Beneficial applications of biofilms.

Many microorganisms live in the form of a biofilm. Although they are feared in the medical sector, biofilms that are composed of non-pathogenic organisms can be highly beneficial in many applications, including the production of bulk and fine chemicals. Biofilm systems are natural retentostats in which the biocatalysts can adapt and optimize their metabolism to different conditions over time. The adherent nature of biofilms allows them to be used in continuous systems in which the hydraulic retention time is much shorter than the doubling time of the biocatalysts. Moreover, the resilience of organisms growing in biofilms, together with the potential of uncoupling growth from catalytic activity, offers a wide range of opportunities. The ability to work with continuous systems using a potentially self-advancing whole-cell biocatalyst is attracting interest from a range of disciplines, from applied microbiology to materials science and from bioengineering to process engineering. The field of beneficial biofilms is rapidly evolving, with an increasing number of applications being explored, and the surge in demand for sustainable and biobased solutions and processes is accelerating advances in the field. This Review provides an overview of the research topics, challenges, applications and future directions in beneficial and applied biofilm research.

Electron transfer of extremophiles in bioelectrochemical systems.

The interaction of bacteria and archaea with electrodes is a relatively new research field which spans from fundamental to applied research and influences interdisciplinary research in the fields of microbiology, biochemistry, biotechnology as well as process engineering. Although a substantial understanding of electron transfer processes between microbes and anodes and between microbes and cathodes has been achieved in mesophilic organisms, the mechanisms used by microbes under extremophilic conditions are still in the early stages of discovery. Here, we review our current knowledge on the biochemical solutions that evolved for the interaction of extremophilic organisms with electrodes. To this end, the available knowledge on pure cultures of extremophilic microorganisms has been compiled and the study has been extended with the help of bioinformatic analyses on the potential distribution of different electron transfer mechanisms in extremophilic microorganisms.

Nanowired electrodes as outer membrane cytochrome-independent electronic conduit in Shewanella oneidensis.

Extracellular electron transfer (EET) from microorganisms to inorganic electrodes is a unique ability of electrochemically active bacteria. Despite rigorous genetic and biochemical screening of the c-type cytochromes that make up the EET network, the individual electron transfer steps over the cell membrane remain mostly unresolved. As such, attempts to transplant entire EET chains from native into non-native exoelectrogens have resulted in inferior electron transfer rates. In this study we investigate how nanostructured electrodes can interface with Shewanella oneidensis to establish an alternative EET pathway. Improved biocompatibility was observed for densely packed nanostructured surfaces with a low cell-nanowire load distribution during applied external forces. External gravitational forces were needed to establish a bioelectrochemical cell-nanorod interface. Bioelectrochemical analysis showed evidence of nanorod penetration beyond the outer cell membrane of a deletion mutant lacking all outer membrane cytochrome encoding genes that was only electroactive on a nanostructured surface and under external force.

Accelerated Electro-Fermentation of Acetoin in Escherichia coli by Identifying Physiological Limitations of the Electron Transfer Kinetics and the Central Metabolism.

Anode-assisted fermentations offer the benefit of an anoxic fermentation routine that can be applied to produce end-products with an oxidation state independent from the substrate. The whole cell biocatalyst transfers the surplus of electrons to an electrode that can be used as a non-depletable electron acceptor. So far, anode-assisted fermentations were shown to provide high carbon efficiencies but low space-time yields. This study aimed at increasing space-time yields of an Escherichia coli-based anode-assisted fermentation of glucose to acetoin. The experiments build on an obligate respiratory strain, that was advanced using selective adaptation and targeted strain development. Several transfers under respiratory conditions led to point mutations in the pfl, aceF and rpoC gene. These mutations increased anoxic growth by three-fold. Furthermore, overexpression of genes encoding a synthetic electron transport chain to methylene blue increased the electron transfer rate by 2.45-fold. Overall, these measures and a medium optimization increased the space-time yield in an electrode-assisted fermentation by 3.6-fold.

Genetic engineering for enhanced productivity in bioelectrochemical systems.

A shift from petrochemical processes toward a bio-based economy is one of the most advocated developments for a sustainable future. To achieve this will require the biotechnological production of platform chemicals that can be further processed by chemical engineering. Bioelectrochemical systems (BESs) are a novel tool within the biotechnology field. In BESs, microbes serve as biocatalysts for the production of biofuels and value-added compounds, as well as for the production of electricity. Although the general feasibility of bioelectrochemical processes has been demonstrated in recent years, much research has been conducted to develop biocatalysts better suited to meet industrial demands. Initially, mainly natural exoelectrogenic organisms were investigated for their performance in BESs. Driven by possibilities of recent developments in genetic engineering and synthetic biology, the spectrum of microbial catalysts and their versatility (substrate and product range) have expanded significantly. Despite these developments, there is still a tremendous gap between currently achievable space-time yields and current densities on the one hand and the theoretical limits of BESs on the other. It will be necessary to move the performance of the biocatalysts closer to the theoretical possibilities in order to establish viable production routines. This review summarizes the status quo of engineering microbial biocatalysts for anode-applications with high space-time yields. Furthermore, we will address some of the theoretical limitations of these processes exemplarily and discuss which of the present strategies might be combined to achieve highly synergistic effects and, thus, meet industrial demands.

Addition of Riboflavin-Coupled Magnetic Beads Increases Current Production in Bioelectrochemical Systems via the Increased Formation of Anode-Biofilms.

Shewanella oneidensis is one of the best-understood model organisms for extracellular electron transfer. Endogenously produced and exported flavin molecules seem to play an important role in this process and mediate the connection between respiratory enzymes on the cell surface and the insoluble substrate by acting as electron shuttle and cytochrome-bound cofactor. Consequently, the addition of riboflavin to a bioelectrochemical system (BES) containing S. oneidensis cells as biocatalyst leads to a strong current increase. Still, an external application of riboflavin to increase current production in continuously operating BESs does not seem to be applicable due to the constant washout of the soluble flavin compound. In this study, we developed a recyclable electron shuttle to overcome the limitation of mediator addition to BES. Riboflavin was coupled to magnetic beads that can easily be recycled from the medium. The effect on current production and cell distribution in a BES as well as the recovery rate and the stability of the beads was investigated. The addition of synthesized beads leads to a more than twofold higher current production, which was likely caused by increased biofilm production. Moreover, 90% of the flavin-coupled beads could be recovered from the BESs using a magnetic separator.

Electron transfer process in microbial electrochemical technologies: The role of cell-surface exposed conductive proteins.

Electroactive microorganisms have attracted significant interest for the development of novel biotechnological systems of low ecological footprint. These can be used for the sustainable production of energy, bioremediation of metal-contaminated environments and production of added-value products. Currently, almost 100 microorganisms from the Bacterial and Archaeal domains are considered electroactive, given their ability to efficiently interact with electrodes in microbial electrochemical technologies. Cell-surface exposed conductive proteins are key players in the electron transfer between cells and electrodes. Interestingly, it seems that among the electroactive organisms identified so far, these cell-surface proteins fall into one of four groups. In this review, the different types of cell-surface conductive proteins found in electroactive organisms will be overviewed, focusing on their structural and functional properties.