Master of Science / Department of Biology / David A. Haukos / Abstract–Duckling Survival Lesser scaup (scaup) populations have been experiencing continent-wide decline since the 1980s. It is important to have complete understanding of the critical factors influencing population change (e.g., duckling survival, nesting success, and health) to advance our understanding of population dynamics and improving species conservation. Duckling survival is a primary driver of scaup demography. I conducted a capture-mark-recapture study using Cormack-Jolly-Seber models in Program MARK to compute apparent daily survival and recapture probabilities for 3256 individually marked ducklings with 620 recaptures during 2010 to 2013. The most parsimonious survival model based on a priori hypotheses found that Julian hatch date squared was the most significant predictor of survival and consistent through all four years. Mass at hatch also was significant as a quadratic effect. Duckling survival to 30 days ranged from 29.0 to 80.0. During this study, stabilizing selection played a significant role in duckling survival, which indicates that there was trade-offs for selection of an optimal timing of hatch on survival and a cost associated with hatching too early or too late and being too heavy or to light. / Abstract–Nest-Site Selection There is a hierarchical process of behavioral and environmental processes that influence habitat selection, which inherently influences the survival and fitness of that individual and contributes to population growth. I investigated nest fate, spatial attributes, and all relationships between high and low-water levels with habitat attributes (distance to upland, distance to open water, nearest neighbor distance) of located nests using general linear models in SAS, t-tests in R, and Hot Spot Analysis in ArcGIS of 481 nests over eight years. In low-water years, successful nests (X̅ = 1153 m) were located 22.0% farther from upland than unsuccessful nests (X̅ = 944 m), but support for a similar relationship was lacking in high-water years. Successful nests were located 21.0% and 23.0% (i.e., 49 and 50 m) closer to conspecific nests than unsuccessful nests in low and high-water levels, respectively. In both high and low-water level years, clusters of nests initiated later in the season coincided with Hot Spots for nest fate (i.e., high-quality habitat patches, clusters of successful nests), whereas areas that tended to be selected first, evidenced by clusters of nests initiated earlier, tended to overlap with clusters of Cold Spots for nest fate. The core Hot Spot for nest fate was in the same spot in both water level conditions and located in flooded emergent vegetation in the heart of Lower Red Rock Lake furthest from any upland habitat. Three out of six Cold Spots for nest fate both in high and low-water years were located in emergent vegetation on the perimeter of Lower Red Rock Lake adjacent to uplands. Density-dependence seems to be a factor affecting late-nesting scaup females that are apparently cuing in on the reproductive performance of conspecifics when determining where to nest. Therefore, management actions focused on survival and reproductive success of scaup should consider managing water levels and habitat for later nesting scaup to increase adult survival and ultimately recruitment of ducklings. / Abstract – Blood Parasite Prevalence Blood parasites, per se, do not lead to direct mortality, but instead reduce the health of individual birds, which may ultimately lead to decreased reproductive success. Evidence has shown that presence of blood parasites can reduce fitness, body condition, and reproductive success of waterfowl. For many avian species, the cost of reproduction is manifested as a negative relationship between female breeding effort and breeding season survival, with trade-offs occurring when these adaptive choices become detrimental to future reproductive performance. Blood was drawn for parasite load determination from 112 individual adult scaup captured from 2011to 2012 via spotlighting and drive-trapping. Parasite prevalence was determined through blood assays that were created using a two-slide wedge technique. Relationships among seasonal heterophile:lymphocyte ratio (a proxy for health), body mass at time of capture (throughout pre-, during, and post-breeding periods), breeding status (females only), and Julian date of capture (date of capture) of capture with parasite prevalence were analyzed using linear (lm) regression models in R 2.15.2. The blood parasite infection rate was 5.0%, with prevalence differing by gender with 33.3% of males positive for blood parasites compared to 1.0% of females. The presence of blood parasites did not affect health, fitness, or breeding status of scaup. A quadratic relationship was found with body mass and date of capture, indicating that body mass increased from pre-breeding period to the breeding period and decreased significantly at the end of the summer during molt. A negative relationship between the heterophile:lymphocyte ratio of female scaup and date of capture (i.e., the health of scaup females was greatest during the pre-breeding period after which it consistently decreased until the molting period). A strong negative correlation between heterophile:lymphocyte ratio and body mass was found in both genders, which indicated that scaup in poor body condition were also in poor health at the end of the breeding season.

