XLD Agar- Composition, Principle, Preparation, Results, Uses

XLD Agar is a type of culture medium that is used in microbiology to isolate and identify certain bacteria that cause infections in humans and animals. The name XLD stands for Xylose Lysine Deoxycholate, which are the main ingredients of the medium. XLD Agar is designed to selectively grow Gram-negative bacteria, especially Salmonella and Shigella species, while inhibiting the growth of Gram-positive bacteria and most other Gram-negative bacteria. XLD Agar also allows the differentiation of Salmonella and Shigella from other bacteria based on their ability to ferment sugars, decarboxylate lysine, and produce hydrogen sulfide.

XLD Agar contains the following ingredients per liter of distilled water:

• Yeast extract: 3 g
• L-Lysine hydrochloride: 5 g
• Xylose: 3.75 g
• Lactose: 7.5 g
• Sucrose: 7.5 g
• Sodium chloride: 5 g
• Sodium deoxycholate: 2.5 g
• Sodium thiosulfate: 6.8 g
• Ferric ammonium citrate: 0.8 g
• Phenol red: 0.08 g
• Agar: 15 g

The final pH of the medium is 7.4 ± 0.2 at 25°C.

The composition of XLD Agar is designed to achieve both selective and differential properties for the isolation and identification of enteric pathogens, especially Salmonella and Shigella species.

The yeast extract provides nitrogen and vitamins required for bacterial growth.

The sodium deoxycholate is a bile salt that inhibits the growth of gram-positive bacteria and some gram-negative bacteria, such as Proteus and Pseudomonas.

The xylose, lactose, and sucrose are fermentable carbohydrates that serve as sources of energy and carbon for the bacteria. They also help to differentiate the bacteria based on their ability to ferment these sugars and produce acid.

The L-lysine hydrochloride is an amino acid that can be decarboxylated by some bacteria, such as Salmonella, to produce alkaline products and increase the pH of the medium.

The sodium chloride maintains the osmotic balance of the medium.

The sodium thiosulfate and ferric ammonium citrate are components of the hydrogen sulfide indicator system. Some bacteria, such as Salmonella, can reduce thiosulfate to sulfide, which reacts with ferric ions to form a black precipitate.

The phenol red is a pH indicator that changes color from red to yellow in acidic conditions and from yellow to red in alkaline conditions.

The agar is a solidifying agent that provides a firm surface for bacterial growth and colony formation.

XLD Agar is both a selective and differential medium. It allows the growth of gram-negative bacteria while inhibiting the growth of gram-positive bacteria. It also distinguishes between different types of gram-negative bacteria based on their ability to ferment sugars, decarboxylate lysine, and produce hydrogen sulfide.

The medium contains four main components that are responsible for its selective and differential properties:

• Xylose: This is a sugar that can be fermented by most enteric bacteria, except Shigella. The fermentation of xylose lowers the pH of the medium and causes the color change of the indicator phenol red from red to yellow. Therefore, xylose fermentation helps to differentiate Shigella (red colonies) from other enteric bacteria (yellow colonies).
• Lysine: This is an amino acid that can be decarboxylated by some enteric bacteria, such as Salmonella. The decarboxylation of lysine raises the pH of the medium and reverses the color change of phenol red from yellow to red. Therefore, lysine decarboxylation helps to differentiate Salmonella (red colonies with black centers) from other enteric bacteria (yellow colonies or red colonies without black centers).
• Lactose and sucrose: These are sugars that can be fermented by some enteric bacteria, such as Escherichia coli and Klebsiella. The fermentation of lactose and sucrose produces more acid than xylose fermentation and prevents the reversal of pH by lysine decarboxylation. Therefore, lactose and sucrose fermentation helps to differentiate E. coli and Klebsiella (yellow colonies) from Salmonella (red colonies with black centers).
• Sodium thiosulfate and ferric ammonium citrate: These are compounds that act as a hydrogen sulfide indicator system. Some enteric bacteria, such as Salmonella and Proteus, can reduce sodium thiosulfate to hydrogen sulfide gas. The hydrogen sulfide gas reacts with ferric ammonium citrate to form a black precipitate of ferric sulfide. Therefore, hydrogen sulfide production helps to differentiate Salmonella and Proteus (black colonies or black centers) from other enteric bacteria (no blackening).

By combining these four components, XLD Agar can effectively isolate and identify Salmonella and Shigella species from clinical samples and food products.

To prepare XLD Agar, you will need the following ingredients and equipment:

• XLD Agar powder (56.68 grams)
• Distilled water (1000 ml)
• A large beaker or flask
• A stirring rod or a magnetic stirrer
• A thermometer
• A water bath
• Sterile Petri plates
• A laminar flow hood or a sterile workbench

Follow these steps to prepare XLD Agar:

1. Measure 1000 ml of distilled water and pour it into the beaker or flask.
2. Add 56.68 grams of XLD Agar powder and stir well to dissolve. You can use a magnetic stirrer to speed up the process.
3. Heat the mixture with frequent agitation until it boils. Do not autoclave or overheat the medium, as this may cause precipitation or loss of selective properties.
4. As soon as the medium boils, transfer it immediately to a water bath set at 50°C. This will prevent the medium from solidifying and maintain its sterility.
5. After cooling, pour the medium into sterile Petri plates under a laminar flow hood or a sterile workbench. You can use a sterile pipette or a dispenser to measure the volume of medium for each plate. The recommended volume is 15-20 ml per plate.
6. Allow the plates to solidify completely before storing them in a refrigerator at 2-8°C. Do not invert the plates, as this may cause condensation to drip onto the surface of the medium and affect its performance.
7. Use the plates within two weeks of preparation. Before use, check the plates for any signs of contamination, discoloration, or deterioration.

You have successfully prepared XLD Agar for your microbiological experiments! 😊

The results of XLD Agar depend on the type and characteristics of the bacteria that grow on the medium. The following table summarizes the typical reactions of some common enteric pathogens and normal flora on XLD Agar:

The colony color is determined by the fermentation of xylose, lactose, and sucrose, and the decarboxylation of lysine by the bacteria. The H2S production is indicated by the formation of black precipitate in the colonies or the surrounding medium.

The following images show some examples of XLD Agar results:

XLD Agar is a selective differential medium for the isolation of Gram-negative enteric pathogens from fecal specimens and other clinical material. It is especially suitable for the isolation of Shigella and Salmonella species, which are significant causative agents of gastrointestinal infections.

XLD Agar can also be used for microbiological testing of foods, water, and dairy products. It can help detect the presence of Salmonella and Shigella in these samples, which may pose a risk to public health and food safety.

XLD Agar is a valuable tool for the identification and differentiation of enteric pathogens based on their biochemical characteristics. It can help distinguish between Salmonella, Shigella, coliforms, and other Gram-negative bacteria based on their ability to ferment xylose, lactose, and sucrose, decarboxylate lysine, and produce hydrogen sulfide.

XLD Agar is one of the most widely used media for the isolation of Salmonella and Shigella from clinical and non-clinical specimens. It has a high sensitivity and specificity for these organisms and can provide reliable results in a short time. XLD Agar is also easy to prepare and interpret, making it a convenient and cost-effective medium for microbiology laboratories.

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