Antibiotic tolerance of biofilms emerging from multicellular effects of antibiotic efflux

• Applicants: Professorin Dr. Rosalind Allen; Dr. Frank Schreiber
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms ; Microbial Ecology and Applied Microbiology
• Term: since 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 503989054

Biofilms are multicellular assemblies of bacteria in which cell-cell interactions via metabolism and signalling lead to emergent structural and functional properties. Biofilms are notoriously difficult to kill with antimicrobial substances, a phenomenon that is known as tolerance. This is an emergent multicellular trait, since biofilms show much greater tolerance than planktonic cells. Tolerance is distinct from antimicrobial resistance, but it can be a stepping stone on the pathway to resistance. Our project focuses on the molecular mechanisms that lead to emerging antimicrobial tolerance. Previous studies suggest a key role for multi-drug efflux pumps, both via efflux-mediated spatial interactions and via a coupling between efflux and the generation of non-growing persister cells. We aim to develop a predictive model for efflux-mediated antimicrobial tolerance in bacterial multicellular assemblies. Our central hypothesis is that efflux pump activity causes emergent antibiotic tolerance of multicellular bacterial populations, through the interplay of efflux-mediated spatial interactions and efflux-linked persistence. To achieve a detailed, yet global, quantitative understanding, we will combine microbiological experiments and mathematical modelling, and we will integrate information from 3 types of multicellular assembly: colonies, cell-to-cell interactions in a microfluidic device, and 3D flow chamber biofilms. First, we will quantify efflux-mediated spatial structure and persistence in bacterial colonies and determine the consequences for antimicrobial tolerance. We will measure colony morphology, spatial patterns of efflux expression and colony tolerance, for strains that differ in their levels of efflux. A microhabitat-based mathematical model that accounts for efflux-mediated neighbour inhibition and persister formation will rationalise our observations. Second, we will quantify efflux-mediated cell-cell interactions and persistence at the single-cell level in a microfluidic growth setup. We will measure the dependence of efflux pumping and persister formation on nutrient conditions, the correlation between efflux and persisters, and the spatial range of efflux-mediated interactions. Using this information as input parameters, we will finally predict and measure efflux-mediated spatial structure and persistence, and the emergent antibiotic tolerance, in complex 3D biofilms. Using individual-based modelling, we will predict biofilm spatial structure development, patterns of efflux and persister formation and tolerance. These predictions will be directly tested with flow chamber biofilm experiments. Our project will contribute to the central aspects of the SPP2389. It will elucidate the physiological benefits and molecular mechanisms of an emergent function – here tolerance - and the underlying architecture, dynamics and biophysical properties of the multicellular form that gives rise to this emergent function.

• DFG Programme: Priority Programmes
• Subproject of: SPP 2389: Emergent Functions of Bacterial Multicellularity