A Biomimetic Gellan-Based Hydrogel as a Physicochemical Biofilm Model

Biofilm-forming microorganisms are ubiquitous, but continuous cultivation of these microorganisms with predictable biofilm growth and structural properties remains challenging. The development of a reliable simulated biofilm has been limited by a lack of information about the microorganism subpopulations and fluid-structure interactions involved in biofilm formation and detachment due to mechanical stress. This paper presents a gellan-based hydrogel as an alternative material for a simulated physicochemical biofilm. The mechanical properties of the hydrogel in terms of the storage (G') and loss (G'') moduli can be tuned and adapted to imitate biofilms of different strengths by changing the concentration of gellan and mono(Na+) or divalent (Mg2+) ions. The storage modulus of the hydrogel ranges from 2 to 20 kPa, and the loss modulus ranges from 0.1 to 2.0 kPa. The material constants of the hydrogels and biofilms of Pseudomonas putida KT2440 were experimentally determined by rheometric analysis. A simplified biofilm imitate based on highly hydrolyzed gellan hydrogels was established by using experimental design techniques that permitted independent analyses regardless of growth. This model system design was compared to real biofilms and was adapted to mimic the mechanical properties of biofilms by changing the hydrogel composition, resulting in biofilm-like viscoelastic behavior. The use of a gellan-based hydrogel enables the imitation of biofilm behavior in the absence of growth effects, thus simplifying the system. Biofilm characterization tools can be tested and verified before their application to the measurement of slow-growing, highly variable biofilms to estimate system errors, which are often smaller than the biological variations. In general, this method permits faster and more reliable testing of biofilm mechanical properties.