Efficacy of Entomopathogenic Nematodes and Entomopathogenic Fungi against Masked Chafer White Grubs, Cyclocephala spp. (Coleoptera: Scarabaeidae) Shaohui Wu Dissertation submitted to the faculty of the Virginia Polytechnic Institute and State University In partial fulfillment of the requirements for the degree of Doctor of Philosophy in Entomology Dr. Roger R. Youngman, Co-Chair Dr. Loke T. Kok, Co-Chair Dr. James M. Goatley Dr. Douglas G. Pfeiffer Dr. Sally L. Paulson April 23, 2013 Blacksburg, Virginia Keywords: Entomopathogenic Nematode, Entomopathogenic Fungus, Masked Chafer, Cyclocephala spp., White Grub Copyright 2013, Shaohui Wu Efficacy of Entomopathogenic Nematodes and Entomopathogenic Fungi against Masked Chafer White Grubs, Cyclocephala spp. (Coleoptera: Scarabaeidae) Shaohui Wu ABSTRACT Entomopathogenic nematodes (EPN) (Heterorhabditis bacteriophora and H. megidis) and entomopathogenic fungi (EPF) (Metarhizium anisopliae and Beauveria bassiana) were evaluated for efficacy against masked chafer white grub, Cyclocephala spp., under laboratory and greenhouse conditions, as well as their efficacy against various grub stages in the field. Under both laboratory and greenhouse conditions, additive interactions were found between EPN and EPF in their combined application against Cyclocephala spp., except a few observations that showed antagonism or synergism. Significantly greater control occurred from the combination of a nematode and a fungus compared with a fungus alone, but not compared with a nematode alone. The combined effect did not differ significantly for nematode and fungi applied simultaneously or at different times. EPF had no significant impact on EPN infection and production of infective juveniles (IJs) in grub carcasses. Nematodes alone or in combination with fungi were comparable to the insecticide Merit 75 WP (imidacloprid) against 3rd instar Cyclocephala spp in the greenhouse. Efficacy of EPF and EPN varied dramatically between field sites and conditions; EPN and EPF applied alone or in combination were less effective than Merit 75 WP in >50% field trials, but some EPN + EPF treatments were more effective than the insecticide in reducing grub numbers. EPN and EPF showed better potential than insecticides for providing extended control of white grubs in the subsequent generation. In addition, the sub-lethal effects of EPF on southern masked chafer, C. lurida, were investigated. Neither M. anisopliae nor B. bassiana had a sub-lethal effect on grub weight gain, adult longevity, oviposition, pupation and eclosion. Finally, interaction between H. bacteriophora and M. anisopliae was examined to determine the potential of the nematode in improving fungal distribution in soil. H. bacteriophora enhanced fungal distribution in sandy loam soil without grass thatch, but not in sandy soil with thatch. In both soil types, soil depths significantly affected nematode and fungal distribution. In water profile, M. anisopliae conidia germinated hyphae that attached to sheath of H. bacteriophora IJs, which molted to detach from the fungus. IJs mortality and virulence were not affected by the presence of M. anisopliae. DEDICATION This is dedicated to my beloved husband Feng Li & daughter Sophie Li, for their unwavering love and support, to make my accomplishment possible. Thank you. iii ACKNOWLEDGEMENTS I would first and foremost like to thank my families who supported me to accomplish my work. Thanks to my dear husband Feng Li for his companionship through the hard times, and sharing all my tears and joys. I would also like to thank my mom Feng-E Wu for without her support and assistance with taking care of my new-born daughter I could not have been able to finish my dissertation. To my extended family, my dad Yan-Liang Rao, grandpa Ting-Kai Wu, brothers Shao-Hu Wu and Shao-Feng Wu, thank you for your spiritual support and warm care of my life. I am also very grateful to my grandma Chun-Ying Lei, who passed away in summer 2012 when I was away from home. Special thanks to my advisors and committee members: my advisor Dr. Roger R. Youngman who directed me with all endeavor even in his sickness, and whose humor made my graduate studies more enjoyable; my co-advisor Dr. Loke T. Kok who provided invaluable advice and suggestions, helped me with the English language and guided me through the difficulties when I first came to this country and during my studies; Dr. James M. Goatley who helped with turfgrass problems I encountered in research; Dr. Sally L. Paulson who impressed me with her kindness and wisdom; Dr. Douglas G. Pfeiffer who enriched me the knowledge of insects and IPM. Many thanks to: Curt A. Laub, who helped with all my field work and equipment supplies in my research, and taught me slang English and American customs that made me laugh and enjoy the long trips to field sites; Dr. Jarrod E. Leland from Novozymes Biologicals, Inc. (Salem, VA) for providing technical support and supplies of Metarhizium anisopliae; Dr. Jonathan D. Eisenback of the Department of Plant Pathology, Physiology & Weed Science, for his technical assistance with nematodes; my lab-mate and friend William Kuhn who helped with my field work and also helped me to better adapt to American life; my lab-mate Sudan Gyawaly for his kindness and assistance with my field work; Becker Underwood Co. (Ames, IA) for providing nematodes in my research; VT turf team who has been very helpful in assisting my field studies; golf course superintendants in Tazewell Country Club, Virginian Country Club, Blacksburg Country Club, and VT Golf Course for providing sites for field research and grub collection. I am very grateful to the Department of Entomology faculty and staff who provided assistance in my research/studies. These include but are not limited to: Dr. Donald E. Mullins, Sandra Gabbert, Dr. Carlyle C. Brewster, Dr. Scott Salom, Dr. Thomas P. Kuhar, Scotty Bolling, Ryan Mays, Dr. Richard D. Fell. Special thanks to Kathy Shelor, Sarah Kenley, Denise Dodd, Mary Parks and Robin Williams, for their personal friendship and kindness. Last but certainly not least, I would like to thank my friends and fellow graduate students who made my times here more enjoyable and easier, listed in no particular order: Lisa Moore, Brenna Traver, Reina iv Koganamura, Heather Andrew, Heather Story, Matt Reis, Andrea Polanco, Melissa Fisher, Serena Ciparis, Azadeh Aryan, Tim Jordan, Maryam Kamali, Meredith Shrader, Ligia Cota Vieira, James Wilson, Fan Yang, Ying Jiang, Wanqi Hu, Gina Davis, Nancy Adamson, Amy Snyder, Laura Maxey, Anne Jones, Edward Mead and Ai Xia. v TABLE OF CONTENTS LIST OF FIGURES ............................................................................................................................... xii LIST OF TABLES ................................................................................................................................ xv CHAPTER 1 Literature Review ............................................................................................................... 1 Abstract ............................................................................................................................................... 1 Introduction ......................................................................................................................................... 1 I. White Grub Life Cycle & Infestation ............................................................................................ 1 II. Management ............................................................................................................................... 3 Entomopathogenic Nematodes ............................................................................................................. 4 I. Taxonomy & Biology .................................................................................................................. 4 II. Efficacies ................................................................................................................................... 7 III. Factors Affecting Efficacy ...................................................................................................... 12 Biotic factors ............................................................................................................................. 12 Abiotic factors ........................................................................................................................... 14 IV. Commercialization & Future Prospects ................................................................................... 16 Entomopathogenic Fungi ................................................................................................................... 17 I. History & Taxonomy ................................................................................................................. 17 II. Pathogenicity & Safety ............................................................................................................. 19 III. Environmental Constraints ...................................................................................................... 21 IV. Commercialization & Future Prospects ................................................................................... 23 Conclusion ........................................................................................................................................ 24 References ........................................................................................................................................