Name

Name: Colleen Lim Student ID: 29880092 Partner: Chai Jun Kai

Investigating Bacterial Antibiotic Sensitivity
Introduction
Many pathogens are microorganisms. There are five main types of
microorganisms, which are the viruses, bacteria, prions, fungi and the protists.
Most of the microorganisms cause health diseases but some of them are
beneficial to us.
Microorganisms, such as moulds and other fungi, make antibiotics naturally to
help the body’s natural immune system to treat bacterial infection. Antibiotics
are a group of medicines which work effectively by fighting against bacteria
cells. Antibiotics have no effect on viruses, fungi and parasites, hence cannot
be used in treating viral, fungal and parasitic infections. Antibiotics that
actually kill bacteria are called bactericidal whereas antibiotics that inhibit
bacterial reproduction without killing them directly are called bacteriostatic.
The function of antibiotics besides killing bacteria include interfering with their
DNA replication and their protein synthesis. This essentially inhibits their
growth and metabolism, causing them unable to replicate or even leading them
to death. Different antibiotics are effective against different types of bacteria.
Broad spectrum antibiotics are effective against a wide range of bacteria
whereas the narrow spectrum antibiotics are only effective against a few.
Bacteria can become resistant to a specific antibiotic over time by undergoing
mutation. Once the bacteria is resistant and not sensitive to an antibiotic,
becoming a drug-resistant, then the bacterial disease can be difficult to treat.
The bacteria, Escherichia coli can be generally found in the digestive system of
warm-blooded organisms whereas the bacteria, Staphylococcus albus, an
anaerobic bacteria, is part of the human’s natural skin flora. In this experiment,
we will discover the effectiveness of antibiotics on the bacterium by observing
the presence of bacteria growth and measuring the zones of inhibition on
bacterial (Escherichia coli and Staphylococcus albus) culture plates after being
incubated for 24 hours. The strains of bacteria tested in this experiment are
non-pathogenic, which means they are harmless and do not cause disease.

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Aim
This experiment is carried out to explore the effectiveness of antibiotics on the
bacterium by measuring the zones of inhibition on bacterial (Escherichia coli
and Staphylococcus albus) culture plates after being incubated for 24 hours.

Hypothesis
If the greater the diameter of the zones of inhibition, then the more effective of
the antibiotics on the bacteria Escherichia coli and Staphylococcus albus.

Materials and equipment
See practical activity handout sheet, Investigating Bacterial Antibiotic Sensitivity.

Method / Procedure
See practical activity handout sheet, Investigating Bacterial Antibiotic Sensitivity.

Results
1. Data Presentations:
Table 1: The effectiveness of six different types of antibiotics on the
bacterium Escherichia coli and Staphylococcus albus by measuring the diameter
of the inhibition zones in (mm) under a black tile after being incubated
Zone Of Inhibition (mm)
Group data Class data
Escherichia
coli
Staphylococcus
albus
Escherichia
coli
Staphylococcus
albus
Grey AP
(Ampicillin)
22.0 57.0 23.7 52.3
Green C
(Chloramphenicol)
29.0 38.0 28.3 35.3
Pink PG
(Penicillin G)
0.0 41.0 0.0 36.7
White S
(Streptomycin)
24.0 20.0 23.4 20.5
Purple ST
(Sulphatriad)
0.0 0.0 0.0 0.0
Brown T
(Tetracycline)
28..0 40.0 27.6 36

Figure 1: The effectiveness of six different types of antibiotics on the
bacterium Escherichia coli and Staphylococcus albus by measuring the diameter
of the inhibition zones in (mm) under a black tile after being incubated

2. Sample Calculations:
To calculate the class data for each inhibition zones of the antibiotics
As we have eight groups in total, we need to sum up every group data
and divided by the number of groups (which is eight), lastly we will get
an average which will be our class data.
For the effect of antibiotic Grey AP (Ampicillin) on Escherichia coli,
Average = ����� �� ����� ������ �� ����
������ �� ������
= ����+���+���+���+���+���+���+����
��
= 23.7mm

