Medical Microbiology Commons^™

Pilz Domain Protein Flgz Mediates Cyclic Di-Gmp-Dependent Swarming Motility Control In Pseudomonas Aeruginosa, Amy Baker, Andreas Diepold, Sherry Kuchma, Jessie Scott, Dae Gon Ha, Giulia Orazi, Judith Armitage, George A. O'Toole

Dartmouth Scholarship

The second messenger cyclic diguanylate (c-di-GMP) is an important regulator of motility in many bacterial species. In Pseudomonas aeruginosa, elevated levels of c-di-GMP promote biofilm formation and repress flagellum-driven swarming motility. The rotation of P. aeruginosa's polar flagellum is controlled by two distinct stator complexes, MotAB, which cannot support swarming motility, and MotCD, which promotes swarming motility. Here we show that when c-di-GMP levels are elevated, swarming motility is repressed by the PilZ domain-containing protein FlgZ and by Pel polysaccharide production. We demonstrate that FlgZ interacts specifically with the motility-promoting stator protein MotC in a c-di-GMP-dependent manner and that a …

Moving Towards Effective Therapeutic Strategies For Neuronal Ceroid Lipofuscinosis., Ryan D. Geraets, Seung Yon Koh, Michelle L. Hastings, Tammy Kielian, David A. Pearce, Jill M. Weimer

Journal Articles: Pathology and Microbiology

The Neuronal Ceroid Lipofuscinoses (NCLs) are a family of autosomal recessive neurodegenerative disorders that annually affect 1:100,000 live births worldwide. This family of diseases results from mutations in one of 14 different genes that share common clinical and pathological etiologies. Clinically, the diseases are subcategorized into infantile, late-infantile, juvenile and adult forms based on their age of onset. Though the disease phenotypes may vary in their age and order of presentation, all typically include progressive visual deterioration and blindness, cognitive impairment, motor deficits and seizures. Pathological hallmarks of NCLs include the accumulation of storage material or ceroid in the lysosome, …

A Subset Of Latency-Reversing Agents Expose Hiv-Infected Resting Cd4+ T-Cells To Recognition By Cytotoxic T-Lymphocytes., R. Brad Jones, Stefanie Mueller, Rachel O'Connor, Katherine Rimpel, Derek D Sloan, Allison S. Thomas, Sara Karandish, Szu-Han Huang, +13 More

Microbiology, Immunology, and Tropical Medicine Faculty Publications

Resting CD4+ T-cells harboring inducible HIV proviruses are a critical reservoir in antiretroviral therapy (ART)-treated subjects. These cells express little to no viral protein, and thus neither die by viral cytopathic effects, nor are efficiently cleared by immune effectors. Elimination of this reservoir is theoretically possible by combining latency-reversing agents (LRAs) with immune effectors, such as CD8+ T-cells. However, the relative efficacy of different LRAs in sensitizing latently-infected cells for recognition by HIV-specific CD8+ T-cells has not been determined. To address this, we developed an assay that utilizes HIV-specific CD8+ T-cell clones as biosensors for HIV antigen expression. By testing …

Intimin Likely Used To Cause Disease During Competition With Commensal Escherichia Coli, Dominique J. Richburg

Senior Honors Theses

The intimin gene in the Locus of Enterocyte Effacement (LEE) island of pathogenicity is the primary attachment mechanism in Citrobacter rodentium. Intimin is a bacterial adhesin (protein) that attaches to obtain a niche/nutrient and thrive within the intestine. Intimin was deleted within C. rodentium to study colonization and pathogenesis in the murine intestine. Additionally, C. rodentium is an attaching/effacing pathogen, and a useful murine model in understanding Enterohemorrhagic Escherichia coli (EHEC) infection in humans. E. coli and C. rodentium cause gastroenteritis in humans and mice, respectively. C. rodentium is a murine pathogen commonly used to model gastrointestinal disease because …

