群感效应对电活性微生物胞外电子传递的影响

群感效应（quorum sensing,QS）是微生物之间以信号分子受体蛋白感知信号浓度变化,从而调控菌群的行为及功能,使其适应环境变化的信号通讯机制。电活性微生物（electroactive microorganisms,EAMs）能进行胞外电子传递,在可再生能源利用和环境修复方面具有广阔的应用前景。近年来,关于QS在EAMs胞外电子传递中的作用的研究日益增多。本文总结了QS对纯EAMs或混合产电菌群的直接或间接电子传递的影响效应及机制,阐述了基于QS的EAMs逻辑与门的构建及其应用前景,并从机制研究的角度展望其未来发展方向。     

Effect of oxygen on the per‐cell extracellular electron transfer rate of Shewanella oneidensis MR‐1 explored in bioelectrochemical systems

Extracellular electron transfer (EET) is a mechanism that enables microbes to respire solid‐phase electron acceptors. These EET reactions most often occur in the absence of oxygen, since oxygen can act as a competitive electron acceptor for many facultative microbes. However, for Shewanella oneidensis MR‐1, oxygen may increase biomass development, which could result in an overall increase in EET activity. Here, we studied the effect of oxygen on S. oneidensis MR‐1 EET rates using bioelectrochemical systems (BESs). We utilized optically accessible BESs to monitor real‐time biomass growth, and studied the per‐cell EET rate as a function of oxygen and riboflavin concentrations in BESs of different design and operational conditions. Our results show that oxygen exposure promotes biomass development on the electrode, but significantly impairs per‐cell EET rates even though current production does not always decrease with oxygen exposure. Additionally, our results indicated that oxygen can affect the role of riboflavin in EET. Under anaerobic conditions, both current density and per‐cell EET rate increase with the riboflavin concentration. However, as the dissolved oxygen (DO) value increased to 0.42 mg/L, riboflavin showed very limited enhancement on per‐cell EET rate and current generation. Since it is known that oxygen can promote flavins secretion in S. oneidensis, the role of riboflavin may change under anaerobic and aerobic conditions. Biotechnol. Bioeng. 2017;114: 96–105. © 2016 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.     

黄素介导的胞外电子转移研究与工程改造*

产电微生物的胞外电子转移在能源、环境等诸多领域有着非常重要的应用价值。希瓦氏菌(Shewanella oneidensis)作为模式产电微生物,其电催化系统引起了广泛的研究。黄素作为S. oneidensis重要的电子载体,其介导的胞外电子转移是电子传递过程中的一个限速步骤。然而自然环境中野生型S. oneidensis的黄素分泌量极低,对其工程改造也存在一定的局限性,因而严重阻碍了胞外电子的传递过程,这已成为限制其电子转移的主要瓶颈。基于S. oneidensis黄素介导的电子转移机制,系统地从黄素的合成路径及转录调控的角度阐明了黄素合成的调控因素,并综述近年来利用代谢工程、合成生物学以及电极材料修饰等方法来提高黄素介导电子转移的工程化策略,未来可利用一些系统的研究方法和表达工具来加速产电微生物黄素介导的胞外电子转移。     

微生物种间直接电子传递机理及应用研究进展

微生物胞内产生的电子转移到其他电子受体而获得能量的过程称为微生物胞外电子传递,其中,另一微生物作为电子受体时发生的电子传递称为微生物种间电子传递。根据微生物种间电子传递机制,可分间接种间电子传递和种间直接电子传递。由于种间直接电子传递不需要其他物质介导,因此较间接种间电子传递效率更高、能量利用更高。本文系统阐述了微生物进行胞外电子传递的机理及应用,重点分析了种间直接电子传递机理,并概述种间直接电子传递应用领域,为寻找更多电连接的微生物群落以及应用微生物提供参考。     

3D Macroporous Nitrogen‐Enriched Graphitic Carbon Scaffold for Efficient Bioelectricity Generation in Microbial Fuel Cells

