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A Comparative Analysis of the Moose Rumen Microbiota and the Pursuit of Improving Fibrolytic Systems

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A Comparative Analysis of the Moose Rumen Microbiota and the Pursuit of Improving Fibrolytic Systems University of Vermont ScholarWorks @ UVM Graduate College Dissertations and Theses Dissertations and Theses 2015 A Comparative Analysis Of The oM ose Rumen Microbiota And The Pursuit Of Improving Fibrolytic Systems. Suzanne Ishaq Pellegrini University of Vermont Follow this and additional works at: https://scholarworks.uvm.edu/graddis Part of the Animal Sciences Commons, Microbiology Commons, and the Molecular Biology Commons Recommended Citation Pellegrini, Suzanne Ishaq, "A Comparative Analysis Of The oosM e Rumen Microbiota And The urP suit Of Improving Fibrolytic Systems." (2015). Graduate College Dissertations and Theses. 365. https://scholarworks.uvm.edu/graddis/365 This Dissertation is brought to you for free and open access by the Dissertations and Theses at ScholarWorks @ UVM. It has been accepted for inclusion in Graduate College Dissertations and Theses by an authorized administrator of ScholarWorks @ UVM. For more information, please contact [email protected]. A COMPARATIVE ANALYSIS OF THE MOOSE RUMEN MICROBIOTA AND THE PURSUIT OF IMPROVING FIBROLYTIC SYSTEMS. A Dissertation Presented by Suzanne Ishaq Pellegrini to The Faculty of the Graduate College of The University of Vermont In Partial Fulfillment of the Requirements For the Degree of Doctor of Philosophy Specializing in Animal, Nutrition and Food Science May, 2015 Defense Date: March 19, 2015 Dissertation Examination Committee: André-Denis G. Wright, Ph.D., Advisor Indra N. Sarkar, Ph.D., MLIS, Chairperson John W. Barlow, Ph.D., D.V.M. Douglas I. Johnson, Ph.D. Stephanie D. McKay, Ph.D. Cynthia J. Forehand, Ph.D., Dean of the Graduate College ABSTRACT The goal of the work presented herein was to further our understanding of the rumen microbiota and microbiome of wild moose, and to use that understanding to improve other processes. The moose has adapted to eating a diet of woody browse, which is very high in fiber, but low in digestibility due to the complexity of the plant polysaccharides, and the presence of tannins, lignin, and other plant-secondary compounds. Therefore, it was hypothesized that the moose would host novel microorganisms that would be capable of a wide variety of enzymatic functions, such as improved fiber breakdown, metabolism of digestibility-reducing or toxic plant compounds, or production of functional metabolites, such as volatile fatty acids, biogenic amines, etc. The first aim, naturally, was to identify the microorganisms present in the rumen of moose, in this case, the bacteria, archaea, and protozoa. This was done using a variety of high-throughput techniques focusing on the SSU rRNA gene (see CHAPTERS 2-5). The second aim was to culture bacteria from the rumen of the moose in order to study their biochemical capabilities (see CHAPTERS 6-7). The final aim was to apply those cultured bacterial isolates to improve other systems. Specifically, bacteria from the rumen of the moose was introduced to young lambs in order to colonize the digestive tract, speed the pace of rumen development, and improve dietary efficiency (see CHAPTER 8). CITATIONS Material from this dissertation has been published in the following form: Ishaq, S.L. and Wright, A-D.G.. (2012). Insight into the bacterial gut microbiome of the North American moose (Alces alces). BMC Microbiology, 12:212. Ishaq, S.L. and Wright, A-D.G.. (2013). Terrestrial Vertebrate Animal Metagenomics, Wild Ruminants. In: Nelson K. (Ed.) Encyclopedia of Metagenomics: Springer Reference. Springer-Verlag Berlin Heidelberg. Ishaq, S.L. and Wright, A-D.G.. (2013). Wild Ruminants. In: Rumen Microbiology – Evolution to Revolution. AK Puniya, R Singh, DN Kamra (Eds). Springer Publishing. Ishaq, S.L. and Wright, A-D.G.. (2014). High-throughput DNA sequencing of the ruminal bacteria from moose (Alces alces) in Vermont, Alaska, and Norway. Microbial Ecology, 68(2):185-195. Ishaq, S.L. and Wright, A-D.G.. (2014). Design and validation of four new primers for next-generation sequencing to target the 18S rRNA gene of gastrointestinal ciliate protozoa. Applied and Environmental Microbiology, 80(17): ahead of print. Material from this dissertation has been submitted for publication to the International Journal of Systemic and Evolutionary Microbiology on December 22, 2014 in the following form: Ishaq, S.L., Reis, D., Lachance, H., and Wright, A-D.G.. (2015). Descriptions of Streptococcus alcis sp. nov. and Streptococcus vermontensis, sp. nov., and proposal of three new subspecies of Streptococcus gallolyticus isolated from the rumen of moose (Alces alces) in Vermont. Material from this dissertation has been submitted for publication to BMC Microbiology on February 