Glacial meltwater determines the balance between autotrophic and heterotrophic processes in a Greenland fjord.

Global warming accelerates melting of glaciers and increases the supply of meltwater and associated inorganic particles, nutrients, and organic matter to adjacent coastal seas, but the ecosystem impact is poorly resolved and quantified. When meltwater is delivered by glacial rivers, the potential impact could be a reduction in light and nutrient availability for primary producers while supplying allochthonous carbon for heterotrophic processes, thereby tipping the net community metabolism toward heterotrophy. To test this hypothesis, we determined physical and biogeochemical parameters along a 110-km fjord transect in NE Greenland fjord, impacted by glacial meltwater from the Greenland Ice Sheet. The meltwater is delivered from glacier-fed river outlets in the inner parts of the fjord, creating a gradient in salinity and turbidity. The planktonic primary production was low, 20-45 mg C m-2 d-1, in the more turbid inner half of the fjord, increasing 10-fold to around 350 mg C m-2 d-1 in the shelf waters outside the fjord. Plankton community metabolism was measured at three stations, which displayed a transition from net heterotrophy in the inner fjord to net autotrophy in the coastal shelf waters. Respiration was significantly correlated to turbidity, with a 10-fold increase in the inner turbid part of the fjord. We estimated the changes in meltwater input and sea ice coverage in the area for the last 60 y. The long-term trend and the observed effects demonstrated the importance of freshwater runoff as a key driver of coastal ecosystem change in the Arctic with potential negative consequences for coastal productivity.

Continuous daylight in the high-Arctic summer supports high plankton respiration rates compared to those supported in the dark.

Plankton respiration rate is a major component of global CO2 production and is forecasted to increase rapidly in the Arctic with warming. Yet, existing assessments in the Arctic evaluated plankton respiration in the dark. Evidence that plankton respiration may be stimulated in the light is particularly relevant for the high Arctic where plankton communities experience continuous daylight in spring and summer. Here we demonstrate that plankton community respiration evaluated under the continuous daylight conditions present in situ, tends to be higher than that evaluated in the dark. The ratio between community respiration measured in the light (Rlight) and in the dark (Rdark) increased as the 2/3 power of Rlight so that the Rlight:Rdark ratio increased from an average value of 1.37 at the median Rlight measured here (3.62 µmol O2 L-1 d-1) to an average value of 17.56 at the highest Rlight measured here (15.8 µmol O2 L-1 d-1). The role of respiratory processes as a source of CO2 in the Arctic has, therefore, been underestimated and is far more important than previously believed, particularly in the late spring, with 24 h photoperiods, when community respiration rates are highest.

Sea ice and primary production proxies in surface sediments from a High Arctic Greenland fjord: Spatial distribution and implications for palaeoenvironmental studies.

In order to establish a baseline for proxy-based reconstructions for the Young Sound-Tyrolerfjord system (Northeast Greenland), we analysed the spatial distribution of primary production and sea ice proxies in surface sediments from the fjord, against monitoring data from the Greenland Ecosystem Monitoring Programme. Clear spatial gradients in organic carbon and biogenic silica contents reflected marine influence, nutrient availability and river-induced turbidity, in good agreement with in situ measurements. The sea ice proxy IP25 was detected at all sites but at low concentrations, indicating that IP25 records from fjords need to be carefully considered and not directly compared to marine settings. The sea ice-associated biomarker HBI III revealed an open-water signature, with highest concentrations near the mid-July ice edge. This proxy evaluation is an important step towards reliable palaeoenvironmental reconstructions that will, ultimately, contribute to better predictions for this High Arctic ecosystem in a warming climate.

Ammonium and nitrite oxidation at nanomolar oxygen concentrations in oxygen minimum zone waters.

