Nitrogen Fixation in the Open Ocean: Assessing the Role of Recently Discovered Diazotrophs

可获得的营养物质，主要是磷、氮和铁，限制了海洋的生产力。氮通常被认为是一个重要的，如果不是限制，营养物在少营养的海洋。氮以二氮气体的形式存在，是地球大气的主要组成部分，在海水中以溶解气体的形式大量存在。然而，大多数生物可以利用硝酸盐、铵或有机氮形式的氮，但不能直接利用二氮。这种形式的氮只能被重氮营养体使用，重氮营养体可以通过酶氮酶的作用将氮“固定”成铵。不同的原核生物分类群具有固定氮的能力，包括异养细菌、蓝藻和古细菌。尽管可以固定氮的微生物种类繁多，但以前很少有来自开放海洋的固定氮分类群的报道。最近在大西洋和太平洋的研究表明，固氮在氮动力学中比以前认为的更重要，并且可能存在大量未被解释的氮通量进入混合层，这可能是由于固氮。已知重氮营养菌（蓝藻菌、毛菌和内共生体黎切菌）对固氮的估计无法解释这一估计的N通量。利用分子方法，最近已经表明，在开放的海洋环境中有多种微生物，它们具有固氮的遗传能力。在这个项目中，这些微生物的生态和生物学意义将通过分子方法、培养努力、固氮率测量和稳定同位素测量来研究。RNA技术（nifH逆转录酶聚合酶链反应，RT-PCR）的最新发展使得在季节性研究和实验操作中检查独特的氮酶种型的表达成为可能。这些研究将集中在HOT站点，以识别和表征新型固氮蓝藻，确定控制其生长和活动的因素，并使用稳定同位素技术量化其对当地氮收支的贡献。与此同时，我们将通过N-15示踪剂测量和RT-PCR来研究与无脊椎动物相关的固氮活性实验，以确定产生固氮活性的固氮微生物类型。最后，新型重氮营养体将通过检查固氮装置（除nifH外的氮酶基因）来确定这些生物的固氮装置是否不寻常，这可能具有生物技术应用和进化意义。这些新型固氮微生物的鉴定和表征对于评估海洋的氮收支具有及时和潜在的重要意义。

Zehr 9977460 and Montoya 9977528The availability of nutrients, primarily phosphorus, nitrogen and iron, limits the productivity of the oceans. Nitrogen is often believed to be an important, if not limiting, nutrient in oligotrophic oceans. Nitrogen, in the form of dinitrogen gas, is a major component of the Earth's atmosphere and is abundant in seawater as dissolved gas. However, most organisms can use nitrogen in the form of nitrate, ammonium, or organic nitrogen, but cannot directly use dinitrogen. This form of nitrogen can be used only by diazotrophs, which can "fix" nitrogen into ammonium through the action of the enzyme nitrogenase. Diverse prokaryotic taxa have the ability to fix nitrogen, including heterotrophic bacteria, cyanobacteria and Archaea. Despite the diversity of types of microorganisms that can fix nitrogen, very few nitrogen--fixing taxa have previously been reported from the open ocean. Recent studies in the Atlantic and Pacific Oceans indicate that nitrogen fixation is more important in nitrogen dynamics than previously believed, and that there may be a large, unaccounted for flux of nitrogen into the mixed layer, which may be due to nitrogen fixation. Estimates of the nitrogen fixation by known diazotrophs, the cyanobacterium Trichodesmium and endosymbiont Richelia, cannot account for this estimated N flux.Using a molecular approach, it has very recently been shown that there are diverse microorganisms in the open ocean environment, which have the genetic capacity for nitrogen fixation. In this project, the ecological and biological significance of these microorganisms will be examined using molecular approaches, cultivation efforts, nitrogen fixation rate measurements, and stable isotope measurements. Recent developments in RNA technology (nifH reverse-transcriptase polymerase chain reaction, RT-PCR) make it possible to examine the expression of the unique nitrogenase phylotypes in seasonal studies, and in experimental manipulations. These studies will focus at the HOT site in order to identify and characterize the novel nitrogen-fixing cyanobacterium, to determine the factors controlling its growth and activity, and to quantify its contribution to the local nitrogen budget using stable isotope techniques. In parallel, experiments on nitrogen-fixation activity associated with invertebrates will be investigated by N-15 tracer measurements and RT-PCR to identify the types of nitrogen-fixing microorganisms responsible for observed nitrogen fixation activity. Finally, the novel diazotroph will be characterized by examining the nitrogen fixation apparatus (nitrogenase genes other than nifH) to determine whether the nitrogen fixation apparatus of these organisms is unusual, which may have biotechnological applications as well as evolutionary implications. The identification and characterization of these novel nitrogen-fixing microorganisms in the open ocean is timely and potentially important for evaluating the nitrogen budget of the sea.