The anaerobic oxidation of short-chain alkanes in hydrothermal vents Citation Adams, Melissa Marie. 2014. The anaerobic oxidation of short-chain alkanes in hydrothermal vents. Doctoral dissertation, Harvard University. Permanent link http://nrs.harvard.edu/urn-3:HUL.InstRepos:12274143 Terms of Use This article was downloaded from Harvard University’s DASH repository, and is made available under the terms and conditions applicable to Other Posted Material, as set forth at http:// nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of-use#LAA Share Your Story The Harvard community has made this article openly available. Please share how this access benefits you. Submit a story . Accessibility The anaerobic oxidation of short-chain alkanes in hydrothermal vents A dissertation presented by Melissa Marie Adams to The Department of Organismic and Evolutionary Biology in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the subject of Biology Harvard University Cambridge, Massachusetts May 2014 ©2014 – Melissa Marie Adams All rights reserved. Advisor: Peter R. Girguis Melissa Marie Adams The anaerobic oxidation of short-chain alkanes in hydrothermal vents Abstract Microorganisms are central to the cycling of methane on Earth. The anaerobic oxidation of methane (AOM) is a globally important process in anoxic marine sediments, which is often coupled to the reduction of sulfate by anaerobic methanotroph (ANME) archaea and sulfate reducing bacteria (SRB). However, the environmental and geochemical conditions that constrain these microbial communities remain largely uncharacterized. In this dissertation, I present evidence that methane and C -C alkanes are substantial sources 2 4 of metabolic energy in sedimented hydrothermal vent systems. Furthermore, these studies demonstrate that AOM and C -C alkane oxidation linked to sulfate reduction (SR) are 2 4 governed by temperature and substrate availability. Using continuous-flow sediment bioreactors and batch incubations, rate measurements revealed a striking uncoupling of AOM from SR across the thermal gradient characteristic of hydrothermal vent sediments, with AOM occurring at the highest temperature (90°C) in the absence of SR. I discovered that ANMEs were present in sediments at all temperatures investigated, including a unique thermophilic ANME clade, while SRB were only detected in cooler regimes. Next, I present data from batch incubations demonstrating for the first time that substantial C -C alkane oxidation occurs 2 4 ii i over a broad temperature range (25 - 75°C) in hydrothermal vent sediments and is coupled to SR up to 75°C with maximum rates at 55°C. Furthermore, there was preferential degradation of C -C alkanes at 55°C, indicating that the active alkane oxidizers are 2 4 thermophilic. At the rate maxima, 16S rRNA pyrotag sequence data revealed that a novel SRB lineage was the likely phylotype mediating the anaerobic oxidation of C -C alkanes. 2 4 Finally, I present a comparative study of methane- and sulfur-cycling ecotypes in geographically separated hydrothermal vent and hydrocarbon seep sediments. By employing high throughput sequencing of 16S rRNA genes coupled to geochemical data, I was able to establish the environmental constraints that determine ANME and SRB ecotype diversity and distribution in these biogeochemically distinct deep sea habitats. In summary, this dissertation sheds light on the pervasive effects of temperature, substrate availability, and habitat type on these metabolic processes that are critical for the cycling of carbon and sulfur in deep sea hydrothermal vent sediments. iv The anaerobic oxidation of short-chain alkanes in hydrothermal vents Table of Contents Acknowledgements ........................................................................................... vi Chapter 1 ......................................................................................................... 1 Introduction Chapter 2 ....................................................................................................... 20 Anaerobic methane oxidation in metalliferous hydrothermal sediments: influence on carbon flux and decoupling from sulfate reduction Chapter 3 ....................................................................................................... 36 Anaerobic oxidation of short-chain alkanes in hydrothermal sediments: potential influences on sulfur cycling and microbial diversity Chapter 4 ....................................................................................................... 48 Environmental distribution of anaerobic methanotrophic and sulfate reducing ecotypes in hydrothermally influenced sediments Chapter 5 ....................................................................................................... 90 Conclusions and Future Directions Appendix 1 ..................................................................................................... 