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Abstract
The aim of this study was to demonstrate the presence of yeast and bacterial biofilms on the surface of tracheoesophageal voice prostheses (TVPs) by a double-staining technique with confocal laser scanning microscopy (CLSM). Biofilms of 12 removed TVPs were visualized by scanning electron microscopy, then stained with ConA-FITC and propidium iodide for CLSM. Microbial identification was by partial 16S rRNA gene analysis and ITS-2 sequence analysis. Microbial biofilms on the TVPs consisted of bacteria and filamentous cells. Bacterial cells were attached to the filamentous and unicellular yeast cells, thus forming a network. Sequence analyses of six voice prostheses identified the presence of a variety of bacterial and yeast species. In vivo studies showed that Klebsiella oxytoca and Micrococcus luteus efficiently attached to Candida albicans. CLSM with double fluorescence staining can be used to demonstrate biofilm formations composed of a mixture of yeast and bacterial cells on the surface of TVPs.
Acknowledgements
The authors are grateful to the following attending otorhinolaryngologist head and neck surgeons (Drs Baatenburg de Jong, Frijns, Brenkman, Otten, Jansen, Peek, Rus, Bonnet, Schmitt) who helped collect specimens. They are also grateful to Marc Arenthorst for help in isolating fungal DNA. This study was supported by an unrestricted research grant from Sanofi Synthelabo Aventis (Prix de Recherche Perspective en ORL 2004). The authors are also grateful to the NRJ foundation (Institut de France) for its financial support.
