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SBL 1023
EXPERIMENT 5 : MICROBIOLOGY
NO MATRIC: E20161015640
GROUP : B
Experiment 5: Microbiology
Introduction
The streak plate method is a rapid qualitative isolation method. The techniques
commonly used for isolation of discrete colonies initially require that the number of
organisms in the inoculums be reduced. It is essentially a dilution technique that involves
spreading a loopful of culture over the surface of an agar plate. The resulting diminution of
the population size ensures that, following inoculation, individual cells will be sufficiently far
apart on the surface of the agar medium to effect a separation of the different species present.
Although many type of procedures are performed, the four ways or quadrant streak is mostly
done.
The hands are the parts of the human body that are in most contact with the outside
world. People use their hands for a variety of activities everyday. It it extremely easy to come
in contact with different microbes and to transfer them to other objects and mybe even
people. Handwashing is thought to be effective for the prevention of transmission of
pathogens. There are many products in the market claim to be able to kill the germs. However
it is not conclusive that handwashing with soap or other cleaning products is more effective at
reducing bacteria contamination than using water only.
Objective
Methods
C. Gram staining
1. The inoculating loop was used and 1 drop of sterile water was added to the slide.
Prepare a smear of:
a) Escherichia coli and b) staphylococcus aures
2. Air dry and heat fix.
3. The smear was covered with Crystal Violet (primary stain) for 1 min
4. The slide was washed with water.
5. Grams’Iodine (mordant) was added for 1 min.
6. Then wash with water.
7. Decolorize with 95% ethanol. This is the “tricky” step. Stop decolorizing with
alcohol as soon as the purple color has stopped leaching off the slide then
immediately wash with water.
8. The smear was covered with safranin for 30 seconds.
9. Both top and bottom of the slide was washed with water.
10. The slide was bloted.
11. The smear was observed using the light microscope. The smear was viewed up to
100x immersion oil.
Results
a) Escherichia coli
b) Staphylococcus aureus
Discussion
Streak plate technique is used to grow bacteria on a growth media surface so that
individual bacterial colonies are isolated and sampled. Isolated colonies indicate a clone of
cells, being derived from a single precursor cell. When the selected culture media is
inoculated using a single isolated colony, the resulting culture grows from that selected single
clone. The modern streak plate method has evolved from the efforts by Robert Koch and
other microbiologists to obtain pure culture of bacteria in order to study them. The dilution or
isolation by streaking procedure was originally developed by Loeffler and Gaffky in Koch's
laboratory, which involves the dilution of bacteria by systematically streaking them over the
surface of the agar in a petri dish to obtain isolated colonies which will subsequently grow
into mass of cells, or isolated colonies. If the agar surface grows microorganisms which are
all the genetically same, the culture is then considered as a pure culture..
Streaked plate are incubated at 37°C for 24 hours. The colonies grown in the plate
was examined carefully. Based ob my result, the water and hand sanitizer do not show any
effect of bacteria on thumb. In generally, hand sanitizer is the least effect of bacteria on
thumb. Control, soap and water section were effected by colonies of bacteria. So, the most
effective handwashing is sanitizer.
Gram-negative bacteria have a thinner layer of peptidoglycan (10% of the cell wall)
and lose the crystal violet-iodine complex during decolorization with the alcohol rinse, but
retain the counter stain Safranin, thus appearing reddish or pink. They also have an additional
outer membrane which contains lipids, which is separated from the cell wall by means of
periplasmic space.Based on this experiment, E coli show the gram-negative bacteria while
staphylococcus aureusis is gram-positive bacteria.
Conclusion
Reference
https://serc.carleton.edu/microbelife/research_methods/microscopy/gramstain.html
http://vlab.amrita.edu/?sub=3&brch=73&sim=208&cnt=1
https://en.wikipedia.org/wiki/Streaking_(microbiology)
Reflection
This experiment, I have been done from the previous semester. So I have an
experience in handling this experiment. The worst part is we need to careful while handling e
coli because e coli is bacteria that can harmful human. After making this experiment, I have
observed that thumb print without washing with water has more bacteria compare to thumb
that has been wash with sanitizer. So the lesson learn is we must wash hand before eat to
prevent food poising.