Doctor of Philosophy / Department of Biology / Samantha Wisely / Emerging infectious diseases play an increasingly critical role in many biological fields, including conservation biology and public health. Many emerging diseases originate in wildlife, most are caused by viruses, and often emergence is due to adaptation to and amplification in a new host, frequently in areas where ecological transformation is occurring. These emergence patterns suggest that the complex interactions among host, virus, and landscape drive disease emergence. Terrestrial rabies in striped skunks (Mephitis mephitis) in the central Great Plains is an excellent model system to investigate the interactions among the components of disease emergence: host ecology, pathogen properties, and landscape features. Striped skunks are not only numerous in the central Great Plains, they are also the reservoir for two genetically distinct rabies strains that co-occur in the region. Additionally, the landscape in the central Great Plains has undergone significant land use change over the last 70 years through increased urbanization and industrial agriculture practices. I used a combination of molecular and spatial techniques to investigate the interactions among host, pathogen, and landscape. Molecular epidemiology results indicated that rabies strains in the central Great Plains exhibit different epidemiological properties, while population genetic analyses indicated that striped skunks in the region are highly admixed and comprise a single population. Spatial analysis revealed that landscape features such as rivers are not a barrier to striped skunk dispersal, but differentially influence the movement of the two rabies strains. Because striped skunks are reservoirs for many diseases other than rabies and are ubiquitous throughout North America, I also examined the historical movements and distribution of striped skunks in North America using a phylogeographic approach. Results revealed that a combination of multiple Pleistocene dispersal events and Holocene admixture are responsible for the contemporary population structure of striped skunks in North America, and allowed me to place my regional-scale striped skunk rabies study into a larger biogeographic context. My results support the use of a holistic approach for studying emerging infectious diseases that includes studies of viral characteristics, host ecology and biogeography, and spatial features.

Bladder obstruction (neurogenic or anatomic) induces strain injury in detrusor smooth muscle cells. Signaling via strain injury in other systems has been highly studied, while in bladder obstruction, it has been quite limited to a small number of pathways. In our study we have examined the effects of strain injury using a combination of in vivo, ex vivo and in vitro models, with the aim of understanding disease pathogenesis in the bladder. Using a combination of literature searches, phospho-protein screens and pathway analysis, we uncovered three pathways activated by mechanical strain, ERK, STAT3 and mTOR, with potential for changing not only the way we understand but also the way we treat obstructive myopathies of the bladder. We found that not only were these pathways activated in response to strain and distension injury of BSMC, but they were also responsible for proliferation and sometimes de-differentiation. Included herein are three chapters, published in 2006 and 2010, on the role of ERK, STAT3 and mTOR pathways in bladder smooth muscle cell proliferation and differentiation, 8 Appendices containing the first pages of other papers and reviews published during the course of my studies.

Doctor of Philosophy / Division of Biology / Theodore Morgan / Temperature is a critical environmental parameter and thermal variation has significant effects on local adaptation and species distributions in nature. This is especially true for organisms that are isothermal with their environment. Variation in temperature imposes stress and directly influences physiology, behavior, and fitness. Thus, to thrive across a range of thermal environments populations must contain sufficient genetic variation, the capacity to respond plastically, or some combination of both genetic and plastic responses. In this work I first quantified patterns of phenotypic and genetic variation in nature and then dissected the genetic basis of variation in thermal traits. In the first aim I used natural populations of Drosophila melanogaster collected from a latitudinal transect in Argentina to investigate variation in heat stress resistance and cold plasticity within and among populations. I found heat stress resistance was highly variable within populations, but was strongly associated with the monthly maximum average temperature of each site. For cold plasticity I was able to demonstrate significant variation in plasticity within and among populations, however the among population variation was best explained by the altitude of each site. I hypothesized that this was caused by a difference in temperature fluctuations at high altitude sites relative to low altitude sites. To evaluate this hypothesis I paired our study with existing laboratory data that demonstrated significant fitness differences between high and low plasticity (and altitude) sites when these populations were reared in variable thermal environments. Thus, cold plasticity is an adaptive response to environmental variation. The final project focused on understanding the genetic basis of thermal variation. I fine-mapped a single co-localized heat and cold tolerance QTL via deficiency and mutant complementation mapping to identify four novel thermal candidate genes. There was no overlap of the deficiencies or genes associated with cold or heat stress resistance. Sequence analysis of each gene identified the polymorphisms that differentiate the lines. To test for independent associations between these polymorphisms and variation in nature the Drosophila Genome Reference Panel was used to confirm associations between allelic variation and cold tolerance in nature.

