|Microfracture Surface Characterizations: Implications for In Situ Remedial Methods in Fractured Rock, Bedrock Bioremediation Center Final Report (EPA/600/R-05/121) June 2006
The Bedrock Bioremediation Center (BBC) at the University of New Hampshire specializes in multidisciplinary research on bioremediation of organically contaminated bedrock aquifers. The focus of its current work is a field research-based program conducted at Site 32 at the Pease International Tradeport (formerly Pease Air Force Base) in Portsmouth, New Hampshire.
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Site 32 contains a contaminant plume of trichloroethylene (TCE) and its transformation products dichloroethylene (DCE), and vinyl chloride (VC). These are the principal contaminants. The site is situated on a variable thickness upper sand layer overlying a marine clay layer overlying a variable thickness lower sand layer. These unconsolidated layers are situated over the Kittery Formation, a tightly folded, biotite-grade, partially metamorphosed sandstone and shale, which is crosscut by numerous porphryitic diabase dikes. The contaminant plume extends downward and laterally northeast approximately 0.5 kilometers via migration through weathered and competent bedrock. The ground water in the bedrock is predominately contaminated with cis-DCE (280–440 micrograms per liter [µg/L]) with some trans-DCE (26–48 µg/L), TCE (24–59 µg/L), and VC (8–22 µg/L). Since 1997, the overburden has been managed using a sheet-pile containment system coupled with pump and treat. The bedrock ground water zone was given a technical impracticability waiver.
The questions addressed by this portion of the project relate to possible relationships among microfracture networks in the bedrock, the surface geochemistry of these microfractures, and the ecology and metabolic activity of attached microbes relative to terminal electron accepting processes and TCE biodegradation. Questions include:
Eleven microfractures were studied. They were extracted from competent bedrock cores from two wells at Site 32 (BBC5 and BBC6) in order to characterize (with a variety of surface spectroscopic and microbial techniques) the relationship among microfracture surface geochemistry, the ecology, and the metabolic activity of attached microbial populations relative to terminal electron-accepting processes or to chlorinated solvent biodegradation. Results are relative to host rock and microfracture mineralogy and geochemistry, ground water geochemistry, microfracture microbiology, and terminal electron accepting processes.
A variety of spectroscopic techniques are needed to characterize the mineralogy and chemical speciation of the host rock and the minerals coating the microfracture surfaces:
Microfracture samples were taken at depths greater than 21.3 meters (70 feet) below ground and within the contaminant plume. Using MIP, the partially metamorphosed sandstones and shales were found to be very impermeable.
The host rock had three nominal pore throat sizes (131.1, 1.136, and 0.109 micrometers [µm]), a porosity of 0.8 percent, and a permeability of less than 1 microDarcy. The host rock mineralogy was typical of metasandstones and metashales (quartz, feldspar, white mica, chlorite, and biotite).
Carbonate minerals and quartz were the dominant microfracture surface precipitates. Oxidized and reduced iron species were identified on the microfracture surfaces with XPS, including siderite (FeCO3), pyrrhotite (FeS), wüstite (FeO), goethite (α-FeOOH), hematite (Fe2O3), aged hydrous ferric oxide (Fe2O3 • 1.57 H2O), limonite (Fe2O3 • nH2O), and magnetite (Fe3O4). Carbon functional groups characteristic of humic substances and aquatic natural organic matter (NOM) were also identified with XPS.
SIMS mass fragment fingerprints revealed chlorinated carbon fragments, which suggested that TCE or perhaps its transformation products were partitioned to the NOM on the microfracture surfaces. The level of spatial resolution of this technique was on the order of 10s of µm. Heterogeneity in mineral abundance on the microfracture surfaces was seen at that level.
