L-Ornithine Decarboxylase Broth

L-Ornithine Decarboxylase Broth

M12231

Intended use

Recommended for detection of the ability of microorganisms to decarboxylate ornithine. The composition and performance of this media are as per specifications laid down in in ISO/TS 22964: 2017.

Composition**

ISO/TS 22964: 2017 Specification-L-Ornithine Decarboxylase Broth

M1223I-L-Ornithine Decarboxylase Broth

**Formula adjusted, standardized to suit performance parameters

Directions

Suspend 14.02 grams in 1000 ml purified / distilled water. Heat if necessary to dissolve the medium completely. Dispense in test tubes and sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. After inoculation overlay the tubes with 2-3 ml mineral oil.

Principle And Interpretation

Decarboxylation is the process in which bacteria that possess specific decarboxylase enzyme attack amino acids at their carboxyl end (-COOH) to yield an amine or a diamine and carbon dioxide (1). The amino acid L-ornithine is decarboxylate by the enzyme ornithine decarboxylase to yield the diamine putrescine and carbon dioxide (2,3).

Ornithine Decarboxylase Broth is based on the Taylors modification (4). It is recommended by the ISO Committee (5) for the detection of ornithine decarboxylation by Cronobacter sakazakii (5).

Yeast extract in the medium provides nitrogen and other nutrients necessary to support bacterial growth. The amino acid L-ornithine is added to detect the production of ornithine decarboxylase. Glucose is the fermentable carbohydrate, which during the initial stages of incubation, is fermented by the organisms with acid production, which results in colour change of the pH indicator (BCP) to yellow. The acidic condition also stimulates decarboxylase activity. If the organism produces the appropriate enzyme, i.e. decarboxylase, the amino acid (ornithine) in the medium is degraded, yielding a corresponding amine. Decarboxylation of ornithine yields putrescine. The production of this amine elevates the pH of the medium towards alkalinity, changing the color of the indicator from yellow to purple or violet. If the organism does not produce the appropriate enzyme, the medium remains acidic or yellow in colour.

Type of specimen

Food and dairy samples

Specimen Collection and Handling:

For food and dairy samples, follow appropriate techniques for sample collection and processing as per guidelines (6,7,8). 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. Overlaying the medium with mineral oil is essential to avoid erroneous results.

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 Light yellow to light green homogeneous free flowing powder

Colour and Clarity of prepared medium Dark purple coloured clear solution without any precipitae

Reaction Reaction of 1.4% aqueous solution at 25°C. pH : 6.8±0.2

pH 6.60-7.00

Cultural Response Cultural characteristics observed after an incubation at 35-37°C for 18-24 hours. Inoculated tubes are overlayed with mineral oil.

Key: *: Corresponding WDCM numbers

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 sample must be decontaminated and disposed of in accordance with current laboratory techniques (9,10).

Reference

1. Smith D.T., Coant N.F., Willett H.P., Zinssers Microbiology, 14th Ed., New York: Appleton-Century-Crofts, 1968:118-119
2. MacFaddin J.F., 2000, Biochemical Tests for Identification of Medical Bacteria, 3rd Ed., Lippincott, Williams and Wilkins, Baltimore.
3. Moeller V., 1955, Acta Pathol. Microbiol. Scand. 36 (2): 158-172
4. Taylor W.I., 1961, Appl. Microbiol., 9:487.
5. International Organization for Standardization. Microbiology of the food chain- Horizontal method for the detection of Cronobacter spp. Draft ISO/TS 22964, 2017 (E).
6. American Public Health Association, Standard Methods for the Examination of Dairy Products, 1978, 14th Ed., Washington D.C.
7. 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.
8. Wehr H. M. and Frank J. H., 2004, Standard Methods for the Microbiological Examination of Dairy Products, 17th Ed., APHA Inc., Washington, D.C.
9. Isenberg, H.D. Clinical Microbiology Procedures Handbook. 2nd Edition.
10. 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.