To investigate the role and potential mechanisms of oropharyngeal microbiota dysbiosis in the development and progression of post-coronavirus disease 2019 (COVID-19) pulmonary fibrosis (PCPF).

A cross-sectional case-control study was conducted, enrolling recovered patients who had been hospitalized for COVID-19 in Xuanwu Hospital Capital Medical University, from December 2022 to May 2023, and had completed a 2-year follow-up after discharge. According to high-resolution computed tomography findings during follow-up, patients were classified into fibrosis group and non-fibrosis group. The α-diversity of the oropharyngeal microbiota between the two groups were compared by Mann-Whitney U test; β-diversity was assessed by non-metric multidimensional scaling (NMDS) and principal coordinates analysis (PCoA). Differences in taxonomic abundance at the phylum, genus and species levels were analyzed by Benjamini-Hochberg method with false discovery rate (FDR) correction. Spearman’s rank correlation was applied to evaluate the associations between microbial taxa and clinical symptoms as well as exercise capacity.

Total of 83 patients were enrolled, 43 cases in fibrosis group and 40 cases in non-fibrosis group. During the 2-year follow-up, patients in fibrosis group had more respiratory symptoms and anxiety. Patients in fibrosis group showed significantly higher scores than non-fibrosis group in mMRC scale (U=676.0, P=0.008), BCSS score (U=625.0, P=0.029), CAT score (t=2.127, P=0.036), CET score (U=538.5, P=0.002) and Hamilton Anxiety Rating Scale score (U=634.0, P=0.039); The 6-minute walking distance of patients in fibrosis group was shorter (U=598.5, P=0.017), all with significant differences. Microbiome analysis revealed significantly elevated α-diversity of oropharyngeal microbiota in fibrosis group (Chao1 index: U=580.5, P=0.010; Shannon index: U=557.0, P=0.005; Simpson index: U=513.0, P=0.001; Faith’s phylogenetic diversity: U=579.0, P=0.010; Simpson’s evenness index: U=577.0, P=0.010; Heip’s evenness index: U=529.0, P=0.035), with significant differences. β-diversity analysis showed distinct separation of microbial community structure between the two groups. Taxonomic analysis demonstrated that the relative abundances of potential pro-inflammatory taxa, including Prevotella (U=358.0, P<0.001), Veillonella (U=418.0, P<0.001) and Streptococcus (U=538.0, P=0.003), were significantly increased in fibrosis group, whereas Neisseria (U=226.0, P<0.001) was significantly decreased. Correlation analysis indicated that the abundance of Streptococcus parasanguinis was positively associated with mMRC (r=0.24, P=0.04) and CAT (r=0.27, P=0.02). In contrast, Fusobacterium periodonticum abundance was negatively correlated with CAT (r=－0.30, P<0.01) and Warrick score (r=－0.26, P=0.02), but positively correlated with 6MWD (r=0.26, P=0.02). Furthermore, Veillonella rogosae and Streptococcus infantis were both significantly and negatively correlated with Warrick score (r=－0.30, P<0.01; r=－0.28, P=0.01).

Patients with post-COVID-19 pulmonary fibrosis exhibited persistent oropharyngeal microbial dysbiosis, which is closely associated with clinical symptoms and functional impairment. The oropharyngeal microbiome might serve as a potential non-invasive biomarker for risk assessment and a novel therapeutic target for post-COVID-19 pulmonary fibrosis.