Theses and Dissertations at Montana State University (MSU)
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Item Comparing juvenile physiology and morphology of two high-elevation pines, Pinus albicaulis and Pinus balfouriana(Montana State University - Bozeman, College of Letters & Science, 2023) Sparks, Katherine Elizabeth; Chairperson, Graduate Committee: Danielle UlrichWhitebark pine (Pinus albicaulis, PIAL) and foxtail pine (P. balfouriana, PIBA) are slow-growing, high-elevation, five needled ("high five") white pines and are foundation and keystone species in alpine and subalpine environments, providing essential resources and habitat for many species including the Clark's nutcracker and grizzly bears. In recent years, PIAL has experienced significant decline due to an amalgamation of climate change, white pine blister rust, and mountain pine beetle. As a result, PIAL is listed as endangered under the Canadian Species at Risk Act and threatened under the United States Endangered Species Act. Conversely, PIBA has experienced minimal decline. PIBA also exists in two disjunct populations, one in southern California (PIBAS) and one in northern California (PIBA N), resulting in the species being split into two sub-species (P. balfouriana subsp. austrina and balfouriana). Our study compared the physiology and morphology of the two species (PIAL and PIBA) and the two foxtail populations (PIBA N and PIBAS) to better understand how they interact with and respond to abiotic and biotic stressors in their high-elevation environments. We grew four-year-old PIAL and PIBA juveniles in a common greenhouse environment. In total, we measured 159 traits describing their morphology, biomass, stomata, xylem, budburst phenology, physiology, whole plant Volatile Organic Compounds (wpVOCs), phloem volatile resin (PVR) compounds, and Non-Structural Carbohydrates (NSCs). We found that PIAL and PIBA displayed different suites of traits that enable them to persist in their high elevation habitats, characterized by similar abiotic stressors (cold temperatures, high winds, summer drought) and biotic stressors (white pine blister rust, bark beetle). The two foxtail populations were similar for most traits except for wpVOC concentration and composition where PIBAS had significantly higher wpVOC concentration than PIBA N. For most traits, PIAL was most similar to PIBA N and differed the most with PIBAS while PIBA N was the intermediate being more similar to both groups, especially in wpVOC composition and concentration.Item Rhizobiome dynamics in plant growth promotion and abiotic stress response(Montana State University - Bozeman, College of Agriculture, 2023) Goemann, Hannah Marie; Chairperson, Graduate Committee: Brent M. PeytonSoil microorganisms play vital roles in global nutrient cycling. Understanding the complex relationships between plants and soil microbes and their implications is one of the greatest challenges facing microbial ecology today. Soil microbes can play beneficial roles in supporting plant growth by increasing access to nutrients, water, and decreasing plant stress signaling under abiotic stresses such as drought and heat. With increasing climate variability due to climate change, it is imperative to make scientifically informed management decisions to best support global biodiversity and plant productivity in natural and agroecosystsms. In this dissertation I summarize four separate investigations of plant-microbe interactions. The first is using nitrogen-fixing cyanobacterial biofertilizers to promote plant growth of perennial second generation bioenergy crops switchgrass (Panicum virgatum) and tall wheatgrass (Agropyrun elongatum). The second and third studies seek to better understand plant-microbe carbon exchange under drought stress in the native North American prairie grass blue grama (Bouteloua gracilis). The final study explores the potential microbial contribution to heat tolerance of panic grass (Dichanthelium lanuginosum) across a natural soil temperature gradient in Yellowstone National Park. Next-generation amplicon sequencing using the Illumina Miseq platform is the primary technique utilized across the three studies to investigate microbial community dynamics. The main results of the biofertilizer study were that tall wheatgrass is better suited to the SW Montana growing season than switchgrass, and similar plant yields were achieved with the cyanobacterial biofertilizer as with urea chemical fertilizer without negatively impacting the microbial community diversity. The first blue grama study found that severe and mild drought had distinct, phylogenetically linked responses within the blue grama rhizobiome with Planctomycetes, Thermoproteota (ammonia-oxidizing archaea), and Glomeromycetes (arbuscular mycorrhizal fungi) exhibiting notably altered relative abundances. The second blue grama study found that climate legacy plays an important role in shaping blue grama drought response. Finally, from the D. lanuginosum study in Yellowstone National Park we learned that pH and temperature both strongly influence community composition, and that D. lanuginosum selects for unique community members in its rhizosphere at higher temperatures. Collectively, these studies contribute to furthering our understanding of the dynamics of plant-associated microbiomes.