Microbiological diversity of peri-implantitis biofilm by Sanger sequencing

Background and objective: To examine the microbial diversity associated with implants with or without peri-implantitis and to identify differences between the bacterial communities in these clinical conditions. Material and methods: Twenty subjects were assigned to a Control group consisting of subjects with healthy implants and a Test group consisting of subjects with peri-implantitis sites, as well as a healthy implant site (n = 10/group). In the Test group, subgingival biofilm samples were taken from the deepest sites of the diseased implant. In both groups, samples were collected from one site around a healthy implant. DNA was extracted and the 16S rRNA gene was amplified and sequenced, and the phylotypes were identified by comparison with known 16S rRNA sequences. Results: The phylogenetic identity of 1387 16S rRNA gene clones was determined. Healthy implants demonstrated higher proportions of Actinomyces, Atopobium, Gemella, Kingella and Rothia and lower levels of Campylobacter, Desulfobulbus, Dialister, Eubacterium, Filifactor, Mitsukella, Porphyromonas and Pseudoramibacter (Mann-Whitney U-test; P < 0.05). Fusobacterium nucleatum, Dialister invisus, Streptococcus sp. human oral taxon (HOT) 064, Filifactor alocis and Mitsuokella sp. HOT 131 presented a higher mean proportion, while Veillonella dispar, Actinomyces meyeri, Granulicatella adiacens showed lower mean proportions in the peri-implantitis sites when compared with healthy implants in both the Control and Test groups (Mann-Whitney U-test; P < 0.05). Conclusion: Marked differences were observed in the composition of the subgingival biofilm between healthy and diseased implants. The biofilm associated with peri-implantitis harbored more pathogenic bacterial species from the orange complex and other unusual putative pathogens, such as F. alocis, D. invisus and Mitsuokella sp. HOT 131.