Bacterial productivity of samples from three stations in the Western North Atlantic aboard R/V Atlantic Explorer cruise AE2413 during May 2024

Website: https://www.bco-dmo.org/dataset/963407
Data Type: Cruise Results, Other Field Results
Version: 1
Version Date: 2025-06-06

Project
» Collaborative Research: Pressure effects on microbially-catalyzed organic matter degradation in the deep ocean (Pressure effects on microbes)
ContributorsAffiliationRole
Arnosti, CarolUniversity of North Carolina at Chapel Hill (UNC-Chapel Hill)Principal Investigator
Ghobrial, SherifUniversity of North Carolina at Chapel Hill (UNC-Chapel Hill)Scientist
Mickle, AudreyWoods Hole Oceanographic Institution (WHOI BCO-DMO)BCO-DMO Data Manager

Abstract
Heterotrophic bacteria and archaea (here: microbes) are critical drivers of the ocean’s biogeochemical cycles, active throughout the depth of the ocean. Their capabilities and limitations help determine the rates and locations at which carbon and nutrients are regenerated, as well as the extent to which organic matter is preserved (Hedges 1992). In the deep ocean, at bathy- and abyssopelagic depths (ca. 1000-6000m), these communities are dependent upon the sinking flux of particulate organic matter (POM) from the surface ocean (Bergauer et al. 2018). This dependence means that heterotrophic microbial communities must produce the extracellular enzymes required to solubilize and hydrolyze high molecular weight (HMW) POM to sizes substrates suitable for cellular uptake. A recent global-scale investigation of deep-sea microbes in fact found that the genetic potential for exported (extracellular) enzymes among bacteria in deep waters was far greater than for communities in surface or mesopelagic waters (Zhao et al. 2020). We have new evidence that a substantial fraction of bacteria in bottom water from the North Atlantic Ocean use a specialized set of extracellular enzymes to rapidly take up HMW polysaccharides (Giljan et al. 2022), a substrate processing mechanism that would not be detected with the low molecular weight substrates used in most prior studies of microbial activity in the deep ocean (Nagata et al. 2010).   Through our collaboration with the Danish Center for Hadal Research, we were able to use pressurization systems and in situ specialized equipment to investigate the effects of pressures characteristic of bathy- and abyssopelagic depths on microbial communities and their extracellular enzymes in the open North Atlantic Ocean.      Here we present the measurement of 3H-leucine incorporation by heterotrophic bacteria using a cold trichloroacetic acid (TCA) and microcentrifuge extraction method (Kirchman, 2001) at different sites in the Western North Atlantic aboard R/V Atlantic Explorer during during the research cruise AE2413 (2024-05-09 to 2024-05-28).  All work and incubations were performed in a UNOLS isotope lab, or within designated areas at the University of North Carolina at Chapel HIll post cruise. This dataset contains collection metadata, environmental conditions, sample types and treatments, incubation conditions, substrate types, radioactivity measurements, and calculated incorporation rates of 3H-leucine.


Coverage

Location: Three stations in the Western North Atlantic. Water samples were taken at multiple depths. Total water column depths were 4260m, 5280m, and 4310m, respectively.
Spatial Extent: N:42.17828 E:-60.0479 S:34.98703 W:-73.03333
Temporal Extent: 2024-05-10 - 2024-05-19

Methods & Sampling

Water for incubation  was collected via Niskin bottles mounted on a rosette, equipped with a CTD.  Bulk water was collected into an acid washed 50 mL Falcon tube directly from niskin bottles. 

