Osteopontin Reduces Biofilm Formation in a Multi-Species Model of Dental Biofilm

<div><p>Background</p><p>Combating dental biofilm formation is the most effective means for the prevention of caries, one of the most widespread human diseases. Among the chemical supplements to mechanical tooth cleaning procedures, non-bactericidal adjuncts that target the mechanisms of bacterial biofilm formation have gained increasing interest in recent years. Milk proteins, such as lactoferrin, have been shown to interfere with bacterial colonization of saliva-coated surfaces. We here study the effect of bovine milk osteopontin (OPN), a highly phosphorylated whey glycoprotein, on a multispecies <i>in vitro</i> model of dental biofilm. While considerable research effort focuses on the interaction of OPN with mammalian cells, there are no data investigating the influence of OPN on bacterial biofilms.</p><p>Methodology/Principal Findings</p><p>Biofilms consisting of <i>Streptococcus oralis, Actinomyces naeslundii, Streptococcus mitis, Streptococcus downei</i> and <i>Streptococcus sanguinis</i> were grown in a flow cell system that permitted <i>in situ</i> microscopic analysis. Crystal violet staining showed significantly less biofilm formation in the presence of OPN, as compared to biofilms grown without OPN or biofilms grown in the presence of caseinoglycomacropeptide, another phosphorylated milk protein. Confocal microscopy revealed that OPN bound to the surface of bacterial cells and reduced mechanical stability of the biofilms without affecting cell viability. The bacterial composition of the biofilms, determined by fluorescence <i>in situ</i> hybridization, changed considerably in the presence of OPN. In particular, colonization of <i>S. mitis</i>, the best biofilm former in the model, was reduced dramatically.</p><p>Conclusions/Significance</p><p>OPN strongly reduces the amount of biofilm formed in a well-defined laboratory model of acidogenic dental biofilm. If a similar effect can be observed <i>in vivo</i>, OPN might serve as a valuable adjunct to mechanical tooth cleaning procedures.</p>