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Characterization, Distribution in Ohio and Resistance Reactions to Two

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Characterization, Distribution in Ohio and Resistance Reactions to Two PATHOGENIC CHARACTERIZATION, DISTRIBUTION IN OHIO AND WHEAT GENOTYPE REACTIONS TO STAGONOSPORA NODORUM AND PYRENOPHORA TRITICI-REPENTIS DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Jessica S. Engle, M. S. The Ohio State University 2005 Dissertation Committee: Approved by Professor Patrick E. Lipps, Adviser Professor Laurence V. Madden ________________________________ Professor Clay Sneller Professor Margaret Redinbaugh Advisor Professor Sophien Kamoun Plant Pathology Graduate Program ABSTRACT Two fungal pathogens, Stagonospora nodorum and Pyrenophora tritici- repentis infect wheat leaves, the spike and impact seed quality. Both fungi produce host specific phytotoxins that elicit distinct lesion types. The distribution and pathogenic characterization of these pathogens was determined in seven wheat growing regions of Ohio. Both pathogens were recovered from wheat flag leaves from all seven regions, but S. nodorum was most prevalent. There was an association between S. nodorum isolate aggressiveness and region of the state. Isolates from North Eastern Ohio were less aggressive than those from the remaining 6 regions, while isolates from Central North West Ohio were more aggressive. P. tritici-repentis exists in the U.S. as a population of races, with the most common being races 1 - 4. These races are characterized by the toxin(s) produced causing specific reactions on a differential set of wheat cultivars and lines. Isolates of P. tritici-repentis in Ohio were determined to be races 1, 2 and rarely race 3, representing 60%, 35% and 5% of the population, respectively. Seedling reactions of 15 wheat cultivars and one breeding line to races 1 and 2 of P. tritici-repentis indicated that some cultivars were more susceptible to race 2 ii than race 1. This same set of cultivars was inoculated with isolates of S. nodorum producing either predominately phytotoxins SnTox1 or SnTox2. Most genotypes were more sensitive to S. nodorum isolate producing predominately SnTox2. Spike and flag leaf reactions to S.nodorum of 13 wheat cultivars were examined in the greenhouse and field. Cultivar Coker 9663 was identified as having moderately high resistance to both leaf blotch and glume blotch. Seed harvested from field plots were examined for pathogen infestation. S. nodorum was the predominant pathogen on seed obtained from both inoculated and non- inoculated field plots. P. tritici-repentis and Bipolaris sorikiniana were also present on seed from non-inoculated plots. This same set of cultivars was examined for P. tritici-repentis resistance in the greenhouse and field. The resistance levels to P. tritici-repentis of the 13 cultivars varied in the field with no significant differences detected among cultivar reactions. Two greenhouse experiments examined the response of wheat cultivars to P. tritici-repentis on adult flag leaves. One experiment used 10 isolates of P. tritici-repentis obtained from Ohio and the other experiment used either a race 1 or race 2 isolate from North Dakota. Cultivar reactions were similar for these two greenhouse experiments regardless of origin of the isolate or isolate race. Cultivar reactions in the field and greenhouse experiments were correlated, and disease severities among cultivars were significantly different. Coker 9663 was more susceptible than the other cultivars tested to P. tritici-repentis. All cultivars tested were sensitive to P. tritici-repentis and S. nodorum toxins. These results indicated that S. nodorum is the predominate leaf blotching pathogen in Ohio, but P. tritici- iii repentis may be impacting leaf disease levels in the state. Moderately high resistance to S. nodorum and P. tritici-repentis was detected in several wheat cultivars indicating that breeding for resistance to both pathogens is possible. iv Dedicated to my parents. v ACKNOWLEDGMENTS Dr. Patrick Lipps not only provided the financial support for this dissertation, but scholarly guidance and support. Dr. Larry Madden gave me all of the necessary statistical analysis advice. He also lent an ear when needed. Drs. Redinbaugh, Kamoun and Sneller provided invaluable advice throughout my project. The editorial comments on my dissertation were very much appreciated. Mr. Richard J. Minyo, Jr. provided the opportunity for natural inoculum plots across Ohio. Mr. Larry Herald taught me not only how to cross wheat, but also how to deal with problems on the job. Mrs. Lynn West was helpful during work and in life. Ms. Audrey L. Johnston helped with multiple technical details. vi VITA Born: June 17, 1978 in Warren, Ohio EDUCATIONAL BACKGROUND M. S. (Plant Pathology)- The Ohio State University, June 2002 Advisor – Dr. Patrick E. Lipps Thesis Title – Evaluation and Characterization of Resistance to Fusarium graminearum in Winter Wheat B. S. (Plant Pathology)- The Ohio State University, June 1999 Minor Plant Biology PROFESSIONAL EXPERIENCE 2004-2005………....Collaborative work with Dr. Tim Freisen’s laboratory on race determination of Pyrenophora tritici-repentis and Stagonospora nodorum toxin screening. 