The subgingival microbiome associated with periodontitis in type 2 diabetes mellitus

Abstract

Type 2 diabetes mellitus (T2DM) is a systemic disease, predisposing patients to other inflammatory conditions including periodontitis. The subgingival microbiome, a key player in periodontitis pathogenesis, is not well characterized in T2DM population. To better understand whether the subgingival microbiome is different between T2DM and systemically healthy, nondiabetic (ND) subjects, we performed a longitudinal analysis of the subgingival microbiome in T2DM patients (n = 15) compared with ND subjects (n = 16). Using metagenomic shotgun sequencing, we investigated the microbiome in the healthy periodontal state, periodontitis state, and resolved state after treatment. We found that in the periodontitis state, the shift in the subgingival microbiome from the healthy state was less prominent in T2DM compared with ND subjects, yet the clinical signs of disease were similar for both. Furthermore, we revealed highly correlated presence of pathogenic species in relative abundance not only in the periodontitis state, but also in the healthy state in T2DM, suggesting an elevated risk of progression to periodontitis in this cohort. We further investigated the functional potentials of the subgingival microbiome and identified a set of microbial marker genes associated with the clinical states. These genes were significantly enriched in 21 pathways, some of which are associated with periodontitis and some potentially link T2DM and periodontitis. This study identified the longitudinal changes of the subgingival microbiome associated with periodontitis in T2DM and suggests that T2DM patients are more susceptible to shifts in the subgingival microbiome toward dysbiosis, potentially due to impaired host metabolic and immune regulation.

Fig. 1

Subgingival microbiome comparison among periodontal states in T2DM and ND subjects. Principal coordinate analysis (PCoA) based on weighted UniFrac distance shows no significant difference between the healthy state and resolved state in both T2DM and ND subjects, however, the extent of the microbiome shift from the healthy to the periodontitis state was less in T2DM (p = 0.26) compared with ND subjects (p < 0.001). The same PCoA plot with subject IDs labeled is shown in Supplementary Fig. 2 for visualization of related samples

Fig. 2

The microbiome index indicates subgingival microbiome differences among periodontal states and between T2DM and ND subjects. The microbiome index was represented in a violin plot in each clinical state in T2DM and ND subjects. In T2DM, the microbiome in the periodontitis state had not shifted to the extent as observed in the disease state of ND subjects, while the periodontal tissue clinically manifested signs of periodontitis, suggesting that T2DM subjects are more susceptible to shifts in the subgingival microbiome toward the disease state

Fig. 3

Comparison of the relative abundances of the prevalent subgingival bacterial species among clinical states and between T2DM and ND subjects. The average relative abundances of the 51 prevalent bacterial species are shown for each clinical state in T2DM and ND. In ND, 15 species indicated by the red bar on the right were significantly more abundant in periodontitis state than in the healthy state, while five species indicated by the green bar were significantly less abundant. In T2DM, the differences between the periodontitis and healthy states had a similar trend as seen in ND, but none of the species differed with statistical significance

Fig. 4

Correlations in relative abundance between subgingival species. The correlations in relative abundance among prevalent species (see list in Supplementary Table 1) were calculated based on Pearson correlation coefficients. In each heatmap, the species are listed in the same order in both horizontal and vertical axes, and the colored bars indicate the bacterial complexes based on Socransky’s designation [37] (more details in Supplementary Table 1). Black boxes in the heatmaps indicate the bacterial clusters that comprise red complex species with an average Pearson correlation coefficients >0.7 between the species. Hierarchical clustering was applied to the correlation matrices

Fig. 5

KO genes encoded in the subgingival microbiome with different prevalence among clinical states and between T2DM and ND subjects. A total of 373 KO genes with different prevalence among the clinical states in T2DM and ND are shown. The bacterial species, which the KO genes belong to, are indicated by the color bars on the right according to Socransky’s designation of the bacterial complexes [37]. The KO genes labeled in brown are identified in the species from more than one microbial complex (more details in Supplementary Table 2)

Fig. 6

Microbial pathways associated with periodontitis and T2DM. Twenty-one microbial pathways were significantly enriched with KO genes that were different in prevalence between clinical states in T2DM and ND. The proportion of known virulence factors identified in each pathway is shown to the left of the heatmap

Fig. 7

Dynamics of the subgingival microbiome associated with periodontitis in T2DM and ND subjects. A schematic quantitatively illustrates the dynamic changes in the subgingival microbiome composition among the healthy state, periodontitis state, and resolved state in T2DM and ND. The health- and disease-associated species are depicted according to the subgingival complexes designated by Socransky et al. [37] and the knowledge of bacterial interactions from the literature [39]. The proportions of the species are scaled based on their relative abundances in our data from this study