The molecular ecology of arsenic; probing the biogeochemical basis of a humanitarian disaster

饮用和灌溉用含高浓度砷的地下水正在毒害全球数百万人。例如，在西孟加拉邦和孟加拉国，砷的含量可以达到每升水含砷毫克量，这导致了被描述为“人类历史上最严重的大规模中毒”。尽管迫切需要从沉积物向水中释放砷的机制的基本信息，但这场人道主义灾难的原因仍然存在争议。几种可能的机制可能会释放吸附到沉积物中的矿物中的砷，所有这些机制都得到了激烈的辩论。这些变化包括含水层上部富砷黄铁矿氧化引起的沉积物矿物变化，或富砷Fe(III)氧化氢氧化物在沉积物深处还原条件下的分解，而水中的其他离子也可以活化吸附的砷，例如磷酸盐或碳酸盐。尽管这些变化是化学变化，但越来越多的人达成共识，认为沉积物中的微生物可能很好地推动了这些反应。事实上，我们实验室最近的结果表明，专门的地下微生物将有毒的砷吸附到沉积物中的矿物上。这些“金属还原细菌”在无氧条件下生长，从吸附的As(V)的还原和有机物的氧化中获得能量。随着其他实验室的研究越来越多地支持，现在的共识是，这种形式的微生物新陈代谢在控制全世界含水层沉积物中的砷浓度方面发挥着关键作用。然而，关于负责细菌的身份的信息很少，也没有模式生物可以作为恒河三角洲含水层中砷减少和动员机制的地球微生物研究的基础。迫切需要这些信息来支持补救工作或帮助制定更安全的用水做法，因为如果不详细了解原因，显然很难解决环境问题。这项工作的目的是解决我们对含水层中砷动员机制的理解的局限性，方法是对孟加拉西部这样一个含水层的微生物进行详细的、早就应该进行的研究，同时应用最先进的分子生物学技术来识别参与从沉积物中释放砷的基因和蛋白质。通过向微生物群落提供同位素标记的有机物质(醋酸盐和乳酸盐作为新有机物的替代品，通过抽水进入含水层，石油是较深沉积物中的电子供体)，我们将分离由沉积物中活跃的细菌合成的标记核酸，并使用遗传指纹技术来鉴定这些“活跃的”细菌，以及它们所包含的相应的砷降低/动员基因。由于许多砷基因将是新的，我们将在“元基因组库”中寻找它们，其中将包含来自沉积物的DNA大片段，这些DNA片段编码As(V)还原酶基因和其他高度保守的标记基因，我们可以用来准确地鉴定细菌。这将是第一次在这一领域使用元基因组文库的构建和筛选来鉴定潜在的新的AS(V)-呼吸细菌，而不需要培养它们。最后，为了更好地了解这些生物转化在砷循环中的作用，我们还将利用最先进的矿物学和地球化学技术研究矿物相和地下水成分，同时监测微生物群落和它们所表达的基因的变化。这将使我们能够开发一张详细的分子尺度图，了解微生物代谢对水和矿物结合形式砷的影响。

The use of groundwaters containing high concentrations of arsenic for drinking and irrigation is poisoning millions worldwide. For example, in West Bengal and Bangladesh arsenic levels can reach mg quantities of arsenic per litre of water, and this has led to what has been described as 'the worst mass poisoning in human history'. Despite the urgent need for fundamental information in the mechanism of arsenic release from sediments into water, the causes of this humanitarian disaster remain controversial. Several possible mechanisms may release arsenic sorbed to minerals in the sediments, and all have been debated vigorously. These include changes in the sediment minerals from the oxidation of arsenic-rich pyrite in the upper regions of the aquifers or the breakdown of arsenic-rich Fe(III) oxyhydroxides under reducing conditions deeper in the sediments, while other ions in the water could also mobilise sorbed arsenic e.g. phosphate or carbonate. Although these changes are chemical, there is a growing consensus that microorganisms in the sediments may well drive these reactions. Indeed, recent results from our laboratory have shown that specialist subsurface microorganisms mobilise the toxic arsenic sorbed to minerals in the sediments. Growing in the absence of oxygen, these 'metal-reducing bacteria' gain energy from the reduction of sorbed As(V) coupled to the oxidation of organic matter. With growing support from studies in other laboratories, there is now a consensus that this form of microbial metabolism plays a critical role in controlling arsenic concentrations in aquifer sediments worldwide. There is, however, little information on the identity of the bacteria responsible, and no model organisms on which to base 'geomicrobiological' studies on the mechanism of arsenic reduction and mobilisation in aquifers in the Ganges delta. This information is needed urgently to underpin remediation efforts or help develop safer practices for water use, as it is clearly very difficult to solve an environmental problem without a detailed understanding of the cause. The aim of this work is to address our limitations of the understanding of the mechanism of As mobilisation in aquifers by conducting a detailed and long overdue study of the microbiology of such an aquifer in W. Bengal, alongside the application of state of the art molecular biology techniques to identify the genes and proteins involved in arsenic release from the sediments. By feeding microbial communities with isotopically labeled organic matter (acetate and lactate as proxies for new organic matter drawn into the aquifers by water abstraction and petroleum which is an electron donor in deeper sediments), we will isolate the labeled nucleic acids synthesized from bacteria that are active in the sediments when arsenic is mobilised, and use genetic fingerprinting techniques to identify these 'active' bacteria, and the corresponding arsenic reducing/mobilising genes that they contain. As many of the arsenic genes will be novel, we will look for them in 'metagenomic libraries' which will contain large fragments of DNA from the sediments that encode both the As(V) reductase genes and other highly conserved marker genes that we can use to identify the bacteria accurately. This will be the first time that metagenomic library construction and screening has been used in this field to identify potentially novel As(V)-respiring bacteria without the need to culture them. Finally, so that we can gain a better picture of the role of these biological transformations in the arsenic cycle, we will also study the mineral phases and groundwater composition using state of the art mineralogical and geochemical techniques, while we are monitoring changes in the microbial communities and the genes that they are expressing. This will allow us to develop a detailed molecular-scale picture of the impact of microbial metabolism on the aqueous and mineral-bound forms of arsenic.