A versatile micromodel technology to explore biofilm development in porous media flows.

Bacterial biofilms that grow in porous media are critical to ecosystem processes and applications ranging from soil bioremediation to bioreactors for treating wastewater or producing value-added products. However, understanding and engineering the complex phenomena that drive the development of biofilms in such systems remains a challenge. Here we present a novel micromodel technology to explore bacterial biofilm development in porous media flows. The technology consists of a set of modules that can be combined as required for any given experiment and conveniently tuned for specific requirements. The core module is a 3D-printed micromodel where biofilm is grown into a perfusable porous substrate. High-precision additive manufacturing, in particular stereolithography, is used to fabricate porous scaffolds with precisely controlled architectures integrating flow channels with diameters down to several hundreds of micrometers. The system is instrumented with: ultraviolet-C light-emitting diodes; on-line measurements of oxygen consumption and pressure drop across the porous medium; camera and spectrophotometric cells for the detection of biofilm detachment events at the outlet. We demonstrate how this technology can be used to study the development of Pseudomonas aeruginosa biofilm for several days within a network of flow channels. We find complex dynamics whereby oxygen consumption reaches a steady-state but not the pressure drop, which instead features a permanent regime with large fluctuations. We further use X-ray computed microtomography to image the spatial distribution of biofilms and computational fluid dynamics to link biofilm development with local flow properties. By combining the advantages of additive manufacturing for the creation of reproducible 3D porous microarchitectures with the flow control and instrumentation accuracy of microfluidics, our system provides a platform to study the dynamics of biofilm development in 3D porous media and to rapidly test new concepts in process engineering.

Ultraviolet control of bacterial biofilms in microfluidic chips.

Polydimethylsiloxane (PDMS) microfluidic systems have been instrumental in better understanding couplings between physical mechanisms and bacterial biofilm processes, such as hydrodynamic effects. However, precise control of the growth conditions, for example, the initial distribution of cells on the substrate or the boundary conditions in a flow system, has remained challenging. Furthermore, undesired bacterial colonization in crucial parts of the systems, in particular, in mixing zones or tubing, is an important factor that strongly limits the duration of the experiments and, therefore, impedes our ability to study the biophysics of biofilm evolving over long periods of time, as found in the environment, in engineering, or in medicine. Here, we develop a new approach that uses ultraviolet-C (UV-C) light-emitting diodes (LEDs) to confine bacterial development to specific zones of interest in the flow channels. The LEDs are integrated into a 3D printed light guide that is positioned upon the chip and used to irradiate germicidal UV-C directly through the PDMS. We first demonstrate that this system is successful in controlling undesired growth of Pseudomonas aeruginosa biofilm in inlet and outlet mixing zones during 48 h. We further illustrate how this can be used to define the initial distribution of bacteria to perturb already formed biofilms during an experiment and to control colonization for seven days-and possibly longer periods of time-therefore opening the way toward long-term biofilm experiments in microfluidic devices. Our approach is easily generalizable to existing devices at low cost and may, thus, become a standard in biofilm experiments in PDMS microfluidics.