raichardt, Leandro; Melo, Itamar S; Moraes, Luiz A B

2013-08-01

273

Total protein extraction and 2-D gel electrophoresis methods for Burkholderia species.  

PubMed

The investigation of the intracellular protein levels of bacterial species is of importance to understanding the pathogenic mechanisms of diseases caused by these organisms. Here we describe a procedure for protein extraction from Burkholderia species based on mechanical lysis using glass beads in the presence of ethylenediamine tetraacetic acid and phenylmethylsulfonyl fluoride in phosphate buffered saline. This method can be used for different Burkholderia species, for different growth conditions, and it is likely suitable for the use in proteomic studies of other bacteria. Following protein extraction, a two-dimensional (2-D) gel electrophoresis proteomic technique is described to study global changes in the proteomes of these organisms. This method consists of the separation of proteins according to their isoelectric point by isoelectric focusing in the first dimension, followed by separation on the basis of molecular weight by acrylamide gel electrophoresis in the second dimension. Visualization of separated proteins is carried out by silver staining. PMID:24192802

Velapatiño, Billie; Zlosnik, James E A; Hird, Trevor J; Speert, David P

2013-01-01

274

Enhanced Bioconversion of Ethylene Glycol to Glycolic Acid by a Newly Isolated Burkholderia sp. EG13.  

PubMed

Burkholderia sp. EG13 with high ethylene glycol-oxidizing activity was isolated from soil, which could be used for the synthesis of glycolic acid from the oxidation of ethylene glycol. Using the resting cells of Burkholderia sp. EG13 as biocatalysts, the optimum reaction temperature and pH were 30 °C and 6.0, respectively. After 24 h of biotransformation, the yield of glycolic acid from 200 mM ethylene glycol was 98.8 %. Furthermore, an integrated bioprocess for the production of glycolic acid which involved in situ product removal (ISPR) was investigated. Using fed-batch method with ISPR, a total of 793 mM glycolic acid has been accumulated in the reaction mixture after the 4th feed. PMID:25123362

Gao, Xiaoxin; Ma, Zhengfei; Yang, Limin; Ma, Jiangquan

2014-10-01

275

Nitrogen-fixing bacterium Burkholderia brasiliensis produces a novel yersiniose A-containing O-polysaccharide.  

PubMed

Burkholderia brasiliensis, a Gram-negative diazotrophic endophytic bacterium, was first isolated from roots, stems, and leaves of rice plant in Brazil. The polysaccharide moiety was released by ammonolysis from the B. brasiliensis lipopolysaccharide (LPS), allowing the unambiguous characterization of a 3,6-dideoxy-4-C-(1-hydroxyethyl)-D-xylo-hexose (yersiniose A), an uncommon feature for Burkholderia LPS. The complete structure of the yersiniose A-containing O-antigen was identified by sugar and methylation analyses and NMR spectroscopy. Our results show that the repeating oligosaccharide motif of LPS O-chain consists of a branched tetrasaccharide with the following structure:-->2-alpha-d-Rhap-(1-->3)-[alpha-YerAp-(1-->2)]-alpha-D-Rhap-(1-->3)-alpha-D-Rhap-(1-->. PMID:15509723

Mattos, Katherine A; Todeschini, Adriane R; Heise, Norton; Jones, Christopher; Previato, Jose O; Mendonça-Previato, Lucia

2005-03-01

276

Characterization of Antimicrobial and Structural Metabolites from Burkholderia gladioli pv. agaricicola  

Microsoft Academic Search

Burkholderia gladioli pv. agaricicola, the causal agent of the soft rot disease of the cultivated mushroom Agaricus bitorquis, was demonstrated to produce in culture metabolites with antimicrobial activity, which were extracted by acid water-saturated\\u000a n-butanol and then purified by HPLC. Furthermore, this paper reports on the structural determination, based on the extensive\\u000a use of chemical analyses and spectroscopic techniques (essentially

A. Cimmino; P. Lo Cantore; G. Karapetyan; Z. Kaczynski; Nicola Sante Iacobellis; O. Holst; A. Evidente

277

Pathogenicity, virulence factors, and strategies to fight against Burkholderia cepacia complex pathogens and related species  

Microsoft Academic Search

The Burkholderia cepacia complex (Bcc) is a group of 17 closely related species of the ?-proteobacteria subdivision that emerged in the 1980s as important\\u000a human pathogens, especially to patients suffering from cystic fibrosis. Since then, a remarkable progress has been achieved\\u000a on the taxonomy and molecular identification of these bacteria. Although some progress have been achieved on the knowledge\\u000a of

Jorge H. Leitão; Sílvia A. Sousa; Ana S. Ferreira; Christian G. Ramos; Inês N. Silva; Leonilde M. Moreira

2010-01-01

278

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

Microsoft Academic Search

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.

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

2005-01-01

279

Burkholderia grimmiae sp. nov., isolated from a xerophilous moss (Grimmia montana).  

PubMed

A Gram-staining-negative, rod-shaped, non-spore-forming bacterium, designated strain R27(T), was isolated from the moss Grimmia montana, collected from Beijing Songshan National Nature Reserve, China, and characterized by using a polyphasic taxonomic approach. The predominant fatty acids of strain R27(T) were C18:1?7c (33.6%), C16:0 (16.3%), summed feature 3 (C16:1?7c and/or C16:1?6c; 15.8%) and C17:0 cyclo (8.7%) and its major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, three uncharacterized aminolipids and an unknown phospholipid. Strain R27(T) contained Q-8 as the dominant isoprenoid quinone and the G+C content of its genomic DNA was 64.6 mol%. On the basis of 16S rRNA gene sequence comparison, strain R27(T) showed 99.1% similarity to the closest related type strain, Burkholderia zhejiangensis OP-1(T), and 97.6% similarity to Burkholderia glathei ATCC 29195(T). However, the DNA-DNA relatedness between strain R27(T) and B. zhejiangensis CCTCC AB 2010354(T) and B. glathei ATCC 29195(T) was 10.2 and 14.9%, respectively. Based on 16S rRNA and rpoB gene sequence similarities and phenotypic and chemotaxonomic data, strain R27(T) is considered to represent a novel species of the genus Burkholderia, for which the name Burkholderia grimmiae sp. nov. is proposed. The type strain is R27(T) (=CGMCC 1.11013(T) =DSM 25160(T)). PMID:23087167

Tian, Yang; Kong, Bi He; Liu, Su Lin; Li, Chun Li; Yu, Rong; Liu, Lei; Li, Yan Hong

2013-06-01

280

Expression and characterization of penicillin-binding proteins in Burkholderia cenocepacia.  

PubMed

Putative penicillin-binding proteins (PBPs) were identified in the genome of the Burkholderia cenocepacia strain J2315 based on homology to E. coli PBPs. The three sequences identified as homologs of E. coli PBP1a, BCAL2021, BCAL0274, and BCAM2632, were cloned and expressed as His(6)-tagged fusion proteins in E. coli. The fusion proteins were isolated and shown to bind beta-lactams, indicating these putative PBPs have penicillin-binding activity. PMID:19924480

Specht, Kimberly Musa; Sheetz, Kyle H; Alexander, Courtney M; Lamech, Lilian T; O'Connor, Lauren H; Walker, Dawn M; Stevenson, Hilary P

2010-04-01

281

Aquifer Protist Response and the Potential for TCE Bioremediation with Burkholderia cepacia G4 PR1  

Microsoft Academic Search

Bacterivorous protists have been recovered from pristine and contaminated aquifer environments, but the ecological role of\\u000a these organisms in bioremediation strategies has not been well defined. Burkholderia cepacia G4 PR1 constitutively expresses a toluene ortho-monooxygenase (tom) due to a secondary transposition of a Tn5 transposable element in a trichloroethylene (TCE) degradative plasmid (TOM). Groundwater and sediment from a potential site

R. A. Snyder; J. D. Millward; W. S. Steffensen

2000-01-01

282

A novel strategy for the isolation and identification of environmental Burkholderia cepacia complex bacteria  

Microsoft Academic Search

The purpose of this study was to develop a novel strategy for the isolation and identification of Burkholderia cepacia complex bacteria from the home environment of cystic fibrosis (CF) patients. Water and soil samples were enriched in a broth containing 0.1% l-arabinose, 0.1% l-threonine, and a mixture of selective agents including 1?gml?1 C-390, 600Uml?1 polymyxin B sulfate, 10?gml?1 gentamycin, 2?gml?1

Elke Vanlaere; Tom Coenye; Emly Samyn; Caroline Van den Plas; John Govan; Frans De Baets; Kris De Boeck; Christiane Knoop; Peter Vandamme

2005-01-01

283

Efficacy of Burkholderia cepacia MCI 7 in disease suppression and growth promotion of maize  

Microsoft Academic Search

Repeated greenhouse experiments were performed to evaluate the ability of a maize-rhizosphere isolate of Burkholderia cepacia, applied as a seed coating, to promote maize growth in both uninfested soil and soil infested with a maize pathogenic strain\\u000a of Fusarium moniliforme, and to displace or negatively affect the population of F. moniliforme throughout plant growth. Results demonstrated that the B. cepacia

A. Bevivino; C. Dalmastri; S. Tabacchioni; L. Chiarini

2000-01-01

284

Short Chain N-acyl Homoserine Lactone Production by Soil Isolate Burkholderia sp. Strain A9  

PubMed Central

In the bacteria kingdom, quorum sensing (QS) is a cell-to-cell communication that relies on the production of and response to specific signaling molecules. In proteobacteria, N-acylhomoserine lactones (AHLs) are the well-studied signaling molecules. The present study aimed to characterize the production of AHL of a bacterial strain A9 isolated from a Malaysian tropical soil. Strain A9 was identified as Burkholderia sp. using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry and 16S rDNA nucleotide sequence analysis. AHL production by A9 was detected with two biosensors, namely Chromobacterium violaceum CV026 and Escherichia coli [pSB401]. Thin layer chromatography results showed N–hexanoylhomoserine lactone (C6-HSL) and N–octanoylhomoserine lactone (C8-HSL) production. Unequivocal identification of C6-HSL and C8-HSL was achieved by high resolution triple quadrupole liquid chromatography-mass spectrometry analysis. We have demonstrated that Burkholderia sp. strain A9 produces AHLs that are known to be produced by other Burkholderia spp. with CepI/CepR homologs. PMID:24084115

Chen, Jian Woon; Koh, Chong-Lek; Sam, Choon-Kook; Yin, Wai-Fong; Chan, Kok-Gan

2013-01-01

285

MtvR Is a Global Small Noncoding Regulatory RNA in Burkholderia cenocepacia  

PubMed Central

Burkholderia cenocepacia J2315 is a highly epidemic and transmissible clinical isolate of the Burkholderia cepacia complex (Bcc), a group of bacteria causing life-threatening respiratory infections among cystic fibrosis patients. This work describes the functional analysis of the 136-nucleotide (nt)-long MtvR small noncoding RNA (sRNA) from the Bcc member B. cenocepacia J2315, with homologues restricted to the genus Burkholderia. Bioinformatic target predictions revealed a total of 309 mRNAs to be putative MtvR targets. The mRNA levels corresponding to 17 of 19 selected genes were found to be affected when MtvR was either overexpressed or silenced. Analysis of the interaction between MtvR and the hfq mRNA, one of its targets, showed that the sRNA binds exclusively to the 5? untranslated region (UTR) of the hfq mRNA. This interaction resulted in decreased protein synthesis, suggesting a negative regulatory effect of MtvR on the RNA chaperone Hfq. Bacterial strains with MtvR silenced or overexpressed exhibited pleiotropic phenotypes related to growth and survival after several stresses, swimming and swarming motilities, biofilm formation, resistance to antibiotics, and ability to colonize and kill the nematode Caenorhabditis elegans. Together, the results indicate that the MtvR sRNA is a major posttranscriptional regulator in B. cenocepacia. PMID:23729649

Ramos, Christian G.; Grilo, Andre M.; da Costa, Paulo J. P.; Feliciano, Joana R.

2013-01-01

286

Draft Genome Sequence of Burkholderia sordidicola S170, a Potential Plant Growth Promoter Isolated from Coniferous Forest Soil in the Czech Republic  

PubMed Central

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, the biological control of plant diseases, and green remediation technologies. PMID:25125648

Xu, Zhuofei; S?rensen, S?ren J.; Baldrian, Petr

2014-01-01

287

Draft Genome Sequence of Burkholderia sordidicola S170, a Potential Plant Growth Promoter Isolated from Coniferous Forest Soil in the Czech Republic.  

PubMed

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, the biological control of plant diseases, and green remediation technologies. PMID:25125648

Lladó, Salvador; Xu, Zhuofei; Sørensen, Søren J; Baldrian, Petr

2014-01-01

288

Burkholderia oxyphila sp. nov., a bacterium isolated from acidic forest soil that catabolizes (+)-catechin and its putative aromatic derivatives.  

PubMed

A novel bacterium, designated strain OX-01(T), was isolated from acidic soil, taxonomically investigated and identified as an agent that catabolizes (+)-catechin into taxifolin. Strain OX-01(T) is a Gram-reaction-negative, aerobic, non-sporulating, non-motile and rod-shaped bacterium. 16S rRNA gene sequence analysis identified this strain as a member of the genus Burkholderia and occupying a phylogenetic position closest to, but clearly distinct from, Burkholderia sacchari. Strain OX-01(T) does not have any nif genes, which are required for N(2)-fixation, in its genome, a feature that is similar to B. sacchari, which lacks nifH, but is distinct from the N(2)-fixing features of many other phylogenetically related taxa, such as Burkholderia ferrariae, B. heleia, B. mimosarum, B. nodosa, B. silvatlantica, B. tropica and B. unamae. Strain OX-01(T) has the following chemotaxonomic characteristics: the major ubiquinone is Q-8, the DNA G+C content is 64 mol% and the major fatty acids are C(16?:?0), C(17?:?0) cyclo and C(18?:?1)?7c. It also has a unique profile of carbohydrate utilization among other species of the genus Burkholderia. The strain cannot assimilate many pentoses, hexoses and oligosaccharides, whereas it can catabolize (+)-catechin and its putative aromatic derivatives, such as 4-hydroxy-3-methoxycinnamic acid, protocatechuic acid, p-hydroxybenzoic acid, trans-p-coumaric acid and vanillic acid. Based on its morphological, physiological and chemotaxonomic characteristics, together with DNA-DNA relatedness values and 16S rRNA gene sequence comparison data, we show that strain OX-O1(T) represents a novel species of the genus Burkholderia, for which the name Burkholderia oxyphila sp. nov. is proposed. The type strain is OX-01(T) (=NBRC 105797(T) =DSM 22550(T)). PMID:20207808

Otsuka, Yuichiro; Muramatsu, Yuki; Nakagawa, Yasuyoshi; Matsuda, Motoki; Nakamura, Masaya; Murata, Hitoshi

2011-02-01

289

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

PubMed Central

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

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

2013-01-01

290

Versatility of the Burkholderia cepacia Complex for the Biosynthesis of Exopolysaccharides: A Comparative Structural Investigation  

PubMed Central

The Burkholderia cepacia Complex assembles at least eighteen closely related species that are ubiquitous in nature. Some isolates show beneficial potential for biocontrol, bioremediation and plant growth promotion. On the contrary, other strains are pathogens for plants and immunocompromised individuals, like cystic fibrosis patients. In these subjects, they can cause respiratory tract infections sometimes characterised by fatal outcome. Most of the Burkholderia cepacia Complex species are mucoid when grown on a mannitol rich medium and they also form biofilms, two related characteristics, since polysaccharides are important component of biofilm matrices. Moreover, polysaccharides contribute to bacterial survival in a hostile environment by inhibiting both neutrophils chemotaxis and antimicrobial peptides activity, and by scavenging reactive oxygen species. The ability of these microorganisms to produce exopolysaccharides with different structures is testified by numerous articles in the literature. However, little is known about the type of polysaccharides produced in biofilms and their relationship with those obtained in non-biofilm conditions. The aim of this study was to define the type of exopolysaccharides produced by nine species of the Burkholderia cepacia Complex. Two isolates were then selected to compare the polysaccharides produced on agar plates with those formed in biofilms developed on cellulose membranes. The investigation was conducted using NMR spectroscopy, high performance size exclusion chromatography, and gas chromatography coupled to mass spectrometry. The results showed that the Complex is capable of producing a variety of exopolysaccharides, most often in mixture, and that the most common exopolysaccharide is always cepacian. In addition, two novel polysaccharide structures were determined: one composed of mannose and rhamnose and another containing galactose and glucuronic acid. Comparison of exopolysaccharides obtained from cultures on agar plates with those extracted from biofilms on cellulose membranes showed important differences, thus suggesting that extrapolating data from non-biofilm conditions might not always be applicable. PMID:24722641

