Multi-omics integration of gut microbiome and plasma metabolome in MASLD: a Hainan-based observational case-control study

Metabolic dysfunction-associated steatotic liver disease (MASLD) affects nearly one-quarter of adults worldwide and has emerged as a leading cause of chronic liver disease. This study aimed to characterize gut microbial composition and plasma metabolite alterations in MASLD patients. This observational case-control study enrolled 60 patients with MASLD, including 30 Han and 30 Li ethnic individuals, along with 60 age, ethnicity, and sex-matched healthy controls (HC), between October 2022 and July 2023 in Hainan Province, China. Analysis of gut microbiota composition and plasma metabolic profiles was conducted using metagenome sequencing and non-targeted liquid chromatography-mass spectrometry (LC-MS). Significant alterations in plasma metabolite composition and intestinal microbiota structure were observed in MASLD patients compared to HC. LEfSe analysis in unadjusted comparisons identified three species ( Bacteroides uniformis , Faecalibacterium prausnitzii , and Lachnospira eligens ) as candidate taxa. However, in multivariable models adjusting for age, sex, BMI, and recruitment site, the disease-group associations for these taxa were not statistically significant (q > 0.05), indicating that these findings are exploratory and require validation in larger cohorts. Untargeted metabolomics detected 14,041 features, of which 1,370 met the screening criteria. After Benjamini-Hochberg false discovery rate (FDR) correction, 236 features remained statistically significant (q < 0.05). Lipid metabolism was the most prominently affected pathway among the FDR-significant metabolites. Faecalibacterium prausnitzii showed a strong positive correlation with 3-Oxo-5β-cholanoic acid and D-Serine. Additionally, exploratory subgroup analysis across Han (Haikou-recruited) and Li (Wuzhishan-recruited) populations identified Bacteroides caccae and Haemophilus parainfluenzae as candidate species associated with the respective ethnicity/site groups. However, because ethnicity and recruitment site are completely confounded, these findings should be interpreted as exploratory and hypothesis-generating. Decreases in Bacteroides uniformis , Faecalibacterium prausnitzii , and Lachnospira eligens were associated with MASLD in unadjusted analyses, though these associations were not significant after multivariable adjustment. The observed weak-to-moderate correlation between Faecalibacterium prausnitzii and 3-oxo-5β-cholanoic acid or D-Serine represents a hypothesis-generating finding that warrants independent validation in larger cohorts and experimental models before any clinical translation can be considered.