Microbial fuel cell (MFC) can generate electricity based on oxidation of organic compounds by exoelectogens, giving rise to a promising potential for recovering electrical energy from organic wastewater. The structure and property of anode materials have inherent impact to extracellular electron transfer (EET), an interfacial process that greatly limits bioelectricity production of MFC. Herein, a three dimensional (3D) macroporous nitrogen‐enriched graphitic carbon (NGC) scaffold is fabricated from commercially available melamine foam using facile pyrolysis method. The NGC electrode is demonstrated to promote EET ef?ciently, achieving a power density of 750 mW m^?2 based on pure cultured Shewanella oneidensis MR‐1 in acetate‐feeding MFC. The unique 3D open‐cell structure not only offers habitats for colonization of electroactive bio?lm up to a maximal density but also provides macroporous architecture for internal mass transfer without concern of bio‐blocking and bio‐fouling. Additionally, nitrogen incorporation also plays a signi?cant role in enhancing EET, where pyrrolic nitrogen is much more active than graphitic and pyridinic nitrogen as indicated by density functional theory calculation. This work provides a proof‐of‐concept demonstration of a high‐ef?ciency, cost‐effective, easily scaling‐up, and environmentally friendly anode material of bioelectrochemical systems for electricity generation, hydrogen production, and pollutant degradation.     

Rewiring the respiratory pathway of Lactococcus lactis to enhance extracellular electron transfer

Lactococcus lactis, a lactic acid bacterium with a typical fermentative metabolism, can also use oxygen as an extracellular electron acceptor. Here we demonstrate, for the first time, that L. lactis blocked in NAD⁺ regeneration can use the alternative electron acceptor ferricyanide to support growth. By electrochemical analysis and characterization of strains carrying mutations in the respiratory chain, we pinpoint the essential role of the NADH dehydrogenase and 2-amino-3-carboxy-1,4-naphtoquinone in extracellular electron transfer (EET) and uncover the underlying pathway systematically. Ferricyanide respiration has unexpected effects on L. lactis, e.g., we find that morphology is altered from the normal coccoid to a more rod shaped appearance, and that acid resistance is increased. Using adaptive laboratory evolution (ALE), we successfully enhance the capacity for EET. Whole-genome sequencing reveals the underlying reason for the observed enhanced EET capacity to be a late-stage blocking of menaquinone biosynthesis. The perspectives of the study are numerous, especially within food fermentation and microbiome engineering, where EET can help relieve oxidative stress, promote growth of oxygen sensitive microorganisms and play critical roles in shaping microbial communities.     

Enzymatic biofuel cell based on anode and cathode powered by ethanol

Enzymatic biofuel cell based on enzyme modified anode and cathode electrodes are both powered by ethanol and operate at ambient temperature is described. The anode of the presented biofuel cell was based on immobilized quino-hemoprotein-alcohol dehydrogenase (QH-ADH), while the cathode on co-immobilized alcohol oxidase (AOx) and microperoxidase (MP-8). Two enzymes AOx and MP-8 acted in the consecutive mode and were applied in the design of the biofuel cell cathode. The ability of QH-ADH to transfer electrons directly towards the carbon-based electrode and the ability of MP-8 to accept electrons directly from the same type of electrodes was exploited in this biofuel cell design. Direct electron transfer (DET) to/from enzymes was the basis for generating an electric potential between the anode and cathode. Application of immobilized enzymes and the harvesting of the same type of fuel at both electrodes (cathode and anode) avoided the compartmentization of enzymatic biofuel cell. The maximal open circuit potential of the biofuel cell was 240mV.     

Improved fuel cell and electrode designs for producing electricity from microbial degradation

A new one-compartment fuel cell was composed of a rubber bunged bottle with a center-inserted anode and a window-mounted cathode containing an internal, proton-permeable porcelain layer. This fuel cell design was less expensive and more practical than the conventional two-compartment system, which requires aeration and a ferricyanide solution in the cathode compartment. Three new electrodes containing bound electron mediators including a Mn(4+)-graphite anode, a neutral red (NR) covalently linked woven graphite anode, and an Fe(3+)-graphite cathode were developed that greatly enhanced electrical energy production (i.e., microbial electron transfer) over conventional graphite electrodes. The potentials of these electrodes measured by cyclic voltametry at pH 7.0 were (in volts): +0.493 (Fe(3+)-graphite); +0.15 (Mn(4+)-graphite); and -0.53 (NR-woven graphite). The maximal electrical productivities obtained with sewage sludge as the biocatalyst and using a Mn(4+)-graphite anode and a Fe(3+)-graphite cathode were 14 mA current, 0.45 V potential, 1,750 mA/m(2) current density, and 788 mW/m(2) of power density. With Escherichia coli as the biocatalyst and using a Mn(4+)-graphite anode and a Fe(3+)-graphite cathode, the maximal electrical productivities obtained were 2.6 mA current, 0.28 V potential, 325 mA/m(2) current density, and 91 mW/m(2) of power density. These results show that the amount of electrical energy produced by microbial fuel cells can be increased 1,000-fold by incorporating electron mediators into graphite electrodes. These results also imply that sewage sludge may contain unique electrophilic microbes that transfer electrons more readily than E. coli and that microbial fuel cells using the new Mn(4+)-graphite anode and Fe(3+)-graphite cathode may have commercial utility for producing low amounts of electrical power needed in remote locations.     

