Microbiology Commons^™

Iron-Regulated Cyanobacterial Predominance And Siderophore Production In Oligotrophic Freshwater Lakes, Ryan J. Sorichetti

Electronic Thesis and Dissertation Repository

The frequency and intensity of cyanobacterial blooms (cyanoblooms) is increasing globally. Contrary to existing phosphorus (P) and nitrogen (N) paradigms describing cyanobloom proliferation in eutrophic (nutrient-rich) freshwater lakes, many of the recent cyanobloom reports pertain to oligotrophic (nutrient-poor) freshwater lakes with no prior history of cyanobloom occurrence. There exists a critical research need to re-visit existing conceptual models, identify regulating factors currently unaccounted for and improve our ability to effectively detect and measure cyanobacterial toxins (cyanotoxins) in lakes. Iron (Fe) is required in nearly all pathways of cyanobacterial macronutrient use, though its direct role in regulating cyanobacterial biomass is not …

Heterologous Production And Characterization Of Cell Wall Degrading Enzymes Using Plants As A Bioreactor, Eridan Orlando Rodrigues Pereira

Electronic Thesis and Dissertation Repository

Plants are wonderful living organisms. They are able to store solar energy into carbohydrates by fixing CO₂ through photosynthesis which can be subsequently harvested and used for fuel production. However, one of the major limitations for transforming these carbohydrates into liquid fuels is the recalcitrance of the plant cell wall. Although microorganisms have evolved a series of cell wall degrading enzymes to harvest efficiently this energy and are considered the main source of these biocatalysts, harnessing these microorganisms for the production of enzymes is a costly process and a major factor limiting the commercialization of lignocellulosic biomass-to-ethanol processes. The …

Pegylation As A Novel Tool To Investigate The Topology Of Escherichia Coli Weca, A Membrane Enzyme Involved In Lipopolysaccharide O Antigen Initiation, Stéphanie L. Lamothe

Electronic Thesis and Dissertation Repository

O-antigen, the most surface exposed moiety of bacterial lipopolysaccharide (LPS), plays several roles in pathogenicity. The biosynthesis of O-antigen starts by the formation of a phosphoanhydride bond linking a sugar phosphate with a membrane isoprenoid lipid phosphate. Two distinct families of integral membrane proteins catalyze this reaction. The protein WecA is the prototypic member of one of these families, termed the polyisoprenyl-phosphate N-acetylaminosugar-1-phosphate transferase (PNPT) family. Because the donor nucleotide sugar is only available in the cytosol, cytosolic exposed regions of WecA are expected to be critical for the catalytic activity of the enzyme. Therefore, elucidating an accurate topological …

Characterization Of Staphylococcus Aureus Lipase, Vithooshan Vijayakumaran

Electronic Thesis and Dissertation Repository

USA300, a strain of community-associated methicillin resistant Staphylococcus aureus (CA-MRSA), has become prevalent in the community. Colonization of human skin requires mechanisms that allow this bacterium to overcome the innate immune defenses on the skin, including secretion of antimicrobial lipids. Antimicrobial lipids inhibit S. aureus growth and induce the staphylococcal proteolytic cascade, producing aureolysin (Aur) which processes the lipase glycerol ester hydrolase (Geh). Nearly all S. aureus strains secrete Geh, yet little information exists concerning its function. Using purified Aur and Geh we confirm that aureolysin processes proGeh to Geh. We then confirmed that geh was required for lipase activity …

Role Of Branched-Chain Amino Acid Transporters In Staphylococcus Aureus Virulence, Sameha Omer

Electronic Thesis and Dissertation Repository

Branched-chain amino acids (BCAAs) act as effector molecules that signal a global transcriptional regulator, CodY, to regulate virulence factors in nutrient depleted environments. Staphylococcus aureus contains three putative BCAA transporters (BrnQ1, BrnQ2, BrnQ3) whose role in BCAA uptake is unknown. We hypothesize that BrnQ transporters are involved in BCAA uptake and contribute to virulence in S. aureus by modulating CodY activity. Results from radioactive uptake assays indicate that BrnQ1 is the predominant BrnQ transporter of isoleucine, valine and leucine. Meanwhile, BrnQ2 is more specific for isoleucine. Furthermore, only the lack of BrnQ1 hinders growth of S. aureus in chemically-defined media …