99 Chapter 2 Supplemental Material v Acknowledgements We shall not cease from exploration And the end of exploration Will be to arrive where we started And know that place for the first time. -T.S. Eliot, Four Quarters (1943) Growing up in rural North Carolina, I was always fascinated by the diversity of life and the natural environment around me. I never imagined that this desire to explore the world would allow me to go to graduate school and have the incredible opportunity to learn from and work with top scientists at Harvard University. The kind and generous efforts of many people have enabled me to complete this dissertation, and I would like to express my gratitude to some of these key individuals. First and foremost, I would like to express my deepest thanks to my advisor, Professor Peter Girguis. I truly appreciate his ability to think outside the box and to take someone with such a diverse background as myself under his wing. Pete has been a mentor and friend that has taught me to be a better scientist. I have learned the vital skill of how to bridge different scientific fields in my research, which will be a great asset in my future career. Pete has also shown me that it is possible to have a thriving career, a terrific lab group, a wonderful family, an impressive whiskey collection, and a great outlook on life. Next I would like to thank the wonderful members of the Girguis lab. A special thanks to Scott Wankel, Arpita Bose, Dave Johnston, Mark Nielson, and Ulli Jaekel for sharing their expertise and providing invaluable guidance and mentorship. The former members of “Team Mud” have taught me how to be both a geochemist and microbiologist. I have received amazing support from Heather Olins in my graduate cohort and Emily v i Gardel that joined the lab with me. I have also greatly appreciated the efforts of Jennifer Delaney and Stephanie Hillsgrove for taking care of many logistical tasks behind the scenes that made the lab a better place to work. It was a pleasure mentoring Emma Dillon, Adrienne Hoarfrost, and Jaemin Jang in the lab and Ryan Lee and Claire Stolz in the classroom. I look forward to following their future careers. I thank my thesis committee, Colleen Cavanaugh, Andrew Knoll, and Dave Johnston for their time, patience, and intellectual contributions that were invaluable to my thesis and development as a scientist. My interdisciplinary research has also allowed me to work with fantastastic colloborators, particularly Jen Biddle, Mandy Joye, and Frank Stewart. My projects would not have been possible without each of you. I acknowledge the R/V Atlantis crew and team of the DSV Alvin for deep sea collections that were critcal for my research projects. I would like to thank the Center for Dark Energy Biosphere Investigations (C-DEBI) and their Fellowship program for funding my graduate study. Outside of my research and teaching experiences, I had incredible support from the Office of Career Services at Harvard University. Heather Law and Laura Stark provided wonderful guidance and were encouraging throughout the entire process of choosing the next steps in my career. Through the Association for Women in Science (AWIS) in Boston, I also had the opportunity to work with a phenomenal mentoring group and form friendships with inspiring female scientist from other universities. I thank Hila Baher, Danielle Feldman, Carrie Hui, Melissa Labonty, and Angela Seliga for their energy, thoughtfulness, and inspiration. vi i I would like to mention the incredible group of friends that were always there for me throughout this journey. In particular, Poem Turner for her sharp wit and appreciation of the finer things in life, Vanessa Guzman and Laura Campbell for keeping me young, and Melissa Kern and Bridgett Payne for providing humor and perspective when things were tough. The ladies of Redline Fight Sports were also a supportive backbone for me throughout graduate school and made sure I had fun whether it was at our weekend brunch or crafting nights. In particular, I thank Ariela Weinbach, Tara Jennings, Jennifer Marchio, and Kristy Nordstrom. You are some of the bravest people I know and have been an incredible influence on me. I would also like to acknowledge Jen Dziura, who taught me to stay bullish by working hard, reserving time for things that are completely awesome, and ignoring everything in between. My thanks to the beautiful music of Sigur Rós, Grimes, and the Magnetic Fields that made writing my thesis more enjoyable. Finally, I thank my family for serving as a source of great comfort and encouragement throughout graduate school. My mother and father have provided unwavering support and love in all challenges that I have faced. My mother has taught me to be kind and appreciate everything I have in this life. I credit my father for instilling an ambitious drive in me and also a love of understanding the world. He was my first science teacher, explaining to a five-year old how a battery worked. My brother Matt and his wife Ellen are both great inspirations to me. Ruthann and Chuck Stiles were also very supportive with incredible kindness, positivity, wisdom, and humor. To my partner Dylan Stiles, words cannot express how blessed I feel to have you in my life. We jump out of planes and venture into unchartered territory together. Thank you to everyone that has been there for me in this great exploration. vi ii Chapter 1 Introduction