Project description:Multiple levels of interkingdom signaling have been implicated in maintaining the ecological balance between Candida albicans and commensal streptococci to assure a state of oral health. To better understand the molecular mechanisms involved in the initial streptococcal response to the presence of C. albicans that can initiate oral surface colonization and biofilm formation, hypha-forming cells were incubated with Streptococcus gordonii cells for 30 minutes to assess the streptococcal transcriptome response. A genome wide microarray analysis and quantitative PCR validation of S. gordonii transcripts identified a number of genes, the majority of which were involved in metabolic functions, that were differentially expressed in the presence of hyphae. The fruR, fruB and fruA genes encoding the transcriptional regulator, fructose-1-phosphate kinase, and fructose-specific permease, respectively, of the phosphoenolpyruvate-dependent fructose phosphotransferase system, were consistently up-regulated. An S. gordonii mutant in which these genes were deleted by allelic replacement, formed an architecturally-distinct, less robust biofilm with C. albicans than did parental strain cells. Complementing the mutant with plasmid borne fruR, fruB and fruA genes caused phenotype reversion, indicating that the genes in this operon played a role in dual species biofilm formation. This genome wide analysis of the S. gordonii transcriptional response to C. albicans has identified several genes that have potential roles in interkingdom signaling and responses.
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:Periodontal diseases are one of the most common human maladies and appear to be caused by the interaction of proximal pathogens such as Porphyromonas gingivalis but only as part of the polymicrobial community known as dental plaque. Streptococcus gordonii is an early colonizing oral organism that binds to oral surfaces and provides adherence for organisms such as P. gingivalis. Together P. gingivalis and S. gordonii form one of the simplest models of potentially pathogenic dental plaque. We used RNA sequencing to monitor the transcriptome of P. gingivalis over time in a biofilm model both in the presence and absence of S. gordonii. Samples were taken at 5, 30, 120, 240, and 360 minutes after shifing from planktonic to sessile conditions and growth media to PBS. When compared to planktonic cells increased transcripts were found for stress, amino acid catabolism, and comeptence and decreased transcripts for DNA replication. The presence of S. gordonii resulted in fewer changes from planktonic cells implying physiological support to Pl gingivalis making the transition from planktonic to sessile easier.
Project description:Recent studies have shown phenotypic and metabolic heterogeneity in related species including Streptococcus oralis, a typical oral commensal bacterium, Streptococcus mutans, a cariogenic bacterium, and Streptococcus gordonii, which functions as an accessory pathogen in periodontopathic biofilm. In this study, metabolites characteristically contained in the saliva of individuals with good oral hygiene were determined, after which the effects of an identified prebiotic candidate, D-tagatose, on phenotype, gene expression, and metabolic profiles of those three key bacterial species were investigated. Examinations of the saliva metabolome of 18 systemically healthy volunteers identified salivary D-tagatose as associated with lower dental biofilm abundance in the oral cavity (Spearman’s correlation coefficient; r = -0.603, p = 0.008), then the effects of D-tagatose on oral streptococci were analyzed in vitro. In chemically defined medium (CDM) containing D-tagatose as the sole carbohydrate source, S. mutans and S. gordonii each showed negligible biofilm formation, whereas significant biofilms were formed in cultures of S. oralis. Furthermore, even in the presence of glucose, S. mutans and S. gordonii showed growth suppression and decreases in the final viable cell count in a D-tagatose concentration-dependent manner. In contrast, no inhibitory effects of D-tagatose on the growth of S. oralis were observed. To investigate species-specific inhibition by D-tagatose, the metabolomic profiles of D-tagatose-treated S. mutans, S. gordonii, and S. oralis cells were examined. The intracellular amounts of pyruvate-derived amino acids in S. mutans and S. gordonii, but not in S. oralis, such as branched-chain amino acids and alanine, tended to decrease in the presence of D-tagatose. This phenomenon indicates that D-tagatose inhibits growth of those bacteria by affecting glycolysis and its downstream metabolism. In conclusion, the present study provides evidence that D-tagatose is abundant in saliva of individuals with good oral health. Additionally, experimental results demonstrated that D-tagatose selectively inhibits growth of the oral pathogens S. mutans and S. gordonii. In contrast, the oral commensal S. oralis seemed to be negligibly affected, thus highlighting the potential of administration of D-tagatose as an oral prebiotic for its ability to manipulate the metabolism of those targeted oral streptococci.
Project description:The mechanisms through which oral commensal bacteria mitigates uncontrolled inflammatory responses in the oral mucosa remain unknown. Here we evaluated the ability of S. gordonii to stimulate the expression of miRNAs in oral epithelial cells with potential to target chemokine expression. The human oral epithelial cell line (OKF6) was exposed to different MOIs of S. gordonii for 24h and expression analysis of miRNAs performed using the Affymetrix platform.
