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Dehydrated Culture Media

PEPTONE WATER

Code: CM0009

A basal medium to which carbohydrates and indicator may be added for fermentation studies.

Typical Formula*

gm/litre

Peptone

10.0

Sodium chloride

5.0

pH 7.2 ± 0.2

* Adjusted as required to meet performance standards

Directions
Dissolve 15g in 1 litre of distilled water. Mix well and distribute into final containers. Sterilise by autoclaving at 121°C for 15 minutes.
When sterile solutions are to be added after autoclaving, reduce the volume of water for reconstitution by an equal amount.

Description
Peptone Water may be used as a growth medium or as the basis of carbohydrate fermentation media, whilst a pure culture in Peptone Water is a convenient inoculum for a series of fermentation tubes or other diagnostic media.

Peptone Water, adjusted to pH 8.4, is suitable for the cultivation and enrichment of Vibrio cholerae from infected material1.
The medium was formerly used for the performance of the indole test, but now better results can be obtained by the use of Tryptone Water CM0087.

Peptone Water may be modified for use in carbohydrate fermentation tests by the addition of Andrade’s indicator. The indicator which is pink at pH 5.0 and yellow at pH 8.0 is prepared by adding sodium hydroxide to acid fuchsin until it becomes yellow. When added to Peptone Water it is colourless to slightly pink. Filter-sterilised `sugar’ solutions are added to the base medium after sterilisation. These solutions are usually at 10% w/v concentrations and it is important to allow for dilution of the Peptone Water when making up the initial volume of medium. A final concentration of 1% w/v sugar in Peptone Water is normally used but more expensive sugar can be used at 0.5%.

Andrade’s indicator may be made by adding 1N sodium hydroxide to a 0.5% solution of fuchsin until the colour just becomes yellow.
Appropriate safety precautions must be taken to avoid inhalation of, and skin contact with, acid fuchsin.

Both Peptone Water and Andrade Peptone Water are prepared and sterilised in the same manner except that an inverted fermentation tube (Durham tube) to detect gas production is included in Andrade Peptone Water containing glucose. Some organisms will utilise carbohydrate to produce acid only without gas formation. It is unnecessary to add Durham tubes to Peptone Water sugars other than glucose.

Precautions for Andrade Peptone Water sugars
Make sure that each individual bottle of Peptone Water sugar is correctly coded for the contained sugar.

Andrade Peptone Water is reddish-pink when hot; it should return to a colourless or a slightly pink colour when cooled to room temperature.
Some sugar solutions may affect the pH of the Peptone Water; check and correct if so.

Sub-cultures may be necessary to ensure purity of the inoculant. Mixed or contaminated cultures will give false reactions.
Andrade indicator will fade on prolonged storage; do not use beyond the expiry date.

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 medium at room temperature.

Appearance
Dehydrated medium: Straw coloured, free-flowing powder
Prepared medium: Light straw coloured solution

Quality Control
Maintain stock cultures of organisms which have known positive and negative reactions in each sugar. Using fresh sub- cultures, test each batch of sugar medium with the appropriate organisms.

Positive control:

Expected results

Escherichia coli ATCC® 25922 *

Turbid growth

Negative control:

 

Uninoculated medium.

No change

* This organism is available as a Culti-Loop®

Reference
1. Cruickshank R. (1968) `Medical Microbiology’ 11th ed., Livingstone Ltd., London, p. 268.

 
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