Life Sciences Commons^™

Use Of Transposon Screening For Salicylic Acid-Assisted Desiccation Killing In Salmonella, Shannon D. Elliott

Electronic Theses and Dissertations

Salmonella enterica serovar Typhimurium is one of the most prevalent food-borne pathogens, affecting millions around the world every year, making it a threat to global health. Salmonella possesses the ability to survive the normally lethal condition of desiccation, however, discovery of the genes and mechanisms behind this phenomenon are still ongoing. Using a transposon mutagenesis approach to construct a broad transposon library, this study aimed to uncover genes that may be contributing to changes in Salmonella’s survivability under desiccation, particularly when exposed to the antimicrobial molecule salicylic acid. Building on previous findings showing salicylic acid can alter cell viability …

The Stringent Response In Pseudomonas Aeruginosa Influences The Phenotypes Controlled By The Gac/Rsm System, Michael Shawn Hooker

Electronic Theses and Dissertations

Pseudomonas aeruginosa is a ubiquitous, opportunistic pathogen that causes acute and chronic infections. Infection is typically initiated via motile and virulent strains. After exposure to stressors, acute infections make both genotypic and phenotypic switches to a chronic, sessile strain. This is due to intricate regulatory networks directing gene expression in response to stressors. One network, GacA/GacS, has been established to control virulence factors. The stringent response of bacteria is mediated by alarmones produced primarily by RelA which responds to starvation.

To study the effect of the stringent response on the virulence switch. A series of experiments were run in both …

Determining The Effects Of Maternal Adiposity On Preterm Neonatal Microbiome And Short Chain Fatty Acid Profiles, Dalton James, William A. Clark Phd, Kristy L. Thomas

Undergraduate Honors Theses

The gut microbiota and its metabolites have vast impacts on the human digestive system, immune system, and health outcomes. Short chain volatile fatty acids (SCVFAs) present in feces can be representative of the interactions of the microbiota present in the gut. Low microbiota diversity in the human gut is highly associated with obesity and adverse health outcomes. Furthermore, the maternal microbiome has a direct impact on neonatal microbiota through various pathways such as environment, skin flora, breast milk composition, and vaginal secretions. This study is aimed to further understand the associations between various factors (maternal adiposity, gestational time, length of …

Investigating The Pi3k/Akt/Atm Pathway, Telomeric Dna Damage, T Cell Death, And Crispr/Cas9-Mediated Gene Editing During Acute And Chronic Hiv Infection, Sushant Khanal

Electronic Theses and Dissertations

Human Immunodeficiency Virus (HIV) infection initiates major metabolic and cell- survival complications. Anti-retroviral therapy (ART) is the current approach to suppress active HIV replication to a level of undetected viral load, but it is not a curative approach. Newer and sophisticated gene editing technologies could indeed be a potent antiviral therapy to achieve a clinical sterilization/cure of HIV infection. Chronic HIV patients, even under a successful ART regimen, exhibit a low-grade inflammation, immune senescence, premature aging, telomeric DNA attrition, T cell apoptosis, and cellular homeostasis. In this dissertation, we investigated CD4 T cell homeostasis, degree of T cell apoptosis, an …

Cyclic Di-Gmp Regulates Motility, Biofilm Formation, And Desiccation Tolerance In Acinetobacter Baumannii, Garrett Reynolds

Electronic Theses and Dissertations

Acinetobacter baumannii is an increasingly multidrug-resistant pathogen contributing to hospital-acquired infections necessitating the discovery of novel treatments. A bacterial second messenger, cyclic diguanosine monophosphate (cyclic di-GMP), can regulate various persistence factors that are potentially advantageous for survival in hospital environments. Cyclic di-GMP–modulating enzymes and cyclic di-GMP–binding effectors predictively are encoded in the Acinetobacter baumannii genome. I hypothesized that cyclic di-GMP controls motility, biofilm formation, and desiccation tolerance in Acinetobacter baumannii. Disrupting cyclic di-GMP–modulating enzymes or cyclic di-GMP–binding effectors should alter the regulatory effectiveness of these phenotypes. I tested the multidrug-resistant isolate Acinetobacter baumannii strain AB5075 and identified several transposon …

Characterizing The Interaction Between Candida Albicans And Two Enterobacter Species, Abigail Cornett

Electronic Theses and Dissertations

Candida albicans is the most common human fungal pathogen. The relationship between C. albicans and Enterobacter bacteria have yet to be explored. The hypothesis of this study is that C. albicans and both E. aerogenes and E. cloacae have a positive relationship and work together to infect the host. In this study, the physical cell-to-cell interaction, molecular components of said interaction, and the impact of the interaction on a live organism were explored. Results indicate that Enterobacter adheres to C. albicans and inhibits growth with unidentified secreted molecules. Als1p has potential involvement in the attachment of E. cloacae to C. …