Master of Science / Department of Grain Science & Industry / Xiuzhi Susan Sun / Bio-based protein adhesives derived from renewable resources, especially soy protein are becoming more significant due to the concerns about environment and health related issues and the limit of petroleum recourses. However, the relatively poor water resistance of soy-based protein adhesives limits its wide applications. The goal of this study was to improve the water resistance performance of soy-based protein adhesives by chemical modification. The specific objectives are 1) to modify soy protein with calcium carbonate (CaCO[subscript]3) and sodium bisulfite (NaHSO[subscript]3) as the denaturing agents; 2) to investigate the effects of calcium carbonate (CaCO[subscript]3) concentrations, curing time and curing temperatures on adhesion performance of the modified soy-based protein adhesives. In this study, the co-effects of NaHSO[subscript]3 and CaCO[subscript]3 on adhesion properties of soy protein adhesives were investigated. NaHSO[subscript]3 was added to soy flour slurry at constant concentration 6g/L, while concentration of CaCO[subscript]3 was chosen in the range of 0 to 23g/L. Soy protein adhesives modified with 4g/L and 16g/L CaCO[subscript]3 were selected to characterize the adhesion performance on 3 ply yellow pine plywood using the Response Surface Method (RSM). The effects of curing temperature and curing time on the adhesion properties were also studied. The major findings are 1) 4g/L CaCO[subscript]3, 6g/L NaHSO[subscript]3 modified soy protein adhesives (MSPA) had better adhesion performance (both dry and wet) than 16g/L CaCO[subscript]3, 6g/L NaHSO[subscript]3 MSPA; 2) Higher temperature (170°C) resulted in higher wet shear adhesion strengths; 3) Longer hot press time had positive impact on wet adhesion shear strength; and 4) 4g/L CaCO[subscript]3, 6g/L NaHSO[subscript]3 MSPA showed better adhesion shear strength after 2 weeks storage than 16g/L CaCO[subscript]3, 6g/L NaHSO[subscript]3 MSPA. In general, 4g/L CaCO[subscript]3, 6g/L NaHSO[subscript]3 MSPA, under longer hot press time and higher temperature would lead to a better adhesion performance.

Doctor of Philosophy / Division of Biology / Carolyn J. Ferguson / The genus Lappula Moench is a diverse group of herbaceous plants in the large, cosmopolitan family Boraginaceae. Over sixty species are recognized in Eurasia; many are found in cold deserts, steppes, and semi-deserts of Central Asia. Fewer species were described from western North America, and compared with the Asian species they are poorly known. Various North American taxa have been placed into synonomy under Eurasian species, and complex patterns of variation have made species circumscription challenging. The goal of this dissertation was to explore phylogenetic relationships between North American and Eurasian species and to revise the taxonomy of the North American species. A molecular phylogenetic study was initiated in order to infer patterns of relationships among the North American species relative to Eurasian diversity. Samples were collected from throughout the western United States and from Siberia. Additional samples of Eurasian species were taken from herbarium specimens. Sequences were generated for three DNA regions (the ITS region of the nuclear ribosomal DNA, and chloroplast intergenic spacers trnS-trnG and rpl32-trnL) and phylogenies were generated using parsimony and Bayesian analyses. Results were in general agreement among all analyses. The genus Lappula was recovered as a monophyletic group, with the exception of the morphologically anomalous L. sessiliflora (Boiss.) Gürke, which was sister to the genus Rochelia Rchb. The native North American species of Lappula and L. redowskii (Hornem.) Greene formed a clade. Samples of L. squarrosa (Retz.) Dumort., a Eurasian steppe plant with a wide introduced range in North America, grouped with samples from Eurasia. Sampled species of Hackelia Opiz, a genus sometimes treated as part of Lappula, formed a clade separate from the Lappula species sampled here. Herbarium and field studies resulted in a revised taxonomy for the North American Lappula. Nomenclatural problems were resolved; nine native species and four varieties were recognized along with the introduced Eurasian species L. squarrosa.

