INVITRO DETERMINATION OF BACTERICIDAL EFFECTS OF GARLIC ON STAPHYLOCCUS AUREUS, STREPTOCOCCUS SPP., ESCHERICHIA COLI AND SALMONELLA SPP.

ABSTRACT

This study was to determine the bactericidal effects of Garlic (Allium sativum) on Staphylococcus aureus, Streptococcus spp. Escherichia coli and Salmonella spp. The fresh Garlic (Allium sativum), in bulbs forms were obtained from Ahieke Market. The test organisms used in this research consisted of two Gram-positive and two Gram-negative bacteria isolates obtained from stool, sputum samples, and skin swabs. The test organisms used included; Escherichia coli, Staphylococcus aureus, Salmonella species and Streptococcus species. Three (3) solvents (hot water, cold water and methanol) were used to get a comparative result. Agar well diffusion method was used to determine the bactericidal effects of the Garlic (Allium sativum) on the test organisms. The methanolic extract of Garlic (Allium sativum) was found to exhibit high degrees of antibacterial activities against the test isolates. This was shown by the clear zones of inhibition produced by the methanolic extract on the test microorganisms. The highest in-vitro antibacterial activity is 16mm which was exhibited by the methanolic extract at the highest concentration of 200mg/ml against Staphylococcus aureus followed by Salmonella species 14(200mg/ml), while no antibacterial activity (0.0mm) was exhibited by the Methanolic extract at the lowest concentration of 50mg/ml against all the test organisms. It was also observed in this study that the Garlic (Allium sativum) extract had both a bacteristatic and bactericidal activity when tested in vitro using methanolic preparation of garlic extract. Therefore, garlic may be used successfully for treating food poisoning causative agent like Staphylococcus aureus. It may be effective on other microbes on gram positive and gram-negative bacteria, so that further in vivo and in vitro studies are necessary. The standard antibiotics (Ciprofloxacin and Chloramphenicol) used as control inhibited the test bacteria and fungi respectively; it had higher inhibitory activity compared to the Garlic (Allium sativum) extracts. More importantly, there is need for detailed scientific study of traditional medical practices to ensure that valuable therapeutic knowledge of some plants is preserved and also to provide scientific evidence for their efficacies.

TABLE OF CONTENTS

Title Page                                                                                                                                  i

Certification                                                                                                                           iii

Dedication                                                                                                                              iv

Acknowledgement                                                                                                                  v

Table of Contents                                                                                                                   vi

List of Tables                                                                                                                          vii

Abstract                                                                                                                                  ix

CHAPTER ONE

1.0       Introduction                                                                                                                1

1.1       Background of Study                                                                                                  1

1.2       Aim of Study                                                                                                              2

1.3       Specific Objectives                                                                                                     2

CHAPTER TWO

2.0       Literature Review                                                                                                       3

2.1       Brief Description of Garlic (Allium sativum)                                                             3

2.2       Brief History of Garlic (Allium sativum)                                                                    3

2.3       Chemical Constituents of Garlic (Allium sativum)                                                     4

2.4       Medicinal Values of Garlic (Allium sativum)                                                             5

2.4.1    Treatment Cardiovascular Disease                                                                             5

2.4.2    Reduces High Blood Pressure/Hypertension                                                             6

2.4.4    Natural Blood Thinner                                                                                                6

2.4.5    Prevents Diabetes                                                                                                       7

2.5       Antimicrobial Properties of Garlic (Allium sativum)                                                 7

2.5.1    Antiviral Effect                                                                                                           8

2.5.2    Antibacterial Effect                                                                                                    8

2.5.3    Adverse Effects of Garlic                                                                                           9

2.6       Classification of Phytochemicals                                                                               9

2.6.1    Tannins                                                                                                                       9

2.6.2    Flavonoids                                                                                                                  10

2.6.3    Organic Acids                                                                                                             10

2.6.4    Phenols                                                                                                                       10

2.6.5    Nitrogenous Compounds                                                                                            10

2.6.6    Terpenoids                                                                                                                  11

2.7       Biology and Pathogenecity of Test Organisms                                                          11

2.7.1    Salmonella                                                                                                                  11

2.7.2    Staphylococcus aureus                                                                                               12

2.7.2.1 Source and Transmission of Staphylococcus aureus                                                  12

2.7.2.2 Diseases Caused by Staphylococcus aureus                                                               13

2.7.3    Escherichia coli                                                                                                          13

2.7.3.1 Diseases Caused by Escherichia coli O157: H7                                                        14

2.7.4    Streptococcus spp                                                                                                       14

2.7.4.1 Streptococcus pneumonia                                                                                           15

CHAPTER THREE

3.0       Materials and Methods                                                                                               17

