Matching Items (190)
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Description
This dissertation explores the use of bench-scale batch microcosms in remedial design of contaminated aquifers, presents an alternative methodology for conducting such treatability studies, and - from technical, economical, and social perspectives - examines real-world application of this new technology. In situ bioremediation (ISB) is an effective remedial approach for

This dissertation explores the use of bench-scale batch microcosms in remedial design of contaminated aquifers, presents an alternative methodology for conducting such treatability studies, and - from technical, economical, and social perspectives - examines real-world application of this new technology. In situ bioremediation (ISB) is an effective remedial approach for many contaminated groundwater sites. However, site-specific variability necessitates the performance of small-scale treatability studies prior to full-scale implementation. The most common methodology is the batch microcosm, whose potential limitations and suitable technical alternatives are explored in this thesis. In a critical literature review, I discuss how continuous-flow conditions stimulate microbial attachment and biofilm formation, and identify unique microbiological phenomena largely absent in batch bottles, yet potentially relevant to contaminant fate. Following up on this theoretical evaluation, I experimentally produce pyrosequencing data and perform beta diversity analysis to demonstrate that batch and continuous-flow (column) microcosms foster distinctly different microbial communities. Next, I introduce the In Situ Microcosm Array (ISMA), which took approximately two years to design, develop, build and iteratively improve. The ISMA can be deployed down-hole in groundwater monitoring wells of contaminated aquifers for the purpose of autonomously conducting multiple parallel continuous-flow treatability experiments. The ISMA stores all sample generated in the course of each experiment, thereby preventing the release of chemicals into the environment. Detailed results are presented from an ISMA demonstration evaluating ISB for the treatment of hexavalent chromium and trichloroethene. In a technical and economical comparison to batch microcosms, I demonstrate the ISMA is both effective in informing remedial design decisions and cost-competitive. Finally, I report on a participatory technology assessment (pTA) workshop attended by diverse stakeholders of the Phoenix 52nd Street Superfund Site evaluating the ISMA's ability for addressing a real-world problem. In addition to receiving valuable feedback on perceived ISMA limitations, I conclude from the workshop that pTA can facilitate mutual learning even among entrenched stakeholders. In summary, my doctoral research (i) pinpointed limitations of current remedial design approaches, (ii) produced a novel alternative approach, and (iii) demonstrated the technical, economical and social value of this novel remedial design tool, i.e., the In Situ Microcosm Array technology.
ContributorsKalinowski, Tomasz (Author) / Halden, Rolf U. (Thesis advisor) / Johnson, Paul C (Committee member) / Krajmalnik-Brown, Rosa (Committee member) / Bennett, Ira (Committee member) / Arizona State University (Publisher)
Created2013
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Description
Reductive dechlorination by members of the bacterial genus Dehalococcoides is a common and cost-effective avenue for in situ bioremediation of sites contaminated with the chlorinated solvents, trichloroethene (TCE) and perchloroethene (PCE). The overarching goal of my research was to address some of the challenges associated with bioremediation timeframes by improving

Reductive dechlorination by members of the bacterial genus Dehalococcoides is a common and cost-effective avenue for in situ bioremediation of sites contaminated with the chlorinated solvents, trichloroethene (TCE) and perchloroethene (PCE). The overarching goal of my research was to address some of the challenges associated with bioremediation timeframes by improving the rates of reductive dechlorination and the growth of Dehalococcoides in mixed communities. Biostimulation of contaminated sites or microcosms with electron donor fails to consistently promote dechlorination of PCE/TCE beyond cis-dichloroethene (cis-DCE), even when the presence of Dehalococcoides is confirmed. Supported by data from microcosm experiments, I showed that the stalling at cis-DCE is due a H2 competition in which components of the soil or sediment serve as electron acceptors for competing microorganisms. However, once competition was minimized by providing selective enrichment techniques, I illustrated how to obtain both fast rates and high-density Dehalococcoides using three distinct enrichment cultures. Having achieved a heightened awareness of the fierce competition for electron donor, I then identified bicarbonate (HCO3-) as a potential H2 sink for reductive dechlorination. HCO3- is the natural buffer in groundwater but also the electron acceptor for hydrogenotrophic methanogens and homoacetogens, two microbial groups commonly encountered with Dehalococcoides. By testing a range of concentrations in batch experiments, I showed that methanogens are favored at low HCO3 and homoacetogens at high HCO3-. The high HCO3- concentrations increased the H2 demand which negatively affected the rates and extent of dechlorination. By applying the gained knowledge on microbial community management, I ran the first successful continuous stirred-tank reactor (CSTR) at a 3-d hydraulic retention time for cultivation of dechlorinating cultures. I demonstrated that using carefully selected conditions in a CSTR, cultivation of Dehalococcoides at short retention times is feasible, resulting in robust cultures capable of fast dechlorination. Lastly, I provide a systematic insight into the effect of high ammonia on communities involved in dechlorination of chloroethenes. This work documents the potential use of landfill leachate as a substrate for dechlorination and an increased tolerance of Dehalococcoides to high ammonia concentrations (2 g L-1 NH4+-N) without loss of the ability to dechlorinate TCE to ethene.
ContributorsDelgado, Anca Georgiana (Author) / Krajmalnik-Brown, Rosa (Thesis advisor) / Cadillo-Quiroz, Hinsby (Committee member) / Halden, Rolf U. (Committee member) / Rittmann, Bruce E. (Committee member) / Stout, Valerie (Committee member) / Arizona State University (Publisher)
Created2013
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Description
Synechocystis sp PCC 6803 is a photosynthetic cyanobacterium that can be easily transformed to produce molecules of interest; this has increased Synechocystis’ popularity as a clean energy platform. Synechocystis has been shown to produce and excrete molecules such as fatty acids, isoprene, etc. after appropriate genetic modification. Challenges faced for

