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:All organisms throughout the tree of life sense and respond to their surface environments. To discriminate from among mucosal surface environmental cues, we grew Streptococcus gordonii biofilms over night at 37C on surfaces coated with the salivary mucin MUC5B, or a low density protein fraction derived from human saliva.
Project description:To investigate the effect on LL-2 conditional medium on bone marrow-derived macrophages, equal volum of LL-2 conditional medium and fresh RPMI 1640 was used to culture bone marrow-derived macrophages for 24 hours. The cells were then lysed for RNA-seq.
Project description:Saliva based diagnostics is a rapidly evolving field due to the large potential of saliva and the simple sample collection. A systematic comparison of IgG antibody profiles in saliva and plasma is currently lacking in scientific literature. Our hypothesis is that IgG profiles are equal in blood and saliva. By showing the equality of the profiles and relative IgG antigenic reactivities towards proteins and peptides we provide evidence that plasma IgG reactivities can be inferred from saliva IgG reactivities. IgG antibodies were isolated from human saliva and plasma samples. The reactivities of IgG isolates were analysed on peptide microarrays displaying linear epitopes of EBV (EBNA1 protein) and HBV (Large envelope protein) virus. Peptide arrays were printed by JPT Peptide Technologies (Berlin, Germany). We show high similarity of saliva and plasma IgG profiles on these two platforms and argue for generalisation from this subset to the whole immunological IgG antibody profile.
Project description:Saliva based diagnostics is a rapidly evolving field due to the large potential of saliva and the simple sample collection. A systematic comparison of IgG antibody profiles in saliva and plasma is currently lacking in scientific literature. Our hypothesis is that IgG profiles are equal in blood and saliva. By showing the equality of the profiles and relative IgG antigenic reactivities towards proteins and peptides we provide evidence that plasma IgG reactivities can be inferred from saliva IgG reactivities. IgG antibodies were isolated from human saliva and plasma samples. The reactivities of IgG isolates were analysed on peptide microarrays displaying linear epitopes of EBV (EBNA1 protein) and HBV (Large envelope protein) virus. Peptide arrays were printed by JPT Peptide Technologies (Berlin, Germany). We show high similarity of saliva and plasma IgG profiles on these two platforms and argue for generalisation from this subset to the whole immunological IgG antibody profile.
Project description:Secreted antimicrobial peptides (AMPs) are an important part of the human innate immune system and prevent local and systemic infections by inhibiting bacterial growth in a concentration dependent manner. In the respiratory tract, the cationic peptide LL-37 is one of the most abundant AMPs and capable of building pore complexes in usually negatively charged bacterial membranes, leading to destruction of bacteria. However, adaptation mechanisms of several pathogens to LL-37 are already described and are known to weaken the antimicrobial effect of the AMP, for instance, by repulsion, export or degradation of the peptide. This study examines proteome wide changes in Streptococcus pneumoniae D39, the leading cause of bacterial pneumonia, in response to physiological concentrations of LL-37 by high resolution mass spectrometry. Our data indicate that pneumococci may use some of the known adaptation mechanisms to reduce the effect of LL-37 on their physiology, too. Additionally, several proteins seem to be involved in resistance to AMPs which have not been related to this process before, such as the teichoic acid flippase TacF (SPD_1128). Understanding colonization and infection relevant adaptations of the pneumococcus to AMPs, especially LL-37, could finally uncover new drug targets to weaken the burden of this widespread pathogen.
Project description:The microbiota plays a crucial role in protecting plants from pests and pathogens. The protection provided by the microbiota constitutes not just the plant’s first line of defense, but possibly its most potent one, as experimental disruptions to the microbiota cause plants to succumb to otherwise asymptomatic infections. To understand how microbial plant defense is deployed, we applied a complex and tractable plant-soil-microbiome microcosm. This system, consisting of Arabidopsis plants and a 150-member bacterial synthetic community, provides a platform for the discovery of novel bacterial plant-beneficial traits, under a realistically complex microbial community context. To identify which components of the plant microbiota are critical for plant defense, we deconstructed this microcosm top-down, removing different microbial groups from the community to examine their protective effect on the plant when challenged with the leaf pathogen Pseudomonas syringae. This process of community deconstruction revealed a critical role for the genus Bacillus in protecting the plant from infection. Using plant RNA-seq and bacterial co-culturing experiments, we demonstrated that Bacillus-provided plant protection is independent of plant immune system activation. We also show that the level of plant protection is strongly dependent on the diversity of the protective inoculum. We show that deconstructing the microbiome top-down is a powerful tool for identifying and prioritizing microbial taxa with specific functions within it.
Project description:The objective of the study was to evaluate transcriptional response of endotoxin-stimulated human monocytic cells in presence or absence of host defense peptide LL-37 at low physiologically relevant concentrations. Human monocytic cells THP-1 were stimulated with LPS (10ng/ml) in presence or absence of LL-37 (5 ug/ml), as well as with the peptide alone, for 4 hours. The trends of LPS-induced altered gene expression in presence of the peptide were further validated by quantitative real-time PCR.