Supplementary Material for: Dairy product intake alters the correlations between circulating bile acids and short-chain fatty acids with the bacterial taxa Roseburia, Faecalibacterium, Flavonifractor and Verrucomicrobia

Introduction: Type 2 diabetes (T2D) risk factors are associated with gut microbiota dysregulation that can alter circulating metabolites levels such as bile acids (BAs) and short-chain fatty acids (SCFAs). The objective is to investigate how the high dairy intake (HD) (≥ 4 servings/day) product intake compared to adequate dairy (AD) (≤ 2 servings/day) intake influences the correlations between Roseburia, Faecalibacterium, Flavonifractor as well as Verrucomicrobia and circulating BAs and SCFAs in subjects at risk of T2D. Methods: In a randomized crossover trial, 10 hyperinsulinemic adults were randomized to HD or AD for 6 weeks separated by a 6-week washout period. Gut microbiota were measured with 16S rRNA-based high-throughput sequencing in the gut. BA profiling in plasma was performed by high-performance liquid chromatography-tandem mass spectrometry. Serum SCFAs were determined using headspace gas-chromatography. Results: No significant differences were observed in mean circulating BA or SCFA levels between AD and HD consumption. Verrucomicrobia and Flavonifractor showed positive correlations with secondary BAs, following HD and AD intake, respectively. Additionally, Flavonifractor correlated positively with acetic and propionic acids after HD intake. Roseburia correlated positively with primary BAs, propionate and butyrate after HD intake. Faecalibacterium was positively correlated with cholic acid after AD intake and with hexanoic acid after HD intake. Conclusion: These findings suggest that high dairy intake may modulate microbiota–metabolite interactions without altering circulating metabolite concentrations, highlighting a potential role for dietary patterns in shaping gut-derived metabolic signals in individuals at risk of T2D.