Description: The extracellular concentration of H2O2 in surface aquatic environments is controlled by a balance between photochemical production and the microbial synthesis of catalase and peroxidase enzymes to remove H2O2 from solution. In any kind of incubation experiment, the formation rates and equilibrium concentrations of reactive oxygen species (ROSs) such as H2O2 may be sensitive to both the experiment design, particularly to the regulation of incident light, and the abundance of different microbial groups, as both cellular H2O2 production and catalase–peroxidase enzyme production rates differ between species. Whilst there are extensive measurements of photochemical H2O2 formation rates and the distribution of H2O2 in the marine environment, it is poorly constrained how different microbial groups affect extracellular H2O2 concentrations, how comparable extracellular H2O2 concentrations within large-scale incubation experiments are to those observed in the surface-mixed layer, and to what extent a mismatch with environmentally relevant concentrations of ROS in incubations could influence biological processes differently to what would be observed in nature. Here we show that both experiment design and bacterial abundance consistently exert control on extracellular H2O2 concentrations across a range of incubation experiments in diverse marine environments. During four large-scale (〉1000 L) mesocosm experiments (in Gran Canaria, the Mediterranean, Patagonia and Svalbard) most experimental factors appeared to exert only minor, or no, direct effect on H2O2 concentrations. For example, in three of four experiments where pH was manipulated to 0.4–0.5 below ambient pH, no significant change was evident in extracellular H2O2 concentrations relative to controls. An influence was sometimes inferred from zooplankton density, but not consistently between different incubation experiments, and no change in H2O2 was evident in controlled experiments using different densities of the copepod Calanus finmarchicus grazing on the diatom Skeletonema costatum (〈1 % change in [H2O2] comparing copepod densities from 1 to 10 L−1). Instead, the changes in H2O2 concentration contrasting high- and low-zooplankton incubations appeared to arise from the resulting changes in bacterial activity. The correlation between bacterial abundance and extracellular H2O2 was stronger in some incubations than others (R2 range 0.09 to 0.55), yet high bacterial densities were consistently associated with low H2O2. Nonetheless, the main control on H2O2 concentrations during incubation experiments relative to those in ambient, unenclosed waters was the regulation of incident light. In an open (lidless) mesocosm experiment in Gran Canaria, H2O2 was persistently elevated (2–6-fold) above ambient concentrations; whereas using closed high-density polyethylene mesocosms in Crete, Svalbard and Patagonia H2O2 within incubations was always reduced (median 10 %–90 %) relative to ambient waters.

Description: Eastern boundary upwelling systems (EBUS) are among the most productive marine ecosystems on Earth. The production of organic material is fueled by upwelling of nutrient-rich deep waters and high incident light at the sea surface. However, biotic and abiotic factors can modify surface production and related biogeochemical processes. Determining these factors is important because EBUS are considered hotspots of climate change, and reliable predictions of their future functioning requires understanding of the mechanisms driving the biogeochemical cycles therein. In this field experiment, we used in situ mesocosms as tools to improve our mechanistic understanding of processes controlling organic matter cycling in the coastal Peruvian upwelling system. Eight mesocosms, each with a volume of ∼55 m3, were deployed for 50 d ∼6 km off Callao (12∘ S) during austral summer 2017, coinciding with a coastal El Niño phase. After mesocosm deployment, we collected subsurface waters at two different locations in the regional oxygen minimum zone (OMZ) and injected these into four mesocosms (mixing ratio ≈1.5 : 1 mesocosm: OMZ water). The focus of this paper is on temporal developments of organic matter production, export, and stoichiometry in the individual mesocosms. The mesocosm phytoplankton communities were initially dominated by diatoms but shifted towards a pronounced dominance of the mixotrophic dinoflagellate (Akashiwo sanguinea) when inorganic nitrogen was exhausted in surface layers. The community shift coincided with a short-term increase in production during the A. sanguinea bloom, which left a pronounced imprint on organic matter C : N : P stoichiometry. However, C, N, and P export fluxes did not increase because A. sanguinea persisted in the water column and did not sink out during the experiment. Accordingly, export fluxes during the study were decoupled from surface production and sustained by the remaining plankton community. Overall, biogeochemical pools and fluxes were surprisingly constant for most of the experiment. We explain this constancy by light limitation through self-shading by phytoplankton and by inorganic nitrogen limitation which constrained phytoplankton growth. Thus, gain and loss processes remained balanced and there were few opportunities for blooms, which represents an event where the system becomes unbalanced. Overall, our mesocosm study revealed some key links between ecological and biogeochemical processes for one of the most economically important regions in the oceans.