Synechocystis sp PCC 6803 is a photosynthetic cyanobacterium that can be easily transformed to produce molecules of interest; this has increased Synechocystis’ popularity as a clean energy platform. Synechocystis has been shown to produce and excrete molecules such as fatty acids, isoprene, etc. after appropriate genetic modification. Challenges faced for large–scale growth of modified Synechocystis include abiotic stress, microbial contamination and high processing costs of product and cell material. Research reported in this dissertation contributes to solutions to these challenges. First, abiotic stress was addressed by overexpression of the heat shock protein ClpB1. In contrast to the wild type, the ClpB1 overexpression mutant (Slr1641+) tolerated rapid temperature changes, but no difference was found between the strains when temperature shifts were slower. Combination of ClpB1 overexpression with DnaK2 overexpression (Slr1641+/Sll0170+) further increased thermotolerance. Next, we used a Synechocystis strain that carries an introduced isoprene synthase gene (IspS+) and that therefore produces isoprene. We attempted to increase isoprene yields by overexpression of key enzymes in the methyl erythritol phosphate (MEP) pathway that leads to synthesis of the isoprene precursor. Isoprene production was not increased greatly by MEP pathway induction, likely because of limitations in the affinity of the isoprene synthase for the substrate. Finally, two extraction principles, two–phase liquid extraction (e.g., with an organic and aqueous phase) and solid–liquid extraction (e.g., with a resin) were tested. Two–phase liquid extraction is suitable for separating isoprene but not fatty acids from the culture medium. Fatty acid removal required acidification or surfactant addition, which affected biocompatibility. Therefore, improvements of both the organism and product–harvesting methods can contribute to enhancing the potential of cyanobacteria as solar–powered biocatalysts for the production of petroleum substitutes.
ContributorsGonzalez Esquer, Cesar Raul (Author) / Vermaas, Willem (Thesis advisor) / Chandler, Douglas (Committee member) / Bingham, Scott (Committee member) / Nielsen, David (Committee member) / Arizona State University (Publisher)
Created2013
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Description
Human activity has increased loading of reactive nitrogen (N) in the environment, with important and often deleterious impacts on biodiversity, climate, and human health. Since the fate of N in the ecosystem is mainly controlled by microorganisms, understanding the factors that shape microbial communities becomes relevant and urgent. In arid

