Enterococcus Selective Broth (Emterococcosel Broth)

Casein and soy peptones provide essential nutrients for growth. Dextrose is the fermentable carbohydrate energy source. Sodium chloride maintains the osmotic balance. Sodium citrate provides additional carbon. Sodium azide is an inhibitor. Sodium sulfite when reduced produces H₂S. L-Cystine lowers the oxidation-reduction potential by removing oxygen to maintain a low pH. Crystal violet is a pH indicator.

Samples are inoculated into this selective medium and tubes are incubated at 35 °C for 18-24 hours in a normal atmosphere. The growth of streptococci can be determined by the formation of a granular precipitate at the bottom of the tube, with the liquid above being clear or slightly turbid. At this point, perform a Gram stain and restreak on Trypticasein Soy Agar (LC-1068) blood plates or Blood Agar Base (LC-1108) to determine the type of hemolysis and to purify the culture.

The presence of variable length chains of Gram-positive cocci inhibited by bacitracin in a low concentration, catalase negative and insoluble in bile or bile salts, constitutes a valid presumptive identification of Group A beta-hemolytic streptococci. The definitive identification of the streptococci groups can be made by performing other biochemical tests such as esculin hydrolysis, pyruvate hydrolysis, etc. Also, serological typing, using Lancefield antisera methods, or more conveniently, the techniques of co-agglutination of Edwards and Larson can be performed.

Typical formula in g/L, adjusted and/or supplemented as required to meet performance criteria.

Preparation

Suspend 30.6 grams of the medium in one liter of distilled water. Mix well and dissolve by heating with frequent agitation. Boil for one minute until complete dissolution. Dispense in 10 mL screw-capped tubes and sterilize in autoclave at 118 °C for 15 minutes. DO NOT OVERHEAT as the medium will become too inhibitory.

Instructions for use

To differentiate streptococci and pneumococci, place bacitracin and optochin discs in the area of the inoculum on the Blood Agar plates and incubate for 18-24 hours at 35 ± 2 °C under the recommended conditions.

Perform a Gram stain, catalase and bile solubility tests on characteristic colonies taken from the Blood Agar plate or from the growth obtained from the broth.

• Washington, D.C. 2nd Ed., 1974
• Facklam and Carly, 1985, Manual of Clinical Microbiology, Lennette and others (Eds). 4th Ed. ASM, Washington DC.