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Phenolic and Lignin Concentration As an Indicator of Resistance to Phytophthora Cinnamomi in Banksia Species

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Phenolic and Lignin Concentration As an Indicator of Resistance to Phytophthora Cinnamomi in Banksia Species Edith Cowan University Research Online Theses : Honours Theses 1996 Phenolic and lignin concentration as an indicator of resistance to phytophthora cinnamomi in banksia species Sandra Jane Nicoski Edith Cowan University Follow this and additional works at: https://ro.ecu.edu.au/theses_hons Part of the Plant Biology Commons Recommended Citation Nicoski, S. J. (1996). Phenolic and lignin concentration as an indicator of resistance to phytophthora cinnamomi in banksia species. https://ro.ecu.edu.au/theses_hons/313 This Thesis is posted at Research Online. https://ro.ecu.edu.au/theses_hons/313 Edith Cowan University Copyright Warning You may print or download ONE copy of this document for the purpose of your own research or study. The University does not authorize you to copy, communicate or otherwise make available electronically to any other person any copyright material contained on this site. You are reminded of the following: Copyright owners are entitled to take legal action against persons who infringe their copyright. A reproduction of material that is protected by copyright may be a copyright infringement. Where the reproduction of such material is done without attribution of authorship, with false attribution of authorship or the authorship is treated in a derogatory manner, this may be a breach of the author’s moral rights contained in Part IX of the Copyright Act 1968 (Cth). Courts have the power to impose a wide range of civil and criminal sanctions for infringement of copyright, infringement of moral rights and other offences under the Copyright Act 1968 (Cth). Higher penalties may apply, and higher damages may be awarded, for offences and infringements involving the conversion of material into digital or electronic form. USE OF THESIS The Use of Thesis statement is not included in this version of the thesis. PHENOLIC AND LIGNIN CONCENTRATION AS AN INDICATOR OF RESISTANCE TO PHYTOPHTHORA CINNAMOMI IN BANKS/A SPECIES Sandra Jane Nicoski Thesis submitted in partial fulfilment of the requirement for the award of B. Sc.(Biological Science) Honours Department of Applied Science Edith Cowan University December 1996 "Extinction undoubtedly is being played out as we gather our thoughts today." - S. Hopper 1994 ABSTRACT Phytophthora cinnamomi Rands, a soilborne fungus, is regarded as one of the most devastating pathogens yet recorded in natural ecosystems. The disease, caused by P. cinnamomi, poses an enormous threat to the flora of the southwest. The Proteaceae, in particular, contribute considerably to species richness and provide the fundamental floristic structure of many south-western plant communities. The genus Banksia, a member of the Proteaceae, is very susceptible to disease caused by P. cinnamomi. Three spec1es, namely B. attenuata, B. grandis and B. menziesii. were established in an aeropomcs system under glasshouse conditions. Trials were conducted to optimise growing conditions in these systems and to determine the most effective method of inoculation. Shoots were treated with a 0.5% phosphonate foliar spray and roots were dipped in P. cinnamomi zoospore suspensions. The temporal, spatial and treatment changes in the concentrations of total soluble phenolics and lignin in roots, four and twelve days after inoculation, were measured. Total soluble phenolics decreased in inoculated treatments compared with controls. In addition, phenolic levels were even lower by twelve days after inoculation. In comparison, lignin concentrations did not ii have a significant effect on these compounds in B. menziesii, however, it increased in inoculated plants above control levels. Phosphonate application B. grandis produced increased levels of total soluble phenolics with phosphonate application. This study aimed at developing an understanding of the biochemical defences of Banksia species to Phytophthora, enables the resistance of these species to be compared and assessed. iii DECLARATION I certify that this thesis does not incorporate without acknowledgment any mate· rial previously submitted for a degree or diploma in any institution of higher education; and that to the best of my knowledge and belief it does not contain any material previously published or written by any other person except where due reference is made in the text. Sandra Jane Nicoski 6 December 1996 IV ACKNOWLEDGMENTS Firstly I would like to thank my supervisor Dr Ian Bennett for his guidance, patience