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Activity 1.2.2 Which Antibiotic Is the Best Choice? (Optional) Introduction In the last activity you looked at the various classes of antibiotics. You learned that different classes of antibiotics effect bacterial cells in different ways. Therefore, each class of antibiotics has a varying level of effectiveness in treating specific strains of bacteria. You investigated the structure of Neisseria menigitidis bacteria, the bacteria infecting Sue Smith, in order to determine which class of antibiotics would be best at treating Sue’s infection. When a patient is diagnosed with a bacterial infection, the physician will often take a sample of the bacteria from the infection site and test several different antibiotics to find the best one to use. The activity you will be completing is the same one performed daily in hospitals to find the most effective antibiotic to treat an infection. You will test several different antibiotics to determine which one is the most effective at preventing the growth of a strain of bacteria.

Equipment 

Carolina Antibiotic Sensitivity BioKit® o Plate culture of Bacillus cereus o Plate culture of Escherichia coli o Antibiotic disks o Control disks

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Forceps Alcohol in beaker Alcohol lamp Matches Permanent marker (Sharpie fine point) Disinfectant in squirt bottle Incubator at 37º C (optional) Safety glasses Latex or Nitrile exam gloves Roll of transparent tape (e.g. Scotch tape) Laboratory Journal

Procedure 1. Put on safety glasses and exam gloves. 2. Spray disinfectant on the area you will be working, and wipe it clean. 3. Get one Petri dish containing Bacillus cereus and another containing Escherichia coli. These dishes contain live bacterial cultures. You cannot see the bacteria

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growing because the cultures are less than two hours old and the bacteria have not multiplied sufficiently to be visible. 4. Write your initials or first names on the outside edge of each Petri dish. Don’t write too large or your writing will interfere with your observation of the results. 5. Place one Petri dish over the diagram below.

6. Use the technique demonstrated by your teacher to sterilize the forceps with flaming alcohol. 7. Use the sterile forceps to place the control disk in the center position on the Petri dish. 8. Expel one disk of each of the six types of antibiotic from its vial onto the agar in the Petri dish. Use the circles on the diagram to position the disks. 9. Repeat steps 5 to 8 with the second Petri dish culture. 10. Seal the Petri dishes using transparent tape. 11. Spray disinfectant on the work area and wipe it clean. 12. Remove your gloves and dispose of them as directed by your teacher. 13. Wash your hands and forearms thoroughly with soap and warm water. 14. Make a copy of the diagram in your laboratory notebook for each culture and record which disk you placed at each position. 15. Make a data table in your Laboratory Journal similar to one shown below with enough rows for each of the antibiotics. DISK Antibiotic

Color-Code

Diameter of Clear Area (mm) 24 Hours B. cereus

E. coli

48 Hours B. cereus

E. coli

16. Complete the first two columns of the data table.

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17. Incubate the bacterial cultures as directed by your teacher. 18. Observe the cultures after 24 hours. 19. Draw properly labeled diagrams of each culture in your laboratory journal. 20. Measure the diameter of the clear area surrounding each disk from one edge across the disk to the opposite edge. The clear area is the area where the bacteria were not able to grow or survive. It is called the zone of inhibition. 21. Record the measurement of the zone of inhibition in millimeters in the Data Table in your laboratory journal. 22. Record a value of zero if there is no clear area around a disk. 23. Repeat steps 18 to 21 after the cultures have incubated for 48 hours.

Conclusion 1 Why is the clear area surrounding an antibiotic disk called the zone of inhibition?

24. Which antibiotic would be the most effective treatment for an infection with the E. coli? Explain your response.

25. Which antibiotic would be the most effective treatment for an infection with the B. cereus? Explain your response.

26. Which antibiotic would be the most effective treatment for a person with an infection with both B. cereus and E. coli? Explain your response.

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27. How were the results after 48 hours different from the results after 24 hours?

28. What was the purpose of the control disk?

29. Propose reasons why an antibiotic may not be as effective at inhibiting E. coli as it is against B. cereus.

30. Classify each of the tested antibiotics as limited-spectrum or broad spectrum. Explain your responses.

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