A major percentage of fixed nitrogen (N) loss in the oceans occurs within nitrite-rich oxygen minimum zones (OMZs) via denitrification and anammox. It remains unclear to what extent ammonium and nitrite oxidation co-occur, either supplying or competing for substrates involved in nitrogen loss in the OMZ core. Assessment of the oxygen (O2) sensitivity of these processes down to the O2 concentrations present in the OMZ core (<10 nmol⋅L(-1)) is therefore essential for understanding and modeling nitrogen loss in OMZs. We determined rates of ammonium and nitrite oxidation in the seasonal OMZ off Concepcion, Chile at manipulated O2 levels between 5 nmol⋅L(-1) and 20 µmol⋅L(-1) Rates of both processes were detectable in the low nanomolar range (5-33 nmol⋅L(-1) O2), but demonstrated a strong dependence on O2 concentrations with apparent half-saturation constants (Kms) of 333 ± 130 nmol⋅L(-1) O2 for ammonium oxidation and 778 ± 168 nmol⋅L(-1) O2 for nitrite oxidation assuming one-component Michaelis-Menten kinetics. Nitrite oxidation rates, however, were better described with a two-component Michaelis-Menten model, indicating a high-affinity component with a Km of just a few nanomolar. As the communities of ammonium and nitrite oxidizers were similar to other OMZs, these kinetics should apply across OMZ systems. The high O2 affinities imply that ammonium and nitrite oxidation can occur within the OMZ core whenever O2 is supplied, for example, by episodic intrusions. These processes therefore compete with anammox and denitrification for ammonium and nitrite, thereby exerting an important control over nitrogen loss.

Phytoplankton Productivity in an Arctic Fjord (West Greenland): Estimating Electron Requirements for Carbon Fixation and Oxygen Production.

Accurate quantification of pelagic primary production is essential for quantifying the marine carbon turnover and the energy supply to the food web. Knowing the electron requirement (Κ) for carbon (C) fixation (ΚC) and oxygen (O2) production (ΚO2), variable fluorescence has the potential to quantify primary production in microalgae, and hereby increasing spatial and temporal resolution of measurements compared to traditional methods. Here we quantify ΚC and ΚO2 through measures of Pulse Amplitude Modulated (PAM) fluorometry, C fixation and O2 production in an Arctic fjord (Godthåbsfjorden, W Greenland). Through short- (2h) and long-term (24h) experiments, rates of electron transfer (ETRPSII), C fixation and/or O2 production were quantified and compared. Absolute rates of ETR were derived by accounting for Photosystem II light absorption and spectral light composition. Two-hour incubations revealed a linear relationship between ETRPSII and gross 14C fixation (R2 = 0.81) during light-limited photosynthesis, giving a ΚC of 7.6 ± 0.6 (mean ± S.E.) mol é (mol C)-1. Diel net rates also demonstrated a linear relationship between ETRPSII and C fixation giving a ΚC of 11.2 ± 1.3 mol é (mol C)-1 (R2 = 0.86). For net O2 production the electron requirement was lower than for net C fixation giving 6.5 ± 0.9 mol é (mol O2)-1 (R2 = 0.94). This, however, still is an electron requirement 1.6 times higher than the theoretical minimum for O2 production [i.e. 4 mol é (mol O2)-1]. The discrepancy is explained by respiratory activity and non-photochemical electron requirements and the variability is discussed. In conclusion, the bio-optical method and derived electron requirement support conversion of ETR to units of C or O2, paving the road for improved spatial and temporal resolution of primary production estimates.

Oxygen at nanomolar levels reversibly suppresses process rates and gene expression in anammox and denitrification in the oxygen minimum zone off northern Chile.

