COMPARATIVE STUDY OF DISINFECTANT EFFICACY OF BLEACH, PHENOL AND ETHANOL ON PSEUDOMONAS AERUGINOSA AND STAPHYLOCOCCUS AUREUS

ABSTRACT

The antibacterial effectiveness of three selected disinfectants (household bleach, phenol and ethanol) was determined using agar well diffusion method. Phenol coefficient was used to compare the effectiveness of the disinfectants against phenol under experimental conditions. The test organisms include Staphylococcus aureus and Pseudomonas aeruginosa isolated from both clinical and environmental samples. The undiluted concentrations of the disisnfectants showed varying zones of inhibition against the test organisms, on Staphylococcus aureus it ranged from 41.00±2.83mm (bleach) to 46.50±4.95mm (phenol), and on Pseudomonas aeruginosa it ranged from 40.00±0.00mm (bleach) to 43.50±2.12mm (phenol). No activity was observed with the absolute ethanol. The diluted concentrations of the other disinfectants also showed varying zones of inhibition, On Staphylococcus aureus the mean zone diameter range from 10.00±0.00mm with 38%v/v bleach to 43.50±2.12mm with 75%v/v phenol. Pseudomonas aeruginosa exhibited inhibitions with zone diameter as low as 15.00±0.00mm with 28.5%v/v bleach to as high as 41.00±1.41mm with 75%v/v of phenol. The phenol coefficient test also showed varying results with bleach being the most efficient on the test organisms. The result qualifies these chemical solutions a good disinfectant hence their use externally as a sterilizing agent should be encouraged to avoid contraction of related infections caused by these test organisms.

TABLE OF CONTENTS

Title page                                                                                                                                          i

Certification                                                                                                                                     ii

Dedication                                                                                                                                       iii

Acknowledgement                                                                                                                          iv                                                                                                                                           

Table of contents                                                                                                                              v                                                                                                                             

List of tables                                                                                                                                 viii

List of figures                                                                                                                                  ix                                                                                                                                

Abstract                                                                                                                                           x                                                                                                                     

CHAPTER ONE

1.0  Introduction                                                                                                                                1

1.1 Aim of study                                                                                                                              3

1.2 Objectives of the study                                                                                                               3

CHAPTER TWO

2.0 Literature review                                                                                                                        4

2.1 Brief history of disinfectants                                                                                                      7

2.2 Disinfecting agents                                                                                                                     8

2.2.1 Alcohols                                                                                                                                  8

2.2.2 Halogens                                                                                                                                 9

2.2.3 Phenols and Phenolics                                                                                                           10

2.3 Factors affecting the efficacy of disinfection and sterilization                                                11

2.3.1 Number and location of Microorganisms                                                                             11

2.3.2 Innate Resistance of Microorganisms                                                                                   11

2.3.3 Concentration and potency of disinfectants                                                                          12

2.3.4 Physical and chemical factors                                                                                               13

2.3.5 Organic and inorganic matter                                                                                                14

2.3.6 Duration of exposure                                                                                                             14

2.3.7 Biofilms                                                                                                                                15

2.4 Test organisms                                                                                                                         16

CHAPTER THREE

3.0 Materials and methods                                                                                                             18

3.1 Source of microorganisms                                                                                                       18

3.3.0 Confirmatory test for isolates                                                                                                18

3.3.1 Gram staining                                                                                                                        18

3.3.2 Catalase test                                                                                                                          18

3.3.3 Coagulase test                                                                                                                       19

3.3.4 Oxidase test                                                                                                                          19

3.4 Test organism suspension                                                                                                        20

3.5 Inoculation of the test organisms                                                                                             20

3.6 Preparation of the disinfectants                                                                                               20

3.7 Efficacy testing                                                                                                                        20

3.7.1 Pouring of the disinfectant and incubation                                                                           21

3.8 Determination of the phenol coefficient of the disinfectants                                                  21

CHAPTER FOUR

4.0 Results                                                                                                                                      22

CHAPTER FIVE

5.0 Discussion and conclusion                                                                                                       29

5.1 Discussion                                                                                                                                29

5.2 Conclusion                                                                                                                               31

References                                                                                                                                      32

Appendix

                                                LIST OF TABLES

Table 1: Morphological and Biochemical characteristics of the test isolates                                24                               

Table 2: Phenol coefficient test of the disinfectants against the test organisms                            28

