THE INTESTINAL MICROBIOME IN INFLAMMATORY BOWEL DISEASE THE INTESTINAL MICROBIOME IN INFLAMMATORY BOWEL DISEASE AND ITS RESPONSE TO THERAPY By JOSIE LIBERTUCCI, M.Sc., B.Sc. A Thesis Submitted to the School of Graduate Studies in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy McMaster University © Copyright by Josie Libertucci, December 2016 PhD Thesis - J. Libertucci; McMaster University - Biochemistry and Biomedical Sciences DESCRIPTIVE NOTE Doctor of Philosophy (2016) McMaster University, Hamilton, Ontario (Biochemistry and Biomedical Sciences) TITLE The intestinal microbiome in inflammatory bowel disease and its response to therapy AUTHOR Josie Libertucci, MSc SUPERVISOR Dr. Michael G. Surette NUMBER OF PAGES xvi, 207 ii PhD Thesis - J. Libertucci; McMaster University - Biochemistry and Biomedical Sciences ABSTRACT Inflammatory bowel diseases, consisting of both Crohn’s disease (CD) and ulcerative colitis (UC), occur when a susceptible host mounts a chronic aberrant immune response to its commensal microbial community resulting in chronic inflammation. CD and UC may exhibit similar clinical manifestations that oscillate between active disease and remission, but these diseases are distinct, and are characterized by specific pathophysiology. The incidence and prevalence of IBD has been increasing over time, with the highest rates occurring in Europe and North America. Although many genome- wide association studies (GWAS) on CD and UC have found loci associated with these diseases, studies conducted on monozygotic twins show low concordance rates, suggesting environmental causes play an important role in the onset of IBD. Current therapies for treatment of IBD focus on suppressing the immune response, but remission rates remain low and relapses occur often. There is a dire need for new treatment options that involve suppressing the immune system stimuli – the commensal gut microbiota. The commensal microbiota triggers the mucosal immune system to drive chronic inflammation in IBD, and studies have shown that patients display a gut dysbiosis compared to healthy individuals. Modulation of the disease through modulation of the gut microbiome, has recently gained interest as a potential treatment option. Little is known about how the microbial community in IBD patients will respond to therapies that attempt to modulate the community. Thus, the overall goal of this thesis was to characterize the IBD microbiome in relation to disease status, severity, and response to therapy. Together these results, (a) provide an in-depth characterization of the intestinal microbiome in IBD, iii PhD Thesis - J. Libertucci; McMaster University - Biochemistry and Biomedical Sciences (b) demonstrate the response of the gut microbiome in UC patients to FMT, providing insight into the successful mechanism, and (c) provide evidence for inflammation related differences in CD giving insight into the mechanism of pathogenesis. These results not only further our understanding regarding dysbiosis in IBD, but also can be used to modify our current treatment protocols to increase efficacy. iv PhD Thesis - J. Libertucci; McMaster University - Biochemistry and Biomedical Sciences ACKNOWLEDGEMENTS Firstly, I would like to thank my supervisor Dr. Michael G. Surette. I have learned so much from him over these past years, and I have very much enjoyed my time in his lab. I appreciate all of his guidance and support throughout my degree. It was an absolute pleasure to work with a supervisor so passionate and excited about the wonderful microbes that live in us and around us! But most of all, I am thankful for how much he has challenged me throughout these past years. He constantly pushed me to be a better scientist, and now I can proudly say that I have accomplished things that I did not even know I was capable of doing! I would also like to thank my supervisory committee, Dr. Lori Burrows and Dr. Elena F. Verdu. Thank for challenging me to think critically about my work. Your insight and guidance were invaluable. Most of all, thank your for being strong role models that I had the pleasure of looking up to. I was lucky to have a truly wonderful committee. Specifically, I am truly grateful for all the time Elena spent mentoring me throughout the past years. She helped me develop professionally as a scientist, and guided me throughout my most difficult times that I encountered in my degree. To all of the Surette lab members, both past and present, thank you for all the help you have given me throughout these years. It was amazing to have the opportunity to work with others that are so very passionate about science! I learned a lot from the members in the lab. I would like to specifically thank Laura Rossi and Michelle Shah for all of their help in the lab. I would like to thank all the Farncombe Institute members, trainees, scientists and clinician scientists. To all the trainees, thank you for making the Farncombe Institute such an amazing place to work! I have had the wonderful opportunity to work with so many talented individuals. Specifically I would like to thank our collaborators Dr. Paul Moayyedi, Melanie Wolfe, Dr. Christine Lee, Catherine Onishi, and Dr. David Armstrong. I would like to thank my family and friends for their support and encouragement. But most of all I would like to thank my parents for teaching me the value of hard work. And lastly, I would like to thank my amazing and very supportive husband. v PhD Thesis - J. Libertucci; McMaster University - Biochemistry and Biomedical Sciences TABLE OF CONTENTS THE INTESTINAL MICROBIOME IN INFLAMMATORY BOWEL DISEASE AND ITS RESPONSE TO THERAPY ............................................................................. i DESCRIPTIVE NOTE ..................................................................................................... ii ABSTRACT ...................................................................................................................... iii ACKNOWLEDGEMENTS ............................................................................................... v TABLE OF CONTENTS ................................................................................................. vi LIST OF FIGURES ........................................................................................................... x LIST OF TABLES .......................................................................................................... xii LIST OF ABBREVIATIONS AND SYMBOLS .......................................................... xiv CHAPTER 1 INTRODUCTION ............................................................................................................ 17 1.1 The human intestinal microbiome .............................................................................. 2 1.1.1 Defining a healthy microbiome ..................................................................... 2 1.1.1.1 Colonization resistance .............................................................................. 3 1.1.1.2 Short chain fatty acid production ............................................................. 4 1.2 Maintaining intestinal homeostasis ......................................................................... 5 1.2.1 Intestinal barrier function .................................................................................... 6 1.2.2 The early life intestinal microbiome in humans and factors that shape the immune response ............................................................................................................ 7 1.2.3 The microbiota and its influences on the immune system ................................. 9 1.3 Investigating the gut microbiome ......................................................................... 10 1.3.1 16S rRNA marker gene sequencing .................................................................. 11 1.3.2 Shotgun metagenomics ....................................................................................... 13 1.4 Inflammatory bowel disease .................................................................................. 13 1.4.1 Ulcerative Colitis .............................................................................................. 14 1.4.2 Crohn’s Disease ................................................................................................ 15 1.5 Modulating the microbiome via fecal microbiota transplantation to modulate disease activity in inflammatory bowel disease ............................................................. 17 1.5.1 History of FMT .................................................................................................... 19 1.5.2 FMT efficacy in Clostridium difficile infection ................................................ 20 1.5.3 FMT in CD and associated microbial changes ................................................. 23 1.5.4 FMT in UC Cohort Studies and associated microbial changes ...................... 24 1.5.5 Donor specificity for treatment of CDI ............................................................. 25 1.5.6 Anaerobic considerations for FMT ................................................................... 26 vi PhD Thesis - J. Libertucci; McMaster University - Biochemistry and Biomedical Sciences 1.5.7 Mechanism of Action in FMT – Engraftment .................................................. 27 1.5.8 Mechanism of Action in FMT – Other possibilities ......................................... 28 1.6 Thesis Scope ...............................................................................................................