2006 Annual Report 4d.Progress report. This report serves to document research conducted under a trust agreement between ARS and Sterilex Corporation through and NRI grant. Our focus for the second year of this three-year project has been to further study with baseline data established during the first year. Methods that were developed during the first year to produce Listeria monocytogenes biofilms and test Sterilex technology were used to complete chemical screening of potential product components for biofilm removal activity. We will use the results of the chemical screening data to optimize product formulations for efficacy against L. monocytogenes biofilms. Selection of test components varied the calcium sequestrants and nonionic surfactants in the formulations. Colorimetric measurement of dye staining and extractions of L. monocytogenes biofilm were used as the biofilm screening test agent. Four strains of Listeria biofilms were grown and tested: two laboratory strains and two field strains. Field strains appeared to be more susceptible to the chemistry than laboratory strains. Six concentrations of 18 product components have been screened against the four strains of Listeria for three time periods. Each variable was tested against L. monocytogenes biofilms in two trials, and duplicate samples were measured within each trial. Data from the experiments are currently undergoing analysis. The data will allow us to quantify the percent reduction in biofilm between treated and untreated samples, as well as to determine the standard deviation of the efficacy measure. The analysis will determine the role of phase-transfer in antibiofilm activity and if high pH is a prerequisite for efficacy. Finally, the data will be used to determine the mechanism of action of current technology in conjunction with the chemical screening to formulate two compounds to be further tested for efficacy. The final product will be developed in the third year of research to meet USDA 'zero tolerance' regulations for L. monocytogenes biofilms, and its activity will be confirmed by the AOAC use-dilution test. Expected Impact Protocol development and validation is critical to the establishment of standards for registration of EPA biofilm claims for used in food processing. Such claims will educate end users about biofilms and the important of utilizing products that can address the problem of biofilms in food processing environments. Understanding the mechanism of action underlying the effectiveness of the current Sterilex technology is critical for the future development of effective products for use against biofilms containing Listeria monocytogenes and other harmful pathogens. The results of these experiments will yield an optimized formulation of a disinfectant for meat and poultry RTE food processing and production equipment that is more effective in reducing Listeria monocytogenes biofilm growth than currently used disinfectants, thus minimizing the risks to food safety from pathogenic contamination. 7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Arnold, J.W., Zambelli-Weiner, A., deLaubenfels, E., and Kramer, S. 2006. Quantitative assessment of hard surface disinfectant activity against the food-borne pathogen, Listeria monocytogenes. J AOAC Int.