ALBERT

Description: We performed N budgets at three stations in the western tropical South Pacific (WTSP) Ocean during austral summer conditions (Feb. Mar. 2015) and quantified all major N fluxes both entering the system (N2 fixation, nitrate eddy diffusion, atmospheric deposition) and leaving the system (PN export). Thanks to a Lagrangian strategy, we sampled the same water mass for the entire duration of each long duration (5 days) station, allowing to consider only vertical exchanges. Two stations located at the western end of the transect (Melanesian archipelago (MA) waters, LD A and LD B) were oligotrophic and characterized by a deep chlorophyll maximum (DCM) located at 51 ± 18 m and 81 ± 9 m at LD A and LD B. Station LD C was characterized by a DCM located at 132 ± 7 m, representative of the ultra-oligotrophic waters of the South Pacific gyre (SPG water). N2 fixation rates were extremely high at both LD A (593 ± 51 µmol N m−2 d−1) and LD B (706 ± 302 µmol N m−2 d−1), and the diazotroph community was dominated by Trichodesmium. N2 fixation rates were lower (59 ± 16 µmol N m−2 d−1) at LD C and the diazotroph community was dominated by unicellular N2-fixing cyanobacteria (UCYN). At all stations, N2 fixation was the major source of new N (〉 90 %) before atmospheric deposition and upward nitrate fluxes induced by turbulence. N2 fixation contributed circa 8–12 % of primary production in the MA region and 3 % in the SPG water and sustained nearly all new primary production at all stations. The e-ratio (e-ratio = PC export/PP) was maximum at LD A (9.7 %) and was higher than the e-ratio in most studied oligotrophic regions (~ 1 %), indicating a high efficiency of the WTSP to export carbon relative to primary production. The direct export of diazotrophs assessed by qPCR of the nifH gene in sediment traps represented up to 30.6 % of the PC export at LD A, while there contribution was 5 and

Description: We performed nitrogen (N) budgets in the photic layer of three contrasting stations representing different trophic conditions in the western tropical South Pacific (WTSP) Ocean during austral summer conditions (February–March 2015). Using a Lagrangian strategy, we sampled the same water mass for the entire duration of each long-duration (5 days) station, allowing us to consider only vertical exchanges for the budgets. We quantified all major vertical N fluxes both entering (N2 fixation, nitrate turbulent diffusion, atmospheric deposition) and leaving the photic layer (particulate N export). The three stations were characterized by a strong nitracline and contrasted deep chlorophyll maximum depths, which were lower in the oligotrophic Melanesian archipelago (MA, stations LD A and LD B) than in the ultra-oligotrophic waters of the South Pacific Gyre (SPG, station LD C). N2 fixation rates were extremely high at both LD A (593 ± 51 µmol N m−2 d−1) and LD B (706 ± 302 µmol N m−2 d−1), and the diazotroph community was dominated by Trichodesmium. N2 fixation rates were lower (59 ± 16 µmol N m−2 d−1) at LD C, and the diazotroph community was dominated by unicellular N2-fixing cyanobacteria (UCYN). At all stations, N2 fixation was the major source of new N (〉 90 %) before atmospheric deposition and upward nitrate fluxes induced by turbulence. N2 fixation contributed circa 13–18 % of primary production in the MA region and 3 % in the SPG water and sustained nearly all new primary production at all stations. The e ratio (e ratio = particulate carbon export ∕ primary production) was maximum at LD A (9.7 %) and was higher than the e ratio in most studied oligotrophic regions (

Description: Here we report N2 fixation rates from a ∼ 4000 km transect in the western and central tropical South Pacific, a particularly undersampled region in the world ocean. Water samples were collected in the euphotic layer along a west to east transect from 160∘ E to 160∘ W that covered contrasting trophic regimes, from oligotrophy in the Melanesian archipelago (MA) waters to ultra-oligotrophy in the South Pacific Gyre (GY) waters. N2 fixation was detected at all 17 sampled stations with an average depth-integrated rate of 631 ± 286 µmolNm-2d-1 (range 196–1153 µmolNm-2d-1) in MA waters and of 85 ± 79 µmolNm-2d-1 (range 18–172 µmolNm-2d-1) in GY waters. Two cyanobacteria, the larger colonial filamentous Trichodesmium and the smaller UCYN-B, dominated the enumerated diazotroph community (〉 80 %) and gene expression of the nifH gene (cDNA 〉 105 nifH copies L−1) in MA waters. Single-cell isotopic analyses performed by nanoscale secondary ion mass spectrometry (nanoSIMS) at selected stations revealed that Trichodesmium was always the major contributor to N2 fixation in MA waters, accounting for 47.1–83.8 % of bulk N2 fixation. The most plausible environmental factors explaining such exceptionally high rates of N2 fixation in MA waters are discussed in detail, emphasizing the role of macro- and micro-nutrient (e.g., iron) availability, seawater temperature and currents.

