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Urinary Tract Infections Through the Lens of the Urinary Microbiome
No full textThe misconception of a sterile urinary tract has been overturned in the past 20 years, leading to a burgeoning field of urinary microbiome research. While Escherichia coli is the predominant cause of urinary tract infections (UTIs), a diverse range of bacteria can also inhabit the urinary tract and cause UTIs. Non-E. coli members of the urinary microbiota remain understudied. Similarly, standardized methods for studying the urinary microbiome remain lacking. This dissertation integrates bioinformatics and bacterial pathogenesis approaches to investigate the urinary microbiome and specific commensal and pathogenic bacteria. Using clinical samples, I examine the composition and functional roles of urinary microbiota in health and disease. Chapter 2 establishes methodologies for studying the infant urinary microbiome and characterizes the Actinotignum genera, an opportunistic urinary pathogen. Chapter 3 links urinary microbiota composition to symptoms in patients with interstitial cystitis/bladder pain syndrome. In Chapter 4, I describe urinary and fecal microbiota in children with spina bifida, identifying mechanisms of bacterial evolutionary adaptation to the urinary tract. These evolutionary changes offer intriguing targets for therapeutic development against urinary pathogens. Chapter 5 explores the pathogenesis of Pseudomonas aeruginosa, revealing its intracellular behavior and respiratory strategies in bladder infection. Collectively, this dissertation integrates sequencing- and culture-based methods to characterize the urinary microbiome across diverse urologic diseases, revealing microbial colonization patterns, bacterial adaptation mechanisms, and host-pathogen interactions relevant to UTI pathogenesis. These findings establish a methodological and conceptual foundation for future mechanistic studies and therapeutic development targeting urinary tract diseases
Intrinsic and Environmental Drivers of Thyroid Cancer
No full textThyroid cancer is predicted to be the fourth most common cancer diagnosis come 2030. We were interested in understanding current drivers of the increase in cases and potential biomarkers of disease. To do so, we first investigated the role of per- and polyfluoroalkyl substances to further understand the alterations in thyroid histology and function. We also investigated the role of Tenascin-C (TNC), a secreted extracellular matrix protein, in facilitating ligand-dependent Wnt signaling in thyroid cancer. To understand the effects of PFAS on thyroid histology, we used a PFAS feeding model of a combination of three PFAS compounds (PFOS, PFOA, and GenX). This treatment of animal models showed altered thyroid architecture and cell structure following PFAS exposure. RNA-sequencing data reveled that the gene Klhl23 and multiple signaling pathways were dysregulated, including actin polymerization. Understanding the role of PFAS-mediated toxicity in the thyroid is critically important for prevention of thyroid disease in the population. Next, we utilized bulk RNA-sequencing, RNA in situ hybridization, and in vitro and in vivo models to investigate TNC. We found that TNC expression was associated with aggressive thyroid cancer behavior. Spatial localization of TNC in patient tissue demonstrated a dramatic increase in expression within cancer cells along the invasive edge, adjacent to Wnt ligand-producing fibroblasts. In vitro, TNC bound Wnt ligands and potentiated Wnt signaling. Finally, in an ATC mouse model, TNC increased Wnt signaling, tumor burden, invasion, and metastasis. Understanding the role of TNC and its interaction with Wnt ligands could lead to the development of novel biomarkers and targeted therapeutics for thyroid cancer
Population Colonization Dynamics in Urinary Tract Infection
No full textUropathogenic Escherichia coli (UPEC), the primary causative agent of urinary tract infection (UTI), is thought to reside in the gut before ascension into the urinary tract, yet a reproducible model for its transit between these host environments has never been developed. Here, we demonstrate that intra-species competition plays a key role in UPEC gut colonization. Resident Enterobacterales, particularly commensal E. coli, impose colonization resistance, whereas their absence or elimination enables stable UPEC persistence. In mouse lineages in which UPEC stably colonizes the gut, we reveal that spontaneous bacteriuria is regularly detected, providing a novel framework for studying UPEC movement along the gut-urogenital axis. Additionally, we show that active UTI promotes UPEC gut colonization, overcoming colonization resistance, and we provide