Outer-inner membrane vesicles naturally secreted by gram-negative pathogenic bacteria

PLoS One. 2015 Jan 12;10(1):e0116896. doi: 10.1371/journal.pone.0116896. eCollection 2015.


Outer-inner membrane vesicles (O-IMVs) were recently described as a new type of membrane vesicle secreted by the Antarctic bacterium Shewanella vesiculosa M7T. Their formation is characterized by the protrusion of both outer and plasma membranes, which pulls cytoplasmic components into the vesicles. To demonstrate that this is not a singular phenomenon in a bacterium occurring in an extreme environment, the identification of O-IMVs in pathogenic bacteria was undertaken. With this aim, a structural study by Transmission Electron Microscopy (TEM) and Cryo-transmission electron microscopy (Cryo-TEM) was carried out, confirming that O-IMVs are also secreted by Gram-negative pathogenic bacteria such as Neisseria gonorrhoeae, Pseudomonas aeruginosa PAO1 and Acinetobacter baumannii AB41, in which they represent between 0.23% and 1.2% of total vesicles produced. DNA and ATP, which are components solely found in the cell cytoplasm, were identified within membrane vesicles of these strains. The presence of DNA inside the O-IMVs produced by N. gonorrhoeae was confirmed by gold DNA immunolabeling with a specific monoclonal IgM against double-stranded DNA. A proteomic analysis of N. gonorrhoeae-derived membrane vesicles identified proteins from the cytoplasm and plasma membrane. This confirmation of O-IMV extends the hitherto uniform definition of membrane vesicles in Gram-negative bacteria and explains the presence of components in membrane vesicles such as DNA, cytoplasmic and inner membrane proteins, as well as ATP, detected for the first time. The production of these O-IMVs by pathogenic Gram-negative bacteria opens up new areas of study related to their involvement in lateral gene transfer, the transfer of cytoplasmic proteins, as well as the functionality and role of ATP detected in these new vesicles.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Outer Membrane Proteins / metabolism*
  • Biological Transport / physiology
  • Cell Membrane / metabolism
  • Cytoplasm / metabolism
  • DNA / metabolism
  • Gram-Negative Bacteria / metabolism*
  • Secretory Vesicles / metabolism*
  • Shewanella / metabolism


  • Bacterial Outer Membrane Proteins
  • DNA

Grant support

This work received funding from the following sources: Government of Spain (CICYT grant CTQ 2010-21183-C02-01/PPQ)EM CP-C, Autonomous Government of Catalonia (grant 2009SGR1212)EM CL-I, and Fellowship FFAR2012.3 from the University of Barcelona (CP-C). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.