Molecular underpinnings for microbial extracellular electron transfer during biogeochemical cycling of earth elements

Microbial extracellular electron transfer(EET) is electron exchanges between the quinol/quinone pools in microbial cytoplasmic membrane and extracellular substrates. Microorganisms with EET capabilities are widespread in Earth hydrosphere, such as sediments of rivers, lakes and oceans, where they play crucial roles in biogeochemical cycling of key elements, including carbon,nitrogen, sulfur, iron and manganese. Over the past 12 years, significant progress has been made in mechanistic understanding of microbial EET at the molecular level. In this review, we focus on the molecular mechanisms underlying the microbial ability for extracellular redox transformation of iron, direct interspecies electron transfer as well as long distance electron transfer mediated by the cable bacteria in the hydrosphere.     

促进电活性微生物产电呼吸能力的方法研究进展

产电呼吸是指电活性微生物(electroactive microorganisms,EAMs)以胞外固体电极作为电子受体的一种呼吸形式,在可再生能源利用和环境修复方面具有广阔的应用前景。能否进一步提高EAMs的产电呼吸能力是相关技术能否从实验室走向实际应用的核心,而提高产电呼吸能力的关键是加强EAMs与胞外固体电极间的电子传递能力。目前总结如何促进EAMs产电呼吸能力的综述文献极少。因此,本文从投加化学试剂、施加物理作用及改造生物基因3个方面总结了现有的促进EAMs产电呼吸能力的方法,介绍了每种方法的优势与缺陷,重点阐述了每种手段的作用机理及促进效果,并从实际应用和机理研究的角度展望了今后的研究方向。     

Initial development and structure of biofilms on microbial fuel cell anodes

Background  

Microbial fuel cells (MFCs) rely on electrochemically active bacteria to capture the chemical energy contained in organics and convert it to electrical energy. Bacteria develop biofilms on the MFC electrodes, allowing considerable conversion capacity and opportunities for extracellular electron transfer (EET). The present knowledge on EET is centred around two Gram-negative models, i.e. Shewanella and Geobacter species, as it is believed that Gram-positives cannot perform EET by themselves as the Gram-negatives can. To understand how bacteria form biofilms within MFCs and how their development, structure and viability affects electron transfer, we performed pure and co-culture experiments.     

Microbial electrocatalysis: Redox mediators responsible for extracellular electron transfer

Redox mediator plays an important role in extracellular electron transfer (EET) in many environments wherein microbial electrocatalysis occurs actively. Because of the block of cell envelope and the low difference of redox potential between the intracellular and extracellular surroundings, the proceeding of EET depends mainly on the help of a variety of mediators that function as an electron carrier or bridge. In this Review, we will summarize a wide range of redox mediators and further discuss their functional mechanisms in EET that drives a series of microbial electrocatalytic reactions. Studying these mediators adds to our knowledge of how charge transport and electrochemical reactions occur at the microorganism-electrode interface. This understanding would promote the widespread applications of microbial electrocatalysis in microbial fuel cells, bioremediation, bioelectrosynthesis, biomining, nanomaterial productions, etc. These improved applications will greatly benefit the sustainable development of the environmental-friendly biochemical industries.     

Increased power production from a sediment microbial fuel cell with a rotating cathode

The application of a rotating cathode in a river sediment microbial fuel cell increased the oxygen availability to the cathode, and therefore improved the cathode reaction rate, resulting in a higher power production (49 mW/m²) compared to a nonrotating cathode system (29 mW/m²). The increased dissolved oxygen in the water of our lab-scale sediment MFC, however, resulted in a less negative anode potential and a higher anodic charge transfer resistance, which constrained the maximum power density. Thus, an optimum balance between the superior cathode reaction rates and the inferior anode reaction rates due to higher dissolved oxygen levels must be ascertained.     

