Microbiology Commons^™

Characterization Of Silicon Ion Exposure On Deinococcus Radiodurans, Richard F. Daughtry

Theses and Dissertations

Deinococcus radiodurans is a robust bacterium that is known for its extraordinary resistance to ionizing radiation. In general, many of the investigations of this bacterium's resistance have revolved around low linear energy transfer radiation, such as gamma and electron radiation. This study explored Deinococcus radiodurans's ability to survive high linear energy transfer radiation, specifically proton and neutron radiation. Deinococcus radiodurans was dehydrated to reduce the effects of low linear energy transfer radiation. The bacteria were exposed to both neutron and proton radiation of varying amounts and rehydrated. The resulting colonies were counted and compared to colonies of non-irradiated control samples …

Characterization Of The Effects Of Heavy Charged Particle Exposure On The Radiation Resistant Bacterium Deinococcus Radiodurans, Todd A. Bryant

Theses and Dissertations

Although not an extremophile, Deinococcus radiodurans has proven to have exceptional resistance to ionizing radiation, specifically via gamma and X-ray photons. To date, no known experiments have bombarded the bacterium with charged particles larger than hydrogen. This study explored the effects on the organism’s ability to survive high linear energy transfer heavy-charged particle exposures of oxygen ions. It also investigated the effects of low linear energy transfer ultraviolet radiation on various mutants. Two uvrB mutants were created to ascertain the importance of the gene in single-strand break repair following gamma irradiation and two recF mutants were created to explore the …

Assessment Of A Predicted Diguanylate Cyclase In Treponema Denticola, Dhara T. Patel

Theses and Dissertations

Periodontal disease is a progressive inflammatory condition that is characterized by the reabsorption of alveolar bone, the destruction of connective tissue, and edentulism. It is caused by a dysbiosis in the oral microbiome as a result of a shift from a Gram-positive aerobic bacterial population, to one that becomes more Gram-negative and anaerobic. Treponema denticola is thought to drive this diseased state based on its role as a keystone periopathogen. A major component of T.denticola’s invasiveness is its motility, which allows the spirochete to penetrate and disseminate through tissues. This motility, which has been seen to be crucial to …

Acid Sphingomyelinase Is Essential For Vacuolar Development Of A. Phagocytophilum., Chelsea Cockburn

Theses and Dissertations

Obligate intracellular bacteria are significant causes of morbidity and mortality with over two hundred and fifty million infections worldwide annually. One such bacterium, Anaplasma phagocytophilum is the etiologic agent of human granulocytic anaplasmosis (HGA), a tick-transmitted febrile illness. Previous studies have shown that A. phagocytophilum lacks genes for cholesterol biosynthesis and solely relies on Niemann Pick protein type C (NPC)1-mediated low density lipoprotein (LDL)-derived cholesterol to complete its infection cycle.Acid sphingomyelinase (ASMase) is a lysosomal enzyme that is essential for diverse cellular processes including liberation of LDL-derived cholesterol from the lysosome. By first studying A. phagocytophilum, we found that …

Orientia Tsutsugamushi Modulates Endoplasmic Reticulum Stress To Benefit Its Intracellular Growth And Targets Nlrc5 To Inhibit Major Histocompatibility Complex I Expression, Kyle G. Rodino

Theses and Dissertations

Scrub typhus, caused by the obligate intracellular bacterium Orientia tsutsugamushi, afflicts one million people annually. Despite being a global health threat, little is known about O. tsutsugamushi pathogenesis. Here, we demonstrate that O. tsutsugamushi modulates the ER and ER-associated processes as mechanisms of nutritional virulence and immune evasion. To obtain amino acids to fuel replication, O. tsutsugamushi simultaneously induces ER stress and the unfolded protein response (UPR) while inhibiting ER-associated degradation (ERAD) during early infection time points. During exponential growth, the bacterium releases the ER bottleneck, resulting in generation of ERAD-derived amino acids that it parasitized for replication. …