Master of Science / Department of Biology / Theodore J. Morgan / The organismal response to temperature represents one of the most ubiquitous processes that occur in the natural world, and this response is critical for survival in most habitats. Increased attention should be focused on how organisms cope with temperature extremes, either through adaptation, plasticity, or a combination of both, as climate models predict increased variations in temperature accompanied by novel thermal extremes. Drosophila melanogaster is an excellent resource for answering questions pertaining to how organisms persist in environmental extremes because they originated in central tropical Africa and have since colonized nearly the entire globe, exposing them to many novel thermal stressors. In this work I elucidated regions of the genome contributing to phenotypic variation in cold tolerance and thermal plasticity. A quantitative trait locus (QTL) approach was used, which involved phenotyping roughly 400 recombinant inbred lines (RILs) of D. melanogaster from the Drosophila Synthetic Population Resource (DSPR). The DSPR captures genetic variation from around the globe, allowing for precision mapping of cold tolerance and thermal plasticity QTL, while simultaneously determining the frequency of the QTL alleles. Upon development at both 18°C and 25°C, RILS were measured for a common cold tolerance metric, chill-coma recovery time (CCR), and a plasticity value was derived as the change in CCR between environments. Analysis of variance revealed significant effects of sex, line (RIL), treatment (temperature), and line by treatment interaction (GxE). Mapped QTL for chill-coma recovery time at 18°C and 25°C spanned the same regions as several studies previously reported, validating the automated phenotyping method used and the mapping power of the DSPR. QTL between CCR at 18°C and 25°C overlapped significantly, and QTL for thermal plasticity shared the similar regions as QTL for CCR, but also exhibited two non-overlapping QTL on the left arm of the third chromosome. This study demonstrated the tremendous amount of variation present in cold tolerance phenotypes and identified candidate regions of the genome that contribute to thermal plasticity and require further investigation.

Master of Science / Department of Biology / Kathrin Schrick / Steryl glucosides (SGs) are a common conjugate of sterols found in the plasma membranes of most plants and fungi, but their cellular functions remain largely unknown. Glycosylation of the C3 hydroxyl group of the sterol nucleus is catalyzed by UDP-glucose:steryl glucosyltransferase 80 (UGT80) enzymes. Two genes encoding UGT80A2 and UGT80B1 are responsible for most SG production in Arabidopsis thaliana, while UGT80C1 presents a putative third enzyme. In Arabidopsis, seed imbibition signals the epidermal seed coat cells to secrete an encapsulating mucilage that consists primarily of hydrated polysaccharides. Cellulose has been identified in the inner layer of the mucilage, providing a convenient model to study cellulose synthesis since seed mucilage is dispensable for viability and pectin and cellulose staining dyes are readily available. A reverse genetics and biochemical approach was used to characterize the role of UGT80 enzymes and their impact on cellulose synthesis in seed mucilage. ugt80B1 mucilage was found to have elongated cellulosic rays, but no defects in pectin synthesis. A double mutant of ugt80B1 and mum3-1, a mutant allele of CELLULOSE SYNTHASE 5 (CESA5), displays a novel phenotype with irregular cellulose patterning and extreme shedding of the pectinaceous layer surrounding the seed coat. The observed mucilage defects may be indicative of disrupted cellulose synthesis and a mechanistic relationship between SGs and the cellulose synthase machinery. UGT80A2 and B1 demonstrate glycosylation activity with a multitude of plant sterols. The two enzymes do display some substrate specificity, however, with UGT80A2 producing the large majority of sitosterol and stigmasterol glucoside compared to B1. UGT80C1 shows little or no sterol glucosyltransferase activity in vitro or in vivo and likely has evolved a different function from the two other genes. GFP:UGT80C1 expressed either from the constitutive 35S or from its native promoter was localized to lipid droplets and possibly chloroplasts as well, creating a new perspective on the role of the protein in plant lipid metabolism. This study extends the currently limited view of SGs as ubiquitous components of the plasma membrane to active regulators of cellulose synthesis in seeds. Evidence presented here changes the perceived role of the plant conserved protein, UGT80C1, from a putative steryl glucosyltransferase enzyme to having a function in intracellular lipid droplets.