Silipo, Alba; Lanzetta, Rosa; Liut, Gianfranco; Rizzo, Roberto; Cescutti, Paola

2014-01-01

291

Evolution of an endofungal Lifestyle: Deductions from the Burkholderia rhizoxinica Genome  

PubMed Central

Background Burkholderia rhizoxinica is an intracellular symbiont of the phytopathogenic zygomycete Rhizopus microsporus, the causative agent of rice seedling blight. The endosymbiont produces the antimitotic macrolide rhizoxin for its host. It is vertically transmitted within vegetative spores and is essential for spore formation of the fungus. To shed light on the evolution and genetic potential of this model organism, we analysed the whole genome of B. rhizoxinica HKI 0454 - a type strain of endofungal Burkholderia species. Results The genome consists of a structurally conserved chromosome and two plasmids. Compared to free-living Burkholderia species, the genome is smaller in size and harbors less transcriptional regulator genes. Instead, we observed accumulation of transposons over the genome. Prediction of primary metabolic pathways and transporters suggests that endosymbionts consume host metabolites like citrate, but might deliver some amino acids and cofactors to the host. The rhizoxin biosynthesis gene cluster shows evolutionary traces of horizontal gene transfer. Furthermore, we analysed gene clusters coding for nonribosomal peptide synthetases (NRPS). Notably, B. rhizoxinica lacks common genes which are dedicated to quorum sensing systems, but is equipped with a large number of virulence-related factors and putative type III effectors. Conclusions B. rhizoxinica is the first endofungal bacterium, whose genome has been sequenced. Here, we present models of evolution, metabolism and tools for host-symbiont interaction of the endofungal bacterium deduced from whole genome analyses. Genome size and structure suggest that B. rhizoxinica is in an early phase of adaptation to the intracellular lifestyle (genome in transition). By analysis of tranporters and metabolic pathways we predict how metabolites might be exchanged between the symbiont and its host. Gene clusters for biosynthesis of secondary metabolites represent novel targets for genomic mining of cryptic natural products. In silico analyses of virulence-associated genes, secreted proteins and effectors might inspire future studies on molecular mechanisms underlying bacterial-fungal interaction. PMID:21539752

2011-01-01

292

Purification and characterization of fenitrothion hydrolase from Burkholderia sp. NF100.  

PubMed

The organophosphorus pesticide hydrolase was purified to homogeneity from Burkholderia sp. NF100 by detergent extraction of the cell membrane fraction, anion-exchange, chromatofocusing, and gel filtration chromatographies. The purified enzyme had a molecular mass of 55 kDa and a pI 5.8, and the hydrolase activity was strongly inhibited by EDTA, dithiothreitol (DTT), Hg2+ and 1,10-phenanthroline. The optimum pH and temperature for the enzyme activity were 8.0 and 40 degrees C, respectively. The enzyme hydrolyzed five organophosphorus pesticides. PMID:16503297

Tago, Kanako; Yonezawa, Sumiko; Ohkouchi, Toshihide; Hashimoto, Masayuki; Hayatsu, Masahito

2006-01-01

293

Eradication of Burkholderia cepacia Using Inhaled Aztreonam Lysine in Two Patients with Bronchiectasis  

PubMed Central

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.

Iglesias, A.; Artiles, I.; Cabanillas, J. J.; Alvarez-Sala, R.; Prados, C.

2014-01-01

294

The chlorobenzoate dioxygenase genes of Burkholderia sp. strain NK8 involved in the catabolism of chlorobenzoates  

Microsoft Academic Search

Burkholderia sp. NK8 grows abundantly on 3-chlorobenzoate (3CB), 4-chlorobenzoate (4CB) and benzoate. The genes encoding the oxidation of (chloro)benzoates (cbeABCD) and catechol (catA, catBC), the LysR-type regulatory gene cbeR and the gene cbeE with unknown function, all of which form a single cluster in NK8, were cloned and analysed. The protein sequence of chlorobenzoate 1,2-dioxygenase (CbeABC) is 50-65% identical to

Perigio B. Francisco Jr; Naoto Ogawa; Katsuhisa Suzuki; Kiyotaka Miyashita

295

The Promise of Bacteriophage Therapy for Burkholderia cepacia Complex Respiratory Infections  

PubMed Central

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 investigation is not only comprised of phage isolation, in vitro phage characterization and assessment of in vivo infection model efficacy, but also adapting aerosol drug delivery techniques to aerosol phage formulation delivery and storage. PMID:22919592

Semler, Diana D.; Lynch, Karlene H.; Dennis, Jonathan J.

2012-01-01

296

ACC (1-Aminocyclopropane-1-Carboxylate) Deaminase Activity, a Widespread Trait in Burkholderia Species, and Its Growth-Promoting Effect on Tomato Plants?  

PubMed Central

The genus Burkholderia includes pathogens of plants and animals and some human opportunistic pathogens, such as the Burkholderia cepacia complex (Bcc), but most species are nonpathogenic, plant associated, and rhizospheric or endophytic. Since rhizobacteria expressing ACC (1-aminocyclopropane-1-carboxylate) deaminase may enhance plant growth by lowering plant ethylene levels, in this work we investigated the presence of ACC deaminase activity and the acdS gene in 45 strains, most of which are plant associated, representing 20 well-known Burkholderia species. The results demonstrated that ACC deaminase activity is a widespread feature in the genus Burkholderia, since 18 species exhibited ACC deaminase activities in the range from 2 to 15 ?mol of ?-ketobutyrate/h/mg protein, which suggests that these species may be able to modulate ethylene levels and enhance plant growth. In these 18 Burkholderia species the acdS gene sequences were highly conserved (76 to 99% identity). Phylogenetic analysis of acdS gene sequences in Burkholderia showed tight clustering of the Bcc species, which were clearly distinct from diazotrophic plant-associated Burkholderia species. In addition, an acdS knockout mutant of the N2-fixing bacterium Burkholderia unamae MTl-641T and a transcriptional acdSp-gusA fusion constructed in this strain showed that ACC deaminase could play an important role in promotion of the growth of tomato plants. The widespread ACC deaminase activity in Burkholderia species and the common association of these species with plants suggest that this genus could be a major contributor to plant growth under natural conditions. PMID:19700546

Onofre-Lemus, Janette; Hernandez-Lucas, Ismael; Girard, Lourdes; Caballero-Mellado, Jesus

2009-01-01

297

South African Papilionoid Legumes Are Nodulated by Diverse Burkholderia with Unique Nodulation and Nitrogen-Fixation Loci  

PubMed Central

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

Beukes, Chrizelle W.; Venter, Stephanus N.; Law, Ian J.; Phalane, Francina L.; Steenkamp, Emma T.

2013-01-01

298

Suppression of Bacterial Wilt and Fusarium Wilt by a Burkholderia nodosa Strain Isolated from Kalimantan Soils, Indonesia.  

PubMed

A trial was conducted to suppress bacterial wilt of tomato (BWT) caused by Ralstonia solanacearum using biocontrol agents (BCAs) isolated from soils in Kalimantan, Indonesia. Five isolates were selected from 270 isolates as better performing BCAs through screening four times using a pumice medium. The isolates selected were identified as Burkholderia nodosa, Burkholderia sacchari, Burkholderia pyrrocinia and Burkholderia terricola according to 16S rDNA sequences, fatty acid composition and carbon source utilization patterns. Among them, B. nodosa G5.2.rif1 had significant suppressive effects on Fusarium wilt of tomato (FWT) and spinach (FWS) as well as BWT. When B. nodosa G5.2rif1 was inoculated into a pumice medium in combination with sucrose, it showed even more stable disease suppression for BWT, but not for FWS. This suppression was considered to mainly occur through competition for nutrients. In two times greenhouse experiments for BWT using pots comparable in size to those used commercially, B. nodosa G5.2rif1 significantly suppressed the disease index by 33-79%, with no inhibitory effects on the growth, yield and quality of tomatoes. PMID:21558699

Nion, Yanetri Asi; Toyota, Koki

2008-01-01

299

Culture-Based and Non-Growth-Dependent Detection of the Burkholderia cepacia Complex in Soil Environments  

Microsoft Academic Search

Burkholderia cepacia complex (Bcc) bacteria reside in soil, plant rhizospheres, and water, but their preva- lence and distribution in outdoor environments is not clear. We sampled a variety of soil and rhizosphere environments with which people may have contact: playgrounds, athletic fields, parks, hiking trails, residential yards, and gardens. A total of 91 sites was sampled in three large U.S.

Suzanne C. M. Miller; John J. LiPuma; Jennifer L. Parke

2002-01-01

300

Extraction of Antifungal Substances from Burkholderia cepacia with Antibiotic Activity against Colletotrichum gloeosporioides on Papaya (Carica papaya)  

Microsoft Academic Search

An experiment was conducted to extract and determine the nature of antifungal substances produced by Burkholderia cepacia strain B23 that were inhibitory towards Colletotrichum gloeosporioides. In addition, the effect of different culture media on the production of antifungal substances by B. cepacia B23 was also investigated to have improved efficacy of this biocontrol agent. B. cepacia B23 grew faster in

J. KADIR; M. A. RAHMAN; T. M. M. MAHMUD; R. ABDUL RAHMAN; M. M. BEGUM

301

An efficient preparative scale resolution of 3-phenylbutyric acid by lipase from Burkholderia cepacia (Chirazyme L1)  

Microsoft Academic Search

Lipase from Burkholderia cepacia (Chirazyme L1) catalysed the highly enantioselective hydrolysis of racemic methyl 3-phenylbutyrate to afford (R)-(?)-methyl 3-phenylbutyrate of >98% ee (E>50). The resolution was performed at 150 g scale yielding 68.7 g of (R)-(?)-methyl 3-phenylbutyrate (>98% ee, 92% yield on enantiomer) and (S)-(+)-3-phenylbutyric acid of 89% ee.

Govindarajulu Varadharaj; Keith Hazell; Christopher D Reeve

1998-01-01

302

Degradation of Chlorobenzenes at Nanomolar Concentrations by Burkholderia sp. Strain PS14 in Liquid Cultures and in Soil  

PubMed Central

The utilization of 1,2,4,5-tetrachloro-, 1,2,4-trichloro-, the three isomeric dichlorobenzenes and fructose as the sole carbon and energy sources at nanomolar concentrations was studied in batch experiments with Burkholderia sp. strain PS14. In liquid culture, all chlorobenzenes were metabolized within 1 h from their initial concentration of 500 nM to below their detection limits of 0.5 nM for 1,2,4,5-tetrachloro- and 1,2,4-trichlorobenzene and 7.5 nM for the three dichlorobenzene isomers, with 63% mineralization of the tetra- and trichloroisomers. Fructose at the same initial concentration was, in contrast, metabolized over a 4-h incubation period down to a residual concentration of approximately 125 nM with 38% mineralization during this time. In soil microcosms, Burkholderia sp. strain PS14 metabolized tetrachlorobenzene present at 64.8 ppb and trichlorobenzene present at 54.4 ppb over a 72-h incubation period to below the detection limits of 0.108 and 0.09 ppb, respectively, with approximately 80% mineralization. A high sorptive capacity of Burkholderia sp. strain PS14 for 1,2,4,5-tetrachlorobenzene was found at very low cell density. The results demonstrate that Burkholderia sp. strain PS14 exhibits a very high affinity for chlorobenzenes at nanomolar concentrations. PMID:10347041

Rapp, Peter; Timmis, Kenneth N.

1999-01-01

303

Comparative Evaluation of the BD Phoenix and VITEK 2 Automated Instruments for Identification of Isolates of the Burkholderia cepacia Complex  

Microsoft Academic Search

We evaluated two new automated identification systems, the BD Phoenix (Becton Dickinson) and the VITEK 2 (bioMérieux), for identification of isolates of the Burkholderia cepacia complex (BCC). The test sample included 42 isolates of the highly virulent and epidemic genomovar III, 45 isolates of B. multivorans, and 47 isolates of other members of the BCC. Rates of correct identification by

Sylvain Brisse; Stefania Stefani; Jan Verhoef; Alex Van Belkum; Peter Vandamme; Wil Goessens

2002-01-01

304

Comparative Genome Sequence Analysis Reveals the Extent of Diversity and Conservation for Glycan-Associated Proteins in Burkholderia spp.  

PubMed Central

Members of the Burkholderia family occupy diverse ecological niches. In pathogenic family members, glycan-associated proteins are often linked to functions that include virulence, protein conformation maintenance, surface recognition, cell adhesion, and immune system evasion. Comparative analysis of available Burkholderia genomes has revealed a core set of 178 glycan-associated proteins shared by all Burkholderia of which 68 are homologous to known essential genes. The genome sequence comparisons revealed insights into species-specific gene acquisitions through gene transfers, identified an S-layer protein, and proposed that significantly reactive surface proteins are associated to sugar moieties as a potential means to circumvent host defense mechanisms. The comparative analysis using a curated database of search queries enabled us to gain insights into the extent of conservation and diversity, as well as the possible virulence-associated roles of glycan-associated proteins in members of the Burkholderia spp. The curated list of glycan-associated proteins used can also be directed to screen other genomes for glycan-associated homologs. PMID:22991502

Ong, Hui San; Mohamed, Rahmah; Firdaus-Raih, Mohd

2012-01-01

305

Studies revealing bioremediation potential of the strain Burkholderia sp. GB-01 for abamectin contaminated soils.  

PubMed

Burkholderia sp. GB-01 strain was used to study different factors affecting its growth for inoculum production and then evaluated for abamectin degradation in soil for optimization under various conditions. The efficiency of abamectin degradation in soil by strain GB-01 was seen to be dependent on soil pH, temperature, initial abamectin concentration, and inoculum size along with inoculation frequency. Induction studies showed that abamectin depletion was faster when degrading cells were induced by pre-exposure to abamectin. Experiments performed with varying concentrations (2-160 mg Kg(-1)) of abamectin-spiked soils showed that strain GB-01 could effectively degrade abamectin over the range of 2-40 mg Kg(-1). The doses used were higher than the recommended dose for an agricultural application of abamectin, taking in account the over-use or spill situations. A cell density of approximately 10(8) viable cells g(-1) dry weight of soil was found to be suitable for bioremediation over a temperature range of 30-35°C and soil pH 7.5-8.5. This is the first report on bacterial degradation of abamectin in soil by a Burkholderia species, and our results indicated that this bacterium may be useful for efficient removal of abamectin from contaminated soils. PMID:22806778

Ali, Shinawar Waseem; Yu, Fang-bo; Li, Lian-tai; Li, Xiao-hui; Gu, Li-feng; Jiang, Jian-dong; Li, Shun-peng

2012-01-01

306

Burkholderia (Pseudomonas) cepacia epidemiology in a cystic fibrosis population: a genome finger-printing study.  

PubMed

In patients with cystic fibrosis, infection with Pseudomonas cepacia is associated with poor outcomes. However, the epidemiology of Burkholderia cepacia is still unclear. The aim of this study was to investigate the epidemiology of Burkholderia (Pseudomonas) cepacia colonization among cystic fibrosis patients attending the Verona CF Centre, a large specialized unit to which patients from different parts of Italy are admitted. We used a genome finger-printing system to analyse the nucleotidic structure of B. cepacia isolates from 60 colonized cystic fibrosis patients. Forty-two different finger-printing patterns were identified. Thirty-two patients were colonized by individual B. cepacia strains (53.3%). The remaining 28 subjects were divided into 10 different subgroups, each exhibiting a distinct strain of B. cepacia (46.7%). Nevertheless, direct, person-to-person transmission was evident in only 10 cases (16.7%). The stability up to 12 months, of the B. cepacia colonizing strain was documented in 36 individuals. Consistent with other reports, risk of B. cepacia transmission between cystic fibrosis patients through intimacy or nosocomial contact was found in our study. However, besides low contagiousness, our data suggest that the environmental reservoir of B. cepacia outside the hospital seems to play an important role in B. cepacia infection of our cystic fibrosis population. PMID:8827098

Cazzola, G; Amalfitano, G; Tonolli, E; Perazzoli, C; Piacentini, I; Mastella, G

1996-05-01

307

Polyhydroxyalkanoate biosynthesis and simultaneous remotion of organic inhibitors from sugarcane bagasse hydrolysate by Burkholderia sp.  

