Malonate Broth, Ewing Modified

Principle And Interpretation

Leifson developed a synthetic liquid medium, which differentiated Aerobacter (now Enterobacter) from Escherichia species based on their ability to utilize malonate (1) where Enterobacter utilizes malonate and Escherichia does not. Ewing et al further modified this medium by the incorporation of yeast extract and dextrose (2). The addition of yeast extract, a source of vitamins, and a relatively small amount of dextrose, a minimal carbon source, is included in Ewings modification to stimulate the growth of some organisms. The medium, therefore, will support the growth of organisms that cannot utilize malonate or ammonium salt. Any spontaneous alkalinization produced by such organisms is buffered by the phosphate system and counteracted by the acid produced by the fermentation of the small amount of dextrose (3). An alkaline reaction (blue color) is produced in this medium by organisms capable of utilizing malonate and ammonium sulfate.

An organism that can simultaneously utilize sodium malonate as its carbon source and ammonium sulfate as its nitrogen source produces alkalinity due to the formation of sodium hydroxide (3). The alkali changes the color of the bromothymol blue indicator in the medium to light blue and finally to prussian blue. The color of the medium remains unchanged in the presence of an organism that cannot utilize these substances. Some malonate-negative strains produce a yellow color due to the fermentation of dextrose only, which results in increased acidity causing the pH indicator to change to yellow at a pH of 6.0. Also some malonate-positive organisms produce only a slight alkalinity that causes the results to be difficult to interpret. Therefore these tubes should be compared with an un-inoculated malonate tube (3).