Re:Re:Re:Mannitol-Egg-yolk-Polymyxine-Agar (MYP-agar)

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Cereus Selective Agar Base acc. to MOSSEL

Mannitol-Egg-yolk-Polymyxine-Agar (M.Y.P.-agar)

Cat. No. 1.05267.0500 (500 g)

Medium proposed by MOSSEL et al. (1967) for the enumeration, detection and isolation of Bacillus cereus in foodstuffs.

The German Regulations for Dietetic Foodstuffs (Verordnung über diätetische Lebensmittel (Diätverordnung)) stipulate that foodstuffs should be tested for Bacillus cereus. The medium furthermore complies with the German DIN-Norm 10198 for the investigation of milk and food and to the requirements acc. to § 35 LMBG (00.00/25) and 01.00/53) for the investigation of food.

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

ChemDAT®

Mode of Action
This culture medium is highly adapted to the properties of Bac. cereus.

a)	Bac. cereus is mannitol-negative. The mannitol content of the medium thus allows differentiation of the accompanying mannitol-positive microbial flora which are identified by a change in colour of the indicator phenol red to yellow.
b)	Bac. cereus is not affected by concentrations of polymyxin which inhibit the common accompanying microbial flora (DONOVAN 1958). Addition of polymyxin is necessary, however, if the sample material is suspected to contain high numbers of accompanying microorganisms.
c)	Bac. cereus produces lecithinase. The insoluble degradation products of egg-yolk lecithin accumulate around the Cereus colonies to form a white precipitate. A lecithinase reaction occurs very early in many strains, Cereus colonies can, therefore, often be rapidly identified before accompanying polymyxin-resistant microorganisms have had a chance to fully develop.

Typical Composition (g/litre)
Peptone from casein 10.0; meat extract 1.0; D(-)mannitol 10.0; sodium chloride 10.0; phenol red 0.025; agar-agar 12.0
Also to be added (per litre of medium):
egg-yolk emulsion 100 ml; polymyxin B sulfate 100,000 IU = Bacillus cereus Selective Supplement.


Preparation

Suspend 21.5 g in 450 ml demin. water, autoclave (15 min
at 121 °C). Cool to about 45 to 50 °C, add 50 ml (this volume can be varied depending on the degree of turbidity desired) of sterile egg-yolk emulsion and the contents of 1 vial Bacillus cereus Selective Supplement, mix. Pour plates.
pH: 7.2 ± 0.2 at 25 °C.
The plates (incl. egg-yolk) are evenly turbid and slightly orange (red without egg-yolk).


Experimental Procedure and Eval‎uation
Inoculate the plates by spreading the sample on the surface of the medium.
Incubation: 18-40 hours at 32 °C.

Bac. cereus appears as rough, dry colonies with a pink to purple base which are surrounded by a ring of dense precipitate. Colonies surrounded by a yellow or a clear zone are not Bacillus cereus. Further tests should be performed to confirm‎ the identity of Bacillus cereus (anaerobic degradation of D(+)glucose, degradation of gelatin, positive nitrate reduction) (BROWN et al. 1958).

Quality control (spiral plating method)

Test strains	Inoculum (cfu/ml)	Recovery rate (%)	Colony colour	Precipitate
Bacillus cereus ATCC 11778	103-105	≥ 40	red	+
Bacillus cereus var. mycoides	103-105	≥ 40	red	+
Bacillus subtilis ATCC 6051	103-105	≥ 40	yellow / pink	+
Escherichia coli ATCC 11775	103-105	≥ 0.01	-
Pseudomonas aeruginosa ATCC 25668	> 105	≥ 0.01	-
Proteus mirabilis ATCC 29906	> 105	Not limited!	red	-
Staphylococcus aureus ATCC 6538-P	103-105	Not limited!	yellow	+

Additives

Merck Cat.No.	Product	Pack Size
1.03784.0001	Egg-yolk emulsion sterile	10 x 100 ml
1.09875.0001	Bacillus cereus Selective Supplement (Polymyxin B; 50,000 IU)	16 vials

Bacillus cereus, Staphylococcus aureus

Literature

BROWN, E.R., MOODY, M.D., TREECE, E.L., a. SMITH, C.W.: Differenzial diagnosis of Bacillus cereus, Bacillus anthracis and Bacillus cereus var. mycoides. – J. Bact., 75; 499-509 (1958).
DONOVAN, K.O.: A selective medium for Bacillus cereus in milk. – J. Appl. Bact., 21; 100-103 (1958).
INAL, T.; Vergleichende Untersuchungen über die Selektivmedien zum qualitativen und quantitativen Nachweis von Bacillus cereus in Lebensmitteln.
I. Mitteilung. – Fleischwirtsch. 51; 1629-1632 (1971).
IV. Mitteilung. – Fleischwirtsch. 52; 1160-1162 (1972).
Bundesgesundheitsamt: Amtliche Sammlung von Untersuchungsverfahren nach § 35 LMBG. – Beuth Verlag Berlin, Köln.
DIN Deutsches Institut für Normierung e.V.: DIN 10198 Teil 1 und 2.
MOSSEL, D.A.A., KOOPMANN, M.J., a. JONGERIUS, E.: Enumeration of Bacillus cereus in foods. – Appl. Microbiol., 15; 650-653 (1967).
NYGREN: Phospholipase C-producing bacteria and food poisoning. An experimental study on Clostridium perfringens and Bacillus cereus. – Acta path. microbiol. scand., 56; Suppl. 1-160 (1962).

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© 2002 Merck KGaA, Darmstadt, Germany

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