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Biological Physics of Living Active Nematics

Periodic Reporting for period 1 - BPLAN (Biological Physics of Living Active Nematics)

Reporting period: 2019-10-01 to 2021-09-30

On surfaces, bacterial micro-colonies first develop a monolayer of cells before a bacterium leaves the plane of the monolayer and initiates the formation of a second layer. This transition is a major event in biofilm formation since this extra layer offers shelter for bacteria underneath. Understanding the determinants of this transition will help design new strategies for hampering biofilm development.

When growing in monolayers, rod-shaped bacteria behave as active nematics. But unlike most nematics, which are active due to particle motility, nematics made of sessile growing bacteria are active because of cell elongation. In addition, bacterial nematics of such a kind do not only interact by steric repulsion but bacteria also adhere to the substrate and to their neighbours. Bacterial adhesion therefore contributes to the formation of topological defects in bacterial nematics. However, we do not yet know how topological defects are coupled to the distribution of adhesions within the monolayer and which of these two contributions account for the formation of the second layer.

The Marie-Curie Action has allowed to unravel the coupling between cell-cell adhesion and topological defects. Our results indicate that the rate of defect generation has a direct impact on the shape of the microcolony. The higher this rate, the rounder the micro-colonies. Surprisingly, we showed that increasing cell-cell adhesion reduces the rate of topological defects generation, resulting in more elongated micro-colonies, which are more exposed to the environment. Our results therefore suggest that treatment including biochemical agents capable of gluing cells together may constitute a strategy to expose more bacteria to antibiotic treatments. Indeed, such treatments will slow down the dynamics topological defect formation and thus increase the exposure of micro-colonies to the environment.
We perfomed TFM microscopy experiments. Unfortunately, it turned out that the rigidity of the PAA gel was a limiting technical factor for the study of topological defects. Therefore, we opted for an alternative strategy based on fluorescent reporters that also allows to study the coupling between bacterial adhesion and topological defects. For this purpose, we developed a protocol to perform live microscopy while visualizing adhesion proteins (TIRF setup). We focused on Ag43, which mediates cell-cell adhesion through homophilic interactions. Through the TIRF observations, we could visualize the distribution of the protein Ag43, and, for different conditions (temperature, cell-cell adhesion over-expression), we measured the dynamics of topological defects during microcolony growth.

In the WT strain, we first showed that the rate of topological defect formation increased with growth rate, i.e. with the temperature, while the lifetime of topological defects decreased with growth rate. We then showed that the lifetime of the -1/2-charged defects and the rate of defect generation were positively and negatively correlated to the level of cell-cell adhesion, respectively. Our results also clearly establish that topological defects influence neither the size of microcolony at second layer formation nor the location of the second layer initiation site. However, our results indicate that the rate of defects generation has a direct impact on the shape of the microcolony. The higher this rate, the rounder the micro-colonies. Surprisingly, we observed that topological defects were generated at a lower rate upon overexpression of cell-cell adhesion. Consistently, we observed that micro-colonies with higher cell-cell adhesion are more elongated.

We published the part related to the methodology (Chekli et al. Sci. Rep. 2020). We will soon publish the part related to the coupling between adhesion and defects.
We discovered that slowing down the dynamics of topological defect formation is a way to make microcolonies more elongated, which means more exposed to their environment and thus more susceptible to antibiotics. Our results therefore suggest that a treatment including biochemical agents to glue cells together may constitute a strategy to expose more bacteria to antibiotic treatments.

During the course of the project, a group of journalism students visited the lab and observed our daily practice to deliver a visual of the research process (Collaboration with the school of graphic artl: EPSAA). We also started a collaboration with a design lab at the ENSAD (Ecole Nationale Superieure des Arts Décoratifs). Finally, several students with a physics or biology background joined the group for periods from 3 to 6 months, being exposed to and actively applying the research methods directly on the field.
-1/2 topological defect and localization of cell-cell adhesin Ag43