Associations of gut microbial and serum metabolic features with metabolic comorbidities in patients with NAFLD

Introduction Non-alcoholic fatty liver disease (NAFLD) coexists with dyslipidemia or impaired fasting glucose (IFG), but gut microbial and serum metabolic features remain incompletely characterized. Objectives We aimed to compare gut microbial beta diversity, taxonomic profiles, predicted microbial functions, and serum metabolites across healthy controls (HCs) and NAFLD subgroups; evaluate omics-based classifiers and exploratory integrated classifiers distinguishing subgroups from HCs; and assess metabolic responses to Dialister invisus supplementation in mice. Methods Fecal microbiota and serum metabolites were profiled by 16S rRNA gene sequencing and targeted UPLC–MS/MS, respectively, in 41 HCs and 222 NAFLD patients. Patients were stratified into three subgroups by dyslipidemia and IFG: NAFLD_N (neither dyslipidemia nor IFG), NAFLD_DLP (dyslipidemia without IFG), and NAFLD_IFG (IFG irrespective of lipid status; HbA1c/OGTT unavailable). Random-forest classifiers were developed for each omics layer and for post hoc integrated multi-omics panels to distinguish each NAFLD subgroup from HCs, with performance evaluated in the held-out set. Dialister invisus was administered to high-calorie diet-fed mice, and metabolic and hepatic outcomes were assessed. Results NAFLD patients showed lower Dialister invisus abundance and higher abundance of the GTDB placeholder taxon Phascolarctobacterium_A sp900545025, with increased predicted menaquinone biosynthetic potential. Microbial and serum metabolite profiles differed significantly among subgroups. Microbiome classifiers achieved AUROCs of 0.71, 0.58, and 0.73 in the held-out internal evaluation, whereas metabolome classifiers achieved AUROCs of 0.89, 0.83, and 0.93 for NAFLD_N, NAFLD_DLP, and NAFLD_IFG. Post hoc integrated panels yielded descriptive AUROCs of 0.92, 0.89, and 0.96 in that subset. Dialister invisus supplementation reduced body weight, serum ALT, AST, triglycerides, fasting blood glucose, and ipGTT AUC in high-calorie diet-fed mice. Conclusion This study identifies shared gut microbial and serum metabolic alterations across NAFLD patients irrespective of IFG or dyslipidemia, alongside subgroup-specific signatures. The Dialister invisus findings support its potential role in metabolic regulation and warrant mechanistic investigation.