Life Sciences Commons^™

Contraction Analysis Of Functional Competitive Lotka-Volterra Systems: Understanding Competition Between Modified Bacteria And Plasmodium Within Mosquitoes., Nickolas Goncharenko

Electronic Thesis and Dissertation Repository

We propose and analyze an extension to the classic Competitive Lotka-Volterra (CLV) model. The goal is to model competition between species, with a response from the environment. This response is a function of the population of all species and can represent numerous physical phenomena including resource limitation and immune response of a host due to infection. We name this new system a Functional Competitive Lotka-Volterra (FCLV) model. We mainly use the construction of contraction metrics, to determine global properties of the model. We use this result to analyze the competition between Plasmodium sp. and genetically engineered bacteria within the midgut …

Designing A Novel Hiv-1 Candidate Vaccine, Rahul Pawa

Electronic Thesis and Dissertation Repository

Currently no vaccine has been developed that can prevent the spread of HIV-1. During sexual transmission, a single viral variant called the Transmitted/Founder (T/F) purportedly with unique physical properties, establishes infection in 70-80% of individuals. Unlike previous studies that have tried to identify T/F viruses based on their structure glycan composition and amino acid sequence, we have analyzed the RNA sequences of HIV-1 to help identify T/F variants. Using a combination of both in silico data analysis and in vitro assays, we have identified that T/F viruses have higher numbers of immunostimulatory motifs than HIV virions that fail to infect. …

Utilizing Fiv (Feline Immunodeficiency Virus) To Develop A Novel Animal Model To Study Hiv (Human Immunodeficiency Virus), Ankita Suryakant Kambli

Electronic Thesis and Dissertation Repository

This project sought to perform the in vitro work needed to accomplish the long-term vision of harnessing the similarities between HIV (Human Immunodeficiency Virus) and FIV (Feline Immunodeficiency Virus) to develop an animal model whereby cats can be used to study HIV pathogenesis and therapeutics. We transfected CRFK (Crandell Rees Feline Kidney) fibroblasts with plasmids that could express human or feline CD4, CCR5, or both, and determined receptor surface expression through flow cytometry. We discovered that HIV envelope expressed on 293T can fuse with huCD4/huCCR5 on CRFK. These cat cell lines were also capable of supporting HIV infection. Additionally, we …

Encountering Lectins In The Recipient Mucosa: Implications Of N-Linked Glycosylation On Hiv-1 Transmission, Adam Meadows

Electronic Thesis and Dissertation Repository

Although several studies have determined key differences in envelope motifs between TF and chronic HIV-1, it is still not known what the overall glycosylation profile is that is selected for in a transmission event, as well as what contributes to this selection. Using a bottom-up approach of modifying specific viruses, determining their transmission fitness in an ex vivo tissue explant assay, and determining their glycan content, we have laid the basis for determining the overall glycan structure which is selected for in TF HIV-1. Preliminarily, we have shown that C-type lectins represent a stringent barrier to transmission and have several …

Determining The Relative Transmission Fitness Of Hiv-1 Subtypes A, B, C, And D, Spencer Yeung

Electronic Thesis and Dissertation Repository

There is in vivo evidence that suggests the genetic diversity of HIV-1 subtypes influence heterosexual transmission efficiency. To recapitulate sexual transmission in vitro, blocks of genital tissue were exposed to mixtures of genetically different subtype viruses. Migrating immune cells were collected and co-cultured with a CD4+ T-cell line permissive to HIV infection (PM1) to measure dendritic cell virus transfer; HIV-exposed tissues were cultured separately. Next generation sequencing (NGS) of HIV-1 DNA was used to quantify relative infection rates of the various challenge viruses, and to assess fitness differences in infection of the tissue vs. migratory/T cell co-cultures. Our results …

Metabolomic And Genomic Analysis Of Lactobacillus Crispatus Vaginal Isolates, Emiley Watson

Electronic Thesis and Dissertation Repository

Lactobacillus crispatus is the dominant species in the vagina of many women. To add to our knowledge of its properties, the metabolic readout of twenty strains was analyzed using liquid chromatography-mass spectrometry (LC-MS). This led to a separation of the strains into two groups, Groups A and B. Notably, five Group B strains produced tyramine, known to raise vaginal pH, and make the environment more favourable to vaginal pathogens. The origin of Group B strains did not correlate with their host having bacterial vaginosis (BV), a condition associated with elevated pH. Five bacteriocins were detected in the genomes. Therefore, differences …

Endogenous, Controlled Expression Of Anti-Hiv-1 Broadly Neutralizing Antibody, Darshit Patel

