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Item Understanding hydrogeomorphic influences on stream network denitrification and temperature dynamics(Montana State University - Bozeman, College of Agriculture, 2020) Carlson, Samuel Paul; Chairperson, Graduate Committee: Geoffrey PooleThe removal of nitrate from stream networks through the process of denitrification is an important component of local and regional nutrient cycles, but the controls on stream network denitrification rates remain poorly understood. Previous work has demonstrated general effects of stream channel size and nitrate loading rates on network-scale denitrification rates, but has been unable to elucidate connections between the complex environmental template of streams, and resulting denitrification rates. Understanding links between land use and management practices, physical characteristics of streams, and stream denitrification rates is critical to interpreting observed patterns of nitrate in freshwater systems and forming holistic management strategies for reducing the negative effects of elevated nitrate concentrations. To address these critical uncertainties, I developed a stream network simulation model that incorporates the effects of whole-stream aerobic respiration on biotic denitrification demand. This model is applied to a small, subalpine stream network under scenarios designed to explore: 1) the implications of temperature-controlled, network scale patterns of respiration rates on the distribution and overall magnitude of stream network denitrification, and 2) the effect of logging-induced channel simplification on whole network denitrification rates. The first analysis is complimented by an evaluation of controls on stream temperature across this network, revealing the spatially and temporally variable influence of in-network lakes on stream temperatures. Results from the first analysis suggest that reach- and network-scale denitrification rates are strongly influenced by respiration rate and temperature when nitrate supplies are high relative to removal rates, indicating an increased contribution of lower, warmer streams to whole-network denitrification. The second analysis reveals that historical logging has caused a ~30% loss of stream network denitrification capacity, which is manifested as a corresponding reduction in whole-network denitrification rates when nitrate supplies are not limiting. In sum, this work emphasizes the diverse set of factors that influence reach- and watershed-scale biogeochemical characteristics and processes, and suggests that land management actions which influence stream morphology may also alter stream denitrification rates.Item Influence of saturation on denitrification in a two-stage, vertical flow treatment wetland at Bridger Bowl ski area(Montana State University - Bozeman, College of Engineering, 2018) Woodhouse, Shayla Lee; Chairperson, Graduate Committee: Otto SteinA full-scale two-stage vertical flow treatment wetland (VF TW) pilot system was installed at the Bridger Bowl Ski Area in 2013 to test its capability as a secondary wastewater treatment system. Water quality was monitored throughout the 2015-2016 and 2016-2017 ski seasons with the primary objective to optimize the system for total nitrogen removal. Eight different combinations (schemes) of daily hydraulic and nutrient loading, dose frequency, effluent recycle, and depth of saturation of the first stage were tested. Average system removal was 93% and 95% for chemical oxygen demand (COD) and 70% and 75% for total nitrogen (TN) in 2016 and 2017, respectively, despite elevated influent concentrations of 930 mg x L -1 COD and 195 mg x L -1 TN. In addition, the system converted virtually all influent TN to nitrate. Both 1:1 and 2:1 (recycle to influent) ratios were effective as were saturation depths of 53 and 71 cm. At the 2:1 recycle ratio, the higher saturation level was superior at light hydraulic and constituent mass loadings while the lower saturation level was superior at higher loadings, but the differences were small. Overall, TN removal and nitrogen species transformations were linearly related to the mass load applied (surface area basis) over the range evaluated. In addition, the maximum removal capacity of the system was not exceeded during any scheme, thus the inherent removal capacity of the system is greater than evaluated. Nevertheless, the removal efficiencies of the VF TW from the past two seasons has shown that this technology can successfully perform secondary wastewater treatment in cold climates with efficient nitrogen removal and can exceed the regulatory requirements under which Bridger Bowl operates.Item Denitrification at the microscale in treatment wetlands(Montana State University - Bozeman, College of Engineering, 2015) Spengler, Justin Warren; Chairperson, Graduate Committee: Robin Gerlach; Anne Camper (co-chair)Treatment wetlands (TWs) have