Detection Method for Histamine-Producing, Dairy-Related Bacteria using Diamine Oxidase and Leucocrystal Violet.

A detection method for histamine-producing, dairy-related bacteria was developed that involves a two-step sequential enzyme system. First, isolated bacteria are incubated in MRS broth or trypticase soy broth fortified with histidine. The histamine formed during this incubation period is reacted with diamine oxidase, which catalyzes the oxidation of histamine to form imidazole acetaldehyde, ammonia, and hydrogen peroxide. The hydrogen peroxide is then detected by the formation of crystal violet from the leuco base in the presence of horseradish peroxidase. Liquid culture medium containing bacteria that produce greater than 1200 nmole histamine per ml will develop a positive purple color. Cultures containing bacteria that produce little or no histamine will not develop a purple color. Other amines often found in cheese, such as tyramine, cadaverine, or putrescine, will not interfere with the color formation.