Microbiological characteristics of different tongue coatings in adults

Abstract

Background: Tongue coating is an important health indicator in traditional Chinese medicine (TCM). The tongue coating microbiome can distinguish disease patients from healthy controls. To study the relationship between different types of tongue coatings and health, we analyzed the species composition of different types of tongue coatings and the co-occurrence relationships between microorganisms in Chinese adults. From June 2019 to October 2020, 158 adults from Hangzhou and Shaoxing City, Zhejiang Province, were enrolled. We classified the TCM tongue coatings into four different types: thin white tongue fur (TWF), thin yellow tongue fur (TYF), white greasy tongue fur (WGF), and yellow greasy tongue fur (YGF). Tongue coating specimens were collected and used for 16S rRNA gene sequencing using the Illumina MiSeq system. Wilcoxon rank-sum and permutational multivariate analysis of variance tests were used to analyze the data. The microbial networks in the four types of tongue coatings were inferred independently using sparse inverse covariance estimation for ecological association inference.

Results: The microbial composition was similar among the different tongue coatings; however, the abundance of microorganisms differed. TWF had a higher abundance of Fusobacterium periodonticum and Neisseria mucosa, the highest α-diversity, and a highly connected community (average degree = 3.59, average closeness centrality = 0.33). TYF had the lowest α-diversity, but the most species in the co-occurrence network diagram (number of nodes = 88). The platelet-to-lymphocyte ratio (PLR) was associated with tongue coating (P = 0.035), and the YGF and TYF groups had higher PLR values. In the co-occurrence network, Aggregatibacter segnis was the "driver species" of the TWF and TYF groups and correlated with C-reactive protein (P < 0.05). Streptococcus anginosus was the "driver species" in the YGF and TWF groups and was positively correlated with body mass index and weight (P < 0.05).

Conclusion: Different tongue coatings have similar microbial compositions but different abundances of certain bacteria. The co-occurrence of microorganisms in the different tongue coatings also varies. The significance of different tongue coatings in TCM theory is consistent with the characteristics and roles of the corresponding tongue-coating microbes. This further supports considering tongue coating as a risk factor for disease.

Keywords: Co-occurrence networks; Disease prevention; Driver species; Microbiome; Tongue coating.

Conflict of interest statement

All authors have no conflicts of interest to declare.

© 2022. The Author(s).

Figures

Fig. 1

Chinese medicine tongue coating types and photos. Tongue coating: a layer of moss-like material covering the tongue. TWF:thin white tongue fur; TYF:thin yellow tongue fur; WGF:white greasy tongue fur; YGF:yellow greasy tongue fur

Fig. 2

Analysis of microbial a-diversity of four different types of tongue coating. A Rarefaction curve: the abscissa is the number of sequencing strips randomly selected from a sample, and the ordinate is the number of OTUs that can be constructed based on the number of sequencing strips, which is used to reflect the sequencing depth. Different samples are represented by curves of different colors. B Venn diagram of OTUs with four different tongue coatings.The Veen diagram depicts the number of OTUs specific and shared by each of the four tongue coatings TWF, TYF, YGF, and WGF. C and D Results of a-diversity analysis of four tongue coatings. C Simpson index; D Shannon index

Fig. 3

Analysis of microbial β-diversity of four different types of tongue coatings A and B Results of β-diversity analysis of four tongue coatings. A unweighted Unifrac distances and (B) weighted Unifrac distances. C and D Cumulative graph of microorganisms with percentages greater than 1 in four different tongue coatings. C Genus level and (D) Species level. E and F The differences in microbial abundance between TWF and the remaining three tongue coating types at the species level showed statistically significant results. E TWF-TYF, (F) TWF-YGF and (G) TWF-WGF

Fig. 4

Discrepancies of co-occurrence networks of the four different tongue coating types A, B, C and D The complex nature of inter microbial interactions in the ecological community of each tongue coating was characterized by co-occurrence networks using graphs. Main groups of co-occurrence species are presented in different colours, and smaller groups are shown in grey. Networks of species in the four tongue coatings are identified by positive and negative correlations among the dominant bacteria. Red line indicates positive correlation and blue line indicates negative correlation. Only the bacterial connections (edges) larger than cut-offs (correlation values > 0.4) are retained. Each node in the network indicates a species. The size of each node is proportional to the relative abundance of each species. Nodes in red color show driver microbes which significantly contributed to the separation of the networks (NESH-score value of > 2). E, F and G Network view: All nodes belonging to the same community are randomly assigned similar colors, and gray nodes represent nodes existing in both case and control subnets. The node size is directly proportional to its nesh score. If the nesh score of a node is case > control, it will be marked in red. Therefore, a large red node can be regarded as a driving node. For edges, red indicates the interaction that exists only in the case subnet, green indicates the interaction that exists only in the control subnet, and blue indicates the interaction that exists in both subnets

Fig. 5

A The effect of clinical indicators and demographic variables on the microbial a-diversity of the tongue, with red representing positive correlations and blue representing negative correlations.” + ” indicates p < 0.05,” +  + ” indicates p < 0.001. B Contributions of clinical and demographic variables to the differences in relative abundances of key distinguishing bacterial species for four tongue coatings based on Spearman correlation coefficients and best multiple regression model. Color bar shows correlation values, where red color indicates positive association, blue color negative association, and only significant correlations were shown.” + ” indicates p < 0.05,” +  + ” indicates p < 0.001