What is XLD agar?

Xylose Lysine Deoxycholate (XLD) Agar is a selective culture medium used for the isolation of Salmonella and Shigella species from clinical and food samples. XLD Agar was developed by Taylor for the differentiation, isolation and identification of enteric pathogens and to support the growth of more fastidious enteric organisms. XLD Agar was specially designed to support the growth of Shigella species and is a proven medium for the isolation of this organism. It has also been found to be an excellent medium for isolating Salmonella species. It has a pH of about 7.4, leaving it looking bright pink or red due to the phenol red indicator.

Sugar fermentation lowers the pH and the phenol red indicator registers this by turning yellow. Most intestinal bacteria, including Salmonella, can ferment the sugar xylose to produce acid; Shigella colonies cannot do this and therefore remain red. After depleting the xylose supply, Salmonella colonies will decarboxylate lysine, raising the pH once again to alkaline and mimicking red Shigella colonies. Salmonellae metabolize thiosulfate to produce hydrogen sulfide, leading to the formation of colonies with black centres and distinguishing them from similarly coloured Shigella colonies.

XLD agar principle

XLD Agar is both a selective and differential medium. The medium contains yeast extract, which provides nitrogen and vitamins necessary for growth. It uses sodium deoxycholate as a selective agent and thus inhibits gram-positive microorganisms. Although the sugars xylose, lactose, and sucrose provide fermentable carbohydrate sources, xylose is primarily incorporated into the medium as it is not fermented by Shigella but by virtually all enterics. This helps in the differentiation of Shigella species. Sodium chloride maintains the osmotic balance of the medium. Lysine is included to differentiate the Salmonella group from non-pathogens. Salmonella rapidly ferments xylose and depletes the supply.

Lysine is subsequently decarboxylated by the enzyme lysine decarboxylase to form amines with reversion to alkaline pH mimicking the Shigella reaction. However, to prevent this reaction by lysine-positive coliforms, lactose and sucrose are added to produce excess acid. The degradation of xylose, lactose, and sucrose to acid causes the phenol red indicator to change its colour to yellow. Bacteria that decarboxylate lysine to cadaverine can be recognized by the appearance of a red colouration around the colonies due to an increase in pH.

These reactions can occur simultaneously or successively, and this can cause the pH indicator to show various shades of colour or change colour from yellow to red with prolonged incubation. To add to the differentiation ability of the formulation, an H2S indicator system, consisting of sodium thiosulfate and ferric ammonium citrate, is included for visualization of hydrogen sulfide produced, resulting in the formation of colonies with black centres. Nonpathogenic H2S producers do not decarboxylate lysine; therefore, the acid reaction they produce prevents the blackening of the colonies.

Preparation of XLD Agar

  • Suspend 56.68 grams in 1000 ml of distilled water.
  • Heat with frequent stirring until the medium boils.
  • Immediately transfer to a 50°C water bath.
  • After cooling, pour into sterile Petri dishes.
  • It is advisable not to prepare large volumes that require prolonged heating, producing precipitates, producing precipitates.

XLD Agar Uses

  • XLD Agar is a selective differential medium for the isolation of gram-negative enteric pathogens from faecal samples and other clinical material.
  • It is especially suitable for the isolation of Shigella and Salmonella species.
  • Microbiological analysis of food, water and dairy products.

Limitations of XLD agar

  • Some strains of Proteus can give a red to yellow colouration and most colonies develop black centres, leading to false-positive reactions.
  • Non-enteric organisms such as Pseudomonas and Providencia may exhibit red colonies.
  • S. Paratyphi A, S. Choleraesuis, S. Pullorum, and S. Gallinarum can form red colonies without H2S, resembling Shigella.
  • Incubation for more than 48 hours may lead to false-positive results.
  • Biochemical, immunological, molecular, or mass spectrometry tests on colonies from pure cultures are recommended for complete identification.