Bacteriophage infection of Escherichia coli leads to the formation of membrane vesicles via both explosive cell lysis and membrane blebbing.

Pappu K Mandal,Laura M Nolan,Giulia Ballerin,Nicola K Petty,Cynthia B Whitchurch

doi:10.1099/mic.0.001021

Pappu K Mandal, Laura M Nolan + Show 3 more

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https://doi.org/10.1099/mic.0.001021

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Abstract

Membrane vesicles (MVs) are membrane-bound spherical nanostructures that prevail in all three domains of life. In Gram-negative bacteria, MVs are thought to be produced through blebbing of the outer membrane and are often referred to as outer membrane vesicles (OMVs). We have recently described another mechanism of MV formation in Pseudomonas aeruginosa that involves explosive cell-lysis events, which shatters cellular membranes into fragments that rapidly anneal into MVs. Interestingly, MVs are often observed within preparations of lytic bacteriophage, however the source of these MVs and their association with bacteriophage infection has not been explored. In this study we aimed to determine if MV formation is associated with lytic bacteriophage infection. Live super-resolution microscopy demonstrated that explosive cell lysis of Escherichia coli cells infected with either bacteriophage T4 or T7, resulted in the formation of MVs derived from shattered membrane fragments. Infection by either bacteriophage was also associated with the formation of membrane blebs on intact bacteria. TEM revealed multiple classes of MVs within phage lysates, consistent with multiple mechanisms of MV formation. These findings suggest that bacteriophage infection may be a major contributor to the abundance of bacterial MVs in nature.

Highlights

Membrane vesicles (MVs) are non-replicating, membrane- bound spherical nanostructures that prevail in all the three domains of life [1]
We have demonstrated that T4 and T7 phage infection of E. coli generates MVs via both explosive cell lysis and membrane blebbing
Using 3D-SIM we observed a range of sizes of MVs in phage-infected cultures, which was observed in phage lysates using Transmission electron microscopy (TEM)

Summary

Introduction

Membrane vesicles (MVs) are non-replicating, membrane- bound spherical nanostructures that prevail in all the three domains of life [1]. Bacterial MVs are involved in diverse biological processes such as pathogenicity [2], horizontal gene transfer [3], biofilm formation [4], and to serve as decoys to defend bacteria from antibiotics [5], antimicrobial peptides [6] and bacteriophage predation [7]. In Gram-negative bacteria MVs are thought to be produced via membrane blebbing, which involves protrusion and budding of the outer membrane resulting in the formation of MVs, which are often referred to as outer membrane vesicles (OMVs). Membrane blebbing occurs as a result of cell-envelope disturbances caused by either imbalance of peptidoglycan biosynthesis or intercalation of hydrophobic molecules into the outer membrane [9]. In Pseudomonas aeruginosa explosive cell lysis occurs as a consequence of degradation of peptidoglycan by the muralytic endolysin, Lys, similar to those used by lytic bacteriophage to lyse its bacterial host. Lys is encoded in the R- and F-pyocin gene cluster which produces tailocins related to P2 and lambda bacteriophage [10,11,12]

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