Table of Contents

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

Sampling

Glycans were profiled in bulk and high molecular weight dissolved organic matter (HMWDOM) and particulate organic matter (POM) in the deep chlorophyll max (DCM) and bottom water at three stations in the western North Atlantic Ocean. 

Sampling and processing of HMWDOM and POM

Seawater was collected using 30 L Niskin bottles on-board the R/V Endeavor. Niskin bottles were attached to rosettes equipped with conductivity, temperature and depth (CTD) sensors. Physical parameters were recorded using a Seabird 911+ CTD profiler. Data were processed and binned with SBE Data Processing software (v7.26.7). Collected seawater was transferred into acid-washed carboys before processing on-board. HMWDOM was concentrated to ~300 mL by tangential flow filtration (TFF). The TFF system (Sartorius <>) was run with 3 filter cassettes (1 kDa). HMWDOM samples were further concentrated back in the lab by freeze-drying and resuspension in lower volumes of MilliQ-water; for example, 10 mL of HMWDOM was freeze-dried and resuspended in 2.5 to 3mL. These concentrated HMWDOM samples were used for acid hydrolysis and microarray printing. POM samples were taken using in situ pumps onboard R/V Endeavor (EN683). At station 21 ~393 L were filtered in surface waters and 1276 L between 2800 and 3100 m depth. At station 22 ~214 L were sampled at DCM and 931 L at depths between 3321 and 3694 m. At station 23 the in situ pumps were deployed at 110 m (DCM) and 5100 m (depth) and pumped 515 L and 1196 L, respectively. 

Sequential extraction of polysaccharides from GF/F filters 

Seven punchouts of 11 m diameter were made from GF/F filters and sequentially extracted using MilliQ-water, 0.3M EDTA and 4M NaOH + 0.1% NaBH4 (Vidal-Melgosa et al., 2021). Subsequently solvents were added to the filter pieces, vortexed and incubated for 2 h at 60°C at 650 rpm in a heat block. Extracts were centrifuged at 6,000 x g for 15 min at 15°C and the supernatant was transferred to a new tube. NaOH extracts were neutralized with 4M HCl. 

Quantification of monosaccharides 

Polysaccharides of HMWDOM and POM samples were hydrolysed into monosaccharides. Freeze dried and concentrated HMWDOM samples (500 µL) were acid hydrolysed with 500 µL of 2M HCl. Additionally, 10 x 5 mm diameter punchouts from all GF/F filters were directly transferred to ampoules and acid hydrolysed in 750 µL 1M HCl. All hydrolyses were run at 100°C for 24 hrs. The supernatant of acid hydrolysed samples was dried in an acid-resistant vacuum concentrator (Martin Christ Gefriertrocknungsanlagen GmbH, Germany). Hydrolysates were reconstituted in MilliQ-water and pH adjusted to >7 with 0.1 M NaOH. Monosaccharide standards and hydrolysates were spiked with 13C-labelled glucose, galactose and mannose before derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP) (Rühmann et al., 2014). PMP-derivatives were separated on a Agilent 1290 Infinity II LC system equipped with a Waters CORTECS UPLC C18 column and measured on a SCIEX qTRAP5500 by multiple reaction monitoring (MRM) (Xu et al., 2018). Signal intensities were normalised to 13-labelled standards and calibrated against standard curves.

Data processed using Microsoft Excel.

- Imported "20250925_EN683_Monosaccharides_DOM_POM_LV_BCODMO.csv" into the BCO-DMO system
- Combined "date" and "time" (local EST time) and converted the value into a UTC ISO datetime value
- Renamed fields to comply with BCO-DMO naming guidelines, removing spaces, units, and parentheses
- Exported file as "987279_v1_en683_monosaccharides_dom_pom_lv.csv"

Substrate Structural Complexity and Abundance Control Distinct Mechanisms of Microbially-Driven Carbon Cycling in the Ocean

Almost half of the organic carbon produced in the ocean is processed by bacteria. Bacteria use extracellular (outside the cell) enzymes to break down large organic molecules to small sizes that can be transported into their cells. It has recently been discovered that bacteria use extracellular enzymes in two ways: ‘selfish uptake’ and ‘external hydrolysis’. External hydrolysis releases low molecular weight products to the environment where they can be used by other organisms. ‘Selfish uptake’ releases little or no products. This research will determine the extent and location of ‘selfish uptake’ in ocean waters. This process affects the distribution of organic carbon in the ocean, the flow of small organic molecules to feed a wider range of bacteria, and the composition and dynamics of the bacterial community. Recent results show that ‘selfish’ bacteria are active in deep ocean waters, where they take up complex polysaccharides (sugars) that are not hydrolyzed externally. These results inspired a new model that links ‘selfish uptake’ and external hydrolysis to the amount and complexity of the organic matter that is used by bacteria. This project will test the model by describing the polysaccharide fraction of marine organic matter, and studying the relationships between organic matter abundance, structural complexity, and extracellular enzyme use. Graduate and undergraduate students will participate in the project as members of the research team in the field and in the laboratory.

This research will test the hypothesis that the mechanism of polysaccharide processing is related to the cost to a cell of producing the enzymes required for its hydrolysis, and the probability that a cell will receive sufficient return on investment for producing the enzymes. The conceptual model that will be tested suggests that external hydrolysis is favored when organic matter is abundant, or when enzyme production costs can be shared (e.g., on particles, in biofilms); selfish uptake would be a better strategy when high molecular weight (HMW) organic matter is scarce, and particularly when the HMW organic matter is very complex. This study will test this model by characterizing the structure of polysaccharide-containing components of dissolved organic matter (DOM) and particulate organic matter (POM) collected from the ocean, by determining the extent of selfish uptake and rates of external hydrolysis of different polysaccharides by natural microbial communities from the surface and the deep ocean, and by incubation experiments that control for the abundance of polysaccharides of different structural complexity. This project will be carried out in collaboration with colleagues at the Max Planck Institute for Marine Microbiology, whose expertise in carbohydrate chemistry and structural analyses, and in advanced microscopy and analysis of complex microbial communities, are central to the project.

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.