Metagenomic analysis of river microbial Title: Metagenomic analysis of river microbial communities Examining

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  • Metagenomic analysis of river microbial communities

    by

    Thea Van Rossum

    Bachelor of Science, University of British Columbia, 2010

    Thesis Submitted in Partial Fulfillment of the

    Requirements for the Degree of

    Doctor of Philosophy

    in the

    Department of Molecular Biology and Biochemistry

    Faculty of Science

    © Thea Van Rossum

    SIMON FRASER UNIVERSITY

    Fall 2017

    Copyright in this work rests with the author. Please ensure that any reproduction or re-use is done in accordance with the relevant national copyright legislation.

  • ii

    Approval

    Name: Thea Van Rossum

    Degree: Doctor of Philosophy

    Title: Metagenomic analysis of river microbial

    communities

    Examining Committee: Chair: Dr. Michel Leroux Professor

    Dr. Fiona Brinkman Senior Supervisor Professor

    Dr. William Davidson Supervisor Professor

    Dr. Margo Moore Supervisor Professor

    Dr. Ryan Morin Internal Examiner Assistant Professor

    Dr. Steven Hallam External Examiner Associate Professor Microbiology & Immunology University of British Columbia

    Date Defended/Approved: September 22, 2017

  • iii

    Abstract

    As concern over the availability of freshwater increases, so does the interest in river

    microorganisms due to their importance in drinking water safety and signalling environmental

    contamination. However, foundational understanding of their variability in rivers is lacking,

    especially for viruses. Here, I present work to improve the understanding of planktonic microbial

    communities in rivers over time in the context of varying environmental conditions and

    contrasting land use. DNA-sequencing based metagenomic and phylogenetic marker gene

    (16S, 18S, g23) approaches were used to profile microbial communities, coupled with measures

    of environmental and chemical conditions. I analysed microbial community profiles from monthly

    samples collected over one year from three watersheds with agricultural, urban, or minimal land

    use. Viral, bacterial, and microeukaryotic planktonic communities were synchronous overall, but

    had contrasting geographic patterns and the strength of their synchrony, as well as their

    relationships with environmental conditions, were heterogenous across sampling sites. These

    differences illustrated that bacteria are important yet insufficient representatives of microbial

    community dynamics despite their prevalence in microbiome research. However, this emphasis

    on bacteria has produced richer reference databases, which enabled a gene-specific analysis.

    Using a reference-based approach, I found that communities with lower water quality due to

    agricultural activity had higher abundances of nutrient metabolism and bacteriophage gene

    families. Based on these water quality associated findings and on complementary analyses, I

    identified potential biomarkers to demonstrate that bacterial river metagenome data could

    feasibly support the development of new assays for water quality monitoring. To complement

    these studies of anthropogenic contamination, I studied bacteria in river biofilms across a

    natural gradient of metal concentrations at a potential mining site. Clear relationships among

    metal concentrations, pH, and microbiomes were evident and this study provided fundamental

    knowledge of microbial communities at a potential mine site before disruption from

    development. Throughout these studies, the scarcity of reference information for microbial

    communities in lotic freshwater provided an opportunity to identify weaknesses in popular

    microbiome analysis methods and present approaches better suited to poorly characterised

    environments. Overall, my work aims to improve the understanding of planktonic river microbial

    community variability, both for the advancement of basic science and to support future

    development of more effective water quality monitoring approaches.

  • iv

    Keywords: metagenomics; microbiome; rivers; microbial ecology; water quality

  • v

    Acknowledgements

    I would like to acknowledge Dr. Fiona Brinkman for being a wonderful leader, teacher,

    supervisor, and mentor. It has been my privilege to receive her support in every endeavour I

    have pursued. I would also like to acknowledge my committee members, Dr. Margo Moore and

    Dr. Willie Davidson for their thoughtful discussions and support throughout my graduate studies.

    A large section of my research was within the context of the GC Watershed Project in

    collaboration with the BCCDC. I would like to acknowledge the supportive and dynamic

    atmosphere the leaders of this project created and the cooperative spirit that all members of the

    project brought to the research. I also enjoyed an excellent collaboration with Dr. Chris Kennedy

    and his lab members. Throughout my time in the Brinkman lab, my lab mates have provided

    support, creative input and fun whenever possible. I would especially like to acknowledge Dr.

    Mike Peabody and Dr. Emma Griffiths for their contributions to my thesis research and Geoff

    Winsor for guidance in my very first bioinformatics project. Finally, I would also like to

    acknowledge all the funding agencies that have supported my thesis work: NSERC, Simon

    Fraser University, Genome Canada, Genome BC, Canadian Water Resources Association, and

    the MBB department.

    To my wonderful parents, I will never be able to say thank you often enough. But I can

    try: thank you. Your love, confidence and guidance encouraged me to never doubt I could

    achieve something if I worked consistently and creatively enough. Your unwavering support

    gave me the courage to risk failing. Your trust in me inspired me to trust in myself. Brett, my

    brilliant partner, your passion for science and your long view of its ups and downs kept me going

    strong throughout my graduate studies. Thank you for always reminding me to celebrate the

    wins, both big and small, and being a calming presence when I needed reassurance. To the rest

    of my family and friends, through the celebrations and commiserations, you’ve helped me smile,

    laugh, and keep moving forward. Thank you all for your wit, kindness, and encouragement.

  • vi

    Table of Contents

    Approval...................................................................................................................................... ii

    Abstract...................................................................................................................................... iii

    Acknowledgements ..................................................................................................................... v

    Table of Contents ....................................................................................................................... vi

    List of Tables .............................................................................................................................. x

    List of Figures ............................................................................................................................ xi

    List of Acronyms ....................................................................................................................... xii

    Chapter 1. Introduction ......................................................................................................... 1

    1.1. Brief overview of river microbial ecology ............................................................................ 1

    1.1.1. Importance of riverine microbial communities ........................................................... 1

    1.1.2. Ecological shaping forces in riverine microbial communities ..................................... 2

    1.1.2.1. General ecological shaping forces in microbial communities .................................. 2

    1.1.2.2. Shaping forces of free-floating microbial communities in rivers .............................. 4

    1.2. Motivation to improve knowledge of microbial communities in rivers ................................. 8

    1.2.1. Lack of foundational characterisation of riverine microplankton ................................ 8

    1.2.2. Towards the development of improved water quality monitoring techniques ............. 9

    1.2.3. The Applied Metagenomics of the Watershed Microbiome project (“GC Watershed Project”) ................................................................................................................................11

    1.3. Microbiome profiling methods based on DNA sequencing ............................................... 12

    1.3.1. Phylogenetic marker gene profiling ..........................................................................13

    1.3.1.1. Phylogenetic marker sequence choice ..................................................................13

    1.3.1.2. Community fingerprinting / gel-based methods .....................................................13

    1.3.1.3. DNA-sequencing-based methods ..........................................................................14

    1.3.2. Shotgun metagenomics .................................