Description: Biological dinitrogen (N2) fixation provides the major source of new nitrogen (N) to the open ocean, contributing more than atmospheric deposition and riverine inputs to the N supply. Yet the fate of the diazotroph-derived N (DDN) in the planktonic food web is poorly understood. The main goals of this study were (i) to quantify how much of DDN is released to the dissolved pool during N2 fixation and how much is transferred to bacteria, phytoplankton and zooplankton, and (ii) to compare the DDN release and transfer efficiencies under contrasting N2 fixation activity and diversity in the oligotrophic waters of the western tropical South Pacific (WTSP) Ocean. We used nanometre-scale secondary ion mass spectrometry (nanoSIMS) coupled with 15N2 isotopic labelling and flow cytometry cell sorting to track the DDN transfer to plankton, in regions where the diazotroph community was dominated by either Trichodesmium or by UCYN-B. After 48 h, ∼ 20–40 % of the N2 fixed during the experiment was released to the dissolved pool when Trichodesmium dominated, while the DDN release was not quantifiable when UCYN-B dominated; ∼ 7–15 % of the total fixed N (net N2 fixation + release) was transferred to non-diazotrophic plankton within 48 h, with higher transfer efficiencies (15 ± 3 %) when UCYN-B dominated as compared to when Trichodesmium dominated (9 ± 3 %). The pico-cyanobacteria Synechococcus and Prochlorococcus were the primary beneficiaries of the DDN transferred (∼ 65–70 %), followed by heterotrophic bacteria (∼ 23–34 %). The DDN transfer in bacteria was higher (34 ± 7 %) in the UCYN-B-dominating experiment compared to the Trichodesmium-dominating experiments (24 ± 5 %). Regarding higher trophic levels, the DDN transfer to the dominant zooplankton species was less efficient when the diazotroph community was dominated by Trichodesmium (∼ 5–9 % of the DDN transfer) than when it was dominated by UCYN-B (∼ 28 ± 13 % of the DDN transfer). To our knowledge, this study provides the first quantification of DDN release and transfer to phytoplankton, bacteria and zooplankton communities in open ocean waters. It reveals that despite UCYN-B fix N2 at lower rates compared to Trichodesmium in the WTSP, the DDN from UCYN-B is much more available and efficiently transferred to the planktonic food web than the DDN originating from Trichodesmium.

Description: In oligotrophic tropical and subtropical oceans, where strong stratification can limit the replenishment of surface nitrate, dinitrogen (N2) fixation by diazotrophs can represent a significant source of nitrogen (N) for primary production. The VAHINE experiment was designed to examine the fate of diazotroph derived nitrogen (DDN) in such ecosystems. In austral summer 2013 three large (~ 50 m3) in situ mesocosms were deployed for 23 days in the New Caledonia lagoon, an ecosystem that typifies the low-nutrient, low-chlorophyll environment, to stimulate diazotroph production. The zooplankton component of the study aimed to measure the incorporation of DDN into zooplankton biomass, and assess the role of direct diazotroph grazing by zooplankton as a DDN uptake pathway. Inside the mesocosms the diatom-diazotroph association (DDA) het-1 predominated during day 5-15 while the unicellular diazotrophic cyanobacteria UCYN-C predominated during day 15-23. A Trichodesmium bloom was observed in the lagoon (outside the mesocosms) towards the end of the experiment. The zooplankton community was dominated by copepods (63 % of total abundance) for the duration of the experiment. Using two source N isotope mixing models we estimated a mean ~ 30 % contribution of DDN to zooplankton biomass at the start of the experiment, indicating that the natural summer peak of N2 fixation in the lagoon was already contributing significantly to the zooplankton. Stimulation of N2 fixation BNF in the mesocosms corresponded with a generally low level enhancement of DDN contribution to zooplankton biomass, but with a peak of ~ 70 % in Mesocosm 1 following the UCYN-C bloom. qPCR analysis targeting four of the common diazotroph groups present in the mesocosms (Trichodesmium, het-1, het-2, UCYN-C) demonstrated that all were ingested by copepod grazers and that target abundance generally corresponded with their in situ abundance. 15N2 labeled grazing experiments provided evidence for direct ingestion and assimilation of UCYN-C-derived N by the zooplankton, but not for het-1 and Trichodesmium, supporting an important role of secondary pathways of DDN to the zooplankton for the latter groups, i.e., DDN contributions to the dissolved N pool and uptake by non-diazotrophs. This study appears to provide the first evidence of direct UCYN-C grazing by zooplankton, and indicates that UCYN-C-derived N contributes significantly to the zooplankton food web in the New Caledonia lagoon though a combination of direct grazing and secondary pathways.