                                                    LIST OF FIGURES

Fig. 1: Zone of inhibition of the diluted concentrations of the test disinfectants on S. aureus                               25

Fig. 2: Zone of inhibition of the diluted concentrations of the test disinfectants on P. aeruginosa 26

Fig. 3: Zone of inhibition of the undiluted concentrations of the disinfectants on the test organisms 27

CHAPTER ONE

1.0 INTRODUCTION

The term control of microorganisms refers to the reduction in number or activity of the total microbial load. The principal reasons for controlling micro organisms are as follows; to prevent transmission of diseases and infection, to prevent contamination by the growth of undesirable microorganisms, to prevent deterioration and spoilage of materials by microorganisms(Pelezar et al., 2006; Willey et al., 2008). The control directed at destroying harmful microorganisms is called disinfection. It usually refers to the destruction of vegetative (non-endospore forming) pathogens.

The term most commonly applies to the use of a disinfectant to free an inert surface or substance. When this treatment is directed at living tissue, it is called antisepsis, and the chemical is then called antiseptic. Therefore, in practice the same chemical might be called a disinfectant for one use and antiseptic for another (Tortora et al., 2004; Ryan and Ray 2004). According to W.H.O and Brooks et al., disinfectants are toxic not only for microbial pathogens but for host cells as well and because of this, they can only be used to inactivate microorganisms in the inanimate environment or to a limited extent, on skin surfaces but cannot be administered systemically.

Antiseptics are biocides or products that destroy or inhibit the growth of microorganisms in or on living tissue (e.g. health care personnel hand washes and surgical scrubs); and disinfectants are similar but generally are products or biocides that are used on inanimate objects or surfaces. Disinfectants can be sporostatic but are not necessarily sporicidal (Johnston et al., 2002).

A wide variety of active chemical agents (or “biocides”) are found in antiseptics and disinfectants. Depending on the chemical nature of disinfectants and antiseptics they can be categorized into several groups. They are alcohols, phenolics, halogens, Quaternary

Ammonium Compounds (QACs) and aldehydes. The mode of action of disinfectants and antiseptics differ greatly according to the chemical substance present. The choice of the disinfectant to be used depends on a particular situation. Some disinfectants have a wide spectrum (kill nearly all microorganisms), whilst others kill a smaller range of disease-causing organisms, but are preferred for other properties (they may be non-corrosive, non-toxic, or inexpensive) (Pelczar et.al., 1993).

The content of many chemical agents can be expressed by more than one notation. In dilutions, a small volume of the liquid chemical (solute) is diluted in a large volume of solvent to achieve a certain ratio. In general, solutions of low dilution or high percentage have more of the active chemical (are more concentrated) and tend to be more biocide, but expense and potential toxicity can necessitate using the minimum strength that is effective (Talaro, 2003).

The effectiveness of a given disinfectant can be evaluated using the Phenol co-efficient test which is the best known disinfectant screening test in which the potency of a disinfectant is compared with that of phenol.  The basic principle now widely accepted is that, the antimicrobial efficiency of a disinfectant or an antiseptic is examined at three stages of testing (Pelczar et al., 1993).

The first stage concerns laboratory tests in which it is verified whether a chemical compound or a preparation possesses antimicrobial activity. For these preliminary screening tests, suspension tests are considered. In the second stage of tests, disinfection procedures and not disinfectants are examined. It is determined under which conditions and at which use-dilution for a given application the preparation is active: the tests simulate real-life situations; such tests are carrier tests for the disinfection of materials by submersion and surface disinfection tests. The last stage takes place in the field, and comprises the in-situ tests which examine whether, after a normal

period of use, germs are still killed by the disinfectant solution.

1.1 AIM OF THE STUDY

The aim of the study was to compare the efficacy of three used disinfectant, bleach, phenol and ethanol on two bacterial isolates, Pseudomonas aeruginosa and Staphylococcus aureus; to know if the disinfectants have any antimicrobial activity against the test microorganisms; and to know the susceptibility of the test gram positive and gram negative bacteria to the test disinfectants.

1.2 OBJECTIVE OF THE STUDY

The objectives of the study were;

·       To isolate the test organisms

·       To evaluate the potentiality of the different disinfectants on the test organism

·       To determine the Phenol Coefficient of the disinfectants

·       To compare the effectivity of the different disinfectants using the test organisms

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