http://lod.bco-dmo.org/id/dataset/887562
eng; USA
utf8
dataset
Highest level of data collection, from a common set of sensors or instrumentation, usually within the same research project
Biological and Chemical Oceanography Data Management Office (BCO-DMO)
Unavailable
508-289-2009
WHOI MS#36
Woods Hole
MA
02543
USA
info@bco-dmo.org
http://www.bco-dmo.org
Monday - Friday 8:00am - 5:00pm
For questions regarding this resource, please contact BCO-DMO via the email address provided.
pointOfContact
2023-01-20
ISO 19115-2 Geographic Information - Metadata - Part 2: Extensions for Imagery and Gridded Data
ISO 19115-2:2009(E)
DOC uptake rates by coccolithophores and scintillation counts from field experiments in the North Atlantic during R/V Endeavor cruise EN616 in July 2018
2023-03-22
publication
2023-03-22
revision
Marine Biological Laboratory/Woods Hole Oceanographic Institution Library (MBLWHOI DLA)
2023-03-23
publication
https://doi.org/10.26008/1912/bco-dmo.887562.1
William M. Balch
Bigelow Laboratory for Ocean Sciences
principalInvestigator
Stephen D. Archer
Bigelow Laboratory for Ocean Sciences
principalInvestigator
David T. Drapeau
Bigelow Laboratory for Ocean Sciences
principalInvestigator
Jelena Godrijan
Bigelow Laboratory for Ocean Sciences
principalInvestigator
Biological and Chemical Oceanography Data Management Office (BCO-DMO)
Unavailable
508-289-2009
WHOI MS#36
Woods Hole
MA
02543
USA
info@bco-dmo.org
http://www.bco-dmo.org
Monday - Friday 8:00am - 5:00pm
For questions regarding this resource, please contact BCO-DMO via the email address provided.
publisher
Cite this dataset as: Balch, W. M., Godrijan, J., Drapeau, D. T., Archer, S. D. (2023) DOC uptake rates by coccolithophores and scintillation counts from field experiments in the North Atlantic during R/V Endeavor cruise EN616 in July 2018. Biological and Chemical Oceanography Data Management Office (BCO-DMO). (Version 1) Version Date 2023-03-22 [if applicable, indicate subset used]. doi:10.26008/1912/bco-dmo.887562.1 [access date]
Dataset Description: <p>This dataset is part of a larger study with the following goals:</p>
<ul>
<li>&nbsp;<strong>Goal #1</strong>: measure the mixotrophic uptake and assimilation of <sup>14</sup>C-acetate, <sup>14</sup>C-mannitol and <sup>14</sup>C-glycerol as a carbon source by natural assemblages of coccolithophores and compare it to their autotrophic uptake and assimilation of DIC. These three organics were chosen due to their high potential for significant osmotrophy by coccolithophores as seen in previous culture studies (Godrijan et al. (2020) and related&nbsp;dataset:&nbsp;https://www.bco-dmo.org/dataset/858771).&nbsp;The design of these experiments used radiochemical and single cell/flow cytometer methods to distinguish osmotrophy by coccolithophores from that of&nbsp;other naturally-occurring microalgae.</li>
<li><strong>Goal #2</strong>: test for the fixation of <sup>14</sup>C-labeled organics into both POC and PIC fractions in natural populations of coccolithophores, in order to examine the potential role of coccolithophore osmotrophy in the&nbsp;biological carbon pump and alkalinity carbon pump paradigms.</li>
</ul> Methods and Sampling: <p>Coccolithophores are typically thought of as photoautotrophs yet they are known to inhabit sub-euphotic environments with insufficient light for photosynthesis. Field experiments were performed in the NW Atlantic to determine the potential uptake of dissolved organic carbon (DOC) by coccolithophores to compare with the bicarbonate uptake associated with standard photosynthesis and calcification.&nbsp; The cruise sampled portions of the Gulf of Maine, Georges Bank, continental shelf and slope waters of the northern part of the mid-Atlantic Bight plus Sargasso Sea, during <em>R/V Endeavor</em> cruise EN616 in July&nbsp;2018.&nbsp; Natural phytoplankton populations were incubated with radiolabeled DOC compounds, <sup>14</sup>C-acetate, <sup>14</sup>C-mannitol and <sup>14</sup>C-glycerol.&nbsp; Following the 24 hour incubations at simulated <em>in situ</em> conditions of light and temperature, coccolithophores were sorted from these natural populations using flow cytometry (based on their chlorophyll fluorescence and birefringence signatures).&nbsp; The sorts were collected and the uptake rate of these compounds into both organic and inorganic fractions was subsequently measured using the microdiffusion technique coupled with high-precision scintillation counting.