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Effects of Temperature on Wheat-Pathogen Interactions

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Effects of Temperature on Wheat-Pathogen Interactions Effects of temperature on wheat-pathogen interactions Ruth Bryant A thesis submitted to the University of East Anglia for the degree of Doctor of Philosophy John Innes Centre September 2013 © This copy of the thesis has been supplied on condition that anyone who consults it is understood to recognise that its copyright rests with the author and that no quotation from the thesis, nor any other information derived there from, may be published without the author’s prior, written consent 1 Abstract Climate change is affecting UK agriculture, and research is needed to prepare crops for the future. Wheat is the UK’s most important crop, and needs to be protected from losses caused by disease. While direct effect of the environment on pathogen spread is often reported, effect of the environment on host defence is not. Many wheat resistance genes are temperature sensitive and these were used as a starting point to investigate defence temperature sensitivity in wheat starting with yellow rust resistance gene Yr36, previously shown to be temperature-sensitive. The effect of temperature on resistance was shown to be independent of Yr36 in breeding line UC1041, and was more likely to be due to a previously-uncharacterised background temperature sensitivity. These results suggest that temperature changes, rather than thresholds, might influence some disease resistance mechanisms. Understanding this phenomenon could enable the breeding of more stable defence in crops. In order to gain further insight into how temperature changes influence resistance, plants were grown under different thermoperiods and challenged with different types of pathogens; Results showed that resistance to multiple pathogens in one cultivar Claire was enhanced under variable temperatures, compared to constant temperatures. Taken together, the research presented revealed that defence temperature sensitivity in plants is much more complex than previously thought, considering that both temperature changes and different thermoperiods can influence aspects of wheat defence. To ascertain which research approaches will be most valuable in preparing for climate change, the effect of the environment on take-all was also investigated. Vulnerable periods for wheat from the threat of take-all development were identified by analysing historical datasets, and controlled environment experiments. Results showed a relationship between initial post-sowing temperatures and spring take-all levels in 2nd 3rd or 4th winter wheats, depending on the location. The work on yellow rust resistance and take-all both identify vulnerable periods for wheat caused by the environment, be it weakening of host defence responses, or increased threat from disease pressure. Further characterisation and understanding of vulnerable periods will be essential to control disease outbreaks under an increasingly unstable climate. 2 Acknowledgements I’d firstly like to thank my supervisors, Chris Ridout and Steve Dorling for their support over the last four years. Thanks also go to Lesley Boyd for her yellow rust knowledge and especially Graham McGrann for much scientific discussion and guidance. In addition I am obliged to Manda Sansom of Monsanto Ltd for providing me with data from take-all trials and The electronic Rothamsted Archive (e-RA) for providing me with meteorological records. Thanks to Henry Creissen and Chris Burt for stats help and all the JI horticultural staff and technicians who do a wonderful job, and so did you in summer 2012 Alice Mitchell! I am also very grateful to Henk-jan who helped me in the lab on a day to day basis. Thank you Henkie for being both a friend and a very nice colleague, with all the time in the world to listen to my science woes. This also applies to Gary who made me feel welcome in the lab and office when I was new. Thank you for taking me to tea for the first time, and both you and Rob for taking me to tea on the many occasions after that. I’m grateful to my conference companions who were common to many of my congress goings and made them fun as well as a learning experience; Philippa, Nikolai, Nick, Laura and Albor. Although to this day I still insist that you didn’t wait for me on that train from London to Norwich, not the other way round. Thanks again go to Nikolai, and also Pauline, for the PhD Plant science retreat 2012. We were a good team and so were we Dan Tromans, during the SVC coalition. Thank you to all my family for your support and taking my mind off things. This is also true of friends, Philippa, Simon, Ellen, Rob, Lucy, Alice, Alastair, Doreen and Hadrien to name just a few. And Mike, right at the end, thank you for seeing me through the final months. The Norwich lindy hoppers also got me though some tough Mondays and will be missed, along with my allotment. And lastly I want to especially thank someone who was predominant in getting me here and for making me willing to try anything at least once. They know who they are. 3 Contents Contents Abbreviations ........................................................................................................................... 9 Chapter 1: General introduction ............................................................................ 11 1.1 The disease triangle .................................................................................................. 11 1.2 Plant disease and climate change ............................................................................. 12 1.3 Climate change in the UK .......................................................................................... 14 1.4 Effect of environment on plant defence ................................................................... 16 1.5 Rust resistance as a model for investigating effect of temperature on wheat defence ..................................................................................................................... 21 1.6 Effect of environment on plant pathogens ............................................................... 24 1.7 Research aims ........................................................................................................... 27 1.7.1 Effect of temperature changes in defence against Pst ................................ 28 1.7.2 Impact of climate change on the incidence of take-all ................................ 29 Chapter 2: Materials and Methods ........................................................................ 30 2.1 Materials ................................................................................................................... 30 2.1.1 Plants ........................................................................................................... 30 2.1.2 Bacterial Strains ........................................................................................... 30 2.1.3 Fungal isolates ............................................................................................. 30 2.1.4 Chemicals and antibiotics ............................................................................ 30 2.1.5 Pathogen culture media and buffers ........................................................... 32 2.1.6 Plant growth media...................................................................................... 33 2.2 Methods .................................................................................................................... 34 2.2.1 Plant growth ................................................................................................ 34 2.2.2 Temperature regimes .................................................................................. 34 2.2.3 Pathogen isolation and maintenance .......................................................... 35 2.2.3.1 Yellow rust (Pst) .............................................................................. 35 2.2.3.2 Take-all (Ggt) .................................................................................. 35 2.2.3.3 Powdery mildew (Bgt) .................................................................... 36 2.2.3.4 P. syringae ....................................................................................... 36 2.2.3.5 Fusarium culmorum ........................................................................ 37 2.2.4 Microscopy ................................................................................................... 37 2.2.5 Sampling tissue, storage and grinding ......................................................... 37 4 Contents 2.2.6 Wheat Genotyping ....................................................................................... 37 2.2.7 RT-qPCR ........................................................................................................ 38 2.2.8 Primers ......................................................................................................... 39 2.2.9 Data analysis ................................................................................................ 42 Chapter 3: Exploring that nature of temperature sensitive resistance to yellow rust in wheat ................................................................................................. 43 3.1 Aim: .......................................................................................................................... 43 3.2 Introduction .............................................................................................................. 43 3.3 Methods ...................................................................................................................
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