evidence that the vaginal space is a crucial intermediary in UPEC transition from the gut to the urinary tract environment. Our findings suggest that UPEC expansion in these alternative host niches contributes to UTI recurrence. This is particularly problematic if antibiotic resistant populations colonize multiple host environments, as even susceptible populations fail to be cleared from the gut following UTI treatment. To further investigate UPEC’s adaptive strategies, we examined fosfomycin resistance mechanisms, identifying common and novel FosR mutations that link resistance to metabolic rewiring. Contrary to prior assumptions that FosR mutations elicit biological cost to the pathogen, we observed that uhp and pyk mutants establish infection comparably to wild type strains, and in some cases mice infected with the FosR mutants resolve UTIs less rapidly, suggesting a selective advantage in maintaining these resistant subpopulations. Our findings support a re-evaluation of fosfomycin clinical susceptibility standards, in which the observation of FosR subpopulations has previously been ignored. Collectively, this work underscores the need for further investigations into UPEC’s metabolic and ecological adaptations, informing treatment strategies and improving UTI management
The Power to Invade: Armed Primacy and Congressional Conflict, 1977-1987
No full textThe Power to Invade: Armed Primacy and Congressional Conflict, 1977-1987 highlights the critical importance of the U.S.’s pursuance of a strategy of armed primacy (leadership, supremacy, and/or dominance) during the Carter and Reagan administrations which was driven by a competition with the Soviet Union in the Cold War. This book argues that armed primacy was a unifying historiographic throughline in Cold War Presidential administrations. During the Carter and Reagan years (1977-1987), the pursuit of this strategy, especially in U.S. foreign policy actions in Latin America, led to conflict between the branches of the U.S. government in foreign policy making and growth in Presidential power
Here for the Humor: Exploring the Role of Humor in Multilingual K-12 Settings
No full textThis three-paper dissertation contributes to the fields of teaching and learning, particularly in contexts with multilingual learners (MLs) and teachers of multilingual students. The research is especially fruitful to both teachers and teacher educators, acknowledging humor and its impact on the classroom. Paper 1, a literature review of humor in multilingual educational settings sets the stage for what we know about the topic, humor’s potential in ML contexts, and its role to humanize or dehumanize by putting humor and humanizing pedagogies literatures in conversation. The work then offers the conceptualization of humanizing humor, which is operationalized by five synthesized tenets: cultural, interactional, affective, instructional, and motivational. This foundation led to empirical pieces in elementary school settings. Building further on the affective tenet of humanizing humor from Paper 1, we employed affect as the theoretical lens in Paper 2. The empirical research involved qualitative data collection and discourse analytic methods that were affectively situated, originating from a larger pre-existing study in partnership with Dr. Emily Phillips-Galloway. The concepts focused on in the discourse interaction episodes were embodiment, power shifts and norm violations with taboo, and affective spread. Our findings amplify playful translanguaging and three of its features as one way to rehumanize language spaces. This contributes to the small but mighty corpus of humor studies in elementary contexts with linguistic humor and further develops playful translingual spaces by focusing on the concept of playful translanguaging through discourse episodes. There is a need for humor studies in elementary schools and the following empirical work strives to elevate the authentic uses of humor in ML settings. Paper 3, an ethnographic dual case study partnering with Kindergarten and 1st grade teachers of multilingual students engaged qualitative, affective, and collaborative methods. The analyses share four uses of humor and their impacts on the holistic development of a student as a learner and a person. This manuscript furthers the evolution of humor scholarship in the between moments and proclaims some of the many ways humor is infused in and impacting our classrooms
Characterization of the Calcium-Mediated Tetramerization of Human Calprotectin