3. Summary
Based on the data shown above, the main difference between Escherichia
coli and Staphylococcus albus is the effectiveness of the six different types
of antibiotics. The Escherichia coli is sensitive to four types of antibiotics
which are the Grey AP (Ampicillin), Green C (Chloramphenicol), White S
(Streptomycin) and Brown T (Tetracycline) but it is resistant to both
antibiotics Pink PG (Penicillin G) and Purple ST (Sulphatriad). In
contrast, the Staphylococcus albus is sensitive to five types of antibiotics
which are the Grey AP (Ampicillin), Green C (Chloramphenicol), Pink PG
(Penicillin G), White S (Streptomycin) and Brown T (Tetracycline) but it is
resistant to the antibiotic Purple ST (Sulphatriad). Besides that, the
sensitivity of each antibiotic on both the bacteria Escherichia coli and
Staphylococcus albus is different. For instance, the Escherichia coli is the
most sensitive to Green C (Chloramphenicol), which its diameter of
inhibition zone is 29.0mm whereas the Staphylococcus albus is the most
sensitive to Grey AP (Ampicillin), which has 57.0mm diameter of the
zone of inhibition. Therefore, based on the results we have observed and
recorded, the descending order of the zones of inhibition for both
bacteria Escherichia coli and Staphylococcus albus with the presence of six
different types of antibiotics can be shown like this as below.
For Escherichia coli, the descending order of our group result is
��?��������?��������l (29.0mm)>��������������������� (28.0��)>
�������������������(24.0��)>��������������� (22.0��)

Note: Penicillin G and Sulphatriad have no effect as Escherichia coli is
resistant to them.

For Staphylococcus albus, the descending order of our group result is
��������������� (57.0mm)>���������������� �� (41.0��)
>��������������������� (40.0��)>��?��������?��������l(38.0��)
>�������������������(20.0��)
Note: Sulphatriad has no effect as Staphylococcus albus is resistant to it.

4. Anomalous data: No anomalous data was observed.

Analysis and discussion
1. Yes, the most effective antibiotics identified for Escherichia coli and
Staphylococcus albus of our group data are similar to the class data. The
antibiotic disc with the greatest diameter of inhibition zone, is the most
effective. The most effective antibiotic identified for Escherichia coli in our
group is Green C (Chloramphenicol) which the diameter of its zone of
inhibition is 29.0mm, whereas the overall average data obtained by our
class is approximately 28.3mm. For the bacteria Staphylococcus albus,
based on our group data, the most effective antibiotic identified for it is
Grey AP (Ampicillin). The diameter of its zone of inhibition is the largest
among all antibiotics tested, which is 57.0mm, and also the overall
average class data is about 52.3mm. Hence, it is obviously seen that
Green C (Chloramphenicol) and Grey AP (Ampicillin) are undoubtedly the
most effective antibiotics for the bacteria Escherichia coli and
Staphylococcus albus respectively. This is because both Green C
(Chloramphenicol) and Grey AP (Ampicillin), have the ability to fight
against the bacteria Escherichia coli and Staphylococcus albus respectively as
well as restrict the growth of bacteria in the agar, thus there are more
clear areas (called the zones of inhibition) around the antibiotic disc of
Green C (Chloramphenicol) on the Escherichia coli agar plate and more
clear areas around the antibiotic disc of Grey AP (Ampicillin) on the
Staphylococcus albus agar plate.

2. Of the two species of bacteria we investigated in this activity,
Staphylococcus albus would be more easily treated with antibiotics. This is
because bacteria Staphylococcus albus is sensitive to more antibiotics than
Escherichia coli. Moreover, the diameter of inhibition zone around the
antibiotic discs on the Staphylococcus albus agar plate is much larger
compared to the antibiotic discs on the Escherichia coli, which means
there is more clear areas where the growth of bacteria Staphylococcus albus
is absent.

