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Characterization of four TCE-dechlorinating microbial enrichments grown with different cobalamin stress and methanogenic conditions

ABSTRACT: Four stable and robust TCE-dechlorinating microbial communities were enriched from TCE-contaminated groundwater under four different conditions exploring two parameters, high and low methanogenic activity (Meth and NoMeth), with and without vitamin B12 supplement (MethB12 and NoMethB12, Meth and NoMeth, respectively). Identical amounts of lactate (2.7 mmol) and TCE (20 μl) were supplied as electron donor and electron acceptor. All four cultures were capable of reductively dechlorinating TCE to VC and ethene. Genomic DNA of the four enrichments was applied on a quad-Dhc-genome microarray in order to characterize the gene content of Dehalococcoides species present in the four enrichments

ORGANISM(S): Dehalococcoides mccartyi BAV1 Dehalococcoides mccartyi 195 Dehalococcoides mccartyi CBDB1 Dehalococcoides mccartyi VS environmental samples

PROVIDER: GSE33530 | GEO | 2013/06/30

SECONDARY ACCESSION(S): PRJNA148909

REPOSITORIES: GEO

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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).

Project description:The genomes of three newly isolated Dehalococcoides strains (11a, 11a5 and MB) were compared against known genomes in the Dehalococcoides genus via a microarray targeting four sequenced Dehalococcoides strains (195, CBDB1, BAV1, and VS). All three strains exhibit different dechlorination patterns, with strains 11a dechlorinating TCE to ethene, 11a5 dechlorinating TCE to VC and MB dechlorinating PCE only to isomers of DCE. Hybridization of their respective genomic DNA to the microarrays showed that the genomes of strains 11a and 11a5 show great similarity to each other and to strains CBDB1 and BAV1 of the Pinellas subgroup, while strain MB shows strong genome similarity to members of the Cornell subgroup. All genes within the respective subgroups that were not detected by microarray are within the respective high plasticity regions or integrated elements of the sequenced strains. A large number of reductive dehalogenase (RDase)-encoding genes are present within each genome, and the presence of the vcrA and tceA genes in strains 11a and 11a5 respectively, and the absence of any of the four functionally-characterized chlorinated ethene RDases (pceA, tceA, vcrA, bvcA) within strain MB appear to dictate chlorinated ethene usages regardless of the respective core genome phylogeny of the three strains. Considering the current data set together with previous comparative genomics results from application of the Dehalococcoides genus microarray to two other un-sequenced strains, the observed incongruence between the core genome phylogeny and chlorinated ethene usage of Dehalococcoides strains is likely driven by horizontal gene transfer of functional RDases. The other genomic features that are repeatedly observed in the microarray analyses of all five un-sequenced Dehalococcoides strains as well as the environmental implications on this work are presented in this study. The genomic DNA (gDNA) of each culture was analyzed in triplicate. gDNA from the two newly isolated Dehalococcoides strains 11a and 11a5 were analyzed.

Project description:A microarray targeting four sequenced strains in the Dehalococcoides (Dhc) genus was used to analyze gene expression in a robust long-term trichloroethene (TCE)-degrading microbial community (designated ANAS) during feeding cycles that involve conditions of periodic substrate supply. The Dhc transcriptome was examined at three time-points throughout a batch feeding cycle: T1 (27 h) when TCE, dichloroethene (DCE), and vinyl chloride (VC) were present; T2 (54 h) when only VC remained; and T3 (13 d) when Dhc had been starved of substrate for nine days. 90% of the Dhc ORFs that were detected in the ANAS DNA were found to be expressed as RNA sometime during the time course, demonstrating extraordinary utilization of the streamlined genome. 97% of these transcripts were differentially expressed during the time course, indicating efficiency of transcription through regulation in Dhc. Most Dhc genes were significantly down-regulated at T3, responding to a lack of substrate as would be expected. The tceA and vcrA genes, which code for proteins with known chlorinated ethene reduction functions, were highly expressed at both T1 and T2, whereas two other putative reductive dehalogenase genes (DET0173 and DET1545) were most highly expressed at T2, likely in response to the presence of VC. Hydrogenases were most highly expressed at T1, reflecting their important role in accumulating electrons used to initiate reductive dechlorination and other biosynthesis pathways. Cobalamin transport genes were preferentially expressed at T2, reflecting an increase in corrinoid transport as chloroethenes were degraded and a decrease in activity of the transport system after dehalogenation was complete. This is the first application of a microarray targeting a known genus, including both core genomes and identified strain-specific genes, applied to improve our understanding of transcriptional dynamics within an undefined microbial community. Replicate samples were independently collected, and simultaneously but individually extracted, fragmented, labeled, and hybridized to arrays. Three DNA samples (one from each of the three replicate cycles) and nine RNA samples (one from each of the three time-points in each of the three replicate cycles) were prepared for microarray analysis.

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Characterization of four TCE-dechlorinating microbial enrichments grown with different cobalamin stress and methanogenic conditions

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