Packer sampling techniques were used to collect ground water samples from packer intervals associated with some of the collected microfracture samples in boreholes BBC5 and BBC6. The water collected in the packer intervals (termed packer water) were characterized using various field and laboratory techniques to describe pH, alkalinity, dissolved gases (hydrogen and oxygen), and dissolved geochemical constituents. The analyses were then used to model and interpret (subject to limitations) the geochemistry of the packer waters with the thermodynamic equilibrium model Visual MINTEQ. Given the volume of the microfracture network relative to the open fracture system, the samples were expected to reflect more from the composition of the open fractures.
Packer waters were alkaline (131–190 milligrams per liter [mg/L] as CaCO3, pH 8.8 to 9.6), mildly reducing (Eh of -208 to 160 millivolts, dissolved oxygen of 0.4 to 2.5 mg/L), with low nonpurgeable dissolved organic carbon values (0.8 to 1.7 mg/L), and measurable iron (II) (0.1 mg/L) and iron (III) (0.02 to 0.3 mg/L). Hydrogen was present in a number of the BBC wells at the site (2.2–7.3 Newton meters [nM]). These levels are capable of supporting reductive dechlorination and are indicative of sulfate reduction as a dominant terminal electron accepting process; however, sulfate was the dominant anion in the packer sample water (110–120 mg/L), and no sulfide was detected.
In addition, no fixed nitrogen was detected. The packer waters were in apparent pseudo-equilibrium with many of the observed major mineral phases (carbonates and iron oxides) in the host rock and on the microfracture surfaces. Estimations of Eh using the Nernst equation and activities of Fe2+ and Fe3+ suggested that the dominant redox couple was iron (II)/iron (III). Estimated values were similar to those measured with a polished platinum inert redox probe and reference Ag/AgCl electrode.
The microbiology of the microfracture surfaces was investigated using SEM, transmission electron microscopy (TEM), and a number of molecular biology techniques. SEM of microfracture surfaces revealed occasional biopatches of attached microbes. The biopatches were located in small depressions, cracks, or crevices on the microfracture surfaces. The microbes were predominantly rod-shaped (1.0 µm in diameter by 2.0 µm in length). In some instances, the bacteria had possible extracellular polymeric substances associated with them. In other cases, the microbes appeared encased in a film of organic material or surface precipitate-like material.
TEM micrographs of soft calcite surface precipitate samples from one microfracture revealed more diverse prokaryotic morphologies (e.g., spirilla, stalked bacteria, filaments). In some cases, flagella and possible cell division septa may have been present. Many cells contained large clear organelles and small dark organelles. These may have been storage bodies. Amplification with specific primer sets of microfractures from borehole BBC5 showed the presence of both bacteria and Archaea (which includes methanogens) in all of the borehole BBC5 microfracture samples. Positive results were also observed for dehalorespirers (Dehalococcoides sp.), sulfate-reducing bacteria, and iron-reducing bacteria (specifically the Geobacteraceae). Denaturing gradient gel electrophoresis community profiles of the polymerase chain reaction-amplified bacterial 16S rDNA showed between 7 and 27 band, indicating significant population diversity of the microfracture surfaces. Dendograms showed that two of seven of the microfractures tested were similar.
All other samples showed significantly different banding patterns, indicating the bacterial communities on the fracture surfaces were, in most cases, compositionally unique. Microfracture porewater likely differed from packer water in composition as the microfracture network may have been more reducing than the open fracture system based on the presence of obligate anaerobes found on the microfracture surfaces.
It can be inferred that there are possible terminal electron-accepting processes occurring in the open fracture system and the microfracture networks. By virtue of its smaller volume, the microfracture network reduced communication with the open fracture system, and mass transfer limitations probably did not significantly contribute to the contaminant or biogeochemical signatures seen in the packer waters collected under fairly transmissive conditions for fractured bedrock at the site. In terms of identification of likely terminal electron accepting processes in the open fracture system, the H2 values observed for borehole BBC6 suggested sulfate reduction. However, high levels of sulfate and the nondetection of sulfide in the packer water samples suggested that sulfate reduction was not dominant; rather, iron (III) reduction might have been the dominant terminal electron accepting process. Iron was a dominant microfracture surface element. Both iron (II) and iron (III) candidate minerals were observed on the microfracture surfaces. The spatial prevalence of iron, as well as its situation in the top few nM of the microfracture surface, suggested that iron (III) was available for iron-reducing bacteria. The spectroscopic characterization of the microfracture surfaces points to iron (III) reduction as perhaps a dominant process in the microfracture network.
