BILE ESCULIN AGAR (BEA)

Cat. no. G12 Bile Esculin Agar, 15x100mm Plate, 18ml 10 plates/bag
Cat. no. J38 Blood Agar / Bile Esulin Agar, 15x100mm Biplate, 10ml/10ml 10 plates/bag
Cat. no. J165 Bile Esculin Agar / Sodium Chloride (NaCl) 6.5% without Indicator, 15x100mm Biplate, 10ml/10ml 10 plates/bag
Cat. no. L10 Bile Esculin Agar, 16x100mm Tube, 5.5ml Slant 20 tubes/box

INTENDED USE

Hardy Diagnostics Bile Esculin Agar (BEA) is recommended for use as a differential medium in the isolation and presumptive identification of enterococci/group D streptococci.

SUMMARY

Esculin hydrolysis was first described by Rochaix in 1924.(8) Swan first introduced the use of Bile Esculin Agar in 1954.(9) In 1970, Facklam and Moody determined that the use of the bile esculin test was a reliable way of identifying group D streptococci from non-group D streptococci.(3) When using BEA in biochemical testing of group D streptococci, they found that all group D streptococci will blacken this medium.(3) Other researchers have used BEA for the presumptive identification of Enterobacter spp., Klebsiella spp., and Serratia spp., among the Enterobacteriaceae.

This medium contains esculin, ferric citrate to provide ferric ions, and 4% oxbile to inhibit most other strains of non-group D streptococci. Esculin is hydrolyzed by group D streptococci to form dextrose and esculetin. This compound reacts with the ferric ions contained within the medium, turning the medium from its original amber color to a dark brown to black. Thus the tolerance to the presence of bile and the hydrolysis of esculin provide the means to presumptively identify group D streptococci.

FORMULA

Ingredients per liter of deionized water:*

Oxbile (Oxgall) 40.0gm
Pancreatic Digest of Gelatin 5.0gm
Beef Extract 3.0gm
Esculin 1.0gm
Ferric Citrate 0.5gm
Agar 15.0gm

Final pH 6.6 +/- 0.2 at 25ºC.

* Adjusted and/or supplemented as required to meet performance criteria.

STORAGE AND SHELF LIFE

Storage: Upon receipt store plated media (Cat. no. G12 and J38) at 2-8ºC. away from direct light. Store tubed media (Cat. no. L10) at 2-30ºC. away from direct light. Media should not be used if there are any signs of deterioration (shrinking, cracking, or discoloration), contamination, or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat and freezing.

PRECAUTIONS

PROCEDURE

Specimen Collection: This product is not intended for primary isolation of patient specimens. It should be used only with cultures of isolated organism. This product is used in conjunction with other biochemical tests to identify cultures of isolated organism.

Method of Use: Allow the BEA medium warm to room temperature before use. Inoculate and streak the medium with one isolated pure colony. Incubate in an aerobic atmosphere at 35ºC for 24-48 hours. Observe for growth and blackening of the medium.

INTERPRETATION OF RESULTS

Esculin hydrolysis is indicated by a blackening of the media around the colonies. Consult listed references for the identification of colony morphology and further biochemical tests required for identification. (1,2.4,6)

LIMITATIONS

Some strains of Staphylococcus, Aerococcus , and Listeria monocytogenes may grow in the presence of bile and hydrolyze esculin. L. monocytogenes will form minute black colonies.

A heavy inoculum on BEA may cause interpretation of the bile esculin test difficult to read. Excess inoculum decreases the ability of the bile to inhibit growth of other gram-positive organisms that may hydrolyze esculin.

There are a few streptococci that do not hydrolyze esculin but will grow in the presence of bile. Growth without blackening of this medium does not constitute a positive test.

BEA does not contain azide; as a result, gram-negative rods will grow on this medium. Many of these organisms may hydrolyze esculin.

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, slides, staining supplies, other culture media, microscope, incinerator, and incubators, as well as serological and biochemical reagents, are not provided.

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Enterococcus faecalis **
ATCC ® 29212
A 18-48hr 35°C Aerobic Growth; blackening of media around colonies
Streptococcus pyogenes **
ATCC ® 19615
B 18-48hr 35°C Aerobic Partial to complete inhibition

** Recommended QC strains for User Quality Control according to the CLSI document M22 when applicable.

User Quality Control

PHYSICAL APPEARANCE

Bile Esculin Agar should appear clear, and light amber with a bluish-tinge in color.

E. faecalis growing on BEA

Enterococcus faecalis (ATCC ® 29212) colonies growing on Bile Esculin Agar (Cat no. G12). Incubated aerobically for 24 hours at 35ºC.

S. pyogenes inhibited on BEA

Streptococcus pyogenes (ATCC ® 19615) growth inhibited on Bile Esculin Agar (Cat no. G12). Incubated aerobically for 24 hours at 35ºC.

REFERENCES

1. Anderson, N.L., et al. Cumitech 3B; Quality Systems in the Clinical Microbiology Laboratory, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C.

2. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

3. Facklam, R.R., and M.D. Moody. 1970. Appl. Microbiol., 20:245

4. Isenberg, H.D. Clinical Microbiology Procedures Handbook, Vol. I, II & III. American Society for Microbiology, Washington, D.C.

5. MacFaddin, J.F. 1985. Media for Isolation, Cultivation, Identification, Maintenance of Bacteria, Vol. I. Williams & Wilkins, Baltimore, MD.

6. Jorgensen., et al. Manual of Clinical Microbiology, American Society for Microbiology, Washington, D.C.

7. Quality Assurance for Commercially Prepared Microbiological Culture Media, M22. Clinical and Laboratory Standards Institute (CLSI - formerly NCCLS), Wayne, PA.

8. Rochaix. 1924. Cr. Soc. Biol., Paris; 90:771.

9. Swan, A. 1954. J. Clin. Path.; 7:160-163.


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121415hh