Burkholderia cocovenenans (van Damme et al. 1960) Gillis et al. 1995 and Burkholderia vandii Urakami et al. 1994 are junior synonyms of Burkholderia gladioli (Severini 1913) Yabuuchi et al. 1993 and Burkholderia plantarii (Azegami et al. 1987) Urakami et al. 1994, respectively.
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Reference strains of Burkholderia cocovenenans and Burkholderia vandii were compared with strains of other Burkholderia species using SDS-PAGE of whole-cell proteins, DNA-DNA hybridization and extensive biochemical characterization. Burkholderia gladioli and B. cocovenenans were indistinguishable in the chemotaxonomic and biochemical analyses. Burkholderia plantarii and B. vandii had indistinguishable whole-cell protein patterns but the B. vandii type strain differed from B. plantarii strains in several biochemical tests. The DNA-DNA binding levels (higher than 70%) indicated that (i) B. gladioli and B. cocovenenans, and (ii) B. plantarii and B. vandii each represent a single species. It is concluded that B. cocovenenans and B. vandii are junior synonyms of B. gladioli and B. plantarii, respectively. (+info)
All in the family: structural and evolutionary relationships among three modular proteins with diverse functions and variable assembly.
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The crystal structures of three proteins of diverse function and low sequence similarity were analyzed to evaluate structural and evolutionary relationships. The proteins include a bacterial bleomycin resistance protein, a bacterial extradiol dioxygenase, and human glyoxalase I. Structural comparisons, as well as phylogenetic analyses, strongly indicate that the modern family of proteins represented by these structures arose through a rich evolutionary history that includes multiple gene duplication and fusion events. These events appear to be historically shared in some cases, but parallel and historically independent in others. A significant early event is proposed to be the establishment of metal-binding in an oligomeric ancestor prior to the first gene fusion. Variations in the spatial arrangements of homologous modules are observed that are consistent with the structural principles of three-dimensional domain swapping, but in the unusual context of the formation of larger monomers from smaller dimers or tetramers. The comparisons support a general mechanism for metalloprotein evolution that exploits the symmetry of a homooligomeric protein to originate a metal binding site and relies upon the relaxation of symmetry, as enabled by gene duplication, to establish and refine specific functions. (+info)
Discrimination of Burkholderia multivorans and Burkholderia vietnamiensis from Burkholderia cepacia genomovars I, III, and IV by PCR.
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We present a PCR procedure for identification of Burkholderia cepacia, Burkholderia multivorans, and Burkholderia vietnamiensis. 16S and 23S ribosomal DNAs (rDNAs) of B. multivorans and B. vietnamiensis were sequenced and aligned with published sequences for definition of species-specific 18-mer oligonucleotide primers. Specific antisense 16S rDNA primers (for B. cepacia, 5'-AGC ACT CCC RCC TCT CAG-3'; for B. multivorans, 5'-AGC ACT CCC GAA TCT CTT-3') and 23S rDNA primers (for B. vietnamiensis, 5'-TCC TAC CAT GCG TGC AA-3') were paired with a general sense primer of 16S rDNAs (5'-AGR GTT YGA TYM TGG CTC AG-3') or with a sense primer of 23S rDNA (5'-CCT TTG GGT CAT CCT GGA-3'). PCR with these primers under optimized conditions is appropriate to specifically and rapidly identify B. multivorans, B. vietnamiensis, and B. cepacia (genomovars I, III, and IV are not discriminated). In comparison with the polyphasic taxonomic analyses presently necessary for species and genomovar identification within the B. cepacia complex, our procedure is more rapid and easier to perform and may contribute to clarifying the clinical significance of individual members of the complex in cystic fibrosis. (+info)
Expression and functional analysis of an N-truncated NifA protein of Herbaspirillum seropedicae.
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In Herbaspirillum seropedicae, an endophytic diazotroph, nif gene expression is under the control of the transcriptional activator NifA. We have over-expressed and purified a protein containing the central and C-terminal domains of the H. seropedicae NifA protein, N-truncated NifA, fused to a His-Tag sequence. This fusion protein was found to be partially soluble and was purified by affinity chromatography. Band shift and footprinting assays showed that the N-truncated NifA protein was able to bind specifically to the H. seropedicae nifB promoter region. In vivo analysis showed that this protein activated the nifH promoter of Klebsiella pneumoniae in Escherichia coli only in the absence of oxygen and this activation was not negatively controlled by ammonium ions. (+info)
Burkholderia caribensis sp. nov., an exopolysaccharide-producing bacterium isolated from vertisol microaggregates in Martinique.
