Transcriptional expression of the tceA gene in a Dehalococcoides-containing microbial enrichment.
Ontology highlight
ABSTRACT:
SUBMITTER: Johnson DR
PROVIDER: S-EPMC1287711 | biostudies-literature | 2005 Nov
REPOSITORIES: biostudies-literature
Publications
Transcriptional expression of the tceA gene in a Dehalococcoides-containing microbial enrichment.
Johnson David R DR Lee Patrick K H PK Holmes Victor F VF Fortin Alexander C AC Alvarez-Cohen Lisa L
Applied and environmental microbiology 20051101 11
Dynamic changes in the transcriptional expression of the tceA gene, which encodes a trichloroethene reductive dehalogenase, were characterized in a Dehalococcoides-containing microbial enrichment culture. Expression was quantified by real-time PCR as the number of tceA transcripts per tceA gene. Expression of tceA increased 40-fold after chlorinated ethene-starved cells were exposed to trichloroethene (TCE), cis-dichloroethene (DCE), or 1,1-DCE but did not increase after exposure to tetrachloroe ...[more]
Similar Datasets
Project description:Chlorinated solvents are among the most prevalent groundwater contaminants in the industrialized world. Biodegradation with Dehalococcoides-containing mixed cultures is an effective remediation technology. To elucidate transcribed genes in a Dehalococcoides-containing mixed culture, a shotgun metagenome microarray was created and used to investigate gene transcription during vinyl chloride (VC) dechlorination and during starvation (no chlorinated compounds) by a microbial enrichment culture called KB-1. In both treatment conditions, methanol was amended as an electron donor. Subsequently, spots were sequenced that contained the genes most differentially transcribed between the VC-degrading and methanol-only conditions, as well as spots with the highest intensities. Sequencing revealed that during VC degradation Dehalococcoides genes involved in transcription, translation, metabolic energy generation, and amino acid and lipid metabolism and transport were overrepresented in the transcripts compared to the average Dehalococcoides genome. KB-1 rdhA14 (vcrA) was the only reductive dehalogenase homologous (RDH) gene with higher transcript levels during VC degradation, while multiple RDH genes had higher transcript levels in the absence of VC. Numerous hypothetical genes from Dehalococcoides also had higher transcript levels in methanol-only treatments, indicating that many uncharacterized proteins are involved in cell maintenance in the absence of chlorinated substrates. In addition, microarray results prompted biological experiments confirming that electron acceptor limiting conditions activated a Dehalococcoides prophage. Transcripts from Spirochaetes, Chloroflexi, Geobacter, and methanogens demonstrate the importance of non-Dehalococcoides organisms to the culture, and sequencing of identified shotgun clones of interest provided information for follow-on targeted studies.
| S-EPMC3273020 | biostudies-literature
Project description:Tetrachloroethene (PCE) and trichloroethene (TCE) are prevalent groundwater contaminants that can be completely reductively dehalogenated by some "Dehalococcoides" organisms. A Dehalococcoides-organism-containing microbial consortium (referred to as ANAS) with the ability to degrade TCE to ethene, an innocuous end product, was previously enriched from contaminated soil. A whole-genome photolithographic microarray was developed based on the genome of "Dehalococcoides ethenogenes" 195. This microarray contains probes designed to hybridize to >99% of the predicted protein-coding sequences in the strain 195 genome. DNA from ANAS was hybridized to the microarray to characterize the genomic content of the ANAS enrichment. The microarray results revealed that the genes associated with central metabolism, including an apparently incomplete carbon fixation pathway, cobalamin-salvaging system, nitrogen fixation pathway, and five hydrogenase complexes, are present in both strain 195 and ANAS. Although the gene encoding the TCE reductase, tceA, was detected, 13 of the 19 reductive dehalogenase genes present in strain 195 were not detected in ANAS. Additionally, 88% of the genes in predicted integrated genetic elements in strain 195 were not detected in ANAS, consistent with these elements being genetically mobile. Sections of the tryptophan operon and an operon encoding an ABC transporter in strain 195 were also not detected in ANAS. These insights into the diversity of Dehalococcoides genomes will improve our understanding of the physiology and evolution of these bacteria, which is essential in developing effective strategies for the bioremediation of PCE and TCE in the environment.
