ollow steps 1 through 4, above, except adjust the pH to the range of 6.8 to 7.0 with hydrochloric acid before autoclaving, cooling, and pouring.
Either Sabouraud agar or its Emmons version can be made more selective by adding antibiotics. Commonly used are gentamicin, which inhibits gram-negative bacteria, and/or chloramphenicol, which inhibits a wide range of gram-positives and gram-negatives, and cycloheximide, which inhibits primarily saprophytic fungi but not dermatophytes or yeasts (3). Chloramphenicol and gentamicin are used at 50 mg/liter (dissolved in 10 ml of 95% ethanol before adding to molten media) and cycloheximide at 0.5 g/liter (dissolved in 2 ml of acetone first) (2). Antibiotics should only be added after media has been autoclaved and then cooled to ~45 to 50°C. Keep all plates at 4°C until they are used, regardless of whether they contain antibiotics.
Sabouraud agar plates can be inoculated by streaking (see the Streak Plate Protocol for an explanation of this method), as with standard bacteriological media, or by exposing the medium to ambient air. Typically, molds are incubated at room temperature (22 to 25°C) and yeasts are incubated at 28 to 30°C or 37°C if suspected of being dimorphic fungi. Incubation times will vary, from approximately 2 days for the growth of yeast colonies such as Malasezzia, to 2 to 4 weeks for growth of dermatophytes or dimorphic fungi such as Histoplasma capsulatum. Indeed, the incubation time required to acquire fungal growth is one diagnostic indicator used to identify or confirm fungal species.