Isotopically diluted L-[3,4,5-3H(N)]-Leucine (Revvity, NET460250UC, specific activity of 3.811 TBq/mmol) was added to 1.7ml triplicate subsamples and one TCA killed control (20 nM final concentration).  Samples and killed control were incubated between 5 and 48 hours at nearly in-situ temperature or 4°C.  Following incubation, live samples were killed with 100% (w/v) TCA and all samples were centrifuged (10,000 rpm at 4°C for 10 min) to pelletize cell material.  The supernatant liquid was removed and 1 mL of ice cold 5% (w/v) TCA solution was added, followed by vortex mixing and centrifugation.  Supernatant removal, mixing, and centrifugation were repeated using 1 mL of ice cold 80% ethanol solution.  Again, the supernatant liquid was removed and each sample was left to dry in a hood overnight.  After drying, 1 mL of scintillation cocktail (ScintiSafe 30% Cocktail, Fisher SX23-5) was added and left overnight so that precipitated proteins dissolve into scintillation fluid.  Incorporated radioactivity was measured using a PerkinElmer Tri-Carb 2910TR LSA scintillation counter.  Radioactivity was compared to 1 mL of scintillation cocktail spiked with an identical amount of isotopically diluted L-[3,4,5-3H(N)]-Leucine that was added to samples.  Incorporation rate was calculated by dividing sample radioactivity by incubation time.


Data Processing Description

Data were processed using Microsoft Excel.


BCO-DMO Processing Description

- Imported "20250424_BCODMO_AE2413 Bulk water_3H-Leucine_incorporation_csv.csv" into BCO-DMO system
- Removed two rows where time is NULL at submitter's request
- Converted "date" to YYYY-MM-DD format
- Created new ISO formatted datetime in UTC
- Removed string "MPa" from the "incubation_pressure" values
- Renamed fields to remove spaces and units from parameters to comply with BCO-DMO system and style requirements
- Exported file as "963407_v1_bacterial_productivity.csv"


Problem Description

Station 26 DCM was not measured due to sampling error.

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Data Files

File
963407_v1_bacterial_productivity.csv
(Comma Separated Values (.csv), 4.61 KB)
MD5:21929134d2f9af0ab5b0bb77a9ab70ea
Primary data file for dataset ID 963407, version 1

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Related Publications

Bergauer, K., Fernandez-Guerra, A., Garcia, J. A. L., Sprenger, R. R., Stepanauskas, R., Pachiadaki, M. G., Jensen, O. N., & Herndl, G. J. (2017). Organic matter processing by microbial communities throughout the Atlantic water column as revealed by metaproteomics. Proceedings of the National Academy of Sciences, 115(3). https://doi.org/10.1073/pnas.1708779115
Methods
Giljan, G., Arnosti, C., Kirstein, I. V., Amann, R., & Fuchs, B. M. (2022). Strong seasonal differences of bacterial polysaccharide utilization in the North Sea over an annual cycle. Environmental Microbiology, 24(5), 2333–2347. Portico. https://doi.org/10.1111/1462-2920.15997
Methods
Hedges, J. I. (1992). Global biogeochemical cycles: progress and problems. Marine Chemistry, 39(1–3), 67–93. https://doi.org/10.1016/0304-4203(92)90096-s https://doi.org/10.1016/0304-4203(92)90096-S
Methods
Kirchman, D. (2001). Measuring bacterial biomass production and growth rates from leucine incorporation in natural aquatic environments. Marine Microbiology, 227–237. doi:10.1016/s0580-9517(01)30047-8 https://doi.org/10.1016/S0580-9517(01)30047-8
Methods
Nagata, T., Tamburini, C., Arístegui, J., Baltar, F., Bochdansky, A. B., Fonda-Umani, S., Fukuda, H., Gogou, A., Hansell, D. A., Hansman, R. L., Herndl, G. J., Panagiotopoulos, C., Reinthaler, T., Sohrin, R., Verdugo, P., Yamada, N., Yamashita, Y., Yokokawa, T., & Bartlett, D. H. (2010). Emerging concepts on microbial processes in the bathypelagic ocean – ecology, biogeochemistry, and genomics. Deep Sea Research Part II: Topical Studies in Oceanography, 57(16), 1519–1536. https://doi.org/10.1016/j.dsr2.2010.02.019
Methods
Zhao, Z., Baltar, F., & Herndl, G. J. (2020). Linking extracellular enzymes to phylogeny indicates a predominantly particle-associated lifestyle of deep-sea prokaryotes. Science Advances, 6(16). https://doi.org/10.1126/sciadv.aaz4354
Methods

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Parameters

ParameterDescriptionUnits
deployment

Cruise ID

unitless
station

Station number 24, 25, 26.

unitless
latitude_n

Latitude of sampling site, south is negative.

