Evaluating models and assessment techniques for understanding oral biofilm complexity

Abstract

Oral biofilms are three‐dimensional (3D) complex entities initiating dental diseases and have been evaluated extensively in the scientific literature using several biofilm models and assessment techniques. The list of biofilm models and assessment techniques may overwhelm a novice biofilm researcher. This narrative review aims to summarize the existing literature on biofilm models and assessment techniques, providing additional information on selecting an appropriate model and corresponding assessment techniques, which may be useful as a guide to the beginner biofilm investigator and as a refresher to experienced researchers. The review addresses previously established 2D models, outlining their advantages and limitations based on the growth environment, availability of nutrients, and the number of bacterial species, while also exploring novel 3D biofilm models. The growth of biofilms on clinically relevant 3D models, particularly melt electrowritten fibrous scaffolds, is discussed with a specific focus that has not been previously reported. Relevant studies on validated oral microcosm models that have recently gaining prominence are summarized. The review analyses the advantages and limitations of biofilm assessment methods, including colony forming unit culture, crystal violet, 2,3‐bis‐(2‐methoxy‐4‐nitro‐5‐sulfophenyl)‐2H‐tetrazolium‐5‐carboxanilide inner salt assays, confocal microscopy, fluorescence in situ hybridization, scanning electron microscopy, quantitative polymerase chain reaction, and next‐generation sequencing. The use of more complex models with advanced assessment methodologies, subject to the availability of equipment/facilities, may help in developing clinically relevant biofilms and answering appropriate research questions.

Rhamnolipid 89 Biosurfactant Is Effective against Streptococcus oralis Biofilm and Preserves Osteoblast Behavior: Perspectives in Dental Implantology

“…Biofilm formation progress by co-aggregation of a number of different additional species, depending on the surface properties, nutrient quality and availability, oxygen levels and microbial interactions including intra and inter-regnum interactions [ 53 ]. Although polymicrobial in-vitro models are reported in the literature [ 53 , 54 , 55 ], major technical limitations occur in mimicking the complexity of the peri-implant microbiota using reliable and reproducible in vitro biofilm models, encompassing the stringent culturing conditions of most of the oral microorganisms [ 56 , 57 ] and moving from two-dimensional to novel three-dimensional biofilm models [ 58 ]. Overall, further studies are needed to evaluate the long-term effectiveness and safety of R89BS coating in more complex environments, in in vivo conditions and finally in clinical settings.…”

Biofilm Formation on Hybrid, Resin-Based CAD/CAM Materials for Indirect Restorations: A Comprehensive Review

“…Attempting to imitate oral conditions, most in vitro studies incorporate in their microbiological protocol the immersion of samples in mucin containing artificial saliva or whole mouth saliva, secreted from a volunteer, to form the acquired pellicle. Colony-forming unit counting (CFU/mL), combined with SEM investigations and confocal laser scanning microscopy (CLSM), is used to perform qualitative and quantitative evaluations of bacterial formation [106]. SEM and CLSM have limitations, including the high cost and complexity of their protocols, the inability of CLSM to discriminate strains, the inability of SEM to discriminate live and dead bacteria, and the fact that only a specific selected area of the substrate may be evaluated [107].…”