Identification Of Genetic Elements Involved In Alcaligenes Faecalis’ Inhibitory Mechanism Against Polymicrobial Species, Abigail Mathis

Undergraduate Honors Theses

The rise of antibiotic resistance in common human pathogens and the lack of development of novel therapeutic treatments has created a threat to global health. A unique source for potential novel treatments are from microorganisms, particularly within the complex, antagonistic polymicrobial interactions that take place in microbial communities. These unique mechanisms utilized by microorganisms to fight each other could potentially identify novel therapeutic targets for use at a clinical level, however, there is a lack of research in this area to determine its applicability. Alcaligenes faecalis is a Gram-negative bacterium that seldom causes human disease and has been observed in …

The Role Of Stm1987 And Arti In Arginine Response Of Salmonella Typhimurium, Deeba Mohseni

Undergraduate Honors Theses

Cyclic-di-GMP, a common bacterial second messenger, has been thought to help develop virulence and biofilms in bacteria, most specifically in Salmonella Typhimurium. By being able to dysregulate cyclic-di-GMP production, virulence may be better combatted. STM1987, an L-arginine-responsive diguanylate cyclase with a periplasmic sensory domain, dimerizes and generates the bacterial second messenger cyclic-di-GMP in response to the amino acid L-arginine in a pathway that also requires the periplasmic L-arginine-binding protein ArtI. Their biochemical responses to L-arginine and when they dimerize could help clarify this pathway, so I sought to develop a periplasmic dimerization sensor to better monitor these biochemical interactions. Similar …

Trained Immunity Enhances The Immune Response And Maintains Microbiome Diversity In Aging And Sepsis, P. Spencer Gill

Electronic Theses and Dissertations

The global population is rapidly aging. It is estimated that over the next thirty years, the number of individuals >60 years of age will increase by over a billion, and the number of individuals over age 80 may increase by 300 million. As humans age, our immune system becomes progressively weaker through a process called immune senescence. This age-related decrease in immune function increases susceptibility to infection and chronic diseases. Sepsis is a leading cause of death worldwide. Over the past two decades, there has been an increased incidence of sepsis which is due, in part, to our aging population …

Genital Chlamydia Infection Is Influenced By The Female Sex Hormones Estrogen And Progesterone In Vivo, Amy Gail Gravitte

Electronic Theses and Dissertations

Chlamydia is the most common bacterial sexually transmitted infection in the United States and worldwide. It often goes unnoticed due to lack of symptoms and left untreated it can ascend the female genital tract to cause sequelae like pelvic inflammatory disease and irreversible tubal infertility. In reproductive-aged women, female sex hormones estrogen (E2) and progesterone (P4) concentrations fluctuate during the menstrual cycle and are influenced by hormonal contraceptives and hormone replacement therapy. E2 and P4 influence genital Chlamydia infection in women and mice, but these multifactorial interactions are not entirely mapped out. The complex interplay of E2 and P4 with …

Neuroinflammation, Peripheral Inflammation And Gut Microbiome Profiles In Male Mice From Two Proposed Mouse Models Of Social Behavior Deficits, Sarah Parkinson

Electronic Theses and Dissertations

Autism spectrum disorder (ASD) is a neurodevelopmental disorder marked by social deficits and repetitive actions. A communication pathway exists between the brain and gut called the gut-brain axis. It is thought that gut bacteria can secrete signaling molecules, triggering inflammation across the body. These studies attempt to determine if markers are expressed in two mouse models of ASD behaviors, BTBR and a valproic acid model. Immunohistochemistry of ionized calcium binding adaptor molecule 1 from male mouse brain tissue showed no microglial activation in any group. Cytokine analysis did exhibit an increase in interleukin 1 (IL-1) in male adult mice only. …

Partial Expression Of The Vbss Gene In Rhizobium Leguminosarum Atcc 14479 And In-Silico Analysis Of The Vbs Gene Cluster In Various Microorganisms, Afreen Siddiqui

Undergraduate Honors Theses

Iron is extremely important for many organisms. Despite its abundancy, it exists in insoluble forms that makes its usability difficult. Some organisms secrete siderophores, low molecular weight compounds, that can chelate iron and convert it into usable forms for cells. One such organism, Rhizobium leguminosarum, is a nitrogen fixing symbiont proteobacteria that infects leguminous plants. The genome of Rhizobium leguminosarum ATCC 14479, which infects the red clover, Trifoli pratense, has previously been completely sequenced in our lab. Our lab has identified several genes in this strain involved in the biosynthesis of a siderophore, vicibactin. The protein product of one of …