Evolution And Niche Specialization Of Microbial Taxa In Vaginal Infection And Pregnancy, Abigail L. Glascock

Theses and Dissertations

The vaginal microbiome plays an important role in reproductive health and pregnancy. It has coevolved with humans and has direct effects on reproductive success, rendering selective pressure more pronounced at this site. Herein, we probe coevolution of the vaginal microbiome using a systems-level approach. In Chapter 2, we examine the evolutionary trajectory of two vaginal Veillonellaceae phylotypes evolved from an ancestral gastrointestinal lineage to inhabit the vaginal niche. We present evidence of their divergence and subniche specification and describe their differential associations with vaginal infection and pregnancy. In Chapter 3, we identify ten bacterial taxa, predicted to contribute to the …

Influence Of Epstein-Barr Virus On Systemic Lupus Erythematosus Disease Development And The Role Of Depression On Disease Progression, Caleb Cornaby

Theses and Dissertations

Systemic Lupus Erythematosus (SLE) is an autoimmune disease affecting 20 to 250 individuals per 100,000 worldwide. Symptomology includes dermatological manifestations such as discoid lesions, acute cutaneous rashes, and oral and nasal ulcers, along with musculoskeletal, pulmonary, and renal complications. Abnormal T and B lymphocyte function and apoptosis, immune complex clearance, complement function, and nucleosome processing are typical of disease pathophysiology. SLE is the result of both environmental and genetic factors, which together create the conditions leading to disease onset and progression. Of these environmental factors, Epstein-Barr virus (EBV) infection is known to cause the genesis of cross-reactive antibodies in SLE …

Acetobacter Fabarum Genes Influencing Drosophila Melanogaster Phenotypes, Kylie Makay White

Theses and Dissertations

Research in our lab has predicted hundreds of bacterial genes that influence nine different traits in the fruit fly, Drosophila melanogaster. As a practical alternative to creating site-directed mutants for each of the predicted genes, we created an arrayed transposon insertion library using a strain of Acetobacter fabarum DsW_054 isolated from fruit flies. Creation of the Acetobacter fabarum DsW_054 gene knock-out library was done through random transposon insertion, combinatorial mapping and Illumina sequencing. Successful mapping of transposon insertion was achieved for 6418 mutants with hits within 63% of annotated genes within Acetobacter fabarum DsW_054. Insertion sites were verified in …

Phosphate Signaling Through Alternate Conformations Of The Pstscab Phosphate Transporter, Ramesh Krishna Vuppada

Theses and Dissertations

Phosphate is an essential compound for life. Escherichia coli employs a signal transduction pathway that controls the expression of genes that are required for the high-affinity acquisition of phosphate and the utilization of alternate sources of phosphorous. These genes are only expressed when environmental phosphate is limiting. The seven genes for this signaling pathway encode the two-component regulatory proteins PhoB and PhoR, as well as the high-affinity phosphate transporter PstSCAB and an auxiliary protein called PhoU. As the sensor kinase PhoR has no periplasmic sensory domain, the mechanism by which these cells sense environmental phosphate is not known. This paper …

The Effects Of Nucleosome Positioning And Chromatin Architecture On Transgene Expression, Colton E. Kempton

Theses and Dissertations

Eukaryotes use proteins to carefully package and compact their genomes to fit into the nuclei of their individual cells. Nucleosomes are the primary level of compaction. Nucleosomes are formed when DNA wraps around an octamer of histone proteins and a nucleosome's position can limit access to genetic regulatory elements. Therefore, nucleosomes represent a basic level of gene regulation. DNA and its associated proteins, called chromatin, is usually classified as euchromatin or heterochromatin. Euchromatin is transcriptionally active with loosely packed nucleosomes while heterochromatin is condensed with tightly packed nucleosomes and is transcriptionally silent. In order to become active, heterochromatin must first …

