Mechanisms of electron acceptor utilization: implications for simulating anaerobic biodegradation.

Simulation of biodegradation reactions within a reactive transport framework requires information on mechanisms of terminal electron acceptor processes (TEAPs). In initial modeling efforts, TEAPs were approximated as occurring sequentially, with the highest energy-yielding electron acceptors (e.g. oxygen) consumed before those that yield less energy (e.g., sulfate). Within this framework in a steady state plume, sequential electron acceptor utilization would theoretically produce methane at an organic-rich source and Fe(II) further downgradient, resulting in a limited zone of Fe(II) and methane overlap. However, contaminant plumes often display much more extensive zones of overlapping Fe(II) and methane. The extensive overlap could be caused by several abiotic and biotic processes including vertical mixing of byproducts in long-screened monitoring wells, adsorption of Fe(II) onto aquifer solids, or microscale heterogeneity in Fe(III) concentrations. Alternatively, the overlap could be due to simultaneous utilization of terminal electron acceptors. Because biodegradation rates are controlled by TEAPs, evaluating the mechanisms of electron acceptor utilization is critical for improving prediction of contaminant mass losses due to biodegradation. Using BioRedox-MT3DMS, a three-dimensional, multi-species reactive transport code, we simulated the current configurations of a BTEX plume and TEAP zones at a petroleum-contaminated field site in Wisconsin. Simulation results suggest that BTEX mass loss due to biodegradation is greatest under oxygen-reducing conditions, with smaller but similar contributions to mass loss from biodegradation under Fe(III)-reducing, sulfate-reducing, and methanogenic conditions. Results of sensitivity calculations document that BTEX losses due to biodegradation are most sensitive to the age of the plume, while the shape of the BTEX plume is most sensitive to effective porosity and rate constants for biodegradation under Fe(III)-reducing and methanogenic conditions. Using this transport model, we had limited success in simulating overlap of redox products using reasonable ranges of parameters within a strictly sequential electron acceptor utilization framework. Simulation results indicate that overlap of redox products cannot be accurately simulated using the constructed model, suggesting either that Fe(III) reduction and methanogenesis are occurring simultaneously in the source area, or that heterogeneities in Fe(III) concentration and/or mineral type cause the observed overlap. Additional field, experimental, and modeling studies will be needed to address these questions.

Nitrate-enhanced bioremediation of BTEX-contaminated groundwater: parameter estimation from natural-gradient tracer experiments.

Two natural-gradient pulse tracer tests were conducted in a petroleum-contaminated aquifer to evaluate the potential for benzene, toluene, ethylbenzene, and xylenes (BTEX) biodegradation under enhanced nitrate-reducing conditions. Addition of nitrate resulted in loss of toluene, ethylbenzene, and m,p-xylenes (TEX) after an initial lag period of approximately 9 days. Losses of benzene were not observed over the 60-day monitoring period. Tracer breakthrough curves (BTCs) were analyzed to derive transport and biodegradation parameters, including advective velocities, retardation factors, dispersion coefficients, biodegradation rate constants, and nitrate utilization ratios. Using the parameters derived from the BTC analysis, numerical simulations of one of the tracer experiments were conducted using BIONAPL/3D [Molson, J., BIONAPL/3D User Guide, A 3D Coupled Flow and Multi-Component Reactive transport model. University of Waterloo, Waterloo, Ontario, Canada]. Simulations using the BTC-derived transport and biodegradation parameters successfully reproduced benzene, TEX, and nitrate concentrations measured during the tracer experiment. Comparisons of observed and simulated nitrate concentrations indicate that the mass ratio of nitrate-N utilized to TEX degraded increased over time during the experiment, reaching values many times that expected based on stoichiometry of TEX oxidation coupled to nitrate reduction. Excess nitrate loss is likely due to oxidation of other organics in addition to TEX.

Molecular analysis of microbial community structures in pristine and contaminated aquifers: field and laboratory microcosm experiments.

This study used phylogenetic probes in hybridization analysis to (i) determine in situ microbial community structures in regions of a shallow sand aquifer that were oxygen depleted and fuel contaminated (FC) or aerobic and noncontaminated (NC) and (ii) examine alterations in microbial community structures resulting from exposure to toluene and/or electron acceptor supplementation (nitrate). The latter objective was addressed by using the NC and FC aquifer materials for anaerobic microcosm studies in which phylogenetic probe analysis was complemented by microbial activity assays. Domain probe analysis of the aquifer samples showed that the communities were predominantly Bacteria; Eucarya and Archaea were not detectable. At the phylum and subclass levels, the FC and NC aquifer material had similar relative abundance distributions of 43 to 65% beta- and gamma-Proteobacteria (B+G), 31 to 35% alpha-Proteobacteria (ALF), 15 to 18% sulfate-reducing bacteria, and 5 to 10% high G+C gram positive bacteria. Compared to that of the NC region, the community structure of the FC material differed mainly in an increased abundance of B+G relative to that of ALF. The microcosm communities were like those of the field samples in that they were predominantly Bacteria (83 to 101%) and lacked detectable Archaea but differed in that a small fraction (2 to 8%) of Eucarya was detected regardless of the treatment applied. The latter result was hypothesized to reflect enrichment of anaerobic protozoa. Addition of nitrate and/or toluene stimulated microbial activity in the microcosms, but only supplementation of toluene alone significantly altered community structures. For the NC material, the dominant subclass shifted from B+G to ALF, while in the FC microcosms 55 to 65% of the Bacteria community was no longer identifiable by the phylum or subclass probes used. The latter result suggested that toluene exposure fostered the proliferation of phylotype(s) that were otherwise minor constituents of the FC aquifer community. These studies demonstrated that alterations in aquifer microbial communities resulting from specific anthropogenic perturbances can be inferred from microcosm studies integrating chemical and phylogenetic probe analysis and in the case of hydrocarbon contamination may facilitate the identification of organisms important for in situ biodegradation processes. Further work integrating and coordinating microcosm and field experiments is needed to explore how differences in scale, substrate complexity, and other hydrogeological conditions may affect patterns observed in these systems.