DIVERSITY AND BIOGEOGRAPHY OF FUNGI WITHIN THE HOT SPRINGS OF SODA LAKES IN KENYA ODILIA ATAMBA SALANO DOCTOR OF PHILOSOPHY (Biotechnology) JOMO KENYATTA UNIVERSITY OF AGRICULTURE AND TECHNOLOGY 2020 Diversity and Biogeography of Fungi within the Hot Springs of Soda Lakes in Kenya Odilia Atamba Salano A Thesis Submitted in Fulfillment for the Degree of Doctor of Philosophy in Biotechnology in the Jomo Kenyatta University of Agriculture and Technology 2020 DECLARATION This thesis is my original work and has not been presented for a degree in any other University. Signature……………………………. Date…………..…… Odilia Atamba Salano This thesis has been submitted for examination with our approval as the university supervisors Signature………………… Date……………….. Prof. Hamadi Iddi Boga Ministry of Agriculture and Irrigation, Kenya Signature…….. ………………………. Date…17/09/2020…….. …… Dr. Remmy Kasili, PhD JKUAT, Kenya Signature……………………………. Date…………..…… Dr. Huxley Mae Makonde, PhD TUM, Kenya ii DEDICATION This work is dedicated to my family. My dear parents; Mrs. Wilfrida Tero and the late Ernest Tero Lihanda, dear husband Fred Manoa Salano, children; Debbie Elabonga, Becken Joseck and Gullit Nelson Salano. Without your encouragement and support this journey would have been long and tough. You put your heart in all that I did, supported, facilitated, encouraged and prayed for me. iii ACKNOWLEDGEMENT I thank God Almighty for bringing me this far from the commencement of this long and challenging journey. I wish to thank the following institutions for their assistance in this research. The National Commission for Science, Technology and Innovation (NACOSTI), for a research grant and permit; Kenya Wildlife Service (KWS) for access permit to the sampling area in Lake Magadi, Lake Bogoria, Lake Elmenteita and Little Magadi; and Tata Chemicals Ltd for providing invaluable research guidance. My most sincere gratitude goes to my supervisors Prof Hamadi Boga, Dr. Remmy Kasili and Dr. Huxley Mae Makonde. I express more gratitude to the director Institute for Biotechnology Research, JKUAT Prof. B. Nyende for granting me access to the institutions facilities to do my research. No words can describe the appreciation I have for my dear friend and mentor, Dr. Anne Kelly Kambura for her encouragement, counseling and great contribution towards the preparation of my two manuscripts and overall writing of this thesis. Thank you for your assistance particularly in using QIIME and R programming. Particular recognition goes to my colleagues, Grace Mungai, Amanuel Menghs, Johnstone Neondo and Richard Wekesa all of the Institute for Biotechnology Research at JKUAT, for their enormous support in the laboratory and Dr. June Jeophita Mwajuma of the Technical University of Mombasa (TUM). Special appreciation goes to my family friend Edward Nderitu, for his much contribution towards this success and more especially in PowerPoint presentations, Mr. Rotich, for his intellectual ability and ensuring exact sampling techniques were achieved for this research. iv Exceptional appreciation goes to my loving husband Fred Manoa Salano for encouraging me to go for this degree, providing financial support and for accompanying me during sampling in the four soda lakes. MAY GOD BLESS YOU ALL! v TABLE OF CONTENTS DECLARATION .............................................................................................................. ii DEDICATION ............................................................................................................... III ACKNOWLEDGEMENT ............................................................................................ IV TABLE OF CONTENTS ............................................................................................. VI LIST OF TABLES ..................................................................................................... XIII LIST OF FIGURES ....................................................................................................XVI LIST OF APPENDICES ............................................................................................XIX LIST OF ABBREVIATIONS AND ACRONYMS ..................................................XXI ABSTRACT ............................................................................................................. XXIII CHAPTER ONE .............................................................................................................. 1 INTRODUCTION ............................................................................................................ 1 1.1 General Introduction ............................................................................................... 1 1.2 Formation of soda lakes .......................................................................................... 2 1.3 Soda lakes in Africa ................................................................................................ 3 1.4 Distribution of soda lakes in Kenya ........................................................................ 5 1.5 Biogeography .......................................................................................................... 8 1.6 Statement of the problem ........................................................................................ 9 vi 1.7 Justification ............................................................................................................. 9 1.8 Hypotheses ............................................................................................................ 10 1.9 Objectives .............................................................................................................. 10 1.9.1 General objective ............................................................................................ 10 1.9.2 Specific objectives .......................................................................................... 11 CHAPTER TWO ........................................................................................................... 12 LITERATURE REVIEW ............................................................................................. 12 2.1 Microbial diversity of soda lakes .......................................................................... 12 2.2 Microbial diversity and ecology of the soda lakes in Kenya ................................ 12 2.3 Fungal diversity in soda Lakes in the world ......................................................... 14 2.4 Influence of Salinity, pH and Temperature on Fungal diversity ........................... 18 2.5 Fungi in Biotechnology ......................................................................................... 18 2.6 Microbial communities in Hot springs .................................................................. 20 2.7 Culture dependent and culture independent methods for studying fungal diversity ...................................................................................................................................... 21 CHAPTER THREE ....................................................................................................... 23 GENERAL MATERIALS AND METHODS ............................................................. 23 3.1 Research authorization .......................................................................................... 23 vii 3.2 Study sites ............................................................................................................. 23 3.3 Measurements of physicochemical parameters at sampling sites ......................... 24 3.4 Sample Collection ................................................................................................. 26 3.5 Preparation of culture media ................................................................................. 26 3.6 Isolation of fungi from sediments and mats .......................................................... 26 3.7 Morphological characterization of fungal isolates ................................................ 27 3.8 Physiological characterization of fungal isolates .................................................. 27 3.8.1 Determination of optimum temperature for growth ........................................ 27 3.8.2 Determination of optimum pH for growth ...................................................... 27 3.8.3 Growth on different media .............................................................................. 28 3.8.4 Determination of optimum salt concentration for growth .............................. 28 3.9 Enzymatic characterization of fungal isolates ....................................................... 28 3.9.1 Determination of amylase activity .................................................................. 28 3.9.2 Determination of esterase activity................................................................... 29 3.9.3 Determination of lipase activity ...................................................................... 29 3.9.4 Determination of protease activity .................................................................. 29 3.9.5 Determination of cellulase activity ................................................................. 29 3.9.6 Determination of xylanase activity ................................................................. 30 viii 3.10 Molecular characterization .................................................................................. 30 3.10.1 Total DNA extraction ..................................................................................