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Item type:Publication, Access status: with files , Living electronics: coupling S. oneidensis MtrC-SpyTag cells to SpyCatcher-functionalized electrodes for direct electron transfer(Elsevier, 2026-08-15); ; ; ; This work set out to establish an easily applicable system to improve a broad range of bioelectronic devices using the SpyTag-SpyCatcher crosslinking system together with one of the model organisms for extracellular electron transport, Shewanella oneidensis . Therefore, the surface-displayed c -type cytochrome MtrC was equipped with an accessible SpyTag and coupled to SpyCatcher-functionalized surfaces. A transposon screen followed by nanopore sequencing was conducted in order to identify integration positions which facilitate MtrC functionality while the SpyTag is surface accessible. Three integration positions (W314, Y417, T603) were chosen for further characterization. Expression of the MtrC-SpyTag constructs in a S. oneidensis strain lacking all outer membrane cytochromes restored the ability to reduce an extracellular electron acceptor. Two of the three strains reached reduction rates at the wildtype MtrC level proving that the integrated SpyTag does not hamper extracellular electron transfer. In vivo, all three constructs showed significantly better binding properties to SpyCatcher functionalized magnetic beads than the wildtype MtrC control. The two most promising candidates were coupled to conductive, magnetic gold nanoparticles and directed towards a screen-printed electrode showcasing how MtrC-SpyTag expression can improve bioelectronic devices. A significantly higher charge transfer compared to the wildtype control was reached in linear sweep voltammetry and chronoamperometry experiments. Moreover, a shift towards direct electron transfer was observed which reduces the problem of redox shuttle washout in flowthrough systems. Direct binding of the cells to SpyCatcher functionalized electrodes enabled robust current production even after thorough washing of the electrodes while control cells failed to produce current under these conditions. The versatile SpyCatcher toolbox can be used together with the here reported strains to eliminate bottlenecks in bioelectronics like poor biofilm formation or the production of an insulating extracellular polymer matrix.Publicationtype: Journal Article47 11 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Tailored enzyme expression modifies Shewanella oneidensis biofilms and increases current density(Elsevier, 2025-06-09); ; ; ; Biofilm formation is the most effective pathway for electron transfer to anodes in bioelectrochemical systems. However, the mechanisms triggering biofilm formation under anoxic conditions, as well as the architectural and compositional factors that positively influence current generation, are not well understood. Recent findings have shown that riboflavin can function similarly to a quorum sensing molecule in the γ-proteobacterium Shewanella oneidensis. Enhanced biofilm formation induced by riboflavin correlates with increased current densities. Only a limited number of candidate proteins were found to have altered concentrations due to this quorum sensing mechanism. This study demonstrates that the catalytic functions of the UDP-N-acetylglucosamine C4 epimerase WbpP and UDP-N-acetyl-D-glucosamine 6-dehydrogenase WbpA affect biofilm formation and lead to increased current density. Using optical coherence tomography, we found that the expression of each protein individually causes specific, quantifiable changes in biofilm architecture, including biovolume, height, and porosity. However, the current density did not significantly differ when these proteins were expressed alone compared to the control. In contrast, co-expression of WbpP and WbpA resulted in a doubling of current density, closely resembling the increases observed with riboflavin-mediated quorum sensing. We hypothesize that riboflavin-based quorum sensing may lead, through several intermediary steps, to the overproduction of WbpA and WbpP, resulting in better attachment to graphite anodes and thus higher current densities.Publicationtype: Journal Article9 18 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Comparative analysis of the influence of BpfA and BpfG on biofilm development and current density in Shewanella oneidensis under oxic, fumarate- and anode-respiring conditions(Nature Publishing Group, 2024-10-05); ; ; ; Biofilm formation by Shewanella oneidensis has been extensively studied under oxic conditions; however, relatively little is known about biofilm formation under anoxic conditions and how biofilm architecture and composition can positively influence current generation in bioelectrochemical systems. In this study, we utilized a recently developed microfluidic biofilm analysis setup with automated 3D imaging to investigate the effects of extracellular electron acceptors and synthetic modifications to the extracellular polymeric matrix on biofilm formation. Our results with the wild type strain demonstrate robust biofilm formation even under anoxic conditions when fumarate is used as the electron acceptor. However, this pattern shifts when a graphite electrode is employed as the electron acceptor, resulting in biofilm formation falling below the detection limit of the optical coherence tomography imaging system. To manipulate biofilm formation, we aimed to express BpfG with a single amino acid substitution in the catalytic center (C116S) and to overexpress bpfA. Our analyses indicate that, under oxic conditions, overarching mechanisms predominantly influence biofilm development, rather than the specific mutations we investigated. Under anoxic conditions, the bpfG mutation led to a quantitative increase in biofilm formation, but both strains exhibited significant qualitative changes in biofilm architecture compared to the controls. When an anode was used as the sole electron acceptor, both the bpfA and bpfG mutations positively impacted mean current density, yielding a 1.8-fold increase for each mutation.Publicationtype: Journal Article11 9 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Magnetic, conductive nanoparticles as