Project description:Small distortions in transcriptional networks might lead to drastic phenotypical changes, especially in cellular developmental programs such as competence for natural transformation. Here, we report a pervasive circuitry rewiring for competence and predation interplay in commensal streptococci. Canonically, in model species of streptococci such as Streptococcus pneumoniae and Streptococcus mutans, the pheromone-based two-component system BlpRH is a central node that orchestrates the production of antimicrobial compounds (bacteriocins) and incorporates signal from the competence activation cascade. However, the human commensal Streptococcus salivarius does not contain a functional BlpRH pair and in this species, the competence signaling system ComRS directly couples bacteriocin production and competence commitment. This network shortcut might account for an optimal reaction against microbial competitors and could explain the high prevalence of S. salivarius in the human digestive tract. Moreover, the broad spectrum of bacteriocin activity against pathogenic bacteria showcases the commensal and genetically tractable S. salivarius species as a user-friendly model for natural transformation and bacterial predation.
Project description:To investigate the subspecies level differences in the expression profiles of Streptococcus salivarius strains, we conducted the RNA sequencing analysis of Streptococcus salivarius strains.
Project description:The human oral cavity is one of the most competing environments, considering the extent and diversity of the bacterial community that colonizes it, as well as the multiple stresses to which it is subjected. The commensal species Streptococcus salivarius is one of the first colonizers of the oral mucosa (tongue, gums, inner cheeks and tooth enamel) in infants and remains predominant throughout the life of an adult. In order to adapt to diverse and variable environmental conditions, S. salivarius triggers in a coordinated way two bacterial developmental processes: competence (acquisition of extracellular DNA and new genetic traits) and predation (secretion of cytotoxic molecule), via ComR, a transcription factor activatable by a "pheromone" peptide. Nevertheless, the activation of other transcriptional regulators can uncouple these 2 mechanisms to allow, for example, only the production of toxins without triggering the competence phase.
Project description:Identification of proteins from extracellular vesicles isolated from Streptococcus salivarius. This work was financially supported by the National Science Centre, Poland (grant number 2021/43/D/NZ6/01464).
Project description:Human bronchial epithelial cells (16HBE14O-) were stimulated with the commensal bacterium S. salivarius K12, or the pathogens S. aureus, P. aeruginosa, or S. enteritidis (subtype Typhimurium) for 1 hour.
Project description:M cells are the main site of bacterial translocation in the intestine. We used the in vitro M cell model to study the effect of the commensal bacteria; Lactobacillus salivarius, Eschericha coli and Bacteroides fragilis, on M cell gene expression. Bacterial translocation across the gut mucosa has traditionally been based on the detection of commensals in the mesenteric lymph node. Differential rates of commensal translocation have been reported in vivo, however fewer studies have examined translocation of commensals at the level of the gut epithelial M cell. In this study we employed an in vitro M cell model to quantify translocation of various bacteria. C2BBe1 cells were differentiated into M cells and the gene expression profile and transport kinetics of different bacterial strains, namely Lactobacillus salivarius, Escherichia coli, and Bacteroides fragilis, was assessed. For comparison with M cell uptake, the THP-1 monocytic cell line was used to analyze bacterial internalization and resulting cytokine production. The commensal bacterial strains were translocated across M cells with different efficiencies; E. coli and B. fragilis translocated with equal efficiency while L. salivarius translocated with less efficiency. In contrast, L. salivarius was internalized by THP-1 cells to a higher degree than B. fragilis or E. coli and was associated with a different cytokine profile. Microarray analysis showed both common and differential gene expression amongst the bacteria and control polystyrene beads. In the presence of bacteria, but not beads, upregulated genes were mainly involved in transcription regulation and dephosphorylation, e.g. EGR1, JUN; whereas proinflammatory and stress response genes were primarily upregulated by E. coli and B. fragilis, but not L. salivarius nor beads, e.g. IL8, TNFAIP3. These results demonstrate that M cells have the ability to discriminate between different commensal bacteria and modify subsequent immune responses. C2bbe1 cells were converted to M cells (C2M) following 21 days of culture on Transwells in the presence of Raji B cells. C2M cells were co-cultured alone, Lactobacillus salivarius, Eschericha coli, Bacteroides fragilis and control beads. Total RNA was extracted and processed for Affymetrix array hybridisation
