Life Sciences Commons^™

Developing Regulated Crispr Systems To Control Bacterial Microbiomes, Gregory M. Pellegrino

Electronic Thesis and Dissertation Repository

Changes to the human microbiome’s composition and metabolome are associated with numerous diseases and alterations to xenobiotic metabolism. As such, targeting the human microbiome is an increasingly popular option for therapeutic interventions. However, traditional therapies that target the microbiome such as antibiotics lack specificity, which can affect the beneficial species of the microbiome and cause adverse health outcomes such as the rise of antimicrobial-resistant bacteria. Therefore, the research and development of specific, targeted antimicrobial therapies is crucial to effectively treating microbiome dysbioses.
CRISPR and CRISPRi provide easily modifiable, RNA-guided mechanisms mediated by the Cas9 or dCas9 enzymes to induce sequence-specific …

Adaptation Reshapes The Distribution Of Fitness Effects, Diego Tenoch Morales Lopez

Electronic Thesis and Dissertation Repository

The process of adaptation has been of interest since the XIX century, when Darwin first proposed the idea of natural selection. Since then, there has been a myriad of theoretical and empirical works that have expanded the field. From the many evolutionary insights these works have produced, a foundational idea is that spontaneous mutations in the genome of organisms can produce changes to their reproductive success that might confer an advantage for the mutant organisms with respect to their peers. Therefore, mutations drive adaptive evolution by virtue of their heritable effects on fitness. Empirical measures of the distribution of these …

Investigating The Soil Mycobiome Of American Ginseng, Rachel Amelia Rajsp

Electronic Thesis and Dissertation Repository

American ginseng (Panax quinquefolius L.) is a perennial herbaceous plant cultivated for its medicinal properties. Growers report that when ginseng is replanted in a field previously used to cultivate ginseng, it soon succumbs to disease, known as ginseng replant disease. I examined changes in composition in the ginseng mycobiome throughout cultivation in a newly planted garden and beyond (i.e., 3–14 years post-harvest) with a third-generation metabarcoding approach (Pacific Biosciences, single-molecule real-time sequencing). Amplicons of about 600 nucleotides from the nuclear ribosomal internal transcribed spacer (ITS) region and translation elongation factor 1-α gene were chosen to help discriminate between closely …

Freezing Tolerance Of Herbaceous Legumes Within Southwestern Ontario: Evidence Of Disproportionate Freezing Sensitivity, Samuel L. Rycroft

Electronic Thesis and Dissertation Repository

Legumes (Fabaceae) represent a diverse and ecologically significant plant taxon; most described legumes form mutualisms with diazotrophic rhizobia, potentially fixing substantial quantities of nitrogen within habitats where they are well-established. Stressors causing lethal or sub-lethal impacts in legumes or rhizobial symbionts may therefore impact the nitrogen dynamics of such habitats. In recent decades, variability of winter temperatures, precipitation, and soil freeze-thaw cycling has increased in temperate regions. Without adequate snow cover to insulate roots and shoot bases, herbaceous plants will likely be exposed to more frequent or severe freezing. In southwestern Ontario, a pattern of disproportionate freezing sensitivity relative to …

Sufu In Shh Signalling Mediated Myogenesis, Suleyman Abdullah

Electronic Thesis and Dissertation Repository

Myogenesis is defined as the formation of skeletal muscle tissue during embryonic development and involves a multitude of cellular signalling pathways. Among these include the Sonic hedgehog (Shh) signalling pathway which must be deactivated for differentiation into muscle cells to occur. However, less is known regarding the pathways operation during cell differentiation and whether Suppressor of Fused (SUFU), the protein inhibitor of Shh signalling, plays a role. To address this, mouse C2C12 myoblast cells were utilized as a model and differentiated into muscle cells to identify the presence of SUFU during this time. Experiments in qRT-PCR show a decrease in …

Elucidating The Relationships Between Spider Size, Joint Stiffness, And The Mechanical Frequency Response Of The Body, Reese L. Gartly

Electronic Thesis and Dissertation Repository

Spiders use vibrations to sense their surroundings. It has been suggested that the vibration perception in spiders may be altered by the mechanics of the body. I studied the biomechanics of spiders, at the level of leg joints and the whole body. To study joints, I quantified the allometry of leg joint stiffness in spiders. I found that the stiffness of spider joints increased nearly isometrically with increasing body mass, partly by having shorter and thicker leg segments and also by other unknown means. Using these data, I developed empirically validated biomechanical models which predicted the effects of mechanics on …

