Impact of multi-species mixtures and edaphic context on soil functioning in sown grasslands: a mesocosm experiment

Impact of multi-species mixtures and edaphic context on soil functioning in sown grasslands: a mesocosm experiment | 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 Research Article Impact of multi-species mixtures and edaphic context on soil functioning in sown grasslands: a mesocosm experiment Robin Tersago, Jeroen Meersmans, Camille Van Eupen, Jan Willem van Groenigen, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8965719/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background and aims Multi-species grassland mixtures are gaining popularity in managed grasslands to maintain yields under a changing climate, yet their effect on belowground functioning of the soil remains poorly quantified. This study aimed to determine the combined effects of plant community, edaphic context and management intensity on productivity, greenhouse gas (GHG) effluxes and soil microbial functioning. Methods We conducted a full-factorial mesocosm experiment comparing a perennial ryegrass monoculture with a multi-species mixture across two edaphic contexts and three management intensities. Above- and belowground productivity, GHG effluxes (CO 2 , CH 4 and N 2 O) and functional catabolic diversity of soil microbes were analysed using ANOVA, linear mixed models and random forests with SHAP values. Results Multi-species grass mixtures consistently increased yield and belowground carbon inputs compared to monocultures and affected all three GHG effluxes. Both plant community and edaphic context significantly shaped functional microbial diversity, with plant community explaining comparable amounts of variance as the edaphic context. SHAP values further identified threshold values for the effects of soil temperature and moisture on GHG effluxes, going beyond the effects captured by the linear mixed models. Conclusion Our results show that sowing a multi-species grass mixture can consistently enhance belowground soil functioning and can promote the delivery of crucial ecosystem services such as climate regulation, biomass production and nutrient cycling. These findings underline the potential of multi-species grassland mixtures to improve productivity and soil processes future multifunctional grassland systems. Multi-species grassland mixtures greenhouse gas effluxes mesocosms context-dependency Biolog EcoPlates Full Text Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-8965719","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":600499572,"identity":"a3846036-354e-44ba-bd00-9d66dd1a4295","order_by":0,"name":"Robin Tersago","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABDUlEQVRIiWNgGAWjYFACHoYDCE5FAgMflJlAQIsBlHMmgYGNGC0McC2MbURoMW/vPXi4gOGPnHn/4ccvPs5LS2xj7zGT+LiDIY8fhxaZM+cSDs9gMDCWuZFmZjlzW05iG88ZM8mZZxiKJRuwa5GQyDE4zMNgkDhDgsHMmHdbRWKbRI6xMW8bQ+KGA/i11M/gP/7NmHcOUIv8G2Pjv0At+wloSZBgyDF+zNsAdJgEj+FjRpAtOPwiwQP0C4+BseEMiZwyxhnH0ozbeNIKH/a2SRRL4LKFvffwZ54KOXkJ/uObP3yoSZbtZz+84cDPNps8fhzehwBItLBJIJuFTz0cMH8gStkoGAWjYBSMOAAA/fdV8TokBAUAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0003-3832-5823","institution":"KU Leuven: Katholieke Universiteit Leuven","correspondingAuthor":true,"prefix":"","firstName":"Robin","middleName":"","lastName":"Tersago","suffix":""},{"id":600499573,"identity":"5ebc833c-30be-4b53-8982-4c87aad70234","order_by":1,"name":"Jeroen Meersmans","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Jeroen","middleName":"","lastName":"Meersmans","suffix":""},{"id":600499574,"identity":"2a0b8a26-737d-4de7-8180-5aadcd3378bd","order_by":2,"name":"Camille Van Eupen","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Camille","middleName":"Van","lastName":"Eupen","suffix":""},{"id":600499575,"identity":"c61be025-6819-4571-9e2e-c049b71a39dc","order_by":3,"name":"Jan Willem van Groenigen","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Jan","middleName":"Willem van","lastName":"Groenigen","suffix":""},{"id":600499576,"identity":"e703fdf3-42d7-4e6d-b62b-a42627b5155c","order_by":4,"name":"Ben Aernouts","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Ben","middleName":"","lastName":"Aernouts","suffix":""},{"id":600499577,"identity":"5f75f6a7-f063-41f7-8484-423a03a2f1b5","order_by":5,"name":"Ellen Desie","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Ellen","middleName":"","lastName":"Desie","suffix":""},{"id":600499578,"identity":"36d99d84-47b8-440a-9b66-c4d04f9c52d5","order_by":6,"name":"Karen Vancampenhout","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Karen","middleName":"","lastName":"Vancampenhout","suffix":""}],"badges":[],"createdAt":"2026-02-25 09:22:06","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8965719/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8965719/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":107704402,"identity":"ed5088a9-bf57-474e-812c-57c9b76d32d8","added_by":"auto","created_at":"2026-04-24 08:45:13","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1845045,"visible":true,"origin":"","legend":"","description":"","filename":"ManuscriptPlantandSoilTersagoetal.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8965719/v1_covered_81dd24d3-51d9-4a37-b753-c3c7db05250e.pdf"}],"financialInterests":"","formattedTitle":"Impact of multi-species mixtures and edaphic context on soil functioning in sown grasslands: a mesocosm experiment","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Multi-species grassland mixtures, greenhouse gas effluxes, mesocosms, context-dependency, Biolog EcoPlates","lastPublishedDoi":"10.21203/rs.3.rs-8965719/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8965719/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground and aims\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMulti-species grassland mixtures are gaining popularity in managed grasslands to maintain yields under a changing climate, yet their effect on belowground functioning of the soil remains poorly quantified. This study aimed to determine the combined effects of plant community, edaphic context and management intensity on productivity, greenhouse gas (GHG) effluxes and soil microbial functioning.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe conducted a full-factorial mesocosm experiment comparing a perennial ryegrass monoculture with a multi-species mixture across two edaphic contexts and three management intensities. Above- and belowground productivity, GHG effluxes (CO\u003csub\u003e2\u003c/sub\u003e, CH\u003csub\u003e4\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO) and functional catabolic diversity of soil microbes were analysed using ANOVA, linear mixed models and random forests with SHAP values.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMulti-species grass mixtures consistently increased yield and belowground carbon inputs compared to monocultures and affected all three GHG effluxes. Both plant community and edaphic context significantly shaped functional microbial diversity, with plant community explaining comparable amounts of variance as the edaphic context. SHAP values further identified threshold values for the effects of soil temperature and moisture on GHG effluxes, going beyond the effects captured by the linear mixed models.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOur results show that sowing a multi-species grass mixture can consistently enhance belowground soil functioning and can promote the delivery of crucial ecosystem services such as climate regulation, biomass production and nutrient cycling. These findings underline the potential of multi-species grassland mixtures to improve productivity and soil processes future multifunctional grassland systems.\u003c/p\u003e","manuscriptTitle":"Impact of multi-species mixtures and edaphic context on soil functioning in sown grasslands: a mesocosm experiment","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-03-09 20:41:20","doi":"10.21203/rs.3.rs-8965719/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"0ca8dab6-bcf6-4f5b-b31d-1c44a508740a","owner":[],"postedDate":"March 9th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-04-21T15:43:43+00:00","versionOfRecord":[],"versionCreatedAt":"2026-03-09 20:41:20","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8965719","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8965719","identity":"rs-8965719","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}