TSI (Triple Sugar Iron Agar) is a type of agar that contains three sugars (Purpose)
The Triple Sugar Iron Agar (TSIA) is mainly used to distinguish members of the Enterobacteriaceae family from other Gram-negative rods such as Pseudomonas.
((Principle))\s— TSIA is a rich media that allows bacteria to be distinguished based on their ability to ferment glucose, lactose, sucrose, and sulfur. In addition to the three carbs, the medium contains carbon and nitrogen sources such as beef extract, yeast extract, and peptone, as well as a supply of reducible sulfur in the form of sodium thiosulfate.
The pH indication is phenol red, while the hydrogen sulfide indicator is iron in ferrous sulfate. The medium is made up of a shallow agar slant with a deep butt, which allows for both aerobic and anaerobic growth. A stab in the agar butte is followed by a fishtail streak of the slant to inoculate it.
— Carbohydrate fermentation takes 18 to 24 hours to complete, but hydrogen sulfide reactions might take up to 48 hours.
— Acid products drop the pH and color the entire medium yellow within a few hours when TSIA is inoculated with a glucose-only fermenter.
— Glucose will be depleted in roughly 12 hours because it is in small supply (0.1 percent).
— As the glucose in the aerobic zone (slant) is depleted, organisms in the aerobic region (slant) begin to break down available amino acids, creating NH3 and raising the pH.
— A reversion is a process that takes 18 to 24 hours to complete and occurs exclusively in the slant due to the anaerobic conditions in the butt.
After a 24-hour incubation period, a TSIA with a red slant and yellow butt indicates that the organism ferments glucose but not lactose.
— Organisms that can ferment glucose, lactose, and/or sucrose turn a medium yellow color all over. Because the concentrations of lactose and sucrose are ten times larger than that of glucose, more acid is produced, and both slant and butt will remain yellow after 24 hours.
- As a result, at 24 hours, a TSIA with a yellow slant and butt indicates that the organism ferments glucose plus one or both of the other sugars.
— Gas created during carbohydrate fermentation will cause fissures in the medium or raise the agar off the tube's bottom.
— Hydrogen sulfide (H2S) is created when thiosulfate in the medium is reduced or cysteine in the peptone is broken down.
The H2S combines with the ferrous sulfate in the medium to generate a black precipitate, which is commonly seen in the butt.
— Because thiosulfate reduction requires acidic conditions, black precipitate in the medium indicates sulfur reduction and fermentation. The color of the slant determines which carbs have been fermented (red slant = glucose fermentation, yellow slant = glucose and lactose and/or sucrose fermentation) if the black precipitate obscures the color of the butt.
An organism that uses peptone and amino acids rather than fermenting carbohydrates will alkalinize the medium and colour it red.
— The slant and butt will be red if the organism can utilise the peptone both aerobically and anaerobically. Only the slant will turn red when an obligatory aerobe is present.
— When it comes to reading TSIA results, timing is crucial. Early readings may display yellow across the medium, leading to the incorrect conclusion that the organism is a lactose or sucrose fermenter when it just hasn't exhausted the glucose.
— After lactose and sucrose have been depleted, a yellow butt and red slant may appear, leading to the incorrect conclusion that the organism is a glucose-only fermenter.
Tubes that have been evaluated for carbohydrate fermentation but have not shown evidence of sulfur reduction can be re-incubated for 24 hours before H2S analysis.
0 Comments