Electronic Thesis and Dissertation Repository

Recently, researchers have identified a number of anti-HIV broadly neutralizing antibodies (bNAbs), such as VRC01 and N6, capable of targeting a broad range of HIV-1 strains. Passive immunization using these patient-derived bNAbs could provide temporary protection but are limited by the short antibody half-life. While current gene transfer technology allows sustained bNAb expression, it lacks the ability to control bNAb production in vivo resulting in possible autoimmunity. To address this issue of achieving controlled bNAb expression in vivo, we hypothesize that bNAb expression from transduced Flu-specific B cells can be activated and modulated by subsequent Flu immunizations in the …

Hiv-1 Group M Subtype Fitness, Disease Progression, And Entry Efficiency, Colin M. Venner

Electronic Thesis and Dissertation Repository

Human immunodeficiency virus type 1 (HIV-1) emerged in the human population shortly after the turn of the 19th century. Distribution of HIV-1 across the globe over the past 30–35 years can be traced to founder events with primordial HIV strains from sub-Saharan Africa. Even considering the burden of HIV in Africa, our knowledge of HIV-1 disease is still largely limited to subtype B HIV-1, a strain responsible for 3 million infections in North America and Europe as compared to the 33 million that are infected with HIV-1 subtypes A, C, D, and circulating and unique recombinant forms.

This dissertation analyzes …

The Group A Streptococci Bacteriocins Facilitate A Competitive Advantage During Nasopharyngeal Infection, Lana Estafanos

Electronic Thesis and Dissertation Repository

Pathogenic streptococci have evolved specific systems to eliminate bacterial competitors within their biological niche. In microbial environments, niche competition is often driven by the production of short antimicrobial peptides called bacteriocins; this provides a mechanism by which Streptococcus pyogenes may compete for ecological stability and establish infection. Recent findings from our laboratory have identified two novel Class IIb bacteriocin systems – Streptococcus pyogenes bacteriocin (Spb) JK and MN in the M18 serotype S. pyogenes strain MGAS8232 – that may contribute to nasopharyngeal infection. Here, we show that galactose and CO₂ are distinct regulatory cues which induce antimicrobial activity. Under …

Novel Insights Into The Genomic Integration Site Landscape Of Hiv-1 And Other Retrovirus Genera, Hinissan P. Kohio

Electronic Thesis and Dissertation Repository

An important event during infection by retroviruses such as human immunodeficiency virus type 1 (HIV-1) is the permanent integration of the viral genome into the host genome. This event leads to life-long infection and is accompanied by a period of quiescence/latency ranging from a few years to >10 years where HIV-1 expression is barely detectable or undetectable. Despite the use of combination antiretroviral therapy (cART) which controls HIV-1 infection, quiescent/latent virus presents a major obstacle towards a functional cure. Integration site location in the genome is thought to contribute to latent infections and has the potential to confound anti-latency treatments, …

Determining The Molecular Mechanisms Of Pacs-1-Mediated Protein Sorting, Brennan S. Dirk

Electronic Thesis and Dissertation Repository

Membrane trafficking events are required to direct proteins to their precise subcellular locations. The cellular Phosphofurin Acidic Cluster Sorting protein – 1 (PACS-1) has emerged as a protein of interest in controlling the localization of a multitude of cellular and viral proteins. Specifically, PACS-1 is hijacked by type-1 Human Immunodeficiency Virus (HIV-1) to contribute to immune evasion in addition to regulating neuroendocrine hormone storage and release. To accomplish this, PACS-1 connects the cytoplasmic tail of cellular receptors to the heterotetrameric adaptor proteins (APs) to form a functional trafficking unit. Throughout this dissertation, I explored the role of PACS-1 and AP-1 …

Interferon-Induced Herc5 Is A Novel And Potent Inhibitor Of Ebola Virus-Like Particle Production, Nina Hunt

Electronic Thesis and Dissertation Repository

Ebola’s severe pathogenesis can be attributed to its suppression of the Type I interferon (IFN) response, suggesting this pathway plays a role in restricting viral replication. One IFN-induced protein, HERC5, warrants further investigation as it inhibits the replication of evolutionarily diverse viruses. We showed that HERC5 drastically reduces the expression of structural protein VP40 at the protein and mRNA level. Mutagenesis of HERC5 demonstrated that the RCC-1-like domain is necessary and sufficient for restriction. This domain is also responsible for HERC5’s interaction with ZAP, a protein required for VP40 mRNA degradation. Finally, we showed that Ebola GP antagonizes HERC5 activity. …

The Role Of The Hiv-1 Accessory Proteins Nef And Vpu In Subverting Host Cellular Immunity, Emily N. Pawlak

