Decarboxylation Test

The decarboxylase test is used to differentiate the Enterobacteriaceae.

Decarboxylase broth tests for the production of the enzyme decarboxylase, which removes the carboxyl group from an amino acid. Each decarboxylase enzyme produced by an organism is specific to the amino acid on which it acts. We test the ability of organisms to produce arginine decarboxylase, lysine decarboxylase, and ornithine decarboxylase using three different but very similar media.

Decarboxylase broth contains nutrients, dextrose (a fermentable carbohydrate), pyridoxal (an enzyme cofactor for decarboxylase), and the pH indicators bromcresol purple and cresol red. Bromcresol purple turns purple above pH 6.8 and yellow below pH 5.2. Cresol red turns red above pH 8.8 yellow below pH 7.2. 

We also add a single amino acid to each batch of decarboxylase broth; that amino acid may be arginine, lysine, or ornithine. 

If the organism is unable to ferment dextrose, there will be no color change in the medium. If an organism is able to ferment the dextrose, acidic byproducts are formed, and the media turns yellow. As the organisms ferment the dextrose, the media initially turns yellow, even when it has been inoculated with a decarboxylase-positive organism. The low pH and the presence of the amino acid will cause the organism to begin decarboxylation.

If an organism is able to decarboxylate the amino acid present in the medium, alkaline byproducts are then produced. Arginine is hydrolyzed to ornithine and is then decarboxylated. Ornithine decarboxylation yields putrescine. Lysine decarboxylation results in cadaverine. These byproducts are sufficient to raise the pH of the media so that the broth turns purple. 

When you retrieve your decarboxylase broths from the rack on the side counter, be sure to label them immediately! The broths look the same, and if you become confused as to which amino acid is in which tube, your results will be useless.

After you inoculate the decarboxylase broth but before you incubate the tubes, add an overlay of sterile mineral oil to each tube. You do not need a thick layer of oil; you only need enough to cover the surface of the medium. This promotes fermentation by locking out oxygen, and it also prevents false alkalinization at the surface of the medium.

If the inoculated medium is yellow, or if there is no color change, the organism is decarboxylase-negative for that amino acid. If the medium turns purple, the organism is decarboxylase-positive for that amino acid.