Human activity has increased loading of reactive nitrogen (N) in the environment, with important and often deleterious impacts on biodiversity, climate, and human health. Since the fate of N in the ecosystem is mainly controlled by microorganisms, understanding the factors that shape microbial communities becomes relevant and urgent. In arid land soils, these microbial communities and factors are not well understood. I aimed to study the role of N cycling microbes, such as the ammonia-oxidizing bacteria (AOB), the recently discovered ammonia-oxidizing archaea (AOA), and various fungal groups, in soils of arid lands. I also tested if niche differentiation among microbial populations is a driver of differential biogeochemical outcomes. I found that N cycling microbial communities in arid lands are structured by environmental factors to a stronger degree than what is generally observed in mesic systems. For example, in biological soil crusts, temperature selected for AOA in warmer deserts and for AOB in colder deserts. Land-use change also affects niche differentiation, with fungi being the major agents of N2O production in natural arid lands, whereas emissions could be attributed to bacteria in mesic urban lawns. By contrast, NO3- production in the native desert and managed soils was mainly controlled by autotrophic microbes (i.e., AOB and AOA) rather than by heterotrophic fungi. I could also determine that AOA surprisingly responded positively to inorganic N availability in both short (one month) and long-term (seven years) experimental manipulations in an arid land soil, while environmental N enrichment in other ecosystem types is known to favor AOB over AOA. This work improves our predictions of ecosystem response to anthropogenic N increase and shows that paradigms derived from mesic systems are not always applicable to arid lands. My dissertation also highlights the unique ecology of ammonia oxidizers and draws attention to the importance of N cycling in desert soils.
ContributorsMarusenko, Yevgeniy (Author) / Hall, Sharon J (Thesis advisor) / Garcia-Pichel, Ferran (Thesis advisor) / Mclain, Jean E (Committee member) / Schwartz, Egbert (Committee member) / Arizona State University (Publisher)
Created2013
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Description
Microbial electrochemical cells (MXCs) are promising platforms for bioenergy production from renewable resources. In these systems, specialized anode-respiring bacteria (ARB) deliver electrons from oxidation of organic substrates to the anode of an MXC. While much progress has been made in understanding the microbiology, physiology, and electrochemistry of well-studied model ARB

Microbial electrochemical cells (MXCs) are promising platforms for bioenergy production from renewable resources. In these systems, specialized anode-respiring bacteria (ARB) deliver electrons from oxidation of organic substrates to the anode of an MXC. While much progress has been made in understanding the microbiology, physiology, and electrochemistry of well-studied model ARB such as Geobacter and Shewanella, tremendous potential exists for MXCs as microbiological platforms for exploring novel ARB. This dissertation introduces approaches for selective enrichment and characterization of phototrophic, halophilic, and alkaliphilic ARB. An enrichment scheme based on manipulation of poised anode potential, light, and nutrient availability led to current generation that responded negatively to light. Analysis of phototrophically enriched communities suggested essential roles for green sulfur bacteria and halophilic ARB in electricity generation. Reconstruction of light-responsive current generation could be successfully achieved using cocultures of anode-respiring Geobacter and phototrophic Chlorobium isolated from the MXC enrichments. Experiments lacking exogenously supplied organic electron donors indicated that Geobacter could produce a measurable current from stored photosynthate in the dark. Community analysis of phototrophic enrichments also identified members of the novel genus Geoalkalibacter as potential ARB. Electrochemical characterization of two haloalkaliphilic, non-phototrophic Geoalkalibacter spp. showed that these bacteria were in fact capable of producing high current densities (4-8 A/m2) and using higher organic substrates under saline or alkaline conditions. The success of these selective enrichment approaches and community analyses in identifying and understanding novel ARB capabilities invites further use of MXCs as robust platforms for fundamental microbiological investigations.
ContributorsBadalamenti, Jonathan P (Author) / Krajmalnik-Brown, Rosa (Thesis advisor) / Garcia-Pichel, Ferran (Committee member) / Rittmann, Bruce E. (Committee member) / Torres, César I (Committee member) / Vermaas, Willem (Committee member) / Arizona State University (Publisher)
Created2013
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Description
Random peptide microarrays are a powerful tool for both the treatment and diagnostics of infectious diseases. On the treatment side, selected random peptides on the microarray have either binding or lytic potency against certain pathogens cells, thus they can be synthesized into new antimicrobial agents, denoted as synbodies (synthetic antibodies).