been in use for over three decades for wastewater treatment, agricultural water treatment, and some industrial wastes. Thousands of TWs exist for treating wastewater globally, but the microbial processes and controls in situ primarily responsible for water treatment are poorly understood. In this study, 16 separate model TW columns consisting of three plant groups and one non-planted group were fed synthetic post-secondary wastewater with half receiving no added carbon and half receiving 0.391 g L -1 as sucrose. Core samples were taken from each of the TW columns and separated into three distinct habitats (roots, gravel, particulates). Each habitat was assayed for its ability to produce N 2O, consume N 2O, and emit N 2O, as well as for denitrification gene abundances (nirS, nirK, and nosZ) and bacterial gene abundance (16S rDNA). The addition of organic carbon to the wetland was found to increase denitrification activity and gene copy abundance in non-root fractions, but organic carbon addition did not affect the root fraction. Plant presence within the TW was found to increase gas assay and gene abundance values in non-root habitats. Differences between three plant species were minor compared to differences attributed to carbon addition and plant presence. Of all habitats, gravel was found to have the highest denitrification activity and denitrification gene copy abundance relative to the number of 16S rDNA copies, as well as the highest ratio of N 2O produced to N 2O emitted. Implications for this study suggest the gravel and root fractions should be studied in further detail for their ability to accommodate denitrifying microbes.Item The role of fixX in electron bifurcation(Montana State University - Bozeman, College of Letters & Science, 2016) Miller, Jacquelyn Marie; Chairperson, Graduate Committee: John W. PetersTwo known methods of physiological energy conservation are substrate level phosphorylation and electron transfer phosphorylation. Recently, electron bifurcation has been established as a third and key mechanism of energy conservation in biological processes. This coupling of endergonic and exergonic reactions allows for utilization of reducing potential to perform energetically expensive physiological reactions. A significant and energetically expensive physiological reaction is nitrogen fixation, which provides a substantial portion of the bioavailable nitrogen that life requires. Electron bifurcation is utilized by the FixABCX system that is up regulated during diazotrophic growth and is suggested to bifurcate electrons from NADH to quinone of the electron transport chain through high potential electron transfer proteins and to nitrogenase though low potential electron transfer proteins. The determination of how cellular mechanisms overcome the energy barriers of high potential electron transfers through electron bifurcation is crucial for our fundamental understanding of energy transfer and energy conservation. The work presented in this thesis aims to progress the present knowledge in this third mechanism of energy conservation and shows support for a protein in the FixABCX complex, FixX, as the low potential electron acceptor in the complex. Numerous organisms were investigated as potential model systems for FixABCX with varying degrees of success. The genome of the organism, Roseiflexus castenholzii, contains both the nitrogenase and fixABCX genes and has successfully been used to obtain FixX. This protein shows homology to ferredoxin, a physiological reductant of the nitrogenase Fe protein in some organisms. EPR spectroscopy and sequence analysis suggests FixX contains 2 [4Fe-4S] clusters, while a potentiometric titration shows the clusters to have highly negative mid-point potentials. The preliminary evidence supports FixX of the FixABCX system to be a low potential electron transfer protein.Item Nitrogen removal and associated greenhouse gas production in laboratory-scale treatment wetlands(Montana State University - Bozeman, College of Engineering, 2016) Allen, Christopher Robert; Chairperson, Graduate Committee: Otto Stein; Mark D. Burr, Anne K. Camper, Jefferson J. Moss and Otto R. Stein were co-authors of the article, 'Plant influence on denitrification in treatment wetlands' submitted to the journal 'Water research' which is contained within this thesis.; Mark D. Burr, Anne K. Camper, Jefferson J. Moss and Otto R. Stein were co-authors of the article, 'Influence of plants and organic carbon addition on greenhouse gas emissions from model treatment wetlands' submitted to the journal 'Environmental science and technology' which is contained within this thesis.; Otto R. Stein was a co-author of the article, 'Empirical modeling of plant influence and organic carbon addition on nitrogen removal in treatment wetlands' submitted to the journal 'Ecological engineering' which is contained within this thesis.Treatment wetlands (TWs) are