Moeller Decarboxylase Broth w/ Ornithine HCl

Intended Use

Recommended for differentiation of bacteria on the basis of their ability to decarboxylate L-Orinithine hydrochloride.

Composition**

Final pH ( at 25°C): 6.0±0.2

**Formula adjusted, standardized to suit performance parameters

# Equivalent to Beef extract

Directions

Suspend 20.52 grams in 1000 ml purified/distilled water. Heat if necessary to dissolve the medium completely. Dispense in 5 ml amounts in screw capped tubes and sterilize by autoclaving at 15lbs pressure (121°C) for 10 minutes. Cool the tubes in an upright position. Inoculate the tubes and overlay with 2-3 ml of sterile mineral oil.

Principle And Interpretation

Moeller Decarboxylase Broth with Ornithine hydrochloride is used for differentiating gram-negative enteric bacilli on the basis of their ability to decarboxylate L-Ornithine hydrochloride. Decarboxylase Broth was introduced by Moeller for detecting the production of lysine and ornithine decarboxylase and arginine dihydrolase (1). Prior to Moellers work, bacterial amino acid decarboxylases were studied by Gale (2) and Gale and Epps (3). Production of ornithine decarboxylase is helpful criterion in differentiating Klebsiella and Enterobacter species. Klebsiella are nonmotile and do not produce ornithine decarboxylase while Enterobacter are motile and produce ornithine decarboxylase except Enterobacter agglomerans (4). Decarboxylase media are also recommended by standard methods for identification of bacteria (5,6,7,8). This medium contains HM peptone B and peptone which provide nitrogenous nutrients for the growth of bacteria. Dextrose is the fermentable carbohydrate and pyridoxal is the co-factor for the decarboxylase enzyme. Bromo cresol purple and cresol red are the pH indicators in this medium. When the medium is inoculated with the dextrose fermenting bacteria, the pH is lowered due to acid production which changes the colour of the indicator from purple to yellow. Acid produced stimulates decarboxylase enzyme. Putrescine is produced due to ornithine decarboxylation. Formation of amine putrescine increases the pH of the medium, changing the colour of the indicator from yellow to purple. If the organisms do not produce the appropriate enzyme, the medium remains acidic, yellow in colour. Each isolate to be tested should also be inoculated into the basal medium tube lacking the amino acid. After incubation, a decarboxylase test may show two layers of different colours, yellow and purple. Shake the tube gently before interpreting the results (4).

Type of specimen

Isolated Microorganism from clinical and non-clinical samples

Specimen Collection and Handling

For clinical samples follow appropriate techniques for handling specimens as per established guidelines (5,6).

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 specimens. Safety guidelines may be referred in individual safety data sheets.

Limitations

1. Inoculated tubes must be protected from air with a layer of sterile mineral oil. Exposure to air may cause alkalinization at the surface of the medium which makes the test invalid.

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 greenish yellow homogeneous free flowing powder

Colour and Clarity of prepared medium: Purple coloured clear solution without any precipitate

Reaction: Reaction of 2.05% w/v aqueous solution at 25°C. pH : 6.0±0.2

pH: 5.80-6.20

Cultural Response: Cultural characteristics observed after an incubation at 35-37°C for upto 4 days (Inoculated tubes are overlaid with sterile mineral oil).

Key : *Corresponding WDCM numbers.
$ Formerly known as Proteus vulgaris
^ Formerly known as Pseudomonas aeruginosa

Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 15-30°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 (5,6).

Reference

1. Lipps WC, Braun-Howland EB, Baxter TE,eds. Standard methods for the Examination of Water and Wastewater, 24th ed. Washington DC:APHA Press; 2023.
2. FDA Bacteriological Analytical Manual, 8th Ed., AOAC International, Gaithersburg, Md.
3. Gale G. F., 1940, Biochem. J., 34:392.
4. Gale and Epps, 1943, Nature, 152:327.
5. Isenberg (Ed.), 1992, Clinical Microbiology Procedures Handbook, Vol. I, ASM, Washington, D. C.
6. 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.
7. MacFaddin J., 2000, Biochemical Tests for Identification of Medical Bacteria, 3rd ed., Williams and Wilkins, Baltimore.
8. Moeller V., 1955, Acta Pathol. Microbiol. Scand. 36:158.
9. Salfinger Y., and Tortorello M.L. Fifth (Ed.), 2015, Compendium of Methods for the Microbiological Examination of Foods, 5th Ed., American Public Health Association, Washington, D.C.