Functional remodeling of the gut microbiome and metabolome in primary idiopathic male infertility

BACKGROUND: Primary idiopathic male infertility (PIMI) is a complex condition with unclear biological mechanisms. Increasing evidence indicates that gut microbiome-derived functional and metabolic alterations can influence host physiological processes, yet microbiome-associated functional changes in PIMI remain poorly characterized. METHODS: In this case–control study, fecal shotgun metagenomics and untargeted liquid chromatography-tandem mass spectrometry (LC–MS/MS) metabolomics were performed in 19 men with PIMI and 12 fertile controls, alongside computer-assisted semen analysis. The study workflow integrated differential analyses, correlation analyses among key microbial species, metabolites, and clinical traits, and Random Forest modeling to derive a microbial-metabolic panel. RESULTS: Compared with fertile controls, infertile men exhibited selective functional remodeling of gut microbial pathways and fecal metabolic profiles, accompanied by reduced sperm concentration and progressive motility and increased round cell counts. Although overall microbial diversity was broadly comparable between groups, 23 differentially abundant species and 53 altered Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were identified by metagenomic profiling. Untargeted metabolomics annotated 4,434 metabolites and identified 780 differential metabolites, with enrichment of 29 KEGG pathways. Eight key microbial species and eight key metabolites mapped to sperm- and testis-related pathways showed coordinated correlations with semen parameters. An integrated Random Forest model incorporating microbial and metabolic features demonstrated robust discrimination between infertile and fertile men, with optimal performance achieved using six top-ranked features. CONCLUSIONS: PIMI is associated with selective gut microbial functional shifts and fecal metabolic disturbances that correlate with semen quality. Multi-omics integration highlights coordinated microbiome-metabolome alterations, providing insights into host-associated microbial functional dysregulation in male infertility.