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Comparison of Analytical Profile Index, Microscan Walkaway 40 Plus and Bruker Maldi Biotyper Systems for Identification of Enteric Pathogens in Kenya

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Comparison of Analytical Profile Index, Microscan Walkaway 40 Plus and Bruker Maldi Biotyper Systems for Identification of Enteric Pathogens in Kenya COMPARISON OF ANALYTICAL PROFILE INDEX, MICROSCAN WALKAWAY 40 PLUS AND BRUKER MALDI BIOTYPER SYSTEMS FOR IDENTIFICATION OF ENTERIC PATHOGENS IN KENYA RONO FRANKLIN KIPYEGON (Bsc Medical Microbiology) Reg; I56/23061/2011 A thesis submitted in partial fulfilment of the requirements for the award of the Degree of Master of Science (Biotechnology) in the School of Pure and Applied Sciences of Kenyatta University July 2017 ii DECLARATION This thesis is my original work and has not been presented for a degree in any other university or any other award. RONO FRANKLIN KIPYEGON I56/23061/2011 Signature ………………………………….. Date ……………………………… Department of Biochemistry and Biotechnology We confirm that the work reported in this thesis was carried out by the student under our supervision. Supervisors: Dr. David Mburu Department of Biochemistry and Biotechnology Kenyatta University Signature………………………………… Date………………………………… Dr. Brook Danboise Clinical Research Centre/Microbiology Hub Laboratory KEMRI/US Army Medical Research Unit-Kenya, Kericho Signature………………………………… Date………………………………… iii DEDICATION I dedicate this project to my family members: my mother Grace Teigut, my wife Sarah Jepkemboi, my children Sharon Lee Chepkorir, Edrian Kibet and Emanuel Kiplangat. My dedication also goes to my fellow colleagues, supervisors and Walter Reed fraternity as a whole for their continued support, understanding and encouragement during the entire period to completion of the course. iv ACKNOWLEDGEMENT Above all, I thank God for the opportunity, grace and provision throughout the course. I wish to acknowledge my supervisors Dr. David Mburu and Dr. Brooke Danboise for the unreserved direction throughout the development of this proposal, data collection, report writing and presentation just to mention a few. I thank the Director KEMRI/Walter Reed Project for the authorization and the invaluable support that made it possible for me to carry out the study at the facility. I am also grateful to Elizabeth’s contribution, all my colleagues as well as upcoming scientists for their support. Thank you all for the positive criticism. May I also give my gratitude to the following people for their tremendous guidance and inspiration during my entire period of my work: Gilbert Kikwai, Research officer at KEMRI/CDC Virology Laboratory; my brother Hillary Rono, Laboratory Technologist at Matter Hospital; and Josphat Tanui, Microbiologist at National Tuberculosis Reference Laboratory. Thank you for your valuable support which not only made the completion of this research study possible but has left me with a great sense of fulfillment and achievement which will continue to influence my confidence and enthusiasm in research work. To my wife, Sarah Jepkemboi our children Sharon Lee Chepkorir and Edrian Kibet this period has been difficult time for you all and I want to express my appreciation for your love and encouragement. It was certainly not in vain. v TABLE OF CONTENTS DECLARATION ......................................................................................................... ii DEDICATION ............................................................................................................ iii ACKNOWLEDGEMENT .......................................................................................... iv TABLE OF CONTENTS ............................................................................................. v LIST OF TABLES ...................................................................................................... ix LIST OF FIGURES .................................................................................................... ix LIST OF APPENDICES ........................................................................................... xiii ABBREVIATIONS AND ACRONYMS .................................................................. xii ABSTRACT .............................................................................................................. xiii CHAPTER ONE .......................................................................................................... 1 INTRODUCTION ....................................................................................................... 1 1.1 Background information ....................................................................................... 1 1.2 Statement of the problem ....................................................................................... 4 1.3 Justification of the study ........................................................................................ 4 1.4 Research questions ................................................................................................. 5 1.5 Hypothesis.............................................................................................................. 5 1.6 Objectives .............................................................................................................. 5 1.6.1 General objective .................................................................................................. 5 1.6.2 Specific objectives ................................................................................................ 6 CHAPTER TWO ......................................................................................................... 7 LITERATURE REVIEW ............................................................................................ 7 2.1 Diarrhea disease ..................................................................................................... 7 2.1.1 Definition.............................................................................................................. 7 2.1.2 Etiology ................................................................................................................ 7 vi 2.1.3 Enterobacteriaceae………………………………………..……………..…....8 2.1.4 Yeast……………………………………………………………...………...…...9 2.2 Epidemiology ...................................................................................................... 11 2.3 Diagnosis.............................................................................................................. 12 2.4 Analytical profile index ....................................................................................... 14 2.4.1 Analytical profile index 20 E ............................................................................ 15 2.4. 2 Analytical profile index 20 C AUX ................................................................ 15 2.5 MicroScan WalkAway 40 plus ............................................................................ 16 2.6 Matrix-assisted laser desorption ionization–time of flight .................................. 17 2.7 Polymerase chain reaction ................................................................................... 19 2.8 American Type Culture Collection (ATCC)........................................................ 20 2.9 Management ......................................................................................................... 20 2.10 Prevention .......................................................................................................... 21 CHAPTER THREE ................................................................................................... 23 MATERIALS AND METHODS ............................................................................... 23 3.1 Study site .............................................................................................................. 23 3.2 Study design ......................................................................................................... 23 3.3 Sample size determination ................................................................................... 23 3.4 Inclusion criteria .................................................................................................. 24 3.5 Exclusion criteria ................................................................................................. 24 3.6 Sampling technique .............................................................................................. 24 3.7 Methods and techniques of identification ............................................................ 24 3.7.1 Colony morphology ............................................................................................ 26 3.7.2 Gram stain ............................................................................................................ 28 3.7.3 Biochemical characterization ............................................................................ 28 3.7.4 The Analytical Profile Index 20 E (API 20 E) ................................................ 29 3.7.5 The Analytical Profile Index API 20 C AUX ................................................. 30 vii 3.7.6 Serotyping ............................................................................................................ 31 3.7.7 Germ tube test ..................................................................................................... 31 3.7.8 MicroScan WalkAway analysis ................................................................. 32 3.7.9 Matrix-assisted laser desorption ionization–time of flight ........................... 33 3.7.10 Polymerase chain reaction ............................................................................... 35 3.7.11 Determination of turnaround time and cost ................................................... 36 3.8 Data analysis.................................................................................................................
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