Abundance and diversity of archaeal ammonia oxidizers in a coastal groundwater system

Author Posting. © The Author(s), 2010. This is the author's version of the work. It is posted here by permission of American Society for Microbiology for personal use, not for redistribution. The definitive version was published in Applied and Environmental Microbiology 76 (2010): 7938-7948, doi:10.1128/AEM.02056-09.Nitrification, the microbially-catalyzed oxidation of ammonia to nitrate, is a key process in the nitrogen cycle. Archaea have been implicated in the first part of the nitrification pathway (oxidation of ammonia to nitrite), but the ecology and physiology of these organisms remain largely unknown. This work describes two different populations of sediment-associated ammonia-oxidizing archaea (AOA) in a coastal groundwater system on Cape Cod, Massachusetts. Sequence analysis of the ammonia monooxygenase subunit A gene (amoA) shows that one population of putative AOA inhabits the upper meter of the sediment where they may experience frequent ventilation with tidally-driven overtopping and infiltration of bay water supplying dissolved oxygen, ammonium and perhaps organic carbon. A genetically distinct population occurs deeper in the sediment, in a mixing zone between a nitrate- and oxygen-rich freshwater zone and a reduced, ammonium-bearing salt water wedge. Both of these AOA populations are coincident with increases in the abundance of Group I crenarchaeota 16S rRNA gene copies.Funding was provided by WHOI’s Coastal Ocean Institute to DRR and KLC, as well as funding from NSF/OCE project #05-24994 to KLC

Oxygen isotopic composition of nitrate and nitrite produced by nitrifying cocultures and natural marine assemblages

Author Posting. © Association for the Sciences of Limnology and Oceanography, 2012. This article is posted here by permission of Association for the Sciences of Limnology and Oceanography for personal use, not for redistribution. The definitive version was published in Limnology and Oceanography 57 (2012): 1361-1375, doi:10.4319/lo.2012.57.5.1361.The δ18O value of nitrate produced during nitrification (δ18ONO3,nit) was measured in experiments designed to mimic oceanic conditions, involving cocultures of ammonia-oxidizing bacteria or ammonia-oxidizing archaea and nitrite-oxidizing bacteria, as well as natural marine assemblages. The estimates of ranged from −1.5‰ ± 0.1‰ to +1.3‰ ± 1.4‰ at δ18O values of water (H2O) and dissolved oxygen (O2) of 0‰ and 24.2‰ vs. Vienna Standard Mean Ocean Water, respectively. Additions of 18O-enriched H2O allowed us to evaluate the effects of oxygen (O) isotope fractionation and exchange on . Kinetic isotope effects for the incorporation of O atoms were the most important factors for setting overall values relative to the substrates (O2 and H2O). These isotope effects ranged from +10‰ to +22‰ for ammonia oxidation (O2 plus H2O incorporation) and from +1‰ to +27‰ for incorporation of H2O during nitrite oxidation. values were also affected by the amount and duration of nitrite accumulation, which permitted abiotic O atom exchange between nitrite and H2O. Coculture incubations where ammonia oxidation and nitrite oxidation were tightly coupled showed low levels of nitrite accumulation and exchange (3% ± 4%). These experiments had values of −1.5‰ to +0.7‰. Field experiments had greater accumulation of nitrite and a higher amount of exchange (22% to 100%), yielding an average value of +1.9‰ ± 3.0‰. Low levels of biologically catalyzed exchange in coculture experiments may be representative of nitrification in much of the ocean where nitrite accumulation is low. Abiotic oxygen isotope exchange may be important where nitrite does accumulate, such as oceanic primary and secondary nitrite maxima.This research was funded by the National Science Foundation Chemical Oceanography grants 05-26277 and 09- 610998 to K.L.C

Nitrogen cycling in the secondary nitrite maximum of the eastern tropical North Pacific off Costa Rica

