Some Studies on Graminicolous Didymella spp. in New Zealand A Thesis submitted in partial fulfilment of the requirements for the degree of Doctor of Philosophy at the University Of Canterbury by Michelle Alexandra Mace-::;: 1999 To my family and Nathalie Table of Contents TABLE OF CONTENTS Table of Contents ......................................................................................................... i Table of Figures and Plates .......................................................................................... vii Table of Tables ............................................................................................................ x List of Abbreviations ................................................................................................. xviii Acknowledgements ...................................................................................................... xix Abstract ........................................................................................................................ xxi 1.0 Introduction 1.01 Hosts .................................................................................................................... 1 1.02 Taxonomy of Didymella ...................................................................................... 2 1.021 Appearance of D. exitialis on host substrate and in culture ........................ 2 1.022 Disease impact on wheat and endophytic nature of D. exitialis ................. 3 1.03 Didymella species in the Canterbury and Southland Regions ............................. 5 1.04 Life cycle ............................................................................................................. 7 1.05 The infection process ........................................................................................... 7 1.06 Control .................................................................................................................. 7 1 .07 Resistance ............................................................................................................ 8 1.08 The role of D. exitialis in late summer asthma .................................................... 8 1.09 Aims and objectives of the study ......................................................................... 10 2.0 Field investigations of Didymella spp. 2.01 Introduction .......................................................................................................... 11 2.02 Resistance ............................................................................................................ 11 2.03 Fungicides ............................................................................................................ 11 2.04 Endophytes ........................................................................................................... 12 2.05 Sporobolomyces ................................................................................................... 12 2.06 Latent infections ................................................................................................... 13 2.07 The confusion between the distinction of a latent and an endophytic nature ...... 14 2.08 Yield, endophytes, environment, fungicides and disease .................................... 15 2.09 Aims ..................................................................................................................... 16 2.10 Materials and methods ......................................................................................... 17 2.101 Trials and assessments ................................................................................ 17 2.1011 1995/96 .............................................................................................. 17 2.1012 1996197 .............................................................................................. 18 2.1013 1997/98 .............................................................................................. 18 2.102 Assessments of disease, plant pathogens and senescence in the trials ....... 18 2.1021 1995/96 .............................................................................................. 18 2.1022 1996197 .............................................................................................. 19 2.1023 1997/98 .............................................................................................. 19 2.1 03 Isolations from surface sterilised leaves ..................................................... 19 2.1031 1995/96 and 1996197 ......................................................................... 19 2.104 Enumeration ofleaf surface microflora ...................................................... 20 ii 2.1041 1995/96 .............................................................................................. 20 2.1042 1996/97 .............................................................................................. 21 2.105 Direct counts of micro-organisms present in the phylloplane .................... 22 2.106 High humidity and leaf imprinting ............................................................. 22 2.107 Field survey ................................................................................................. 23 2.1071 1995/96 .............................................................................................. 23 2.1072 1996/97 .............................................................................................. 23 2.108 The effects of fungicide applications on the Sporobolomyces populations .................................................................................................. 24 2.1081 1996/97 and 1997/98 ......................................................................... 24 2.109 Resistance ................................................................................................... 24 2.1091 1995/96 and 1996/97 ......................................................................... 24 2.110 Fungicide experiment ................................................................................. 24 2.20 Results .................................................................................................................. 27 2.201 Disease ........................................................................................................ 27 2.2011 1995/96 .............................................................................................. 27 2.2012 1996/97 .......................................................... , ................................... 27 2.202 Isolations from surface sterilised leaves ..................................................... 28 2.2021 Levels of Didymella spp., Sporobolomyces spp., and bacterial species cultured from wheat leaves from the trials carried out during 1995/96 and 1996/97 .............................................................. 28 2.20211 1995/96 ................................................................................... 28 2.20212 1996/97 ................................................................................... 28 2.2022 Levels of Didymella isolated from tebuconazole and azoxystrobin treated leaves ................................................................ 29 2.20221 1995/96 ................................................................................... 29 2.20222 1996/97 ................................................................................... 29 2.203 Field survey ................................................................................................. 48 2.2031 1995/96 ............................................... :.............................................. 48 2.2032 1996/97 .............................................................................................. 48 2.2033 Grasses present at the crop margins ................................................... 49 2.2034 Resistance .......................................................................................... 50 2.20341 1995/96 and 1996/97 .............................................................. 50 2.2035 Sporulation of Didymella on senesced leaf material ......................... 50 2.20351 1995/96 ................................................................................... 50 2.20352 1996/97 ................................................................................... 52 2.2036 Presence of Sporobolomyces in 1996/97 and 1997/98 ...................... 53 2.2037 Seed plating ....................................................................................... 54 2.2038 Antagonism ................................................................ : ....................... 54 2.2039 High humidity and leaf imprinting .................................................... 54 2.2040 Embedded leaves ............................................................................... 55 2.2041 Leaf clearing ...................................................................................... 56 2.205 Indirect method of enumeration of phylloplane micro-organisms ............. 56 2.2051 Leaf washings .................................................................................... 56 2.20511 1995/96 ................................................................................... 57 2.20512 1996/97 ................................................................................... 59 2.206 The effect of fungicide application on the duration of green flag leaf area ......................................................................................................