Triple sugar iron agar

• Contains three sugars (lactose, sucrose, and glucose) as well as iron, within a solid agar medium
  • Peptone as a source of nitrogen
  • Medium is originally reddish due to the phenol red indicator, which turns yellow in acidic conditions
  • Tubes contain a well-oxygenated slant and a poorly-oxygenated butt which need to be interpreted separately
  • Inoculated with a stab down towards the butt followed by streaking the slant
• After 18 to 24 hours at 35°C with a loose cap, the final colour depends on the metabolism of the four components
  • Glucose alone fermented: butt turns yellow (acid), slant stays red (alkaline) (Alk/A)
  • Lactose or sucrose fermented: both butt and slant turn yellow (acid) (A/A)
  • No fermentation of glucose, lactose, or sucrose: both butt and slant stay red (Alk/Alk)
  • Iron: if hydrogen sulfide (H₂S) is produced, the iron will turn black
  • Gas may be formed with active fermentation of either glucose or lactose/sucrose

Interpretation

• Blackening suggests H₂S production

Slant	Butt	Gas	H2S	Fermentation	Examples
Red	Red	No	No	—	Pseudomonas
Red	Yellow	No	No	G	Shigella, Serratia
Yellow	Yellow	Yes	No	G/L/S	Escherichia, Klebsiella, Enterobacter
Yellow	Yellow	Yes	Yes	G/L/S
Red	Yellow	Yes	Yes	G	Salmonella, Proteus

Further Reading

• Microbe Online. Triple Sugar Iron Agar (TSI): Principle, Procedure and Interpretation