3.1 Mechanical properties of synthetic and P. aeruginosabiofilms
Rheometry is commonly used to measure the mechanical properties of viscoelastic materials. It is also widely used to measure biofilm mechanical properties, as it is suitable for viscoelastic solids and non-Newtonian fluids. Averaged properties of a sample are measured, providing the relationship between force, deformation, and time. Our goal was to obtain the biofilm mechanical parameters as inputs for the computational model. As a first step, alginate with embedded microbial cells was used as a synthetic biofilm, since it is mechanically more homogenous and it can be formed into a regular geometry. A regular geometry is desirable for both rheometry analysis and for simulations. We also tested a homogenized P. aeruginosa biofilm.
In order to confirm the homogeneity of synthetic biofilms, G′ and G′′ from strain sweep test were discussed (Fig. 3). The storage modulus G′, representing elasticity, and loss modulus G′′, representing viscosity, were calculated. Averaged G′ and G′′ from synthetic biofilm were 251.3 Pa ± 7.1 Pa and 18.2 Pa ± 5.5 Pa, respectively. A higher G′ compared to G′′ also revealed that the synthetic biofilm samples were more elastic than viscous. Error bars in Fig. 3 show the standard deviations from three different locations of the same sample. The small standard errors indicated a small magnitude of the bias, which demonstrated that the alginate is mechanically homogeneous and can provide a simplified control.
The values of G′ and G′′ for the P. aeruginosa biofilm were 858 ± 36 Pa and 98 ± 12 Pa. These moduli were in the same order of magnitude as those of the synthetic biofilm sample, which indicates that the alginate sample was a reasonable surrogate of a real biofilm. Previous studies on different types of biofilms also suggested a wide range G′ and G′′ from rheometry tests, ranging from 101-103 Pa for both G′ and G′′ (Böl et al., 2012). The synthetic biofilm and P. aeruginosa biofilm in this study were within the range of reported data, with a relatively small standard deviation.
Stress relaxation tests were performed with both synthetic and P. aeruginosa biofilm samples (Fig. 4). The stress relaxation curve (black-square markers) demonstrates that after an initial stress response (elastic response), the viscous response recovers the stress over time when constant strain was applied on the sample. The shape of both curves indicated that both samples behaved as typical viscoelastic materials.
The fitted Maxwell model was also plotted as a green line in Fig. 4. The residual sum of squares (RSS) were compared from both samples in nonlinear least-square fitting. The RSS of synthetic biofilm was 45.4 while the RSS of homogenized P. aeruginosa biofilm was 152. The alginate sample had a better fit than the real biofilm. For a more accurate fitting, more elements (springs and dashpots) can be added to the Maxwell model (Peterson et al., 2013).