Plot No. C40, Road No. 21Y, MIDC, Wagle Industrial Estate,
Thane(W)-400604, MH, India.
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Intended use
Recommended for selective detection and enumeration of Campylobacter species from food chain. The composition and performance criteria of this medium are as per the specifications laid down in ISO 10272-1:2017 & 10272-2:2017.
Composition**
ISO 10272-1 and ISO 10272-2 Specification - mCCD agar
| Ingredients | Gms / Litre |
|---|---|
| Meat extract | 10.000 |
| Enzymatic digest of animal tissues | 10.000 |
| Enzymatic digest of casein | 3.000 |
| Sodium chloride | 5.000 |
| Sodium deoxycholate | 1.000 |
| Iron (II) sulfate, hydrate | 0.250 |
| Sodium pyruvate | 0.250 |
| Activated charcoal | 4.000 |
| Agar | 8.0-18.0 |
mCCD agar M8871
| Ingredients | Gms / Litre |
|---|---|
| HM Extract # | 10.000 |
| Peptone ## | 10.000 |
| Tryptone ### | 3.000 |
| Sodium chloride | 5.000 |
| Sodium deoxycholate | 1.000 |
| Iron (II) sulfate, hydrate | 0.250 |
| Sodium pyruvate | 0.250 |
| Activated charcoal | 4.000 |
| Agar | 12.000 |
Supplements to be added after autoclaving
| CCDA Selective Supplement-FD135 (2 vials) | ||
|---|---|---|
| Cefoperazone | 0.032 g | 16 mg |
| Amphotericin B | 0.01 g | 5mg |
Final pH (at 25°C): 7.4±0.2
**Formula adjusted, standardized to suit performance parameters
# Equivalent to Meat extract
## Equivalent to Enzymatic digest of animal tissues
### Equivalent to Enzymatic digest of casein
Directions
Either streak, inoculate or surface spread the test inoculum (50-100 CFU) aseptically on the plate.
Principle And Interpretation
Campylobacters are carried in the intestinal tract of animal and therefore contaminate foods of animal origin (1). Campylobacter causes intestinal upset or abortion in animals. It is also one of the most important causes of human gastroenteritis, particularly in children. Initially blood was used in the isolation of Campylobacter. But, later it was reported by Bolton et al (2) that charcoal can be effectively used in place of blood. This rules out the variability obtained due to the use of blood.
Modified Charcoal Cefoperazone Deoxycholate Agar Base is formulated as per APHA (1) and is also recommended by the ISO Committee (3,4) for detection and enumeration of Campylobacter spp. from food and environmental samples in food production area. Cephalothin in the original formulation was replaced by Cefoperazone as the selective agent since the latter gave better selectivity (5). Campylobacter species are highly resistant to cefoperazone, an antibiotic which effectively suppresses growth of Pseudomonas and Enterobacteriaceae (6,7,8). Addition of cefoperazone increases the selectivity of the medium. Due to this addition, the medium is also known as Campylobacter Charcoal Differential Agar (CCDA).
Charcoal, sodium pyruvate and iron sulfate reduces the aero tolerance of medium by quenching photo chemically generated toxic oxygen derivatives (8). Peptone, Tryptone and HM extract serve as sources of carbon, nitrogen, long chain amino acids and essential nutrients. Sodium chloride maintains osmotic balance. Additional Amphotericin B suppresses the growth of yeast and mold contaminants.
Type of specimen
Food samples
Specimen Collection and Handling:
Processesing: (3,4)
- Test portion and initial suspension:
Selective Enrichment A: To prepare the initial suspension, combine a quantity of 10g or 10ml of the test portion with 90ml of the enrichment medium Bolton Broth (M1592) from cooked and frozen food. Incubate the initial suspension in a microaerobic atmosphere at 37°C for 4 to 6 hours and then at 41.5°C for 44 hours ± 4 hours.
Selective Enrichment B: To prepare the initial suspension, combine a quantity of 10g or 10ml of the test portion with 90ml of the enrichment medium Preston Broth (M899I) from raw meats or raw milk. Incubate the initial suspension in a microaerobic atmosphere at 41.5°C for 24 hours ± 2 hours.
- Plating out : Using the culture obtained in the enrichment medium, inoculate with a sterile 10 µl loop on the surface of mCCD Agar. Incubate the plates at 41.5°C in a microaerobic atmosphere for 44 hours ± 4 hours.
- Selective Enrichment C: Directly swab or streak on mCCD agar from caecal or faecal samples by using a loop or a sterile swab and second medium can be optional. Incubate the plates at 41.5°C in a microaerobic atmosphere for 44 hours ± 4 hours.
- Confirmation: Biochemical and serological tests are performed for confirmation.
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 to individual safety data sheets.
Limitations
- Individual organisms differ in their growth requirement and may show variable growth patterns on the medium
- Each lot of the medium has been tested for the organisms specified on the COA. It is recommended to users to validate themedium for any specific microorganism other than mentioned in the COA based on the user's unique requirement.
- It is recommended to store the plates at 24-30°C to avoid minimum condensation.
- Further biochemical tests must be carried out for confirmation.
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
Sterile Modified Charcoal Cefoperazone Deoxycholate Agar in 90mm disposable plates with smooth surface and absence of black particles/ cracks/bubbles.
Colour of medium
Black coloured medium
Quantity of medium
25ml of medium in 90mm plate
pH
7.20-7.60
Sterility Check
Passes release criteria
Cultural Response
Cultural characteristics observed after an incubation at 41.5°C ± 1°C for 40 hours under microaerobic atmosphere.
Recovery is considered as 100% on Blood Agar.
| Organism | Inoculum (CFU) | Growth | Recovery | Colour of colony |
|---|---|---|---|---|
| Productivity | ||||
| Campylobacter coli ATCC 33559 (00004*) | 50-100 | good-luxuriant | >=50% | greyish, flat colonies, may have metallic sheen |
| Campylobacter jejuni ATCC 29428 (00005*) | 50-100 | good-luxuriant | >=50% | sheen greyish, flat colonies, may have metallic sheen |
| Selectivity | ||||
| Escherichia coli ATCC 25922 (00013)* | 50-100 | total or partial inhibition | <=10% | |
| Escherichia coli ATCC 8739 (00012*) | 50-100 | total or partial inhibition | <=10% | |
| Staphylococcus aureus subsp. aureus ATCC 25923 (00034*) | >=104 | inhibited | ||
Key: * - Corresponding WDCM numbers
Storage and Shelf Life
On receipt store between 2-8°C Use before expiry date on the label. Product performance is best if used within stated expiry period.
Disposal
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 (9,10).
| Product Name | Modified Charcoal Cefoperazone Deoxycholate Agar Plate |
|---|---|
| SKU | MP887I |
| Customized Product Available | No |
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