Project description:Streptococcus gordonii, an important primary colonizer of dental plaque biofilm, specifically binds to salivary amylase via the surface-associated amylase-binding protein A (AbpA). We hypothesized that amylase binding to S. gordonii modulates expression of chromosomal genes, which could influence bacterial survival and persistence in the oral cavity. Gene expression profiling by microarray analysis was performed to detect differentially expressed genes in S. gordonii strain CH1 in response to the binding of purified human salivary amylase as compared to exposure to heat-denatured amylase. Selected genes found to be differentially expressed were validated by qRT-PCR. Five genes from the fatty acid synthesis (FAS) cluster were highly (10-35 fold) up-regulated in amylase treated S. gordonii CH1 cells compared to the denatured-amylase treated cells. An abpA-deficient strain of S. gordonii exposed to amylase did not show a similar response in FAS gene expression as observed in the parental strain. Predicted phenotypic effects of amylase binding to S. gordonii strain CH1 associated with increased expression of FAS genes leading to changes in fatty acid synthesis were noted, as evidenced by increased bacterial growth, survival at low pH, and resistance to triclosan. These changes were not observed in the amylase exposed abpA-deficient strain, suggesting for the role of AbpA in amylase-induced phenotype. These results provide evidence that the binding of salivary amylase elicits a differential gene response in S. gordonii, resulting in a phenotype adjustment that is potentially advantageous for bacterial survival in the oral environment.
Project description:In the oral biofilm, the mitis streptococci are among the first group of organisms to colonize the tooth surface. Their proliferation is thought to be an important factor required for antagonizing the growth of cariogenic species such as Streptococcus mutans. In this study, we used a 3-species mixed culture to demonstrate that another ubiquitous early colonizing species, Veillonella parvula, could greatly impact the competitive outcome of a mixed culture of S. mutans and S. gordonii. Transcriptome analysis further revealed that S. mutans responds differentially to its friend (V. parvula) and foe (S. gordonii). In the mixed culture with S. gordonii, all but one S. mutans sugar uptake and metabolic genes were down-regulated, while genes for alternative energy source utilization and H2O2 tolerance were up-regulated, resulting in a slower but persistent growth. In contrast, when cultured with V. parvula, S. mutans grew equally well or better than in monoculture and exhibited relatively few changes within its transcriptome. When V. parvula was introduced into the mixed culture of S. mutans and S. gordonii, it rescued the growth inhibition of S. mutans. In this 3-species environment, S. mutans increased the expression of genes required for the uptake and metabolism of minor sugars, while genes required for oxidative stress tolerance were down-regulated. We conclude that the major factors affecting the competition between S. mutans and S. gordonii are carbohydrate utilization and H2O2 resistance. The presence of V. parvula in the tri-species culture mitigates these two major factors and allows S. mutans to proliferate, despite the presence of S. gordonii. In this study, we used microarrays to investigate how S. mutans responds to different species. S. mutans was grown as either monospecies, dual-species cultures with S. gordonii or Veillonella, or tri-species cultures with S. gordonii and Veillonella. The transcriptional profile of the whole genome was examined with microarray.
Project description:Interspecies coaggregation promotes transcriptional changes of oral bacteria, contributing to the development of structurally balanced biofilms as well as oral diseases such as periodontitis. Streptococcus gordonii (S. gordonii) is an early colonizer of the oral cavity, and Fusobacterium nucleatum (F. nucleatum) may act as a bridge adhering to both early and late oral colonizers. These two species were commonly detected in healthy and periodontitis-diseased oral sites and could interact with immune cells such as macrophages. However, little research explored how intergeneric coaggregation affected transcriptional changes in S. gordonii and F. nucleatum subsp. polymorphum and how these gene changes might affect both species’ pathogenicity. The present study investigated transcriptional changes of both species in response to dual-species physical association using dual RNA-seq. Results indicated that after 30-min dual-species coaggregation, 148 genes were significantly up-regulated, and 124 genes were significantly down-regulated in S. gordonii. A total of 154 genes were significantly down-regulated, and 10 genes were significantly up-regulated in F. nucleatum subsp. polymorphum. A majority of up-regulated S. gordonii genes were involved in the biosynthesis and export of cell-wall proteins and the pathway of carbohydrate metabolism, and a group of down-regulated S. gordonii genes were associated with fatty acid biosynthesis and peptidoglycan biosynthesis. The transcriptome profiles indicated that the interspecies coaggregation led to a reduced level of DNA repair and lipopolysaccharides virulence in F. nucleatum subsp. polymorphum. The present study revealed that dual-species coaggregation induced a wide array of gene changes in S. gordonii and F. nucleatum subsp. polymorphum, enhancing S. gordonii’s adherence ability and attenuating F. nucleatum subsp. polymorphum's ability to produce LPS.
Project description:Streptococcus gordonii is a primary colonizer of the multispecies biofilm on tooth surfaces forming dental plaque, and a potential agent of endocarditis. The recent completion of the genome sequence of the naturally competent strain Challis allowed the design of a spotted oligonucleotide microarray to examine a genome-wide response of this organism to environmental signals. Based on temporal responses to synthetic competence signaling peptide (CSP) as indicated by transformation frequencies, the S. gordonii transcriptome was monitored at increments after CSP exposure. Microarray analysis identified 35 candidate early genes and 127 candidate late genes that were up-regulated at 5 and 15 minutes, respectively; these genes were often grouped in clusters. Findings supported published literature on the S. gordonii competence response, with up-regulation of most, but not all, genes that have been reported to affect this species' transformation frequencies. The CSP-induced transcriptomes of S. gordonii were compared to those of published S. pneumoniae strains. Both conserved and species-specific genes were identified. Putative intergenic regulatory sites such as the conserved combox sequence thought to be a binding site for competence sigma factor, were found preceding S. gordonii late responsive genes. In contrast, S. gordonii early CSP-responsive genes were not preceded by S. pneumoniae conserved direct repeats. These studies provide the first insights into a genome-wide transcriptional response of an oral commensal organism. They offer an extensive analysis of transcriptional changes that accompany competence in S. gordonii and form a basis for future intra- and inter-species comparative analyses of this ecologically important phenotype. Keywords: gene expression design