VictorJuiz/thesis huggingface dataset:
Revealing Phylogenetic Relationships and CRISPR-Cas Functionality in Legionella pneumophila using Next Generation Sequencing

Legionella pneumophila is a Gram-negative intracellular pathogen of diverse protozoan hosts in the aquatic environment. The generalist evolutionary strategy enables the bacterium to also replicate within human macrophages and cause Legionnaires' disease. The environmental challenges that shape L. pneumophila evolution, however, had not been well characterized. In my first data chapter, I employed next generation sequencing approaches to reveal the evolutionary relationships of L. pneumophila isolates. Two laboratory lineages were domesticated from the clinical strain collected during the eponymous 1976 Philadelphia outbreak of Legionnaires' disease. Through whole genome sequencing, I reconstructed the phylogenies of these commonly used laboratory strains and showed that certain laboratory manipulations would render both desired and unexpected mutations. Next, I characterized the genomes of a 2005 Toronto outbreak strain and several local strains of a similar genotype. Using comparative genomics and plasmid transformation, I demonstrated for the first time an active genome defense role of CRISPR-Cas systems in L. pneumophila. CRISPR-Cas is known as an adaptive immune system in prokaryotes against mobile genetic elements. Using CRISPR-Cas as a window into the environmental challenges to the bacterium, I revealed that L. pneumophila employs multiple distinct CRISPR-Cas systems to protect against a recurrent invasive DNA termed LME-1. To combat the mutational escape of mobile genetic elements, CRISPR-Cas can rapidly renew its immunological arsenal by acquiring secondary spacers through a priming process. In my last data chapter, using deep sequencing approaches I characterized the molecular patterns of primed spacer acquisition in L. pneumophila type I-C CRISPR-Cas. My examination of spacer dynamics suggested a close interplay between CRISPR-Cas interference and adaptation machineries under priming. Taken together, my thesis provided insights into L. pneumophila adaptations in the laboratory and to the environmental life style that involves generalist parasitism and extensive horizontal gene transfers.

Keywords: comparative genomics
CRISPR-Cas
Legionella
mobile genetic elements
next-generation sequencing
Advisor: Ensminger, Alexander W
The Role of Environmental Cues in Gamma-Delta T-Cell Lineage Choice and Functional Programming

The thymic landscape provides an array of spatially and temporally controlled cues that guide the development of functionally distinct effector immune cells. This thesis focuses on how signals downstream of the T cell receptor (TCR), Notch and critical cytokine receptors, dictate lineage choice between αβ and γδ T cells and, furthermore, can combine to play a role in programming γδ T cell effector function. The signal strength hypothesis posits that the quantity of the signal downstream of the TCR can direct lineage choice through the variable accumulation of inhibitor of DNA (deoxyribonucleic acid) binding 3 (Id3) proteins downstream of the TCR-induced ERK (extracellular signal-regulated kinase)- early growth response (Egr) pathway. To test this hypothesis, we assessed the differentiation outcome of Rag2-/- (Recombination-activating gene) DN3 (double negative 3) progenitors transduced to co-express both TCR and γδ TCRs, followed by co-culture with cells expressing a weak or strong binding γδ TCR ligand in the presence or absence of Notch ligands. In the presence of Notch and weak γδ ligands, a percentage of progenitors expressing only the γδ TCR matured towards the CD4+ CD8+ double positive (DP) stage of T cell development. Importantly, progenitors co-expressing TCR and γδ TCR cultured under the same conditions developed towards the γδ lineage as a result of higher accumulation of Id3 downstream of each of the two receptors, hence lending further support for the signal strength hypothesis. With this in mind, we set out to describe the role of these and other signals necessary for the development of functional γδ T cell subsets as defined by differential cytokine production. To this end, Rag2-/- DN3 progenitors were transduced with a transgenic KN6 γδ TCR, and co-cultured on various stromal cells engineered to express Notch, KN6 TCR ligands and supplemented with combinations of cytokines, such as IL-1β (Interluekin-1 β), IL-7, IL-21 and IL-23. Our results showed that specific combinations of these signals are required to program IFN-γ (Interferon-γ), IL-17, and IL-22 producing KN6 γδ T cell subsets. More specifically, I established that weak but not absent TCR-ligand interactions are required for γδ17 development, and Notch enables the survival of γδ T cells in an apparent inflammatory milieu. This was shown to be relevant in the fetal thymus as blockade of IL-1β, IL-21 and IL-23 in hanging drop cultures resulted in the absence of γδ17 cells. This work provides a framework for the integration of signals downstream of the Notch, TCR, and cytokine receptors as they lead to the development of γδ T cell functional subsets that can play a fundamental role in regulating health and disease.

