Showing papers on "Dehalococcoides published in 2014"

Genomic characterization of three unique Dehalococcoides that respire on persistent polychlorinated biphenyls

Abstract: Fastidious anaerobic bacteria play critical roles in environmental bioremediation of halogenated compounds. However, their characterization and application have been largely impeded by difficulties in growing them in pure culture. Thus far, no pure culture has been reported to respire on the notorious polychlorinated biphenyls (PCBs), and functional genes responsible for PCB detoxification remain unknown due to the extremely slow growth of PCB-respiring bacteria. Here we report the successful isolation and characterization of three Dehalococcoides mccartyi strains that respire on commercial PCBs. Using high-throughput metagenomic analysis, combined with traditional culture techniques, tetrachloroethene (PCE) was identified as a feasible alternative to PCBs to isolate PCB-respiring Dehalococcoides from PCB-enriched cultures. With PCE as an alternative electron acceptor, the PCB-respiring Dehalococcoides were boosted to a higher cell density (1.2 × 10(8) to 1.3 × 10(8) cells per mL on PCE vs. 5.9 × 10(6) to 10.4 × 10(6) cells per mL on PCBs) with a shorter culturing time (30 d on PCE vs. 150 d on PCBs). The transcriptomic profiles illustrated that the distinct PCB dechlorination profile of each strain was predominantly mediated by a single, novel reductive dehalogenase (RDase) catalyzing chlorine removal from both PCBs and PCE. The transcription levels of PCB-RDase genes are 5-60 times higher than the genome-wide average. The cultivation of PCB-respiring Dehalococcoides in pure culture and the identification of PCB-RDase genes deepen our understanding of organohalide respiration of PCBs and shed light on in situ PCB bioremediation.

Bioremediation of Chlorinated Ethenes in Fractured Bedrock and Associated Changes in Dechlorinating and Nondechlorinating Microbial Populations

Abstract: The use of enhanced in situ anaerobic bioremediation (EISB) and bioaugmentation in fractured bedrock is limited compared to its use in granular media. We evaluated EISB for the treatment of trichloroethene (TCE)-impacted groundwater in fractured carbonate rock at a site in Southern Ontario, Canada, with cool average groundwater temperature (∼ 13 °C). Borehole-connectivity, contaminant concentrations, and groundwater properties were investigated. Changes in dechlorinating and nondechlorinating populations (fermenters, acetogens, methanogens, and sulfate reducers) were assessed via quantitative PCR (qPCR). During biostimulation with ethanol, concentrations of TCE daughter products cis-dichloroethene (cDCE) and vinyl chloride (VC) decreased in association with an enrichment of vcrA (VC reductive dehalogenase)-carrying Dehalococcoides, whereas ethene production was only moderate. Following bioaugmentation with the mixed dechlorinating culture KB-1, greater concentrations of chloride-a product of dechlorination-was observed in most wells; in addition, ethene production increased significantly in monitoring well locations that had strong hydraulic connectivity to the groundwater recirculation system, while Dehalococcoides and vcrA concentrations did not appreciably vary. Interestingly, increases of 3-4 orders of magnitude of an ethanol-fermenting Bacteroidetes population also present in KB-1 were correlated to improved conversion to ethene, an observation which suggests there could be a causal relationship-for example, better syntrophy and/or synergy among bacterial populations.

Current trends in trichloroethylene biodegradation: a review.

Abstract: Over the past few years biodegradation of trichloroethylene (TCE) using different microorganisms has been investigated by several researchers. In this review article, an attempt has been made to present a critical summary of the recent results related to two major processes--reductive dechlorination and aerobic co-metabolism used for TCE biodegradation. It has been shown that mainly Clostridium sp. DC-1, KYT-1, Dehalobacter, Dehalococcoides, Desulfuromonas, Desulfitobacterium, Propionibacterium sp. HK-1, and Sulfurospirillum bacterial communities are responsible for the reductive dechlorination of TCE. Efficacy of bacterial communities like Nitrosomonas, Pseudomonas, Rhodococcus, and Xanthobacter sp. etc. for TCE biodegradation under aerobic conditions has also been examined. Mixed cultures of diazotrophs and methanotrophs have been used for TCE degradation in batch and continuous cultures (biofilter) under aerobic conditions. In addition, some fungi (Trametes versicolor, Phanerochaete chrysosporium ME-446) and Actinomycetes have also been used for aerobic biodegradation of TCE. The available information on kinetics of biofiltration of TCE and its degradation end-products such as CO2 are discussed along with the available results on the diversity of bacterial community obtained using molecular biological approaches. It has emerged that there is a need to use metabolic engineering and molecular biological tools more intensively to improve the robustness of TCE degrading microbial species and assess their diversity.

