微生物厌氧呼吸与有机污染水体沉积物修复

水体沉积物有机污染是当前全球关注的重要环境问题。微生物具有呼吸和代谢多样性,能以多种污染物作为厌氧呼吸的电子供体或受体,与周围环境中的生物和非生物因素组成代谢网络耦合有机污染物降解转化,是有机污染水体沉积物修复的重要驱动者。本文重点综述了微生物厌氧呼吸、电子传递网络及其对有机污染水体沉积物的修复机制研究进展,并对有机污染水体沉积物微生物修复理论和技术研究的问题和挑战进行了探讨。     

腐殖质呼吸作用及其生态学意义

腐殖质呼吸是厌氧环境中普遍存在的一种微生物呼吸代谢模式.自1996年发现以来,日益成为生态学与环境科学领域的研究热点.在厌氧条件下,一些微生物能以腐殖质作为唯一电子受体,氧化环境中的有机质或者甲苯等环境有毒物质,产生CO2,参与碳循环;同时,腐殖质呼吸作用产生的还原态腐殖质可以还原环境中的一些氧化态物质,如Fe(III)、Mn(IV)、Cr(VI)、U(VI) 、硝基芳香化合物和多卤代污染物.因此,腐殖质呼吸能够影响环境中C、N、Fe、Mn以及一些痕量金属元素的生物地球化学循环,并且能够促进重金属以及有机污染物的脱毒,在水体自净、污染土壤原位修复、污水处理等方面具有积极作用.     

Inorganic types of fermentation and anaerobic respirations in the evolution of energy-yielding metabolism

We proposed long ago the following sequence as one of the main pathways in the evolution of energy-yielding metabolism: fermentation→nitrate fermentation→nitrate respiration→oxygen respiration. In the present report our concept is presented in a more general form: (1) fermentation→ →(2) fermentation with H₂ release→(3) inorganic types of fermentation→(4) anaerobic respirations →(5) oxygen respiration, based upon recent biological and physical information. The energy-yielding efficiency increased gradually together with the evolution. (2) is characterized by the participation of ferredoxin, (3) by the establishment of electron transfer chain, and (4) by the participation of cytochrome and oxidative phosphorylation. The close relationship between the primary structure of ferredoxins of anaerobic bacteria and that of a cytochrome (cytochromec ₃) was demonstrated. It reveals that the transition from inorganic types of fermentation to anaerobic respirations was direct and accompanied by the transition from ferredoxins to cytochromes, and it further supports our concept that the cytochrome system, and consequently the oxidative phosphorylation, were induced at this evolutionary step. Our concept based upon biological observations is consistent with a physical theory recently proposed by M. Shimizu.     

腐殖质在环境污染物生物降解中的作用研究进展

腐殖质物质在地球的生态环境中大量存在,它不仅可以在有毒化合物的生物降解和生物转化过程中起到氧化还原中间体的作用,加速有毒物质的降解和转化。也可以作为唯一末端电子受体,接受来自一些有机酸或者甲苯等环境中有毒物质提供的电子,偶联能量的产生,支持菌体的生长,形成一种新的细菌厌氧呼吸形式——腐殖质呼吸。因此,对腐殖质在环境有毒物质的生物降解和生物转化过程中的作用进行研究,不仪对于深入理解细菌呼吸的本质具有重要的理论意义,而且对于环境有毒物质的降解和转化以及元素的生物地球化学循环具有重要的生态学意义,同时对地球表面的有毒物质进行更有效的生物降解具有重要的现实意义。     

细菌产黄素类化合物介导的电子传递及对环境污染物的厌氧生物转化研究进展

氧化还原介体能够加速有毒环境污染物的厌氧生物转化。黄素类化合物是一类微生物自身合成分泌的氧化还原介体,其应用可有效地避免外源性介体带来的成本较高及造成二次污染的问题,因此引起了广泛的关注。研究表明,细菌合成的微量黄素类化合物不仅能够作为黄素蛋白的辅酶因子参与偶氮染料、铬酸盐和硝基芳烃等污染物的厌氧生物转化,并且还可以分泌到胞外将电子传递给固态电子受体如含铁矿物和电极等来参与生物修复过程。根据黄素类化合物的功能,本文综述了黄素类化合物的合成与分泌,及其介导的胞内外电子传递和对环境污染物厌氧生物转化的影响,以促进其在实际环境污染物处理中的应用。     