Sex Comparisons Of External, Anatomical And Methodological Factors On Motor Unit Firing Behavior, Michael James Marsala

Electronic Thesis and Dissertation Repository

Motor unit (MU) recruitment and firing rate (MUFR) modulation are the primary factors governing force control. Differences in the anatomical distributions of α-motoneurons, muscle properties, and perception of fatigue between males and females may cause sexually dimorphic MU firing behaviors. The purpose of this dissertation was to further examine external, anatomical, and methodological factors that may influence observations of sex-related differences in neuromuscular function. In chapter 2, sex differences in how mental fatigue may influence MU firing behaviors were examined. Mental fatigue did not significantly alter MU firing behavior, maximal force, or force tracing ability in either males or females. …

Characterizing The Spatial Distribution Of Inhibitory Interneurons Across Early Sensory And Association Areas In Callithrix Jacchus, Nika Khajehdehi

Electronic Thesis and Dissertation Repository

The prefrontal cortex (PFC) is one of the cortical areas responsible for complex cognitive abilities, a function that is believed to arise from increased persistent activity within its microcircuits. Activity within microcircuits is regulated by parvalbumin-containing (PV), calbindin-containing (CB), and calretinin-containing (CR) inhibitory interneurons (INs). It remains unclear how the distribution of activity-regulating INs differs across cortical areas such that persistent activity increases specifically within association areas, allowing for their complex functions. This thesis aims to address this gap by characterizing the spatial distributions and differences in relative proportions of INs across early sensory areas and association areas of the …

The Introduction Of Nursery Seedlings And Their Fungi To A Spruce-Fir Forest In Newfoundland, Alicia G. Banwell

Electronic Thesis and Dissertation Repository

A few species of boreal trees form symbiotic relationships with many ectomycorrhizal (ECM) fungi that allow trees to thrive on nutrient poor soils in Canada. A small number of ECM species grow on tree seedlings raised in nurseries for reforestation of clearcut sites. The native ECM fungal community composition of a mixed spruce-fir forest in Newfoundland was determined through next generation sequencing. With the introduction of nursery seedlings, the transfer of native ECM fungi to seedlings and nursery-established ECM fungi to roots was investigated over 20 months. ECM community composition was found to be similar to that of other boreal …

Investigating Task-Free Functional Connectivity Patterns In Newborns Using Functional Near-Infrared Spectroscopy, Homa Vahidi

Electronic Thesis and Dissertation Repository

Robust functional connectivity in sensorimotor resting-state network (RSN) has been linked to positive neurodevelopmental outcomes in neonates. In the current study, we aimed to map the developmental trajectory of sensorimotor RSN in awake neonates using functional near-infrared spectroscopy (fNIRS). We acquired fNIRS resting-state data from 41 healthy newborns (17 females, gestational age range=36+0 to 42+1 weeks) within the first 48 hours after birth. At the group level, we observed robust positive connectivity in numerous channel-pairs across the sensorimotor network, especially in the left hemisphere. Next, we examined the relationship between functional connectivity, gestational age and postnatal age, while controlling for …

Differential Migration Timing And The Form And Function Of Avian Wings, Jessica E. Deakin

Electronic Thesis and Dissertation Repository

A common differential migration strategy in birds is protandry, whereby males arrive earlier than females. The probable causes of protandry are well studied from the perspective of innate and physical environmental cues, but the influence of the social environment and wing morphology are less known. Theoretical models propose that sex ratio influences protandry; male-biased sex ratios are predicted to advance timing in males due to increased intrasexual competition. To empirically test this, I investigated the spring migration traits of male Yellow-rumped Warblers Setophaga coronata under differing sex ratios. An integrated automated telemetry approach was used, where locomotor movement of captive …

The Adaptor Protein P66shc Governs Central Nervous System Cell Metabolism And Resistance To Aβ Toxicity, Asad Lone

Electronic Thesis and Dissertation Repository

Alzheimer’s disease (AD), a progressive and irreversible neurodegenerative disorder, and is the leading cause of dementia worldwide. It has been posited that AD is caused by the gradual deposition of toxic amyloid-b (Ab) plaques in the brain- that cause oxidative stress and eventually leads to neuronal death and synaptic loss. However, multiple therapies that either interfere with the production, or enhance the removal of Ab from the brain, have ultimately failed to slow or prevent AD. With the ever-increasing burden of AD worldwide, there exists an urgent need for novel therapeutic targets. The adult human brain is an energy demanding …

Neural Responses To Magnetic Orientation Information In Songbirds, Madeleine Ir Brodbeck

