In 1984, Bolton et al. Bolton et al. (1984) suggested that charcoal could be used to replace blood in a medium for the isolation of Campylobacter species. Endtz et al. Later, it was confirmed that Campylobacter isolates at a higher rate when using charcoal selective media.
Principle of Charcoal Selective Medium
The medium is made up of beef extract, which provides nitrogen, vitamins, and other nutrients necessary for Campylobacter growth. Casein peptones and gelatin provide nutrients in form of amino acids or peptides. The charcoal, hematin, sodium pyruvate, and ferrous sulfate improve the aerotolerance of Campylobacter species; it has been suggested that these supplements act as quenching agents of photochemically-produced toxic oxygen derivatives. Charcoal acts as a detoxifier and decreases oxygen tension. Sodium chloride is an essential electrolyte that maintains osmotic equilibrium and thus preserves the integrity of cells. Sodium desoxycholate, a selective medium that inhibits certain bacteria, is available. Cefoperazone, a cephalosporin antibiotic, inhibits some bacteria. Vancomycin, a glycopeptide antibiotic, inhibits many types of gram-positive bacteria. Cycloheximide, an antifungal agent, inhibits yeast growth and mold.
Composition of Charcoal Selective Medium
| Ingredients | Gm/Liter |
| Beef extract | 10.0g |
| Gelatin Peptone | 10.0g |
| Sodium Chloride | 5.0g |
| Casein Peptone | 3.0g |
| Charcoal | 4.0g |
| Sodium desoxycholate | 1.0g |
| Ferrous sulfate | 0.25g |
| Sodium Pyruvate | 0.25g |
| Cycloheximide | 100.0mg |
| Cefoperazone | 32.0mg |
| Hematin | 32.0mg |
| Vancomycin | 20.0mg |
| Agar | 12.0g |
pH 7.4± 0.2 (at 25°C)
Preparation of Charcoal Selective Medium
- You can add components to deionized/distilled water to increase volume to 1000ml
- Combine thoroughly. Mix well. Heat gently and boil, stirring frequently.
- Autoclave for 15 minutes at 15 psi pressure at 120°C.
- Keep it at 45-50 degrees Celsius
- Pour into sterile Petri dishes or distribute into tubes.
- To keep charcoal suspended, shake the flask.
Result Interpretation on Charcoal Selective Medium
- Two types of colonies are produced by Campylobacter Jejuni
- One is small, raised and grayish-brown. The other is smooth and glistening, with a translucent entire edge.
- Another colony type is flat and mucoid, translucent, grayish, and has an irregular edge.
- Fresh medium can be used to observe flat, irregular or spreading colonies.
- Some strains form thin films on the agar, while others form colonies that trail along the streaking.
- Colonies are formed on less fresh media and are approximately 1-2mm in diameter. They are round, convex, and glistening. Non-hemolytic colonies can be pinkish or yellowish in color, and they are not hemolytic.
Uses of Charcoal Selective Medium
- It is used to presumptive identify and selectively isolate Campylobacter species using human feces specimens and food.
Limitations of Charcoal Selective Medium
- For complete identification, it is recommended that colonies grown from pure culture be subject to biochemical, immunological and molecular testing.
- The Campylobacter isolate rate may be increased by extending the incubation time to 72 hours
- Cephalosporins inhibit Campylobacter coli and Campylobacter fetus, as well as some Campylobacter jejuni strains.
- Because Campylobacter jejuni is a thermophilic organism, it should be incubated at 42°C. Higher temperatures promote Campylobacter jellyjuni’s growth by enhancing selectivity and inhibiting the microflora.
- A selective medium can inhibit certain strains of the desired species, or allow growth of a species it was not designed to prevent. This is especially true if large quantities of the species are present in the specimens. To obtain additional information and to ensure the recovery of pathogens, specimens that have been cultured in selective media should be cultured in nonselective media.