No full textCalprotectin (CP) is an S100 protein that plays a role in the inflammatory response by acting as a ligand for the receptor for advanced glycation end-products (RAGE) and Toll-like receptor 4 (TLR-4). Activation of these receptors leads to upregulation of inflammatory cytokines, chemokines, and CP through the NF-κB pathway. In the case of people with irritable bowel disease (IBD) these pathways are stimulated which leads to damage of the gastrointestinal tract and disease symptoms. CP is a heterodimer of the S100A8 and S100A9 EF-hand calcium binding proteins. Like all S100 proteins, calcium-induced conformational changes in CP are required for binding to partner proteins. However, in the case of CP, the addition of calcium correlates with self-association and the formation of a dimer of heterodimers (heterotetramer). Ligand induced receptor oligomerization has been proposed as a mechanism of receptor activation. Conversely, it has also been suggested that CP tetramerization can inhibit binding to receptors and serve as an autoinhibitory mechanism to modulate the inflammatory response. In order to investigate the functional relevance of CP tetramerization and facilitate in-depth biophysical and structural analysis, I prepared single-site mutations of two hydrophobic residues mediating the tetramer interface (Ile60, Ile73) and a double-site mutant (I160K,170K). With the goal of maximally destabilizing the tetramer interface, these Ile residues were mutated to Lys (I60K, I73K). This work reports biophysical characterization of the three tetramer-deficient mutants using calorimetric, scattering, and structural approaches. Dynamic light scattering, small-angle x-ray scattering, and nuclear magnetic resonance show that the CP tetramer-deficient mutants remain as dimers in solution even in the presence of 40-fold excess calcium. The crystal structure of CP I73K mutant was determined to atomic level resolution and confirms there are virtually no differences in the structure of the heterodimer. Together, these results indicate that the mutants will be useful reagents for discerning the functional role of CP oligomerization in activation of cell-surface receptors. The implications of these results are discussed in the context of the complexity of CP receptor signaling
Mechanisms of Cell-Free Hemoglobin-Mediated Injury to the Endothelial Glycocalyx During Sepsis
No full textSepsis is a critical public health issue with high morbidity and mortality. A major reason for the substantial burden of sepsis is an insufficient understanding of the biologic mechanisms that potentiate its pathogenesis. One of the hallmarks of sepsis is endothelial glycocalyx degradation, which occurs due to the activation of heparanase, a glycocalyx-cleaving enzyme. This manifests as endothelial barrier hyperpermeability that leads to organ dysfunction including acute respiratory distress syndrome (ARDS). However, much remains unknown regarding the molecular pathways that underlie sepsis-induced heparanase upregulation. This dissertation presents findings that address this key knowledge gap, centering on the concept that increased heparanase activity during sepsis is in part driven by reduction-oxidation (redox) cycling of hemoglobin that is released into the circulation (i.e., cell-free hemoglobin, CFH). Utilizing plasma from sepsis patients, I demonstrate that concentrations of circulating heparanase and heparan sulfate fragments correlate with worse clinical prognosis. My findings also reveal a direct association between plasma CFH, heparanase, and heparan sulfate degradation products in human sepsis. Further, in experimental sepsis models, I show that CFH stimulates heparanase transcription and activation, and that this is mitigated by acetaminophen, an inhibitor of CFH redox cycling. Moreover, in wildtype septic mice, CFH treatment enhances endothelial glycocalyx shedding, inflammation, and illness severity – effects that are not observed in mice lacking endothelial heparanase. Together, these data suggest CFH upregulates active heparanase, exacerbating endothelial glycocalyx breakdown and worsening outcomes. This work thus uncovers a previously unrecognized pathway of endothelial injury, providing novel insight into sepsis pathogenesis
Maximizing Statistical Efficiency in Clinical Trials: Ordinal Longitudinal Models and Two-Phase Designs
