Award: OCE-1357024

Nitrification--the microbial oxidation of ammonia to nitrate via a nitrite intermediate--plays a critical role in the global nitrogen cycle and especially within marine ecosystems, where Thaumarchaeota are thought to be responsible for most of the ammonia oxidation in the water column. The overarching goal of this project was to examine the relationships between the diversity and abundance of ammonia-oxidizing archaea (AOA) and nitrification activity, as well as physical/chemical gradients, across multiple depths at two stations (M1 and M2) within the Monterey Bay Time Series (MBTS). Over a two-year, near-monthly time series at M1 and M2, we observed repeatable seasonal and depth-based patterns of Thaumarchaeota ecotype abundance that highlighted a clear delineation between populations in shallow euphotic (≤ 50 m) versus deeper mesopelagic (60-500 m) depths. Euphotic depths show greater seasonality and influence from light, while mesopelagic waters have trends based on water mass and other covarying features with depth. Three major ecotypes, previously defined based on functional gene diversity, were recovered within our massive 16S rRNA (V4-V5) amplicon sequence dataset: a Nitrosopumilus-like (NP) group, a Nitrosopelagicus-like ecotype containing 'shallow' water column A (WCA) members, and an ecotype affiliated with the 'deep' water column B (WCB) Thaumarchaeota. Together, these ecotypes comprised 7.1% of the total overall community (up to 27% of a given sample) and appeared to occupy different habitats within the Monterey Bay water column. Interestingly, only the NP ecotype showed a correlation with measured nitrification rates. Our results support the importance of ecotype-based analysis of Thaumarchaeota and show that their abundance and distribution are controlled based on their water column position, with a distinct shift at 50 m between euphotic and mesopelagic depths. Co-occurrence networks inferred using the 16S rRNA dataset were used to explore potential associations involving Thaumarchaeota and other members of the microbial community. Networks systematically reflected depth-related patterns in the abundances of ecotype populations, suggesting thaumarchaeal ecotypes as keystone members of the microbial community below the euphotic zone. Differential environmental controls on the ecotype populations were further evident in subnetwork modules showing preferential co-occurrence of operational taxonomic units (OTUs) belonging to the same ecotype cluster. Correlated abundances of Thaumarchaeota and heterotrophic bacteria indicated potential reciprocal interactions via dissolved organic matter transformations. Notably, the networks recovered ecotype-specific associations between OTUs of Thaumarchaeota and Nitrospina--the major nitrite-oxidizing bacteria in the ocean. Even at depths where WCB-like Thaumarchaeota dominated, Nitrospina OTUs were found to preferentially co-occur with WCA-like and Nitrosopumilus-like thaumarchaeal OTUs, highlighting the need to investigate the ecological implications of the composition of both ammonia- and nitrite-oxidizing microbial assemblages in marine waters. Although the distribution, diversity, and abundance of AOA has been studied extensively for the last 15 years--primarily using the amoA gene (encoding ammonia monooxygenase subunit A) as a molecular marker--significant gaps still remain in our understanding of thaumarchaeal ecology and metabolism. Furthermore, despite multiple lines of evidence pointing to a central role for copper‐containing nitrite reductase (NirK) in AOA metabolism, the thaumarchaeal nirK gene has rarely been studied in the environment. In this project, we examined the diversity of nirK in the marine pelagic environment, in light of previously described ecological patterns of pelagic thaumarchaeal populations. Phylogenetic analyses reveal that nirK better resolves diversification patterns of marine Thaumarchaeota, compared to the conventionally used amoA gene. We demonstrated that the three major phylogenetic clusters of marine nirK correspond to the three 'ecotype'populations of pelagic Thaumarchaeota described above. In this context, we also examined the relative distributions of the three variant groups in metagenomes and metatranscriptomes representing two depth profiles (from M1 and M2) in Monterey Bay. Our results reveal that nirK effectively tracks the dynamics of thaumarchaeal ecotype populations, particularly finer‐scale diversification patterns within major lineages. Finally, we have assembled and binned 10 metagenomes representing two depth profiles at stations M1 and M2, respectively. Over 400 draft quality metagenome-assembled genomes (MAGs) have been obtained from diverse marine bacteria and archaea, including a number of novel Thaumarchaeota. In addition to aiding in better elucidating the biogeochemistry of Monterey Bay, these MAGs will serve as a valuable resource for the field of marine microbial ecology in general. Overall, this project has provided valuable new insights into how the diversity, abundance, and activity of AOA communities are influenced by complex and fluctuating environmental conditions in Monterey Bay--one of the most productive and biologically diverse regions of the global ocean. This project also led to the training and mentorship of Stanford undergraduate and graduate students, as well as postdoctoral researchers. Last Modified: 07/29/2019 Submitted by: Christopher A Francis