Peptone Water: Introduction, Principle, Composition, Preparation, Procedure, Colony Morphology, Uses and Keynotes

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4 years ago

Peptone Water: Introduction, Principle, Composition, Preparation, Procedure, Colony Morphology, Uses and Keynotes

Introduction of Peptone Water

Peptone water is the formulation of Shread, Donovan, and Lee. It is the simplest broth medium used for the growth of the organism and a base for determining carbohydrate fermentation patterns of non-fastidious micro-organisms. It is also used for the detection of indole production by the bacteria after the addition of Kovacs or Ehlrich reagent.

Principle of Peptone Water

Water is the source of hydrogen and oxygen. Sodium chloride is the source of electrolytes. Peptone is a complex mixture of partially digested proteins. It contains proteoses, amino acids, polypeptides, phosphates, minerals (K, Mg), and accessory growth factors like nicotinic acid and riboflavin. Peptone used in this medium is rich in tryptophan content. The presence of indole can be detected using either Kovacs or Ehlrich reagent. Peptone Water is also applied as a base for carbohydrate fermentation studies with the addition of sugar and indicators such as bromothymol blue, bromocresol purple, or phenol red. Gas formation is achieved using Durham’s tube.

Composition of Peptone Water

Ingredients Gms / Litre

Peptone: 10.0
Sodium chloride: 5.0
Final pH (at 25°C) 7.2±0.2

Preparation of Peptone Water

Suspend 15.0 grams of the dehydrated medium in 1000 ml purified/distilled water.
Heat to boiling to dissolve the medium completely.
Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes i.e. validated cycle.

Storage and Shelf life of Peptone Water

Store at 2-8ºC and away from direct light.
Media should not be used if there are any signs of deterioration ( discoloration), contamination (turbidity).
The product is light and temperature-sensitive; protects from light, excessive heat, moisture, and freezing

Test Requirements for Peptone Water

Test specimen
Peptone water
Inoculating loop/ dropper
Bunsen burner
Incubator
Control strains ( Staphylococcus aureus ATCC V25923, Escherichia coli ATCC 25922, and uninoculated medium)
Waste bin

Procedure of Peptone Water

Allow the medium to warm at 37°C or to room temperature before inoculating.
Inoculate the specimen/ test organism.
Incubate it aerobically at 35-37°C for 24 hours ( or depending on the nature of suspected organisms).
Examine for turbidity.

Result Interpretation

Growth: Turbidity

No growth: Lacking turbidity

Control strain

Staphylococcus aureus ATCC V25923: Luxuriant growth

Escherichia coli ATCC 25922: Luxuriant growth with a red ring at the interface of the medium on the addition of Kovac’s reagent

Uninoculated tube: No growth

Visible growth of various organisms in Peptone Water

Turbidity – Aerobic gram-negative bacilli
Pellicle formation – Bacillus species, Pseudomonas species, Yeast cells
Clotting – Staphylococcus aureus
Visible colonies ( puffballs) -Staphylococci

Uses of Peptone Water

Peptone water is the simplest medium and it is used as a growth medium as a base for carbohydrate fermentation media.
It is a liquid medium and also suitable for the detection of indole.
It is a minimal growth medium that is used for determining carbohydrate fermentation patterns of non-fastidious organisms (E. coli. Klebsiella, Proteus, etc).
It is useful as a diluent or for making suspensions of non-fastidious organisms for antimicrobial sensitivity testing, motility test, microbial enumeration procedures, and so on.
Converting peptone water in the alkaline zone ( pH 8.4) is suitable for the cultivation and enrichment of Vibrio cholerae (etiological agent of cholera).
This medium is also useful for the detection of gas formation (using Durham’s tube)

Limitations of Peptone Water

This is the simplest medium due to its low nutrient content and this is not useful for the growth or maintenance of the fastidious organism.
pH determination is mandatory since some sugar solutions may affect the pH of the Peptone Water and hence checking must be done.
Subcultures on a solid medium may be necessary to ensure the purity of the inoculant. Mixed or contaminated growth will give false reactions.
Some strains may show poor growth because of nutritional variations.
The organism cannot be confirmed by this medium and thus biochemical, immunological, molecular, or mass spectrometry testing be performed on colonies from pure culture for complete identification.

Keynotes

Peptone water is a simple medium but even useful for various purposes like motility testing, antimicrobial sensitivity testing ( as a diluent/suspension), indole formation, oxidation-fermentation (base) test, enrichment medium (pH sift) for Vibrio species.
It is also the base of various culture media.