Dehalogenation of chlorobenzenes, dichlorotoluenes, and tetrachloroethene by three Dehalobacter spp.

Abstract: Three enrichment cultures containing Dehalobacter spp. were developed that dehalogenate each of the dichlorobenzene (DCB) isomers to monochlorobenzene (MCB), and the strains using 1,2-DCB (12DCB1) or 1,3-DCB (13DCB1) are now considered isolated, whereas the strain using 1,4-DCB (14DCB1) is considered highly enriched. In this study, we examined the dehalogenation capability of each strain to use chlorobenzenes with three or more chlorines, tetrachloroethene (PCE), or dichlorotoluene (DCT) isomers. Strain 12DCB1 preferentially dehalogenated singly flanked chlorines, but not doubly flanked or unflanked chlorines. It dehalogenated pentachlorobenzene to MCB with little buildup of intermediates. Strain 13DCB1, which could use either 1,3-DCB or 1,2-DCB, demonstrated the widest dehalogenation spectrum of electron acceptors tested, and dehalogenated every chlorobenzene isomer except 1,4-DCB. Notably, strain 13DCB1 dehalogenated the recalcitrant 1,3,5-trichlorobenzene isomer to MCB, and qPCR of 16S rRNA genes indicated that strain 13DCB1 grew. Strain 14DCB1 exhibited the narrowest range of substrate utilization, but was the only strain to dehalogenate para-substituted chlorines. Strains 12DCB1 and 13DCB1 dehalogenated PCE to cis-dichloroethene, and all strains dehalogenated 3,4-DCT to monochlorotoluene. These findings show that Dehalobacter spp., like Dehalococcoides spp., are versatile dehalogenators and should be considered when determining the fate of chlorinated organics at contaminated sites.

Selective Enrichment Yields Robust Ethene-Producing Dechlorinating Cultures from Microcosms Stalled at cis-Dichloroethene

Abstract: Dehalococcoides mccartyi strains are of particular importance for bioremediation due to their unique capability of transforming perchloroethene (PCE) and trichloroethene (TCE) to non-toxic ethene, through the intermediates cis-dichloroethene (cis-DCE) and vinyl chloride (VC). Despite the widespread environmental distribution of Dehalococcoides, biostimulation sometimes fails to promote dechlorination beyond cis-DCE. In our study, microcosms established with garden soil and mangrove sediment also stalled at cis-DCE, albeit Dehalococcoides mccartyi containing the reductive dehalogenase genes tceA, vcrA and bvcA were detected in the soil/sediment inocula. Reductive dechlorination was not promoted beyond cis-DCE, even after multiple biostimulation events with fermentable substrates and a lengthy incubation. However, transfers from microcosms stalled at cis-DCE yielded dechlorination to ethene with subsequent enrichment cultures containing up to 109 Dehalococcoides mccartyi cells mL−1. Proteobacterial classes which dominated the soil/sediment communities became undetectable in the enrichments, and methanogenic activity drastically decreased after the transfers. We hypothesized that biostimulation of Dehalococcoides in the cis-DCE-stalled microcosms was impeded by other microbes present at higher abundances than Dehalococcoides and utilizing terminal electron acceptors from the soil/sediment, hence, outcompeting Dehalococcoides for H2. In support of this hypothesis, we show that garden soil and mangrove sediment microcosms bioaugmented with their respective cultures containing Dehalococcoides in high abundance were able to compete for H2 for reductive dechlorination from one biostimulation event and produced ethene with no obvious stall. Overall, our results provide an alternate explanation to consolidate conflicting observations on the ubiquity of Dehalococcoides mccartyi and occasional stalling of dechlorination at cis-DCE; thus, bringing a new perspective to better assess biological potential of different environments and to understand microbial interactions governing bioremediation.

