INFECTIVITY OF VERTICILLIUM DAHLIAE ISOLATES ON WEEDY HOSTS, LITCHI TOMATO, AND TEFF, AND THE EFFECT OF ALFALFA RESIDUE INCORPORATION ON THE NUMBER OF VERTICILLIUM DAHLIAE MICROSCLEROTIA, AND SOIL BACTERIAL METAGENOMICS By ZACHARY ANDREW FREDERICK A dissertation submitted in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY WASHINGTON STATE UNIVERSITY Department of Plant Pathology MAY 2017 © Copyright by ZACHARY ANDREW FREDERICK, 2017 All Rights Reserved © Copyright by ZACHARY ANDREW FREDERICK, 2017 All Rights Reserved To the Faculty of Washington State University: The members of the Committee appointed to examine the dissertation of ZACHARY ANDREW FREDERICK find it satisfactory and recommend that it be accepted. ___________________________________ Dennis A. Johnson, Ph.D, Chair. ___________________________________ Mark J. Pavek, Ph.D. ___________________________________ Debra A. Inglis, Ph.D. ___________________________________ Weidong Chen, Ph.D. ii ACKNOWLEDGMENTS I thank Dr. Dennis A. Johnson for the opportunity to pursue the study of plant pathology, cooperative extension, and potato disease at Washington State University through his program. I also thank Thomas F. Cummings for instruction and support of establishing trials, as well as guidance on statistical analyses. I wish to thank my committee members, Drs. Mark J. Pavek, Debra A. Inglis, and Weidong Chen for their critiques and guidance. I am grateful for my present and former members of my laboratory workgroup, including David Wheeler and Dr. Lydia Tymon for direction and toleration of my contributions to entropy, as well as Dr. Jeremiah Dung for his isolates and copious notes left behind. Would you kindly join me in extending special thanks to Dr. Kerik Cox, who continues to serve as an additional adviser. Last, but certainly not least, I give thanks for family and my fiancée for reminding me that there is always a man, always a lighthouse, and always a city. iii INFECTIVITY OF VERTICILLIUM DAHLIAE ISOLATES ON WEEDY HOSTS, LITCHI TOMATO, AND TEFF, AND THE EFFECT OF ALFALFA RESIDUE INCORPORATION ON THE NUMBER OF VERTICILLIUM DAHLIAE MICROSCLEROTIA, AND SOIL BACTERIAL METAGENOMICS Abstract by Zachary Andrew Frederick, Ph.D. Washington State University May 2017 Chair: Dennis A. Johnson Verticillium wilt, caused by Verticillium dahliae, is an important disease of many dicotyledonous crops due to a wide host range and the long-term survival of microsclerotia in soil for up to 14 years. Some V. dahliae isolates are aggressive on a specific plant host, such as potato, but can still infect a range of crops. Isolates of V. dahliae that are aggressive on potato are referred to as the potato pathotype. Litchi tomato (Solanum sisymbriifolium) has been grown as a trap crop for the pale cyst nematode in Idaho and teff (Eragrostis tef) could be a short- season rotation crop in the northwestern United States. It is unknown if litchi tomato, teff, or weeds could serve as sources of inoculum for the potato pathotype of V. dahliae. When sixteen weeds were evaluated for V. dahliae, black nightshade (Solanum nigrum) had significantly more microsclerotia of the V. dahliae potato pathotype compared to the other isolates in three of four greenhouse trials (second trial P < 0.0158, third trial P < 0.0264, fourth trial P < 0.0193). There were no differences in numbers of microsclerotia between isolates of V. dahliae in infected teff, iv while on litchi tomato the potato pathotype of V. dahliae produced greater numbers of microsclerotia than other isolates in one of six trials (first trial FDR-adjusted P < 0.0149). Soil incorporation of alfalfa residues prior to planting potato could be a Verticillium wilt management strategy by reducing the number of viable microsclerotia in field soil. The impact of incorporating alfalfa residue on soil metagenomics is unknown. The number of V. dahliae microsclerotia in soil where alfalfa was incorporated was greater than when residue was not incorporated (P = 0.0003) when field soils were subject to soil fumigation with chloropicrin. The soil bacterial metagenome in field soils subjected to alfalfa residue incorporation did not differ from those not subject to residue incorporation after one year. Alfalfa residue incorporation alone did not reduce V. dahliae microsclerotia in the soil or impact soil metagenomics; the practice by itself was not an effective strategy to induce a disease-suppressive soil on short rotations. v TABLE OF CONTENTS Page ACKNOWLEDGMENTS…………………..…………………………………………………....iii ABSTRACT ………………………………………………………………………….…….….iv-v LIST OF TABLES ………….……….…………………………………………...….….…....ix-xii LIST OF FIGURES …………………..………………….…………………………....