Theses and Dissertations at Montana State University (MSU)
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Item Implementing metabolic monitoring in second-generation antipsychotic use: a quality improvement project(Montana State University - Bozeman, College of Nursing, 2024) Stone, Amanda Marie; Chairperson, Graduate Committee: Lindsey Davis; This is a manuscript style paper that includes co-authored chapters.Background: Roughly one in five adults in the United States live with a psychiatric disorder, including major depressive disorder, bipolar disorder, and schizophrenia. Despite their effectiveness in treating psychiatric disorders, second-generation antipsychotics (SGAs) are associated with an increased rate of metabolic side effects (MSEs). To reduce the impact of MSEs and the potential development of metabolic syndrome, individuals on SGA need routine metabolic screening. Local problem: At an outpatient mental health clinic, 21% of the patients were prescribed SGAs; however, the clinic lacked a standardized protocol to assess MSEs. This project aimed to increase the completion rate of metabolic monitoring and improve patient health outcomes in patients diagnosed with a psychiatric disorder. Methods: A multifaceted approach was created for providers to increase adherence to metabolic screening in SGA use. Interventions: Three interventions were initiated: an electronic health record macro was created to promote consistent provider documentation, a standardized metabolic monitoring process, and educational in-services performed to encourage screening adherence. Results: The project aims were achieved with the clinic maintaining an 80% blood pressure (BP) completion rate. Documentation of body mass index (BMI) increased by 72%. Appropriate lab monitoring improved from an 8.5% rate during the preintervention phase to a rate of 33% postintervention. There were no changes in provider perceptions and practices regarding metabolic monitoring in SGA use. Conclusion: The project effectively increased the rate of metabolic monitoring in individuals prescribed an SGA.Item Microbial adaptation to cultivation stress using storage compounds(Montana State University - Bozeman, College of Agriculture, 2022) Arnold, Adrienne Dale; Chairperson, Graduate Committee: Ross Carlson; This is a manuscript style paper that includes co-authored chapters.Methanotrophs and green algae are microorganisms that grow on single carbon substrates. Methanotrophs are bacteria that use methane as their carbon source, and green algae are eukaryotic phototrophs that grow on CO 2. They are of interest both as primary producers in the environment and as biological catalysts for the conversion of greenhouse gases into value-added compounds. Understanding how methanotrophs and green algae adapt to cultivation stresses is key to understanding carbon cycling in the environment and in industrial settings. This work uses stoichiometric metabolic modeling to investigate the role of carbon storage compounds in the metabolism of C1-utilizing organisms. Storage compounds are accumulated as intracellular reserves of polysaccharides or lipids, which can be catabolized under stress conditions to provide carbon and energy to the cell. Catabolism of carbon storage compounds often results in the excretion of multi-carbon organic compounds that can be utilized as carbon substrates by other members of the microbial community. In silico metabolic models were developed for methanotroph and algal systems and used to examine the breakdown of storage compounds in response to common cultivation stresses. For the aerobic methanotrophs, predictions focused on the use of polyhydroxybutyrate and glycogen in adaptation to O 2 limitation. For the green algae, starch and triacylglycerol reserves are analyzed as sources for compatible solutes, which are produced by cells in response to high salinity conditions. Metabolic modeling of storage compound utilization by methanotrophs and algae helps elucidate the role of these organisms as primary producers and presents an opportunity for industrial production of multi-carbon compounds from single carbon substrates.Item Genomic composition of green algae grown in high alkaline conditions(Montana State University - Bozeman, College of Agriculture, 2023) Goemann, Calvin Lee Cicha; Chairperson, Graduate Committee: Blake Wiedenheft; This is a manuscript style paper that includes co-authored chapters.Algae are responsible for 50% of global oxygen production and sequestration of CO 2 from the atmosphere. Algal photosynthesis plays a critical role in all aquatic ecosystems converting sunlight and CO2 into usable biomass. Algal growth and biomass production can be coopted to produce industrially relevant bioproducts like triacylglycerol (TAGs) that can be converted into biodiesel and provide a sustainable carbon-neutral alternative to fossil fuels. In high-stress environments, algae produce high levels of TAGs. Multiple