ABSTRACT We used a novel atomic force microscopy (AFM)-based technique to compare the local viscoelastic properties of individual gram-negative ( Escherichia coli ) and gram-positive ( Bacillus subtilis ) bacterial cells. We found that the viscoelastic properties of the bacterial cells are well described by a three-component mechanical model that combines an instantaneous elastic response and a delayed elastic response. These experiments have allowed us to investigate the relationship between the viscoelastic properties and the structure and composition of the cell envelope. In addition, this is the first report in which the mechanical role of Lpp, the major peptidoglycan-associated lipoprotein and one of the most abundant outer membrane proteins in E. coli cells, has been quantified. We expect that our findings will be helpful in increasing the understanding of the structure-property relationships of bacterial cell envelopes.