The subsurface is a system comprising organic, inorganic, and microbiological components, which function together to support processes including nutrient cycling, carbon sequestration, and water filtration. Past research focuses on understanding the interplay of these dynamic microbiological, geochemical, and physical processes. Contamination of the subsurface due to urban and industrial activities represents a serious environmental risk, requiring the development of efficient remediation technologies that can restore these sites into functional areas. However, the presence and treatment of these contaminants also offer a scientifically advantageous system for investigation of more fundamental topics of microbial ecology and soil geochemistry. This dissertation investigates in situ chemical oxidation (ISCO) and in situ bioremediation (ISB) of organic contaminants, with a primary focus on illuminating the dynamic microbiological and geochemical processes that occur during remediation (Chapter 1).
Strong oxidants applied during ISCO significantly change contaminant and soil characteristics, and influence microbial community size, structure, and function. When properly applied, ISCO can be a beneficial pretreatment prior to ISB. A review of previous research indicates that investigations focused on (1) optimizing the chemical oxidant treatment to aid a subsequent biodegradation step, and (2) determining amendments required to support bioremediation following ISCO (Chapter 2). In some cases, full bioremediation is inhibited either by bioavailability of the contaminant or oxidation-reduction potential of the subsurface, both of which can be ameliorated with ISCO treatments. A survey of bioremediation efficiency is performed using multiple diesel-contaminated soils comprising four soil matrix types (Chapter 3). By incubating soils under optimized biodegradation conditions, performing molecular analysis of microbial biodegradation capacity based on an alkane monooxygenase gene (alkB), and monitoring diesel bioavailability, it is demonstrated that full biodegradation is most likely inhibited by bioavailability rather than nutrient abundance, electron acceptor availability, or microbial capacity. Additionally, analysis of microbial community composition based on pyrosequencing of the 16S rRNA gene fragment is performed to better understand the interplay among soil geochemical parameters, diesel characteristics, and microbial diversity at a contaminated location (Chapter 4). Results reveal a significant reduction in diversity in contaminated samples, and a strong association between the presence of diesel and the abundance of strict anaerobes, indicating that oxidation-reduction potential forms a notable hurdle for natural attenuation. Together, these sections provide convincing support for the importance of applying ISCO prior to ISB.
The coupling of chemical oxidation with aerobic bioremediation of diesel-contaminated soil is investigated in both laboratory and field experiments. Treatment with Fenton’s reagent and modified Fenton’s reagent focuses on understanding the geochemical changes occurring due to chemical oxidation that impact a subsequent bioremediation step (Chapter 5). Chemical oxidation degrades soil organic matter, thereby releasing dissolved nutrients and organic carbon, the latter of which competes with diesel as a substrate during biodegradation. Changes in microbial community structure and diesel degradation capacity are investigated during bioremediation following treatment with Fenton’s reagent, modified Fenton’s reagent, permanganate, and persulfate in two different diesel-contaminated soils (Chapter 6). Overall remediation efficiencies are dependent on the soil type; mobilization of diesel due to chemical oxidation causing reduced remediation efficiencies in peat, as compared to fill, where limited contaminant mobilization is observed. DGGE fingerprinting indicates the development of dissimilar microbial communities between treatments, with quantification of 16S rRNA and alkB gene abundance showing enrichment in biodegradation capacity in chemically treated soils, as compared to the biotic control. Finally, microbiological and geochemical dynamics of ISCO with persulfate followed by a nutrient-stimulated ISB are investigated in a diesel contaminated field location (Chapter 7). Groundwater parameters such as pH and oxidation-reduction potential indicate inhospitable conditions for microbial growth following ISCO, which is reflected in a 1-2 orders of magnitude decrease in both microbial community size (16S rRNA gene) and biodegradation capacity (alkB). While there is attenuation of acidic groundwater and regeneration of the microbial community, alterations to soil organic matter structure and oxidation of metal sulfides are irreversible changes to soil components.
The coupling of a chemical oxidation step with anaerobic bioremediation for the treatment of chlorinated solvents is investigated in laboratory and field experiments. Organohalide respiration (OHR) rates are researched in liquid microcosm experiments with permanganate treatment of tetrachloroethene (PCE; Chapter 8). Results indicate that mild permanganate treatments appear to slightly stimulate biodegradation, as determined by OHR rates, while strong permanganate doses disrupt PCE degradation to vinyl chloride and cause a 2-4 orders of magnitude decrease in the abundance of organohalide respiring bacteria (OHRB) and reductive dehalogenase genes (rdh). Microbial community composition based on pyrosequencing of 16S rRNA gene fragments indicates that chemical oxidation appears to select for Deltaproteobacteria, especially Geobacter, and Epsilonproteobacteria, especially Sulfurospirillum. The abundance of OHRB and rdh gene abundance is also followed during ISCO treatment with persulfate at a PCE and trichloroethene (TCE) contaminated location (Chapter 9). While natural attenuation capacity is measured prior to treatment, ISCO caused a significant drop in the abundance of OHRB and rdh genes, most likely due to groundwater acidification (pH<3) and high oxidation-reduction potential (>500 mV). Though regeneration of the OHRB community is observed six months after ISCO, Dehalococcoides mccartyi and rdh genes are absent, indicating a long-term disruption of full microbial natural attenuation capacity.
The outcomes of this dissertation provide insight into the dynamics of subsurface microbiological and geochemical processes (Chapter 10). The value of coupling ISCO with ISB is supported by the improved remediation efficiencies achieved when chemical oxidation is applied rather than bioremediation alone. Studies with chemical treatment illuminate microbial resilience, as microbes are not only able to regenerate following harsh oxidation, but in many cases a slight enrichment in biodegradation capacity is also observed. Chemical oxidation causes changes to subsurface geochemistry, including organic matter structure, mineralogy, and groundwater composition, which can have irreversible implications on microbial community structure and function. Therefore, molecular tools are required in order to properly illuminate the impact of contamination and remediation on microbial communities. Such knowledge on the resilience and sensitivity of soil and groundwater ecosystems to stress conditionsthus furthers the field of microbial ecology. Properly executed research on these subsurface systems will go beyond contributing to the field of soil remediation to provide a meaningful exploration of essential microbiological and geochemical processes in the environment. The results of this dissertation are an important new step towards examining contaminated locations and remediation from a fundamental scientific perspective.