UNLABELLED: A major percentage (20 to 40%) of global marine fixed-nitrogen loss occurs in oxygen minimum zones (OMZs). Concentrations of O2 and the sensitivity of the anaerobic N2-producing processes of anammox and denitrification determine where this loss occurs. We studied experimentally how O2 at nanomolar levels affects anammox and denitrification rates and the transcription of nitrogen cycle genes in the anoxic OMZ off Chile. Rates of anammox and denitrification were reversibly suppressed, most likely at the enzyme level. Fifty percent inhibition of N2 and N2O production by denitrification was achieved at 205 and 297 nM O2, respectively, whereas anammox was 50% inhibited at 886 nM O2. Coupled metatranscriptomic analysis revealed that transcripts encoding nitrous oxide reductase (nosZ), nitrite reductase (nirS), and nitric oxide reductase (norB) decreased in relative abundance above 200 nM O2. This O2 concentration did not suppress the transcription of other dissimilatory nitrogen cycle genes, including nitrate reductase (narG), hydrazine oxidoreductase (hzo), and nitrite reductase (nirK). However, taxonomic characterization of transcripts suggested inhibition of narG transcription in gammaproteobacteria, whereas the transcription of anammox narG, whose gene product is likely used to oxidatively replenish electrons for carbon fixation, was not inhibited. The taxonomic composition of transcripts differed among denitrification enzymes, suggesting that distinct groups of microorganisms mediate different steps of denitrification. Sulfide addition (1 µM) did not affect anammox or O2 inhibition kinetics but strongly stimulated N2O production by denitrification. These results identify new O2 thresholds for delimiting marine nitrogen loss and highlight the utility of integrating biogeochemical and metatranscriptomic analyses. IMPORTANCE: The removal of fixed nitrogen via anammox and denitrification associated with low O2 concentrations in oceanic oxygen minimum zones (OMZ) is a major sink in oceanic N budgets, yet the sensitivity and dynamics of these processes with respect to O2 are poorly known. The present study elucidated how nanomolar O2 concentrations affected nitrogen removal rates and expression of key nitrogen cycle genes in water from the eastern South Pacific OMZ, applying state-of-the-art (15)N techniques and metatranscriptomics. Rates of both denitrification and anammox responded rapidly and reversibly to changes in O2, but denitrification was more O2 sensitive than anammox. The transcription of key nitrogen cycle genes did not respond as clearly to O2, although expression of some of these genes decreased. Quantifying O2 sensitivity of these processes is essential for predicting through which pathways and in which environments, from wastewater treatment to the open oceans, nitrogen removal may occur.

Vertical partitioning of nitrogen-loss processes across the oxic-anoxic interface of an oceanic oxygen minimum zone.

We investigated anammox, denitrification and dissimilatory reduction of nitrite to ammonium (DNRA) activity in the Eastern Tropical South Pacific oxygen minimum zone (OMZ) off northern Chile, at high-depth resolution through the oxycline into the anoxic OMZ core. This was accompanied by high-resolution nutrient and oxygen profiles to link changes in nitrogen transformation rates to physicochemical characteristics of the water column. Denitrification was detected at most depths, but anammox was the most active N2 -producing process, while DNRA was not detectable. Anammox and denitrification were mainly active in the anoxic OMZ core while activity was low to not detectable in the oxycline, except in association with an intrusion of OMZ core water. This indicates that continuous exposure to even submicromolar oxygen levels inhibits the processes either directly or through nitrite limitation. Anammox activity did not peak at the oxic-anoxic boundary but 20-50 m below matching the salinity maximum of the Equatorial Subsurface Water. This suggests that water history plays a major role for anammox activity possibly due to slow growth of anammox bacteria. Denitrification peaked deeper than anammox, likely reflecting a shift in the balance between this process and nitrate reduction to nitrite, governed by the relative availability of nitrate and nitrite.

Experimental incubations elicit profound changes in community transcription in OMZ bacterioplankton.

