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Design and testing of PCR-based diagnostics generated from NGS for distinguishing among Monilinia species of economic importance to US agriculture.

Jeremy Sutherland: USDA-APHIS-PPQ-S&T-CPHST

<div>Fungi in the genus <em>Monilinia</em> cause a devastating brown rot disease of stone and pome fruits. <em>Monilinia fructicola </em>and<em> M. laxa</em> are established in the USA, while <em>M. fructigena </em>and<em> M. polystroma</em> are pathogens of regulatory significance. We have completed draft genomes of all 4 <em>Monilinia</em> species using Illumina Next Generation Sequencing (NGS) platform. Prior to our work, limited genomic information of <em>M. fructigena, M. laxa </em>and <em>M. polystroma</em> were available to use in diagnostics development. These draft genome assemblies are between 39-45 Mb and consistent with the genome sizes of related fungal species. We have developed comparative genomics pipelines that are allowing us to screen this new wealth of NGS data for candidate genomic targets that can then be tested on a broader set of strains from these 4 <em>Monilinia</em> species. Our goal for the current phase of this project is use these comparative genomics pipelines to identify candidate targets, to design PCR primers targeting these, and to evaluate and validate species-specific molecular markers for the detection and differentiation of <em>Monilinia</em> species of concern (<em>M. fructigena</em> and <em>M. polystroma</em>) from other <em>Monilinia</em> species already established within the US. We will discuss these pipelines and progress toward testing, optimizing and validating molecular methods that can distinguish among these species of regulatory concern.</div>