EAT TO REPRODUCE: THE ROLE OF DIET QUANTITY ON HONEY BEE (APIS MELLIFERA) CASTE DETERMINATION A Thesis Submitted to the Graduate Faculty of the North Dakota State University of Agriculture and Applied Science By Garett Paul Slater In Partial Fulfillment of the Requirements for the Degree of MASTER OF SCIENCE Major Department: Biological Sciences March 2017 Fargo, North Dakota North Dakota State University Graduate School Title EAT TO REPRODUCE: THE ROLE OF DIET QUANTITY ON HONEY BEE (APIS MELLIFERA) CASTE DETERMINATION By Garett Paul Slater The Supervisory Committee certifies that this disquisition complies with North Dakota State University’s regulations and meets the accepted standards for the degree of MASTER OF SCIENCE SUPERVISORY COMMITTEE: Julia Bowsher Chair George Yocum Kendall Swanson Approved: 04/11/2017 Wendy Reed Date Department Chair ABSTRACT The received wisdom is that diet quality drives this queen-worker dichotomy, even though diet quantity differs significantly between queen and worker diet. Diet quantity likely determines honey bee caste; yet it has never been explicated tested. In this paper, we tested the hypothesis of diet quantity induced caste determination by; first, determine the ubiquity of quantity induced caste systems among highly-related taxa; second, determining the relative contributions of diet quantity vs. diet quality on adult caste in honey bees; and third, determine the influence of diet quantity on important cellular and physiological pathways known during caste determination. I found that: 1) diet quantity determines caste in honey bees and other eusocial hymenoptera, and 2) honey bees do not have a critical weight. This research likely will move forward the field of honey bee caste determination, which is a model for high-order sociality and phenotypic plasticity. iii ACKNOWLEDGEMENTS I would first like to thank my thesis advisor Dr. Julia H. Bowsher of the Department of Biology at North Dakota State University. The door to Prof. Bowsher office was always open whenever I ran into a trouble spot or had a question about my research or writing. She consistently allowed this paper to be my own work, but steered me in the right the direction whenever she thought I needed it. I would also like to thank the experts who were involved in this research project: Dr. George Yocum, Dr. Joe Rinehart, Dr. Arun Rajmahoen and Dr. Kendra Greenlee. Without their passionate participation and input, this project could not have been successfully conducted. I would also like to acknowledge my lab members, undergraduates, and other graduate students who were involved in this research project: Bryan Helm, Alex Torson, Dacotah Melicher, Marnie Larson, Raphael Royaute, Courtney Grulu, Kevin Cortez, Heather North, Micki Palmerson, Elizabeth Jensen, Lincoln Christenson, and Sean Nash. I am gratefully indebted to them for their valuable comments and input on this thesis. Finally, I must express my very profound gratitude to my parents and to my Stephanie St. Aubin for providing me with unfailing support and continuous encouragement throughout my years of study and through the process of researching and writing this thesis. This accomplishment would not have been possible without them. Thank you. iv DEDICATION To Stephanie for her unending support, level-headedness, and love. I wouldn’t have gotten through my thesis without her. To Wes for teaching me everything I know about honey bees. My thesis is impossible without him. v TABLE OF CONTENTS ABSTRACT ................................................................................................................................... iii ACKNOWLEDGEMENTS ........................................................................................................... iv DEDICATION .................................................................................................................................v LIST OF TABLES ...........................................................................................................................x LIST OF FIGURES ....................................................................................................................... xi LIST OF ABBREVIATIONS ...................................................................................................... xiii LIST OF APPENDIX TABLES .................................................................................................. xiv CHAPTER 1: AN OVERVIEW OF CASTE DETERMINATION ................................................1 1.1. Significance............................................................................................................................1 1.2. What Determines Caste in Other Eusocial Hymenoptera? ....................................................1 1.3. Diet Quality Vs. Quantity ......................................................................................................3 1.4. The Physiological Basis of Caste Determination in Honey Bee ............................................6 1.5. Cellular Mechanisms .............................................................................................................8 1.6. Conclusion .............................................................................................................................9 CHAPTER 2: DIET QUANTITY AND CASTE DETERMINATION IN EUSOCIAL HYMENOPTERA: META-ANALYSIS .......................................................................................11 2.1. Abstract ................................................................................................................................11 2.2. Introduction ..........................................................................................................................12 2.3. Materials and Methods .........................................................................................................14 2.3.1. Literature Search ............................................................................................................14 2.3.2. Data Extraction and Calculation of Effect Sizes ............................................................16 2.3.3. Data Grouping and Analysis ..........................................................................................17 2.3.4. Phylogeny ......................................................................................................................18 2.4. Results ..................................................................................................................................19 2.4.1. Diet Quantity ..................................................................................................................19 vi 2.4.2. Genetic ...........................................................................................................................21 2.4.3. Honey Bees ....................................................................................................................22 2.4.4. Phylogeny ......................................................................................................................24 2.5. Discussion ............................................................................................................................25 2.5.1. How Can Quantitative Caste Determination Arise? ......................................................26 2.5.2. How Can a Genetic Basis of Caste Determination Arise? .............................................28 2.5.3. Are Honey Bees Truly an Anomaly? .............................................................................29 2.5.4. Evolution of Multiple Cues ............................................................................................30 2.5.5. Framework for Eusociality ............................................................................................31 CHAPTER 3: DIET QUANTITY AND CASTE DETERMINATION IN HONEY BEES (APIS MELLIFERA) ......................................................................................................................32 3.1. Abstract ................................................................................................................................32 3.2. Introduction ..........................................................................................................................33 3.3. Materials and Methods .........................................................................................................35 3.3.1. Artificial Rearing ...........................................................................................................35 3.3.2. Diet Treatments ..............................................................................................................36 3.3.3. Determination of Protein, Carbohydrate, Lipid and Water Contents in Artificial Diets ...............................................................................................................................37 3.3.4. Diet Quantities ...............................................................................................................39 3.3.5. Morphometrics ...............................................................................................................39 3.3.6. Data Analysis and Presentation of Data.........................................................................40 3.3.6.1 Principal Component Analysis .................................................................................41 3.3.6.2. Clustering Analysis ..................................................................................................41