Multiparametric Sensing of Outer Membrane Vesicle-Derived Supported Lipid Bilayers Demonstrates the Specificity of Bacteriophage Interactions

Abstract

The use of bacteriophages, viruses that specifically infect bacteria, as antibiotics has become an area of great interest in recent years as the effectiveness of conventional antibiotics recedes. The detection of phage interactions with specific bacteria in a rapid and quantitative way is key for identifying phages of interest for novel antimicrobials. Outer membrane vesicles (OMVs) derived from Gramnegative bacteria can be used to make supported lipid bilayers (SLBs) and therefore in vitro membrane models that contain naturally occurring components of the bacterial outer membrane. In this study, we employed Escherichia coli OMV derived SLBs and use both fluorescent imaging and mechanical sensing techniques to show their interactions with T4 phage. We also integrate these bilayers with microelectrode arrays (MEAs) functionalized with the conducting polymer PEDOT:PSS and show that the pore forming interactions of the phages with the SLBs can be monitored using electrical impedance spectroscopy. To highlight our ability to detect specific phage interactions, we also generate SLBs using OMVs derived from Citrobacter rodentium, which is resistant to T4 phage infection, and identify their lack of interaction with the phage. The work presented here shows how interactions occurring between the phages and these complex SLB systems can be monitored using a range of experimental techniques. We believe this approach can be used to identify phages that work against bacterial strains of interest, as well as more generally to monitor any pore forming structure (such as defensins) interacting with bacterial outer membranes, and thus aid in the development of next generation antimicrobials.

In Vitro Models for Investigating Intestinal Host–Pathogen Interactions

“…[ 288 ] Additionally, optical fluorescence microscopy and electrochemical impedance spectroscopy (EIS) provide tools to characterize membrane integrity and to study binding events as well as membrane‐pathogen interactions. [ 293 ]…”

“…[288] Additionally, optical fluorescence microscopy and electrochemical impedance spectroscopy (EIS) provide tools to characterize membrane integrity and to study binding events as well as membranepathogen interactions. [293] Examples of host-pathogen interaction studies featuring supported lipid bilayers comprise interactions of bacterial toxins with cell membranes and virus fusion events. Since bacterial toxins mainly act on the cell membrane by introducing pores or hydrolyzing lipids, [294][295][296] electrochemical impedance spectroscopy can be leveraged to monitor changes in resistance due to the incorporation of pores or disturbances in membrane integrity.…”

Droplet Polymer Bilayers for Bioelectronic Membrane Interfacing

“…SLBs on solid supports generally have a lower membrane resistance (1 kΩ cm 2 to 1 MΩ cm 2 ) because of interactions with the underlying substrate. , Further, while soft, hygroscopic polymer substrates can cushion fluid membranes, their rough surfaces can dissuade SLB formation and generate new sources of membrane defects. − Previous works creating SLBs on conducting polymers have innovated in polymer surface treatments − and membrane formation techniques to overcome this barrier. Despite this, SLBs on conducting polymer electrodes have displayed particularly low electrical sealing to nonspecific ionic currents, with normalized membrane resistances of <1 kΩ*cm 2 , suggesting incomplete surface coverage and the presence of significant membrane defects that may hinder future investigation of ion transport processes. ,,,− …”

“…Compared to traditional metal electrodes, organic conducting polymers can intimately interface with a biological environment by directly transducing ionic fluxes to an electronic current. , Their low impedance vastly improves sensitivity to small and otherwise undetectable biological signals . Conducting polymer electronics have so far been used to interface with increasingly complex membranes to investigate ion channel activity, the impact of antimicrobials, , and mechanisms of virus–host entry …”