Efficient Bioelectrochemical Conversion of Industrial Wastewater by Specific Strain Isolation and Community Adaptation

The aim of this study was the development of a specifically adapted microbial community for the removal of organic carbon from an industrial wastewater using a bioelectrochemical system. In a first step, ferric iron reducing microorganisms were isolated from the examined industrial wastewater. In a second step, it was tested to what extent these isolates or a cocultivation of the isolates with the exoelectrogenic model organism Geobacter sulfurreducens (G. sulfurreducens) were able to eliminate organic carbon from the wastewater. To establish a stable biofilm on the anode and to analyze the performance of the system, the experiments were conducted first under batch-mode conditions for 21 days. Since the removal of organic carbon was relatively low in the batch system, a similar experiment was conducted under continuous-mode conditions for 65 days, including a slow transition from synthetic medium to industrial wastewater as carbon and electron source and variations in the flow rate of the medium. The overall performance of the system was strongly increased in the continuous- compared to the batch-mode reactor and the highest average current density (1,368 mA/m2) and Coulombic efficiency (54.9%) was measured in the continuous-mode reactor inoculated with the coculture consisting of the new isolates and G. sulfurreducens. The equivalently inoculated batch-mode system produced only 82-fold lower current densities, which were accompanied by 42-fold lower Coulombic efficiencies

Biochemical characterization of recombinant isocitrate dehydrogenase and its putative role in the physiology of an acidophilic micrarchaeon

Despite several discoveries in recent years, the physiology of acidophilic Micrarchaeota, such as “Candidatus Micrarchaeum harzensis A_DKE”, remains largely enigmatic, as they highly express numerous genes encoding hypothetical proteins. Due to a lacking genetic system, it is difficult to elucidate the biological function of the corresponding proteins and heterologous expression is required. In order to prove the viability of this approach, A_DKE’s isocitrate dehydrogenase (MhIDH) was recombinantly produced in Escherichia coli and purified to electrophoretic homogeneity for biochemical characterization. MhIDH showed optimal activity around pH 8 and appeared to be specific for NADP$^{+}$ yet promiscuous regarding divalent cations as cofactors. Kinetic studies showed K$_{M}$-values of 53.03 ± 5.63 µM and 1.94 ± 0.12 mM and k$_{cat}$-values of 38.48 ± 1.62 and 43.99 ± 1.46 s$^{-1}$ resulting in k$_{cat}$/K$_{M}$-values of 725 ± 107.62 and 22.69 ± 2.15 mM$^{-1}$ s$^{-1}$ for DL-isocitrate and NADP$^{+}$, respectively. MhIDH’s exceptionally low affinity for NADP$^{+}$, potentially limiting its reaction rate, can likely be attributed to the presence of a proline residue in the NADP$^{+}$ binding pocket, which might cause a decrease in hydrogen bonding of the cofactor and a distortion of local secondary structure

Influence of the Potential Carbon Sources for Field Denitrification Beds on Their Microbial Diversity and the Fate of Carbon and Nitrate

Nitrogen based eutrophication of ecosystems is a global problem that gains momentum through a growing global population. The water quality of nitrate or ammonium contaminated rivers and streams cannot always be amended in centralized waste water treatment plants. Field denitrification plants were suggested as a solution for a decentralized reduction of nitrate to dinitrogen. Here, stable and cheap organic carbon sources serve as carbon and electron source for a microbial community. Still, our knowledge on the impact of these organic carbon sources on the development and diversity of these cultures is sparse. Moreover, the stability of these denitrification plants at different nitrate loading rates especially in the higher concentration regime were not tested so far. In this study, we compare the fate of carbon and nitrogen as well as the microbial community of wood pellet (WP) (pressed sawdust), wheat straw, and wood chips (WC) based laboratory denitrification reactors. Our study reveals that the diversity and composition of the community is strongly dependent on the carbon source. The diversity decreased in the order WC, wheat straw, and WPs. The three reactor types were characterized by different nitrate reduction kinetics and were affected differently by high nitrate loading rates. While the nitrate reduction kinetics were negatively influenced by higher nitrate doses in the wheat straw reactors, WPs as carbon source sustained the opposite trend and WC lead to an overall slower but concentration independent nitrate reduction rate. Counterintuitively, the concentration of soluble organic carbon was highest in the WP reactors but methane emission was not detectable. This is corroborated by the microbial diversity data in which methanogenic species were highly underrepresented compared to the other two reactor types. In contrary, the methane emissions in the wheat straw and WC reactors were comparable to each other

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

GraftThis: Modernizing Organic Farming

Traditional plant characteristic improvement techniques are accompanied by certain disadvantages. Most notably, it takes several seasons in order to breed for favorable characteristics such as crop yield and disease resistance via deliberate selection. In order to expedite this improvement process, grafting - a method in which two separate plants with individual, desirable qualities are physically combined, may be used. This report explores the development of a regulated environment as a solution to many of the problems associated with the grafting of plants during their recovery stage. The grafting chamber model necessitates: (1) an ability to maintain the specific temperature, humidity and ambient light necessary to produce healthy grafts, (2) ease of use including setup and operation, (3) system portability and an extended lifetime. A review of scientific literature discussing the current state of typical grafting processes yields multiple requirements. The first of these requirements is a regulated environment that maintains a temperature between 72°F and 85 °F and a humidity between 85% and 95%. Also, the environment must be very dark and gradually increase to ambient conditions. The light, temperature, and humidity need to slowly taper to ambient conditions over the ten-day healing period in order to match the external environment. Due to the specificity of these conditions, grafting has historically been a labor-intensive process. Small-scale agricultural operations, those with less than 10,000 USD in annual sales, are impacted most severely by these constraints. The agricultural industry is often space-sensitive so collapsibility and portability are significant concerns. Here, the proof of concept of a grafting chamber capable of satisfying the market need defined by the above, is developed and presented. Stages of the product design process are evaluated individually within the scope of the project

Improving the Cathodic Biofilm Growth Capabilities of Kyrpidia spormannii EA-1 by Undirected Mutagenesis

The biotechnological usage of carbon dioxide has become a relevant aim for future processes. Microbial electrosynthesis is a rather new technique to energize biological CO$_{2}$ fixation with the advantage to establish a continuous process based on a cathodic biofilm that is supplied with renewable electrical energy as electron and energy source. In this study, the recently characterized cathodic biofilm forming microorganism Kyrpidia spormannii strain EA-1 was used in an adaptive laboratory evolution experiment to enhance its cathodic biofilm growth capabilities. At the end of the experiment, the adapted cathodic population exhibited an up to fourfold higher biofilm accumulation rate, as well as faster substratum coverage and a more uniform biofilm morphology compared to the progenitor strain. Genomic variant analysis revealed a genomically heterogeneous population with genetic variations occurring to various extends throughout the community. Via the conducted analysis we identified possible targets for future genetic engineering with the aim to further optimize cathodic growth. Moreover, the results assist in elucidating the underlying processes that enable cathodic biofilm formation