Comparison Of Cytokine Expression And Bacterial Growth During Periparturient And Mid Lactation Mastitis In A Mouse Model, Rhonda Nicole Chronis

Theses and Dissertations

Clinical cases of bovine mastitis are most severe in the early stages of lactation. The causes of this increased propensity for severe mastitis during early lactation, compared to mid and late lactation are unclear. In order to better understand the early lactation immune response to mastitis, a murine model of mastitis was employed. Intramammary inoculation of a mastitis causing Escherichia coli strain was performed in lactating mice at various stages of lactation to model the immune response seen in cows during lactation. In our experiments, mice in the early stages of lactation exhibited altered mRNA expression of cytokines IL-1β, IL-6, …

Characterization Of Neuronal Nicotinic Acetylcholine Receptors And Their Positive Allosteric Modulators, Doris Clark Jackson

Theses and Dissertations

Neuronal nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels that are necessary in memory and cognition. They are pentameric and consist of α and β subunits. They are most commonly heteromeric but, can sometimes be homomeric. nAChRs are activated by many ligands including nicotine (exogenous) and acetylcholine (endogenous).nAChRs are located on hippocampal interneurons. The interneurons, although sparse, control the synchronous firing of the pyramidal cells. However, the hippocampal interneuron structure and function is quite diverse and not fully characterized. Therefore, we sought to quantify nAChR subunit mRNA levels using real-time PCR of CA1 hippocampal interneurons.Surprisingly we found that the α3 …

The Effects Of The Hiv-1 Tat Protein And Morphine On The Structure And Function Of The Hippocampal Ca1 Subfield, William D. Marks

Theses and Dissertations

HIV is capable of causing a set of neurological diseases collectively termed the HIV Associated Neurocognitive Disorders (HAND). Worsening pathology is observed in HIV+ individuals who use opioid drugs. Memory problems are often observed in HAND, implicating HIV pathology in the hippocampus, and are also known to be exacerbated by morphine use. HIV-1 Tat was demonstrated to reduce spatial memory performance in multiple tasks, and individual subsets of CA1 interneurons were found to be selectively vulnerable to the effects of Tat, notably nNOS+/NPY- interneurons of the pyramidal layer and stratum radiatum, PV+ neurons of the pyramidal layer, and SST+ neurons …

Orientia Tsutsugamushi Secretes Two Ankyrin Repeat-Containing Effectors Via A Type 1 Secretion System To Inhibit Host Nf-Κb Function, Sean M. Evans

Theses and Dissertations

Scrub typhus is a potentially fatal infection that threatens one billion persons in the Asia-Pacific region and is caused by the obligate intracellular bacterium, Orientia tsutsugamushi. How this organism facilitates its intracellular survival and pathogenesis is poorly understood. Intracellular bacterial pathogens utilize the Type 1 (T1SS) or Type 4 secretion system (T4SS) to translocate ankyrin repeat-containing proteins (Anks) into the host cell to modulate host cell processes. The O. tsutsugamushi genome encodes one of the largest known bacterial Ank libraries as well as Type 1 and Type 4 secretion systems (T1SS and T4SS), which are expressed during infection. In …

Contribution Of A Class Ii Ribonucleotide Reductase To The Manganese Dependence Of Streptococcus Sanguinis, John L. Smith

Theses and Dissertations

Manganese-deficient Streptococcus sanguinis mutants exhibit a dramatic decrease in virulence for infective endocarditis and in aerobic growth in manganese-limited media. Loss of activity of a manganese-dependent, oxygen-dependent ribonucleotide reductase (RNR) could explain the decrease in virulence. When the genes encoding this RNR are deleted, there is no growth of the mutant in aerobic broth culture or in an animal model. Testing the contribution of the aerobic RNR to the phenotype of a manganese transporter mutant, a heterologous class II RNR from Lactobacillus leichmannii called NrdJ that requires B₁₂ rather than manganese as a cofactor was previously introduced into an …

