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Detection of Zoonotic Bacterial Pathogens in Multiple Hosts Using a Microbiome Approach and Molecular Characterization of Anaplasma Phagocytophilum

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Detection of Zoonotic Bacterial Pathogens in Multiple Hosts Using a Microbiome Approach and Molecular Characterization of Anaplasma Phagocytophilum Detection of zoonotic bacterial pathogens in multiple hosts using a microbiome approach and molecular characterization of Anaplasma phagocytophilum A thesis submitted in fulfilment of the requirements for the degree Doctor Philosophiae in the Department of Veterinary Tropical Diseases, Faculty of Veterinary Science, University of Pretoria By AGATHA ONYEMOWO KOLO SUPERVISOR: PROF MARINDA OOSTHUIZEN CO-SUPERVISORS: PROF KELLY BRAYTON DR NICOLA COLLINS July 2019 Declaration I hereby declare that this thesis which I submit for the degree Philosophiae Doctor at the Department of Veterinary Tropical Diseases, Faculty of Veterinary Science, University of Pretoria, South Africa is my original research work and has not been submitted by me to any institution or University for the conferment of an award. ____________________ Agatha Onyemowo Kolo July 2019 i Dedication This thesis is dedicated to the memory of my Father, Sir Augustine Ola Abah who passed away to be with our Lord last year, thank you Daddy for all your selfless love and support, and all the sacrifice you went through to make my life better, thank you for believing in the education of the girl child. Thank you for instilling in me the belief that nothing is impossible, if you can dream it, you can achieve it! ii Acknowledgements I wish to extend my heartfelt gratitude and appreciation to the following people for the dedication, service and impact they have made upon my life and career during the course of this project. First to God Almighty, my rock, my fortress, my help in time of need. You are the vine and I am your branch, cut off from you I wither and dry. To my Husband Francis Babaman, the love of my life, the one who holds my heart, my greatest cheerleader, the General! Kolo thank you for loving me deeply, madly, passionately. Thank you for dedicated hours drawing maps upon maps on QGIS. To my Kids, Daisy Kaka, Sophie Hauwa and Daniel Nmadu, my precious ones whom I adore, my not so little angels anymore thank you for loving mummy unconditionally and unreservedly and being a great blessing to my life. To my Supervisor, Prof Marinda Oosthuizen, a true description of a mentor: kind, supportive and selfless who has guided and moulded me to be a better scientist, to think outside the box, I am so grateful for all the support you have rendered to me. I have really learnt a lot from you. When I grow up, I want to be just like you! To my Co-supervisor, Prof Kelly Brayton, who challenges me to critical thinking whose wry humour brings out the best of every situation, whose kindness, assistance and support enveloped me during the course of this journey, thank you for the guidance. To my Co-supervisor, Dr Nicola Collins, thank you for the being the thorough researcher that you are. For the expertise and keenness, you displayed during the duration of this project, I am deeply grateful. To my darling Mom Christiana Ada Abah, my siblings: Angela Ogodo, Constance Onyego, Augustine Ola, James Ode, Francis Adah, Fatima Ene, John Ejembi and Emmanuella Anyaoyi, thank you for loving me from my beginning through thick and thin, through lack and plenty, through valleys and mountain tops, you were all there. To my Mum-in-law Mrs Christiana Kaka Kolo, Brothers in law: Emmanuel, Moses, and Nathaniel Kolo, Sisters in law, nieces and nephews thank you for your support, Indeed Family is everything. iii To the support staff at the Department of Veterinary Tropical Diseases, Mrs Milana Troskie, Ms Ilse Vorster and Ms Anna-Mari Bosman. Thank you for lending a helping hand anytime I popped by your office. To Dr Charles Byaruhanga and Dr Zama Khumalo thank you for your help in sample collection from rodents and all the adventures in the Kruger and Dr Charles for running models used for phylogenetic analysis. To the University of Pretoria, for the PhD Research Support bursary and the travel abroad exchange bursary to undergo training at the Washington State University, Pullman, USA. The expertise I have garnered I will treasure for a lifetime. To the South African National Research Foundation (NRF) (grants 92739, 110448 and 109350 to Marinda Oosthuizen), the University of Pretoria Institutional Research Theme on Animal and Zoonotic Diseases grant (awarded to Marinda Oosthuizen), and the Belgian Directorate General for Development Co-operation Framework (ITM/DGCD): Thank you for the research funds allocated to this project. I acknowledge Dr Erich Zweygarth (Freie Universitat, Berlin, Germany) for providing control DNA, and Prof Sonja Matthee (Stellenbosch University, South Africa) and Prof Luis Neves (University of Pretoria) for expertise provided with wild rodent trapping. I acknowledge Ms. Jeanette Wentzel of the Hans Hoheisen Wildlife research station, University of Pretoria for the logistic support she rendered throughout my research visits to the Mnisi community area. I also acknowledge the technical assistance and support of Derek Pouchnik and Mark Wildung of the Genomics Sequencing core at Washington State University, Pullman, USA. I acknowledge Prof Lucille Blumberg, Prof John Frean and Dr Jennifer Rossouw of the National Institute of Communicable Diseases (NICD) Johannesburg, South Africa for the blood samples collected from febrile patients that was used in this study. I thank Prof Celia Abolnik of the Faculty of Veterinary Science, University of Pretoria for the use of the BSL-3 laboratory at the Poultry Research unit. I also acknowledge Prof Armanda Bastos of the Department of Zoology & Entomology, University of Pretoria for the molecular profiling of the rodents used in this study. I thank Prof Eric Etter of the Department of Production Animal Studies, Faculty of Veterinary Science, University of Pretoria for the assistance and insights he provided into the statistical analysis of microbiome data. I acknowledge the environmental monitors of the Mnisi community Programme University of Pretoria who assisted with sample and ectoparasite collection from dogs and cattle. I acknowledge Estelle Mayhew for the production of the map of the study area. To the staff and all postgraduate students of the Department of Veterinary Tropical Diseases, thank you for the conducive environment, the support and the laughter you have provided during the years of my stay at the Department. iv Table of contents Declaration .............................................................................................................................................. i Dedication .............................................................................................................................................. ii Acknowledgements ............................................................................................................................. iii Table of contents .................................................................................................................................. v List of Tables .................................................................................................................................... xviii List of Abbreviations ......................................................................................................................... xix Thesis Summary ................................................................................................................................ xxi CHAPTER 1 ............................................................................................................................................ 1 General Introduction ............................................................................................................................. 1 1.1 Introduction ..................................................................................................................................... 1 1.2 Research Hypothesis...................................................................................................................... 2 1.3 Research aims and objectives ....................................................................................................... 2 1.4 Thesis overview .............................................................................................................................. 3 1.5 References ....................................................................................................................................... 5 CHAPTER 2 ............................................................................................................................................ 7 Literature Review .................................................................................................................................. 7 2.1 Microbiome analysis ....................................................................................................................... 7 2.1.1 Methods to study microbial communities ................................................................................... 7 2.1.2 Statistical approaches in community analyses ........................................................................... 9 2.1.3 Tick microbiome studies ............................................................................................................. 9 2.1.4 Other microbiome studies ........................................................................................................ 10 2.2 Acute febrile illness ...................................................................................................................... 11 2.2.1 Anaplasma
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