The description, pathogenicity and epidemiology of Phytophthora boodjera, a new nursery pathogen of Eucalyptus from Western Australia by Agnes Virginia Simamora B.Sc. Agriculture (Universitas Nusa Cendana) MCP (Adelaide University) The thesis is submitted for the degree of Doctor of Philosophy School of Veterinary and Life Sciences Murdoch University Perth, Western Australia October 2016 Declaration I hereby declare that the work in this thesis is my own account of my research and contains as its main content work, which has not previously been submitted for a degree at any tertiary education institution. To the best of my knowledge, all work performed by others, published or unpublished, has been acknowledged. Agnes V. Simamora October 2016 ii Acknowledgments First and foremost I praise the Almighty God, my creator, the one who gives me strength and knows the plans intended for me; thank you for your graciousness and love. You are the one who gives me power to be successful. Many people have vitally assisted in making this PhD possible and pleasurable and I will be eternally grateful to you all. It is my great pleasure to express my gratitude to all those who have supported me and helped me during the past four years. Personally, I would like to especially thank my three supervisors (Prof. Giles Hardy, Assoc. Prof. Treena Burges, and Mike Stukely), whom I think had the hardest job. Thank you for giving me this opportunity, providing me with countless valuable guidance, and supporting me on my experiments and writing skills especially in times of adversity. Thank you so much for being patient with me when I still had much to learn. Your belief in me has kept me going and allowed me to reach this goal. For these gifts no words could ever express my gratitude. I also express my gratitude to the government of Indonesia for providing the Indonesian Higher Education scholarship, and to my University, Universitas Nusa Cendana, for study leave. I would like to especially thank all the staff and students at Murdoch University, the Centre of Phytophthora Science and Management, who helped with my experiments and training. To Dr. Michael Crone and Dr. Sonia Aghighi, I really appreciate your valuable help. I hope to work with you again throughout our careers. I am indebted to Ms. Diane White for helping me with my molecular analysis. Special thanks go to Dr. Trudy Paap for helping me in many ways throughout my project. Thank you dearly to Briony Williams, Dr. Bill Dunstan, and Janet Box for all your assistance. Also I thank Ian McKernan and Jose Minetto for assisting me in my glasshouse trials. iii I would like to thank all the administration staff at Murdoch University and in the School of Biological Sciences and Biotechnology for all your help over the years. To my dear friends Emma and Louise, your kind friendships, warm kind personalities and elated spirits have helped me throughout my journey; I could not have got this far without you. Emma, thank you for being my carer at the time of trouble. Louise, thank you for being my intercessor. To all the wonderful women in the office, thank you for sharing my PhD journey, your kind friendships, sisterhood and for all the fun we have had throughout this journey. Thank you Sarah, Manisha, Eman, Rajah, Louise, Emma, Nuk, and Tuyet for being my friends. I can’t wait for our reunion day. I also respectfully acknowledge Parnell’s nursery for the help throughout my project. It has been such a pleasure to work with you and your help on my project was invaluable. A huge thank you to Jodi Burgess for helping me to get some very stunning pictures for my thesis. I also place on record, my sense of gratitude to one and all, who directly or indirectly, have lent their hands in this venture. At last, but by no means least I would like to thank all my family and friends for their love, prayers and support throughout the years, with a special thanks to my dear late father, my mother (Ny. R. Simamora Bait), my brothers (in laws) and sisters (in laws), my nieces and nephews (Ucok, Martha, Victor, Daniel, David, Aya, Cilli, and Nova). I would not have accomplished so much or be where I am today without your love and prayers. I would especially like to thank my beloved husband, Sondang Siallagan; my precious son, Firman Siallagan, for their understanding, support and love, which has been irreplaceable throughout my journey. God bless you. iv Abstract When this study was commenced in November 2011, a recent outbreak of damping-off diasese in a Western Australia (WA) nursery had indicated the presence of a new Phytophthora species. Despite industry standard hygiene, the disease continued across years. The impact of this new disease in the nursery caused a great concern as the plants grown there were intended for environmental plantings and hence posed the risk of introducing a new species into the natural environment. This raised many questions to be cleared: (1) was this pathogen a new species, (2) was it