The Homophilic Domain – An Immunological Archetype, Heinz Kohler, Jagadeesh Bayry, Srinivas V. Kaveri

Microbiology, Immunology, and Molecular Genetics Faculty Publications

The homophilic potential emerges as an important biological principle to boost the potency of immunoglobulins. Since homophilic antibodies in human and mouse sera exist prior environmental exposure, they are part of the natural antibody repertoire. Nevertheless, hemophilic properties are also identified in induced antibody repertoire. The use of homophilicity of antibodies in the adaptive immunity signifies an archetypic antibody structure. The unique feature of homophilicity in the antibody repertoire also highlights an important mechanism to boost the antibody potency to protect against infection and atherosclerosis as well to treat cancer patients.

The Inhibitory Site Of A Diguanylate Cyclase Is A Necessary Element For Interaction And Signaling With An Effector Protein, Kurt M. Dahlstrom, Krista M. Giglio, Holger Sondermann, George A. O'Toole

Dartmouth Scholarship

Many bacteria contain large cyclic diguanylate (c-di-GMP) signaling networks made of diguanylate cyclases (DGCs) and phosphodiesterases that can direct cellular activities sensitive to c-di-GMP levels. While DGCs synthesize c-di-GMP, many DGCs also contain an autoinhibitory site (I-site) that binds c-di-GMP to halt excess production of this small molecule, thus controlling the amount of c-di-GMP available to bind to target proteins in the cell. Many DGCs studied to date have also been found to signal for a specific c-di-GMP-related process, and although recent studies have suggested that physical interaction between DGCs and target proteins may provide this signaling fidelity, the importance …

Effects Of A Mindfulness-Based Weight Loss Intervention In Adults With Obesity: A Randomized Clinical Trial., Jennifer Daubenmier, Patricia J Moran, Jean Kristeller, Michael Acree, Peter Bacchetti, Margaret E Kemeny, Douglas F. Nixon, +11 Additional Authors

Microbiology, Immunology, and Tropical Medicine Faculty Publications

OBJECTIVE: To determine whether adding mindfulness-based eating and stress management practices to a diet-exercise program improves weight loss and metabolic syndrome components.

METHODS: In this study 194 adults with obesity were randomized to a 5.5-month program with or without mindfulness training and identical diet-exercise guidelines. Intention-to-treat analyses with multiple imputation were used for missing data. The primary outcome was 18-month weight change.

RESULTS: Estimated effects comparing the mindfulness to control arm favored the mindfulness arm in (a) weight loss at 12 months, -1.9 kg (95% CI: -4.5, 0.8; P = 0.17), and 18 months, -1.7 kg (95% CI: -4.7, 1.2; …

Lipidomic Dataset Of Plasma From Patients Infected With Wild Type And Nef-Deficient Hiv-1 Strain., Peter Meikle, Hann Low, Melissa J Churchill, Michael Bukrinsky, Dmitri Sviridov

Microbiology, Immunology, and Tropical Medicine Faculty Publications

Previous in vitro and in vivo studies demonstrated that HIV protein nef plays a key role in impairing cellular and systemic cholesterol metabolism in HIV disease, but clinical support for these findings is lacking. Here we present the data of comparative lipidomic analysis (330 lipid species) of plasma samples from HIV-negative subjects, patients infected with WT HIV-1 strain and patients infected with nef-deficient strain of HIV-1. We determine which effects of HIV on plasma lipidome are explained by the presence of nef. The data can be used to evaluate cardiovascular risk in HIV disease and to assess the role of …

Herpes Simplex Virus And Interferon Signaling Induce Novel Autophagic Clusters In Sensory Neurons, Sarah Katzenell, David A. Leib