Author Posting. © American Geophysical Union, 2015. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Global Biogeochemical Cycles 29 (2015): 2061–2081, doi:10.1002/2015GB005187.Nitrite is a central intermediate in the marine nitrogen cycle and represents a critical juncture where nitrogen can be reduced to the less bioavailable N2 gas or oxidized to nitrate and retained in a more bioavailable form. We present an analysis of rates of microbial nitrogen transformations in the oxygen deficient zone (ODZ) within the eastern tropical North Pacific Ocean (ETNP). We determined rates using a novel one-dimensional model using the distribution of nitrite and nitrate concentrations, along with their natural abundance nitrogen (N) and oxygen (O) isotope profiles. We predict rate profiles for nitrate reduction, nitrite reduction, and nitrite oxidation throughout the ODZ, as well as the contributions of anammox to nitrite reduction and nitrite oxidation. Nitrate reduction occurs at a maximum rate of 25 nM d−1 at the top of the ODZ, at the same depth as the maximum rate of nitrite reduction, 15 nM d−1. Nitrite oxidation occurs at maximum rates of 10 nM d−1 above the secondary nitrite maximum, but also in the secondary nitrite maximum, within the ODZ. Anammox contributes to nitrite oxidation within the ODZ but cannot account for all of it. Nitrite oxidation within the ODZ that is not through anammox is also supported by microbial gene abundance profiles. Our results suggest the presence of nitrite oxidation within the ETNP ODZ, with implications for the distribution and physiology of marine nitrite-oxidizing bacteria, and for total nitrogen loss in the largest marine ODZ.National Science Foundation. Grant Numbers OCE 05-26277, OCE 09-610998; WHOI Coastal Ocean Institute2016-06-1

Nitrite cycling in the primary nitrite maxima of the eastern tropical North Pacific

The primary nitrite maximum (PNM) is a ubiquitous feature of the upper ocean, where nitrite accumulates in a sharp peak at the base of the euphotic zone. This feature is situated where many chemical and hydrographic properties have strong gradients and the activities of several microbial processes overlap. Near the PNM, four major microbial processes are active in nitrite cycling: ammonia oxidation, nitrite oxidation, nitrate reduction and nitrite uptake. The first two processes are mediated by the nitrifying archaeal/bacterial community, while the second two processes are primarily conducted by phytoplankton. The overlapping spatial habitats and substrate requirements for these microbes have made understanding the formation and maintenance of the PNM difficult. In this work, we leverage high-resolution nutrient and hydrographic data and direct rate measurements of the four microbial processes to assess the controls on the PNM in the eastern tropical North Pacific (ETNP). The depths of the nitrite maxima showed strong correlations with several water column features (e.g., top of the nitracline, top of the oxycline, depth of the chlorophyll maximum), whereas the maximum concentration of nitrite correlated weakly with only a few water column features (e.g., nitrate concentration at the nitrite maximum). The balance between microbial production and consumption of nitrite was a poor predictor of the concentration of the nitrite maximum, but rate measurements showed that nitrification was a major source of nitrite in the ETNP, while phytoplankton release occasionally accounted for large nitrite contributions near the coast. The temporal mismatch between rate measurements and nitrite standing stocks suggests that studies of the PNM across multiple timescales are necessary.</p

Multiple Metabolisms Constrain the Anaerobic Nitrite Budget in the Eastern Tropical South Pacific