………..xiii CHAPTER ONE: Introduction........................................................................................................1 1. Solanaceae and Solanum origin…………………...…………………….......………….1 2. How is potato production important to Washington State? ………………………........3 3. Verticillium – taxonomy, symptoms, and life cycle………………………….………...4 4. Verticillium wilt management…………...……………………...….…………………11 5. Weedy hosts as sources of Verticillium dahliae inoculum……………………………15 6. Black dot – problem, symptoms, and biology…………..………………............…….16 7. Black dot – management………………………...……………………...........……......18 8. Research Objectives.......................................................................................................19 9. Literature cited…………………………………...………………..………………......23 CHAPTER TWO: Evaluation of Solanum sisymbriifolium as a Potential Inoculum Source of Verticillium dahliae and Colletotrichum coccodes………………………………………………39 1. ABSTRACT……………….………………...………………………………………...39 2. INTRODUCTION ………………………………....……………………………........40 3. MATERIALS AND METHODS……………………………………………………...43 4. RESULTS………………………………………………………………………..........49 5. DISCUSSION…………………………………………………………………………51 vi 6. ACKNOWLEDGMENTS..…………………………………………………………...55 7. LITERATURE CITED…………………………...……………............……………...57 8. TABLES………………………………………………………………………………61 CHAPTER THREE: Susceptibility of Weedy Hosts from Pacific Northwest Potato Production Systems to Crop-Aggressive Isolates of Verticillium dahliae………….......………...………….68 1. ABSTRACT…………………………………………………………………………...68 2. INTRODUCTION.………………………………………………………………........69 3. MATERIALS AND METHODS……………………………………………………...72 4. RESULTS…………………………………………………………………………......77 5. DISCUSSION…………………………………………………………………………81 6. ACKNOWLEDGEMENTS…………………………………………………………...83 7. LITERATURE CITED…………………………………….......................…………...86 8. TABLES……………………………....………………………………………………90 9. FIGURE………………………………….………………………………………........99 CHAPTER FOUR: The Effect of Alfalfa Residue Incorporation on Soil Bacterial Communities and the Quantity of Verticillium dahliae Microsclerotia in Potato Fields in the Columbia Basin of Washington State, USA...............................................................................................................103 1. ABSTRACT……………………………………………………………………….....103 2. INTRODUCTION ………………………………………………………………......104 3. MATERIALS AND METHODS…………………………………………………….108 4. RESULTS……………………………………………………...………………….....116 5. DISCUSSION………………………………………………………………………..120 6. ACKNOWLEDGMENTS..………………………………………………………….122 vii 7. LITERATURE CITED…………………………………….......................………….123 8. TABLES…………………………………………………………………………..…128 CHAPTER FIVE: The Low Potential of Teff (Eragrostis tef) as an Inoculum Source for Verticillium dahliae.....................................................................................................................142 1. ABSTRACT……………………………………………………………………….....142 2. INTRODUCTION ………………………………………………………………......143 3. MATERIALS AND METHODS…………………………………………………….146 4. RESULTS……………………………………………………...………………….....149 5. DISCUSSION………………………………………………………………………..151 6. ACKNOWLEDGMENTS.………………….……………………………………….152 7. LITERATURE CITED…………………………………….......................………….153 8. TABLE……………………………...……………………………………………..…157 9. FIGURES………………………………………………………………..……….......158 CHAPTER SIX: Conclusions......................................................................................................160 1. LITERATURE CITED................................................................................................165 viii LIST OF TABLES 1. Table 1. Verticillium dahliae and Colletotrichum coccodes isolates used to confirm host status of litchi tomato (Solanum sisymbriifolium)..........................................................………….……61 2. Table 2. Verticillium dahliae potato pathotype microsclerotia counts (CFU/g) from eggplant, potato, or litchi tomato (Solanum sisymbriifolium) stems at senescence by initial V. dahliae colony forming units (CFU)/g potting medium………………………………………………….62 3. Table 3. Mean number of Verticillium dahliae microsclerotia from stems of three potato cultivars Alturas, Russet Norkotah, and Ranger Russet, and litchi tomato in a greenhouse experiment in 2013………………………………………………………………………………63 4. Table 4. Mean number of Verticillium dahliae CFU from stems and roots of three potato cultivars Alturas, Russet Norkotah Ranger Russet, and litchi tomato (Solanum sisymbriifolium) in a greenhouse in 2014……………………………………...……………………………….64-65 5. Table 5: Mean number of Colletotrichum coccodes CFU from stems and roots of three potato cultivars Alturas, Russet Norkotah Ranger Russet, and litchi tomato (Solanum sisymbriifolium) in a greenhouse in 2013 and 2014……………………………………………….…........……….66 ix 6. Table 6: Mean number of Verticillium