Download Lethal Effects of Temperature

The Lethal Effects of High Temperature on Bacteria
Each bacterial species possesses the ability to grow over a range of temperatures, but if the maximum
temperature for growth is exceeded, a killing effect will be observed. The susceptibility of different
organisms to high temperature can be measured as the Thermal Death Point or the Thermal Death Time.
The former is the lowest temperature at which a suspension of bacteria is killed in 10 minutes; while the later
is the time required to kill a finite number of cells or spores at a given temperature.
Wet heat is often used as a means of killing bacteria. How effective this is as a means of destroying
organisms depends on the type of bacteria. Generally, Gram negative organisms, which are usually found
inside other organisms (such as people) are the most sensitive to temperature. Gram positive organisms,
which are often found free in the environment, are a little less sensitive to temperature. Endospore-formers
(Bacillus and Clostridium) are the most resistant, due to the highly resistant endospore. It is important to note
that the vegetative cell of a spore former is only as resistant as other Gram positive organisms.
In this exercise, students will expose a bacterial suspension to temperatures of 60°C, 80°C, and boiling water.
At intervals of 5, 10 and 20 minutes (total time) a sample of the suspension will be removed and plated out to
test for viability. We will compare results from a gram positive organism, a gram negative organism and a
spore-forming organism.
Materials:
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Three broth cultures of the organism to be studied
Three Tryptic Soy Agar plates
Water baths set for 60°C, 80°C, and boiling water
Inoculating loop
Procedure:
1. Work in groups of three. Students in a group will work with the same organism, but each student will
expose the organism to a different temperature. Different groups will work with different organisms.
2. Obtain a Tryptic Soy Agar plate, invert it, and use a marking pen to divide the plate into quadrants. Label
the quadrants "0", "5", "10", and "20".
3. Inoculate the zero time quadrant of the plate with a loopful of your test culture. Streak the loop in a "Z"
pattern. This will serve as a control to demonstrate organism viability prior to exposure to high temperature.
4. Place the culture in the appropriate water bath. After 5, 10, and 20 minutes (total time), mix your heated
specimen by gently tapping the tube and then obtain a loopful of inoculum. Streak the appropriate quadrant
of your plate. Work quickly so that cooling does not occur while the sample is being obtained, and return
your culture to the bath after the sample has been collected.
5. Incubate plates as directed until the next laboratory period.
7. At the next laboratory period, examine each plate for growth and attempt to determine the Thermal Death
Time of your test specimen for each temperature studied.
8. Compare your results with those obtained by the other groups in the class.
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