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Dehydrated Culture Media

DESOXYCHOLATE AGAR

Code: CM0163

A differential medium for the enumeration of coliforms in dairy products. It may be employed as a non-selective medium for the isolation of enteric pathogens.

Typical Formula*

gm/litre

Peptone

10.0

Lactose

10.0

Sodium desoxycholate

1.0

Sodium chloride

5.0

Dipotassium hydrogen phosphate

2.0

Ferric citrate

1.0

Sodium citrate

1.0

Neutral red

0.03

Agar

15.0

pH 7.1 ± 0.2 @ 25°C

 
* Adjusted as required to meet performance standards

Directions
Suspend 45g in 1 litre of distilled water. Bring to the boil over gauze and flame to dissolve the medium completely. Agitate to prevent charring.
THIS MEDIUM IS HEAT SENSITIVE. AVOID EXCESSIVE OR PROLONGED HEATING DURING RECONSTITUTION. DO NOT AUTOCLAVE OR REMELT.

Description
Desoxycholate Agar is a differential medium for the direct count of coliforms in dairy products (American Public Health Association1). It may also be employed for the isolation of enteric pathogens from rectal swabs, faeces, or other specimens.

The medium may be used in a `pour-plate’ technique or as a surface inoculated medium. A thin layer of uninoculated desoxycholate agar poured over the surface of a gelled `pour-plate’ assists subsequent counting.

Technique
Enumeration of Coliforms in Milk and Cream (APHA1)
1. Pipette 1-4ml of the sample (or decimal dilution of the sample) into a sterile Petri dish.
2. Cool freshly prepared Desoxycholate Agar to 42-44°C and add 10-20ml to each dish.
3. Mix the contents of the dishes by gentle tilting and rotation.
4. Allow the plates to solidify and pour on an overlay of 3-4ml of uninoculated Desoxycholate Agar.
5. When the overlay has set, invert the plates and incubate them for 18-24 hours at 35°C.
6. Count all dark red colonies measuring at least 0.5mm in diameter, and calculate the number of coliform colonies per millilitre or gram of original sample.

Isolation of Enterobacteriaceae
It is advisable to use Desoxycholate Agar in parallel with other plating media for this purpose.

Lightly inoculate a Desoxycholate Agar plate with faeces, rectal swab, or enrichment culture. Incubate for 18-24 hours at 35°C and examine. Non-lactose fermenters of enteric origin form colourless colonies. Non-lactose fermenters which are not of enteric origin are generally inhibited by the sodium desoxycholate in the medium. Identify suspect colonies in the usual manner.

Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
Store the prepared agar plates at 2-8°C.

Appearance
Dehydrated medium: Straw-pink coloured, free-flowing powder
Prepared medium: Pink coloured gel

Quality control

Positive controls:

Expected results

Lactose Fermenters 
Escherichia coli ATCC® 25922*

Good growth; pink coloured colonies with
bile precipitation

Non-Lactose Fermenters 

Shigella sonnei ATCC® 25931

Good growth; straw coloured colonies

Negative control:  
Staphylococcus aureus ATCC® 25923*Inhibited
* This organism is available as a Culti-Loop®

Precautions
As with all desoxycholate media, this medium is heat sensitive. Observe the precautions stated under Directions.

Reference
1. American Public Health Association (1978) `Standard Methods for the Examination of Dairy Products’ 14th ed,. APHA Inc., New York, pp. 58-59.

 
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