In 1911, Russell described the use of two sugars in a medium to differentiate Gram-negative organisms of intestinal origin.1 Lead or iron salts were added to Russell’s medium to detect the presence of hydrogen sulfide. Kligler added lead acetate to Russell Double Sugar Agar, resulting in a medium capable of differentiating typhoid, paratyphoid, and dysentery.2,3 A modification of this medium was developed, Kligler Iron Agar, using Phenol Red as an indicator and iron salts to detect hydrogen sulfide production. In 1940, Sulkin and Willett described a triple sugar ferrous sulfate medium for use in identification of enteric organisms.4 Triple Sugar Iron Agar is essentially the formula originally described by Sulkin and Willett.4
Triple Sugar Iron Agar is recommended for differentiation of enteric, Gram-negative bacilli from clinical specimens, dairy samples, and food products.5-7 Casein peptone, mixed peptone and yeast extract act as a source of nitrogen, sulphur, carbon, vitamin and minerals. Lactose, Sucrose and Glucose are the fermentable carbohydrate. Due to the building of acid during fermentation the pH falls. In case of oxidative decarboxylation of peptone alkaline products are built and the pH rises. This is indicated by the phenol red, which changes its colour in acidic surroundings from red-orange to yellow, on alkalinization it turns deep red. Sodium chloride is for the osmotic balance. Gas production (CO2) is detected by the presence of cracks or bubbles in the medium, when the accumulated gas escapes.
The H2S-positive bacteria reduce the sulfide in the culture medium to sulfide, which reacts with iron. Due to this reaction a black precipitate (FeS) is formed. Triple Sugar Iron Agar should be used in parallel with Urea Broth to distinguish between Salmonella and Proteus species.
- 1. Russell, F. F. 1911. The isolation of typhoid bacilli from urine and feces with the description of a new double sugar tube medium. J. Med. Res. 25:217.
- 2. Kligler, I. J. 1917. A simple medium for the differentiation of members of the typhoid-paratyphoid group. Am. J. Public Health 7:1042-1044.
- 3. Kligler, I. J. 1918. Modifications of culture media used in the isolation and differentiation of typhoid, dysentery, and allied bacilli. J. Exp. Med. 28:319-322.
- 4. Sulkin, S. E., and J. C. Willett. 1940. A triple sugar-ferrous sulfate medium for use in identification of enteric organisms. J. Lab. Clin. Med. 25:649-653.
- 5. P. R. Murray, E. J. Baron, M. A. Pfaller, F. C. Tenover, and R. H. Yolken (eds.). 1995. Manual of clinical microbiology, 6th ed. American Society for Microbiology, Washington, D.C.
- 6. Marshall, R. T. (ed.). 1992. Standard methods for the examination of dairy products, 16th ed. American Public Health Association, Washington, D.C.
- 7. US Food and Drug Administration. 1995. Bacteriological analytical manual, 8th ed. AOAC International, Gaithersburg, M.D.
- 8. J. MacFaddin, Media for lsolation-Cultivation-Identification-Maintenance of Medical Bacteria, Vol. 1, Williams and WiIkins, Baltimore (1985).
- 9. International Organization for Standardization (ISO), Draft ISO/DIS 6579 (1993).
- 10. Bundesgesundheitsamt: Amtliche Sammlung von Untersuchungsverfahren nach § 35 LMBG. Beuth Verlag Berlin, Köln.
- 11. Deutsches Arzneibuch, 10. Auflage, Chapter VIII, 10.
- 12. DIN Deutsches Institut für Normung: Untersuchung von Fleisch und Fleischerzeugnissen, Nachweis von Salmonellen (Referenzverfahren), DIN 10160.
- DIN Deutsches Institut für Normung e.V.: Mikrobiologische Milchuntersuchung. Nachweis von Salmonellen. Referenzverfahren.