Visualization of Charge Transfer from Bacteria to a Self-Doped Conjugated Polymer Electrode Surface Using Conductive Atomic Force Microscopy.

In this work, we aim to provide a better understanding of the reasons behind electron transfer inefficiencies between electrogenic bacteria and the electrode in microbial fuel cells. We do so using a self-doped conjugated polyelectrolyte (CPE) as the electrode surface, onto which Geobacter sulfurreducens is placed, then using conductive atomic force microscopy (C-AFM) to directly visualize and quantify the electrons that are transferring from each bacterium to the electrode, thereby helping us gain a better understanding for the overpotential losses in MFCs. In doing so, we obtain images that show G. sulfurreducens can directly transfer electrons to an electrode surface without the use of pili, and that overpotential losses are likely due to cell death and poor distribution or performance of individual bacterium's OmcB cytochromes. This unique combination of CPEs with C-AFM can also be used for other studies where electron transfer loss mechanisms need to be understood on the nanoscale, allowing for direct visualization of potential issues in these systems.

Publisher Correction: Achieving high permeability and enhanced selectivity for Angstrom-scale separations using artificial water channel membranes.

The original version of this Article contained an error in the spelling of the author Woochul Song, which was incorrectly given as Woochul C. Song. This has been corrected in both the PDF and HTML versions of the Article.

Achieving high permeability and enhanced selectivity for Angstrom-scale separations using artificial water channel membranes.

Synthetic polymer membranes, critical to diverse energy-efficient separations, are subject to permeability-selectivity trade-offs that decrease their overall efficacy. These trade-offs are due to structural variations (e.g., broad pore size distributions) in both nonporous membranes used for Angstrom-scale separations and porous membranes used for nano to micron-scale separations. Biological membranes utilize well-defined Angstrom-scale pores to provide exceptional transport properties and can be used as inspiration to overcome this trade-off. Here, we present a comprehensive demonstration of such a bioinspired approach based on pillar[5]arene artificial water channels, resulting in artificial water channel-based block copolymer membranes. These membranes have a sharp selectivity profile with a molecular weight cutoff of ~ 500 Da, a size range challenging to achieve with current membranes, while achieving a large improvement in permeability (~65 L m-2 h-1 bar-1 compared with 4-7 L m-2 h-1 bar-1) over similarly rated commercial membranes.

Membrane Protein Insertion into and Compatibility with Biomimetic Membranes.

Membrane protein and membrane protein-mimic functionalized materials are rapidly gaining interest across a wide range of applications, including drug screening, DNA sequencing, drug delivery, sensors, water desalination, and bioelectronics. In these applications, material performance is highly dependent on activity-per-protein and protein packing density in bilayer and bilayer-like structures collectively known as biomimetic membranes. However, a clear understanding of, and accurate tools to study these properties of biomimetic membranes does not exist. This paper presents methods to evaluate membrane protein compatibility with biomimetic membrane materials. The methods utilized provide average single protein activity, and for the first time, provide experimentally quantifiable measures of the chemical and physical compatibility between proteins (and their mimics) and membrane materials. Water transport proteins, rhodopsins, and artificial water channels are reconstituted into the full range of current biomimetic membrane matrices to evaluate the proposed platform. Compatibility measurement results show that both biological and artificial water channels tested largely preserve their single protein water transport rates in biomimetic membranes, while their reconstitution density is variable, leading to different overall membrane permeabilities. It is also shown that membrane protein insertion efficiency inversely correlates with both chemical and physical hydrophobicity mismatch between membrane protein and the membrane matrix.

Influence of molecular structure on the antimicrobial function of phenylenevinylene conjugated oligoelectrolytes.

Conjugated oligoelectrolytes (COEs) with phenylenevinylene (PV) repeat units are known to spontaneously intercalate into cell membranes. Twelve COEs, including seven structures reported here for the first time, were investigated for the relationship between their membrane disrupting properties and structural modifications, including the length of the PV backbone and the presence of either a tetraalkylammonium or a pyridinium ionic pendant group. Optical characteristics and interactions with cell membranes were determined using UV-Vis absorption and photoluminescence spectroscopies, and confocal microscopy. Toxicity tests on representative Gram-positive (Enterococcus faecalis) and Gram-negative (Escherichia coli) bacteria reveal generally greater toxicity to E. faecalis than to E. coli and indicate that shorter molecules have superior antimicrobial activity. Increased antimicrobial potency was observed in three-ring COEs appended with pyridinium ionic groups but not with COEs with four or five PV repeat units. Studies with mutants having cell envelope modifications indicate a possible charge based interaction with pyridinium-appended compounds. Fluorine substitutions on COE backbones result in structures that are less toxic to E. coli, while the addition of benzothiadiazole to COE backbones has no effect on increasing antimicrobial function. A weakly membrane-intercalating COE with only two PV repeat units allowed us to determine the synthetic limitations as a result of competition between solubility in aqueous media and association with cell membranes. We describe, for the first time, the most membrane disrupting structure achievable within two homologous series of COEs and that around a critical three-ring backbone length, structural modifications have the most effect on antimicrobial activity.