Sequencing of microbial community RNA (metatranscriptome) is a useful approach for assessing gene expression in microorganisms from the natural environment. This method has revealed transcriptional patterns in situ, but can also be used to detect transcriptional cascades in microcosms following experimental perturbation. Unambiguously identifying differential transcription between control and experimental treatments requires constraining effects that are simply due to sampling and bottle enclosure. These effects remain largely uncharacterized for "challenging" microbial samples, such as those from anoxic regions that require special handling to maintain in situ conditions. Here, we demonstrate substantial changes in microbial transcription induced by sample collection and incubation in experimental bioreactors. Microbial communities were sampled from the water column of a marine oxygen minimum zone by a pump system that introduced minimal oxygen contamination and subsequently incubated in bioreactors under near in situ oxygen and temperature conditions. Relative to the source water, experimental samples became dominated by transcripts suggestive of cell stress, including chaperone, protease, and RNA degradation genes from diverse taxa, with strong representation from SAR11-like alphaproteobacteria. In tandem, transcripts matching facultative anaerobic gammaproteobacteria of the Alteromonadales (e.g., Colwellia) increased 4-13 fold up to 43% of coding transcripts, and encoded a diverse gene set suggestive of protein synthesis and cell growth. We interpret these patterns as taxon-specific responses to combined environmental changes in the bioreactors, including shifts in substrate or oxygen availability, and minor temperature and pressure changes during sampling with the pump system. Whether such changes confound analysis of transcriptional patterns may vary based on the design of the experiment, the taxonomic composition of the source community, and on the metabolic linkages between community members. These data highlight the impressive capacity for transcriptional changes within complex microbial communities, underscoring the need for caution when inferring in situ metabolism based on transcript abundances in experimental incubations.

Construction of STOX oxygen sensors and their application for determination of O2 concentrations in oxygen minimum zones.

Until recently, it has not been possible to measure O(2) concentrations in oxygen minimum zones (OMZs) with sufficient detection limits and accuracy to determine whether OMZs are anoxic or contain 1-2 µM O(2). With the introduction of the STOX (switchable trace oxygen) sensor, the level for accurate quantification has been lowered by a factor of 1000. By analysis with STOX sensors, O(2) can be prevented from reaching the sensing cathode by another cathode (front guard cathode), and it is the amplitude in signal by polarization/depolarization of this front guard that is used as a measure of the O(2) concentration. The STOX sensors can be used in situ, most conveniently connected to a conventional CTD (conductivity, temperature, and depth analyzer) along with a conventional oxygen sensor, and they can be used for monitoring O(2) dynamics during laboratory incubations of low-O(2) media such as OMZ water. The limiting factors for use of the STOX sensors are a relatively slow response, with measuring cycle of at least 30 s with the current design, and fragility. With improved procedures for construction, the time for a complete measuring cycle is expected to come down to about 10 s.

A cryptic sulfur cycle in oxygen-minimum-zone waters off the Chilean coast.

Nitrogen cycling is normally thought to dominate the biogeochemistry and microbial ecology of oxygen-minimum zones in marine environments. Through a combination of molecular techniques and process rate measurements, we showed that both sulfate reduction and sulfide oxidation contribute to energy flux and elemental cycling in oxygen-free waters off the coast of northern Chile. These processes may have been overlooked because in nature, the sulfide produced by sulfate reduction immediately oxidizes back to sulfate. This cryptic sulfur cycle is linked to anammox and other nitrogen cycling processes, suggesting that it may influence biogeochemical cycling in the global ocean.

Effects of fish farm waste on Posidonia oceanica meadows: synthesis and provision of monitoring and management tools.

This paper provides a synthesis of the EU project MedVeg addressing the fate of nutrients released from fish farming in the Mediterranean with particular focus on the endemic seagrass Posidonia oceanica habitat. The objectives were to identify the main drivers of seagrass decline linked to fish farming and to provide sensitive indicators of environmental change, which can be used for monitoring purposes. The sedimentation of waste particles in the farm vicinities emerges as the main driver of benthic deterioration, such as accumulation of organic matter, sediment anoxia as well as seagrass decline. The effects of fish farming on P. oceanica meadows are diverse and complex and detected through various metrics and indicators. A safety distance of 400 m is suggested for management of P. oceanica near fish farms followed by establishment of permanent seagrass plots revisited annually for monitoring the health of the meadows.

