Development of a Microbioreactor for Bacillus subtilis Biofilm Cultivation

To improve our understanding of <i>Bacillus subtilis</i> growth and biofilm formation under different environmental conditions, two versions of a microfluidic reactor with two channels separated by a polydimethylsiloxane (PDMS) membrane were developed. The gas phase was introduced into the channel above the membrane, and oxygen transfer from the gas phase through the membrane was assessed by measuring the dissolved oxygen concentration in the liquid phase using a miniaturized optical sensor and oxygen-sensitive nanoparticles. <i>B. subtilis</i> biofilm formation was monitored in the growth channels of the microbioreactors, which were designed in two shapes: one with circular extensions and one without. The volumes of these microbioreactors were (17 ± 4) μL for the reactors without extensions and (28 ± 4) μL for those with extensions. The effect of microbioreactor geometry and aeration on <i>B. subtilis</i> biofilm growth was evaluated by digital image analysis. In both microbioreactor geometries, stable <i>B. subtilis</i> biofilm formation was achieved after 72 h of incubation at a growth medium flow rate of 1 μL/min. The amount of oxygen significantly influenced biofilm formation. When the culture was cultivated with a continuous air supply, biofilm surface coverage and biomass concentration were higher than in cultivations without aeration or with a 100% oxygen supply. The channel geometry with circular extensions did not lead to a higher total biomass in the microbioreactor compared to the geometry without extensions.