University of Tennessee, Knoxville TRACE: Tennessee Research and Creative Exchange Masters Theses Graduate School 12-2015 Environmental Controls on the Diversity and Distribution of Endosymbionts Associated with Phacoides pectinatus (Bivalvia: Lucinidae) from Shallow Mangrove and Seagrass Sediments, St. Lucie County, Florida Thomas Walters Doty University of Tennessee - Knoxville, [email protected] Follow this and additional works at: https://trace.tennessee.edu/utk_gradthes Part of the Biodiversity Commons, Biogeochemistry Commons, Geochemistry Commons, Marine Biology Commons, Molecular Biology Commons, and the Terrestrial and Aquatic Ecology Commons Recommended Citation Doty, Thomas Walters, "Environmental Controls on the Diversity and Distribution of Endosymbionts Associated with Phacoides pectinatus (Bivalvia: Lucinidae) from Shallow Mangrove and Seagrass Sediments, St. Lucie County, Florida. " Master's Thesis, University of Tennessee, 2015. https://trace.tennessee.edu/utk_gradthes/3548 This Thesis is brought to you for free and open access by the Graduate School at TRACE: Tennessee Research and Creative Exchange. It has been accepted for inclusion in Masters Theses by an authorized administrator of TRACE: Tennessee Research and Creative Exchange. For more information, please contact [email protected]. To the Graduate Council: I am submitting herewith a thesis written by Thomas Walters Doty entitled "Environmental Controls on the Diversity and Distribution of Endosymbionts Associated with Phacoides pectinatus (Bivalvia: Lucinidae) from Shallow Mangrove and Seagrass Sediments, St. Lucie County, Florida." I have examined the final electronic copy of this thesis for form and content and recommend that it be accepted in partial fulfillment of the equirr ements for the degree of Master of Science, with a major in Geology. Annette Engel, Major Professor We have read this thesis and recommend its acceptance: Michael McKinney, Andrew Steen Accepted for the Council: Carolyn R. Hodges Vice Provost and Dean of the Graduate School (Original signatures are on file with official studentecor r ds.) Environmental Controls on the Diversity and Distribution of Endosymbionts Associated with Phacoides pectinatus (Bivalvia: Lucinidae) from Shallow Mangrove and Seagrass Sediments, St. Lucie County, Florida A Thesis Presented for the Master of Science Degree The University of Tennessee, Knoxville Thomas Walters Doty December 2015 Acknowledgments I would like to start by thanking the National Science Foundation and Department of Earth and Planetary Sciences for their finical support. I would also like to sincerely thank the staff throughout The University of Tennessee that continuously supported and assisted me. Dr. Annette Engel, thank you for encouraging me to enter and continue through with graduate school. I have undergone quite a transformation as a scientist, and certainly owe much of that to your dedication and efforts. I would also like to thank my committee members, Mike McKinney and Drew Steen. Furthermore, I greatly appreciate the assistance and support of Kathleen Brannen-Donnelly, Terri Brown, Caroline Dietz, Chanda Drennen, Aaron Goemann, Brendan Headd, Hunter Johnson, Sarah Keenan, Audrey Paterson, and the other friends and individuals that have helped me along the way. I appreciate Laurie Anderson, Brooke Long, Jean Lim, and Barbara Campbell for their time and efforts while completing field work in Florida. Finally, Mom and Dad, thank you so much for making sure that my education was always a priority. It’s been a long journey, and I could not have done it without you. Beth and John, you are both amazing students and individuals, and I am always amazed and inspired at the way you work and live. ii Abstract Lucinid bivalves are capable of colonizing traditionally inhospitable shallow marine sediments due to the chemosynthetic metabolic functions of bacterial endosymbionts located within their gills. Because lucinids can often be the dominant sediment infauna species, defining their abundances and roles in sediment and pore fluid biogeochemical cycling is necessary to understand coastal biological diversity and the sensitivity of threatened coastal ecosystems over time. However, there has been limited research done to evaluate the diversity and distribution of many lucinid chemosymbiotic systems. The goals of this study were to access the distribution of the lucinid Phacoides pectinatus and its endosymbiont communities from Ft. Pierce, St. Lucie County, Florida, and to define the environmental controls on potential free-living P. pectinatus endosymbionts to understand symbiont distribution patterns and host acquisition. Nearly all of the encountered P. pectinatus hosts were confined to within one meter of the mangrove canopy. P. pectinatus distribution correlated to concentrations of organic carbon in the sediments, but not to total dissolved sulfide concentrations or sea grass vegetation densities. Sequencing of bacterial 16S rRNA genes from sediment and pore fluids from the P. pectinatus habitat revealed that Proteobacteria dominated the communities, including Alpha-, Delta-, and Gammaproteobacteria classes. Other major phyla included the Bacteroidetes, Chloroflexi, Planctomycetes, and Spirochaetae. 