Francisella tularensis is an intracellular bacterial pathogen, and is a category A bioterrorism agent. Within quiescent human macrophages, the F.tularensis pathogenicity island (FPI) is essential for bacterial growth within quiescent macrophages. The F.tularensis -containing phagosome matures to a late endosome-like stage that does not fuse to lysosomes for 1-8h, followed by gradual bacterial escape into the macrophage cytosol. Here we show that the FPI protein IglD is essential for intracellular replication in primary human monocyte-derived macrophages (hMDMs). While the parental strain replicates robustly in pulmonary, hepatic and splenic tissues of BALB/c mice associated with severe immunopathologies, the isogenic iglD mutant is severely defective. Within hMDMs, the iglD mutant-containing phagosomes mature to either a late endosome-like phagosome, similar to the parental strain, or to a phagolysosome, similar to phagosomes harbouring the iglC mutant control. Despite heterogeneity and alterations in phagosome biogenesis, the iglD mutant bacteria escape into the cytosol faster than the parental strain within hMDMs and pulmonary cells of BALB/c mice. Co-infections of hMDMs with the wild-type strain and the iglD mutant, or super-infection of iglD mutant-infected hMDMs with the wild-type strain show that the mutant strain replicates robustly within the cytosol of hMDMs coinhabited by the wild strain. However, when the wild-type strain-infected hMDMs are super-infected by the iglD mutant, the mutant fails to replicate in the cytosol of communal macrophages. This is the first demonstration of a F.tularensis novel protein essential for proliferation in the macrophage cytosol. Our data indicate that F.tularensis transduces signals to the macrophage cytosol to remodel it into a proliferative niche, and IglD is essential for transduction of these signals.
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