Drivers of Fungal Community Composition and Function In
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DRIVERS OF FUNGAL COMMUNITY COMPOSITION AND FUNCTION IN TEMPERATE FORESTS A dissertation submitted to Kent State University in partial fulfillment of the requirements for the degree of Doctor of Philosophy By Matthew D. Gacura December 2018 © Copyright All rights reserved Except for previously published materials i Dissertation written by Matthew David Gacura B.S., Youngstown State University, 2007 M.S., Youngstown State University, 2009 Ph.D., Kent State University, 2018 Approved by Christopher B. Blackwood, Ph.D. , Chair, Doctoral Dissertation Committee Mark W. Kershner, Ph.D. , Members, Doctoral Dissertation Committee Xiaozhen Mou, Ph.D. Mandy J. Munro-Stasiuk, Ph.D. Abdul Shakoor, Ph.D. Accepted by Laura G. Leff, Ph.D. , Chair, Department of Biological Sciences James L. Blank, Ph.D. , Dean, College of Arts and Sciences ii TABLE OF CONTENTS TABLE OF CONTENTS…………………………………………….…………………………...iii LIST OF FIGURES…………………………….………………….………………………………v LIST OF TABLES……………….………………………………………………………………..x ACKNOWLEDGEMENTS……………………………………………………………………...xii I. CHAPTER 1: INTRODUCTION………………………..……………………………1 REFERENCES……………………..………………………………………………..20 II. CHAPTER 2: NICHE VS NEUTRAL: FACTORS INFLUENCING THE STRUCTURE OF SAPROTROPHIC FUNGAL COMMUNITIES AT FINE AND LARGE SPATIAL SCALES……………...…………………………………………35 ABSTRACT………………………………………………………………………….35 INTRODUCTION…………………..……………………………………………….36 MATERIALS AND METHODS…………...……………………..…………………40 RESULTS……………………..……………………………………………………..47 DISCUSSION……………..……………………………………………………..…..51 ACKNOWLEDGEMENTS………………………………………………………….60 REFERENCES……..………………………………………………………………..71 III. CHAPTER 3: THE ROLE OF PRIORITY EFFECTS IN SAPROTROPHIC FUNGAL COMMUNITIES IN TEMPERATE FORESTS………………………...……..…….84 ABSTRACT………………………………………………………………………….84 INTRODUCTION…………………..……………………………………………….85 MATERIALS AND METHODS……...………..………………………………...….89 RESULTS……………………………………………..………………………….….95 iii DISCUSSION……………………………………………..…………………...…….98 ACKNOWLEDGEMENTS……………………………………………………...…104 REFERENCES…………………………………..…………………………………117 IV. CHAPTER 4: COMPARISON OF PECTIN-DEGRADING FUNGAL COMMUNITIES IN TEMPERATE FORESTS USING GLYCOSYL HYDROLASE FAMILY 28 PECTINASE PRIMERS TARGETING ASCOMYCETE FUNGI..….130 ABSTRACT………………………………………………………………………...130 INTRODUCTION………..………………………………………………….……..131 MATERIALS AND METHODS…………..……...………………………..………133 RESULTS…………………..………………………………………………...…….137 DISCUSSION…………………..………………………………………….……….139 ACKNOWLEDGEMENTS………………………………………………………...142 REFERENCES………..……………………………………………………………149 V. CHAPTER 5: SYNTHESIS………...……………………………………….……...157 REFERENCES……………………..………………………………………………165 VI. APPENDIX I………………..……………………………...…………….………..171 APPENDIX I REFERENCES………………….………………..………………....176 iv LIST OF FIGURES Figure 1. Map of Manistee National Forest MI. Indicated on this map are important glacial landforms and other geologic structures that determine soil characteristics and ecosystem type. Nine sites were utilized in this study and their spatial locations can be observed from the figure. These sites are also color coded by ecosystem type as follows: Black = BOWO, Red = SMRO, and Green = SMBW. Map generated using ArcGIS; glacial land systems taken from Michigan Department of Natural Resources (generated April 2017)……………………………...………...18 Figure 2. Map of Manistee National Forest MI. Indicated on this are the major soil suborders found in Manistee that play a role in ecosystem type. Nine sites were utilized in this study and their spatial locations can be observed from the figure. These sites are also color coded by ecosystem type as follows: Black = BOWO, Red = SMRO, and Green = SMBW. Map generated using ArcGIS; Soil data taken from Michigan Department of Natural Resources (generated April 2017)……………………………………………………………………..………………………19 Figure 3. Map of Manistee National Forest. Included are locations for each site used, geographic features, locations of largest cities, and a scale bar indicating distances. Sites are separated by between 3 and 56 km. These sites are also color coded for which ecosystem type that they are classified as. Ecosystem classifications are as follows: Black = BOWO, Red = SMRO, and Green = SMBW. Map was generated using ArcGIS, geologic features, cities, and other structures were taken from Michigan Department of Natural Resources (generated April 2017)……………..…..61 Figure 4. Plots of average Jaccard transformed community distance (based on T-RFLP profiles) at increasing spatial distance (number of steps), for all April sites. Each point indicates an average community distance of all leaves at that distance, with the size of each point proportional to the v number of sample pairs (ranging from 1-50 pairs per step). Error bars are included for each point indicating standard error. Summarized on each plot are the site number and ecosystem. * P < 0.05…………………………………………………………………………………………….