Rapid spot test for the determination of esculin hydrolysis

Abstract

Esculin hydrolysis is a useful test in the differentiation of gram-positive and gram-negative bacteria covering a wide spectrum of aerobes, facultative anaerobes and anaerobes. Commonly utilized methods require a minimum of 18 h incubation in broth or agar medium and utilize the production of a brown-black compound, due to the combination of ferric ions with the hydrolysis product esculetin, as indicator. A procedure is presented that requires 15-30 min for completion and utilizes fluorescence loss as the indicator of hydrolysis. Esculin fluoresces at 366 nm; the hydrolysis product esculetin does not. More than 1400 strains of gram-positive and gram-negative bacteria [Enterobacteriaceae and Listeria members] were tested. There was 98.4% correlation between the spot test and esculin broth and 97% correlation with the bile-esculin agar.