Phylogenetic analysis of bacterial and archaeal species in symptomatic and asymptomatic endodontic infections

Abstract

Phylogenetic analysis of bacterial and archaeal 16S rRNA was used to examine polymicrobial communities within infected root canals of 20 symptomatic and 14 asymptomatic patients. Nucleotide sequences from ∼750 clones amplified from each patient group with universal bacterial primers were matched to the Ribosomal Database Project II database. Phylotypes from 37 genera representing Actinobacteria, Bacteroidetes, Firmicutes, Fusobacteria and Proteobacteria were identified. Results were compared to those obtained with species-specific primers designed to detect Prevotella intermedia, Porphyromonas gingivalis, Porphyromonas endodontalis, Peptostreptococcus micros, Enterococcus sp., Streptococcus sp., Fusobacterium nucleatum, Tannerella forsythensis and Treponema denticola. Since members of the domain Archaea have been implicated in the severity of periodontal disease, and a recent report confirms that archaea are present in endodontic infections, 16S archaeal primers were also used to detect which patients carried these prokaryotes, to determine if their presence correlated with severity of the clinical symptoms. A Methanobrevibacter oralis-like species was detected in one asymptomatic and one symptomatic patient. DNA from root canals of these two patients was further analysed using species-specific primers to determine bacterial cohabitants. Trep. denticola was detected in the asymptomatic but not the symptomatic patient. Conversely, Porph. endodontalis was found in the symptomatic but not the asymptomatic patient. All other species except enterococci were detected with the species-specific primers in both patients. These results confirm the presence of archaea in root canals and provide additional insights into the polymicrobial communities in endodontic infections associated with clinical symptoms.

Miniprimer PCR, a New Lens for Viewing the Microbial World

“…The primers 27F-P and 1492R-P are "first-generation" nondegenerate universal bacterial primers that remain in widespread use today (1,5,18,19,39), and 27F-HT and 1492R-HT are more recently published primers that are based on the original first-generation primers and include several degenerate positions to broaden their scope of targets (37). These primers produce nearly full-length 16S rRNA gene sequences and thus provide a fair assessment of the miniprimer method relative to standard methods in common use; duplicate libraries were constructed with each of the primer pairs ( Table 2).…”

Exploring the oral bacterial flora: current status and future directions

“…16S rDNA sequencing has shown that the infected canals contain a more diverse flora than previously recognized (Vickerman et al, 2007), but surprisingly symptomatic root canal lesions harbor a less diverse flora than do asymptomatic lesions. Certain Streptococcus spp.…”

“…Methanobrevibacter oralis-like phylotypes are found in oral niches but their pathological associations are yet to be determined (Lepp et al, 2004;Vickerman et al, 2007) DNA microarrays to study the diversity of subgingival plaque microflora…”

The Etio-Pathogenesis of Periodontal Disease

“…It can also be detected in root-canal samples. [10] Archaea have been isolated from the human oral cavity, gut, colon, and vagina; [3,9] but have not been established as causes of human disease. [3] It has been detected that interactions between viruses and bacteria within periodontal pockets is as a process of etiopathogenesis of periodontal diseases.…”