16S rRNA genes obtained from P. pectinatus gills and feet were closely related to novel bacterial communities comprised of Sedimenticola, Kistimonas, Methylomarinum, and Spirochaeta spp., unclassified Rickettsiales (Alphaproteobacteria) and unclassified Lentisphaerae. This level of potential endosymbiont diversity has not been previously reported for lucinids. Moreover, potential endosymbiont populations differed by lucinid location, as gills iii containing higher Rickettsiales abundances occurred in areas with the lowest clam density and hosts from deeper sediments contained a higher proportion of Methylomarinum. Despite broad sediment and pore water bacterial diversity, no genetically similar sequences that matched those from P. pectinatus hosts were retrieved from the environment. These results, while strengthening our understanding of a lucinid-symbiont system, still highlight how limited our knowledge is about these systems. These results provide new directions for future research and can be used to help understand how sensitive these systems are to environmental change. iv Table of Contents CHAPTER ONE: INTRODUCTION ................................................................................. 1 Background ..................................................................................................................... 2 Lucinid Symbiont Acquisition, Identity, and Ecology ................................................... 4 Phacoides pectinatus ...................................................................................................... 7 Objectives and Hypothesis .............................................................................................. 7 CHAPTER TWO: MATERIALS AND METHODS ......................................................... 9 Study Site and Sample Collection................................................................................... 9 Oceanographic and Meteorological Data ...................................................................... 13 DNA Extraction, Amplicon Pyrosequencing, and Analysis ......................................... 14 Statistical Analyses ....................................................................................................... 16 CHAPTER THREE: RESULTS ....................................................................................... 18 Aqueous and Sediment Geochemistry .......................................................................... 18 Fauna and Vegetation of Wildcat Cove ........................................................................ 19 Lucinid Gill and Foot Bacterial Diversity .................................................................... 21 Bacterial Populations in Environmental Water and Sediment ...................................... 24 CHAPTER FOUR: DISCUSSION ................................................................................... 27 CHAPTER FIVE: CONCLUSION................................................................................... 34 REFERENCES ................................................................................................................. 36 APPENDIX ....................................................................................................................... 48 VITA ............................................................................................................................... 141 v List of Tables Table 1 - Field physiochemical parameters from Wildcat Cove, St. Lucie County, Florida ........50 Table 2 - Cation and anion concentrations (mmol/L) at Wildcat Cove, St. Lucie County, Florida ....................................................................................................................................51 Table 3 - Dissolved gases measured by gas chromatography and fluorescence spectroscopy indices for water for Wildcat Cove, St. Lucie County, Florida. ............................................52 Table 4 - Average percent organic carbon based on loss-on-ignition (LOI) measurements and P. pectinatus abundancesa (in parentheses) from Wildcat Cove sediments ....................53 Table 5 - Organic carbon content measured by Loss on Ignition (LOI) ......................................54 Table 6 - Summary of 454 tag pyrosequencing