ISOLATION AND IDENTIFICATION OF MICROORGANIMS FROM PALM WINE SOLD IN UMUAHIA

ABSTRACT

A total of ten (10) fresh palm wine samples were collected at guided random from local palm wine tappers in Umuahia, Abia State Nigeria. A total of 29 microbial species made up of 21 bacterial species and 8 fungal species were isolated from fresh palm wine samples. The said isolates includes Bacillus spp (2), Staphylococcus aureus (3), Escherichia coli (4), Lactobacillus spp (6), Micrococcus spp (2), Serratia spp (4), Saccharomyces spp (5), and Candida spp (3). The total viable microbial counts of fresh palm wine samples ranged from 2.04×103cfu/g to 7.71×103cfu/g, total coliform plate count (TCPC) from 2.04×103cfu/g to 5.37×103cfu/g, total lactic acid bacterial plate count (TLPC) from 5.44×103cfu/g to 7.71×103cfu/g, total bacterial plate count (TBPC) from 2.21×103cfu/g to 4.24×103cfu/g and total fungal plate count (TFPC) from 3.45×103cfu/g to 6.16×103cfu/g. From the findings in this study, it was observed that Lactobacillus spp is the most frequently occurring isolates with a percentage occurrence of 6(20.7%), followed by Saccharomyces spp 5(17.2%), then Serratia spp and Escherichia coli 4(13.8%), Candida spp and Staphylococcus aureus 3(10.3%), and finally Micrococcus spp and Bacillus spp 2(2.9%). The presence of these organisms in the wine is an indication of the poor hygienic state of the tappers, materials used and the method of tapping involved. This occurrence is of public health importance as the wine is consumed in almost every part of the country due to its nutritional, health and social significance. There is therefore the need for public awareness in the consumption of these wines to help promote the quality of these products to enhance their nutritional as well as avoiding the health risk that may be associated with the consumption of contaminated palm wine.

TABLE OF CONTENTS

Title Page                                                                                                                                i

Certification                                                                                                                           iii

Dedication                                                                                                                              iv

Acknowledgement                                                                                                                  v

Table of Contents                                                                                                                   vi

List of Tables                                                                                                                          vii

Abstract                                                                                                                                  x

CHAPTER ONE

1.0       Introduction                                                                                                                1         

1.1       Aim and Objectives                                                                                                    2

CHAPTER TWO

2.0       Literature Review                                                                                                       4

2.1       Microbial Communities in Palm Wine                                                                       4

2.2       Biochemical Constituents of Palm Wine                                                                   6

2.3       Nutrient Composition                                                                                                 8

2.4       Derived Products                                                                                                        8

2.5       Health Benefits                                                                                                           8

2.5.1    Electrolytes                                                                                                                 9

2.5.2    Magnesium                                                                                                                 9

2.5.3    Iron                                                                                                                             10

2.5.4    Manganese                                                                                                                  11

2.5.5    Zinc                                                                                                                             11

2.6       Water Soluble Vitamins, Sucrose and Proteins of Palm Sap                                     11

2.6.1    Thiamine                                                                                                                    12

2.6.2    Riboflavin                                                                                                                   12

2.6.3    Sucrose                                                                                                                       12

2.6.4    Protein                                                                                                                        13

2.7       Methods Employed For Identification of Microorganisms In Palm Wine               13

CHAPTER THREE

3.0       Materials and Methods                                                                                               16

3.1       Collection of Samples                                                                                                16

3.2       Sterilization of Materials                                                                                            16

3.3       Preparation of Culture Media                                                                                     16

3.4       Isolation of Bacteria and Yeast                                                                                  16

3.5       Purification of Isolates                                                                                               17

3.6       Identification of the Isolates                                                                                       17

3.7       Identification of Bacterial Isolates                                                                             17

3.7.1    Gram Staining                                                                                                            17

3.7.2    Biochemical Tests                                                                                                      18

3.7.2.1 Catalase Test                                                                                                               18

3.7.2.2 Indole Test                                                                                                                  18

3.7.2.3 Citrate Utilization Test                                                                                               18

3.7.2.4 Hydrogen Sulphide (H₂S) Production Test                                                                19

3.7.2.5 Starch Hydrolysis                                                                                                       19

