Nanowire Arrays as Cell Force Sensors To Investigate Adhesin-Enhanced Holdfast of Single Cell Bacteria and Biofilm Stability

Abstract

Surface attachment of a planktonic bacteria, mediated by adhesins and extracellular polymeric substances (EPS), is a crucial step for biofilm formation. Some pathogens can modulate cell adhesiveness, impacting host colonization and virulence. A framework able to quantify cell-surface interaction forces and their dependence on chemical surface composition may unveil adhesiveness control mechanisms as new targets for intervention and disease control. Here we employed InP nanowire arrays to dissect factors involved in the early stage biofilm formation of the phytopathogen Xylella fastidiosa. Ex vivo experiments demonstrate single-cell adhesion forces up to 45 nN, depending on the cell orientation with respect to the surface. Larger adhesion forces occur at the cell poles; secreted EPS layers and filaments provide additional mechanical support. Significant adhesion force enhancements were observed for single cells anchoring a biofilm and particularly on XadA1 adhesin-coated surfaces, evidencing molecular mechanisms developed by bacterial pathogens to create a stronger holdfast to specific host tissues.

Nanowire Arrays as Force Sensors with Super‐Resolved Localization Position Detection: Application to Optical Measurement of Bacterial Adhesion Forces

“…Therefore, we decide to calculate displacements using a Gaussian fitting of nanowire apex deflections instead of Feret diameters . We can observe from Figure B,C that indeed the Gaussian function properly describes our data, providing the center, linewidth, and height of the reflection pattern.…”

“…A second requirement is the material system chosen for the nanowires. Semiconductor surfaces, such as Si and InP, have been shown as suitable supports for bacterial adhesion in many studies . In fact, Si nanowires have been used to demonstrate that Shewanella oneidensis MR‐1 cells can recognize nanoscale structures; their swimming patterns and initial attachment locations are strongly influenced by the presence of nanowires on a surface .…”

“…In a previous report using this technique, the positions of all nanowire tips in a video frame were tracked by considering them as solid circular particles (Figure B); the center of the circle was computed from the position of horizontal and vertical Feret diameters of a circular post. This procedure was automatically carried out after appropriately choosing the region of interest in the particle analysis plugin of ImageJ2/Fiji .…”

“…Therefore, an AFM similar concept of optically measuring a cantilever deflection to probe forces, was recently introduced using polymeric or semiconductor nanopillar arrays, capable to acquire vector force maps in parallel and real time . Several groups have demonstrated the use of force sensors based on vertical nanopillars or nanowires for mammalian cell studies . The use of crystalline materials with well‐defined elastic properties, and stiffer than the elastomeric materials originally used for that purpose, improves the method accuracy.…”

Nanoneedle-Based Sensing in Biological Systems

“…In comparison, optical tweezers measure forces down to a single pN 72 , but only for a few points simultaneously. Nanoneedles can also monitor the forces exerted on the substrate by individual bacterial cells and biofilms 73 ( Figure 4B). Nanowire arrays (InP) with 500nm spacing can monitor the forces generated by X. fastidiosa seeded on them by electron microscopy or by confocal microscopy.…”