XLD (Xylose Lysine Deoxycholate) Agar

XLD Agar

EMD Cat. No. 1.05287.0500/5007

500 g, 5 kg

Medium proposed by TAYLOR (1965), TAYLOR and HARRIS (1965, 1967) and TAYLOR and SCHELHART (1967) for the isolation and differentiation of pathogenic Enterobacteriaceae, especially of Shigella and Salmonella species.

This culture medium complies with the recommendations of the United States Pharmacopeia XXIII (1995), the European Pharmacopeia II and the APHA (1992).

Mode of Action	Preparation
Experimental Procedure and Evaluation	Quality Control
Typical Composition (g/liter)	Literature
Picture/Image

ChemDat®

Mode of Action
Degradation of xylose, lactose and sucrose to acid causes phenol red to change its color to yellow. Production of hydrogen sulfide is indicated by thiosulfate and iron(III) salt, which react to form a precipitate of black iron sulfide in the colonies. Bacteria which decarboxylate lysine to cadaverine can be recognized by the appearance of a purple coloration around the colonies due to an increase in pH.
These reactions can proceed simultaneously or successively, this may cause the pH indicator to exhibit various shades of color or it may change its color from yellow to red on prolonged incubation. The culture medium is weakly inhibitory.

Preparation

1.	Weigh out 55 g of XLD Agar.
2.	Add 50 ml of purified water to a flask
3.	Transfer 55 g of XLD Agar gently to flask with swirling.
4.	Mix thoroughly, add remaining 950 ml purified water, until completely suspended. Check for lumps. If present repeat mixing.
5.	Heat to boiling to dissolve completely.
6.	Immediately cool the medium to about 47-50 °C in a waterbath set at this temperature. Agitate flask to cool rapidly.
7.	Pour plates.
8.	Dry plates and check for sterility prior to use.

Note: preparation of large volumes, overheating and prolonged storage in water bath (47-50 °C) should be avoided.

Do not autoclave.
pH: 7.4 ± 0.2 at 25 °C.
The plates are clear and red.
Crystalline precipitate of salts may occur. To avoid this, the liquid medium needs to be filtered through a flute-formed filter.

Experimental Procedure and Evaluation
Inoculate by spreading the material thinly on the surface of the plates.
Incubation: up to 48 hours at 35 °C aerobically.

Further tests should be performed in order to identify the colonies.

Appearance of Colonies	Microorganisms
Yellow, surrounded by yellow zones, opaque with precipitation zones	Escherichia coli, Enterobacter, Aeromonas
Yellow, surrounded by yellow zones, opaque, mucoid with precipitation zones	Klebsiella
Yellow, surrounded by yellow zones, opaque, sometimes with a black centre	Citrobacter (lactose-positive strains)
Yellow, surrounded by yellow zones, opaque,	Serratia, Hafnia
Yellow, surrounded by yellow zones, translucent, black centre	Proteus vulgaris, most Proteus mirabilis
Colonies have the same color as the culture medium, translucent, sometimes with a black centre	Salmonella
Colonies have the same color as the culture medium, translucent	Shigella, Providencia, Pseudomonas
Orange, slightly opaque	Salmonella typhosa (xylose-positive strains)

According to EMD internal data, E. coli generally does not grow on XLD when inoculated directly from lyophilized forms that are commercially available (e.g., Quanti-Cult®, Kwik-Stik™, BioBall™, etc). In order to see any E. coli growth on XLD, it is necessary to: 1) Pre-enrich the E. coli overnight (18-48 hours) in Tryptic Soy Broth or similar, then 2) Dilute the enriched sample to reach the desired inoculum level (10 - 100 cfu/ml acc. to USP 30) and then 3) Inoculate the XLD using this diluted E. coli culture.

Quality control (spiral plating method)

Test strains	Inoculum (cfu/ml)	Recovery rate %	Colony color	Black centre	color change of medium
Escherichia coli ATCC 25922	> 10⁵	not limited	yellow	-	yellow + precipitate
Enterobacter cloacae ATCC 13047	10³-10⁵	30	yellow	-	yellow + precipitate
Klebsiella pneumoniae ATCC 13883	10³-10⁵	30	yellow	-	yellow + precipitate
Shigella flexneri ATCC 12022	10³-10⁵	10	colorless	-
Shigella sonnei ATCC 11060	10³-10⁵	10	colorless	-
Salmonella typhimurium ATCC 14028	10³-10⁵	30	colorless	+	-

Quality control (spiral plating method)

Test strains	Inoculum (cfu/ml)	Recovery rate %	Colony color	Black centre	color change of medium
Salmonella enteritidis NCTC 5188	10³-10⁵	30	colorless	+	-
Proteus mirabilis ATCC 14273	10³-10⁵	30	yellow	+	yellow / orange
Enterococcus faecalis ATCC 11700	> 10⁵	0.01		-

Typical Composition (g/liter)
Yeast extract 3.0; sodium chloride 5.0; D(+)xylose 3.75; lactose 7.5; sucrose 7.5; L(+)lysine 5.0; sodium deoxycholate 1.0; sodium thiosulfate 6.8; ammonium iron(III) citrate 0.8; phenol red 0.08; agar-agar 14.5.

Literature

American Public Health Association. Compendium of Methods for the microbiological Examination of Foods. 3rd ed. (1992).
BHAT, P., a. RAIAN, D.: Comparative evaluation of deoxycholate citrate medium and xylose lysine deoxycholate medium in the isolation of shigellae. Am. J. Clin. Pathol.,
64; 99-404 (1975).
DUNN, C., a. MARTIN, W.J.: Comparison of media for isolation of Salmonellae and Shigellae from fecal specimens. Appl. Microbiol., 22; 17-22 (1971).
European Pharmacopeia II, Chapter VIII, 10.
ROLLENDER, W., BECKFORD, O., BELSKY, R.D., a. KOSTROFF, B.: Comparison of xylose lysine deoxycholate agar and MacCONKEY Agar for the isolation of Salmonella and Shigella from clininal specimens. Am. J. Clin. Pathol., 51/2; 284-386 (1969).
TAYLOR, W.J.: Isolation of Shigellae. I. Xylose lysine agars: new media for isolation of enteric pathogens. Am. J. Clin. Path., 44; 471-475 (1965).
TAYLOR, W.J., a. HARRIS, B.: Isolation of Shigellae. II. Comparison of plating media and enrichment broths. Am. J. Clin. Path., 44X 476-479 (1965).
TAYLOR, W.J., a. HARRIS, B.: Isolation of Shigellae. III. Comparison of new and traditional media with stool specimens. Amer. J. Clin. Pathol., 48; 350-355 (1967).
TAYLOR, W.J., a. SCHELHART, D.: Isolation of Shigellae. IV. Comparison of plating media with stools. Amer. J. Clin. Pathol., 48; 356-362 (1967).
TAYLOR, W.J., a. SCHELHART, D.: Isolation of Shigellae. V. Comparison of enrichment broth with stools. Appl. Microbiol., 16; 1383-1386 (1968).
United States Pharmacopeia XXIII, Chapter "Microbioal Limit Tests", 1995.

BioBall is a trademark of BTF Pty Ltd, NSW, Australia
Quanti-Cult is a registered trademark of Remel Inc.
KWIK-STIK is a trademark of MicroBioLogics, Inc., St. Cloud, MN

Salmonella Typhimurium ATCC 14028

XLD Agar - Klebsiella pneumoniae