&nbsp; Bulk samples were also filtered from these incubated natural populations in which radiolabeled DOC was taken-up and assimilated into the particulate organic carbon of general phytoplankton (not just coccolithophores) as well as the PIC of the broader assemblage of calcifiers (likely dominated by coccolithophores based on the small sample volumes).&nbsp; &nbsp;The bulk samplings had higher signal-to-noise for radioactivity measurements of DOC uptake into coccolithophore PIC due to the increased sample size, whereas the flow cytometer sorts had higher specificity for isolating coccolithophores, which allowed better discerning the fixation of <sup>14</sup>C-labeled DOC into both organic tissues and calcium carbonate (particulate inorganic carbon) of coccolithophores, despite the lower signal-to-noise.&nbsp;DOC uptake rates were measurable but slow relative to bicarbonate uptake rates and the resultant growth rates on these compounds were low, suggesting osmotrophy plays more of a survival strategy in low-light situations. Notably, a significant amount of the assimilated DOC was found in both particulate organic carbon and calcite coccoliths (particulate inorganic carbon) within 24 hours, suggesting that osmotrophic uptake of dissolved organics into calcite of coccolithophores is a small but notable&nbsp;part of both the biological carbon pump and alkalinity pump paradigms.</p>
<p><em><strong>Incubation experiments</strong></em><br />
A total of nine stations were visited during <em>R/V Endeavor</em> cruise EN616 in the northwest Atlantic during July 2018, but only four were used for incubation experiments (stations 1, 3, 5, and 9). The depth of maximum coccolithophore concentration was determined by sampling at eight depths and performing filter transfer freeze (FTF) coccolithophore counts (Hewes &amp; Holm-Hansen, 1983; Balch et al., 2023).&nbsp;The FTF technique is a semi-quantitative technique for microscopy and enumeration of phytoplankton onboard the ship&nbsp;to determine rough depth profiles of phytoplankton concentration (different from the more-quantitative birefringence counts done ashore, postcruise. See Related Dataset below).</p>
<p>Once the depth of&nbsp; maximum coccolithophore concentration was found, the Niskin bottles from that&nbsp;depth were drained and combined into a single, acid-cleaned polycarbonate carboy for the 24 hour incubation experiments with natural populations.&nbsp;At each station, 1 to 2 liters&nbsp;of ambient seawater from the combined carboy water sample were poured into 16 intravenous or “IV” flexible plastic bags (Thermo Fisher LabTainer&nbsp;BioProcess Container). Four bags each were used for incubations with each of the four&nbsp;radiolabeled compounds (16 bags total):</p>
<ul>
<li><sup>14</sup>C-acetate (specific activity = 1.924 MBq μmol-1 544; inoculum = 120μL per bag at 1.923 mM stock concentration),</li>
<li><sup>14</sup>C-mannitol (specific activity =&nbsp;2.146 MBq μmol-1; inoculum = 120μL per bag at 1.724 mM stock concentration),</li>
<li><sup>14</sup>C-glycerol&nbsp;(specific activity = 5.92 M Bq mol-1 547; inoculum = 120μL per bag at 0.625mM stock concentration), and</li>
<li><sup>14</sup>C-bicarbonate (specific activity =2.146 MBq μmol-1 548; inoculum = 200μL per bag at&nbsp;17.24mM stock concentration) (PerkinElmer, Waltham, MA, USA).</li>
</ul>
<p>See Balch et al. (2023) for detailed methodology, as well as final concentrations of radiolabeled and ambient DOI compounds for each experiment.&nbsp;&nbsp;</p>
<p>Following inoculation, all bags were gently mixed to disperse the radiolabeled compound.&nbsp;The bags were placed in an incubator with neutral-density screening to achieve the ambient light&nbsp;level from the collection depth and maintained at the collection temperature with day-night cycle&nbsp;adjusted for the collection location. One of each of the four replicate bags received 5% final&nbsp;concentration of buffered formalin and served as a killed control. Bags were incubated for 24 hours then brought into the darkened radioisotope van (with only dim red light). Bags were hung and&nbsp;100-200mL were withdrawn and immediately filtered as “bulk samples” on a 25mm diameter,&nbsp;0.4μm pore-size polycarbonate filter and subsequently processed using the microdiffusion&nbsp;technique (Balch et al., 2000)&nbsp;to determine the C14 activity in POC and PIC of the particles (Balch et al., 2023).&nbsp;</p>