Dartmouth Scholarship

Herpes simplex virus 1 (HSV-1) establishes lifelong infection in the neurons of trigeminal ganglia (TG), cycling between productive infection and latency. Neuronal antiviral responses are driven by type I interferon (IFN) and are crucial to controlling HSV-1 virulence. Autophagy also plays a role in this neuronal antiviral response, but the mechanism remains obscure. In this study, HSV-1 infection of murine TG neurons triggered unusual clusters of autophagosomes, predominantly in neurons lacking detectable HSV-1 antigen. Treatment of neurons with IFN-β induced a similar response, and cluster formation by infection or IFN treatment was dependent upon an intact IFN-signaling pathway. The autophagic …

The Significance Of A Common Idiotype (1f7) On Antibodies Against Human Immune Deficiency Virus Type 1 And Hepatitis C Virus, Sybille Muller, Matthew S. Parsons, Heinz Kohler, Michael Grant

Microbiology, Immunology, and Molecular Genetics Faculty Publications

In this review, we trace the concept and potential functional role of regulatory idiotypes in the immune response to human immunodeficiency virus type 1 (HIV-1), simian immunodeficiency virus, and hepatitis C virus (HCV). A major idiotype involved in these viral infections is recognized and defined by a murine monoclonal antibody (1F7). Antibodies expressing the idiotype defined by 1F7 are dominant in HIV-1 infection and are also found on many broadly neutralizing antibodies against HIV-1. This regulatory idiotypic axis offers opportunities for exploitation in vaccine development for HIV-1, HCV, and other chronic viral infections.

Structural And Evolutionary Analyses Show Unique Stabilization Strategies In The Type Iv Pili Of Clostridium Difficile, Kurt H. Piepenbrink, Grace A. Maldarelli, Claudia F. Martinez De La Pena, Tanis C. Dingle, George L. Mulvey, Amanda Lee, Erik Von Rosenvinge, Glen D. Armstrong, Michael S. Donnenberg, Eric J. Sundberg

Food for Health: Publications

Type IV pili are produced by many pathogenic Gram-negative bacteria and are important for processes as diverse as twitching motility, biofilm formation, cellular adhesion and horizontal gene transfer. However, many Gram-positive species, including C. difficile, also produce Type IV pili. Here, we identify the major subunit of the Type IV pili of C. difficile, PilA1, and describe multiple three-dimensional structures of PilA1, demonstrating the diversity found in three strains of C. difficile. We also model the incorporation of both PilA1 and a minor pilin, PilJ, into the pilus fiber. Although PilA1 contains no cysteine residues, and therefore cannot form the …

The Gras Sensor In Staphylococcus Aureus Mediates Resistance To Host Defense Peptides Differing In Mechanisms Of Action, Siyang Chaili, Ambrose L. L. Cheung, Arnold S. Bayer, Yan Q. Xiong, Alan Waring, Guido Memmi, Niles Donegan

Dartmouth Scholarship

Staphylococcus aureus uses the two-component regulatory system GraRS to sense and respond to host defense peptides (HDPs). However, the mechanistic impact of GraS or its extracellular sensing loop (EL) on HDP resistance is essentially unexplored. Strains with null mutations in the GraS holoprotein (ΔgraS) or its EL (ΔEL) were compared for mechanisms of resistance to HDPs of relevant immune sources: neutrophil α-defensin (human neutrophil peptide 1 [hNP-1]), cutaneous β-defensin (human β-defensin 2 [hBD-2]), or the platelet kinocidin congener RP-1. Actions studied by flow cytometry included energetics (ENR); membrane permeabilization (PRM); annexin V binding (ANX), and cell death protease activation (CDP). …

Cd147 And Downstream Adamtss Promote The Tumorigenicity Of Kaposi's Sarcoma-Associated Herpesvirus Infected Endothelial Cells, Lu Dai, Jimena Trillo-Tinoco, Yihan Chen, Karlie Bonstaff, Luis Del Valle, Chris Parsons, Augusto C. Ochoa, Jovanny Zabaleta, Bryan P. Toole, Zhiqiang Qin

