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Understanding the Microbiome of Puget Prairies: Community Composition of Bacteria in a Hemiparasitic Plant System Victoria G. J

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Understanding the Microbiome of Puget Prairies: Community Composition of Bacteria in a Hemiparasitic Plant System Victoria G. J Understanding the Microbiome of Puget Prairies: Community composition of bacteria in a hemiparasitic plant system Victoria G. J. Fox A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science University of Washington 2020 Committee: Jonathan D. Bakker Sharon L. Doty Program Authorized to Offer Degree: Environmental and Forest Sciences © Copyright 2020 Victoria G. J. Fox University of Washington Abstract Understanding the microbiome of Puget prairies: community composition of bacteria in a hemiparasitic plant system Victoria G. J. Fox Chair of the Supervisory Committee: Professor Jonathan D. Bakker School of Environmental and Forest Sciences Recent advances in the field of metagenomics have allowed for a boom of research in the field of microbial community ecology. Using DNA extraction techniques, Illumina sequencing, and advanced statistical software, scientists are now able to examine the community composition of microbiomes existing throughout the world. My research examines the microbial communities of Puget prairie plants, which have remained largely unexplored until now. I performed a field study to identify the bacterial communities that comprise the stem microbiomes of 16 native prairie plant species. I discovered that the bacterial communities within Puget prairie plants often differ significantly between plant species, but plant species belonging to the same family often have similar bacterial communities. Additionally, I discovered that bacterial communities differed between samples taken from different sampling locations. I also found that bacterial communities are only affected by disturbances applied several years prior to sampling, and in disturbance regimes applied continuously to research plots, for Cerastium arvense. I explored the theory that the bacterial community within Puget prairie plants could be influenced by parasitic root connections established by Castilleja levisecta, a hemiparasitic plant that attaches root connections to other prairie plants. Testing all samples that could be assigned to trios regardless of species sample size, I found that plant parasitism significantly affects the bacterial communities of Puget prairie plants overall. Testing individual species with large sample sizes, I found an effect of plant parasitism on the microbiomes of parasitic plant C. levisecta for Eriophyllum lanatum and Lomatium utriculatum, and further study of this system with larger sample sizes could reveal an effect of parasitism for Balsamorhiza deltoidea and Festuca roemeri. This research provides valuable information about the types of bacteria that exist within the stem tissues of native Puget prairie plants, and insights into the role that parasitic plants may play in the colonization of bacteria across the Puget prairie ecosystem. 1 Table of Contents List of Abbreviations ...................................................................................................................... 3 List of Figures ................................................................................................................................. 4 List of Tables .................................................................................................................................. 5 Acknowledgements ......................................................................................................................... 7 Introduction ..................................................................................................................................... 8 Literature Cited ......................................................................................................................... 15 Chapter 1: Bacterial Composition of Puget Prairie Plants ............................................................ 18 Abstract ..................................................................................................................................... 18 Introduction ............................................................................................................................... 20 Methods..................................................................................................................................... 26 Results ....................................................................................................................................... 46 Differences between Plant Species ....................................................................................... 49 Effect of Site Location: Glacial Heritage Preserve and Smith Prairie .................................. 55 Initial Restoration Treatments............................................................................................... 58 Disturbance Regime .............................................................................................................. 62 Discussion ................................................................................................................................. 66 Literature Cited ......................................................................................................................... 77 Appendix 1.A: ....................................................................................................................... 84 Appendix 1.B: ....................................................................................................................... 86 Appendix 1.C: ....................................................................................................................... 87 Appendix 1.D: ....................................................................................................................... 88 Appendix 1.E: ....................................................................................................................... 89 Appendix 1.F: ....................................................................................................................... 93 Appendix 1.G: ....................................................................................................................... 94 Chapter 2: Transfer of Bacteria between Castilleja Levisecta and Host Plants .......................... 97 Abstract ..................................................................................................................................... 97 Introduction ............................................................................................................................... 99 Methods................................................................................................................................... 104 Results ..................................................................................................................................... 115 Discussion ............................................................................................................................... 121 Literature Cited ....................................................................................................................... 129 2 Appendix 2 .......................................................................................................................... 132 Conclusion .................................................................................................................................. 135 Appendix 3 .......................................................................................................................... 138 3 List of Abbreviations Abbreviation Explanation GHP............................................. Glacial Heritage Preserve ID................................................. Identity NMDS.......................................... Non-Metric Multidimensional Scaling OTU.............................................. Operational Taxonomic Unit PERMANOVA............................. Permutational Multivariate Analysis of Variance SM................................................ Smith Prairie UFWS........................................... U.S. Fish and Wildlife Service UMGC.......................................... University of Minnesota Genomics Center UW............................................... University of Washington 4 List of Figures Chapter 1 Figure 1.1: Map of Washington State Counties.................................................................26 Figure 1.2: Map of Glacial Heritage Preserve research plots............................................27 Figure 1.3: Map of Smith Prairie research plots................................................................28 Figure 1.4: Box and whisker plot of the number of bacterial OTUs derived from each species, after removal of potentially contaminating bacterial OTUs.................................47 Figure 1.5: Average abundance of bacterial OTU reads representing each bacterial phyla within a plant species.........................................................................................................49 Figure 1.6: Three-dimensional NMDS ordination of bacterial OTU abundance with plant species overlay...................................................................................................................53 Figure 1.7: Three-dimensional NMDS ordination of bacterial OTU abundance with plant family overlay....................................................................................................................54 Figure 1.8: Two-dimensional NMDS ordination of bacterial OTU abundance of Achillea millefolium,
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