B.C.Y.E. AGAR

Technical Data #1251a / 98.05.01

B.C.Y.E. Agar is a solid selective medium used for the isolation and identification of Legionella pneumophila and other Legionella sp. from clinical specimens and environmental samples. L-Cysteine, ferric pyrophosphate, alpha-ketoglutarate and yeast extract are the growth factors.

FORMULA
in grams per liter purified filtered water

Yeast Extract	11,5
Charcoal, activated	1,5
ACES Buffer	6,0
alpha-Ketoglutarate	1,0
Agar	17,0
Growth Factors
L-Cysteine HCl	0,4
Ferric Pyrophosphate	0,25

pH 6,9 +/- 0,1 à 25º C

This approximate formula may be adjusted and/or enriched to obtain best results.

PRECAUTIONS
This medium is for IN VITRO diagnostic use only.

STORAGE
Store prepared media at 2-8º C protected from direct light and dehydrated powder, in a dry place, in tightly-sealed containers at 2-25º C.

SIGN OF DETERIORATION
Media should not be used if the expiry date has passed. Prepared media should not be used if there are signs of contamination or deterioration (shrinking, cracking, evaporation or discoloration). Do not use dehydrated media if it is caked.

DIRECTIONS
Suspend 37 g of dehydrated media in 1000 ml of purified filtered water. Do not heat prior to sterilization. Sterilize at 121º C for 15 minutes. Cool to 45-50º C. Aseptically add 2x5 ml of Legionella Supplement (8708), adjust pH with sterile solution of KOH 1N. Mix gently and dispense into sterile Petri dishes.

PROCEDURE
Prior to inoculate, the prepared media should be brought to room temperature.

1. Directly, inoculate plates with specimens and streak for isolation.
2. Incubate at 35º C in an humidified atmosphere for a minimum of 3 days.
3. Examine daily for growth.

QUALITY CONTROL
Results after 72 hrs at 35ºC under humid atmosphere

Organisms	ATCC	Growth
Legionella pneumophila	33152	+
Legionella bozemanii	33217	+
Legionella dumoffii	33278	+

Colonies of L. pneumophila are gray-white on this medium and give yellow-green fluorescence under UV light.

LIMITATIONS OF METHOD
This medium is only a part of the identification. Other tests may be required.

REFERENCES

1. Balows, A., W.J. Hausler Jr., K.L. Herrmann, H.D. Isenberg, J. Shadomy. 1991 Manual of Clinical Microbiology 5th ed. American Society for Microbiology. Washington, D.C.
2. Buesching, W.J., R.A. Brust, L.W. Ayers. 1983 Enhanced Primary Isolation of Legionella pneumophila from Clinical specimens by Low-pH Treatment. J. Clin. Microbiol. 17:1153-1155.
3. Edelstein, P., 1981 Improved Semisective Medium for Isolation of Legionella pneumophila from Contaminated Clinical and Environmental Specimens. J. Clin. MIcrobiol. 14:298-303.
4. Feeley, J.C., G.W. Gorman, R.E. Weaver, D.C. Mackel, H.W. Smith. 1978 Primary Isolation Media for Legionnaires Disease Bacterium J. Clin. Microbiol. 8:320-325.
5. Feeley, J.C., R.J. Gibson, G.W. Gorman, N.C. Langford, J.R. Rasheed, D.C. Mackel, W.B. Baine 1979 Charcoal-Yeast Extract Agar: Primary Isolation Medium for Legionella pneumophila. J. Clin. Microbiol. 10:437-441.
6. Ristroph, J.D., K.W. Hedlund, S. Gowda. 1981 Chemically Defined Medium for Legionella pneumophila Growth. J. Clin. Microbiol. 13:115-119.

CATALOGUE NUMBERS

Dehydrated media	500 g	QB-39-2420
Prepared media
Plates, 100x15 mm	10/pkg	1251
Mono-Plates, 100x15 mm	10/pkg	1251M
Legionella Supplement, 5 ml	10/pkg	8708

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