Project description:<p>Bacterial metabolism in oral biofilms is comprised of complex networks of nutritional chains and biochemical regulations. These processes involve both intraspecies and interspecies networks as well as interactions with components from host saliva, gingival crevicular fluid, and dietary intake. In a previous paper, a large salivary glycoprotein, mucin MUC5B, was suggested to promote a dental health-related phenotype in the oral type strain of <em>Streptococcus gordonii</em> DL1, by regulating bacterial adhesion and protein expression. In this study, nuclear magnetic resonance-based metabolomics was used to examine the effects on the metabolic output of monospecies compared to dual species early biofilms of two clinical strains of oral commensal bacteria, <em>S. gordonii</em> and <em>Actinomyces naeslundii</em>, in the presence of MUC5B. The presence of <em>S. gordonii</em> increased colonization of <em>A. naeslundii</em> on salivary MUC5B, and both commensals were able to utilize MUC5B as a sole nutrient source during early biofilm formation. The metabolomes suggested that the bacteria were able to release mucin carbohydrates from oligosaccharide side chains as well as amino acids from the protein core. Synergistic effects were also seen in the dual species biofilm metabolome compared to the monospecies, indicating that <em>A. naeslundii</em> and <em>S. gordonii</em> cooperated in the degradation of salivary MUC5B. A better understanding of bacterial interactions and salivary-mediated regulation of early dental biofilm activity is meaningful for understanding oral biofilm physiology and may contribute to the development of future prevention strategies for biofilm-induced oral disease.</p>
Project description:Biofilms are matrix-encased microbial communities that increase the environmental fitness and infectivity of many human pathogens including Vibrio cholerae. Biofilm matrix assembly is essential for biofilm formation and function. Known components of the V. cholerae biofilm matrix are the polysaccharide VPS, matrix proteins RbmA, RbmC, Bap1, and extracellular DNA, but the majority of the protein composition is uncharacterized. This study comprehensively analyzed the biofilm matrix proteome and revealed the presence of outer membrane proteins (OMPs). Outer membrane vesicles (OMVs) were also present in the V. cholerae biofilm matrix and were associated with OMPs and many biofilm matrix proteins suggesting that they participate in biofilm matrix assembly. Consistent with this, OMVs had the capability to alter biofilm structural properties depending on their composition. OmpU was the most prevalent OMP in the matrix, and its absence altered biofilm architecture by increasing VPS production. Using single-cell force spectroscopy, we further showed that OmpU, the matrix proteins RbmA, RbmC, and Bap1, and VPS contribute to cell-surface adhesion forces, which are critical for biofilm formation. Our findings provide new insights into the molecular mechanisms underlying biofilm matrix assembly in V. cholerae, which may open up new opportunities to develop inhibitors that specifically alter biofilm matrix properties and, thus, affect either the environmental survival or pathogenesis of Vibrio cholerae.
Project description:Biofilms are matrix-encased microbial communities that increase the environmental fitness and infectivity of many human pathogens including Vibrio cholerae. Biofilm matrix assembly is essential for biofilm formation and function. Known components of the V. cholerae biofilm matrix are the polysaccharide VPS, matrix proteins RbmA, RbmC, Bap1, and extracellular DNA, but the majority of the protein composition is uncharacterized. This study comprehensively analyzed the biofilm matrix proteome and revealed the presence of outer membrane proteins (OMPs). Outer membrane vesicles (OMVs) were also present in the V. cholerae biofilm matrix and were associated with OMPs and many biofilm matrix proteins suggesting that they participate in biofilm matrix assembly. Consistent with this, OMVs had the capability to alter biofilm structural properties depending on their composition. OmpU was the most prevalent OMP in the matrix, and its absence altered biofilm architecture by increasing VPS production. Single-cell force spectroscopy revealed that proteins critical for biofilm formation, OmpU, the matrix proteins RbmA, RbmC, Bap1, and VPS, contribute to cell-surface adhesion forces at differing efficiency, with VPS showing the highest efficiency whereas Bap1 showing the lowest efficiency. Our findings provide new insights into the molecular mechanisms underlying biofilm matrix assembly in V. cholerae, which may provide new opportunities to develop inhibitors that specifically alter biofilm matrix properties and, thus, affect either the environmental survival or pathogenesis of V. cholerae.
Project description:This study investigate how soluble by-products derived from a four-species endodontic biofilm model impacts upon the viability, transcriptomic profile, and inflammatory response of human dental pulp stem cells (DPSCs). The sterile-filtered supernatant was extracted from an established interkingdom endodontic biofilm model comprising Streptococcus gordonii, Fusobacterium nucleatum, Porphyromonas gingivalis, and Candida albicans. DPSCs were exposed to the microbial biofilm supernatant (BSN) for 4 and 24 hours. Cellular responses were evaluated via MTT, CCK-8, LDH assays, and Annexin V/PI staining. Whole transcriptomic sequencing was performed to assess gene expression dynamics, with GO and KEGG pathway enrichment analyses. IL6 and IL8 expression was validated by qPCR and ELISA.It was found that BSN significantly suppressed DPSC metabolic activity without inducing apoptosis or necrosis. RNA-seq revealed 723 significantly differentially expressed genes at 4 h and 1667 at 24 h. Early responses were dominated by upregulation of inflammatory mediators (e.g., CXCL8, IL6, TNFAIP2), with enrichment of TNF, NF-κB, and JAK-STAT signalling pathways. At 24 h, the expression profile shifted toward redox regulation and metabolic suppression, including downregulation of glycolytic and purine metabolism pathways. IL6 and IL8 expression was markedly increased at both transcript and protein levels. Indeed, soluble factors produced by a biofilm model representative of deep caries and carious pulp exposures are capable of reprogramming DPSC function in a time-dependent manner. The soluble factors initiate a biphasic response characterized by early immune activation and later metabolic adaptation. The findings underscore the potential role of biofilm-derived products associated with deep caries and carious pulp exposures in compromising DPSC function. In addition, they emphasise the need for the development of vital pulp therapy strategies that not only neutralise microorganisms but also their secreted products.