PubMed

Burkholderia sp. F24, originally isolated from soil, was capable of growth on xylose and removed organic inhibitors present in a hemicellulosic hydrolysate and simultaneously produced poly-3-hydroxybutyrate (P3HB). Using non-detoxified hydrolysate, Burkholderia sp. F24 reached a cell dry weight (CDW) of 6.8 g L(-1), containing 48 % of P3HB and exhibited a volumetric productivity (PP3HB) of 0.10 g L(-1) h(-1). Poly-3-hydroxybutyrate-co-3-hydroxyvalerate copolymers (P3HB-co-3HV) were produced using xylose and levulinic acid (LA) as carbon sources. In shake flask cultures, the 3HV content in the copolymer increased from 9 to 43 mol% by adding LA from 1.0 to 5.0 g L(-1). In high cell density cultivation using concentrated hemicellulosic hydrolysate F24 reached 25.04 g L(-1) of CDW containing 49 % of P3HB and PP3HB of 0.28 g L(-1 )h(-1). Based on these findings, second-generation ethanol and bioplastics from sugarcane bagasse is proposed. PMID:25059637

Lopes, Mateus Schreiner Garcez; Gomez, José Gregório Cabrera; Taciro, Marilda Keico; Mendonça, Thatiane Teixeira; Silva, Luiziana Ferreira

2014-09-01

308

Screening a mushroom extract library for activity against Acinetobacter baumannii and Burkholderia cepacia and the identification of a compound with anti-Burkholderia activity  

PubMed Central

Background Acinetobacter baumannii and species within the Burkholderia cepacia complex (BCC) are significant opportunistic bacterial pathogens of humans. These species exhibit a high degree of antibiotic resistance, and some clinical isolates are resistant to all currently available antimicrobial drugs used for treatment. Thus, new drugs are needed to treat infections by these species. Mushrooms could be a potential source for new drugs to treat A. baumannii and BCC infections. Methods The aim of this study was to screen a library of crude extracts from 330 wild mushrooms by disk diffusion assays for antibacterial activity against A. baumannii and Burkholderia cepacia in the hope of identifying a novel natural drug that could be used to treat infections caused by these species. Once positive hits were identified, the extracts were subjected to bioassay-guided separations to isolate and identify the active drug molecules. MICs were performed to gauge the in vitro activity of the purified compounds. Results Only three crude extracts (0.9%) had activity against A. baumannii and B. cepacia. Compounds from two of these extracts had MICs greater than 128 ?g/ml, and further analyses were not performed. From the third extract, prepared from Leucopaxillus albissimus, 2-aminoquinoline (2-AQ) was isolated. This compound exhibited a modest MIC in vitro against strains from nine different BCC species, including multi-drug resistant clinical isolates (MIC = 8-64 ?g/ml), and a weak MIC (128 ?g/ml) against A baumannii. The IC50 against a murine monocyte line was 1.5 mg/ml. Conclusion The small number of positive hits in this study suggests that finding a new drug from mushrooms to treat Gram-negative bacterial infections may be difficult. Although 2-AQ was identified in one mushroom, and it was shown to inhibit the growth of multi-drug resistant BCC isolates, the relatively high MICs (8-128 ?g/ml) for both A. baumannii and BCC strains suggests that 2-AQ is not suitable for further drug development in its current form. PMID:20092635

2010-01-01

309

IN SITU BIOREMEDIATION OF TRICHLOROETHYLENE USING BURKHOLDERIA CEPACIA G4 PR1: ANALYSIS OF MICROBIAL ECOLOGY PARAMETERS FOR RISK ASSESSMENT (RESEARCH BRIEF)  

EPA Science Inventory

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) ...

310

Burkholderia dabaoshanensis sp. nov., a Heavy-Metal-Tolerant Bacteria Isolated from Dabaoshan Mining Area Soil in China  

PubMed Central

Heavy-metal-tolerant bacteria, GIMN1.004T, was isolated from mine soils of Dabaoshan in South China, which were acidic (pH 2–4) and polluted with heavy metals. The isolation was Gram-negative, aerobic, non-spore-forming, and rod-shaped bacteria having a cellular width of 0.5?0.6 µm and a length of 1.3?1.8 µm. They showed a normal growth pattern at pH 4.0–9.0 in a temperature ranging from 5°C to 40°C.The organism contained ubiquinone Q-8 as the predominant isoprenoid quinine, and C16?0, summed feature 8 (C18?1?7c and C18?1?6c), C18?0, summed feature 3 (C16?1?7c or iso-C15?0 2-OH), C17?0 cyclo, C18?1?9c, C19?0 cyclo ?8c, C14?0 as major fatty acid. These profiles were similar to those reported for Burkholderia species. The DNA G+C % of this strain was 61.6%. Based on the similarity to 16S rRNA gene sequence, GIMN1.004T was considered to be in the genus Burkholderia. The similarities of 16S rRNA gene sequence between strain GIMN1.004T and members of the genus Burkholderia were 96?99.4%, indicating that this novel strain was phylogenetically related to members of that genus. The novel strain showed the highest sequence similarities to Burkholderia soli DSM 18235T (99.4%); Levels of DNA-DNA hybridization with DSM 18235T was 25%. Physiological and biochemical tests including cell wall composition analysis, differentiated phenotype of this strain from that closely related Burkholderia species. The isolation had great tolerance to cadmium with MIC of 22 mmol/L, and adsorbability of 144.94 mg/g cadmium,and it was found to exhibit antibiotic resistance characteristics. The adsorptive mechanism of GIMN1.004T for cadmium depended on the action of the amide,carboxy and phosphate of cell surface and producing low-molecular-weight (LMW ) organic acids to complex or chelated Cd2+.Therefore, the strain GIMN1.004T represented a new cadmium resistance species, which was tentatively named as Burkholderia dabaoshanensis sp. nov. The strain type is GIMN1.004T (?=?CCTCC M 209109T?=? NRRL B-59553T ). PMID:23226514

Zhu, Honghui; Guo, Jianhua; Chen, Meibiao; Feng, Guangda; Yao, Qing

2012-01-01

311

Biodiesel production from Jatropha oil catalyzed by immobilized Burkholderia cepacia lipase on modified attapulgite.  

PubMed

Lipase from Burkholderia cepacia was immobilized on modified attapulgite by cross-linking reaction for biodiesel production with jatropha oil as feedstock. Effects of various factors on biodiesel production were studied by single-factor experiment. Results indicated that the best conditions for biodiesel preparation were: 10 g jatropha oil, 2.4 g methanol (molar ratio of oil to methanol is 1:6.6) being added at 3h intervals, 7 wt% water, 10 wt% immobilized lipase, temperature 35°C, and time 24h. Under these conditions, the maximum biodiesel yield reached 94%. The immobilized lipase retained 95% of its relative activity during the ten repeated batch reactions. The half-life time of the immobilized lipase is 731 h. Kinetics was studied and the Vmax of the immobilized lipases were 6.823 mmol L(-1). This immobilized lipase catalyzed process has potential industrial use for biodiesel production to replace chemical-catalyzed method. PMID:24055964

You, Qinghong; Yin, Xiulian; Zhao, Yuping; Zhang, Yan

2013-11-01

312

Identification of aldolase and ferredoxin reductase within the dbt operon of Burkholderia fungorum DBT1.  

PubMed

Burkholderia fungorum DBT1, first isolated from settling particulate matter of an oil refinery wastewater, is a bacterial strain which has been shown capable of utilizing several polycyclic aromatic hydrocarbons (PAHs) including dibenzothiophene (DBT). In particular, this microbe is able to efficiently degrade DBT through the Kodama pathway. Previous investigations have lead to the identification of six genes, on a total of eight, required for DBT degradation. In the present study, a combined experimental/computational approach was adopted to identify and in silico characterize the two missing genes, namely a ferredoxin reductase and a hydratase-aldolase. Thus, the finding of all enzymatic components of the Kodama pathway in B. fungorum DBT1 makes this bacterial strain amenable for possible exploitation in soil bioremediation protocols. PMID:23686744

Piccoli, Stefano; Andreolli, Marco; Giorgetti, Alejandro; Zordan, Fabio; Lampis, Silvia; Vallini, Giovanni

2014-05-01

313

Identification of two new sets of genes for dibenzothiophene transformation in Burkholderia sp. DBT1.  

PubMed

A novel genotype for the initial steps of the oxidative degradation of dibenzothiophene (DBT) is described in a Burkholderia sp. strain isolated from a drain receiving oil refinery wastewater. The strain is capable of transforming DBT with significant efficiency when compared to other microorganisms. Its genotype was discovered by investigating insertional mutants of genes involved in DBT degradation by the Kodama pathway. The cloned dbt genes show a novel genomic organization when compared to previously described genes capable of DBT catabolism in that they constitute two distinct operons and are not clustered in a single transcript. Sequence analysis suggests the presence of a sigma54-dependent positive transcriptional regulator that may be involved in the control of the transcription of the two operons, both activated by DBT. The achieved results suggest the possibility of novel features of DBT biotransformation in nature. PMID:15068372

Di Gregorio, Simona; Zocca, Chiara; Sidler, Stephan; Toffanin, Annita; Lizzari, Daniela; Vallini, Giovanni

2004-04-01

314

Optimization of Lipase Production by Burkholderia sp. Using Response Surface Methodology  

PubMed Central

Response surface methodology (RSM) was employed to optimize the extracellular lipase production by Burkholderia sp. HL-10. Preliminary tests showed that olive oil, tryptone and Tween-80 exhibited significant effects on the lipase production. The optimum concentrations of these three components were determined using a faced-centered central composite design (FCCCD). The analysis of variance revealed that the established model was significant (p < 0.01). The optimized medium containing 0.65% olive oil (v/v), 2.42% tryptone (w/v) and 0.15% Tween-80 (v/v) resulted in a maximum activity of 122.3 U/mL, about three fold higher than that in basal medium. Approximately 99% of validity of the predicted value was achieved. PMID:23203100

Lo, Chia-Feng; Yu, Chi-Yang; Kuan, I-Ching; Lee, Shiow-Ling

2012-01-01

315

Antimicrobial properties of an oxidizer produced by Burkholderia cenocepacia P525.  

PubMed

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 single major peak. Size exclusion chromatography suggests a molecular mass of ~1,150 and UV spectroscopy suggests the presence of a polyene structure consisting of as many as six conjugated double bonds. Biological studies indicate that the compound is bacteriostatic. Enterobacter soli and E. aerogenes cells incubated with the compound exhibit a longer lag phase of growth. The bacteriostatic activity is greater at pH 3 than at pH 5. Bacteria such as B. cenocepacia strain P525 may have value in the agricultural industry as biocontrol agents. PMID:24384626

Hunter, William J; Manter, Dan K

2014-05-01

316

Genome sequence of Burkholderia mimosarum strain LMG 23256T, a Mimosa pigra microsymbiont from Anso, Taiwan  

PubMed Central

Burkholderia mimosarum strain LMG 23256T is an aerobic, motile, Gram-negative, non-spore-forming rod that can exist as a soil saprophyte or as a legume microsymbiont of Mimosa pigra (giant sensitive plant). LMG 23256T was isolated from a nodule recovered from the roots of the M. pigra growing in Anso, Taiwan. LMG 23256T is highly effective at fixing nitrogen with M. pigra. Here we describe the features of B. mimosarum strain LMG 23256T, together with genome sequence information and its annotation. The 8,410,967 bp high-quality-draft genome is arranged into 268 scaffolds of 270 contigs containing 7,800 protein-coding genes and 85 RNA-only encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

Willems, Anne; Tian, Rui; Brau, Lambert; Goodwin, Lynne; Han, James; Liolios, Konstantinos; Huntemann, Marcel; Pati, Amrita; Woyke, Tanja; Mavrommatis, Konstantinos; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; Reeve, Wayne

2013-01-01

317

Identification by replicon-typing of indigenous plasmids of Burkholderia (Pseudomonas) solanacearum.  

PubMed

Fifteen strains of Burkholderia (Pseudomonas) solanacearum were isolated, identified from different parts of India and screened for plasmid profile. They harboured 1-3 plasmids, but most of the isolates contained a single plasmid. Plasmids were classified on the basis of replicon typing by Southern hybridization using incompatible specific rep probes. Plasmids of 9 strains hybridized with repP probe, plasmids of 5 strains hybridized with repW probe and one plasmid hybridized with repQ probe. Fingerprint and cluster analyses of plasmids from 10 strains of B. solanacearum digested with Eco RI revealed that plasmids of race 3 strains clustered in one group and the plasmids of race 1 strains clustered in another group. PMID:9315243

Elumalai, R P; Mahadevan, A

1997-04-01

318

Immobilization of Burkholderia sp. lipase on a ferric silica nanocomposite for biodiesel production.  

PubMed

In this work, lipase produced from an isolated strain Burkholderia sp. C20 was immobilized on magnetic nanoparticles to catalyze biodiesel synthesis. Core-shell nanoparticles were synthesized by coating Fe(3)O(4) core with silica shell. The nanoparticles treated with dimethyl octadecyl [3-(trimethoxysilyl) propyl] ammonium chloride were used as immobilization supporters. The Burkholderia lipase was then bound to the synthesized nanoparticles for immobilization. The protein binding efficiency on alkyl-functionalized Fe(3)O(4)-SiO(2) was estimated as 97%, while the efficiency was only 76% on non-modified Fe(3)O(4)-SiO(2). Maximum adsorption capacity of lipase on alkyl-functionalized Fe(3)O(4)-SiO(2) was estimated as 29.45 mg g(-1) based on Langmuir isotherm. The hydrolytic kinetics (using olive oil as substrate) of the lipase immobilized on alkyl-grafted Fe(3)O(4)-SiO(2) followed Michaelis-Menten model with a maximum reaction rate and a Michaelis constant of 6251 Ug(-1) and 3.65 mM, respectively. Physical and chemical properties of the nanoparticles and the immobilized lipase were characterized by Brunauer-Emmett-Teller (BET) analysis, scanning electron microscope (SEM), and Fourier transform infrared spectroscopy (FT-IR). Moreover, the immobilized lipase was used to catalyze the transesterification of olive oil with methanol to produce fatty acid methyl esters (FAMEs), attaining a FAMEs conversion of over 90% within 30 h in batch operation when 11 wt% immobilized lipase was employed. The immobilized lipase could be used for ten cycles without significant loss in its transesterification activity. PMID:22306108

Tran, Dang-Thuan; Chen, Ching-Lung; Chang, Jo-Shu

2012-04-15

319

Novel Burkholderia mallei virulence factors linked to specific host-pathogen protein interactions.  

PubMed

Burkholderia mallei is an infectious intracellular pathogen whose virulence and resistance to antibiotics makes it a potential bioterrorism agent. Given its genetic origin as a commensal soil organism, it is equipped with an extensive and varied set of adapted mechanisms to cope with and modulate host-cell environments. One essential virulence mechanism constitutes the specialized secretion systems that are designed to penetrate host-cell membranes and insert pathogen proteins directly into the host cell's cytosol. However, the secretion systems' proteins and, in particular, their host targets are largely uncharacterized. Here, we used a combined in silico, in vitro, and in vivo approach to identify B. mallei proteins required for pathogenicity. We used bioinformatics tools, including orthology detection and ab initio predictions of secretion system proteins, as well as published experimental Burkholderia data to initially select a small number of proteins as putative virulence factors. We then used yeast two-hybrid assays against normalized whole human and whole murine proteome libraries to detect and identify interactions among each of these bacterial proteins and host proteins. Analysis of such interactions provided both verification of known virulence factors and identification of three new putative virulence proteins. We successfully created insertion mutants for each of these three proteins using the virulent B. mallei ATCC 23344 strain. We exposed BALB/c mice to mutant strains and the wild-type strain in an aerosol challenge model using lethal B. mallei doses. In each set of experiments, mice exposed to mutant strains survived for the 21-day duration of the experiment, whereas mice exposed to the wild-type strain rapidly died. Given their in vivo role in pathogenicity, and based on the yeast two-hybrid interaction data, these results point to the importance of these pathogen proteins in modulating host ubiquitination pathways, phagosomal escape, and actin-cytoskeleton rearrangement processes. PMID:23800426

Memisevi?, Vesna; Zavaljevski, Nela; Pieper, Rembert; Rajagopala, Seesandra V; Kwon, Keehwan; Townsend, Katherine; Yu, Chenggang; Yu, Xueping; DeShazer, David; Reifman, Jaques; Wallqvist, Anders

2013-11-01

320

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

NASA Astrophysics Data System (ADS)

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.