Use of silicate minerals for pH control during reductive dechlorination of chloroethenes in batch cultures of different microbial consortia

Abstract: In chloroethene-contaminated sites undergoing in situ bioremediation, groundwater acidification is a frequent problem in the source zone, and buffering strategies have to be implemented to maintain the pH in the neutral range. An alternative to conventional soluble buffers is silicate mineral particles as a long-term source of alkalinity. In previous studies, the buffering potentials of these minerals have been evaluated based on abiotic dissolution tests and geochemical modeling. In the present study, the buffering potentials of four silicate minerals (andradite, diopside, fayalite, and forsterite) were tested in batch cultures amended with tetrachloroethene (PCE) and inoculated with different organohalide-respiring consortia. Another objective of this study was to determine the influence of pH on the different steps of PCE dechlorination. The consortia showed significant differences in sensitivities toward acidic pH for the different dechlorination steps. Molecular analysis indicated that Dehalococcoides spp. that were present in all consortia were the most pH-sensitive organohalide-respiring guild members compared to Sulfurospirillum spp. and Dehalobacter spp. In batch cultures with silicate mineral particles as pH-buffering agents, all four minerals tested were able to maintain the pH in the appropriate range for reductive dechlorination of chloroethenes. However, complete dechlorination to ethene was observed only with forsterite, diopside, and fayalite. Dissolution of andradite increased the redox potential and did not allow dechlorination. With forsterite, diopside, and fayalite, dechlorination to ethene was observed but at much lower rates for the last two dechlorination steps than with the positive control. This indicated an inhibition effect of silicate minerals and/or their dissolution products on reductive dechlorination of cis-dichloroethene and vinyl chloride. Hence, despite the proven pH-buffering potential of silicate minerals, compatibility with the bacterial community involved in in situ bioremediation has to be carefully evaluated prior to their use for pH control at a specific site.

Successful operation of continuous reactors at short retention times results in high-density, fast-rate Dehalococcoides dechlorinating cultures

Abstract: The discovery of Dehalococcoides mccartyi reducing perchloroethene and trichloroethene (TCE) to ethene was a key landmark for bioremediation applications at contaminated sites. D. mccartyi-containing cultures are typically grown in batch-fed reactors. On the other hand, continuous cultivation of these microorganisms has been described only at long hydraulic retention times (HRTs). We report the cultivation of a representative D. mccartyi-containing culture in continuous stirred-tank reactors (CSTRs) at a short, 3-d HRT, using TCE as the electron acceptor. We successfully operated 3-d HRT CSTRs for up to 120 days and observed sustained dechlorination of TCE at influent concentrations of 1 and 2 mM TCE to ≥ 97 % ethene, coupled to the production of 10(12) D. mccartyi cells Lculture (-1). These outcomes were possible in part by using a medium with low bicarbonate concentrations (5 mM) to minimize the excessive proliferation of microorganisms that use bicarbonate as an electron acceptor and compete with D. mccartyi for H2. The maximum conversion rates for the CSTR-produced culture were 0.13 ± 0.016, 0.06 ± 0.018, and 0.02 ± 0.007 mmol Cl(-) Lculture (-1) h(-1), respectively, for TCE, cis-dichloroethene, and vinyl chloride. The CSTR operation described here provides the fastest laboratory cultivation rate of high-cell density Dehalococcoides cultures reported in the literature to date. This cultivation method provides a fundamental scientific platform for potential future operations of such a system at larger scales.

DNA extraction-free quantification of Dehalococcoides spp. in groundwater using a hand-held device.

Abstract: Nucleic acid amplification of biomarkers is increasingly used to measure microbial activity and predict remedial performance in sites with trichloroethene (TCE) contamination. Field-based genetic quantification of microorganisms associated with bioremediation may help increase accuracy that is diminished through transport and processing of groundwater samples. Sterivex cartridges and a previously undescribed mechanism for eluting biomass was used to concentrate cells. DNA extraction-free loop mediated isothermal amplification (LAMP) was monitored in real-time with a point of use device (termed Gene-Z). A detection limit of 10(5) cells L(–1) was obtained, corresponding to sensitivity between 10 to 100 genomic copies per reaction for assays targeting the Dehalococcoides spp. specific 16S rRNA gene and vcrA gene, respectively. The quantity of Dehalococcoides spp. genomic copies measured from two TCE contaminated groundwater samples with conventional means of quantification including filtration, DNA extraction, purification, and qPCR was comparable to the field ready technique. Overall, this method of measuring Dehalococcoides spp. and vcrA genes in groundwater via direct amplification without intentional DNA extraction and purification is demonstrated, which may provide a more accurate mechanism of predicting remediation rates.

