Phylogenetic Characterization of Microbial Communities That Reductively Dechlorinate TCE Based upon a Combination of Molecular Techniques

Abstract

An anaerobic microbial consortium (referred to as ANAS) that reductively dechlorinates trichloroethene (TCE) completely to ethene with the transient production of cisdichloroethene (cDCE) and vinyl chloride was enriched from contaminated soil obtained from Alameda Naval Air Station. ANAS uses lactate as its electron donor and has been functionally stable for over 2 years. Following a brief exposure to oxygen, a subculture (designated VCC) derived from ANAS could dechlorinate TCE only to vinyl chloride with lactate as its electron donor. Three molecular methods were used concurrently to characterize the community structure of ANAS and VCC: clone library construction/clone sequencing, terminal restriction fragment length polymorphism (T-RFLP) analysis, and fluorescent in situ hybridization (FISH) with rRNA probes. The community structure of ANAS did not change significantly over the course of a single feeding/dechlorination cycle, and only minor fluctuations occurred over many feeding cycles spanning the course of 1 year. Clone libraries and T-RFLP analyses suggested that ANAS was dominated by populations belonging to three phylogenetic groups: Dehalococcoides species, Desulfovibrio species, and members of the Clostridiaceae (within the low G + C Gram-positives). FISH results suggest that members of the Cytophaga/Flavobacterium/Bacteroides (CFB) cluster and high G + C Gram-positives (HGCs) were numerically important in ANAS despite their under-representation in the clone libraries. Parallel analyses of VCC samples suggested that Dehalococcoides species and Clostridiaceae were only minor populations in this community. Instead, VCC had increased populations of organisms in the beta and gamma subclasses of the Proteobacteria as well as significant populations of organisms in the CFB cluster. It is possible that symbiotic interactions are occurring between some of ANAS's phylogenetic groups under the enrichment conditions, including interspecies hydrogen transfer from Desulfovibrio species to Dehalococcoides species. However, the nucleic acid-based analyses performed here would need to be supplemented with chemical species data in order to test any hypotheses about functional roles of various community members. Additionally, these results suggest that an organism outside the Dehalococcoides genus may be capable of dechlorinating cDCE to vinyl chloride.

Metagenomic analysis of a stable trichloroethene-degrading microbial community

“…The description of the community composition derived from metagenomic analysis is generally consistent with those of previous 16S clone library studies (Richardson et al, 2002;Lee et al, 2006) and the PhyloChip study presented here. Overall, data from the clone libraries and metagenome sequencing agreed on the most abundant bacterial taxa, which were also detected by the PhyloChip.…”

“…ANAS enrichment culture and DNA sample preparation Culture conditions and maintenance procedures for ANAS have been described previously (Richardson et al, 2002). Briefly, 350 ml of culture was grown at 25-28 1C and 1.8 atm with a N 2 -CO 2 (90:10) headspace in a 1.5 l continuously stirred semi-batch reactor.…”

“…This culture was derived from sediment collected at the Alameda Naval Air Station and is referred to as ANAS (Richardson et al, 2002). The phylogenetic composition of ANAS has been studied using clone libraries (Richardson et al, 2002;Lee et al, 2006), and the Dhc strains in ANAS have been analyzed using quantitative PCR and whole-genome microarrays West et al, 2008;Lee et al, 2011).…”

Effects of the antimicrobial tylosin on the microbial community structure of an anaerobic sequencing batch reactor

“…DNA extracts were visualized using electrophoresis on a 0.8% agarose gel. PCR targeting the 16S rRNA gene of all bacteria was performed as described by Dojka et al (2000) using primers 8f and 1492r (Richardson et al, 2002). Triplicate PCR products were pooled, purified by gel electrophoresis, and extracted using a MinElute Gel Extraction Kit (QIAGEN, Inc., Valencia, CA).…”

Sustainable syntrophic growth of Dehalococcoides ethenogenes strain 195 with Desulfovibrio vulgaris Hildenborough and Methanobacterium congolense: global transcriptomic and proteomic analyses

“…(Richardson et al, 2002;Ritalahti and Löffler, 2004;Duhamel and Edwards, 2006;Lee et al, 2006), which are able to ferment organic substrates into hydrogen and acetate. Studies of a previously described dechlorinating microbial community, ANAS, showed that it was dominated by three bacterial groups: Dehalococcoides, Desulfovibrio and Clostridium, with the hydrogenotrophic methanogen Methanobacterium congolense (MC) as the dominant archaeal species (Richardson et al, 2002). Previous studies have shown that Desulfovibrio vulgaris Hildenborough (DVH) can transform lactate to acetate and H 2 when grown syntrophically with a H 2 -utilizing methanogen (McInerney and Bryant, 1981;Schink, 1997).…”