…63 Figure 5. RDA ordination plots for hellinger transformed T-RFLPs profiles in A) April and B) August. Colors represent each site, with centroids labeled with each ecosystem type: BW=BOWO, SB = SMBW and RS = SMRO. Shapes indicate different leaf species and are indicated by a key. Percent varience explained by RDAs are indicated for each season is indicated by the bar graph found in panel C…………………………………………………...…………………………..…65 Figure 6. Percent varience explained by RDAs for hellinger transformed T-RFLPs profiles for each season. Each color indicates a separate explanatory factor as indicated by side panel………66 Figure 7. Abundance of taxa found on each leaf type. Classification for each taxa is the highest reliable resolution given for an OTU. Classes of each type of fungi are given with orders found below them also included……………………………………………………………………..….67 Figure 8. Amount of variation explained by each factor for all pyrosequencing OTUs, functional groups pooled together and each functional group individually. Functional groups are abbreviated as follows: Mycop./Yeast=mycoparasite/yeast, Necrotroph=nectrophic plant pathogen, Ectomyco=ectomycorrhizal, Arbuscular=arbuscular mycorrhizal, PrimSap=primary saprotroph and Whiterot=white rot saprotroph. Designations for each factor are found on the key on the side…………………………………………...…………………………………………………..68 Figure 9. RDA ordination plot of pyrosequencing OTUs. Colors represent each site, with centroids labeled with each ecosystem type: BW=BOWO, SB = SMBW and RS = SMRO. Shape indicate leaf species as indicated by the key found on the plot………………………………………...…..69 vi Figure 10. Average percent abundances for each functional group for each leaf type. Functional groups are identified by a key on the side. Various includes all functional groups that were less 1% combined with taxa that were found to be responsible for multiple functions………............…….70 Figure 11. Map of Manistee National Forest, MI with sites and ecosystem type indicated. Each ecosystem is indicated by a separate color: green indicates SMBW, black indicates BOWO, and red indicates SMRO. An example of the experimental set up for each site is indicated. Three subsites within each site are indicated, with each separated by 500 m. Map was generated utilizing the program ArcGIS……………………………………………………...……………..………106 Figure 12. Percent variance explained (Y-axis), as indicated by Redundancy Analysis, for T-RFLP profiles at 1-Month and 5-Months in the field. The interactions category includes interactions between the factors: leaf type, isolate, and ecosystem………………………….……………….109 Figure 13. Ordination plots of T-RFLP community profiles separated by ecosystem and time point. Shape indicates litter bag leaf type (circles = oak leaves; triangle = maple leaves). Color indicates subsites found within each site (nine subsites across the three sites for each ecosystem at each time point)…………………………………………………………………………………110 Figure 14. Significant factors that influence percent mass loss as indicated by Redundancy Analysis, for each data set at 1-month and 5-months. Variance explained for each significant factor is indicated on the y axis. The interactions category includes interactions between the factors: leaf type, isolate, and ecosystem…………………………………………………………………….111 Figure 15. Average decomposition of leaf litter, as indicated by percent mass loss (y axis), for treatments at 1-month and 5-months. Treatments are noted by what ecosystem they were placed in vii and what leaf type litter bags consisted of. Isolated type is indicated by bar color and pattern. Error bars are utilized to indicate standard error…………………………………...………………….112 Figure 16. Variance explained (Y axis) for enzyme profiles per sample and each separate enzyme functional group at 1-month (A) and 5-months (B). Laccase produced at 1 month was very low or absent across all samples. The interactions category includes interactions between the factors: leaf type, isolate, and ecosystem……………………………………………...……………………..113 Figure 17. Ordination plots for enzyme profiles at 1-month and 5-months. Each ecosystem is separated by panel, with three replicate sites found on each. Shape indicates litter bag leaf type, circles indicate oak leaf bags and triangles indicate maple leaf bags. Color indicates isolate type original colonized onto litter bag with: red indicating isolate 1, blue indicating isolate 2, and black indicating isolate 3. Amount of variation explained by RDA vectors is indicated on the X and Y axis…………………………………………………………………...…………………………114 Figure 18. Extracellular enzyme activity after 1-Month. Enzyme activity has been categorized based upon enzyme function: A. phosphatase activity,