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 Evaluating the genetic and phenotypic responses of Camelina sativa to heat stress(Montana State University - Bozeman, College of Agriculture, 2023) Smith, Brian Edward; Chairperson, Graduate Committee: Chaofu LuCamelina sativa (L.) Crantz is a low-input oilseed crop with a unique fatty acid profile in its seed oil. Camelina oil can be used in biodiesels, jet fuels, and industrial lubricants. Improving the abiotic stress tolerance of camelina is a crucial step for increasing agronomic viability. Climate change is threatening production of camelina with rising global temperatures and shorter growing seasons. Elucidating the phenotypic and genetic responses to high temperatures is essential for successful breeding of heat tolerant camelina varieties. Three experiments were conducted to understand these responses. Two genotypes, Suneson (MT5) and Pryzeth (MT102) were exposed to a transient 14-day heat stress during the reproductive stage and evaluated for agronomic and seed quality traits along the main stem. Next, a mapping population consisting of 257 recombinant inbred lines (RILs) were grown under the same temperature regimes for 14 days beginning at the onset of the reproductive stage. Finally, reproductive tissues undergoing heat stress from two genotypes with contrasting heat responses, RIL23 and RIL167, were examined with RNA sequencing, and the phenotypes along the main stem were compared. From the phenotype evaluation, both MT5 and MT102 were significantly impacted by heat. Both genotypes experienced reductions in seed and pod size, seed weight, and total oil contents. As reported in other oilseed crops, camelina is negatively affected by heat, characterized primarily by lower yield and reduced oil content. The QTL analysis identified several key gene regions with co-located traits on chromosomes 8, 10, and 12. This demonstrates the ability to identify heat-responsive gene regions via phenotyping along the main stem. The transcriptomes of RILs 23 and 167 contrasted in both sampled tissue types, with RIL23 appearing more responsive to heat. Phenotypic analysis of these genotypes confirmed the transcriptional differences, as RIL23 was more resistant for several traits associated with fertility. These studies provide resources and protocols for future studies that may assist in improving the heat-tolerance of camelina.Item Characteristics of whitebark pine (Pinus albicaulis) growth & defense in disturbance-prone, high-elevation, montane ecosystems of the northern Rocky Mountains(Montana State University - Bozeman, College of Letters & Science, 2022) Kichas, Nickolas Earl; Chairperson, Graduate Committee: David McWethy; This is a manuscript style paper that includes co-authored chapters.Whitebark pine (Pinus albicaulis) is a high-elevation conifer, recognized as a foundation species due to the numerous ecological benefits it provides in subalpine environments. In whitebark pine and other conifers, resin-based defenses have long been recognized as the primary mechanism by which trees respond to bark beetle attacks and several studies have linked resin duct properties to survivorship during periods of increased beetle activity. Utilizing a unique dataset of whitebark pine collected on the Flathead Indian Reservation in northwestern Montana, we set out to investigate the following research questions: (1) Are there differences in physiology (tree growth and resin duct anatomy) between trees that persisted through recent mountain pine beetle outbreaks and trees that died? (2) Does constitutive resin chemistry differ between whitebark and co-occurring lodgepole pine and are there relationships between tree growth, resin duct anatomy and resin chemistry? (3) Does competition influence constitutive resin chemistry in either whitebark or lodgepole pine? and (4) Is whitebark pine growth and/or resin duct anatomy constrained by warmer and/or regionally drier conditions? We found that whitebark pine trees that have persisted through recent stand-level disturbance produced fewer but larger resin duct structures with greater duct area compared to trees that died. We also detected important differences in the chemical composition of resin between whitebark and lodgepole pine that generally support field observations, whereby under endemic scenarios mountain pine beetle preferentially select lodgepole pine, while under outbreak scenarios, beetles successfully colonize whitebark pine trees. We found complex relationships between tree growth, resin duct anatomy and constitutive resin chemistry that present beetles with many permutations of resin-based defenses, while competition, particularly with Engelmann spruce (Picea engelmannii) can further influence constitutive resin chemistry. Lastly, we found that whitebark pine across our study sites are experiencing increased growth and defense under warmer and regionally drier conditions. Whitebark pine at our study sites exhibit differing strategies in the allocation of resources toward growth and defense. Our results support the idea that maintaining genetic variability promotes diverse response strategies to a complex array of biophysical stressors that might leave a species vulnerable to extinction across its range.Item Investigating the ability of arbuscular mycorrhizal fungi to mitigate the negative effects of warming and drought on native perennial forbs(Montana State University - Bozeman, College of Letters & Science, 2022) Eggers, Jessica Avery; Chairperson, Graduate Committee: Laura Burkle; This is a manuscript style paper that includes co-authored chapters.The ability of arbuscular mycorrhizal fungi (AMF) to mitigate the negative effects of warming and drought on plant hosts is known for crop species but