Chlorobaculum tepidum Outer Membrane Vesicles Are Likely a Significant Route for Biogenic Sulfur Transport

Outer membrane-derived vesicles (OMVs) have been studied in different phyla of Gram-negative bacteria, most extensively in the Pseudomonadota, where they have been shown to participate in diverse biological and environmental processes. To date, the production of OMVs has not been reported in the Chlorobiaceae within the phylum Chlorobiota. Chlorobaculum. tepidum is the model organism for the Chlorobiaceae that synthesizes and consumes insoluble extracellular sulfur (S(0)) globules by an unknown mechanism. Here, we report evidence implicating outer membrane vesicles in biogenic S(0) globule synthesis. First we demonstrate that Cba. tepidum secretes OMVs in the extracellular milieu, and that OMV yield and size vary with growth conditions, particularly sulfide concentration. A core of 28 proteins involved in diverse biological processes such as cell wall biogenesis, inorganic ion transport and metabolism were found to be shared between OMVs, extracellular S(0) globules and Cba. tepidum intact cells. Multiple analytical methods indicated that OMVs contain S(0) and that OMVs and biogenic S(0) globules share protein and polysaccharide signatures, including lipooligosaccharides. Together these lines of evidence indicate that Cba. tepidum ’s OMVs are one component of sulfur transport between cells and extracellular sulfur globules.