An Exploratory Longitudinal Analysis of Salivary Host, Bacterial and Fungal Extracellular Vesicles Following 6‐Month Non‐Surgical Periodontal Therapy in Periodontitis

AIM: To evaluate the longitudinal changes in salivary host and non-host extracellular vesicles (EVs) following non-surgical periodontal therapy (NSPT). MATERIALS AND METHODS: Unstimulated saliva and plaque (from a deep pocket) were collected from 25 Stage III/IV periodontitis patients at baseline and at 3 and 6 months post NSPT. Host salivary EVs and non-host EVs were isolated using immunoaffinity, and size-exclusion chromatography, respectively. Host EV cytokines were assessed by multiplex flow cytometry, and non-host EVs, saliva and plaque were profiled by 16S rRNA and fungal internal transcribed spacer (ITS) sequencing. RESULTS: Periodontal parameters significantly decreased at 3 and 6 months post NSPT (vs. baseline), accompanied by increased host-EVs associated cytokines interleukin-4 (IL-4), IL-12 and IL-9 (not whole saliva). At both timepoints, bacterial EVs also declined, including the genera Filifactor, Parvimonas and Lentimicrobium and the species Fusobacterium nucleatum, Filifactor alocis, Tannerella forsythia, Treponema denticola, Porphyromonas gingivalis, P. endodontalis, P. micra and Lachnospiraceae bacterium; similar reductions were observed in plaque and saliva taxa, including Treponema, Tannerella, Parvimonas, T. forsythia, T. socranskii and P. micra. At 6 months, fungal EVs from Malassezia arunalokei were decreased and the fungal Candida parapsilosis EVs were increased. CONCLUSIONS: NSPT led to changes in salivary host and non-host EV cargoes, suggesting that they may serve as a non-invasive tool for disease treatment monitoring in periodontitis.