There was generally good agreement between SEM-EDAX, XRD, and XPS about identification of carbon, sulfur, and iron within the microfracture surface precipitates and on their surfaces. However, the observed population diversity cannot be related to the speciation of any of the three elements on the microfracture surfaces. The spatial heterogeneity of minerals was quite high on the microfracture surfaces. Mineral grain sizes were on the order of µm. While minerals may have been common to all observed microfracture surfaces, their relative spacing and proximity to each other and to surface topography were quite varied. It may be that the biopatches that were observed with SEM reflect more localized microbial population response to microfracture surface mineral speciation. The level of resolution of SEM, SEM-EDAX, and XPS, however, was not high enough to discern such spatial relationships, though such relations are likely.
The presence of transformation products of dehalorespiration, as well as hydrogen concentrations, supported the role of Dehalococcoides sp. in dehalorespiration in the microfracture networks under conditions where iron (III) reduction was strongly correlated to the presence of oxidized iron species on the microfracture surfaces. Other means of TCE biodegradation, including abiotic as well as aerobic and anaerobic respiratory and cometabolic processes, cannot be excluded.
The bulk of the data suggested that the microfracture networks supported diverse microbial communities. The communities differed spatially and were not similar to open fracture system planktonic population compositions. The adherent populations were patchy and associated with microfracture topography. Microbes were also found within the microfracture surface precipitates themselves, suggesting a more complex mineral-microbe spatial relationship. The dominant mineralogy on the microfracture surfaces (iron [II] and iron [III] oxides and carbonates) was related to the microbial metabolism of some of the identified isolates, notably iron reducers. However, other types, including obligate anaerobes, suggested that the microfracture network was perhaps more reducing than the open fracture system, perhaps particularly within the microfracture surface precipitate structure. Dehalococcoides sp. was a predominant component on the microfracture microbial population and suggested that reductive dechlorination was one principal process whereby TCE was transformed.
A number of follow-up activities are suggested. Methods to collect and characterize microfracture porewaters may help to better describe terminal electron accepting processes and may elaborate on real differences with packer sample composition. The relative absence of NOM in the system, as well as the concentration of NOM on microfracture surfaces, deserves further examination. Understanding NOM bioavailability on microfracture surfaces may help to explain the phylogenetic and metabolic diversity seen on the microfracture surfaces. Studies looking at partitioning of TCE and transformation products to partitioned NOM under controlled isotherm conditions may help to better describe partitioning with respect to microfracture surface organic carbon fractions, particularly if more sensitive SIMS methods (such as time of flight SIMS) are used.
Understanding the spatial proximity of adhering microbes of terminal electron accepting process activity to minerals necessary to that terminal electron accepting process may help to describe the heterogeneous nature of terminal electron accepting processes in the microfracture network and at the microscale within the formation. Determining the extent of the microfracture specific surface area relative to that of the open fracture network would help in determining the role of microfracture in terminal electron accepting process and biodegradative processes within contaminated bedrock aquifers. The role of mass transfer between the open fracture system and the microfracture network, as well as redox zonations that might develop relative to proximity to the open fractures might be subjected to mass transfer and reaction path modeling exercises.
Additional work defining the complex microbial communities, their metabolic interactions, and their possible syntrophy with respect to TCE degradation may help to explain observed accumulations of transformation products. Further, the expression of enzymatic activity relative to terminal electron accepting processes and TCE biodegradation would help determine the metabolic activity on microfracture surfaces and why these might differ from those occurring in the open fracture ground water.
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