2004-2005…………Collaborative work on a regional Stagonospora nodorum leaf blotch and glume blotch screening nursery and field work associated with that nursery. 2002 – 2005……….Graduate Research Assistant in the laboratory of Dr. Patrick E. Lipps at The Ohio State University. Duties included vii technical assistance with on-going research projects in the greenhouse and field; collection of field samples from across the state for species identification; assembly of a field disease screening nursery for breeding purposes, creating a breeding population for QTL analysis for Stagonospora nodorum resistance; hand spraying of fungicides; assistance with biological agent application; purchasing lab equipment; use of common lab equipment; attending several extension meetings dealing with field crops; writing extension bulletins; assisting in managing summer help. 2004………………..Participation in the graduate student seminar exchange program. Presented seminar at Cornell University. 1999 – 2002……….Graduate Research Assistant in the laboratory of Dr. Patrick E. Lipps at The Ohio State University. Duties included technical assistance with on-going research projects in the greenhouse and field; assembly of a field nursery for disease screening; hand spraying of fungicides; assistance with biological agent applications; purchasing lab equipment; use of common lab equipment such as balances, pH meter, microscopes, autoclaves and flow hoods; incorporating serial dilutions of endogenous wheat spike compounds into media; attending several extension meetings dealing with field crops; writing two extension bulletins; assisting in managing summer help. 2000…………………Teaching Assistant at The Ohio State University. Taught a laboratory section of PP401 Introductory Plant Pathology. viii 1998 – 1999……….Student worker in Dr. Terrance Graham’s laboratory in the department of plant pathology at The Ohio State University. Work involved maintaining Arabidopsis thaliana plants and visual evaluation of populations for mutations that allowed growth in the presence of Rose Bengal. Mutants were then analyzed using high pressure liquid chromatography (HPLC). 1997 – 1998……….Student worker in Dr. Fred Sack’s laboratory in the department of plant biology at The Ohio State University. Work involved maintaining Arabidopsis thaliana plants for genetic experiments and visual evaluation of populations for stomotal mutations. PUBLICATIONS REFERED JOURNALS 1. Engle, J. S., Lipps, P. E., Graham, T. L. and Boehm, M. J. 2003. Effects of choline, betaine and wheat floral extracts on growth of Fusarium graminearum. Plant Dis. 88:175-180. 2. Engle, J. S., Madden, L. V. and Lipps, P. E. 2003. Evaluation of Inoculation Methods to Determine Resistance Reactions of Wheat to Fusarium graminearum. Plant Dis. 87:1530-1535. ix EXTENSION PAPERS 1. Engle, J. S., De Wolf, E. D. and Lipps, P. E. 2001. A visual scale for estimating damage to soft red winter wheat kernels by Fusarium Head Blight. The Ohio State University. Ohio State University Extension. 2. Engle, J. S., Lipps, P. E. and Mills, D. 2003. Fusarium Head Blight severity visual scale of non-awned wheat. The Ohio State University. Ohio State University Extension. Fact Sheet AC-49-03. 3. Engle, J. S., Lipps, P. E. and Mills, D. 2004. Tan spot: yellow leaf spot or blotch. The Ohio State University. Ohio State University Extension. Fact Sheet:AC-50-04. 4. Engle, J. S., Lipps, P. E. and Mills, D. 2005. Spot blotch and common root rot. The Ohio State University. Ohio State University Extension. Fact Sheet:AC-51-05. FIELDS OF STUDY Major Field: Plant Pathology x TABLE OF CONTENTS Page Abstract………………………………………………………………………………...…ii Dedication………………………………………………………………………………..v Acknowledgments………………………………………………………………………vi Vita……………………………………………………………………………………….vii List of Tables………………………………………………………………………..…xvii List of Figures………………………………………………………………………….xxi Chapters: 1. Distribution and Pathogenic Characterization of Pyrenophora tritici- repentis and Stagonospora nodorum in Ohio………………………..………1 1.1 Abstract………………………………………………………………..……1 1.2 Introduction……………………………………………………………...….2 1.3 Objectives……………………………………………………………..……6 1.4 Materials and Methods…………………………………………..………..7 1.4.1 Collection of samples………………………………………...….7 1.4.2 Isolation of pathogens……………………..…………………….8 1.4.3 Single spore isolation and storage……………………….…..10 1.4.4 P. tritici-repentis race determination……………………….…11
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