Scanning electron microscopy approach to observe bacterial adhesion to dental surfaces

AIM: To describe the in vitro bacterial adhesion protocol of Streptococcus mutans and Lactobacillus casei on dental surfaces for a qualitative approach by Scanning Electron Microscopy (SEM) observations. A control and Amelogenesis Imperfecta (AI) affected teeth were used to validate the protocol. METHOD DETAILS: Eight teeth were collected and fixed in 10% formalin during 10 days. Crowns were fragmented into 4 parts and kept in the freshly prepared artificial saliva. For the preparation of bacterial suspensions, bacterial strains (S. mutans and L. casei) were incubated in a freshly prepared culture medium. After two successive cultures at 37 degrees C and 3 rinces, bacterial suspensions were prepared in artificial saliva and adjusted to correspond to 10(8) CFU ml(-1). Bacterial adhesion was carried out by sedimentation. Dental fragments were immersed in bacterial suspensions and rinsed with PBS to remove non adherent bacteria. Adherent bacteria were fixed with glutaraldehyde. Finally, teeth samples were dehydrated, coated, dried and observed using high-resolution SEM (JEOL, JSM-5400). RESULTS: SEM observations showed adherence of spheric stuctures, identified as S. mutans and bacilic structures identified as L. casei. CONCLUSION: Adhesion of bacteria could be observed by SEM and depends on the quality of dental mineralized tissues. (C) 2020 The Authors. Published by Elsevier B.V.