Triple Sugar Iron Agar (TSI) is recommended by ISO 6579 and ISO 19250 for the biochemical confirmation of Salmonella. Peptone and the beef extract provide nitrogen, vitamins, minerals and amino acids essential for growth. Yeast extract is source of vitamins, particularly the B-group. The three carbohydrates (glucose, sucrose and lactose) are the fermentable carbohydrates providing carbon and energy. When these are fermented the acid production is indicated by the phenol red indicator, being the color changes yellow for acid production and red for alkalinization. Sodium thiosulfate is reduced to hydrogen sulfide, which reacts with the iron salt to give the black iron sulfide. The ferric ammonium citrate is a H2S indicator. Sodium chloride supplies essential electrolytes for transport and osmotic balance. Bacteriological agar is the solidifying agent. The glucose concentration in the medium is one-tenth the concentration of lactose or sucrose in order to facilitate the detection of organisms that only ferment glucose. The fermentation of glucose produces a small amount of acid in the inclination of the tube, which is rapidly oxidized and the medium remains red or revert to an alkaline pH. On the other hand, the same acid reaction in the butt of the tube keep the acid pH (yellow) due to the lower oxygen tension. When all glucose is used, organisms able to ferment lactose or glucose will begin to utilize them. In order to enhance the free exchange of air in the slant of the tube, the tube cap must be closed loosely. The addition of 1% Sucrose in the TSI Agar allows differentiating between Proteus and Salmonella. The fermentation of the sucrose by Proteus turns the color of the Phenol red indicator in the slant from red to yellow. Dextrose positive and lactose negative members of the genus Salmonella, all cause a reddening of the slant and acidify the depths of the agar tubes.
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