Effects of varying growth conditions on stable carbon isotope fractionation of trichloroethene (TCE) by tceA-containing Dehalococcoides mccartyi strains.

To quantify in situ bioremediation using compound specific isotope analysis (CSIA), isotope fractionation data obtained from the field is interpreted according to laboratory-derived enrichment factors. Although previous studies that have quantified dynamic isotopic shifts during the reductive dechlorination of trichloroethene (TCE) indicate that fractionation factors can be highly variable from culture-to-culture and site-to-site, the effects of growth condition on the isotope fractionation during reductive dechlorination have not been previously examined. Here, carbon isotope fractionation by Dehalococcoides mccartyi 195 (Dhc195) maintained under a variety of growth conditions was examined. Enrichment factors quantified when Dhc195 was subjected to four suboptimal growth conditions, including decreased temperature (-13.3 ± 0.9‰), trace vitamin B12 availability (-12.7 ± 1.0‰), limited fixed nitrogen (-14.4 ± 0.8‰), and elevated vinyl chloride exposure (-12.5 ± 0.4‰), indicate that the fractionation is similar across a range of tested conditions. The TCE enrichment factors for two syntrophic cocultures, Dhc195 with Desulfovibrio vulgaris Hildenborough (-13.0 ± 2.0‰) and Dhc195 with Syntrophomonas wolfei (-10.4 ± 1.2‰ and -13.3 ± 1.0‰), were also similar to a control experiment. In order to test the stability of enrichment factors in microbial communities, the isotope fractionation was quantified for Dhc-containing groundwater communities before and after two-year enrichment periods under different growth conditions. Although these enrichment factors (-8.9 ± 0.4‰, -6.8 ± 0.8‰, -8.7 ± 1.3‰, -9.4 ± 0.7‰, and -7.2 ± 0.3‰) were predominantly outside the range of values quantified for the isolate and cocultures, all tested enrichment conditions within the communities produced nearly similar fractionations. Enrichment factors were not significantly affected by changes in any of the tested growth conditions for the pure cultures, cocultures or the mixed communities, indicating that despite a variety of temperature, nutrient, and cofactor-limiting conditions, stable carbon isotope fractionations remain consistent for given Dehalococcoides cultures.

Characterization of four TCE-dechlorinating microbial enrichments grown with different cobalamin stress and methanogenic conditions.

To investigate the important supportive microorganisms responsible for trichloroethene (TCE) bioremediation under specific environmental conditions and their relationship with Dehalococcoides (Dhc), four stable and robust enrichment cultures were generated using contaminated groundwater. Enrichments were maintained under four different conditions exploring two parameters: high and low TCE amendments (resulting in inhibited and uninhibited methanogenic activity, respectively) and with and without vitamin B12 amendment. Lactate was supplied as the electron donor. All enrichments were capable of reductively dechlorinating TCE to vinyl chloride and ethene. The dechlorination rate and ethene generation were higher, and the proportion of electrons used for dechlorination increased when methanogenesis was inhibited. Biologically significant cobalamin biosynthesis was detected in the enrichments without B12 amendment. Comparative genomics using a genus-wide microarray revealed a Dhc genome similar to that of strain 195 in all enrichments, a strain that lacks the major upstream corrin ring biosynthesis pathway. Seven other bacterial operational taxonomic units (OTUs) were detected using clone libraries. OTUs closest to Pelosinus, Dendrosporobacter, and Sporotalea (PDS) were most dominant. The Clostridium-like OTU was most affected by B12 amendment and active methanogenesis. Principal component analysis revealed that active methanogenesis, rather than vitamin B12 limitation, exerted a greater effect on the community structures even though methanogens did not seem to play an essential role in providing corrinoids to Dhc. In contrast, acetogenic bacteria that were abundant in the enrichments, such as PDS and Clostridium sp., may be potential corrinoid providers for Dhc.