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

2014-09-01

321

Transports of acetate and haloacetate in Burkholderia species MBA4 are operated by distinct systems  

PubMed Central

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. PMID:23167477

2012-01-01

322

Novel Burkholderia mallei Virulence Factors Linked to Specific Host-Pathogen Protein Interactions*  

PubMed Central

Burkholderia mallei is an infectious intracellular pathogen whose virulence and resistance to antibiotics makes it a potential bioterrorism agent. Given its genetic origin as a commensal soil organism, it is equipped with an extensive and varied set of adapted mechanisms to cope with and modulate host-cell environments. One essential virulence mechanism constitutes the specialized secretion systems that are designed to penetrate host-cell membranes and insert pathogen proteins directly into the host cell's cytosol. However, the secretion systems' proteins and, in particular, their host targets are largely uncharacterized. Here, we used a combined in silico, in vitro, and in vivo approach to identify B. mallei proteins required for pathogenicity. We used bioinformatics tools, including orthology detection and ab initio predictions of secretion system proteins, as well as published experimental Burkholderia data to initially select a small number of proteins as putative virulence factors. We then used yeast two-hybrid assays against normalized whole human and whole murine proteome libraries to detect and identify interactions among each of these bacterial proteins and host proteins. Analysis of such interactions provided both verification of known virulence factors and identification of three new putative virulence proteins. We successfully created insertion mutants for each of these three proteins using the virulent B. mallei ATCC 23344 strain. We exposed BALB/c mice to mutant strains and the wild-type strain in an aerosol challenge model using lethal B. mallei doses. In each set of experiments, mice exposed to mutant strains survived for the 21-day duration of the experiment, whereas mice exposed to the wild-type strain rapidly died. Given their in vivo role in pathogenicity, and based on the yeast two-hybrid interaction data, these results point to the importance of these pathogen proteins in modulating host ubiquitination pathways, phagosomal escape, and actin-cytoskeleton rearrangement processes. PMID:23800426

Memiševi?, Vesna; Zavaljevski, Nela; Pieper, Rembert; Rajagopala, Seesandra V.; Kwon, Keehwan; Townsend, Katherine; Yu, Chenggang; Yu, Xueping; DeShazer, David; Reifman, Jaques; Wallqvist, Anders

2013-01-01

323

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

PubMed Central

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). PMID:24854358

Goh, Share Yuan; Tan, Wen-Si; Khan, Saad Ahmed; Chew, Hooi Pin; Kasim, Noor Hayaty Abu; Yin, Wai-Fong; Chan, Kok-Gan

2014-01-01

324

Colonization behavior of bacterium Burkholderia cepacia inside the Oryza sativa roots visualized using green fluorescent protein reporter  

Microsoft Academic Search

Colonization behavior of endophytic bacteria Burkholderia cepacia strains RRE-3 and RRE-5 was studied in the seedlings of rice variety NDR97 using confocal laser scanning microscopy under\\u000a controlled laboratory and greenhouse conditions. For studying colonization pattern, bacterial strains were tagged with pHRGFPGUS\\u000a plasmid. The role of bacterial strains (both gfp\\/gus-tagged and untagged) in growth promotion was also studied. After coming into

Shiveta Sharma; Shailendra Sharma; Ramesh K. Singh; Akhauri Vaishampayan

2008-01-01

325

Genome Sequence of the 2,4,5-Trichlorophenoxyacetate-Degrading Bacterium Burkholderia phenoliruptrix Strain AC1100  

PubMed Central

Burkholderia phenoliruptrix strain AC1100 (ATCC 53867) degrades a variety of recalcitrant xenobiotics, including 2,4,5-trichlorophenoxyacetate. The molecular mechanism of 2,4,5-trichlorophenoxyacetate degradation has been extensively studied. Here we present a 7.8-Mb assembly of the genome sequence of this 2,4,5-trichlorophenoxyacetate-degrading strain, which may provide useful information related to the degradation of chlorinated aromatic compounds. PMID:23929488

Yu, Hao; Chakrabarty, Ananda M.; Xun, Luying

2013-01-01

326

Evaluation of Trichoderma virens and Burkholderia cepacia for antagonistic activity against root-knot nematode, Meloidogyne incognita  

Microsoft Academic Search

Summary - The bacterium Burkholderia cepacia (strain Bc-2) and the fungus Trichoderma virens (strain Gl-3) were investigated for activity against the nematode Meloidogyne incognita . Culture é ltrates from Bc-2 and Gl-3 contained extracellular factors that inhibited egg hatch and second-stage juvenile (J2) mobility. Size fractionation results and lack of detectable chitinase or protease activities from Bc-2 and Gl-3 culture

Susan L. F. Meyer; Samia I. Massoud; David J. Chitwood; Daniel P. Roberts

2000-01-01

327

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

PubMed Central

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

Kikuchi, Yoshitomo; Fukatsu, Takema

2014-01-01

328

Saturation Mutagenesis of Toluene ortho-Monooxygenase of Burkholderia cepacia G4 for Enhanced 1Naphthol Synthesis and Chloroform Degradation  

Microsoft Academic Search

Directed evolution of toluene ortho-monooxygenase (TOM) of Burkholderia cepacia G4 previously created the hydroxylase -subunit (TomA3) V106A variant (TOM-Green) with increased activity for both trichloroethylene degradation (twofold enhancement) and naphthalene oxidation (six-times-higher activity). In the present study, saturation mutagenesis was performed at position A106 with Escherichia coli TG1\\/pBS(Kan)TOMV106A to improve TOM activity for both chloroform degradation and naphthalene oxidation. Whole

Lingyun Rui; Young Man Kwon; Ayelet Fishman; Kenneth F. Reardon; Thomas K. Wood

2004-01-01

329

Assessment of three Resistance-Nodulation-Cell Division drug efflux transporters of Burkholderia cenocepacia in intrinsic antibiotic resistance  

Microsoft Academic Search

BACKGROUND: Burkholderia cenocepacia are opportunistic Gram-negative bacteria that can cause chronic pulmonary infections in patients with cystic fibrosis. These bacteria demonstrate a high-level of intrinsic antibiotic resistance to most clinically useful antibiotics complicating treatment. We previously identified 14 genes encoding putative Resistance-Nodulation-Cell Division (RND) efflux pumps in the genome of B. cenocepacia J2315, but the contribution of these pumps to

Silvia Buroni; Maria R Pasca; Ronald S Flannagan; Silvia Bazzini; Anna Milano; Iris Bertani; Vittorio Venturi; Miguel A Valvano; Giovanna Riccardi

2009-01-01

330

Exploring the metabolic network of the epidemic pathogen Burkholderia cenocepacia J2315 via genome-scale reconstruction  

Microsoft Academic Search

Background  \\u000a Burkholderia cenocepacia is a threatening nosocomial epidemic pathogen in patients with cystic fibrosis (CF) or a compromised immune system. Its high\\u000a level of antibiotic resistance is an increasing concern in treatments against its infection. Strain B. cenocepacia J2315 is the most infectious isolate from CF patients. There is a strong demand to reconstruct a genome-scale metabolic network\\u000a of B.

Kechi Fang; Hansheng Zhao; Changyue Sun; Carolyn M C Lam; Suhua Chang; Kunlin Zhang; Gurudutta Panda; Miguel Godinho; Vítor A P Martins dos Santos; Jing Wang

2011-01-01

331

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

Microsoft Academic Search

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

Nelly Dubarry; Franck Pasta

2006-01-01

332

Identification of volatile compounds produced by the bacterium Burkholderia tropica that inhibit the growth of fungal pathogens  

PubMed Central

It has been documented that bacteria from the Burkholderia genera produce different kinds of compounds that inhibit plant pathogens, however in Burkholderia tropica, an endophytic diazotrophic and phosphate-solubilizing bacterium isolated from a wide diversity of plants, the capacity to produce antifungal compounds has not been evaluated. In order to expand our knowledge about Burkholderia tropica as a potential biological control agent, we analyzed 15 different strains of this bacterium to evaluate their capacities to inhibit the growth of four phytopathogenic fungi, Colletotrichum gloeosporioides, Fusarium culmorum, Fusarium oxysporum and Sclerotium rolffsi. Diverse analytical techniques, including plant root protection and dish plate growth assays and gas chromatography-mass spectroscopy showed that the fungal growth inhibition was intimately associated with the volatile compounds produced by B. tropica and, in particular, two bacterial strains (MTo293 and TTe203) exhibited the highest radial mycelial growth inhibition. Morphological changes associated with these compounds, such as disruption of fungal hyphae, were identified by using photomicrographic analysis. By using gas chromatography-mass spectroscopy technique, 18 volatile compounds involved in the growth inhibition mechanism were identified, including ?-pinene and limonene. In addition, we found a high proportion of bacterial strains that produced siderophores during growth with different carbon sources, such as alanine and glutamic acid; however, their roles in the antagonism mechanism remain unclear. PMID:23680857

Tenorio-Salgado, Silvia; Tinoco, Raunel; Vazquez-Duhalt, Rafael; Caballero-Mellado, Jesus; Perez-Rueda, Ernesto

2013-01-01

333

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

PubMed

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. PMID:24701243

Maida, Isabel; Lo Nostro, Antonella; Pesavento, Giovanna; Barnabei, Martina; Calonico, Carmela; Perrin, Elena; Chiellini, Carolina; Fondi, Marco; Mengoni, Alessio; Maggini, Valentina; Vannacci, Alfredo; Gallo, Eugenia; Bilia, Anna Rita; Flamini, Guido; Gori, Luigi; Firenzuoli, Fabio; Fani, Renato

2014-01-01

334

Burkholderia cepacia Complex Infection in Italian Patients with Cystic Fibrosis: Prevalence, Epidemiology, and Genomovar Status  

PubMed Central

The prevalence, epidemiology, and genomovar status of Burkholderia cepacia complex strains recovered from Italian cystic fibrosis (CF) patients were investigated using genetic typing and species identification methods. Four CF treatment centers were examined: two in Sicily, one in central Italy, and one in northern Italy. B. cepacia complex bacteria were isolated from 59 out of 683 CF patients attending these centers (8.6%). For the two geographically related treatment centers in Sicily, there was a high incidence of infection caused by a single epidemic clone possessing the cblA gene and belonging to B. cepacia genomovar III, recA group III-A, closely related to the major North America-United Kingdom clone, ET12; instability of the cblA sequence was also demonstrated for clonal isolates. In summary, of all the strains of B. cepacia encountered in the Italian CF population, the genomovar III, recA group III-A strains were the most prevalent and transmissible. However, patient-to-patient spread was also observed with several other genomovars, including strains of novel taxonomic status within the B. cepacia complex. A combination of genetic identification and molecular typing analysis is recommended to fully define specific risks posed by the genomovar status of strains within the B. cepacia complex. PMID:11474009

Agodi, Antonella; Mahenthiralingam, Eshwar; Barchitta, Martina; Giannino, Viviana; Sciacca, Agata; Stefani, Stefania

2001-01-01

335

Lung transplant for a patient with cystic fibrosis and active Burkholderia cenocepacia pneumonia.  

PubMed

Lung transplant for cystic fibrosis has been considered contraindicated in patients who have Burkholderia Cenocepacia infection. A 24-year-old white woman who had cystic fibrosis presented with respiratory failure caused by B. Cenocepacia pneumonia. She was treated with broad-spectrum antibiotics and a double-lung transplant. The chest cavity and both bronchi were irrigated with 0.5% povidone-iodine solution. For immunosuppression, she received induction therapy with alemtuzumab (15 mg) and methylprednisolone and maintenance therapy with tacrolimus, mycophenolate mofetil, and prednisone (5 mg daily). Postoperative antibiotics included intravenous meropenem for 3 weeks; vancomycin for 10 days; and inhaled ceftazidime, oral trimethoprim-sulfamethoxazole, and doxycycline for several months. Follow-up at 25 months after transplant showed that chest radiographs were clear and lung function was normal. At 6 years after transplant, she was working full time and had no recurrence of infection from B. Cenocepacia. This case suggests that patients who have cystic fibrosis and active B. Cenocepacia pneumonia may be successfully treated with a lung transplant. PMID:25299375

Salizzoni, Stefano; Pilewski, Joseph; Toyoda, Yoshiya

2014-10-01

336

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

PubMed Central

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. PMID:24701243

Lo Nostro, Antonella; Calonico, Carmela; Perrin, Elena; Chiellini, Carolina; Fondi, Marco; Mengoni, Alessio; Vannacci, Alfredo; Bilia, Anna Rita; Gori, Luigi

2014-01-01

337

A two-tier model of polymyxin B resistance in Burkholderia cenocepacia.  

PubMed

Burkholderia cenocepacia is an environmental bacterium causing serious human opportunistic infections and is extremely resistant to multiple antibiotics including antimicrobial peptides, such as polymyxin B (PmB). Extreme antibiotic resistance is attributed to outer membrane impermeability ('intrinsic' resistance). Previous work showed that production of full-length lipopolysaccharide (LPS) prevents surface binding of PmB. We hypothesized that two tiers of resistance mechanisms rendering different thresholds of PmB resistance exist in B. cenocepacia. To test this notion, candidate genes were mutated in two isogenic strains expressing full-length LPS or truncated LPS devoid of heptose ('heptoseless LPS') respectively. We uncovered various proteins required for PmB resistance only in the strain with heptoseless LPS. These proteins are not involved in preventing PmB binding to whole cells or permeabilization of the outer membrane. Our results support a two-tier model of PmB resistance in B. cenocepacia. One tier sets a very high threshold mediated by the LPS and the outer membrane permeability barrier. The second tier sets a lower threshold that may play a role in PmB resistance only when outer membrane permeability is compromised. This model may be of general applicability to understanding the high antimicrobial peptide resistance of environmental opportunistic pathogens. PMID:23761261

Loutet, Slade A; Mussen, Lauren E; Flannagan, Ronald S; Valvano, Miguel A

2011-04-01

338

Oxidation of aminonitrotoluenes by 2,4-DNT dioxygenase of Burkholderia sp. strain DNT.  

