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Microbial consortia development and microcosm and column experiments for enhanced bioremediation of chlorinated volatile organic compounds, West Branch Canal Creek wetland area, Aberdeen Proving Ground, Maryland

April 4, 2008

Chlorinated solvents, including 1,1,2,2-tetrachloroethane, tetrachloroethene, trichloroethene, carbon tetrachloride, and chloroform, are reaching land surface in localized areas of focused ground-water discharge (seeps) in a wetland and tidal creek in the West Branch Canal Creek area, Aberdeen Proving Ground, Maryland. In cooperation with the U.S. Army Garrison, Aberdeen Proving Ground, Maryland, the U.S. Geological Survey is developing enhanced bioremediation methods that simulate the natural anaerobic degradation that occurs without intervention in non-seep areas of the wetland. A combination of natural attenuation and enhanced bioremediation could provide a remedy for the discharging ground-water plumes that would minimize disturbance to the sensitive wetland ecosystem. Biostimulation (addition of organic substrate or nutrients) and bioaugmentation (addition of microbial consortium), applied either by direct injection at depth in the wetland sediments or by construction of a permeable reactive mat at the seep surface, were tested as possible methods to enhance anaerobic degradation in the seep areas. For the first phase of developing enhanced bioremediation methods for the contaminant mixtures in the seeps, laboratory studies were conducted to develop a microbial consortium to degrade 1,1,2,2-tetrachloroethane and its chlorinated daughter products under anaerobic conditions, and to test biostimulation and bioaugmentation of wetland sediment and reactive mat matrices in microcosms. The individual components required for the direct injection and reactive mat methods were then combined in column experiments to test them under groundwater- flow rates and contaminant concentrations observed in the field. Results showed that both direct injection and the reactive mat are promising remediation methods, although the success of direct injection likely would depend on adequately distributing and maintaining organic substrate throughout the wetland sediment in the seep area.

For bioaugmentation, two mixed anaerobic cultures, named the 'West Branch Consortia' (WBC-1 and WBC-2), were developed by enrichment of wetland sediment collected from two contaminated sites in the study area where rapid and complete reductive dechlorination naturally occurs. WBC are capable of degrading 1,1,2,2-tetrachloroethane, 1,1,2-trichloroethane, 1,2-dichloroethane, tetrachloroethene, trichloroethene, cis- and trans-1,2-dichloroethene, and vinyl chloride to the non-chlorinated end-products ethene and ethane. In addition, the column experiments showed that the consortia could completely degrade carbon tetrachloride and chloroform, although they were not grown on these contaminants. No other cultures are known that can degrade the broad mixture of chlorinated alkanes, alkenes, and methanes as shown for WBC. WBC-2 (suspended in the culture media) is capable of complete dechlorination of 50 micromolar 1,1,2,2-tetrachloroethane to ethene in 1 to 2 days with little transient accumulation of chlorinated daughter products. Only about 5 percent of the clones sequenced from WBC-1 and WBC-2 were related to dechlorinating bacteria that have been studied previously in culture, indicating the presence of unknown dechlorinators. Dehalococcoides spp. comprised about 1 percent of WBC-1 and WBC-2, which is minor compared to the population size of about 30 percent in other dechlorinating consortia for chlorinated alkenes. Although both WBC-1 and WBC-2 showed efficient degradation in laboratory tests in this study, long-term cultivation of WBC-1, which was developed using hydrogen as the organic substrate, was determined to be infeasible. Thus, WBC-2, cultivated with lactate as the organic substrate, would be used in future tests.

Nutrient (ammonia and phosphate mixture) addition to anaerobic microcosms constructed with wetland sediment and ground water collected from the study area showed some enhancement in the degradation rate of 1,1,2,2-tetrachloroethane, but degrada