The Interaction Of The Human Adenovirus E1a Protein With The Human Dref Transcription Factor, Kris M. James

Electronic Thesis and Dissertation Repository

The human adenovirus (HAdV) E1A protein is the first protein produced post-HAdV infection, and serves two main functions. The first is to modulate host and viral transcription. The second is to induce host cell cycle progression to S phase, to promote an optimal environment for viral replication. E1A performs its functions by binding and manipulating over 50 cellular factors. Interestingly, I found that E1A is capable of interacting with the poorly characterized human DNA replication-related element-binding factor (hDREF). hDREF is a transcription factor associated with the expression of several genes related to the cell cycle. I hypothesized that the interaction …

Human Adenovirus E1a Binds And Retasks Cellular Hbre1, Blocking Interferon Signalling And Activating Virus Early Gene Transcription, Gregory J. Fonseca

Electronic Thesis and Dissertation Repository

Upon infection, human adenovirus (HAdV) must block interferon signaling and activate the expression of its early genes to reprogram the cellular environment to support virus replication. During the initial phase of infection, these processes are orchestrated by the first HAdV gene expressed during infection, early region 1A (E1A). E1A binds and appropriates components of the cellular transcriptional machinery to modulate cellular gene transcription and activate viral early genes transcription. We have identified hBre1/RNF20 as a novel target of E1A. hBre1 is an E3 ubiquitin ligase which acts with the Ube2b E2 conjugase and accessory factors RNF40 and WAC1 to monoubiquitinate …

Systematic Analysis Of Residues In Conserved Region 3 Of The Human Papillomavirus 16 E7 Oncoprotein, Biljana Todorovic

Electronic Thesis and Dissertation Repository

Although remarkable biological diversity is exhibited by viruses, as obligate intracellular parasites, they rely on host cell functions. As such, viruses typically must overcome a set of host barriers that prevent infection. For human papillomaviruses (HPV) one of these barriers is the state of terminal differentiation of the host cell. For that purpose HPVs encode two major oncoproteins, E6 and E7, which combine their efforts to effectively uncouple cellular differentiation from the cell cycle arrest. The E7 proteins have no intrinsic enzymatic activity or DNA binding ability, but they bind and manipulate numerous host proteins. E7 is a modular oncoprotein …

An Investigation Of Plant-Microbe Interactions Under Cadmium Stress In Agar-Based Medium, Hydroponics, And Soil Studies, Melanie P. Columbus

Electronic Thesis and Dissertation Repository

This thesis investigated plant-microbe-metal interactions at two scales: a single plant-microbe system and an agricultural rhizobacterial community. The first objective was to investigate the effectiveness of a plant growth-promoting rhizobacterium (PGPR) on mediating cadmium stress in a plant model system. Arabidopsis thaliana Col-0 was inoculated with Pseudomonas putida UW4, which in its wild type form has been reported to reduce plant stress by simultaneously metabolizing the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC) with the enzyme ACC deaminase and stimulating plant growth through the production of indole-3-acetic acid (IAA). A mutant strain that lacks ACC deaminase and a no bacteria treatment were used …

Staphylococcus Aureus Response To Long Chain Antimicrobial Fatty Acids, Benjamin Arsic

Electronic Thesis and Dissertation Repository

Staphylococcus aureus is a common pathogen that has the ability to infect virtually every tissue and organ system of the body. Despite its propensity to cause invasive infection, S. aureus is also a commensal organism, asymptomatically colonizing ~25% of the population. Much research has gone into resolving this paradox, focusing on both human and bacterial factors that may contribute to colonization. Antimicrobial fatty acids present on the skin and in the nasal mucosa are important components of the innate immune system, and thus we undertook to further understand how S. aureus responds to these fatty acids, and how this response …

