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We present a study about AFM imaging of living, moving or self-immobilized,bacteria in their genuine physiological liquid medium. No externalimmobilization protocol, neither chemical nor mechanical, was needed. For thefirst time, the native gliding movements of Gram-negative Nostoc cyanobacteriaupon the surface, at speeds up to 900microns/h, were studied by AFM. This waspossible thanks to an improved combination of a gentle sample preparationprocess and an AFM procedure based on fast and complete force-distance curvesmade at every pixel, drastically reducing lateral forces. No limitation inspatial resolution or imaging rate was detected. Gram-positive and non-motileRhodococcus wratislaviensis bacteria were studied as well. From the approachcurves, Young modulus and turgor pressure were measured for both strains atdifferent gliding speeds and are ranging from 20 to 105MPa and 40 to 310kPadepending on the bacterium and the gliding speed. For Nostoc, spatially limitedzones with higher values of stiffness were observed. The related spatial periodis much higher than the mean length of Nostoc nodules. This was explained by aninhomogeneous mechanical activation of nodules in the Nostoc. We also observedthe presence of a soft extra cellular matrix (ECM) around the Nostoc bacterium.Both strains left a track of polymeric slime with variable thicknesses. ForRhodococcus, it is equal to few hundreds of nm, likely to promote its adhesionto the sample. While gliding, the Nostoc secretes a slime layer the thicknessof which is in the nanometer range and increases with the gliding speed. Theseresults open a large window on new studies of both dynamical phenomena ofpractical and fundamental interests such as the formation of biofilms anddynamic properties of bacteria in real physiological conditions.

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