Abstract Body: Background: American Indians experience a markedly high prevalence of type 2 diabetes (T2D). Emerging evidence links the gut microbiome to T2D, yet data in this high-risk population are scarce.
Objective: To identify specific gut microbial taxa and functional pathways associated with T2D, and to assess their mediating role in the relationships between diet and T2D among American Indians in the Strong Heart Study (SHS).
Methods: We conducted deep shotgun metagenomic sequencing (~40 million reads/sample) on stool samples from 971 American Indian adults (mean age 57 years, 68% women) enrolled in the SHS Phase 7 (2022–2025). Taxonomic and functional profiles were generated using MetaPhlAn4 and HUMAnN3, respectively. Diet was assessed via a culturally tailored food frequency questionnaire. Mixed-effects logistic regression models were used to identify microbial species and functional pathways associated with T2D, adjusting for age, sex, study center, BMI, diet control (yes/no), antibiotic use, and Bristol stool scale (with random effects for family relatedness and sequencing batch). Mediation analyses then quantified microbial contributions to diet–T2D associations.
Results: T2D prevalence was 37.4% (n=363). Family clustering (R²=10.3%, P=0.001) was the strongest predictors of β-diversity (Bray-Curtis), followed by age (R²=1.6%, P=0.001), study center (R²=0.78%, P=0.001), sex (R²=0.60%, P=0.001), and T2D status (R²=0.43%, P=0.003). Furthermore, we identified 9 species (e.g., Escherichia coli, Enterocloster bolteae, Dielma fastidiosa) and 7 metabolic pathways (e.g., glycolysis, thiamine phosphate formation) significantly associated with T2D (false discovery rate (FDR)<0.05). Individuals with T2D reported significantly lower consumption of several key nutrients, including fiber (odds ratio (OR)=0.98, P=0.006), vitamin A (OR=0.99, P=0.007), vitamin D (OR=0.93, P=0.001), and vitamin B12 (OR=0.97, P=0.03). Critically, several species (e.g., Escherichia coli, Phocaeicola vulgatus) were found to mediate 7%–20% of the associations between these dietary nutrients and T2D (P<0.05).
Conclusion: Gut microbial composition and function are robustly associated with T2D and significantly mediate the effects of diet on T2D in American Indians. These findings highlight the gut microbiome as a novel and viable target for T2D prevention and treatment in this high-risk population.