Our randomized, placebo-controlled trial showed resistant starch (RS), a type of prebiotic, has therapeutic effects in metabolic dysfunction-associated steatotic liver disease (MASLD). Here, we observed its heterogeneous efficacy, where 30% of participants exhibited limited benefits, which was replicated in a multi-center trial (ChiCTR2300074588). Multi-omics analysis and fecal microbiota transplantation identified baseline microbiota as a dominant contributor of response. Further population stratification and network analysis combined with in vitro and in vivo experiments revealed Prevotella as the key cause of low response by inhibiting RS-degrading bacteria, thereby impairing RS utilization. Conversely, Bifidobacterium pseudocatenulatum RRP01, a strain isolated from our cohort, restored RS degradation and improved Prevotella-attenuated RS response. Furthermore, we developed a predictive model integrating baseline microbial and clinical features (area under the curve [AUC] = 0.74–0.87), enabling stratification for personalized interventions. Our study indicates that gut microbiota determines the heterogeneity in RS efficacy and offers possibilities for novel microbiota- oriented precision therapeutics for MASLD.