Table of Contents

• Coverage
• Dataset Description
  • Methods & Sampling
  • Data Processing Description
  • BCO-DMO Processing Description
• Data Files
• Supplemental Files
• Related Publications
• Parameters
• Instruments
• Deployments
• Project Information
• Funding

Cruises STING I and STING II were conducted in Feb and July of 2023 along the West Florida Shelf (WFS), a coastal oligotrophic region characterized by low inorganic nutrient concentrations. Water was collected using towfish, stationary teflon pump, and ctd. 

To measure ¹⁵N₂ uptake, triplicate polycarbonate bottles (0.5-4.5 L) were rinsed with whole water then filled, ensuring that no bubbles remained. ¹⁵N₂ gas (~99%, Cambridge Isotope Laboratories) was added using a gas-tight syringe (VICI Precision Sampling) through a flexible silicon septa; samples were gently rocked for 15 min as described by Selden et al. (2019). Any remaining bubble was subsequently removed to ensure constant isotopic enrichment over the incubation period, and bottles were incubated on-deck for approximately 24 hours. On-deck incubators were constructed from clear acrylic and covered with two layers of screening to reduce light to approximately surface seawater conditions and were plumbed with flowing surface seawater to maintain temperature. Triplicate initial PN and N isotope samples (0.5-4.5 L) were vacuum-filtered from whole water onto 25 mm GF/F (nominal pore size 0.7 µm) pre-combusted (450 ^oC for 2 hours) filters and frozen at -20 ^oC until pelletizing and analysis.   

At the end of the incubation period, a 6 ml sample aliquot was collected from each bottle and transferred to a helium-purged Exetainer^TM to which 50 μl zinc chloride (50% w/v) was added. These samples were stored at room temperature, upside down, in 15 ml Falcon^TM tubes, submerged in ultrapure water, until analysis at the UC Davis Stable Isotope Facility using a ThermoScientific GasBench + Precon gas concentration system interfaced to a ThermoScientific Delta V Plus isotope-ratio mass spectrometer. The remaining incubation volume was then filtered as described above for initial PN samples. All PN samples were dried at 50 ^oC at least overnight and for no longer than a week. Once dry, the filters were pelletized into 30 mm-diameter tin disks (EA Consumables). Enriched and non-enriched pelletized samples were stored in separate dedicated desiccators until analysis. Initial and final incubation blank PN isotopic composition and concentration were analyzed at the University of South Florida on a Thermo Delta+XL IRMS w/ continuous-flow inlet coupled to a Carlo-Erba 1108 EA for NC isotope (15N, 13C) analysis. Final enriched PN isotopic composition and concentration samples were analyzed at the UC Davis Stable Isotope Facility using their Elementar Vario EL Cube or Micro Cube elemental analyzer interfaced with a PDZ Europa 20-20 isotope ratio mass spectrometer. Low mass samples (<10 µg N) were excluded from downstream analysis as IRMS response can become non-linear at low mass (White et al., 2020). 

Specific N₂ uptake rates (SUR) were calculated following Montoya et al. (1996):

SUR= [(A­_PN_tf -A_PN_t0)/(A_N2 – A_PN_t0)] x (1/time)                                                 Eqn. 1

where A represents the atom-% enrichment of the initial (t=0) or final (t=f) PN pool, or the N₂ pool. Across all experiments, A_N2_t0 averaged 0.367 ± 0.001 %. Experiments with <1% enrichment were excluded from downstream analysis. The specific uptake rate represents the relative contribution of diazotroph-derived N to PN turnover within a given sample. Absolute N₂ fixation rates (NFR) were calculated as:

NFR= SUR x [PN]                                                                                                     Eqn. 2

where [PN] indicates the PN concentration at t=f. Limits of detection and quantification were calculated per incubation by propagating the minimum detectable difference between initial and final ¹⁵N atom-% enrichment of the PN pool (3σ and 10σ, respectively, n≥6 N standards between 9-16 µg N measured daily) through Eqn. 2. Station rate estimates were calculated by taking the mean of the replicate incubations. For the station rate estimates, a rate was only considered detectable if two of three replicate incubations yielded detectable rates.