Diversity of dechlorination pathways and organohalide respiring bacteria in chlorobenzene dechlorinating enrichment cultures originating from river sludge

Abstract: Anaerobic reductive dechlorination of hexachlorobenzene (HCB) and three isomers of tetrachlorobenzene (TeCB) (1,2,3,4-, 1,2,3,5- and 1,2,4,5-TeCB) was investigated in microcosms containing chloroaromatic contaminated river sediment. All chlorobenzenes were dechlorinated to dichlorobenzene (DCB) or monochlorobenzene. From the sediment, a methanogenic sediment-free culture was obtained which dechlorinated HCB, pentachlorobenzene, three TeCB isomers, three trichlorobenzene (TCB) isomers (1,2,3-, 1,2,4- and 1,3,5-TCB) and 1,2-DCB. Dechlorination involved multiple pathways including the removal of doubly flanked, singly flanked and isolated chlorine substituents. 454-pyrosequencing of partial bacterial 16S rRNA genes amplified from selected chlorobenzene dechlorinating sediment-free enrichment cultures revealed the presence of a variety of bacterial species, including Dehalobacter and Dehalococcoides mccartyi, that were previously documented as organohalide respiring bacteria. A genus with apparent close relationship to Desulfitobacterium that also has been associated with organohalide respiration, composed the major fraction of the operational taxonomic units (OTUs). Another major OTU was linked with Sedimentibacter sp., a genus that was previously identified in strict co-cultures of consortia reductively dehalogenating chlorinated compounds. Our data point towards the existence of multiple interactions within highly chlorinated benzene dechlorinating communities.

Effects of bioaugmentation on enhanced reductive dechlorination of 1,1,1-trichloroethane in groundwater - a comparison of three sites

Abstract: Microcosm studies investigated the effects of bioaugmentation with a mixed Dehalococcoides (Dhc)/Dehalobacter (Dhb) culture on biological enhanced reductive dechlorination for treatment of 1,1,1-trichloroethane (TCA) and chloroethenes in groundwater at three Danish sites. Microcosms were amended with lactate as electron donor and monitored over 600 days. Experimental variables included bioaugmentation, TCA concentration, and presence/absence of chloroethenes. Bioaugmented microcosms received a mixture of the Dhc culture KB-1 and Dhb culture ACT-3. To investigate effects of substrate concentration, microcosms were amended with various concentrations of chloroethanes (TCA or monochloroethane [CA]) and/or chloroethenes (tetrachloroethene [PCE], trichloroethene [TCE], or 1,1-dichloroethene [1,1-DCE]). Results showed that combined electron donor addition and bioaugmentation stimulated dechlorination of TCA and 1,1-dichloroethane (1,1-DCA) to CA, and dechlorination of PCE, TCE, 1,1-DCE and cDCE to ethane. Dechlorination of CA was not observed. Bioaugmentation improved the rate and extent of TCA and 1,1-DCA dechlorination at two sites, but did not accelerate dechlorination at a third site where geochemical conditions were reducing and Dhc and Dhb were indigenous. TCA at initial concentrations of 5 mg/L inhibited (i.e., slowed the rate of) TCA dechlorination, TCE dechlorination, donor fermentation, and methanogenesis. 1 mg/L TCA did not inhibit dechlorination of TCA, TCE or cDCE. Moreover, complete dechlorination of PCE to ethene was observed in the presence of 3.2 mg/L TCA. In contrast to some prior reports, these studies indicate that low part-per million levels of TCA (<3 mg/L) in aquifer systems do not inhibit dechlorination of PCE or TCE to ethene. In addition, the results show that co-bioaugmentation with Dhc and Dhb cultures can be an effective strategy for accelerating treatment of chloroethane/chloroethene mixtures in groundwater, with the exception that all currently known Dhc and Dhb cultures cannot treat CA.