Keywords: cytokines
gamma delta t cell
IL-17
regenerative medicine
t cell development
t cells
Advisor: Zúñiga-Pflücker, Juan Carlos
Ecosystem Functioning in the Real World

Ecosystem functions are components of an ecosystem that change over time. We rely on many of these functions, such as carbon or water cycling, for survival. For over twenty years, ecologists have been vigorously testing the effects of biodiversity loss on ecosystem functioning. These experiments have nearly all been done either in growth chambers or carefully cultivated field communities. As ecologists frequently acknowledge, it remains to be determined how accurately the results of these studies portray the effects of biodiversity on ecosystem functioning in nature. Ecosystem functioning research has also emphasized biodiversity as the key variable, while other community properties and processes have gone largely ignored. I studied carbon cycling in Arctic tundra plants by focusing on the ecophysiology of respiration rates rather than biodiversity. I also conducted a biodiversity and ecosystem functioning experiment using natural, unmanipulated communities. Finally, I used the data from both of these experiments to quantify intraspecific variation in functional traits, a crucial tool in studies of ecosystem functions. I found that respiration rates in the unique constant daylight environment in the Arctic are much higher than would be expected from studies in temperate environments. In the biodiversity and ecosystem functioning experiment, I was unable to find any measurable effect of biodiversity on biomass production, marking a stark contrast from previous results in controlled experiments. I also found extensive intraspecific variation in many common functional traits, highlighting the importance of including these measurements in similar studies.

Advisor: Cadotte, Marc
Assessing Glycaemic Index Utility: From Bench to Bedside

Glycaemic index (GI) categorizes carbohydrate-containing food according to postprandial glycaemic effect. The Canadian Diabetes Association supports integration of GI-education into standard care for type 2 diabetes mellitus (T2DM). Notwithstanding, most Canadian RDs identify as “non-users”; citing a need for studies examining GI-mechanism and GI-efficacy and a lack of reliable educational materials as barriers to GI-utility. These barriers highlight practice-based research opportunities spanning bench-to-bedside, such as: The examination of novel metabolic pathways and evaluation of low GI-education/ diet effectiveness using established markers of utility. The overall purpose of this dissertation was to address, experimentally, gaps in the literature, highlighted by educators as barriers to GI-utility. METHODS: Three studies were completed. Study one examined the effect of slowing carbohydrate absorption on postprandial oxidative stress (novel GI-mechanism) in overweight/obese participants (n=18). Study two involved the development and evaluation of a questionnaire designed to evaluate an evidence-based GI-education platform for DM (n = 29), using a one-armed pre/post-intervention pre-test design. Study three (RCT; n=99) was performed to evaluate the effect of the low GI-education platform on glycaemic control in women with GDM using the questionnaire pre-tested in study two. RESULTS: Sipping 75g dextrose solution slowly over 3.5h reduced postprandial oxidative stress compared to consuming it over 5min. Studies two and three showed that participants who received GI-education were satisfied with the education, showed a significant increase in GI-knowledge score (36%; p

Keywords: Diabetes Mellitus
Education
Glycaemic Index
Intervention
Knowledge Translation
Nutrition
Advisor: Wolever, Thomas MS
Uncovering a Forgotten Italian Renaissance Comedy: A Commented Critical Edition of Francesco Antonio Rossi's Capriccio

Comedy is an essential literary genre for any culture and any period. It provides both a representation and criticism of society, societal norms and societal institutions, and, as such, allows scholars to reconstruct societies and regain knowledge from centuries ago. As Riccardo Scrivano argues, “with the audacity of its fictions, its hypotheses, its capacity to cancel the usual modes of conduct, it reveals to us different ways of organizing knowledge, and thus different ways of knowing” (12). Each and every play and playwright add a new dimension of knowledge and a new means of knowing to our established repertoire of understanding. Consequently, every play, comedy in this instance, contributes to the process of understanding and influencing the society in which it is set, as well as those that follow it and those from which it borrowed. Douglas Radcliff-Umstead observes that “Italian playwrights turned to Roman antiquity to find a dramatic technique, but they combined literary traditions and the contemporary reality of their own country to create a comic theatre that helped determine the course of modern European drama” (Modern Comedy 243). Every play and playwright, including Francesco Antonio Rossi and his Capriccio, perform an important role in this process and should not be overlooked. The central objective of this dissertation is to produce a commented critical edition of Francesco Antonio Rossi’s Capriccio in accordance with the standard methods and practices laid down by modern scholarship in textual criticism. Until Rossi’s Capriccio is given new life in the form of a critical edition, academia’s comprehension of this author and his work will be incomplete, leaving Rossi’s comedy to sit on library shelves, never having had its significance realized.

