WATER PLATE COUNT AGAR (ISO)

CODE: CM1012

A medium for the enumeration of culturable micro-organisms from water using a colony count technique.

Typical Formula*	gm/litre
Tryptone	6.0
Yeast Extract	3.0
Agar	15.0
pH 7.2 ± 0.2 @ 25°C

* Adjusted as required to meet performance standards 

Directions
Suspend 24g of Water Plate Count Agar (ISO) in 1 litre of distilled water and bring gently to the boil to dissolve. If desired, distribute volumes of 15ml to 20ml in tubes, bottles or other containers. Sterilise by autoclaving at 121°C for 15 minutes. After sterilisation, allow to cool and maintain at 45 ± 1°C using a water bath.

Description
A nutritious non-selective medium for all culturable organisms in water which conforms to ISO 6222:1999 (E) Water quality – Enumeration of culturable micro-organisms – Colony count by inoculation in a nutrient agar culture medium.

Waters of all kinds invariably contain a variety of micro-organisms derived from various sources, such as soil and vegetation. Estimation of the overall numbers provides useful information for the assessment and surveillance of water quality. Separate counts are usually made of the micro-organisms which are able to grow and form colonies on nutrient agar media at 36°C and 22°C.

Colony counts are useful for assessing the integrity of ground water sources and the efficiency of water treatment processes such as coagulation, filtration and disinfection and provide an indication of the cleanliness and integrity of the distribution system. They can also be used to assess the suitability of a water supply for the preparation of food and drink to avoid contaminating the product with spoilage organisms.

The main value of colony counts lies in the detection of changes from expected counts, based on frequent, long term monitoring. Any sudden increase in the count can be an early warning of serious pollution and a call for immediate investigation¹.

Technique
Prepare the sample, in accordance with current procedures, and make dilutions if necessary. Place a volume of the test sample (or its dilution), not exceeding 2ml, in a Petri dish. Add 15ml to 20ml of molten Water Plate Count Agar (ISO) and mix carefully by gentle rotation. Allow the medium to set.

The time between addition of the test sample (or its dilution) and the addition of the molten medium shall not exceed 15 minutes. Inoculate at least one plate for incubation at each temperature.

Invert the plates and incubate one set at 36°C ± 2°C for 44 ± 4 hours. Incubate the other set at 22°C ± 2°C for 68 ± 4 hours. Examine the plates immediately after incubation. If this is not possible refrigerate the plates and examine within 48 hours. Reject any plate with confluent growth. 
For each temperature of incubation, count the colonies present in each plate and calculate the estimated number of colony forming units (cfu) present in 1ml of sample.
Express the results as the number of colony forming units per millilitre (cfu/ml) of the sample for each temperature of incubation.

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 plates of medium at 2-8°C.

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

 Quality control

Positive control:	Expected results
Escherichia coli ATCC® 25922 *	Good growth: straw coloured colonies
Negative control:
Uninoculated medium	No change