XLD Agar - Hardy Diagnostics - G65

XLD AGAR Cat. no. G65 XLD Agar, 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 and other specimens. 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. 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 Final pH 7.4 +/- 0.2 at 25 degrees C. * Adjusted and/or supplemented as required to meet performance criteria.

STORAGE AND SHELF LIFE Storage: Upon receipt store at 2-8 degrees C. 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. The expiration date applies to the product in its intact packaging when stored as directed. This product has the following shelf life from the date of manufacture: 90 Days: G65 XLD Agar Refer to the keyword ""Storage"", in the Hardy Diagnostics software program HUGO™, for more information on storing culture media.

PRECAUTIONS This product is for in vitro diagnostic use only and is to be used only by adequately trained and qualified laboratory personnel. Observe approved biohazard precautions and aseptic techniques. All laboratory specimens should be considered infectious and handled according to ""standard precautions"". The ""Guideline for Isolation Precautions"" is available from the Centers of Disease Control and Prevention at www.cdc.gov/ncidod/dhqp/gl_isolation.html. For additional information regarding specific precautions for the prevention of the transmission of all infectious agents from laboratory instruments and materials, and for recommendations for the management of exposure to infectious disease, refer to CLSI document M29. Sterilize all biohazard waste before disposal. Refer to the keyword ""Precautions"", in the Hardy Diagnostics software program HUGO™, for more information regarding general precautions when using culture media. Refer to the keyword ""MSDS"", in the Hardy Diagnostics software program HUGO™, for more information on handling potentially hazardous material. 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 35-37 degrees C. 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 It is recommended that biochemical and/or serological tests be performed on colonies from pure culture for complete identification, as 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. Refer to the keyword ""Limitations"", in the Hardy Diagnostics software program HUGO™, for more information regarding general limitations on culture media. 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 The following organisms are routinely used for testing at Hardy Diagnostics: Test Organisms Inoculation Method* Incubation Results Time Temperature Atmosphere Salmonella typhimurium 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 Check for signs of contamination and deterioration. Users of commercially prepared media may be required to perform quality control testing with at least one known organism to demonstrate growth or a positive reaction; and at least one organism to demonstrate inhibition or a negative reaction (where applicable). Refer to the following keywords, in the Hardy Diagnostics software program HUGO™, for more information on QC: ""Introduction to QC"", ""QC of Finished Product"", and ""The CLSI (NCCLS) Standard and Recommendations for User QC of Media"". Also see listed references for more information.(1-3,5,7) * Refer to the keyword ""Inoculation Procedures"", in the Hardy Diagnostics software program HUGO™, for a description of inoculation procedures. PHYSICAL APPEARANCE XLD Agar should appear a clear, and red in color. Salmonella typhimurium (ATCC® 14028) colonies growing on XLD Agar (Cat. no. G65). Incubated aerobically for 24 hours at 35 deg. C. Shigella flexneri (ATCC® 12022) colonies growing on XLD Agar (Cat. no. G65). Incubated aerobically for 24 hours at 35 deg. C. Enterococcus faecalis (ATCC® 29212) growth inhibited on XLD Agar (Cat. no. G65). Incubated aerobically for 24 hours at 35 deg. C. REFERENCES 1. August, M.J., et al. 1990. Cumitech 3A; Quality Control and Quality Assurance Practices in Clinical Microbiology, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C. 2. Forbes, B.A., et al. 1998. Bailey and Scott's Diagnostic Microbiology, 10th ed. C.V. Mosby Company, St. Louis, MO. 3. Isenberg, H.D. Clinical Microbiology Procedures Handbook, Vol. I & II. 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. Murray, P.R., et al. 1995. Manual of Clinical Microbiology, 6th ed. 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-A2, Vol. 16, No. 16. 1996. Clinical Laboratory Standards Institute (CLSI - formerly NCCLS), Villanova, PA.