SBI3U1Name: __________________________

Bacteria Lab II: Effect of Bacteriocides

Purpose: To test the ability of various household cleaners to act as bacteriocides to inhibit bacterial growth.

Materials and Methods: (wear safety glasses, tie back long hair, never leave flame unattended)

  1. Obtain your inoculated and un-inoculated agar plates. Examine both for evidence of bacterial growth. You should see a number of colonies of bacteria (and mold) growing on the inoculated plate. You may see some bacterial growth on the other plate since it was not sterilized.

  2. Using a permanent marker, divide the bottom of the un-inoculated agar plate into 4 quadrants. Label the quadrants: 1, 2, 3 and C (for control).

  3. Obtain a clean, dry test tube and add 3-4 mL of distilled water.

  4. Set up a Bunsen burner with a small blue flame and heat the tip of a wire loop to sterilize it.

  5. Cautiously lift the lid of the inoculated agar plate just enough to allow you to scoop up a small amount of a non-shiny bacterial colony with the wire loop. DO NOT PENETRATE THE SURFACE. Swirl the wire loop in the distilled water in the test tube to dislodge the bacteria.

  6. Using a clean dropper, mix the bacteria and distilled water and then fill up the dropper with the bacterial suspension.

  7. Lift the lid of the un-inoculated agar plate enough to allow you to empty the dropper's contents onto the surface of the agar. Very gently swirl the petri dish to spread the suspension evenly on the surface. Let the suspension settle for a few minutes before you go on to the next step.

  8. Soak a filter disc in one of the bacteriocide solutions available. Using forceps to handle the disc, place it on the surface of the agar in quadrant 1 and very gently press it down. Write the quadrant number and the identity of the bacteriocide in a table (NOTE: tables are to be included with this lab under the heading OBSERVATIONS).

  9. Repeat with 2 other discs, soaking them in different bacteriocides and placing them in quadrant 2 and 3. For quadrant 4, soak the disc in distilled water (control). Complete your table.

  10. Re-seal both petri dishes with tape and leave them to incubate for a few days. Wash all equipment and your lab bench. Lastly, wash your hands.

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  12. After a few days, examine the agar plate with the bacteriocide discs. Look for a circular region around each disc where bacterial growth was inhibited (ie. the zone of inhibition).

  13. Using a ruler, measure the zone of inhibition for each disc. Measure from the centre of the disc to the periphery of the inhibition (ie. to where bacterial growth begins again). Record your measurements in a table.

  14. Make a labelled drawing of your petri dish showing: quadrants, filter discs (including the names of all bacteriocides and control), bacterial colonies, and zones of inhibition (NOTE: drawing goes in OBSERVATIONS section below table).

Questions:

In an experiment, the zones of inhibition around 4 discs are: (1) Saniflush - 2.3 mm, (2) Listerine - 5.1 mm, (3) Mr. Clean - 3.5 mm, (C) Distilled Water - 1.6 mm.

  1. Order the liquids in the experiment from strongest to weakest bacteriocide.

  2. There was a small zone of inhibition around the control. Does this completely invalidate the experiment? Explain.

  3. Would the bacteriocide that worked best in the above experiment necessarily be the best for your experiment? Explain.

  4. How might a bacterial mutation aid certain colonies of bacteria which are exposed to a particular bacteriocide repeatedly?

  5. Suggest one way your bacteriocide experiment could be modified or extended?