Membrane Vesicles Shed byLegionella pneumophilaInhibit Fusion of Phagosomes with Lysosomes

Abstract

When cultured in broth to the transmissive phase, Legionella pneumophila infects macrophages by inhibiting phagosome maturation, whereas replicative-phase cells are transported to the lysosomes. Here we report that the ability of L. pneumophila to inhibit phagosome-lysosome fusion correlated with developmentally regulated modifications of the pathogen's surface, as judged by its lipopolysaccharide profile and by its binding to a sialic acid-specific lectin and to the hydrocarbon hexadecane. Likewise, the composition of membrane vesicles shed by L. pneumophila was developmentally regulated, based on binding to the lectin and to the lipopolysaccharidespecific monoclonal antibody 3/1. Membrane vesicles were sufficient to inhibit phagosome-lysosome fusion by a mechanism independent of type IV secretion, since only ϳ25% of beads suspended with or coated by vesicles from transmissive phase wild type or dotA secretion mutants colocalized with lysosomal probes, whereas ϳ75% of beads were lysosomal when untreated or presented with vesicles from the L. pneumophila letA regulatory mutant or E. coli. As observed previously for L. pneumophila infection of mouse macrophages, vesicles inhibited phagosome-lysosome fusion only temporarily; by 10 h after treatment with vesicles, macrophages delivered ϳ72% of ingested beads to lysosomes. Accordingly, in the context of the epidemiology of the pneumonia Legionnaires' disease and virulence mechanisms of Leishmania and Mycobacteria, we discuss a model here in which L. pneumophila developmentally regulates its surface composition and releases vesicles into phagosomes that inhibit their fusion with lysosomes.To exploit macrophages as a replication niche, L. pneumophila, Coxiella burnetii, and Leishmania spp. apply a similar strategy (69). When ingested, each pathogen differentiates from a transmissive form that inhibits phagosome-lysosome fusion for several hours to a cell type fit for intracellular replication (20, 35, 37, 63, 67). Differentiation of L. pneumophila can be modeled in broth, where exponential (E)-phase bacteria switch to the transmissive phenotype as their nutrient supply wanes (11). As L. pneumophila enters the postexponential (PE) phase, the alarmone ppGpp triggers a regulatory cascade mediated in part by the two component system LetA/LetS and the sigma factors RpoS and FliA to coordinate expression of motility, cytotoxicity, stress resistance, factors that inhibit phagosome-lysosome fusion, and other traits likely to promote transmission to a new host cell (55, 56).To establish a replication niche in macrophages, L. pneumophila utilizes type IV secretion to deliver substrates that alter host membrane traffic (14, 75). Instead of merging with lysosomes, the L. pneumophila vacuole becomes surrounded by membranes derived from the early secretory pathway by a process that in A/J mouse macrophages resembles autophagy (3,19,34,42,70,72). The pathogen persists for several hours within an ER-like compartment, where it differentiates to the replicative form. Nevertheless, muta...

Multigenome analysis identifies a worldwide distributed epidemic Legionella pneumophila clone that emerged within a highly diverse species

“…Thus, they represent interesting marker genes for rapid Sg1 identification. As LPS is not only the basis for the classification of serogroups, but is also a major immunodominant antigen of L. pneumophila, and membrane vesicles shed by virulent L. pneumophila containing LPS have been shown to be sufficient to inhibit phagosome-lysosome fusion (Fernandez-Moreira et al 2006), our results suggest that LPS of Sg1 is implicated in the predominance of Sg1 strains in human disease Doleans et al 2004) compared with other serogroups of L. pneumophila and other Legionella species. …”

Polyketide synthase (PKS) reduces fusion of Legionella pneumophila-containing vacuoles with lysosomes and contributes to bacterial competitiveness during infection

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Mucosal immunization with Shigella flexneri outer membrane vesicles induced protection in mice

“…Currently, there have been described many functions for this blebbing process. Functions proposed vary from facilitating the intracellular bacterial growth within phagocytes [11], to the delivery of effectors molecules critical for pathogen dissemination such as pathogen-associated molecular patterns (PAMPs) and other virulence factors to host cells [12][13][14].…”