PubMed

Aminonitrotoluenes form rapidly from the reduction of dinitrotoluenes (DNTs) which are priority pollutants and animal carcinogens. For example, 4-amino-2-nitrotoluene (4A2NT) and 2A4NT accumulate from the reduction of 2,4-DNT during its aerobic biodegradation. Here, we show that 2,4-DNT dioxygenase (DDO) from Burkholderia sp. strain DNT oxidizes the aminonitrotoluenes 2A3NT, 2A6NT, 4A3NT, and 5A2NT to 2-amino-3-nitrobenzylalcohol, 2-amino-4-nitro-m-cresol and 3-amino-5-nitro-p-cresol, 4-amino-3-nitrobenzylalcohol and aminonitrocresol, and 2-amino-5-nitro-o-cresol, respectively. 2A5NT and 3A4NT are oxidized to aminonitrocresols and/or aminonitrobenzylalcohols, and 4A2NT is oxidized to aminonitrocresol. Only 2A4NT, a reduced compound derived from 2,4-DNT, was not oxidized by DDO or its three variants. The alpha subunit mutation I204Y resulted in two to fourfold faster oxidization of the aminonitrotoluenes. Though these enzymes are dioxygenases, they acted like monooxygenases by adding a single hydroxyl group, which did not result in the release of nitrite. PMID:16315327

Leungsakul, Thammajun; Keenan, Brendan G; Mori, Masa-aki; Morton, Martha D; Stuart, James D; Smets, Barth F; Wood, Thomas K

2006-02-01

339

Biodegradation of Mono-Hydroxylated PCBs by Burkholderia xenovorans LB400  

PubMed Central

Three hydroxylated derivatives of PCBs, including 2?-hydroxy-4-chlorobiphenyl (2?-OH-4-CB), 3?-hydroxy-4-chlorobiphenyl (3?-OH-4-CB), and 4?-hydroxy-4-chlorobiphenyl (4?-OH-4-CB), were transformed by the PCB degrader, Burkholderia xenovorans LB400. When the bacterium was growing on biphenyl (biphenyl pathway-inducing conditions), all three hydroxylated isomers were significantly transformed. On the contrary, only 2?-OH-4-CB was transformed by the bacterium growing on succinate (conditions non-inductive of the biphenyl pathway). Gene expression analyses showed a strong induction of key genes of the biphenyl pathway (bph) when cells were grown on biphenyl, which is consistent with the transformation of the three isomers by biphenyl-grown cells. When cells were grown on succinate, only exposure to 2?-OH-4-CB resulted in expression of biphenyl pathway genes, which suggests that this isomer was capable of inducing the biphenyl pathway. These results provide the first evidence that bacteria are able to metabolize PCB derivatives hydroxylated on the non-chlorinated ring. PMID:22918793

Tehrani, Rouzbeh; Lyv, Monica M.; Kaveh, Rashid; Schnoor, Jerald L.; Aken, Benoit Van

2013-01-01

340

Requirement of DNA Repair Mechanisms for Survival of Burkholderia cepacia G4 upon Degradation of Trichloroethylene  

PubMed Central

A Tn5-based mutagenesis strategy was used to generate a collection of trichloroethylene (TCE)-sensitive (TCS) mutants in order to identify repair systems or protective mechanisms that shield Burkholderia cepacia G4 from the toxic effects associated with TCE oxidation. Single Tn5 insertion sites were mapped within open reading frames putatively encoding enzymes involved in DNA repair (UvrB, RuvB, RecA, and RecG) in 7 of the 11 TCS strains obtained (4 of the TCS strains had a single Tn5 insertion within a uvrB homolog). The data revealed that the uvrB-disrupted strains were exceptionally susceptible to killing by TCE oxidation, followed by the recA strain, while the ruvB and recG strains were just slightly more sensitive to TCE than the wild type. The uvrB and recA strains were also extremely sensitive to UV light and, to a lesser extent, to exposure to mitomycin C and H2O2. The data from this study establishes that there is a link between DNA repair and the ability of B. cepacia G4 cells to survive following TCE transformation. A possible role for nucleotide excision repair and recombination repair activities in TCE-damaged cells is discussed. PMID:11722883

Yeager, Chris M.; Bottomley, Peter J.; Arp, Daniel J.

2001-01-01

341

Experimental Adaptation of Burkholderia cenocepacia to Onion Medium Reduces Host Range ? † ‡  

PubMed Central

It is unclear whether adaptation to a new host typically broadens or compromises host range, yet the answer bears on the fate of emergent pathogens and symbionts. We investigated this dynamic using a soil isolate of Burkholderia cenocepacia, a species that normally inhabits the rhizosphere, is related to the onion pathogen B. cepacia, and can infect the lungs of cystic fibrosis patients. We hypothesized that adaptation of B. cenocepacia to a novel host would compromise fitness and virulence in alternative hosts. We modeled adaptation to a specific host by experimentally evolving 12 populations of B. cenocepacia in liquid medium composed of macerated onion tissue for 1,000 generations. The mean fitness of all populations increased by 78% relative to the ancestor, but significant variation among lines was observed. Populations also varied in several phenotypes related to host association, including motility, biofilm formation, and quorum-sensing function. Together, these results suggest that each population adapted by fixing different sets of adaptive mutations. However, this adaptation was consistently accompanied by a loss of pathogenicity to the nematode Caenorhabditis elegans; by 500 generations most populations became unable to kill nematodes. In conclusion, we observed a narrowing of host range as a consequence of prolonged adaptation to an environment simulating a specific host, and we suggest that emergent pathogens may face similar consequences if they become host-restricted. PMID:20154121

Ellis, Crystal N.; Cooper, Vaughn S.

2010-01-01

342

Bioactive and Structural Metabolites of Pseudomonas and Burkholderia Species Causal Agents of Cultivated Mushrooms Diseases1  

PubMed Central

Pseudomonas tolaasii, P. reactans and Burkholderia gladioli pv. agaricicola, are responsible of diseases on some species of cultivated mushrooms. The main bioactive metabolites produced by both Pseudomonas strains are the lipodepsipeptides (LDPs) tolaasin I and II and the so called White Line Inducing Principle (WLIP), respectively, LDPs which have been extensively studied for their role in the disease process and for their biological properties. In particular, their antimicrobial activity and the alteration of biological and model membranes (red blood cell and liposomes) was established. In the case of tolaasin I interaction with membranes was also related to the tridimensional structure in solution as determined by NMR combined with molecular dynamic calculation techniques. Recently, five news minor tolaasins, tolaasins A–E, were isolated from the culture filtrates of P. tolaasii and their chemical structure was determined by extensive use of NMR and MS spectroscopy. Furthermore, their antimicrobial activity was evaluated on target micro-organisms (fungi—including the cultivated mushrooms Agaricus bisporus, Lentinus edodes, and Pleurotus spp.—chromista, yeast and bacteria). The Gram positive bacteria resulted the most sensible and a significant structure-activity relationships was apparent. The isolation and structure determination of bioactive metabolites produced by B. gladioli pv. agaricicola are still in progress but preliminary results indicate their peptide nature. Furthermore, the exopolysaccharide (EPS) from the culture filtrates of B. gladioli pv. agaricicola, as well as the O-chain and lipid A, from the lipopolysaccharide (LPS) of the three bacteria, were isolated and the structures determined. PMID:19787100

Andolfi, Anna; Cimmino, Alessio; Cantore, Pietro Lo; Iacobellis, Nicola Sante; Evidente, Antonio

2008-01-01

343

A Murine Model for Infection with Burkholderia cepacia with Sustained Persistence in the Spleen  

PubMed Central

Burkholderia cepacia is an opportunistic pathogen that causes severe systemic infections in patients with chronic granulomatous disease (CGD) or with cystic fibrosis (CF), but its mechanisms of virulence are poorly understood. We developed a murine model of systemic infection in wild-type (WT) and gamma interferon knockout (GKO) BALB/c mice to facilitate dissection of components of pathogenicity and host defense. Both WT and GKO mice were susceptible to chronic splenic infection with B. cepacia, but not with Pseudomonas aeruginosa. B. cepacia strains from patients with CGD persisted longer than those from CF patients. C57BL/6 mice were the most susceptible murine strain; bacteria persisted in the spleen for 2 months. DBA/2, BALB/c, and A/J strains of mice were relatively resistant to infection. Certain strains of B. cepacia complex can persist in the murine spleen after systemic infection; this may provide clues to its virulence in compromised hosts, such as those with CGD and CF. PMID:10417170

Speert, David P.; Steen, Barbara; Halsey, Keith; Kwan, Eddie

1999-01-01

344

Burkholderia cenocepacia conditional growth mutant library created by random promoter replacement of essential genes  

PubMed Central

Identification of essential genes by construction of conditional knockouts with inducible promoters allows the identification of essential genes and creation of conditional growth (CG) mutants that are then available as genetic tools for further studies. We used large-scale transposon delivery of the rhamnose-inducible promoter, PrhaB followed by robotic screening of rhamnose-dependent growth to construct a genomic library of 106 Burkholderia cenocepacia CG mutants. Transposon insertions were found where PrhaB was in the same orientation of widely conserved, well-characterized essential genes as well as genes with no previous records of essentiality in other microorganisms. Using previously reported global gene-expression analyses, we demonstrate that PrhaB can achieve the wide dynamic range of expression levels required for essential genes when the promoter is delivered randomly and mutants with rhamnose-dependent growth are selected. We also show specific detection of the target of an antibiotic, novobiocin, by enhanced sensitivity of the corresponding CG mutant (PrhaB controlling gyrB expression) within the library. Modulation of gene expression to achieve 30–60% of wild-type growth created conditions for specific hypersensitivity demonstrating the value of the CG mutant library for chemogenomic experiments. In summary, CG mutants can be obtained on a large scale by random delivery of a tightly regulated inducible promoter into the bacterial chromosome followed by a simple screening for the CG phenotype, without previous information on gene essentiality. PMID:23389959

Bloodworth, Ruhi A M; Gislason, April S; Cardona, Silvia T

2013-01-01

345

Involvement of toxin-antitoxin modules in Burkholderia cenocepacia biofilm persistence.  

PubMed

Biofilms are involved in the recalcitrance of infections due to the presence of persister cells. Although the molecular basis of persistence is still largely unknown, toxin-antitoxin modules (TA) are thought to play a role in this process. In this study, we investigated whether TA modules contribute to persistence toward antibiotics in Burkholderia cenocepacia J2315. Sixteen pairs of genes were identified based on their apparent similarity to TA modules. Overexpression of the putative toxins had various effects on growth, persistence, and biofilm formation. Toxins, whose overexpression resulted in growth inhibition, often increased the number of surviving persisters; in contrast, overexpression of putative toxins showing no effects on growth had no positive influence on the number of surviving persisters. Furthermore, the expression of the TA modules was compared between treated and untreated sessile and planktonic wild-type cultures. For 10 toxin-encoding genes, the expression was higher in untreated sessile cells than in untreated planktonic cells. Nine toxin-encoding genes were upregulated after treatment with tobramycin, but none after treatment with ciprofloxacin. These results indicate that most, but not all TA modules contribute to persistence in B. cenocepacia J2315 and that this contribution depends on the mode of growth and the antibiotic used. PMID:24719230

Van Acker, Heleen; Sass, Andrea; Dhondt, Inne; Nelis, Hans J; Coenye, Tom

2014-08-01

346

Disinfection of Burkholderia cepacia complex from non-touch taps in a neonatal nursery.  

PubMed

Burkholderia cepacia complex (Bcc) comprises nine closely related species or genomovars. It is an important causative agent of opportunistic infections and waterborne nosocomial infections. B. cepacia (formerly genomovar I) was identified from the blood culture of a baby in our neonatal unit (NU) in March 2005. B. cepacia was isolated four times from clinical specimens since the introduction of non-touch taps in the NU from 2000 to 2005 and only once from 1994 to 2000. Environmental samples were collected from the NU, including tap water from non-touch taps. Clinical and environmental isolates of Bcc were characterized using molecular identification and strain typing. A literature review was undertaken to delineate a method for eradication of Bcc. Several variations for hot water eradication of the organism from the taps were attempted. Genotyping and molecular analysis revealed that tap water isolates were B. cenocepacia which was a different species from the B. cepacia isolated from blood cultures of the neonate. However, B. cenocepacia has been known to cause nosocomial outbreaks and it was eventually eradicated from the NU by using repeated thermal shock (hot water at 65 degrees C for 10 min), changing taps and decolonizing sinks with hypochlorite. Molecular typing is useful in assisting the investigation of Bcc nosocomial infections. PMID:18576933

Kotsanas, Despina; Brett, Judith; Kidd, Tim J; Stuart, Rhonda L; Korman, Tony M

2008-01-01

347

In Vitro Antifungal Activity of Burkholderia gladioli pv. agaricicola against Some Phytopathogenic Fungi  

PubMed Central

The trend to search novel microbial natural biocides has recently been increasing in order to avoid the environmental pollution from use of synthetic pesticides. Among these novel natural biocides are the bioactive secondary metabolites of Burkholderia gladioli pv. agaricicola (Bga). The aim of this study is to determine antifungal activity of Bga strains against some phytopathogenic fungi. The fungicidal tests were carried out using cultures and cell-free culture filtrates against Botrytis cinerea, Aspergillus flavus, Aspergillus niger, Penicillium digitatum, Penicillium expansum, Sclerotinia sclerotiorum and Phytophthora cactorum. Results demonstrated that all tested strains exert antifungal activity against all studied fungi by producing diffusible metabolites which are correlated with their ability to produce extracellular hydrolytic enzymes. All strains significantly reduced the growth of studied fungi and the bacterial cells were more bioactive than bacterial filtrates. All tested Bulkholderia strains produced volatile organic compounds (VOCs), which inhibited the fungal growth and reduced the growth rate of Fusarium oxysporum and Rhizoctonia solani. GC/MS analysis of VOCs emitted by strain Bga 11096 indicated the presence of a compound that was identified as 1-methyl-4-(1-methylethenyl)-cyclohexene, a liquid hydrocarbon classified as cyclic terpene. This compound could be responsible for the antifungal activity, which is also in agreement with the work of other authors. PMID:23208371

Elshafie, Hazem S.; Camele, Ippolito; Racioppi, Rocco; Scrano, Laura; Iacobellis, Nicola S.; Bufo, Sabino A.

2012-01-01

348

Molecular and catalytic properties of 2,4'-dihydroxyacetophenone dioxygenase from Burkholderia sp. AZ11.  

PubMed

The gene dad encoding 2,4'-dihydroxyacetophenone (DHAP) dioxygenase was cloned from Burkholderia sp. AZ11. The initiation codon GTG was converted to ATG for high-level expression of the enzyme in Escherichia coli. The enzyme was moderately thermostable, and the recombinant enzyme was briefly purified. The enzyme (M(r)=90 kDa) was a homotetramer with a subunit M(r) of 23 kDa. It contained 1.69 mol of non-heme iron, and had a dark gray color. On anaerobic incubation of it with DHAP, the absorption at around 400 nm increased due to the formation of an enzyme-DHAP complex. Multiple sequence alignment suggested that His77, His79, His115, and Glu96 in the cupin fold were possible metal ligands. The apparent K(m) for DHAP and the apparent V(max) were estimated to be 1.60 µM and 6.28 µmol/min/mg respectively. 2-Hydroxyacetophenone was a poor substrate. CuCl(2) and HgCl(2) strongly inhibited the enzyme, while FeSO(4) weakly activated it. PMID:22451401

Enya, Mayu; Aoyagi, Keiko; Hishikawa, Yoshihiro; Yoshimura, Azusa; Mitsukura, Koichi; Maruyama, Kiyofumi

2012-01-01

349

Sorption versus Biomineralization of Pb(II) within Burkholderia cepacia Biofilms  

SciTech Connect

X-ray spectroscopy measurements have been combined with macroscopic uptake data and transmission electron microscopy (TEM) results to show that Pb(II) uptake by Burkholderia cepacia is due to simultaneous sorption and biomineralization processes. X-ray microprobe mapping of B. cepacia biofilms formed on -Al2O3 surfaces shows that Pb(II) is distributed heterogeneously throughout the biofilms because of the formation of Pb "hot spots". EXAFS data and TEM observations show that the enhanced Pb accumulation is due to the formation of nanoscale crystals of pyromorphite (Pb5(PO4)3(OH)) adjacent to the outer-membrane of a fraction of the total population of B. cepacia cells. In contrast, B. cepacia cell suspensions or biofilms that were heat-killed or pretreated with X-rays do not form pyromorphite, which suggests that metabolic activity is required. Precipitation of pyromorphite occurs over several orders of magnitude in [H+] and [Pb] and accounts for approximately 90% of the total Pb uptake below pH 4.5 but only 45-60% at near-neutral pH because of the formation of additional Pb(II) adsorption complexes. Structural fits of Pb LIII EXAFS data collected for heat-treated cells at near-neutral pH suggest that Pb(II) forms inner-sphere adsorption complexes with carboxyl functional groups in the biofilms.

Templeton, Alexis S.; Trainor, T. P.; Spormann, Alfred M.; Newville, Mathew; Sutton , Steven R.; Dohnalkova, Alice; Gorby, Yuri A.; Brown, Gordon E.

2003-01-15

350

High-temperature-induced transposition of insertion elements in burkholderia multivorans ATCC 17616.  

