Carbon vegetal

Charcoal Agar

Procedure
1. Streak a sample of the specimen onto the surface of the agar. Make several stabs into the medium along the streak. 2. Incubate, aerobically, at 30 ± 2°C for up to 48 hours. 3. Examine for growth and the presence or absence of clear zones around colonies. 4. To determine mannitol fermentation, add a few drops of bromcresol purple to areas on the medium from whichcolonies have been removed. Any change in color of the indicator, compared with that of the uninoculated medium, indicates fermentation of mannitol.

aureus. White or nonpigmented colonies, with or without a clear zone, are probably S. epidermidis.

Limitations of the Procedure
1. Confirm the presumptive identification of pathogenic staphylococci with additional tests, such as coagulaseactivity. 2. 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.3

C

Expected Results
Mannitol fermentation: Positive = change in color of the indicator to yellow. Gelatinase activity: Positive Stone reaction = formationof clear zones around the colonies. Any mannitol-positive, yellow or orange colonies surrounded by a clear zone are presumptively identified as Staphylococcus

References
1. Chapman. 1948. Food Res. 13:100. 2. Chapman. 1946. J. Bacteriol. 51:409. 3. MacFaddin. 1985. Media for isolation-cultivation-identification-maintenance of medical bacteria. Williams & Wilkins, Baltimore, Md.

AvailabilityDifco™ Chapman Stone Medium
Cat. No. 211805 Dehydrated – 500 g

Charcoal Agar
Intended Use
Charcoal Agar is used for cultivating fastidious organisms, especially Bordetella pertussis, for vaccine production and stock culture maintenance. The genus Bordetella consists primarily of four species: Bordetella pertussis, B. parapertussis, B. bronchiseptica and B. avium; additional species haverecently been described.2 All Bordetella are respiratory pathogens, residing on the mucous membranes of the respiratory tract. B. pertussis is the major cause of whooping cough or pertussis. B. parapertussis is associated with a milder form of the disease.3 B. bronchiseptica is an opportunistic human pathogen associated with both respiratory and non-respiratory infections, often occurring inpatients having close contact with animals.2 B. bronchiseptica
Uninoculated Plate Bordetella bronchiseptica ATCC™ 4617

Summary and Explanation
Charcoal Agar is prepared according to the method of Mishulow, Sharpe and Cohen.1 The authors found this medium to be an efficient substitute for Bordet-Gengou Agar in the production of B. pertussis vaccines.

User Quality Control
Identity SpecificationsDifco™ Charcoal Agar
Dehydrated Appearance: Solution: Gray, free-flowing, homogeneous. 6.25% solution, soluble in purified water upon boiling. Solution is black, opaque with a precipitate. Black, opaque. pH 7.3 ± 0.2

Prepared Appearance: Reaction of 6.25% Solution at 25°C:

Cultural Response
Difco™ Charcoal Agar
Prepare the medium per label directions. Inoculate and incubate at 35 ± 2°Cunder 5-10% CO2 for 18-72 hours.
ORGANISM ATCC™ INOCULUM CFU RECOVERY

Bordetella bronchiseptica Bordetella parapertussis Bordetella pertussis

4617 15237 8467

102-103 102-103 102-103

Good Good Good

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Section III C Charcoal Agar, cont.

has not been reported to cause pertussis. There have been no reports of recovery of B. avium from humans.2 Charcoal Agar supplemented withHorse Blood is used for the cultivation and isolation of Haemophilus influenzae.4

3. Autoclave at 121°C for 15 minutes. 4. Mix thoroughly during dispensing to uniformly distribute the charcoal. 5. Test samples of the finished product for performance using stable, typical control cultures.

Principles of the Procedure
Infusion from beef heart and peptone provide the nitrogen, carbon and amino...