Background
The oral microbiome plays a key role in the production of nitric oxide (NO), a crucial signalling molecule involved in cardiovascular health and exercise performance (Bryan et al., 2022). NO is synthesised through the bacterial reduction of nitrate (NO₃⁻) to nitrite (NO₂⁻) in the oral cavity (Burleigh et al., 2018). Previous studies suggest that physical activity is linked to oral health (Tripodi et al., 2021) and NO bioavailability (Oral, 2021), but the impact of exercise on the oral microbiome remains unclear. This study assessed differences in the oral microbiome composition and NO₃⁻/NO₂⁻ levels between competitive athletes and inactive controls.
Methods
Ten highly trained/national athletes (Tier 3) and ten inactive controls (Tier 0) were recruited (McKay et al., 2022). V̇O2max was confirmed via a graded exercise test to exhaustion, habitual diet and weekly exercise volume were recorded using diaries, oral health status was measured via a dental examination. Samples of saliva, plasma, supragingival plaque, and the tongue dorsum microbiome were collected. Microbial composition was analysed using long-read 16S rRNA sequencing. NO₃⁻ and NO₂⁻ levels were measured using ozone-based chemiluminescence. Differences in microbial diversity and abundance were assessed using beta diversity metrics, and correlations were evaluated with heatmaps and confirmed with Spearman rho. Independent t-tests, Wilcoxon, Fisher’s exact tests, and ANCOM-BC2 and FDR filters were applied where appropriate.
Results
Athletes engaged in significantly more weekly exercise (median 484, IQR 382–787 minutes/week) than controls (median 12, IQR 0–60 minutes/week) (W = 0, p < 0.0001). V̇O2max was significantly higher in athletes (61.4 ± 8.8 mL/kg/min) compared to controls (38.6 ± 7.8 mL/kg/min) (t = 6.127, p < 0.001, 95% CI 15.0–30.5 mL/kg/min). No differences were observed in macronutrient intake, nitrate levels, or markers of oral health (all p > 0.05). Beta diversity of the tongue dorsum microbiome differed between groups (p = 0.046), with athletes showing a higher abundance of NO₃⁻-reducing bacteria, Rothia mucilaginosa and unclassified Gemella species. No significant differences were found in the supragingival plaque microbiome (all p > 0.05).
Athletes had higher salivary NO₃⁻ (p = 0.003) and NO₂⁻ (p = 0.03). Plasma NO2– concentration was also higher in the trained group (t=3.439, p=0.003, 95% CI 52.7 – 220.0 nM), indicating enhanced NO bioavailability. Positive correlations were found between the abundance of R. mucilaginosa and Gemella species and V̇O2max (R. mucilaginosa (ρ=0.68, p=0.02), Gemella species (ρ=0.79, p=0.002)). Training volume was also associated with higher levels of these species (R. mucilaginosa (ρ=0.63, p=0.03), and Gemella species (ρ=0.66, p=0.02)).
Conclusions
This pilot study suggests that exercise influences the oral microbiome in ways that support both oral and systemic health, as well as athletic performance. The higher abundance of NO₃⁻-reducing bacteria in athletes, coupled with elevated NO₃⁻ and NO₂⁻ levels, indicates that regular physical activity may enhance NO production, which is known to improve cardiovascular function and performance. While further research with larger samples is necessary to confirm these findings and further investigate the underlying mechanisms, this data provides the first evidence linking exercise-induced changes in the oral microbiome to enhanced nitric oxide bioavailability.