ASU Electronic Theses and Dissertations
This collection includes most of the ASU Theses and Dissertations from 2011 to present. ASU Theses and Dissertations are available in downloadable PDF format; however, a small percentage of items are under embargo. Information about the dissertations/theses includes degree information, committee members, an abstract, supporting data or media.
In addition to the electronic theses found in the ASU Digital Repository, ASU Theses and Dissertations can be found in the ASU Library Catalog.
Dissertations and Theses granted by Arizona State University are archived and made available through a joint effort of the ASU Graduate College and the ASU Libraries. For more information or questions about this collection contact or visit the Digital Repository ETD Library Guide or contact the ASU Graduate College at gradformat@asu.edu.
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- All Subjects: Biology
- Creators: Wang, Xuan
Description
The Multiple Antibiotic Resistance Regulator Family (MarR) are transcriptional regulators, many of which forms a dimer. Transcriptional regulation provides bacteria a stabilized responding system to ensure the bacteria is able to efficiently adapt to different environmental conditions. The main function of the MarR family is to create multiple antibiotic resistance from a mutated protein; this process occurs when the MarR regulates an operon. We hypothesized that different transcriptional regulator genes have interactions with each other. It is known that Salmonella pagC transcription is activated by three regulators, i.e., SlyA, MprA, and PhoP. Bacterial Adenylate Cyclase-based Two-Hybrid (BACTH) system was used to research the protein-protein interactions in SlyA, MprA, and PhoP as heterodimers and homodimers in vivo. Two fragments, T25 and T18, that lack endogenous adenylate cyclase activity, were used for construction of chimeric proteins and reconstruction of adenylate cyclase activity was tested. The significant adenylate cyclase activities has proved that SlyA is able to form homodimers. However, weak adenylate cyclase activities in this study has proved that MprA and PhoP are not likely to form homodimers, and no protein-protein interactions were detected in between SlyA, MprA and PhoP, which no heterodimers have formed in between three transcriptional regulators.
ContributorsTao, Zenan (Author) / Shi, Yixin (Thesis advisor) / Wang, Xuan (Committee member) / Bean, Heather (Committee member) / Arizona State University (Publisher)
Created2018
Description
Lignocellulosic biomass represents a renewable domestic feedstock that can support large-scale biochemical production processes for fuels and specialty chemicals. However, cost-effective conversion of lignocellulosic sugars into valuable chemicals by microorganisms still remains a challenge. Biomass recalcitrance to saccharification, microbial substrate utilization, bioproduct titer toxicity, and toxic chemicals associated with chemical pretreatments are at the center of the bottlenecks limiting further commercialization of lignocellulose conversion. Genetic and metabolic engineering has allowed researchers to manipulate microorganisms to overcome some of these challenges, but new innovative approaches are needed to make the process more commercially viable. Transport proteins represent an underexplored target in genetic engineering that can potentially help to control the input of lignocellulosic substrate and output of products/toxins in microbial biocatalysts. In this work, I characterize and explore the use of transport systems to increase substrate utilization, conserve energy, increase tolerance, and enhance biocatalyst performance.
ContributorsKurgan, Gavin (Author) / Wang, Xuan (Thesis advisor) / Nielsen, David (Committee member) / Misra, Rajeev (Committee member) / Nannenga, Brent (Committee member) / Arizona State University (Publisher)
Created2018
Description
The ability to tolerate bouts of oxygen deprivation varies tremendously across the animal kingdom. Adult humans from different regions show large variation in tolerance to hypoxia; additionally, it is widely known that neonatal mammals are much more tolerant to anoxia than their adult counterparts, including in humans. Drosophila melanogaster are very anoxia-tolerant relative to mammals, with adults able to survive 12 h of anoxia, and represent a well-suited model for studying anoxia tolerance. Drosophila live in rotting, fermenting media and a result are more likely to experience environmental hypoxia; therefore, they could be expected to be more tolerant of anoxia than adults. However, adults have the capacity to survive anoxic exposure times ~8 times longer than larvae. This dissertation focuses on understanding the mechanisms responsible for variation in survival from anoxic exposure in the genetic model organism, Drosophila melanogaster, focused in particular on effects of developmental stage (larval vs. adults) and within-population variation among individuals.
Vertebrate studies suggest that surviving anoxia requires the maintenance of ATP despite the loss of aerobic metabolism in a manner that prevents a disruption of ionic homeostasis. Instead, the abilities to maintain a hypometabolic state with low ATP and tolerate large disturbances in ionic status appear to contribute to the higher anoxia tolerance of adults. Furthermore, metabolomics experiments support this notion by showing that larvae had higher metabolic rates during the initial 30 min of anoxia and that protective metabolites were upregulated in adults but not larvae. Lastly, I investigated the genetic variation in anoxia tolerance using a genome wide association study (GWAS) to identify target genes associated with anoxia tolerance. Results from the GWAS also suggest mechanisms related to protection from ionic and oxidative stress, in addition to a protective role for immune function.