Electronic Thesis and Dissertation Repository

The HIV-1 accessory proteins Nef and Vpu have no known enzymatic activity, yet these viral proteins are critical for HIV-1 viral spread and pathogenicity. To mediate this, these viral proteins bind to and hijack host cellular proteins, including proteins implicated in the membrane trafficking machinery. Key to the effects of these proteins is their ability to alter the subcellular localization of a plethora of cell surface receptors, including the key immune cell receptorsCD28 and MHC-I.However,the mechanism utilized by Nef and Vpu to modulate these proteins is not fully understood. Herein, we demonstrated that Nef and Vpu mediate a decrease in …

Understanding The Impact Of Hiv-1 Genetic Diversity On The Function Of Nef And Its Role In Serinc5 Antagonism, Aaron Leslie Johnson

Electronic Thesis and Dissertation Repository

HIV-1 Nef is a key pathogenic protein, allowing HIV-1 to evade the host immune system by downregulating MHC-I and CD4. Furthermore, it was recently discovered that Nef counteracts the host factor SERINC5 to increase HIV-1 infectivity, but the mechanistic details of the Nef:SERINC5 interaction still need to be explored. Throughout this dissertation, I will explore the hypothesis that the genetic diversity that defines HIV-1 has a pronounced effect on the HIV-1 protein Nef, altering its function between and within group M subtypes. To address this hypothesis I investigated how MHC-I and CD4 downregulation differ among all non-recombinant group M subtypes. …

The Surreptitious Survival Of The Emerging Pathogen Staphylococcus Lugdunensis In Macrophages Enhances S. Aureus Infection, David Watson

Electronic Thesis and Dissertation Repository

Staphylococcus lugdunensis is an opportunistic pathogen that can cause invasive infections suggesting an ability to circumvent host immunity. S. lugdunensis was shown to resist killing and persist within macrophages and acetylation of its peptidoglycan is important for this survival. This was consistent in vivo, as S. lugdunensis resides inside Kupffer cells for at least 16 hours post-infection in mice. Despite its capability for survival, S. lugdunensis is unable to replicate within phagolysosomes. Inhibiting phagolysosomal effectors allows S. lugdunensis to initiate replication, after which the bacteria escape phagosomal containment. Moreover, intracellular S. lugdunensis augments the growth S. aureus during co-infection. Eight …

Effect Of Estrogen And Glucocorticoid Signaling On Th2 Cells – Implications For Severe Asthma, Meerah Vijeyakumaran

Electronic Thesis and Dissertation Repository

T helper (Th2) cells are increased in asthma and mediate allergic inflammation through production of type 2 cytokines (interleukin-4, -5 and -13). One pathway to activate Th2 cells is through chemoattractant-homologous receptor expressed on Th2 cells (CRTh2), a receptor for prostaglandin D₂ (PGD₂). Glucocorticoids (GCs) are the main treatment for allergic disease, due to their ability to suppress type 2 cytokine production and induce apoptosis. Incidence and severity of asthma is greater in women than men, which may be related to sex-specific efficacy of GCs. Recently, our laboratory found that severe asthmatic women had more circulating Th2 …

Functional And Structural Mimicry Of A-Kinase Anchoring Proteins By Human Adenovirus E1a, Cason R. King

Electronic Thesis and Dissertation Repository

As an obligate intracellular parasite, human adenovirus (HAdV) must utilize host factors for survival and replication. Early during infection, its multifunctional E1A protein interacts with an impressive range of cellular target proteins to exert control over the cellular environment. Through these virus-host interactions, E1A massively reprograms both viral and cellular transcription to activate the other HAdV genes, downregulate the host’s immune response, and induce the cell cycle. Consequently, E1A converts the infected cell into a compliant state more amenable for HAdV replication, resulting from its numerous protein-protein interactions. I sought to examine E1A’s interaction with cellular protein kinase A (PKA), …

Iron And Copper Homeostasis In Staphylococcus Aureus, Holly A. Laakso

Electronic Thesis and Dissertation Repository

All microorganisms require transition metals for key metabolic processes, thus during infection microbial access to essential metals is tightly regulated by the host in a process termed nutritional immunity. Iron acquisition is critical to the pathogenesis of the formidable human pathogen, Staphylococcus aureus, which utilizes heme-uptake systems and two high-affinity iron-scavenging siderophores, staphyloferrin A (SA) and staphyloferrin B (SB) for iron acquisition. In this study, I identify sbnI as encoding a transcription factor required for expression of genes in the sbn operon, the biosynthetic operon for SB synthesis. I also show that SbnI is a novel hemoprotein, where binding …