Anaerobic ammonium-oxidizing bacteria in marine environments: widespread occurrence but low diversity.

Laboratory and field studies have indicated that anaerobic ammonium oxidation (anammox) is an important process in the marine nitrogen cycle. In this study 11 additional anoxic marine sediment and water column samples were studied to substantiate this claim. In a combined approach using the molecular methods, polymerase chain reaction (PCR), qualitative and quantitative fluorescence in situ hybridization (FISH), as well as (15)N stable isotope activity measurements, it was shown that anammox bacteria were present and active in all samples investigated. The anammox activity measured in the sediment samples ranged from 0.08 fmol cell(-1) day(-1) N(2) in the Golfo Dulce (Pacific Ocean, Costa Rica) sediment to 0.98 fmol cell(-1) day(-1) N(2) in the Gullmarsfjorden (North Sea, Sweden) sediment. The percentage of anammox cell of the total population (stained with DAPI) as assessed by quantitative FISH was highest in the Barents Sea (9% +/- 4%) and in most of the samples well over 2%. Fluorescence in situ hybridization and phylogenetic analysis of the PCR products derived from the marine samples indicated the exclusive presence of members of the Candidatus'Scalindua' genus. This study showed the ubiquitous presence of anammox bacteria in anoxic marine ecosystems, supporting previous observations on the importance of anammox for N cycling in marine environments.

Effects of specific inhibitors on anammox and denitrification in marine sediments.

The effects of three metabolic inhibitors (acetylene, methanol, and allylthiourea [ATU]) on the pathways of N2 production were investigated by using short anoxic incubations of marine sediment with a 15N isotope technique. Acetylene inhibited ammonium oxidation through the anammox pathway as the oxidation rate decreased exponentially with increasing acetylene concentration; the rate decay constant was 0.10+/-0.02 microM-1, and there was 95% inhibition at approximately 30 microM. Nitrous oxide reduction, the final step of denitrification, was not sensitive to acetylene concentrations below 10 microM. However, nitrous oxide reduction was inhibited by higher concentrations, and the sensitivity was approximately one-half the sensitivity of anammox (decay constant, 0.049+/-0.004 microM-1; 95% inhibition at approximately 70 microM). Methanol specifically inhibited anammox with a decay constant of 0.79+/-0.12 mM-1, and thus 3 to 4 mM methanol was required for nearly complete inhibition. This level of methanol stimulated denitrification by approximately 50%. ATU did not have marked effects on the rates of anammox and denitrification. The profile of inhibitor effects on anammox agreed with the results of studies of the process in wastewater bioreactors, which confirmed the similarity between the anammox bacteria in bioreactors and natural environments. Acetylene and methanol can be used to separate anammox and denitrification, but the effects of these compounds on nitrification limits their use in studies of these processes in systems where nitrification is an important source of nitrate. The observed differential effects of acetylene and methanol on anammox and denitrification support our current understanding of the two main pathways of N2 production in marine sediments and the use of 15N isotope methods for their quantification.

Anaerobic ammonium oxidation (anammox) in the marine environment.

Anammox, anaerobic ammonium oxidation with nitrite, is now recognized as an important process in the marine nitrogen cycle. The bacteria conducting anammox are highly specialized and appear to belong to the Planctomycetales. The process has now been found in a range of environments including marine sediments, sea ice and anoxic water columns, and it may be responsible for up to 50% of the global removal of fixed nitrogen from the oceans.

N2 production by the anammox reaction in the anoxic water column of Golfo Dulce, Costa Rica.