<p><strong><em>Bulk phytoplankton measurements</em></strong></p>
<p>Bulk water samples from the I.V. bags were filtered onto a 0.4um-poresize polycarbonate filter, held in a 12-place Millipore filter tub under &lt;5mm Hg vacuum.&nbsp;&nbsp;Following filtration, each filter was rinsed three times with filtered seawater and then given a gentle “rim rinse” following the removal of the top filter holder, in order to remove any remaining dissolved&nbsp;<sup>14</sup>C activity from the moist filter.&nbsp;&nbsp;Each filter was then removed for the microdiffusion protocol&nbsp;(Balch et al., 2000; Paasche and Brubak, 1994).&nbsp;&nbsp;Briefly, each filter was placed on the bottom of a clean scintillation vial, sealed with a rubber septum which also held a suspended bucket containing a GFA filter saturated with 0.2 mL of phenethylamine (PEA is a CO<sub>2</sub>&nbsp;absorbent organic compound).&nbsp;&nbsp;One mL of 1% (by volume)&nbsp;&nbsp;phosphoric acid was injected through the rubber septum, past the suspended bucket, onto the original sample filter on the bottom of the scintillation vial to dissolve the PIC, converting it to CO<sub>2</sub>&nbsp;gas, which diffused into the headspace.&nbsp;&nbsp;This&nbsp;<sup>14</sup>C-CO<sub>2</sub>&nbsp;was absorbed onto the filter in the suspended bucket over the next 24 hours as the sealed scintillation vials were gently shaken on a shaker table.&nbsp;&nbsp;After 24 hours shaking, the vials were removed, septa opened within a fume hood, and the bucket containing the GFA filter with absorbed&nbsp;<sup>14</sup>C-CO<sub>2</sub>&nbsp;(originally&nbsp;<sup>14</sup>C-PIC) was snipped into a new, clean scintillation vial, to which scintillation cocktail was added (Ecolume; MP Biomedicals). Sample radioactivity was measured within two months following the cruise.</p>
<p><strong><em>Sample radioactivity measurements</em></strong></p>
<p>The radioactivity of the filters&nbsp;was measured using a Tri-Carb 3110TR time-resolved liquid scintillation counter (PerkinElmer), set for transformed spectral index of external standards. The instrument was coupled to automatic efficiency correction as well as background subtraction from each of the three spectral counting regions of the counter.&nbsp;&nbsp;Static electricity associated with each vial was eliminated by the counter.&nbsp;&nbsp;True decay events from the sample were defined as being within 18 ns&nbsp;(nanosecond)&nbsp;time difference for the two PMTs (photomultiplier tubes)&nbsp;monitoring a sample vial.&nbsp;&nbsp;The time period that the detector looks for additional pulses after the initial pulse (termed “after pulses”) was set to 75ns.&nbsp;&nbsp;Beyond this “after-pulse” time, the scintillation events were considered to be associated with background counts.&nbsp;&nbsp;&nbsp;To increase the precision of the radioactivity measurements, ten replicate counts were performed for each sample.&nbsp;&nbsp;Replicate sample counts continued until the final average count rate had an overall 95% confidence limit &lt;= ±0.5%.&nbsp;&nbsp;This was achieved after a total of 160,000 accumulated scintillation counts. If the accumulated counts did not reach that level in 10 minutes, counting of that replicate was terminated (but the count was still tabulated along with its confidence limit).&nbsp; This ten-replicate approach allowed an increase in precision, reducing the standard errors of the radioactivity measurements (in units of disintegrations per minute or DPMs) by a factor of 3 (=square root of (10-1)).</p>
<p>Uptake was calculated as the difference between the sample average count and its associated formalin-killed blank. Uptake was calculated as:</p>
<p>Uptake = [(DPM<sub>(s)</sub>-DPM<sub>(b)</sub>) x 1.05] /[DPM<sub>(tot)</sub>&nbsp;x (V<sub>(s)</sub>/V<sub>(tot)</sub>) x T<sub>(elap)</sub>]&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;</p>
