Altered Gut Microbiota and Metabolites in Children with Non-organic Anorexia: A Multi-omics Integration Study

Altered Gut Microbiota and Metabolites in Children with Non-organic Anorexia: A Multi-omics Integration Study | 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 Article Altered Gut Microbiota and Metabolites in Children with Non-organic Anorexia: A Multi-omics Integration Study Zonglong Li, Qiong Zhang, Jin Yang, Rui Lei, Wei Lu This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8679173/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 14 You are reading this latest preprint version Abstract Objective While alterations in gut microbiota have been implicated in childhood eating disorders, a comprehensive understanding of the functional and metabolic consequences in non-organic anorexia (NOA) remains limited. This study aimed to employ an integrated multi-omics approach to systematically characterize the gut microbial composition, functional potential, and associated metabolic profiles in children with NOA compared to healthy controls. Methods A case-control study was conducted involving 88 children aged 1–5 years (48 NOA, 40 healthy controls). Gut microbiota composition was assessed via 16S rRNA gene sequencing of all fecal samples. Subsequently, the five most representative samples from each group were selected for deep shotgun metagenomic sequencing and liquid chromatography-mass spectrometry (LC-MS) based non-targeted metabolomics. Results 16S rRNA sequencing revealed significantly higher microbial richness and diversity (Chao1 and Shannon indices, P < 0.001) in the NOA group. Taxonomic analysis showed a distinct structural shift: the NOA group exhibited increased relative abundances of the phyla Firmicutes and Bacteroidota, and the genera Bacteroides, Faecalibacterium, Subdoligranulum, and Roseburia, while Actinobacteriota, Bifidobacterium, and Enterococcus were decreased. Metagenomic analysis indicated significant alterations in key metabolic pathways, including a notable downregulation of riboflavin (Vitamin B2) metabolism ( P < 0.05) and upregulation of pathways related to fat digestion and absorption in the NOA group. Metabolomic profiling identified 33 significantly altered fecal metabolites. Specifically, levels of L-carnitine derivatives were lower, whereas tyramine glucuronide was higher in the NOA group and was enriched in bile secretion-related pathways. Spearman correlation analysis demonstrated significant associations between these differential metabolites and the altered bacterial genera. Conclusion Our integrated multi-omics analysis demonstrates that NOA in children is associated with a specific gut ecosystem characterized by altered microbiota structure, perturbed microbial metabolic functions (particularly riboflavin metabolism), and corresponding host-microbiota co-metabolic disturbances. These findings provide novel evidence for the disrupted "microbiota-metabolite" axis in NOA, offering new mechanistic insights and potential targets for intervention. Health sciences/Gastroenterology Biological sciences/Microbiology Non-organic anorexia Children Gut microbiota Metagenomics Metabolomics Riboflavin metabolism Full Text Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 14 Apr, 2026 Reviewers agreed at journal 13 Apr, 2026 Reviews received at journal 12 Apr, 2026 Reviews received at journal 11 Apr, 2026 Reviewers agreed at journal 11 Apr, 2026 Reviews received at journal 25 Mar, 2026 Reviewers agreed at journal 25 Mar, 2026 Reviewers agreed at journal 19 Mar, 2026 Reviewers agreed at journal 19 Mar, 2026 Reviewers invited by journal 19 Mar, 2026 Editor assigned by journal 19 Mar, 2026 Editor invited by journal 09 Mar, 2026 Submission checks completed at journal 03 Mar, 2026 First submitted to journal 03 Mar, 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. 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-8679173","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":608979382,"identity":"ea1688ad-f672-4003-8ef2-17754b8162a6","order_by":0,"name":"Zonglong Li","email":"","orcid":"","institution":"Guizhou Branch of Shanghai Children’s Medical Center，Shanghai Jiao Tong University School of Medicine","correspondingAuthor":false,"prefix":"","firstName":"Zonglong","middleName":"","lastName":"Li","suffix":""},{"id":608979383,"identity":"2155bcb6-a179-40a1-a78e-7959f7c4286a","order_by":1,"name":"Qiong Zhang","email":"","orcid":"","institution":"The First People’s Hospital of Qingzhen City","correspondingAuthor":false,"prefix":"","firstName":"Qiong","middleName":"","lastName":"Zhang","suffix":""},{"id":608979384,"identity":"3d1acfa8-a7af-4884-8c8a-06fe7b649a6f","order_by":2,"name":"Jin Yang","email":"","orcid":"","institution":"Department of Pediatrics, Affiliated Hospital of Zunyi Medical University/Guizhou Children's Hospital","correspondingAuthor":false,"prefix":"","firstName":"Jin","middleName":"","lastName":"Yang","suffix":""},{"id":608979385,"identity":"1add9478-9da9-488c-ae87-08ad1efc955a","order_by":3,"name":"Rui Lei","email":"","orcid":"","institution":"Department of Pediatrics, Affiliated Hospital of Zunyi Medical University/Guizhou Children's Hospital","correspondingAuthor":false,"prefix":"","firstName":"Rui","middleName":"","lastName":"Lei","suffix":""},{"id":608979386,"identity":"2f59497c-dbee-447f-9a61-574880566362","order_by":4,"name":"Wei Lu","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA5klEQVRIiWNgGAWjYBACNvbG9p8fKmrq29gbGx8kVNQQ1sLHc/iAtMSZY4z9PIcPGzw4c4ywFjmJtAQJ3jZmxpkz3NIkH7YwE+EwnjMGBpJtbMwGN3jMKhIb2Bj427sTCPilxyCh4JwMm8HtHrMbiTtkGCTOnN1A0JYDEmVsPAZ3zgC1nGFjMJDIJaBFIsewgYeNWcLgRo5ZQWIbMzFa0pIZeNqYDSRnpKUxEKeF5/AxZmAgJ/ADA1ki4cwxHoJ+kW9vbGMERmUCME4bP/6oqJHjb+/FrwUD8JCmfBSMglEwCkYBVgAAfphLCN/AUxgAAAAASUVORK5CYII=","orcid":"","institution":"Department of Pediatrics, Affiliated Hospital of Zunyi Medical University/Guizhou Children's Hospital","correspondingAuthor":true,"prefix":"","firstName":"Wei","middleName":"","lastName":"Lu","suffix":""}],"badges":[],"createdAt":"2026-01-23 12:24:46","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8679173/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8679173/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":105327009,"identity":"facca9f6-238b-4fa0-89ae-0d40bc08dbae","added_by":"auto","created_at":"2026-03-24 19:20:58","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":952393,"visible":true,"origin":"","legend":"","description":"","filename":"Article.docx2026.01.28.