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Title: A SENSITIVE KINETIC MICROPLATE METHOD FOR QUANTIFYING THE ANTIBACTERIAL ACTIVITY OF BIOLOGICAL FLUIDS

Author
item HOLOWACHUK, S - MISS STATE UNIVERSITY
item El Balaa, Mohamad
item BUDDINGTON, R - MISS STATE UNIVERSITY

Submitted to: Journal of Microbiological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/19/2003
Publication Date: 9/1/2003
Citation: Holowachuk, S.A., El Balaa, M.F., Buddington, R.K. 2003. A kinetic microplate method for quantifying the antibacterial properties of biological fluids. Journal of Microbiological Methods 55:441-446.

Interpretive Summary: This paper describes the development of an assay to measure antibacterial activity in swine pancreatic juice. Increasing reports of microbial resistance to antibiotics calls for intensifying the search for new antibiotic compounds. Of specific interest in this research are gastrointestinal secretions and their potential role in suppressing infections caused by gastrointestinal and food borne pathogens. The new assay developed here will permit determining the effects of diet changes and potential other treatments on increasing the production of antibacterial activity in pancreatic juice. This method provides a tool for measuring swine pancreatic juice activity in hopes of determining factors that could boost the natural defenses to enhance food safety and increase health in animals and reduce levels of potential human pathogens in animal manure.

Technical Abstract: We describe a microtiter broth dilution method for measuring antibacterial activity of secretory fluids that incorporates a standard curve to convert observed activity to units of gentamcin. The approach used Staphylococcus aureus subsp. Aureus strain ATCC® Escherichia Coli ATCC 25922, and Lactobacillus spp, but is suitable for other indicator organisms and antibiotics. Assay sensitivity is dependent on the species and density of indicator organism, assay condition (e.g., type and concentration of growth medium pH. A linear standard curve generated by plotting the area under the growth curve (AUGC) versus gentamicin concentration (log transformed) was used to quantify antibacterial activity in antibiotic units. The inclusion of an internal antibiotic standard allows for comparisons among samples analyzed on different days, from different studies, and from different tissues or organs. This adaptation of the microtiter broth method will be useful for investigating the antibacterial activity of natural and man-made compounds under different conditions, and for understanding how age, dietary intake, and health and disease states influence antibacterial activity of secretory fluids.