is poorly understood for native, perennial forbs. Examining the indirect influence of AMF on forbs' responses to these stressors will provide a more complete understanding of how native forbs will be affected by climate change. In an experimental greenhouse study, we inoculated two native forb species (Achillea millefolium and Linum lewisii) with three separate AMF species (Rhizophagus clarus, Claroideoglomus etunicatum, and Gigaspora rosea), then exposed plants, including an uninoculated control treatment, to varying degrees of drought and heat stress in a factorial design. We tested the effects of warming or drought treatments on plants' physical, floral, phenological, and physiological traits, including biomass, height, floral abundance, flower size, first date of flowering, floral scent, and photosynthetic performance. For both forbs, AMF ameliorated the negative effects of drought and warming on plant survival and vegetative growth, but the magnitude of effect was specific to the forb species, climate treatment, and AMF inoculant. AMF also produced changes in forb phenology, floral scent (volatile organic compounds), and flowering success and duration, which have broad implications for plant-pollinator interactions and the links between belowground and aboveground symbioses. Together, these results indicate that AMF can assist native forbs in surviving, growing, and reproducing in a warmer and drier climate.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 Physiological traits and stress tolerances of three high-elevation pine species(Montana State University - Bozeman, College of Letters & Science, 2023) Wasteneys, Chloe Elizabeth; Chairperson, Graduate Committee: Danielle UlrichThe increasing frequency and intensity of drought and heat events are increasing forest mortality rates worldwide. To improve restoration and management effort effectiveness for at- risk tree species and to accurately predict how these species will respond to future climates, the physiological mechanisms of seedling establishment and survival need to be fully understood. Using a greenhouse common garden approach, we compared the physiological traits and stress tolerances of three high-elevation five-needle pine species, Pinus albicaulis (PIAL), P. flexilis (PIFL), and P. longaeva (PILO). We measured a suite of physiological response curves to determine photosynthetic capacity, high-light tolerance, drought tolerance, and heat tolerance traits. We compared these traits among the three species (Q1), among three different ages of PIAL seedlings (2-, 3-, and 5-years-old; Q2), and among seven families of 2-year-old PIAL originating from different locations (Q3). Among species (Q1), only one heat tolerance trait (Tcrit, the temperature at which minimal fluorescence begins to increase) significantly differed and increased from PILO to PIFL to PIAL, suggesting that PIAL had greater heat tolerance than PIFL and PILO. Among PIAL ages (Q2), two drought tolerance traits, saturated water content (SWC) and leaf mass per area (LMA), and two heat tolerance traits, Tcrit and T50_EL (temperature that results in a 50% increase in electrolyte leakage), significantly differed among ages. SWC decreased, LMA and Tcrit increased, and T50_EL partially increased with increasing age, suggesting that older PIAL seedlings may be more drought and heat tolerant than younger PIAL seedlings. Among PIAL families (Q3), no traits significantly differed. However, we observed four significant correlations between our measured traits and growing season mean temperature and vapor pressure deficit of the seven PIAL families. Families from relatively cooler, wetter locations tended to have greater high-light tolerance and greater heat tolerance (based on T50_EL) while families from relatively warmer, drier locations tended to have greater heat tolerances (based on Tcrit and T50 measured with chlorophyll fluorescence). Together, these findings improve our understanding of physiological mechanisms underlying seedling establishment and our ability to predict how these species may be affected by future climates.Item The relationship between physiological stress response and variation in omics data(Montana State University - Bozeman, College of Letters & Science, 2021) Steward, Katherine Fay; Chairperson, Graduate Committee: Brian Bothner; This is a manuscript style paper that includes co-authored chapters.Omics analysis is the cornerstone of systems biology. It offers comprehensive assessments of stress, interaction networks and connections to phenotype. Defining a stressed phenotype can be challenging, however, as stress response mechanisms can arise from a range of environmental conditions and experimental perturbations. Previous work from our lab noted the possibility of a relationship between stress in omics data and the variation of that data. This connection has yet to be clearly defined, and the cellular mechanisms responsible for the canalization of omics data remain a mystery. In this work I have taken advantage of the sensitivity of metabolomics and proteomics to detect cellular stress and characterize its relationship to variation. By utilizing coefficient of variation (CV) as a statistic of merit, the depth of the relationship between stress and variation can be uncovered. Once the model was clearly defined, a proteomics dataset with a large proportion of protein coverage was utilized to investigate what pathways might be responsible for the metabolite and protein canalization.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.