2007 Annual Report 1a.Objectives (from AD-416) To evaluate, validate, and where necessary, develop new innovative, robust and valid predictive models for the responses of microbial pathogens, including foodborne threat agents, in select food matrices, as a function of: temperature, food formulation, competitive microflora, physiological history, and surface transfer. To develop novel approaches to assess model performance and robustness, leading to more efficient strategies for producing and extrapolating models to different classes of food. To determine the probability distribution of lag phase duration (LPD) for foodborne pathogens, as a function of the previous bacterial physiological history, to allow risk managers to estimate worst-and best-case scenarios for pathogen behavior, depending on likely sources of contamination; To identify molecular markers that discriminate bacterial lag, growth and stationary phases, thus leading to more mechanistic models and greater certainty for LPD prediction. 1b.Approach (from AD-416) Quantitative data will be collected for the effects of selected environmental parameters on foodborne pathogen growth, survival and inactivation. Relevant environmental conditions will include food formulation, native microbial flora, inoculum level, bacterial history, and the effects of food process operations. Priority pathogen-food combinations will be identified through stakeholder interactions and by identifying sensitive data gaps in microbial risk assessment. Experimental data will be used to confirm and where necessary produce primary growth and inactivation models, as well as probabilistic models for growth/no growth interfaces and microbial transfer among food processing surfaces. Model performance will be described using independent validation data from ongoing experiments with food matrices and microbiology databases such as ComBase. The resulting technologies will be transferred to stakeholders vis the ARS Pathogen Modeling Program and process risk model software. 3.Progress Report Research is also conducted under the following Agreements: 1935-42000-057-01G "Eliminating Listeria Monocytogenes from Ready-to-Eat Products" A General Assistance Grant with University of Arkansas. 1935-42000-057-02G "Dose Response of Listeria Monocytogenes in Pregnant Guinea Pigs for Use In Risk Assessment" a grant with the University of Georgia 1935-42000-057-03G "Predictive Models for Thermal Inactivation of Listeria Mnocytogenes on the Surface of Hot Dogs." A General Assistance Grant with Purdue University. 1935-42000-0570-04G "Transfere Coefficients for L. Monocytogenes to and from food contact surfaces found in meat fabrication facilities."A General Assistance Grant with Michigan University. 1935-42000-057-05R - "HACCP Assistance for Small and Very Small Processors with Development and Validation of meat chilling Rates." A Reimbursable Agreement with University of Nebraska.1935-42000-057-07R - "HACCP Training and Research to Assist Meat Processors with Process Deviations for Lethality and Stabilization." A Reimbursable Agreement with the University of Nebraska. 1935-42000-057-08R - "HACCP Assistance for Small and Very Small Meat Processors: Challenge Studies and Predictive Modeling for Validation of Critical Limits." A Reimbursable Agreement with the University of Wisconsin. 1935-42000-057-09R - "An Internet-Based Portal to Assist Small and Very Small Food Processors in Meeting Food Safety Regulations." A Reimbursable Agreement with Food Safty & Inspection Service (FSIS), USDA. 1935-42000-10R - "Study on Survivability of Yersinia Pestis and Bacillus Anthracis in Raw and Cooked Ground Beef." A Reimbursable Agreement with the Food Safety and Inspection Service (FSIS), USDA. 1935-42000-057-11S - "Acetate, Lactate, and Diacetate Levels in RTE Processed Meat & Poultry Products Collected At Retail & Correlation with Occurrence of L. Mono." A Specific Cooperative Agreement with the University of Tennessee. 1935-42000-057-12S "Listeria Monocytogenes Contamination of Deli Meat Slicers: Risk and Communication." A Specific Cooperative Agreement with Michigan State. 1935-42000-057-13S - "An Internet-Based Predictive Microbiology Portal to Assist Small and Very Small Food Processors in Meeting Food Safety Regulations." A Specific Cooperative Agreement with Rutgers University. 1935-42000-057-14S - "Construction and Characterization of Reporter Strains for the Identification of Physiochemical Parameters Influencing...of Escherichia Coli." A Specific Cooperative Agreement with the University of Wisconsin. 1935-42000-057-15N - "Transfer of Listeria Monocytogenes Among Salmon and Slicing Machines. A Geneal Assistance Grant with VTT Echnical Research Center - Finland. 1935-42000-057-16G - "Support for the International Conference On Microbial Risk Assessment (PA06-15)." A General Assistance type of Agreement between ARS Office of Technlogy Transfer. Additional details can be found in the specific reports for these agreements. The agreements were monitored via regular meetings, emails or conference calls. 