stresses including nitrogen limitation and high pH impact algae physiology, but little is known about how algae shift their metabolism to produce TAGs in response to these stresses. This topic remains relatively unexplored due to the limited availability of complete algae genomes. Here we sequence and annotate the complete telomere-to-telomere genome of an alkali-tolerant green algae Chlorella sp. SLA-04. Genomic analysis supports a reclassification of Chlorophyta green algae and illuminates how SLA-04 adapts to diverse environmental conditions. Additionally, transcriptomic analysis revealed how Chlorella sp. SLA-04 rewires carbon metabolism in high alkaline and nutrient-deplete conditions to produce TAGs while minimizing photosynthetic oxidative stress. Together, we double the amount of publicly available telomere-to-telomere green algal genomes and use this resource to explore how algae respond to diverse environmental conditions in their native and industrial settings.Item Identification of cellulolytic hot spring organisms through bioorthogonal labeling(Montana State University - Bozeman, College of Letters & Science, 2021) Reichart, Nicholas John; Chairperson, Graduate Committee: Roland Hatzenpichler; This is a manuscript style paper that includes co-authored chapters.Microbial physiology is the study of the metabolism and function of microorganisms. The recent expansion of genomic diversity has outpaced the description of physiology. To better understand microbial metabolisms and environmental processes, more detailed research is needed for both novel and undescribed microbes. While many new methods are being developed to describe in situ microbial activity, this dissertation implements bioorthogonal non-canonical amino acid tagging as a proxy to track metabolic activity of microbes under close to environment conditions. Using differential analyses on hot spring microbial communities, we were able to show that certain microbial taxa had preferential activity towards specific incubation amendments. Previous activity-based studies had shown that hot springs were a unique environment for discovering cellulolytic microbes that could be used in industrial processing of plant biomass. Herein, we used computational analysis to screen publicly available metagenomic datasets to identify the enzymatic potential of hot springs worldwide. The wide diversity of taxa and biomass degrading enzymes were investigated and hot springs were further highlighted as a system that could be used to find improvement for the industry of plant biomass degradation and processing. To build upon the cellulolytic potential found in hot spring metagenomic datasets, bioorthogonal non-canonical amino acid tagging coupled with fluorescence-activated cell sorting was applied to the biotechnological relevant field of plant biomass degradation to identify microbes involved in the cellulolytic process. Examination of the active microbes revealed difference in the community when supplemented with cellulose. Taken together, the work in this dissertation served to expand and apply the recent development of activity-based studies used to describe environmental microbial populations, with a focus on plant biomass degradation.Item Managing metabolic side effects: a process improvement(Montana State University - Bozeman, College of Nursing, 2022) Scally, Shannon St. Onge; Chairperson, Graduate Committee: Lindsay BenesOne in 25 people live with severe mental illness (SMI) including schizophrenia, bipolar disorder, and major depressive disorder. People with SMI have an earlier mortality rate by up to 25 years. This is, in part, due to treatment with second-generation antipsychotics (SGAs). SGAs treat positive and negative symptoms but are associated with metabolic side effects (MSEs) such as increases in weight, body mass index (BMI), waist circumference, blood pressure, cholesterol, and blood glucose levels. These MSEs place a person at risk for metabolic syndrome (MetS). To mitigate the impact of MSEs and the development of MetS, patients on SGAs need to have regular screening of MSEs. At a mental health clinic in Western Montana, 32% of patients were prescribed SGAs, yet the clinic had no standardized process to address MSEs. Therefore, this project implemented a standardized process to address the management of MSEs. For each SGA prescription or renewal, patients received an MSE risk discussion, MSE screening, and education regarding lifestyle and diet strategies to prevent MSEs. The process-improvement method used was Plan-Do-Study-Act (PDSA). The results revealed an overall range of adherence to the standardized process: 43% of patients with a new or renewed SGA order had a documented MSE risk discussion and MSE screening ordered. Additionally, 89% of patients with an SGA order received education regarding lifestyle and diet management of MSEs by a nurse health educator. The success of this aspect of the process was