LIFE HISTORY INVESTIGATIONS OF HEMIPTERANS SELECTED FOR NON-TARGET HOST-SUITABILITY STUDIES OF THE SPOTTED LANTERNFLY (LYCORMA DELICATULA) BIOCONTROL AGENT ANASTATUS ORIENTALIS, WITH MOLECULAR INVESTIGATIONS OF LOCAL ANASTATUS TAXA by Tyler Hagerty A thesis submitted to the Faculty of the University of Delaware in partial fulfillment of the requirements for the degree of Master of Science in Entomology Summer 2020 © 2020 Tyler Hagerty All Rights Reserved LIFE HISTORY INVESTIGATIONS OF HEMIPTERANS SELECTED FOR NON-TARGET HOST-SUITABILITY STUDIES OF THE SPOTTED LANTERNFLY (LYCORMA DELICATULA) BIOCONTROL AGENT ANASTATUS ORIENTALIS, WITH MOLECULAR INVESTIGATIONS OF LOCAL ANASTATUS TAXA by Tyler Hagerty Approved: __________________________________________________________ Charles Bartlett, Ph.D. Professor in charge of thesis on behalf of the Advisory Committee Approved: __________________________________________________________ Jacob L. Bowman, Ph.D. Chair of the Department of Entomology and Wildlife Ecology Approved: __________________________________________________________ Mark W. Rieger, Ph.D. Dean of the College of Agriculture and Natural Resources Approved: __________________________________________________________ Douglas J. Doren, Ph.D. Interim Vice Provost for Graduate and Professional Education and Dean of the Graduate College ACKNOWLEDGMENTS I would first like to thank my advisor, Dr. Charles Bartlett, for his continuous help and support through my time at the University of Delaware. His help and guidance helped shape and push my research forward, and his knowledge of Hemiptera was indispensable. His door was always open, and he was happy to listen to me complain about insects not laying eggs whenever I needed to. I would like to thank my committee members, Dr. Debbie Delaney of the University of Delaware, and Dr. Kim Hoelmer of the USDA ARS. Each helped me in numerous ways and offered assistance and support whenever I needed it. This project would not have been possible without their guidance. In addition I would like to thank the USDA (especially the USDA ARS BIIRU facility in Newark, DE and the Otis Laboratory in Buzzards Bay, MA) and the Farm Bill for providing funding for this research and allowing me to work with such interesting species. I would also like to thank Dr. Hannah Broadley and Dr. Juli Gould (USDA APHIS) for their help with multiple aspects of this research. Finally, I would like to thank all my colleagues and family, especially my wife Michelle, for their constant support and encouragement throughout my years of research. This thesis would not have been possible without them. iii TABLE OF CONTENTS LIST OF TABLES ........................................................................................................ vi LIST OF FIGURES ...................................................................................................... vii ABSTRACT .................................................................................................................. ix Chapter 1 THE ECOLOGY AND LIFE HISTORY OF THE INVASIVE SPOTTED LANTERNFLY (LYCORMA DELICATULA) ................................................... 1 2 GENETIC ANALYSIS OF THE GENUS ANASTATUS (HYMENOPTERA: EUPELMIDAE) ................................................................ 9 2.1 Literature Review for Anastatus ................................................................ 9 2.2 Methods ................................................................................................... 12 2.3 Results ..................................................................................................... 18 2.4 Discussion ................................................................................................ 20 3 TAXONOMY OF FOCAL HEMIPTERA AND REARING TECHNIQUES OF SEVERAL NATIVE HEMIPTERAN SPECIES TO BE USED IN NON-TARGET BIOLOGICAL CONTROL TESTS ...................................... 22 3.1 Family Choices for Non-Targets species ................................................ 22 3.2 Selection of Non-target Species for Collection and Evaluation .............. 23 3.3 Life History and Ecology of Non-Target Taxa ....................................... 25 3.4 Materials and Methods ............................................................................ 33 3.5 Results ..................................................................................................... 36 3.6 Discussion ................................................................................................ 54 REFERENCES ............................................................................................................. 67 Appendix A QIAGEN DNEASY BLOOD AND TISSUE KIT ........................................... 