Multi-kingdom profiling reveals altered gut phage-bacteria-metabolite interactions in MASLD

Metabolic dysfunction-associated steatotic liver disease (MASLD) is increasingly linked to gut microbial dysbiosis, but most studies have focused on bacteria, neglecting viruses and fungi, and their interactions. Here we show that MASLD is characterized by coordinated disruption of bacterial, viral and fungal communities and by a disturbed phage-bacteria-metabolite axis associated with disease-related bile acid changes. Integrating shotgun metagenomics, fungal ITS2 sequencing, fecal metabolomics and clinical profiling in 210 patients with MASLD and 210 age- and gender-matched healthy controls, we find reduced microbial diversity and extensive remodeling of cross-kingdom ecological networks in MASLD. Ruminococcus gnavus emerges as an enriched central hub, while Faecalibacterium prausnitzii and its associated bacteriophages are depleted. Phage-host analyses further reveal reduced lytic activity against R. gnavus and loss of sulfur amino acid metabolism-related auxiliary metabolic genes, which may impair F. prausnitzii fitness. Diminished phage control may facilitate R. gnavus expansion, alongside increased fecal isodeoxycholic acid, a secondary bile acid implicated in hepatic steatosis. A diagnostic classifier integrating bacterial and viral features with clinical parameters distinguish MASLD from controls in our cohort and maintain predictive performance in two external datasets. Together, these findings uncover a disrupted phage-bacteria-metabolite axis in MASLD and provide a multi-kingdom framework for non-invasive biomarker discovery and microbiome-targeted therapies.