Development, Expansion And Role Of Myeloid-Derived Suppressor Cells In Post-Sepsis Immune Suppression, Tuqa Alkhateeb

Electronic Theses and Dissertations

Myeloid-derived suppressor cells (MDSCs) numbers increase significantly in sepsis and are associated with high mortality rates. These myeloid cell precursors promote immunosuppression, especially in the late (post sepsis) stage. However, the mechanisms that underlie MDSC expansion and programming are not completely understood. To investigate these mechanisms, we used a cecal-ligation and puncture (CLP) mouse model of polymicrobial sepsis that progresses from an early/acute proinflammatory phase to a late/chronic immunosuppressive phase. Previous studies in our laboratory showed that microRNA (miR)-21 and miR-181b elevate levels of the transcription factor nuclear factor 1 (NFI-A) that promotes MDSC expansion. We report here that miR-21 …

A Novel Mode Of Action Of C-Reactive Protein In Protecting Against Streptococcus Pneumoniae Infection And Synergy With Antibiotics, Donald Ngwa

Electronic Theses and Dissertations

C-reactive protein (CRP) is a part of the innate immune system, is synthesized in the liver, its blood level increases in inflammatory states, and it binds to Streptococcus pneumoniae. The conformation of CRP is altered under conditions mimicking an inflammatory milieu and this non-native CRP also binds to immobilized/aggregated/pathogenic proteins. Experiments in mice have revealed that one of the functions of CRP is to protect against pneumococcal infection. For protection, CRP must be injected into mice within two hours of administering pneumococci, thus, CRP is protective against early-stage infection but not against late-stage infection. It is unknown how CRP protects …

Synergistic Inhibition Of Resistant Enterobacteriaceae Using A Possible Klebsiella Secreted Bacteriocin With Broad-Spectrum Antibiotic, Andrew Robbins

Undergraduate Honors Theses

Due to the increasing prevalence of multi-drug resistant (MDR) bacteria, it is now important to begin the search for novel means of defending against such resistant infections. Enterobacteriaceae is a clinically relevant family of bacteria that has shown extensive resistance to many antibiotics, especially after biofilm formation. Inhibitory poly-microbial interactions within this family have been observed. It is known that Citrobacter freundii (CF) growth is significantly inhibited by Klebsiella pneumoniae (KP) through a secreted protein. In this study, the potential KP bacteriocin was screened for its inhibitory effects on CF at various phases of biofilm development. The suspected KP bacteriocin …

Gut Pathophysiology In Mouse Models Of Social Behavior Deficits, Kyla Scott

Undergraduate Honors Theses

Autism spectrum disorders (ASD) encompass neurodevelopment disorders characterized by atypical patterns of development that impact multiple areas of functioning beginning in early childhood. The etiology of ASD is unknown and there are currently no preventative treatment options. Gastrointestinal symptoms are commonly associated comorbidities. The microbiota-gut-brain axis is a multidirectional communication chain that connects the central and enteric nervous system that relates brain function to peripheral intestinal functions. Changes within this axis have been postulated in ASD. For example, the “leaky gut theory” proposes that chronic inflammation is linked to alterations in the bacterial profiles of the gut microbiome and subsequent …

Characterization Of The Broad-Spectrum Inhibitory Capability Of Alcaligenes Faecalis And A. Viscolactis Against Potential Pathogenic Microorganisms, Andrew Fuqua

Undergraduate Honors Theses

The recent rise of multidrug resistant microorganisms has grown from an isolated concern to a massive public health crisis. It has become imperative that scientists look for new ways to combat this issue. Due to the selective pressures of competition, bacteria and other microbes possess a host of defenses and weapons designed to exploit vulnerabilities in other microorganisms. Consequently, the study of these systems and microbial interactions has much to reveal in the search for novel antimicrobial treatments. Previous research from our laboratory has discovered that both Alcaligenes faecalis and Alcaligenes viscolactis, two rarely studied and generally non-virulent bacteria, …

The Effects Of Farnesol, A Quorum Sensing Molecule From Candida Albicans, On Alcaligenes Faecalis, Savannah Hutson

Undergraduate Honors Theses

Quorum sensing molecules have become a recent focus of study to learn if and how they can be used, both on their own and in conjecture with current antimicrobial methods, as a means of bacterial control. One such quorum sensing molecule is the sesquiterpene alcohol, Farnesol, which is synthesized and released by the fungus, Candida albicans. In most in-vivo cases, our laboratory has shown that Alcaligenes faecalis overtakes C. albicans, preventing its growth. However, as a way to counteract this inhibitory effect, Farnesol may be one way that Candida has found to fight back. In this study, we …