Protein Body Formation In Stable Transgenic Plants Of Nicotiana Tabacum Expressing Elastin-Like Polypeptide And Hydrophobin Fusion Proteins, Sonia P. Gutierrez

Electronic Thesis and Dissertation Repository

Plants are recognized as an efficient and inexpensive system to produce valuable recombinant proteins. However, the use of plants still faces two main limitations: the low accumulation levels of some recombinant proteins and the lack of efficient protein purification methods. Two fusion partners, elastin-like polypeptides (ELP) and hydrophobin I (HFBI) were found to increase the accumulation of recombinant proteins and induce the formation of protein bodies (PBs) when targeted to the ER in transient expression assays. In this study I examined the effect of these tags in stable transgenic plants of two Nicotiana tabacum cultivars when fused to green fluorescent …

Sequesteration Of Lead, Cadmium And Arsenic By Lactobacillus Species And Detoxication Potential, Marc A. Monachese

Electronic Thesis and Dissertation Repository

Toxic metals are a class of elements with no biological role but with extreme toxicity. On average only 50% of ingested toxins are absorbed into the human body, for reasons still unknown. It was hypothesized that the gut microbiota plays a role in reducing toxin absorbance. The aim of this study was to determine if constituents of the gut, namely Lactobacillus species, are able to sequester arsenic, lead and cadmium from the environment. Lactobacilli were incubated with the metals, both in vitro and with a Caco-2 cell line. Analysis of metal concentrations was conducted to determine if these were reduced …

Colonization Of Zea Mays By The Nitrogen Fixing Bacterium Gluconacetobacter Diazotrophicus, Nikita Eskin

Electronic Thesis and Dissertation Repository

To achieve high yields, corn fields are supplemented with nitrogen fertilizers. Nitrogen fertilizers account for a significant portion of production costs, and are harmful to the environment. Gluconacetobacter diazotrophicus, an endophytic nitrogen-fixing bacterium supplies its natural host sugarcane with a significant amount of nitrogen. This study investigated the colonization of G. diazotrophicus in seven different corn genotypes consisting of sweet and grain corn via three different methods of inoculation: soil drench, root dip, and aseptic inoculation. Sucrose content of the corn genotypes and nitrogenase activity were also analysed. Colonization was confirmed by PCR analysis. G. diazotrophicus colonization was detected using …

Restriction Of Hiv-1 Replication By Unique Trim22 Isoforms., Clayton Hattlmann

Electronic Thesis and Dissertation Repository

Understanding how the immune system reacts to HIV infection and why normal antiviral defenses are insufficient to fight infection is a key step towards creating better therapies. Several interferon-induced proteins, such as the tripartite motif protein TRIM22, are capable of restricting HIV-1 replication; however single nucleotide polymorphisms (SNPs) can dramatically impact the actions of these proteins. While the trim22 gene contains numerous SNPs, no study has addressed how these may affect TRIM22 functions. Here we provide the first direct comparison of two TRIM22 unique isoforms. Through confocal microscopy we observed these isoforms exhibit different patterns of localization. In vitro studies …

The Intracellular Behaviour Of Burkholderia Cenocepacia In Murine Macrophages, Jennifer S. Tolman

Electronic Thesis and Dissertation Repository

Burkholderia cenocepacia is an opportunistic pathogen causing life-threatening infections in cystic fibrosis and other immunocompromised patients. The bacterium survives within macrophages by interfering with typical endocytic trafficking, resulting in delayed maturation of a B. cenocepacia-containing phagosome. We hypothesize that B. cenocepacia alters gene expression after internalization by macrophages, inducing genes involved in intracellular survival and host adaptation. Furthermore, we hypothesize that specialized bacterial secretion systems are involved in the interactions between intracellular bacteria and macrophages. In this work, we characterize later-stage infection of macrophages by B. cenocepacia, showing replication within an acidified endosomal compartment suggestive of a phagolysosome. …