facilitated by the nurse health educator and demonstrated a positive step in the sustainability of lifestyle and diet support as a first-line intervention in the prevention of MSEs. The organization now has a standardized process to manage MSEs and a new clinical workflow to support patients in mitigating MSEs. As the organization continues improving the process by adding the EHR automation and continuing support of the nurse health educator role in lifestyle and diet follow-up, the future impact of reducing morbidity and mortality of the SMI population is set in motion.Item The influence of external load during hiking on markers of joint stress and movement efficacy(Montana State University - Bozeman, College of Education, Health & Human Development, 2022) Rowland, Isaac Franco; Chairperson, Graduate Committee: Mary P. Miles; Mary Miles, David Graham, Ron June, Brian Bothner and Hunter Fausset were co-authors of the article, 'The influence of external load during hiking on markers of joint stress and movement efficacy' which is contained within this thesis.PURPOSE: Complications to lower extremity joints, including injury and inflammation, are prevalent issues that arise during prolonged external load-bearing exercise. Metabolomic blood analysis can provide insight into the metabolic processes that occurs during this type of exercise. METHODS: Eight healthy, active men and women participated in a series of blood sample collections and motion capture recording before and after completing a 7.2-mile hiking protocol under two separate conditions. Blood was collected prior to hiking, 15-20 minutes after hiking, 8-hours after, 24-hours after, and 48 hours after. Movement coordination and efficacy was measured with a motion capture system while performing the y-balance test and an obstacle clearance task. Hiking conditions were randomized as backpacking with 20% of body mass external load or daypacking with minimal external load for each participant and separated by two weeks. Serum was analyzed to detect differences in metabolite upregulation between conditions. Biomechanical data were analyzed for inter- and intra-differential values relevant to fatigue between conditions. RESULTS: Analysis found clear differences between conditional metabolite upregulation at all post-hike timepoints. The upregulation of cortisol was significant in backpacking conditions at the post-hike timepoint. Glycerophospholipids were significantly upregulated in backpacking at 8-hours post-hike All significant metabolite upregulation switched to daypacking conditions at 24-hours post-hike. Significant metabolite upregulation varied between conditions at 48-hours post-hike. The only findings of significance in movement coordination and efficacy were between the y-balance lateral leg movement. CONCLUSIONS: The presence of cortisol is consistent with the physiological and mental stress of external loadcarriage and alludes to exposure that can lead to decreased bone mineral content. Glycerophospholipid metabolism pathways play an important role in joint degradation, which could explain their upregulation in backpacking conditions. Ceramide, omega-3s, and fatty acid/triglyceride cycling are functions of cell proliferation and turnover which may be upregulated with more efficiency in daypacking conditions. Upregulation of anserine at 48-hours post-hike in daypacking supports the idea of more efficient exercise recovery occurring in this condition. While significant differences were not clear in motor control measures, the findings show potential reliability for future study designs.Item Quantitative 1 H NMR analyses of immunometabolic modulation in human macrophages(Montana State University - Bozeman, College of Letters & Science, 2019) Fuchs, Amanda Lee; Chairperson, Graduate Committee: Valerie Copie; Sage M. Schiller was an author and Wyatt J. Keegan, Mary Cloud B. Ammons, Brian Eilers, Brian Tripet and Valerie Copie were co-authors of the article, 'Quantitative 1 H NMR metabolomics reveals distinct metabolic adaptations in human macrophages following differential activation' in the journal 'Metabolites' which is contained within this dissertation.; Sage M. Schiller was an author and Isaac R. Miller, Mary Cloud B. Ammons, Brian Eilers, Brian Tripet and Valerie Copie were co-authors of the article, 'Pseudomonas aeruginosa planktonic- and biofilm-conditioned media elicit divergent responses in human macrophages' submitted to the journal 'PLoS pathogens' which is contained within this dissertation.Macrophages are innate immune cells that are found ubiquitously in nearly all human tissues, where they support host innate and adaptive immune responses in an effort to maintain systemic homeostasis. They are inherently plastic in nature and can dramatically modulate their functional phenotype according to pathogen and microenvironmental stimuli. Previous studies have shown that macrophages are particularly important for the resolution