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Here we introduce PeakForce Tapping non-resonance atomic force microscopy for imaging and nanomechanical mapping of Caenorhabditis elegans nematodes. The animals were imaged both in air and water at nanoscale resolution. Layer-by-layer glass surface modification was employed to secure the worms for imaging in water. Microtopography of head region, annuli, furrows, lateral alae and tail region was visualized. Analysis of nanoscale surface features obtained during AFM imaging of three larval and adult hermaphrodite nematodes in natural environment allowed for numerical evaluation of annuli periodicity, furrows depth and annuli roughness. Nanomechanical mapping of surface deformation, Young modulus and adhesion confirms that the mechanical properties of the nematode cuticle are non-uniform. Overall, PeakForce Tapping AFM is a robust and simple approach applicable for nanoscale three …