Description: N2 fixation rates were measured daily in large (~ 50 m3) mesocosms deployed in the tropical South West Pacific coastal ocean (New Caledonia) to investigate the spatial and temporal dynamics of diazotrophy and the fate of diazotroph-derived nitrogen (DDN) in a low nutrient, low chlorophyll ecosystem. The mesocosms were intentionally fertilized with ~ 0.8 μM dissolved inorganic phosphorus (DIP) to stimulate diazotrophy. Bulk N2 fixation rates were replicable between the three mesocosms, averaged 18.5 ± 1.1 nmol N L−1 d−1 over the 23 days, and increased by a factor of two during the second half of the experiment (days 15 to 23) to reach 27.3 ± 1.0 nmol N L−1 d−1. These rates are higher than the upper range reported for the global ocean, indicating that the waters surrounding New Caledonia are particularly favourable for N2 fixation. During the 23 days of the experiment, N2 fixation rates were positively correlated with seawater temperature, primary production, bacterial production, standing stocks of particulate organic carbon, nitrogen and phosphorus, and alkaline phosphatase activity, and negatively correlated with DIP concentrations, DIP turnover time, nitrate, and dissolved organic nitrogen and phosphorus concentrations. The fate of DDN was investigated during the bloom of the unicellular diazotroph, UCYN-C, that occurred during the second half of the experiment. Quantification of diazotrophs in the sediment traps indicates that ~ 10 % of UCYN-C from the water column were exported daily to the traps, representing as much as 22.4 ± 5.5 % of the total POC exported at the height of the UCYN-C bloom. This export was mainly due to the aggregation of small (5.7 ± 0.8 μm) UCYN-C cells into large (100–500 μm) aggregates. During the same time period, a DDN transfer experiment based on high-resolution nanometer scale secondary ion mass spectrometry (nanoSIMS) coupled with 15N2 isotopic labelling revealed that 16 ± 6 % of the DDN was released to the dissolved pool and 21 ± 4 % was transferred to non-diazotrophic plankton, mainly picoplankton (18 ± 4 %) followed by diatoms (3 ± 2 %) within 24 h of incubation. This is consistent with the observed dramatic increase in picoplankton and diatom abundances, primary production, bacterial production and standing stocks of particulate organic carbon, nitrogen and phosphorus during the second half of the experiment in the mesocosms. These results offer insights into the fate of DDN during a bloom of UCYN-C in low nutrient, low chlorophyll ecosystems.

Description: N2 fixation fuels ~ 50 % of new primary production in the oligotrophic South Pacific Ocean. The VAHINE mesocosm experiment designed to track the fate of diazotroph derived nitrogen (DDN) in the New Caledonia lagoon. Here, we examined the temporal dynamics of heterotrophic bacterial production during this experiment. Three replicate large-volume (~ 50 m3) mesocosms were deployed and were intentionally fertilized with dissolved inorganic phosphorus (DIP) to stimulate N2 fixation. We specifically examined relationships between N2 fixation rates and primary production, determined bacterial growth efficiency and established carbon budgets of the system from the DIP fertilization to the end of the experiment (days 5–23). Heterotrophic bacterioplankton production (BP) and alkaline phosphatase activity (APA) were statistically higher during the second phase of the experiment (P2: days 15–23), when chlorophyll biomass started to increase compared to the first phase (P1: days 5–14). Among autotrophs, Synechococcus abundances increased during P2, possibly related to its capacity to assimilate leucine and to produce alkaline phosphatase. Bacterial growth efficiency based on the carbon budget was notably higher than generally cited for oligotrophic environments (27–43 %), possibly due to a high representation of proteorhodopsin-containing organisms within the picoplanctonic community. The carbon budget showed that the main fate of gross primary production (particulate + dissolved) was respiration (67 %), and export through sedimentation (17 %). BP was highly correlated with particulate primary production and chlorophyll biomass during both phases of the experiment but slightly correlated, and only during P2 phase, with N2 fixation rates. Our results suggest that most of the DDN reached the heterotrophic bacterial community through indirect processes, like mortality, lysis and grazing.