<p>where 1.05 was the isotope discrimination factor for&nbsp;<sup>14</sup>C compared to&nbsp;<sup>12</sup>C, DPM<sub>(tot)</sub>&nbsp;was the radioactivity of the total counts added to the experimental sample, V<sub>(s)&nbsp;</sub>and V<sub>(tot)</sub>&nbsp;were the volumes of the experimental sample and the subsample for determination of the total activity, respectively, and T<sub>(elap)</sub>&nbsp;was the elapsed time between the moment the isotope was added until the sample was filtered.&nbsp;&nbsp;The radioactivity for the three experimental bags was then averaged within a treatment and standard deviation calculated.&nbsp; Cellular uptake into PIC for bulk phytoplankton samples was calculated by dividing the uptake rates into PIC (i.e. calcification rate) from the equation above&nbsp;by the concentration of coccolithophores in the sample.&nbsp;&nbsp;For flow cytometer samples, the coccolithophore-specific POC and PIC production rates were calculated by taking the results of the above equation and dividing by the numbers of coccolithophores sorted by the flow cytometer for each&nbsp;&nbsp;sample. For flow cytometry results, see the Related Datasets section below.</p>
<p>&nbsp;</p>
Funding provided by NSF Division of Ocean Sciences (NSF OCE) Award Number: OCE-1635748 Award URL: https://www.nsf.gov/awardsearch/show-award?AWD_ID=1635748
completed
William M. Balch
Bigelow Laboratory for Ocean Sciences
207-315-2567
60 Bigelow Drive
East Boothbay
ME
04544
USA
bbalch@bigelow.org
pointOfContact
Stephen D. Archer
Bigelow Laboratory for Ocean Sciences
207-350-0791
60 Bigelow Drive
East Boothbay
ME
04554
USA
sarcher@bigelow.org
pointOfContact
David T. Drapeau
Bigelow Laboratory for Ocean Sciences
207-315-2567
60 Bigelow Drive
East Boothbay
ME
04544
USA
ddrapeau@bigelow.org
pointOfContact
Jelena Godrijan
Bigelow Laboratory for Ocean Sciences
+385-1-456-1148
Ruder Boskovic Institute Bijenicka cesta 54
10000 Zagreb
Croatia
jelena.godrijan@irb.hr
pointOfContact
asNeeded
Dataset Version: 1
Unknown
Start_Time_UTC
End_Time_UTC
Latitude
Longitude
Experiment_ID
Sample_num
Compound
Incubation
Type
SMPL_ID
Count_Time
CPMA
DPM1
Count_Precision
Coeff_Var
Calcification
Comments
Start_Time_EDT
End_Time_EDT
Tri-Carb 3110TR time-resolved liquid scintillation counter
shaker table
theme
None, User defined
date
latitude
longitude
experiment id
sample identification
sample type
radioactive_isotope_activity
No BCO-DMO term
flag
comments
featureType
BCO-DMO Standard Parameters
Liquid Scintillation Counter
Shaker
instrument
BCO-DMO Standard Instruments
EN616
service
Deployment Activity
Gulf of Maine and North West Atlantic
place
Locations
otherRestrictions
otherRestrictions
Access Constraints: none. Use Constraints: Please follow guidelines at: http://www.bco-dmo.org/terms-use Distribution liability: Under no circumstances shall BCO-DMO be liable for any direct, incidental, special, consequential, indirect, or punitive damages that result from the use of, or the inability to use, the materials in this data submission. If you are dissatisfied with any materials in this data submission your sole and exclusive remedy is to discontinue use.
Coccolithophore Mixotrophy
https://osprey.bco-dmo.org/project/648016
Coccolithophore Mixotrophy
<p><strong>NSF Award Abstract</strong><br />
Coccolithophores are single-cell algae that are covered with limestone (calcite) plates called coccoliths. They may make up most of the phytoplankton biomass in the oceans. Coccolithophores are generally considered to be autotrophs, meaning that they use photosynthesis to fix carbon into both soft plant tissue and hard minerogenic calcite, using sunlight as an energy source ("autotrophic"). However, there is an increasing body of evidence that coccolithophores are "mixotrophic", meaning that they can fix carbon from photosynthesis as well as grow in darkness by engulfing small organic particles plus taking up other simple carbon molecules from seawater. The extent to which Coccolithophores engage in mixotrophy can influence the transfer of carbon into the deep sea. This work is fundamentally directed at quantifying coccolithophore mixotrophy -- the ability to use dissolved and reduce carbon compounds for energy -- using lab and field experiments plus clarifying its relevance to ocean biology and chemistry. This work will generate broader impacts in three areas: 1) Undergraduate training: Two REU undergraduates will be trained during the project. The student in the second year will participate in the research cruise. 2) Café Scientifique program: This work will be presented in Bigelow Laboratory’s Café Scientifique program. These are free public gatherings where the public is invited to join in a conversation about the latest ideas and issues in ocean science and technology. 3) Digital E-Book: We propose to make a digital E-book to specifically highlight and explain mixotrophy within coccolithophores. Images of mixotrophic coccolithophores would be the primary visual elements of the book. The E-book will be publicly available and distributed to our educational affiliate, Colby College. The goal of the book is to further communicate the intricacies of the microbial world, food web dynamics, plus their relationship to the global carbon cycle, to inspire interest, education, and curiosity about these amazing life forms.</p>
