A Novel Clinically Relevant Multi-Species Biofilm Model of Endodontic Infections

Objectives: Background: The strategies used to evaluate new treatment modalities are usually small scale in vitro studies employing single or dual-species biofilm models. Because of substantial variation in experimental parameters and models, it is difficult to compare results between studies and to translate bench discovery to bedside application.

Aim: to develop a reproducible and clinically relevant complex multi-species biofilm model of root canal infections.
Methods: Methods: Species were selected based on literature search; i.e., taxa frequently reported in clinical studies. We anaerobically grew suspensions of Streptococcus sanguinis (ATCC #10556), Actinomyces viscosus (ATCC #43146), Fusobacterium nucleatum (ATCC #25586), Parvimonas micra (ATCC #33270) and Prevotella nigrescens (ATCC #33563). Their co-occurrence was explored in silico using Correlation Network Analysis (CNA) to analyze an existing collection 298 root canal samples. Interspecies relationships were assessed in vitro using a 5X5 interaction plate experiment by using growth inhibition spot diffusion assay, where zones of inhibition were recorded. For biofilm development taxa were anaerobically grown and mixed suspensions of the five selected species identified prepared for inoculation in extracted human premolar teeth for 7 and 14 days. Then, the effectiveness of different laser technologies (EdgePro™, EdgeEndo®) in biofilm removal were assessed and compared with that of the standard of care. The speciespresence, abundance and architecture were determined using microbial cultivation, SEM and checkerboard DNA-DNAhybridization.
Results: Results: CNA and growth inhibition spot diffusion assay experiments confirmed the frequent co-occurrence of the selected species the absence of negative relationships among them. The complexity and composition of the biofilms developed on extracted teeth were confirmed. Certain laser technologies presented promising results regarding biofilm removal.
Conclusions: Conclusions: A new, complex and clinically relevant biofilm model for root canal infections was developed. It demonstrated to be a useful tool to evaluate new strategies for the treatment of root canal infections.