Electronic Thesis and Dissertation Repository

Effective navigation and orientation is essential for animals to survive. The study of migratory birds provides insights into these processes, particularly through mechanisms like the geomagnetic compass, which uses Earth's magnetic field for directional information. A brain region in songbirds relevant to this is cluster N, known for its potential role in processing geomagnetic information. Notably, cluster N appears exclusive to nocturnally migratory birds, being active solely at night, and lesion studies reveal that an intact cluster N is necessary for geomagnetic compass orientation. However, given the scarcity of empirical data concerning cluster N, substantial questions persist regarding its function. …

Identification Of Early Diagnostic And Predictive Biomarkers Of Cisplatin-Induced Acute Kidney Injury Using Metabolomics, Yong Jin Lim

Electronic Thesis and Dissertation Repository

Cisplatin is an effective chemotherapeutic agent used for the treatment of a wide variety of solid tumors and hematologic cancers. Despite its effectiveness, cisplatin is associated with several toxic effects to non-cancerous tissues. Nephrotoxicity is the main dose-limiting toxicity in cisplatin therapy, manifesting as acute kidney injury (AKI) in approximately one-third of patients receiving cisplatin. AKI is defined as an abrupt decline in kidney function and is associated with several short- and long-term adverse outcomes, including chronic kidney disease, cardiovascular disease, and mortality. AKI is currently diagnosed by increased serum creatinine (SCr) concentrations. However, SCr is a marker of functional …

Acetylation Regulates Thioredoxin Reductase Activity And Oligomerization, David E. Wright

Electronic Thesis and Dissertation Repository

The Thioredoxin (Trx) system provides the cell with robust defense against oxidative stress and regulates the function of nearly every cellular process through the reduction-oxidation (redox) regulation of proteins. The Trx system is involved in the development of many diseases ranging from cancer to cardiovascular disorders. Thioredoxin reductase (TrxR) is the key enzyme in the Trx system and contains the 21^st genetically encoded amino acid, selenocysteine (Sec). There were multiple experimentally identified TrxR acetylation sites with an unknown effect on TrxR activity. My thesis tested the hypothesis that programmed protein acetylation will enhance the activity of TrxR1. I used …

Aging Effects On The Motivational Consequences Of Unsuccessful Memory Recall, Rachel Sargeson

Electronic Thesis and Dissertation Repository

The present thesis addressed whether experiences of failed recall for names of familiar faces could induce states of curiosity and drive memory benefits. Experiment 1 investigated whether older adults exhibit the same familiarity preference in information seeking, following an unsuccessful recall attempt for names of previously studied faces, as seen in younger adults. Experiment 2 considered whether acting on the curiosity induced by unsuccessful recall of names associated with familiar faces could provide benefits for the relearning of those names in both age groups. The older adults displayed a similar, if not more pronounced, positive relationship between familiarity and subsequent …

Assessing Copd Care Quality In A Rural Ontario Primary Care Clinic: A Retrospective Chart Review, Daniel Henke

Electronic Thesis and Dissertation Repository

Background: Primary chronic obstructive pulmonary disease (COPD) care may be worse in rural versus more urban environments.

Purpose: To evaluate the quality of COPD care in a rural Ontario primary care clinic.

Methods: A 12-month retrospective chart review study was conducted between June 2022 and January 2023 at a Goderich, Ontario primary care clinic. Electronic medical records (EMRs) were randomly selected for inclusion. Baseline sociodemographic (e.g., occupation) and health (e.g., smoking status) characteristics were extracted as well as information regarding COPD care received (i.e., Health Quality Ontario [HQO] Quality Standard [QS] indicators; n=33. The primary study outcome was overall proportion …

Kinetic Analyses Of Sacas9[D10e] In Vitro, Claire Zhang

Electronic Thesis and Dissertation Repository

CRISPR is a well-known adaptive defense mechanism that gained attention through its ability to be easily reprogrammable. Over time, the native CRISPR-Cas system evolved to tolerate mismatches, broadening its cleavage preferences; however, this development poses concerns with off-target cleavage in gene editing. Here, we introduced a D10E mutation into SaCas9 to potentially reduce off-target cleavage. To characterize SaCas9[D10E] and its tolerance for mutations, we designed 21 different substrates that each contained a single transversion in a therapeutically relevant EMX1-1gene. Through a series of in vitro cleavage assays, SaCas9[WT] and SaCas9[D10E] were compared across these substrates. A kinetic analysis of SaCas9[D10E] …

