MIP: Re-evaluating Ammonia Oxidation in Coastal Estuarine Sediments: Assessing the Relative Diversity, Abundance, and Activity of Ammonia-oxidizing Archaea and Bacteria

硝化作用，微生物介导的铵（NH 4+）通过亚硝酸盐（NO2-）转化为硝酸盐（NO3-），在整个海洋的氮循环中起着重要的作用。 在沿海/河口系统中，这一过程通过消耗氨和氧，通过产生强有力的温室气体N2 O，并通过将含氮有机物的分解与通过反硝化作用（微生物将NO3-转化为N2气体）的氮损失联系起来，发挥着特别重要的作用。尽管它们具有巨大的生物地球化学重要性，但令人惊讶的是，关于负责河口硝化作用的微生物群落，或者关键环境因素（例如，氨、氧、盐度、金属等）梯度影响它们的分布、多样性和活性。虽然硝化的第一步和限速步骤，氨氧化（由氨单加氧酶催化），长期以来一直被认为是由细菌（氨氧化细菌，AOB）的成员专门进行的，但最近发现，一些嗜温Crenarchaeota（古细菌代表地球上最普遍和丰富的原核生物群之一）也可能是重要的氨氧化剂。氨氧化古菌（AOA）的发现代表了全球氮循环中一个令人惊讶的新“转折”，因此我们对硝化作用的微生物生态学的理解需要仔细重新评估。该项目的总体目标是比较AOA和AOB社区的相对多样性，丰度和活动的埃尔克霍恩斯劳，一个小的农业影响沿海加州河口，开放到蒙特雷湾。细菌和古细菌amoA基因，编码氨单加氧酶的催化亚基，将被用作分子标记，以表征这两个明显的“功能等效”组在多个日期和采样点沿着河口梯度。为了编译一个新的河口氨氧化剂，其中几乎没有什么是目前已知的文化收集，也将进行密集的培养努力。埃尔克霍恩斯劳是一个主要的湿地复杂的加州中部，被认为是最重要的生态河口在该州，证明了其罕见的国家河口研究保护区（NERR）的地位。与大多数河口一样，硝化作用在埃尔克霍恩斯劳河口的氮循环和去除中起着关键作用。 该项目将提供有关硝化群落的多样性和功能如何受到复杂的物理/化学梯度和环境扰动（例如，N径流）。由此收集的关于已开垦和未开垦河口AOB，特别是AOA的遗传、功能和生物地球化学信息将是广泛科学学科的宝贵资源（例如，微生物学、海洋学、地球生物学、环境工程）。 与埃尔克霍恩斯劳NERR合作，本研究的相关数据将以公众友好的形式提供给保护区协调的教育和推广计划。最后，这个项目将帮助支持，培训和教育研究生，本科生和高中生，以及博士后研究人员。

Nitrification, the microbially-mediated conversion of ammonium (NH4+) to nitrate (NO3-) via nitrite (NO2-), plays a quantitatively important role in nitrogen cycling throughout the ocean. In coastal/estuarine systems, this process plays a particularly critical role by consuming ammonia and oxygen, via production of the potent greenhouse gas, N2O, and by linking the decomposition of nitrogenous organic matter to N loss via denitrification (microbial conversion of NO3- to N2 gas). Despite their tremendous biogeochemical importance, surprisingly little is known regarding the microbial communities responsible for estuarine nitrification, or how key environmental factors (e.g., ammonia, oxygen, salinity, metals, etc.) and gradients influence their distribution, diversity, and activity. Although the first and rate-limiting step of nitrification, ammonia oxidation (catalyzed by the enzyme ammonia monooxygenase), has been long believed to be carried out exclusively by members of the Bacteria (ammonia-oxidizing bacteria, AOB), it has recently been discovered that some mesophilic Crenarchaeota (archaea representing one of the most ubiquitous and abundant prokaryotic groups on the planet) may also be significant ammonia-oxidizers. The discovery of ammonia-oxidizing archaea (AOA) represents a surprising new 'twist' within the global N cycle, and our understanding of the microbial ecology of nitrification therefore requires careful re-evaluation. The overarching goal of this project is to compare the relative diversity, abundance, and activity of AOA and AOB communities in Elkhorn Slough, a small agriculturally-impacted coastal California estuary that opens into Monterey Bay. Bacterial and archaeal amoA genes, encoding the catalytic subunit of ammonia monooxygenase, will be used as molecular markers to characterize these two apparently 'functionally-equivalent' groups at multiple dates and sampling sites along the estuarine gradient. In order to compile a culture collection of novel estuarine ammonia-oxidizers, for which virtually nothing is currently known, an intensive cultivation effort will also be undertaken.Elkhorn Slough is a principal wetland complex in central California and is considered one of the most ecologically important estuaries in the state, as evidenced by its rare National Estuarine Research Reserve (NERR) status. As in most estuaries, nitrification plays a pivotal role in nitrogen cycling and removal in the Elkhorn Slough estuary. This project will provide critical information regarding how the diversity and functioning of nitrifying communities are influenced by complex physical/chemical gradients and environmental perturbations (e.g., N runoff). The resultant collection of genetic, functional, and biogeochemical information regarding cultivated and uncultivated estuarine AOB and, especially, AOA will be a valuable resource to a wide range of scientific disciplines (e.g., microbiology, oceanography, geobiology, environmental engineering). In collaboration with the Elkhorn Slough NERR, relevant data from this study will be made available in a public-friendly format for education and outreach programs coordinated by the Reserve. Finally, this project will help support, train, and educate graduate, undergraduate, and high school students, as well as a postdoctoral researcher.