Bile Broth Base

Intended Use

Recommended for cultivation of members of the Enterobacteriaceae and in culture of blood clots from patients with suspected enteric fever.

Composition**

Final pH (at 25°C): 7.6±0.2

**Formula adjusted, standardized to suit performance parameters

Directions

Suspend 30.0 grams in 1000 ml distilled/purified water. Heat if necessary to dissolve the medium completely. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. Cool to 45-50°C and add 1 ml of Streptokinase solution (100000 units/ml). Mix well and dispense into tubes or flasks as desired.

Principle And Interpretation

Enterobacteriaceae inhabit a wide variety of niches that include the human gastrointestinal tract and various environmental niches. When blood samples from a patient with suspected enteric fever is submitted for the widal test, it is useful as a routine to culture the clot after separation of serum (1). If it is known that the blood has been withdrawn with strict aseptic precautions, the clot may be placed in a wide tube half-filled with broth, or in a wide mouth screw-capped bottle containing 80 ml of broth. When there is any doubt regarding the presence of contaminating organisms, and this is always a possibility when blood specimens are sent to the laboratory from a distance, the clot should be transferred directly to a tube of sterile ox bile and disintegrated with aseptic precautions. After overnight incubation the bile culture is examined for enteric organism in the usual manner. A method of clot culture with Streptokinase has been recommended (2). Blood is allowed to clot in 5 ml quantities in sterile screw-capped universal containers. The separated serum is removed and 15 ml of 0.5% Bile Broth Base with Streptokinase 100 units/ml is added to each bottle. The streptokinase causes rapid clot lysis with release of bacteria trapped in the clot (2).

Peptone serves as a source of nitrogen, carbon, long chain amio acids and other essential amino acids. Sodium taurocholate inhibits majority of Gram-positive species. Sodium chloride maintains the isotonicity of the medium whereas addition of streptokinase solution causes rapid clot lysis with release of bacteria trapped in the clot (2).

Type of specimen

Clinical samples - Blood clot

Specimen Collection and Handling:

For clinical samples follow appropriate techniques for handling specimens as per established guidelines (3,4).

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 :

1. Further biochemical and serological tests must be carried out for complete identification.
2. Individual organisms differ in their growth requirement and may show variable growth patterns on the medium.
3. Each lot of the medium has been tested for the organisms specified on the COA. It is recommended to users to validate the medium for any specific microorganism other than mentioned in the COA based on the user's unique requirement.

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

Colour and Clarity of prepared medium Yellow coloured, clear solution without any haziness

Reaction Reaction of 3.0% w/v aqueous solution at 25°C. pH: 7.6±0.2

pH 7.40-7.80

Cultural Response Cultural characteristics observed after an incubation at 35-37°C for 18-48 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 2-8°C. Use before expiry date on the label. On opening, product should be properly stored dry, after tightly capping the bottle inorder 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 (3,4).

Reference

1. Colle, J.G., Duguid J.P., Fraser A.G. and Marmion, B.P. (Eds.) 1989 Mackie and McCartney Practical Medical Microbiology, Vol. 2, p:134 Longman Group, UK.
2. Watson, K.C. 1955, Isolation of Salmonella Typhi from the blood stream. J. of ,Lab and Clinical Medicine 46:128-134.
3. Isenberg, H.D. Clinical Microbiology Procedures Handbook 2nd Edition.
4. 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.