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Item Microbially induced calcium carbonate precipitation: meso-scale optimization and micro-scale characterization(Montana State University - Bozeman, College of Engineering, 2020) Zambare, Neerja Milind; Chairperson, Graduate Committee: Robin Gerlach and Ellen G. Lauchnor (co-chair); Ellen Lauchnor and Robin Gerlach were co-authors of the article, 'Controlling the distribution of microbially precipitated calcium carbonate in radial flow environments' in the journal 'Environmental science and technology' which is contained within this dissertation.; Robin Gerlach and Ellen Lauchnor were co-authors of the article, 'Spatio-temporal dynamics of strontium partitioning with microbially induced calcium carbonate precipitation in porous media flow cells' submitted to the journal 'Environmental science & technology' which is contained within this dissertation.; Robin Gerlach and Ellen Lauchnor were co-authors of the article, 'Co-precipitation of strontium and barium' submitted to the journal 'Environmental science & technology' which is contained within this dissertation.; Nada Naser, Robin Gerlach and Connie Chang were co-authors of the article, 'Visualizing microbially induced mineral precipitation from single cells using drop-based microfluidics' submitted to the journal 'Nature methods' which is contained within this dissertation.Microorganisms have the potential to impact processes on a scale orders of magnitude larger than their size. For example, microbe-mineral interactions at the micro-scale can drive macro-scale processes such as rock formation and weathering. Many bioremediation technologies derive inspiration from microbial mineralization processes. Microbially induced calcium carbonate precipitation (MICP) can produce calcium carbonate (CaCO 3) precipitates which can be utilized as a biological cement to strengthen porous media by reducing fluid permeability in subsurface fractures or as an immobilization matrix to remove metal contaminants dissolved in groundwater. To make MICP a feasible and successful bioremediation technology in the world outside the lab, it is necessary to bridge the gap between the meso-scale research studies and macro-scale applications. This thesis focuses on such meso-scale studies but also contributes to bridging the gap in the other direction, i.e., meso-scale to micro-scale to gain a fundamental understanding of the cellular level processes behind MICP. The research presented here investigates two applications of MICP with a focus on controlling precipitate distribution and process efficiency in target environments. Subsurface precipitate distribution and metal partitioning during MICP were studied in novel reactive transport systems that mimic application-environment conditions. A radial flow reactor was used to study the spatial distribution of precipitates in conditions similar to subsurface injection well environments. The distribution and degree of metal partitioning during MICP was investigated in batch reactors and porous media flow cells to study kinetics and reactive transport effects on kinetics. In the radial flow environment, more precipitates formed away from the center injection zone. Results showed that longer reactant residence times and an equimolar ratio of calcium to urea were able to maximize precipitation efficiency. Metal partitioning could be maximized at low reactant flow rates and low metal concentrations. The novel flow cell set up used revealed a spatial decoupling between ureolysis and precipitation. A micro-scale investigation of the fundamental MICP process itself is presented wherein microbe-mineral interactions are observed at the cell level. A semi-correlative approach to investigating individual precipitates in microdroplets is presented, using a multitude of microscopy and microanalysis techniques. The presented studies capture MICP across a range of scales.Item Nuclear magnetic resonance studies of biofilm - porous media systems(Montana State University - Bozeman, College of Engineering, 2017) Kirkland, Catherine Mullinnix; Chairperson, Graduate Committee: Sarah L. Codd; Joseph D. Seymour (co-chair); Sarah L. Codd was a co-author of the article, 'Low-field borehole NMR applications in the near subsurface environment' submitted to the journal 'Vadose zone journal' which is contained within this thesis.; Randy Hiebert, Adrienne Phillips, Elliot Grunewald, David O. Walsh, Joseph D. Seymour and Sarah L. Codd were co-authors of the article, 'Biofilm detection in a model well-bore environment using low-field NMR' in the journal 'Groundwater monitoring and remediation' which is contained within this thesis.; Maria P. Herrling, Randy Hiebert, Andrew T. Bender, Elliot Grunewald, David O. Walsh and Sarah L. Codd were co-authors of the article, 'In-situ detection of subsurface biofilm using low-field NMR - a field study' in the journal 'Environmental science and technology' which is contained within this thesis.; Sam Zanetti, Elliot Grunewald, David O. Walsh, Sarah L. Codd and Adrienne J. Phillips were co-authors of the article, 'Detecting microbially-induced calcite precipitation (MICP) in a model well-bore using downhole low-field NMR' in the journal 'Environmental science and technology' which is contained within this thesis.; Jessica Weisbrodt, Catherine M. Kirkland, Nathan H. Williamson, Susanne Lackner, Sarah L. Codd, Joseph D. Seymour, Gisela Guthausen and Harald Horn were co-authors of the article, 'NMR investigation of water diffusion in different biofilm structures' submitted to the journal 'Biotechnology and bioengineering' which is contained within this thesis.Nuclear magnetic resonance (NMR) allows for in-situ non-invasive studies of opaque systems over a wide range of length and time scales, making the method uniquely suited to studies of biofilms and porous media. The research comprising this thesis uses NMR to explore biophysical, chemical, and transport properties within heterogeneous porous media systems at both a macro- and micro-scale. The macro-scale projects validate a low-field borehole NMR instrument to monitor field-scale environmental engineering applications like subsurface biofilms and microbially-induced calcite precipitation (MICP). Subsurface biofilms are central to bioremediation of chemical contaminants in soil and groundwater whereby micro-organisms degrade or sequester environmental pollutants like nitrate, hydrocarbons, chlorinated solvents and heavy metals. When composed of ureolytic microbes, subsurface biofilms