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Microbial Pollutants in Stagnant Water in Rr Section, Khayelitsha, Western Cape, South Africa

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Microbial Pollutants in Stagnant Water in Rr Section, Khayelitsha, Western Cape, South Africa MICROBIAL POLLUTANTS IN STAGNANT WATER IN RR SECTION, KHAYELITSHA, WESTERN CAPE, SOUTH AFRICA by QENEHELO ALICE LEUTA Dissertation submitted in partial fulfilment of the requirements for the degree Master of Technology: Environmental Management in the Faculty of Applied Science at the Cape Peninsula University of Technology Supervisor: Dr. A.N. Paulse Co-supervisor: Prof. J.P. Odendaal Cape Town April 2015 CPUT copyright information The dissertation may not be published either in part (in scholarly, scientific or technical journals), or as a whole (as a monograph), unless permission has been obtained from the University DECLARATION I, Qenehelo Alice Leuta, declare that the contents of this dissertation/thesis represent my own unaided work, and that the dissertation/thesis has not previously been submitted for academic examination towards any qualification. Furthermore, it represents my own opinions and not necessarily those of the Cape Peninsula University of Technology. Signed Date ii ABSTRACT Greywater is domestic wastewater from daily kitchen, laundry, bath, shower, hand washing practices and does not include wastewater from the toilet. Greywater from informal settlement has been identified as important environmental pollution sources. Inadequate sanitation and poor drainage in informal settlements result in greywater being stagnant at the base of communal taps. This water has a potential to cause health problems to those who come in contact with it. Studies of greywater quality in informal settlements in South Africa tend to concentrate on physico-chemical analysis and microbial indicator organisms. In order to adequately manage greywater in informal settlements there is a need to understand the microbial pathogens present in such water. Therefore this study is aimed at determining the level of microbial contamination of stagnant greywater in the RR Section of Khayelitsha, Western Cape. Six sampling sites were identified and sampling of stagnant greywater was conducted twice a month (from January to May 2013) from the base of six communal taps, which served as the sampling sites. The microbial enumeration techniques employed in this study were the Most Probable Number (MPN) techniques, the Heterotrophic Plate Count (HPC) technique and the Flow Cytometric (FCM) technique. The API 20E and the RapID™ ONE systems were used to identify possible pathogenic Gram-negative microorganisms, while possible pathogenic Gram-positive microorganisms were identified with the BBL Crystal™ Gram Positive (GP) Identification (ID) system. The highest MPN counts were 1.6 x 108 microorganisms/100mℓ recorded at Site A (weeks 3 and 5) as well as at Site B (week 5). The corresponding highest faecal coliform count was 4.7 x 106 microorganisms/100mℓ obtained at Site B (week 5). The highest E. coli count observed was 1.8 x 106 microorganisms/100mℓ recorded at Site A (week 5) and Site F (week 5). In comparison, the highest HPC count was 2.9 x105 microorganisms/mℓ recorded at Site C in week 4. The results obtained by the MPN and HPC techniques were significantly (p < 0.05) higher than the water quality standards by Department of Water Affairs and Forestry (DWAF) (1996a; 1996b) and the SABS (2011). The highest total FCM and viable FCM counts were 3.4 x 107 microorganisms/mℓ and 3.1 x 107 microorganisms/mℓ, respectively recorded at Site A in week 5. The FCM technique displayed significantly (p < 0.05) higher results than both the MPN and HPC techniques, which highlighted its reliability in obtaining more accurate enumeration results. The RapID™ ONE and the API 20E identification systems mostly identified Escherichia coli, Klebsiella pneumonia, K. oxytoca, Acinetobacter baumannii/calcoaceticus and Enterobacter cloacae, while the organisms more commonly identified by the BBL Crystal™ Gram Positive (GP) Identification (ID) system, were the Corynebacterium species, iii and Bacillus cereus. The presence of these organisms raises health concern to the community of RR Section, as some are known to cause waterborne diseases, while others are known to cause nosocomial infections. iv ACKNOWLEDEMENTS I wish to thank: . First and foremost my mother, Makhotso Cecilia Leuta, for instilling in me the value of education. You gave up everything you had in order for me to acquire my education. I hope to pass everything you have taught me on to my son. Dr. Arnelia Paulse, my supervisor, to whom I owe my deepest gratitude for the hard work, dedication, guidance and support. Prof. James Odendaal, my co-supervisor for all the assistance, guidance and provision of laboratory space to conduct my research. Håvard Ovesen, my husband, and Odin Thabo