Abstract
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.
Highlights
The disposition of biofilms, which are composed of dynamic substratum-attached microbial networks, is an everyday phenomenon in nature (Hall-Stoodley et al, 2004; Marsh, 2005)
Levels of F. nucleatum ssp. vincenti, F. nucleatum ssp. polymorphum, F. naviforme, as well as that of F. periodonticum were significantly lower in the biofilm than the F. nucleatum ssp. nucleatum control, which is used in the standard biofilm
Box plots represent the colony forming units (CFU) determined by selective agar plating, while horizontal lines indicate their median values
Summary
The disposition of biofilms, which are composed of dynamic substratum-attached microbial networks, is an everyday phenomenon in nature (Hall-Stoodley et al, 2004; Marsh, 2005). More than 700 different bacterial species embedded in extracellular polysaccharide-affluent matrix account for the formation of the multispecies oral communities (Foster and Kolenbrander, 2004; Keijser et al, 2008). The latter encounter 1,000-times more resistance to antimicrobial agents; host immunity; nutrient restriction; alternating oxygen-, nitrogen-, and carbon dioxide supply; and abrupt temperature fluctuation (Davies, 2003; Welin-Neilands and Svensater, 2007; Marsh and Devine, 2011). Streptococci are found to prevail the first 6–48 h, whereas F. nucleatummediated coaggregation takes place after the first 48 h of oral biofilm formation (Dige et al, 2007)
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