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

Water was collected via Niskin bottles mounted on a rosette, equipped with a CTD.

For mesocosm (large volume) incubation experiments (referred to as “LV” incubations), seawater was transferred to 20 L carboys that were rinsed three times with water from the sampling depth and then filled with seawater from a single Niskin bottle, using silicone tubing that had been acid washed then rinsed with distilled water prior to use. Four carboys were filled at each depth from bottom water, water from the depth at which oxygen showed a minimum, and deep chlorophyll maximum (DCM) water, according to the CTD. Triplicate 20L carboys were amended with ca. 500 mg (exact mass was recorded for each addition) of HMW Thalassiosira; unamended single carboys were used for controls. All mesocosms were incubated in the dark at near in-situ temperatures. Mesocosms were sub-sampled at the start of incubation (0 days), and then after at approximately 2 days, 6 or 7 days, 11 days, and 16 days for multiple assays including total cell abundance.

Total cell abundance was quantified following Giljan et al. (2023). In brief, 25-50 mL of water was fixed with a final concentration of 1% formaldehyde and subsequently filtered onto 0.2 µm polycarbonate filters (Millipore). The DNA of filtered cells were simultaneously counterstained and mounted using a 1 ng/µL working solution of 4’,6-diamidin-2-phenylindol (DAPI) mixed with Citifluor/VectaShield (4:1) solution. A minimum of 45 microscopic images were acquired per sample using an automated imaging system (Zeiss AxioImager.Z2 microscope stand; 63x magnification oil immersion plan apochromatic objective with 1.4 NA, Carl Zeiss). Final cell abundance was determined using ACMETOOL 3 (http://www.tchnobiology.ch and Max Planck Institute of Marine Microbiology, Bremen). Quantification of DAPI-stained cells (signal:background > 1.5) is reported as total cell abundance. Note that for the amended mesocosms, in many cases the cells were too dense to count automatically. For these samples, 15 microscopic images were manually counted using the ACMETOOL 3 software; a minimum of 12 of 4510 images are needed to accurately capture the total cell abundance per filter. In some cases, moreover, there were not enough cells to calculate total cell abundance or the filters needed to determine cell abundance were not available (i.e., samples were lost during transit; some filters were dropped during experimental subsampling), so the data presented for each timepoint in several cases originates from different replicate mesocosms during the time course of incubation.

Image analysis was performed with ACMETOOL (Zeder, M. 2005-2021, Software for Biology, http://www.technobiology.ch and Max Planck Institute for marine microbiology, Bremen, version 3) and Zen software package (Carl Zeiss).

- Loaded CSV file "20251114_EN638_CellCounts_Processed_BCODMO.csv" with header row 1; treated empty strings and "nd" as missing values
- Extracted non-numeric values from cell_abundance into a new flag field cell_abundance_flag, to capture symbols that were present in the "cell_abundance" values ("-" indicating lost samples or missing images, "*" indicating too few cells to quantify) without corrupting the numeric column
- Applied find/replace to time_sampled to normalize timestamps lacking seconds by appending ":00" to values matching the pattern m/d/yyyy h:mm
- Converted time_sampled from format "%m/%d/%Y %H:%M:%S" to ISO 8601 format "%Y-%m-%dT%H:%M" as a string field
- Renamed field "in-situ_temp" to "in_situ_temp" to remove the hyphen
- Exported file as "994900_v1_en638_lv_cell_counts.csv"

NSF Award Abstract:
Marine dissolved organic matter (DOM) is one of the largest actively-cycling reservoirs of organic carbon on the planet, and thus a major component of the global carbon cycle. The high molecular weight (HMW) fraction of DOM is younger in age and more readily consumed by microbes than lower molecular weight (LMW) fractions of DOM, but the reasons for this difference in reactivity between HMW DOM and LMW DOM are unknown. Two factors may account for the greater reactivity of HMW DOM: (i) targeted uptake of HMW DOM by specific bacteria, a process the PI and her collaborators at the Max Planck Institute for Marine Microbiology (MPI) recently identified in surface ocean waters; and (ii) a greater tendency of HMW DOM to aggregate and form gels and particles, which can be colonized by bacteria that are well-equipped to breakdown organic matter. Scientists and students from the University of North Carolina (UNC) - Chapel Hill will collaborate with researchers at the MPI for Marine Microbiology (Bremen, Germany) to investigate this breakdown of HMW DOM by marine microbial communities. These investigations will include a field expedition in the North Atlantic, during which HMW DOM degradation rates and patterns will be compared in different water masses and under differing conditions of organic matter availability. DOM aggregation potential, and degradation rates of these aggregates, will also be assessed. Specialized microscopy will be used in order to pinpoint HMW DOM uptake mechanisms and rates. The work will be complemented by ongoing studies of specific bacteria that breakdown HMW DOM, their genes, and their proteins. Graduate as well as undergraduate students will participate as integral members of the research team in all aspects of the laboratory and field work; aspects of the project will also be integrated into classes the scientist teaches at UNC.

The existence of a size-reactivity continuum of DOM - observations and measurements showing that HMW DOM tends to be younger and more reactive than lower MW DOM - has been demonstrated in laboratory and field investigations in different parts of the ocean. A mechanistic explanation for the greater reactivity of HMW DOM has been lacking, however. This project will investigate the mechanisms and measure rates of HMW DOM degradation, focusing on identifying the actors and determining the factors that contribute to rapid cycling of HMW DOM. Collaborative work at UNC and MPI-Bremen recently identified a new mechanism of HMW substrate uptake common among pelagic marine bacteria: these bacteria rapidly bind, partially hydrolyze, and transport directly across the outer membrane large fragments of HMW substrates that can then be degraded within the periplasmic space, avoiding production of LMW DOM in the external environment. This mode of substrate processing has been termed selfish, since targeted HMW substrate uptake sequesters resources away from other members of microbial communities. Measurements and models thus must account for three modes of substrate utilization in the ocean: selfish, sharing (external hydrolysis, leading to low molecular weight products), and scavenging (uptake of low molecular weight hydrolysis products without production of extracellular enzymes). Using field studies as well as mesocosm experiments, the research team will investigate the circumstances and locations at which different modes of substrate uptake predominate. A second focal point of the project is to determine the aggregation potential and microbial degradation of aggregated HMW DOM. Preliminary studies have demonstrated that particle-associated microbial communities utilize a broader range of enzymatic capabilities than their free-living counterparts. These capabilities equip particle-associated communities to effectively target a broad range of complex substrates. The project will thus focus on two key aspects of HMW DOM - the abilities of specialized bacteria to selectively sequester HMW substrates, as well as the greater potential of HMW substrates to aggregate ? and will quantify these factors at different locations and depths in the ocean. The project will thereby provide a mechanistic underpinning for observations of the DOC size-reactivity continuum, an essential part of developing an overall mechanistic understanding of organic matter degradation in the ocean.