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Clubroot in Canola and Cabbage in Relation to Soil Temperature, Plant Growth and Host Resistance

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Clubroot in Canola and Cabbage in Relation to Soil Temperature, Plant Growth and Host Resistance Clubroot in canola and cabbage in relation to soil temperature, plant growth and host resistance By Thomas Vinzenz Gludovacz A Thesis presented to The University of Guelph In partial fulfillment of requirements for the degree of Master of Science in Plant Agriculture Guelph, Ontario, Canada © Thomas Vinzenz Gludovacz, May, 2013 ABSTRACT CLUBROOT IN CANOLA AND CABBAGE IN RELATION TO SOIL TEMPERATURE, PLANT GROWTH AND HOST RESISTANCE Thomas Vinzenz Gludovacz Advisors: University of Guelph, 2013 Dr. Mary Ruth McDonald Dr. Bruce D. Gossen The effects of diurnal temperature fluctuation and the utility of degree days for modeling clubroot on canola (Brassica napus L.) caused by Plasmodiophora brassicae Woronin were assessed using microscopy and qPCR, and in field trials. Temperature fluctuation had little effect on pathogen development. The optimal temperature for root hair infection was 25° C. Air and soil degree days and rainfall were used as metrics for estimating clubroot development, with only limited success. Several cultivars of cabbage (Brassica oleracea L. var. capitata) with unknown clubroot resistance mechanism(s) were assessed using staining and microscopy, and qPCR. In field trials, ‘Bronco’ was susceptible to clubroot (100 DSI), ‘Kilaherb’ was resistant (0 DSI), and ‘B-2819’ was intermediate (53 DSI). Plasmodiophora brassicae was present in cortical tissue of all cultivars. A delayed disease phenotype in ‘B-2819’ may indicate a quantitative resistance genotype that could be exploited in research on resistance genes and breeding. ACKNOWLEDGEMENTS Completing my Masters of Science degree has been the most challenging undertaking of the first 24 years of my life. It has been an honour to spend a few years in the Department of Agriculture at the University of Guelph. I would like to thank my advisors Dr. Mary Ruth McDonald and Dr. Bruce Gossen for their guidance, encouragement to succeed, and for challenging me to be a better scientist. I would like to thank my committee member Dr. Sean Westerveld for providing me with very valuable criticism and suggestions for my research. I owe a great deal of gratitude to Dr. Kalpana Sharma, Dr. Abhinandan Deora, Dr. Monica Parker, Hema Kasinathan, Nael Thaher and the rest of my lab for their mentorship, guidance, and assistance in my research, I learned so much about agriculture, plant pathology, and research from them. I am particularly grateful for the training in molecular biology given by Dr. Rachid Lahlali and for mentoring me throughout my program. Assistance provided by Dr. Michael Tesfaendrias and Dr. Cezarina Kora in helping to setup and troubleshoot the ThetaProbe for my field studies was greatly appreciated. I wish to acknowledge the help provided by Laura, Shawn, Michael, Mitchigo and Dennis from the Muck Station, as well as Ken Bassendowski, and Linda McGregor from the Saskatoon Research Centre (AAFC) for their technical assistance and field research training. I am indebted to Ken for his help in operating the thermal gradient plates. Thanks to the Animal Health Laboratory at the University of Guelph for preparing the cortical tissue slides for Chapter 4, and to Chris Granger for helping to setup the Real- Time PCR machine in the Crop Science building. Special thanks to Kalpana Adhikari and iii Dr. Sean Westerveld again for loaning the field data that was incorporated into the clubroot prediction models in Chapter 3. My thanks are extended to the staff of the Crop Science building and the Department of Plant Agriculture for working behind the scenes and doing all the paper work to make everyone’s research and programs of study run more smoothly. Thank you to Christophe Liseron-Monfils and Jerlene Nessia for their encouragement and advice in my research and studies. I would like to express my very great appreciation to Stephanie Khurana for her support during this program and for the countless hours of editing of this thesis and other course work. Editorial advice given by Alixandra Bamford has been a great help in improving my writing. Finally, I would like to thank my mother, father, and friends for their support throughout my program. iv TABLE OF CONTENTS ABSTRACT ........................................................................................................................ ii ACKNOWLEDGEMENTS ............................................................................................... iii TABLE OF CONTENTS .................................................................................................... v LIST OF TABLES ........................................................................................................... viii LIST OF FIGURES ........................................................................................................... ix CHAPTER ONE LITERATURE REVIEW ....................................................................... 1 1.1 Agricultural significance of canola and other Brassica spp. .................................... 1 1.1.1 Canola (B. napus and B. rapa) ........................................................................... 1 1.1.2 Head cabbage (B. oleracea var. capitata) and other Brassica vegetables ......... 3 1.1.3 Diseases of Brassica spp. ................................................................................... 5 1.2 Clubroot of Brassica ................................................................................................. 8 1.2.1 Significance ........................................................................................................ 8 1.2.2 Plasmodiophora brassicae ................................................................................. 9 1.2.3 Characterization and distribution of P. brassicae populations ........................ 18 1.3 Factors affecting clubroot severity ......................................................................... 20 1.3.1 Temperature ..................................................................................................... 21 1.3.2 Soil pH ............................................................................................................. 24 1.3.3 Soil moisture .................................................................................................... 25 1.3.4 Spore load ........................................................................................................ 26 1.3.5 Light intensity .................................................................................................. 27 1.4 Clubroot management ............................................................................................. 27 1.4.1 Cultural controls ............................................................................................... 28 1.4.2 Biocontrols ....................................................................................................... 34 1.4.3 Fungicide management .................................................................................... 36 1.4.4 Host resistance ................................................................................................. 40 1.5 Techniques for quantifying clubroot development ................................................. 45 1.5.1 Microscopy ...................................................................................................... 45 1.5.2 Molecular techniques ....................................................................................... 47 1.5.3 Clubroot symptoms .......................................................................................... 48 v 1.6 Summary and objectives ......................................................................................... 49 CHAPTER TWO EFFECT OF CONSTANT AND FLUCTUATING TEMPERATURES ON THE INCIDENCE AND SEVERITY OF CLUBROOT ........................................... 52 2.1 Introduction ............................................................................................................. 52 2.2 Materials and methods ............................................................................................ 54 2.2.1 Constant and fluctuating temperatures trials .................................................... 54 2.2.2 Range of temperature fluctuation ..................................................................... 58 2.2.3 Statistical analysis ............................................................................................ 58 2.3 Results ..................................................................................................................... 60 2.3.1 Root hair infection ........................................................................................... 60 2.3.2 Molecular quantification of in planta colonization of root hairs ..................... 63 2.4 Discussion ............................................................................................................... 68 CHAPTER THREE DEGREE DAY MODELING OF CLUBROOT INCIDENCE AND SEVERITY ON CANOLA ............................................................................................... 76 3.1 Introduction ............................................................................................................. 76 3.2 Materials and methods ............................................................................................ 79 3.2.1 Seeding date trial .............................................................................................. 79 3.2.2 Degree day calculation ....................................................................................
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