Keywords: commented critical edition
European theatre
Italian comedy
literary criticism
philology
Renaissance
Advisor: Lettieri, Michael
Conjunctive Coding of Complex Object Features

Critical to perceiving an object is the ability to bind its constituent features into a cohesive representation, yet the manner by which the visual system integrates object features to yield a unified percept remains unknown. Another important aspect of object perception is the ability to represent complex objects across different viewing angles, despite the drastic variability in appearance caused by shifting viewpoints. This dissertation describes a novel application of multivoxel pattern analysis (MVPA) of neuroimaging data that allows a direct investigation of whether neural representations integrate object features into a whole that is different from the sum of its parts and whether these representations are view invariant. Results showed that patterns of activity throughout the ventral visual stream (VVS), extending anteriorly into the perirhinal cortex (PRC), discriminated between the same sets of features when combined into different objects. Despite this sensitivity to the unique conjunctions of features, this activity was invariant to the viewpoints from which the conjunctions were presented. These results suggest that the manner in which our visual system processes complex objects depends on the explicit coding of the feature conjunctions comprising them, and that these representations are invariant to large changes in visual appearance caused by shifting viewpoints. To test whether experience with the objects changed the pattern of conjunctive representations across the VVS, participants were scanned in a separate fMRI study following an extensive behavioural training period. Results showed that both view-dependent and view-invariant conjunctive representations shifted to more posterior regions of the VVS, suggesting that training affected how unique feature conjunctions are represented across the VVS. These results support the representational-hierarchical view of cortical organization and contribute to our understanding of object perception and memory.

Keywords: Experience
fMRI
Memory
Object perception
Perirhinal Cortex
Ventral Visual Stream
Advisor: Barense, Morgan D
'This Place Should (Not) Exist': An Ethnography of Shelter Workers and the In-Between

This dissertation draws on twenty-one months of participant observation at The Salvation Army Gateway, a shelter for men in Toronto. It pays particular attention to the shelter’s thirty-two workers, to their efforts to be both “the best shelter in the city” and to “befriend the poor.” The story told here employs Victor and Edith Turner’s theory of liminality as both prompt and foil. The understanding of the liminal it employs and the rationale for deploying it in relation to the Gateway are the subject of Part I. Aspects of the liminal (in-between) stage/state have been paired with various aspects of shelter life – certain repertoires, objects, ideas. There are eighteen such pairings, each provoking a short, ethnographically driven chapter. The interplay between the shelter and the liminal is at times illuminating, ironic, even incomplete. The chapters are clustered according to four themes – also called the coefficients of life at the shelter: stuckedness, subjunctivity, immaturity, irony. These four are simultaneously plotted in such a way as to lead the reader deeper into Gateway life. Part II Stuckedness, discusses contexts that surround the Gateway and hold it in place. Part III Subjunctivity, enters the shelter by way of the workers’ own ideas, desires, and intentions for the way the Gateway operates, especially its friendliness. Part IV Immaturity, tells the story of any-given day at the Gateway in order to consider its ambient youthfulness, and the significance of such for the formation of alternative community there. Part V Irony, unpacks the irreconcilable ironies that the workers live with. Understanding the Gateway as liminal space offers opportunity to think about whether or not the shelter system ought to exist in a city like Toronto and how all of us are implicated in sustaining it. While this thesis tries to preserve some of the undecideability of such a question, it also seeks to complicate the notion that people ought always to leave shelters in favour of “housing” or having their “own home.” This dissertation ultimately offers “homefulness” as a better measure for what is and is not lacking among the so-called “homeless.”

Keywords: christianity
ethnography
homelessness
liminality
Advisor: Klassen, Pamela E.
Prior-knowledge Effects on Associative Memory Processing

Forming new associations is a fundamental process in building our knowledge system and creating episodic memories. How prior-knowledge influences acquisition of novel associations has not been thoroughly investigated. Based on recent cognitive neuroscience literature on multiple-component memory processing, I hypothesize that prior-knowledge can trigger additional evaluative, semantic, or episodic processes, supported by the ventral medial prefrontal cortex (vmPFC), anterior temporal pole regions (aTPL), and hippocampus (HPC), respectively, to facilitate new associative learning. To test this hypothesis, I first developed a face-house associative memory task in Experiment 1, in which famous faces were used to elicit multiple-component prior-knowledge, such as social emotional, semantic, and episodic memories. Using this task, I found that recollection-based, not familiarity-based, associative memory was indeed facilitated by this type of prior-knowledge. Then, I used this task in two fMRI studies to examine how prior-knowledge could affect brain activity during associative encoding (Experiment 2) and post-encoding rests (Experiment 3). Consistent with my hypotheses, in Experiment 2, I found that the vmPFC and HPC, as well as the parahippocampal place area (PPA) and fusiform face area (FFA), showed stronger activation when famous, compared to nonfamous, faces were involved. Importantly, the vmPFC, aTPL, and HPC also exhibited stronger activation when famous faces elicited stronger prior emotions and memories. Experiment 3 showed that HPC connectivity with FFA and vmPFC during rest was stronger following encoding of associations with famous than non-famous faces. Post-encoding connectivity between the HPC and FFA, and between the aTPL and PPA/FFA, also predicted later associative, but not item, memory, only in the famous condition. These results indicate that when prior-knowledge is involved, the HPC, vmPFC, and aTPL, which support prior episodic, social-evaluative, and semantic memories, respectively, continue to interact with each other and the posterior perceptual brain regions (e.g., the PPA and FFA) during the post-encoding rest to facilitate off-line processing of the newly formed memory and enhance it. Taken together, these findings support my hypotheses that prior-knowledge reconfigures memory processing among different brain systems, by triggering additional evaluative, semantic, or episodic processes, to facilitate new associative memory formation during both encoding phase and post-encoding rests.

