Biological Control of Paropsis charybdis Stål (Coleoptera: Chrysomelidae) and the Paropsine Threat to Eucalyptus in New Zealand A Thesis submitted in fulfilment of the requirements for the Degree of Doctor of Philosophy in the University of Canterbury by Brendan Dene Murphy New Zealand School of Forestry University of Canterbury 2006 TABLE OF CONTENTS ABSTRACT v ACKNOWLEDGEMENTS vi ERRATA vii CHAPTERS Chapter 1. Biological Control of Paropsis charybdis Stål and the Paropsine Threat to Eucalyptus in New Zealand.................................................................................................... 1 Chapter 2. The Collection, Importation, and Release of Tasmanian Enoggera nassaui for Biological Control of Paropsis charybdis............................................................................. 8 Chapter 3. Molecular Detection of Enoggera nassaui Strains using the Mitochondrial DNA Gene, Cytochrome Oxidase I ............................................................................................... 22 Chapter 4. Field and Bioassay Assessment of the Host Range .................................................. 32 Chapter 5. Phylogenetic Reconstruction of Tasmanian Chrysophtharta ..................................45 Chapter 6. Assessment of Paropsine Fecundity as an Indicator................................................. 59 Chapter 7. Testing the Parasitoid Host Range and Reproductive Output Hypotheses against Dicranosterna semipunctata ................................................................................................ 78 Chapter 8. Discussion ................................................................................................................... 86 Chapter 9. Conclusions ................................................................................................................. 97 Chapter 10. References ................................................................................................................. 99 Chapter 11. Appendices..............................................................................................................119 FIGURES Figure 1. Paropsis charybdis adult...................................................................................................... 2 Figure 2. CLIMEX generated climate match of Busselton (Western Australia) ...................... 14 Figure 3. CLIMEX generated climate match of Florentine Valley (Tasmania) ....................... 14 Figure 4. Sources of Tasmanian E. nassaui strains.....................................................................16 Figure 5. Release sites of Tasmanian E. nassaui strains in the central North Island................ 17 Figure 6. Neighbour-joining dendogram of Enoggera nassaui sequences using Kimura 2- Parameter substitution model............................................................................................... 26 Figure 7. Enoggera nassaui. ............................................................................................................. 32 Figure 8. Mean one hour parasitism rate of Enoggera nassaui on Chrysophtharta, Paropsis and Trachymela hosts ........................................................................................................... 36 Figure 9. Scanning Electron Micrographs of Chrysophtharta and Paropsis egg chorion ....... 37 Figure 10. Chrysophtharta phylogeny using MP and K2P substitution model. ....................... 51 Figure 11. Chrysophtharta phylogeny using UPGMA and K2P substitution model. .............. 52 ii Figure 12. Chrysophtharta phylogeny using NJ methods and K2P substitution model........... 53 Figure 13. The chosen Chrysophtharta phylogeny, using NJ and JC69 substitution model ...53 Figure 14. Chorion analysis of Chrysophtharta phylogeny topology. ...................................... 54 Figure 15. Mean ± SE Chrysophtharta fecundity....................................................................... 65 Figure 16. Mean ± SE Paropsis fecundity................................................................................... 65 Figure 17. Mean ± SE Chrysophtharta oviposition rate............................................................. 66 Figure 18. Mean ± SE Paropsis oviposition rate ........................................................................ 66 Figure 19. Mean ± SE Chrysophtharta egg batch size ............................................................... 67 Figure 20. Mean ± SE Paropsis egg batch size........................................................................... 67 Figure 21. Correlation between fecundity and oviposition rate for paropsine species............. 68 Figure 22. Chrysophtharta phylogeny and mean fecundity....................................................... 69 Figure 23. Chrysophtharta phylogeny and oviposition rate....................................................... 70 Figure 24. Dicranosterna semipunctata egg. ..................................................................................... 79 TABLES Table 1. Summary of biological control attempts for Paropsis charybdis.................................. 3 Table 2. Field Collections of Tasmanian egg parasitoids on paropsine species ....................... 15 Table 3. New Zealand Release locations and quantities of Tasmanian E. nassaui strains....... 16 Table 4. Field Collections of Enoggera nassaui from Tasmanian Chrysophtharta and Paropsis.................................................................................................................................35 Table 5. Paropsine study species, host species, and numbers of replicates............................... 63 Table 6. Mean fecundity and oviposition rates of D. semipunctata and P. charybdis. ............ 81 Table 7. Mean E. nassaui parasitism rate (eggs per hour-1) and time to acceptance (minutes) on D. semipunctata and P. charybdis.................................................................................. 81 Table 8. Ranking of paropsine threat using reproductive output ............................................... 96 APPENDICES Appendix 1. Genus Chrysophtharta Weise...............................................................................119 Appendix 2. Genus Paropsis Olivier .........................................................................................120 Appendix 3. Tasmanian egg parasitoid egg collection records................................................121 Appendix 4. Enoggera nassaui COI sequence data..................................................................123 Appendix 5. Paropsine COI sequence data................................................................................126 Appendix 6. Chrysophtharta fecundity data .............................................................................130 Appendix 7. Paropsis and miscellaneous paropsine fecundity data........................................131 Appendix 8. Chrysophtharta oviposition rate data ...................................................................132 Appendix 9. Paropsis and miscellaneous species oviposition rate data..................................133 iii Appendix 10. Chrysophtharta mean egg batch size data .........................................................134 Appendix 11. Paropsis and miscellaneous paropsine mean egg batch size data ....................135 Appendix 12. Parasitism rates for Enoggera nassaui...............................................................136 Appendix 13. Published paper....................................................................................................137 Appendix 14. Published paper....................................................................................................141 Appendix 15. Published paper....................................................................................................146 iv ABSTRACT Ineffective biological control of the Eucalyptus pest Paropsis charybdis Stål (Coleoptera: Chrysomelidae: Paropsini) in cold areas of New Zealand was believed to be caused a climatic mismatch of the egg parasitoid Enoggera nassaui Girault (Hymenoptera: Pteromalidae). Two Tasmanian strains of the parasitoid were introduced to test climate-matching theory in 2000, with approximately 7000 wasps released. Establishment of the Florentine Valley strain was detected in 2002 using the Mitochondrial (mtDNA) gene Cytochrome Oxidase I (COI) as a strain specific marker. The hyperparasitoid Baeoanusia albifunicle Girault (Hymenoptera: Encyrtidae) and primary parasitoid Neopolycystus insectifurax Girault (Hymenoptera: Pteromalidae) were detected for the first time in New Zealand. As paropsines have proven highly invasive internationally, a risk assessment of the paropsine threat to New Zealand was undertaken by evaluating the host range of E. nassaui and a reproductive assessment of 23 paropsine species in the genera Dicranosterna Motschulsky, Chrysophtharta Weise, Paropsis Olivier, Paropsisterna Motschulsky and Trachymela Weise. Enoggera nassaui proved polyphagous, but bioassay results proved that Paropsis species were significantly more susceptible to the egg parasitoid than Chrysophtharta species. Resistance within Chrysophtharta was attributed to spine-like chorion modifications. A COI derived Chrysophtharta phylogeny divided the genus into two distinct groupings, which was supported by chorion morphology. Paropsine reproductive output was tested for key parameters indicating pest potential. Pest species displayed fecundity exceeding 600 eggs at an oviposition