Quantitative analysis of the effects of essential oil mouthrinses on clinical plaque microbiome: a parallel-group, randomized trial.

BACKGROUND: The rich diversity of microorganisms in the oral cavity plays an important role in the maintenance of oral health and development of detrimental oral health conditions. Beyond commonly used qualitative microbiome metrics, such as relative proportions or diversity, both the species-level identification and quantification of bacteria are key to understanding clinical disease associations. This study reports the first-time application of an absolute quantitative microbiome analysis using spiked DNA standards and shotgun metagenome sequencing to assess the efficacy and safety of product intervention on dental plaque microbiome. METHODS: In this parallel-group, randomized clinical trial, essential oil mouthrinses, including LISTERINE® Cool Mint Antiseptic (LCM), an alcohol-containing prototype mouthrinse (ACPM), and an alcohol-free prototype mouthrinse (AFPM), were compared against a hydroalcohol control rinse on clinical parameters and the oral microbiome of subjects with moderate gingivitis. To enable a sensitive and clinically meaningful measure of bacterial abundances, species were categorized according to their associations with oral conditions based on published literature and quantified using known amounts of spiked DNA standards. RESULTS: Multivariate analysis showed that both LCM and ACPM shifted the dysbiotic microbiome composition of subjects with gingivitis to a healthier state after 4 weeks of twice-daily use, resembling the composition of subjects with clinically healthy oral conditions recruited for observational reference comparison at baseline. The essential oil-containing mouthrinses evaluated in this study showed statistically significant reductions in clinical gingivitis and plaque measurements when compared to the hydroalcohol control rinse after 6 weeks of use. CONCLUSIONS: By establishing a novel quantitative method for microbiome analysis, this study sheds light on the mechanisms of LCM mouthrinse efficacy on oral microbial ecology, demonstrating that repeated usage non-selectively resets a gingivitis-like oral microbiome toward that of a healthy oral cavity. TRIAL REGISTRATION: The trial was registered on ClinicalTrials.gov on 10/06/2021. The registration number is NCT04921371.

Targeting macrophages and their recruitment in the oral cavity using swellable (+) alpha tocopheryl phosphate nanostructures.

The phosphorylation of (+) alpha tocopherol produces adhesive nanostructures that interact with oral biofilms to restrict their growth. The aim of this work was to understand if these adhesive (+) alpha tocopheryl phosphate (α-TP) nanostructures could also control macrophage responses to the presence of oral bacteria. The (+) α-TP planar bilayer fragments (175 nm ±â€¯21 nm) formed in a Trizma®/ethanol vehicle swelled when exposed to the cell lines (maximum stabilized size = 29 µm). The swelled (+) α-TP aggregates showed selective toxicity towards THP-1 macrophages (LD50 = 304 µM) compared to human gingival fibroblasts (HGF-1 cells; LD50 > 5 mM), and they inhibited heat killed bacteria stimulated MCP-1 production in both macrophages (control 57.3 ±â€¯18.1 pg/mL vs (+) α-TP 6.5 ±â€¯3.2 pg/mL) and HGF-1 cells (control 673.5 ±â€¯133 pg/mL vs (+) α-TP - 463.9 ±â€¯68.9 pg/mL).

In vitro efficacy of essential oil mouthrinse versus dentifrices.

OBJECTIVES: To compare the antimicrobial efficacy and kill penetration of essential oils (EO) mouthrinse versus stannous fluoride, and triclosan dentifrice slurries on saliva-derived biofilms using confocal laser scanning microscopy (CLSM). METHODS: Saliva-derived biofilms were grown for 48h on hydroxyapatite discs using pooled, homogenized saliva from 8 healthy volunteers as the inoculum. The mean thickness of these biofilms was 84µm (range, 23-241µm). CLSM with viability mapping was used to visualize the antimicrobial kill penetration of each treatment regime within a biofilm. RESULTS: At 30s treatment durations, CLSM imaging revealed greater antimicrobial activity and kill penetration of EO mouthrinse compared to sodium fluoride-, stannous fluoride-, and triclosan-containing dentifrice slurries. Quantification of biovolume revealed that EO mouthrinse treatment at 30s resulted in a greater non-viable biovolume proportion (84.6%±15.0%) than other treatment groups. Increasing the treatment duration of the triclosan dentifrice (to 60 and 120s) resulted in better penetration and an increased reduction of viable cells, comparable to EO mouthrinse treatment at 30s duration. Further, CLSM imaging showed that the combined treatment of a non-antimicrobial dentifrice (45s) with EO mouthrinse (30s) showed superior antimicrobial activity (96.2%±3.7%) compared to the antimicrobial triclosan-containing dentifrice used without a mouthrinse step (26.0%±32.0%). CONCLUSIONS: Within typical exposure times, the EO-containing mouthrinse can penetrate deep into the accumulating plaque biofilm compared to the chemotherapeutic dentifrice slurries, and may provide an efficacious alternative to triclosan, when used as an adjunct with a mechanical oral care regimen. CLINICAL SIGNIFICANCE: Using viability mapping and CLSM, this study demonstrated that EO-containing mouthrinse penetrates and kills microorganisms deeper and more effectively in plaque biofilm in typical exposure times when compared to dentifrice chemotherapeutic agents, providing an efficacious alternative to triclosan or stannous fluoride when used as an adjunct to mechanical oral care.