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The oceanic dissolved organic phosphorus (DOP) pool is mainly composed of P-esters and, to a lesser extent, equally abundant phosphonate and P-anhydride molecules. In phosphate-limited ocean regions, diazotrophs are thought to rely on DOP compounds as an alternative source of phosphorus (P). While both P-esters and phosphonates effectively promote dinitrogen (N2) fixation, the role of P-anhydrides for diazotrophs is unknown. Here we explore the effect of P-anhydrides on N2 fixation at two stations with contrasting biogeochemical conditions: one located in the Tonga trench volcanic arc region (“volcano,” with low phosphate and high iron concentrations), and the other in the South Pacific Gyre (“gyre,” with moderate phosphate and low iron). We incubated surface seawater with AMP (P-ester), ATP (P-ester and P-anhydride), or 3polyP (P-anhydride) and determined cell-specific N2 fixation rates, nifH gene abundance, and transcription in Crocosphaera and Trichodesmium. Trichodesmium did not respond to any DOP compounds added, suggesting that they were not P-limited at the volcano station and were outcompeted by the low iron conditions at the gyre station. Conversely, Crocosphaera were numerous at both stations and their specific N2 fixation rates were stimulated by AMP at the volcano station and slightly by 3polyP at both stations. Heterotrophic bacteria responded to ATP and 3polyP additions similarly at both stations, despite the contrasting phosphate and iron availability. The use of 3polyP by Crocosphaera and heterotrophic bacteria at both low and moderate phosphate concentrations suggests that this compound, in addition to being a source of P, can be used to acquire energy for which both groups compete. P-anhydrides may thus leverage energy restrictions to diazotrophs in the future stratified and nutrient-impoverished ocean.

© The Author(s), 2020. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Sutherland, K. M., Grabb, K. C., Karolewski, J. S., Plummer, S., Farfan, G. A., Wankel, S. D., Diaz, J. M., Lamborg, C. H., & Hansel, C. M. Spatial heterogeneity in particle-associated, light-independent superoxide production within productive coastal waters. Journal of Geophysical Research: Oceans, 125(10), (2020): e2020JC016747, https://doi.org/10.1029/2020JC016747., In the marine environment, the reactive oxygen species (ROS) superoxide is produced through a diverse array of light‐dependent and light‐independent reactions, the latter of which is thought to be primarily controlled by microorganisms. Marine superoxide production influences organic matter remineralization, metal redox cycling, and dissolved oxygen concentrations, yet the relative contributions of different sources to total superoxide production remain poorly constrained. Here we investigate the production, steady‐state concentration, and particle‐associated nature of light‐independent superoxide in productive waters off the northeast coast of North America. We find exceptionally high levels of light‐independent superoxide in the marine water column, with concentrations ranging from 10 pM to in excess of 2,000 pM. The highest superoxide concentrations were particle associated in surface seawater and in aphotic seawater collected meters off the seafloor. Filtration of seawater overlying the continental shelf lowered the light‐independent, steady‐state superoxide concentration by an average of 84%. We identify eukaryotic phytoplankton as the dominant particle‐associated source of superoxide to these coastal waters. We contrast these measurements with those collected at an off‐shelf station, where superoxide concentrations did not exceed 100 pM, and particles account for an average of 40% of the steady‐state superoxide concentration. This study demonstrates the primary role of particles in the production of superoxide in seawater overlying the continental shelf and highlights the importance of light‐independent, dissolved‐phase reactions in marine ROS production., This work was funded by grants from the Chemical Oceanography program of the National Science Foundation (OCE‐1355720 to C. M. H. and C. H. L.), NASA Earth and Space Science Fellowship (Grant NNX15AR62H to K. M. S.), Agouron Institute Postdoctoral Fellowship (K. M. S.), NSF GRFPs (2016230268 to K. C. G. and 2017250547 to S. P.), and a Sloan Research Fellowship (J. M. D.). The Guava flow cytometer was purchased through an NSF equipment improvement grant (1624593).

