WBC2 Development
This consortium of bacteria and methanogens has been developed in our U.S. Geological Survey lab at Reston, VA. This consortium, in development since 2002, was started using Aberdeen sediment, and enriched and grown in a medium with chlorinated compounds as terminal electron acceptors.
1,1,2,2 tetrachloroethane degradation in wetland sediment (WBCC)
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1,1,2,2 tetrachloroethane (TeCA) 1,1,2 trichloroethane (TCA) 1,2 dichloroethane (DCA) Chloroethane (CA) Trichloroethene (TCE) Dichloroethene (DCE) Vinyl chloride (VC) |
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*EPA states that exposure increases risk of cancer |
TeCA, TCE, TCA, DCA, DCE and VC have been observed during degradation studies in West Branch Canal Creek sediment. CA, ethene and ethane were theoretical reductive dechlorination products only.
Enriching a dechlorinating microbial consortium from contaminated sediment
Sediments from 2 sites (WB23 and WB30) in West Branch Canal Creek, MD were incubated anaerobically with 1,1,2,2 tetrachloroethane (TeCA) for a month to build up the dechlorinating population. The sediments were then split into smaller bottles and incubated with a TeCA daughter product in order to enrich (selectively stimulate) organisms capable of metabolizing the entire pathway. After several months of enrichment, the sediments (10%) were added into medium (90%). WBC-1 was sufficiently enriched to use as a sediment amendment at this time. WBC-1 was cultivated with H2 and acetate as electron donors and TeCA provided the electron acceptors, but activity was gradually lost in the culture. WBC-2 was tested with different electron donors for support of TCA, DCE and VC reduction, and lactate was chosen for cultivation. WBC-2 was cultivated with TeCA or with TeCA, TCA and DCE and parallel cultures were also grown on TCA or DCE. Culture with TeCA alone did about as well as culture with all three compounds added and also degraded TCA as well as the TCA culture and DCE as well as the DCE culture. WBC-1 was cloned and sequenced immediately after the transfer into medium (*) and had a high diversity, whereas WBC-2 was analyzed after 3 transfers into fresh medium (11 months), and is less diverse.
WBC1 Bacteria
WBC-1 was cultivated with H2 and acetate as electron donors and 1122 Tetrachloroethane as an electron acceptor. The clone library was constructed from DNA sampled soon after sediment enrichments were added to medium. Clones related to known dechlorinators Dehalobacter restrictus and Dehalococcoides strain FL2 were sequenced. The culture was successfully used to enhance dechlorination in sediment. Unfortunately the medium did not support the culture in the long term and over time dechlorination abilities were lost, resulting in an accumulation of VC and DCA. [note GNS = green non-sulfur of which Dehalococcoides is a sub-group]
WBC2 Bacteria
WBC-2, analyzed after cultivation with lactate and a chlorinated mixture for several transfers, is related to a subset of the bacteria observed in the WBC-1 clone library. Acetobacterium and Clostridium are both members of the Order Clostridiales. Acetobacteria can produce acetate from H2 and CO2, and has also been shown to dechlorinate. The Clostridium in WBC-2 is very closely to a Clostridium found in a dechlorinating system. Dehelobacter restrictus can dechlorinate TCA to VC and TCE to DCE. Several strains of CFB (cytophaga/flavobacterium/bacteriodes) were cloned and these are related to CFB found at other dechlorinating sites. The three delta Proteobacter clones were not closely related to each other: a Geobacter sp. was closely related to Geobacter Lovleyi, which can dechlorinate TCE to DCE; and a sulfate reducer related to Desulfobulbus and Syntrophus was present. The epsilon Proteobacteria were a tight group related to Arcobacter and Sulfurospirillum (not closely related to the dechlorinating Sulfurospirillum). The gamma proteobacters were related to Pseudomonas stutzeri and P. chlorodismutans. Although Dehalococcoides was not detected in the clone library, direct quantification using qPCR indicated it was present (<1%).