| S-EPMC2423027 | biostudies-literature
Project description:In order to elucidate interactions between sulfate reduction and dechlorination, we systematically evaluated the effects of different concentrations of sulfate and sulfide on reductive dechlorination by isolates, constructed consortia, and enrichments containing Dehalococcoides sp. Sulfate (up to 5 mM) did not inhibit the growth or metabolism of pure cultures of the dechlorinator Dehalococcoides mccartyi 195, the sulfate reducer Desulfovibrio vulgaris Hildenborough, or the syntroph Syntrophomonas wolfei In contrast, sulfide at 5 mM exhibited inhibitory effects on growth of the sulfate reducer and the syntroph, as well as on both dechlorination and growth rates of D. mccartyi Transcriptomic analysis of D. mccartyi 195 revealed that genes encoding ATP synthase, biosynthesis, and Hym hydrogenase were downregulated during sulfide inhibition, whereas genes encoding metal-containing enzymes involved in energy metabolism were upregulated even though the activity of those enzymes (hydrogenases) was inhibited. When the electron acceptor (trichloroethene) was limiting and an electron donor (lactate) was provided in excess to cocultures and enrichments, high sulfate concentrations (5 mM) inhibited reductive dechlorination due to the toxicity of generated sulfide. The initial cell ratio of sulfate reducers to D. mccartyi (1:3, 1:1, or 3:1) did not affect the dechlorination performance in the presence of sulfate (2 and 5 mM). In contrast, under electron donor limitation, dechlorination was not affected by sulfate amendments due to low sulfide production, demonstrating that D. mccartyi can function effectively in anaerobic microbial communities containing moderate sulfate concentrations (5 mM), likely due to its ability to outcompete other hydrogen-consuming bacteria and archaea.IMPORTANCE Sulfate is common in subsurface environments and has been reported as a cocontaminant with chlorinated solvents at various concentrations. Inconsistent results for the effects of sulfate inhibition on the performance of dechlorination enrichment cultures have been reported in the literature. These inconsistent findings make it difficult to understand potential mechanisms of sulfate inhibition and complicate the interpretation of bioremediation field data. In order to elucidate interactions between sulfate reduction and reductive dechlorination, this study systematically evaluated the effects of different concentrations of sulfate and sulfide on reductive dechlorination by isolates, constructed consortia, and enrichments containing Dehalococcoides sp. This study provides a more fundamental understanding of the competition mechanisms between reductive dechlorination by Dehalococcoides mccartyi and sulfate reduction during the bioremediation process. It also provides insights on the significance of sulfate concentrations on reductive dechlorination under electron donor/acceptor-limiting conditions during in situ bioremediation applications. For example, at a trichloroethene-contaminated site with a high sulfate concentration, proper slow-releasing electron donors can be selected to generate an electron donor-limiting environment that favors reductive dechlorination and minimizes the sulfide inhibition effect.
