Limitations of 16S rRNA gene as phylogenetic marker: a large-scale meta-omics analysis of plaque microbiota in periodontal diseases

Abstract

In the literature, 16S rRNA gene sequencing is the most widely used technology for studying the periodontal microbiota. However, there is no evidence on how methodological aspects such as primer coverage, detection of matching amplicons (MAs), and clustering into operational taxonomic units (OTUs) could influence the results obtained for the oral niche. Furthermore, the comparison of 16S sequencing-based studies on periodontal microbiota is controversial due to significant methodological differences. Therefore, meta-omics analyses would favour the accuracy of phylogenetic data associated with different periodontal conditions. In the present Thesis, we analysed in silico 1) the coverage of primers employed in sequencing-based studies of the mouth microbiota using oral-specific databases containing bacterial and archaeal 16S rRNA gene sequences; 2) the number of 16S rRNA genes in the complete genomes of bacterial and archaeal species inhabiting the human mouth, and how the use of different primers would affect the detection of MAs from different taxa; and 3) the performance of different primers to detect distinct oral species with 16S rRNA gene amplicon similarity ≥97%, identifying the taxa that may be erroneously grouped into the same OTU.