Amixicile Inhibits Anaerobic Bacteria Within An Oral Microbiome Derived From Patients With Chronic Periodontitis, Kane Ramsey

Theses and Dissertations

Periodontitis is a chronic inflammatory disease caused by pathogenic bacteria residing in a complex biofilm within a susceptible host. Amixicile is a non-toxic, readily bioavailable novel antimicrobial that targets strict anaerobes through inhibition of the activity of Pyruvate Ferredoxin Oxidoreductase (PFOR), a major enzyme mediating oxidative decarboxylation of pyruvate. Our study aimed to evaluate the efficacy of amixicile, when compared to metronidazole, in inhibiting the growth of bacteria present in a microbiome harvested from patients with chronic periodontitis.

Plaque samples were harvested from patients with severe chronic periodontitis and cultured under anaerobic conditions. The microbiomes were grown in the presence …

Come Fly With Me: Using Amixicile To Target Periodontal Pathogens And Elucidating The Innate Immune Response In Drosophila Melanogaster, Kathryn Sinclair

Theses and Dissertations

Periodontal diseases (PD) affect 46% of American adults over age 30. These diseases cause symptoms including bleeding and swelling of the gums, bone resorption, and tooth loss, that affect quality of life and have a high economic burden. Periodontal diseases are caused by an imbalance in the oral microbiome, from a healthy state that contains anti-inflammatory commensals like Streptococcus gordonii and mitis, to a diseased state that has pro-inflammatory anaerobic pathogens including Porphyromonas gingivalis, Prevotella intermedia, Fusobacterium nucleatum, and Tannerella forsythia. The latter initiate disease progression in the oral cavity. However, it’s the host immune …

Discovery And Characterization Of Bile Acid And Steroid Metabolism Pathways In Gut-Associated Microbes, Spencer Harris

Theses and Dissertations

The human gut microbiome is a complex microbial ecosystem residing in the lumen of our gastrointestinal tract. The type and amounts of microbes present in this ecosystem varies based on numerous factors, including host genetics, diet, and environmental factors. The human gut microbiome plays an important role in normal host physiological functions, including providing energy to colonocytes in the form of short-chain fatty acids. However, gut microbial metabolites have also been associated with numerous disease states. Current tools for analyzing the gut microbiome, such as high-throughput sequencing techniques, are limited in their predictive ability. Additionally, “-omic” approaches of studying the …

Hub Proteins, Paralogs, And Unknown Proteins In Bacterial Interaction Networks, Neha Sakhawalkar

Theses and Dissertations

Proteins are the functional units of cells. However, a major portion of the proteome does not have a known functional annotation. This dissertation explores protein -protein interactions, involving these uncharacterized or unknown function proteins. Initially, protein – protein interactions were tested and analyzed for paralogous proteins in Escherichia coli. To expand this concept further and to get an overview, protein – protein interactions were analyzed using ‘comparative interactomics’ for four pathogenic bacterial species including Escherichia coli, Yersinia pestis, Vibrio cholerae and Staphylococcus aureus. This approach was used to study unknown function protein pairs as well as to …

In Search Of A Function For An Uncharacterized Conserved Protein In Streptococcus Sanguinis Sk36, Ayana Scott-Elliston

Theses and Dissertations

With the number of fully sequenced bacterial genomes increasing in the past 7 years, it has been discovered that a large percentage of the putative protein coding genes have no known function. This lack of knowledge leaves scientists with an incomplete understanding of bacteria. In this study, conserved hypothetical protein mutants from Streptococcus sanguinis SK36 were screened on solid media with various environmental conditions. From these screens, the candidate protein, SSA_2372, displayed a sensitivity to acidic conditions. Its homolog in Bacillus subtilis 168, BSU00030, also displayed a sensitivity to pH conditions at its acid tolerance extremes unlike its other homolog …

A Systems Biology Approach For Predicting Essential Genes And Deciphering Their Dynamics Under Stress In Streptococcus Sanguinis, Fadi El-Rami

Theses and Dissertations

Infectious diseases are the top leading cause of death worldwide. Identifying essential genes, genes indispensable for survival, has been proven indispensable in defining new therapeutic targets against pathogens, major elements of the minimal set genome to be harnessed in synthetic biology, and determinants of evolutionary relationships of phylogenetically distant species. Thus, essentiality studies promise valuable revenues that can decipher much of biological complexities.

