What is the purpose of TSI agar?

Answer 1

TSI means Triple sugar Iron. To identify gram negative entric bacilli on the basis od carbohydrate fermentation and hydrogen sulphide production. The TSI agar contains three sugars dextrose, lactose and sucrose and indicator phenol red. Carbohydrate fermentation is idicated by the release of gas and change in colour from red to yellow. Hydrogen sulphide production is seen blackening of butt. For the detection of organisms that only ferment dextrose, the dextrose concentration is one-tenth the concentration of lactose or sucrose. The small amount of acid produced in the slant of the tube during dextrose fermentation oxidizes rapidly, causing the medium to remain red or revert to an alkaline pH. In contrast, the acid reaction (yellow) is maintained in the butt of the tube because it is under lower oxygen tension. Once the dextrose is depleted the organism starts to utilise lactose or sucrose.

Answer 2

Triple Sugar-Iron Agar TestObjective

1. To differentiate among members of the Enterobacteriaceae.

2. Distinguish between the gram-negativeenteric bacilli from other groups of intestinal bacilli based on carbohydrate fermentation and the production of hydrogen sulfide. Principle:

The triple sugar- iron agar test is designed to differentiate among the different groups or genera of the Enterobacteriaceae, which are all gram negative bacilli capable of fermenting glucose with the production of acid, and to distinguish them from other gram negative intestinal bacilli. This differentiation is based on the differences in carbohydrate fermentation patterns and hydrogen sulfide production by the various groups of intestinal organisms. Carbohydrate fermentation is detected by the presence of gas and a visible colour change (from red to yellow) of the pH indicator, phenol red. The production of hydrogen sulfide is indicated by the presence of a precipitate that blackens the medium in the butt of the tube.

To facilitate the observation of carbohydrate utilization patterns, TSI Agar contains three fermentative sugars, lactose and sucrose in 1% concentrations and glucose in a concentration of 0.1%. Due to the building of acid during fermentation, the pH falls. The acid base indicator Phenol red is incorporated for detecting carbohydrate fermentation that is indicated by the change in colour of the medium from orange red to yellow in the presence of acids. In case of oxidativedecarboxylation of peptone, alkaline products are built and the pH rises. This is indicated by the change in colour of the medium from orange red to deep red. Sodium thiosulfate and ferrous ammonium sulfate present in the medium detects the production of hydrogen sulfide (indicated by blackening in the butt of the tube).

Carbohydrate fermentation is indicated by the production of gas and a change in the colour of the pH indicator from red to yellow. To facilitate the detection of organisms that only ferment glucose, the glucose concentration is one-tenth the concentration of lactose or sucrose. The small amount of acid produced in the slant of the tube during glucose fermentation oxidizes rapidly, causing the medium to remain orange red or revert to an alkaline pH. In contrast, the acid reaction (yellow) is maintained in the butt of the tube because it is under lower oxygen tension.

After depletion of the limited glucose, organisms able to do so will begin to utilize the lactose or sucrose. To enhance the alkaline condition of the slant, free exchange of air must be permitted by closing the tube cap loosely. If the tube is tightly closed, an acid reaction (caused solely by glucose fermentation) will also involve the slant.

Materials Required:

Culture:

24 hour typticase soy broth culture

Media:

Triple sugar-iron agar slants

Equipment:

Bunsen burner

Inoculating needle

Test tubes

Marking pen

Media preparation: Triple sugar-iron agar (pH 7.4):

Add 3.0 gram of Beef extract,3 gram of yeast extract,15 gram of peptone,5 grams of protease peptone,10.0 grams of lactose,10.0 gram of saccharose,1.0 gram of glucose,0.2 gram of ferrous sulphate,5.0 gram of sodium chloride,0.3 gram of sodium thiosulphate,0.024 gram of phenol red and 12 gram of agar and make up to 1000ml with distilled water.

The Peptone mixture and the Beef and Yeast extracts provide the nutrients essential for growth. Sodium chloride maintains the osmotic balance of the medium. The Bacteriological agar is the solidifying agent.