Broad-range 16S rDNA sequencing and quantitative bacteriome profiling: a small-cohort feasibility study in...

Many bacteriome studies employing next-generation sequencing (NGS) have focused on the partial sequencing of the 16S rRNA gene (rDNA). This limits both species-level resolution and quantitative interpretation of bacterial abundance, which have been addressed in this study.

Nanopore sequencing was used to analyze nearly the entire 16S rDNA sequence for species-level classification, and quantitative PCR for bacterial quantification. This approach was applied in a small-cohort feasibility study to investigate oral bacterial changes in orthodontic patients. Saliva samples were collected from 10 patients before and 1, 3, and 6 months after the initiation of orthodontic treatment.

Dental caries was not detected during the study, and periodontal pockets ≥4 mm were rarely observed. However, all patients exhibited changes in bleeding on probing (BOP), which is indicative of early stage gingivitis. Although the compositional analysis did not reveal any significant association between specific bacterial species and BOP changes, the quantitative analysis showed a positive correlation between BOP changes and two bacterial species in the phyla Candidatus Saccharibacteria: Candidatus Saccharimonas aalborgensis and Candidatus Saccharibacteria bacterium oral taxon TM7x. No correlation was observed with representative periodontal disease-associated bacteria.

These findings support the feasibility of species-level and quantitative bacteriome analysis in a small cohort and highlight the bacterial species potentially linked to early stage gingivitis.