Table of Contents

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

Seawater for nutrients and dissolved organic carbon (DOC) analyses were typically collected from the CTD rosette in triplicate from different rosette bottles from two or three depths per cast.

For dissolved organic carbon (DOC) concentration measurements, seawater was collected directly from CTD bottles into a filtration apparatus consisting of a glass syringe connected to a filter unit loaded with 25 mm carbon-cleaned GF/F filter  (combusted at 450 °C for 4 hours.) 2–30 mL of seawater was filtered into a 40 mL glass TOC “EPA” pre-chilled glass vials (about ½ to 3/4 full). Vial moisture/condensation was wiped off of the outside of the vial and immediately placed in the freezer (upright). Samples were shipped frozen to the University of Washington Marine Chemistry Laboratory for analysis.

Seawater samples for nutrient concentration analysis (phosphate, silicate, nitrate, nitrite, and ammonium) were collected from CTD bottles into 60 ml plastic syringes loaded with a surfactant-free cellulose membrane (25 mm, 0.45 micron pore size, Nalgene). Syringes and 60 ml HDPE sample collection bottles were rinsed twice with sample water. About 45-50 ml of seawater was filtered into the collection bottles after rinsing again with 5 mL of filtered water. Samples were frozen for storage.

- Loaded "RC0078_bottle_nutrients_doc.csv" as resource "rc0078_bottle_nutrients_doc" (CSV format, row 1 headers), treating "", "nd", and "NA" as missing values
- Renamed fields: "year_month_day" to "Date", "Beam.Attenuation" to "Beam_Attenuation", "Beam.Transmission" to "Beam_Transmission"
- Reordered fields to: Date, Cruise, Station, LAT, LON, Cast, DOC, PO4, SiO4, NO3, NO2, NH4, PRS, Depth, TMP, SAL, OXY, Beam_Attenuation, Beam_Transmission, Fluorescence, PAR, pH
- Output to "997299_v1_rc0078_bottle_nutrients_doc.csv"

NSF Award Abstract:
Photosynthetic microbes provide food for nearly all other life in the ocean. Their metabolism produces organic molecules called metabolites that can leak out of cells, be intentionally excreted into seawater, or be released during cell death. Once outside the cell, these metabolites are the basis for specific interactions among microbes and determine community structure and activity. Yet, current understanding of metabolites in the ocean is limited by a historical lack of ability to measure them. The work proposed here will expand current knowledge of metabolite structures, concentrations, and production rates using recently developed analytical methods. These methods have already led to the discovery that homarine, a substituted pyridine first found in lobster in 1933, is the most abundant detectable metabolite in microbial communities of the North Pacific Ocean. While homarine is known as a predator deterrent, osmoprotectant, methyl donor, and antibiofouling agent, studies of its role in microbial community dynamics are lacking. The work proposed will clarify the role of homarine in the ocean’s microbial communities. This work will create an open-source metabolite database that will serve the broader field of metabolomics, a growing area in environmental, engineering, and medical sciences. This collaboration will also promote the careers of a graduate student and a postdoctoral researcher as well as an early career professor from an underrepresented group at a primarily undergraduate institution (PUI). Undergraduates from both institutions will contribute to project development and implementation, local cruises on the R.V. Carson, lab work, and dissemination of results. This research will be integrated into a curriculum-based research experience for undergraduates in a 200-level genetics course at the PUI, University of Puget Sound.

The proposed work will carry out field studies and laboratory experiments to test the hypothesis that metabolites are quantitatively significant forms of carbon and nitrogen flowing through microbial communities. The identity, quantity, and production rates of metabolites will also be determined. For homarine, the enzymes and organisms responsible for its transformations will be determined. Specific proposed activities will 1) Quantify nitrogenous metabolite pools and their net production rates (particulate and dissolved) in phytoplankton cultures and in marine surface water communities; 2) Isolate homarine consuming heterotrophic bacteria and use mutagenesis techniques, transcriptomics, and stable isotope assisted metabolomics to annotate genes and characterize the biochemical reactions involved in the degradation of homarine; 3) Carry out incubations of stable isotope labeled homarine in phytoplankton cultures, heterotrophic bacterial cultures sensitive to homarine, and natural communities to quantitatively evaluate the effect of homarine on growth, track homarine through metabolic pathways, and determine the kinetics of homarine uptake; 4) Identify homarine consumers and biochemical pathways for homarine use in the environment by mining existing environmental metatranscriptomes for homarine catabolism genes. The combination of these approaches will provide better understanding of the flow of nitrogen containing metabolites through marine microbial ecosystems. Results from this work will be disseminated through peer reviewed open-source publications as well as presentations to the scientific community and the general public.

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.