Real-time tracking of bacterial membrane vesicles reveals enhanced membrane traffic upon antibiotic exposure.

Julia Bos,Luis H Cisneros,Didier Mazel

doi:10.1126/sciadv.abd1033

Abstract

Membrane vesicles are ubiquitous carriers of molecular information. A broad understanding of the biological functions of membrane vesicles in bacteria remains elusive because of the imaging challenges during real-time in vivo experiments. Here, we provide a quantitative analysis of the motion of individual vesicles in living microbes using fluorescence microscopy, and we show that while vesicle free diffusion in the intercellular space is rare, vesicles mostly disperse along the bacterial surfaces. Most remarkably, when bacteria are challenged with low doses of antibiotics, vesicle production and traffic, quantified by instantaneous vesicle speeds and total traveled distance per unit time, are significantly enhanced. Furthermore, the enhanced vesicle movement is independent of cell clustering properties but rather is associated with a reduction of the density of surface appendages in response to antibiotics. Together, our results provide insights into the emerging field of spatial organization and dynamics of membrane vesicles in microcolonies.

Highlights

Membrane vesicles (MVs) are small membrane blebs containing biological material derived from their parental cell
We demonstrate that MVs undergo several dynamical states, including arrested motion, modest subdiffusive motion constrained to the surface of cell membranes, and occasional random walks in the intercellular space
To capture E. coli MVs’ dynamics inside the microcolony and examine the effect of antibiotic stressors on their movement, we used high-magnification wide-field fluorescence microscopy combined with a scientific complementary metal–oxide–semiconductor camera (Hamamatsu ORCA-Flash 4.0) capable of low read noise at 20 frames per second

Summary

Introduction

Membrane vesicles (MVs) are small membrane blebs containing biological material derived from their parental cell. While MVs are generally produced by fit viable cells [6, 8,9,10], mechanisms of cell death have been shown to mediate vesiculation in both bacteria [e.g., prophage-induced lysis [11], explosive lysis [12], and cell disintegration [13]] and eukaryotic organisms [e.g., apoptosis [14, 15] and necrosis [16]] Other inducing factors, such as antibiotics [11, 17,18,19,20,21], chemotherapy drugs [22], intrinsic envelop stress [23], and environmental stresses [24, 25] or DNA damage [11, 12, 26], have been shown to stimulate vesiculation, suggesting that MVs may be involved in some form of yet undiscovered general biological stress response

Methods

Results

Conclusion