Bacterial Volatile Organic Compounds as Potential Caries and Periodontitis Disease Biomarkers

Abstract

Oral diseases represent a significant global health and economic burden, necessitating the development of effective diagnostic tools. This study investigates the volatile organic compound (VOC) profiles of bacteria associated with dental caries and periodontal disease to explore their potential as diagnostic biomarkers. Four microbial strains-Streptococcus mutans (700610), Streptococcus sanguis (NCO 2863), Porphyromonas gingivalis (ATCC 33277), and Fusobacterium nucleatum (PK1594)-were cultured (N = 24), alongside intraoral samples (N = 60), from individuals with common oral diseases. Headspace VOCs were analyzed using gas chromatography-mass spectrometry (GC-MS), and statistical analyses were conducted by applying non-parametric Wilcoxon and Kruskal-Wallis tests. VOC identification was performed using the NIST14 database. Strain-specific VOC signatures were identified, with P. gingivalis and F. nucleatum exhibiting distinct profiles from each other and from Streptococcus strains. Comparative analysis of disease cohorts revealed statistically significant differences at multiple retention times between caries, gingivitis, and periodontitis. These findings suggest that VOC profiling enables differentiation between bacterial strains and disease phenotypes, supporting their potential application as diagnostic biomarkers for oral diseases. This study establishes a foundational framework for VOC-based diagnostic methodologies in dental pathology.

Keywords: Fusobacterium nucleatum; Porphyromonas gingivalis; Streptococcus mutans; Streptococcus sanguis; gas chromatography–mass spectrometry; metabolomics; oral diseases.

Figure 1

Venn diagram of signature VOCs of Fusobacterium nucleatum, Porphyromonas gingivalis, Streptococcus sanguis, and Streptococcus mutans.

Figure 2

Box-plots illustrating a comparative analysis of volatile organic compounds (VOCs) observed in cultured intraoral clinical samples of common oral diseases, considering only compounds with a cut-off of 70% or above, and excluding room control or empty tubes. (A) RT 16 32327. (B) RT 21.22148. (C) RT 24.18885 (D) RT 30.01467 (E) RT 32.85932 (F) RT 38.41207. Each box-plot corresponds to a specific compound and is associated with its respective retention time (RT), illustrating the relative abundance of the compounds quantified by the area under the curve (AUC). The median of the data is represented by the black line within each box. Outliers are displayed as follows: ◦ = Mild outlier (1.5 to 3 times the interquartile range [IQR] from Q1 or Q3); * = Extreme outlier (greater than 3 times the IQR from Q1 or Q3). The whiskers extending from the upper to lower quartiles provide a visual representation of the variability in the data.

Figure 3

Stage I: Four cariogenic and periopathogenic reference microbial strains underwent cultivation (A_I) in hermetically sealed 500 cm³ bottles, incubated at 37 °C for 48 h to promote bacterial growth (B_I). The headspaces above the colonies were transferred to adsorbent tubes (C). The collected samples were desorbed (D) and injected into a GC-MS system (E). Statistical analysis was conducted on the peaks and areas under the curve of each sample’s chromatograms (F). Stage II: Adult participants (N = 60) were clinically categorized into three groups—caries, gingivitis, and periodontitis. Microbial specimens were obtained via intraoral swabbing of each participant (A_II). The intraoral samples were cultured in sealed 500 cm³ bottles and incubated under anaerobic conditions at 37 °C for 72 h to facilitate microbial growth (B_II). Samples were processed and analyzed under similar conditions as Stage 1.