First Cultivation of Health-Associated Tannerella sp. HOT-286 (BU063).

Despite significant advances in recent years in culture-independent molecular microbiology methods, the detailed study of individual bacterial species still relies on having pure cultures in the laboratory. Yet, more than a third of the approximately 700 bacterial taxa found in the human oral cavity are as yet uncultivated in vitro. One such taxon, Tannerella sp. HOT-286 (phylotype BU063), is the focus of much interest since it is associated with periodontal health, while Tannerella forsythia, its closest phylogenetic neighbor, is strongly associated with periodontal disease. HOT-286, however, has remained uncultivated despite the efforts of several research groups, spanning over a decade. The aim of this study was to cultivate Tannerella sp. HOT-286. A heavily diluted sample of subgingival plaque was inoculated onto culture plates supplemented with siderophores (pyoverdines-Fe complex or desferricoprogen) or a neat plaque suspension. After 8 d of anaerobic incubation, microcolonies and colonies showing satellitism were passaged onto fresh culture plates cross-streaked with potential helper strains or onto cellulose-acetate membranes placed over lawn cultures of helper strains. Subcultured colonies were identified by 16S rRNA gene sequencing, and purity was confirmed by sequencing 20 clones per library prepared from a single colony. Three colonies of interest (derived from pyoverdines- and plaque-supplemented plates) were identified as Tannerella sp. HOT-286. The isolates were found to be incapable of independent growth, requiring helpers such as Propionibacterium acnes and Prevotella intermedia for stimulation, with best growth on membranes over "helper" lawns. A representative isolate was subjected to phenotypic characterization and found to produce a range of glycosidic and proteolytic enzymes. Further comparison of this novel "periodontal health-associated" taxon with T. forsythia will be valuable in investigating virulence factors of the latter and possible health benefits of the former.

Comparison of the possible protective effect of the salivary pellicle of individuals with and without erosion.

The acquired pellicle adheres to tooth surfaces and has been suggested to provide differing degrees of protection against acidic erosion. This study investigated whether pellicle formed on enamel blocks in patients suffering dietary dental erosion modified the effect of an in vitro simulated dietary challenge, in comparison with pellicle formed on enamel blocks in healthy subjects and to no-pellicle enamel samples. Sixty subjects recruited from dental erosion clinics were compared to healthy age-matched controls. Subjects wore a custom-made maxillary splint holding human enamel blocks for 1 h during which the acquired enamel pellicle was formed. Enamel blocks were removed from the splints and a simulated dietary erosive challenge of 10 min was performed. In addition the challenge was performed on 30 enamel samples without pellicle. Profilometry showed no statistical difference between samples from the erosion subjects with a mean step height of 1.74 µm (SD 0.88) and median roughness (Sa) of 0.39 µm (interquartile range, IQR 0.3-0.56) and the controls with 1.34 µm (SD 0.66) and 0.33 µm (IQR 0.27-0.38), respectively. The control samples without pellicle had Sa of 0.44 µm (IQR 0.36-0.69) and these differences were statistically significant compared to those from the healthy subjects (p = 0.002). Mean (SD) microhardness reduction with a 100-gram load for the erosion group was 113.5 (10) KHN, for healthy subjects was 93 (15.4) KHN and for the enamel samples without pellicle 139.6 (21.8) KHN and all groups were statistically different. The microhardness and roughness data suggested the pellicle influenced erosion under these study conditions.