Predominance of Dehalococcoides in the presence of different sulfate concentrations

Abstract: This is the first study that investigates in detail the effect of different sulfate concentrations on trichloroethene-dechlorinating microbial communities, both in terms of dechlorinating performance and microbial composition. The study used a series of Dehalococcoides-containing trichloroethene-dechlorinating microbial communities, which operated for more than 800 days in the presence of different sulfate concentrations and limiting-electron donor conditions. This study proves the ability of Dehalococcoides spp., the only genus able to completely dechlorinate trichloroethene, to predominate in mixed anaerobic microbial communities regardless of the magnitude of sulfate concentration, even under limiting-electron donor conditions. Although other microorganisms, such as the Sulfurospirillum spp. bacteria and members of the sulfate-reducing bacteria group were able to thrive, they were not able to predominate in such a competitive environment. However, this picture was not reflected in reductive dechlorination, which demonstrated a much better performance under methanogenic conditions or in the presence of low sulfate concentration (30 mg/l) than in the presence of higher sulfate concentrations (>400 mg/l). Therefore, different species of Dehalococcoides or other dechlorinating bacteria, which are not able to thrive in the presence of high sulfate concentrations (>400 mg/l), are possibly responsible for the higher dechlorination efficiency that was observed under methanogenic conditions.

Biostimulation of indigenous communities for the successful dechlorination of tetrachloroethene (perchloroethylene)-contaminated groundwater

Abstract: Chlorinated ethenes are of environmental concern with most reports of successful microbial-mediated remediation being associated with major dechlorinating groups such as Dehalococcoides (Dhc) species. However, limited information is available on the community dynamics and dechlorinating activities of indigenous non-Dhc groups. Here, we present evidence of dechlorination of tetrachloroethene (perchloroethylene, PCE) in groundwater samples by indigenous microbial communities. 100 % PCE conversion to ethene was observed in acetate-stimulated 24 week-microcosms (controls; 15 %). Microbial community profiles showed dominance by groups such as Proteobacteria, Spirochaetes, Firmicutes, Methanomicrobiaceae and Methanosarcinaceae. Pareto-Lorenz (PL) analyses suggested an adapted (45 % PL value) but variable bacterial community (55.5 % Δt(week)) compared to Archaea (25 % PL value; 46.9 % Δt(week)). Our findings provide evidence of dechlorinating potential of indigenous microorganisms and useful information on their dynamics which may be exploited for in situ groundwater bioremediation.

Molecular signatures for members of the genus Dehalococcoides and the class Dehalococcoidia.

Abstract: The bacteria belonging to the class Dehalococcoidia , due to their ability to dehalogenate chlorinated compounds, are of much interest for bioremediation of contaminated sites. We report here comparative analyses on different genes/proteins from the genomes of members of the class Dehalococcoidia . These studies have identified numerous novel molecular markers in the forms of conserved signature indels (CSIs) in broadly distributed proteins and conserved signature genes/proteins (CSPs), which are uniquely found in members of the class Dehalococcoidia , but except for an isolated exception, they are not found in other sequenced bacterial genomes. Of these molecular markers, nine CSIs in divergent proteins and 19 CSPs are specific for members of the genera Dehalococcoides and Dehalogenimonas , providing potential molecular markers for the bacterial class Dehalococcoidia . Additionally, four CSIs in divergent proteins and 28 CSPs are only found in all members of the genus Dehalococcoides for which genome sequences are available, but they are absent in Dehalogenimonas lykanthroporepellens and in other bacteria. The gene sequences of several of these CSPs exhibiting specificity for the genus Dehalococcoides or the class Dehalococcoidia are highly conserved and PCR primers based upon them provide a novel means for identification of other related bacteria. Two other CSIs identified in this study in the SecD and aspartate carbomyltransferase proteins weakly support an affiliation of the class Dehalococcoidia with the other members of the phylum Chloroflexi.

Detection of Dehalococcoides spp. by peptide nucleic acid fluorescent in situ hybridization.

