Metagenomic Sequencing for Identification of Xylella fastidiosa from Leaf Samples

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dc.contributor.authorRoman-Reyna, Veronicaen
dc.contributor.authorDupas, Enoraen
dc.contributor.authorCesbron, Sophieen
dc.contributor.authorMarchi, Guidoen
dc.contributor.authorCampigli, Saraen
dc.contributor.authorHansen, Mary Annen
dc.contributor.authorBush, Elizabethen
dc.contributor.authorPrarat, Melanieen
dc.contributor.authorShiplett, Katherineen
dc.contributor.authorIvey, Melanie L. Lewisen
dc.contributor.authorPierzynski, Joyen
dc.contributor.authorMiller, Sally A.en
dc.contributor.authorHand, Francesca Pedutoen
dc.contributor.authorJacques, Marie-Agnesen
dc.contributor.authorJacobs, Jonathan M.en
dc.date.accessioned2022-03-30T19:01:40Zen
dc.date.available2022-03-30T19:01:40Zen
dc.date.issued2021-09en
dc.description.abstractXylella fastidiosa (Xf) is a globally distributed plant-pathogenic bacterium. The primary control strategy for Xf diseases is eradicating infected plants; therefore, timely and accurate detection is necessary to prevent crop losses and further pathogen dispersal. Conventional Xf diagnostics primarily relies on quantitative PCR (qPCR) assays. However, these methods do not consider new or emerging variants due to pathogen genetic recombination and sensitivity limitations. We developed and tested a metage-nomics pipeline using in-house short-read sequencing as a complementary approach for affordable, fast, and highly accurate Xf detection. We used metagenomics to identify Xf to the strain level in single-and mixed-infected plant samples at concentrations as low as 1 pg of bacterial DNA per gram of tissue. We also tested naturally infected samples from various plant species originating from Europe and the United States. We identified Xf sub-species in samples previously considered inconclusive with real-time PCR (quantification cycle [C-q], > 35). Overall, we showed the versatility of the pipeline by using different plant hosts and DNA extraction methods. Our pipeline provides taxonomic and functional infor-mation for Xf diagnostics without extensive knowledge of the disease. This pipeline demonstrates that metagenomics can be used for early detection of Xf and incorporated as a tool to inform disease management strategies. IMPORTANCE Destructive Xylella fastidiosa (Xf) outbreaks in Europe highlight this pathogen's capacity to expand its host range and geographical distribution. The current disease diag-nostic approaches are limited by a multiple-step process, biases to known sequences, and detection limits. We developed a low-cost, user-friendly metagenomic sequencing tool for Xf detection. In less than 3 days, we were able to identify Xf subspecies and strains in field-collected samples. Overall, our pipeline is a diagnostics tool that could be easily extended to other plant-pathogen interactions and implemented for emerging plant threat surveillance.en
dc.description.notesWe are grateful for funding support from the Ohio Department of Agriculture Specialty Crops block grant (AGR-SCG-19-03) and USDA NIFA FACT (2021-67021-34343) to J.M.J. Sara Campigli was financed by a grant from the Phytosanitary Service of the Tuscany Region (Italy).en
dc.description.sponsorshipOhio Department of Agriculture Specialty Crops block grant [AGR-SCG-19-03]; USDA NIFA FACT [2021-67021-34343]; Phytosanitary Service of the Tuscany Region (Italy)en
dc.description.versionPublished versionen
dc.format.mimetypeapplication/pdfen
dc.identifier.doihttps://doi.org/10.1128/mSystems.00591-21en
dc.identifier.issn2379-5077en
dc.identifier.issue5en
dc.identifier.othere00591-21en
dc.identifier.pmid34698548en
dc.identifier.urihttp://hdl.handle.net/10919/109507en
dc.identifier.volume6en
dc.language.isoenen
dc.rightsCreative Commons Attribution 4.0 Internationalen
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/en
dc.subjectXylella fastidiosaen
dc.subjectmetagenomicsen
dc.subjectdiagnosticsen
dc.subjectshort-read sequencingen
dc.titleMetagenomic Sequencing for Identification of Xylella fastidiosa from Leaf Samplesen
dc.title.serialmSystemsen
dc.typeArticle - Refereeden
dc.type.dcmitypeTexten

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