Development & Application of Molecular Diagnostics for Fe & P Limitation of Trichodesmium Nitrogen Fixation & Productivity

海洋中二氧化碳的光合固定约占初级生产总量的一半。 这种初级生产的一个重要组成部分是由于海洋蓝藻的生长，例如固氮菌Trichodesmium。 例如，在北大西洋热带和亚热带地区，Trichodesmium spp。被认为是最重要的初级生产者。 此外，在该地区，毛藻将最大部分的新氮引入到透光区（~30mg N/m2/天），该值超过了跨温跃层硝酸盐的估计通量。 尽管有许多生物地球化学因素可能影响毛藻生产力和 N2 固定（例如养分、光照、温度），但最近的研究强调了 P 和 Fe 生物利用度的至关重要性。 为了加深我们对这些元素的生物利用度如何限制初级生产和 N2 固定的理解，这些 PI 开发了针对 Trichodesmium 中 Fe 和 P 限制的特定分子诊断方法。 PI 已确定 P 胁迫的 Trichodesmium 菌落的无菌培养物诱导酶 PhoA，而 Fe 胁迫的菌落表达蛋白质 IdiA。 PI 还证明他们可以检测野外群体中的 PhoA 活性和 IdiA 表达。在该项目中，实验室研究将用于确定这些诊断与毛藻生产力和氮固定能力下降之间的关系。 这项工作将导致两种经过充分测试的分子诊断方法的开发和应用，以确定 P、Fe 或两者是否限制 Trichodesmium C 和 N2 的固定。物种特异性 PhoA 活性的测定称为酶标记荧光，利用市售试剂，并且与在大肠杆菌中异源表达的 IdiA 一样，PI 将为其他人提供 IdiA 蛋白或抗血清。 这项研究的影响将是研究人员可以使用工具来快速、轻松地识别任何环境中的毛藻物种中的 P 和 Fe 限制。

Photosynthetic fixation of CO2 in the oceans accounts for approximately half of total primary production. A significant component of this primary production is due to the growth of oceanic cyanobacteria, such as the diazotroph Trichodesmium. For example, in the tropical and subtropical North Atlantic, Trichodesmium spp. are thought to be the most significant primary producers. Additionally, in this region Trichodesmium introduces the largest fraction of new nitrogen to the euphotic zone (~30mg N/m2/day), a value that exceeds the estimated flux of nitrate across the thermocline. Although there are many biogeochemical factors that may impact Trichodesmium productivity and N2 fixation (e.g. nutrients, light, temperature), recent studies have emphasized the critical importance of P and Fe bioavailability. To increase our understanding of how the bioavailability of these elements may constrain primary production and N2 fixation, these PIs developed specific molecular diagnostics for Fe and P limitation in Trichodesmium. The PIs have determined that axenic cultures of P-stressed Trichodesmium colonies induce the enzyme PhoA and Fe-stressed colonies express the protein IdiA. The PIs have also demonstrated that they can detect PhoA activity and IdiA expression in field populations. In this project, laboratory studies will be used to determine how these diagnostics correlate with decreases in both Trichodesmium productivity and N2 fixation. This work will result in the development and application of two well-tested molecular diagnostics for determining if P, Fe, or both, limit Trichodesmium C and N2 fixation. The assay for species-specific PhoA activity, called Enzyme Labeled Fluorescence, utilizes commercially available reagents, and as with IdiA heterologously-expressed in Escherichia coli, the PIs will provide the IdiA protein or antiserum to others. The impact of this research will be tools that can be used by investigators to quickly and easily identify P and Fe limitation in Trichodesmium species from any environment.