Investigation of genetically-distinct populations within Fusobacterium nucleatum using 16S rRNA gene sequence comparisons

Objective: The aim of the current study was to further investigate the existence of these two groups within F. nucleatum oral and extraoral isolates using partial 16S rRNA gene sequence comparisons. The 16S rRNA gene was used because it is highly conserved and often used for phylogenetic investigations of bacterial species.

Methods: A 490-bp section of the 16S rRNA gene was sequenced from 12 clinical isolates from various oral and extraoral sites used in previous allozyme electrophoretic analyses. The PCR primers were designed from alignments of all available Fusobacterium species Genebank sequences. The sequences obtained were aligned with the Genebank sequences using Clustal X (1.64b) and phylogenetically analysed using the Phylip version 3.6 (alpha 2) package (J. Felsenstein).

Results: All F. nucleatum sequences formed a distinct cluster from other Fusobacterium species, supporting their inclusion into a single species. Most of the relationships between isolates agreed with those obtained using allozyme electrophoresis. There were a number of distinct clusters identified within F. nucleatum, some associated with particular sites of infection. These clusters do not support the proposed subspecies within F. nucleatum, as had also been concluded from the allozyme electrophoresis study.

Conclusion: Although the 16S rRNA gene is highly conserved within F. nucleatum, phylogenetic analysis of a number of clinical isolates using a variable region of the gene have suggested the existence of a number of distinct lineages within the species, perhaps constituting subspecies. These subspecies, however, conflict with those previously described.