EE Broth, Modified

Intended Use

Recommended for selective enrichment of Enterobacteriaceae in the bacteriological examination of food and animal feed stuffs.

Composition**

Final pH (at 25°C): 7.2±0.2

# Equivalent to Ox bile, dried @ Equivalent to Pancreatic digest of gelatin **Formula adjusted, standardized to suit performance parameters

Directions

Suspend 42.93 grams (the equivalent weight of dehydrated medium per litre) in 1000 ml purified / distilled water. Distribute in 120 ml amounts in 250 ml flasks or 9 ml in tubes. Stopper with cotton plugs or loose fitting caps. Heat in free flowing steam or boiling water for 30 minutes. Avoid overheating of the medium. Cool to 45-50°C rapidly in cold running water. DO NOT AUTOCLAVE.

Principle And Interpretation

EE Broth, Modified is recommended as an enrichment medium for Enterobacteriaceae in bacteriological examination of foods and animal feed stuffs. Gelatin peptone, glucose allow the growth of most of the members of Enterobacteriaceae. Incorporation of brilliant green and Bile, dried makes the medium, inhibitory to non-Enterobacteriaceae. Lactose non fermenters, anaerogenic lactose fermenters or late lactose fermenters may be missed by the standard ‘Coli-aerogenes’ test. To overcome this problem, lactose is replaced by glucose in the media.

The cells damaged due to drying or exposure to low pH are resuscitated in well-aerated Tryptone Soya Broth (M011) for 2 hours at 25°C prior to enrichment. The resuscitation procedure is recommended for dried foods (3), animal feeds (4) and semi-preserved foods (5). This medium should be used in conjunction with Violet Red Bile Glucose Agar without Lactose (M581). Subcultures must be made onto lactose differential media as MacConkey Agar w/ 0.15% Bile salts, CV and NaCl (M081) and Deoxycholate Citrate Agar (M065).

Type of specimen

Food samples and animal feed stuff

Specimen Collection and Handling

For food samples, follow appropriate techniques for sample collection and processing as per guidelines (6). 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.

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 coloured homogeneous free flowing powder

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

Reaction: Reaction of 4.3% w/v aqueous solution at 25°C. pH : 7.2±0.2

pH: 7.00-7.40

Cultural Response: Cultural characteristics observed after an incubation at 35-37°C for 18-24 hours.

Key : (*) Corresponding WDCM numbers. (#) Formerly known as Enterobacter aerogenes

Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 15 - 25°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 (1,2).

Reference

1. Isenberg, H.D. Clinical Microbiology Procedures Handbook 2nd Edition.
2. 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.
3. Mossel D.A.A. and Ratto M.A., 1970, Appl. Microbiol., 20:273.
4. Mossel D.A.A. and Shennan J.L. and Clare V., 1973, J. Sci. Fd. Agric., 24 : 499.
5. Mossel D.A.A., Ratto M.A., 1973, J. Fd. Technol., 8 : 97.
6. 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.