4.Accomplishments Safe time/temperature for cooling of cooked foods. Improper storage and/or inadequate cooling practices in retail food operations have been cited as the cause of food poisoning for 97% of Clostridium perfringens outbreaks. Mathematical models were developed to predict the relative growth of C. perfringens from spores at temperatures applicable to the cooling of cooked uncured chicken. The growth data /predictive models on the safe cooling rate of meat will enable the food industry to assure that cooked products remain pathogen-free. The research is relevant to Component 1.2 (Pathogen Reduction Postharvest), Problem Statement 1.2.7 (Risk Assessment), in the 2006-2010 NP 108 Action Plan. Behavior of L. monocytogenes in Cooked Ham Containing Lactate and Diacetate. The effect of lactate and diacetate on L. monocytogenes in ham at temperatures that the product is likely to be exposed to during manufacturing and distribution is lacking. Predictive model describing the behavior of L. monocytogenes in ham containing sodium lactate (1.0-4.2%) and sodium diacetate (0.05-0.2%) at storage temperatures of 0°-45°C were developed. The model elucidates the effects of both additives and storage temperature on L. monocytogenes, and enables the producers to select the concentrations of lactate and diacetate that are able to control the growth of L. monocytogenes in ham products. The research is relevant to Component 1.2 (Pathogen Reduction Postharvest), Problem Statement 1.2.7 (Risk Assessment), in the 2006-2010 NP 108 Action Plan. Microbiological Safety of Fermented Sausages. The use of raw meat in the manufacturing of fermented dry and semidry sausage may introduce L. monocytogenes and Salmonella spp into the finished product. Understanding the survivability of both pathogens in sausage during the manufacturing processes allows processing/product parameters be selected to minimize the presence of both pathogens in finished products. Studies conducted to collect the inactivation data of L. monocytogenes and S. typhymurium in sausage were completed. Models were developed to describe the rates of inactivation of both pathogens during sausage manufacturing. The results will be used by the manufacturers to determine the process/product parameters to meet the food safety requirement. The research is relevant to Component 1.2 (Pathogen Reduction Postharvest), Problem Statement 1.2.7 (Risk Assessment), in the 2006-1010 NP 108 Action Plan. Transfer of E. coli O157:H7 during mechanical slicing of deli meats. Food safety managers currently lack the ability to predict the microbial pathogen transfer in slicing operation for ready-to-eat foods. Models were developed for pathogen transfer prediction during mechanical slicing for RTE deli meats. Predictive models will be useful in developing HACCP plans and in risk assessment development for ready-to-eat meats. By understanding the surface transfer, the production or retail (Franchise) equipment and operations for RTE deli meat may be further improved and therefore, reduce the possibility of outbreaks. Finally, the models will be available through the PMP and ComBase. The research is relevant to Component 1.2 (Pathogen Reduction Postharvest), Problem Statement 1.2.7 (Risk Assessment), in the 2006-2010 NP 108 Action Plan. Expression of 70-kb virulence plasmid (pYV/pCD) phenotypes in Yersinia species. Risk assessors in food safety management need to predict the fate of Yersinia pestis for scenarios where bulk foods could be contaminated, thereby exposing a relatively large number of individuals. Therefore, diagnostic markers were developed to predict the growth of Y. pestis and Y. pseudotuberculosis in ground beef and ground pork. This accomplishment addressed on the phenotypic expression of the 70-kb virulence plasmid (pYV/pCD)-associated genes in Yersinia pestis and Y. pseudotuberculosis. The pYV/pCD-encoded phenotypes expressed can be used as virulence markers for detection of these pathogens in foods. This will assist the government risk assessor and food companies in detecting and determining the fate of Y. pestis in contaminated foods, thus improve exposure assessment and aid in designing more effective food safety controls. The research is relevant to Component 1.2 (Pathogen Reduction Postharvest), Problem Statement 1.2.7 (Risk Assessment) and 1.2.9 (Food Security), in the 2006-2010 NP 108 Action Plan. 5.Significant Activities that Support Special Target Populations Collaborated with a small producer of fermented sausage named Mediterranean Foods, a company with 2 employees and an annual sale of less $200,000, located in Upper Darby, Pennsylvania, to examine the behavior of E. coli O157:H7, Salmonella, and L. monocytogenes on fermented, semi-dry sausage introduced by post-processing contamination. The producer provides finished commercial products, and ARS conducts the challenge study. 6.Technology Transfer Number of web sites managed 3 Number of non-peer reviewed presentations and proceedings 9 Review Publications Hwang, C., Marmer, B.S. Growth of listeria monocytogenes in egg salad and pasta salad formulated with mayonnaise of various ph and stored at refrigerated and abuse temperatures. Food Microbiology. 24:211-218. Juneja, V.K., Melendres, M.V., Huang, L., Gumudavelli, V., Subbia, J., Thippareddi, H. 2007. Modeling the effect of temperature on growth of salmonella in chicken. Food Microbiology. 24:328-335. Oscar, T.P. 2007. Predictive model for growth of salmonella typhimurium dt104 from low initial and high initial density on ground chicken with a natural microflora. Food Microbiology. 24:640-651. Velugoti, P.R., Lalit, B.K., Juneja, V.K., Thippareddi, H. 2007. Inhibition of germination and outgrowth of clostridium perfringens spores by lactic acid salts during cooling of cooked ground turkey. Journal of Food Protection. 70:923-929. Juneja, V.K., Friedman, M. 2007. Carvacrol, cinnamaldehyde, oregano oil, and thymol inhibit clostridium perfringens spore germination and outgrowth in ground turkey during chilling. Journal of Food Protection. 70:218-222. Juneja, V.K., Bari, L.M., Inatsu, Y., Kawamoto, S., Friedman, M. 2007. Control of Clostridium perfringens Spores by Green Tea Leaf Extracts During Cooling of Cooked Ground Beef, Chicken, and Pork. Journal of Food Protection. 70(6):1429-1433.