A Method for Isolation of Ascorbic Acid Fermenting Bacteria

The objectives of this preliminary communication are two: to describe a procedure for the isolation of ascorbic acid fermenting bacteria, from suitable material, and to describe briefly the results obtained with this procedure. This preliminary communication does not presume completeness either with respect to the types or strains of bacteria which may actually utilize ascorbic acid for energy, or with regard to the patterns of fermentation of this substance by specific strains of bacteria.

Plain, nutrient broth is reinforced with ascorbic acid.∗ Fifteen to 20 mg. per 10 cc. of broth is a preferable amount, both because this is sufficient to indicate unequivocally that decomposition has taken place, and also because within these limits the initial reaction of the broth is not brought below pH 6.0, a point at which the bacteria thus far encountered grow readily. Also, a reaction slightly on the acid side is conducive to the stability of ascorbic acid. The broth, thus prepared is inoculated with stomach contents, feces, or other suitable material and incubated at 37° for 18-30 hours. Then, if ascorbic fermenting bacteria are present, a considerable growth will be noticed. From this first, or preliminary culture, a second broth culture is made, using 2 to 5 loops for the inoculum. After incubation and growth, plates are made either upon plain, or, preferably, glucose agar. (Ascorbic acid agar is not so well suited for this purpose.) Colonies are picked off, tested for purity and eventually for ascorbic acid utilization.