School of Medicine Faculty Publications

Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiologic agent of several human cancers, including Kaposi's sarcoma (KS), which preferentially arise in immunocompromised patients and lack effective therapeutic options. We have previously shown that KSHV or viral protein LANA up-regulates the glycoprotein CD147, thereby inducing primary endothelial cell invasiveness. In the current study, we identify the global network controlled by CD147 in KSHV-infected endothelial cells using Illumina microarray analysis. Among downstream genes, two specific metalloproteases, ADAMTS1 and 9, are strongly expressed in AIDS-KS tissues and contribute to KSHV-infected endothelial cell invasiveness through up-regulation of IL-6 and VEGF. By using a KS-like nude …

The 40-Residue Insertion In Vibrio Cholerae Fadr Facilitates Binding Of An Additional Fatty Acyl-Coa Ligand, Wei Shi, Gabriela Kovacikova, Wei Lin, Ronald. K. Taylor, Karen Skorupski, F. Jon Kull

Dartmouth Scholarship

FadR is a master regulator of fatty acid metabolism and influences virulence in certain members of Vibrionaceae. Among FadR homologues of the GntR family, the Vibrionaceae protein is unusual in that it contains a C-terminal 40-residue insertion. Here we report the structure of Vibrio cholerae FadR (VcFadR) alone, bound to DNA, and in the presence of a ligand, oleoyl-CoA. Whereas Escherichia coli FadR (EcFadR) contains only one acyl-CoA-binding site in each monomer, crystallographic and calorimetric data indicate that VcFadR has two. One of the binding sites resembles that of EcFadR, whereas the other, comprised residues from the insertion, has not …

Gene Deletion By Fluorescence-Reported Allelic Exchange Mutagenesis In Chlamydia Trachomatis, Konrad E. Mueller, Katerina Wolf, Kenneth A. Fields

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Although progress in Chlamydia genetics has been rapid, genomic modification has previously been limited to point mutations and group II intron insertions which truncate protein products. The bacterium has thus far been intractable to gene deletion or more-complex genomic integrations such as allelic exchange. Herein, we present a novel suicide vector dependent on inducible expression of a chlamydial gene that renders Chlamydia trachomatis fully genetically tractable and permits rapid reverse genetics by fluorescence-reported allelic exchange mutagenesis (FRAEM). We describe the first available system of targeting chlamydial genes for deletion or allelic exchange as well as curing plasmids from C. trachomatis …

Characterization Of Rna Helicase Csha And Its Role In Protecting Mrnas And Small Rnas Of Staphylococcus Aureus Strain Newman, Samin Kim, Anna-Rita Corvaglia, Stefano Léo, Ambrose Cheung, Patrice Francois

Dartmouth Scholarship

The toxin MazF_sa in Staphylococcus aureus is a sequence-specific endoribonuclease that cleaves the majority of the mRNAs in vivo but spares many essential mRNAs (e.g., secY mRNA) and, surprisingly, an mRNA encoding a regulatory protein (i.e., sarA mRNA). We hypothesize that some mRNAs may be protected by RNA-binding protein(s) from degradation by MazF_sa. Using heparin-Sepharose-enriched fractions that hybridized to sarA mRNA on Northwestern blots, we identified among multiple proteins the DEAD box RNA helicase CshA (NWMN_1985 or SA1885) by mass spectroscopy. Purified CshA exhibits typical RNA helicase activities, as exemplified by RNA-dependent ATPase activity and unwinding of …

Rnai-Mediated Gene Knockdown By Microinjection In The Model Entomopathogenic Nematode Heterorhabditis Bacteriophora., Ramesh Ratnappan, Jonathan Vadnal, Melissa Keaney, Ioannis Eleftherianos, Damien O'Halloran, John M. Hawdon

