In-situ determination of the mechanical properties of gliding or non-motile bacteria by Atomic Force Microscopy under physiological conditions without immobilization.

51 pages; 15figures; 3 tables, accepted for publication in PloS ONEInternational audienceWe 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 900microns/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 R. 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±5MPa and 40±5 to 310±30kPa 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 Nostoc. We also observed the presence of a soft extra cellular matrix (ECM) around the Nostoc. Both strains left a track of polymeric slime with variable thicknesses. For R. wratislaviensis, 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