Optimized bio-fertilization drives the synergistic recovery of vegetation productivity and soil multifunctionality by reshaping fungal interaction patterns in degraded alpine mines

Optimized bio-fertilization drives the synergistic recovery of vegetation productivity and soil multifunctionality by reshaping fungal interaction patterns in degraded alpine mines | 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 Optimized bio-fertilization drives the synergistic recovery of vegetation productivity and soil multifunctionality by reshaping fungal interaction patterns in degraded alpine mines Zongcheng Cai, Shouquan Fu, Fayi Li, Yunfei Xing, Jun Ma, Enqi Yang, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8837993/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 02 May, 2026 Read the published version in BMC Microbiology → Version 1 posted 10 You are reading this latest preprint version Abstract Restoring degraded alpine mining ecosystems is critically constrained by soil infertility and the functional decoupling of plant–soil–microbe interactions. While optimized bio-fertilization represents a promising restoration strategy, the deterministic mechanisms linking vegetation recovery, fungal community assembly, and soil multifunctionality (SMF) remain poorly understood. This study investigated the effects of a three-year restoration experiment at the Muli coal mine on the Qinghai–Tibet Plateau, evaluating the integrated response of plant communities and soil functional networks to various fertilization regimes. Our results demonstrated that the optimized regime (W3J1, comprising 375 kg·hm⁻² of forage-specific fertilizer and 350 kg·hm⁻² of microbial inoculant) elicited the most robust ecological recovery, increasing plant height to 35.87 cm and aboveground biomass to 352.67 g·m⁻², which represents a significant increase of 80.23% compared to the control (195.67 g·m⁻²). Concurrently, the SMF index in W3J1 reached a peak of 0.95, effectively reversing the functional impairment observed in the degraded control (SMF = -2.32). Regarding fungal α-diversity, the W3J1 treatment significantly enhanced community complexity, recording a peak Shannon index of 3.50 and Pielou’s evenness of 0.55, compared to 3.01 and 0.48 in the control, respectively. Furthermore, the W1J3 treatment achieved the highest observed OTU richness (693) and Chao1 index (822), representing significant increases of approximately 33%-37% relative to the control. Notably, excessive nutrient inputs in the W3J3 treatment failed to yield additional benefits, as SMF and fungal diversity indices stabilized or declined due to resource imbalances. Random Forest modeling identified vegetation density as the paramount predictor of fungal diversity, significantly outperforming soil physicochemical variables.Structural Equation Modeling (SEM) further elucidated a significant cascading pathway (R² = 0.67) where fertilization directly promoted fungal diversity ( β = 0.76) and initiated vegetation establishment, which subsequently facilitated the deterministic restructuring of the fungal community. In conclusion, the optimized co-application of microbial agents and fertilizers facilitates ecosystem reconstruction by orchestrating the coupling between vegetation density and fungal stability, providing a theoretical basis for the sustainable restoration of severely degraded alpine mines. Muli coal mine on the Qinghai-Tibet Plateau soil multifunctionality composite microbial inoculant fungal co-occurrence network vegetation-soil-microbe coupling alpine ecosystem restoration Full Text Additional Declarations No competing interests reported. Supplementary Files Graphicalabstract.jpg SupplementaryInformation.docx Cite Share Download PDF Status: Published Journal Publication published 02 May, 2026 Read the published version in BMC Microbiology → Version 1 posted Editorial decision: Revision requested 12 Mar, 2026 Reviews received at journal 12 Mar, 2026 Reviews received at journal 11 Mar, 2026 Reviewers agreed at journal 21 Feb, 2026 Reviewers agreed at journal 18 Feb, 2026 Reviewers invited by journal 18 Feb, 2026 Editor assigned by journal 18 Feb, 2026 Editor invited by journal 16 Feb, 2026 Submission checks completed at journal 14 Feb, 2026 First submitted to journal 14 Feb, 2026 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. 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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-8837993","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":593830988,"identity":"ef0e14c2-0360-489a-bbdd-a896809a8ac6","order_by":0,"name":"Zongcheng Cai","email":"","orcid":"","institution":"Qinghai University","correspondingAuthor":false,"prefix":"","firstName":"Zongcheng","middleName":"","lastName":"Cai","suffix":""},{"id":593830989,"identity":"1083cddc-1d6f-4a20-82d7-ff443f9dfafd","order_by":1,"name":"Shouquan Fu","email":"","orcid":"","institution":"Qinghai 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mines","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"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":"bmc-microbiology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mcro","sideBox":"Learn more about [BMC Microbiology](http://bmcmicrobiol.