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Bacterial biofilms in porous structures: from biomechanics to control

Periodic Reporting for period 1 - BEBOP (Bacterial biofilms in porous structures: from biomechanics to control)

Reporting period: 2019-01-01 to 2020-06-30

The key ideas motivating this project are that: 1) precise control of the properties of porous systems can be obtained by exploiting bacteria and their fantastic abilities; 2) conversely, porous media (large surface to volume ratios, complex structures) could be a major part of bacterial synthetic biology, as a scaffold for growing large quantities of microorganisms in controlled bioreactors.

The main scientific obstacle to precise control of such processes is the lack of understanding of biophysical mechanisms in complex porous structures, even in the case of single-strain biofilms. The central hypothesis of this project is that a better fundamental understanding of biofilm biomechanics and physical ecology will yield a novel theoretical basis for engineering and control.

The first scientific objective is thus to gain insight into how fluid flow, transport phenomena and biofilms interact within connected multiscale heterogeneous structures - a major scientific challenge with wide-ranging implications. To this end, we combine microfluidic and 3D printed micro-bioreactor experiments; fluorescence and X-ray imaging; high performance computing blending CFD, individual-based models and pore network approaches.

The second scientific objective is to create the primary building blocks toward a control theory of bacteria in porous media and innovative designs of microbial bioreactors. Building upon the previous objective, we first aim to extract from the complexity of biological responses the most universal engineering principles applying to such systems. We will then design a novel porous micro-bioreactor to demonstrate how the permeability and solute residence times can be controlled in a dynamic, reversible and stable way - an initial step toward controlling reaction rates.

We envision that this will unlock a new generation of biotechnologies and open the way toward completely different approaches to engineering with a variety of societal applications, for example in wastewater processing, bioremediation of soils or creating smart construction materials that self-regulate their properties. Further, this project may also lead to novel bioreactor designs enabling translation from proof-of-concept synthetic microbiology to industrial processes.

BEBOP is therefore structured around two scientific objectives (SOs) and two methodological objectives (MOs) as follows.

SO1. Advance our understanding of single-strain biofilms in connected, multiphase, heterogeneous structures - focusing on aspects that are either fundamental (e.g. couplings & regimes) or promising for engineering bacteria in porous media (e.g. biomass control by chemicals or predation).

SO2. Advance toward a control theory of biofilms in porous media and develop a demonstration micro-bioreactor with an innovative design allowing us to control - in a dynamic, stable and reversible way - two outputs: the permeability and solute residence times.

MO1. Develop a multiscale, multimodal imaging framework for biofilms in porous media combining (microfluidics + fluorescence imaging) and (3D printing + functionalized X-ray microtomography).

MO2. Develop a multiscale (cell- to bioreactor-scale) modelling framework for biofilms in porous media combining individual-based models, computational fluid dynamics and network/graph approaches.
During this first part of the project, advances have been primarily focused on methodological aspects (MO1 and MO2) with preliminary results regarding the fundamentals of biofilm growth in porous media (SO1).

We have established the most important part of the microfluidics/microscopy setup, which has allowed us to start exploring the impact of flow and of ecological interactions on Pseudomonas aeruginosa biofilm-induced clogging in model network structures.

We have modified the microbioreactor system with better confinement of biological activity to the porous structure, improved measurements and more precise control of oxygenation levels at the inlet. The bioreactor designs have also been upgraded and fabricated using 3D printing approaches such as photolithography. Preliminary results have been obtained for using nanoparticles as an X-ray tomography contrast agent for imaging biofilm development in the bioreactors.

We have used individual-based models to explore different aspects of bacterial ecology in dense populations, notably the impact of competition mechanisms (paper submitted) and of solid obstacles on cellular mixing within the population. A network approach has also been implemented to simulate biofilm growth under flow in large porous networks.
Through a better understanding of bacterial biomechanics and physical ecology in porous media, we aim to generate a paradigm shift for process design and control of bacteria in porous media. Indeed, much emphasis has recently been given to engineering consortia and to structural design, but not to understanding fundamental mechanisms.

To do so, we have focused on developing two novel experimental setups in MO1, combining observations in (microfluidics + fluorescence imaging) and (3D printing + functionalized X-ray microtomography). This combination in our lab is beyond the state of the art and is allowing us to study biofilm growth in a wide variety of regimes for both confined quasi-2D microfluidics and 3D microbioreactors.

We have also layed the groundwork for MO2, developing a network approach that will be used to model biofilm development in large porous structures. At smaller scales, individual-based models have further been used to answer new questions on how competition mechanisms operate in dense populations and on the impact of solid obstacles, more generally of heterogeneities, on the development of biofilms.

Ultimately, we aim at using this fundamental knowledge to create the primary building blocks toward a control theory of bacteria in porous media and innovative designs of microbial bioreactors. We envision that this will unlock a new generation of biotechnologies and open the way toward completely different approaches to engineering with a variety of societal applications, for example in wastewater processing, bioremediation of soils or creating smart construction materials that self-regulate their properties. This project may also lead to novel bioreactor designs enabling translation from proof-of-concept synthetic microbiology to industrial processes
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