3.1       Sources of Materials                                                                                                   17

3.2       Sterilization of Materials                                                                                            17

3.3       Media Used                                                                                                                 17

3.4       Preparation of Garlic Extracts                                                                                    17

3.4.1    Preparation of Different Concentration of Extract                                                     18

3.4.1.1 Cold Water Extract                                                                                                     18

3.4.1.2 Hot Water Extract                                                                                                       18

3.4.1.3 Methanol Extract                                                                                                        18

3.4.3    Media Preparation                                                                                                      18

3.4.4    Agar Well Diffusion Method                                                                                      18

3.5       Determination of Antibacterial Activity                                                                    19

3.6       Determination of Minimum Inhibitory Concentration (MIC)                                    19

3.6.1    Agar Well Diffusion Method                                                                                     19

3.6.2    Disc Diffusion Method                                                                                               20

3.7       Determination of Minimum Bactericidal Concentration (MBC)                                    21

CHAPTER FOUR

4.0 Results                                                                                                                              21

CHAPTER FIVE

5.0       Discussion, Conclusion and Recommendation                                                          27

5.1       Discussion                                                                                                                   27

5.2       Conclusion                                                                                                                  29

5.3       Recommendations                                                                                                      30

            References

LIST OF TABLES

CHAPTER ONE

1.0       INTRODUCTION

1.1       BACKGROUND OF STUDY

Herbs and spices are parts of plants from indigenous or exotic origin are essential part of human diet as they improve taste, color and aroma of foods (Venugopal et al., 2009). In addition they act as preservatives in many foods; they also have antioxidant (Karuppiah and Rajaram, 2012) and antimicrobial properties (Singh et al., 2008). Herbs have also been utilized in human and veterinary medicine (Alsaid et al., 2010).

Garlic belongs to a family of Alliaceae and its scientific name is Allium sativum. Other members of the family include onion, leek, shallot and leek. Garlic is widely used in culinary and medicine (Karuppiah and Rajaram, 2012). It has a pungent hot flavor but mellows and improves with cooking. It has been utilized to fight infections such as cold, cough, asthma, diarrhea, flu, headache, sore throat, abdominal discomfort and respiratory tract infections (Shobana et al., 2009).

The antibacterial properties of crushed garlic have been known for a long time. Various garlic preparations have been shown to exhibit a wide spectrum of antibacterial activity against Gram-negative and Gram-positive bacteria including species of Escherichia, Salmonella, Staphylococcus, Streptococcus, Klebsiella, Proteus, Bacillus, and Clostridium. Even acid-fast bacteria such as Mycobacterium tuberculosis are sensitive to garlic. Analysis of steam distillations of crushed garlic cloves performed over a century ago showed a variety of allyl Sulfides which are isolated and identified as the component responsible for the remarkable antibacterial activity of crushed garlic cloves. The compound turned out to be an oxygenated sulfur compound which they termed allicin from the Latin name of the garlic plant, Allium sativum (Rahman, 2001).

Allicin is the main biologically and biochemically active component of freshly crushed garlic which can easily be decomposed under the influence of heat and time into stable compounds. 2-propenesulfenic acid and tioacrolein are two important molecules derived from garlic, which in turn result in the formation of greater molecules includ­ing diallyl trisulfide and dithiin (Guo et al., 2012). Generally, garlic is a potential antibiotic and shows an effective performance against the bacteria resistant to pharmaceutical antibiot­ics. In 1970’s, Europeans tested the garlic extract on 10 spe­cies resistant to bacteria and yeasts and found it very re­sistant to Salmonella, S. aureus, Mycobacterium and Proteus species (Yin et al., 2002).

Food borne pathogens are widely distributed in the environment and may be a significant cause of mortality and morbidity in the population (Indu et al., 2006). Escherichia coli (EHEC) is a significant foodborne hazard in many countries around the world. Infection often causes haemorrhagic diarrhoea, and occasionally to kidney failure and death. Salmonella is another bacterium that is the cause of foodborne illness mainly from foods of animal origin throughout the world. Staphylococcus aureus and Bacillus cereus cause foodborne illness due to their ability to form heat stable toxins (WHO, 2007).

1.2       AIM OF STUDY

To determine the effect of garlic (Allium sativum) on selected pathogenic microorganisms (Escherichia coli, Salmonella spp, Staphylococcus aureus, and Streptococcus species)

1.3       SPECIFIC OBJECTIVES

·       To isolate test organisms from clinical samples (stool, sputum and skin swab)

·       To determine the antibacterial effects of garlic (Allium sativum) extract on selected pathogens

·       To determine the MIC of the garlic (Allium sativum) extract on selected pathogens

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