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Temporal and spatial trigger of post-exponential virulence-associated regulatory cascades by Legionella pneumophila after bacterial escape into the host cell cytosol

Authors

  • Maëlle Molmeret,

    1. Department of Microbiology and Immunology, Room MS-410, University of Louisville College of Medicine, Louisville, KY 40292, USA.
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  • Snake Jones,

    1. Department of Microbiology and Immunology, Room MS-410, University of Louisville College of Medicine, Louisville, KY 40292, USA.
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  • Marina Santic,

    1. Department of Microbiology and Immunology, Room MS-410, University of Louisville College of Medicine, Louisville, KY 40292, USA.
    2. Department of Microbiology and Parasitology, University of Rijeka, Rijeka, Croatia.
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  • Fabien Habyarimana,

    1. Department of Microbiology and Immunology, Room MS-410, University of Louisville College of Medicine, Louisville, KY 40292, USA.
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  • Maria Teresa Garcia Esteban,

    1. Department of Microbiology and Immunology, Room MS-410, University of Louisville College of Medicine, Louisville, KY 40292, USA.
    2. Centro Nacional de Microbiologi'a, Instituto de Salud Carlos II, Madrid, Spain.
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  • Yousef Abu Kwaik

    Corresponding author
    1. Department of Microbiology and Immunology, Room MS-410, University of Louisville College of Medicine, Louisville, KY 40292, USA.
      *E-mail abukwaik@louisville.edu; Tel. (+1) 502 852 4117; Fax (+1) 502 852 7531.
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*E-mail abukwaik@louisville.edu; Tel. (+1) 502 852 4117; Fax (+1) 502 852 7531.

Summary

During late stages of infection and prior to lysis of the infected macrophages or amoeba, the Legionella pneumophila-containing phagosome becomes disrupted, followed by bacterial escape into the host cell cytosol, where the last few rounds of bacterial proliferation occur prior to lysis of the plasma membrane. This coincides with growth transition into the post-exponential (PE) phase, which is controlled by regulatory cascades including RpoS and the LetA/S two-component regulator. Whether the temporal expression of flagella by the regulatory cascades at the PE phase is exhibited within the phagosome or after bacterial escape into the host cell cytosol is not known. We have utilized fluorescence microscopy-based phagosome integrity assay to differentiate between vacuolar and cytosolic bacteria/or bacteria within disrupted phagosomes. Our data show that during late stages of infection, expression of FlaA is triggered after bacterial escape into the macrophage cytosol and the peak of FlaA expression is delayed for few hours after cytosolic residence of the bacteria. Importantly, bacterial escape into the host cell cytosol is independent of flagella, RpoS and the two-component regulator LetA/S, which are all triggered by L. pneumophila upon growth transition into the PE phase. Disruption of the phagosome and bacterial escape into the cytosol of macrophages is independent of the bacterial pore-forming activity, and occurs prior to the induction of apoptosis during late stages of infection. We conclude that the temporal and spatial engagement of virulence-associated regulatory cascades by L. pneumophila at the PE phase is temporally and spatially triggered after phagosomal escape and bacterial residence in the host cell cytosol.

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