<p>Coccolithophores can significantly affect the draw-down of atmospheric CO2 and they can transfer CO2 from the surface ocean and sequester it in the deep sea via two carbon pump mechanisms: (1) The "alkalinity pump" (also known as the calcium carbonate pump), where coccolithophores in the surface ocean take up dissolved inorganic carbon (DIC; primarily a form called bicarbonate, a major constituent of ocean alkalinity). They convert half to CO2, which is either fixed as plant biomass or released as the gas, and half is synthesized into their mineral coccoliths. Thus, coccolithophore calcification can actually increase surface CO2 on short time scales (i.e. weeks). However, over months to years, coccoliths sink below thousands of meters, where they dissolve and release bicarbonate back into deep water. Thus, sinking coccoliths essentially "pump" bicarbonate alkalinity from surface to deep waters, where that carbon remains isolated in the abyssal depths for thousands of years. (2) The "biological pump", where the ballasting effect of the dense limestone coccoliths speeds the sinking of organic, soft-tissue debris (particulate organic carbon or POC), essentially "pumping" this soft carbon tissue to depth. The biological pump ultimately decreases surface CO<sub>2</sub>. The soft-tissue and alkalinity pumps reinforce each other in maintaining a vertical gradient in DIC (more down deep than at the surface) but they oppose each other in terms of the air-sea exchange of CO<sub>2</sub>. Thus, the net effect of coccolithophores on atmospheric CO2 depends on the balance of their CO<sub>2</sub>-raising effect associated with the alkalinity pump and their CO2-lowering effect associated with the soft-tissue biological pump. It is virtually always assumed that coccolith particulate inorganic carbon (PIC) originates exclusively from dissolved inorganic carbon (DIC, as bicarbonate), not dissolved organic carbon (DOC). The goal of this proposal is to describe a) the potential uptake and assimilation of an array of DOC compounds by coccolithophores, b) the rates of uptake, and potential incorporation of DOC by coccolithophores into PIC coccoliths, which, if true, would represent a major shift in the alkalinity pump paradigm. This work is fundamentally directed at quantifying coccolithophore mixotrophy using lab and field experiments plus clarifying its relevance to ocean biology and chemistry. There have been a number of technological advances to address this issue, all of which will be applied in this work. The investigators will: (a) screen coccolithophore cultures for the uptake and assimilation of a large array of DOC molecules, (b) perform tracer experiments with specific DOC molecules in order to examine uptake at environmentally-realistic concentrations, (c) measure fixation of DOC into organic tissue, separately from that fixed into PIC coccoliths, (d) separate coccolithophores from other phytoplankton and bacteria using flow cytometry and e) distinguish the modes of nutrition in these sorted coccolithophore cells. This work will fundamentally advance the state of knowledge of coccolithophore mixotrophy in the sea and address the balance of carbon that coccolithophores derived from autotrophic versus heterotrophic sources.</p>
Cocco-Mix
largerWorkCitation
project
eng; USA
biota
oceans
Gulf of Maine and North West Atlantic
-72.92708
-66.51748
36.98572
43.71835
2018-07-05
2018-07-13
Partially lab-based, with field sites in Gulf of Maine and NW Atlantic between the Gulf of Maine and Bermuda
0
BCO-DMO catalogue of parameters from DOC uptake rates by coccolithophores and scintillation counts from field experiments in the North Atlantic during R/V Endeavor cruise EN616 in July 2018
Biological and Chemical Oceanography Data Management Office (BCO-DMO)
Unavailable
508-289-2009
WHOI MS#36
Woods Hole
MA
02543
USA
info@bco-dmo.org
http://www.bco-dmo.org
Monday - Friday 8:00am - 5:00pm
For questions regarding this resource, please contact BCO-DMO via the email address provided.
