2,3-Butandiol production using biofilm clusters bound to magnetic particles

• Applicants: Professor Dr. Johannes Gescher; Professorin Dr. Karin Jacobs; Professor Dr. Rainer Udo Meckenstock
• Subject Area: Biological Process Engineering
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 559358135

Despite their potential, several challenges hinder the implementation of biofilm processes in white biotechnology. Key challenges include the unpredictable retention of biomass in biofilms, the absence of established model processes, and limited knowledge about the genetic stability of genetically modified strains in continuous biofilm-based processes. Furthermore, it will be crucial to minimize diffusion limitations in order to successfully optimize biofilm processes. We aim to address these challenges by using the biofilm-based production of the platform chemical 2,3-butanediol as a model process. We seek to develop Shewanella oneidensis biofilms that produce 2,3-butanediol and are bound to magnetic nanoparticles. These nanoparticles will enable the manipulation of biofilm density, morphology, and movement in response to applied magnetic fields, as well as facilitate the transport of electrons to oxygen-rich areas within the reactor. We also aim to genetically control the properties of the biofilm and stabilize the cells genetically through transposase deactivation. The planned experiments will allow us to answer a series of fundamental questions regarding the application of biofilms in production processes and evaluate engineering solutions for existing problems. We have organized the necessary work into six interconnected work packages (WPs): - WP1: We will develop S. oneidensis strains with a predictable and robust extracellular polymeric matrix, analyzing the surface characteristics of these strains using single-cell force spectroscopy. The cells will be specifically connected to magnetic conductive nanoparticles, which will aid in steering the biofilm architecture using permanent magnets. - WP2: We will enhance genetic stability by deactivating transposases and analyze the correlation between transposase deactivation and production stability. Additionally, we will compare long-term productivity in biofilm and planktonic systems to understand how the form of cell application influences production stability. - WP3: We will improve our robotic infrastructure by integrating a robotic arm that allows for programmable movement of permanent magnets to modulate biofilm architecture, as well as facilitate reproducible placement of the setups on a microscope stage for fluorescence microscopy inspection. - WP4: We will analyze biofilm productivity under different movement regimes of the permanent magnets and investigate methods to increase productivity through modulation of biofilm structure. - WP5: We will utilize Raman micro-spectroscopy to analyze the three-dimensional profile of metabolic activity in the biofilms developed in WP4 and compare overall metabolic productivity with the productivity of 2,3-butanediol. - WP6: We will develop a Raman-assisted cell sorting method that will be coupled with transcriptomic and genomic sequencing approaches to analyze how metabolic activity correlates with the genetic changes that occur over time.

• DFG Programme: Priority Programmes
• Subproject of: SPP 2494: Productive Biofilm Systems
• Major Instrumentation: Fluoreszenzmikroskop
• Instrumentation Group: 5000 Labormikroskope