ISOLATION AND CHARACTERIZATION OF BIOFILM FORMING BACTERIA IN DRINKING WATER AT MALE AND FEMALE HOSTELS IN MICHEAL OKPARA UNIVERSITY OF AGRICULTURE, UMUDIKE

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ABSTRACT


The present study was carried out to determine the prevalence of bacterial organisms isolated from biofilms of water system. Our results clearly demonstrated the overall prevalence raie of the microorganism was 100%. The frequency of occurrence of the bacterial isolates were; Staphylococcus 100%, Streptococcus 73.33%, Enterobacter 66.66%, E. coli 60% and Klebsiella 53.33%. The physiochemical analysis result obtained were all within the WHO standards for drinking water quality, the physiochemical parameters were within acceptable limits. The results were compared with WHO. The investigation shows that the boreholes water at male and female hostels met with WHO Standards for drinking water. Biofilms in drinking water distribution system could provide nutrients for microbial growth, bio systems and proliferation. It is necessary to treat water distribution plants and provide adequate public health education in order to safe guard human health and animal health especially in developing countries.







TABLE OF CONTENTS


Title                                                                                                                                        i

Certification                                                                                                                            ii

Dedication                                                                                                                              iii

Acknowledgement                                                                                                                  iv

Table of contents                                                                                                                    v

List of tables                                                                                                                           ix

List of figure                                                                                                                           x

Abstract                                                                                                                                  xi


CHAPTER ONE: INTRODUCTION

1.1       Background Studies                                                                                                    2

1.2       Aim                                                                                                                             3                                                                                                                                 

1.3       Objectives                                                                                                                   3                                                                                                                                                                     

1.4       Problem Statement                                                                                                     3                                                                                                                                                         

CHAPTER TWO: LITERATURE REVIEW

2.1       Biofilm                                                                                                                        4         

2.2       Development of Biofilms                                                                                           7         

2.3       Environmental Factors Influencing Biofilm Development                                        8

2.3.1    Effect of pH                                                                                                                8

2.3.2    Rheological and Adhesive Properties of Biofilms                                                     9         

2.3.3    Effect of Temperature                                                                                                9

2.4       Biofilms Formation                                                                                                    10

2.4.1    Factors Affecting Biofilms Formation                                                                       11       

2.4.2    Substratum Effect                                                                                                       11       

2.4.3    Conditioning Film                                                                                                      11

2.4.4    Hydrodynamics                                                                                                          12                                                                                                                   

2.4.5    Characteristics of Aqueous Medium                                                                          12

2.4.6    Horizontal Gene Transfer                                                                                           12

2.4.7    Quorum Sensing                                                                                                         13

2.5       Biofilm Structure and Function                                                                                  14

2.5.1    Extracellular Polymeric Substances                                                                           14

2.5.2    Protein                                                                                                                        15       

2.5.3    Interaction of Particles                                                                                                16

2.6       Water                                                                                                                          16

2.6.1    Sources of Water                                                                                                        17

2.6.1.1 Surface Water                                                                                                             17       

2.6.1.2 Ground Water                                                                                                             17       

2.6.1.3 Sources of Water Contamination                                                                               17       

2.7       Bacteria That Can Be Found In Water                                                                       18       

2.7.1    Escherichia Coli                                                                                                         18

2.7.2    Staphylococcus aureus                                                                                               19

2.7.3    Klebsiella Species                                                                                                       19       

2.7.4    Streptococcus Species                                                                                                20

2.7.5    Enterobacter Species                                                                                                  20

2.8       Physiochemical Quality of Water                                                                               21

2.8.1    Temperature                                                                                                                21

2.8.2    Total Dissolved Solids (TDS mg/c)                                                                           21

2.8.3    Hydrogen Ion Concentration                                                                                      21

2.8.4    Alkalinity (Mg/V)                                                                                                       22

2.8.5    Total Hardness                                                                                                            22

2.8.6    Turbidity                                                                                                                     22       

2.8.7    Conductivity                                                                                                               22

2.8.8    Colour                                                                                                                         23

2.8.9    Odour                                                                                                                          23       

2.9       Biofilms in Drinking Water Distribution Systems                                                     23

2.10     Effect of Long Storage of Water on Biofilm Formation                                            24


CHAPTER THREE: MATERIALS AND METHODS

3.1       Study Area                                                                                                                  25

3.2       Collection of Samples                                                                                                25       

3.3       Preparation of Culture Media                                                                                     25

3.4       Bacteria Inoculation and Isolation                                                                              26

3.5       Isolation of Biofilm Producing Bacteria                                                                    26

3.6       Determination of Biofilm Formation of Isolates                                                        27

3.7       Characterization and Identification of Organisms                                                      27       

3.8       Gram Stain                                                                                                                  27

3.9       Biochemical Tests                                                                                                      28

3.9.1    Catalase Test                                                                                                               28

3.9.2    Coagulase Test                                                                                                           28       

3.9.3    Citrate Utilization Test                                                                                               29

3.9.4    Urease Test                                                                                                                 29       

3.9.5    Indole Test                                                                                                                  29

3.9.6    Oxidase Test                                                                                                               30       

3.9.7    Methyl Red Test                                                                                                         30       

3.9.8    Motility Test                                                                                                               30

3.9.9    Sugar Fermentation Test                                                                                            31

3.10     Methods for the Physico-Chemical Analysis of the Water                                        31

3.10.1 pH                                                                                                                               31       

3.10.2 Determination of Conductivity                                                                                  31       

3.10.3 Determination of Temperature                                                                                   32

3.10.4  Determination of Turbidity                                                                                        32