The Eastern Tropical South Pacific is one of the three major oxygen deficient zones (ODZs) in the global ocean and is responsible for approximately one third of marine water column nitrogen loss. It is the best studied of the ODZs and, like the others, features a broad nitrite maximum across the low oxygen layer. How the microbial processes that produce and consume nitrite in anoxic waters interact to sustain this feature is unknown. Here we used 15N-tracer experiments to disentangle five of the biologically mediated processes that control the nitrite pool, including a high-resolution profile of nitrogen loss rates. Nitrate reduction to nitrite likely depended on organic matter fluxes, but the organic matter did not drive detectable rates of denitrification to N2. However, multiple lines of evidence show that denitrification is important in shaping the biogeochemistry of this ODZ. Significant rates of anaerobic nitrite oxidation at the ODZ boundaries were also measured. Lodate was a potential oxidant that could support part of this nitrite consumption pathway. We additionally observed N2 production from labeled cyanate and postulate that anammox bacteria have the ability to harness cyanate as another form of reduced nitrogen rather than relying solely on ammonification of complex organic matter. The balance of the five anaerobic rates measured—anammox, denitrification, nitrate reduction, nitrite oxidation, and dissimilatory nitrite reduction to ammonium—is sufficient to reproduce broadly the observed nitrite and nitrate profiles in a simple one-dimensional model but requires an additional source of reduced nitrogen to the deeper ODZ to avoid ammonium overconsumption. ©2017. American Geophysical Union. All Rights Reserved

Protocols for Assessing Transformation Rates of Nitrous Oxide in the Water Column

Nitrous oxide (N2O) is a potent greenhouse gas and an ozone destroying substance. Yet, clear step-by-step protocols to measure N2O transformation rates in freshwater and marine environments are still lacking, challenging inter-comparability efforts. Here we present detailed protocols currently used by leading experts in the field to measure water-column N2O production and consumption rates in both marine and other aquatic environments. We present example 15N-tracer incubation experiments in marine environments as well as templates to calculate both N2O production and consumption rates. We discuss important considerations and recommendations regarding (1) precautions to prevent oxygen (O2) contamination during low-oxygen and anoxic incubations, (2) preferred bottles and stoppers, (3) procedures for 15N-tracer addition, and (4) the choice of a fixative. We finally discuss data reporting and archiving. We expect these protocols will make 15N-labeled N2O transformation rate measurements more accessible to the wider community and facilitate future inter-comparison between different laboratories

Inverse kinetic isotope fractionation during bacterial nitrite oxidation

Author Posting. © Elsevier B.V., 2009. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Geochimica et Cosmochimica Acta 73 (2009): 2061-2076, doi:10.1016/j.gca.2008.12.022.Natural abundance stable isotopes in nitrate (NO3-), nitrite (NO2-), and nitrous oxide (N2O) have been used to better understand the cycling of nitrogen in marine and terrestrial environments. However, in order to extract the greatest information from the distributions of these isotopic species, the kinetic isotope effects for each of the relevant microbial reactions are needed. To date, kinetic isotope effects for nitrite oxidation and anaerobic ammonium oxidation (anammox) have not been reported. In this study, the nitrogen isotope effect was measured for microbial nitrite oxidation to nitrate. Nitrite oxidation is the second step in the nitrification process, and it plays a key role in the regeneration of nitrate in the ocean. Surprisingly, nitrite oxidation occurred with an inverse kinetic isotope effect, such that the residual nitrite became progressively depleted in 15N as the reaction proceeded. Three potential explanations for this apparent inverse kinetic isotope effect were explored: 1) isotope exchange equilibrium between nitrite and nitrous acid prior to reaction, 2) reaction reversibility at the enzyme level, and 3) true inverse kinetic fractionation. Comparison of experimental data to ab initio calculations and theoretical predictions leads to the conclusion that the fractionation is most likely inverse at the enzyme level. Inverse kinetic isotope effects are rare, but the experimental observations reported here agree with kinetic isotope theory for this simple N-O bond-forming reaction. Nitrite oxidation is therefore fundamentally different from all other microbial processes in which N isotope fractionation has been studied. The unique kinetic isotope effect for nitrite oxidation should help to better identify its role in the cycling of nitrite in ocean suboxic zones, and other environments in which nitrite accumulates.Funding from NSF award OCE 05-26277 to KLC is also gratefully acknowledged

