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Bacteriocins of Bovine Non-Aureus Staphylococci

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Bacteriocins of Bovine Non-Aureus Staphylococci University of Calgary PRISM: University of Calgary's Digital Repository Graduate Studies The Vault: Electronic Theses and Dissertations 2017 Bacteriocins of bovine non-aureus staphylococci Carson, Domonique Carson, D. (2017). Bacteriocins of bovine non-aureus staphylococci (Unpublished master's thesis). University of Calgary, Calgary, AB. doi:10.11575/PRISM/25092 http://hdl.handle.net/11023/4124 master thesis University of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission. Downloaded from PRISM: https://prism.ucalgary.ca UNIVERSITY OF CALGARY Bacteriocins of bovine non-aureus staphylococci by Domonique Carson A THESIS SUBMITTED TO THE FACULTY OF GRADUATE STUDIES IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE GRADUATE PROGRAM IN VETERINARY MEDICAL SCIENCES CALGARY, ALBERTA SEPTEMBER, 2017 © Domonique Carson 2017 ii Abstract The non-aureus staphylococci (NAS) species are among the most prevalent isolated from bovine milk and have been reported to inhibit major mastitis pathogens, likely by producing bacteriocins. This thesis is comprised of two sections, focusing on in vitro inhibition assays and in silico identification of bacteriocin gene clusters and bacteriocin resistance genes in NAS and Staphylococcus aureus, using isolates obtained from the Canadian Bovine Mastitis and Milk Quality Research Network. The first part determined the inhibitory capability of 441 bovine NAS isolates (comprising 25 species) against bovine S. aureus and human methicillin-resistant S. aureus (MRSA) and determined the presence of bacteriocin biosynthetic gene clusters in NAS whole genomes. Overall, 40 isolates from 9 species (S. capitis, S. chromogenes, S. epidermidis, S. pasteuri, S. saprophyticus, S. sciuri, S. simulans, S. warneri, and S. xylosus) inhibited growth of S. aureus in vitro; of which, 23 isolates (from S. capitis, S. chromogenes, S. epidermidis, S. pasteuri, S. simulans, and S. xylosus) also inhibited MRSA. 105 putative bacteriocin gene clusters encompassing 6 different subclasses (lanthipeptides, sactipeptides, lasso peptides, class IIa, class IIc, and class IId) in 95 whole genomes from 16 species were identified. The second part of the thesis determined the susceptibility of 139 bovine S. aureus isolates to a bacteriocin producing S. chromogenes isolate and identified and described the distribution of genes potentially associated with susceptibility and resistance in S. aureus whole genomes. Overall, 90 S. aureus isolates (65%) were resistant to inhibition by the S. chromogenes isolate. We identified 77 genes that were associated with an isolate being resistant. We also identified 76 genes that were associated with an isolate being susceptible to the S. chromogenes. Bacteriocin susceptibility and resistance seems to be linked to a large number of genes, the majority of which iii are annotated as hypothetical proteins and will need further assessment to determine their role in S. aureus susceptibility. Overall, bacteriocins may be a potential source of novel antimicrobials and this thesis represents the foundation to explore novel NAS bacteriocins. iv Acknowledgements Firstly, I would like to thank my supervisors Herman Barkema and Jeroen De Buck for all of their patience and support during this project. Thank you to Herman for the opportunity to attend so many conferences and speak about my research. Thank you to Jeroen for all your direction with my projects. I also would like to thank my committee members for their support and Dr. John Kastelic for his edits of my manuscript and for his scientific writing courses. I would also like to thank Uliana Kanevets and Aaron Lucko for their help in the lab. Thank you to Matthew Workentine for all his help with bioinformatics. Thank you to the original CNS crew, Larissa Condas and Diego Nobrega for everything. Larissa, thank you for showing me the CNS ropes, I came into this project with little lab experience and your expertise was essential for my success. Your endless encouragement and honest life talks during your time here were also so appreciated- “Everything is AWESOME”. Diego, thank you for all your help, from whole genome sequencing to talking through absolutely everything with me, I could not have done it without you. I am also so grateful for the rest of the CNS crew that has joined us. Thank you Ali Naqvi for helping me with statistics and thank you Ana Paula Monteiro Alves for being my conference buddy and letting me be the absolute introvert that I am. Lastly, thank you to Sohail Naushad, without you I would have not been able to do this. Thank you for the countless conversations about genes, bioinformatics, and how to interpret my results. Thank you for the coffee dates and the pep talks and for always being on my team. Thank you for all of your help with organizing genomes, creating trees, running jobs, and giving me feedback whenever I ask. v Thank you to all my fellow graduate students. Thank you to Emily Morabito for beginning this program at the same time as me and becoming such a huge support as we figured this all out together. Thank you to Caroline Corbett for being a rock in all of this. You were always there to talk and were always able to provide valuable feedback on my research. Thank you for being such a good friend to me. Also thank you Casey Jacobs for always being there for me (usually with a beer and chocolate ready). Our daily dog walks became so special and without them life would have been much more difficult. I would also like to thank Dr. Keliesha Roth and Amy Stanley for being the best friends a girl could ask for in this life. Keliesha, thank you for always making me feel like I could do this. Thank you for letting me come work at your house, for taking such good care of me after my surgery this spring, for always taking the dogs for me whenever they needed somewhere to go, and for bringing Took into my life. You are one of the most amazing people I’ve ever met and a huge reason that I am the person I am today. Amy, thank you for being the most understanding and amazing human. You supported me through this every way a person possibly could… you ran away to the mountains with me when I needed that, you exercised the dogs for me when I couldn’t, you provided hours of deep conversations about life, and you have always just wanted what is best for me. Thank you to you both. Thank you to Reid Anderson. You came into my life on that airplane at the exact moment I needed you to. You have been my strength in times of weakness, my never ending support system, my inspiration to keep working towards my goals, and (most importantly) the best dog dad I could ever ask for. You mean more to me than you’ll ever know. Lastly, thank you to my family for the endless support and understanding. Thank you for answering all of my phone calls and always giving me words of encouragement. It has been hard vi to be so far apart but it never felt too far. Thank you for instilling in me a passion for learning and for sticking things out. vii Dedication To my family, in particular my parents and my chosen family who have always supported me and to my dogs who have kept me sane. viii Table of Contents Abstract .......................................................................................................................................... ii Acknowledgements ...................................................................................................................... iv Dedication .................................................................................................................................... vii Table of Contents ....................................................................................................................... viii List of Tables ................................................................................................................................ xi List of Figures and Illustrations ................................................................................................ xii List of Symbols, Abbreviations and Nomenclature ................................................................ xiv Preface .......................................................................................................................................... xv Chapter One: General Introduction ........................................................................................... 1 1.1 Mastitis in the Canadian dairy industry ........................................................................... 1 1.2 Non-aureus staphylococci ................................................................................................... 2 1.3 Bacteriocins ......................................................................................................................... 4 1.4 Classification of bacteriocins ............................................................................................. 5 1.4.1 Lanthipeptides ................................................................................................................ 7 1.4.2 Sactipeptides ................................................................................................................ 12 1.4.3 Lasso Peptides .............................................................................................................
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