of inflammation in acute wound healing, which is marked by a phenotypic transition of wound macrophages from pro-inflammatory to anti-inflammatory. Chronic, or non-healing, wounds, such as diabetic, pressure, and venous leg ulcers, feature a prolonged host inflammatory response due in part to aberrant wound macrophage behavior. Non-healing in chronic wounds has also been shown to be dependent upon the establishment of pathogenic biofilms, which are more resistant to host defense mechanisms than planktonic, or free-floating, bacteria. Therefore, investigating macrophage dysregulation in the presence of bacterial biofilms has gained considerable interest. Here, 1D 1 H NMR-based metabolomics was utilized to identify metabolic pathways that are differentially modulated following primary human monocyte-derived macrophage activation with pro-inflammatory or anti-inflammatory stimuli relative to resting macrophages. Metabolic profiling of inflammatory macrophages indicated a substantial increase in oxidative stress as well as a decrease in mitochondrial respiration. These metabolic profiles also provided evidence that inflammatory macrophages divert metabolites from de novo glycerophospholipid synthesis to inhibit oxidative phosphorylation. In addition, we investigated which metabolic pathways are differentially modulated following primary human monocyte-derived macrophage exposure to Pseudomonas aeruginosa planktonic- and biofilm-conditioned media. Metabolic profiling of PCM- and BCM-exposed macrophages indicated a significant depletion of intracellular glucose without elevation of downstream glycolytic products. These metabolic patterns suggest that PCM- and BCM-exposed macrophages potentially divert glycolytic intermediates towards inositol phosphate metabolism. Overall, our studies provide additional support to previous findings, generate novel results regarding metabolic modulation of human macrophages following activation and exposure to planktonic- vs. biofilm-conditioned media, and contribute new insight to the field of immunometabolism.Item Proteomics analysis of the metabolic transition between aerobic and anaerobic conditions in Escherichia coli(Montana State University - Bozeman, College of Letters & Science, 2019) Refai, Mohammed Yahya; Chairperson, Graduate Committee: Brian Bothner; Nina Paris, Hunter Fausset, Monika Tokmina Lukaszewska were co-authors of the article, 'Proteomics analysis of the transition between aerobic and anaerobic growth conditions in Escherichia coli' submitted to the journal 'Biochimica et biophysica acta' which is contained within this dissertation.As a facultative anaerobe, Escherichia coli has the ability to grow in anaerobic and aerobic environments. Despite detailed characterizations of this model organism in the presence and absence of oxygen, an in-depth understanding of changes to the proteome during transitions from aerobic to anaerobic growth is lacking. This thesis work focuses on elucidating how protein thiol oxidation and reduction change during a facultative anaerobe's transition from aerobic to anaerobic growth conditions, and pathways of thiol-mediated cell signaling. Redox driven changes in cysteine oxidation involved in signaling are referred to as 'thiol switches'. These modulate diverse biological activities ranging from gene expression and protein synthesis to environmental stress response. Surprisingly, little is known about the role of thiol switches during microbial transitions from aerobic and anaerobic growth conditions. To explore this uncharted territory, a mass-spectrometry (MS)-based proteomics workflow was developed and refined. Following extensive protocol optimization for high-throughput MS data processing, normalization, and pattern matching, the analytical pipeline was fine-tuned for the specific proteome-wide analysis of cysteine chemical modifications in E. coli. The approach was based on open-source software and publicly accessible databases, creating a transparent, reproducible, and easily sharable proteomics approach. Herein, the redox state and chemical forms of protein-based thiol switches in E. coli were characterized over time as the bacterium reversibly transitioned between aerobic and anaerobic growth conditions. Unexpectedly, differential alkylation analysis of cysteine-containing E. coli proteins revealed a higher degree of protein thiol oxidation under anaerobic growth conditions, a result not reported for E. coli or any other facultative anaerobe. Our proteome-wide analysis also revealed that cysteine redox potentials vary widely, and several specific E. coli proteins contain highly reactive thiols. These findings provide strong evidence for thiol-based signaling in E. coli in response to environmental changes such as aerobic to anaerobic growth transitions. Characterization of specific redox switches underlying metabolic changes associated with oxygen availability has uncovered a previously unknown E. coli