Description: The VAHINE mesocosm experiment was designed to trigger a diazotroph bloom and to follow the subsequent transfer of diazotroph derived nitrogen (DDN) in the rest of the foodweb. Three mesocosms (50 m3) located inside the Nouméa lagoon (New Caledonia, South West Pacific) were enriched with dissolved inorganic phosphate (DIP) in order to promote N2 fixation in these Low Nutrient Low Chlorophyll (LNLC) waters. Initial diazotrophic community were dominated by diatom diazotroph associations (DDAs), mainly by Rhizosolenia/Richelia intracellularis, and by Trichodesmium which fueled enough DDN to sustain the growth of other diverse diatom species and Synechococcus populations, that were well adapted to limiting DIP levels. After DIP fertilization (1 mM) on day 4, an initial lag time of 10 days was necessary for the mesocosm ecosystems to start building up biomass. Yet changes in community structure were already observed during this first period, with a significant drop of both Synechococcus and diatom populations, while Prochlorococcus benefited from DIP-addition. At the end of this first period, corresponding to when most added DIP was consumed, the diazotroph community changed drastically and became dominated by UCYN-C populations, which were accompanied by a monospecific bloom of the diatom Cylindrotheca closterium. During the second period, biomass increased sharply together with primary production and N2 fixation fluxes near tripled. Diatom populations, as well as Synechococcus and nano-phytoeukaryotes showed a re-increase towards the end of the experiment, showing efficient transfer of DDN to non diazotrophic phytoplankton.

Description: The globally important marine diazotrophic cyanobacterium Trichodesmium blooms regularly in the New Caledonian lagoons (Sowthwestern Pacific). We exploited the development of a Trichodesmium bloom in the lagoon waters outside the enclosed VAHINE mesocosms to specifically investigate the cellular processes mediating its decline. Trichodesmium cells (and associated microbiota) were sampled from the time of surface accumulation to biomass demise using a series of enclosed incubations to elucidate the stressors and subcellular underpinning of rapid (~ 24 h) biomass demise and disappearance. The development and decline of Trichodesmium populations was rapid with extensive surface accumulations (blooms) appearing within 24 h on the surface waters of the lagoon. Rapid decline of 〉 90 % biomass after 24 h of peak accumulation was observed in p opulations that were collected and incubated under ambient conditions. Metatranscriptomic profiling of Trichodesmium biomass 8 h and 22 h after bottle incubation of surface accumulations revealed evidence for phosphorus (P) and iron (Fe) stress, with upreg ulation of genes required to increase their availability and transport. In contrast, genes responsible for nutrient storage were downregulated. Total viral abundance, assessed by SYBR-green staining and analytical flow cytometry, oscillated throughout the experiment and showed no significant relationship with Trichodesmium bloom development or demise. Enhanced caspase-specific activity and upregulation of a suite of metacaspase genes during bloom demise implicated autocatalytic programmed cell death (PCD) as the mechanistic cause. At the same time, genes associated with buoyancy and gas-vesicle production were strongly downregulated concomitant with high concentrations of transparent exopolymeric particles (TEP), greatly aiding aggregation and expediting vertical flux to depth. Our results demonstrate that the rapid demise of this high-density, Trichodesmium surface bloom over 24 h was not caused by specific lytic infection but was rather induced by PCD in response to combined nutrient and oxidative stressors.

Description: In marine ecosystems, N2 fixation provides the predominant external source of nitrogen (N) (140 ± 50 Tg N yr−1), contributing more than atmospheric and riverine inputs to the N supply. Yet the fate and magnitude of the newly-fixed N, or diazotroph-derived N (hereafter named DDN) in marine ecosystems is poorly understood. Moreover, it remains unclear whether the DDN is preferentially directly exported out of the photic zone, recycled by the microbial loop, and/or transferred into larger organisms, subsequently enhancing indirect particle export. These questions were investigated in the framework of the VAHINE (VAriability of vertical and tropHIc transfer of diazotroph derived N in the south wEst Pacific) project. Triplicate large volume (~50 m3) mesocosms were deployed in the tropical South West Pacific coastal ocean (New Caledonia) to maintain a stable water-mass without disturbing ambient light and temperature conditions. The mesocosms were intentionally fertilized with ~0.8 μM dissolved inorganic phosphorus (DIP) at the start of the experiment to stimulate diazotrophy. A total of 47 stocks, fluxes, enzymatic activities and diversity parameters were measured daily inside and outside the mesocosms by the 40 scientists involved in the project. The experiment lasted for 23 days and was characterized by two distinct and successive diazotroph blooms: a dominance of diatom-diazotroph associations (DDAs) during the first half of the experiment (days 2–14) followed by a bloom of UCYN-C during the second half of the experiment (days 15–23). These conditions provided a unique opportunity to compare the DDN transfer and export efficiency associated with different diazotrophs. Here we summarize the major experimental and modelling results obtained during the project and described in the VAHINE Special issue, in particular those regarding the evolution of the main standing stocks, fluxes and biological characteristics over the 23-days experiment, the contribution of N2 fixation to export fluxes, the DDN released to dissolved pool and its transfer to the planktonic food web (bacteria, phytoplankton, zooplankton). We then apply our Eco3M modelling platform further to infer the fate of DDN in the ecosystem and role of N2 fixation on productivity, food web structure and carbon export. Recommendations for future work are finally provided in the conclusion section.