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All cultures should be examined at least weekly for growth and should be held for several weeks before being reported as negative.

Depending on the clinical diagnosis and the agents suspected to cause the infection, other media should be included.

For isolation of fungi causing systemic mycoses and the isolation of aerobic Actinomycetales, two sets of media should be inoculated, with one set incubated at 25 to 30☌ and a duplicate set at 35 to 37☌.

Incubate the plates at 25 to 30☌ in an inverted position with increased humidity.

For isolation of fungi from potentially contaminated specimens, a selective medium should be inoculated along with the nonselective medium.

Streak the specimen as soon as possible after it is received in the laboratory using a sterile inoculating loop to obtain isolated colonies.Note: If desired, 20 units of Penicillin and 40 µg Streptomycin per ml of the medium may be added to make the medium selective for fungi. Mix well and pour into sterile Petri plates.Sterilize by autoclaving at 15 lbs pressure (121☌) for 15 minutes.Heat to boiling to dissolve the medium completely.Suspend 52.0 grams in 1000 ml distilled water.Preparation and Method of Use of Brain Heart Infusion (BHI) Agar BHI Agar is thus a solid medium recommended for the cultivation of fastidious pathogenic bacteria, yeasts, and molds from clinical and non-clinical samples.A more selective formulation containing chloramphenicol and cycloheximide is also available that will allow the recovery of pathogenic fungi while inhibiting a wide range of bacteria and saprophytic fungi.BHI Agar supplemented with (5 to 10%) defibrinated sheep blood is used extensively for the recovery of dimorphic fungi such as Histoplasma capsulatum and other pathogenic fungi such as Coccidioides immitis.Brain Heart Infusion (BHI) Agar is a highly nutritious base that meets the growth requirements of many types of microorganisms including bacteria, yeasts, and molds.Our current formulation contains infusion from calf brain in place of brain tissue and disodium phosphate has replaced calcium carbonate.Rosenow’s formulation was later modified by Hayden who found the addition of crushed marble resulted in favorable growth of dental pathogens.In 1919, Rosenow devised an excellent medium for culturing streptococci by using a dextrose broth supplemented with brain tissue.