硝酸盐还原促进毒害性有机污染物降解的研究进展

大量具有高毒性、持久性和生物蓄积性的有机污染物被排放到环境中,对生态环境和人类健康造成了严重威胁。近年来,利用硝酸盐作电子受体在厌氧条件下降解毒害性有机污染物,已取得一定的进展。本文综述了硝酸盐还原体系中几种典型毒害性有机污染物(多环芳烃、单环或杂环芳烃类有机物及卤代有机物)的厌氧降解研究进展。在此基础上,提出了硝酸盐还原促进毒害性有机污染物降解研究中存在的主要问题及其在加速污染环境净化方面的应用前景。     

厌氧条件下希瓦氏菌腐殖质还原对偶氮还原的影响

以希瓦氏菌属的3个代表种为研究对象,研究了在厌氧条件下腐殖质的存在对偶氮还原的影响。实验结果表明:3个代表菌株在厌氧条件下都有高效的偶氮还原和腐殖质还原功能,1mmol/L偶氮染料在24h内完全脱色,并且偶氮还原与电子供体氧化存在着紧密的偶联关系。腐殖质物质模式物2-磺酸蒽醌AQS在小于1~2mmol/L条件下能显著加速偶氮还原,12h就完全脱色,3mmol/L时18h完全脱色。但当浓度大于3mmol/L时则对偶氮还原产生明显抑制作用。另一腐殖质模式物2,6-双磺酸蒽醌AQDS其浓度在1~3mmol/L以内亦使脱色在12h内完成,4~6mmol/L时15h左右完成脱色。7~12mmol/L仍有一定的脱色促进作用,但随着浓度的提高,其促进作用也逐渐减弱。这说明腐殖质的确可以作为氧化还原中间体穿梭于电子供体与染料的偶氮双键之间促进偶氮还原。但当其浓度达到某一阈值时它就显出与偶氮键竞争电子的本质,从而使偶氮还原速率下降。原因在于他们的氧化还原电势的差异,导致细菌呼吸链的电子递体对腐殖质物质和偶氮键的亲和力不同,从而使不同腐殖质浓度对偶氮键还原产生了不同的影响。     

一株促甲烷氧化假单胞菌Pseudomonas putida P7的分离及电活性特征

微生物胞外呼吸是厌氧环境中控制性能量代谢方式,直接驱动着C、N、S、Fe等关键元素的生物地球化学循环。微生物纳米导线（Microbial nanowires）的发现,被认为是微生物胞外呼吸的里程碑事件,推动了电微生物学（Electromicrobiology）的形成与发展。微生物纳米导线是一类由微生物合成的,具有导电性的纤维状表面附属结构。通过细菌纳米导线,微生物胞内代谢产生的电子可以长距离输送到胞外受体或其他微生物,改变了电子传递链仅仅局限于细胞胞内的认识,从而大大拓展了微生物-胞外环境互作的范围。微生物纳米导线的良好导电性,赋予了其作为天然纳米材料的广阔应用前景。目前,微生物纳米导线的导电机制、生态功能及其在生物材料、生物能源、生物修复及人体健康多领域的应用,已经成为新兴电微生物学的前沿与热点。然而,微生物纳米导线的生物学、生态学功能尚不清楚,它的电子传递机制仍存在分歧。本文在系统性总结微生物纳米导线性质、功能的基础上,以Geobacter sulfurreducens和Shewanella oneidensis纳米导线为模型,详细阐述了纳米导线的组成与结构、表征与测量方法、导电理论（类金属导电学说与电子跃迁学说）及其潜在的应用,最后提出了未来微生物纳米导线研究的重点方向、挑战与机遇。     

