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UNIVERSITY of CALIFORNIA RIVERSIDE The UNIVERSITY OF CALIFORNIA RIVERSIDE The Geographic Distribution, Genetic Structure and Cultural Management of Waitea circinata var. circinata, the Causal Agent of Brown Ring Patch Isolated from Turf Grass in the U.S. A Dissertation submitted in partial satisfaction of the requirements for the degree of Doctor of Philosophy in Plant Pathology by Chi-Min Chen August 2011 Dissertation Committee: Dr. Greg W. Douhan, Chairperson Dr. Frank P. Wong Dr. Michael E. Stanghellini Copyright by Chi-Min Chen 2011 The Dissertation of Chi-Min Chen is approved: __________________________________________________________ __________________________________________________________ __________________________________________________________ Committee Chairperson University of California, Riverside Acknowledgements First and foremost I want to thank my advisor Prof. Frank Wong. It has been an honor to be his Ph.D. student. He has always encouraged me and has taught me, both consciously and un-consciously, how a research should be done. I appreciate all his contributions of time, ideas, and funding to make my Ph.D. experience productive, diverse and stimulating. And I will always remember all of those drawings after discussion, those extreme early field trips and the most important question of the day. Special thanks are due to my co-advisor and the chair of my dissertation committee, Prof. Greg Douhan, who not only supported me with resources, but he encouraged and challenged me through my academic program. I appreciate all his time and teaching which lead my successfully solve the problems I had during the experiments. I also wish to express my appreciation to Prof. Michael Stanghellini for his support through the program and as my oral defense and dissertation committee. I would also like to thank the other four members of my oral defense committee, Prof. Deborah M. Mathews, Prof. Michael D. Coffey, Prof. Wenbo Ma, and Prof. Philip Roberts, for their time and insightful questions. The generous support from Dr. Larry Stowell is greatly appreciated. I appreciate his contribution of time and ideas on my PhD research. Special thanks to Wayne carpenter, Candice Combs, John Maddern, and Mark Woodward from Torrey Pines Municipal Golf Course and Adam Kloster from Industry Hills Golf Club for their help and assistance in conducting this research. iv I want to thank all my labmates at the turf disease diagnostic laboratory. In particular, I would like to thank Juanita Corza Rios for her friendship and help in the past five years. Special thanks to Dr. Naveen hyder on behalf of devoting her precious time and made many valuable suggestions which indeed helped improve of this thesis. Thanks to Karla de la Cerda for sharing her passion and all of her knowledge of Wcc with me. I want to thank Erica Serna, Arjun Avila, Roni Garlington, and Jim Huang, I treasured all precious moments we shared and would really like to thank them. In my later work of AFLP and population study, I thank Dr. Deborah Pagliaccia for helpful discussion and her friendship. I gratefully acknowledge the funding sources that made my Ph.D. work possible. I was funded by the Department of Plant Pathology fellowship for my first 2 years. My work was also supported by the GCSAA Environmental Institute for Golf, California State GCSA, Southern and Northern California, San Diego, Hi-Lo, and Sierra Nevada GCSAs. My time at UC Riverside was made enjoyable in large part due to the many friends and groups that became a part of my life. I am grateful for time spent with roommates, Cindy Chou and Szu-Ching Chang, and my starbucks friends and our memorable trips into the national parks, and for many other people and memories. Lastly, I would like to thank my family for all their love and encouragement. For my parents who raised me with a love of nature and supported me in all my pursuits. For all of the encouragement and support from my brother and my in-laws. And most of all v for my loving, supportive, encouraging, and patient husband Ni-Chun whose faithful support during the final stages of this Ph.D. is so appreciated. Thank you. Chi-Min Chen University of California, Riverside August 2011 vi ABSTRACT OF THE DISSERTATION The Geographic Distribution, Genetic Structure and Cultural Management of Waitea circinata var. circinata, the Causal Agent of Brown Ring Patch of Turf Grass in the U.S. by Chi-Min Chen Doctor of Philosophy, Graduate Program in Plant Pathology University of California, Riverside, August 2011 Dr. Greg W. Douhan, Chairperson Waitea circinata var. circinata (Wcc), the causal agent of brown ring patch, is an emergent disease of turfgrass in the U. S. It causes serious problems on golf course putting greens by disrupting the uniformity and aesthetic value. Based on morphological and molecular data, we identified that Wcc has a broad host range and is present in multiple states across the U.S. Besides annual bluegrass (Poa annua L.), Wcc was also found as a pathogen on rough bluegrass (Poa trivialis L.) and creeping bentgrass (Agrostis palustris). A new variety, W. circinata var. prodigus, was also reported in 2011 causing basal leaf blight of kikuyugrass in southern California. To demonstrate the distribution of Wcc in the U.S., ribosomal DNA internal transcribed spacer regions and 5.8S region (ITS) of 42 Wcc isolates collected from annual bluegrass obtained from nine States were sequenced. A total of 17 ITS haplotypes were observed and there was no obvious relationship between ITS haplotype and the geographic distribution of the isolates. This preliminary data showed that Wcc in the U.S. may be genetic diverse. To vii further demonstrate the genetic variation of Wcc and examine how Wcc exchange genes, a 3 year study was conducted at a golf course in San Diego, California. High genotypic diversity and few clonal genotypes were found among the 116 isolates collected from two putting greens. Results suggest that the pathogen may be sexually reproducing in the field and that inbreeding could account for the gametic disequilibrium detected. Before the above studies had been conducted, very little information about the epidemiology of brown ring patch was known. The disease was confused with another turf disease called yellow patch that produced similar yellow rings on greens. Attempts to manage brown ring patch similar to yellow patch makes the disease more severe. There was also concern that using trinexapac-ethyl (TE), a commonly used plant growth regulator to control turfgrass height, might be increasing disease severity. Our study showed that three nitrogen sources (nitrate, ammonium and urea) decreased brown ring patch severity. No difference in disease severity was found between treatments containing TE and the control plots that received no treatment. Azoxystrobin plus nitrogen or azoxystrobin plus nitrogen plus TE showed significant effect in alleviating brown ring patch severity in this study. To accurately evaluate disease severity of brown ring patch, digital image analysis (DIA) was compared to visual rating. Linear regression analysis showed that a bias existed between different raters regardless of the rater experience. Overall DIA and visual rating showed similar disease trends and cameras from digital camera to digital single- lens reflex cameras yielded similar results. This study showed that Wcc is a pathogen with high genetic diversity and the sexual stage may play an important role in epidemiology. Brown ring patch could be detected without using expensive equipment viii and this technique could help to develop a model for predicting this important turfgrass disease. Although similar to other turf grass diseases, this disease needs to be managed differently. ix Table of Contents Acknowledgements………………………………………………………………………iv Abstract…………………………………………………………………………………..vii List of tables…………………………………………..……………………….……..… xii List of figures…………………………….…………….………………………………...xv Chapter 1: Geographic Distribution and ribosomal DNA internal transcribed spacer (rDNA-ITS) Region Sequence Diversity of Waitea circinata var. circinata Isolated from Annual Bluegrass in the U.S .................................................................................1 Introduction ....................................................................................................................2 Materials and Methods ...................................................................................................4 Results ............................................................................................................................7 Discussion ......................................................................................................................9 Literature Cited ............................................................................................................14 Chapter 2: Response of Putting Greens Infected by Brown Ring Patch to Nitrogen, Trinexapac –Ethyl, and Azoxystrobin .........................................................................24 Introduction ..................................................................................................................25 Materials and Methods .................................................................................................28 Results ..........................................................................................................................30 Discussion ....................................................................................................................34
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