Nutritional Regulation Of Sulfonamide Antibiotic Biodegradation By Microbacterium Sp. Strain C448, Tonya Malcolm

Electronic Thesis and Dissertation Repository

Sulfonamide antibiotics are frequently released into the environment due to their widespread use in livestock production, imposing a potential selection pressure for the development and dissemination of sulfonamide resistance. Recently, a sulfonamide-degrading bacterium Microbacterium sp. Strain C448 was discovered, whose activity has the potential to be used as a sulfonamide removal strategy from agricultural soils. This research gains insight into the sulfonamide biodegradation pathway by 1) evaluating the nutritional regulation of sulfamethazine biodegradation and 2) undertaking a comparative proteomic analysis of cells grown in the presence and in the absence of sulfamethazine. The nutritional analyses revealed that increasing amounts of …

Characterizing The Interaction Between Human Adenovirus E1a And Sting, Jessica Hill

Electronic Thesis and Dissertation Repository

When challenged by viral DNA, the cytoplasmic DNA sensor cyclic GMP-AMP synthase (cGAS) signals through the adaptor protein stimulator of interferon genes (STING) to induce a primary type I IFN response. Studies from recent years have also revealed shared architecture between metabolism and innate immunity. Viruses have evolved to counteract these mechanisms. Human adenovirus (HAdV) early region 1A (E1A) protein antagonizes the cGAS-STING pathway to prevent an innate immune response by physically interacting with STING. I hypothesize that the interaction between E1A and STING is mediated through several motifs and involves ribosomal protein S6 kinase beta-1 (S6K1). Using a series …

Identification And Characterization Of Farr And Fare As A Regulator And Effector Of Fatty Acid Resistance In Staphylococcus Aureus, Heba Alnaseri

Electronic Thesis and Dissertation Repository

Although Staphylococcus aureus is exposed to antimicrobial fatty acids on the skin, in nasal secretions and in abscesses, specific mechanisms for regulating gene expression and intrinsic resistance in response to these fatty acids have not been reported. Through in vitro selection for increased resistance of S. aureus to linoleic acid, I identified fatty acid resistant clone FAR7, where a single nucleotide polymorphism caused a His₁₂₁Tyr substitution in an uncharacterized member of the TetR family of transcriptional regulators, which is divergently transcribed from a gene encoding a member of the resistance-nodulation-division superfamily of multi-drug efflux pumps. I named these …

Urea As An Effective Nitrogen Source For Cyanobacteria, Kevin J. Erratt

Electronic Thesis and Dissertation Repository

Urea use has grown substantially in the past half-century, with urea now accounting for > 50% of nitrogen fertilizer applications worldwide. The shift from inorganic nitrogen fertilizers to urea-based sources has coincided with the reappearance of cyanobacteria blooms in freshwaters. Here, we examined urea as a nitrogen source for three bloom-forming cyanobacteria species. We found that (1) urea was consumed more rapidly relative to inorganic nitrogen substrates, suggesting that cyanobacteria exhibit a preference for urea; (2) urea was consumed in excess of cellular requirements; and (3) urea may offer cyanobacteria a competitive edge over eukaryotic algae by enhancing light absorption capabilities. …

The Effect Of Diet On Midgut And Resulting Changes In Infectiousness Of Acmnpv Baculovirus In Trichoplusia Ni, Elizabeth Chen

Electronic Thesis and Dissertation Repository

The cabbage looper, Trichoplusia ni, a global generalist lepidopteran pest, has developed resistance to many synthetic and biological insecticides, requiring effective and environmentally acceptable alternatives. One possibility is the Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV). This baculovirus is highly infectious for T. ni, with potential as a biocontrol agent, however, its effectiveness is strongly influenced by dietary context. In this study, microscopy and transcriptomics were used to examine how the efficacy of this virus was affected when T. ni larvae were raised on different diets. Larvae raised on potato host plants had lower chitinase and chitin deacetylase transcript levels …

Examination Of Bcaa Transport And Synthesis In Staphylococcus Aureus, Julienne Kaiser

Electronic Thesis and Dissertation Repository

Staphylococcus aureus is a human pathogen capable of causing infections that range from mild skin and soft tissue infections to severe infections of the bone, muscle, heart, and lung. To survive and thrive in such diverse host environments, S. aureus must maintain sufficient levels of metabolites and cofactors to support virulence determinant production and replication. The branched-chain amino acids (BCAAs; Ile, Leu, Val) represent an important group of nutrients for S. aureus metabolism, as they are required for protein synthesis and synthesis of membrane branched-chain fatty acids (BCFAs), which are important for S. aureus environmental adaptation. Moreover, the BCAAs are …

