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Isolation and Characterization of Novel Bacteria and Bacteriophages from the Haloalkaline Lake Elmenteita Juliah Khayeli Akhwale A thesis submitted in fulfillment for the degree of Doctor of Philosophy in Biotechnology in the Jomo Kenyatta University of Agriculture and Technology 2015 1 DECLARATION This thesis is my original work and has not been presented for a degree in any other university Signature. Date.....................15 June 2015........................ Juliah Khayeli Akhwale This thesis has been submitted with our approval as University Supervisors: Signature Date.....................15 June 2015........................ Prof. Hamadi Iddi Boga Taita Taveta University College, Kenya Signature... ..Date......................31 May 2015........................ Prof. Hans-Peter Klenk Newcastle University, Newcastle upon Tyne, United Kingdom Formerly of DSMZ-German Collection of Microorganisms and Cell Cultures ii DEDICATION To my parents who made great sacrifices to make me who I am today, for their advice and guidance. To all my family members for moral support all through my studies. God bless you all. iii ACKNOWLEDGEMENT This study was carried out under the supervision of Prof. Hans-Peter Klenk in Leibniz-Institute DSMZ- Braunschweig, Germany, Prof. Hamadi Iddi Boga of Taita Taveta University College. I would like to pass my sincere gratitude to both my supervisors for the insightful ideas, guidance, suggestions and encouragement they provided all the way from proposal development to the end of the project. I wish to further express my gratitude to Prof. Hans-Peter Klenk for accepting to host me in Germany, for providing me with excellent working facilities at the DSMZ. I am greatful to the German Academic Exchange Service (DAAD) for awarding me a PhD scholarship to carry out research in Germany and The National Commission for Science, Technology and Innovation (NACOSTI) for authorizing the research. The Kenya wildlife services (KWS) and National Environmental Management Authority (NEMA) are acknowledged for permits and other necessary documents for sample collection in Kenya and shipment of samples to Germany. Special thanks to the Bacteriophage team at DSMZ headed by Dr. Christine Rhode for their guidance and in advising me on experimental procedures. I am highly indebted to Gabrielle Pöter and Marlen Jando for their excellent technical support, guidance and in making my stay in the laboratory fruitful and a success, thank you. My profound appreciation extends to the entire DSMZ community at Stockheim, Braunschweig, Germany for providing a friendly working environment conducive for research and there prompt assistance when conducting this study. I would like to thank Mrs. Anne Kelly Nderitu and Mr. Josephat Muthanga from JKUAT for their assistance with sample collection and preparation. Lastly, all this work was possible because of God's grace. I thank God for good health, wisdom and understanding during this course. iv TABLE OF CONTENTS DECLARATION........................................................................................................ii DEDICATION............................................................................................................ii ACKNOWLEDGEMENT........................................................................................iv TABLE OF CONTENTS ........................................................................................... v LIST OF TABLES.....................................................................................................ix LIST OF FIGURES ................................................................................................... x LIST OF PLATES ....................................................................................................xii LIST OF APPENDICES.........................................................................................xiii ABBREVIATIONS AND ACRONYMS................................................................xiv ABSTRACT............................................................................................................xvii CHAPTER ONE ........................................................................................................ 1 1.0 GENERAL INTRODUCTION ........................................................................... 1 1.1 PROBLEM STATEMENT................................................................................. 3 1.2 JUSTIFICATION ............................................................................................... 3 1.3 HYPOTHESIS ................................................................................................... 5 1.4 GENERAL OBJECTIVE................................................................................... 5 1.5 SPECIFIC OBJECTIVES .................................................................................. 5 1.6 EXPECTED OUTPUT ...................................................................................... 5 CHAPTER TWO ....................................................................................................... 6 2.0 LITERATURE REVIEW.................................................................................... 6 2.1 INTRODUCTION ............................................................................................. 6 2.2 STRATEGIES FOR ISOLATING NOVEL BACTERIA .................................. 7 2.3 PHYSICO-CHEMICAL PROPERTIES OF SODA LAKES............................. 8 2.4 PREVIOUS STUDIES ON SODA LAKES ...................................................... 9 2.4.1 Bacterial diversity in Kenyan soda lakes................................................... 9 2.4.2 Mycological diversity in Kenyan soda lakes ............................................11 2.4.3 Archael Diversity in Kenyan soda lakes...................................................11 2.4.4 Phage Diversity in Kenyan soda lakes..................................................... 12 CHAPTER THREE ................................................................................................. 13 3.0 BELLIELLA KENYENSIS SP. NOV., ISOLATED FROM THE v HALOALKALINE LAKE ELMENTEITA IN THE AFRICAN RIFT VALLEY ....................................................................................................................................13 ABSTRACT...........................................................................................................13 3.1 INTRODUCTION............................................................................................14 3.2 SAMPLING SITE ............................................................................................14 3.3 MATERIALS AND METHODS......................................................................15 3.4 RESULTS AND DISCUSSION.......................................................................17 3.5 CONCLUSION ................................................................................................24 CHAPTER FOUR ....................................................................................................25 4.0 STREPTOMYCES ALKALIPHILUS SP. NOV., ISOLATED FROM SEDIMENTS OF LAKE ELMENTEITA IN THE KENYAN RIFT VALLEY..25 ABSTRACT...........................................................................................................25 4.1 INTRODUCTION............................................................................................26 4.2 ISOLATION AND PHENOTYPIC CHARACTERIZATION.........................27 4.3 CHEMOTAXONOMIC PROCEDURES ........................................................29 4.4 GENETIC AND PHYLOGENETIC ANALYSIS............................................29 4.5 RESULTS AND DISCUSSION.......................................................................30 4.6 CONCLUSION ................................................................................................38 CHAPTER FIVE......................................................................................................40 5.0 NOCARDIOPSIS MWATHAE SP. NOV., ISOLATED FROM THE HALOALKALINE LAKE ELMENTEITA, IN THE AFRICAN RIFT VALLEY ....................................................................................................................................40 ABSTRACT...........................................................................................................40 5.1 INTRODUCTION............................................................................................41 5.2 ISOLATION AND PHENOTYPIC CHARACTERIZATION.........................42 5.3 CHEMOTAXONOMIC PROCEDURES ........................................................43 5.4 GENETIC AND PHYLOGENETIC ANALYSIS............................................44 5.5 RESULTS AND DISCUSSION.......................................................................44 CHAPTER SIX.........................................................................................................56 6.0 ISOLATION, CHARACTERIZATION AND COMPARATIVE ANALYSIS OF BACTERIOPHAGES FROM THE HALOALKALINE LAKE ELMENTEITA, KENYA.........................................................................................56 vi ABSTRACT........................................................................................................... 56 6.1 INTRODUCTION ........................................................................................... 57 6.2 MATERIALS AND METHODS...................................................................... 58 6.2.1 Study site and sampling..........................................................................
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