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Applied and Environmental Microbiology, December 2004, p. 7329-7341, Vol. 70, No. 12
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.12.7329-7341.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Molecular Characterization of a Dechlorinating Community Resulting from In Situ Biostimulation in a Trichloroethene-Contaminated Deep, Fractured Basalt Aquifer and Comparison to a Derivative Laboratory Culture

Tamzen W. Macbeth,^1,2, David E. Cummings,^1, Stefan Spring,³ Lynn M. Petzke,¹ and Kent S. Sorenson Jr.^4*

Biotechnology Department, Idaho National Engineering and Environmental Laboratory,¹ North Wind, Inc., Idaho Falls,⁴ Department of Environmental Engineering, Idaho State University, Pocatello, Idaho,² Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Braunschweig, Germany³

Received 11 February 2004/ Accepted 8 August 2004

Sodium lactate additions to a trichloroethene (TCE) residual source area in deep, fractured basalt at a U.S. Department of Energy site have resulted in the enrichment of the indigenous microbial community, the complete dechlorination of nearly all aqueous-phase TCE to ethene, and the continued depletion of the residual source since 1999. The bacterial and archaeal consortia in groundwater obtained from the residual source were assessed by using PCR-amplified 16S rRNA genes. A clone library of bacterial amplicons was predominated by those from members of the class Clostridia (57 of 93 clones), of which a phylotype most similar to that of the homoacetogen Acetobacterium sp. strain HAAP-1 was most abundant (32 of 93 clones). The remaining Bacteria consisted of phylotypes affiliated with Sphingobacteria, Bacteroides, Spirochaetes, Mollicutes, and Proteobacteria and candidate divisions OP11 and OP3. The two proteobacterial phylotypes were most similar to those of the known dechlorinators Trichlorobacter thiogenes and Sulfurospirillum multivorans. Although not represented by the bacterial clones generated with broad-specificity bacterial primers, a Dehalococcoides-like phylotype was identified with genus-specific primers. Only four distinct phylotypes were detected in the groundwater archaeal library, including predominantly a clone affiliated with the strictly acetoclastic methanogen Methanosaeta concilii (24 of 43 clones). A mixed culture that completely dechlorinates TCE to ethene was enriched from this groundwater, and both communities were characterized by terminal restriction fragment length polymorphism (T-RFLP). According to T-RFLP, the laboratory enrichment community was less diverse overall than the groundwater community, with 22 unique phylotypes as opposed to 43 and a higher percentage of Clostridia, including the Acetobacterium population. Bioreactor archaeal structure was very similar to that of the groundwater community, suggesting that methane is generated primarily via the acetoclastic pathway, using acetate generated by lactate fermentation and acetogenesis in both systems.

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* Corresponding author. Present address: CDM Inc., 1331 17th St., Suite 1200, Denver, CO 80202. Phone: (303) 298-1311. Fax: (303) 293-8236. E-mail: sorensonks{at}cdm.com.

Present address: North Wind, Inc., Idaho Falls, ID 83402.

Department of Biology, Point Loma Nazarene University, San Diego, CA 92106.

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Applied and Environmental Microbiology, December 2004, p. 7329-7341, Vol. 70, No. 12
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.12.7329-7341.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

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