Microbiology, Immunology, and Tropical Medicine Faculty Publications

BACKGROUND: Parasitic nematodes threaten the health of humans and livestock and cause a major financial and socioeconomic burden to modern society. Given the widespread distribution of diseases caused by parasitic nematodes there is an urgent need to develop tools that will elucidate the genetic complexity of host-parasite interactions. Heterorhabditis bacteriophora is a parasitic nematode that allows simultaneous monitoring of nematode infection processes and host immune function, and offers potential as a tractable model for parasitic nematode infections. However, molecular tools to investigate these processes are required prior to its widespread acceptance as a robust model organism. In this paper we …

Identification And Spontaneous Immune Targeting Of An Endogenous Retrovirus K Envelope Protein In The Indian Rhesus Macaque Model Of Human Disease., Helen L Wu, Enrique J Léon, Lyle T Wallace, Francesca A Nimiyongskul, Matthew B Buechler, Laura P Newman, Philip A Castrovinci, R Paul Johnson, Robert J Gifford, R. Brad Jones, Jonah B Sacha

Microbiology, Immunology, and Tropical Medicine Faculty Publications

BACKGROUND: Endogenous retroviruses (ERVs) are remnants of ancient retroviral infections that have invaded the germ line of both humans and non-human primates. Most ERVs are functionally crippled by deletions, mutations, and hypermethylation, leading to the view that they are inert genomic fossils. However, some ERVs can produce mRNA transcripts, functional viral proteins, and even non-infectious virus particles during certain developmental and pathological processes. While there have been reports of ERV-specific immunity associated with ERV activity in humans, adaptive immune responses to ERV-encoded gene products remain poorly defined and have not been investigated in the physiologically relevant non-human primate model of …

Cold Atmospheric Plasma Inhibits Hiv-1 Replication In Macrophages By Targeting Both The Virus And The Cells., Olga Volotskova, Larisa Dubrovsky, Michael Keidar, Michael Bukrinsky

Microbiology, Immunology, and Tropical Medicine Faculty Publications

Cold atmospheric plasma (CAP) is a specific type of partially ionized gas that is less than 104°F at the point of application. It was recently shown that CAP can be used for decontamination and sterilization, as well as anti-cancer treatment. Here, we investigated the effects of CAP on HIV-1 replication in monocyte-derived macrophages (MDM). We demonstrate that pre-treatment of MDM with CAP reduced levels of CD4 and CCR5, inhibiting virus-cell fusion, viral reverse transcription and integration. In addition, CAP pre-treatment affected cellular factors required for post-entry events, as replication of VSV-G-pseudotyped HIV-1, which by-passes HIV receptor-mediated fusion at the plasma …

Expression Levels Of Virulence Genes In Group A Streptococci: A Response To Aerosolized Propylene Glycol, Michael S. Costello

Honors Undergraduate Theses

Electronic cigarette usage is becoming increasingly prevalent among school age children and young adults. A known bactericidal agent, propylene glycol, is often used as a carrier for nicotine, flavoring, and additional constituents of electronic cigarette juice. This study examined the relationship between propylene glycol and virulence gene expression in Streptococcus pyogenes, a respiratory tract pathogen commonly found in school-age individuals. A variety of virulence genes controlled by the three stand alone regulators mga, RofA, and Rgg/RopB were sampled in an effort to understand the pathway by which virulence is affected. The genes chosen encode C5a peptidase, fibronectin binding protein, hyaluronate …

Coordination Of Rna Polymerase Ii Pausing And 3' End Processing Factor Recruitment With Alternative Polyadenylation, Becky Fusby, Soojin Kim, Benjamin Erickson, Hyunmin Kim, Martha L. Peterson, David L Bentley

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Most mammalian genes produce transcripts whose 3' ends are processed at multiple alternative positions by cleavage/polyadenylation (CPA). Poly(A) site cleavage frequently occurs cotranscriptionally and is facilitated by CPA factor binding to the RNA polymerase II (Pol II) C-terminal domain (CTD) phosphorylated on Ser2 residues of its heptad repeats (YS₂PTSPS). The function of cotranscriptional events in the selection of alternative poly(A) sites is poorly understood. We investigated Pol II pausing, CTD Ser2 phosphorylation, and processing factor CstF recruitment at wild-type and mutant IgM transgenes that use alternative poly(A) sites to produce mRNAs encoding the secreted and membrane-bound forms of …

