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... Show moreThe 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. Show less
Ruellan, K.; Frijns, J.H.M.; Bloemberg, G.V.; Hautefort, C.; Abbeele, T. van den; Lamers, G.E.M.; ... ; Kania, R.E. 2010
Objective: To investigate the formation of bacterial biofilms on the surface of the electrode array of cochlear implants (CI) explanted because of device failure, without evidence of infection, by... Show moreObjective: To investigate the formation of bacterial biofilms on the surface of the electrode array of cochlear implants (CI) explanted because of device failure, without evidence of infection, by use of scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Study Design: Prospective study. Setting: Patients from 2 tertiary-care referral centers. Patients and Methods: CIs were explanted from 9 patients because of device failure. Specimens were immediately snapfrozen in cold isopenthane, stored at -80 degrees and examined with SEM and CLSM by 3 investigators. Main Outcome Measure: Presence of bacterial biofilm ascertained by SEM and CSLM. Results: One specimen showed the formation of a bacterial biofilm on the middle ear part of the electrode array. No biofilm formation was found in the inner-ear part of electrode arrays. In the middle-ear part of the electrode array, a cylindrical cover of human muscular tissue was seen plugging the cochleostomy. Conclusion: This is the first study demonstrating that bacterial biofilms may exist on the surface of the electrode array of CIs explanted because of device failure but not infection. We found 1 case of biofilm formation in 9 explanted CIs. Further studies with larger series of CIs are required to investigate biofilm formation on the surface of CI electrode arrays to address both the pathophysiology of bacterial biofilms and prevention of device-related infections in CI patients. Show less
Kania, R.E.; Lamers, G.E.M.; Laar, N. van de; Dijkhuizen, M.; Lagendijk, E.; Tran Ba Huy, P.; ... ; Bloemberg, G.V. 2010
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... Show moreThe 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. Show less
Objectives: To demonstrate the presence of mucosal biofilm in adenoid tissue using double staining for visualization of both the bacterial matrix and the bacterial cells. To identify bacterial... Show moreObjectives: To demonstrate the presence of mucosal biofilm in adenoid tissue using double staining for visualization of both the bacterial matrix and the bacterial cells. To identify bacterial species present on the surface of the studied adenoids.Study Design: Prospective study.Methods: A total of 39 specimens of adenoidectomy were removed from children with chronic and/or recurrent otitis media. The specimens were prepared for light microscopy using Gram staining, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Double staining was performed with CLSM to visualize both the bacteria and the glycocalyx matrix. Nine adenoids on which bacterial biofilms were visualized with CLSM were used for identification of bacterial species by 16S-DNA polymerase chain reaction (PCR) amplification and homology analysis.Results: Of the 39 adenoids investigated, 22 (54%) showed evidence of mucosal biofilms. Gram staining, SEM and CLSM showed the presence of bacterial cells, organized in bacterial microcolonies. CLSM with double staining demonstrated mucosal biofilms by showing the presence of both bacteria and the glycocalyx. The use of 16S-DNA polymerase chain reaction (PCR) amplification and subsequent sequence analyses identified the presence of Corynebacterium argentoratense, Streptococcussalivarius, Micrococcusluteus, and Staphylococcus aureus.Conclusions: This study demonstrates that adenoid tissue in children with chronic or/and recurrent otitis media contains mucosal biofilms in 54% of the cases. The existence of living bacteria has been demonstrated. Further studies are required to describe the panel of bacteria that can be harbored in the biofilms present in adenoids and the mechanisms involved in the physiopathology of otitis prone children. Show less
