ABSTRACT: Next Generation Sequencing approaches such as Illumnia HiSeq2000 and 454 were used to study diversity and gene expression profiles in human oral biofilms.
Project description:Within the mouth bacteria are starved of saccharides as their main nutrient source between meals and it is unclear what drives their metabolism. Previously oral in vitro biofilms grown in saliva have shown proteolytic degradation of salivary proteins and increased extracellular proline. Although arginine and glucose have been shown before to have an effect on oral biofilm growth and activity, there is limited evidence for proline. Nuclear magnetic resonance (NMR) spectroscopy was used to identify extracellular metabolites produced by bacteria in oral biofilms grown on hydroxyapatite discs. Biofilms were inoculated with whole mouth saliva and then grown for 7 days using sterilised whole mouth saliva supplemented with proline, arginine and glucose as a growth-medium. Overall proline had a beneficial effect on biofilm growth – with significantly fewer dead bacteria present by biomass and surface area of the biofilms (p <0.05). Where arginine and glucose significantly increased and decreased pH, respectively, the pH of proline supplemented biofilms remained neutral at pH 7.3-7.5. SDS-polyacrylamide gel electrophoresis of the spent saliva from proline and arginine supplemented biofilms showed inhibition of salivary protein degradation of immature biofilms. NMR analysis of the spent saliva revealed that proline supplemented biofilms were metabolically similar to unsupplemented biofilms, but these biofilms actively metabolised proline to 5-aminopentanoate, butyrate and propionate, and actively utilised glycine. This study shows that in a nutrient limited environment, proline has a beneficial effect on in vitro oral biofilms grown from a saliva inoculum.
Project description:An organotypic gingiva model RHG was constructed by immortalized human gingiva cell lines (keratinocytes and fibroblasts) in vitro. The RHG model was exposed to either control medium or one of the three multi-species oral biofilms (commensal, gingivitis and cariogenic) for 24 hours. We used Qiagen RT² Profiler PCR array to analyse the expression of genes which are related to Toll-like receptor signaling pathway from the unexposed (control) and differently exposed (test) keratinocyte layer of the RHG model. Reference for RHG model and oral biofilms culture: J. K. Buskermolen et al., J Dent Res 97 (2018): 201-208. doi: 10.1177/0022034517729998.
Project description:Next generation sequencing was perfomed to identify differentially expressed micro-RNA in HNSCC tissues, normal oral epithelial tissues and CAFs.
Project description:Oral in vitro biofilms were established from human saliva and grown as described in Edlund et al. 2013 (Microbiome, doi:10.1186/2049-2618-1-25). Replicate samples were collected for small molecule- and metatranscriptome analyses at pH 7 (prior to glucose amendment), pH 4.2 (6 hrs after glucose amendment) and pH 5.2 (9 hrs after glucose amendment). Other information: 16S 454-sequencing analyses exist for these communities as well as metagenomics data analyses.
Project description:These biofilms were established from saliva samples as described in Edlund et al. 2013 (Microbiome, doi:10.1186/2049-2618-1-25). After starving the biofilms in minimal medium anaerobiaclly glucose was added to the cultures. Samples for metatranscriptomics and metabolomics were collected in parallel at pH 7 (prior to glucose amendment), pH 4.2 (6 hours after glucose amendment) and pH 5.2 (9 hours after pH amendment). Secreted small molecules were extracted from biofilms (cells were not lysed) at the differnet pH stages.
Project description:Expression of miRNA analysis in human lecukocytes was analysed using Next-generation sequencing (NGS) method in Oral Submucosis Firbrosis condition.
Project description:To combat dental implant-associated infections, there is a need for novel materials which effectively inhibit bacterial biofilm formation. In the present study, a titanium surface functionalization based on the “slippery liquid-infused porous surfaces” (SLIPS) principle was analyzed in an oral flow chamber system. The immobilized liquid layer was stable over 13 days of continuous flow. With increasing flow rates, the surface exhibited a significant reduction in attached biofilm of both the oral initial colonizer Streptococcus oralis and an oral multi-species biofilm composed of S. oralis, Actinomyces naeslundii, Veillonella dispar and Porphyromonas gingivalis. Using single cell force spectroscopy, reduced bacterial adhesion forces on the lubricant layer could be measured. Gene expression patterns in biofilms on SLIPS, on control surfaces and planktonic cultures were also compared. For this purpose, the genome of S. oralis strain ATCC® 9811TM was sequenced using PacBio Sequel technology. Even though biofilm cells showed clear changes in gene expression compared to planktonic cells, no differences could be detected between bacteria on SLIPS and on control surfaces. Therefore, it can be concluded that the ability of liquid-infused titanium to repel biofilms is solely due to weakened bacterial adhesion to the underlying liquid interface.