Complete detoxification of short chain chlorinated aliphatics: Isolation of halorespiring organisms and biochemical studies of the dehalogenating enzyme systems. 1997 annual progress report
'The objectives of the research within this grant are: (1) Isolation and characterization of chlororespiring organisms responsible for the complete dehalogenation of chlorinated ethenes and propanes. (2) Development of conditions that yield high cell densities and induce dechlorinating activity. (3) Development of assay systems to detect the dechlorinating activity in cell-free extracts. (4) Purification and characterization of the dehalogenating enzymes. Anaerobic microcosms were obtained from a variety of geographically different sediment samples. In several microcosms complete dechlorination of tetrachloroethene (PCE) to ethene (ETH), and 1,2-dichloropropane ( 1,2-D) and/or 1,2,3-trichloropropane to propene was observed. Upon subsequent transfers to anaerobic medium, sediment-free, methanogenic enrichment cultures were obtained that dechlorinated PCE to ETH, and 1,2-D to propene, respectively. 2-Bromoethanesulfonate (BES), a well known inhibitor of methanogens, did not inhibit the dechlorination of 1,2-D to propene and the dechlorination of PCE to cis-dichloroethene (cis-DCE). However,-the complete dechlorination of PCE to vinyl chloride (VC) and ETH was severely inhibited. The authors could show that BES inhibited the dechlorination of chloroethenes in cultures not containing methanogens. Previous to this study, BES was believed to be aspecific inhibitor of methanogens and the inhibitory effect of BES on declorination was explained by the involvement of methanogens in the dechlorination process. The non-methanogenic cultures obtained after the BES treatment were subsequently transferred to medium riot containing BES and complete dechlorination of PCE to ETH was observed as was in the original microcosms. Subcultures were further enriched with PCE, cis-DCE, VC, or 1,2-D as the only available electron acceptor and acetate, or acetate plus hydrogen as the only available electron donor(s). To date these cultures have undergone up to 45 transfers. Interestingly, two cultures that originally dechlorinated PCE to ETH, but were then enriched with cis-DCE or VC, lost their ability to-dechlorinate PCE or TCE. This finding indicates that different populations are involved in the complete dechlorination of PCE to ETH in these cultures. In contrast, one culture that was enriched with cis-DCE and VC, respectively, maintained its ability to dechlorinate PCE. Using molecular tools ({sup 16}S rDNA targeted PCR,TA cloning of {sup 1}6S rDNA genes, ARDRA analysis and sequencing) they showed that this culture consisted of three distinct organisms. Two of them could be isolated in pure Culture but neither of them showed any dechlorinating activity, indicating that one organism was responsible for the complete dechlorination of PCE to ETH in the mixed culture. The authors are currently focusing on the isolation and phylogenetic characterization of this organism.'
- Research Organization:
- Michigan State Univ., East Lansing, MI (US)
- Sponsoring Organization:
- USDOE Office of Environmental Management (EM), Office of Science and Risk Policy
- DOE Contract Number:
- FG07-96ER62319
- OSTI ID:
- 13697
- Report Number(s):
- EMSP-55105-97; ON: DE00013697
- Country of Publication:
- United States
- Language:
- English
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