THE EFFECTS OF AN ARTIFICIAL HOLDING DIET ON THE FECUNDITY OF THE ECTOPARASITIC WASP TAMARIXIA RADIATA (WATERSTON) (HYMENOPTERA: EULOPHIDAE) A Thesis Presented to the Faculty of California State Polytechnic University, Pomona In Partial Fulfillment Of the Requirements for the Degree Master of Science In Plant Science By Danielle Renay Ruais 2018 SIGNATURE PAGE DISSERTATION: THE EFFECTS OF AN ARTIFICIAL HOLDING DIET ON THE FECUNDITY OF THE ECTOPARASISTIC WASPTAMARIXIA RADIATA (WATERSTON) (HYMENOPTERA: EULOPHIDAE) AUTHOR: Danielle Renay Ruais DATE SUBMITTED: Spring 2018 College of Agriculture Dr. Valerie J. Mellano _________________________________________ Department Chair Plant Sciences Dr. Anna L. Soper _________________________________________ Committee Chair Plant Sciences Dr. Robert L. Green _________________________________________ Adjunct Professor Plant Sciences Dr. David J.W.Morgan _________________________________________ CDFA Environmental Program Manager ii ACKNOWLEDGEMENTS Thank you to the ARI for providing funding for this research under grant #15-04- 219. Thank you to the project director Valerie J. Mellano, PhD, Chair of the Plant Sciences Department at California State Polytechnic University, Pomona. As well as the project collaborators Anna L. Soper, PhD, Assistant Professor of Plant Sciences at California State Polytechnic University, Pomona, Brian Taylor of the California Citrus Research Board, and David J. W. Morgan, PhD, Environmental Program Manager at the California Department of Food and Agriculture at Mt. Rubidoux, Riverside, California. Thank you to the California Citrus Research Board and California Citrus Mutual for your contributions. Thank you to California State Polytechnic University, Pomona and my committee for your guidance in developing the process; Robert L. Green, PhD, Lecturer of Plant Science at California State Polytechnic University, Pomona, David J. W. Morgan, PhD, Environmental Program Manager at the California Department of Food and Agriculture at Mt. Rubidoux, Riverside, California, and my Committee Chair Anna L. Soper, PhD, Assistant Professor of Plant Science at California State Polytechnic University, Pomona. Thank you to the California Department of Food and Agriculture at Mt. Rubidoux, Riverside, California for providing the Tamarixia for my experiments. I would like to especially thank David J. W. Morgan, PhD, Environmental Program Manager, and Environmental Scientists Grace Radabaugh, Alex Muniz, and Judith Herreid. Thank you Hoon Kim, PhD, Professor of Statistics and Co-Director of the Consulting Center for Statistics and Applied Mathematics for your help with providing iii the statistical analysis for my experiments. Thank you also to the Spring 2017 Advanced Statistics 559 course for your work with my data. Thank you David W. Still, PhD, Executive Director of the California State University Agricultural Research Institute for your guidance and direction during the early part of the process. I would like to thank Benjamin J. Lehan, MS, Lecturer at California State Polytechnic University, Pomona, for managing the Cal Poly Pomona Entomology Laboratory and making sure that I always had everything I needed to make my work go smoothly. I thank the following Plant Science undergraduate students for your help with the seemingly never-ending dissections and laboratory work. Thank you Christian Buelna. You are a reliable man and intelligent problem solver. Thank you Kevin Gonzalez for always making yourself available to me even when in the middle of your own important projects. Also thank you to University of California, Berkley undergraduate student Cindy E. Hsu for your help with the Tamarixia project. Thank you to Robert R. Shortell, PhD for your unwavering belief in my abilities and for giving me that shove when I was too hesitant to take the leap. Thank you David H. Headrick, PhD, Professor in the Horticulture and Crop Science Department at California State Polytechnic University, San Luis Obispo, for your wise counsel and for making me realize that bugs have feelings too. Thank you Paul Fletcher for introducing me to the love I feel for the trees, the bugs, and the soil. Thank you for cultivating my inquisitive spirit and connection to the earth. You have made a deep impact on how I view the world. I thank my parents Lynn and Raymond Ruais for their continuous love and support along this journey. To my sister Brie Ruais, you have been my steadfast iv supporter who kept my mind on the big picture. And to Niles Ruais for causing me to realize my own strengths. Thank you to Enrique Wallace IV who supported my ambitions and encouraged me to persevere. To Haley E. McCown, MS, thank you for your positive attitude and love along the way. Thank you to Haley Chappell and Elliott Popel for your encouragement throughout the process. Thank you Heidi Arndt for your brilliance. And last but certainly not the least, thank you Rebecca A. Morris, MSG, MPH, for reminding me to believe in myself and for helping me to make my master’s degree a reality. v ABSTRACT Since the detection of the invasive Asian Citrus Psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera: Psyllidae)—vector of the lethal citrus disease Huanglongbing (HLB) Candidatus Liberibacter asiaticus—in LA County in 2008, millions of Tamarixia radiata (Hymenoptera: Eulophidae) have been reared and released as an augmentative biological control agent to mitigate the ACP threat to California’s massive commercial citrus production. T. radiata is an effective ectoparasitoid because it establishes quickly under optimal release conditions and the adult female wasp has dual impact on ACP mortality: death by host-feeding on all nymphal instars and through parasitisation of later instars. As a synovigenic species, the female T. radiata wasp can emerge with a few eggs and after those eggs have been deposited onto ACP nymphs, she can produce new eggs. However, the formation of these new eggs is contingent on the type of nutrients, or diet, available to her as an adult. Increasing her egg load through diet during storage periods prior to release has the potential to monstrously affect the threat of ACP. Mass rearing these insects for release by the millions necessitates an efficient production system that includes manipulation of rearing conditions and storage for large populations of T. radiata. In this study, wasps received various diets containing various proteins, sugars, and amino acids for 24-hour increments prior to dissection to record EL. This information will aid in optimizing T. radiata rearing and storage protocol, which allows for wasp peak performance in field releases throughout California. vi TABLE OF CONTENTS Signature Page .................................................................................................................... ii Acknowledgments.............................................................................................................. iii Abstract .............................................................................................................................. vi List of Figures .................................................................................................................... ix Literature Review. A Fatal Disease, Its Vector, and the Heroine ........................................1 Background on Biological Control ..........................................................................1 The World Citrus Economy .....................................................................................2 Huanglongbing (HLB) .............................................................................................4 Asian Citrus Psyllid, Diaphorina citri (Kuwayama) (Hemiptera: Psyllidae) ........14 Tamarixia radiata (Waterston, 1922) (Hymenoptera: Eulophidae) ......................22 Mass-Rearing the Star of Biological Control .........................................................34 Experiment One. Regeneration of Egg Load .....................................................................37 Introduction. A Synovigenic Solitary Ectoparasitic Wasp ....................................37 Objective. Egg Regeneration through Diet ............................................................38 Materials and Methods. Age, Exposure Lengths, and Diet Treatments ................42 Results. It’s All About the Egg Load .....................................................................52 Discussion. Age, Body Size, and Diet Treatment ..................................................57 Conclusion. The Ultimate Factor ...........................................................................61 Experiment Two. CPP Citrus Grove Trials .......................................................................63 Introduction. Exposed to the Elements. .................................................................63 Objective. Observations of Wasp Activity ............................................................66 vii Materials and Methods. The Quest for ACP ..........................................................68 Results. To Host-Feed or to Parasitize ...................................................................74 Discussion. In-field Diet Treatment Performance .................................................75 Conclusion. Limitations and Future Directions .....................................................76 References ..........................................................................................................................80 Appendix A. Contacts