Exocellular electron transfer in anaerobic microbial communities

Exocellular electron transfer plays an important role in anaerobic microbial communities that degrade organic matter. Interspecies hydrogen transfer between microorganisms is the driving force for complete biodegradation in methanogenic environments. Many organic compounds are degraded by obligatory syntrophic consortia of proton-reducing acetogenic bacteria and hydrogen-consuming methanogenic archaea. Anaerobic microorganisms that use insoluble electron acceptors for growth, such as iron- and manganese-oxide as well as inert graphite electrodes in microbial fuel cells, also transfer electrons exocellularly. Soluble compounds, like humic substances, quinones, phenazines and riboflavin, can function as exocellular electron mediators enhancing this type of anaerobic respiration. However, direct electron transfer by cell-cell contact is important as well. This review addresses the mechanisms of exocellular electron transfer in anaerobic microbial communities. There are fundamental differences but also similarities between electron transfer to another microorganism or to an insoluble electron acceptor. The physical separation of the electron donor and electron acceptor metabolism allows energy conservation in compounds as methane and hydrogen or as electricity. Furthermore, this separation is essential in the donation or acceptance of electrons in some environmental technological processes, e.g. soil remediation, wastewater purification and corrosion.     

Bacterial extracellular electron transfer in bioelectrochemical systems

Bioelectrochemical systems (BES), typically microbial fuel cells (MFCs), have attracted increasing attention in the past decade due to their promising applications in many fields, such as bioremediation, energy generation and biosynthesis. Current-generating microorganisms play a key role in BES. The process of transferring electrons to electrode has been considered as a novel anaerobic bacteria respiration, and more and more bacteria capable of exchanging electrons with electrodes have been isolated. Among those bacteria, Shewanella and Geobacter genera are the most frequently used model organisms in the studies of BES, as well as the bacteria-electrode electron transfer mechanisms. Many significant new findings in the field of the bacterial extracellular electron transfer in BES have been reported recently. A better understanding of the mechanisms of bacterial extracellular electron transfer would provide more efficient strategies to enhance the applicability of BES. This review summarizes the recent advances of extracellular electron transfer mechanisms with foci on Shewanella and Geobacter species in BES.     

Humic substances act as electron acceptor and redox mediator for microbial dissimilatory azoreduction by Shewanella decolorationis S12

The potential for humic substances to serve as terminal electron acceptors in microbial respiration and the effects of humic substances on microbial azoreduction were investigated. The dissimilatory azoreducing microorganism Shewanella decolorationis S12 was able to conserve energy to support growth from electron transport to humics coupled to the oxidation of various organic substances or H2. Batch experiments suggested that when the concentration of anthraquinone-2-sulfonate (AQS), a humics analog, was lower than 3 mmol/l, azoreduction of strain S12 was accelerated under anaerobic condition. However, there was obvious inhibition to azoreduction when the concentration of the AQS was higher than 5 mmol/l. Another humics analog, anthraquinone-2-sulfonate (AQDS), could still prominently accelerate azoreduction, even when the concentration was up to 12 mmol/l, but the rate of acceleration gradually decreased with the increasing concentration of the AQDS. Toxic experiments revealed that AQS can inhibit growth of strain S12 if the concentration past a critical one, but AQDS had no effect on the metabolism and growth of strain S12 although the concentration was up to 20 mmol/l. These results demonstrated that a low concentration of humic substances not only could serve as the terminal electron acceptors for conserving energy for growth, but also act as redox mediator shuttling electrons for the anaerobic azoreduction by S. decolorationis S12. However, a high concentration of humic substances could inhibit the bacterial azoreduction, resulting on the one hand from the toxic effect on cell metabolism and growth, and on the other hand from competion with azo dyes for electrons as electron acceptor.     