Investigating Soil Microbiome Changes During Panax Quinquefolius Cultivation In New Gardens, Pok Man Paul Wan

Electronic Thesis and Dissertation Repository

Ginseng (Panax quinquefolius) is susceptible to ginseng replant disease (GRD), resulting in root rot and decreased yield when ginseng is grown in soils previously used for ginseng crops. Although fungal and oomycete pathogens are implicated, GRD is a complex disease and the changes in the soil when initially cultivating ginseng that lead to GRD remain unclear. This thesis reported changes in the soil microbiome from three new gardens in Norfolk County, Ontario over three years starting from seeding with ginseng. Over this period of cultivation, metabarcoding of the V4 region of the 16S ribosomal RNA gene clearly showed …

Invariant Object Recognition In Deep Neural Networks And Humans, Haider Al-Tahan

Electronic Thesis and Dissertation Repository

Invariant object recognition, a cornerstone of human vision, enables recognizing objects despite variations in rotations, positions, and scales. To emulate human-like generalization across object transformations, computational models must perform well in this aspect. Deep neural networks (DNNs) are popular models for human ventral visual stream processing, though their alignment with human performance remains inconsistent. We examine object recognition across transformations in human adults and pretrained feedforward DNNs. DNNs are grouped in model families by architecture, visual diet, and learning goal. We focus on object rotation in depth, and observe that object recognition performance is better preserved in humans than in …

Short-Term Versus Long-Term Effects Of Nitrogen Addition And Warming On Ecosystem N Dynamics In A Grass-Dominated Temperate Old Field, Benjamin F. A. Souriol

Electronic Thesis and Dissertation Repository

Increased atmospheric nitrogen (N) deposition and climate warming are both anticipated to influence the ecosystem N dynamics of northern temperate ecosystems substantially over the next century. Nevertheless, in field experiments with N addition and warming treatments, temporal scale can play an important role in determining the extent of treatment effects on N dynamics, and it is unclear to what extent the results of short-term studies can be extrapolated to responses over longer time scales. I compared the short-term versus long-term effects of N addition and warming on net N mineralization, N leaching, and N retention in a grass-dominated old field. …

Cortical Activation During Mobility In An Indoor Real-World Environment: A Mobile Eeg Study, Sam Marshall

Electronic Thesis and Dissertation Repository

Human mobility requires neurocognitive inputs to safely navigate the environment. Previous research has examined neural processes that underly walking using mobile neuroimaging technologies, yet few studies have incorporated true real-world methods without a specific task imposed on participants (e.g., dual-task, motor demands). The present study utilized mobile electroencephalography to examine and compare theta, alpha, and beta frequency band power (μV2) in young adults during sitting and walking in laboratory and real-world environments. Our findings support that mobility and environment may modulate neural activity, as we observed increased brain activation for walking compared to sitting, and for real-world walking compared to …

Smartphones For Smart Eating: Evaluating The Effectiveness Of Smartphone Applications On Improving Adolescent Food Knowledge, Heather A. Jantzi

Electronic Thesis and Dissertation Repository

Many Canadian adolescents have unhealthy eating habits which are risk factors for developing chronic diseases later in life. Food literacy plays a role in healthy eating habits but teens in Ontario currently lack sufficient nutrition education learning opportunities in home and school environments. Across health research, smartphones have been utilized as a novel medium to promote learning and alter behaviour. This thesis seeks to understand the effectiveness of smartphone applications at improving food knowledge in adolescents through (i) a systematic review of published interventions, and (ii) by investigating the efficacy of a novel location-based smartphone application. Findings from this thesis …

Characterizing Sacas9 And Sacas9[D10e] Tolerance To Mismatches Using Directed Evolution Using Fluorescence Activated Cell Sorting (Deufacs), Olha Haydaychuk

Electronic Thesis and Dissertation Repository

CRISPR-Cas9 nucleases are commonly used for genome editing but are limited by their off- target effects, which must be reduced for therapeutic use. Current strategies for engineering higher fidelity variants do not directly compare on- versus off-target activity. I establish Directed Evolution Using Fluorescence Activated Cell Sorting (DEUFACS), a bacterial dual- fluorophore system for characterizing and altering the sequence preferences of nucleases. I aimed to use DEUFACS to identify SaCas9 variants with increased on-target and decreased off-target activity. I found that D10E substitution in the RuvC domain of SaCas9 improved discrimination and was superior to other RuvC active site variants …

Structural Control Of Metallothionein Metallation And Oxidation, Natalie C. Korkola

