Phase-contrast microtomography unveils mechanisms of root colonization by a vascular fungal pathogen

Phase-contrast microtomography unveils mechanisms of root colonization by a vascular fungal pathogen | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Biological Sciences - Article Phase-contrast microtomography unveils mechanisms of root colonization by a vascular fungal pathogen David Turrà, Valentino Maria Guastaferro, Stefania Vitale, Lorenzo D'Amico, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6567296/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted You are reading this latest preprint version Abstract Soil-borne vascular pathogens pose serious threats to agriculture with complex invasion strategies that remain poorly characterized compared to foliar pathogens 1 . While foliar pathogens like Magnaporthe oryzae employ specialized appressoria to penetrate plant surfaces through a combination of mechanical force and enzymatic degradation 2 , the invasion mechanisms of vascular pathogens that lack classical appressoria have remained largely theoretical. The nanoscale processes governing root penetration and colonization by these pathogens are particularly challenging to visualize due to technical limitations of conventional microscopy 3-5 . Here we show, using phase-contrast X-ray computed microtomography and advanced microscopy, that Fusarium oxysporum ( Fo ) employs distinct mitogen-activated protein kinase (MAPK) cascades to orchestrate root invasion through unprecedented morphological plasticity. We discovered previously undocumented appressoria-like structures that facilitate physical penetration, while demonstrating that Fo exhibits remarkable cellular adaptability, reducing hyphal diameter by more than 20-fold (from 5 μm to 220 nm) to navigate confined plant spaces 6-7 , a dramatic morphological transition previously thought impossible. By using cellulase-deficient mutants 8 , we demonstrate that cellulolytic activity is dispensable for surface breach and submicrometric hyphal colonization, establishing that mechanical force generation rather than enzymatic degradation is the primary determinant of successful host penetration 9-10 . Three-dimensional reconstruction reveals a quantitative correlation between fungal proliferation and progressive embolism formation, with distinct MAPK pathways differentially regulating penetration force generation (Fmk1), osmotic adaptation during apoplastic colonization (Hog1), and directional growth toward vascular tissues (Mpk1). These findings provide a mechanistic framework for vascular wilt pathogenesis and reveal potential targets for controlling these economically devastating plant diseases. Biological sciences/Microbiology/Pathogens Biological sciences/Developmental biology/Differentiation Earth and environmental sciences/Ecology/Microbial ecology Full Text Additional Declarations There is NO Competing Interest. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6567296","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Biological Sciences - Article","associatedPublications":[],"authors":[{"id":450609846,"identity":"abaa82bd-d6a8-4d02-ab10-4abf96236d85","order_by":0,"name":"David 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pathogen","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"nature-portfolio","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"","title":"Nature Portfolio","twitterHandle":"","acdcEnabled":false,"dfaEnabled":false,"editorialSystem":"ejp","reportingPortfolio":"","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-6567296/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6567296/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Soil-borne vascular pathogens pose serious threats to agriculture with complex invasion strategies that remain poorly characterized compared to foliar pathogens\u003csup\u003e1\u003c/sup\u003e. While foliar pathogens like \u003ci\u003eMagnaporthe oryzae\u003c/i\u003e employ specialized appressoria to penetrate plant surfaces through a combination of mechanical force and enzymatic degradation\u003csup\u003e2\u003c/sup\u003e, the invasion mechanisms of vascular pathogens that lack classical appressoria have remained largely theoretical. The nanoscale processes governing root penetration and colonization by these pathogens are particularly challenging to visualize due to technical limitations of conventional microscopy\u003csup\u003e3-5\u003c/sup\u003e. Here we show, using phase-contrast X-ray computed microtomography and advanced microscopy, that \u003ci\u003eFusarium oxysporum\u003c/i\u003e (\u003ci\u003eFo\u003c/i\u003e) employs distinct mitogen-activated protein kinase (MAPK) cascades to orchestrate root invasion through unprecedented morphological plasticity. We discovered previously undocumented appressoria-like structures that facilitate physical penetration, while demonstrating that \u003ci\u003eFo\u003c/i\u003e exhibits remarkable cellular adaptability, reducing hyphal diameter by more than 20-fold (from 5 μm to 220 nm) to navigate confined plant spaces\u003csup\u003e6-7\u003c/sup\u003e, a dramatic morphological transition previously thought impossible. By using cellulase-deficient mutants\u003csup\u003e8\u003c/sup\u003e, we demonstrate that cellulolytic activity is dispensable for surface breach and submicrometric hyphal colonization, establishing that mechanical force generation rather than enzymatic degradation is the primary determinant of successful host penetration\u003csup\u003e9-10\u003c/sup\u003e. Three-dimensional reconstruction reveals a quantitative correlation between fungal proliferation and progressive embolism formation, with distinct MAPK pathways differentially regulating penetration force generation (Fmk1), osmotic adaptation during apoplastic colonization (Hog1), and directional growth toward vascular tissues (Mpk1). These findings provide a mechanistic framework for vascular wilt pathogenesis and reveal potential targets for controlling these economically devastating plant diseases.","manuscriptTitle":"Phase-contrast microtomography unveils mechanisms of root colonization by a vascular fungal pathogen","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-05-12 03:21:20","doi":"10.21203/rs.3.rs-6567296/v1","editorialEvents":[],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"nature-communications","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"NCOMMS","sideBox":"Learn more about [Nature Communications](http://www.nature.com/ncomms/)","snPcode":"","submissionUrl":"https://mts-ncomms.nature.com/","title":"Nature Communications","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature Communications","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"f04c406f-a5b9-45fa-8ba3-1dea40123786","owner":[],"postedDate":"May 12th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[{"id":47945308,"name":"Biological sciences/Microbiology/Pathogens"},{"id":47945309,"name":"Biological sciences/Developmental biology/Differentiation"},{"id":47945310,"name":"Earth and environmental sciences/Ecology/Microbial ecology"}],"tags":[],"updatedAt":"2025-06-19T08:36:52+00:00","versionOfRecord":[],"versionCreatedAt":"2025-05-12 03:21:20","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6567296","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6567296","identity":"rs-6567296","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}