Streak the specimen onto prepared media with a sterile inoculating loop to obtain isolated colonies. When used for determining yeast and mold counts, the medium should be adjusted to a pH of approximately 3.5 with sterile tartaric aid and used in the standard pour plate technique. Incubate the plates at 25-30°C in an inverted position (agar side up) with increased humidity.

Tubed slants are used primarily for the cultivation and maintenance of pure cultures. They should be inoculated with an inoculating loop and incubated under the same conditions as the plated medium.

For the isolation of fungi from potentially contaminated specimens, a selective medium should be inoculated along with the nonselective medium. For isolation of fungi causing systemic mycoses, two sets of media should be inoculated, with one set incubated at 25-30°C and a duplicate set at 35 ± 2°C. All cultures should be examined at least weekly for fungal growth and should be held for 4-6 weeks before being reported as negative.

Inoculation of Potato Dextrose Broth with pure cultures of yeasts can assist in their identification.

Difco Potato Dextrose Agar – 39 g; Difco Potato Dextrose Broth – 24 g. Mix thoroughly.

2.      Heat with frequent agitation and boil for 1 minute to completely dissolve the powder.

3.      Autoclave at 121°C for 15 minutes.

4.      To alter the reaction of the agar medium to pH 3.5, cool the base to 45-50°C and aseptically add an appropriate amount of sterile 10% tartaric acid to each liter of medium. Mix well. Do not reheat the medium.

5.      Test samples of the finished product for performance using stable, typical control cultures.

Potato Dextrose Agar is not a differential medium. Perform microscopic examination and biochemical tests to identify isolates to genus and species if necessary.

Growth from tubes inoculated with pure cultures may be used for biochemical and/or serological testing.

For broth, observe cultures for surface growth and pellicle formation.

2.   Marshall, (ed.). 1993. Standard methods for the examination of dairy products, 16th ed. American Public Health Association, Washington, D.C.

3.   United States Pharmacopeial Convention, Inc. 2001. The United States pharmacopeia 25/The national formulary 20 – 2002. United States Pharmacopeial Convention, Inc., Rockville, Md.

4.   MacFaddin. 1985. Media for isolation-cultivation-identification-maintenance of medical bacteria, vol. 1. Williams & Wilkins, Baltimore, Md.

5.   Murray, Baron, Pfaller, Tenover, and Yolken (ed.). 1999. Manual of clinical microbiology, 7th ed. American Society for Microbiology, Washington, D.C.

Isenberg (ed.). 1992. Clinical microbiology procedures handbook, vol. 1. American Society for Microbiology, Washington, D.C.