Bacteriology 102:
Custom Formulation of Media
Based on Kligler Iron Agar

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Modifying KIA
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Modified:
4/9/01

How to "program" such a medium to screen for certain physiological types of bacteria.
(A "KIA Tutorial.")


Recall from our discussions about the triple differentiating feature of Kligler Iron Agar (KIA) in that it tests whether or not an organism can (1) ferment glucose, (2) ferment lactose, and (3) produce hydrogen sulfide (H2S) from the reduction of thiosulfate. (This medium is also explained on this page of our Differential Media Site, along with its variant, TSI Agar.) Whether or not there is enough acid produced from the fermentation of one or more sugars to permeate the medium and over-neutralize the alkaline reaction of the slant (which is caused by the aerobic deamination of amino acids) is the key differential feature in distinguishing between those organisms able to ferment just the "minor" sugar (present in 0.1% concentration – i.e., glucose in KIA) and those able to ferment both the "minor" and the "major" sugars (the latter present in 1.0% concentration – i.e., lactose in KIA). The additional "major" sugar in TSI Agar is sucrose.

Conceivably this medium could be altered to test for fermentation with other sugar combinations, and one could also think about adding a specific amino acid to see if the inoculated organism can decarboxylate it. Such amino acids (often lysine, ornithine and arginine) are usually present in decarboxylation test media in a 0.5 to 1.0% concentration, and an anaerobic, alkaline reaction is indicative of decarboxylation.


Consider the diagram on the right which applies to KIA and related media such as TSI Agar. The influence of decarboxylation of an optionally -added amino acid is also indicated.

Decarboxylation of the added amino acid will result in an alkaline "butt" if it is not overneutralized by excess acid formation by fermentation of the major sugar(s). Gas produced during fermentation will show up as cracks in the medium. Also, H2S production will be evident by the black color of ferrous sulfide (FeS) in the butt of the tube – if the pH of the medium is not too low from excess fermentation, as the FeS can dissolve and disappear under such conditions.


Here is a question from a recent (Spring, 2000) Bacteriology 102 take-home quiz: (Looks like another one of our thought questions doesn't it?)

We would like to be able to differentiate the enteric genera Sorgobacter, Splammobacter and Wiscobacter in a KIA-based medium – not only from each other but also from all other enterics and also Pseudomonas. Consider the reactions in the following table while answering the questions below.

organism fermentation of sugars: decarboxylation of
amino acids:
H2S pro-
duction
glucose fructose galactose lactose mannitol ornithine lysine
enterics Sorgobacter + + +
Splammobacter + + + +
Wiscobacter + + +
all other genera + + + +or– +or– +or– +or– +or–
Pseudomonas ?

a.  As you know, KIA contains glucose and lactose as the only fermentable sugars, and it has a H2S indicator system, and it does not contain any extra added ornithine or lysine. What would you add to KIA (in relatively large amounts) to differentiate Sorgobacter, Splammobacter and Wiscobacter from each other and also other enterics and Pseudomonas?

Choose two:  FRUCTOSE  GALACTOSE  MANNITOL  ORNITHINE  LYSINE

b.  Show how these organisms would appear in this modified KIA medium (i.e., color or pH of slant region, color or pH of butt region, and whether or not H2S is indicated in the butt region).

Think about it. Once you have it figured out, compare your answers to those given here along with a few take-home lessons.

Remember that many of the organisms specified on our "scenario questions" are fictional as is the case here (except for Pseudomonas), in our quizzes and exams, and in some of our "thought questions." (That's about as far as I go in writing science fiction.) Of course, all organisms mentioned on our bacterial identification pages are the real thing.


GO 
TO:
• Bacteriology 102 Home Page.
• Differential Media Site.
• Bacterial Nutrition and Cultivation.
• Bact. 102 Thought Questions.
• Site Outline of related pages.

Page last modified on 4/9/01 at 2:45 PM, CDT.
John Lindquist, Department of Bacteriology,
University of Wisconsin – Madison