Cholera Medium Base

Intended Use

Recommended for selective isolation of Vibrio species from specimens heavily contaminated with Enterobacteriaceae.

Composition

Final pH (at 25°C): 8.5±0.2

**Formula adjusted, standardized to suit performance parameters

Directions

Suspend 65.1 grams in 1000 ml purified/distilled water. Heat to boiling to dissolve the medium completely. DO NOT AUTOCLAVE. Cool to 70°C and add 2 ml of sterile 1% Potassium Tellurite Solution (FD052) and 5 ml of sterile defibrinated blood. Maintain at 70°C for a few minutes. Cool to 45-50°C. Mix well and pour into sterile Petri plates.

Principle And Interpretation

Vibrio cholerae is the etiological agent of cholera in humans in which the disease is caused not by tissue invasion of microorganisms but through the production of toxins that interrupt normal intra-intestinal exchanges of water and electrolytes. Vibrios grow readily on most isolation media. Adding sodium chloride to the medium enhances growth of all species. Cholera Medium Base is a selective medium used for the isolation of Vibrio species from specimens contaminated with enteric bacteria. It is based on the formulation described by Felsenfeld and Watanabe (1) for the isolation of V. cholerae and similar Vibrios from specimens contaminated with Enterobacteriaceae.

Peptone and HM peptone B provide nitrogenous nutrients whereas sucrose serves as the fermentable carbohydrate source for the metabolism of Vibrios. Sodium lauryl sulphate inhibits many contaminating organisms. Potassium tellurite also inhibits many gram-positive and gram-negative bacteria except Vibrios. Sodium chloride maintains osmotic equilibrium.

Type of specimen

Clinical samples- faeces; Food samples.

Specimen Collection and Handling

For clinical samples follow appropriate techniques for handling specimens as per established guidelines (2,3).

For food samples, follow appropriate techniques for sample collection and processing as per guidelines (4).

After use, contaminated materials must be sterilized by autoclaving before discarding.

Warning and Precautions

In Vitro diagnostic Use. For professional use only. 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 clinical specimens. Safety guidelines may be referred in individual safety data sheets.

Limitations

1. Slight colour variation may be observed depending upon strains.
2. 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: Cream to yellow homogeneous free flowing powder

Gelling: Firm, comparable with 1.0% Agar gel.

Colour and Clarity of prepared medium: Basal medium: Yellow coloured clear to slightly opalescent gel. After Addition of blood & Tellurite and on heating : Brownish red coloured opaque gel forms in Petri plates.

Reaction: Reaction of 6.5% w/v aqueous solution at 25°C. pH: 8.5±0.2

pH: 8.30-8.70

Cultural Response: Cultural characteristics observed with added 1% Potassium Tellurite Solution(FD052) and sterile defibrinated blood, after an incubation at 35-37°C for 18-24 hours.

Key: *Corresponding WDCM numbers. **Formerly known as Bacillus subtilis subsp. spizizenii

Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 2 - 8°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. 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 clinical sample must be decontaminated and disposed of in accordance with current laboratory techniques (2,3).

Reference

1. Felsenfeld O. and Watanabe Y., 1958, U.S. Armed Forces Med. J., 9 (7) : 975.
2. Isenberg, H. Clinical Microbiology Procedures Handbook. 2nd Edition.
3. Jorgensen, J.H., Pfaller, M.A., Carroll, K.C., Funke, G., Landry, M.L., Richter, S.S and Warnock., D.W. (2015) Manual of Clinical Microbiology, 11th Edition. Vol. 1.
4. Salfinger Y., and Tortorello M.L., 2015, Compendium of Methods for the Microbiological Examination of, Foods, 5th Ed., American Public Health Association, Washington, D.C.