A STUDY OF METHANE-RELATED PROCESSES IN FRESHWATER ECOSYSTEMS: METHANOGENESIS, ANAEROBIC METHANE OXIDATION, AND INTERACTIONS WITH OTHER TERMINAL METABOLISMS by KATHERINE EOWYN SEGARRA (Under the Direction of Samantha B. Joye) ABSTRACT This dissertation seeks to understand the seasonal controls of methane cycling in freshwater sediments. Using a combination of field measurements, radiotracer incubations, porewater characterization, lipid biomarker analysis, and stable carbon isotopes, pronounced seasonal variations in microbial carbon turnover were documented in a freshwater sediment and in two peat wetlands. Constraints of the methane budget in shallow (< 40 cm) sediments revealed a seasonal imbalance between methane fluxes and methane production that may be relieved through tidal pumping of methane-laden porewaters derived from adjacent high marsh through the creekbank. Rate measurements of sulfate reduction and the anaerobic oxidation of methane (AOM), two processes not typically considered relevant in low salinity habitats, revealed their importance in freshwater settings. Seasonal variations in AOM may be driven by fluctuations in hydrogen and acetate dynamics generated by variations in other microbial metabolisms (e.g. sulfate reduction and methanogenesis). Lipid biomarker analysis revealed the presence of sulfate-reducing bacteria and archaea associated with methane cycling. However, seasonal variations in microbial metabolisms were not associated with changes in the lipid distribution. Stable carbon isotope analyses revealed the imprint of AOM on the signatures of methane and dissolved inorganic carbon. The influence of methanotrophy, however, was not as pronounced in the microbial lipid signatures. A potential AOM isotopic signal may have been diluted by methanogenesis and other autotrophic and heterotrophic processes, which may mask a clear methanotrophic signature. While sulfate reduction activity is sufficient to support all observed AOM activity, no conclusive evidence was found to link these processes. Long-term enrichments of coastal sediments with various electron acceptors demonstrated a positive influence of sulfate and ferric citrate additions on AOM. Other electron acceptors such as nitrate and manganese may also support AOM in these coastal settings. These studies advance the understanding of the seasonal controls on methane emissions, methane production, and methane consumption via AOM in freshwater ecosystems. Future efforts are aimed at closer examinations of these mediating factors, especially temperature changes and substrate availability. INDEX WORDS: methane, sulfate, biogeochemistry, AOM, methanogenesis, freshwater, wetland, lipid A STUDY OF METHANE-RELATED PROCESSES IN FRESHWATER ECOSYSTEMS: METHANOGENESIS, ANAEROBIC METHANE OXIDATION, AND INTERACTIONS WITH OTHER TERMINAL METABOLISMS by KATHERINE EOWYN SEGARRA BS, Brown University, 2002 A Dissertation Submitted to the Graduate Faculty of The University of Georgia in Partial Fulfillment of the Requirements for the Degree DOCTOR OF PHILOSOPHY ATHENS, GEORGIA 2012 © 2012 Katherine Eowyn Segarra All Rights Reserved A STUDY OF METHANE-RELATED PROCESSES IN FRESHWATER ECOSYSTEMS: METHANOGENESIS, ANAEROBIC METHANE OXIDATION, AND INTERACTIONS WITH OTHER TERMINAL METABOLISMS by KATHERINE EOWYN SEGARRA Major Professor: Samantha B. Joye Committee: Christof Meile Vladimir Samarkin Kai-Uwe Hinrichs William ‘Barny’ Whitman Electronic Version Approved: Maureen Grasso Dean of the Graduate School The University of Georgia May 2012 ACKNOWLEDGEMENTS Above all I must thank my advisor, Mandy Joye. Her support, both intellectual and financial, made this work possible. I am in her debt for her guidance, for providing quality feedback and insight, and for always challenging me to reach beyond my limits. I would also like to thank my other committee members, for whom I have a great deal of respect and admiration. Vladimir Samarkin, who was very giving of his time and knowledge, always had a kind face and a good joke. I appreciate the intellectual and emotional support he has provided over the years. Christof Meile has been very supportive during my PhD tenure. He is a wonderful editor and provided creative solutions to problems I faced. William ‘Barny’ Whitman was both a great teacher and committee member. I appreciate his wisdom and thoughtful comments. And last, but not least, I thank Kai-Uwe Hinrichs, who is both a great scientist and a great advisor. I am so grateful to him for making my time in Bremen an unforgettable experience. Working in the Joye Lab has been an amazing experience and I wish to thank all of its members, both past and present, especially Laura Polomo, Christelle Hyacinthe, Melitza Crespo-Medina, Charles Schutte, Chris Comerford, and