Pathogenic Microbiology Commons^™

Understanding The Role Of Toxs As A Bile Sensing Component Of The Toxrs Virulence Regulatory System In Vibrio Spp., Minje Kim

Dartmouth College Ph.D Dissertations

Pathogenic Vibrio bacteria, such as V. cholerae and V. parahaemolyticus, colonize in the human small intestine to cause severe gastrointestinal disease. During the infection, these bacteria encounter bile salts, antimicrobial cholesterol metabolites secreted into the intestine. Pathogenic Vibrio species have evolved to utilize bile salts as signals to regulate virulence. The signaling depends in part on ToxRS, a conserved co-component transmembrane transcription regulator. ToxRS consists of the transcription factor ToxR and its membrane-tethered binding partner ToxS. ToxS is required for ToxR stability and full transcriptional activity. Although bile salts are known to influence ToxRS-dependent virulence gene expression, the molecular …

Evolution Of Antibiotic Responses During Chronic P. Aeruginosa Infections In The Human Lung, David Frank Ritz

Dartmouth College Ph.D Dissertations

Typical antibiotic susceptibility testing (AST) of microbial samples is performed in homogeneous cultures in batch environments, which does not account for the highly heterogeneous and dynamic nature of antibiotic responses. The most common mutation found in P. aeruginosa lineages evolved during chronic cystic fibrosis infections in the human lung, a loss of function of repressor MexZ, increases basal levels of multidrug efflux MexXY, but does not increase resistance by traditional minimal inhibitory concentration (MIC) assays. Here, we use single cell microfluidics to show that P. aeruginosa’s response to aminoglycosides is highly heterogeneous, with only a subpopulation of cells surviving exposure. …

The Impact Of Polymycovirus 1 On Aspergillus Fumigatus Pathogenesis During Lung Infection, Alexander W. Rapp

Dartmouth College Ph.D Dissertations

The genus Aspergillus comprises over 250 species of fungi. Within this genus, only a handful of species cause disease in humans. One species is Aspergillus fumigatus (A.f.), an environmentally ubiquitous, saprophytic fungal pathogen. Lung infections due to A.f. range from invasive infection to allergic forms of the disease. Allergic bronchopulmonary aspergillosis (ABPA) is one of the most severe forms and presents as a hypersensitivity that often results in off-target damage to the surrounding airways. Patients with ABPA are sensitized to fungal antigens and this is commonly seen in cystic fibrosis and genetic predispositions. Recent data from our lab shown how …

Biochemical And Structural Insights Into The Yersinia Effector Yopm And Its Negative Regulation Of The Pyrin Inflammasome, Bethany Wairimu Mwaura

Dartmouth College Ph.D Dissertations

Bacteria utilize sophisticated secretion systems to inject effectors into host cells in order to facilitate their survival and replication in the host. Some pathogenic species of Gram-negative bacteria use a conserved contact-dependent T3SS to inject a wide array of effectors. While the effectors diverge in their structures, functions and cell-localization, the T3SS is well conserved. Several effectors discussed in this thesis target key host inflammasome responses. The focus of this work is how the Yersinia effector YopM inhibits the pyrin inflammasome. During Yersinia infection, two effectors, YopE and YopT inadvertently activate the pyrin inflammasome while targeting RhoA to avoid phagocytosis. …

Neutrophil Inflammatory Death Regulation By Bacterial Type Iii Secreted Effectors, Arianna D. Reuven

Dartmouth College Ph.D Dissertations

Yersinia and Pseudomonas use a T3SS to secrete effectors into host cells that inhibit phagocytosis and modulate immune responses, such as inflammasome activation. Inflammasomes promote the activation of the proinflammatory protease caspase-1. The activation of an inflammasome sensor in response to a signal or ligand initiates the assembly of the inflammasome leading to activation of caspase-1 which cleaves GSDMD and pro-inflammatory cytokines IL-1β and IL-18 into their mature forms. As neutrophils are recruited in large quantities to sites of infection, they are excellent candidates to drive inflammasome dependent responses. Neutrophils have been shown to produce multiple inflammasomes, like pyrin and …

Biochemical And Molecular Analysis Of Anr-Dependent Microoxic Fitness Factors In Pseudomonas Aeruginosa, Stacie E. Balsam

Dartmouth College Ph.D Dissertations

Pseudomonas aeruginosa is a prominent opportunistic pathogen that is often found in microoxic environments like sites of infection and colony biofilms. P. aeruginosa can grow at oxygen (O2) at concentrations as little as 3 μM and also grows anaerobically by generating energy using nitrate as an alternate electron acceptor. An important way P. aeruginosa adapts to low O2 is through Anr, a transcription factor that upregulates many genes important for microoxic and anoxic fitness.