Abstract: Chlorinated solvents including tetrachloroethene (perchloroethene and trichloroethene), are widely used industrial solvents. Improper use and disposal of these chemicals has led to a widespread contamination. Anaerobic treatment technologies that utilize Dehalococcoides spp. can be an effective tool to remediate these contaminated sites. Therefore, the aim of this study was to develop, optimize and validate peptide nucleic acid (PNA) probes for the detection of Dehalococcoides spp. in both pure and mixed cultures. PNA probes were designed by adapting previously published DNA probes targeting the region of the point mutations described for discriminating between the Dehalococcoides spp. strain CBDB1 and strain 195 lineages. Different fixation, hybridization and washing procedures were tested. The results indicated that the PNA probes hybridized specifically and with a high sensitivity to their corresponding lineages, and that the PNA probes developed during this work can be used in a duplex assay to distinguish between strain CBDB1 and strain 195 lineages, even in complex mixed cultures. This work demonstrates the effectiveness of using PNA fluorescence in situ hybridization to distinguish between two metabolically and genetically distinct Dehalococcoides strains, and they can have strong implications in the monitoring and differentiation of Dehalococcoides populations in laboratory cultures and at contaminated sites.

Methods, Systems, and Culture Medium for Production of Dechlorinating Microorganisms

Abstract: Methods, systems, and compositions for growing high density cultures of dechlorinating microorganisms, such as the bacteria Dehalococcoides . Dechlorinating cultures are grown in continuous flow stirred-tank reactors at short hydraulic retention time, resulting in improved batch production. For some cultures, a culture medium including chlorine containing compounds, bicarbonate, and HEPES utilized.

DEVELOPMENT AND EVALUATION OF AN ENRICHMENT CULTURE FOR REDUCTIVE DECHLORINATION OF TETRACHLOROETHENE UNDER LOW pH CONDITIONS

Abstract: Perchloroethene (PCE) is a pollutant of major environmental concern at hazardous waste sites worldwide. PCE and trichloroethene (TCE) are suspected carcinogens and are ranked 16 and 31, respectively, on the Environmental Protection Agency’s priority list for hazardous substances, developed under the Comprehensive Environmental Response, Compensation, and Liability Act. As a consequence of the widespread use of chlorinated solvents (including PCE and TCE) for dry cleaning, chemical feedstocks, metal degreasing and other purposes, chloroethenes are widely distributed in the environment. Many soils and groundwater throughout the world are contaminated by chloroethenes. Therefore, further improvements are needed in clean-up methods. Bioaugmentation has been used extensively to treat aquifers contaminated with chlorinated ethenes at sites that lack the microbes needed to accomplish reductive dechlorination at a reasonable rate. However, a major limitation to bioaugmentation has been aquifer pH. Dehalococcoides are required for achieving complete dechlorination to ethene, yet their reported pH optimum is approximately 6.5 to 7.5. To account for this in aquifers with a lower pH level, buffers have been added prior to injection of culture. However, buffer addition can lead to clogging by precipitates, the chemical costs can be substantial, and achieving homogenous distribution is very challenging. One alternative is to use bioaugmentation cultures that are able to function at lower pH levels. The observation of complete dechlorination of PCE and TCE at some sites with pH levels below 6 suggest this should be achievable. However, very limited information is

New microorganism that dechlorinate volatile organochlorine compounds

Abstract: PROBLEM TO BE SOLVED: To provide a method for quickly and completely decontaminating a polluted environment of which remediation has not been possible due to the absence of useful microorganisms, by isolating and using microorganisms that can completely dechlorinate chlorinated ethylenes and chlorinated ethanes to ethylene.SOLUTION: Provided is a new microorganism of Dehalococcoides having an ability to completely dechlorinate chlorinated ethylenes and chlorinated ethanes. Also provided is a new microorganism of Sulfurospirillum having an ability to enhance the dechlorination reaction of the above Dehalococcoides microorganism to dechlorinate chlorinated ethylenes and chlorinated ethanes. Also provided is a method for decontaminating the environment using these microorganisms.

A systems-level understanding of electron flow in TCE-dechlorinating microbial communities using modeling and molecular biology tools

