In situ analysis of multispecies biofilm formation on customized titanium surfaces

Many studies to identify surfaces that enhance the incorporation of dental implants into bone and soft-tissue have been undertaken previously. However, to succeed in the clinical situation, an implant surface must not support development of microbial biofilms with a pathogenic potential. As a first step in investigating this, we used two-species and three-species biofilm models with 16S ribosomal RNA fluorescence in situ hybridization and confocal laser scanning microscopy to examine the effect of surface characteristics on biofilm formation by species that can colonize titanium implants in vivo: Streptococcus sanguinis, Actinomyces naeslundii and Lactobacillus salivarius. Surfaces blasted with Al2O3 (S-a = 1.0-2.0 mu m) showed a seven-fold higher bacterial adhesion after 2 h than turned surfaces (S-a = 0.18 mu m) whereas porous surfaces, generated by anodic oxidation (S-a = 0.4 mu m), showed four-fold greater adhesion than turned surfaces. Hence, increased roughness promoted adhesion, most likely through protection of bacteria from shear forces. Chemical modification of the blasted and oxidized surfaces by incorporation of Ca2+ ions reduced adhesion compared with the corresponding non-modified surfaces. After 14 h, biofilm growth occurred in the three-species model but not in the two-species consortium (containing S. sanguinis and A. naeslundii only). The biofilm biovolume on all surfaces was similar, suggesting that the influence of surface characteristics on adhesion was compensated for by biofilm development.