Julia Slaughter. Thanks to Kim Hunter for keeping things running smoothly and efficiently. Special thanks to Bill Porubsky for getting my feet wet in the field of biogeochemistry and navigating the science world. A big thank you to my undergraduate helpers, especially Ruthie Taylor, Laura Potter, Leia Edenfelder, and Will Spence. iv There is a small but special community of people in Athens that made this journey all the better. A big hug to Megan ‘Chubi’ Machmuller, Peter Baas, and Andrew Binderup – thanks for the camaraderie and love. I am grateful to Beth! Orcutt for being a real friend in the science world. You are an inspiration to me – kein Fleisch! Eric King was an invaluable colleague whether we were pondering data or driving up and down the east coast in a windowless van. Thanks for all the Dexters and the Weezer snuggie. My time at the MARUM in Bremen, Germany were the most rewarding months of my PhD program. I thank the A.G. Hinrichs for making me feel very welcome and for their generosity of knowledge, time, and energy. I especially thank Florence Schubotz for her tireless enthusiasm, and for her wonderful analytical and editorial assistance. A big thanks to my other Bremen-based co-authors, Marcos Yoshinaga, and Verena Heuer. A special thanks to Julius Lipp, Matthias Kellerman, Marcus Elvert, Xavier Prieto, Eoghan Reeves, and Travis Meador for teaching me the ins and outs of lipid analysis, for hot mensa dates, and for making those long days huffing solvents even more enjoyable. I thank my family (Mom, Dad, and Ben), whose generosity, support, and unconditional love have been wondrous gifts that I may never repay. And finally, I thank Noah, my light when all other lights have gone out. This dissertation project was primarily funded by a NSF DEB award. The LTER sites at the Georgia Coastal Ecosystems and the Florida Coastal Everglades provided field support and personnel. I especially want to thank Daniel Saucedo and Franco Tobias, two very capable and hardworking captains. Additional financial support was provided by a Boyd Scholarship, The Alfred E. Brown Scholarship, and a recruitment fellowship from the Graduate School at the University of Georgia. v TABLE OF CONTENTS Page ACKNOWLEDGEMENTS ............................................................................................... iv CHAPTER 1 INTRODUCTION ...................................................................................................1 2 SEASONAL VARIATIONS OF METHANE FLUXES FROM A TIDAL, FRESHWATER WETLAND (HAMMERSMITH CREEK, GA) ..........................................................11 3 TIDAL, FRESHWATER SEDIMENTS SUPPORT HIGH RATES OF ANAEROBIC OXIDATION OF METHANE ...................................................................................44 4 SEASONAL PATTERNS IN METHANE CYCLING IN FLORIDA AND MAINE PEAT WETLANDS ........................................................................................................94 5 IMPACT OF ELECTRON ACCEPTOR AVAILABILITY ON THE ANAEROBIC OXIDATION OF METHANE IN COASTAL FRESHWATER AND BRACKISH INTERTIDAL SEDIMENTS ..................................................................................186 6 CONCLUSIONS AND OUTLOOK ........................................................................250 vi CHAPTER 1 INTRODUCTION “There is something fascinating about science. One gets such wholesale returns of conjecture out of such a trifling investment of fact. — Mark Twain, Life on the Mississippi Despite anthropogenic changes, microbial processes continue to play a key role in regulating the Earth’s atmosphere and climate. This PhD dissertation explored pathways of microbially-mediated carbon flow in freshwater environments. In these chapters, I examined the factors, both biotic (e.g., microbial community composition) and abiotic (e.g., temperature), that regulate Figure 1.1: Atmospheric methane concentrations at the terminal metabolism of organic carbon in anoxic Mona Loa, Hawaii observatory from 1983 to 2002. From sediments, with a specific focus on seasonal Canfield et al. (2005). variations on rates and pathways of methane (CH4) cycling in wetland ecosystems. CH4, a very potent greenhouse gas, has a heat-trapping potential twenty-five times that of carbon dioxide (BLAKE and ROWLAND, 1988). Increases of global atmospheric 1 methane levels (about 1% per year) contribute to present-day global warming (Fig. 1.1; IPCC, 2007; KHALIL and RASMUSSEN, 1990). Atmospheric methane concentrations are currently estimated at 1.8 ppm (IPCC, 2007). Sources of this methane include a variety of natural and anthropogenic (Fig. 1.2), the largest of which is natural wetlands (BROOK et al., 2008). A large uncertainty is associated with this source due to the high spatial variability of these