© The Author(s), 2019. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Diaz, J. M., Plummer, S., Hansel, C. M., Andeer, P. F., Saito, M. A., & McIlvin, M. R. NADPH-dependent extracellular superoxide production is vital to photophysiology in the marine diatom Thalassiosira oceanica. Proceedings of the National Academy of Sciences of the United States of America, 116 (33), (2019): 16448-16453, doi: 10.1073/pnas.1821233116., Reactive oxygen species (ROS) like superoxide drive rapid transformations of carbon and metals in aquatic systems and play dynamic roles in biological health, signaling, and defense across a diversity of cell types. In phytoplankton, however, the ecophysiological role(s) of extracellular superoxide production has remained elusive. Here, the mechanism and function of extracellular superoxide production by the marine diatom Thalassiosira oceanica are described. Extracellular superoxide production in T. oceanica exudates was coupled to the oxidation of NADPH. A putative NADPH-oxidizing flavoenzyme with predicted transmembrane domains and high sequence similarity to glutathione reductase (GR) was implicated in this process. GR was also linked to extracellular superoxide production by whole cells via quenching by the flavoenzyme inhibitor diphenylene iodonium (DPI) and oxidized glutathione, the preferred electron acceptor of GR. Extracellular superoxide production followed a typical photosynthesis-irradiance curve and increased by 30% above the saturation irradiance of photosynthesis, while DPI significantly impaired the efficiency of photosystem II under a wide range of light levels. Together, these results suggest that extracellular superoxide production is a byproduct of a transplasma membrane electron transport system that serves to balance the cellular redox state through the recycling of photosynthetic NADPH. This photoprotective function may be widespread, consistent with the presence of putative homologs to T. oceanica GR in other representative marine phytoplankton and ocean metagenomes. Given predicted climate-driven shifts in global surface ocean light regimes and phytoplankton community-level photoacclimation, these results provide implications for future ocean redox balance, ecological functioning, and coupled biogeochemical transformations of carbon and metals., This work was supported by a postdoctoral fellowship from the Ford Foundation (to J.M.D.), the National Science Foundation (NSF) under grants OCE 1225801 (to J.M.D.) and OCE 1246174 (to C.M.H.), a Junior Faculty Seed Grant from the University of Georgia Research Foundation (to J.M.D.), and a National Science Foundation Graduate Research Fellowship (to S.P.). The FIRe was purchased through a NSF equipment improvement grant (1624593).The authors thank Melissa Soule for assistance with LC/MS/MS analysis of peptide samples.

© The Author(s), 2019. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Plummeer, S., Taylor, A. E., Harvey, E. L., Hansel, C. M., & Diaz, J. M. Dynamic regulation of extracellular superoxide production by the coccolithophore Emiliania huxleyi (CCMP 374). Frontiers in Microbiology, 10, (2019): 1546, doi: 10.3389/fmicb.2019.01546., In marine waters, ubiquitous reactive oxygen species (ROS) drive biogeochemical cycling of metals and carbon. Marine phytoplankton produce the ROS superoxide (O2−) extracellularly and can be a dominant source of O2− in natural aquatic systems. However, the cellular regulation, biological functioning, and broader ecological impacts of extracellular O2− production by marine phytoplankton remain mysterious. Here, we explored the regulation and potential roles of extracellular O2− production by a noncalcifying strain of the cosmopolitan coccolithophorid Emiliania huxleyi, a key species of marine phytoplankton that has not been examined for extracellular O2− production previously. Cell-normalized extracellular O2− production was the highest under presumably low-stress conditions during active proliferation and inversely related to cell density during exponential growth phase. Removal of extracellular O2− through addition of the O2− scavenger superoxide dismutase (SOD), however, increased growth rates, growth yields, cell biovolume, and photosynthetic efficiency (Fv/Fm) indicating an overall physiological improvement. Thus, the presence of extracellular O2− does not directly stimulate E. huxleyi proliferation, as previously suggested for other phytoplankton, bacteria, fungi, and protists. Extracellular O2− production decreased in the dark, suggesting a connection with photosynthetic processes. Taken together, the tight regulation of this stress independent production of extracellular O2− by E. huxleyi suggests that it could be involved in fundamental photophysiological processes., This research was supported by a Junior Faculty Seed Grant from the University of Georgia Research Foundation (JD), a National Science Foundation (NSF) Graduate Research Fellowship (SP), and NSF grant OCE-1355720 (CH). The FlowCam® and FIRe were purchased through a NSF Equipment Improvement Grant (1624593).

