SCREENING METHODS: Inoculum

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Inoculum

Mueller-Hinton medium allows good growth of most non-fastidious bacteria and is generally low in antagonists. It also meets the requirements of the NCCLS standard and is recommended as reference medium for agar and broth-dilution tests (Anon, 2003).

The level of infection, i.e. inoculum concentration can have a profound influence on the antibacterial and antifungal potency of a sample, endorsing the need for standardization of inoculates (Anon., 2003).

In dilution methods, an inoculum of about 105 CFU/ml is adequate for most bacterial species while for yeasts and fungi between 103 and 104 CFU/ml is sufficient (Hadacek and Greger, 2000). A too low inoculum size (e.g. 102 CFU/ml) will create many false-positives, while a too high inoculum size (e.g. 107 CFU/ml) will hamper endpoint reading and increase the chances for false-negatives. Bacterial or yeast inoculates can be prepared from overnight cultures or from existing biofreeze stocks. It is recommended to collect from cultures during the logarithmic growth phase and always to take four or five colonies of a pure culture on agar to avoid selecting an atypical variant (Anon., 2003).

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