Description: Upwelling of nutrient-rich waters into the sunlit surface layer of the ocean supports high primary productivity in eastern boundary upwelling systems (EBUSs). However, subsurface waters contain not only macronutrients (N, P, Si) but also micronutrients, organic matter and seed microbial communities that may modify the response to macronutrient inputs via upwelling. These additional factors are often neglected when investigating upwelling impacts on surface ocean productivity. Here, we investigated how different components of upwelled water (macronutrients, organic nutrients and seed communities) drive the response of surface plankton communities to upwelling in the Peruvian coastal zone. Results from our short-term (10 d) study show that the most influential drivers in upwelled deep water are (1) the ratio of inorganic nutrients (NOx : PO) and (2) the microbial community present that can seed heterogeneity in phytoplankton succession and modify the stoichiometry of residual inorganic nutrients after phytoplankton blooms. Hence, this study suggests that phytoplankton succession after upwelling is modified by factors other than the physical supply of inorganic nutrients. This would likely affect trophic transfer and overall productivity in these highly fertile marine ecosystems.

Description: Upwelling of nutrient-rich deep waters make eastern boundary upwelling systems (EBUSs), such as the Humboldt Current system, hot spots of marine productivity. Associated settling of organic matter to depth and consecutive aerobic decomposition results in large subsurface water volumes being oxygen depleted. Under these circumstances, organic matter remineralisation can continue via denitrification, which represents a major loss pathway for bioavailable nitrogen. Additionally, anaerobic ammonium oxidation can remove significant amounts of nitrogen in these areas. Here we assess the interplay of suboxic water upwelling and nitrogen cycling in a manipulative offshore mesocosm experiment. Measured denitrification rates in incubations with water from the oxygen-depleted bottom layer of the mesocosms (via 15N label incubations) mostly ranged between 5.5 and 20 nmol N2 L−1 h−1 (interquartile range), reaching up to 80 nmol N2 L−1 h−1. However, actual in situ rates in the mesocosms, estimated via Michaelis–Menten kinetic scaling, did most likely not exceed 0.2–4.2 nmol N2 L−1 h−1 (interquartile range) due to substrate limitation. In the surrounding Pacific, measured denitrification rates were similar, although indications of substrate limitation were detected only once. In contrast, anammox (anaerobic ammonium oxidation) made only a minor contribution to the overall nitrogen loss when encountered in both the mesocosms and the Pacific Ocean. This was potentially related to organic matter C / N stoichiometry and/or process-specific oxygen and hydrogen sulfide sensitivities. Over the first 38 d of the experiment, total nitrogen loss calculated from in situ rates of denitrification and anammox was comparable to estimates from a full nitrogen budget in the mesocosms and ranged between ∼ 1 and 5.5 µmol N L−1. This represents up to ∼  20 % of the initially bioavailable inorganic and organic nitrogen standing stocks. Interestingly, this loss is comparable to the total amount of particulate organic nitrogen that was exported into the sediment traps at the bottom of the mesocosms at about 20 m depth. Altogether, this suggests that a significant portion, if not the majority of nitrogen that could be exported to depth, is already lost, i.e. converted to N2 in a relatively shallow layer of the surface ocean, provided that there are oxygen-deficient conditions like those during coastal upwelling in our study. Published data for primary productivity and nitrogen loss in all EBUSs reinforce such conclusion.

Description: The Peruvian upwelling system is a highly productive ecosystem with a large oxygen minimum zone (OMZ) close to the surface. In this work, we carried out a mesocosm experiment off Callao, Peru, with the addition of water masses from the regional OMZ collected at two different sites simulating two different upwelling scenarios. Here, we focus on the pelagic remineralization of organic matter by the extracellular enzyme activity of leucine aminopeptidase (LAP) and alkaline phosphatase activity (APA). After the addition of the OMZ water, dissolved inorganic nitrogen (N) was depleted, but the standing stock of phytoplankton was relatively high, even after N depletion (mostly 〉 4 µg chlorophyll a L−1). During the initial phase of the experiment, APA was 0.6 nmol L−1 h−1 even though the PO concentration was 〉 0.5 µmol L−1. Initially, the dissolved organic phosphorus (DOP) decreased, coinciding with an increase in the PO concentration that was probably linked to the APA. The LAP activity was very high, with most of the measurements in the range of 200–800 nmol L−1 h−1. This enzyme hydrolyzes terminal amino acids from larger molecules (e.g., peptides or proteins), and these high values are probably linked to the highly productive but N-limited coastal ecosystem. Moreover, the experiment took place during a rare coastal El Niño event with higher than normal surface temperatures, which could have affected enzyme activity. Using a nonparametric multidimensional scaling analysis (NMDS) with a generalized additive model (GAM), we found that biogeochemical variables (e.g., nutrient and chlorophyll-a concentrations) and phytoplankton and bacterial communities explained up to 64 % of the variability in APA. The bacterial community best explained the variability (34 %) in LAP. The high hydrolysis rates for this enzyme suggest that pelagic N remineralization, likely driven by the bacterial community, supported the high standing stock of primary producers in the mesocosms after N depletion.

