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University of Florida Thesis Or Dissertation ASSESSING VIRULENCE OF Beauveria bassiana AGAINST HOUSE FLY (Musca domestica) IMMATURES AND ADULTS By ROXIE LOURENE WHITE A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE UNIVERSITY OF FLORIDA 2019 © 2019 Roxie Lourene White To my mother for her never-ending love and support ACKNOWLEDGMENTS Above all, I want to express great appreciation to my main advisor, Dr. Christopher Geden for his support, knowledge, and guidance throughout this process. His encouraging words matched with his eternal patience has made this learning adventure a manageable and pleasurable experience. His expert knowledge helped formulate the methods of my research and he answered my relentless questions in the laboratory. I am greatly appreciative of Dr. Geden for providing the funding necessary for me to further my education as well as employing me as a full-time technician at the USDA. He has taught me many things about life both within and outside of the laboratory and for that, I am thankful. I would also like to show great appreciation to my co-chair, Dr. Phillip Kaufman, for consistently guiding me, providing feedback and for pushing me to take things a step further than the minimally required. Through his weekly writing meetings, my writing has improved tremendously, while being able to edit scientific literature for other students. I am very thankful and appreciative of the lessons learned through Dr. Kaufman’s writing club. Although I may have tried to hide from Dr. Kaufman at one point or another, he has always had an open door and open lab to make me feel very welcome and part of his lab. An extended thank you is due to Dr. Emma Weeks as she has served almost as an unofficial committee member. She has graciously reviewed all my writing and has provided tips, tricks and insights on things I may have overlooked. She has also provided a soft place when things got tough and remained encouraging while not pushing too hard during the sensitive times. 4 Tremendous thanks are extended to the members of the Geden lab. In particular, Dana Johnson has been a very critical aspect of my success. She has taught me most of what I know about fungal culturing, sterile lab procedures and microbiology techniques. Dana has been a huge help in preparing fungal strains, isolating strains for pure cultures and assistance in bioassays. She has been a listening ear when things got tough, always encouraged me to “just keep swimming”, and helped voice some of the hesitations I was too afraid to speak about. Thanks are extended to Lindsey Granko, Katie Carrol and Rachel Dillard for maintaining house fly colonies, keeping the laboratory in tip-top shape and their assistance in bioassays. Great thanks are due to my friends, family, and colleagues for helping me in various ways throughout the last few years. I would like to thank my mother Rhonda White for always encouraging me to become the best while pursuing my dreams. She has remained encouraging and supportive for the duration of my educational journey including the last two years of graduate school. Without her support, graduate school would not have been possible. Lastly, I would like to thank my four-legged children for picking me up daily with their endless wet kisses and wagging tails. Coming home to a happy family of animals was a very enriching part of my day during the stressful parts of graduate school. 5 TABLE OF CONTENTS page ACKNOWLEDGMENTS .................................................................................................. 4 LIST OF TABLES ............................................................................................................ 8 LIST OF FIGURES .......................................................................................................... 9 ABSTRACT ................................................................................................................... 11 CHAPTER 1 LITERATURE REVIEW OF FILTH FLIES AND ENTOMOPATHOGENIC FUNGI FOR HOUSE FLY MANAGEMENT ........................................................................ 13 Introduction ............................................................................................................. 13 House Fly Biology and Life History ......................................................................... 14 Medical, Veterinary and Economic Importance ....................................................... 16 Need for IPM to Manage House Flies ..................................................................... 16 House Fly Parasitoids ............................................................................................. 19 Entomopathogenic Fungi Overview ........................................................................ 22 Beauveria bassiana: Biology, History, and Development........................................ 25 Virulence of Beauveria bassiana and Metarhizium anisopliae against Adult House Flies .......................................................................................................... 27 Research Objectives ............................................................................................... 29 2 EXPOSURE OF HOUSE FLY LARVAE TO Beauveria bassiana ........................... 30 Introduction ............................................................................................................. 30 Materials and Methods............................................................................................ 32 House Fly Rearing ............................................................................................ 32 Beauveria bassiana used in Assays ................................................................. 33 Larval Test 1: Assays with Dry L90 Conidia and BotaniGard® ......................... 35 Larval Test 2: Effect of Host age and Dose on Response to Treatment with B. bassiana. .................................................................................................. 35 Larval Test 3: Effect of Temperature and Host Age on Response to Treatment with B. bassiana. .......................................................................... 37 Larval Test 4: Effect of Medium Composition and Host Age on Response to Treatment with B. bassiana. .......................................................................... 38 Statistical Analysis .................................................................................................. 39 Results .................................................................................................................... 40 Larval Test 1: Assays with Dry L90 Conidia and BotaniGard® ......................... 40 Larval Test 2: Effect of Host age and Dose on Response to Treatment with B. bassiana. .................................................................................................. 41 6 Larval Test 3: Effect of Temperature and Host Age on Response to Treatment with B. bassiana. .......................................................................... 42 Larval Test 4: Effect of Medium Composition and Host Age on Response to Treatment with B. bassiana. .......................................................................... 42 Discussion .............................................................................................................. 43 3 SELECTION FOR A FASTER-KILLING Beauveria bassiana STRAIN ................... 51 Introduction ............................................................................................................. 51 Materials and Methods............................................................................................ 55 Sources of Flies and B. bassiana ..................................................................... 55 Selection for Early-dying Flies .......................................................................... 56 Comparing Virulence of Selected and Unselected B. bassiana strain NFH10.. 58 Statistical Analysis .................................................................................................. 61 Results .................................................................................................................... 61 Discussion .............................................................................................................. 62 4 COMPARISON OF VIRULENCE OF Metarhizium anisopliae AND FOUR STRAINS OF Beauveria bassiana AGAINST ADULT HOUSE FLIES .................... 75 Introduction ............................................................................................................. 75 Materials and Methods............................................................................................ 77 Sources, Isolation and Propagation of Fungal Strains ...................................... 78 Harvesting of Conidia and Preparation of Inocula for Testing .......................... 81 Bioassay Method .............................................................................................. 82 Statistical Analysis .................................................................................................. 84 Results .................................................................................................................... 84 Discussion .............................................................................................................. 85 5 DISCUSSION OF FINDINGS AND FUTURE RESEARCH DIRECTIONS FOR USING Beauveria bassiana AGAINST HOUSE FLY (Musca domestica)
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