The assembly and functions of microbial communities on complex substrates by Xiaoqian Yu B.S./M.S., Molecular Biophysics and Biochemistry Yale University (2011) Submitted to the Biology Graduate Program in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy at the MASSACHUSETTS INSTITUTE OF TECHNOLOGY September 2019 2019 Massachusetts Institute of Technology. All rights reserved. Signature of Author: ____________________________________________________________________ Xiaoqian Yu Department of Biology Certified by: __________________________________________________________________________ Eric J. Alm Professor of Biological Engineering Professor of Civil and Environmental Engineering Thesis Advisor Accepted by: _________________________________________________________________________ Amy Keating Professor of Biology Co-Director, Biology Graduate Committee The assembly and functions of microbial communities on complex substrates by Xiaoqian Yu Submitted to the Department of Biology on August 5th, 2019 in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in Biology Abstract Microbes form diverse and complex communities to influence the health and function of all ecosystems on earth. However, key ecological and evolutionary processes that allow microbial communities to form and maintain their diversity, and how this diversity further affects ecosystem function, are largely underexplored. This is especially true for natural microbial communities that harbor large numbers of species whose interactions are often the result of long term evolutionary processes of co-occurring organisms. In this thesis, I make use of “common garden experiments” — introducing varying microbial communities to the same environments — to investigate how the assembly and functions of natural microbial communities are affected by the diversity of communities, as well as the chemical nature of substrates that they assemble on. In the first project, I present an experimental workflow that streamlines the generation of self-assembled microbial communities with a wide range of diversity, measurements of community function in “common gardens”, followed by subsequent isolation of the most abundant taxa from these communities via dilution-to-extinction. This high-throughput workflow is applied to assess how interactions scale with organismal diversity to affect the function of microbial communities from the coastal ocean. In the second project, I use a combination of theoretical models and an ex vivo experimental framework to examine how the volume and content of gas produced by gut microbiota assembling on different prebiotic substrates (“gardens”) are influenced by the chemical nature of the substrate and the composition of the gut microbiota itself. As a whole, this body of work represents a small step towards finding common organization principles in microbial community assembly and their functional consequences. Thesis Supervisor: Eric J. Alm Title: Professor of Biological Engineering, Professor of Civil and Environmental Engineering 2 Acknowledgements As the long journey of graduate school finally comes to a conclusion, despite it has taken many more twists and turns than I expected, I look back but still cannot help say it was a joyful and satisfying experience. I owe this to the mentors that have guided me along the way, and my friends and family who were always encouraging, patient and supportive. My first thanks must, of course, go to my advisor, Eric Alm. Eric was brave enough to take me into his lab despite I knew nothing about the microbiome, genomics, or statistics. Transitioning from a protein biochemist to a microbial ecologist was hard, and I definitely had a long lag phase in doing so, but Eric was always positive, optimistic and encouraging, pulling me out of the weeds enough times before I eventually learned to not get lost in them so frequently. And as I became more scientifically mature, I was given all the freedom and trust to carry my projects in directions that I desired, which was a great boost to my confidence. Eric was also always able to come to the rescue when I got confused with math – any improvement in my quantitative skills I had over the past years, I would attribute to Eric. I am also deeply thankful to Martin Polz, who have effectively been my co-advisor despite not signing any official papers. Little did I know when I first met with Martin because I “did a rotation on Vibrios” how lucky I would be to have him as a mentor, always being there when I needed help or support. Through working with Martin I learned how to treat scientific results with both rigor and respect, and how to not be intimidated by the imperfectness of experimental results. He also exemplified to me how novelty in science is deeply rooted in logic and knowledge, as well as how doing research is a life-long learning process. These are values that I will carry on for life. I am also grateful to my committee