Effects of constant or dynamic low anode potentials on microbial community development in bioelectrochemical systems.

In bioelectrochemical systems, exoelectrogenic bacteria respire with anode electrodes as their extracellular electron acceptor; therefore, lower anode potentials can reduce the energy gain to each microbe and select against ones that are not able to respire at a lower potential range. Often fully developed anode communities are compared across bioelectrochemical systems with set anode potentials or fixed external resistances as different operational conditions. However, the comparative effect of the resulting constantly low versus dynamically low anode potentials on the development of anode microbial communities as well as the final cathode microbial communities has not been directly demonstrated. In this study, we used a low fixed anode potential of -250 mV and a higher-current control potential of -119 mV vs. Standard Hydrogen Electrode to approximately correspond with the negative peak anode potential values obtained from microbial fuel cells operated with fixed external resistances of 1 kΩ and 47 Ω, respectively. Pyrosequencing data from a 2-month time series show that a lower set anode potential resulted in a more diverse community than the higher- and variable-potential systems, likely due to the hindered enrichment of a Geobacter-dominated community with limited energy gain at this set potential. In this case, it appears that the selective pressure caused by the low set potential was counteracted by the low energy gain over a 2-month time scale. The air cathode microbial community with constant low anode potentials showed delayed enrichment of denitrifiers or perchlorate-reducing bacteria compared to the fixed external resistance condition.

Membrane-intercalating conjugated oligoelectrolytes: impact on bioelectrochemical systems.

Conjugated oligoelectrolytes (COEs), molecules that are defined by a π-delocalized backbone and terminal ionic pendant groups, have been previously demonstrated to effectively reduce charge-injection/extraction barriers at metal/organic interfaces in thin-film organic-electronic devices. Recent studies demonstrate a spontaneous affinity of certain COEs to intercalate into, and align within, lipid bilayers in an ordered orientation, thereby allowing modification of membrane properties and the functions of microbes in bioelectrochemical and photosynthetic systems. Several reports have provided evidence of enhanced current generation and bioproduction. Mechanistic approaches suggest that COEs influence microbial extracellular electron transport to abiotic electrode surfaces via more than one proposed pathway, including direct electron transfer and meditated electron transfer. Molecular dynamics simulations as a function of molecular structure suggest that insertion of cationic COEs results in membrane thinning as the lipid phosphate head groups are drawn toward the center of the bilayer. Since variations in molecular structures, especially the length of the conjugated backbone, distribution of ionic groups, and hydrophobic substitutions, show an effect on their antimicrobial properties, preferential cell localization, and microbial selection, it is promising to further design novel membrane-intercalating molecules based on COEs for practical applications, including energy generation, environmental remediation, and antimicrobial treatment.

Two-dimensional protein crystals for solar energy conversion.

Two-dimensional photosynthetic protein crystals provide a high density of aligned reaction centers. We reconstitute the robust light harvesting protein Photosystem I into a 2D crystal with lipids and integrate the crystals into a photo-electrochemical device. A 4-fold photocurrent enhancement is measured by incorporating conjugated oligoelectrolytes to form a supporting conductive bilayer in the device which produces a high photocurrent of ∼600 µA per mg PSI deposited.

Enhanced nitrogen removal in single-chamber microbial fuel cells with increased gas diffusion areas.

Single-chamber microbial fuel cells (MFCs) with nitrifiers pre-enriched at the air cathodes have previously been demonstrated as a passive strategy for integrating nitrogen removal into current-generating bioelectrochemical systems. To further define system design parameters for this strategy, we investigated in this study the effects of oxygen diffusion area and COD/N ratio in continuous-flow reactors. Doubling the gas diffusion area by adding an additional air cathode or a diffusion cloth significantly increased the ammonia and COD removal rates (by up to 115% and 39%), ammonia removal efficiency (by up to 134%), the cell voltage and cathode potentials, and the power densities (by a factor of approximately 2). When the COD/N ratio was lowered from 13 to 3, we found up to 244% higher ammonia removal rate but at least 19% lower ammonia removal efficiency. An increase of COD removal rate by up to 27% was also found when the COD/N ratio was lowered from 11 to 3. The Coulombic efficiency was not affected by the additional air cathode, but decreased by an average of 11% with the addition of a diffusion cloth. Ammonia removal by assimilation was also estimated to understand the ammonia removal mechanism in these systems. These results showed that the doubling of gas diffusion area enhanced N and COD removal rates without compromising electrochemical performance.