Electronic Thesis and Dissertation Repository

Metallothioneins (MT) are a family of cysteine-rich metal-binding intrinsically disordered proteins that are ubiquitous across life. The proposed functions of MTs are to maintain Zn(II) homeostasis, participate in heavy metal detoxification, and protect against oxidative stress by binding to a variety of metals in a range of stoichiometries. However, due to their disordered nature and ability to form multiple metallated species, these diverse structures are not well characterized. Electrospray ionization mass spectrometry (ESI-MS) is a technique that allows quantification of heterogeneous metallation states through analysis of the speciation distributions. In this thesis, ESI-MS in combination with other spectroscopic techniques is …

Mechanism Of Permissive Cleavage Activity Of Tevcas12a, Alexa White

Electronic Thesis and Dissertation Repository

We created a novel dual endonuclease called TevCas12a. TevCas12a is a fusion of the I-TevI nuclease and linker domains to Cas12a. The goal was to create a dual endonuclease that made precise double-strand breaks leaving non-compatible ends for exogenous DNA insertion and specific deletions. TevCas12a was expected to make two double-strand breaks, one by I-TevI and the other from Cas12a. However, when tested in vitro, TevCas12a cut the DNA substrate multiple times, rapidly degrading the substrate. We call this activity permissive cleavage. TevCas12a activity is guide RNA dependent, but not guide RNA sequence specific, it can degrade any DNA substrate …

Multi-Omics Analysis Of Epithelial Ovarian Cancer Metastasis, Mallory I. Frederick

Electronic Thesis and Dissertation Repository

Epithelial ovarian cancer (EOC) is a high-risk cancer with heterogeneous tumors. The high incidence of EOC metastasis from primary tumors to nearby tissues and organs is a major driver of EOC lethality. The phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT1) pathway regulates cell survival and is over-activated in most human cancers, including EOC. Growth factor stimulation activates AKT1 by phosphorylation at T308 and S473. In my cell-based metastasis model, adherent cells model primary tumors, spheroids represent metastatic spread, and re-adherent spheroid cells represent secondary tumors. In spheroids, I show reduced S473 and T308 phosphorylation of AKT, along with significantly increased levels of …

Characterization Of Dna Regulatory Elements Of The Highly Abundant Secreted Protein 1 (Hasp1) Promoter And Analysis Of Transcriptomic Responses Under Phosphate Depletion In The Diatom Phaeodactylum Tricornutum, Mahsa Farmanbar

Electronic Thesis and Dissertation Repository

Phosphorus (P) is essential for all life. The bioavailability of phosphorus in oceans impacts diatoms like Phaeodactylum tricornutum (P. tricornutum). P. tricornutum is extensively used in transcriptomic studies to understand pathways involved in P-acquisition. However, activation mechanisms and roles of regulatory elements in P-acquisition responses remains unclear. Here, I deleted predicted phosphate regulatory sites in P.tricornutum’s HASP1 promoter to create different HASP1-eGFP constructs. Under P-depletion, two constructs showed increased eGFP secretion. Additionally, a HASP1 knockout strain was grown under different phosphorus sources to determine if HASP1 is a phytase. Cells grown in full phosphorus utilized organic …

Protein Stability In Solution And In The Gas Phase., Yousef Haidar

Electronic Thesis and Dissertation Repository

Electrospray Ionization mass spectrometry (ESI-MS) is widely used for probing proteins, yet many aspects of this technique remain elusive. Using MS, ion mobility spectrometry (IMS), and circular dichroism (CD) spectroscopy, this thesis sheds light on the stability differences of proteins in the gas phase and solution. After a general introduction (Chapter 1), Chapter 2 scrutinizes some aspects of native ESI. Our data highlight the significance of cone voltage in maintaining a native-like fold and show the advantage of using NH₄Ac in protein experiments. Chapter 3 focuses on hydrogen/deuterium exchange (HDX)-MS. Several studies have reported that D₂O …

Selective Recruitment Of Cerebellum In Cognition, Ladan Shahshahani

Electronic Thesis and Dissertation Repository

Previous studies of cerebellar function in humans have shown that it is activated by a myriad of tasks ranging from motor learning and language to working memory and more. These studies have prompted a deviation from the traditional view of the cerebellum as a purely motor structure. However, the precise contribution of the cerebellum to these tasks remains ambiguous.

A prevalent assumption in fMRI studies is interpreting BOLD activation as evidence of the cerebellum's involvement in specific tasks. However, this interpretation is potentially misleading, especially considering that the BOLD signal predominantly represents cerebellar input, with output activity largely absent. Consequently, …