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PRACTICAL 2
TITLE: Examination of Living Bacteria.
OBJECTIVE
1. To prepare and observe wet-mount slides and hanging-drop slides.
2. To distinguish between true motility and Brownian movement.
INTRODUCTION
A significant portion of prokaryotic microorganisms are bacteria. Bacteria are prokaryotic, tiny,
unicellular creatures. They are categorized under the domain "Prokaryote" with Archae because
they lack genuine nuclei and membrane-bound cell organelles. Bacteria come in a variety of
shapes, ranging from spheres to rods and spirals, and are typically only a few micrometers long.
Organelles like chloroplasts and mitochondria are absent in bacteria. Instead, a nucleoid contains
their continuous, circular double-stranded DNA. The term nucleoid is an area with an atypical
form and no nuclear membrane. Additionally, bacteria have a cell membrane and a wall that is
frequently composed of peptidoglycan. The cell envelope is the collective name for the cell
membrane and cell wall. Numerous bacteria require cell walls to survive. The genetic material of
bacteria normally consists of a single circular bacterial chromosome of DNA, which is found in
the cytoplasm in a body. The chromosome, together with the accompanying proteins and RNA,
is found within the nucleoid.
In this experiment, hanging drops technique will be conducted to observe Staphylococcus

aureus under the microscopes. The hanging drop slide method is used to examine the motility or
mobility of bacteria and view live cultures. To do this, add a drop of fluid to a slide that has a
dimple in the center, then top it off with a cover slide to suspend the culture in the concave
region. This method is frequently used while researching the behaviour and specifics of bacteria.
The liquid element makes it simpler to observe how bacteria typically migrate and how this is
essential to the organism's survival. The tentacle-like appendage, known as a flagellum, can also
be seen in a more natural condition because the movement is less constrained. This is necessary
to comprehend the biology of these life forms and how it actually functions. From this
experiment, the size and shape of the microorganisms when seen down under the microscope has
been incredible. The shape of Staphylococcus aureus was cocci which was like a sphere and the
size was smaller. However, some other shape of Staphylococcus aureus cannot be seen because it
exploded due to some errors occurring during an experiment. Next, microorganisms are alive
because they sustain essential life functions. The movement of bacteria towards nutrients is
known as motility. The hypothesis of the experiment is the bacteria Staphylococcus aureus can
be seen.
MATERIALS
Compound microscope
Cavity glass slides
Cover slips
Inoculating loop
Paper towels
Applicator sticks
Disposable pipettes
24-hours nutrient broth cultures of Staphylococcus aureus
METHODOLOGY
Hanging Drop Techniques
1. Cavity glass slides were obtained and cleaned.

2. Small amount of Staphylococcus aureus was picked up on the applicator stick.
3. The petroleum jelly was carefully touched to all four edges of a coverslip. A small rim of
Staphylococcus aureus was obtained around the entire coverslip by using the technique
demonstrated by the instructor. Staphylococcus aureus have got carefully on the other
side of the coverslip.
4. The coverslip was placed on a paper towel, with the Staphylococcus aureus -sided up.
5. A small drop of the bacterial broth was transferred onto the coverslip by using an
inoculating loop.
6. A cavity slide was placed over the drop, the paper towels were scooted to the edge of the
table, the slide over was flipped carefully and quickly. The drop of fluid was suspended
upside down from the coverslip into the space provided by the cavity glass slide.
7. The drop was examined under low power by locating the edge of the drop and the slide
moved therefore, the edge of the drop crossed the center of the field.
8. The light was reduced with the iris diaphragm and focus.
9. The different sizes, shapes, and types of movement were observed.
10. The high-power was switched and observations were recorded.
11. The procedure was repeated with the culture of another bacterial broth by using a new
coverslip. The observations were recorded.
12. Microscope was cleaned and returned to the proper cupboard after completely finished.
DATA AND ANALYSIS
Figure 1: Image of Staphylococcus aureus under the microscope