In oxygen-depleted zones of the open ocean, and in anoxic basins and fjords, denitrification (the bacterial reduction of nitrate to give N2) is recognized as the only significant process converting fixed nitrogen to gaseous N2. Primary production in the oceans is often limited by the availability of fixed nitrogen such as ammonium or nitrate, and nitrogen-removal processes consequently affect both ecosystem function and global biogeochemical cycles. It was recently discovered that the anaerobic oxidation of ammonium with nitrite--the 'anammox' reaction, performed by bacteria--was responsible for a significant fraction of N2 production in some marine sediments. Here we show that this reaction is also important in the anoxic waters of Golfo Dulce, a 200-m-deep coastal bay in Costa Rica, where it accounts for 19-35% of the total N2 formation in the water column. The water-column chemistry in Golfo Dulce is very similar to that in oxygen-depleted zones of the oceans--in which one-half to one-third of the global nitrogen removal is believed to occur. We therefore expect the anammox reaction to be a globally significant sink for oceanic nitrogen.

Factors controlling anaerobic ammonium oxidation with nitrite in marine sediments.

Factors controlling the anaerobic oxidation of ammonium with nitrate and nitrite were explored in a marine sediment from the Skagerrak in the Baltic-North Sea transition. In anoxic incubations with the addition of nitrite, approximately 65% of the nitrogen gas formation was due to anaerobic ammonium oxidation with nitrite, with the remainder being produced by denitrification. Anaerobic ammonium oxidation with nitrite exhibited a biological temperature response, with a rate optimum at 15 degrees C and a maximum temperature of 37 degrees C. The biological nature of the process and a 1:1 stoichiometry for the reaction between nitrite and ammonium indicated that the transformations might be attributed to the anammox process. Attempts to find other anaerobic ammonium-oxidizing processes in this sediment failed. The apparent K(m) of nitrite consumption was less than 3 microM, and the relative importance of ammonium oxidation with nitrite and denitrification for the production of nitrogen gas was independent of nitrite concentration. Thus, the quantitative importance of ammonium oxidation with nitrite in the jar incubations at elevated nitrite concentrations probably represents the in situ situation. With the addition of nitrate, the production of nitrite from nitrate was four times faster than its consumption and therefore did not limit the rate of ammonium oxidation. Accordingly, the rate of this process was the same whether nitrate or nitrite was added as electron acceptor. The addition of organic matter did not stimulate denitrification, possibly because it was outcompeted by manganese reduction or because transport limitation was removed due to homogenization of the sediment.

Production of N(2) through anaerobic ammonium oxidation coupled to nitrate reduction in marine sediments.

In the global nitrogen cycle, bacterial denitrification is recognized as the only quantitatively important process that converts fixed nitrogen to atmospheric nitrogen gas, N(2), thereby influencing many aspects of ecosystem function and global biogeochemistry. However, we have found that a process novel to the marine nitrogen cycle, anaerobic oxidation of ammonium coupled to nitrate reduction, contributes substantially to N(2) production in marine sediments. Incubations with (15)N-labeled nitrate or ammonium demonstrated that during this process, N(2) is formed through one-to-one pairing of nitrogen from nitrate and ammonium, which clearly separates the process from denitrification. Nitrite, which accumulated transiently, was likely the oxidant for ammonium, and the process is thus similar to the anammox process known from wastewater bioreactors. Anaerobic ammonium oxidation accounted for 24 and 67% of the total N(2) production at two typical continental shelf sites, whereas it was detectable but insignificant relative to denitrification in a eutrophic coastal bay. However, rates of anaerobic ammonium oxidation were higher in the coastal sediment than at the deepest site and the variability in the relative contribution to N(2) production between sites was related to large differences in rates of denitrification. Thus, the relative importance of anaerobic ammonium oxidation and denitrification in N(2) production appears to be regulated by the availability of their reduced substrates. By shunting nitrogen directly from ammonium to N(2), anaerobic ammonium oxidation promotes the removal of fixed nitrogen in the oceans. The process can explain ammonium deficiencies in anoxic waters and sediments, and it may contribute significantly to oceanic nitrogen budgets.