PubMed

An efficient and quantitative method to analyze the transposition of various insertion sequence (IS) elements in Burkholderia multivorans ATCC 17616 was devised. pGEN500, a plasmid carrying a Bacillus subtilis-derived sacB gene, was introduced into ATCC 17616 cells, and 25% of their sucrose-resistant derivatives were found to carry various IS elements on pGEN500. A PCR-based experimental protocol, in which a mixture of several specific primer pairs was used, revealed that pGEN500 captured, in addition to five previously reported IS elements (IS401, IS402, IS406, IS407, and IS408), three novel IS elements, ISBmu1, ISBmu2, and ISBmu3. The global transposition frequency of these IS elements was enhanced more than sevenfold under a high-temperature condition (42 degrees C) but not under oxidative stress or starvation conditions. To our knowledge, this is the first report demonstrating the elevated transposition activities of several IS elements at a high temperature. The efficient experimental protocol developed in this study will be useful in quantitatively and simultaneously investigating various IS elements, as well as in capturing novel functional mobile elements from a wide variety of bacteria. PMID:15812007

Ohtsubo, Yoshiyuki; Genka, Hiroyuki; Komatsu, Harunobu; Nagata, Yuji; Tsuda, Masataka

2005-04-01

351

Screening of Burkholderia sp. WGB31 producing anisic acid from anethole and optimization of fermentation conditions.  

PubMed

Anisic acid, the precursor of a variety of food flavors and industrial raw materials, can be bioconversed from anethole which extracted from star anise fruits. WGB31 strain with anisic acid molar production rate of 10.25% was isolated and identified as Burkholderia sp. Three significant influential factors, namely, glucose concentration, initial pH value, and medium volume were selected and their effects were evaluated by Box-Behnken Design (BBD). Regression analysis was performed to determine response surface methodology and the significance was tested to obtain the process model of optimal conditions for producing anisic acid. The fermentation conditions at the stable point of the model were obtained: glucose 6?g?L(-1) , pH 6.2, culture medium volume 61?mL in a triangular flask with 250?ml volume. Verification test indicated that the production rate of anisic acid was 30.7%, which was three times of that before optimizing. The results provide a basis and reference for producing anisic acid by microbial transformation. PMID:25100156

Shen, Peihong; Song, Zhangyang; Zhang, Zhenyong; Zeng, Huahe; Tang, Xianlai; Jiang, Chengjian; Li, Junfang; Wu, Bo

2014-11-01

352

Localization of Burkholderia cepacia Complex Bacteria in Cystic Fibrosis Lungs and Interactions with Pseudomonas aeruginosa in Hypoxic Mucus.  

PubMed

The localization of Burkholderia cepacia complex (Bcc) bacteria in cystic fibrosis (CF) lungs, alone or during coinfection with Pseudomonas aeruginosa, is poorly understood. We performed immunohistochemistry for Bcc and P. aeruginosa bacteria on 21 coinfected or singly infected CF lungs obtained at transplantation or autopsy. Parallel in vitro experiments examined the growth of two Bcc species, Burkholderia cenocepacia and Burkholderia multivorans, in environments similar to those occupied by P. aeruginosa in the CF lung. Bcc bacteria were predominantly identified in the CF lung as single cells or small clusters within phagocytes and mucus but not as "biofilm-like structures." In contrast, P. aeruginosa was identified in biofilm-like masses, but densities appeared to be reduced during coinfection with Bcc bacteria. Based on chemical analyses of CF and non-CF respiratory secretions, a test medium was defined to study Bcc growth and interactions with P. aeruginosa in an environment mimicking the CF lung. When test medium was supplemented with alternative electron acceptors under anaerobic conditions, B. cenocepacia and B. multivorans used fermentation rather than anaerobic respiration to gain energy, consistent with the identification of fermentation products by high-performance liquid chromatography (HPLC). Both Bcc species also expressed mucinases that produced carbon sources from mucins for growth. In the presence of P. aeruginosa in vitro, both Bcc species grew anaerobically but not aerobically. We propose that Bcc bacteria (i) invade a P. aeruginosa-infected CF lung when the airway lumen is anaerobic, (ii) inhibit P. aeruginosa biofilm-like growth, and (iii) expand the host bacterial niche from mucus to also include macrophages. PMID:25156735

Schwab, Ute; Abdullah, Lubna H; Perlmutt, Olivia S; Albert, Daniel; Davis, C William; Arnold, Roland R; Yankaskas, James R; Gilligan, Peter; Neubauer, Heiner; Randell, Scott H; Boucher, Richard C

2014-11-01

353

Bacterial Cell Wall Synthesis Gene uppP Is Required for Burkholderia Colonization of the Stinkbug Gut  

PubMed Central

To establish a host-bacterium symbiotic association, a number of factors involved in symbiosis must operate in a coordinated manner. In insects, bacterial factors for symbiosis have been poorly characterized at the molecular and biochemical levels, since many symbionts have not yet been cultured or are as yet genetically intractable. Recently, the symbiotic association between a stinkbug, Riptortus pedestris, and its beneficial gut bacterium, Burkholderia sp., has emerged as a promising experimental model system, providing opportunities to study insect symbiosis using genetically manipulated symbiotic bacteria. Here, in search of bacterial symbiotic factors, we targeted cell wall components of the Burkholderia symbiont by disruption of uppP gene, which encodes undecaprenyl pyrophosphate phosphatase involved in biosynthesis of various bacterial cell wall components. Under culture conditions, the ?uppP mutant showed higher susceptibility to lysozyme than the wild-type strain, indicating impaired integrity of peptidoglycan of the mutant. When administered to the host insect, the ?uppP mutant failed to establish normal symbiotic association: the bacterial cells reached to the symbiotic midgut but neither proliferated nor persisted there. Transformation of the ?uppP mutant with uppP-encoding plasmid complemented these phenotypic defects: lysozyme susceptibility in vitro was restored, and normal infection and proliferation in the midgut symbiotic organ were observed in vivo. The ?uppP mutant also exhibited susceptibility to hypotonic, hypertonic, and centrifugal stresses. These results suggest that peptidoglycan cell wall integrity is a stress resistance factor relevant to the successful colonization of the stinkbug midgut by Burkholderia symbiont. PMID:23747704

Kim, Jiyeun Kate; Lee, Ho Jin; Kikuchi, Yoshitomo; Kitagawa, Wataru; Nikoh, Naruo

2013-01-01

354

Exceptionally High Representation of Burkholderia cepacia among B. cepacia Complex Isolates Recovered from the Major Portuguese Cystic Fibrosis Center?  

PubMed Central

Burkholderia cepacia, a species found infrequently in cystic fibrosis (CF), was isolated from 85% of patients infected with bacteria of the B. cepacia complex that visited the major Portuguese CF center, in Lisbon, during 2003 to 2005. A detailed molecular analysis revealed that this was mainly due to two B. cepacia clones. These clones were indistinguishable from two strains isolated from intrinsically contaminated nonsterile saline solutions for nasal application, detected during routine market surveillance by the Portuguese Medicines and Health Products Authority. PMID:17360834

Cunha, Monica V.; Pinto-de-Oliveira, Ana; Meirinhos-Soares, Luis; Salgado, Maria Jose; Melo-Cristino, Jose; Correia, Susana; Barreto, Celeste; Sa-Correia, Isabel

2007-01-01

355

Draft genomes of three Antarctic Psychrobacter strains producing antimicrobial compounds against Burkholderia cepacia complex, opportunistic human pathogens.  

PubMed

Herein we present the draft genomes of three Psychrobacter strains isolated from Antarctic sponges and able to inhibit the growth of bacteria belonging to the Burkholderia cepacia complex, responsible for infections of the respiratory system in patients affected by Cystic Fibrosis. The comparative analysis of the annotated genomes of these Psychrobacter strains highlighted their differences in terms of overall genomic content (e.g. shared gene sets) and allowed the identification of gene clusters hypothetically involved in the biosynthesis of antimicrobial compounds. PMID:24401162

Fondi, Marco; Orlandini, Valerio; Perrin, Elena; Maida, Isabel; Bosi, Emanuele; Papaleo, Maria Cristiana; Michaud, Luigi; Lo Giudice, Angelina; de Pascale, Donatella; Tutino, Maria Luisa; Liò, Pietro; Fani, Renato

2014-02-01

356

Tom, a new aromatic degradative plasmid from Burkholderia (Pseudomonas) cepacia G4  

SciTech Connect

Burkholderia (Pseudomonas) cepacia PR1{sub 23} has been shown to constitutively express a toluene catabolic pathway distinguished by a unique toluene ortho-monooxygenase (Tom). This strain has also been shown to contain two extrachromosomal elements of <70 and> 100 kb. A derivative strain cured of the largest plasmid, PR1{sub 23} Cure, was unable to grow on phenol or toluene as the sole source of carbon and energy, which requires expression of the Tom pathway. Transfer of the larger plasmid from strain G4 J(the parent strain inducible for Tom) enabled PR1{sub 23} Cure to grow on toluene or phenol via inducible Tom pathway expression. Conjugal transfer of TOM{sub 23c} from PR1{sub 23} to an antibiotic-resistant derivative of PR1{sub 23} Cure enabled the transconjugant to grow with either phenol or toluene as the sole source of carbon and energy through constitutive expression of the Tom pathway. A cloned 11.2-kb EcoRI restriction fragment of Tom{sub 23c} resulted in the expression of both Tom and catechol 2,3-dioxygenase in Escherichia coli, as evidenced by its ability to oxidize trichloroethylene, toluene, m-cresol, o-cresol, phenol, and catechol. The largest resident plasmid of PR1 was identified as the source of these genes by DNA hybridization. These results indicate that the genes which encode Tom and catechol 2,3-dioxygenase are located on TOM, an approximately 108-kb degradative plasmid of B. cepacia G4. 35 refs., 3 figs., 3 tabs.

Shields, M.S.; Reagin, J.J.; Campbell, R. [Univ. of West Florida, Pensacola, FL (United States)] [and others

1995-04-01

357

Tracking the Response of Burkholderia cepacia G4 5223-PR1 in Aquifer Microcosms  

PubMed Central

The introduction of bacteria into the environment for bioremediation purposes (bioaugmentation) requires analysis and monitoring of microbial population dynamics to define persistence and activity from both efficacy and risk assessment perspectives. Burkholderia cepacia G4 5223-PR1 is a Tn5 insertion mutant which constitutively expresses a toluene ortho-monooxygenase that degrades trichloroethylene (TCE). This ability of G4 5223-PR1 to degrade TCE without aromatic induction may be useful for bioremediation of TCE-containing aquifers and groundwater. Thus, a simulated aquifer sediment system and groundwater microcosms were used to monitor the survival of G4 5223-PR1. The fate of G4 5223-PR1 in sediment was monitored by indirect immunofluorescence microscopy, a colony blot assay, and growth on selective medium. G4 5223-PR1 was detected immunologically by using a highly specific monoclonal antibody which reacted against the O-specific polysaccharide chain of the lipopolysaccharides of this organism. G4 5223-PR1 survived well in sterilized groundwater, although in nonsterile groundwater microcosms rapid decreases in the G4 5223-PR1 cell population were observed. Ten days after inoculation no G4 5223-PR1 cells could be detected by selective plating or immunofluorescence. G4 5223-PR1 survival was greater in a nonsterile aquifer sediment microcosm, although after 22 days of elution the number of G4 5223-PR1 cells was low. Our results demonstrate the utility of monoclonal antibody tracking methods and the importance of biotic interactions in determining the persistence of introduced microorganisms. PMID:16534928

Winkler, J.; Timmis, K. N.; Snyder, R. A.

1995-01-01

358

X-Ray crystallographic and mutational studies of fluoroacetate dehalogenase from Burkholderia sp. strain FA1.  

PubMed

Fluoroacetate dehalogenase catalyzes the hydrolytic defluorination of fluoroacetate to produce glycolate. The enzyme is unique in that it catalyzes the cleavage of a carbon-fluorine bond of an aliphatic compound: the bond energy of the carbon-fluorine bond is among the highest found in natural products. The enzyme also acts on chloroacetate, although much less efficiently. We here determined the X-ray crystal structure of the enzyme from Burkholderia sp. strain FA1 as the first experimentally determined three-dimensional structure of fluoroacetate dehalogenase. The enzyme belongs to the alpha/beta hydrolase superfamily and exists as a homodimer. Each subunit consists of core and cap domains. The catalytic triad, Asp104-His271-Asp128, of which Asp104 serves as the catalytic nucleophile, was found in the core domain at the domain interface. The active site was composed of Phe34, Asp104, Arg105, Arg108, Asp128, His271, and Phe272 of the core domain and Tyr147, His149, Trp150, and Tyr212 of the cap domain. An electron density peak corresponding to a chloride ion was found in the vicinity of the N(epsilon1) atom of Trp150 and the N(epsilon2) atom of His149, suggesting that these are the halide ion acceptors. Site-directed replacement of each of the active-site residues, except for Trp150, by Ala caused the total loss of the activity toward fluoroacetate and chloroacetate, whereas the replacement of Trp150 caused the loss of the activity only toward fluoroacetate. An interaction between Trp150 and the fluorine atom is probably an absolute requirement for the reduction of the activation energy for the cleavage of the carbon-fluorine bond. PMID:19218394

Jitsumori, Keiji; Omi, Rie; Kurihara, Tatsuo; Kurata, Atsushi; Mihara, Hisaaki; Miyahara, Ikuko; Hirotsu, Ken; Esaki, Nobuyoshi

2009-04-01

359

Survey of Bartonella spp. in U.S. Bed Bugs Detects Burkholderia multivorans but Not Bartonella  

PubMed Central

Bed bugs (Cimex lectularius L.) have resurged in the United States and globally. Bed bugs are hematophagous ectoparasites of humans and other animals, including domestic pets, chickens, and bats, and their blood feeding habits contribute to their potential as disease vectors. Several species of Bartonella are re-emergent bacterial pathogens that also affect humans, domestic pets, bats and a number of other wildlife species. Because reports of both bed bugs and Bartonella have been increasing in the U.S., and because their host ranges can overlap, we investigated whether the resurgences of these medically important pathogens and their potential vector might be linked, by screening for Bartonella spp. in bed bugs collected from geographic areas where these pathogens are prevalent and from bed bugs that have been in culture in the laboratory for several years. We screened a total of 331 bed bugs: 316 bed bugs from 36 unique collections in 29 geographic locations in 13 states, 10 bed bugs from two colonies maintained in the laboratory for 3 yr, and 5 bed bugs from a colony that has been in culture since before the recent resurgence of bed bugs. Bartonella spp. DNA was screened using a polymerase chain reaction assay targeting the 16S–23S rRNA intergenic transcribed spacer region. Bartonella DNA was not amplified from any bed bug, but five bed bugs from four different apartments of an elderly housing building in North Carolina contained DNA sequences that corresponded to Burkholderia multivorans, an important pathogen in nosocomial infections that was not previously linked to an arthropod vector. PMID:24040015

Saenz, Virna L.; Maggi, Ricardo G.; Breitschwerdt, Edward B.; Kim, Jung; Vargo, Edward L.; Schal, Coby

2013-01-01

360

Fosmidomycin decreases membrane hopanoids and potentiates the effects of colistin on Burkholderia multivorans clinical isolates.  

PubMed

Burkholderia cepacia complex (Bcc) pulmonary infections in people living with cystic fibrosis (CF) are difficult to treat because of the extreme intrinsic resistance of most isolates to a broad range of antimicrobials. Fosmidomycin is an antibacterial and antiparasitic agent that disrupts the isoprenoid biosynthesis pathway, a precursor to hopanoid biosynthesis. Hopanoids are involved in membrane stability and contribute to polymyxin resistance in Bcc bacteria. Checkerboard MIC assays determined that although isolates of the Bcc species B. multivorans were highly resistant to treatment with fosmidomycin or colistin (polymyxin E), antimicrobial synergy was observed in certain isolates when the antimicrobials were used in combination. Treatment with fosmidomycin decreased the MIC of colistin for isolates as much as 64-fold to as low as 8 ?g/ml, a concentration achievable with colistin inhalation therapy. A liquid chromatography-tandem mass spectrometry technique was developed for the accurate quantitative determination of underivatized hopanoids in total lipid extracts, and bacteriohopanetetrol cyclitol ether (BHT-CE) was found to be the dominant hopanoid made by B. multivorans. The amount of BHT-CE made was significantly reduced upon fosmidomycin treatment of the bacteria. Uptake assays with 1-N-phenylnaphthylamine were used to determine that dual treatment with fosmidomycin and colistin increases membrane permeability, while binding assays with boron-dipyrromethene-conjugated polymyxin B illustrated that the addition of fosmidomycin had no impact on polymyxin binding. This work indicates that pharmacological suppression of membrane hopanoids with fosmidomycin treatment can increase the susceptibility of certain clinical B. multivorans isolates to colistin, an agent currently in use to treat pulmonary infections in CF patients. PMID:24957830

Malott, Rebecca J; Wu, Chia-Hung; Lee, Tracy D; Hird, Trevor J; Dalleska, Nathan F; Zlosnik, James E A; Newman, Dianne K; Speert, David P

2014-09-01

361

Family shuffling of soil DNA to change the regiospecificity of Burkholderia xenovorans LB400 biphenyl dioxygenase.  