Leuta Ovesen, my son, thank you for being there for me. My husband, for encouraging me to go on even during those moments I wanted to quit. My son, for sacrificing those moments he wanted to be with me in order for me to do my work. Thabo, your inquisitive mind and interest in my work always humbled me and made me creative when explaining what my work is all about. Dustin Kramer, for introducing me to Social Justice Coalition. The Social Justice Coalition, for taking the time to assist and accompany me every time I went sampling. Special thanks to Luthando Tokota of Social Justice Coalition who always accommodated my sampling needs within his busy schedule. He is one of the most reliable people I know, and his dedication to his work serves as a good example to all of us. Thank you for making me understand the complexities around water and sanitation issues in RR Section. Thando Ndlovu, for the assistance provided whenever I need one. My mother and father in law, Gerd Karin and Egil Øivind Ovesen, thank you for your love and constant support. Vincent Zungu, thank you very much for the time you took to assist me to develop my maps and also for the constant support in my research. All the postgraduate students at Cape Peninsula University of Technology – Cape Town campus, who became my friends. They provided emotional support and served as unofficial psychologists throughout this tough journey. Cape Peninsula University of Technology and the financial assistance of the National Research Foundation towards this research is acknowledged. Opinions expressed in this thesis and the conclusions arrived at, are those of the author, and are not necessarily to be attributed to the National Research Foundation. v DEDICATION This dissertation is dedicated to Håvard Ovesen and Odin Thabo Leuta Ovesen, without whom I am nothing. vi TABLE OF CONTENTS DECLARATION……………………………………………………………………………...ii ABSTRACT…………………………..………………………………………………...……iii ACKNOWLEDGEMENTS………………………..…………………………………………v DEDICATION…………………………………..…………………………………….……...vi TABLE OF CONTENTS………………………..…………………………………….……vii LIST OF TABLES ………………………………...……………………..……………….....x LIST OF FIGURES……………………………………………………..…………….……..xi APPENDICES……………………………………………………….……………..……….xiii GLOSSARY…………………………………………………………..………………….….xiv REFERENCE MAP………………………………………………..……………………..…xv CHAPTER ONE: LITERATURE REVIEW 1 1.1 INTRODUCTION 1 1.2 BACKGROUND TO THE STUDY AREA 2 1.3 INFORMAL SETTLEMENTS AND THE PROVISION OF WATER AND SANITATION IN SOUTH AFRICA 5 1.4 THE SITUATION OF GREYWATER IN INFORMAL SETTLEMENTS OF SOUTH AFRICA 7 1.4.1 Greywater 7 1.4.2 Characteristics of greywater 8 1.4.3 Microbial quality and health risks of greywater 8 1.4.4 City of Cape Town greywater guidelines 9 1.5 WATER QUALITY 10 1.6 WATERBORNE PATHOGENS 11 1.6.1 Bacteria 13 1.6.1.1 Salmonella 13 1.6.1.2 Helicobacter pylori 13 1.6.1.3 Vibrio cholerae 14 1.6.1.4 Legionella 14 1.6.1.5 Pseudomonas aeruginosa 14 1.6.1.6 Campylobacter 15 1.6.1.7 Yersinia 15 vii 1.6.1.8 Indicator organisms 15 1.6.1.8.1 Total coliform bacteria 16 1.6.1.8.2 Faecal coliform bacteria 17 1.6.1.8.3 Escherichia coli 17 1.6.1.8.4 Heterotrophic bacteria 18 1.6.1.9 Clostridium spp 19 1.6.1.10 Staphylococcus spp 19 1.6.1.11 Bacillus spp 19 1.6.1.12 Listeria monocytogenes 20 1.6.1.13 The Viable-but-non-culturable (VBNC) state of bacteria 20 1.7 DETECTION, ISOLATION, ENUMERATION AND IDENTIFICATION OF WATERBORNE ORGANISMS 21 1.7.1 Heterotrophic Plate Count 21 1.7.2 Most Probable Number (MPN) technique 21 1.7.3 Flow Cytometric Analysis (FCM) and LIVE/DEAD® BacLight™ Viability Probe 22 1.7.4 API 20E system 23 1.7.5 RapID™ ONE system 23 1.7.6 BBL Crystal™ Gram-positive Identification system 24 1.8 OBJECTIVES OF RESEARCH 25 CHAPTER TWO: MATERIALS AND METHODS 26 2.1 SAMPLING SITES 26 2.2 SAMPLING 26 2.3 ENUMERATION TECHNIQUES 26 2.3.1 Heterotrophic Plate Count 26 2.3.2 Most Probable Number (MPN) technique 28 2.3.3 Flow Cytometric Analysis (FCM) and LIVE/DEAD® BacLight™ Viability Probe 32 2.3.4 Statistical analysis 33 2.4 IDENTIFICATION TECHNIQUES 33 2.4.1 API 20E system 33 2.4.2 RapID™ ONE system 34 2.4.3 BBL Crystal™ Gram-positive Identification system 34 viii CHAPTER THREE: RESULTS AND DISCUSSION 35 3.1 PHYSICAL PARAMETERS 35 3.2 ENUMERATION OF BACTERIAL CONTAMINANTS IN STAGNANT WATER POOLS 36 3.2.1 Most Probable Number (MPN) technique 36 3.2.2 Heterotrophic Plate Count 38 3.2.3 Flow Cytometric Analysis (FCM) and LIVE/DEAD® BacLight™ Viability Probe 42 3.3 IDENTIFICATION OF GRAM-NEGATIVE BACTERIA 51 3.3.1 Identification of isolates by means of the API 20E and RapID™ ONE identification systems 55 3.3.1.1 Health concerns linked to isolated microorganisms 59 3.3.1.2 Comparison of the API 20E and RapID™ ONE systems 64 3.4 IDENTIFICATION OF GRAM-POSITIVE BACTERIA 65 3.4.1 Health concerns linked to isolated microorganisms 68 CHAPTER FOUR: GENERAL CONCLUSIONS AND RECOMMENDATIONS 72 4.1
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