Nitrogen removal in a single-chamber microbial fuel cell with nitrifying biofilm enriched at the air cathode.

Nitrogen removal is needed in microbial fuel cells (MFCs) for the treatment of most waste streams. Current designs couple biological denitrification with side-stream or combined nitrification sustained by upstream or direct aeration, which negates some of the energy-saving benefits of MFC technology. To achieve simultaneous nitrification and denitrification, without extra energy input for aeration, the air cathode of a single-chamber MFC was pre-enriched with a nitrifying biofilm. Diethylamine-functionalized polymer (DEA) was used as the Pt catalyst binder on the cathode to improve the differential nitrifying biofilm establishment. With pre-enriched nitrifying biofilm, MFCs with the DEA binder had an ammonia removal efficiency of up to 96.8% and a maximum power density of 900 ± 25 mW/m(2), compared to 90.7% and 945 ± 42 mW/m(2) with a Nafion binder. A control with Nafion that lacked nitrifier pre-enrichment removed less ammonia and had lower power production (54.5% initially, 750 mW/m(2)). The nitrifying biofilm MFCs had lower Coulombic efficiencies (up to 27%) than the control reactor (up to 36%). The maximum total nitrogen removal efficiency reached 93.9% for MFCs with the DEA binder. The DEA binder accelerated nitrifier biofilm enrichment on the cathode, and enhanced system stability. These results demonstrated that with proper cathode pre-enrichment it is possible to simultaneously remove organics and ammonia in a single-chamber MFC without supplemental aeration.

Characterization of microbial fuel cells at microbially and electrochemically meaningful time scales.

The variable biocatalyst density in a microbial fuel cell (MFC) anode biofilm is a unique feature of MFCs relative to other electrochemical systems, yet performance characterizations of MFCs typically involve analyses at electrochemically relevant time scales that are insufficient to account for these variable biocatalyst effects. This study investigated the electrochemical performance and the development of anode biofilm architecture under different external loadings, with duplicate acetate-fed single-chamber MFCs stabilized at each resistance for microbially relevant time scales. Power density curves from these steady-state reactors generally showed comparable profiles despite the fact that anode biofilm architectures and communities varied considerably, showing that steady-state biofilm differences had little influence on electrochemical performance until the steady-state external loading was much larger than the reactor internal resistance. Filamentous bacteria were dominant on the anodes under high external resistances (1000 and 5000 Ω), while more diverse rod-shaped cells formed dense biofilms under lower resistances (10, 50, and 265 Ω). Anode charge transfer resistance decreased with decreasing fixed external resistances, but was consistently 2 orders of magnitude higher than the resistance at the cathode. Cell counting showed an inverse exponential correlation between cell numbers and external resistances. This direct link of MFC anode biofilm evolution with external resistance and electricity production offers several operational strategies for system optimization.

Time-course correlation of biofilm properties and electrochemical performance in single-chamber microbial fuel cells.

The relationship between anode microbial characteristics and electrochemical parameters in microbial fuel cells (MFCs) was analyzed by time-course sampling of parallel single-bottle MFCs operated under identical conditions. While voltage stabilized within 4days, anode biofilms continued growing during the six-week operation. Viable cell density increased asymptotically, but membrane-compromised cells accumulated steadily from only 9% of total cells on day 3 to 52% at 6weeks. Electrochemical performance followed the viable cell trend, with a positive correlation for power density and an inverse correlation for anode charge transfer resistance. The biofilm architecture shifted from rod-shaped, dispersed cells to more filamentous structures, with the continuous detection of Geobacter sulfurreducens-like 16S rRNA fragments throughout operation and the emergence of a community member related to a known phenazine-producing Pseudomonas species. A drop in cathode open circuit potential between weeks two and three suggested that uncontrolled biofilm growth on the cathode deleteriously affects system performance.

Bacterial community dynamics in a functionally stable pilot-scale wastewater treatment plant.

To determine whether functional stability was correlated with a stable microbial community structure in a functionally stable pilot-scale wastewater treatment plant, bacterial communities in the system were monitored over a one-year period. Bacterial community dynamics was characterized by the terminal restriction fragment length polymorphism (T-RFLP) of 16S rRNA genes. During the study period, the effluent BOD concentrations were very stable, with the average BOD concentration below 10 mg/L. The effluent TN concentrations were always below 20 mg/L, except for the first 40 days. T-RFLP results showed that, during the test period, the bacterial community structures were not stable, with an average change rate (every 15 days) of 20.4%±11.2%. Based on Lorenz distribution curves, it was observed that 20% of the species corresponded with 40-77% of cumulative relative abundances. Results clearly revealed that, in the pilot-scale wastewater treatment plant, functional stability did not correlate with stable bacterial communities.