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Hoffman Dissertation.Pdf Determining the spatial and seasonal influences of microbial community composition and structure from the Hawaiian anchialine ecosystem by Stephanie Kimie Hoffman A dissertation submitted to the Graduate Faculty of Auburn University in partial fulfillment of the requirements for the Degree of Doctor of Philosophy Auburn, Alabama December 10, 2016 Keywords: anchialine, microbial ecology, Hawaii, laminated mat, spatial variation, seasonal variation Copyright 2016 by Stephanie Kimie Hoffman Approved by Scott R Santos, Chair, Professor of Biological Sciences Mark R Liles, Professor of Biological Sciences Todd D Steury, Associate Professor of Wildlife Sciences Robert S Boyd, Professor and Undergraduate Program Officer of Biological Sciences Abstract Characterized as coastal bodies of water lacking surface connections to the ocean but with subterranean connections to the ocean and groundwater, habitats belonging to the anchialine ecosystem occur worldwide in primarily tropical latitudes. Such habitats contain tidally fluctuating complex physical and chemical clines and great species richness and endemism. The Hawaiian Archipelago hosts the greatest concentration of anchialine habitats globally, and while the endemic atyid shrimp and keystone grazer Halocaridina rubra has been studied, little work has been conducted on the microbial communities forming the basis of this ecosystem’s food web. Thus, this dissertation seeks to fill the knowledge gap regarding the endemic microbial communities in the Hawaiian anchialine ecosystem, particularly regarding spatial and seasonal influences on community diversity, composition, and structure. Briefly, Chapter 1 introduces the anchialine ecosystem and specific aims of this dissertation. In Chapter 2, environmental factors driving diversity and spatial variation among Hawaiian anchialine microbial communities are explored. Specifically, each sampled habitat was influenced by a unique combination of environmental factors that correlated with correspondingly unique microbial communities. Notably, salinity was the one water chemistry factor with strong explanatory power and influence in driving microbial community structure. Chapter 3 examines seasonality in these Hawaiian anchialine microbial communities across an 18-month period. Although there was evidence that microbial community structure varied across the wet and dry seasons, these changes were minimal overall and the greatest shifts were in relative abundance of oxygenic and anoxygenic phototrophs, with oxygenic phototrophs more abundant during wet seasons and anoxygenic phototrophs during dry seasons. The specific microbial consortia found in the four distinct layers composing the unique orange laminated cyanobacterial- bacterial crusts from select Hawaiian anchialine habitats are discussed in Chapter 4. As with laminated ii microbial mats from other ecosystems, greater taxonomic richness within the community occurred deeper within the crust structure, with these crusts apparently, and unusually, oxygenated at both their top and bottom surfaces. Therefore, oxygenic phototrophs were most abundant in the top and bottom of the crusts, with anaerobic metabolisms largely confined to the middle two layers. Finally, Chapter 5 discusses the conclusions of the preceding chapters and future research directions. iii Acknowledgments I owe much to my dissertation committee in supporting and guiding me through this research: Dr. Scott Santos for being willing to take on a deer population ecologist for a microbial project and encouraging me to prepare myself for the future I want, Dr. Mark Liles for help with all thing microbial, Dr. Todd Steury for help with all things statistical, and Dr. Bob Boyd for endless encouragement and cheerleading. I also must thank Dr. Alan Wilson for being willing to serve as my University Reader and providing valuable feedback. I could not have completed this research without help from innumerable other people, including Dr. Pamela Brannock and Dr. Molli Newman for guidance and manuscript editing, Dr. Justin Havird and Dr. David Weese for collecting samples, Kiley Seitz for doing essentially all the sample processing and collecting samples, Damien Waits for scripting help, and Katie Kim for collecting samples. Many thanks also the past and present members of the Molette Lab and the numerous other graduate students with whom my path crossed. I also must thank Bonnie Wilson, the Women in Science and Engineering Institute, and Auburn University’s chapter of Graduate Women in Science for endless encouragement, advice, support, and free meals. I am grateful for the funding support for this work, which came from the National Science Foundation (DEB #0949855 awarded to Dr. Scott Santos). This dissertation could not have come to be without the love and support of my family. There aren’t enough words or pages to express how much they did so I could achieve this. I must especially thank my husband who has tolerated my science-talk, stress, constant preoccupation, and freak-outs. I’m done! iv Table of Contents Abstract ...................................................................................................................................................... ii Acknowledgments ..................................................................................................................................... iv List of Tables ............................................................................................................................................ ix List of Figures ............................................................................................................................................. x Chapter 1. Dissertation Introduction .......................................................................................................... 1 1.1 Introduction to the anchialine ecosystem ................................................................................ 1 1.2 Environmental factors driving spatial variation and diversity among Bacteria and micro- Eukarya communities of the Hawaiian anchialine ecosystem ....................................................... 5 1.3 Seasonal stability in Hawaiian anchialine microbial communities across an 18-month period ............................................................................................................................................. 7 1.4 Comparison of microbial consortia composition in the layers of laminated cyanobacterial- bacterial mats found in select Hawaiian anchialine habitats ........................................................ 10 1.5 References ............................................................................................................................. 12 Chapter 2. Environmental factors driving spatial variation and diversity among Bacteria and micro- Eukarya communities of the Hawaiian anchialine ecosystem ..................................................... 18 2.1 Abstract ................................................................................................................................. 18 2.2 Introduction ........................................................................................................................... 19 2.3 Materials and Methods .......................................................................................................... 21 2.3.1 Sites and Sampling ................................................................................................ 21 2.3.2 Sequence Data Generation .................................................................................... 22 2.3.3 Operational Taxonomic Unit (OTU) Clustering ................................................... 22 2.3.4 Analyses of Community Composition and Environmental Factors ...................... 23 2.3.5 Data Accessibility ................................................................................................. 24 v 2.4 Results ................................................................................................................................... 25 2.4.1 Sites and Sampling ................................................................................................ 25 2.4.2 Sequence Data Generation and OTU Clustering .................................................. 25 2.4.3 Analyses of Community Composition and Environmental Factors ...................... 26 2.5 Discussion ............................................................................................................................. 29 2.5.1 Microbial Diversity of the Hawaiian Anchialine Ecosystem ................................ 29 2.5.2 Environmental Drivers of Microbial Diversity in the Hawaiian Anchialine Ecosystem ....................................................................................................................... 31 2.5.3 Considerations for Conservation Efforts of Hawaiian Anchialine Habitats ......... 32 2.5.4 Conclusions ........................................................................................................... 34 2.6 Acknowledgments................................................................................................................. 34 2.7 References ............................................................................................................................. 35 Chapter 3. Seasonal stability in Hawaiian anchialine microbial communities across an 18-month period ..........................................................................................................................................
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