Vertebrate studies suggest that surviving anoxia requires the maintenance of ATP despite the loss of aerobic metabolism in a manner that prevents a disruption of ionic homeostasis. Instead, the abilities to maintain a hypometabolic state with low ATP and tolerate large disturbances in ionic status appear to contribute to the higher anoxia tolerance of adults. Furthermore, metabolomics experiments support this notion by showing that larvae had higher metabolic rates during the initial 30 min of anoxia and that protective metabolites were upregulated in adults but not larvae. Lastly, I investigated the genetic variation in anoxia tolerance using a genome wide association study (GWAS) to identify target genes associated with anoxia tolerance. Results from the GWAS also suggest mechanisms related to protection from ionic and oxidative stress, in addition to a protective role for immune function.
ContributorsCampbell, Jacob B (Author) / Harrison, Jon F. (Thesis advisor) / Gadau, Juergen (Committee member) / Call, Gerald B (Committee member) / Sweazea, Karen L (Committee member) / Rosenberg, Michael S. (Committee member) / Arizona State University (Publisher)
Created2018
Description
Isolation-by-distance is a specific type of spatial genetic structure that arises when parent-offspring dispersal is limited. Many natural populations exhibit localized dispersal, and as a result, individuals that are geographically near each other will tend to have greater genetic similarity than individuals that are further apart. It is important to identify isolation-by-distance because it can impact the statistical analysis of population samples and it can help us better understand evolutionary dynamics. For this dissertation I investigated several aspects of isolation-by-distance. First, I looked at how the shape of the dispersal distribution affects the observed pattern of isolation-by-distance. If, as theory predicts, the shape of the distribution has little effect, then it would be more practical to model isolation-by-distance using a simple dispersal distribution rather than replicating the complexities of more realistic distributions. Therefore, I developed an efficient algorithm to simulate dispersal based on a simple triangular distribution, and using a simulation, I confirmed that the pattern of isolation-by-distance was similar to other more realistic distributions. Second, I developed a Bayesian method to quantify isolation-by-distance using genetic data by estimating Wright’s neighborhood size parameter. I analyzed the performance of this method using simulated data and a microsatellite data set from two populations of Maritime pine, and I found that the neighborhood size estimates had good coverage and low error. Finally, one of the major consequences of isolation-by-distance is an increase in inbreeding. Plants are often particularly susceptible to inbreeding, and as a result, they have evolved many inbreeding avoidance mechanisms. Using a simulation, I determined which mechanisms are more successful at preventing inbreeding associated with isolation-by-distance.
ContributorsFurstenau, Tara N (Author) / Cartwright, Reed A (Thesis advisor) / Rosenberg, Michael S. (Committee member) / Taylor, Jesse (Committee member) / Wilson-Sayres, Melissa (Committee member) / Arizona State University (Publisher)
Created2015
Description
Emergence of multidrug resistant (MDR) bacteria is a major concern to global health. One of the major MDR mechanisms bacteria employ is efflux pumps for the expulsion of drugs from the cell. In Escherichia coli, AcrAB-TolC proteins constitute the major chromosomally-encoded drug efflux system. AcrB, a trimeric membrane protein is well-known for its substrate promiscuity. It has the ability to efflux a broad spectrum of substrates alongside compounds such as dyes, detergent, bile salts and metabolites. Newly identified AcrB residues were shown to be functionally relevant in the drug binding and translocation pathway using a positive genetic selection strategy. These residues—Y49, V127, D153, G288, F453, and L486—were identified as the sites of suppressors of an alteration, F610A, that confers a drug hypersensitivity phenotype. Using site-directed mutagenesis (SDM) along with the real-time efflux and the classical minimum inhibitory concentration (MIC) assays, I was able to characterize the mechanism of suppression.
Three approaches were used for the characterization of these suppressors. The first approach focused on side chain specificity. The results showed that certain suppressor sites prefer a particular side chain property, such as size, to overcome the F610A defect. The second approach focused on the effects of efflux pump inhibitors. The results showed that though the suppressor residues were able to overcome the intrinsic defect of F610A, they were unable to overcome the extrinsic defect caused by the efflux pump inhibitors. This showed that the mechanism by which F610A imposes its effect on AcrB function is different than that of the efflux pump inhibitors. The final approach was to determine whether suppressors mapping in the periplasmic and trans-membrane domains act by the same or different mechanisms. The results showed both overlapping and distinct mechanisms of suppression.
To conclude, these approaches have provided a deeper understanding of the mechanisms by which novel suppressor residues of AcrB overcome the functional defect of the drug binding domain alteration, F610A.
