Francisella tularensis subsp. holarctica Releases Differentially Loaded Outer Membrane Vesicles Under Various Stress Conditions

Abstract

Francisella tularensis is a Gram-negative, facultative intracellular bacterium, causing a severe disease called tularemia. It secretes unusually shaped nanotubular outer membrane vesicles (OMV) loaded with a number of virulence factors and immunoreactive proteins. In the present study, the vesicles were purified from a clinical isolate of subsp. holarctica strain FSC200. We here provide a comprehensive proteomic characterization of OMV using a novel approach in which a comparison of OMV and membrane fraction is performed in order to find proteins selectively enriched in OMV vs. membrane. Only these proteins were further considered to be really involved in the OMV function and/or their exceptional structure. OMV were also isolated from bacteria cultured under various cultivation conditions simulating the diverse environments of F. tularensis life cycle. These included conditions mimicking the milieu inside the mammalian host during inflammation: oxidative stress, low pH, and high temperature (42°C); and in contrast, low temperature (25°C). We observed several-fold increase in vesiculation rate and significant protein cargo changes for high temperature and low pH. Further proteomic characterization of stress-derived OMV gave us an insight how the bacterium responds to the hostile environment of a mammalian host through the release of differentially loaded OMV. Among the proteins preferentially and selectively packed into OMV during stressful cultivations, the previously described virulence factors connected to the unique intracellular trafficking of Francisella were detected. Considerable changes were also observed in a number of proteins involved in the biosynthesis and metabolism of the bacterial envelope components like O-antigen, lipid A, phospholipids, and fatty acids. Data are available via ProteomeXchange with identifier PXD013074.

Membrane Vesicle Production as a Bacterial Defense Against Stress

“…Both the production rate and content of bacterial membrane vesicles can be altered by pH changes. Several fold increases in vesiculation rates have been observed at low pH for both extracellular and facultative intracellular bacteria, including F. tularensis ( Klimentova et al, 2019 ), Salmonella typhimurium ( Malabirade et al, 2018 ), and S. enteridis ( Chart et al, 1994 ).…”

Fitness of Outer Membrane Vesicles From Komagataeibacter intermedius Is Altered Under the Impact of Simulated Mars-like Stressors Outside the International Space Station

“…OMVs mirrored, to some extent, changes in their outer membrane structure, as well as membrane-associated activities of the K. intermedius re-isolates. An increase in both K. intermedius OMVs size and their number resulted from microbial adaptation to extraterrestrial stressors, apparently via cellular envelope remodeling, which is known from other Gram-negative bacteria exposed to stressful environmental or induced conditions (Sabra et al, 2003;Gerritzen et al, 2019;Klimentova et al, 2019). The increase in envelope thickness and production rate of OMVs with respect to the ground-based reference was previously demonstrated in the spaceflight samples of E. coli under microgravity (Zea et al, 2017).…”

The Translocation and Assembly Module (TAM) of Edwardsiella tarda Is Essential for Stress Resistance and Host Infection

“…pH is known to regulate the expression of membrane proteins, such as porins, that are required for bacterial survival under stress conditions associated with low pH, oxidation, osmotic pressure, and high temperature (Begic and Worobec, 2006;Klimentova et al, 2019). Low pH has been shown to induce conformational changes in the extracellular loop reign of OmpG and influence the transmembrane pore formation (Korkmaz-Ozkan et al, 2010).…”