How do superbugs hide from the host? Understanding the mechanisms of innate immune evasion in the opportunistic pathogen Enterococcus faecalis

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All enterococci produce a surface polysaccharide called the enterococcal polysaccharide antigen (EPA) that plays a key role during pathogenesis (Mistou et al., 2016). EPA is is encoded by two gene clusters: (i) 18 genes extremely conserved, encoding a core synthetic machinery and (ii) 10-20 genes variable from one strain to another, responsible for the decoration of the polysaccharide backbone. Our results indicate that mutants in the decoration genes display morphological defects, an altered susceptibility to beta-lactams and are no longer virulent in a zebrafish model of infection (Prasjnar et al., 2013; Smith et al., 2018). We hypothesize that EPA plays a major role for the cell surface display of proteins involved in cell wall synthesis and that the decoration of this polymer mediates innate immune evasion during pathogenesis.The aim of this project is to investigate how EPA contributes to E. faecalis antimicrobial resistance and virulence. Three objectives will be set.1. Explore the role of EPA in cell growth and resistance to antimicrobials. We will use available E. faecalis recombinant strains which no longer or conditionally produce EPA to investigate the contribution of this polymer to cell wall structure and dynamics. We will also explore how EPA modulates the capacity of antimicrobials targeting the cell envelope (eg., beta-lactams, cationic peptides, bile salts) to bind and penetrate the cell walls.2. Investigate the role of EPA during pathogenesis. We will study (i) how EPA and its decoration modulate recognition by macrophages and neutrophils during pathogenesis, using both in vitro phagocytosis assays and the zebrafish model of infection and (ii) investigate the role of EPA on bacterial population dynamics in the host. We will test if soluble EPA can be used as a decoy molecule to evade phagocytosis.3. Explore the cell signalling pathway triggered by enterococci during infection. We will use in vitro phagocytosis assay in primary human and murine macrophages to identify signal transduction cascades that shape the host response to enterococcal infections.