cell signaling mechanism. Since transitioning between aerobic and anaerobic environments is associated with bacterial virulence, this work opens new avenues to target pathogenic facultative anaerobes and to develop novel thiol-based antibacterial therapies.Item Metabolic interactions and activity partitioning in a methanogenic, interdomain biofilm(Montana State University - Bozeman, College of Letters & Science, 2019) Camilleri, Laura Beth; Chairperson, Graduate Committee: Matthew Fields; Kristopher A. Hunt, Aurelien Mazurie, Jennifer Kuehl, Alex Michaud, James Connolly, Egan Lohman, Zack Miller, Adam M. Deutschbauer and Matthew W. Fields were co-authors of the article, 'Differential gene expression of a bacterial-archaeal interdomain biofilm producing methane' submitted to the journal 'Biofilms' which is contained within this dissertation.; B.P. Bowen, C.J. Petzold, T.R. Northen and M.W. Fields were co-authors of the article, 'Activity partitioning in an archaeal-bacterial biofilm' submitted to the journal 'Letters in applied microbiology' which is contained within this dissertation.; Matthew W. Fields was a co-author of the article, 'Methanococcus maripaludis factor causes slowed growth in Desulfovibrio vulgaris Hildenborough' submitted to the journal 'Letters in applied microbiology' which is contained within this dissertation.; Matthew W. Fields was a co-author of the article, 'Growth effects of sulfopyruvate and sulfoacetate on the sulfate-reducing bacterium, Desulfovibrio vulgaris Hildenborough, and the methanogenic archaeon Methanococcus maripaludis S2' submitted to the journal 'Scientific reports' which is contained within this dissertation.; Matthew W. Fields was a co-author of the article, 'Methane production in Pelosinus fermentans JBW45' submitted to the journal 'Letters in applied microbiology' which is contained within this dissertation.Biofilms are an ancient survival strategy in which communities of organisms can grow as a cohesive unit, generally attached to a surface and/or at interfaces. Despite the paradigm that 99% of microorganisms grow as a biofilm in the environment, current research methods are largely limited to monoculture planktonic studies. Although more investigations are trying to improve culture complexity by evaluating interactions between two or more populations, experiments are still more readily performed with microorganisms in the planktonic growth mode. The research presented here aims to elucidate the complexity of interactions between two microorganisms from different domains of life that results in enhanced metabolism due to localization of cells in close proximity within an anaerobic biofilm. Desulfovibrio vulgaris Hildenborough (DvH) and Methanococcus maripaludis S2 (Mmp) form a syntrophic mutualism when grown in sulfate-limited media that requires electron flux from DvH to Mmp through what is commonly assumed to be interspecies hydrogen transfer, thereby establishing cross-feeding. The biofilm has been shown to promote a stable and more even carrying capacity for both populations that is likely linked to improved hydrogen transfer (and/or other potential carbon and electron co-metabolites) as compared to planktonic populations. Transcriptomic and proteomic analyses, utilizing RNA-seq and deuterated water respectively, were used to elucidate genes and proteins that contribute to the biofilm growth mode that results in a more efficient metabolism for the syntrophic co-culture (defined by biomass per substrate flux). The results demonstrate the expression of many genes with unknown functions, and others that contribute to cell-cell interactions as well as active proteins in electron processing (e.g., lactate oxidation) in DvH and CO2 reduction (e.g., methanogenesis) in Mmp. A metabolic model of the coculture provided reinforcement for transcriptomic assumptions and aided in the identification of a sulfonate and other amino acids as important syntrophic metabolites. Assessment of biofilm co-culture activity utilizing a new method, Biorthogonal Noncanonical Amino Acid Tagging (BONCAT), showed Mmp was less active in the uptake of a methionine analog as compared to DvH. Alternate assessments confirmed that Mmp was in fact active (based upon methane generation) although translational activity was below the detection limit. Further investigation of the system under sulfate stress showed that the metabolic pairing is more stable than previously thought and could indicate survival strategies that drive the seemingly 'mutualistic' relationship as a forced cooperation. The sulfate stress response coincided with observed lags in DvH growth when grown in Mmp spent medium that was associated with a decoupling of lactate-oxidation and sulfate-reduction. Together the results demonstrate metabolic interactions and activity partitioning within a methanogenic archaeal-bacterial biofilm. The dogma of mutualism being synonymous with equal reciprocity is challenged