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Procedures - Day One:
- Review laboratory safety as explain on the Laboratory Safety Sheet
- Review Aseptic Techniques
- Provide an example of a properly labeled plate
- Demonstrate removal of the antibiotic disks from the vials.
- You can have the students flame the alcohol dipped forceps, if you wish. Emphasize safety!
- Designate containers in which the students should place their used swabs.
- Remind students to note on the Data Table which antibiotic they place in which section of the plate.
- Do not store the plates inverted because the disks may fall off of the agar.
Procedures - Day Two:
- Tell the students to keep the plates closed when recording data. You may want to tape them closed to ensure that students do not open them. The measurements can be taken by flipping the plate over.
- You can have the students put the plates back into the incubator for another 24 hours to see if there is a change in the appearance of the zones of inhibition or not.
- Designate a specific area for students to place all plates that need to be autoclaved.
Data Collection / Data Analysis:
- Remind students that measurements should be taken in millimeters.
- Point out that they need to record both quantitative and qualitative data.
- Refer to the literature available from the biological supply company or in microbiology textbooks for the ranges for sensitivity for each antibiotic that you test. For example, Tetracycline 30 microgram is
Resistant if the zone of inhibition is 14 mm or less, Intermediate if the zone of inhibition measures 15 to 18 mm and Sensitive for zones larger than 18mm.
Conclusion:
Have students write conclusions based on what they “Did, Thought, and Got.”
Summary Questions:
- Did you see any differences between S. aureus and E.coli in their resistance to any of the antibiotics? If yes, please state the differences.
ANSWERS WILL VARY.
- Can you tell by using this test whether the sensitive bacteria were killed or just not able to grow?
NO, FURTHER TESTING WOULD BE NEEDED.
- Did you observe any zones with a few colonies of bacteria within the zone of inhibition? If yes, what is a possible explanation for this occurrence.
ANSWERS WILL VARY.
- List some factors that a physician would need to consider other than zone of inhibition size before prescribing an antibiotic.
ADAPTED FROM: Hudson, Barbara K. Microbiology in Today’s World Second Edition. Kendall / Hunt Publishing Company, 1998 and Harley, John P. and Lansing M. Prescott. Laboratory Exercises in
Microbiology Fourth Edition. WCB McGraw-Hill, 1999.
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