members, Penny Chisholm and Jing-ke Weng, who have been with me for six years. Both in and outside of committee meetings, Penny has showered me with gems of wisdom, providing me with both intellectual and emotional support. Jing-ke not only brings a non- ecologist view to my committee meetings, but has been a constant source for practical advice. Outside of my committee, I have largely benefited from the conversations I have had with Serguei Saavedra, which always helped me put my non-systematic knowledge of ecology into a systematic context. Jeff Gore was also pivotal for my decision of switching into the field of microbial ecology. I have been incredibly lucky to have two “homes” for my graduate school: both the Alm lab and the Polz lab were filled with young scientists who were friendly, fun and incredibly helpful. Sarah Spencer was my first window to the Alm lab; this was followed by many conversations over tea in the Tech square kitchen. A lot of my intellectual growth during grad school I owe to the three postdocs that I shared an office with: Sean Gibbons, Tami Lieberman and Fangqiong Ling. They were always patient in answering my questions about data processing, helping me look over my writing, or telling me how to build academic connections. We also had great times together brainstorming about anything related to the microbiome, ecology, or population genomics. Shijie Zhao has also been a constant source of intellectual stimulation, always bringing new ideas and techniques to the table. My collaborators in the lab for the project in chapter three, Thomas Gurry and Tu Nguyen, have always been quick and responsive to answer my questions or provide experimental support. I also greatly enjoyed the times that I 3 would “whine” with Sean Kearney about library preps or funky bacteria, or just anything not working in lab. And as I was rushing to try to finish my projects at the end of grad school, Fuqing Wu and Anni Zhang were never hesitant to lend a helping hand. Shandrina was also a great hero in making sure that I could get facetime with Eric, that is, making things happen. Jan-Hendrik Hehemann was my first mentor in the Polz lab. Also coming from a biochemistry background, he helped me see and learn how genomics, biochemistry and ecology can be brought together into a project and complement each other. He also helped shape some of my earliest research ethics, that is to always be respectful of others’ ideas and contributions. Manoshi Datta also provided me a lot of guidance during my early years in the lab, despite that she was only one year senior than me. Chris Corzett was one of the best collaborators I could have hoped for—responsible and meticulous, I would often trust his experimental results more than my own. Kathryn Kauffman is truly someone that I admire and love: her persistence, kindness, and generosity make her the exemplary lab citizen that I would hope to become one day. Fabiola Miranda is also someone that I would look forward to seeing when I come to work—her energy is infectious like sunshine, and she was never shy to share her knowledge and advice, or even cuddle me a bit when I am low in emotions. And whenever I rushed into the Polz lab office for help, i.e. looking for Joy because I needed help with stats, or Joseph and Dave for bioinformatics, or Fatima for almost anything general in lab, I was able to. Finally, Michael Cutler was essential for helping me in many aspects in the lab, amazingly cobbling things together (often from cardboard!) whenever I needed it. I also owe much of my development to the wonderful scientific community here at MIT. I am deeply grateful for the generosity and supportiveness of Amy Keating, the co-director of the graduate program in the biology department, while I was going through the “bumps” of life and research. Betsey Walsh was also an ever amazing admin that knew the solution to every question or problem that I had. Craig Mclean has been a great friend and collaborator, sharing my passions in connecting chemistry to ecology. I have also benefited greatly from talking to many people in the Gore lab, the Cordero lab, the Chisholm lab and the Saavedra lab: Jonathan Friedman, Arolyn Conwil, Avihu Yona, Daniel Amor, Shaul Pollak, Ali Ebrahimi, Julia Schwartzman, Rogier Braakman, and Chuliang Song. Much of my molecular biology techniques I learned from my mentors and labmates during my short time in the Lindquist lab: Kendra Frederick, Greg Newby, and Can Kayatekin. Finally, I thank my friends and family for keeping me happy and positive during my PhD. Jie Bai and I came to MIT together from college, and we have not only shared rooms and dinners, but also all the ups and downs in graduate school, research and life. My wonderful friends in the “gang of Chedan (bullshit)”, Lixi Wang, Tewei Luo and Xuanzong Guo have traveled far and wide with me, attempted to keep a book blog together, and listened to my rants about difficulties in life.