Taking advantage of the available microbial sequences and the essentiality studies conducted in various microbial models, we proposed a framework for the prediction of essential genes based on our experimentally verified knowledge of the pathways involved in …

Expression Of Matrix Metalloproteinases In Naegleria Fowleri And Their Role In Degradation Of The Extracellular Matrix, Charlton Lam

Theses and Dissertations

Naegleria fowleri is a free-living amoeba found in freshwater lakes and ponds that is the causative agent of Primary Amoebic Meningoencephalitis (PAM). Matrix metalloproteinases (MMPs) have been described in protozoa, such as Plasmodium falciparum, Trypanosoma brucei, and Balamuthia mandrillaris, and have been linked to their increased motility and invasive capability by degrading components of the extracellular matrix (ECM). In addition, MMPs are often upregulated in tumorigenic cells and have been attributed as responsible for the metastasis of certain cancers. In the present study, in vitro experiments indicated that MMPs are linked functionally to the ECM degradation process. …

The Role Of Viral Interleukin-6 In Tumor Development Of Kaposi's Sarcoma-Associated Herpesvirus Lymphomas, Rebecca A. Fullwood

Theses and Dissertations

Kaposi's sarcoma herpesvirus (KSHV) is a cancer-causing virus, primarily affecting AIDS patients. KSHV is found in 3-10% of the U.S. population and can cause a range of cancers in the highly immunosuppressed; these cancers include Kaposi's sarcoma, pleural effusion lymphoma (PEL) and multicentric Castleman's disease (MCD). The current techniques for treating these cancers are relatively ineffective, largely due to their inefficiency at targeting tumors formed by the infection. One protein produced by KSHV, the viral homolog of interleukin-6 (vIL-6), is thought to play a major role in tumor development post-infection. Here a novel animal model is implemented to study the …

Analysis Of Nucleosome Isolation And Recovery: From In Silico Invitrosomes To In Vivo Nucleosomes, Collin Brendan Skousen

Theses and Dissertations

There are a vast number of factors that influence nucleosome formation, and consequently gene regulation. These factors include histone modifications, nucleotide composition, transcriptional region elements, and specific nucleotide motifs, among others. Although the amount we know now is limited, we are creating new techniques and discoveries to assist us in continued understanding of chromatin. To make a significant contribution to the field of chromatin, I conducted two hypothesis driven sets of experiments that address the topic of chromatin structure. First, I created a technique for tissue specific nucleosome isolation with the goal of observing the effect of single nucleotide polymorphisms …

Genetic And Biochemical Analysis Of The Micrococcin Biosynthetic Pathway, Philip Ross Bennallack

Theses and Dissertations

Declining antibiotic discovery and flourishing antibiotic resistance have led to a modern antibiotic crisis which threatens to compromise our ability to treat infectious disease. Consequently, there is significant interest in developing new antibiotics with novel modes of action and chemical properties. Ribosomally synthesized and post-translationally modified peptides (RiPPs) are natural compounds with the appealing attributes of being derived directly from a genetic template while possessing numerous exotic chemical features that contribute to stability and antimicrobial activity. Abundant in nature, their diverse range of biological activities makes them excellent prospects for antibiotic development. Thiopeptides, a RiPP family rich in chemical complexity, …

