The overall objective of this project is to develop commercially viable methods for the control of pathogens in tree nuts (almonds, pistachios and walnuts) and raisins which lead to foodborne illness in humans and animals. This includes both the control of fungal pathogens (mycotoxins) as well as bacterial pathogens such as E. coli and Salmonella. One approach will be the development of biocontrol agents using bacteria, yeast and non-toxigenic Aspergillus carbonarius. In addition, a better understanding of organic and conventional farming systems will provide new insights on mycotoxin control. The specific objectives for the period covered by this project plan are as follows: Objective 1: Define the critical control points for pathogen contamination during the production stream. Place particular emphasis on agricultural water sources including dairy waste water. Using both cultural and non-cultural based methods we will identify the points in the developmental process, as well as the processing cycle where tree nuts are most likely to become contaminated with human pathogenic bacteria and mycotoxigenic fungi. Objective 2: Evaluate the microbial ecology of organic v. conventional practices. Although several surveys have reported that consumers equate organically grown food stuffs with higher levels of food safety, little is known about how these practices affect the microbial population structure or mycotoxin levels in tree nuts. We will address the influence of phyllosphere microbial community on the population diversity of A. flavus in tree nut orchards, and A. carbonarius on grape surfaces in both farming systems. Objective 3: Delineate the factors affecting cross-contamination during processing and develop a potential intervention strategy during storage. A number of experiments point to a strong possibility that cross-contamination of toxigenic fungi is possible during processing and storage, although no research appears to have been done on the transfer in actual (not laboratory) processing and storage conditions. Development of novel approaches to prevent the growth of storage fungi and production of harmful toxins is a high priority in the almond industry. Edible films and coatings (EFC) containing antimicrobial natural compounds will be tested for their efficiency to reduce mycotoxin in stored almonds. Objective 4: Develop biological-control/intervention technologies using competitive or antagonistic microorganisms such as yeasts or bacteria that can be mass-produced and effectively utilized in a variety of pre- or post-harvest environments. Bacterial and yeast biocontrol agents will be tested in almond orchards and vineyards to control A. flavus and A. carbonarius, as well as human pathogenic bacteria, E. coli and Salmonella. EPA registration of the patented yeast, Pichia anomala will be pursued for commercial application. Methods to enhance the biocontrol efficacy will be developed.