building blocks for steerable micropillar-structured anodic biofilmsIn bioelectrochemical systems (BES), biofilm formation and architecture are of crucial importance, especially for flow-through applications. The interface between electroactive microorganisms and the electrode surface plays an important and often limiting role, as the available surface area influences current generation, especially for poor biofilm forming organisms. To overcome the limitation of the available electrode surface, nanoparticles (NPs) with a magnetic iron core and a conductive, hydrophobic carbon shell were used as building blocks to form conductive, magnetic micropillars on the anode surface. The formation of this dynamic three-dimensional electrode architecture was monitored and quantified in situ using optical coherence tomography (OCT) in conjunction with microfluidic BES systems. By cyclic voltammetry the assembled three-dimensional anode extensions were found to be electrically conductive and increased the available electroactive surface area. The NPs were used as controllable carriers for the electroactive model organisms Shewanella oneidensis and Geobacter sulfurreducens, resulting in a 5-fold increase in steady-state current density for S. oneidensis, which could be increased 22-fold when combined with Poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT:PSS) aggregates. In the case of G. sulfurreducens, the steady-state current density was not increased, but was achieved four times faster. The study presents a controllable, scalable and easy-to-use method to increase the electrode surface area in existing BES by applying a magnetic field and adding conductive magnetic NPs. These findings can most likely also be transferred to other electroactive microorganisms.Publicationtype: Journal Article10 21 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Elucidating the development of cooperative anode-biofilm-structures(2024-06-01); ; ; Microbial electrochemical systems are a highly versatile platform technology with a particular focus on the interplay of chemical and electrical energy conversion and offer immense potential for a sustainable bioeconomy. The industrial realization of this potential requires a critical focus on biofilm optimization if performance is to be controlled over a long period of time. Moreover, the aspect and influence of cooperativity has to be addressed as many applied anodic bioelectrochemical systems will most likely be operated with a diversity of interacting microbial species. Hence, the aim of this study was to analyze how interspecies dependence and cooperativity of a model community influence the development of anodic biofilms. To investigate biofilm activity in a spatially resolved manner, a microfluidic bioelectrochemical flow cell was developed that can be equipped with user-defined electrode materials and operates under laminar flow conditions. With this infrastructure, the development of single and co-culture biofilms of the two model organisms Shewanella oneidensis and Geobacter sulfurreducens on graphite electrodes was monitored by optical coherence tomography analysis. The interdependence in the co-culture biofilm was achieved by feeding the community with lactate, which is converted by S. oneidensis into acetate, which in turn serves as substrate for G. sulfurreducens. The results show that co-cultivation resulted in the formation of denser biofilms than in single culture. Moreover, we hypothesize that S. oneidensis in return utilizes the conductive biofilm matrix build by G. sulfurreducens for direct interspecies electron transfer (DIET) to the anode. FISH analysis revealed that the biofilms consisted of approximately two-thirds G. sulfurreducens cells, which most likely formed a conductive 3D network throughout the biofilm matrix, in which evenly distributed tubular S. oneidensis colonies were embedded without direct contact to the anode surface. Live/dead staining shows that the outermost biofilm contained almost exclusively dead cells (98 %), layers near the anode contained 45–56 % and the entire biofilm contained 82 % live cells. Our results exemplify how the architecture of the exoelectrogenic biofilm dynamically adapts to the respective process conditions.Publicationtype: Journal Article5 93 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Automatisierte Charakterisierung und Optimierung elektroaktiver Biofilme(2024); Microbial electrochemical systems (MES) represent a highly versatile platform technology with an immense potential for the development towards a sustainable bioeconomy, whereby the industrial realization of this potential requires a critical focus on biofilm optimization. In order to be able to directly correlate biofilm development and current production as well as spatially resolve biofilm activity, a microfluidic cultivation platform with integrated automated three-dimensional imaging was established. The platform was validated by investigating a model community of G. sulfurreducens and S. oneidensis, revealing a co-dependency in which G. sulfurreducens relies on the nutrient supply from S. oneidensis, while S. oneidensis in turn utilizes the conductive biofilm matrix developed by G. sulfurreducens, facilitating direct interspecies electron transfer (DIET) to the anode. Moreover, the influence of different genetic factors on the biofilm development of \textit{S.