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Tsi test
- 1. TSI TEST TRIPLE SUGARIRON AGAR TEST
- 2. Background • TSI standsfor three sugars – glucose, sucrose and lactose. • Russell – used an agar medium with two sugars to aid in the identification of intestinal gram-negative bacilli • Krunweide and Kohn modified Russell’s Double Sugar agar by adding a third sugar—sucrose • Difco Laboratories, Sulkin and Willet, and Hajna described a similar triple sugar ferrous sulfate medium for the identification of enteric bacilli.
- 3. Major Content • Glucose0.1%(used first) • Sucrose 1%(used second) • Lactose 1% • Proteins used third • Phenol red pH indicator for acid production • Ferric ammonium citrate H2S indicator • NaCl maintains osmotic pressure • Agar solidifying agent
- 4. Purpose • used todetermine carbohydrate fermentation and H2S production in bacteria. • Gas from carbohydrate metabolism can also be detected.
- 5. Principle • Carbohydrates pyruvate(acid)+ CO2 • Peptones NH3 (makes medium alkaline) yellow • Phenol red red • Sodium thiosulfate in the medium is reduced by some bacteria to hydrogen sulfide (H2S), a colorless gas. The hydrogen sulfide will react with ferric ions in the medium to produce iron sulfide, a black insoluble precipitate. fermentation fermentation acid alkali
- 6. Medium TSI Agar (Tubedslant) Formula / Liter Enzymatic Digest of Casein ............................................... 5 g Enzymatic Digest of Animal Tissue................... 5 g Yeast Enriched Peptone ................................... 10 g Dextrose ........................................................... 1 g Lactose ............................................................ 10 g Sucrose ...................................................... ..... 10 g Ferric Ammonium Citrate ................................ 0.2 g Sodium Chloride ............................................... 5 g Sodium Thiosulfate ........................................... 0.3 g Phenol Red ...................................................... 0.025 g Agar ................................................................... 13.5 g Final pH: 7.3 ± 0.2 at 250C
- 7. Theory • Fermentation ofcarbohydrates results in the production of acid which decreases the pH of the medium to change from reddish-orange to yellow. • Utilization of peptones results alkalization of the medium due to the production of NH3. • The production of hydrogen sulfide is indicated by the presence of black ppt. formed by the reaction of H2S with ferric ions. • Slant is aerobic while butt is anaerobic. • Gas production is indicated by the splitting of the agar of lifting of it to the top.
- 8. Interpretation • Based oncarbohydrate utilization and hydrogen sulfide production, a TSI slant can be interpreted in several ways:
- 9. Glucose fermenter • tubereaction ---- alkaline over acid (K/A) • Red slant , yellow butt With H2S production (K/A, H2S +ve) • Red slant, black butt
- 10. Glucose, Lactose and/orSucrose Fermenter • Tube reaction --- acid over acid (A/A) • Yellow slant, yellow butt With H2S production(A/A, H2S +ve): • Yellow slant, black ppt. in the butt
- 11. Glucose, Lactose andSucrose Non-fermenters • Tube reaction: i) alkaline over alkaline(K/K) If the bacteria can metabolize peptones both aerobically and anaerobically. both slant and butt red ii) alkaline over no change (K/NC) If peptones can only be metabolized aerobically slant red, butt no change. With H2S production • alkaline over no change (K/NC, H2S +ve) • black precipitate (H2S) in the butt.
- 15. Expected Cultural Response SlantButt Gas H2S Proteus mirabilis K A - + Pseudomonas aeruginosa K K - - E. coli A A + - Shigella flexneri K A - -