| S-EPMC5377507 | biostudies-literature
Project description:UnlabelledBackgroundThe Dehalococcoides are strictly anaerobic bacteria that gain metabolic energy via the oxidation of H2 coupled to the reduction of halogenated organic compounds. Dehalococcoides spp. grow best in mixed microbial consortia, relying on non-dechlorinating members to provide essential nutrients and maintain anaerobic conditions.A metagenome sequence was generated for the dechlorinating mixed microbial consortium KB-1. A comparative metagenomic study utilizing two additional metagenome sequences for Dehalococcoides-containing dechlorinating microbial consortia was undertaken to identify common features that are provided by the non-dechlorinating community and are potentially essential to Dehalococcoides growth.ResultsThe KB-1 metagenome contained eighteen novel homologs to reductive dehalogenase genes. The metagenomes obtained from the three consortia were automatically annotated using the MG-RAST server, from which statistically significant differences in community composition and metabolic profiles were determined. Examination of specific metabolic pathways, including corrinoid synthesis, methionine synthesis, oxygen scavenging, and electron-donor metabolism identified the Firmicutes, methanogenic Archaea, and the ?-Proteobacteria as key organisms encoding these pathways, and thus potentially producing metabolites required for Dehalococcoides growth.ConclusionsComparative metagenomics of the three Dehalococcoides-containing consortia identified that similarities across the three consortia are more apparent at the functional level than at the taxonomic level, indicating the non-dechlorinating organisms' identities can vary provided they fill the same niche within a consortium. Functional redundancy was identified in each metabolic pathway of interest, with key processes encoded by multiple taxonomic groups. This redundancy likely contributes to the robust growth and dechlorination rates in dechlorinating enrichment cultures.
| S-EPMC3475024 | biostudies-literature
Project description:Tetrachloroethene (PCE) and trichloroethene (TCE) are prevalent groundwater contaminants that can be completely reductively dehalogenated by Dehalococcoides organisms. A Dehalococcoides-containing microbial consortium (ANAS) with the ability to degrade TCE to ethene, an innocuous end-product, was previously enriched from contaminated soil. A whole-genome photolithographic microarray was developed based on the genome of Dehalococcoides ethenogenes 195 (strain 195). This microarray contains probes designed to hybridize to >99% of the predicted protein-coding sequences in the strain 195 genome. DNA from ANAS was hybridized to the microarray to characterize the genomic content of the ANAS enrichment. The microarray revealed that the genes associated with central metabolism including an apparently incomplete carbon fixation pathway, cobalamin salvaging system, nitrogen fixation pathway, and five hydrogenase complexes are present in both strain 195 and ANAS. Although the gene encoding the TCE reductase tceA was detected, 13 of the 19 reductive dehalogenase genes present in strain 195 were not detected in ANAS. Additionally, 88% of the genes in predicted integrated genetic elements in strain 195 were not detected in ANAS, consistent with these elements being genetically mobile. Sections of the tryptophan operon and an operon encoding an ABC transporter in strain 195 were also not detected in ANAS. These insights into the diversity of Dehalococcoides genomes will improve our understanding of the physiology and evolution of these bacteria which is essential in developing effective strategies for bioremediation of PCE and TCE in the environment. Keywords: comparative genomic hybridization
2008-09-01 | GSE9612 | GEO
Project description:Tetrachloroethene (PCE) and trichloroethene (TCE) are prevalent groundwater contaminants that can be completely reductively dehalogenated by Dehalococcoides organisms. A Dehalococcoides-containing microbial consortium (ANAS) with the ability to degrade TCE to ethene, an innocuous end-product, was previously enriched from contaminated soil. A whole-genome photolithographic microarray was developed based on the genome of Dehalococcoides ethenogenes 195 (strain 195). This microarray contains probes designed to hybridize to >99% of the predicted protein-coding sequences in the strain 195 genome. DNA from ANAS was hybridized to the microarray to characterize the genomic content of the ANAS enrichment. The microarray revealed that the genes associated with central metabolism including an apparently incomplete carbon fixation pathway, cobalamin salvaging system, nitrogen fixation pathway, and five hydrogenase complexes are present in both strain 195 and ANAS. Although the gene encoding the TCE reductase tceA was detected, 13 of the 19 reductive dehalogenase genes present in strain 195 were not detected in ANAS. Additionally, 88% of the genes in predicted integrated genetic elements in strain 195 were not detected in ANAS, consistent with these elements being genetically mobile. Sections of the tryptophan operon and an operon encoding an ABC transporter in strain 195 were also not detected in ANAS. These insights into the diversity of Dehalococcoides genomes will improve our understanding of the physiology and evolution of these bacteria which is essential in developing effective strategies for bioremediation of PCE and TCE in the environment. Keywords: comparative genomic hybridization Genomic DNA from each culture was divided into replicate samples which were independently fragmented, labeled, and hybridized to arrays. Two microarrays were processed for the positive control (strain 195), two for the negative control (D. restrictus), and five for the ANAS enrichment culture(two analyses from one biological sample followed one year later by three analyses of a second biological sample).
