Bile Esculin Test

Bile esculin agar (we do not use bile esculin azide agar, as sodium azide is extremely hazardous) is a selective and differential medium which is used to presumptively identify enterococci  and group D streptococci based on the ability of an organism to hydrolyze esculin. 

The bile esculin handout is here.

Bile esculin agar contains oxgall (bile salts) to inhibit the growth of Gram positive organisms other than enterococci and group D streptococci. It also contains nutrients, esculin, and ferric citrate. When an organism hydrolyzes the glycoside esculin to form esculetin and dextrose, the esculetin reacts with the ferric citrate to produce a dark brown or black phenolic iron complex.

If an organism can hydrolyze esculin, the media will turn dark brown or black. However, the test is interpreted as a positive result only if more than half the medium is dark brown or black after incubation. 

On the left, Streptococcus pyogenes does not hydrolyze esculin in the presence of bile. No dark brown complex is formed. This is a negative result. On the right, Enterococcus faecalis hydrolyzes esculin in the presence of bile and turns more than half the medium dark brown. This is a positive result.

The bile esculin test can be used to differentiate Enterococcus spp. and members of the Streptococcus bovis/Streptococcus equinus complex (previously referred to as group D streptococci) from other Streptococcus species.