The Remediation of Chromium (VI)-Contaminated Soils Using Microbial Fuel Cells

Chromium (VI) is a priority pollutant in soil and water and poses serious threats to the environment. Microbial fuel cells (MFCs), as a sustainable technology, have been applied to treat heavy-metal-contaminated wastewater. To study MFC application in soil remediation, red clay soil and fluvo-aquic soil were spiked with Cr(VI) and packed into a cathode chamber of MFCs, which were then operated at external resistances of 100 and 1000 Ω for 16 days, with open circuit condition as a control treatment. After the operation, the concentration of dissolved Cr(VI) in supernatant and total Cr(VI) in soil was decreased. Soil type and external resistance significantly affected the current, removal efficiency of Cr(VI), and cathode efficiency. Reducing external resistance improved the removal efficiency. The red soil generated a higher current of MFCs, but showed a lower removal efficiency and cathode efficiency than fluvo-aquic soil, implying that the red soil may contain more electron acceptors that competed with Cr(VI) reduction reaction. Our study demonstrated that MFC-based technology has the potential to remediate Cr(VI)-contaminated soil; the efficiency varied between soil types and can be improved with high current.     

A kinetic perspective on extracellular electron transfer by anode-respiring bacteria

In microbial fuel cells and electrolysis cells (MXCs), anode-respiring bacteria (ARB) oxidize organic substrates to produce electrical current. In order to develop an electrical current, ARB must transfer electrons to a solid anode through extracellular electron transfer (EET). ARB use various EET mechanisms to transfer electrons to the anode, including direct contact through outer-membrane proteins, diffusion of soluble electron shuttles, and electron transport through solid components of the extracellular biofilm matrix. In this review, we perform a novel kinetic analysis of each EET mechanism by analyzing the results available in the literature. Our goal is to evaluate how well each EET mechanism can produce a high current density (>10 A m⁻²) without a large anode potential loss (less than a few hundred millivolts), which are feasibility goals of MXCs. Direct contact of ARB to the anode cannot achieve high current densities due to the limited number of cells that can come in direct contact with the anode. Slow diffusive flux of electron shuttles at commonly observed concentrations limits current generation and results in high potential losses, as has been observed experimentally. Only electron transport through a solid conductive matrix can explain observations of high current densities and low anode potential losses. Thus, a study of the biological components that create a solid conductive matrix is of critical importance for understanding the function of ARB.     

Molecular design of laccase cathode for direct electron transfer in a biofuel cell

In order to improve the direct electron transfer in enzymatic biofuel cells, a rational design of a laccase electrode is presented. Graphite electrodes were functionalized with 4-[2-aminoethyl] benzoic acid hydrochloride (AEBA). The benzoic acid moiety of AEBA interacts with the laccase T1 site as ligand with an association constant (K(A)) of 6.6×10(-6) M. The rational of this work was to orientate the covalent coupling of laccase molecule with the electrode surface through the T1 site and thus induce the direct electron transfer between the T1 site and the graphite electrode surface. Direct electron transfer of laccase was successfully achieved, and the semi-enzymatic fuel cell Zn-AEBA laccase showed a current density of 2977 μA cm(-2) and a power density of 1190 μW cm(-2) at 0.41 V. The molecular oriented laccase cathode showed 37% higher power density and 43% higher current density than randomly bound laccase cathode. Chronoaperometric measurements of the Zn-AEBA fuel cell showed functionality on 6 h. Thus, the orientation of the enzyme molecules improves the electron transfer and optimizes enzyme-based fuel cells efficiency.     

Microbial extracellular electron transfer and its relevance to iron corrosion

Extracellular electron transfer (EET) is a microbial metabolism that enables efficient electron transfer between microbial cells and extracellular solid materials. Microorganisms harbouring EET abilities have received considerable attention for their various biotechnological applications, including bioleaching and bioelectrochemical systems. On the other hand, recent research revealed that microbial EET potentially induces corrosion of iron structures. It has been well known that corrosion of iron occurring under anoxic conditions is mostly caused by microbial activities, which is termed as microbiologically influenced corrosion (MIC). Among diverse MIC mechanisms, microbial EET activity that enhances corrosion via direct uptake of electrons from metallic iron, specifically termed as electrical MIC (EMIC), has been regarded as one of the major causative factors. The EMIC‐inducing microorganisms initially identified were certain sulfate‐reducing bacteria and methanogenic archaea isolated from marine environments. Subsequently, abilities to induce EMIC were also demonstrated in diverse anaerobic microorganisms in freshwater environments and oil fields, including acetogenic bacteria and nitrate‐reducing bacteria. Abilities of EET and EMIC are now regarded as microbial traits more widespread among diverse microbial clades than was thought previously. In this review, basic understandings of microbial EET and recent progresses in the EMIC research are introduced.     