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/mcro","title":"BMC Microbiology","twitterHandle":"#bmcmicrobiology","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Muli coal mine on the Qinghai-Tibet Plateau, soil multifunctionality, composite microbial inoculant, fungal co-occurrence network, vegetation-soil-microbe coupling, alpine ecosystem restoration","lastPublishedDoi":"10.21203/rs.3.rs-8837993/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8837993/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eRestoring degraded alpine mining ecosystems is critically constrained by soil infertility and the functional decoupling of plant\u0026ndash;soil\u0026ndash;microbe interactions. While optimized bio-fertilization represents a promising restoration strategy, the deterministic mechanisms linking vegetation recovery, fungal community assembly, and soil multifunctionality (SMF) remain poorly understood. This study investigated the effects of a three-year restoration experiment at the Muli coal mine on the Qinghai\u0026ndash;Tibet Plateau, evaluating the integrated response of plant communities and soil functional networks to various fertilization regimes. Our results demonstrated that the optimized regime (W3J1, comprising 375 kg\u0026middot;hm⁻\u0026sup2; of forage-specific fertilizer and 350 kg\u0026middot;hm⁻\u0026sup2; of microbial inoculant) elicited the most robust ecological recovery, increasing plant height to 35.87 cm and aboveground biomass to 352.67 g\u0026middot;m⁻\u0026sup2;, which represents a significant increase of 80.23% compared to the control (195.67 g\u0026middot;m⁻\u0026sup2;). Concurrently, the SMF index in W3J1 reached a peak of 0.95, effectively reversing the functional impairment observed in the degraded control (SMF = -2.32). Regarding fungal α-diversity, the W3J1 treatment significantly enhanced community complexity, recording a peak Shannon index of 3.50 and Pielou\u0026rsquo;s evenness of 0.55, compared to 3.01 and 0.48 in the control, respectively. Furthermore, the W1J3 treatment achieved the highest observed OTU richness (693) and Chao1 index (822), representing significant increases of approximately 33%-37% relative to the control. Notably, excessive nutrient inputs in the W3J3 treatment failed to yield additional benefits, as SMF and fungal diversity indices stabilized or declined due to resource imbalances. Random Forest modeling identified vegetation density as the paramount predictor of fungal diversity, significantly outperforming soil physicochemical variables.Structural Equation Modeling (SEM) further elucidated a significant cascading pathway (R\u0026sup2; = 0.67) where fertilization directly promoted fungal diversity (\u003cem\u003eβ\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.76) and initiated vegetation establishment, which subsequently facilitated the deterministic restructuring of the fungal community. In conclusion, the optimized co-application of microbial agents and fertilizers facilitates ecosystem reconstruction by orchestrating the coupling between vegetation density and fungal stability, providing a theoretical basis for the sustainable restoration of severely degraded alpine mines.\u003c/p\u003e","manuscriptTitle":"Optimized bio-fertilization drives the synergistic recovery of vegetation productivity and soil multifunctionality by reshaping fungal interaction patterns in degraded alpine mines","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-02-20 15:54:49","doi":"10.21203/rs.3.rs-8837993/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-03-12T20:58:03+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-12T08:17:25+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-11T16:07:57+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"232320687788665945023698661568571659390","date":"2026-02-21T09:27:29+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"56469097880017648489874556033772046892","date":"2026-02-19T04:35:07+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-02-18T16:25:04+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-02-18T16:21:23+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2026-02-16T09:44:42+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-02-14T07:25:14+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Microbiology","date":"2026-02-14T07:20:17+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-microbiology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mcro","sideBox":"Learn more about [BMC Microbiology](http://bmcmicrobiol.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/mcro","title":"BMC Microbiology","twitterHandle":"#bmcmicrobiology","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"a55b0f1f-18d1-447e-9392-2c40b5549094","owner":[],"postedDate":"February 20th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-05-05T10:02:35+00:00","versionOfRecord":{"articleIdentity":"rs-8837993","link":"https://doi.org/10.1186/s12866-026-05054-z","journal":{"identity":"bmc-microbiology","isVorOnly":false,"title":"BMC Microbiology"},"publishedOn":"2026-05-02 15:58:25","publishedOnDateReadable":"May 2nd, 2026"},"versionCreatedAt":"2026-02-20 15:54:49","video":"","vorDoi":"10.1186/s12866-026-05054-z","vorDoiUrl":"https://doi.org/10.1186/s12866-026-05054-z","workflowStages":[]},"version":"v1","identity":"rs-8837993","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8837993","identity":"rs-8837993","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}