~oneidensis} was fundamentally investigated. The host strain used for these studies was S. oneidensis MR-1 Δλ, based on the hypothesis that the Δλ-induced reduction of extracellular polymeric substances (EPS) significantly facilitates electron transport through the biofilm. By investigating various genetic factors, it could be shown that under oxic conditions, superordinate mechanisms seem to take effect and determine biofilm development, whereas under anoxic conditions with fumarate as electron acceptor, both qualitative and quantitative differences compared to the control strains could be determined. Using an anode as the terminal electron acceptor, co-overexpression of wbpA and wbpP, overexpression of bpfA, a point-mutated bpfG variant and the simple addition of an empty plasmid vector led to a 1.67 - 1.85-fold increase in mean current density. In summary, this work identified and characterized key factors of anaerobic biofilm formation, providing a basis for the cultivation of biofilms with optimized user-defined properties.Publicationtype: Doctoral Thesis54 94 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Microbe–anode interactions : comparing the impact of genetic and material engineering approaches to improve the performance of microbial electrochemical systems (MES)(2023-02-18); ; Microbial electrochemical systems (MESs) are a highly versatile platform technology with a particular focus on power or energy production. Often, they are used in combination with substrate conversion (e.g., wastewater treatment) and production of value-added compounds via electrode-assisted fermentation. This rapidly evolving field has seen great improvements both technically and biologically, but this interdisciplinarity sometimes hampers overseeing strategies to increase process efficiency. In this review, we first briefly summarize the terminology of the technology and outline the biological background that is essential for understanding and thus improving MES technology. Thereafter, recent research on improvements at the biofilm–electrode interface will be summarized and discussed, distinguishing between biotic and abiotic approaches. The two approaches are then compared, and resulting future directions are discussed. This mini-review therefore provides basic knowledge of MES technology and the underlying microbiology in general and reviews recent improvements at the bacteria–electrode interface.Publicationtype: Journal ArticleScopus© Citations 4 22 81 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Enrichment of phosphate-accumulating organisms (PAOs) in a microfluidic model biofilm system by mimicking a typical aerobic granular sludge feast/famine regime(Springer, 2022-01-15); ; ; ; Wastewater treatment using aerobic granular sludge has gained increasing interest due to its advantages compared to conventional activated sludge. The technology allows simultaneous removal of organic carbon, nitrogen, and phosphorus in a single reactor system and is independent of space-intensive settling tanks. However, due to the microscale, an analysis of processes and microbial population along the radius of granules is challenging. Here, we introduce a model system for aerobic granular sludge on a small scale by using a machine-assisted microfluidic cultivation platform. With an implemented logic module that controls solenoid valves, we realized alternating oxic hunger and anoxic feeding phases for the biofilms growing within. Sampling during ongoing anoxic cultivation directly from the cultivation channel was achieved with a robotic sampling device. Analysis of the biofilms was conducted using optical coherence tomography, fluorescence in situ hybridization, and amplicon sequencing. Using this setup, it was possible to significantly enrich the percentage of polyphosphate-accumulating organisms (PAO) belonging to the family Rhodocyclaceae in the community compared to the starting inoculum. With the aid of this miniature model system, it is now possible to investigate the influence of a multitude of process parameters in a highly parallel way to understand and efficiently optimize aerobic granular sludge-based wastewater treatment systems. Key points • Development of a microfluidic model to study EBPR. • Feast-famine regime enriches polyphosphate-accumulating organisms (PAOs). • Microfluidics replace sequencing batch reactors for aerobic granular sludge research.Publicationtype: Journal ArticleScopus© Citations 5 52 173 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: without files , Cultivation of Exoelectrogenic Bacteria in Conductive DNA Nanocomposite Hydrogels Yields a Programmable Biohybrid Materials System(2020-04-01); ; ; ; The use of living microorganisms integrated within electrochemical devices is an expanding field of research, with applications in microbial fuel cells, microbial biosensors or bioreactors. We describe the use of porous nanocomposite materials prepared by DNA polymerization of carbon nanotubes (CNTs) and silica nanoparticles (SiNPs) for the construction of a programmable biohybrid system containing the exoelectrogenic bacterium Shewanella oneidensis. We initially demonstrate the electrical conductivity of the CNT-containing DNA composite by employment of chronopotentiometry, electrochemical impedance spectroscopy, and cyclic voltammetry. Cultivation of Shewanella oneidensis in the conductive materials shows that the exoelectrogenic bacteria populate the matrix of the conductive composite, while nonexoelectrogenic Escherichia coli remain on its surface. Moreover, the ability to use extracellular electron transfer pathways is positively correlated with the number of cells within the conductive synthetic biofilm matrix. The Shewanella-containing composite remains stable for several days and shows electrochemical activity, indicating that the conductive backbone is capable of extracting the metabolic electrons produced by the bacteria under strictly anoxic conditions and conducting them to the anode. Programmability of this biohybrid material system is demonstrated by on-demand release and degradation induced by a short-term enzymatic stimulus. We believe that the application possibilities of such biohybrid materials could even go beyond microbial biosensors, bioreactors, and fuel cell systems.Publicationtype: Journal ArticleScopus© Citations 31 52 - Some of the metrics are blocked by yourconsent settings
Item type:Publication, Access status: with files , Kultivierung und Analyse von Biofilmen durch automatisierte MikrosystemeBiofilms are the most abundant growth form of microorganisms in nature. To exploit these living systems for applications in biotechnology, we have developed a microfluidic cultivation platform with an integrated automatic sampling robot that enables the characterization of biofilms with high spatiotemporal resolution. Moreover, in situ imaging and anoxic or gas-consumption based cultivation can be applied. Biocatalytic productivity can be assessed using a spectrum of analytical technologies.Publicationtype: Journal Article67 126