In-Situ Determination of the Mechanical Properties of Gliding or Non-Motile Bacteria by Atomic Force Microscopy under Physiological Conditions without Immobilization

Abstract

We present a study about AFM imaging of living, moving or self-immobilized bacteria in their genuine physiological liquid medium. No external immobilization protocol, neither chemical nor mechanical, was needed. For the first time, the native gliding movements of Gram-negative Nostoc cyanobacteria upon the surface, at speeds up to 900 µm/h, were studied by AFM. This was possible thanks to an improved combination of a gentle sample preparation process and an AFM procedure based on fast and complete force-distance curves made at every pixel, drastically reducing lateral forces. No limitation in spatial resolution or imaging rate was detected. Gram-positive and non-motile Rhodococcus wratislaviensis bacteria were studied as well. From the approach curves, Young modulus and turgor pressure were measured for both strains at different gliding speeds and are ranging from 20±3 to 105±5 MPa and 40±5 to 310±30 kPa depending on the bacterium and the gliding speed. For Nostoc, spatially limited zones with higher values of stiffness were observed. The related spatial period is much higher than the mean length of Nostoc nodules. This was explained by an inhomogeneous mechanical activation of nodules in the cyanobacterium. We also observed the presence of a soft extra cellular matrix (ECM) around the Nostoc bacterium. Both strains left a track of polymeric slime with variable thicknesses. For Rhodococcus, it is equal to few hundreds of nanometers, likely to promote its adhesion to the sample. While gliding, the Nostoc secretes a slime layer the thickness of which is in the nanometer range and increases with the gliding speed. This result reinforces the hypothesis of a propulsion mechanism based, for Nostoc cyanobacteria, on ejection of slime. These results open a large window on new studies of both dynamical phenomena of practical and fundamental interests such as the formation of biofilms and dynamic properties of bacteria in real physiological conditions.

Graphene oxide increases the viability of C2C12 myoblasts microencapsulated in alginate

“…Force Mapping was followed by QI imaging for each capsule. We found this method was most appropriate for our samples, given that they are loosely immobilized and the applied force can be easily controlled through this method (Dhahri et al, 2013). Our results showed very homogeneous topographical images for capsules lacking coatings and GO, with more smoothed surfaces after PLL and alginate coating addition.…”

Mechanical and cell-to-cell adhesive properties of aggregated Methanosarcina

“…3. Due to its specific indentation mode, the force-indentation curve is a different shape than is usually observed for the microbial cells (see for instance [33,35]). The three stages in force-indentation response can be observed in Fig.…”

“…For measuring the force-displacement relationship, atomic force microscopy (AFM) technique has been successfully used over the past two decades [29,31]. The use of AFM as a tool to probe the mechanical properties of single bacterial cells as well as their cell-to-surface adhesive properties [21,[32][33][34][35][36][37] as well as adhesion and cohesion in bacterial biofilms [38,39] has been well reported. However, the use of AFM for determination the mechanical properties of archaeal cells has been attracted much less attention (see for instance [40]).…”

Dissipative shocks behind bacteria gliding

“…These latter are filamentous bacteria (blue-green algae) with a number of septa, for which the slime extrusion hypothesis assumes that slime filaments are ejected through the pores that surround each cell septum (see also [9]). Hoiczyk and Baumeister showed in [18] how slime formed bands about the cell surface that could be removed [35]. It should be noted that the resolution of these images is at least 100 times poorer than that of the images shown in figure 1, and so the slime filaments cannot be seen.…”