**4. Factors affecting viscoelasticity of biological materials**

Cell components imparting viscoelastic properties to the entire cell are well studied, but there are very few examples in the literature in which external factors are shown to affect cell viscoelasticity. The structural integrity of biological systems is partially dependent on the degree of hydration. Humidity affects the mechanical properties of biopolymers, but the intensity of this effect depends on the type of biopolymer. For example, effects of humidity on cellulose are much less severe than on peptidoglycan. Thwaites et al. [136] demonstrated that the viscoelastic behavior of *Bacillus subtilis* depends on humidity, which affects the viscoelasticity of the peptidoglycan layer. Increasing humidity gives rise to cell wall hydration, allowing water to form hydrogen bonds with peptides and other cell wall components, making the cell wall more pliable [136]. Bacterial and fungal spores are very rigid under dry conditions, but increased humidity leads to removal of the outer spore hydrophobic layer, making it softer than under dry conditions and leading to changes in viscoelasticity and eventually germination [20, 136].

Cells also require optimal temperature conditions for their survival. Temperature affects the proper function and conformation of biomolecules, and hence indirectly affects cell mechanical properties. The rigidity of *E. coli* has been found to increase as a function of temperature, attributed to the folding of lipoproteins in the outer membrane leading to an increase in turgor pressure [136]. An earlier study showed that a temperature sensitive mutant of *B. subtilis* (indole- and thymine- ) was converted to a spherical shape from the wild type rod shape, attributed to loosening of the cell wall strength [137]. Hochmuth et al. [7] demonstrated that for red blood cells the time dependent viscoelastic behavior was reduced with increasing temperature.

The elastic behavior of the bacterial cell surface depends on the cell volume and ionic strength of the surrounding medium, which is related to its osmolarity. Abu-Lail and Camesano [138] observed that the elasticity of *E. coli* increased with reduced solvent polarity, and that bacteria in the least polar solvent have the highest Young's modulus [138]. Further, the spring constant of bacteria in a high ionic strength solvent of is higher than that in low ionic strength.