Specifically, Anr regulates mhr which encodes a hemerythrin protein (Mhr) that binds O2 with affinities relevant to microoxia and is also necessary for full fitness of …

Targeting Aspergillus Fumigatus Hypoxia Response Pathways For Novel Antifungal Drug Development., Cecilia Gutierrez Perez

Dartmouth College Ph.D Dissertations

Invasive fungal disease present are difficult to diagnose and treat and present a high mortality rate across the world with approximately 1.5 million people a year succumbing to fungal infections worldwide. Currently, there are limited antifungal classes used therapeutically, and antifungal drug resistance on the rise further exacerbates the dire need for novel therapeutics with innovative mode of actions. The filamentous fungi, Aspergillus fumigatus is the causative agent of invasive pulmonary aspergillosis (IPA). It is estimated that contemporary antifungals fail in about 50% of patients with invasive filamentous fungal infections, likely due to the biofilm mode of growth exhibited in …

Mechanistic Insights On Positive Regulation Of The Pyrin Inflammasome In Macrophages, Haleema Sadia Malik

Dartmouth College Ph.D Dissertations

The mammalian innate immune defense employs sophisticated mechanisms, such as effector triggered immunity (ETI), to discern between pathogenic and non-pathogenic bacteria and initiate a protective response. ETI is involved in immune detection of virulent activities of bacterial effector proteins in the host cytosol. Inflammasome sensors, such as pyrin, detect cellular infection or stress as part of ETI, leading to the assembly of multiprotein complexes, caspase-1 activation and proinflammatory cytokine release. Pyrin uniquely initiates inflammatory responses against RhoA-inactivating bacterial toxins and effectors such as Yersinia's YopE and YopT, and C. difficile’s toxins TcdA/B. Understanding pyrin regulation is crucial due to its …

Dpr-Mediated H2o2 Resistance Contributes To Streptococci Survival In A Cystic Fibrosis Airway Model System, Rendi R. Rogers

Dartmouth College Master’s Theses

The cystic fibrosis (CF) lung environment is conducive to the colonization of bacteria as polymicrobial biofilms, which are associated with poor clinical outcomes for persons with CF (pwCF). Streptococcus spp. is highly prevalent in the CF airway, but its role in the CF lung microbiome is poorly understood. Some studies have shown Streptococcus spp. to be associated with better clinical outcomes for pwCF, while others show that high abundance of Streptococcus spp. is correlated with exacerbations. Our lab previously reported a polymicrobial culture system consisting of four CF-relevant pathogens that can be used to study microbial behavior in a more …

Causes And Consequences Of Lasr Mutant Selection In Pseudomonas Aeruginosa Populations, Dallas L. Mould

Dartmouth College Ph.D Dissertations

Change is the only constant in life, and these changes, though random in nature, can have consequences. Quorum sensing is heterogeneous in phenotype and prone to negative selection. In P. aeruginosa, the regulator LasR is frequently non-functional in phylogenetically diverse isolates. Through repeated experimental evolution and mathematical modeling, we show that differences in growth enable lasR mutant evolutionary success and this requires a system enabling metabolic choices, known as carbon catabolite repression (or catabolite repression). The differences in catabolite repression between wild type and lasR mutants enable altered metabolite preferences, and the resulting differences in metabolic states enable intraspecies …

The Immune Modulation On Innate Immunity, From Pathogen Recognition To Fungal Clearance., Ko-Wei Liu

Dartmouth College Ph.D Dissertations

The human lung is not sterile but a complex environment with various microorganisms. Besides commensals in the lung, hundreds to thousands of individual microbiomes enter the lung every day but without causing the symptom. Host innate immunity plays an important role in maintaining homeostasis of the lung environment and as the first defense line against pathogens. Aspergillus fumigatus (A. fumigatus) is a saprophytic filamentous fungus that can cause human disease in immune compromised patients. However, with functional innate immunity, immune cells can quickly recognize pathogen associated molecular patterns (PAMPs) from A. fumigatus through pattern recognition receptors (PRRs). The activation of …

The Role Of L-Alanine Signaling In Aspergillus Fumigatus Biofilm Adherence, Carbon Catabolism, And Echinocandin Susceptibility., Joshua D. Kerkaert

Dartmouth College Ph.D Dissertations

Aspergillus fumigatus is a saprophytic filamentous fungus that participates in environmental carbon and nitrogen cycles through the degradation of complex organic substrates. In addition to its ecological role, A. fumigatus is the primary causative agent of a spectrum of diseases depending on the immune status of the individual, the most lethal of which is invasive aspergillosis (IA). Treatment strategies for IA are limited and far too frequently fail. Despite the high rates of treatment failure, antifungal resistance remains relatively low, albeit rising at a concerning rate. Insights into this discrepancy between the rate of treatment failure and the rate of …

Links Between Electrophilic Stress And Antifungal Resistance In Pathogenic Candida Species, Amy R. Biermann