Description: According to modelling studies, ocean alkalinity enhancement (OAE) is one of the proposed carbon dioxide removal (CDR) approaches with large potential, with the beneficial side effect of counteracting ocean acidification. The real-world application of OAE, however, remains unclear as most basic assumptions are untested. Before large-scale deployment can be considered, safe and sustainable procedures for the addition of alkalinity to seawater must be identified and governance established. One of the concerns is the stability of alkalinity when added to seawater. The surface ocean is already supersaturated with respect to calcite and aragonite, and an increase in total alkalinity (TA) together with a corresponding shift in carbonate chemistry towards higher carbonate ion concentrations would result in a further increase in supersaturation, and potentially to solid carbonate precipitation. Precipitation of carbonate minerals consumes alkalinity and increases dissolved CO2 in seawater, thereby reducing the efficiency of OAE for CO2 removal. In order to address the application of alkaline solution as well as fine particulate alkaline solids, a set of six experiments was performed using natural seawater with alkalinity of around 2400 µmol kgsw−1. The application of CO2-equilibrated alkaline solution bears the lowest risk of losing alkalinity due to carbonate phase formation if added total alkalinity (ΔTA) is less than 2400 µmol kgsw−1. The addition of reactive alkaline solids can cause a net loss of alkalinity if added ΔTA 〉 600 µmol kgsw−1 (e.g. for Mg(OH)2). Commercially available (ultrafine) Ca(OH)2 causes, in general, a net loss in TA for the tested amounts of TA addition, which has consequences for suggested use of slurries with alkaline solids supplied from ships. The rapid application of excessive amounts of Ca(OH)2, exceeding a threshold for alkalinity loss, resulted in a massive increase in TA (〉 20 000 µmol kgsw−1) at the cost of lower efficiency and resultant high pH values 〉 9.5. Analysis of precipitates indicates formation of aragonite. However, unstable carbonate phases formed can partially redissolve, indicating that net loss of a fraction of alkalinity may not be permanent, which has important implications for real-world OAE application. Our results indicate that using an alkaline solution instead of reactive alkaline particles can avoid carbonate formation, unless alkalinity addition via solutions shifts the system beyond critical supersaturation levels. To avoid the loss of alkalinity and dissolved inorganic carbon (DIC) from seawater, the application of reactor techniques can be considered. These techniques produce an equilibrated solution from alkaline solids and CO2 prior to application. Differing behaviours of tested materials suggest that standardized engineered materials for OAE need to be developed to achieve safe and sustainable OAE with solids, if reactors technologies should be avoided.

Description: A natural plankton community from oligotrophic subtropical waters of the Atlantic near Gran Canaria, Spain, was subjected to varying degrees of ocean alkalinity enhancement (OAE) to assess the potential physiological effects, in the context of the application of ocean carbon dioxide removal (CDR) techniques. We employed 8.3 m3 mesocosms with a sediment trap attached to the bottom, creating a gradient in total alkalinity (TA). The lowest point on this gradient was 2400 μmol · L-1, which corresponded to the natural alkalinity of the environment, and the highest point was 4800 μmol · L-1. Over the course of the 33-day experiment, the plankton community exhibited two distinct phases. In phase-I (days 5–20), a notable decline in the photosynthetic efficiency (Fv/Fm) was observed. This change was accompanied by substantial reductions in the abundances of picoeukaryotes, small size nanoeukaryotes (nanoeukaryotes-1), and microplankton. The cell viability of picoeukaryotes, as indicated by fluorescein-di-acetate hydrolysis by cellular esterases (FDA- green fluorescence), slightly increased by the end of phase-I whilst the viability of nanoeukaryotes 1 and Synechococcus spp . did not change. Reactive oxygen species levels (ROS-green fluorescence) showed no significant changes for any of the functional groups. In contrast, in phase-II (days 21–33), a pronounced community response was observed. Increases in Fv/Fm in the intermediate OAE treatments of ∆900 to ∆1800 μmol · L-1 and in chlorophyll-a (Chl-a), chlorophyll-c2 (Chl-c2) , fucoxanthin and divinyl-Chl-a were attributed to the emergence of blooms of large size nanoeukaryotes (nanoeukaryotes-2) from the genera Chrysochromulina, as well as picoeukaryotes. Synechococcus spp. also flourished towards the end of this phase. In parallel, we observed a total 20 % significant change in the metaproteome of the phytoplankton community. This is considered a significant alteration in protein expression, having substantial impacts on cellular functions and the physiology of the organisms. Medium levels of ∆TA showed more upregulated and less downregulated proteins than higher ∆TA treatments. Under these conditions, cell viability significantly increased in pico and nanoeukaryotes-1 in intermediate alkalinity levels, while in Synechococcus spp., nanoeukaryotes-2 and microplankton remained stable. ROS levels did not significantly change in any functional group. The pigment ratios DD+DT : FUCO, and DD+DT : Chl-a increased in medium ∆TA treatments, supporting the idea of nutrient deficiency alleviation and the absence of physiological stress. Taken all data together, this study shows that there is minimal evidence indicating a harmful impact of high alkalinity on the plankton community. The OAE treatments did not result in physiological fitness impairment, thus OAE did not cause cellular stress in the phytoplankton community studied.