Decimal degrees
longitude_e

Longitude of sampling site, west is negative.

Decimal degrees
ISO_DateTime_UTC

Datetime of collection in ISO format, UTC.

unitless
date

Date of sample collection.

unitless
time_local_est

Time of sample collection (local time), US Eastern Time (UTC-05:00)

unitless
cast_number

Cast number (refers to cast of CTD/Niskin bottles on cruise)

unitless
depth_description

Water column feature or oceanic zone sampled DCM (Deep Chlorophyll Maximum), OMZ (Oxygen Minimum Zone), Bathy, or Deep (bottom or near bottom). Station 26 DCM was not measured due to sampling error.

unitless
depth_actual

Actual depth at which water was collected

m
insitu_temp

Temperature of the samples in-situ.

degrees Celsius
sample_type

The type of sample, whether it was incubated using water from the bulk water column, sediments, or amended

unitless
incubation_type

The type of incubation (in epitube or excitaner vial at atmospheric pressure or in excitaner vial in pressure vessel)

unitless
incubation_pressure

Amount of pressure applied during incubation

MPa
incubation_temp

Temperature of 3H-Leucine incubation.

degrees Celsius
unamended_amended

Whether the sample was amended (A) or unamended (U).

unitless
substrate

3H-leucine

unitless
incubation_time

Amount of time in hours samples were incubated with 3H-leu prior to addition of TCA

hours
DPM_Kill

Radioactivity of incorporated 3H-leucine in disintegrations per minute of killed control.

disintegrations per minute (dpm)
DPM_rep1

Radioactivity of incorporated 3H-leucine in disintegrations per minute of replicate 1.

disintegrations per minute (dpm)
DPM_rep2

Radioactivity of incorporated 3H-leucine in disintegrations per minute of replicate 2.

disintegrations per minute (dpm)
DPM_rep3

Radioactivity of incorporated 3H-leucine in disintegrations per minute of replicate 3.

disintegrations per minute (dpm)
Average_incorp

Amount of incorporated 3H-leucine in picomoles per liter (average radioactivity of replicates in excess of radioactivity of killed control relative to the radioactivity of a standard amount of 3H-Leucine).

pmol L-1
H3_Leu

Amount of incorporated 3H-leucine in picomoles per liter per hour.

pmol L-1 h-1
stdev

Standard deviation in the amount of incorporated 3H-leucine in picomoles per liter per hour.

pmol L-1 h-1


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Instruments

Dataset-specific Instrument Name
CTD
Generic Instrument Name
CTD - profiler
Dataset-specific Description
Water for incubation  was collected via Niskin bottles mounted on a rosette, equipped with a CTD.  Bulk water was collected into an acid washed 50 mL Falcon tube directly from niskin bottles. 
Generic Instrument Description
The Conductivity, Temperature, Depth (CTD) unit is an integrated instrument package designed to measure the conductivity, temperature, and pressure (depth) of the water column. The instrument is lowered via cable through the water column. It permits scientists to observe the physical properties in real-time via a conducting cable, which is typically connected to a CTD to a deck unit and computer on a ship. The CTD is often configured with additional optional sensors including fluorometers, transmissometers and/or radiometers. It is often combined with a Rosette of water sampling bottles (e.g. Niskin, GO-FLO) for collecting discrete water samples during the cast. This term applies to profiling CTDs. For fixed CTDs, see https://www.bco-dmo.org/instrument/869934.

Dataset-specific Instrument Name
PerkinElmer Tri-Carb 2910TR LSA scintillation counter
Generic Instrument Name
Liquid Scintillation Counter
Dataset-specific Description
Incorporated radioactivity was measured using a PerkinElmer Tri-Carb 2910TR LSA scintillation counter. 
Generic Instrument Description
Liquid scintillation counting is an analytical technique which is defined by the incorporation of the radiolabeled analyte into uniform distribution with a liquid chemical medium capable of converting the kinetic energy of nuclear emissions into light energy. Although the liquid scintillation counter is a sophisticated laboratory counting system used the quantify the activity of particulate emitting (ß and a) radioactive samples, it can also detect the auger electrons emitted from 51Cr and 125I samples. Liquid scintillation counters are instruments assaying alpha and beta radiation by quantitative detection of visible light produced by the passage of rays or particles through a suitable scintillant incorporated into the sample.