23, 2015 in the following form: Ishaq, S.L., Reis, D., and Wright, A-D.G.. (2015). Fibrolytic bacteria isolated from the rumen of North American moose (Alces alces). ii Material from this dissertation has been submitted for publication to Applied and Environmental Microbiology on February 11, 2015 in the following form: Ishaq, S.L., Sundset, M.A., Crouse, J., and Wright, A-D.G. 2015. High-throughput DNA sequencing of the moose rumen from different geographical location reveals a core ruminal methanogenic archaeal diversity and a differential ciliate protozoal diversity. iii ACKNOWLEDGEMENTS First and foremost, I want to thank my advisor and mentor, Dr. André-Denis Wright, without whom this doctoral project would not have happened. Thanks for supporting me professionally and personally, and especially for endlessly correcting my poor spelling, putting up with my dry sense of humor, and helping me to mature into a professional. Thanks to my other committee members, Dr. John Barlow, Dr. Doug Johnson, Dr. Stephanie McKay, and Dr. Neil Sarkar for their interest in my work, their constructive criticism, helpful suggestions, and for reading every page of this tome. The entire Animal Science Department has my gratitude for too many reasons to list here, but I would like to thank Jane O’Neil and Helen Maciejewski for all of the paperwork they’ve done and phone calls they’ve made on my behalf. I’d like to thank the Wright lab members: the undergraduates who were eager to learn laboratory skills and help with my project, Dr. Benoit St-Pierre and Rachel P. Smith, M.S. for their assistance in technical problems and discussions on analysis, Laura Cersosimo for being my sounding-block and partner in science, and Christina Kim for helping me collect rumen samples from sheep we had to chase down in a field in the hot July sun because literally no one else was available. I’d like to thank my collaborators and those individuals who assisted me in sample collection, as moose parts are not as easy to come by as one would think. iv On a more personal note, I’d like to thank my family for their heroic attempts to understand my work and their unfailing support for my nine grueling years of higher education. I especially need to thank my parents, Jill and Tony Pellegrini, for instilling in me a love of reading, organization, and learning, as those have motivated me and proved to be essential while writing this. I’d like to thank my friends, especially Laura Cersosimo, Rachel Smith, Amanda Ochoa, Aimee Benjamin, Mital Pandya, and Mike Haselton, for giving me something to do outside of work, and for always being willing to inevitably talk about work. I want to thank JP Ishaq for his support for so many years, even when I didn’t make it easy, and for putting up with me smelling like a wide variety of animals after work. I’d like to thank everyone who lent a proverbial shoulder to cry on during this last year, and especially for using those shoulders to move all my stuff multiple times up and down stairs. I want to thank Lee Warren for his support in the last and most stressful year of my Ph.D., especially when the only thing to do was to hand me chocolate and hope for the best, and for being a willing participant in my adventures. Finally, I would like to thank the Windows 7 version of Word for its help formatting this beast, and for not being the Windows 8 version of Word. v TABLE OF CONTENTS CITATIONS…………………………………………………………………………..…..ii ACKNOWLEDGEMENTS…………………………………………...………………….iv LIST OF FIGURES…………………………………………………………………….xiii LIST OF TABLES…………………………………………………………..…………..xv CHAPTER 1 COMPREHENSIVE LITERATURE REVIEW ................................ 1 1.1. Moose .................................................................................................................. 1 1.1.1 Ecology and anatomy ...................................................................................... 1 1.1.2 Diet and Nutrition ........................................................................................... 3 1.2. Microbial phylogeny and the small-subunit rRNA genes................................... 5 1.3. An overview of the rumen microbiome and its role in digestion ....................... 8 1.3.1 Bacteria ........................................................................................................... 8 1.3.2 Archaea and methanogenesis ........................................................................ 11 1.3.3 Protozoa ........................................................................................................ 14 1.3.4 Fungi ............................................................................................................. 16 1.3.5 Dietary components and volatile fatty acids ................................................. 17 1.4. Probiotics and manipulation of the rumen environment ................................... 22 1.5.
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