Developing A Plant Virus-Based Expression System For The Expression Of Vaccines Against Porcine Reproductive And Respiratory Syndrome Virus, Hong Hanh Tran

Electronic Thesis and Dissertation Repository

Virus-based expression systems have been widely exploited for the production of recombinant proteins in plants during the last three decades. Advances in technology have boosted scale-up manufacturing of plant-made pharmaceuticals to high levels, via the complementation of transient expression and viral vectors. This combination allows proteins of interest to be produced in plants within a matter of days and thus, is well suited for the development of plant-made vaccines or therapeutics against emerging infectious diseases and potential bioterrorism agents. Several plant-based products are currently in varying stages of clinical development. To investigate the viability of virus-based expression systems for plant-made …

Disturbance And The Community Composition Of Arbuscular Mycorrhizal Fungi In Ontario Tallgrass Prairies, Sarah N. Allan

Electronic Thesis and Dissertation Repository

Arbuscular mycorrhizal fungi (Glomeromycota) form beneficial symbiotic relationships with the majority of land plants, especially in grasslands. Tallgrass prairies in Ontario are an endangered plant community currently being restored on former agricultural land. The objectives of my study were to determine if there were differences in the communities of Glomeromycota between disturbed and undisturbed tallgrass prairies in Ontario and if there were any potential indicator taxa for the ends of the disturbance spectrum. A molecular approach using DNA derived from soil samples was used to compare species composition between disturbed and undisturbed tallgrass prairies. A total of 177 operational taxonomic …

Characterizing The Interaction Between Human Adenovirus E1a And The Transcriptional Repressor Bs69, Ali Zhang

Electronic Thesis and Dissertation Repository

Protein products of the Early Region 1A (E1A) gene in human adenovirus 5 (HAdV-5) are the first viral proteins expressed upon adenovirus infection. E1A disrupts many cellular physiological events by binding to and regulating an impressive number of host factors. Of particular interest is BS69, a repressor of E1A transactivation. Due to the strong interaction observed between E1A and BS69, I hypothesize that these two proteins function together to disrupt gene expression within an infected cell.

Using in silico modelling and a series of yeast two-hybrid assays, I determined that residues 112-119 of HAdV-5 E1A is the minimal interacting region …

Investigating Adenovirus E1a As An Rna Polymerase Ii C-Terminal Domain Mimic And Its Role In Transcription Activation, Kristianne Jc Galpin

Electronic Thesis and Dissertation Repository

Viruses rely on host cell machinery, often mimicking cellular components, in order to circumvent host cell defenses and hijack cellular processes. DNA viruses, such as human Adenovirus (hAdV), rely on RNA Polymerase II (RNAPII) to transcribe viral genes. RNAPII has a C-terminal domain (CTD), made up of highly conserved heptad repeats of tyrosine-serine-proline-threonine-serine-proline-serine (YSPTSPS). Post-translational modifications of residues within the CTD, including phosphorylation, coordinates the transcription cycle. Several viruses, including Human Immunodeficiency Virus (HIV), Human Cytomegalovirus (hCMV), Epstein-Bar Virus (EBV) and Herpes Simplex Virus (HSV), modify the phosphorylation state of the RNAPII CTD by hijacking cellular cyclin dependent kinases (CDKs) …

Microbial Repopulation Following In Situ Star Remediation, Gavin Overbeeke

Electronic Thesis and Dissertation Repository

In situ STAR (Self-sustaining Treatment for Active Remediation) is an emerging remediation technology which uses smouldering combustion to destroy nonaqueous phase liquid (NAPL) contamination in the subsurface. Since STAR smouldering travels through contaminated soils slowly (~0.5 to 5 m/day) and subjects them to high temperatures (400–1000°C), it is expected that this technology will thoroughly dry and sterilize the zones which it treats. Further, soils surrounding the treatment zone which are not smouldered will be heated, although not smouldered, by virtue of their proximity to STAR, impacting microbial communities within them. Therefore, the objectives of this work are to quantify the …

Species Pluralism: Conceptual, Ontological, And Practical Dimensions, Justin Bzovy

Electronic Thesis and Dissertation Repository

Species are central to biology, but there is currently no agreement on what the adequate species concept should be, and many have adopted a pluralist stance: different species concepts will be required for different purposes. This thesis is a multidimensional analysis of species pluralism. First I explicate how pluralism differs monism and relativism. I then consider the history of species pluralism. I argue that we must re-frame the species problem, and that re-evaluating Aristotle's role in the histories of systematics can shed light on pluralism. Next I consider different forms of pluralism: evolutionary and extra-evolutionary species pluralism, which differ in …