Project description:Bacteria of the group “Dehalococcoides” display the ability to respire recalcitrant chlorinated organic compounds in laboratory and field site applications. Though reductive dehalogenases (RDases) have been shown to directly catalyze dechlorination reactions, the respiratory pathways and function of most genome-encoded RDases in Dehalococcoides strains remain incompletely described. In order to broaden the understanding of the biological organization of “Dehalococcoides”, this study monitored the trancriptomic response of “Dehalococcoides ethenogenes” stain 195 through microarray technology. Batch versus continuously fed cultures were examined and compared. When similarly respiring (~120 μeeq PCE/(L-hr)) batch and pseudo steady-state cultures were contrasted, the reductive dehalogenases (RDases) DET1545 and DET0180 were up-regulated in the PSS system indicating their activity at lower overall electron acceptor concentration.
Project description:Meta-Analyses of Dehalococcoides mccartyi Strain 195 Transcriptomic Profiles Identify a Respiration Rate-Related Gene Expression Transition Point and Interoperon Recruitment of a Key Oxidoreductase Subunit
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
Project description:Dehalococcoides bacteria can reductively dehalogenate a wide range of halogenated organic pollutants. In this study, DNA microarrays were applied to monitor dynamic changes in the transcriptome as Dehalococcoides ethenogenes strain 195 transitioned from exponential growth into the stationary phase. In total, 415 non-redundant genes were identified as differentially expressed. As expected, genes involved with translation and energy metabolism were down-regulated while genes involved with general stress response, transcription, and signal transduction were up-regulated. Unexpected, however, was the 8- to 10-fold up-regulation of four putative reductive dehalogenases (RDases) (DET0173, DET0180, DET1535, and DET1545). Another unexpected finding was the up-regulation of a large number of genes located within integrated elements, including a putative prophage and a muti-copy transposon. Finally, genes encoding the dominant hydrogenase-RDase respiratory chain of this strain (Hup and TceA) were expressed at stable levels throughout the experiment, providing molecular evidence that strain 195 can uncouple dechlorination from net growth. Keywords: time course, stationary phase
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:Development of a Fluorescence-Activated Cell Sorting Method Coupled with Whole Genome Amplification To Analyze Minority and Trace Dehalococcoides Genomes in Microbial Communities
Project description:Transcriptional profiling of the Donna II mixed community containing Dehalococcoides mccartyi strain 195 comparing a batch starved control to the mixed community being fed 1,2,3,4-tetrachlorobenzene as an electron acceptor. The goal was to determine which transcripts are regulated in response to a shift in a different electron acceptor rather than the consistent tetrachloroethene (PCE) that the parent reactor was maintained on.