Figure 2: Image of Staphylococcus aureus under the microscope from the internet
DISCUSSION
Staphylococcus aureus from the nutrient broth cultures are observed under the microscope.
The bacteria Staphylococcus aureus is significant and has a significant effect on human health.
Although it is well known for causing infections of the skin and soft tissues, it has the capacity
to infect almost all organ systems in the human body, frequently with catastrophic results. The
wide variety of virulence factors it produces, many of which are encoded on plasmids,
transposons, prophages, and pathogenicity islands, is largely responsible for this remarkable
adaptability. In figure 1, the cocci shape of Staphylococcus aureus cannot be seen under 4X
magnification (from group 3). This is because students forget to put oil immersion
magnification. The cocci shape of Staphylococcus aureus can be seen under 10X
magnification (from group 3). Meanwhile, under the 40X magnification of the microscope of
Staphylococcus aureus cannot be seen. This is due to bacteria being exploded when students
heated the bacteria for a longer period of time than necessary during an inoculating loop. In
order to get a clear result from an experiment, it is consequently important to avoid making
personal mistakes in the lab. It is important to follow the instructions in the lab manual exactly
when conducting the experiment to prevent errors, such as heating the bacteria in an
inoculating loop incorrectly, which could lead to exploded bacteria.
CONCLUSION
In conclusion, the skills of hanging drop techniques have been prepared to observe the
Staphylococcus aureus under the microscopes. In this experiment, the hanging drops technique
helps in observing the general shape of living bacteria like Staphylococcus aureus and how the
arrangement of bacterial cells changes when they are joined together. The application of this
experiment in real life is since the shape of spiral bacteria distorts when fixed with heat, their
morphology can be specifically investigated using the hanging drop method. Spiral bacteria,
such as spirochetes, must be inspected while they are still alive, and a dark field microscope is
a good tool for examining their structure and arrangement. By using the hanging drop method,
which allows bacteria to move around freely in a liquid medium, it is possible to study the
movement or mobility of bacteria. The hypothesis in this experiment based on data obtained
was rejected because an experiment does not falsify hypothesis in the introduction. The
bacteria Staphylococcus aureus cannot be seen under the microscope when conducting an
experiment.
QUESTION
1. How would you describe in words the size and shape of the microorganisms that you see
down in the microscope?
The size and shape of the microorganisms when seen down under the microscope has
been incredible. The shape of s.aureus was cocci which was like a sphere and the size
was smaller.
2. Do you think microorganisms are alive?
Microorganisms are alive because they sustain essential life functions.
3. The movement of bacteria towards nutrients is known as?
The movement of bacteria towards nutrients is known as motility.

REFERENCE
1. Dahal, P., Joseph, Nelima, C., & Asim, M. (2022, May 03). Bacteria- definition,
structure, shapes, sizes, classification. Retrieved November 24, 2022, from
https://microbenotes.com/bacteria/
2. Ward, B. (2021, May 25). What is the hanging drop slide technique? Retrieved November
24, 2022, from https://microscopeclarity.com/what- is-the-hanging-drop-slide- technique/
3. Editors, B. (2019, October 04). Bacteria - definition, shapes, characteristics, types &
examples. Retrieved November 24, 2022, from https://biologydictionary.net/bacteria/
4. Staphylococcus aureus. (n.d.). Retrieved November 24, 2022, from
https://www.sciencedirect.com/topics/biochemistry-genetics-and- molecular-
biology/staphylococcus-aureus
5. Tracey, A., Taylor, T., & Chandrashekhar G., Unakal. (n.d.). NCBI Bookshelf. Retrieved
November 24, 2022, from https://www.ncbi.nlm.nih.gov/books/NBK441868/

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PRACTICAL 2

TITLE: Examination of Living Bacteria.

In this experiment, hanging drops technique will be conducted to observe Staphylococcus

1. Cavity glass slides were obtained and cleaned.

Figure 1: Image of Staphylococcus aureus under the microscope

2. Do you think microorganisms are alive?

Microorganisms are alive because they sustain essential life functions.

3. The movement of bacteria towards nutrients is known as?

The movement of bacteria towards nutrients is known as motility.

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