Dartmouth College Ph.D Dissertations

Collectively, Candida species are the most prevalent cause of both superficial and invasive fungal infections worldwide. Invasive Candida infections have a high mortality rate and predominantly affect individuals with underlying diseases, such as diabetes, HIV, or cancer. Unfortunately, many invasive Candida infections are recalcitrant to antifungal treatment, while intrinsically multidrug-resistant pathogens, like Candida auris, are increasing in prevalence. Although the canonical mechanisms of antifungal resistance in Candida species are well established, i.e., overexpression of efflux pumps and overexpression of or mutations in genes encoding drug targets, factors affecting the natural evolution and regulation of resistance mechanisms remain poorly understood. …

Deletion Mutant Library For Investigation Of Functional Outputs Of Cyclic Diguanylate Metabolism In Pseudomonas Aeruginosa Pa14, Dae-Gon Ha, Megan E. Richman, George A. O'Toole

Dartmouth Scholarship

We constructed a library of in-frame deletion mutants targeting each gene in Pseudomonas aeruginosa PA14 predicted to participate in cyclic di-GMP (c-di-GMP) metabolism (biosynthesis or degradation) to provide a toolkit to assist investigators studying c-di-GMP-mediated regulation by this microbe. We present phenotypic assessments of each mutant, including biofilm formation, exopolysaccharide (EPS) production, swimming motility, swarming motility, and twitch motility, as a means to initially characterize these mutants and to demonstrate the potential utility of this library.

Ecology And Genetic Structure Of A Northern Temperate Vibrio Cholerae Population Related To Toxigenic Isolates, Brian M. Schuster, Anna L. Tyzik, Rachel A. Donner, Megan J. Striplin, Salvador Almagro-Moreno

Dartmouth Scholarship

Although Vibrio cholerae is an important human pathogen, little is known about its populations in regions where the organism is endemic but where cholera disease is rare. A total of 31 independent isolates confirmed as V. cholerae were collected from water, sediment, and oysters in 2008 and 2009 from the Great Bay Estuary (GBE) in New Hampshire, a location where the organism has never been detected. Environmental analyses suggested that abundance correlates most strongly with rainfall events, as determined from data averaged over several days prior to collection. Phenotyping, genotyping, and multilocus sequence analysis (MLSA) revealed a highly diverse endemic …

Type Ii Toxoplasma Gondii Ku80 Knockout Strains Enable Functional Analysis Of Genes Required For Cyst Development And Latent Infection, Barbara A. Fox, Alejandra Falla, Leah M. Rommereim, Tadakimi Tomita

Dartmouth Scholarship

Type II Toxoplasma gondii KU80 knockouts (Δku80) deficient in nonhomologous end joining were developed to delete the dominant pathway mediating random integration of targeting episomes. Gene targeting frequency in the type II Δku80 Δhxgprt strain measured at the orotate (OPRT) and the uracil (UPRT) phosphoribosyltransferase loci was highly efficient. To assess the potential of the type II Δku80 Δhxgprt strain to examine gene function affecting cyst biology and latent stages of infection, we targeted the deletion of four parasite antigen genes (GRA4, GRA6, ROP7, and tgd057 …

Genome-Wide Transcriptional Profiling Of The Cyclic Amp-Dependent Signaling Pathway During Morphogenic Transitions Of Candida Albicans, Yong-Sun Bahn, Matthew Molenda, Janet F. Staab, Courtney A. Lyman, Laura J. Gordon, Paula Sundstrom

Dartmouth Scholarship

Candida albicans is an opportunistic human fungal pathogen that causes systemic candidiasis as well as superficial mucosal candidiasis. In response to the host environment, C. albicans transitions between yeast and hyphal forms. In particular, hyphal growth is important in facilitating adhesion and invasion of host tissues, concomitant with the expression of various hypha-specific virulence factors. In previous work, we showed that the cyclic AMP (cAMP) signaling pathway plays a crucial role in morphogenic transitions and virulence of C. albicans by studying genes encoding adenylate cyclase-associated protein (CAP1) and high-affinity phosphodiesterase (PDE2) (Y. S. Bahn, J. Staab, and P. Sundstrom, Mol. …

Bacteriophage Migration Via Nematode Vectors: Host-Parasite-Consumer Interactions In Laboratory Microcosms, John J. Dennehy, Nicholas A. Friedenberg, Yul W. Yang, Paul E. Turner

Dartmouth Scholarship

Pathogens vectored by nematodes pose serious agricultural, economic, and health threats; however, little is known of the ecological and evolutionary aspects of pathogen transmission by nematodes. Here we describe a novel model system with two trophic levels, bacteriophages and nematodes, each of which competes for bacteria. We demonstrate for the first time that nematodes are capable of transmitting phages between spatially distinct patches of bacteria. This model system has considerable advantages, including the ease of maintenance and manipulation at the laboratory bench, the ability to observe many generations in short periods, and the capacity to freeze evolved strains for later …