LAB REPORT 3 MIC 254

ENUMERATION OF E. COLI FROM MEAT PRODUCT

By:

Muhammad Ilham Nur Hakim Bin Ramli (2022775425)

Muhammad Nasren Hisyam Bin Saifuddin (2022744737)
Muhammad Idham Bin Abu Bakar (2022340509)
Nadia Yasmin Binti Mohd Zaki (2022777181)
Farahin Nursyahirah Binti Mahthir (2022553425)

Lecturer: Madam Syazuani Mohd Shariff

Date of Experiment: 05/03/2023

Date of Submission: 11/04/2023

Group: AS1143A1
TITLE
Enumeration of E.coli from Meat Product.

OBJECTIVES

To determine the presence of E. coli from meat products.

INTRODUCTION

Bacterial enumeration is the technique of determining the quantity of bacteria in a particular

sample.There are a variety of reasons why researchers must count the quantity of bacteria or
compare the rising population of these germs under specified settings. It is mostly necessary and
routine labour in food, water, and dairy microbiology labs.

The knowledge of the quantities of various microorganisms in our food, including milk,
curd, buttermilk, and water, assists labs in determining whether or not the prepared food or
accessible water is safe to consume. Yet, because these microbes are so minute and undetectable
to our human eyes, a high-powered microscope is necessary to investigate them.

Meat can get infected during slaughter, and organisms can become extensively
incorporated with ground beef. Bacteria found on cow udders or on equipment may enter raw
milk. Consuming meat that has not been cooked well enough to destroy E. coli can lead to
illness. The contaminated meat seems and smells normal. The number of organisms necessary to
induce illness is unknown, however it is thought to be quite tiny.

Eating contaminated food, such as ground beef, is the most common method to obtain an
E. coli infection. E. coli bacteria in cow intestines can be transferred to meat during slaughter
and processing.
MATERIALS

Molten tryptic soy agar

Molten MacConkey agar
Autoclaved distilled water
Sample of meat products
70% alcohol
Petri dish
Micropipette 1000 µ𝐿
Micropipette tips 1000 µ𝐿
Test tube
Test tube rack

PROCEDURE

1. 25 g of the meat product was measured and was added into stomacher bag by using
aseptic technique.
2. 225 ml autoclaved distilled was added into the stomacher bag. Sample solution with a
1
dilution factor 10 was prepared.
3. Stomach for three minutes.
4. Three sterile test tubes were added with 9 ml autoclaved distilled water. 1 ml sample of
solution was inserted into the first tube and homogeneously mixed. The tube that is
2
labelled as T2 with dilution factor 10 . The same procedure was done by using the two
3 4
test tubes which were labelled as T3 and T4 with dilution factor 10 and 10 .
2
5. 100 μl of the homogenised mixture was inoculated from tubes with dilution factor 10 ,
3 4
10 and 10 and was added by molten tryptic soy sugar into an empty petri dish. The
plate was swirled gently by letting the combinations mixed homogeneously and letting
the agar solidify.
6. Step 5 was repeated with molten MacConkey agar.
7. The agar plate was incubated at 37℃ for 48 hours in aerobic condition
 8. The colonies were counted on both types of agar which are dark red colonies on
MacConkey agar and CFU/ml was counted and isolated from the sample.

RESULT

A) Enumeration of bacterial colonies from meat product inoculated using tryptic soy agar.

Dilution Number of colonies

10
2 112

10
3 96

10
4 83

Number of bacteria in original sample (use the formula (C×M)/V)

(112×102 ) + (96×103 ) + (83×104)
= 0.5

1874400
=
3

= 624 800

B) Enumeration of bacterial colonies from meat product inoculated using MacConkey agar.

Dilution Number of colonies

10
2 5

10
3 1

10
4 0
 Number of bacteria in original sample (use the formula (C×M)/V)

(5×102) + (1×103) + (0×104)

=
0.5

3000
=
3

= 1000

DISCUSSION
In this experiment, we used tryptic soy agar and MacConkey agar. The result for tryptic soy agar
showed the number of colonies in T² was 112 meanwhile 96 in T³ and 83 for T⁴.

MacConkey agar shown the number of colonies in T² was 5 and 1 in T³ while 0 in T⁴. These two
types of agar have shown a huge difference in growth of E.coli.

Tryptic soy agar has a lot of colonies that grow on it. It is because of the composition of the agar.
In this agar, E.coli were able to grow because soybean provides a lot of nutrients like vitamins
and minerals that make it suitable for the microorganisms to grow. It also has glucose that can
convert to carbohydrates as an energy source to E.coli. Thus, it is proof that tryptic soy agar has a
lot of factors to the growth of E.coli.

The enumeration in MacConkey agar showed lesser growth of E.coli.The growth factors are
quite similar with tryptic soy agar, so E.coli should have grown normally like the other agar. In
this case, E.coli produces transparent growth that makes it harder to observe it since MacConkey
agar is in pink colour. E.coli only creates colour in some media. Hence, MacConkey is not a
suitable substance to observe the growth of E.coli.

Both agar has similar pH so they weren't any problem with the growth but for some cases, the
colour of MacConkey is the problem since E.coli is a gram-negative type of bacteria.
MacConkey agar usually shows the growth of microorganisms that are gram-positive.
CONCLUSION

In conclusion, the objective of the experiment was achieved. The characteristics of the bacteria
E. coli in meat products have been observed and determined during the experiment. Results from
enumeration of E. coli follow the theoretical result expected. The number of colonies inoculated
decreases for each dilution factor in both tryptic soy agar and MacConkey media. The number of
bacteria in the original sample were found. Therefore the experiment was successful with no
errors or mistakes occurring.

QUESTION

1) Discuss why MacConkey media is categorised as a differential and selective media.

MacConkey media is categorised as a differential and selective media Based on the
ability to ferment lactose, different species will yield colonies in varying appearance on a
MacConkey medium. This gives McConkey agar its differentiating property.

2) Predict the similarities and/or differences in the observations if the plates are
incubated at 30°C.
The observations that can be seen if the plates containing MacConkey agar and tryptic
soy agar that were inoculated with homogeneous mixture would be the same as the result
obtained in this experiment. This is due to the fact that the culture is best incubated at
30°C up to 35°C. Hence, the predicted observations would not be as different as the
observations that can be seen in this experiment.

3) Discuss the differences between results obtained using tryptic soy agar and
MacConkey agar.
The number of colonies counted on the tryptic soy agar plates are significantly larger
compared to the colonies on the MacConkey agar plates. Because E. coli, which is
contained in the meat product, would develop more on the plates with higher nutrients,
which is the tryptic soy agar, there is a large difference in the number of colonies that can
be seen on the two plates.
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