生物工程强化微生物电合成转化CO_2的研究进展

微生物电合成(Microbial electrosynthesis,MES)可直接利用电能驱动微生物还原固定CO_2合成多碳化合物,为可再生新能源转化、精细化学品制备和生态环境保护提供新机遇。但是,微生物吸收胞外电极电子速率慢、产物合成效率低和产品品位不高,限制了MES实现工业化应用。在概述阴极电活性微生物吸收胞外电子的分子机制的基础上,重点综述近5年应用生物工程的理论和技术强化MES用于CO_2转化的策略与研究进展,包括改造和调控胞外电子传递通路和胞内代谢途径以及定向构建有限微生物混合培养菌群三方面,阐明了生物工程可有效突破MES中电子传递慢和可用代谢途径相对单一等瓶颈。针对目前生物工程在改进MES所面临的主要问题,从胞外电子传递机理研究、基因工具箱开发、组学技术与现代分析技术联用等角度展望了今后的研究方向。     

奥奈达希瓦氏菌CctA介导周质甲基橙还原的电子传递机理

电活性微生物奥奈达希瓦氏菌的胞外电子传递（extracellular electron transfer,EET）在污染物降解、环境修复、生物电化学传感、能源利用等方面具有广泛的应用潜力;四血红素细胞色素CctA （small tetraheme cytochrome）是希瓦氏菌周质空间中最丰富的蛋白质之一,能够参与多种氧化还原过程,但目前对CctA在EET中的行为和机理认识仍然有限。【目的】研究阐明CctA蛋白在希瓦氏菌模式菌株MR-1周质空间以偶氮染料作为电子受体的EET中的作用,补充和拓展希瓦氏菌的厌氧呼吸产能机制。【方法】以周质还原型偶氮染料甲基橙（methyl orange,MO）作为电子受体,在mteal reduction （Mtr）蛋白缺失菌株Δmtr中研究MO的周质还原特点,并通过基因敲除和回补表达研究CctA蛋白在周质电子传递中的作用。【结果】在缺失Mtr通道的情况下,细胞色素CctA可以介导周质空间的电子传递而还原MO。重组表达CctA在低水平时,MO在周质空间中的还原速率与其表达水平呈正相关,更高水平的CctA表达无助于进一步提高MO的还原速率。蛋白膜伏安结果展示了CctA与周质空间内其他高电位氧化还原蛋白的显著区别,可能参与构成一条低电位的MO还原通道。【结论】从分子动力学层面揭示了CctA在周质MO还原中的独特电子传递行为,为进一步推进对细菌周质电子传递机制的理解,以及通过合成生物学设计或改造胞外氧化还原系统、强化生物电化学在污染物降解中的应用提供了重要信息。     

Photogeneration of singlet oxygen (1O2) and free radicals (Sen·-, O·-2) by tetra-brominated hypocrellin B derivative

To improve photodynamic activity of the parent hypocrellin B (HB), a tetra-brominated HB derivative (compound 1) was synthesized in high yield. Compared with HB, compound 1 has enhanced red absorption and high molar extinction coefficients. The photodynamic action of compound 1, especially the generation mechanism and efficiencies of active species (Sen^·-, O^·-₂ and ¹O₂) were studied using electron paramagnetic resonance (EPR) and spectrophotometric methods. In the deoxygenated DMSO solution of compound 1, the semiquinone anion radical of compound 1 is photogenerated via the self-electron transfer between the excited and ground state species. The presence of electron donor significantly promotes the reduction of compound 1. When oxygen is present, superoxide anion radical (O^·-₂) is formed via the electron transfer from Sens^·- to the ground state molecular oxygen. The efficiencies of Sens^·- and O^·-₂ generation by compound 1 are about three and two times as much as that of HB, respectively. Singlet oxygen (¹O₂) can be produced via the energy transfer from triplet compound 1 to ground state oxygen molecules. The quantum yield of singlet oxygen (¹O₂) is 0.54 in CHCl₃ similar to that of HB. Furthermore, it was found that the accumulation of Sens^·- would replace that of O^·-₂ or ¹O₂ with the depletion of oxygen in the sealed system.