Random peptide microarrays are a powerful tool for both the treatment and diagnostics of infectious diseases. On the treatment side, selected random peptides on the microarray have either binding or lytic potency against certain pathogens cells, thus they can be synthesized into new antimicrobial agents, denoted as synbodies (synthetic antibodies). On the diagnostic side, serum containing specific infection-related antibodies create unique and distinct "pathogen-immunosignatures" on the random peptide microarray distinct from the healthy control serum, and this different mode of binding can be used as a more precise measurement than traditional ELISA tests. My thesis project is separated into these two parts: the first part falls into the treatment side and the second one focuses on the diagnostic side. My first chapter shows that a substitution amino acid peptide library helps to improve the activity of a recently reported synthetic antimicrobial peptide selected by the random peptide microarray. By substituting one or two amino acids of the original lead peptide, the new substitutes show changed hemolytic effects against mouse red blood cells and changed potency against two pathogens: Staphylococcus aureus and Pseudomonas aeruginosa. Two new substitutes are then combined together to form the synbody, which shows a significantly antimicrobial potency against Staphylococcus aureus (<0.5uM). In the second chapter, I explore the possibility of using the 10K Ver.2 random peptide microarray to monitor the humoral immune response of dengue. Over 2.5 billion people (40% of the world's population) live in dengue transmitting areas. However, currently there is no efficient dengue treatment or vaccine. Here, with limited dengue patient serum samples, we show that the immunosignature has the potential to not only distinguish the dengue infection from non-infected people, but also the primary dengue infection from the secondary dengue infections, dengue infection from West Nile Virus (WNV) infection, and even between different dengue serotypes. By further bioinformatic analysis, we demonstrate that the significant peptides selected to distinguish dengue infected and normal samples may indicate the epitopes responsible for the immune response.
ContributorsWang, Xiao (Author) / Johnston, Stephen Albert (Thesis advisor) / Blattman, Joseph (Committee member) / Arntzen, Charles (Committee member) / Arizona State University (Publisher)
Created2013
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Description
To further the efforts producing energy from more renewable sources, microbial electrochemical cells (MXCs) can utilize anode respiring bacteria (ARB) to couple the oxidation of an organic substrate to the delivery of electrons to the anode. Although ARB such as Geobacter and Shewanella have been well-studied in terms of their

To further the efforts producing energy from more renewable sources, microbial electrochemical cells (MXCs) can utilize anode respiring bacteria (ARB) to couple the oxidation of an organic substrate to the delivery of electrons to the anode. Although ARB such as Geobacter and Shewanella have been well-studied in terms of their microbiology and electrochemistry, much is still unknown about the mechanism of electron transfer to the anode. To this end, this thesis seeks to elucidate the complexities of electron transfer existing in Geobacter sulfurreducens biofilms by employing Electrochemical Impedance Spectroscopy (EIS) as the tool of choice. Experiments measuring EIS resistances as a function of growth were used to uncover the potential gradients that emerge in biofilms as they grow and become thicker. While a better understanding of this model ARB is sought, electrochemical characterization of a halophile, Geoalkalibacter subterraneus (Glk. subterraneus), revealed that this organism can function as an ARB and produce seemingly high current densities while consuming different organic substrates, including acetate, butyrate, and glycerol. The importance of identifying and studying novel ARB for broader MXC applications was stressed in this thesis as a potential avenue for tackling some of human energy problems.
ContributorsAjulo, Oluyomi (Author) / Torres, Cesar (Thesis advisor) / Nielsen, David (Committee member) / Krajmalnik-Brown, Rosa (Committee member) / Popat, Sudeep (Committee member) / Arizona State University (Publisher)
Created2013
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Description
Microbial mat communities that inhabit hot springs in Yellowstone National Park have been studied for their biodiversity, energetics and evolutionary history, yet little is know about how these communities cope with nutrient limitation. In the present study the changes in assimilatory gene expression levels for nitrogen (nrgA), phosphorus (phoA), and

Microbial mat communities that inhabit hot springs in Yellowstone National Park have been studied for their biodiversity, energetics and evolutionary history, yet little is know about how these communities cope with nutrient limitation. In the present study the changes in assimilatory gene expression levels for nitrogen (nrgA), phosphorus (phoA), and iron (yusV) were measured under various nutrient enrichment experiments. While results for nrgA and phoA were inconclusive, results for yusV showed an increase in expression with the addition of N and Fe. This is the first data that shows the impact of nutrients on siderophore uptake regulation in hot spring microbes.
ContributorsThorne, Michele (Author) / Elser, James J (Thesis advisor) / Touchman, Jeffrey (Committee member) / Stout, Valerie (Committee member) / Arizona State University (Publisher)
Created2012
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Description
The prevalence of antibiotic resistant bacterial pathogens has increased since the introduction of penicillin in the 1940s. Insufficient development of novel antibacterial agents is leaving us with a failing arsenal of therapies to combat these pathogenic organisms. We have identified a clay mineral mixture (designated CB) that exhibits in vitro