A Study Of The Polymicrobial Inhibitory Interactions Between Alcaligenes Faecalis And Staphylococcus Aureus, Blakeley Griffin

Undergraduate Honors Theses

Members of the Staphylococcus genus are found as a part of normal microflora in humans and can commonly be found on the skin or in the nasal cavity. However, these microorganisms can cause serious and life-threatening opportunistic infections when there is a break in the physical barrier of skin. These infections have become difficult to treat as resistant strains emerge, particularly Methicillin Resistant Staphylococcus aureus (MRSA). MRSA has become a commonly acquired nosocomial infection which is difficult to treat with conventional antibiotics of the blactam class. Even Vancomycin, a last resort antibiotic, has been ineffective on some infections. Furthermore, …

Second Messenger Cyclic-Di-Gmp Regulation In Acinetobacter Baumannii, Justin Deal

Undergraduate Honors Theses

Over time, “superbugs,” or bacteria that have become resistant to antibiotics, have become a great concern in modern medicine. Viable alternates are currently being looked into as effective and safe ways to prevent or treat infections caused by these superbugs. One such method is through the utilization of the second messenger molecule cyclic-di-GMP (c-di-GMP) that has been shown to regulate phenotypes within other bacteria that may control surface colonization in Acinetobacter baumannii. Through a series of experiments, the active enzymes that create c-di-GMP - diguanylate cyclases - and break down c-di- GMP - phosphodiesterases - have been inactivated in …

Role Of Topoisomerase Ii Alpha In Dna Topology And T Cell Responses During Chronic Viral Infections, Stella Chinyere Ogbu

Electronic Theses and Dissertations

The clearance of viruses is largely dependent upon the activation of T cells to generate a robust immune response. However, host responses are suppressed during chronic viral infections. In this thesis, we explored the role of Top2α in DNA topology in individuals with chronic HBV, HCV, and HIV infections. We found that Top2α protein expression and activity were low in T cells derived from chronically virus-infected individuals compared to healthy subjects. Using CD4⁺ T cells treated with Top2α inhibitor or poisoner as a model, we demonstrated that Top2α inhibition disrupts the DNA topology, suppresses DNA repair kinase (ATM), and …

A Possible Luxr Solo Type Regulator Of An Antibiotic-Like Compound From The Soil Bacterium Rhodococcus, Katelyn Sellick

Electronic Theses and Dissertations

Rhodococcus, a species of bacteria commonly found in the soil, is an under-explored producer of small bioactive compounds including siderophores, pigments and antibiotics. MTM3W5.2 is a strain of Rhodococcus that was previously discovered to produce an antibiotic-like compound that has inhibitory effects on other Rhodococcus strains, including the veterinary pathogen, R. equi. The biosynthetic gene cluster responsible for production of the antibiotic has been identified, and a small gene, BTZ20_3964 at the start of the operon is believed to be a luxR solo regulator of the gene cluster. The goal of this project was to determine this gene’s …

Isolation, Analysis, And Partial Characterization Of An Inhibitor Of Neisseria Gonorrhoeae, Natania Paul

Undergraduate Honors Theses

There is an emerging threat of Neisseria gonorrhoeae strains that are resistant to all antibiotics. Because of this, the purpose of this research is to isolate, analyze, and partially characterize a new inhibitor(s) of N. gonorrhoeae. Since there is an unknown molecule secreted by Candida albicans that inhibits N. gonorrhoeae, this molecule can be partially characterized using ¹H NMR Spectroscopy to assist in the development of a new antibiotic compound. It was hypothesized that quorum-sensing molecules, trans, trans- farnesol, tyrosol, phenylethyl alcohol, and tryptophol, could be possible candidates for the inhibitor. Because of this, ¹H …

The Chelation Of Metal Ions By Vicibactin, A Siderophore Produced By Rhizobium Leguminosarum Atcc 14479, Joshua Stinnett

Undergraduate Honors Theses

Vicibactin is a small, high-affinity iron chelator produced by Rhizobium leguminosarum ATCC 14479. Previous work has shown that vicibactin is produced and secreted from the cell to sequester ferric iron from the environment during iron-deplete conditions. This ferric iron is then transported into the cell to be converted into ferrous iron. This study uses UV-Vis spectroscopy as well as ion trap-time of flight mass spectroscopy to determine that vicibactin does form a complex with copper(II) ions, however, at a much lower affinity than for iron(III). Stability tests have shown that the copper(II)-vicibactin complex is stable over time. The results of …

