MUG Brilliant Green Bile Broth

Intended Use

Recommended for detection of Escherichia coli in water and food samples by a fluorogenic assay method.

Composition

Final pH (at 25°C): 7.2±0.2

Formula adjusted, standardized to suit performance parameters

Equivalent to Oxgall

Directions

Suspend 40.1 grams in 1000 ml purified/distilled water. Heat if necessary to ensure completely solution. Dispense 10 ml amounts in test tubes containing inverted Durham's tubes. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. For testing larger quantities of sample prepare concentrated medium to accommodate volume of the test sample.

Principle And Interpretation

Brilliant Green Bile Broth is one of the most widely used medium for the detection of coliform bacteria in water, wastewater, foods, and milk and dairy products. This medium is formulated as per APHA (1, 6,7) for the presumptive identification and confirmation of coliform bacteria (4, 5).

Gelatin peptone serves as a source of essential nutrients. Lactose is the fermentable carbohydrate. Bile inhibits gram-positive bacteria whereas the gram-negative bacteria are inhibited by brilliant green. Production of gas from lactose fermentation is detected by incorporating inverted Durham's tube, which indicates the positive evidence of faecal coliform since non faecal coliforms growing in this medium do not produce gas. Gram-positive spore formers may produce gas if the bile or brilliant green inhibition is weakened by reaction with food material. The fluorogenic compound, MUG (4-Methylumbelliferyl-ẞ-D-glucuronide) in the medium permits the rapid detection of E.coli which produces a blue fluorescence when hydrolyzed by the enzyme ẞ-glucuronidase and is observed using a long-wave UV light source. During examination of water samples, growth from presumptive positive tubes showing gas in Lactose Broth (M026) or Lauryl Tryptose Broth (M080) is inoculated in Brilliant Green Bile Broth 2% (M121). Gas formation within 48 ± 2 hours confirms the presumptive test (1).

Type of specimen

Food and dairy samples; Water samples

Specimen Collection and Handling:

For food and dairy samples, follow appropriate techniques for sample collection and processing as per guidelines (6,7). For water samples, follow appropriate techniques for sample collection, processing as per guidelines and local standards.(1) 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. Biochemical and serological tests must be carried out for further 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

Light yellow to light green homogeneous free flowing powder

Colour and Clarity of prepared medium

Emerald green coloured clear solution

Reaction

Reaction of 4.01% 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 aerogens

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 (2,3).

Reference

1. Baird R.B., Eaton A.D., and Rice E.W., (Eds.), 2015, Standard Methods for the Examination of Water and Wastewater, 23rd ed., APHA, Washington, D.C.
2. Isenberg, H.D. 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. McCrady and Langerin, 1932, J. Dairy Science, 15:321.
5. McCrady, 1937, Am. J. Publ. Health, 27:1243.
6. 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.
7. Wehr H. M. and Frank J. H., 2004, Standard Methods for the Microbiological Examination of Dairy Products, 17th Ed.,APHA Inc., Washington, D.C.