- Loaded sheet 1 from Excel file "STING_nfix_individual_tosubmit.xlsx" with header row 1; treated "" and "nd" as missing values
- Converted "Date-time_EST" field (format "%m/%d/%y %H:%M", Eastern timezone) to "DateTime_local_Eastern" field with output format "%Y-%m-%dT%H:%M" in Eastern timezone
- Converted "DateTime_local_Eastern" to "DateTime_UTC" with output format "%Y-%m-%dT%H:%MZ", converting from Eastern to UTC timezone
- Deleted original "Date-time_EST" field
- Renamed fields to comply with BCO-DMO naming conventions: Depth_m to Depth, Lat_degN to Lat, Lon_degW to Lon, vol_L to vol, "APN_ t0" to APN_t0
- Output written to "997130_v1_sting_nfix.csv"

NSF Award Abstract:
This project will investigate how groundwater discharge delivers important nutrients to the coastal ecosystems of the West Florida Shelf. Preliminary studies indicate that groundwater may supply both dissolved organic nitrogen (DON) and iron in this region. In coastal ecosystems like the West Florida Shelf that have very low nitrate and ammonium concentrations, DON is the main form of nitrogen available to organisms. Nitrogen cycling is strongly affected by iron availability because iron is essential for both photosynthesis and for nitrogen fixation. This study will investigate the sources and composition of DON and iron, and their influence on the coastal ecosystem. The team will sample offshore groundwater wells, river and estuarine waters, and conduct two expeditions across the West Florida Shelf in winter and summer. Investigators will participate in K-12 and outreach activities to increase awareness of the project and related science. The project will fund the work of six graduate and eight undergraduate students across five institutions, furthering NSF’s goals of education and training.

Motivated by preliminary observations of unexplained, tightly-correlated DON and dissolved iron concentrations across the West Florida Shelf (WFS), the proposed work will quantify the flux and isotopic signatures of submarine groundwater discharge (SGD)-derived DON and iron to the WFS, and evaluate the bioavailability of this temporally-variable source using four seasonal near-shore campaigns sampling offshore groundwater wells, estuarine, and riverine endmembers and two cross-shelf cruises. The work will evaluate whether SGD stimulates nitrogen fixation on the WFS, and the potential for the stimulated nitrogen fixation to further modify the chemistry of DON and dissolved iron in the region. The cross-shelf cruises will investigate hypothesized periods of maximum SGD and Trichodesmium abundance (June), and reduced river discharge and SGD (February), thus comparing two distinct biogeochemical regimes. The concentrations and isotopic compositions of DON and dissolved iron, molecular composition of DON, and the concentration and composition of iron-binding ligands will be characterized. Nitrogen fixation rates and Trichodesmium spp. abundance and expression of iron stress genes will be measured. Fluxes of DON and iron from SGD and rivers will be quantified with radium isotope mass balances. The impacts of SGD on nitrogen fixation and DON/ligand production will be constrained with incubations of natural phytoplankton communities with submarine groundwater amendments. Two hypotheses will be tested: 1) SGD is the dominant source of bioavailable DON and dissolved iron on the WFS, and 2) SGD-alleviation of iron stress changes the dominant Trichodesmium species on the WFS, increases nitrogen fixation rates and modifies DON and iron composition. Overall, the work will establish connections between marine nitrogen and iron cycling and evaluate the potential for coastal inputs to modify water along the WFS before export to the Atlantic Ocean. This study will thus provide a framework to consider these boundary fluxes in oligotrophic coastal systems and the relative importance of rivers and SGD as sources of nitrogen and iron in other analogous locations, such as coastal systems in Australia, India, and Africa, where nitrogen fixation and SGD have also been documented.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.