2007 Annual Report 1a.Objectives (from AD-416) Mass spectrometry methods and technology will be developed for the detection of plant pathogens. These objectives should help accomplish goals to develop alternative technologies that will enable the rapid detection of unknown pathogens. The research will be performed by a multidisciplinary team from the USDA-ARS Plant Sciences Institutes Soybean Genomics and Improvement Laboratory and the Animal and Natural Resources Institutes Biotechnology and Germplasm Laboratory in Beltsville, MD. 1b.Approach (from AD-416) We will first use mass spectrometry to identify proteins from pure cultures of plant pathogenic fungi and determine if these mass spectra match those derived from GenBank or fungal genome/proteome databases. If the mass spectra match, then we will know that this method is suitable for detecting these fungi. If we cannot unambiguously identify a fungal protein with common protein databases, i.e. there is not enough known genome or proteome data for a fungus, then we will use the mass spectra as biomarkers for characterizing a fungus, which will aid in pathogen identification in future assays. Tests will also be performed to establish the lower limits at which fungal proteins that can be detected. Next, we will infect plants with plant pathogenic fungi, including the bioterrorism threat and quarantined soybean rust, and will use mass spectrometry methods and technology for the detection of these pathogens against a plant protein background. We will also mix fungal mycelia and spores into soil to determine if mass spectrometry would be sensitive enough to screen and survey field soil samples. 3.Progress Report This report serves to document research conducted under a reimbursable agreement between ARS and USDA-CSREES via a research grant entitled “Mass Spectrometry for the Proteomic Detection and Analysis of Plant Pathogenic Fungi Including Soybean Rust”. This project supports objectives of parent project 1275-21220-221-00D, “Genomics and Proteomics Approaches to Broadening Resistance of Soybean to Pests and Pathogens” and additional details of research can be found in the report for the parent project. Proteins from Ustilago maydis, Fusarium graminearum, Fusarium oxysporum, Rhizoctonia solani, and the soybean pathogen Phytophthora sojae were identified by mass spectrometry. Bioinformatics and statistical approaches were evaluated to best interpret the results and it was concluded that routine bioinformatics approaches are insufficient when used in conjunction with mass spectrometry to identify pathogens or diagnose disease. A novel bioinformatics approach has been investigated to determine if raw peptide tandem mass spectra can serve as pathogen biomarkers. These have been inserted into a searchable library and can used for pathogen detection. Annual budget reports, annual written progress reports and an annual presentation are used to monitor project progress.