and effort throughout this project and for helping me to develop confidence in my own ability. Also Dr Kim Tynan for her ever helpful advice and for taking the time to teach me new skills. I would also like to thank Mr Andrew Woodward, Ms Danielle Eyre, Ms Debra McDavid, Ms Bernadette Gunn and Ms Meredith Fairbanks for their encouragement, advice and amusing lunch conversations. A big thankyou to Mr Clay Miller, Mr Mark Bannister, Ms Linda Irvine ami especially to Mr Simon Collins for putting up with my constant pestering and for making me computer literate. To Kim Upton for her support, companionship and much valued friendship which made this year so much more enjoyable. Lastly to my mother for her encouragement, love and support for which I am very grateful. v TABLE OF CONTENTS PAGE Abstract 11 Declaration iv Acknowledgments v Table of Contents Vl List of Tables X List of Figures Xl CHAPTER: 1. INTRODUCTION 1 1.1 The Genus Banksia 1 1.1.1 Origins and Distribution 1 1.1.2 Ecological Value and Commercial Uses of Banksia 2 1.1.2.1 Ecological importance 2 1.1.2.2 Commercial uses 3 1.2 The Genus Phytophthora 4 1.2.1 Classification 4 1.2.2 Morphology and Reproduction 5 1.3 Phytophthora cinnamomi 7 1.3.1 Origin and Distribution in Australia 7 1.3.2 Host Range 8 1.3.3 Method oflnfection 10 Vl 1.3.4 Management of P. cinnamomi 11 1.3.4.1 Hygiene 11 1.3.4.2 Biological Control 12 1.3.4.3 Chemical Control: Phosphonates 13 1.4 Banl<sia Species and P. cinnamomi 16 1.4.1 Susceptibility toP. cinnamomi 16 1.4.2 Symptoms of Disease 17 1.5 Mechanisms of Resistance to Fungal Pathogens 19 1.5.1 Host-Pathogen Interactions 19 1.5.2 Mechanisms of Resistance 20 1.5.2.1 Phenolics 20 1.5.2.2 Lignin 22 1.5.3 Phenolics as Biochemical Markers for Resistance 24 1.5.4 Breeding for Resistance 26 1.6 Aims 27 1. 7 Hypotheses 28 2. GENERAL MATERIALS AND METHODS 29 2.1 Plant Material and Maintenance 29 2.2 P. cinnamomi Maintenance and Zoospore Production 31 2.3 Phenolic Analysis 33 2.4 Lignin Analysis 33 2.5 Plating of Roots 35 vii 3. PRELIMINARY INOCULATION TESTS 36 3.1 Introduction 36 3.2 Materials And Methods 38 3.2.1 Trial 1 38 3.2.2 Trial 2 38 3.2.3 Trial 3 39 3.2.4 Trial 4 39 3.2.5 Trial 5 40 3.2.6 Trial 6 40 3.2. 7 Trial 7 40 3.3 Results 41 3.3.1 Triall 41 3.3.2 Trials 2, 3 And 4 41 3.3.3 Trial 5 41 3.3.4 Trial 6 43 3.3.5 Trial 7 43 3.4 Discussion 46 4. LIGNIN AND PHENOLIC ANALYSIS 48 4.1 Introduction 48 4.2 Materials And Methods 50 4.2.1 Phosphonate Application 50 4.2.2 Design Of Biochemical Analysis Experiments 50 4.2.3 Experiment 1 51 Vlll 4.2.4 Experiment 2 51 4.2.5 Experiments 3, 4 and 5 52 4.2.6 Statistical Analysis 52 4.3 Results 52 4.3.1 Experiment 1 52 4.3.2 Experiment 2 54 4.3.3 Experiment 3 54 4.3.4 Experiment 4 58 4.3.5 Experiment 5 60 4.4 Discussion 62 4.4.1 Phenolics 62 4.4.2 Lignin 63 4.4.3 Effect of Phosphonate on Phenolic and Lignin Levels 65 5. GENERAL DISCUSSION 66 References 71 ix LIST OF TABLES TABLE PAGE 3.1 Lesion development in seedlings of B. menziesii Triall (means calculated from three replicates) 42 3.2 Recovery of P. cinnamomi from plated roots of B. menziesii (means calculated from five replicates) 42 3.3 Summary of results from Trials 1 to 6 45 X LIST OF FIGURES FIGURE PAGE 1.1 Lifecycle of P. cinnamomi 6 1.2 Distribution of P. cinnamomi in south.west Australia 9 2.1 Aeroponics system 32 3.1 The effect of plastic bags on mean increase in root length in (a) B. attenuata; (b) B. menziesii and (c) B. grandis. 44 4.1 Mean concentrations of(a) total soluble phenolics; (b) lignin in B. menziesii for treatments at days four and twelve in Experiment 1. 53 4.2 Mean concentration of total soluble phenolics in B. menziesii for treatment at day four in Experiment 2. 55 4.3 Mean concentrations of (a) total soluble phenolics; (b)lignin in B. menziesii for treatments at day four in Experiment 3. 56 4.4 Mean concentrations of (a) total soluble phenolics; (b)lignin in treatments in B. menziesii root segments in Experiment 3. 57 4.5 Mean concentrations of (a) total soluble phenolics; (b) lignin in B. grandis for treatments at day four in Experiment 4. 59 4.6 Mean concentrations of (a) total soluble phenolics; (b) lignin in B. menziesii for treatments at days four and twelve in Experiment 5. 61 Xl 1. INTRODUCTION 1.1 The Genus Banksia 1.1.1 Origins and Distribution The genus name, Banksia, commemorates the botanist Joseph Banks who accompanied Captain James Cook in 1770 to the east coast of Australia (Holliday and Watton, 1975). Of the 75 species of Banksia recognised, 58 are endemic to south-western Australia, and 14 occur in eastern Australia ranging from the Eyre peninsula in south Australia, to Tasmania and north Queensland (George, 1984).
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