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Ecology of Culicoides Biting Midges and Their Role As Vectors of Epizootic Hemorrhagic Disease Virus on Florida Big Game Preserves

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Ecology of Culicoides Biting Midges and Their Role As Vectors of Epizootic Hemorrhagic Disease Virus on Florida Big Game Preserves ECOLOGY OF CULICOIDES BITING MIDGES AND THEIR ROLE AS VECTORS OF EPIZOOTIC HEMORRHAGIC DISEASE VIRUS ON FLORIDA BIG GAME PRESERVES By BETHANY L MCGREGOR A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2019 © 2019 Bethany L McGregor To my family, friends, and mentors who have always supported and encouraged me ACKNOWLEDGEMENTS I thank my family for their encouragement, support, and patience throughout my graduate education. I thank my friends for offering humor and companionship during the most stressful of times. I thank my lab mates for serving as a source of inspiration and a sounding board for ideas. I thank the FMEL lab technicians for their unending help towards completion of this research. Finally, I thank my committee and mentors for their guidance and advice throughout the PhD process. 4 TABLE OF CONTENTS page ACKNOWLEDGEMENTS ............................................................................................... 4 LIST OF TABLES ............................................................................................................ 8 LIST OF FIGURES ........................................................................................................ 10 LIST OF ABBREVIATIONS ........................................................................................... 11 ABSTRACT ................................................................................................................... 12 CHAPTER 1 LITERATURE REVIEW .......................................................................................... 14 2 HOST USE PATTERNS OF CULICOIDES SPP. BITING MIDGES AT A BIG GAME PRESERVE IN FLORIDA, USA .............................................................................. 25 Methods .................................................................................................................. 29 Field Methods .................................................................................................... 29 Laboratory Methods .......................................................................................... 31 Data Analysis .................................................................................................... 33 Results .................................................................................................................... 34 Forage Ratios .................................................................................................... 34 Habitat Associated Host Use ............................................................................. 36 White-tailed Deer Aspiration .............................................................................. 37 Discussion .............................................................................................................. 38 3 VERTICAL STRATIFICATION OF CULICOIDES BITING MIDGES AT A FLORIDA BIG GAME PRESERVE ......................................................................................... 52 Methods .................................................................................................................. 54 Field Collections ................................................................................................ 54 Laboratory Methods .......................................................................................... 56 Data Analysis .................................................................................................... 58 Results .................................................................................................................... 58 Total Collections ................................................................................................ 58 Ground versus Canopy Use by Abundant Species ........................................... 59 Funnel Suction Traps ........................................................................................ 60 Habitat Associations of Abundant Species ........................................................ 60 Physiological Status .......................................................................................... 61 Blood Meal Analysis .......................................................................................... 61 Discussion .............................................................................................................. 63 5 4 IMPLICATING CULICOIDES STELLIFER AND CULICOIDES VENUSTUS (DIPTERA: CERATOPOGONIDAE) AS VECTORS OF EPIZOOTIC HEMORRHAGIC DISEASE VIRUS ........................................................................ 79 Methods .................................................................................................................. 82 Culicoides Sampling and Virus Detection During EHDV Epizootic in Northern Florida ........................................................................................................... 82 White-tailed Deer Sampling and Virus Detection during the 2017 Epizootic ..... 84 Statistical Analysis of Virus Data ....................................................................... 85 Live Animal Aspiration ....................................................................................... 86 Blood Meal Analysis .......................................................................................... 86 Seasonal abundance of C. stellifer and C. venustus ......................................... 87 Results .................................................................................................................... 88 Virus Detection in Field Collected Midges during an EHDV Epizootic ............... 88 Animal Mortality due to EHDV ........................................................................... 89 Live Animal Aspiration ....................................................................................... 91 Blood Meal Analysis .......................................................................................... 91 Seasonal Abundance of C. stellifer and C. venustus ........................................ 92 Discussion .............................................................................................................. 92 5 VECTOR COMPETENCE OF CULICOIDES SONORENSIS FOR EPIZOOTIC HEMORRHAGIC DISEASE VIRUS SEROTYPE 2 STRAINS FROM CANADA AND FLORIDA .............................................................................................................. 106 Methods ................................................................................................................ 108 Culicoides sonorensis colony .......................................................................... 108 Viral Strains ..................................................................................................... 109 Bloodfeeding ................................................................................................... 110 Sample Collection ........................................................................................... 110 Honey Cards ................................................................................................... 111 Sample processing .......................................................................................... 112 Statistical analysis ........................................................................................... 113 Results .................................................................................................................. 114 Infection, Dissemination, and Transmission Rates .......................................... 114 Viral Titer Analysis .......................................................................................... 115 Honey Card Versus Capillary Assay Comparison ........................................... 116 Discussion ............................................................................................................ 117 6 VECTOR COMPETENCE OF CULICOIDES INSIGNIS (DIPTERA: CERATOPOGONIDAE) FOR EPIZOOTIC HEMORRHAGIC DISEASE VIRUS SEROTYPE 2 ....................................................................................................... 127 Methods ................................................................................................................ 130 Blood Feeding Trial Specimen Collection........................................................ 130 Blood Feeding Apparatus Design .................................................................... 130 6 Blood Feeding Trial Variables ......................................................................... 131 Infection Trial Culicoides insignis Collections .................................................. 131 Viral Blood Feeding ......................................................................................... 132 Sample Processing ......................................................................................... 132 RNA Extraction and Virus Detection ............................................................... 133 Statistical Analyses ......................................................................................... 134 Intrathoracic Inoculation .................................................................................. 135 Virus Screening of Field-Collected C. insignis ................................................. 135 Results .................................................................................................................. 136 Blood
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