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38: TTC Motility Agar

Objectives

  • Identify different ways to run motility testing

SIM 

SIM deeps are a multi-test medium comprising 3 tests: sulfide (H2S gas), indole production, and motility. You have already tested for motility via the hanging drop slide, and here is an additional way to determine it, with less muss and fuss. In fact, there is an even better way to determine motility, motility agar with tetrazolium dye, that you will also be using in lab. Both of these agars have advantages. SIM is actually 3 tests in one, but, on the other hand, the motility with tetrazolium is much easier to read for motility. The tetrazolium is a colorless salt which becomes red when reduced, occurring as a result of bacterial metabolism.

Mitochondrial reduction of MTT to blue formazan product Image used with permission from Wikipedia (credit: Rogan Grant).

MATERIALS NEEDED

  •  SIM agar deep per organism
  • 1 TTC motility agar with tetrazolium dye per organism
  • Organisms: Klebsiella, E. coli

THE PROCEDURE

  1. Inoculate into a tube of SIM media with a NEEDLE, all the way to the bottom.
  2. Inoculate into a tube of TTC motility agar with tetrazolium dye with a NEEDLE, all the way to the bottom.
  3. Incubate at 25 or 37 degrees C.

INTERPRETATION

SIM

  1. Hold the tube up to the light and look at the stab line to determine motility.
  2. If NONMOTILE, you will see the intact straight stab line.
  3. If MOTILE, the original stab line will diffuse out into the medium as the bacteria spread throughout. In fact, you may not even be able to see a stab line at all: it may be turbid throughout the medium. FOR THAT REASON, you might want to compare the inoculated SIM with an uninoculated one.

Motility agar with tetrazolium 

The same description from above applies to this medium, BUT with the added, helpful colored dye tetrazolium which turns red as a result of the bacteria metabolizing.

 

 

QUESTIONS

  1. What is the function of tetrazolium?
  2. How do you tell if the organism is motile?

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