2010-06-20 | E-GEOD-9612 | biostudies-arrayexpress
Project description:Many reductive dehalogenases (RDases) have been identified in organohalide-respiring microorganisms, and yet their substrates, specific activities, and conditions for expression are not well understood. We tested whether RDase expression varied depending on the substrate-exposure history of reductive dechlorinating communities. For this purpose, we used the enrichment culture KB-1 maintained on trichloroethene (TCE), as well as subcultures maintained on the intermediates cis-dichloroethene (cDCE) and vinyl chloride (VC). KB-1 contains a TCE-to-cDCE dechlorinating Geobacter and several Dehalococcoides strains that together harbor many of the known chloroethene reductases. Expressed RDases were identified using blue native polyacrylamide gel electrophoresis, enzyme assays in gel slices, and peptide sequencing. As anticipated but never previously quantified, the RDase from Geobacter was only detected transiently at the beginning of TCE dechlorination. The Dehalococcoides RDase VcrA and smaller amounts of TceA were expressed in the parent KB-1 culture during complete dechlorination of TCE to ethene regardless of time point or amended substrate. The Dehalococcoides RDase BvcA was only detected in enrichments maintained on cDCE as growth substrates, in roughly equal abundance to VcrA. Only VcrA was detected in subcultures enriched on VC. Enzyme assays revealed that 1,1-DCE, a substrate not used for culture enrichment, afforded the highest specific activity. trans-DCE was substantially dechlorinated only by extracts from cDCE enrichments expressing BvcA. RDase gene distribution indicated enrichment of different strains of Dehalococcoides as a function of electron acceptor TCE, cDCE, or VC. Each chloroethene reductase has distinct substrate preferences leading to strain selection in mixed communities.
| S-EPMC4551202 | biostudies-literature
Project description:Organohalide-respiring bacteria can be difficult to enrich and isolate, which can limit research on these important organisms. The goal of this research was to develop a method to rapidly (minutes to days) enrich these organisms from a mixed community. The method presented is based on the hypothesis that organohalide-respiring bacteria would be more hydrophobic than other bacteria as they dehalogenate hydrophobic compounds. The method developed tests this hypothesis by separating a portion of putative organohalide-respiring bacteria, those phylogenetically related to Dehalococcoides mccartyi, at the interface between a hydrophobic organic solvent and an aqueous medium. This novel partial separation technique was tested with a polychlorinated biphenyl-enriched sediment-free culture, a tetrachloroethene-enriched digester sludge culture, and uncontaminated lake sediment. Significantly higher fractions, up to 20.4 times higher, of putative organohalide-respiring bacteria were enriched at the interface between the medium and either hexadecane or trichloroethene. The selective partial separation of these putative organohalide-respiring bacteria occurred after 20 min, strongly suggesting that the separation was a result of physical-chemical interactions between the cell surface and hydrophobic solvent. Dechlorination activity postseparation was verified by the production of cis-dichloroethene when amended with tetrachloroethene. A longer incubation time of 6 days prior to separation with trichloroethene increased the total number of putative organohalide-respiring bacteria. This method provides a way to quickly separate some of the putative organohalide-respiring bacteria from other bacteria, thereby improving our ability to study multiple and different bacteria of potential interest and improving knowledge of these bacteria.IMPORTANCE Organohalide-respiring bacteria, bacteria capable of respiring chlorinated contaminants, can be difficult to enrich, which can limit their predictable use for the bioremediation of contaminated sites. This paper describes a method to quickly separate Dehalococcoides-like bacteria, a group of organisms containing organohalide-respiring bacteria, from other bacteria in a mixed community. From this work, Dehalococcoides-like bacteria appear to have a hydrophobic cell surface, facilitating a rapid (20 min) partial separation from a mixed culture at the surface of a hydrophobic liquid. This method was verified in a polychlorinated biphenyl-enriched sediment-free culture, an anaerobic digester sludge, and uncontaminated sediment. The method described can drastically reduce the amount of time required to partially separate Dehalococcoides-like bacteria from a complex mixed culture, improving researchers' ability to study these important bacteria.
