Regulation, ecophysiology, and kinetic parameters of uncultured N-gas flux associated anaerobic microbial communities in agricultural soils: Towards a microscale spatial resolution

反硝化剂（将n-氧化物还原为N2O和/或N2）、非反硝化N2O还原剂（将N2O还原为N2）和异化硝酸盐还原剂（DNRA；将n-氧化物还原为NH4+）影响温室气体N2O的排放以及氮的保留，从而影响植物营养。n2o还原剂和DNRA与反硝化剂竞争。决定其竞争力和产氮能力的生态生理特征，在未培养的分类群中基本上是未知的。因此，以下假设将进一步得到解决：(i)对环境因素的反硝化反应是由不同的微生物群落形成的，包括迄今为止未知的物种，并且可以通过其内在的生态生理特性来解释。（ii）与反硝化、N2O还原和DNRA相关的基因表达和群落结构反映了代谢状态和潜力，从而增加了N2O和N2通量的可预测性。第一阶段的结果总结在两篇出版物和三篇提交的手稿或准备中的手稿中（附）。采用各种条件下的高通量孵育（包括15n示踪剂）结合功能基因表达，以及基于基因和转录本的下一代测序来鉴定表观Michaelis-Menten和生理参数。主要分析反硝化菌的功能基因（编码亚硝酸盐还原酶产生NO的nirK/S，编码N2O还原酶产生N2的nosZ I和II）和DNRA（编码亚硝酸盐还原酶产生NH4+的nrfA）。第二阶段研究pH、温度、植物根系和有机碳对目标群体响应模式的影响。利用基于RNA和DNA的稳定同位素探测技术鉴定活性根渗出物的靶生物。目标群体对环境控制因素的调节将在中生态系统和作为中心实验的实地研究中进行分析。同样也将讨论目标群体的空间分布问题。这些数据将扩展反硝化器-调节表型的概念，并深入了解不同条件下目标群体的生态生理和竞争。将开发基于反硝化剂、DNRA和clade II N2O还原剂的N2O / N2比电位分子微生物社区代理。各种动力学参数、响应函数和微生物群落结构数据将提供反硝化和n -气体通量的建模。

Denitrifiers (reducing N-oxides to N2O and/ or N2), non-denitrifying N2O-reducers (reduce N2O to N2), and dissimilatory nitrate reducers (DNRA; reduce N-oxides to NH4+) impact the emission of the greenhouse gas N2O as well as retention of nitrogen and thus plant nutrition. N2O-reducers and DNRA compete with denitrifiers. Ecophysiological traits determining their competitiveness and N-gas production capabilities and are essentially unknown for uncultured taxa. Thus, the following hypotheses will further be addressed: (i) Denitrification response to environmental factors is shaped by contrasting microbial communities including hitherto unknown species, and can be explained by their intrinsic ecophysiological properties. (ii) Gene expression and community structure associated with denitrification, N2O reduction and DNRA reflect metabolic states and potentials, thus increasing predictability of N2O and N2 fluxes. Outcomes from phase I were summarized in two publications and three submitted manuscripts or manuscripts in preparation (attached). High-throughput incubations under various conditions (including 15N-tracers) combined with functional gene expression, as well as gene and transcript based next generation sequencing will be applied to identify apparent Michaelis-Menten and physiological parameters. Functional genes of denitrifiers (nirK/S encoding nitrite reductases yielding NO; nosZ I and II encoding N2O reductase yielding N2), and DNRA (nrfA encoding nitrite reductases yielding NH4+), will be primarily analyzed. Phase II focuses on the effect of pH, temperature, plant roots and organic carbon on response patterns of target groups. Active root exudate fueled target organisms will be identified by RNA and DNA based stable isotope probing. Regulation of target groups in response to environmental control factors will be analyzed in mesocosms and field studies performed as central experiments. The spatial distribution of target groups will likewise be addressed. Such data will extend the denitrifier-regulatory-phenotype-concept and give insights into the ecophysiology and competition of target groups under various conditions. A molecular microbial community-based proxy for N2O to N2 ratio potentials based on denitrifiers, DNRA, and clade II N2O reducers will be developed. Various kinetic parameters, response function, and microbial community structure data will be provided for modelling of denitrification and N-gas fluxes.