75 B SMALL GEL ELECTROPHORESIS METHODS .......................................... 77 C MEDIUM GEL ELECTROPHORESIS METHODS ...................................... 79 D QIAGEN PCR PURIFICATION METHODS ................................................. 81 E WB2 CO1 SEQUENCE ................................................................................... 82 iv F COLLECTING METHODS ............................................................................. 83 G REARING AND EGG COLLECTION METHODS ....................................... 88 H BEHAVIORAL OBSERVATIONS ................................................................. 94 v LIST OF TABLES Table 1. List of biological control agents for spotted lanternfly (all Hymenoptera) ...... 5 Table 2. Anastatus (Hymenoptera: Eupelmidae) species extracted with method ........ 18 Table 3. Pairwise distance for Anastatus reduvii CO1 sequences ................................ 19 Table 4. Species selected for non-target host suitability studies .................................. 34 Table 5. Rearing techniques summary ......................................................................... 35 Table 6. Collection results for species targeted for host suitability studies ................. 37 vi LIST OF FIGURES Figure 1. Adult spotted lanternfly (Lycorma delicatula) ................................................ 8 Figure 2. Covered Lycorma delicatula egg mass ........................................................... 8 Figure 3. Gel with DNA banding from Anastatus reduvii extraction .......................... 20 Figure 4. Adult female Anastatus reduvii ..................................................................... 21 Figure 5. Adult female Anastatus disparis ................................................................... 21 Figure 6. Adult Poblicia fuliginosa .............................................................................. 54 Figure 7. Adult Scolops sulcipes .................................................................................. 55 Figure 8. Adult Rhynchomitra microrhina on plant material in lab ............................. 55 Figure 9. Adult Flatormenis proxima ........................................................................... 55 Figure 10. Two adult Metcalfa pruinosa photographed in the field ............................. 56 Figure 11. Seven adult Ormenoides venusta photographed by Charles Bartlett .......... 56 Figure 12. Adult Acanalonia conica ............................................................................ 56 Figure 13. Four adult Acanalonia bivittata on Robinia pseudoacacia in rearing cage 57 Figure 14. Adult Aplos simplex .................................................................................... 57 Figure 15. Several Thelia bimaculata on Robinia pseudoacacia in the field in Delaware .................................................................................................. 57 Figure 16. Two adult Anasa tristis mating ................................................................... 58 Figure 17. Strip of Leptoglossus fulvicornis eggs on Magnolia virginiana ................. 58 Figure 18. Rearing cage used for Poblicia fuliginosa .................................................. 58 Figure 19. General rearing cage setup for Acanaloniidae, Issidae, Flatidae and Membracidae ........................................................................................... 59 vii Figure 20. Field collecting site for Poblicia fuliginosa in Bladen County, North Carolina ................................................................................................... 59 Figure 21. Single “favored tree” (Rhus copallinum) of Poblicia fuliginosa in Bladen County, North Carolina ............................................................... 59 Figure 22. Late instar Poblicia fuliginosa on Rhus copallinum with seeping feeding wound ...................................................................................................... 60 Figure 23. Poblicia fuliginosa egg mass laid on a plastic pot ...................................... 60 Figure 24. Lab hatched first instar Poblicia fuliginosa ................................................ 60 Figure 25. Late instar Poblicia fuliginosa nymph hatched in the greenhouse ............. 61 Figure 26. Collecting habitat for Rhynchomitra microrhina in Newark, Delaware ..... 61 Figure 27. Rhynchomitra microrhina egg placed at base of petiole on Solidago ........ 62 Figure 28. Rhynchomitra microrhina egg .................................................................... 62 Figure 29. Rhynchomitra microrhina egg after 210 days ............................................. 62 Figure 30. Rhynchomitra microrhina egg after 211 days ............................................. 63 Figure