Critical Stages In Viral Replication - Entry, Gene Regulation And Egress, Matthew S. Miller

Electronic Thesis and Dissertation Repository

Despite the amazing biological diversity exhibited by viruses, their very existence relies upon their ability to overcome a set of common barriers. The nature of these barriers reflects the nature of viruses themselves. During their extracellular phase, viruses are metabolically inert obligate parasites. Upon encountering a host cell, productive infection necessitates that the virus successfully enter the cell, regulate the expression of its genes, and after assembling new progeny particles, egress such that the cycle of infection can continue. These three basic processes are not only attractive candidates for therapeutic intervention, but also reveal much about virus biology in the …

Structure-Function Analysis Of Enzymes Of The Polyisoprenyl-Phosphate Hexose-1-Phosphate Transferase Family, Kinnari B. Patel

Electronic Thesis and Dissertation Repository

Enzymes of the polyisoprenyl-phosphate hexose-1-phosphate transferase (PHPT) family are integral membrane proteins that initiate the synthesis of glycans by catalyzing the transfer of a hexose-1-phosphate sugar from UDP-hexose to the lipid carrier undecaprenyl phosphate (Und-P). These glycans such as O antigen and exopolysaccharide (EPS) provide bacteria with protection and adaptation to the environment and host immune factors. The role of PHPT proteins in initiation and the absence of any eukaryotic homologues make them an attractive target for novel antimicrobials; however study of these proteins is difficult due to the presence of multiple transmembrane helices. A requirement of the C-terminal domain …

Exploring Bacterial Nanowires: From Properties To Functions And Implications, Kar Man Leung

Electronic Thesis and Dissertation Repository

The discovery of electrically conductive bacterial nanowires from a broad range of microbes provides completely new insights into microbial physiology. Shewanella oneidensis strain MR-1, a dissimilatory metal-reducing bacterium, produces extracellular bacterial nanowires up to tens of micrometers long, with a lateral dimension of ~10 nm. The Shewanella bacterial nanowires are efficient electrical conductors as revealed by scanning probe techniques such as CP-AFM and STM.

Direct electrical transport measurements along Shewanella nanowires reveal a measured nanowire resistivity on the order of 1 Ω∙cm. With electron transport rates up to 10⁹/s at 100 mV, bacterial nanowires can serve as a …

Transactivation By Human Adenovirus Early Region 1a-Conserved Region Three, Jailal Ng Ablack

Electronic Thesis and Dissertation Repository

One of the critical functions of human adenovirus (hAd) early region 1A (E1A) protein is to activate transcription of the early viral genes. The largest isoform of E1A contains a unique region termed conserved region 3 (CR3), which includes a Cysteine-4 (C4) zinc finger domain. This region activates viral gene expression by interacting with and recruiting cellular transcription machinery to the regulatory regions of early viral genes. Although this process has been studied at length with hAd type 5 E1A, far less is known about how the E1A proteins from other hAd types activate transcription. There are dramatic differences in …

The Hiv-1 Tat Protein And Adverse Drug Reactions: A Model System Utilizing Jurkat T Cells And Sulphamethoxazole-Hydroxylamine, Kaothara Adeyanju

Electronic Thesis and Dissertation Repository

In 2009 approximately 2.6 million people became infected with the Human Immunodeficiency Virus (HIV). In addition to the estimated 33.3 million people currently living with the virus, this makes HIV/ AIDS an epidemic of unprecedented scale in modern times. Treatment of HIV infection requires antiretroviral agents as well as a number of other drugs such as antimicrobials. Hypersensitivity adverse drug reactions (ADRs) to a variety of drugs are common in HIV-infected individuals, but the antimicrobial Sulphamethoxazole remains a major culprit. Hypersensitivity ADRs cause significant morbidity, with the skin and liver most commonly affected and are among the top causes of …