XLD AGAR

Cat. no. G245 XLD Agar with Novobiocin, 15x100mm Plate, 18ml 10 plates/bag

INTENDED USE

Hardy Diagnostics XLD Agar is recommended for use as a selective and differential medium for the isolation of gram-negative enteric pathogens from fecal specimens or from food and other samples.

This product is not intended to be used for the diagnosis of human disease.

SUMMARY

Xylose Lysine Deoxycholate (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.(6) XLD Agar was especially designed to allow 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 as well.

The selective agent in XLD Agar is sodium deoxycholate, which inhibits the growth of gram-positive organisms. The carbohydrate source is xylose which is fermented by most enterics except for Shigella species, and these colonies appear red on this medium as a result. A second differential mechanism for Salmonella is employed by the addition of lysine. Lysine decarboxylation reverts the pH of the medium to an alkaline condition. To avoid this reversal to a Shigella reaction, lactose and sucrose are added in excess. The addition of sodium thiosulfate and ferric ammonium citrate as a sulfur source and indicator, respectively, allows hydrogen sulfide forming organisms to produce colonies with black centers, under alkaline conditions. Organisms which ferment xylose, are lysine decarboxylase-negative, and do not ferment lactose or sucrose cause an acid pH in the medium, and form yellow colonies. Examples of such organisms are Citrobacter spp., Proteus spp., and Escherichia coli.

XLD Agar with Novobiocin contains novobiocin, which is commonly used to inhibit the growth of Proteus spp., and helps reduce the potential for false-positives from this organism.

FORMULA

Ingredients per liter of deionized water:*

Lactose 7.5gm
Sucrose 7.5gm
Sodium Thiosulfate 6.8gm
L-Lysine 5.0gm
Sodium Chloride 5.0gm
Xylose 3.75gm
Yeast Extract 3.0gm
Sodium Deoxycholate 2.5gm
Ferric Ammonium Citrate 0.8gm
Phenol Red 0.08gm
Agar 15.0gm
Novobiocin 10.0mg/mL

Final pH 7.4 +/- 0.2 at 25ºC.

* Adjusted and/or supplemented as required to meet performance criteria.

STORAGE AND SHELF LIFE

Storage: Upon receipt store at 2-8ºC. away from direct light. Media should not be used if there are any signs of deterioration (shrinking, cracking, or discoloration), contamination, or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat, moisture, and freezing.

PRECAUTIONS

PROCEDURE

Specimen Collection: Consult listed references for information on specimen collection.(1-3,5) Infectious material should be submitted directly to the laboratory without delay and protected from excessive heat and cold. If there is to be a delay in processing, the specimen should be inoculated onto an appropriate transport media and refrigerated until inoculation.

Method of Use: Allow the plates to warm to room temperature, and the agar surface to dry before inoculating. Inoculate and streak the specimen as soon as possible after collection. If the specimen to be cultured is on a swab, roll the swab over a small area of the agar surface. Streak for isolation with a sterile loop. Incubate plates aerobically at 35-37ºC. for 18-24 hours. Examine colonial morphology, characteristics, and hemolytic reactions.

It is recommended that selective enrichment broths, such as GN Broth or Selenite Cystine Broth (Cat. no. K39, or K69, respectively), be used in conjunction with other selective plating medias, such as HE Agar (Cat. no. G63). This recommendation is made in order to maximize the recovery of enteric pathogens.

INTERPRETATION OF RESULTS

Salmonella spp. appear as red colonies with black centers. Lysine-positive organisms appear red. Shigella spp. also appear red. Other lysine-negative fermenters, such as E. coli, Citrobacter and Proteus spp. appear yellow. Consult listed references for the identification of colony morphology and further biochemical tests required for identification.(1-3,5)

LIMITATIONS

Some species of Salmonella may form red colonies without a black center, which resemble Shigella colonies. In addition, a few species of Shigella ferment lactose, and Salmonella that fail to decarboxylate lysine would not be detected on this medium.

Processing delays of over 2-3 hours of unpreserved stool specimens greatly jeopardizes the recovery of many enteric pathogens, as these organisms are very susceptible to the acidic changes that occur with a temperature drop of the feces.

Red, false-positive colonies may occur with Proteus and Pseudomonas

Incubation in excess of 48 hours may lead to false-positive results.

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, slides, staining supplies, other culture media, microscopes, incinerators, and incubators, etc., as well as serological and biochemical reagents, are not provided.

QUALITY CONTROL

Test Organisms
Inoculation Method*
Incubation
Results
Time
Temperature
Atmosphere
Salmonella enterica
ATCC® 14028
A 24hr 35°C Aerobic Growth; red colonies with black centers
Shigella flexneri
ATCC® 12022
A 24hr 35°C Aerobic Growth; red to pink colonies
Enterococcus faecalis
ATCC® 29212
B 24hr 35°C Aerobic Partial to complete inhibition; clear, pinpoint colonies
Escherichia coli
ATCC® 25922
B 24hr 35°C Aerobic Partial to complete inhibition; yellow to yellow red colonies

User Quality Control

PHYSICAL APPEARANCE

XLD Agar should appear a clear, and red in color, and may have a slight precipitate.

S. typhimurium growing on XLD Agar

Salmonella enterica (ATCC® 14028) colonies growing on XLD Agar. Incubated aerobically for 24 hours at 35ºC.

S. flexneri growing on XLD Agar

Shigella flexneri (ATCC® 12022) colonies growing on XLD Agar. Incubated aerobically for 24 hours at 35ºC.

E. faecalis inhibited on XLD Agar

Enterococcus faecalis (ATCC® 29212) growth inhibited on XLD Agar. Incubated aerobically for 24 hours at 35ºC.

REFERENCES

1. Anderson, N.L., et al. Cumitech 3B; Quality Systems in the Clinical Microbiology Laboratory, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C.

2. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

3. Isenberg, H.D. Clinical Microbiology Procedures Handbook, Vol. I, II & III. American Society for Microbiology, Washington, D.C.

4. MacFaddin, J.F. 1985. Media for Isolation, Cultivation, Identification, Maintenance of Bacteria, Vol. I. Williams & Wilkins, Baltimore, MD.

5. Versalovic, J., et al. Manual of Clinical Microbiology. American Society for Microbiology, Washington, D.C.

6. Taylor, E.I. 1965. Am. J. Clin. Path. ; 44:471.

7. Quality Assurance for Commercially Prepared Microbiological Culture Media, M22. Clinical and Laboratory Standards Institute (CLSI - formerly NCCLS), Wayne, PA.

ATCC is a registered trademark of the American Type Culture Collection.

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