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INVESTIGATOR: Crowell, C. R.

PERFORMING INSTITUTION:
N Y AGRICULTURAL EXPT STATION
GENEVA, NEW YORK 14456

UNDERSTANDING THE EPIDEMIOLOGY AND INFECTION BIOLOGY OF MELAMPSORA AMERICANA WILLOW RUST IN THE NORTHEAST UNITED STATES

NON-TECHNICAL SUMMARY: Bioenergy production in the United Stateshas largely relied on corn and soybean production, both of which have major concerns regarding sustainability. Shrub willow provides an exciting alternative to these traditional fuel sources in the northeast United States because of its favorable carbon emission ratios, ability to grow on marginal lands, and potential for various ecosystem services. However, a fungal disease known as willow rust can significantly reduce yield and can strain the economic success for growers, making this transition from traditional fuel sources a challenge. The use of fungicides are not cost effective for control of this disease, therefore breeding for resistant plants is the most promising route for industrial success. Since this is a poorly understood pathogen, we need to investigate the population and epidemiology of this pathogen in the northeast US to ensure we are accurately and effectively breeding for resistance in new shrub willow plants. Additionally, this type of fungal pathogen is extremely devastating for some of the most important cropping systems around the world, including wheat and coffee, yet is very poorly understood due to the challenges of working with this pathogen. For these reasons, I plan to 1.) investigate the secondary spread of the rust pathogen within a shrub willow field to get a sense of the pathogen epidemiology, and 2.) explore molecules involved in early infection of the rust to explore the genes essential for pathogenicity to serve as targets for future study. Objective 1 will require heavy fugal sampling of a shrub willow field throughout the growing season and DNA sequencing of these collected fungi to look at the differences between individuals. Objective two will require growing a select fungus on a variety of surfaces and performing RNA sequencing to look at how the fungus is responding to the different media. These projects will not only impact directly those involved shrub willow cultivation, including shrub willow breeds and growers, but will have the potential to change our understanding of fungi across the entire field of mycology.

OBJECTIVES: Goal 1.) Foundational population discovery though genotyping-by-sequencing: Initial understanding of the population biology has already been determined, howevera thorough understanding of secondary spread within shrub willow fields is still not understood. For this reason, thorough and systematic sampling within a single field over a growing season is necessary. Objective 1.) Complete isolate collections in Geneva NY and Morgantown WV following strict sampling scheme described in methods. Objective 2.) Generate single pustule isolates from infected leaves collected from the field and collect uredospores for DNA extraction. Objective 3.) Submit DNA for Genotyping-by-sequencing at University of Wisconsin and analyze data using Tassel v5 pipeline.Objective 4.) incorporate in manuscript and present data at scientific meetingsGoal 2.): Identificationof candidate effectors associated with early invasion:Growing knowledge suggests that fungal-plant interactions begin as early as penetration of the fungal spore. In rust systems, nearly all effector biology research has focused on the haustorial stage or rust growth for effector discovery, leaving early penetration as an unknown field of effector research. This gap in fungal effector biology has the potential to reveal mechanisms of infection across all rust fungi, not just the particular rust-host interaction. Objective 1.)Perform proposed appressorium RNAseq project on three select media: polyethylene flat sheet, oil collodion membrane, and leaf tissue. Objective 2.) submit extracted RNA for RNAseq and analyze data subtracting out genes identified and upregulated on the first wo media from the leaf tissue condition. Objective 3.) use smaller subset of RNAseq reads for effector prediction based on predicted secretion signal and length of transcript and qPCR validation. Objective 4.) compare list of candidates to separately in silico prediction of effectors using EffectorP. Objective 5.) Incorporate results into a manuscript and submit for publication; present data at scientific meetings.