Many uranium (U)-contaminated U.S. Department of Energy (DOE) sites are difficult to remediate due to the presence of high nitrate levels and low pH conditions. A combined aboveground/in situ approach is currently being used to remediate one of these sites at the DOE NABIR-Field Research Center in Oak Ridge, TN. An aboveground, anaerobic, fluidized bed reactor (FBR) is initially used to denitrify the groundwater following pH adjustment and pre-treatment. This water is then injected into the subsurface along with ethanol for biostimulation and ultimately U(VI) reduction and precipitation as insoluble U(IV). Most-probable-number analysis was used to enumerate denitrifying, sulfate-reducing, and iron-reducing microorganisms during biostimulation. A functional gene microarray (FGA) containing ~10,000 probes for genes involved in several biogeochemical processes was used to analyze microbial population dynamics in groundwater samples. Nitrate levels decreased following ethanol injection. Groundwater U levels began to decrease within two months after initiation of ethanol injection and eventually fell below 0.1 µM from an original level of ~200 µM. Sulfate consumption and sulfide formation were observed along with U removal. Numbers of denitrifying and sulfate-reducing microorganisms in the groundwater increased to around 105 cells/ml following biostimulation while iron-reducing microorganisms increased to around 103 cells/ml. Changes in denitrifier and sulfate-reducer populations, as indicated by FGA analyses, were associated with decreased nitrate and sulfate levels following biostimulation. These populations also correlated with U removal along with levels of several cytochrome C-type genes including those similar to U-reducing Geobacter spp. These results demonstrate the efficacy of field-scale biostimulation for U(VI) reduction and suggest that specific microbial populations played an integral role in the remediation.
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