A dead end trap? - Functions of bacteriophages bound to outer membrane vesicles in host infection and defence

• Applicants: Professorin Dr. Stefanie Barbirz; Dr. Nina Bröker
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms ; Virology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 548605513

The outer membrane (OM) of Gram-negative bacteria effectively protects the cell from environmental influences. In particular, the OM is the initial contact interface for infection with bacteriophages (phages), the viruses of bacteria. Bacteria dynamically change their OM composition by shedding small, spherical outer membrane vesicles (OMVs) with diameters of 40-400 nm. OMVs are highly relevant in bacterial pathogenesis and molecular exchange between all kingdoms including eukaryotic cells. For example, OMVs can transfer toxins or factors that enhance antibiotic resistance. OMVs compete with the bacterial OM for binding of bacteriophages, as they may contain all or a subset of phage receptors. OMVs thus efficiently reduce the number of infecting particles in a bacterial population. In spite of this highly important cross-kingdom communication via OMVs, not much is known about the molecular details of their bacteriophage interactions and about the functional synergies of OMVs, phages and bacterial hosts. Aim of our project is to characterize the interactions of Salmonella phages with OMVs in the model organism Salmonella Typhimurium. We want to describe the mechanisms by which OMVs inactivate bacteriophages. Using fluorescence spectroscopy, we could show that the model phage P22 injected its DNA into OMVs. However, so far it remains unclear whether OMVs can transfer this externally acquired phage genetic material into bacteria, and thus represent an alternative infection pathway. We will employ in vitro model membrane systems or intact Salmonella and fluorescence microscopy to study the OMV fusion to Gram-negative OMs as a prerequisite for OMV-mediated gene transfer. Bacteria react to bacteriophages with enhanced vesiculation. We will analyze the OMV response of Salmonella to bacteriophages for different strain types and culture conditions. We will establish fluorescence correlation spectroscopy to rapidly quantify OMVs in complex particle mixtures, for example in culture supernatants. Our project will contribute to understanding of bacterial virus-host interactions where extracellular vesicles are ubiquitously present. In particular, eukaryotic cells are able to take up these bacterial nanoparticles. This emphasizes that also bacteriophages interacting with OMVs represent major players in extracellular vesicle-based communication at the interface of microbioms and higher organisms.

• DFG Programme: Priority Programmes
• Subproject of: SPP 2330: New concepts in prokaryotic virus-host interactions - from single cells to microbial communities
• Co-Investigators: Dr. Stephan Block; Professor Dr. Salvatore Chiantia