pointOfContact
http://lod.bco-dmo.org/id/dataset-parameter/892372.rdf
Name: Start_Time_UTC
Units: unitless
Description: <p>Start datetime of incubation experiment (time of inoculation) in UTC with ISO8601 format</p>
http://lod.bco-dmo.org/id/dataset-parameter/892373.rdf
Name: End_Time_UTC
Units: unitless
Description: <p>End datetime of incubation experiment (time of filtration or sorting) in UTC with ISO8601 format</p>
http://lod.bco-dmo.org/id/dataset-parameter/892374.rdf
Name: Latitude
Units: decimal degrees
Description: <p>Latitude of incubation station</p>
http://lod.bco-dmo.org/id/dataset-parameter/892375.rdf
Name: Longitude
Units: decimal degrees
Description: <p>Longitude of incubation station</p>
http://lod.bco-dmo.org/id/dataset-parameter/892376.rdf
Name: Experiment_ID
Units: unitless
Description: <p>Experiment ID</p>
http://lod.bco-dmo.org/id/dataset-parameter/892377.rdf
Name: Sample_num
Units: unitless
Description: <p>Sample number</p>
http://lod.bco-dmo.org/id/dataset-parameter/892378.rdf
Name: Compound
Units: unitless
Description: <p>Organic compound being measured (acetate, mannitol, glycerol, bicarbonate)</p>
http://lod.bco-dmo.org/id/dataset-parameter/892379.rdf
Name: Incubation
Units: unitless
Description: <p>Incubation number where 1 to 3 are replicates and 4 is formalin-treated incubation</p>
http://lod.bco-dmo.org/id/dataset-parameter/892380.rdf
Name: Type
Units: unitless
Description: <p>Type of sample (bulk or flow cytometer sorted)</p>
http://lod.bco-dmo.org/id/dataset-parameter/892381.rdf
Name: SMPL_ID
Units: unitless
Description: <p>Sample ID</p>
http://lod.bco-dmo.org/id/dataset-parameter/892382.rdf
Name: Count_Time
Units: minutes
Description: <p>Scintillation counting time</p>
http://lod.bco-dmo.org/id/dataset-parameter/892383.rdf
Name: CPMA
Units: counts per minute
Description: <p>Counts per minute of activity</p>
http://lod.bco-dmo.org/id/dataset-parameter/892384.rdf
Name: DPM1
Units: disintegrations per minute (DPM)
Description: <p>Disintengrations per minute calculated after taking counting efficiency into consideration</p>
http://lod.bco-dmo.org/id/dataset-parameter/892385.rdf
Name: Count_Precision
Units: percent
Description: <p>A:2S%; Count precision as + or - percent for two sigma error</p>
http://lod.bco-dmo.org/id/dataset-parameter/892386.rdf
Name: Coeff_Var
Units: percent
Description: <p>%CV:DPM1; Coefficient of variation for counts</p>
http://lod.bco-dmo.org/id/dataset-parameter/892387.rdf
Name: Calcification
Units: unitless
Description: <p>Indicator for calcification process. Entries with no C indicate incorporation into organic matter.</p>
http://lod.bco-dmo.org/id/dataset-parameter/892388.rdf
Name: Comments
Units: unitless
Description: <p>Comments about sample, experiment, or other relevant information</p>
http://lod.bco-dmo.org/id/dataset-parameter/892389.rdf
Name: Start_Time_EDT
Units: unitless
Description: <p>Start datetime of incubation experiment (time of inoculation) in EDT time zone</p>
http://lod.bco-dmo.org/id/dataset-parameter/892390.rdf
Name: End_Time_EDT
Units: unitless
Description: <p>End datetime of incubation experiment (time of filtration or sorting) in EDT time zone</p>
GB/NERC/BODC > British Oceanographic Data Centre, Natural Environment Research Council, United Kingdom
Biological and Chemical Oceanography Data Management Office (BCO-DMO)
Unavailable
508-289-2009
WHOI MS#36
Woods Hole
MA
02543
USA
info@bco-dmo.org
http://www.bco-dmo.org
Monday - Friday 8:00am - 5:00pm
For questions regarding this resource, please contact BCO-DMO via the email address provided.
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653665
https://darchive.mblwhoilibrary.org/server/api/core/bitstreams/d2a246ea-b529-4179-939f-c1a65c5ce9e1/content
download
https://doi.org/10.26008/1912/bco-dmo.887562.1
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onLine
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<p>Coccolithophores are typically thought of as photoautotrophs yet they are known to inhabit sub-euphotic environments with insufficient light for photosynthesis. Field experiments were performed in the NW Atlantic to determine the potential uptake of dissolved organic carbon (DOC) by coccolithophores to compare with the bicarbonate uptake associated with standard photosynthesis and calcification.&nbsp; The cruise sampled portions of the Gulf of Maine, Georges Bank, continental shelf and slope waters of the northern part of the mid-Atlantic Bight plus Sargasso Sea, during <em>R/V Endeavor</em> cruise EN616 in July&nbsp;2018.&nbsp; Natural phytoplankton populations were incubated with radiolabeled DOC compounds, <sup>14</sup>C-acetate, <sup>14</sup>C-mannitol and <sup>14</sup>C-glycerol.&nbsp; Following the 24 hour incubations at simulated <em>in situ</em> conditions of light and temperature, coccolithophores were sorted from these natural populations using flow cytometry (based on their chlorophyll fluorescence and birefringence signatures).