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Twenty-one exopolysaccharide-producing strains were isolated from the 5-20 microns fraction of a vertisol in the south-east of the island of Martinique in the French West Indies. Although these strains were phenotypically identified as Burkholderia cepacia or as Burkholderia glathei using BIOLOG microplates, they did not cluster genotypically by amplified rDNA restriction analysis (ARDRA) with any described Burkholderia species. A phylogenetic analysis revealed that the rrs (16S rDNA) sequences of three representative strains clustered in a single branch within the genus Burkholderia and distantly from all of the previously described species of Burkholderia for which rrs sequences were available. DNA-DNA hybridization data as well as phenotypic analyses indicated that the 21 isolates represented a single and new species for which the name Burkholderia caribensis sp. nov. is proposed (type strain MWAP64T = LMG 18531T). (+info)
Degradation of chlorobenzenes at nanomolar concentrations by Burkholderia sp. strain PS14 in liquid cultures and in soil.
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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. (+info)
Identification of Burkholderia spp. in the clinical microbiology laboratory: comparison of conventional and molecular methods.
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Cystic fibrosis (CF) predisposes patients to bacterial colonization and infection of the lower airways. Several species belonging to the genus Burkholderia are potential CF-related pathogens, but microbiological identification may be complicated. This situation is not in the least due to the poorly defined taxonomic status of these bacteria, and further validation of the available diagnostic assays is required. A total of 114 geographically diverse bacterial isolates, previously identified in reference laboratories as Burkholderia cepacia (n = 51), B. gladioli (n = 14), Ralstonia pickettii (n = 6), B. multivorans (n = 2), Stenotrophomonas maltophilia (n = 3), and Pseudomonas aeruginosa (n = 11), were collected from environmental, clinical, and reference sources. In addition, 27 clinical isolates putatively identified as Burkholderia spp. were recovered from the sputum of Dutch CF patients. All isolates were used to evaluate the accuracy of two selective growth media, four systems for biochemical identification (API 20NE, Vitek GNI, Vitek NFC, and MicroScan), and three different PCR-based assays. The PCR assays amplify different parts of the ribosomal DNA operon, either alone or in combination with cleavage by various restriction enzymes (PCR-restriction fragment length polymorphism [RFLP] analysis). The best system for the biochemical identification of B. cepacia appeared to be the API 20NE test. None of the biochemical assays successfully grouped the B. gladioli strains. The PCR-RFLP method appeared to be the optimal method for accurate nucleic acid-mediated identification of the different Burkholderia spp. With this method, B. gladioli was also reliably classified in a separate group. For the laboratory diagnosis of B. cepacia, we recommend parallel cultures on blood agar medium and selective agar plates. Further identification of colonies with a Burkholderia phenotype should be performed with the API 20NE test. For final confirmation of species identities, PCR amplification of the small-subunit rRNA gene followed by RFLP analysis with various enzymes is recommended. (+info)
Differentiation of Burkholderia species by PCR-restriction fragment length polymorphism analysis of the 16S rRNA gene and application to cystic fibrosis isolates.
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Burkholderia cepacia, which is an important pathogen in cystic fibrosis (CF) owing to the potential severity of the infections and the high transmissibility of some clones, has been recently shown to be a complex of five genomic groups, i.e., genomovars I, II (B. multivorans), III, and IV and B. vietnamiensis. B. gladioli is also involved, though rarely, in CF. Since standard laboratory procedures fail to provide an accurate identification of these organisms, we assessed the ability of restriction fragment length polymorphism (RFLP) analysis of amplified 16S ribosomal DNA (rDNA), with the combination of the patterns obtained with six endonucleases, to differentiate Burkholderia species. This method was applied to 16 type and reference strains of the genus Burkholderia and to 51 presumed B. cepacia clinical isolates, each representative of one clone previously determined by PCR ribotyping. The 12 Burkholderia type strains tested were differentiated, including B. cepacia, B. multivorans, B. vietnamiensis, and B. gladioli, but neither the genomovar I and III reference strains nor the genomovar IV reference strain and B. pyrrociniaT were distinguishable. CF clinical isolates were mainly distributed in RFLP group 2 (which includes B. multivoransT) and RFLP group 1 (which includes B. cepacia genomovar I and III reference strains, as well as nosocomial clinical isolates). Two of the five highly transmissible clones in French CF centers belonged to RFLP group 2, and three belonged to RFLP group 1. The remaining isolates either clustered with other Burkholderia species (B. cepacia genomovar IV or B. pyrrocinia, B. vietnamiensis, and B. gladioli) or harbored unique combinations of patterns. Thus, if further validated by hybridization studies, PCR-RFLP of 16S rDNA could be an interesting identification tool and contribute to a better evaluation of the respective clinical risks associated with each Burkholderia species or genomovar in patients with CF. (+info)