Master of Science / Department of Anatomy and Physiology / Lei Wang / Jishu Shi / Interleukin-1 beta (IL-1β) is an important mediator of inflammatory response, and the elevated expression of IL-1β is correlated with tumor growth and metastasis. Epithelial-mesenchymal transition (EMT) is a reversible transition between epithelial phenotype and mesenchymal phenotype. Usually, EMT can be identified by its unique morphology change and expression of EMT markers. In our study, we have found after treated HCT-116, a colon cancer cell line, and human primary colon cancer cells with IL-1β, cells began to display mesenchymal phenotype with highly down-regulated E-cadherin expression and up-regulated ZEB factors expression. For colon cancer cells, sphere formation assay in serum free medium (SFM) with the presence of growth factors is used to identify cancer stem cell population. We have shown that IL-1β can induce colon cancer stem cell proliferation and express stem cell markers (Bmi1, Nanog, and Nestin). In addition, besides the stem cell markers, we also found ZEB factors were highly up-regulated in spheroid cells as well. We silenced Zeb1 expression and investigated the effect of IL-1β on shZeb1 HCT-116 cells. The results indicated Zeb1 knockdown not only inhibited IL-1β-induced EMT but also reduced proliferation of spheroid cells and inhibited Bmi1 expression. Therefore, ZEB factors must play an important role in both EMT process and cancer stem cell development. From our data, we conclude that IL-1β promotes epithelial-mesenchymal transition and a stem cell phenotype in colon cancer via ZEB factors.

Doctor of Philosophy / Department of Biology / Michael Herman / Soil nematode communities are important components of the micro fauna in grassland ecosystems and their interaction with soil microbes affects important ecological processes such as decomposition and nutrient recycling. To study genetic mechanisms underlying ecologically important traits involved in the response of nematode communities to soil microbes, we employed genomic tools available for the model nematode, Caenorhabditis elegans. Previous work identified 204 C. elegans genes that were differentially expressed in response to growth on four different bacteria: Bacillus megaterium, Pseudomonas sp., Micrococcus luteus and Escherichia coli. For many of the genes the degree of differential gene expression between two bacterial environments predicted the magnitude of the effect of the loss of gene function on life-history traits in those environments. Mutations can have differential effects on fitness in variable environments, which can influence their maintenance in a population. Our fitness assays revealed that bacterial environments had varying magnitude of stress, defined as an environment in which the wild-type has a relatively low fitness. We performed fitness assays as part of a comprehensive analysis of life history traits on thirty five strains that contained mutations in genes involved in the C. elegans response to E. coli, B. megaterium, Pseudomonas sp. We found that many of the mutations had conditionally beneficial effects and led to increased fitness when nematodes bearing them were exposed to stressful bacteria. We compared the relative fitness of strains bearing these mutations across bacterial environments and found that the deleterious effects of many mutations were alleviated in the presence of stressful bacteria. Although transcriptional profiling studies can identify genes that are differentially regulated in response to environmental stimuli, how the expressed genes provide functional specificity to a particular environment remains largely unknown. We focused on defense and metabolism genes involved in C. elegans-bacterial interactions and measured the survivorship of loss-of-function mutants in these genes exposed to different bacteria. We found that genes had both bacteria-specific and bacteria-shared responses. We then analyzed double mutant strains and found bacteria-specific genetic interaction effects. Plasticity in gene interactions and their environment-specific modulation have important implications for host phenotypic differentiation and adaptation to changing environments.