2007 Annual Report 1a.Objectives (from AD-416) The goal of the study is to characterize the systemic pathological effect of pyrrocidine A in mice through (A) intraperitoneal injection with increased exposure time; (B) intravenous administration to obviate the potential for metabolic detoxification in the liver; and (C) oral exposure (gavage). 1b.Approach (from AD-416) Pyrrocidine A is a newly discovered antibiotic produced by the 'protective' maize endophyte Acremonium zeae. This antibiotic exhibits potent activity against kernel rotting fungal pathogens of maize such as the mycotoxin producers Aspergillus flavus (aflatoxin) and Fusarium verticillioides (fumonisin), as well as pathogens of humans including Candida albicans and species of Gram-positive bacteria. Research identified in Objective 5 of the ARS parent CRIS will investigate the epidemiology and biocontrol potential of A. zeae in maize. The Preharvest Mycotoxins Panel reviewing this ARS CRIS project posed the following question: "What safety information is available for pyrrocidines that could convince regulators that its levels could be increased in corn grain and not pose a safety risk for consumers?" The in vivo effect of pyrrocidine A and B to animals or humans is not known. An in vitro study performed by the Department of Pathobiology shows that pyrrocidine A and B are cytotoxic to HepG2 cells (a cell line derived from a human hepatocellular carcinoma) and PK15 cells (a cell line derived from a normal pig kidney) within 24 hours of treatment. This warrants a study of the in vivo toxicity of pyrrocidine A in mice that will include a pilot study to determine toxic dose levels followed by a study of potential target organs and dose response for pyrrocidine A toxicity including comprehensive histopathology and clinical pathology. 3.Progress Report This report documents accomplishments under a Specific Cooperative Agreement between Agricultural Research Services (ARS) and the University of Illinois. Additional details of research can be found in the parent project 3620-42000-033-00D entitled "Fungal Endophytes of Maize: Gene Products Conferring Resistance to Aflatoxin and Fumonisin." The Authorized Departmental Officer's Designated Representative monitors this agreement by: e-mails, telephone calls, providing samples for testing, and reviewing text for poster presentation. Pyrrocidine A is a newly discovered antibiotic produced by the ‘protective’ maize endophyte Acremonium zeae. This antibiotic exhibits potent activity against kernel-rotting fungal pathogens of maize including the aflatoxin producer Aspergillus flavus, as well as pathogens of humans including Candida albicans and species of Gram-positive bacteria. The Preharvest Mycotoxins Panel reviewing this project posed the following question: “What safety information is available for pyrrocidines that could convince regulators that its levels could be increased in corn grain and not pose a safety risk for consumers?” A study performed by a university scientist in the Department of Pathobiology, University of Illinois, shows that pyrrocidine A and B are cytotoxic to the human hepatocellular carcinoma cell line and a cell line derived from a normal pig kidney within 24 hours of treatment. These results warranted a study of the potential for whole-animal toxicity. Mice were evaluated for toxicity based on clinical and morphologic evaluation. Selected tissues, including sections of major organs, were examined histologically. Toxicity due to pyrrocidine A, based on clinical signs, organ weights, and gross microscopic lesions, was not identified at doses up to 10 milligram/kilogram body weight. To further examine whether or not pyrrocidine A is toxic to mice in vivo, the oral and intravenous routes of administration will be used and multiple doses of pyrrocidine A will be given to mimic potential natural exposure.