© The Author(s), 2019. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Hansel, C. M., Diaz, J. M., & Plummer, S.. Tight regulation of extracellular superoxide points to its vital role in the physiology of the globally relevant Roseobacter clade. Mbio, 10(2), (2019):e02668-18, doi:10.1128/mBio.02668-18., There is a growing appreciation within animal and plant physiology that the reactive oxygen species (ROS) superoxide is not only detrimental but also essential for life. Yet, despite widespread production of extracellular superoxide by healthy bacteria and phytoplankton, this molecule remains associated with stress and death. Here, we quantify extracellular superoxide production by seven ecologically diverse bacteria within the Roseobacter clade and specifically target the link between extracellular superoxide and physiology for two species. We reveal for all species a strong inverse relationship between cell-normalized superoxide production rates and cell number. For exponentially growing cells of Ruegeria pomeroyi DSS-3 and Roseobacter sp. strain AzwK-3b, we show that superoxide levels are regulated in response to cell density through rapid modulation of gross production and not decay. Over a life cycle of batch cultures, extracellular superoxide levels are tightly regulated through a balance of both production and decay processes allowing for nearly constant levels of superoxide during active growth and minimal levels upon entering stationary phase. Further, removal of superoxide through the addition of exogenous superoxide dismutase during growth leads to significant growth inhibition. Overall, these results point to tight regulation of extracellular superoxide in representative members of the Roseobacter clade, consistent with a role for superoxide in growth regulation as widely acknowledged in fungal, animal, and plant physiology., We thank Mary Ann Moran and Alison Buchan for providing Roseobacter cultures, Kevin Sutherland for providing helpful feedback on the manuscript, and Elizabeth Harvey for use of her flow cytometer. This research was supported by NSF OCE-1355720 and a WHOI Independent Study Award (27005303) to C.M.H., as well as a Junior Faculty Seed Grant from the University of Georgia Research Foundation to J.M.D. and a National Science Foundation Graduate Research Fellowship to S.P.

© The Author(s), 2016. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Frontiers in Marine Science 3 (2016): 232, doi:10.3389/fmars.2016.00232., The reactive oxygen species (ROS) superoxide has been implicated in both beneficial and detrimental processes in coral biology, ranging from pathogenic disease resistance to coral bleaching. Despite the critical role of ROS in coral health, there is a distinct lack of ROS measurements and thus an incomplete understanding of underpinning ROS sources and production mechanisms within coral systems. Here, we quantified in situ extracellular superoxide concentrations at the surfaces of aquaria-hosted Porites astreoides during a diel cycle. High concentrations of superoxide (~10's of nM) were present at coral surfaces, and these levels did not change significantly as a function of time of day. These results indicate that the coral holobiont produces extracellular superoxide in the dark, independent of photosynthesis. As a short-lived anion at physiological pH, superoxide has a limited ability to cross intact biological membranes. Further, removing surface mucus layers from the P. astreoides colonies did not impact external superoxide concentrations. We therefore attribute external superoxide derived from the coral holobiont under these conditions to the activity of the coral host epithelium, rather than mucus-derived epibionts or internal sources such as endosymbionts (e.g., Symbiodinium). However, endosymbionts likely contribute to internal ROS levels via extracellular superoxide production. Indeed, common coral symbionts, including multiple strains of Symbiodinium (clades A to D) and the bacterium Endozoicomonas montiporae LMG 24815, produced extracellular superoxide in the dark and at low light levels. Further, representative P. astreoides symbionts, Symbiodinium CCMP2456 (clade A) and E. montiporae, produced similar concentrations of superoxide alone and in combination with each other, in the dark and low light, and regardless of time of day. Overall, these results indicate that healthy, non-stressed P. astreoides and representative symbionts produce superoxide extern, This work was supported by a Postdoctoral Fellowship from the Ford Foundation (JD), the National Science Foundation under grants OCE 1225801 (JD) and OCE 1233612 (AA), the Ocean and Climate Change Institute of the Woods Hole Oceanographic Institution (CH), a BIOS Grant in aid award (SM), the Sidney Stern Memorial Trust (CH and AA), as well as an anonymous donor.