No full textClinical trials are the primary method used to establish the safety and efficacy of medicines and interventions. Results from clinical trials form the foundation of medical knowledge and improve the lives of millions of patients. However, clinical trials are often expensive to run, slow to enroll participants, and subject to unforeseen delays -- leading to trials that are underpowered to detect a clinically relevant effect. Optimizing statistical efficiency through the use of modern study designs and analysis methods is therefore an important goal. In this work, we advance two Bayesian methods to improve the efficiency of clinical trials. First, we show how Bayesian ordinal transition models (OTMs) can be used to analyze ordinal longitudinal data, an information-rich type of outcome data that has become common in COVID-19 clinical trials, and develop the statistical theory for these models from first principles. We illustrate OTMs using data from ACTT-1, a clinical trial evaluating the use of remdesivir in hospitalized patients with COVID-19. We then compare OTMs to other commonly used models and approaches for analyzing ordinal longitudinal data through a comprehensive simulation study, and demonstrate that OTMs can greatly improve statistical power. Second, we describe a Bayesian factored likelihood approach to account for missing exposure data in two-phase designs using outcome-dependent sampling and BLUP-dependent sampling. We demonstrate how this approach can improve statistical efficiency compared to simple random sampling through simulations, and provide a case study using data from the Lung Health Study clinical trial. Both of these methods can inform clinical trial design to improve statistical efficiency and advance medical knowledge more quickly
Memory for Discourse Context Following Disfluency
No full textThis study investigated how memory for the reparandum (the original, corrected speech) and the repair (the corrected portion of speech) are influenced by speech containing disfluencies, specifically repair disfluencies. A modified version of the paradigm from Lowder and Ferreira (2016) was used, in which participants listened to sentences containing either disfluent repairs (e.g., “salt, uh I mean ketchup”) or coordination structures (e.g., “salt and also ketchup”). The primary goal was to examine how the presence of disfluencies might affect memory for the reparandum and repair, with a particular focus on whether certain types of linguistic structures—such as repairs or coordination structures—might be better remembered. The results revealed that participants were most accurate in recalling the repair portion of the sentence, followed by the reparandum. Additionally, semantically related items were better remembered than unrelated distractors, indicating that meaning plays a significant role in memory encoding. These findings suggest that the reparandum, while often interrupted by disfluency, continues to linger in memory and may influence how subsequent speech is processed and recalled. Interestingly, the length of the editing phase (the pause or hesitation during disfluency) did not have a significant impact on memory performance. Furthermore, no significant differences were found between the disfluency condition and the coordination condition, suggesting that it is the role of prediction mechanisms in memory encoding that is most influential, rather than the presence of disfluency itself. These results highlight the importance of prediction in how speech is processed and remembered
Silicon Meta-optics Enabling Edge Detection and Varifocal Thermal Imaging
No full textOptical metasurfaces have emerged as an exciting small form factor alternative to traditional refractive imaging systems due to their ability to spatially control amplitude, phase, and polarization of incident light. Propagation phase metasurfaces employ sub-wavelength meta-atoms of varying diameters for wavefront control by inducing specific phase delay spatially across the device. Spatial design freedom gives rise to point spread function (PSF) engineering, which can perform information encoding of incoming light. Specifically, I discuss an optimized broadband Laplacian-of-Gaussian edge detector metasurface, which uses birefringent meta-atoms to create PSFs in both polarization directions. I examine testing results captured with a long wave infrared (LWIR) camera, which detects large amounts of signal from objects between 0-150°C. In the future, similar edge detection metasurfaces could serve as a preprocessing step for computer vision neural networks, minimizing computational power requirements. Computer vision object recognition over this temperature range has applications in medical imaging, autonomous vehicles, and security. Next, I review the design and creation of a Moiré metalens system, which provides continuous focal length adjustment in the LWIR. Alignment of two Moiré metasurfaces supplies tunable varifocal imaging via rotation of one metasurface. This system eliminates traditional axial translation in varifocal imaging systems, significantly decreasing the form factor for applications in endoscopy and compact device imaging. A newly developed silicon photolithography and etching method was used to fabricate metasurfaces for both projects. I explore the potential for high-aspect ratio meta-atoms with this new process