Ecological restructuring of the nonbacterial fecal microbiome in obesity across human cohorts.

Obesity is a complex metabolic disorder increasingly linked to alterations in the gut microbiome. While most research has focused on bacterial communities, the contribution of nonbacterial components including viruses, archaea, and eukaryotic microorganisms remains insufficiently characterized. Here, we performed a multicohort analysis to investigate the role of the nonbacterial gut microbiome in obesity across three independent human cohorts. Using compositional analyses adjusted for key covariates and network based approaches, we identified consistent multikingdom alterations associated with obesity. Individuals without obesity showed a reproducible enrichment of methanogenic archaea, particularly Methanobrevibacter smithii and Methanobrevibacter millerae, whereas individuals with obesity were characterized by increased abundance of bacteriophages from the class Caudoviricetes. In an elderly cohort, eukaryotic taxa such as Blastocystis spp. were additionally associated with the without obesity group. These patterns were largely consistent across cohorts and robust to sex stratification. Beyond taxonomic differences, ecological network analyses revealed substantial reorganization of microbial interactions in obesity. The identity and composition of hub taxa differed significantly between obesity and without obesity networks across all cohorts, indicating a shift in the taxa occupying central ecological roles. Notably, these differences were observed even when similar microbial kingdoms were represented, underscoring the importance of species-level resolution. Collectively, our findings demonstrate that obesity is associated with coordinated compositional and ecological alterations across the nonbacterial gut microbiome. This multikingdom perspective expands current understanding of microbiome dysbiosis in metabolic disease and highlights the archaeome and virome as potential contributors to host metabolic health.