as it pertains to this model biofilm system. Moreover, this unique bacterial-archaeal biofilm represents interdomain interactions that could represent systems that contributed shared metabolic processes that lead to the development of eukaryotic life.Item Characterization of metabolic changes in osteoarthritis using global metabolomic profiling(Montana State University - Bozeman, College of Agriculture, 2018) Carlson, Alyssa Kay; Chairperson, Graduate Committee: Ronald K. June II; Rachel A. Rawle, Erik Adams, Mark C. Greenwood, Brian Bothner and Ronald K. June were co-authors of the article, 'Application of global metabolomic profiling of synovial fluid for osteoarthritis biomarkers' in the journal 'Biochemical and biophysical research communications' which is contained within this thesis.; Rachel A. Rawle, Cameron W. Wallace, Erik Adams, Mark C. Greenwood, Brian Bothner and Ronald K. June were co-authors of the article, 'Global metabolomic profiling of human synovial fluid for rheumatoid arthritis biomarkers' in the journal 'Clinical and experimental rheumatology' which is contained within this thesis.; Rachel A. Rawle, Cameron Wallace, Ellen Brooks, Erik Adams, Mark C. Greenwood, Merissa Olmer, Martin K. Lotz, Brian Bothner and Ronald K. June were co-authors of the article, 'Characterization of osteoarthritis phenotypes in human synovial fluid using global metabolomic profiling' submitted to the journal 'Arthritis and rheumatology' which is contained within this thesis.; Rachel A. Rawle, Erica Barboza, Albert Batushansky, Timothy M. Griffin, Brian Bothner and Ronald K. June were co-authors of the article, 'In vivio mechanotransduction: effect of acute exercise on the metabolomic profiles of mouse synovial fluid' submitted to the journal 'Journal of biomechanics' which is contained within this thesis.; Rachel A. Rawle, Erin Hutchison, Joanna Hudson, Brian Bothner, Timothy M. Griffin and Ronald K. June were co-authors of the article, 'The effect of long-term voluntary exercise on the metabolomic profiles of synovial fluid from high-fat diet-induced obese mice' submitted to the journal 'Arthritis and rheumatology' which is contained within this thesis.; Dissertation includes one article of which Alyssa Kay Carlson is not the main author.Osteoarthritis affects over 250 million individuals worldwide. It is a disease of the whole joint, exhibiting heterogenous pathology, and a multifactorial etiology consisting of obesity and joint trauma as important risk factors. This heterogenous nature contributes to the disparity in symptom presentation and response to treatments, presenting challenges for diagnosis and the development of targeted therapies for osteoarthritis phenotypes. Therefore, the goals of this work were to (1) enhance our understanding of osteoarthritis as a heterogenous disease for improved early diagnosis and (2) evaluate the interaction between osteoarthritis risk factors and therapeutic interventions. Because osteoarthritis and its risk factors are associated with aberrant metabolism, liquid chromatography-mass spectrometry-based global metabolomic profiling was employed to investigate changes in small molecules in response to osteoarthritis, risk factors, and therapeutic interventions. The first area of research focused on osteoarthritis diagnosis. The results show that global metabolomic profiling of human osteoarthritic synovial fluid is capable of identifying candidate biomarkers of osteoarthritis and classifying donors into subgroups representative of metabolic phenotypes. Metabolic phenotypes include structural deterioration, oxidative stress, and/or inflammation. The second area of research focused on osteoarthritis risk factors and therapeutic interventions. We investigated the effects of acute exercise in mouse synovial fluid to provide insight into exercise as a nonpharmacologic mechanobiology-based intervention prescribed for osteoarthritis. We found that acute exercise may have beneficial effects in maintaining overall joint health. We expanded on exercise as a nonpharmacologic treatment by investigating the effects of long-term exercise in an obesity-associated osteoarthritis mouse model. Long-term exercise did not exacerbate osteoarthritis in the knee joints of obese mice but did abrogate some obesity-induced metabolic perturbations in the synovial fluid. In addition, a pharmacologic intervention was investigated in posttraumatic osteoarthritis. Inhibition of early response genes by a Cdk9 inhibitor immediately after joint trauma was also capable of reversing a portion of injury-induced metabolic perturbations in whole joints of injured mice. Overall, this work demonstrates that global metabolomic profiling has potential for biomarker discovery and classifying patients into metabolic phenotypes. It also demonstrates the potential for exercise and inhibition of early response genes as therapeutic interventions for obesity-associated and post-traumatic osteoarthritis.