ABSTRACT
Ethanol, Bleach and phenolic are three kinds of disinfectants which have been widely used in common laboratories. In this study, the minimum inhibitory concentration (MIC) experiment on these three disinfectants efficiency was conducted against Staphylococcus aureus using agar hold diffusion method. Different concentrations of bleach (1%, 2% 3%, 4% and 5%) were used on Staphylococcus aureus. Also (50%, 60%, 70%, 85% and 95%) of ethanol as well as (5%, 10%, 20%, 25%, and 30%) phenolic were used. Differences in concentrations tested was because, the original concentrations of the disinfectants differs. After 24 hours of incubation at 370c, the results showed that all the disinfectants inhibited the growth of the test organism in their concentrated forms. The diameter of zone of inhibitions was measured around each well by using a ruler in millimeters, using different concentrations, their efficacies varied. The results showed that 30% phenolic had the best efficiency against the test organism (Staphylococcus aureus) and showed the (MIC) value at 5%, and 5% bleach had a better effect on the test organism than ethanol and showed the MIC value at 4%, ethanol showed least sensitivity. 70% concentration of ethanol gave the highest effect on the test organism, while at 60%conc. of ethanol showed the minimum inhibitory concentration (MIC).
TABLE OF CONTENTS
Title page i
Certification ii
Dedication iii
Acknowledgements iv
Table of Contents v
List of Table vi
List of Figures viiAbstract
viii
CHAPTER
ONE: INTRODUCTION
1.1 Background
of study 1
1.2 Objectives
of the Study 3
1.3 Statement
of Problem 4
1.4 Justification
4
CHAPTER
TWO: LITERATURE REVIEW
2.1 General
Concept 5
2.1 History
of Disinfections 6
2.2 About
Disinfectants 7
2.3 Types
of Disinfectants 7
2.4 Properties
of a Disinfectant 13
2.5 General
Features of Disinfectants 14
2.6 General
Features of the Test Organism 15
2.7 Mechanism
of Actions of Disinfectants against Bacteria 15
2.8 Resistant
Action of Bacteria 16
2.9 Advantages
and Disadvantages of Disinfectants 17
CHAPTER
THREE: MATERIAL
AND METHOD
3.1 Collection
of Materials and Sample 21
3.2 Preparation
of Media 22
3.3 Identification
of Isolates 23
3.4 Biochemical
Test 24
3.5 Antibacterial
Activity Testing 26
3.5.1 Preparation
of Disinfectants 26
3.5.2 Antimicrobial
Susceptibility Testing (Using Kirby Bauer Diffusion Assay Well Method.
3.6 Determination of MinimumInhibitory
Concentration (MIC) Using the Broth Dilution Method 28
CHAPTER
FOUR
4.0 RESULTS 30
CHAPTER
FIVE
5.0 Discussion 34
5.1 Recommendations
37
5.2 Conclusion 37
REFERENCE
APPENDIX 1
APPENDIX
I
APPENDIX
III
LIST OF TABLES
Table Title Page
3.5.1 Preparation
of disinfectant 26
3.5.1 Tube
concentration 27
4.1 Results
of identification tests 30
4.2 Results of diameter of zone of inhibition of
ethanol, phenol and bleach for
Staphylococcus aureus. 31
4.3Staphylococcus
aureus response to ethanol, phenol and bleach 32
4.4 Minimum inhibitory
concentrations of ethanol phenol and bleach against
Staphylococcus aureus 33
LIST OF FIGURES
Figure Title
Page
1: Plate
showing zone of inhibition 50
2: MacConkey media with colonies of Staphylococcus aureus50
3: Some of the used plates 51
4: MIC result tubes 51
CHAPTER ONE
INTRODUCTION
1.1 Background of study
Disinfectant are used extensively in hospitals and other
health care setting for a variety of topical and hard-surface applications .in
particular, they are an essential part of infection control practice and aid in
the prevention of nosocomia infections. Mounting concern over the potential for
microbial contamination and infection risks in the food and general consumer
markets have also led to increased usage of disinfectants by the general
public. Disinfectants generally are products or biocides that are used on
inanimate objects sporicidal (Johnston et
al; 2002).
A
wide variety of active chemical agents (or “biocides”) are found in antiseptic
and disinfectants. Depending on the chemical nature of disinfectants they can
be categorized into several groups.they are aicohols, phenolics, halogens,
Quaternary Ammonium Compounds (QACs) and aldehydes. The mode of action of
disinfectants differs greatly according to the chemical substances present. The
choice of the disinfectant to be used depends on a particular situation. Some
disinfectant have a wide spectrum (kills nearly all microorganisms), whilst
others kills a smaller range of disease- causing organisms, but are preferred
for other properties (they may be non corrosive, non toxic, or inexpensive)
(pelczeret al, 1993).