| S-EPMC5335530 | biostudies-literature
Project description:Transcriptional target genes show functional enrichment of genes. However, how many and how significantly transcriptional target genes include functional enrichments are still unclear. To address these issues, I predicted human transcriptional target genes using open chromatin regions, ChIP-seq data and DNA binding sequences of transcription factors in databases, and examined functional enrichment and gene expression level of putative transcriptional target genes.Gene Ontology annotations showed four times larger numbers of functional enrichments in putative transcriptional target genes than gene expression information alone, independent of transcriptional target genes. To compare the number of functional enrichments of putative transcriptional target genes between cells or search conditions, I normalized the number of functional enrichment by calculating its ratios in the total number of transcriptional target genes. With this analysis, native putative transcriptional target genes showed the largest normalized number of functional enrichments, compared with target genes including 5-60% of randomly selected genes. The normalized number of functional enrichments was changed according to the criteria of enhancer-promoter interactions such as distance from transcriptional start sites and orientation of CTCF-binding sites. Forward-reverse orientation of CTCF-binding sites showed significantly higher normalized number of functional enrichments than the other orientations. Journal papers showed that the top five frequent functional enrichments were related to the cellular functions in the three cell types. The median expression level of transcriptional target genes changed according to the criteria of enhancer-promoter assignments (i.e. interactions) and was correlated with the changes of the normalized number of functional enrichments of transcriptional target genes.Human putative transcriptional target genes showed significant functional enrichments. Functional enrichments were related to the cellular functions. The normalized number of functional enrichments of human putative transcriptional target genes changed according to the criteria of enhancer-promoter assignments and correlated with the median expression level of the target genes. These analyses and characters of human putative transcriptional target genes would be useful to examine the criteria of enhancer-promoter assignments and to predict the novel mechanisms and factors such as DNA binding proteins and DNA sequences of enhancer-promoter interactions.
| S-EPMC5780744 | biostudies-literature
Project description:BackgroundTomato (Solanum lycopersicum) is both an important agricultural product and an excellent model system for studying plant-pathogen interactions. It is susceptible to bacterial wilt caused by Ralstonia solanacearum (Rs), and infection can result in severe yield and quality losses. To investigate which genes are involved in the resistance response to this pathogen, we sequenced the transcriptomes of both resistant and susceptible tomato inbred lines before and after Rs inoculation.ResultsIn total, 75.02 Gb of high-quality reads were generated from 12 RNA-seq libraries. A total of 1,312 differentially expressed genes (DEGs) were identified, including 693 up-regulated and 621 down-regulated genes. Additionally, 836 unique DEGs were obtained when comparing two tomato lines, including 27 co-expression hub genes. A total of 1,290 DEGs were functionally annotated using eight databases, most of which were found to be involved in biological pathways such as DNA and chromatin activity, plant-pathogen interaction, plant hormone signal transduction, secondary metabolite biosynthesis, and defense response. Among the core-enriched genes in 12 key pathways related to resistance, 36 genotype-specific DEGs were identified. RT-qPCR integrated analysis revealed that multiple DEGs may play a significant role in tomato response to Rs. In particular, Solyc01g073985.1 (NLR disease resistance protein) and Solyc04g058170.1 (calcium-binding protein) in plant-pathogen interaction are likely to be involved in the resistance.ConclusionWe analyzed the transcriptomes of both resistant and susceptible tomato lines during control and inoculated conditions and identified several key genotype-specific hub genes involved in a variety of different biological processes. These findings lay a foundation for better understanding the molecular basis by which resistant tomato lines respond to Rs.
| S-EPMC10061692 | biostudies-literature