Effect of salivary antimicrobial factors on microbial composition of tongue coating in patients with coronary heart disease with phlegm-stasis syndrome

Objective: Phlegm-stasis syndrome is one of the most common traditional Chinese medicine (TCM) syndromes and found in 59% of patients with coronary heart disease (CHD) in routine TCM clinical practice in China. One of the diagnostic criteria of phlegm-stasis syndrome is its characteristic white-greasy and thin tongue coating. We have previously reported that different types of tongue coating have different microbiome characteristics that can be used as diagnostic markers. However, the microbial characteristics of tongue coating of CHD patients with phlegm-stasis syndrome, including coating feature and underlying reason of formation, have rarely been reported. Herein, we examined the characteristic microbiome of tongue coating, and discussed the cause of tongue coating formation via salivary proteins in patients with phlegm-stasis syndrome. Methods: We examined white-greasy and thin tongue coatings obtained from 10 patients with CHD having phlegm-stasis syndrome (n = 10), and compared with those of patients with almost no coating – patients with Qi-Yin deficiency syndrome (n = 10) – and healthy controls (n = 10). 16S rRNA sequencing of tongue coating microbiome and isobaric tags for relative and absolute quantitation (iTRAQ)-based quantitative analysis of salivary proteins were used to detect tongue coating and salivary protein separately. Salivary levels of sIgA, lysozyme, and amylase were detected by ELISA. Results: We identified Candidatus_Saccharimonas and Candidate_division_TM7_norank as the prominent members of tongue coating in patients with CHD having phlegm-stasis syndrome. Salivary proteins involved in biological processes, pentose phosphate pathway, and complement and coagulation cascades were among the differentially expressed proteins identified in patients with CHD having phlegm-stasis syndrome on iTRAQ analysis. Moreover, the formation of microbiota in tongue coating was associated with salivary sIgA, lysozyme, and saliva flow rate. Conclusions: We explored the characteristics of microbial composition of tongue coating patients with CHD having phlegm-stasis syndrome and identified correlations between salivary proteins and microbiome formation, providing a theoretical and mechanistic basis for tongue coating formation.