The spread and evolution of RNA viruses among honey bees and the wider insect community with particular emphasis on Deformed wing virus (DWV). Laura Emily BRETTELL Ph.D. Thesis 2017 School of Environment and Life Sciences University of Salford CONTENTS CONTENTS ............................................................................................................................................... 1 LIST OF FIGURES ...................................................................................................................................... 5 General Introduction........................................................................................................................... 5 Chapter 1: A comparison of Deformed wing virus in deformed and asymptomatic honey bees. ..... 5 Chapter 2: Oldest Varroa tolerant honey bee population provides insight into the origins of the global decline of honey bees. ............................................................................................................. 7 Chapter 4: Moku virus; a new Iflavirus found in wasps, honey bees and Varroa. ............................. 8 Chapter 5: Common apiary pests as a potential source of honey bee – associated viruses in a Hawaiian apiary environment. ............................................................................................................ 9 LIST OF TABLES ...................................................................................................................................... 10 Chapter 1: A comparison of Deformed wing virus in deformed and asymptomatic honey bees. ... 10 Chapter 2: Oldest Varroa tolerant honey bee population provides insight into the origins of the global decline of honey bees. ........................................................................................................... 10 Chapter 5: Common apiary pests as a potential source of honey bee – associated viruses in a Hawaiian apiary environment. .......................................................................................................... 10 ACKNOWLEDGEMENTS ......................................................................................................................... 12 DECLARATION ....................................................................................................................................... 13 Chapter 1: A comparison of Deformed wing virus in deformed and asymptomatic honey bees. .. 13 Chapter 2: Oldest Varroa tolerant honey bee population provides insight into the origins of the global decline of honey bees. ........................................................................................................... 14 Chapter 3: Novel RNA virus genome discovered in Ghost ants (Tapinoma melanocephalum) from Hawaii. .............................................................................................................................................. 14 Chapter 4: Moku virus; a new Iflavirus found in wasps, honey bees and Varroa. ........................... 15 Chapter 5: Common apiary pests as potential reservoirs of honey bee – associated viruses in a Hawaiian apiary environment. .......................................................................................................... 15 Appendices ........................................................................................................................................ 16 Appendix 1: Evidence of Varroa-mediated Deformed Wing virus spillover in Hawaii. ................ 16 Appendix 2: Superinfection exclusion and the long-term survival of honey bees in Varroa- infested colonies. .......................................................................................................................... 16 Appendix 3: Covert deformed wing virus infections have long-term deleterious effects on honeybee foraging and survival. ................................................................................................... 17 ABBREVIATIONS .................................................................................................................................... 18 GENERAL ABSTRACT .............................................................................................................................. 19 1 GENERAL INTRODUCTION ..................................................................................................................... 21 The honey bee .................................................................................................................................. 21 Honey bee colony losses ................................................................................................................... 21 Varroa destructor .............................................................................................................................. 24 Honey bee viruses ............................................................................................................................. 26 Deformed Wing Virus ........................................................................................................................ 28 Symptoms ......................................................................................................................................... 29 Transmission ..................................................................................................................................... 30 Varroa destructor as a vector ........................................................................................................... 31 History ............................................................................................................................................... 34 Structure ........................................................................................................................................... 35 Honey bee immune response to DWV transmission by Varroa ....................................................... 37 Resistance ......................................................................................................................................... 39 Honey bee viruses and cross species transmission .......................................................................... 40 Studying insect viruses ...................................................................................................................... 41 Aims .................................................................................................................................................. 43 References ........................................................................................................................................ 45 Chapter 1: A comparison of Deformed wing virus in deformed and asymptomatic honey bees. ....... 59 Abstract ............................................................................................................................................. 59 Introduction ...................................................................................................................................... 59 Materials and Methods ..................................................................................................................... 62 Honeybee Samples ........................................................................................................................ 62 Effect of Viral Load on Wing Deformity ........................................................................................ 62 Next Generation Sequencing, Assembly and Data Normalisation ............................................... 64 Results ............................................................................................................................................... 65 Viral Quantification and Wing Deformity in Honey Bees ............................................................. 65 Next Generation Illumina Sequencing .......................................................................................... 68 Discussion.......................................................................................................................................... 72 Conclusions ................................................................................................................................... 74 References ........................................................................................................................................ 75 Supplementary Information ............................................................................................................. 80 Chapter 2: Oldest Varroa tolerant honey bee population provides insight into the origins of the global decline of honey bees. ............................................................................................................... 84 Abstract ............................................................................................................................................. 84 2 Introduction ...................................................................................................................................... 84 Methods ............................................................................................................................................ 87 Mite & honey bee samples ........................................................................................................... 87 Detection of DWV ........................................................................................................................