PubMed

Previous work has shown that the C-terminal portion of BphA, especially two amino acid segments designated region III and region IV, influence the regiospecificity of the biphenyl dioxygenase (BPDO) toward 2,2'-dichlorobiphenyl (2,2'-CB). In this work, we evolved BPDO by shuffling bphA genes amplified from polychlorinated biphenyl-contaminated soil DNA. Sets of approximately 1-kb DNA fragments were amplified with degenerate primers designed to amplify the C-terminal portion of bphA. These fragments were shuffled, and the resulting library was used to replace the corresponding fragment of Burkholderia xenovorans LB400 bphA. Variants were screened for their ability to oxygenate 2,2'-CB onto carbons 5 and 6, which are positions that LB400 BPDO is unable to attack. Variants S100, S149, and S151 were obtained and exhibited this feature. Variant S100 BPDO produced exclusively cis-5,6-dihydro-5,6-dihydroxy-2,2'-dichlorobiphenyl from 2,2'-CB. Moreover, unlike LB400 BPDO, S100 BphA catalyzed the oxygenation of 2,2',3,3'-tetrachlorobiphenyl onto carbons 5 and 6 exclusively and it was unable to oxygenate 2,2',5,5'-tetrachlorobiphenyl. Based on oxygen consumption measurements, variant S100 oxygenated 2,2'-CB at a rate of 16 +/- 1 nmol min(-1) per nmol enzyme, which was similar to the value observed for LB400 BPDO. cis-5,6-Dihydro-5,6-dihydroxy-2,2'-dichlorobiphenyl was further oxidized by 2,3-dihydro-2,3-dihydroxybiphenyl dehydrogenase (BphB) and 2,3-dihydroxybiphenyl dioxygenase (BphC). Variant S100 was, in addition, able to oxygenate benzene, toluene, and ethyl benzene. Sequence analysis identified amino acid residues M237 S238 and S283 outside regions III and IV that influence the activity toward doubly ortho-substituted chlorobiphenyls. PMID:17142386

Vézina, Julie; Barriault, Diane; Sylvestre, Michel

2007-02-01

362

Genotyping of Burkholderia mallei from an Outbreak of Glanders in Bahrain Suggests Multiple Introduction Events  

PubMed Central

Background 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. Methodology/Principal Findings 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. Conclusion/Significance 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. PMID:25255232

Hornstra, Heidie; Projahn, Michaela; Terzioglu, Rahime; Wernery, Renate; Georgi, Enrico; Riehm, Julia M.; Wagner, David M.; Keim, Paul S.; Joseph, Marina; Johnson, Bobby; Kinne, Joerg; Jose, Shanti; Hepp, Crystal M.; Witte, Angela; Wernery, Ulrich

2014-01-01

363

Key Role for Efflux in the Preservative Susceptibility and Adaptive Resistance of Burkholderia cepacia Complex Bacteria  

PubMed Central

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, 383T (LMG 22485T), 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-?-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. PMID:23587949

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

2013-01-01

364

Homology modeling and protein engineering of alkane monooxygenase in Burkholderia thailandensis MSMB121: in silico insights.  

PubMed

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. PMID:24990796

Jain, Chakresh Kumar; Gupta, Money; Prasad, Yamuna; Wadhwa, Gulshan; Sharma, Sanjeev Kumar

2014-07-01

365

Involvement of hexokinase1 in plant growth promotion as mediated by Burkholderia phytofirmans.  

PubMed

Potato plantlets inoculated with strain PsJN of the bacterium Burkholderia phytofirmans exhibit consistent and significant increases in plant growth under in vitro conditions, when compared with uninoculated plants. The greatest influence on the degree and type of growth enhancement that develops has been shown to be mediated by the sugar concentration in the agar media. Bacterial growth promotion has been suggested in other studies to be regulated by the sugar sensor enzyme hexokinase1, the role of which is activation of glucose phosphorylation. In this present study, we examined the co-relationship between root and stem development in potato plants treated with PsJN and the activity of hexokinase1. Plants grown in the presence of 1.5% and 3% sucrose showed increased levels of hexokinase1 activity only in the roots of inoculated plants, suggesting that the increased enzyme levels may be associated with root growth. Analysis for mRNA using reverse transcriptase did not reveal any significant differences in transcription levels of the gene between inoculated and uninoculated plants. When PsJN-inoculated plants were grown in 1.5% and 3% concentrations of glucose and fructose, stem height and mass, leaf number, root mass, and overall biomass increased. No growth promotion occurred when PsJN-inoculated plants were grown in 3% maltose. Subsequently, a hexokinase1 activity assay showed that PsJN-induced growth of potato plants was found to only occur when plants were grown in the presence of sugars that are recognized by the plant hexokinase1. The results suggest that PsJN may enhance sugar uptake in plants by direct or indirect stimulation of hexokinase1 activity in roots and this results in enhanced overall plant growth. PMID:24849083

Park, Jae Min; Lazarovits, George

2014-06-01

366

Multiple pathways for toluene degradation in Burkholderia sp. strain JS150.  

PubMed Central

Burkholderia (Pseudomonas) sp. strain JS150 uses multiple pathways for the metabolism of catechols that result from degradation of aromatic compounds. This suggests that the strain also uses multiple upstream pathways for the initial hydroxylation of aromatic substrates. Two distinct DNA fragments that allowed Pseudomonas aeruginosa PAO1c to grow with benzene as a sole carbon source were cloned from strain JS150. One of the recombinant plasmids containing the initial steps for the degradative pathway contained a 14-kb DNA insert and was designated pRO2016. We have previously shown that the DNA insert originated from a plasmid carried by strain JS150 and contained genes encoding a multicomponent toluene-2-monooxygenase (tbmABCDEF) as well as the cognate regulatory protein (tbmR) that controls expression of the 2-monooxygenase (G. R. Johnson and R. H. Olsen, Appl. Environ. Microbiol. 61:3336-3346, 1995). Subsequently, we have identified an additional region on this DNA fragment that encodes toluene-4-monooxygenase activity. The toluene-4-monooxygenase activity was also regulated by the tbmR gene product. A second DNA fragment that allowed P. aeruginosa to grow with benzene was obtained as a 20-kb insert on a recombinant plasmid designated pRO2015. The DNA insert contained genes encoding toluene-4-monooxygenase activity but no toluene-2-monooxygenase activity. The pRO2015 insert originated from the chromosome of strain JS150, unlike the region cloned in pRO2016. Southern blots and restriction map comparisons showed that the genes for the individual 4-monooxygenases were distinct from one another. Thus, strain JS150 has been shown to have at least three toluene/benzene monooxygenases to initiate toluene metabolism in addition to the toluene dioxygenase reported previously by others. PMID:9327568

Johnson, G R; Olsen, R H

1997-01-01

367

IFN-? Stimulates Autophagy-Mediated Clearance of Burkholderia cenocepacia in Human Cystic Fibrosis Macrophages  

PubMed Central

Burkholderia cenocepacia is a virulent pathogen that causes significant morbidity and mortality in patients with cystic fibrosis (CF), survives intracellularly in macrophages, and uniquely causes systemic infections in CF. Autophagy is a physiologic process that involves engulfing non-functional organelles and proteins and delivering them for lysosomal degradation, but also plays a role in eliminating intracellular pathogens, including B. cenocepacia. Autophagy is defective in CF but can be stimulated in murine CF models leading to increased clearance of B. cenocepacia, but little is known about autophagy stimulation in human CF macrophages. IFN-? activates macrophages and increases antigen presentation while also inducing autophagy in macrophages. We therefore, hypothesized that treatment with IFN-? would increase autophagy and macrophage activation in patients with CF. Peripheral blood monocyte derived macrophages (MDMs) were obtained from CF and non-CF donors and subsequently infected with B. cenocepacia. Basal serum levels of IFN-? were similar between CF and non-CF patients, however after B. cenocepacia infection there is deficient IFN-? production in CF MDMs. IFN-? treated CF MDMs demonstrate increased co-localization with the autophagy molecule p62, increased autophagosome formation, and increased trafficking to lysosomes compared to untreated CF MDMs. Electron microscopy confirmed IFN-? promotes double membrane vacuole formation around bacteria in CF MDMs, while only single membrane vacuoles form in untreated CF cells. Bacterial burden is significantly reduced in autophagy stimulated CF MDMs, comparable to non-CF levels. IL-1? production is decreased in CF MDMs after IFN-? treatment. Together, these results demonstrate that IFN-? promotes autophagy-mediated clearance of B. cenocepacia in human CF macrophages. PMID:24798083

Assani, Kaivon; Tazi, Mia F.; Amer, Amal O.; Kopp, Benjamin T.

2014-01-01

368

Burkholderia BcpA mediates biofilm formation independently of interbacterial contact dependent growth inhibition  

PubMed Central

SUMMARY Contact dependent growth inhibition (CDI) is a phenomenon in which Gram-negative bacteria use the toxic C-terminus of a large surface-exposed exoprotein to inhibit the growth of susceptible bacteria upon cell-cell contact. Little is known about when and where bacteria express the genes encoding CDI system proteins and how these systems contribute to the survival of bacteria in their natural niche. Here we establish that, in addition to mediating interbacterial competition, the Burkholderia thailandensis CDI system exoprotein BcpA is required for biofilm development. We also provide evidence that the catalytic activity of BcpA and extracellular DNA are required for the characteristic biofilm pillars to form. We show using a bcpA-gfp fusion that within the biofilm, expression of the CDI system-encoding genes is below the limit of detection for the majority of bacteria and only a subset of cells express the genes strongly at any given time. Analysis of a strain constitutively expressing the genes indicates that native expression is critical for biofilm architecture. Although CDI systems have so far only been demonstrated to be involved in interbacterial competition, constitutive production of the system’s immunity protein in the entire bacterial population did not alter biofilm formation, indicating a CDI-independent role for BcpA in this process. We propose, therefore, that bacteria may use CDI proteins in cooperative behaviors, like building biofilm communities, and in competitive behaviors that prevent non-self bacteria from entering the community. PMID:23879629

Garcia, Erin C.; Anderson, Melissa S.; Hagar, Jon A.; Cotter, Peggy A.

2013-01-01

369

Identification, molecular characterisation and antimicrobial susceptibility of genomovars of the Burkholderia cepacia complex in Spain.  

PubMed

Burkholderia spp. strains collected in Spain over a 13-year period from patients with cystic fibrosis (CF) (n = 148), non-CF patients (n = 103) and from environmental sources (n = 64) were characterised. One hundred and forty-one of the examined strains were involved in seven suspected nosocomial disease outbreaks. Strains were identified by their 16s rRNA and recA genes. Their genetic relatedness, the possession of cable pili and the B. cepacia epidemic strain marker (BCESM), and their susceptibility to antimicrobial agents were studied using pulsed-field gel electrophoresis (PFGE), cblA and esmR genes analysis, and by the E-test, respectively. The genomovar distribution for the 315 strains was as follows: B. stabilis 29.5 %, B. cepacia 14.9 %, B. multivorans 11.1 %, B. cenocepacia IIIA 9.5 %, B. vietnamiensis 3.8 %, B. cenocepacia IIIB 3.5 %, and B. ambifaria and B. pyrrocinia 0.3 % each. The genetic diversity of the B. cepacia complex (Bcc) was ample, with 57 different SpeI types, showing a genetic similarity of 36.4-96.6 %. No strain carried cblA, whereas 25 B. cenocepacia genotypes harboured BCESM (23 from patients with CF). Antimicrobial resistance rates to tobramycin (TOB; 86 %) and imipenem (IPM; 67 %) were high. The strains from patients with CF showed significantly greater resistance to piperacillin (PIP), levofloxacin (LVX) and co-trimoxazole (SXT) than those isolated from non-CF patients (p < 0.05). In conclusion, B. cenocepacia was the most prevalent genomovar found in patients with CF (19.1 %), whereas B. cepacia was the most common among non-CF patients (20.7 %). B. stabilis (47.6 %) was the most common environmental genomovar. Susceptibility to antimicrobial agents depended on genomovar status and strain origin. PMID:22855365

Medina-Pascual, M J; Valdezate, S; Villalón, P; Garrido, N; Rubio, V; Saéz-Nieto, J A

2012-12-01

370

The environmental risk factors associated with medical and dental equipment in the transmission of Burkholderia (Pseudomonas) cepacia in cystic fibrosis patients  

Microsoft Academic Search

Transmission of Burkholderia (Pseudomonas) cepacia by close contact with colonized patients is well documented, and minimizing social contact between cystic fibrosis (CF) patients by segregation and cohorting of B. cepacia colonized patients has achieved some success in controlling the nosocomial and community spread of this organism. However, direct and indirect environmental transmission still occurs. There is evidence for transmission of

C. L. Pankhurst; J. Philpott-Howard

1996-01-01

371

Life-Threatening Sepsis Caused by Burkholderia cepacia From Contaminated Intravenous Flush Solutions Prepared by a Compounding Pharmacy in Another State  

Microsoft Academic Search

We report 2 life-threatening cases of Burkholderia cepacia sepsis caused by infusate contamination during compound- ing. Bacterial isolates from the patients' blood cultures and the infusate were indistinguishable by pulsed-field gel electrophoresis. Proper quality controls at a local and national level are important for ensuring safe delivery of compounded medications to patients in all settings, including those outside health care

Melissa R. Held; Elizabeth M. Begier; Diana S. Beardsley; Frederick A. Browne; Richard A. Martinello; Robert S. Baltimore; L. Clifford McDonald; Bette Jensen; James L. Hadler; Louise-Marie Dembry

2010-01-01

372

Draft Genome Sequences of Two Antimicrobial-Producing Burkholderia sp. Strains, MSh1 and MSh2, Isolated from Malaysian Tropical Peat Swamp Forest Soil  

PubMed Central

We report the draft genome sequences of two antimicrobial-producing isolates, Burkholderia sp. strains MSh1 and MSh2, which were isolated from tropical peat swamp forest soil. Putative genes related to different antimicrobial production have been annotated in both genome sequences. PMID:25301661

Aw, Yoong Kit; Gan, Han Ming; Yule, Catherine M.; Lee, Sui Mae

2014-01-01

373

Arabidopsis thaliana and Pisum sativum models demonstrate that root colonization is an intrinsic trait of Burkholderia cepacia complex bacteria.  

PubMed

Burkholderia cepacia complex (Bcc) bacteria possess biotechnologically useful properties that contrast with their opportunistic pathogenicity. The rhizosphere fitness of Bcc bacteria is central to their biocontrol and bioremediation activities. However, it is not known whether this differs between species or between environmental and clinical strains. We investigated the ability of 26 Bcc strains representing nine different species to colonize the roots of Arabidopsis thaliana and Pisum sativum (pea). Viable counts, scanning electron microscopy and bioluminescence imaging were used to assess root colonization, with Bcc bacteria achieving mean (±sem) levels of 2.49±0.23×10(6) and 5.16±1.87×10(6) c.f.u. per centimetre of root on the A. thaliana and P. sativum models, respectively. The A. thaliana rhizocompetence model was able to reveal loss of colonization phenotypes in Burkholderia vietnamiensis G4 transposon mutants that had only previously been observed in competition experiments on the P. sativum model. Different Bcc species colonized each plant model at different rates, and no statistical difference in root colonization was observed between isolates of clinical or environmental origin. Loss of the virulence-associated third chromosomal replicon (>1 Mb DNA) did not alter Bcc root colonization on A. thaliana. In summary, Bcc bacteria possess intrinsic root colonization abilities irrespective of their species or source. As Bcc rhizocompetence does not require their third chromosomal replicon, the possibility of using synthetic biology approaches to engineer virulence-attenuated biotechnological strains is tractable. PMID:24327425

Vidal-Quist, J Cristian; O'Sullivan, Louise A; Desert, Annaëlle; Fivian-Hughes, Amanda S; Millet, Coralie; Jones, T Hefin; Weightman, Andrew J; Rogers, Hilary J; Berry, Colin; Mahenthiralingam, Eshwar

2014-02-01

374

[Bacteria of the genus Burkholderia as a typical component of the microbial community of sphagnum peat bogs].  

