Visualization and characterization of Enterococcus faecalis biofilm structure in bovine dentin using 2D and 3D microscop
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ORIGINAL PAPER
Visualization and characterization of Enterococcus faecalis biofilm structure in bovine dentin using 2D and 3D microscopic techniques Ali Keleş1 · Cangül Keskin1 · Melis Kalkan2 · Yusuf Yakupoğulları3 · Mehmet Gül4 · Hikmet Aydemir1 · Fikrettin Şahin2 Received: 24 April 2020 / Revised: 11 August 2020 / Accepted: 2 September 2020 © Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract Bacterial biofilms are related to various dental and periodontal infectious diseases, and the characterization of this biological structure with micro-computed tomography (micro-CT) may offer valuable information for clinical and research applications. In this study, we aimed to develop a model to visualize three-dimensionally the biofilm structure on dentin using micro-CT. Dentin blocks were prepared and incubated in tryptic soy broth with Enterococcus faecalis (ATCC 29212). The control group did not receive any staining procedure, while groups 1 and 2 were stained with 100% and 50% barium sulfate, respectively. Transmission electron microscopy (TEM) and confocal laser scanning microscopy (CLSM) were used to detect biofilm formation, barium sulfate penetration, and microbial cell density in the biofilm. Micro-computed tomography (micro-CT) (SkyScan 1172, Bruker Co., Belgium) was used to visualize biofilm formation on the dentin blocks. Biofilm thicknesses were measured from 10 different locations on the specimen surfaces, using CTAn v.1.14.4 software. Obtained data were statistically analyzed using Kruskal–Wallis and Dunn’s tests. TEM photomicrographs showed that barium sulfate could penetrate the biofilm structure. CLSM analysis showed that viable and total cell densities were similar between the control and barium sulfate-treated groups (P > 0.05), indicating barium sulfate had no significant influence on cell density. In barium sulfate-treated blocks, biofilm could be discriminated from the dentin, and its thickness could be measured with micro-CT. This study showed that bacterial biofilm on dentin could be characterized by micro-CT after barium sulfate staining without causing any significant side effect on viable and total cell densities. Keywords Biofilm · Dentin · Micro-computed tomography · Transmission electron microscopy
Introduction
Communicated by Erko stackebrandt.
Biofilms are complex polymicrobial structures, which are composed of microbial cells enclosed in a self-produced extracellular polymeric matrix adherent to a surface
* Cangül Keskin [email protected]; [email protected]
Fikrettin Şahin [email protected]
Ali Keleş [email protected]
1
Melis Kalkan [email protected]
Department of Endodontics, Faculty of Dentistry, Ondokuz Mayıs University Samsun, Samsun, Turkey
2
Yusuf Yakupoğulları [email protected]
Department of Genetic and Bioengineering, Faculty of Engineering and Architecture, Yeditepe University, Istanbul, Turkey
3
Mehmet Gül [email protected]
Department of Medical Microbiology, Faculty of Medicine, İ
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