can also induce calcite precipitation. MICP has engineering applications that include soil stabilization and subsurface barriers, as well as sealing of cap rocks and well-bore regions for carbon dioxide sequestration. To meet the design goals of these beneficial applications, subsurface biofilms and MICP must be monitored over space and time - a challenging task with traditional methods. The low-field borehole NMR tool recorded changes in the T 2 relaxation distribution where enhanced relaxation indicated biofilm accumulation in a sand bioreactor and in subsurface soil. Additionally, the tool was able to detect MICP in a sand bioreactor. The changed mineral surface of the sand lead to an increase in T 2 relaxation times. The complementary high-field NMR project investigated micro-scale internal structures and mass transport within biofilm granules used for wastewater treatment. Granular sludge, composed of spherical aggregates of biofilm grown without a carrier, is an innovative biological treatment method with the potential to vastly reduce the cost of wastewater treatment without sacrificing efficiency. Large gaps remain, however, in our understanding of the fundamental formation mechanisms and the factors that control granule activity and stability. Magnetic resonance imaging (MRI) identified heterogeneous internal structures within aerobic granular sludge where relaxation rates and diffusion coefficients vary. Ultimately, these results will help improve modeling for optimization of granular sludge wastewater treatment process design.Item Influence of biosurfactant and non-biosurfactant producing bacteria on phenanthrene removal from model soils(Montana State University - Bozeman, College of Engineering, 1999) Eyre, Julie AnnItem Alkaline hydrolysis of explosives(Montana State University - Bozeman, College of Engineering, 2010) VanEngelen, Catherine Elizabeth; Chairperson, Graduate Committee: Brent M. PeytonIn the United States, ammunitions testing and manufacturing facilities must transform unused explosives into non-hazardous materials for disposal. 2,4,6- trinitrotoluene (TNT) is an explosive that has been found as a soil and groundwater contaminant at numerous ammunitions testing sites. Unused quantities of nitrocellulose (NC), another explosive, have also been accumulating at ammunitions manufacturing facilities. Transformation of both TNT and NC to non-explosive compounds has been studied using either chemical or biological approaches, each with limited success. With respect to TNT, the use of alkaline hydrolysis (degradation at high pH) as a chemical treatment had been tested at room temperature (20°C) under conditions where the hydroxide concentration exceeded that of TNT (pH > 10). These high hydroxide conditions were not directly amenable to biological treatment of the hydrolysis products. This study found that alkaline hydrolysis was effective for complete degradation of TNT at elevated temperatures (60°C and 80°C) when the concentration of TNT was less than the hydroxide concentration (pH 9 and 10). The resulting solution, or hydrolysate, contained no TNT. This hydrolysate was used as the carbon and nitrogen source for an aerobic bacterial enrichment from the Bozeman wastewater treatment plant. With respect to NC, the back-log of accumulated NC necessitates a degradation method that will process high NC concentrations (200g/L). Alkaline hydrolysis at 60°C was used with very high hydroxide concentrations to rapidly degrade high concentrations of NC, producing high nitrate and nitrite concentrations. The NC hydrolysate was neutralized and spiked into a denitrifying culture which was able to reduce both nitrate and nitrite. The goal of this work was to develop a dual component chemical-biological system for complete degradation of the explosives TNT and NC, which was achieved using alkaline hydrolysis as the chemical component and bacterial wastewater treatment enrichments as the biological component.Item Reactive transport in biofouled and biomineralized porous media(Montana State University - Bozeman, College of Engineering, 2010) Schultz, Logan Nicholas; Chairperson, Graduate Committee: Robin GerlachThe geologic subsurface environment contains regions of high porosity where fluid flows both naturally and during engineered technologies such as carbon sequestration, enhanced oil recovery, and bioremediation of contaminants. In these porous media regions, microbes can have both desirable and undesirable effects on the hydrodynamics and fluid chemistry by inducing the formation of microbial aggregates, which can include extracellular polymeric substance and abiotic particles such as mineral precipitates. While not the focus of this research, these analyses are likewise applicable to biomatter control scenarios in filtration systems and other industrial reactors with a high surface area to volume ratio. Microbially-induced ureolytic calcium carbonate precipitation has been suggested as a means to mitigate leakage from geologic CO 2 sequestration sites and as a means to immobilize divalent contaminants such as strontium-90 in remediation scenarios. In this process, microbes hydrolyze urea, increasing the solution pH, generating carbonate ions, and ultimately shifting the saturation state of the fluid and leading to solid calcium carbonate (CaCO 3) formation in a calcium-rich environment. Experiments were conducted to assess the distribution and effects of biofouling and biomineralization in two-dimensional flat plate reactors with 1mm pore spaces simulating a tortuous porous media environment. In biomineralization experiments, calcium carbonate was formed under flow conditions, and strontium was effectively immobilized within the crystal lattice, suggesting the applicability of subsurface biotechnical applications utilizing this technology. Image, residence time distribution, and piezometer analyses of biofouling experiments quantified porosity and hydraulic conductivity reductions. Biofilms were grown under constant flow and head conditions and were shown to be more channeled and evenly distributed along the flow path in constant flow conditions. Biofilms were challenged with chlorinated bleach, which temporarily increased the hydraulic conductivity, yet failed to remove significant biofouling unless coupled with significant fluid shear. In situ methods utilizing stereo and confocal microscopy were developed to visualize and quantify the distribution of biomatter formation and analyze the biological environment at the surface of bio-induced minerals.