Metagenomic Sequencing of Tomato Plants with Wilt Symptoms Allows for Strain-Level Pathogen Identification and...

Diseases that affect the vascular system or the pith are of great economic impact because they can rapidly destroy the affected plants, leading to complete loss in production. Fast and precise identification is thus important to inform containment and management, but many identification methods are slow because they are culture-dependent and do not reach strain resolution. Here we used culture-independent long-read metagenomic sequencing of DNA extracted directly from the stems of two tomato samples that displayed wilt symptoms. We obtained enough sequencing reads to assemble high-quality metagenome-assembled genomes of Ralstonia solanacearum from one sample and of Pseudomonas corrugata from the other. The genome sequences allowed us to identify both pathogens to the strain level using the genomeRxiv platform, perform phylogenetic analyses, predict virulence genes, and infer antibiotic and copper resistance. In the case of R. solanacearum, it was straightforward to exclude the pathogen from being the Select Agent race 3 biovar 2. Using the Branchwater tool, it was also possible to determine the worldwide distribution of both pathogen strains based on public metagenomic sequences. The entire analysis could have been completed within 2 days, starting with sample acquisition. Steps necessary toward establishing metagenomic sequencing as a more routine approach in plant diseases clinics are discussed.