Hydroxyapatite-Based 3D Tooth Models for Investigating Spatially Resolved Analysis of Biofilm Formation Dynamics

Experimental oral biofilms have traditionally been cultured on various substrates. Among them, hydroxyapatite (HA), in the form of discs or beads, has been widely used due to its chemical relevance to natural dental tissues. However, these conventional HA substrates fail to replicate the intricate topography of natural teeth, features that significantly influence microbial adhesion and biofilm development. To address this limitation, we developed morphologically accurate HA-based tooth models, including both intact and restored variants. Premolar and molar tooth models were fabricated from patient-derived computed tomography scans using HA powder via cold isostatic pressing and sintering. Restorations (inlays and onlays) were incorporated using permanent dental resin, which was mounted with clinical-grade dental cement. The resulting tooth models were free of visible cracks or macroscopic defects and demonstrated mechanical stability comparable to that of natural human teeth. Results from biofilm experiments using Streptococcus mutans and Candida albicans revealed significantly higher biofilm accumulation on the occlusal surfaces of the mandibular tooth models. In contrast, maxillary tooth models exhibited greater colonization on lateral surfaces. Restored tooth models showed increased biofilm formation on and around inlay and onlay materials. These morphologically accurate tooth models enable spatially resolved analysis of oral biofilms. This approach may enhance our understanding of microbial colonization and biofilm formation mechanisms.