Development of Biofilm Model for Assessment of Oral Hygiene Products
Objectives: In vitro testing of oral care products is an important and inexpensive tool for assessing safety and efficacy. Current Clinical and Laboratory Standards Institute (CLSI) standards recommend minimum inhibitory concentration (MIC) for antimicrobial susceptibility testing. However, due to the complex nature of oral cavity (hard and soft tissues, multi-species biofilm, salivary constituents, etc.) when MIC’s are used alone there can be large differences between the efficacy and effectiveness. There is a need for development of a clinically relevant model to help narrow the gap between results seen in vitro vs in vivo. Hence, this study is focused on development of an orally relevant multispecies biofilm model and comparing the surface specificity of hydroxyapatite (HA), human dentin/enamel, and polystyrene surfaces.
Methods: ADA-IRB approval was obtained for collection of extracted human teeth and pooled saliva. Commercially available HA (Clarkson Chromatography, PA) were shaped to mimic occlusal surfaces of extracted molars. Tissue treated 24 well plates, HA and teeth were coated in saliva and/or artificial saliva and left to dry. Media, sucrose, and oral bacteria were added in succession to replicate dental plaque progression. After 24H each sample was removed from its respective well and sonicated for biofilm extraction. Total viable counts of bacteria were assessed via CFU/ml and metabolic assays.
Results: Use of pooled human saliva creates a biofilm 1.2x denser than tested formulations of artificial saliva. HA and extracted teeth adhere 10x more CFUs/cm2 than polystyrene. Multispecies biofilm on a clinically relevant surface is comparable to in vivo studies.
Conclusions: This study describes the development of a clinically relevant antimicrobial susceptibility test for antiplaque and antigingivitis products by mimicking the oral environment (relevant surfaces, salivary components and multispecies biofilm). Further analysis of bacterial specificity by RT-PCR will provide in-depth analysis and a standardized in vitro testing model.