The prevalence of antibiotic resistant bacterial pathogens has increased since the introduction of penicillin in the 1940s. Insufficient development of novel antibacterial agents is leaving us with a failing arsenal of therapies to combat these pathogenic organisms. We have identified a clay mineral mixture (designated CB) that exhibits in vitro antibacterial activity against a broad spectrum of bacterial pathogens, yet the antibacterial mechanism of action remains unknown. Antibacterial susceptibility testing of four different clay samples collected from the same source revealed that these natural clays had markedly different antibacterial activity. X-ray diffraction analyses of these minerals revealed minor mineralogical differences across the samples; however, ICP analyses demonstrated that the concentrations of many elements, Fe, Co, Cu, Ni, and Zn in particular, vary greatly across the four clay mixture leachates. Supplementation of a non-antibacterial leachate containing lower concentrations of Fe, Co, Ni, Cu, and Zn to final ion concentrations and a pH equivalent to that of the antibacterial leachate resulted in antibacterial activity against E. coli and MRSA, confirming the role of these ions in the in vitro antibacterial clay mixture leachates. The prevailing hypothesis is that metal ions participate in redox cycling and produce ROS, leading to oxidative damage to macromolecules and resulting in cellular death. However, E. coli cells showed no increase in DNA or protein oxidative lesions and a slight increase in lipid peroxidation following exposure to CB-L. Supplementation of CB-L with ROS scavengers eliminated oxidative damage in E. coli, but did not rescue the cells from killing, indicating that in vitro killing is due to direct metal toxicity and not to indirect oxidative damage. Finally, we ion-exchanged non-antibacterial clays with Fe, Co, Cu, and Zn and established antibacterial activity in these samples. Treatment of MRSA skin infections with both natural and ion-exchanged clays significantly decreased the bacterial load after 7 days of treatment. We conclude that 1) in vitro clay-mediated killing is due to toxicity associated directly with released metal ions and not to indirect oxidative damage and 2) that in vivo killing is due to the physical properties of the clays rather than metal ion toxicity.
ContributorsOtto, Caitin Carol (Author) / Haydel, Shelley (Thesis advisor) / Stout, Valerie (Committee member) / Roberson, Robby (Committee member) / Sandrin, Todd (Committee member) / Rege, Kaushal (Committee member) / Arizona State University (Publisher)
Created2014
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Description
This thesis research focuses on developing a single-cell gene expression analysis method for marine diatom Thalassiosira pseudonana and constructing a chip level tool to realize the single cell RT-qPCR analysis. This chip will serve as a conceptual foundation for future deployable ocean monitoring systems. T. pseudonana, which is a common

This thesis research focuses on developing a single-cell gene expression analysis method for marine diatom Thalassiosira pseudonana and constructing a chip level tool to realize the single cell RT-qPCR analysis. This chip will serve as a conceptual foundation for future deployable ocean monitoring systems. T. pseudonana, which is a common surface water microorganism, was detected in the deep ocean as confirmed by phylogenetic and microbial community functional studies. Six-fold copy number differences between 23S rRNA and 23S rDNA were observed by RT-qPCR, demonstrating the moderate functional activity of detected photosynthetic microbes in the deep ocean including T. pseudonana. Because of the ubiquity of T. pseudonana, it is a good candidate for an early warning system for ocean environmental perturbation monitoring. This early warning system will depend on identifying outlier gene expression at the single-cell level. An early warning system based on single-cell analysis is expected to detect environmental perturbations earlier than population level analysis which can only be observed after a whole community has reacted. Preliminary work using tube-based, two-step RT-qPCR revealed for the first time, gene expression heterogeneity of T. pseudonana under different nutrient conditions. Heterogeneity was revealed by different gene expression activity for individual cells under the same conditions. This single cell analysis showed a skewed, lognormal distribution and helped to find outlier cells. The results indicate that the geometric average becomes more important and representative of the whole population than the arithmetic average. This is in contrast with population level analysis which is limited to arithmetic averages only and highlights the value of single cell analysis. In order to develop a deployable sensor in the ocean, a chip level device was constructed. The chip contains surface-adhering droplets, defined by hydrophilic patterning, that serve as real-time PCR reaction chambers when they are immersed in oil. The chip had demonstrated sensitivities at the single cell level for both DNA and RNA. The successful rate of these chip-based reactions was around 85%. The sensitivity of the chip was equivalent to published microfluidic devices with complicated designs and protocols, but the production process of the chip was simple and the materials were all easily accessible in conventional environmental and/or biology laboratories. On-chip tests provided heterogeneity information about the whole population and were validated by comparing with conventional tube based methods and by p-values analysis. The power of chip-based single-cell analyses were mainly between 65-90% which were acceptable and can be further increased by higher throughput devices. With this chip and single-cell analysis approaches, a new paradigm for robust early warning systems of ocean environmental perturbation is possible.
ContributorsShi, Xu (Author) / Meldrum, Deirdre R. (Thesis advisor) / Zhang, Weiwen (Committee member) / Chao, Shih-hui (Committee member) / Westerhoff, Paul (Committee member) / Arizona State University (Publisher)
Created2013