Changes in the Type 2 diabetes gut mycobiome associate with metformin treatment across populations

Abstract

The human gut teems with a diverse ecosystem of microbes, yet non-bacterial portions of that community are overlooked in studies of metabolic diseases firmly linked to gut bacteria. Type 2 diabetes mellitus (T2D) associates with compositional shifts in the gut bacterial microbiome and fungal mycobiome, but whether T2D and/or pharmaceutical treatments underpin the community change is unresolved. To differentiate these effects, we curated a gut mycobiome cohort to-date spanning 1,000 human samples across 5 countries and a murine experimental model. We use Bayesian multinomial logistic normal models to show that metformin and T2D both associate with shifts in the relative abundance of distinct gut fungi. T2D associates with shifts in the Saccharomycetes and Sordariomycetes fungal classes, while the genera Fusarium and Tetrapisipora most consistently associate with metformin treatment. We confirmed the impact of metformin on individual gut fungi by administering metformin to healthy mice. Thus, metformin and T2D account for subtle, but significant and distinct variation in the gut mycobiome across human populations. This work highlights for the first time that oral pharmaceuticals can confound associations of gut fungi with T2D and warrants the need to consider pharmaceutical interventions in investigations of linkages between metabolic diseases and gut microbial inhabitants.

Keywords: Gut mycobiome; gut microbiome; metagenomics; type 2 diabetes mellitus.

Figure 1:. Characterization of the gut mycobiome from metagenomics data across studies.

(A) The geographic location of each study. The size of the bubbles corresponds with total sample size. (B) An Upset plot demonstrating shared genera between studies. The grey dot matrix shows which study-treatment category combination shared mycobiome features (genera) and the blue bar shows the total number shared. (C-E) Barplots of compositional abundance at the fungal class level from each study, geographic region, and treatment category without adjustment for between-study differences.

Figure 2:. Metformin’s associations with gut fungi in T2D in (A) the total dataset, (B, D) randomized clinical trials (RCTs), and (3) cross-sectional studies.

The black triangle and lines show posterior mean and 95% credible intervals of the aggregate dataset. Colored diamond points show posterior means of individual studies. The arrows show increased relative abundance at baseline or T2D-NOMET (left) or during metformin (MET) treatment (right). The dashed vertical line denotes zero association. (D) In three RCTs with time-series sampling, proportional relative abundance is shown at baseline and after starting metformin treatment for each study. Paired samples from the same individual are shown as neighboring horizontal bars of proportional relative abundance with the duration of metformin treatment shown above the barplot for each study. Taxa are colored as Class:Family, and less relatively abundant taxa are merged in the “other” category.

Figure 3:. Metformin is associated with similar differences in the human and mouse mycobiomes.

Black triangles and line ranges shows posterior mean and posterior 95% credible intervals of the aggregate dataset. Colored diamond points show posterior means of individual studies. Red squares show the posterior means of the mouse study. The arrows show increased abundance in NORM subjects or mice with placebo treatment (left) or T2D-MET subjects and metformin-treated mice (right). The dashed vertical line denotes zero association.

Figure 4:. Metabolic disease is associated with a shift in genera belonging to the Saccharomycetes class.

Subjects with T2D-NOMET were compared to NORM (A), and markers of metabolic disease were compared; (B) fasting blood glucose (FBG), (C) fasting plasma insulin (FPI), and (D) body mass index (BMI). In all panels, black triangles and line ranges shows posterior mean and posterior 95% credible intervals of the aggregate dataset. Colored diamond points show posterior means of individual studies. Arrows show increased fungal abundance. The dashed vertical line denotes zero association.