only a damping-off pathogen or could it infect older seedlings or trees, (3) did it have a narrow or broad host range, (4) how did it get to the nursery and survive from year to year, (5) could it persist in environmental plantings, (6) was it endemic to Western Australia? This project addressed all of these questions. A new species, Phytophthora boodjera was described. It has a relatively narrow host range and is a pathogen of Eucalyptus. P. boodjera is especially a pre and post- emergence damping-off pathogen with host susceptibility decreasing with age. Older seedlings and trees have damaged root systems but did not die. Within the nursery P. boodjera survived between seedings in the debris of the used trays. Immersion in 5% Calcium hypochlorite or using dry heat at 65oC for 2 hours did not eliminate the inoculum. Tracing of infected seedlings from the nursery showed that P. boodjera can persist in the natural environment if introduced. While common in nursery and recovered from urban planting, extensive sampling in natural ecosystems has recovered very few isolates of P. boodjera. Based on this low recovery and the high susceptibility of Eucalyptus species tested I conclude that P. boodjera is not endemic to WA. v Publications and conference presentations Journal paper: Chapter 2 published as: Simamora A, Stukely M, Hardy G, and Burgess TI, 2015. Phytophthora boodjera sp. nov., a damping-off pathogen in production nurseries and from urban and natural landscapes, with an update on the status of P. alticola IMA FUNgUs · 6(2): 319–335 (2015). doi:10.5598/imafungus.2015.06.02.04 Chapter 3 published as : Simamora AV, Stukely MJC, Barber PA, Hardy GES, Burgess TI (2016) Age-related susceptibility of Eucalyptus species to Phytophthora boodjera. Plant Pathology, 10.1111/ppa.12592 Conferences presentations (2012-2015): Simamora AV, Hardy GESJ, Stukely M, and Burgess TI, 2012. More new Phytophthora species from natural ecosystems in Western Australia. Proceedings of the 6th meeting of the International Union of Forest Research Research Organizations (IUFRO ) Working Party 7- 02-09 “Phytophthora in Forests and Natural Ecosystems”. 9-14 September 2012, Cordoba, Spain. Oral presentation. Simamora AV, Paap T, Stukely M, Hardy GESJ and Burgess TI, 2013. A re-evaluation of the Phytophthora alticola-P. arenaria species complex. Dieback Information Group Conference, 28 June 2013 at WA State Library, Perth. Oral presentation. Simamora AV, Hardy GESJ, Stukely M, and Burgess TI, 2013. More new Phytophthora species from natural ecosystems in Western Australia. Australasian Plant Pathology Society (APPS) conference Auckland, NZ, 25-28 Nov 2013. Poster presentation. Simamora AV, Paap T, Stukely M, Hardy GESJ and Burgess TI, 2013. A new Phytophthora disease from nurseries in Western Australia. Australasian Plant Pathology Society (APPS) conference Auckland, NZ, 25-28 Nov 2013. Oral presentation. Simamora AV, Paap T, Stukely M, Hardy GESJ and Burgess TI, 2014. Epidemiology of Phytophthora boodjera prov. nom., a damping-off pathogen in tree production nurseries in Western Australia. Dieback Information Group Conference, 8 July 2014 at Murdoch University, Perth, WA. Oral presentation. Simamora AV, Stukely M, Hardy GESJ and Burgess TI, 2014. Age related susceptibility of Eucalyptus spp. to Phytophthora boodjera prov.nom. International Union of Forest Research Organizations (IUFRO ) Working Party 7-02-09 “Phytophthora in Forests and Natural Ecosystems”. 10-14 November 2014, Esquel, Argentina. Poster and oral presentation. Simamora AV, Paap T, Stukely M, Burgess TI, and Hardy GESJ, 2014. Epidemiology of Phytophthora boodjera prov. nom., a damping-off pathogen in tree production nurseries in vi Western Australia. International Union of Forest Research Organizations (IUFRO ) Working Party 7-02-09 “Phytophthora in Forests and Natural Ecosystems”. 10-14 November 2014, Esquel, Argentina. Poster and oral presentation. Simamora AV, Stukely M, Hardy GESJ and Burgess TI, 2015. Age related susceptibility of Eucalyptus species to Phytophthora boodjera prov.nom. APPS conference, Fremantle WA, 14-16 Sept 2015. Oral presentation. Simamora AV, Stukely M, Hardy GESJ and Burgess TI, 2015. Phytophthora boodjera sp. nov., a damping-off pathogen in production nurseries and from urban and natural landscapes, and an update on the status of P. alticola. Asian Mycological Congress, Goa India, 7-10 Oct 2015. Oral presentation. Murdoch University Poster Day Simamora AV, Paap T, Stukely M, Burgess TI, and Hardy GESJ, 2014. Epidemiology of Phytophthora boodjera prov. nom., a damping-off pathogen in tree production nurseries in Western Australia. 1 November 2014. Simamora AV, Stukely M, Hardy GESJ, and Burgess TI, 2015. Age related susceptibility of Eucalyptus species to Phytophthora boodjera. 6 November 2015. vii Table of contents Thesis title Declaration ..........................................................................................................................