Three approaches were used for the characterization of these suppressors. The first approach focused on side chain specificity. The results showed that certain suppressor sites prefer a particular side chain property, such as size, to overcome the F610A defect. The second approach focused on the effects of efflux pump inhibitors. The results showed that though the suppressor residues were able to overcome the intrinsic defect of F610A, they were unable to overcome the extrinsic defect caused by the efflux pump inhibitors. This showed that the mechanism by which F610A imposes its effect on AcrB function is different than that of the efflux pump inhibitors. The final approach was to determine whether suppressors mapping in the periplasmic and trans-membrane domains act by the same or different mechanisms. The results showed both overlapping and distinct mechanisms of suppression.
To conclude, these approaches have provided a deeper understanding of the mechanisms by which novel suppressor residues of AcrB overcome the functional defect of the drug binding domain alteration, F610A.
ContributorsBlake, Mellecha (Author) / Misra, Rajeev (Thesis advisor) / Stout, Valerie (Committee member) / Wang, Xuan (Committee member) / Arizona State University (Publisher)
Created2016
Description
Leprosy and tuberculosis are age-old diseases that have tormented mankind and left behind a legacy of fear, mutilation, and social stigmatization. Today, leprosy is considered a Neglected Tropical Disease due to its high prevalence in developing countries, while tuberculosis is highly endemic in developing countries and rapidly re-emerging in several developed countries. In order to eradicate these diseases effectively, it is necessary to understand how they first originated in humans and whether they are prevalent in nonhuman hosts which can serve as a source of zoonotic transmission. This dissertation uses a phylogenomics approach to elucidate the evolutionary histories of the pathogens that cause leprosy and tuberculosis, Mycobacterium leprae and the M. tuberculosis complex, respectively, through three related studies. In the first study, genomes of M. leprae strains that infect nonhuman primates were sequenced and compared to human M. leprae strains to determine their genetic relationships. This study assesses whether nonhuman primates serve as a reservoir for M. leprae and whether there is potential for transmission of M. leprae between humans and nonhuman primates. In the second study, the genome of M. lepraemurium (which causes leprosy in mice, rats, and cats) was sequenced to clarify its genetic relationship to M. leprae and other mycobacterial species. This study is the first to sequence the M. lepraemurium genome and also describes genes that may be important for virulence in this pathogen. In the third study, an ancient DNA approach was used to recover M. tuberculosis genomes from human skeletal remains from the North American archaeological record. This study informs us about the types of M. tuberculosis strains present in post-contact era North America. Overall, this dissertation informs us about the evolutionary histories of these pathogens and their prevalence in nonhuman hosts, which is not only important in an anthropological context but also has significant implications for disease eradication and wildlife conservation.
ContributorsHonap, Tanvi Prasad (Author) / Stone, Anne C (Thesis advisor) / Rosenberg, Michael S. (Thesis advisor) / Clark-Curtiss, Josephine E (Committee member) / Krause, Johannes (Committee member) / Arizona State University (Publisher)
Created2017
Characterization and Manipulation of Microbiomes From Arid Landfills for Improved Methane Production
Description
Environmentally harmful byproducts from solid waste’s decomposition, including methane (CH4) emissions, are managed through standardized landfill engineering and gas-capture mechanisms. Yet only a limited number of studies have analyzed the development and composition of Bacteria and Archaea involved in CH4 production from landfills. The objectives of this research were to compare microbiomes and bioactivity from CH4-producing communities in contrasting spatial areas of arid landfills and to tests a new technology to biostimulate CH4 production (methanogenesis) from solid waste under dynamic environmental conditions controlled in the laboratory. My hypothesis was that the diversity and abundance of methanogenic Archaea in municipal solid waste (MSW), or its leachate, play an important role on CH4 production partially attributed to the group’s wide hydrogen (H2) consumption capabilities. I tested this hypothesis by conducting complementary field observations and laboratory experiments. I describe niches of methanogenic Archaea in MSW leachate across defined areas within a single landfill, while demonstrating functional H2-dependent activity. To alleviate limited H2 bioavailability encountered in-situ, I present biostimulant feasibility and proof-of-concepts studies through the amendment of zero valent metals (ZVMs). My results demonstrate that older-aged MSW was minimally biostimulated for greater CH4 production relative to a control when exposed to iron (Fe0) or manganese (Mn0), due to highly discernable traits of soluble carbon, nitrogen, and unidentified fluorophores found in water extracts between young and old aged, starting MSW. Acetate and inhibitory H2 partial pressures accumulated in microcosms containing old-aged MSW. In a final experiment, repeated amendments of ZVMs to MSW in a 600 day mesocosm experiment mediated significantly higher CH4 concentrations and yields during the first of three ZVM injections. Fe0 and Mn0 experimental treatments at mesocosm-scale also highlighted accelerated development of seemingly important, but elusive Archaea including Methanobacteriaceae, a methane-producing family that is found in diverse environments. Also, prokaryotic classes including Candidatus Bathyarchaeota, an uncultured group commonly found in carbon-rich ecosystems, and Clostridia; All three taxa I identified as highly predictive in the time-dependent progression of MSW decomposition. Altogether, my experiments demonstrate the importance of H2 bioavailability on CH4 production and the consistent development of Methanobacteriaceae in productive MSW microbiomes.
