Biogenesis of Outer Membrane Vesicles in Serratia marcescens Is Thermoregulated and Can Be Induced by Activation of the Rcs Phosphorelay System

Abstract

Outer membrane vesicles (OMVs) have been identified in a wide range of bacteria, yet little is known of their biogenesis. It hasbeen proposed that OMVs can act as long-range toxin delivery vectors and as a novel stress response. We have found that theformation of OMVs in the Gram-negative opportunistic pathogen Serratia marcescens is thermoregulated, with a significantamount of OMVs produced at 22 or 30°C and negligible quantities formed at 37°C under laboratory conditions. Inactivation ofthe synthesis of the enterobacterial common antigen (ECA) resulted in a hypervesiculation phenotype, supporting the hypothesisthat OMVs are produced in response to stress. We demonstrate that the phenotype can be reversed to wild-type (WT) levelsupon the loss of the Rcs phosphorelay response regulator RcsB, but not RcsA, suggesting a role for the Rcs phosphorelay in theproduction of OMVs. MS fingerprinting of the OMVs provided evidence of cargo selection within wild-type cells, suggesting apossible role for Serratia OMVs in toxin delivery. In addition, OMV-associated cargo proved toxic upon injection into the haemocoelof Galleria mellonella larvae. These experiments demonstrate that OMVs are the result of a regulated process in Serratiaand suggest that OMVs could play a role in virulence.