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

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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.