Anaerobic biodecolorization mechanism of methyl orange by Shewanella oneidensis MR-1

In this work, we investigated the anaerobic decolorization of methyl orange (MO), a typical azo dye, by Shewanella oneidensis MR-1, which can use various organic and inorganic substances as its electron acceptor in natural and engineered environments. S. oneidensis MR-1 was found to be able to obtain energy for growth through anaerobic respiration accompanied with dissimilatory azo-reduction of MO. Chemical analysis shows that MO reduction occurred via the cleavage of azo bond. Block of Mtr respiratory pathway, a transmembrane electron transport chain, resulted in a reduction of decolorization rate by 80%, compared to the wild type. Knockout of cymA resulted in a substantial loss of its azo-reduction ability, indicating that CymA is a key c-type cytochrome in the electron transfer chain to MO. Thus, the MtrA-MtrB-MtrC respiratory pathway is proposed to be mainly responsible for the anaerobic decolorization of azo dyes such as MO by S. oneidensis.     

Bacterial degradation of aromatic pollutants: a paradigm of metabolic versatility.

Although most organisms have detoxification abilities (i.e mineralization, transformation and/or immobilization of pollutants), microorganisms, particularly bacteria, play a crucial role in biogeochemical cycles and in sustainable development of the biosphere. Next to glucosyl residues, the benzene ring is the most widely distributed unit of chemical structure in nature, and many of the aromatic compounds are major environmental pollutants. Bacteria have developed strategies for obtaining energy from virtually every compound under oxic or anoxic conditions (using alternative final electron acceptors such as nitrate, sulfate, and ferric ions). Clusters of genes coding for the catabolism of aromatic compounds are usually found in mobile genetic elements, such as transposons and plasmids, which facilitate their horizontal gene transfer and, therefore, the rapid adaptation of microorganisms to new pollutants. A successful strategy for in situ bioremediation has been the combination, in a single bacterial strain or in a syntrophic bacterial consortium, of different degrading abilities with genetic traits that provide selective advantages in a given environment. The advent of high-throughput methods for DNA sequencing and analysis of gene expression (genomics) and function (proteomics), as well as advances in modelling microbial metabolism in silico, provide a global, rational approach to unravel the largely unexplored potentials of microorganisms in biotechnological processes thereby facilitating sustainable development.     

金属还原菌希瓦氏菌厌氧呼吸能力及其在环境修复中的研究进展

Shewanella oneidensis MR-1是一种模式金属还原菌,它能够在厌氧条件下,将多种金属化合物和人工合成染料等作为电子受体还原代谢。因此,该菌常常被用于生态修复等研究。厌氧条件下,S.oneidensis MR-1能够将细胞质内或细胞内膜产生的电子通过定位于细胞内膜、细胞膜周质和细胞外膜上的c-血红色素蛋白或还原酶所组成的具有多样性的电子传递系统,最终传递到存在于细菌细胞外环境中的电子受体。通过对多种电子传递过程的介绍,进一步阐明其对污染物修复和纳米材料合成的机理,从而为未来对该类微生物的利用和开发提供更为充分的理论依据。     

Correlation between bacterial haemoglobin gene (vgb) and aeration: their effect on the growth and alpha-amylase activity in transformed Enterobacter aerogenes

AIMS: To evaluate the effects of bacterial haemoglobin on bacterial growth and alpha-amylase formation under different aeration conditions. METHODS AND RESULTS: Enterobacter aerogenes was transformed with the gene encoding Vitreoscilla (bacterial) haemoglobin, vgb. The growth kinetics and ability to synthesize alpha-amylase enzyme were investigated in this transformed Enterobacter strain as well as in two other Enterobacter control strains that do not harbour the vgb gene. Such comparison was made under variable aeration conditions, using the agitation rate as a measure of aeration. The expression of bacterial haemoglobin-supported cell growth determined as O.D.600 and cell viability in addition to the alpha-amylase production. These positive effects of bacterial haemoglobin were observed under both low and high aerations, but at different extents. CONCLUSIONS: In addition to improving cell growth under low aeration, the bacterial haemoglobin is able to promote bacterial cell tolerance during exposure to high oxygen tension. SIGNIFICANCE AND IMPACT OF THE STUDY: The expression of bacterial haemoglobin is advantageous in reducing the burden of certain toxic conditions such as high oxygen levels. It may have the same impact on some environmental toxic substances. This, haemoglobin biotechnology can be extended to induce enzymes of pollutants degradation or production of some useful industrial substances.     

