Cysteine depletion triggers sOMV release.

<p>The effect of cysteine depletion on sOMV release is demonstrated with a repletion experiment in shakeflasks. (A) Pre-cultures are grown on control medium with cysteine (circles). Harvested cells are repleted with control medium (circles) or medium without cysteine (triangles). Shake flasks without cysteine enter the stationary phase, while cysteine repleted flasks continue growth normally. A third group (squares) is repleted with control medium, but growth is repressed with peroxide to mimic oxidative stress and verify the transcriptome data (arrows indicate peroxide additions). Y-axis represents dry biomass concentration (g dry weight per L). (B) Vesicle release in the cultivation supernatant is monitored with a fluorescent probe up to 6 hours after repletion. Shake flasks with control medium grow normally but do not release sOMV, while cysteine depletion triggers a sustained release of vesicles. Oxidative stress has a similar but transient effect, since sOMV are released temporarily after each peroxide addition. Cysteine depletion is therefore an external stimulus for sOMV release, which induces oxidative stress as the intracellular signal. Y-axis represents specific sOMV yield (mg PorA antigen per g dry weight).