Keywords: Associative memory
encoding
fMRI
memory
post-encoding brain activity
prior knowledge effects
Advisor: Moscovitch, Morris
The Large Maf Transcription Factor Traffic Jam Regulates Collective Cell Migration During Drosophila Oogenesis

Collective cell migration is a fundamental process throughout development. The migrating border cell cluster (BCC) in the Drosophila ovary is a good in-vivo model system to study this process. My work focuses on the function of the large Maf transcription factor, Traffic Jam (Tj), in regulating BCC migration. Tj has a dual function in both promoting and inhibiting BCC migration, as reduction or increase of tj expression leads to migration delays. I found a genetic interaction between Tj and three factors known to promote BCC migration: an inhibitor of the JAK/Stat pathway, Socs36E, the transcription factor Slow border cells (Slbo), and the adhesion molecule DEcadherin (DEcad). At high expression level, Tj enhances the expression of Socs36E, which subsequently inhibits JAK/Stat and causes reduction of Slbo. High Tj expression also inhibits DEcad upregulation in the BCC. Activation of Slbo, which is induced by JAK/Stat, leads to a natural downregulation of Tj. This low expression level of Tj maintains Slbo expression in the BCC, which allows for DEcad upregulation. My work has shown that Tj is an important component of this regulatory network, which allows for proper BCC migration. Analysis of BCC morphology reveals that Tj also functions in regulating the formation of actin-based cellular protrusions. These protrusions are normally extended by border cells and are necessary for its movement. Altering Tj expression leads to abnormal organization of the actin cytoskeleton and morphology of the protrusions. This effect does not appear to be mediated through Slbo or DEcad, which are known to regulate actin. A genome-wide microarray screen identified serine/threonine kinase Protein Kinase C δ (PKCδ) as a putative target of Tj. Changing PKCδ expression in the BCC also causes abnormal organization of actin filaments. Genetic interaction studies suggest that Tj regulates actin organization through upregulating PKCδ. Furthermore, altering Tj or PKCδ expression changes the distribution pattern of an actin filament elongation factor, Enabled, in the BCC. My results have identified Tj and its putative downstream effector, PKCδ, as crucial regulators of the actin cytoskeleton and cellular protrusions in the BCC.

Keywords: Border Cell Migration
Large Maf Transcription Factor
PKCdelta
Traffic Jam
Advisor: Godt, Dorothea
Sowing the State: Nationalism, Sovereignty and Agrarian Politics in Venezuela

Sowing the State is an ethnographic account of the remaking of the Venezuelan nation-state at the start of the twenty-first century which underscores the centrality of agriculture to the re-envisioning of sovereignty. The narrative explores the recent efforts of the Venezuelan government to transform the rural areas of the nation into a model of agriculture capable of feeding its mostly urban population as well as the logics and rationales for this particular reform project. The dissertation explores the subjects, livelihoods, and discourses conceived as the proper basis of sovereignty as well as the intersection of agrarian politics with statecraft. In a nation heavily dependent on the export of oil and the import of food, the politics of land and its various uses is central to governance and the rural is a contested field for a variety of social groups. Based on extended fieldwork in El Centro Técnico Productivo Socialista Florentino, a state owned enterprise in the western plains of Venezuela, the narrative analyses the challenges faced by would-be nation builders after decades of neoliberal policy designed to integrate the nation into the global market as well as the activities of the enterprise directed at transcending this legacy. Not merely a restoration of the status quo or reassertion of a prior independence, I argue the Venezuelan nation is being reinvented in this drive for sovereignty and the tensions between peasants, technical experts and workers in the Florentino enterprise reflect the cleavages of an emerging state form.

Keywords: Agrarian Reform
Development
Food Security
Nationalism
Sovereignty
Venezuela
Advisor: Li, Tania