The Angola Gyre is a hotspot of dinitrogen fixation in the South Atlantic Ocean

© The Author(s), 2022. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Marshall, T., Granger, J., Casciotti, K. L., Dahnke, K., Emeis, K.-C., Marconi, D., McIlvin, M. R., Noble, A. E., Saito, M. A., Sigman, D. M., & Fawcett, S. E. The Angola Gyre is a hotspot of dinitrogen fixation in the South Atlantic Ocean. Communications Earth & Environment, 3(1), (2022): 151, https://doi.org/10.1038/s43247-022-00474-x.Biological dinitrogen fixation is the major source of new nitrogen to marine systems and thus essential to the ocean’s biological pump. Constraining the distribution and global rate of dinitrogen fixation has proven challenging owing largely to uncertainty surrounding the controls thereon. Existing South Atlantic dinitrogen fixation rate estimates vary five-fold, with models attributing most dinitrogen fixation to the western basin. From hydrographic properties and nitrate isotope ratios, we show that the Angola Gyre in the eastern tropical South Atlantic supports the fixation of 1.4–5.4 Tg N.a−1, 28-108% of the existing (highly uncertain) estimates for the basin. Our observations contradict model diagnoses, revealing a substantial input of newly-fixed nitrogen to the tropical eastern basin and no dinitrogen fixation west of 7.5˚W. We propose that dinitrogen fixation in the South Atlantic occurs in hotspots controlled by the overlapping biogeography of excess phosphorus relative to nitrogen and bioavailable iron from margin sediments. Similar conditions may promote dinitrogen fixation in analogous ocean regions. Our analysis suggests that local iron availability causes the phosphorus-driven coupling of oceanic dinitrogen fixation to nitrogen loss to vary on a regional basis.This work was supported by the South African National Research Foundation (114673 and 130826 to T.M., 115335, 116142 and 129320 to S.E.F.); the US National Science Foundation (CAREER award, OCE-1554474 to J.G., OCE-1736652 to D.M.S. and K.L.C., OCE-05-26277 to K.L.C.); the German Federal Agency for Education and Research (DAAD-SPACES 57371082 to T.M.); the Royal Society (FLAIR fellowship to S.E.F.); and the University of Cape Town (T.M., J.G., S.E.F.). The authors also recognize the support of the South African Department of Science and Innovation’s Biogeochemistry Research Infrastructure Platform (BIOGRIP)

Enrichment and characterization of ammonia-oxidizing archaea from the open ocean : phylogeny, physiology and stable isotope fractionation

Author Posting. © The Author(s), 2011. This is the author's version of the work. It is posted here by permission of Nature Publishing Group for personal use, not for redistribution. The definitive version was published in The ISME Journal 5 (2011): 1796–1808, doi:10.1038/ismej.2011.58.Archaeal genes for ammonia oxidation are widespread in the marine environment, but direct physiological evidence for ammonia oxidation by marine archaea is limited. We report the enrichment and characterization of three strains of pelagic ammonia-oxidizing archaea (AOA) from the north Pacific Ocean that have been maintained in laboratory culture for over three years. Phylogenetic analyses indicate the three strains belong to a previously identified clade of water column-associated AOA and possess 16S rRNA genes and ammonia monooxygenase subunit a (amoA) genes highly similar (98-99% identity) to those recovered in DNA and cDNA clone libraries from the open ocean. The strains grow in natural seawater-based liquid medium while stoichiometrically converting ammonium (NH4 +) to nitrite (NO2 -). Ammonia oxidation by the enrichments is only partially inhibited by allylthiourea at concentrations known to inhibit cultivated ammonia-oxidizing bacteria. The three strains were used to determine the nitrogen stable isotope effect (15εNH3) during archaeal ammonia oxidation, an important parameter for interpreting stable isotope ratios in the environment. Archaeal 15εNH3 ranged from 13- 41‰, within the range of that previously reported for ammonia-oxidizing bacteria. Despite low amino acid identity between the archaeal and bacterial Amo proteins, their functional diversity as captured by 15εNH3 is similar.This work was supported by a Woods Hole Oceanographic Institution (WHOI) Postdoctoral Scholar fellowship to AES and the WHOI Ocean Life Institute