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8679173/v1_covered_3e88f582-c8a9-4b4d-801c-837b0e56672d.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Altered Gut Microbiota and Metabolites in Children with Non-organic Anorexia: A Multi-omics Integration Study","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":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Non-organic anorexia, Children, Gut microbiota, Metagenomics, Metabolomics, Riboflavin metabolism","lastPublishedDoi":"10.21203/rs.3.rs-8679173/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8679173/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eObjective\u003c/h2\u003e \u003cp\u003eWhile alterations in gut microbiota have been implicated in childhood eating disorders, a comprehensive understanding of the functional and metabolic consequences in non-organic anorexia (NOA) remains limited. This study aimed to employ an integrated multi-omics approach to systematically characterize the gut microbial composition, functional potential, and associated metabolic profiles in children with NOA compared to healthy controls.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eA case-control study was conducted involving 88 children aged 1\u0026ndash;5 years (48 NOA, 40 healthy controls). Gut microbiota composition was assessed via 16S rRNA gene sequencing of all fecal samples. Subsequently, the five most representative samples from each group were selected for deep shotgun metagenomic sequencing and liquid chromatography-mass spectrometry (LC-MS) based non-targeted metabolomics.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003e16S rRNA sequencing revealed significantly higher microbial richness and diversity (Chao1 and Shannon indices, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.001) in the NOA group. Taxonomic analysis showed a distinct structural shift: the NOA group exhibited increased relative abundances of the phyla Firmicutes and Bacteroidota, and the genera Bacteroides, Faecalibacterium, Subdoligranulum, and Roseburia, while Actinobacteriota, Bifidobacterium, and Enterococcus were decreased. Metagenomic analysis indicated significant alterations in key metabolic pathways, including a notable downregulation of riboflavin (Vitamin B2) metabolism (\u003cem\u003eP\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) and upregulation of pathways related to fat digestion and absorption in the NOA group. Metabolomic profiling identified 33 significantly altered fecal metabolites. Specifically, levels of L-carnitine derivatives were lower, whereas tyramine glucuronide was higher in the NOA group and was enriched in bile secretion-related pathways. Spearman correlation analysis demonstrated significant associations between these differential metabolites and the altered bacterial genera.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eOur integrated multi-omics analysis demonstrates that NOA in children is associated with a specific gut ecosystem characterized by altered microbiota structure, perturbed microbial metabolic functions (particularly riboflavin metabolism), and corresponding host-microbiota co-metabolic disturbances. These findings provide novel evidence for the disrupted \"microbiota-metabolite\" axis in NOA, offering new mechanistic insights and potential targets for intervention.\u003c/p\u003e","manuscriptTitle":"Altered Gut Microbiota and Metabolites in Children with Non-organic Anorexia: A Multi-omics Integration Study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-03-24 19:20:38","doi":"10.21203/rs.3.rs-8679173/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-04-14T08:41:53+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"98525328604089820902271570623049564484","date":"2026-04-13T07:03:11+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-04-13T00:55:09+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-04-11T09:46:30+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"69099962107542112374306783505036624588","date":"2026-04-11T08:04:02+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-25T20:57:05+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"25424873320868781633858220202435965666","date":"2026-03-25T18:50:04+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"278868561277960352451569105394393575003","date":"2026-03-19T14:59:14+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"304613560693239838122225917892221884024","date":"2026-03-19T13:06:24+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-03-19T08:15:50+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-03-19T08:12:09+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2026-03-09T12:02:34+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-03-03T05:37:04+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2026-03-03T05:27:34+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"2c5432df-97de-40e5-a40e-00dec4d7b387","owner":[],"postedDate":"March 24th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[{"id":64801118,"name":"Health sciences/Gastroenterology"},{"id":64801119,"name":"Biological sciences/Microbiology"}],"tags":[],"updatedAt":"2026-05-04T05:24:30+00:00","versionOfRecord":[],"versionCreatedAt":"2026-03-24 19:20:38","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8679173","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8679173","identity":"rs-8679173","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}