The research will perform the initial screening of plant extracts/essential oils against a range of organisms including Gram-positive and Gram-negative pathogenic and spoilage bacteria, and fungi. Using automated techniques, large numbers of compounds, at multiple concentrations, can be rapidly screened. This initial screening will provide the foundation for subsequent research on antimicrobial activity of isolated phytochemical chiompounds.
APPROACH: A variety of non-pathogenic indicator organisms will be used for initial screening of phytochemical compounds. Compounds that show anti-microbial activity will be further screened against appropriate microorganisms: spoilage, pathogenic, probiotic, fermentative. Suggested organisms include approximately equal numbers of Gram-positive and Gram-negative bacteria, and fungi. At least 2 strains of each organism will be tested. All cultures will be obtained from culture collections of the Investigators or purchased from ATCC (American Type Culture Collection). Cultures will be checked for purity and identifying characteristics and "freeze-away" stock cultures prepared. Fresh inoculum will be prepared for each experiment from stock cultures as follows: an isolated colony on the appropriate agar medium will be transferred to the recommended broth and grown to early stationary phase (determined based on optical density measurements in preliminary experiments). Stationary phase cultures will be used in assays. The assay system employed to screen for antimicrobial activity will vary according to the hydrophobicity of the phytochemical and will utilize an automated microtiter plate reader. Cell suspensions of the test organism will be prepared in the appropriate liquid growth medium and transferred to microtiter plate wells. Serial dilutions of the phytochemical will be added to sequential wells and microbicidal/microbistatic activity determined. Appropriate positive/negative controls will be run. Inhibitory concentrations and viability assays will be run. Appropriate cell enumeration studies will be conducted.