Chapman Stone Agar
Intended Use:
Recommended for selective isolation of Staphylococci causing food poisoning
Composition
Ingredients Gms / Litre
Tryptone 10.000
Yeast extract 2.500
Gelatin 30.000
D-Mannitol 10.000
Sodium chloride 55.000
Ammonium sulphate 75.000
Dipotassium hydrogen phosphate 5.000
Agar 15.000
Final pH ( at 25°C) 7.0±0.2
Directions
Suspend 20.25 grams in 100 ml purified / distilled water. Heat to boiling to dissolve the medium completely. Sterilize by
autoclaving at 15 lbs pressure (121°C) for 10 minutes. Cool to 45-50°C. Mix well and pour into sterile Petri plates.
Principle And Interpretation
Staphylococcus aureus is one of the pathogens most frequently isolated from clinical specimens. In fact, S.aureus is
currently the most common cause of nosocomial infections (1). Treatment of infection caused by S.aureus has become
more problematic since the development of multiple drug resistant strains. To identify S. aureus from contaminated samples
more easily and reliably, selective media have been developed.
Chapman Stone Agar is a selective media used for the isolation of food poisoning staphylococci. Foods commonly contaminated
with S. aureus included synthetic creams, custards and high-salted food.
Chapman Stone Agar is prepared according to the modification of Staphylococcus Medium 110 described by Chapman (1). It
is similar to Staphylococcus Medium 110, previously described by Chapman (2), except that the sodium chloride
concentration is reduced to 5.5% and additionally ammonium sulfate is included in the formulation. The main modification
consists the inclusion of ammonium sulfate in the medium that allows the direct observation of gelatin hydrolysis, instead of
adding reagents to the plate medium. Chapman Stone Medium is especially recommended for suspected food poisoning
studies involving Staphylococcus (7). It is selective, due to the relatively high salt content, and is differential due to
pigmentation, mannitol fermentation and the presence or absence of gelatin liquefaction.
Tryptone, yeast extract provide nitrogen, carbon, sulphur, vitamin B and trace elements. Sodium chloride acts as a selective
agent, which inhibits most of the bacterial species. Mannitol is the fermentable carbohydrate and its fermentation can be
detected by adding a few drops of bromocresol purple resulting in production of yellow colour. Gelatin hydrolysis is
observed as clear zones around colonies. Due to the presence of ammonium sulphate in the medium itself there is no need to
flood the plate with ammonium sulphate solution for detection of gelatin liquefaction by the isolates, which is known as
Stones method (7). Dipotassium phosphate provides buffering capability. Material under test is inoculated on the surface and
incubated at 30°C for 48 hours to produce separated colonies. After incubation, cream to golden yellow colonies surrounded
by clear zones are presumptively identified as S.aureus. White or non-pigmented colonies, with or without a clear zone, are
presumptively identified as S. epidermidis. Coagulase activity should be performed to confirm the findings.
Enterococci and/or Group D streptococci may exhibit growth on the medium and show slight mannitol fermentation. The
colonies, however, are tiny and can easily be differentiated from staphylococci by gram stain and the catalase test (5).
Type of specimen
Food samples
Specimen Collection and Handling:
For food samples, follow appropriate techniques for sample collection and processing as per guidelines (6).
After use, contaminated materials must be sterilized by autoclaving before discarding.
Warning and Precautions :
Read the label before opening the container. Wear protective gloves/protective clothing/eye protection/ face protection.
Follow good microbiological lab practices while handling specimens and culture. Standard precautions as per
established guidelines should be followed while handling specimens. Safety guidelines may be referred in individual
safety data sheets.
Limitations :
1. Enterococci and/or Group D streptococci may exhibit growth on the medium and show slight mannitol fermentation. The
colonies, however, are tiny and can easily be differentiated from staphylococci by Gram stain and the catalase test.
2. Further biochemical and serological tests must be carried out for further identification.
Performance and Evaluation
Performance of the medium is expected when used as per the direction on the label within the expiry period when stored at
recommended temperature.
Quality Control
Appearance
Cream to yellow coarse free flowing powder
Gelling
Firm, comparable with 1.5% Agar gel and 3.0% Gelatin gel
Colour and Clarity of prepared medium
Light amber coloured, opalescent gel forms in Petri plates
Reaction
Reaction of 20.25% w/v aqueous solution at 25°C. pH : 7.0±0.2
pH
6.80-7.20
Cultural Response
Cultural characteristics observed after an incubation at 25 - 30°C for 18 - 48 hours.
Organism Inoculum
(CFU)
Growth Recovery Mannitol
fermentation
Gelatinase
production
Escherichia coli ATCC
25922 (00013*)
=104
inhibited 0%
Staphylococcus aureus
ATCC 25923 (00034*)
luxuriant =50% positive
reaction,
production of
yellow colour
on addition of
Bromo cresol
purple
positive
reaction,
clearing or halo
Staphylococcus epidermidis
ATCC 12228 (00036*)
50-100 luxuriant =50% negative
reaction, no
production of
yellow colour
on addition of
Bromo cresol
purple
positive
reaction,
clearing or halo
Storage and Shelf Life
Store between 10-30°C in a tightly closed container and the prepared medium at 20-30°C. Use before expiry date on
the label. On opening, product should be properly stored dry, after tightly capping the bottle in order to prevent lump
formation due to the hygroscopic nature of the product. Improper storage of the product may lead to lump formation.
Store in dry ventilated area protected from extremes of temperature and sources of ignition. Seal the container tightly
after use. Use before expiry date on the label.
Product performance is best if used within stated expiry period.
User must ensure safe disposal by autoclaving and/or incineration of used or unusable preparations of this product. Follow
established laboratory procedures in disposing of infectious materials and material that comes into contact with sample must
be decontaminated and disposed of in accordance with current laboratory techniques (3,4).