designed to treat domestic wastewater and water polluted from non-point sources such as agricultural runoff. Because many recent design improvements have increased aerobic removal pathways, nearly complete removal of biological oxygen demand (BOD) and oxidation of ammonium to nitrate in domestic wastewater is possible. These improvements have come at the expense of reducing the TW capacity to remove nitrate. Nitrate is also a main pollutant of concern in many non-point pollution sources. Organic carbon (OC) is a limiting factor for microbial nitrate removal and in wetlands can be supplied externally or provided by plants. Nitrate removal has the potential to release greenhouse gases (GHG) resulting in TWs being capable of acting as a net source or sink for GHG. Increasing our understanding of nitrogen removal and GHG production in TWs is the overarching goal of this project. A multi-year controlled environment greenhouse study measured water quality within 15-day incubations over annual cycles of temperature as well as greenhouse gas production at the seasonal extremes of the annual cycle. The experiment consisted of microcosms planted with either Carex utriculata, Deschampsia cespitosa, Phragmites australis, Schoenoplectus acutus, Typha latifolia or left unplanted. The fully factorial experiment also included three levels of OC addition, ranging from zero to two times the stoichiometric equivalent required for complete nitrogen removal. Nitrogen removal was affected by all experimental factors; plant species, OC addition, and temperature, with plant species mediating the effects of carbon and temperature in some treatments. The three highest performing species, C. utriculata, P. australis and S. acutus, removed nitrogen at an annual rate exceeding 166 g m -2 yr -1, without OC; only C. utriculata showed less N removal in winter. Incubation time series analysis indicated greater total and seasonal removal capacity for these plant treatments. Total GHG emission was dominated by summer CO 2 emission and varied by plant treatment and carbon load. CO 2 emission correlated negatively with OC addition in the high performing species attributable to plant biomass that decreased with OC addition. N 2O production significantly increased with the addition of organic carbon and did not vary significantly by season.Item Ammonia volatilization from native grasslands and forests of southwestern Montana(Montana State University - Bozeman, College of Letters & Science, 1984) Aradottir, Asa LovisaItem Process optimization of an on-site wastewater treatment system for nitrogen removal(Montana State University - Bozeman, College of Engineering, 1997) Blicker, Brian RobertItem An electrochemically enhanced denitrification process using biofilms(Montana State University - Bozeman, College of Engineering, 1997) Tripathi, Vijay K.Item Purification and characterization of gene products of the nitrous oxide reductase gene cluster(Montana State University - Bozeman, College of Letters & Science, 2002) Henery, Shannon MichelleItem Nitrate reduction by denitrifying bacteria within a porous medium(Montana State University - Bozeman, College of Engineering, 1998) Begaye-Ibbotson, Evangeline M.Acid processing of uranium ore resulted in aquifer contamination (nitrate, sulfate, and uranium) of the aquifer associated with the Navajo Sandstone formation at Tuba City, Arizona. The objectives of this study focused on the use of bioprocesses to remediate this aquifer contamination. The bench-scale objective of this study was to evaluate the ability of an indigenous microbial consortium to bioremediate nitrate contamination. The reduction of nitrate typically results in the production of nitrite, which under most conditions is further reduced to dinitrogen gas or ammonia. However, under some conditions inhibitory concentrations of nitrite may accumulate. Sandstone-packed columns fed with aquifer-relevant concentrations of nitrate were used to assess denitrification rates by indigenous bacteria. To enumerate denitrifying consortia used in the column experiments, most probable number (MPN) techniques were used. Sandstone-packed column influent and effluent data for nitrate, nitrite, carbon substrate and biomass concentrations were collected over time. These data were used to assess nitrate reduction rates within a sandstone column. This research demonstrates that with indigenous bacteria with stable conditions nitrate is reduced to dinitrogen forming only minimum levels of nitrite which should not inhibit sulfate-reducing bacteria (SRBs). The results of these studies indicate that bacterial denitrification has good potential as a remediation strategy for nitrate-contaminated groundwater to levels below the established regulatory limits of 44 mg/L. Field tests applications are planned for the Department of Energy UMTRA site in Tuba City, Arizona, using an extensive grid of injection and pumping wells.