COMBINED ISOTOPIC AND MOLECULAR EVIDENCE FOR DEVELOPMENT OF HIGHLY HETEROGENEOUS ZONES OF MICROBIAL ACTIVITY IN A GROUNDWATER UNDERGOING ENHANCED BIOREMEDIATION

For more than 8 years, electron donors (initially Na-lactate, followed by whey) have been added to the groundwater at the source of a 3 km long trichloroethene (TCE) plume at the Test Area North (TAN) site of the Idaho National Laboratory. The electron donors have led to high levels of microbial activity in the groundwater, resulting in anaerobic conditions that have successfully stimulated complete reductive dechlorination of the TCE in the source area of the plume. Anaerobic conditions in the groundwater extend to greater than 150 m down-gradient from the injection well. Within this zone, concentrations of dissolved methane are high (>3 micromolar), whereas nitrate and sulfate concentrations are depleted. However, surveys of the microbial communities in groundwater samples from the anaerobic zone of the plume using high density DNA microarrays identified a range of aerobic microorganisms including methane-oxidizing bacteria in addition to anaerobic organisms such as methanogens and sulfate reducers. Large shifts in the carbon isotope ratios of the methane (from -55‰ in the source area to -28‰ in down-gradient wells) confirm that significant microbial methane oxidation is occurring within the largely anaerobic core of the TCE plume. In addition, the isotope compositions of dissolved inorganic carbon (DIC) decreased from values typical of methanogenic activity (> 8‰) in the source area to much lower values (< -13‰) in the down-gradient anaerobic wells, as would be expected for high levels of methane oxidation. Conversely, down-gradient from the anaerobic core of the plume where the dissolved oxygen contents of the groundwater have rebounded, there is molecular evidence for diverse methanogenic communities. In addition, sulfate reducing and metal reducing microorganisms were identified in the aerobic groundwater. These data indicate that diverse microbial communities exist in the groundwater and that bulk measurements of the groundwater geochemistry may not reflect the geochemical conditions that would be required for some of these communities to be active. Heterogeneous groundwater flow paths coupled with microbial micro-sites (e.g., biofilms in lower permeability zones) are potential explanations for these findings.