© The Author(s), 2016. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Nature Communications 7 (2016): 13801, doi:10.1038/ncomms13801., The reactive oxygen species superoxide (O2·−) is both beneficial and detrimental to life. Within corals, superoxide may contribute to pathogen resistance but also bleaching, the loss of essential algal symbionts. Yet, the role of superoxide in coral health and physiology is not completely understood owing to a lack of direct in situ observations. By conducting field measurements of superoxide produced by corals during a bleaching event, we show substantial species-specific variation in external superoxide levels, which reflect the balance of production and degradation processes. Extracellular superoxide concentrations are independent of light, algal symbiont abundance and bleaching status, but depend on coral species and bacterial community composition. Furthermore, coral-derived superoxide concentrations ranged from levels below bulk seawater up to ∼120 nM, some of the highest superoxide concentrations observed in marine systems. Overall, these results unveil the ability of corals and/or their microbiomes to regulate superoxide in their immediate surroundings, which suggests species-specific roles of superoxide in coral health and physiology., This work was supported by a Postdoctoral Fellowship from the Ford Foundation (J.M.D.), the National Science Foundation under grants OCE 1225801 (J.M.D.) and OCE 1233612 (A.A.), the Ocean and Climate Change Institute of the Woods Hole Oceanographic Institution (C.M.H.), the Sidney Stern Memorial Trust (C.M.H. and A.A.) and an anonymous donor.

Author Posting. © Association for the Sciences of Limnology and Oceanography, 2016. 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 61 (2016): 1188–1200, doi:10.1002/lno.10266., Reactive oxygen species (ROS) are key players in the health and biogeochemistry of the ocean and its inhabitants. The vital contribution of microorganisms to marine ROS levels, particularly superoxide, has only recently come to light, and thus the specific biological sources and pathways involved in ROS production are largely unknown. To better understand the biogenic controls on ROS levels in tropical oligotrophic systems, we determined rates of superoxide production under various conditions by natural populations of the nitrogen-fixing diazotroph Trichodesmium obtained from various surface waters in the Sargasso Sea. Trichodesmium colonies collected from eight different stations all produced extracellular superoxide at high rates in both the dark and light. Colony density and light had a variable impact on extracellular superoxide production depending on the morphology of the Trichodesmium colonies. Raft morphotypes showed a rapid increase in superoxide production in response to even low levels of light, which was not observed for puff colonies. In contrast, superoxide production rates per colony decreased with increasing colony density for puff morphotypes but not for rafts. These findings point to Trichodesmium as a likely key source of ROS to the surface oligotrophic ocean. The physiological and/or ecological factors underpinning morphology-dependent controls on superoxide production need to be unveiled to better understand and predict superoxide production by Trichodesmium and ROS dynamics within marine systems., Major support for this work was provided by NSF OCE- 1246174 to CMH, NSF OCE-1332912 to STD and NSF OCE-13329898 to BASVM.

Biological productivity in many ocean regions is controlled by the availability of the nutrient phosphorus. In the Mediterranean Sea, aerosol deposition is a key source of phosphorus and understanding its composition is critical for determining its potential bioavailability. Aerosol phosphorus was investigated in European and North African air masses using phosphorus near-edge X-ray fluorescence spectroscopy (P-NEXFS). These air masses are the main source of aerosol deposition to the Mediterranean Sea. We show that European aerosols are a significant source of soluble phosphorus to the Mediterranean Sea. European aerosols deliver on average 3.5 times more soluble phosphorus than North African aerosols and furthermore are dominated by organic phosphorus compounds. The ultimate source of organic phosphorus does not stem from common primary emission sources. Rather, phosphorus associated with bacteria best explains the presence of organic phosphorus in Mediterranean aerosols.