Project description:M cells are the main site of bacterial translocation in the intestine. We used the in vitro M cell model to study the effect of the commensal bacteria; Lactobacillus salivarius, Eschericha coli and Bacteroides fragilis, on M cell gene expression. Bacterial translocation across the gut mucosa has traditionally been based on the detection of commensals in the mesenteric lymph node. Differential rates of commensal translocation have been reported in vivo, however fewer studies have examined translocation of commensals at the level of the gut epithelial M cell. In this study we employed an in vitro M cell model to quantify translocation of various bacteria. C2BBe1 cells were differentiated into M cells and the gene expression profile and transport kinetics of different bacterial strains, namely Lactobacillus salivarius, Escherichia coli, and Bacteroides fragilis, was assessed. For comparison with M cell uptake, the THP-1 monocytic cell line was used to analyze bacterial internalization and resulting cytokine production. The commensal bacterial strains were translocated across M cells with different efficiencies; E. coli and B. fragilis translocated with equal efficiency while L. salivarius translocated with less efficiency. In contrast, L. salivarius was internalized by THP-1 cells to a higher degree than B. fragilis or E. coli and was associated with a different cytokine profile. Microarray analysis showed both common and differential gene expression amongst the bacteria and control polystyrene beads. In the presence of bacteria, but not beads, upregulated genes were mainly involved in transcription regulation and dephosphorylation, e.g. EGR1, JUN; whereas proinflammatory and stress response genes were primarily upregulated by E. coli and B. fragilis, but not L. salivarius nor beads, e.g. IL8, TNFAIP3. These results demonstrate that M cells have the ability to discriminate between different commensal bacteria and modify subsequent immune responses.
Project description:Synchronizing production of antibacterial compounds and integration of DNA released by dead cells, competence is one of the most efficient bacterial evolutionary and adaptative strategies. In most streptococci, this tactic is orchestrated by the ComRS system, a pheromone communication device providing competence bimodal initiation and sharp time window of activation. Understanding how this developmental process integrates multiple inputs to fine-tune the adequate response is a long-standing question. Actually, essential genes involved in the regulation of ComRS have been challenging to study. In this work, we built a conditional mutant library using the CRISPR-interference technology and developed three complementary screens to investigate competence genetic regulation in the human commensal Streptococus salivarius. We highlighted that competence increases upon cell-wall impairment, suggesting a connection between cell envelope stress and competence activation. Notably, we report the key role played by StkP, a serine-threonine kinase known to regulate cell-wall assembly. We showed that StkP controls competence by a mechanism responding to peptidoglycan fragments. Together, those data suggest a key cell-wall sensing mechanism coupling competence to cell envelope integrity.
Project description:Lactobacillus salivarius, found in the intestinal microbiota of humans and animals, is studied as an example of sub-dominant intestinal commensals that may impart benefits upon their host. Strains typically harbor at least one megaplasmid which encodes functions contributing to contingency metabolism and environmental adaptation. RNA-seq transcriptomic analysis of L. salivarius strain UCC118 identified the presence of a novel long and unusually abundant non-coding RNA (lancRNA) encoded by the megaplasmid and which represented more than 75% of total RNA-seq reads after depletion of ribosomal RNA species. The expression level of this 550 nt lancRNA in L. salivarius UCC118 exceeded that of the 16S rRNA, it accumulated during growth, was very stable over time, and was also expressed during intestinal transit in a mouse. This lancRNA sequence is specific to the L. salivarius species, however among 45 L. salivarius genomes analyzed, not all (only 34) harbored the sequence for the lancRNA. This lancRNA was produced in 27 tested L. salivarius strains but at strain-specific expression levels. High-level lancRNA expression correlated with high megaplasmid copy number. Transcriptome analysis of a deletion mutant lacking this lancRNA identified altered expression levels of genes in a number of pathways but a definitive function of this new long non-coding RNA was not identified. This lancRNA presents distinctive and unique properties, and suggests potential basic and applied scientific developments of this phenomenon.