Table of Contents

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

An oceanographic survey was performed onboard the RV Sally Ride from Puntarenas, Costa Rica to San Diego, USA. Forty-nine CTD-rosette stations and 8 MOCNESS tows (Multiple Opening-Closing Net and Environmental Sensing System; Wiebe et al. 1985) were conducted along the cruise track (see Figure 1 of Gutiérrez-Bravo, et al. 2024). The CTD-rosette system used for water sampling and water column profiling included a SeaBird SBE9+ CTD and calibrated Seapoint-Fluorescence and SBE 43-DO sensors. A MOCNESS system was used for zooplankton and fish sampling and was equipped with 10 nets of 1 square meter (m^2) mouth opening and 333 micrometer (µm) mesh size, a SeaBird SBE9+ CTD, a SeaBird SBE 43 DO sensor, and flow meter and angle sensors.

The vertical sampling strategy followed five specific oxypleths (referred to as sampling levels) using horizontal tows. The deep level followed the 10 micromoles per kilogram (µmol/kg) oxypleth below the anoxic core (~900 meters (m) depth). The anoxic level followed the center of the anoxic core, with DO <1 µmol/kg and depth of ~450 m. The suboxic and hypoxic levels followed the 10 and 100 µmol/kg oxypleths above the anoxic core, with varying depths. The oxic level followed the >200 µmol/kg oxypleth near the surface (~20 m). Abrupt vertical oxygen gradients and the lagged depth control of the MOCNESS tow caused DO values to be less than half or more than double the desired DO values in seven out of 40 nets (M1-Oxic, M1-Hypoxic, M6-Hypoxic, M3-Suboxic, M6-Suboxic, M7-Suboxic and M8-Anoxic). These nets were considered outliers and were removed from the inter-level comparison statistics.

The zooplankton samples were preserved in ethanol 95%. Fish larvae were separated and identified to the most specific taxonomic level possible. The larval stages (preflexion, flexion, postflexion, and transformation) were defined according to Moser (1996). Juveniles and adults were separated, counted, and identified to the most specific taxonomic level possible. A more comprehensive description of the sample processing is described in Gutiérrez Bravo et al. (2024). The habitat type of each taxon was consulted in specialized literature (Moser 1996; Aceves-Medina et al. 2003; Froese and Pauly 2010).

The stable isotope ratios of carbon (13C/12C) and nitrogen (15N/14N) were measured on 39 samples of zooplankton (sample M5-Suboxic presented issues), 44 samples of fish larvae, and 8 samples of fish adults. For zooplankton, a wide-mouth 1.5 milliliter (mL) pipette was used to separate an aliquot from each of the 40 zooplankton samples. This approach was used to assess the integrated isotopic signature of the whole zooplankton community, and does not resolve species-specific differences. For fish, on the other hand, individuals of the same species, same development stage, and same net, were separated using a stereoscope and tweezer to obtain a critical weight of >2 milligrams (mg). Adjacent development stages were pooled if the sample weight was lower than 2mg.

The samples were rinsed to remove ethanol with deionized water, and then dried at 60 degrees Celsius (°C) for 24 hours. The dried samples were then ground to a fine powder using a mortar and pestle. The samples were loaded into tin cups and weighed to ~2mg using an analytical balance. Carbon and nitrogen isotope ratios were measured at the Boston University Stable Isotope Laboratory using a GV Instruments IsoPrime isotope ratio mass spectrometer coupled with an elemental analyzer.

The results were compared to international standards (Pee Dee Belemnite for 13C and atmospheric nitrogen for 15N) and presented as delta notation. MOCNESS data were analyzed using SBE Data processing.

- Imported original file "Supp material.xlsx" into the BCO-DMO system.
- Renamed fields to comply with BCO-DMO naming conventions.
- Converted Date column to YYYY-MM-DD format.
- Saved the final file as "936689_v1_sr2111_fish_larvae_isotope_data.csv".

NSF Award Abstract:
Several regions of the deep ocean naturally contain almost no oxygen. Because of this lack of oxygen, microbes living in these regions live in ways that differ from those in oxygenated waters consuming nitrate ions instead of oxygen for respiration. Use of nitrate for microbial respiration results in the production of nitrogen gas which is called denitrification. The resulting removal of nitrate has consequences for the whole ocean as nitrogen is an important nutrient controlling plant growth; however, whereas plants can use nitrogen in the form of nitrate, they cannot, with a few exceptions, use nitrogen gas. There remains a number of uncertainties regarding how much denitrification occurs in the ocean, what controls it, and how it varies in time and space. Traditional studies of ocean denitrification have been limited by the time ships can be at sea and the relatively small proportion of the ocean they can observe. Our project plans to remedy this problem by using vehicles called floats that can operate autonomously in the ocean for three years or more as they drift with currents over hundreds of kilometers. We will outfit ten floats with sensors to measure oxygen and nitrogen gas which will be placed throughout the oxygen-depleted region of the Pacific Ocean to the west of Mexico. This is the largest such region in the ocean from which we have two years of results from a prototype float which validated our approach. This study may well transform our understanding of ocean denitrification and ultimately benefit society as a whole through greater confidence in predictions of the ocean's nitrogen cycle and capacity to fix carbon dioxide under current and future conditions. Application and further development of float systems using commercially available technology will directly benefit successor studies, and more broadly showcase the use of water-following platforms to tackle difficult oceanographic problems. Advances from this study are expected to carry over to other disciplines including ocean biogeochemical modeling. Outreach activities, support for an early career scientist, and student training are included in the project. For the outreach activities, the investigators plan to tie into well-established after-school programs serving underrepresented populations in Massachusetts and established opportunities for public presentations using float related display materials at the University of Washington.

Oxygen deficient zones (ODZs), despite constituting a small fraction of total oceanic volume, play important roles in regulating global ocean carbon and nitrogen cycles including hosting 30 to 50% of the global loss of fixed nitrogen. Unfortunately, current uncertainty in ODZ nitrogen loss derives from substantial temporal and spatial variability in rates that remain under-sampled by ship-based measurements. While local regulation of nitrogen loss by oxygen and organic matter availability are well accepted, temporal/spatial variability in the nitrogen flux is likely a result of the influence of physical forcings such as remote ventilation, seasonal variability, and mesoscale eddies. Understanding how the impact of physical forcings on nitrogen loss as mediated through oxygen and organic flux will be required to fully understand the causes and consequences of any future ODZ expansion. To improve our understanding of ODZ nitrogen loss, we will carry out a multiyear, autonomous float-based observational program to address outstanding questions regarding bioavailable nitrogen loss in ODZs. As the largest ODZ and region of our pilot deployments, our operation area will be the Eastern Tropical N. Pacific (ETNP) where our study will determine over a multi-year period, in-situ nM-level oxygen and biogenic nitrogen on float profiles spanning geographic gradients in oxygen and surface productivity. For the first time, our study will also determine in situ nitrogen loss rates from changes in nitrogen concentration during 1 to 2 week Lagrangian float drifts along a constant density surface. A pilot 2 yr float deployment in the ETNP documents our ability to do so. Critically, our float-based approach more closely matches the frequency and distribution of observations to the expected variability in biogenic nitrogen production as compared to prior work and will dramatically increase the data density for this region by acquiring >500 profiles/drifts for nitrogen and >1000 profiles for nM oxygen.

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.