Treponema denticola is considered an important oral pathogen in the development and progression of periodontal diseases. In the present study, the mechanisms of recognition and activation of murine macrophages by T. denticola and its major outer sheath protein (MSP) and lipooligosaccharide (LOS or glycolipid) were investigated. T. denticola cells and the MSP induced innate immune responses through TLR2-MyD88, whereas LOS induced a macrophage response through TLR4-MyD88. The presence of gamma interferon (IFN-␥), or of high numbers of T. denticola, circumvented the requirement for TLR2 for the macrophage response to T. denticola, although the response was still dependent on MyD88. In contrast, synergy with IFN-␥ did not alter the TLR dependence of the response to the T. denticola surface components LOS and MSP, despite enhanced sensitivity. These data suggest that although there is flexibility in the requirements for recognition of T. denticola cells (TLR2 dependent or independent), MyD88 is a requirement for the downstream signaling events that lead to inflammation. We also demonstrate that both outer sheath molecules LOS and MSP induce macrophage tolerance to further stimulation with enterobacterial lipopolysaccharide. Tolerance induced by T. denticola components during mixed infections may represent a general mechanism through which bacteria evade clearance.

Coagreggation of Treponema denticola with either Porphyromonas gingivalis or Fusobacterium nucleatum was characterized and the role of the major outer sheath protein (MSP) in the coaggregation process of these bacteria was evaluated. The MSP of T. denticola was found to be able to bind to P. gingivalis and F. nucleatum cells and this binding could be inhibited by MSP in a concentration-dependent manner. While sodium dodecyl sulfate polyacrylamide gel electrophoresis and Periodic acid-Schiff (PAS) staining of MSP revealed that it is a glycoprotein, monosaccharide analysis showed that MSP contains: Glc (44.4), Gal (20.4%) GlcN (1.3%), GalN (31.6%) and Fuc (9.2%). Peptide N-glycosidase F deglycosylation of MSP was found to inhibit its binding to F. nucleatum but not to P. gingivalis cells. Sugar-binding studies showed that the requirements for the binding of both T. denticola and MSP to F. nucleatum cells are similar to those of the F. nucleatum galactose-binding lectin. These data suggest that MSP acts as an adhesin during the coaggregation process of T. denticola with P. gingivalis and F. nucleatum through its protein and carbohydrate moieties, respectively.