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Lecithinase Activity#CC293D
Intended Use
Used for detecting lecithinase activity of anaerobic microorganisms.
Composition**
| Ingredients | g/L |
|---|---|
| Tryptone | 5.000 |
| Yeast extract | 5.000 |
| Sodium chloride | 2.500 |
| Sodium sulphite | 0.100 |
| L-Cystine | 0.400 |
| L-Tryptophan | 0.200 |
| Dextrose (Glucose) | 2.000 |
| Disodium hydrogen phosphate | 5.000 |
| Magnesium sulphate | 0.010 |
| Hemin | 0.010 |
| Vitamin K1 | 0.010 |
| Agar | 20.000 |
| Final pH (at 25°C) | 7.4±0.2 |
**Formula adjusted, standardized to suit performance parameters
Directions
Suspend 40.23 grams in 900 ml purified/distilled water. Heat to boiling to dissolve the medium completely. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. Cool to 60°C and aseptically add 100 ml sterile Egg Yolk Emulsion (FD045). Mix well and pour into sterile petri plates.
Principle And Interpretation
Organisms that grow in the absence of oxygen are termed as anaerobes. Depending upon their ability to tolerate oxygen, they are classified as either facultative or obligate anaerobes. The anaerobic gram-negative bacteria are part of the normal flora of the upper respiratory tract, mouth, intestinal tract and urinogenital tract of human and animals. The bile-resistant Bacteroides fragilis group is the most commonly recovered anaerobe in clinical specimens and is more resistant to antimicrobial agents than any other anaerobe. Fusobacterium necrophorum is a very virulent anaerobe that may cause severe infections, usually in children or young adults (1).
L. D. Medium or Lombard-Dowell Medium was developed by Dowell and Lombard (2) for the cultivation and identification of fastidious anaerobic bacteria. L. D. Egg Yolk Agar Base, along with egg yolk emulsion is used for the detection of lipase; lecithinase and proteolytic activity of both spore forming and non-spore-forming obligate anaerobes (3). L. D. Agar is essentially a casein digest agar enriched with hemin, vitamin K1, L-cystine and yeast extract (4). This medium contain various nutritious substances, which can promote the growth of fastidious anaerobic bacteria. Tryptone and yeast extract provide the necessary nitrogenous nutrients while hemin and vitamin K1 supply additional growth factors. L-cystine and L-tryptophan serve as the amino acid sources. Sodium sulphite is an antioxidant. Sodium chloride maintains osmotic balance of the medium.
L. D. Egg Yolk Agar with Egg Yolk Supplement contains egg yolk, which is the source of lecithin in the medium. Magnesium sulphate helps in sporulation. Disodium phosphate buffers the medium.
Type of specimen
Clinical samples - wound 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 only. 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
- Further biochemical and serological test must be carried out for complete identification.
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 2.0% agar gel.
Colour and Clarity of prepared medium
Basal medium : Medium amber coloured clear to slightly opalescent gel. After addition of sterile egg yolk emulsion : Yellow coloured opaque gel forms in Petri plates
Reaction
Reaction of 4.02% w/v aqueous solution at 25°C. pH: 7.4±0.2
pH
7.20-7.60
Cultural Response
Cultural characteristics observed under anaerobic condition,with added Egg Yolk Emulsion (FD045), after an incubation at 35-37°C for 24-48 hours.
| Organism | Inoculum (CFU) | Growth | Lecithinase/ Halos | Lipase | Proteolysis |
|---|---|---|---|---|---|
| Clostridium perfringens ATCC 12924 | 50 - 100 | luxuriant | positive reaction, opaque zone of insoluble precipitate | negative reaction | negative reaction |
| Clostridium sporogenes ATCC 11437 | 50 - 100 | luxuriant | negative reaction | positive reaction, irridescent sheen on the colony surface and medium | positive reaction, clear zone surrounding colonies |
| Fusobacterium necrophorum ATCC 25286 | 50 - 100 | luxuriant | negative reaction | positive reaction, irridescent sheen on the colony surface and medium | negative reaction |
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 (5,6).
| Product Name | L.D. Egg Yolk Agar Base |
|---|---|
| SKU | M744 |
| Product Type | Regular |
| Physical Form | Powder |
| Origin | Animal |
| Packaging type | HDPE |
| References | 1. Dowell V. and Lombard G., June 1977, U.S., DHEW, Center for Disease Control (CDC), Atlanta. Ga. 2.MacFaddin J. F., 1985, Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria, Vol. I, Williamsand Wilkins, Baltimore 3.Finegold S. M., Baron E. J., Bailey and Scotts Diagnostic Microbiology, 8th Ed., 1990, The C.V. Mosby Company 4.Murray P. R., Baron J. H., Pfaller M. A., Jorgensen J. H. and Yolken R. H., (Ed.), 2003, Manual of Clinical Microbiology,8th Ed., American Society for Microbiology, Washington, D.C. |
| Customized Product Available | No |
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