Antibiotic Efficacy And Interaction In Escherichia Coli During Varying Nutrient Conditions, Kristina K. Millar

Scripps Senior Theses

Due to the recent rise in antibiotic resistant pathogens, and the difficulties surrounding the quest for new antibiotics, many researchers have started revisiting antibiotic interactions in hopes of finding new treatment options. The primary outcome of this project was to examine the efficacy of concomitant antibiotic use under varying nutrient conditions, to identify variations in antibiotic interactions. Antibiotic interactions were studied, utilizing E. coli as a model bacterial system, grown in four different media types. E. coli cultures were treated with streptomycin, tobramycin, erythromycin, and amikacin individually and in a pairwise fashion at varying doses. We found that at least …

Modulation Of Body Weight By Intestinal Flora In Orphan Nuclear Receptor Shp-/- Mice, Ryan Mifflin

Williams Honors College, Honors Research Projects

The whole-body deletion of small heterodimer partner (SHP) in mice is associated with protection from diet-induced obesity and hepatic steatosis upon feeding of a western diet. This protection was reported to be mediated through decreases in hepatic gene expression for lipogenesis, as well as increases in gene expression for fatty acid oxidation. SHP has been known to regulate the expression of the CYP7A1 gene, encoding the rate-limiting enzyme for bile acid synthesis, thereby altering the bile acid pool. The effects of this altered bile acid profile on the gut microbiome are unknown, as some bacteria in the gut are responsible …

Emerging Pathogens In Cystic Fibrosis Patients At Virginia Commonwealth University Medical Center (Vcumc), Emily M. Hill

Theses and Dissertations

Cystic fibrosis (CF) is an autosomal recessive disorder affecting 70,000 individuals worldwide. This disease is characterized by the buildup of mucus in the airways leading to chronic lung infections resulting in pulmonary failure and death in 95% of CF patients. Routine surveillance of CF pathogens using traditional microbiology culture guides management and treatment of CF patients. Molecular profiling studies have revealed emerging pathogens that may play a role in CF lung disease by either directly causing infection or upregulating the virulence factors of classic CF pathogens, such as P. aeruginosa; however, routine CF culture protocols have not been modified …

Drug And Vaccine Development For Neisseria Gonorrhoeaea, Devin R. Cash

Theses and Dissertations

Neisseria gonorrhoeae, the causative agent of the STI gonorrhea, is not preventable by vaccination and is rapidly developing resistance to antibiotics. One important strategy for gonococcal survival in the host is iron acquisition in the face of nutritional immunity. To overcome iron limitation, the gonococcus expresses TonB dependent transporters (TdTs), outer membrane proteins that facilitate nutrient acquisition. Of the TdTs, the transferrin (Tf), lactoferrin (Lf), and hemoglobin (Hb) receptors hijack iron directly from host proteins, and studies have already shown that the Tf receptor is essential for the initiation of human infection. Given that the TdTs are virulence factors, …

Inhibition Of Bacterial Growth And Prevention Of Bacterial Adhesion With Localized Nitric Oxide Delivery, Julia Osborne

Dissertations, Master's Theses and Master's Reports

Bacterial infections continue to be a problem at the site of an indwelling medical device, and over the years, various bacterial strains have become more resistant to current antibiotic treatments. Bacterial infection at an indwelling medical device can be dangerous and affect the performance of the medical device which can ultimately lead to the failure of the device due to bacterial resistance to treatment.

Nitric Oxide (NO) has been shown to possess antibacterial properties to prevent and inhibit bacterial growth. NO releasing coatings on indwelling medical devices could provide a reduction in bacterial infections that occur at the device site …