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In situ identification of streptococci and other bacteria in initial dental biofilm by confocal laser scanning microscopy and fluorescence in situ hybridization.

Journal article
Authors Irene Dige
Holger Nilsson
Mogens Kilian
Bente Nyvad
Published in European journal of oral sciences
Volume 115
Issue 6
Pages 459-67
ISSN 0909-8836
Publication year 2007
Published at Institute of Neuroscience and Physiology, Department of Physiology
Pages 459-67
Language en
Keywords Adult, Biofilms, DNA Probes, genetics, Dental Plaque, microbiology, Female, Humans, In Situ Hybridization, Fluorescence, methods, Male, Microscopy, Confocal, methods, Oligonucleotide Probes, diagnostic use, RNA, Ribosomal, 16S, analysis, Streptococcus, genetics, isolation & purification, Time Factors
Subject categories Medical and Health Sciences


Confocal laser scanning microscopy (CLSM) has been employed as a method for studying intact natural biofilm. When combined with fluorescence in situ hybridization (FISH) it is possible to analyze spatial relationships and changes of specific members of microbial populations over time. The aim of this study was to perform a systematic description of the pattern of initial dental biofilm formation by applying 16S rRNA-targeted oligonucleotide probes to the identification of streptococci and other bacteria, and to evaluate the usefulness of the combination of CLSM and FISH for structural studies of bacterial populations in dental biofilm. Biofilms were collected on standardized glass slabs mounted in intra-oral appliances and worn by 10 individuals for 6, 12, 24 or 48 h. After intra-oral exposure the biofilms were labelled with probes against either streptococci (STR405) or all bacteria (EUB338) and analysed by CLSM. The current approach of using FISH techniques enabled differentiation of streptococci from other bacteria and determination of their spatio-temporal organization. The presence of chimney-like multilayered microcolonies with different microbial compositions demonstrated by this methodology provided information supplementary to our previous knowledge obtained by classical electron microscopic methods and increased our understanding of the structure of developing biofilms.

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