Virulence Regulation In Pseudomonas Aeruginosa Via The Alginate Regulators, Algu And Algr, The Posttranscriptional Regulator, Rsma, And The Two-Component System, Algz/R, Sean Stacey

Electronic Theses and Dissertations

Pseudomonas aeruginosa is a Gram-negative bacillus able to colonize a wide variety of environments. In the human host, P. aeruginosa can establish an acute infection or persist and create a chronic infection. P. aeruginosa is able to establish a niche and persist in human hosts by using a wide array of virulence factors used for: movement, killing host cells, and evading immune cells and antibiotics. Understanding virulence factors and their regulation has proved to be an important means of combating the morbidity and mortality of P. aeruginosa as well as the ever-increasing threat of drug resistance. By targeting virulence factors …

The Role Of Non-Neuronal Acetylcholine In Urogenital Chlamydial Infection, Jessica R. Lockhart

Electronic Theses and Dissertations

Chlamydia trachomatiscauses a bacterial sexually transmitted infection, Chlamydia, that is often chronic and casues reproductive complications in women. We hypothesized that Chlamydia infection increases local acetylcholine (ACh) production, which regulates the host’s inflammatory response to the infection. Female mice infected with C. muridarumwere sacrificed at days 3, 9, 15, and 21 post-infection, genital tract tissues harvested, and immunohistochemistry performed to enumerate ACh-producing cells. Infection increased the number of ACh-producing cells in cervical tissue at days 3,15, and 21 post-infection (pi), uterine tissue at day 3 and 9 pi, and ovarian tissue day 3, 15, and 21 pi. These …

Maxent Estimation Of Aquatic Escherichia Coli Stream Impairment, Dennis Gilfillan, Timothy A. Joyner, Phillip Scheuerman

ETSU Faculty Works

Background.The leading cause of surface water impairment in United States’ rivers and streams is pathogen contamination. Although use of fecal indicators has reduced human health risk, current approaches to identify and reduce exposure can be improved. One important knowledge gap within exposure assessment is characterization of complex fate and transport processes of fecal pollution. Novel modeling processes can inform watershed decision-making to improve exposure assessment.

Maxent Estimation Of Aquatic Escherichia Coli Stream Impairment, Dennis Gilfillan, Timothy Andrew Joyner, Phillip R. Scheuerman

ETSU Faculty Works

Background: The leading cause of surface water impairment in United States’ rivers and streams is pathogen contamination. Although use of fecal indicators has reduced human health risk, current approaches to identify and reduce exposure can be improved. One important knowledge gap within exposure assessment is characterization of complex fate and transport processes of fecal pollution. Novel modeling processes can inform watershed decision-making to improve exposure assessment.

Methods: We used the ecological model, Maxent, and the fecal indicator bacterium Escherichia coli to identify environmental factors associated with surface water impairment. Samples were collected August, November, February, and May for 8 …

Post-Transcriptional Regulation Of Rsma In Pseudomonas Aeruginosa, Ian Miller

Electronic Theses and Dissertations

Pseudomonas aeruginosa is a Gram-negative bacillus found in numerous environments. Gene regulatory mechanisms such as; Two-Component Systems, transcriptional and post-transcriptional regulators, and small non-coding RNAs control the expression of virulence factors that allow P. aeruginosa to initiate acute infections and persist as a chronic infection. A significant post-transcriptional regulator involved in these regulatory networks is the Regulator of Secondary Metabolites (RsmA). In this study, we investigated the contribution of a putative stem-loop on expression of RsmA. We constructed rsmA leader fusions to measure translation with and without the stem-loop present. Secondly, we introduced point mutations to disrupt the formation …

Identification Of “Fhua” Like Genes In Rhizobium Leguminosarum Atcc 14479 And Its Role In Vicibactin Transport And Investigation Of Heme Bound Iron Uptake System, Sushant Khanal

Electronic Theses and Dissertations

Siderophores are low molecular weight, iron chelating compounds produced by many bacteria for uptake of iron in case of iron scarcity. Vicibactin is a trihydroxamate type siderophore produced by Rhizobium leguminosarum bv. trifolii ATCC 14479. This work focuses on identifying an outer membrane receptor involved in the transport of vicibactin. We have confirmed the presence of the putative fhuA gene in R. leguminosarum bv. trifolii ATCC 14479. This bacteria shows mutualistic symbiosis with the red clover plant Trifoliium prantense. Leghemoglobin, with its cofactor heme is present in the plant root nodules that surrounds the infecting organism present in the nodules. …