ContributorsReynolds, Mark Christian (Author) / Cadillo-Quiroz, Hinsby (Thesis advisor) / Krajmalnik-Brown, Rosa (Thesis advisor) / Wang, Xuan (Committee member) / Kavazanjian, Edward (Committee member) / Arizona State University (Publisher)
Created2022
Description
Biomass synthesis is a competing factor in biological systems geared towards generation of commodity and specialty chemicals, ultimately limiting maximum titer and yield; in this thesis, a widely generalizable, modular approach focused on decoupling biomass synthesis from the production of the phenylalanine in a genetically modified strain of E. coli BW25113 was explored with the use of synthetic trans-encoded small RNA (sRNA) to achieve greater efficiency. The naturally occurring sRNA MicC was used as a scaffold, and combined on a plasmid with a promoter for anhydrous tetracycline (aTc) and a T1/TE terminator. The coding sequence corresponding to the target binding site for fourteen potentially growth-essential gene targets as well as non-essential lacZ was placed in the seed region of the of the sRNA scaffold and transformed into BW25113, effectively generating a unique strain for each gene target. The BW25113 strain corresponding to each gene target was screened in M9 minimal media; decreased optical density and elongated cell morphology changes were observed and quantified in all induced sRNA cases where growth-essential genes were targeted. Six of the strains targeting different aspects of cell division that effectively suppressed growth and resulted in increased cell size were then screened for viability and metabolic activity in a scaled-up shaker flask experiment; all six strains were shown to be viable during stationary phase, and a metabolite analysis showed increased specific glucose consumption rates in induced strains, with unaffected specific glucose consumption rates in uninduced strains. The growth suppression, morphology and metabolic activity of the induced strains in BW25113 was compared to the bacteriostatic additives chloramphenicol, tetracycline, and streptomycin. At this same scale, the sRNA plasmid targeting the gene murA was transformed into BW25113 pINT-GA, a phenylalanine overproducer with the feedback resistant genes aroG and pheA overexpressed. Two induction times were explored during exponential phase, and while the optimal induction time was found to increase titer and yield amongst the BW25113 pINT-GA murA sRNA variant, overall this did not have as great a titer or yield as the BW25113 pINT-GA strain without the sRNA plasmid; this may be a result of the cell filamentation.
ContributorsHerschel, Daniel Jordan (Author) / Nielsen, David R (Thesis advisor) / Torres, César I (Committee member) / Wang, Xuan (Committee member) / Arizona State University (Publisher)
Created2016
Description
Reactive oxygen species (ROS) including superoxide, hydrogen peroxide, and hydroxyl radicals occur naturally as a byproduct of aerobic respiration. To mitigate damages caused by ROS, Escherichia coli employs defenses including two cytosolic superoxide dismutases (SODs), which convert superoxide to hydrogen peroxide. Deletion of both sodA and sodB, the genes coding for the cytosolic SOD enzymes, results in a strain that is unable to grow on minimal medium without amino acid supplementation. Additionally, deletion of both cytosolic SOD enzymes in a background containing the relA1 allele, an inactive version of the relA gene that contributes to activation of stringent response by amino acid starvation, results in a strain that is unable to grow aerobically, even on rich medium. These observations point to a relationship between the stringent response and oxidative stress. To gain insight into this relationship, suppressors were isolated by growing the ∆sodAB relA1 cells aerobically on rich medium, and seven suppressors were further examined to characterize distinct colony sizes and temperature sensitivity phenotypes. In three of these suppressor-containing strains, the relA1 allele was successfully replaced by the wild type relA allele to allow further study in aerobic conditions. None of those three suppressors were found to increase tolerance to exogenous superoxides produced by paraquat, which shows that these mutations only overcome the superoxide buildup that naturally occurs from deletion of SODs. Because each of these suppressors had unique phenotypes, it is likely that they confer tolerance to SOD-dependent superoxide buildup by different mechanisms. Two of these three suppressors have been sent for whole-genome sequencing to identify the location of the suppressor mutation and determine the mechanism by which they confer superoxide tolerance.
ContributorsFlake, Melissa (Author) / Misra, Rajeev (Thesis advisor) / Shah, Dhara (Committee member) / Wang, Xuan (Committee member) / Arizona State University (Publisher)
Created2024