&nbsp; The sorts were collected and the uptake rate of these compounds into both organic and inorganic fractions was subsequently measured using the microdiffusion technique coupled with high-precision scintillation counting.&nbsp; Bulk samples were also filtered from these incubated natural populations in which radiolabeled DOC was taken-up and assimilated into the particulate organic carbon of general phytoplankton (not just coccolithophores) as well as the PIC of the broader assemblage of calcifiers (likely dominated by coccolithophores based on the small sample volumes).&nbsp; &nbsp;The bulk samplings had higher signal-to-noise for radioactivity measurements of DOC uptake into coccolithophore PIC due to the increased sample size, whereas the flow cytometer sorts had higher specificity for isolating coccolithophores, which allowed better discerning the fixation of <sup>14</sup>C-labeled DOC into both organic tissues and calcium carbonate (particulate inorganic carbon) of coccolithophores, despite the lower signal-to-noise.&nbsp;DOC uptake rates were measurable but slow relative to bicarbonate uptake rates and the resultant growth rates on these compounds were low, suggesting osmotrophy plays more of a survival strategy in low-light situations. Notably, a significant amount of the assimilated DOC was found in both particulate organic carbon and calcite coccoliths (particulate inorganic carbon) within 24 hours, suggesting that osmotrophic uptake of dissolved organics into calcite of coccolithophores is a small but notable&nbsp;part of both the biological carbon pump and alkalinity pump paradigms.</p>
<p><em><strong>Incubation experiments</strong></em><br />
A total of nine stations were visited during <em>R/V Endeavor</em> cruise EN616 in the northwest Atlantic during July 2018, but only four were used for incubation experiments (stations 1, 3, 5, and 9). The depth of maximum coccolithophore concentration was determined by sampling at eight depths and performing filter transfer freeze (FTF) coccolithophore counts (Hewes &amp; Holm-Hansen, 1983; Balch et al., 2023).&nbsp;The FTF technique is a semi-quantitative technique for microscopy and enumeration of phytoplankton onboard the ship&nbsp;to determine rough depth profiles of phytoplankton concentration (different from the more-quantitative birefringence counts done ashore, postcruise. See Related Dataset below).</p>
<p>Once the depth of&nbsp; maximum coccolithophore concentration was found, the Niskin bottles from that&nbsp;depth were drained and combined into a single, acid-cleaned polycarbonate carboy for the 24 hour incubation experiments with natural populations.&nbsp;At each station, 1 to 2 liters&nbsp;of ambient seawater from the combined carboy water sample were poured into 16 intravenous or “IV” flexible plastic bags (Thermo Fisher LabTainer&nbsp;BioProcess Container). Four bags each were used for incubations with each of the four&nbsp;radiolabeled compounds (16 bags total):</p>
<ul>
<li><sup>14</sup>C-acetate (specific activity = 1.924 MBq μmol-1 544; inoculum = 120μL per bag at 1.923 mM stock concentration),</li>
<li><sup>14</sup>C-mannitol (specific activity =&nbsp;2.146 MBq μmol-1; inoculum = 120μL per bag at 1.724 mM stock concentration),</li>
<li><sup>14</sup>C-glycerol&nbsp;(specific activity = 5.92 M Bq mol-1 547; inoculum = 120μL per bag at 0.625mM stock concentration), and</li>
<li><sup>14</sup>C-bicarbonate (specific activity =2.146 MBq μmol-1 548; inoculum = 200μL per bag at&nbsp;17.24mM stock concentration) (PerkinElmer, Waltham, MA, USA).</li>
</ul>
<p>See Balch et al. (2023) for detailed methodology, as well as final concentrations of radiolabeled and ambient DOI compounds for each experiment.&nbsp;&nbsp;</p>
<p>Following inoculation, all bags were gently mixed to disperse the radiolabeled compound.&nbsp;The bags were placed in an incubator with neutral-density screening to achieve the ambient light&nbsp;level from the collection depth and maintained at the collection temperature with day-night cycle&nbsp;adjusted for the collection location. One of each of the four replicate bags received 5% final&nbsp;concentration of buffered formalin and served as a killed control. Bags were incubated for 24 hours then brought into the darkened radioisotope van (with only dim red light). Bags were hung and&nbsp;100-200mL were withdrawn and immediately filtered as “bulk samples” on a 25mm diameter,&nbsp;0.4μm pore-size polycarbonate filter and subsequently processed using the microdiffusion&nbsp;technique (Balch et al., 2000)&nbsp;to determine the C14 activity in POC and PIC of the particles (Balch et al., 2023).&nbsp;</p>
<p><strong><em>Bulk phytoplankton measurements</em></strong></p>
