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Fusobacteria are common obligately anaerobic Gram-negative bacteria of the oral cavity that may act as a bridge between early and late colonizing bacteria in dental plaque and have a role in oral and extra-oral infections.  Fusobacterium nucleatum  has a crucial role in oral biofilm structure and ecology, as revealed in experimental and clinical biofilm models. The aim of this study was to investigate the impact of various  Fusobacterium  species on  in vitro  biofilm formation and structure in three different oral biofilm models namely a supragingival, a supragingival “feeding”, and a subgingival biofilm model. The standard six-species supragingival and “feeding” biofilm models employed contained  Actinomyces oris ,  Candida albicans ,  Streptococcus mutans ,  Streptococcus oralis ,  Veillonella dispar , and  Fusobacterium  sp. The subgingival biofilm model contained 10 species ( A. oris ,  Campylobacter rectus ,  F. nucleatum  ssp.  nucleatum ,  Porphyromonas gingivalis ,  Prevotella intermedia ,  Streptococcus anginosus ,  S. oralis ,  Tannerella forsythia ,  Treponema denticola , and  V. dispar ). Six different  Fusobacterium  species or subspecies, respectively, were tested namely  F. nucleatum  ssp.  fusiforme ,  F. nucleatum  ssp.  nucleatum ,  F. nucleatum  ssp.  polymorphum ,  F. nucleatum  ssp.  vincentii ,  F. naviforme , and  F. periodonticum ). Biofilms were grown anaerobically on hydroxyapatite disks in 24-well culture dishes. After 64 h, biofilms were either harvested and quantified by culture analysis or proceeded to fluorescent  in situ  hybridization (FISH) and confocal laser scanning microscopy (CLSM). All  Fusobacterium  species tested established well in the biofilms, with CFUs ranging from 1.4E+04 ( F. nucleatum  ssp.  fusiforme ) to 5.6E+06 ( F. nucleatum  ssp.  nucleatum ). The presence of specific  Fusobacterium  sp./ssp. induced a significant decrease in  C. albicans  levels in the supragingival model and in  V. dispar  levels in the “feeding” supragingival model. In the subgingival model, the counts of  A. oris ,  S. oralis ,  P. intermedia ,  P. gingivalis , and  C. rectus  significantly decreased in the presence of specific  Fusobacterium  sp./ssp. Collectively, this study showed variations in the growing capacities of different fusobacteria within biofilms, affecting the growth of surrounding species and potentially the biofilm architecture. Hence, clinical or experimental studies need to differentiate between  Fusobacterium  sp./ssp., as their biological properties may well vary.

Fusobacterium Species and Subspecies Differentially Affect the Composition and Architecture of Supra- and Subgingival Biofilms Models