PubMed

Bacteria of the genus Burkholderia are a typical component of the microbial complex of sphagnum peat bogs and constitute a substantial portion of the aerobic chemoorganotrophic isolates which are routinely obtained from these environments on acidic nutrient media. The ecophysiological characteristics of the 27 strains of such organisms, which were isolated from the peat of acidic sphagnum bogs of the boreal and tundra zones of Russia, Canada, and Estonia, were investigated in the present paper. The overwhelming majority of the Burkholderia strains isolated from these bogs were phylogenetically close to the species B. glathei, B. phenazinium, B. fungorum, and B. caryophylli, the typical inhabitants of soil and plant rhizosphere. The bog isolates utilized a broad range of substrates as carbon and energy sources, including organic acids, sugars, polyalcohols, and certain aromatic compounds. All the strains studied were capable of growth on nitrogen-free media. They developed in the pH ranges of 3.5 to 7.4 and from 3 to 37 degrees C, with the optima at pH 5-7 and 11-23 degrees C, respectively. They were therefore moderately acidophilic, psychroactive, dinitrogen-fixing microorganisms well adapted to the conditions of acidic northern sphagnum bogs. PMID:16579452

Belova, S E; Pankratov, T A; Dedysh, S N

2006-01-01

375

[Evaluation of commercial systems VITEK 2 and API 20NE for identification of Burkholderia cepacia complex bacteria from clinical samples].  

PubMed

Species belonging to the Burkholderia cepacia complex (BCC) are capable of causing chronic respiratory tract infections in patients suffering from cystic fibrosis as well as in immunocompromised individuals. Most of these species are highly resistant to antibiotic therapy, generating the need for their rapid and accurate detection for the proper treatment and clinical management of these patients. In this work, the polymerase chain reaction (PCR) technique based on the amplification of the recA gene (PCR-recA) was applied for an accurate identification of bacteria belonging to the BCC. Sensitivity (S) and specificity (E) of two biochemically-based commercial automated systems, API 20NE and VITEK 2 (bioMérieux®), and of the most representative biochemical manual tests for the identification of the Burkholderia cepacia complex were herein evaluated. The commercial systems VITEK 2 and API 20NE showed the following sensitivity and specificity vaues for identification to the species level, S: 71.1 %, E: 100 %, S: 69.7 %, E: 90.2 %, respectively. More complex results were observed for phenotypic manual tests, since BCC bacteria can undergo selective pressure to survive in chronic patients causing the loss of their typical phenotypic characteristics. The PCR-recA technique was easy to implement even in medium-complexity clinical diagnostic laboratories. PMID:22430988

Oderiz, Sebastián; Palau, María J; Del Palacio, Paula; Lewis, Miriam C; Bettiol, Marisa P; Martina, Pablo; Bosch, Alejandra; Yantorno, Osvaldo M; Gatti, Blanca M

2011-01-01

376

Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2232 from Karijini National Park, Australia  

PubMed Central

Burkholderia sp. strain WSM2232 is an aerobic, motile, Gram-negative, non-spore-forming acid-tolerant rod that was trapped in 2001 from acidic soil collected from Karijini National Park (Australia) using Gastrolobium capitatum as a host. WSM2232 was effective in nitrogen fixation with G. capitatum but subsequently lost symbiotic competence during long-term storage. Here we describe the features of Burkholderia sp. strain WSM2232, together with genome sequence information and its annotation. The 7,208,311 bp standard-draft genome is arranged into 72 scaffolds of 72 contigs containing 6,322 protein-coding genes and 61 RNA-only encoding genes. The loss of symbiotic capability can now be attributed to the loss of nodulation and nitrogen fixation genes from the genome. This rhizobial genome is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project. PMID:25197442

Walker, Robert; Watkin, Elizabeth; Tian, Rui; Brau, Lambert; O'Hara, Graham; Goodwin, Lynne; Han, James; Reddy, Tatiparthi; Huntemann, Marcel; Pati, Amrita; Woyke, Tanja; Mavromatis, Konstantinos; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; Reeve, Wayne

2013-01-01

377

Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2230 from Karijini National Park, Australia  

PubMed Central

Burkholderia sp. strain WSM2230 is an aerobic, motile, Gram-negative, non-spore-forming acid-tolerant rod isolated from acidic soil collected in 2001 from Karijini National Park, Western Australia, using Kennedia coccinea (Coral Vine) as a host. WSM2230 was initially effective in nitrogen-fixation with K. coccinea, but subsequently lost symbiotic competence. Here we describe the features of Burkholderia sp. strain WSM2230, together with genome sequence information and its annotation. The 6,309,801 bp high-quality-draft genome is arranged into 33 scaffolds of 33 contigs containing 5,590 protein-coding genes and 63 RNA-only encoding genes. The genome sequence of WSM2230 failed to identify nodulation genes and provides an explanation for the observed failure of the laboratory grown strain to nodulate. The genome of this strain is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.

Walker, Robert; Watkin, Elizabeth; Tian, Rui; Brau, Lambert; O'Hara, Graham; Goodwin, Lynne; Han, James; Lobos, Elizabeth; Huntemann, Marcel; Pati, Amrita; Woyke, Tanja; Mavromatis, Konstantinos; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; Reeve, Wayne

2013-01-01

378

Biosynthesis and mobilization of a novel polyhydroxyalkanoate containing 3-hydroxy-4-methylvalerate monomer produced by Burkholderia sp. USM (JCM15050).  

PubMed

We attempted to synthesize a polyhydroxyalkanoate (PHA) containing newly reported 3-hydroxy-4-methylvalerate (3H4MV) monomer by using wild type Burkholderia sp. USM (JCM15050) and its transformed strain harboring the PHA synthase gene of Aeromonas caviae (phaCAc). The introduction of 3H4MV as a second monomer will improve the material properties of 3HB-based polymers. To promote the accumulation of PHA containing 3H4MV monomer, isocaproic acid was provided as co-carbon source. Approximately 1mol% of 3H4MV was detected in wild type Burkholderia sp. cultures when they were fed glucose or fructose together with isocaproic acid. Thus, the wild type strain can synthesize the 3H4MV monomer. High 3H4MV fractions, of about 40mol%, were obtained when the transformed strain was cultivated on glucose or fructose together with isocaproic acid. In addition, the ability of the transformed strain to mobilize accumulated PHA containing 3H4MV monomer was demonstrated in this study. This is the first report on mobilization of the 3H4MV monomer. PMID:20541932

Lau, Nyok-Sean; Chee, Jiun-Yee; Tsuge, Takeharu; Sudesh, Kumar

2010-10-01

379

Mechanism and Inhibition of the FabV Enoyl-ACP Reductase from Burkholderia Mallei†  

PubMed Central

Enoyl-ACP reductases catalyze the final step in the elongation cycle of the bacterial fatty acid biosynthesis (FAS-II) pathway. Currently four distinct enoyl-ACP reductases have been identified, which are the products of the fabI, fabL, fabK and fabV genes. The FabV enoyl-ACP reductase is the most recent member of this enzyme class and was originally identified in Vibrio cholerae by Cronan and coworkers [Massengo-Tiasse and Cronan (2008) Vibrio cholerae FabV defines a new class of enoyl-acyl carrier protein reductase, J. Biol. Chem. 283, 1308–1316]. In the present work a detailed kinetic analysis of the mechanism of the FabV enzyme from Burkholderia mallei (bmFabV) has been undertaken, which reveals that bmFabV catalyzes a sequential Bi Bi mechanism with NADH binding first and NAD+ dissociating last. The enzyme is a member of the short chain dehydrogenase/reductase superfamily in which the catalytic tyrosine (Y235) and lysine (K244) residues are organized in the consensus Tyr-(Xaa)8-Lys motif. The role of these active-site residues has been investigated using site-directed mutagenesis which has shown that both Y235 and K244 are involved in acid/base chemistry during substrate reduction. Sequence alignment and site-directed mutagenesis also identify a second lysine in the active site (K245) that has an important role in binding of the enoyl substrate. Due to interests in developing inhibitors of bmFabV, a detailed analysis of the inhibition of the enzyme by triclosan has been conducted showing that triclosan is a competitive inhibitor with respect to NADH and an uncompetitive inhibitor with respect to the substrate 2-dodecenoyl-CoA (Ki = 0.4 µM). Combined with fluorescence binding experiments, it is concluded that triclosan binds to the enzyme-NAD+ product complex which is in rapid and reversible equilibrium with other intermediates on the reaction pathway. PMID:20055482

Lu, Hao; Tonge, Peter J.

2010-01-01

380

Potential mechanisms underlying the acute lung dysfunction and bacterial extrapulmonary dissemination during Burkholderia cenocepacia respiratory infection  

PubMed Central

Background Burkholderia cenocepacia, an opportunistic pathogen that causes lung infections in cystic fibrosis (CF) patients, is associated with rapid and usually fatal lung deterioration due to necrotizing pneumonia and sepsis, a condition known as cepacia syndrome. The key bacterial determinants associated with this poor clinical outcome in CF patients are not clear. In this study, the cytotoxicity and procoagulant activity of B. cenocepacia from the ET-12 lineage, that has been linked to the cepacia syndrome, and four clinical isolates recovered from CF patients with mild clinical courses were analysed in both in vitro and in vivo assays. Methods B. cenocepacia-infected BEAS-2B epithelial respiratory cells were used to investigate the bacterial cytotoxicity assessed by the flow cytometric detection of cell staining with propidium iodide. Bacteria-induced procoagulant activity in cell cultures was assessed by a colorimetric assay and by the flow cytometric detection of tissue factor (TF)-bearing microparticles in cell culture supernatants. Bronchoalveolar lavage fluids (BALF) from intratracheally infected mice were assessed for bacterial proinflammatory and procoagulant activities as well as for bacterial cytotoxicity, by the detection of released lactate dehydrogenase. Results ET-12 was significantly more cytotoxic to cell cultures but clinical isolates Cl-2, Cl-3 and Cl-4 exhibited also a cytotoxic profile. ET-12 and CI-2 were similarly able to generate a TF-dependent procoagulant environment in cell culture supernatant and to enhance the release of TF-bearing microparticles from infected cells. In the in vivo assay, all bacterial isolates disseminated from the mice lungs, but Cl-2 and Cl-4 exhibited the highest rates of recovery from mice livers. Interestingly, Cl-2 and Cl-4, together with ET-12, exhibited the highest cytotoxicity. All bacteria were similarly capable of generating a procoagulant and inflammatory environment in animal lungs. Conclusion B. cenocepacia were shown to exhibit cytotoxic and procoagulant activities potentially implicated in bacterial dissemination into the circulation and acute pulmonary decline detected in susceptible CF patients. Improved understanding of the mechanisms accounting for B. cenocepacia-induced clinical decline has the potential to indicate novel therapeutic strategies to be included in the care B. cenocepacia-infected patients. PMID:20082687

2010-01-01

381

Advanced amperometric respiration assay for antimicrobial susceptibility testing.  

PubMed

A ferricyanide-based electrochemical cell respiration assay was adapted for use in broad-spectrum antimicrobial susceptibility testing (AST). Total bacterial respiration was converted into faradaic current by electro-oxidation of ferrocyanide, produced when ferricyanide is reduced by bacterial electron-transport. For Escherichia coli (E. coli), the signal was linear with 5-13 × 10(8) colony-forming units in measuring buffer. For AST, test cells were treated with drugs before ferricyanide addition; cell counts from the amperometric assay provided a measure of drug-induced cell death. Initial trials with six antimicrobial agents produced incorrect susceptibility classifications for drugs that were electroactive at the potential used to detect ferrocyanide or which affected cellular respiration rates. We therefore changed the procedure from drug-treatment and assay in the same buffer to sequential drug exposure in treatment buffer, centrifugal separation of surviving cells, cell resuspension, incubation in the presence of ferricyanide and finally ferrocyanide amperometry in drug-free buffer. Data analysis with E. coli led to an activity classification that agreed with cell culture-based ASTs, obtained by a quicker, more convenient procedure. The potential of this approach was confirmed by trials with the highly virulent bacterium Burkholderia pseudomallei, a particularly antimicrobial-resistant pathogen that is the cause of lethal melioidosis in tropical climates and is currently of concern as a potential bioterrorism agent. PMID:25222107

Chotinantakul, Kamonnaree; Suginta, Wipa; Schulte, Albert

2014-10-21

382

Burkholderia terrae BS001 migrates proficiently with diverse fungal hosts through soil and provides protection from antifungal agents  

PubMed Central

Soil bacteria can benefit from co-occurring soil fungi in respect of the acquisition of carbonaceous nutrients released by fungal hyphae and the access to novel territories in soil. Here, we investigated the capacity of the mycosphere-isolated bacterium Burkholderia terrae BS001 to comigrate through soil along with hyphae of the soil fungi Trichoderma asperellum, Rhizoctonia solani, Fusarium oxysporum, F. oxysporum pv lini, Coniochaeta ligniaria, Phanerochaete velutina, and Phallus impudicus. We used Lyophyllum sp. strain Karsten as the reference migration-inciting fungus. Bacterial migration through presterilized soil on the extending fungal hyphae was detected with six of the seven test fungi, with only Phallus impudicus not showing any bacterial transport. Much like with Lyophyllum sp. strain Karsten, intermediate (106–108 CFU g-1 dry soil) to high (>108 CFU g-1 dry soil) strain BS001 cell population sizes were found at the hyphal migration fronts of four fungi, i.e., T. asperellum, Rhizoctonia solani, F. oxysporum and F. oxysporum pv lini, whereas for two fungi, Coniochaeta ligniaria and Phanerochaete velutina, the migration responses were retarded and population sizes were lower (103–106 CFU g-1 dry soil). Consistent with previous data obtained with the reference fungus, migration with the migration-inciting fungi occurred only in the direction of the hyphal growth front. Remarkably, Burkholderia terrae BS001 provided protection from several antifungal agents to the canonical host Lyophyllum sp. strain Karsten. Specifically, this host was protected from Pseudomonas fluorescens strain CHA0 metabolites, as well as from the anti-fungal agent cycloheximide. Similar protection by strain BS001was observed for T. asperellum, and, to a lower extent, F. oxysporum and Rhizoctonia solani. The protective effect may be related to the consistent occurrence of biofilm-like cell layers or agglomerates at the surfaces of the protected fungi. The current study represents the first report of protection of soil fungi against antagonistic agents present in the soil provided by fungal-associated Burkholderia terrae cells.

Nazir, Rashid; Tazetdinova, Diana I.; van Elsas, Jan Dirk

2014-01-01

383

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

PubMed Central

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. PMID:25288974

Jung, Boknam; Lee, Sehee; Ha, Jiran; Park, Jong-Chul; Han, Sung-Sook; Hwang, Ingyu; Lee, Yin-Won; Lee, Jungkwan

2013-01-01

384

Heterologous Production of Glidobactins/Luminmycins in Escherichia coli Nissle Containing the Glidobactin Biosynthetic Gene Cluster from Burkholderia DSM7029.  

PubMed

Natural product peptide-based proteasome inhibitors show great potential as anticancer drugs. Here we have cloned the biosynthetic gene cluster of a potent proteasome inhibitor-glidobactin from Burkholderia DSM7029-and successfully detected glidobactins/luminmycins in E. coli Nissle. We have also improved the yield of glidobactin A tenfold by promoter change in a heterologous host. In addition, two new biosynthetic intermediates were identified by comparative MS/MS fragmentation analysis. Identification of acyclic luminmycin E implies substrate specificity of the TE domain for cyclization. The establishment of a heterologous expression system for syrbactins provided the basis for the generation of new syrbactins as proteasome inhibitors by molecular engineering, but the TE domain's specificity cannot be ignored. PMID:25147087

Bian, Xiaoying; Huang, Fan; Wang, Hailong; Klefisch, Thorsten; Müller, Rolf; Zhang, Youming

2014-10-13