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Intravital Imaging in a Zebrafish Model Elucidates Interactions Between Mucosal Immunity and Pathogenic Fungi Linda S

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Intravital Imaging in a Zebrafish Model Elucidates Interactions Between Mucosal Immunity and Pathogenic Fungi Linda S The University of Maine DigitalCommons@UMaine Electronic Theses and Dissertations Fogler Library Summer 8-23-2019 Intravital Imaging in a Zebrafish Model Elucidates Interactions Between Mucosal Immunity and Pathogenic Fungi Linda S. Archambault University of Maine, [email protected] Follow this and additional works at: https://digitalcommons.library.umaine.edu/etd Part of the Animal Experimentation and Research Commons, Immunity Commons, Immunology of Infectious Disease Commons, Immunopathology Commons, and the Pathogenic Microbiology Commons Recommended Citation Archambault, Linda S., "Intravital Imaging in a Zebrafish Model Elucidates Interactions Between Mucosal Immunity and Pathogenic Fungi" (2019). Electronic Theses and Dissertations. 3066. https://digitalcommons.library.umaine.edu/etd/3066 This Open-Access Thesis is brought to you for free and open access by DigitalCommons@UMaine. It has been accepted for inclusion in Electronic Theses and Dissertations by an authorized administrator of DigitalCommons@UMaine. For more information, please contact [email protected]. INTRAVITAL IMAGING IN A ZEBRAFISH MODEL ELUCIDATES INTERACTIONS BETWEEN MUCOSAL IMMUNITY AND PATHOGENIC FUNGI By Linda S. Archambault B.S. Bates College, 1982 M.A. Boston University, 1986 A DISSERTATION Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy (in Biochemistry) The Graduate School The University of Maine August 2019 Advisory Committee: Robert T. Wheeler, Associate Professor of Microbiology, Advisor Clarissa Henry, Associate Professor of Biological Sciences Julie Gosse, Associate Professor of Biochemistry Paul Millard, Associate Professor of Chemical and Biomedical Engineering Reeta Rao, Associate Professor of Biology and Biotechnology, Worcester Polytechnic Institute, Worcester, Massachusetts. Copyright 2019 Linda S. Archambault ii INTRAVITAL IMAGING IN A ZEBRAFISH MODEL ELUCIDATES INTERACTIONS BETWEEN MUCOSAL IMMUNITY AND PATHOGENIC FUNGI By Linda S. Archambault Dissertation Advisor: Dr. Robert T. Wheeler An Abstract of the Dissertation Presented in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy (in Biochemistry) August 2019 Candida yeasts are common commensals that can cause mucosal disease and life- threatening systemic infections. While many of the components required for defense against Candida albicans infection are Well established, questions remain about hoW various host cells at mucosal sites assess threats and coordinate defenses to prevent normally commensal organisms from becoming pathogenic. Using tWo Candida species, C. albicans and C. parapsilosis, which differ in their abilities to damage epithelial tissues, we used traditional methods (pathogen CFU, host survival, and host cytokine expression) combined With high- resolution intravital imaging of transparent zebrafish larvae to illuminate host-pathogen interactions at the cellular level in the complex environment of a mucosal infection. In zebrafish, C. albicans grows as both yeast and epithelium-damaging filaments, activates the NF-kB pathway, evokes proinflammatory cytokines, and causes the recruitment of phagocytic immune cells. On the other hand, C. parapsilosis remains in yeast morphology and elicits the recruitment of phagocytes Without inducing inflammation. High-resolution mapping of phagocyte-Candida interactions at the infection site revealed that neutrophils and macrophages attack both Candida species, regardless of the cytokine environment. Time-lapse monitoring of single-cell gene expression in transgenic reporter zebrafish revealed a partitioning of the immune response during C. albicans infection: the transcription factor NF-kB is activated largely in cells of the swimbladder epithelium, While the proinflammatory cytokine tumor necrosis factor alpha (TNF-a) is expressed in motile cells, mainly macrophages. Our results point to different host strategies for combatting pathogenic Candida species and separate signaling roles for host cell types. DEDICATION I dedicate this dissertation to my husband, Thomas Archambault, who has been my steadfast supporter through this endeavor and every endeavor I have undertaken since We met. There is no one who has more unselfishly wished for my happiness and success and no one more suited to travel the path With me as I have pursued them. iii ACKNOWLEDGEMENTS My collaborators on this project include University of Maine graduate, Dominika TrZilova, and researchers from University of Minnesota, Dr. Cheryl Gale and Sara Gonia. I would like to thank the Tobin, Huttenlocher, Bagnat, RaWls and Lieschke laboratories for sharing fish lines and am grateful for the exceptional fish husbandry provided by Mark Nilan at the UMaine Zebrafish Facility. I thank members of the Wheeler Lab and Drs. Henry and Rao for their contributions along the Way and comments on the paper manuscript. I especially thank Dr. Remi Gratacap for his excellent training in confocal microscopy and the use of the zebrafish model. Thank-you to Dr. Muse Davis for giving me the opportunity to co-author a review article. Linda Archambault is a Janet Waldron Fellow at University of Maine. Her advisor, Dr. Robert Wheeler, is a Burroughs Wellcome Fund Investigator in the Pathogenesis of Infectious Disease. This Work Was funded by NIH grants R15AI094406 and R15AI133415 and USDA Hatch Grant ME0-H-1-00517-48413. iv TABLE OF CONTENTS DEDICATION ............................................................................................................................... iii ACKNOWLEDGEMENTS ............................................................................................................ iv LIST OF TABLES ......................................................................................................................... ix LIST OF FIGURES ........................................................................................................................ x CHAPTERS 1. INTRODUCTION AND BACKGROUND .................................................................................... 1 1.1. Host-pathogen Interactions .......................................................................................... 1 1.1.1. Human microbiota ............................................................................................ 1 1.1.2. Commensalism vs. disease .............................................................................. 2 1.1.3. Detecting and responding to disease ............................................................... 2 1.2. Host Defenses .............................................................................................................. 2 1.2.1. Barriers ............................................................................................................ 3 1.2.2. Innate immunity ............................................................................................... 3 1.2.3. Adaptive immunity ........................................................................................... 3 1.3. Fungal Pathogens ........................................................................................................ 4 1.3.1. Environmental fungi ......................................................................................... 4 1.3.2. Commensal fungi ............................................................................................. 5 1.3.3. Candida disease states ................................................................................... 6 1.3.3.1. Chronic mucocutaneous candidiasis .................................................... 6 1.3.3.2. Oropharyngeal candidiasis ................................................................... 6 1.3.3.3. Vulvovaginal candidiasis ....................................................................... 7 1.3.3.4. Dermal candidiasis ............................................................................... 7 v 1.3.4. Candida albicans ............................................................................................. 7 1.3.5. Non-albicans Candida species ........................................................................ 8 1.3.6 Candida parapsilosis ........................................................................................ 9 1.4. Virulence Factors ........................................................................................................ 10 1.4.1. Immune evasion ............................................................................................ 10 1.4.2. Dissemination ................................................................................................ 11 1.4.3. Morphogenesis .............................................................................................. 12 1.4.4. Adhesion ........................................................................................................ 13 1.4.5. Invasion ......................................................................................................... 13 1.4.6. Damage ......................................................................................................... 14 1.4.7. Biofilm formation ............................................................................................ 15 1.5. Host Mucosal Immune Defenses ................................................................................ 16 1.5.1. Mucosal adaptive immunity ........................................................................... 16 1.5.2. Mucosal
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