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Characterization of the Stemphylium Blight Pathogens and Their Effect on Lentil Yield

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Characterization of the Stemphylium Blight Pathogens and Their Effect on Lentil Yield Characterization of the stemphylium blight pathogens and their effect on lentil yield A Thesis Submitted to the College of Graduate Studies and Research In Partial Fulfillment of the Requirements For the Degree of Masters of Science In the Department of Plant Sciences University of Saskatchewan Saskatoon By Kiela B. Caudillo-Ruiz © Copyright Kiela B. Caudillo-Ruiz, July, 2016. All rights reserved. PERMISSION TO USE In presenting this thesis in partial fulfillment of the requirements for a postgraduate degree from the University of Saskatchewan, I agree that the libraries of this University may make it freely available for inspection. I further agree that permission for copying of this thesis in any manner, in whole or in part, for scholarly purposes may be granted by the professor or professors who supervised my thesis work or, in their absence, by the Head of the Department or the Dean of the College in which my thesis work was done. It is understood that any copying or publication or use of this thesis or parts thereof for financial gain shall not be allowed without my written permission. It is also understood that due recognition shall be given to me and to the University of Saskatchewan in any scholarly use which may be made of any material in my thesis. Requests for permission to copy or to make other use of material in this thesis in whole or part should be addressed to: Head of the Department of Plant Sciences University of Saskatchewan 51 Campus Drive Saskatoon, Saskatchewan S7N 5A8 i ABSTRACT The disease stemphylium blight has become common in lentil fields in Saskatchewan, but the effect of this disease on developing lentil plants, has not been studied under field conditions. Even though Stemphylium botryosum is suspected to be the pathogen causing stemphylium blight in lentil around the world, there is no confirmation of the pathogenic species involved in stemphylium blight of lentil in Saskatchewan. The objectives of this study were to determine the effect on lentil seed quantity and quality of S. botryosum infection at the seedling, the early- flowering, the mid-flowering or the podding stages, and to characterize the pathogen or pathogens causing stemphylium blight in Saskatchewan lentil fields. Three field experiments were conducted over two years using green polyethylene low tunnels to create conducive environments and control the timing of infection. Cumulative disease severity, measured as area under the disease progress curve, in lentil treatments inoculated at the seedling stage was higher compared to treatments inoculated at later growth stages or to uninoculated control treatments, in which some stemphylium blight developed due to natural inoculum. Neither the amount of harvested seed, nor seed weight or seed size were reduced compared to the uninoculated control, even though disease severity was higher in inoculated versus uninoculated treatments. Seed infection levels of 2.6 to 3.4% in seed harvested from treatments that were inoculated at the seedling, early and mid-flower stages were significantly higher than those observed in seeds from uninoculated control treatments (0.6%) or from treatments inoculated at podding (1.2%). Field isolates of Stemphylium spp. were compared to the ex-type isolate by morphology, and additionally to sequence data of five Stemphylium spp. and one Alternaria sp. obtained from GenBank by molecular phylogenetic analyses of the internal transcribe spacer (ITS) and the glyceraldehyde 3 – phosphate dehydrogenase (gpd) gene regions. Morphology of colony and conidia were not informative since features overlapped except for three isolates. Results of the molecular phylogenetic analyses revealed that S. botryosum is one of two possible Stemphylium spp. involved in the development of stemphylium blight in lentil. The three isolates with different morphology were also consistently clustered as a species distinct from the Stemphylium species. Although yield loss could not be demonstrated here, further studies on the epidemiology of the pathogens causing stemphylium blight in Saskatchewan lentil fields are warranted in view of the fact that more than one candidate species was identified as the causal agent. ii ACKNOWLEDGEMENTS I would like to thank my supervisor Dr. Sabine Banniza for her guidance and support, for sharing her knowledge and for her immense patience during this research. Also, I would like to thank my committee members Drs. Randy Kutcher, Steve Shirtliffe, Yuguang Bai and Dr. Godfrey Chongo who served as external examiner, for their guidance and advice. I am thankful to the Saskatchewan Pulse Growers Association and the Saskatchewan Ministry of Agriculture for providing the funding for the development of this project. I want to also thank the McLean Memorial Bursary that provided me with some financial support. A special thanks to the great team from the Pulse Pathology Laboratory which includes Stephanie Boechler, Cheryl Cho, Vijai Bhadauria, David Congly, Kamal Pathirannehelage, Laura Cox and my friend Nimllash Sivachandrakumar for their help, support, advice and friendship. I want to thank as well the Field Lab Crew under Brent Barlow’s guidance, for their immense help during the field experiments. Finally, I would like to thank my family for their unconditional support, love and prayers from back home; and special thanks to my son Leonardo who inspired and encouraged me every day and made my life better. iii DEDICATION I want to dedicate this thesis to my dad Manuel Caudillo and my mom Dora Ruiz, to my beautiful sisters Manuela, Dora, Olympia and Edelmira and to my son Leonardo iv TABLE OF CONTENTS PERMISSION TO USE ................................................................................................................... i ABSTRACT .................................................................................................................................... ii ACKNOWLEDGEMENTS ........................................................................................................... iii DEDICATION ............................................................................................................................... iv TABLE OF CONTENTS .................................................................................................................v LIST OF TABLES ....................................................................................................................... viii LIST OF FIGURES ....................................................................................................................... ix LIST OF APPENDICES ................................................................................................................ xi LIST OF ABBREVIATIONS ...................................................................................................... xiii 1. INTRODUCTION ....................................................................................................................1 2. LITERATURE REVIEW .........................................................................................................4 2.1. Lens culinaris Medikus ........................................................................................................4 2.1.1 Origins ................................................................................................................................... 4 2.1.2 Taxonomic classification ....................................................................................................... 4 2.1.3 Morphology ........................................................................................................................... 5 2.1.4 Life cycle ............................................................................................................................... 6 2.1.4.1 Flowering process ............................................................................................................. 6 2.1.4.2 Physiology of seed yield ................................................................................................... 7 2.1.5 Diseases ................................................................................................................................. 7 2.2. Importance of Stemphylium species ....................................................................................8 2.2.1 Yield loss due to stemphylium blight in lentil ..................................................................... 11 2.2.2 Stemphylium botryosum ....................................................................................................... 11 2.2.2.1 Classification, taxonomy and identification .................................................................... 11 2.2.2.2 Biology and life cycle ..................................................................................................... 13 2.2.3 Control options and agronomic management ...................................................................... 15 v 3. STEMPHYLIUM BLIGHT INDUCED YIELD LOSS IN LENTIL .....................................17 3.1 Introduction ........................................................................................................................17 3.2 Materials and Methods .......................................................................................................18 3.2.1 Plant material ....................................................................................................................... 18 3.2.2 Determination of the optimal conidial
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