3. The unknown concentration of the antibiotics on the Mastring and paper
discs may influence the presence and size of a zone of inhibition. If the
concentration of antibiotics are low, then it will result in a not significant
and prominent inhibition zone or inhibition zone with a smaller
diameter. By the way, the concentration of the antibiotics does not
matter if the bacteria itself is resistant and not sensitive to the antibiotics.
For instance, the antibiotic Purple ST (Sulphatriad) has no effect on
bacteria Escherichia coli and Staphylococcus albus. Hence, whether the
concentration of the antibiotics on the Mastring and paper disc is higher
or lower, there will be no difference and any effects on the presence and
size of a zone of inhibition.

4. From Table 2, the antibiotics Chloramphenicol, Streptomycin and
Tetracycline would be considered as broad-spectrum antibiotics. This is
because broad-spectrum antibiotics are antibiotics that can act against a
diverse range of disease-causing bacteria, help the body’s natural
immune system to fight bacterial infections. All the nine bacterial species
which are the E.coli, B.subtilis, B.cereus, Sarcina lutea, Erwina, Serratia
marcescens, P.fluorescens, R.rubrum, S.albus, are sensitive to the antibiotics
Chloramphenicol, Streptomycin and Tetracycline, showing us the
characteristics of broad-spectrum antibiotics, the zone of inhibition will
indicate poor growth of the bacteria. It also means that the disease
caused by these nine bacterial species can be treated easily by these
broad-spectrum antibiotics.

5. From Table 2, the bacteria B.cereus and P.fluorescens could be considered
as multiple drug resistant (MDR), which the bacteria are resistant to a
large number of different antibiotics, resulting in difficulties to treat.
Both bacteria B.cereus and P.fluorescens are resistant to three antibiotics
out of six in total, which are the antibiotics Ampicillin, Penicillin and
Sulphatriad.

6. The first and second safety procedures are very important in this
experiment. Lab coats, safety goggles and gloves are to be worn at all
times in order to avoid our skin and eyes exposed to the bacteria
Escherichia coli and Staphylococcus albus because these bacteria can cause
bacterial infectious disease, which negatively impacts on our health.
Also, never have any food or drink at workstation because the bacteria
can spread and transmit to the food or drink, causes it to be
contaminated, thus we will catch a disease after consuming the
contaminated food or drink.

7. One additional safety precaution that we followed but is not listed in the
safety section of the Practical handout is that we sterilized our
workspace and all the equipment with 70% alcohol prior for plating
procedures. This is relevant to this experiment as the experiment carried
out is about the microorganism?bacteria and by sterilizing the
workspace and equipment with 70% alcohol, we can minimize the
possible contamination with the bacterial culture.

8. Improvement:
Personally, I think this experiment can be improved by spreading the
bacteria Escherichia coli and Staphylococcus albus evenly over the entire
surface of the agar in the petri dishes and allow the bacteria to absorb
thoroughly. Also, I think the experiment result can be improved by
measuring the zone of incubation (clear areas) with a transparent ruler
from the back of the plate under a black tile instead of measuring it from
the front of the plate as the diameter of the inhibition zone can be
measured more easily. Lastly, I think the presence of moisture content

on the agar medium can counteract with the accuracy of the
susceptibility testing. It is important to make sure there is no moisture or
liquid present on the agar surface by allowing the plate to dry at room
temperature for a few minutes before placing the antibiotic discs. From
these, we would improve the accuracy, quality and reliability of our
data.

9. Extension of this experiment/ Further research
While this experiment investigates the effectiveness of antibiotics on the
bacterium by measuring the zones of inhibition on bacterial (Escherichia
coli and Staphylococcus albus) culture plates, the temperature of incubation
of this experiment remains consistent, which is 30?. Hence, we can
further extend this experiment on how temperature changes of
incubation affect the effectiveness of antibiotics or the sensitivity of
bacteria to antibiotics.

Conclusion
Our investigation is coherent with our hypothesis formed at the beginning of
the experiment. All in all, to sum up our findings, we discovered that the larger
the diameter of the zones of inhibition or clear areas around the antibiotic discs
on the agar after incubation, indicating that the poorer the growth of the
bacteria, hence the more effective of the antibiotics on the bacteria Escherichia
coli and Staphylococcus albus.

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