Legionella pneumophila-Derived Outer Membrane Vesicles Promote Bacterial Replication in Macrophages

PLoS Pathog. 2016 Apr 22;12(4):e1005592. doi: 10.1371/journal.ppat.1005592. eCollection 2016 Apr.


The formation and release of outer membrane vesicles (OMVs) is a phenomenon of Gram-negative bacteria. This includes Legionella pneumophila (L. pneumophila), a causative agent of severe pneumonia. Upon its transmission into the lung, L. pneumophila primarily infects and replicates within macrophages. Here, we analyzed the influence of L. pneumophila OMVs on macrophages. To this end, differentiated THP-1 cells were incubated with increasing doses of Legionella OMVs, leading to a TLR2-dependent classical activation of macrophages with the release of pro-inflammatory cytokines. Inhibition of TLR2 and NF-κB signaling reduced the induction of pro-inflammatory cytokines. Furthermore, treatment of THP-1 cells with OMVs prior to infection reduced replication of L. pneumophila in THP-1 cells. Blocking of TLR2 activation or heat denaturation of OMVs restored bacterial replication in the first 24 h of infection. With prolonged infection-time, OMV pre-treated macrophages became more permissive for bacterial replication than untreated cells and showed increased numbers of Legionella-containing vacuoles and reduced pro-inflammatory cytokine induction. Additionally, miRNA-146a was found to be transcriptionally induced by OMVs and to facilitate bacterial replication. Accordingly, IRAK-1, one of miRNA-146a's targets, showed prolonged activation-dependent degradation, which rendered THP-1 cells more permissive for Legionella replication. In conclusion, L. pneumophila OMVs are initially potent pro-inflammatory stimulators of macrophages, acting via TLR2, IRAK-1, and NF-κB, while at later time points, OMVs facilitate L. pneumophila replication by miR-146a-dependent IRAK-1 suppression. OMVs might thereby promote spreading of L. pneumophila in the host.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Membrane
  • Enzyme-Linked Immunosorbent Assay
  • Extracellular Vesicles / metabolism*
  • Fluorescent Antibody Technique
  • Humans
  • Legionella pneumophila
  • Legionnaires' Disease / metabolism*
  • Macrophage Activation / physiology
  • Macrophages / metabolism
  • Macrophages / microbiology*
  • Real-Time Polymerase Chain Reaction

Grants and funding

Part of this work has been funded by Bundesministerium für Bildung und Forschung (e:bio miRSys - FKZ 0316175B, e:Med CAPSYS - FKZ 01X1304E; http://www.bmbf.de/) to BS, and Deutsche Forschungsgemeinschaft (SFB/TR-84; http://www.sfb-tr84.de/) and Hessisches Ministerium für Wissenschaft und Kunst (LOEWE Medical RNomics - FKZ 519/03/00.001-(0003); http://www.proloewe.de/medicalrnomics) to BS. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.