The basic principle now widely accepted is
that, the antimicrobial efficiency of a disinfectant 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 posses
antimicrobial activity. For these preliminary screening test, suspension test
are considered. In the second stage of test, 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
stimulate real-life situations; such tests are carrier tests for the
disinfection of material by submersion and surface disinfection tests. The
last stage takes place in the field, and
comprises the in-situ tests which examines whether, after a normal period of
use, germs are still killed by the disinfectant solution.
There are many standards to follow in
carrying out efficiency tests for disinfectant; American standards; European
standard; ISO standards etc. in Sri Lanka the efficiency of disinfectants is
tested using SLS 688: 1985 (Prescribe the requirements and methods of sampling
and test for disinfectants).
Staphylococcus
aureus is the leading cause of human infections in
the skin and soft tissues bones andjoints, abscesses and normal heart valves. Staphylococcusaureus occur in 40-50% of
humans (BhatiaandIcchpujani, (2008). It flourishes in the hospital setting, producing
blood stream and surgical wound infections. Staphylococcus
aureus is now a huge burden for most healthcare institutions around the
world and is by far the most significant antibiotic-resistant hospital acquired
pathogen we have ever encountered. (Earssar, 2005).
Staphylococcus aureus is usually spread
by direct physical contact with those already infected or through indirect
contact by touching objects (towels clothes, sports equipment etc.) that
infected skin has contaminated. Consequently, much heavily trafficked area can
be a source of infection. Many previous studies have evaluated staphylococcus aureus contamination of
various items such as computers and computer keyboards (Devine et al., 2001), pen (Banerijee, et al., 99) television sets (Stacey et al., 1998), stethoscopes (Cohen, et
al., 1997), tourniquets, (Berman, et al,
1986), clothes (Perry, et al, 2001),
mattresses, pillows, chairs, bed-frames, and over bed tables (Blythe, et a.l, 1998). Even all of the
surfaces sampled on chiropractic adjusting tables in one study carried
microorganisms.
Disinfection
protocols must therefore address removal of staphylococci from these surfaces
to prevent horizontal transmission in the outpatient settings (Biffero, et al., 2006). Strange enough, a study
has showed that a hairdresser is a possible source for cross-contamination of Staphylococcus aureus, and that
currently this is not a widelyrecognized potential route. Ruddy, et al., 2001). In another study, the
door handles in 53 (27%) of 196 rooms were contaminated by Staphylococcus aureus. It was detected on door handles of 41 rooms
(20.9%) Staphylococcus aureus were
detected on the door handles. (Sextonamet
al., 2006).
Barrier
precautions and hand disinfection are main methods used to break the chain of
transmission (Flournoy, et al.,
1993).
Previous studies have shown the minimum
inhibitory concentrations (MICs) of the commonly used biguanide disinfectant,
chlorhexidine (HCX), for Staphylococcus
aureus to be 4-fold, (Brumfitt, et al.,
1985). 8 to 16-fold (mycock, et al.,
1985). In this study, disinfectant experiment was conducted using different
concentrations of laboratory ethanol as disinfectant A, household bleach (Jik)
disinfectant B and saponatedbrand of cresol (Isol disinfectant C againstStaphylococcus aureus.
1.2 OBJECTIVES OF THE STUDY
To find out the minimums
inhibitory concentration of disinfectants that will be effective in Gram
positive Staphylococcus aureus.
To investigate their
sterilizing pattern.
To advice the public on the
important of disinfectants and dangers of harmful microorganisms.
1.3 STATEMENT OF PROBLEM
Staphylococcus
aureus is the leading cause of human infection in
the skin and soft tissues, bones and joints, abscesses and normal heart valves
and is found every where. Staphylococcus
aureus is usually spread by direct physical contact with those already
infected or through indirect contact by touching objects examples towels
clothes sports equipment, computer, mattresses, chairs , pen, bed frames,
adjusting tables etc.
Thus this problems lies at the improper use
of these disinfectants (i.e at incorrect concentration) by individuals,
families, hospitals and other laboratories which leads to not achieving
sterility and inadequate activities.
1.4 JUSIFICATION
This is to bring to the knowledge of the
general public that Staphylococcus aureus
are everywhere and is usually spread by direct physical contact with infected
persons or through indirect contact by touching objects. Also that the common
disinfectants we used against the organism can only be effective when it is
used in a correct concentration.
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