Triple Sugar Iron Agar Slant

 

What is this medium used for?

 

Bacteria are identified based largely on what organic compounds they can break down. The range of compounds used depends on the collection of enzymes a species of bacteria can make. TSIA contains three sugars, and thus is used to determine the ability of the microbe to use some or all of these sugars. Additionally, TSIA contains iron compounds that react with hydrogen sulfide (H2S), a gas produced by some microbes. When a microbe produces H2S, a precipitate is formed, turning all or part of the agar black.

 

How is TSIA useful in following fermentation of sugars? 

 

Three sugars are found in TSIA: glucose, sucrose, and lactose. If any of the sugars can be used, the microbe will accumulate acidic byproducts. In a positive test, the pH indicator in the medium changes color from its normal red to yellow, indicating acid production.

If only glucose can be used, it is rapidly depleted in the slant but more slowly in the butt of the tube. Needing more nutrients but being unable to use the other sugars, the bacteria use the other ingredients. This activity turns the slant hot pink or magenta, a color stemming from the alkaline/basic compounds changing the color of the pH indicator. Bacteria using glucose, but not lactose or sucrose, can be expected to have a hot pink slant and a yellow butt.

If either sucrose and/or lactose is used, more sugars are available in the slant and the acid production is much greater. The pH indicator in the medium changes color from its normal red to yellow throughout the tube, yielding a yellow slant and a yellow butt. Bacteria using glucose and either lactose and/or sucrose, can be expected to have a yellow slant and a yellow butt.

 

How is TSIA useful in following production of hydrogen sulfide (H2S)? 

Also present in TSIA is an iron compound. The iron ions (Fe2+) have a high affinity for sulfide ions. The result is that H2S combines with the iron to make FeS, a black compound. In tubes of TSIA containing bacteria producing hydrogen sulfide, the agar turns black from the FeS.

 

What is the content of this medium?  

 

The medium is a nutrient broth to which 0.1% glucose, and 1% each of lactose and sucrose are added. The pH indicator phenol red is red at neutral pH but turns yellow at pH <6.8. It also changes to magenta or hot pink at pH >8.4. Also included is an iron compound so that any H2S production can be detected.

 

How is the test performed?  

 

For information on how to determine the ability of a microbe to use glucose, lactose, or sucrose, refer to the individual tests for those sugars. Interpreting the reactions for these three sugars from the TSIA reaction is not very exact.

For information on how to perform the test for H2S, refer to the hydrogen sulfide production test.