3.7.2.6 Motility, Indole, Urease (MIU)                                                                                  19

3.7.2.7 Coagulase Test                                                                                                           20

3.7.2.8 Oxidase Test                                                                                                               20

3.8       Identification of Fungal Isolates                                                                                 21

3.8.1    Colonial Morphology                                                                                                 21

3.8.2    Yeast Viability Staining                                                                                             21

3.8.3    Sugar Fermentation                                                                                                    21

CHAPTER FOUR

4.0       Results                                                                                                                        22

CHAPTER FIVE

5.0       Discussion and Conclusion                                                                                        28

5.1       Discussion                                                                                                                   28

5.2       Conclusion                                                                                                                  30

            References

LIST OF TABLES

CHAPTER ONE

1.0       INTRODUCTION

Palm wine is the collective name for a group of alcoholic beverages produced by the natural fermentation of the sap obtained from various tropical plants of the Palmae family (Alcántara-Hernández et al., 2010). Palm wine is an alcoholic beverage that is produced and consumed in different regions of the world The sap of the palm trees, which is originally sweet (Amoa-Awua et al., 2007; Naknean et al., 2010; Santiago-Urbina et al., 2013) serves as a rich substrate for the growth of various types of microorganisms. The sap undergoes spontaneous fermentation, which promotes the proliferation of yeasts and bacteria for the conversion of the sweet substrate into several metabolites mainly ethanol, lactic acid and acetic acid (Amoa-Awua et al., 2007; Stringini et al., 2009; Ouoba et al., 2012; Santiago-Urbina et al., 2013). Palm sap fermentation has been reported to be an alcoholic, lactic and acetic fermentation (Amoa-Awua et al., 2007; Stringini et al., 2009; Ouoba et al., 2012; Santiago-Urbina et al., 2013), therefore, yeasts, lactic acid bacteria and acetic acid bacteria are considered to play the most important role in the palm wine production.

The palm sap is obtained through the process known as tapping, which involves a series of operations to stimulate the flow of sap, such as the perforation of the trunk, insertion of a tube in the hole and collection of the sap in a container (gourd, clay pot, plastic container, glass bottle or calabash) (Ouoba et al., 2012). There are diverse ways of tapping palm trees which depend on the locality. In general, two methods are practiced: in the first method the sap is obtained from a live standing tree, such as the Bandji and Toddy production, This process implicates climbing very tall palm trees, and perforate the trunk in the top of the tree for Bandji production (Ouoba et al., 2012), or cutting into the end of spadix from the tender inflorescence of the palm tree (inflorescence tapping) for Toddy production (Mbuagbaw and Noorduyn, 2012). In the second method, the tree is felled or cut down before tapping (stem tapping), such as palm wine from Ghana and Taberna production. The cessation of the flow of palm sap varies according to the palm tree species and from tree to tree; for instance the shorter duration of tapping could be 2 weeks and the longest 8 weeks (Amoa-Awua et al., 2007; Santiago-Urbina et al., 2013). Palm wine is collected twice a day, normally in the morning and the evening, it can be either immediately consumed or stored for later sale (Amoa-Awua et al., 2007; Naknean et al., 2010; Karamoko et al., 2012; Santiago-Urbina et al., 2013). Palm wine from Ghana is distilled for gin production called Akpeteshie (Amoa-Awua et al., 2007); similarly, Toddy is also distilled to produce the spirit known as Arrack. Tapping process from a live standing palm tree such as Bandji production from Borassus akeassii, has been reported to be different from wine production from others types of palm trees where sap is collected from a live upright tree, as the palm wine from Elaeis guineensis produced in Ghana (Amoa-Awua et al., 2007).

This drink has a significant role in several nutritional, medical, religious and social uses such as traditional wedding ceremonies, traditional religious ceremonies or festivals, prayers and it is good for malaria (Olasupo and Obayori, 2003; Chandrasekhar et al., 2012).

1.1       AIM AND OBJECTIVES

The aim of this study was to evaluate the microbial quality of some locally sold palm wine within Umuahia metropolis.

The specific objectives of this study include:

      i.         The isolation and characterization of the microbial flora of the palm wine samples.

     ii.         The determination of the percentage occurrence of the microbial isolates

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