Anaerobic mineralization of toluene by enriched sediments with quinones and humus as terminal electron acceptors

The anaerobic microbial oxidation of toluene to CO(2) coupled to humus respiration was demonstrated by use of enriched anaerobic sediments from the Amsterdam petroleum harbor (APH) and the Rhine River. Both highly purified soil humic acids (HPSHA) and the humic quinone moiety model compound anthraquinone-2,6-disulfonate (AQDS) were utilized as terminal electron acceptors. After 2 weeks of incubation, 50 and 85% of added uniformly labeled [(13)C]toluene were recovered as (13)CO(2) in HPSHA- and AQDS-supplemented APH sediment enrichment cultures, respectively; negligible recovery occurred in unsupplemented cultures. The conversion of [(13)C]toluene agreed with the high level of recovery of electrons as reduced humus or as anthrahydroquinone-2,6-disulfonate. APH sediment was also able to use nitrate and amorphous manganese dioxide as terminal electron acceptors to support the anaerobic biodegradation of toluene. The addition of substoichiometric amounts of humic acids to bioassay reaction mixtures containing amorphous ferric oxyhydroxide as a terminal electron acceptor led to more than 65% conversion of toluene (1 mM) after 11 weeks of incubation, a result which paralleled the partial recovery of electron equivalents as acid-extractable Fe(II). Negligible conversion of toluene and reduction of Fe(III) occurred in these bioassay reaction mixtures when humic acids were omitted. The present study provides clear quantitative evidence for the mineralization of an aromatic hydrocarbon by humus-respiring microorganisms. The results indicate that humic substances may significantly contribute to the intrinsic bioremediation of anaerobic sites contaminated with priority pollutants by serving as terminal electron acceptors.     

Proteome comparison of Vibrio cholerae cultured in aerobic and anaerobic conditions

The pathogen Vibrio cholerae causes severe diarrheal disease in humans. This environmental inhabitant has two distinct life cycles, in the environment and in the human small intestine, in which it differs in its multiplication behavior and virulence expression. Anaerobiosis, limitation of some nutrient elements, and excess burden from host metabolism reactants are the major stresses for V. cholerae living in intestine, in comparison to conditions in the environment and laboratory medium. For an insight into the response of V. cholerae to different microenvironments, we cultured the bacteria in aerobic and anaerobic conditions, and compared the whole cell proteome by two-dimensional electrophoresis. Among the protein spots identified, some protein species involved in aerobic respiration and the nutrient carbohydrate transporters were found to be more abundant in aerobic conditions, and some enzymes for anaerobic respiration and some stress response proteins were found more abundant in anaerobic culture. One spot corresponding to flagellin B subunit was decreased in anaerobic conditions, which suggests correlation with the meticulous regulation of bacterial motility during infection in the host intestine. This proteome analysis is the starting point for in-depth understanding of V. cholerae behavior in different environments.     

Physiology and biochemistry of reduction of azo compounds by Shewanella strains relevant to electron transport chain

Azo dyes are toxic, highly persistent, and ubiquitously distributed in the environments. The large-scale production and application of azo dyes result in serious environmental pollution of water and sediments. Bacterial azo reduction is an important process for removing this group of contaminants. Recent advances in this area of research reveal that azo reduction by Shewanella strains is coupled to the oxidation of electron donors and linked to the electron transport and energy conservation in the cell membrane. Up to date, several key molecular components involved in this reaction have been identified and the primary electron transportation system has been proposed. These new discoveries on the respiration pathways and electron transfer for bacterial azo reduction has potential biotechnological implications in cleaning up contaminated sites.