Dataset-specific Instrument Name
Niskin bottles
Generic Instrument Name
Niskin bottle
Dataset-specific Description
Water for incubation  was collected via Niskin bottles mounted on a rosette, equipped with a CTD.  Bulk water was collected into an acid washed 50 mL Falcon tube directly from niskin bottles. 
Generic Instrument Description
A Niskin bottle (a next generation water sampler based on the Nansen bottle) is a cylindrical, non-metallic water collection device with stoppers at both ends. The bottles can be attached individually on a hydrowire or deployed in 12, 24, or 36 bottle Rosette systems mounted on a frame and combined with a CTD. Niskin bottles are used to collect discrete water samples for a range of measurements including pigments, nutrients, plankton, etc.


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Deployments

AE2413

Website
Platform
R/V Atlantic Explorer
Start Date
2024-05-08
End Date
2024-05-28


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Project Information

Collaborative Research: Pressure effects on microbially-catalyzed organic matter degradation in the deep ocean (Pressure effects on microbes)

Coverage: Western North Atlantic, hadal depths of the Pacific


NSF Award Abstract:
Microbes are important players in the carbon cycle in the ocean. These organisms consume organic carbon and produce carbon dioxide in marine systems. Because the average depth of the ocean is 4000 m, microbes must work at high pressures typical of the deep ocean (>1000 m). Although high pressure is known to affect marine microbes, their carbon cycling activities have mostly been measured at surface ocean pressures. As a result, it remains unknown how closely these measurements reflect the activities of deep-sea microbes at high pressures. As a result of collaborations with scientists in Denmark and Germany, this project will be able to use special equipment to investigate the effects of high pressures on marine microbes and their carbon cycling activities. This work is necessary to quantify rates of carbon cycling and identify the microbes involved, especially in deep waters. The project will provide training for diverse undergraduate and graduate students, and a postdoc who will conduct novel research in the U.S., Denmark, and Germany, both at sea and in the lab. The scientists will also teach middle school students about the role of microbes in the carbon cycle and pressure effects on life in the ocean. The project will provide internships for high school students, focusing on first-generation students who would like to go to college. This work may aid in future efforts to identify enzymes that function well under high pressure.

Heterotrophic microbes (e.g., bacteria and archaea) are found throughout the ocean. Their biogeochemical functions help determine the rates and locations at which carbon and nutrients are regenerated, as well as the extent to which organic matter is preserved. Although research has shown that pressure profoundly affects the activities of marine microbes, most investigations of microbial communities of the deep sea are conducted at atmospheric pressure, due to the limited availability of specialized equipment. In collaboration with the Danish Center for Hadal Research at the University of Southern Denmark, this study will identify the effects of pressure on microbial communities and their extracellular enzymes of pressures characteristic of bathy- and abyssopelagic depths. At sea and in the lab, the scientific team will compare the effects of depressurization on the activities of enzymes produced by microbial communities of the deep ocean, as well as the effects of high pressure on surface-water derived enzymes and communities. Fieldwork will take place in Danish coastal waters, well as in the open North Atlantic and Pacific Oceans. Using pressurization systems and in situ incubations, this study will measure hydrolysis rates of peptides and polysaccharides, two of the major classes of marine organic matter. Project activities will also focus on developing the means to measure enzyme activities in situ in the deep ocean. In collaboration with colleagues from the Max Planck Institute for Marine Microbiology in Germany, this proect will additionally investigate whether pressure affects the selfish uptake of polysaccharides. These studies will provide new insight into understudied but key factors that help determine the fate of organic matter in the deep ocean.

This project is funded by the Biological and Chemical Oceanography Programs.

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.



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Funding

Funding SourceAward
NSF Division of Ocean Sciences (NSF OCE)

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