Advances in Food Mycology

344 Recommended Methods for Food Mycology
Diluents and dilution: The recommended diluent for fungi including
yeasts is 0.1% aqueous peptone. Dilution should be 1:10 (1+9).
Homogenisation: A Coleworth Stomacher or equivalent Bag Mixer
is the preferred type of homogeniser, used for 2 minutes per sample.
Blending for 30-60 sec or shaking in a closed bottle with glass beads
for 2-5 min are less preferable alternatives.
Plating: For mycological studies, spread plates are recommended;
pour plates are less effective. Inocula should be 0.1 ml per plate, spread
with a sterile, bent glass rod.
Incubation: For general purpose enumeration, 5 days incubation as
25°C is recommended. Plates should be incubated upright. A higher
temperature, e.g. 30°C, is suitable in tropical regions.
1.1.2. Direct plating
Direct plating is considered to be the more effective technique for
mycological examination of particulate foods such as grains and nuts.
In most situations, surface disinfection (also commonly referred to as
surface sterilisation) before direct plating is considered essential, to
permit detection and enumeration of fungi actually invading the food.
An exception is to be made for cases where surface contaminants
become part of the downstream mycoflora, e.g. wheat grains to be
used in flour manufacture. In such cases, grains should not be surface
disinfected.
Surface disinfection: Surface disinfect food particles by immersion in
0.4% chlorine solution (household bleach, diluted 1:10) for 2 minutes.
A minimum of 50 particles should be disinfected and plated. The
chlorine solution should be used only once.
Rinse: After pouring off chlorine solution, rinse once in sterilised
distilled or deionised water. Note: this step has not been shown to be
essential, but is generally recommended.
Surface plate: As quickly as possible, transfer food particles with
sterile forceps to previously poured and set plates, at the rate of 5-10
particles per plate, depending on the size of the particles.
Incubation: The standard incubation regimen for general purpose
enumeration is 25°C for 5 days. A higher temperature (30°C) may be
used in the tropics. Plates should be incubated upright.
Results: Express results as per cent of particles infected by fungi.
Differential counting of a variety of genera is possible using a stereomicroscope.