A Cryab Interactome Reveals Clientele Specificity And Dysfunction Of Mutants Associated With Human Disease, Whitney Katherine Hoopes

Theses and Dissertations

Small Heat Shock Proteins (sHSP) are critical molecular chaperones that function to maintain protein homeostasis (proteostasis) and prevent the aggregation of other proteins during cellular stress. Any disruption in the process of proteostasis can lead to prevalent diseases ranging from cancer and cataract to cardiovascular and Alzheimer's disease. CryAB (αB-crystallin, HspB5) is one of ten known human sHSP that is abundant in the lens, skeletal, and cardiac muscle. This protein is required for cardiac function and muscle cell integrity. When the cell experiences physiological stress, including heat shock, CryAB moves to the cytoskeleton to act as a chaperone and prevent …

Membrane Biophysics Of Listeria Monocytogenes: Analysis Of An Alternate Pathway Of Branched-Chain Fatty Acid Biosynthesis And Elasticity Of Fatty Acid Utilization, Laura E. Kuczek

Theses and Dissertations

The membrane homeostasis of the psychrotolerant foodborne pathogen Listeria monocytogenes involves maintaining fluidity in a range of different conditions by incorporation of specific branched-chain fatty acids (BCFAs). BCFAs are synthesized through a well-characterized pathway from branched-chain amino acids into short branched-chain acyl-CoAs followed by elongation by the FAS II system. Branched-chain alpha-keto acid dehydrogenase (Bkd) is a major player in this pathway. MOR401 is a transposon insertion mutant deficient in Bkd and has decreased membrane BFCAs. Low levels of BCFAs in L. monocytogenes are linked to diminished growth, less resistance to antimicrobials, and a severe reduction in virulence. Rescued growth …

Characterizing Interaction Between Pask And Pbp1/ Atxn2 To Regulate Cell Growth And Proliferation, Nidhi Rajan Choksi

Theses and Dissertations

Pbp1 is a component of glucose deprivation induced stress granules and is involved in P-body-dependent granule assembly. We have recently shown that Pbp1 plays an important role in the interplay between three sensory protein kinases in yeast: AMP-regulated kinase (Snf1 in yeast), PAS kinase 1 (Psk1 in yeast), and the target of rapamycin complex 1 (TORC1), to regulate glucose allocation during nutrient depletion. This signaling cascade occurs through the SNF1-dependent phosphorylation and activation of Psk1, which phosphorylates and activates poly(A)- binding protein binding protein 1 (Pbp1), which then inhibits TORC1 through sequestration at stress granules. In this study we further …

The Activity Of Alkaline Glutaraldehyde Against Bacterial Endospores And Select Non-Enveloped Viruses, Justen Thalmus Despain

Theses and Dissertations

Alkaline glutaraldehyde (GTA) has been used as a high level chemical disinfectant and sterilant for many years and is known to kill a broad spectrum of organisms ranging from vegetative eukaryotes to bacterial endospores. Although the mechanism of sporicidal action has been studied on numerous occasions, GTA's exact mechanism(s) of action are still debated. In addition to the uncertainty of GTA's mechanism(s) of action, GTA has also shown significant variability in the time required to kill endospores and naked viruses. A better knowledge of the lethal mechanism(s) of GTA is needed to understand this discrepancy in kill times for GTA …

Synthesis And Biological Activity Of Novel Quorum Sensing Compounds, Joseph Nicholas Capilato

Theses and Dissertations

Bacteria communicate with chemical signals in a process known as quorum sensing. This population density-dependent process involves the bacterial production, release and detection of structurally specific small molecules and enables the bacterial pathogen to regulate its virulence on a population-wide level. Using a variety of chemical and biological techniques, I have studied various quorum sensing systems in several bacteria, including Vibrio cholera and Pseudomonas aeruginosa. A key principle of this research involves the design, synthesis and testing of novel compounds for their biological activity. These molecules are typically based off of an initial lead target, which is often identified …