3.10.5  Determination of Dissolved Solids                                                                             32

3.10.6  Determination of Total Suspended Solids (TSS).                                                      32

3.10.7  Determination of Chloride                                                                                          33

3.10.8  Determination Dissolved Oxygen                                                                              33

3.10.9  Determination of Chemical Oxygen Demand                                                                        33

3.10.10 Determination of Nitrate                                                                                           33       

3.10.11 Determination of Phosphate                                                                                      34

3.10.12 Determination of Sulphate                                                                                        34


CHAPTER FOUR: RESULTS                   

4.0       Results                                                                                                                        35


CHAPTER FIVE: DISCUSSION, CONCLUSION AND RECOMMENDATION

5.1       Discussion                                                                                                                   42

5.2       Conclusion                                                                                                                  43

5.3       Recommendation                                                                                                       44

REFERENCES                                                                                                                     45







 

LIST OF TABLES


TABLE           Title                                                                       Page

1:                     Percentage Occurrence of Bacteria Isolates                                       37

2:                     Conforming of Biofilm Formation                                                     38

3:                     Identification and Characterization of Bacteria Isolates                         39

4:                     The Physio-chemical Results of the Water Samples Analyzed.        40


 






LIST OF FIGURES


Figure             Title                                                                         Page

 I:                    Occurrence of Biofilm Suspect On Slip Film Culture                     41

 


 





 

CHAPTER ONE

1.0       INTRODUCTION

Drinking water is an important resource all-round the globe (Martiny et al., 2003), but water borne disease is still a major cause of death in many parts of the world (Fawell and Nieuwenhuijsen, 2003). Nevertheless, little research has focused on identifying the bacteria in water distribution systems (Martiny et al., 2003).

Microorganisms that grow in the environment may enter the drinking water and attach to and grow on drinking water pipes and other surfaces, forming biofilms (WHO, 2014) as the most common means of non-pathogenic microorganisms of source water (Farkas et al., 2013), however, quite a few opportunistic bacterial pathogens naturally occur in aquatic and soil environments are able to persist and grow in biofilms of drinking water systems (Wingender and Flemming, 2011).

The Contamination of drinking water by Escherichia coli that only come from human and animal faecal waste (USEPA, 2009) is an important aspect of drinking water quality (WHO, 2011).                                                                                                                Five groups of Escherichia coli causing diarrhoea in humans and other warm blooded animals have been identified (Brook et al., 1994 and Wasteson, 2001), that include:                                                                                                                                          Enterotoxingenic Escherichia coli (ETEC)                                                                                                                                                             Enteropathogenic Escherichia coli (EPEC)                                                                                                                                                        Enteroaggregative Escherichia coli (EAEC)                                                                                                                                                            Enteroinvasive Escherichia coli ( EIEC)                                                                                                                                                                    Enterohaemorrhagic Escherichia coli ( EHEC). The latter includes shiga toxin (STX) producing Escherichia coli (STX- EC) (Harake et al., 2006).                                                                                                                                                       

Drinking water is essential to sustain life and a satisfactory (adequate, safe and accessible) supply must be available to all (WHO, 2004). The quality of life depends on the key elements such as the quantity and the quality of water available for consumption. A rational use of resources considering the support capacity of the ecosystems in which we live indicates a sustainable society. Strategies such as management and protection of water sources, effective treatment methods, the proper distribution and handling of water are intended to be implemented worldwide.

Trend to increase productivity are evident in water sectors as well as in food industry, in order to provide indispensable consumer products for a continuously growing population, drinking water is not only a liquid for hydration. It interacts with all industrial, agricultural, economic, social and cultural components characterizing the human society (Ancuta, 2012).

Guidelines recommend monitoring of water in order to characterize bacterial counts and identify organisms (Beale et al., 2013). The primary goal for this monitoring is to identify the conditions that promote biofilm growth and observe changes in bacterial trends. It is considered more important to identify changes in the bacterial composition of the biofilm than to identify increased bacterial numbers (Beale et al., 2012).


1.1       Background Studies

Bacteria pathogens can form biofilms in water, especially in different sources of drinking water which include boreholes, streams, wells, storage (geepee) tanks, pots, bowls etc., when kept for a long period of time. These pathogens could enter the water through the following means; soil erosion, rain water fall, wind erosion, rock weathering, human activities and so on. These pathogens have the potency to cause various life-threatening infections.

Drinking water distribution systems have an intake of suspended bacteria from different sources as such the bacteria present in groundwater and raw surface waters which is not removed even after the treatment, the bacteria yielding from the treatment processes such as bio filters and or bacteria which is introduced by water is obstructed from other aquatic environments. Many problems in drinking water distribution system (DWDS) are microbial in nature, including biofilm growth, nitrification, microbial mediated corrosion and the occurrence and persistence of pathogens (Regan et al., 2003; Beech and Sunner, 2004; Emtiazi et al., 2004).

Microbial adhesion to surfaces and biofilm, formation occur in environment on board ships (portable water systems, water treatment facilities and air conditioning).  


1.2       Aim

To analyse, the quality of water supply at both male and female hostels.


1.3       Objectives

Ø  To determine the various kinds of bacteria pathogens that can be found in the water supply.

Ø  To isolate the bacteria identify and characterize them.

Ø  To evaluate if the bacteria isolates are of public health importance.


1.4       Problem Statement

The problems associated with the presence of biofilms in drinking water sources are as follows.

1)    They may be responsible for a wide range of water quality and operational problems.

2)    They can be responsible for loss of distribution system disinfectant residuals, increased bacterial levels, reduction of dissolved oxygen, taste and odour changes.

3)    Microbial influenced corrosion, hydraulic roughness and reduced materials life.

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