Physico-chemical determinants of bacterial interactions – PHYSCHEMBACT

Scientific context and objectives: The interactions of bacteria with their close environment involve biological and/or physico-chemical phenomena that are difficult to quantify at a molecular scale. Over the past 10 years, most of the studies dedicated to deriving the surface properties of bacterial systems were done according to macroscopic and averaging measurements. Such approaches suffered in many cases from overlooking the intrinsic relationship between bioparticle behavior and the complexity of their interface as governed by hydrodynamic permeability, chemical and structural heterogeneities and mechanical softness. Given these elements, the definition and quantification of cell surface properties in terms of softness, interfacial deformation, flow permeation within the bacteria and bacterial envelop dynamics remain challenging to achieve a consistent physico-chemical description valid at different scales, from the molecular compounds of the cell membrane to the cell population interactions. The aim of this project is the identification and quantitative understanding of the molecular to macromolecular determinants that regulate the interactions between bacteria and its environment under steady and dynamic conditions. The originality of the project relies on the combination between innovative experimental and theoretical approaches that tend to investigate cell properties according to a multiscale approach with the benefit of modulating via genetic means bacterial surface phenotypes of the chosen bacterial model Escherichia coli K12. The project will be carried out in two phases. At first the relationships between structure and interfacial properties will be investigated from physico-chemical measurements under equilibrium or non-equilibrium conditions (Work stage 1 to 4). In a second phase, the modifications of these properties in response to the addition and/or presence of antiadhesive macromolecules or antibiotics will be explored (Work stage 5). This interdisciplinary project coordinated by a young researcher from the LCPME combines the different expertise met in this laboratory (physico-chemists, microbiologists, chemists) and also that of a local partner (LEM) through a young researcher who brings the mandatory theoretical background for quantitatively interpreting the data. Furthermore, the genetic constructions of the different strains will be realized at the GGB from Pasteur Institute. Description and Methodology: The project will include five work stages: WS1: In this work stage, the electro-hydrodynamic properties of bacterial envelopes will be investigated via electrophoretic mobility and electrical conductivity measurements. In particular, the electrostatic field strength and local hydrodynamic permeability distributions across the external structures of the bacteria will be quantified using a theoretical development that takes into account the details of the surface structure as well as its soft and diffuse characters. WS2: Force spectroscopy will be carried out to map the physical properties of cells (mechanical and viscoelastic properties) and the non-specific interactions (electrostatic, hydrophobic/hydrophilic balance) at two different scales, the nanometer scale and the cell scale. The cell-cell interactions will be also explored by attempting the construction of bacterial probes. WS3: The dynamics of bacterial cells will be evaluated from electrical and mechanical perturbation measurements carried out by force spectroscopy and dielectric relaxation. Theoretical approaches will be developed and shall complete those elaborated in WS1 to estimate the electrical double layer and/or the structural polarizations. WS4: Kinetics and strength of bacterial adhesion will be quantified by combining the classical approaches of flow chamber with AFM imaging that will enable to quantify the lateral forces required to detach the individual bacteria from the substrate they have colonized. WS5: Through the different methodologies developed in the four previous working stage, the modifications of the aforementioned parameters at the different scales investigated will be monitored after addition in the bacterial medium of polysaccharidic molecule that exhibits antiadhesive properties (capsular polysaccharide), and antibiotic molecule (ticarcilline). Expected results: A better physico-chemical capture of the mechanisms that govern the interactions between bacteria and other entities (bacterium, substratum, macromolecules) will be achieved following the methods mentioned before. Finally, this project will constitute a milestone toward the understanding of interactions between bacteria and other biological systems such as yeasts or viruses.