Abstract: Author(s): Mao, Xinwei | Advisor(s): Alvarez-Cohen, Lisa | Abstract: Groundwater and soils have been frequently contaminated by trichloroethene (TCE), perchloroethene (PCE) and other chlorinated compounds in the U.S. and worldwide, in spite of their established toxicity and mutagenicity towards many organisms, including humans. In order to protect public health, bioremediation using Dehalococcoides-containing microbial communities is a promising approach to reach ecotoxicological-safety endpoints. The overall goal of this research is to understand electron flows in complex dechlorinating microbial communities, and to develop mathematical models to predict the performance of the microbial communities in different environmental conditions. To accomplish these goals, we first studied the electron flow and material exchange of constructed TCE-dechlorinating consortia. We also applied emerging molecular techniques to study TCE-dechlorinating microbial communities under different remediation conditions. Furthermore, we developed integrated thermodynamic and kinetic models to predict the dechlorination performance and microbial growth of syntrophic consortia under batch and continuous-flow conditions, and the suite of models were further validated using enrichment cultures. The first objective of this research was to understand the material and energy exchange between Dehalococcoides and its supporting syntrophic bacteria. We investigated dechlorination activity, cell growth, cell aggregate formation, and global gene expression of D. mccartyi strain 195 (strain 195) grown with Syntrophomonas wolfei in co-cultures amended with butyrate and TCE. By applying thermodynamically consistent rate laws to study the electrons flows in the co-culture, we found that the growth rates of the two species were strictly coupled by H2 transfer, and that the growth yield of syntrophic bacteria and the ratio maintained in the co-cultures were mainly controlled by thermodynamics. We demonstrated, for the first time, that D. mccartyi could form cell aggregates with its supporting fermenter S. wolfei on butyrate. Furthermore, we found carbon monoxide (CO) may serve as a supplemental energy source for S. wolfei during syntrophic fermentation with strain 195, and that the observed increased cell yields of strain 195 is likely due to the continuous removal of CO in the co-culture. In order to understand the microbial community structure shift from "feast-and-famine" condition (semi-batch) to the continuous feeding of low nutrients condition (completely-mixed flow reactor (CMFR)), molecular techniques based on 16S I-tags and metagenomic sequencing were applied to investigate the dechlorinating community structural shift after transition from semi-batch to a long-term steady-state CMFR condition. A Dehalococcoides genus-wide microarray was also applied to study the transcriptional dynamics of D. mccartyi strains within the CMFR community that was grown in the continuous-flowing diluted, nutrient poor environment. I-tags and metagenomic sequencing analysis revealed that dominant species in the CMFR shifted significantly from the semi-batch culture condition while the ratio of D. mccartyi was maintained at relatively stable levels. Transcriptional analysis identified tceA and vcrA to be among the most expressed genes in the CMFR, hup and vhu were more critical hydrogenases utilized by Dehalococcoides sp. in the continuous-flowing system. In contrast, corrinoid-related uptake and modification genes were expressed at lower levels in the CMFR than in the semi-batch culture during active dechlorination. A systems-level approach was applied to determine accurate kinetic parameters involved in reductive dechlorination from simple constructed syntrophic consoria to complex microbial communities. The results demonstrated that the kinetic parameters involved in reductive dechlorination were in similar ranges for simple and complex Dehalococcoides-containing cultures. Cell growth calculations showed H2 was the most sensitive factor limiting the growth of H2-utilizing microorganisms involved in dechlorinating communities. High concentrations of acetate resulted in slower dechlorination rates by inhibiting the growth of specific fermenting bacteria. High sulfate concentrations also hindered dechlorination performance due to either sulfide inhibition or competition with sulfate reduction. The mechanism for observed slower dechlorination rates with lower bicarbonate concentrations was not clear and further experiments need to be conducted to evaluate the role of bicarbonate in reductive dechlorination communities. Based on the knowledge obtained in the previous studies, an integrated thermodynamic and kinetic model was developed to predict reductive dechlorination and cell growth under batch growth conditions. The model parameters calculated to fit the experimental data were at the same levels as those determined experimentally. The resultant model accurately captured the dechlorination kinetics in two Dehalococcoides-containing syntrphic co-cultures using different fermenting substrates. The model was validated at different donor to acceptor ratios in syntrophic co-cultures and in syntrophic tri-cultures and enrichment cultures performing hydrogenotrophic methanogenesis. The sensitivity of kinetic parameters on model stability was tested. Half velocity constants and inhibition coefficients were found to be the most sensitive factors affecting model predictions.The significance of this research is to provide a more fundamental understanding of the metabolic exchange and energy transfer among the key players of TCE-dechlorinating communities, as well as the physiology of dechlorinating microbial communities experiencing different environmental stresses. The integrated thermodynamic and kinetic models developed in this study could be used as a platform to incorporate more biological processes under different experimental conditions. The knowledge developed in this research will aid practitioners to better design, monitor and optimize future in situ bioremediation systems.