<p>Bulk water samples from the I.V. bags were filtered onto a 0.4um-poresize polycarbonate filter, held in a 12-place Millipore filter tub under &lt;5mm Hg vacuum.&nbsp;&nbsp;Following filtration, each filter was rinsed three times with filtered seawater and then given a gentle “rim rinse” following the removal of the top filter holder, in order to remove any remaining dissolved&nbsp;<sup>14</sup>C activity from the moist filter.&nbsp;&nbsp;Each filter was then removed for the microdiffusion protocol&nbsp;(Balch et al., 2000; Paasche and Brubak, 1994).&nbsp;&nbsp;Briefly, each filter was placed on the bottom of a clean scintillation vial, sealed with a rubber septum which also held a suspended bucket containing a GFA filter saturated with 0.2 mL of phenethylamine (PEA is a CO<sub>2</sub>&nbsp;absorbent organic compound).&nbsp;&nbsp;One mL of 1% (by volume)&nbsp;&nbsp;phosphoric acid was injected through the rubber septum, past the suspended bucket, onto the original sample filter on the bottom of the scintillation vial to dissolve the PIC, converting it to CO<sub>2</sub>&nbsp;gas, which diffused into the headspace.&nbsp;&nbsp;This&nbsp;<sup>14</sup>C-CO<sub>2</sub>&nbsp;was absorbed onto the filter in the suspended bucket over the next 24 hours as the sealed scintillation vials were gently shaken on a shaker table.&nbsp;&nbsp;After 24 hours shaking, the vials were removed, septa opened within a fume hood, and the bucket containing the GFA filter with absorbed&nbsp;<sup>14</sup>C-CO<sub>2</sub>&nbsp;(originally&nbsp;<sup>14</sup>C-PIC) was snipped into a new, clean scintillation vial, to which scintillation cocktail was added (Ecolume; MP Biomedicals). Sample radioactivity was measured within two months following the cruise.</p>
<p><strong><em>Sample radioactivity measurements</em></strong></p>
<p>The radioactivity of the filters&nbsp;was measured using a Tri-Carb 3110TR time-resolved liquid scintillation counter (PerkinElmer), set for transformed spectral index of external standards. The instrument was coupled to automatic efficiency correction as well as background subtraction from each of the three spectral counting regions of the counter.&nbsp;&nbsp;Static electricity associated with each vial was eliminated by the counter.&nbsp;&nbsp;True decay events from the sample were defined as being within 18 ns&nbsp;(nanosecond)&nbsp;time difference for the two PMTs (photomultiplier tubes)&nbsp;monitoring a sample vial.&nbsp;&nbsp;The time period that the detector looks for additional pulses after the initial pulse (termed “after pulses”) was set to 75ns.&nbsp;&nbsp;Beyond this “after-pulse” time, the scintillation events were considered to be associated with background counts.&nbsp;&nbsp;&nbsp;To increase the precision of the radioactivity measurements, ten replicate counts were performed for each sample.&nbsp;&nbsp;Replicate sample counts continued until the final average count rate had an overall 95% confidence limit &lt;= ±0.5%.&nbsp;&nbsp;This was achieved after a total of 160,000 accumulated scintillation counts. If the accumulated counts did not reach that level in 10 minutes, counting of that replicate was terminated (but the count was still tabulated along with its confidence limit).&nbsp; This ten-replicate approach allowed an increase in precision, reducing the standard errors of the radioactivity measurements (in units of disintegrations per minute or DPMs) by a factor of 3 (=square root of (10-1)).</p>
<p>Uptake was calculated as the difference between the sample average count and its associated formalin-killed blank. Uptake was calculated as:</p>
<p>Uptake = [(DPM<sub>(s)</sub>-DPM<sub>(b)</sub>) x 1.05] /[DPM<sub>(tot)</sub>&nbsp;x (V<sub>(s)</sub>/V<sub>(tot)</sub>) x T<sub>(elap)</sub>]&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp;</p>
<p>where 1.05 was the isotope discrimination factor for&nbsp;<sup>14</sup>C compared to&nbsp;<sup>12</sup>C, DPM<sub>(tot)</sub>&nbsp;was the radioactivity of the total counts added to the experimental sample, V<sub>(s)&nbsp;</sub>and V<sub>(tot)</sub>&nbsp;were the volumes of the experimental sample and the subsample for determination of the total activity, respectively, and T<sub>(elap)</sub>&nbsp;was the elapsed time between the moment the isotope was added until the sample was filtered.&nbsp;&nbsp;The radioactivity for the three experimental bags was then averaged within a treatment and standard deviation calculated.&nbsp; Cellular uptake into PIC for bulk phytoplankton samples was calculated by dividing the uptake rates into PIC (i.e. calcification rate) from the equation above&nbsp;by the concentration of coccolithophores in the sample.&nbsp;&nbsp;For flow cytometer samples, the coccolithophore-specific POC and PIC production rates were calculated by taking the results of the above equation and dividing by the numbers of coccolithophores sorted by the flow cytometer for each&nbsp;&nbsp;sample. For flow cytometry results, see the Related Datasets section below.</p>
<p>&nbsp;</p>
Specified by the Principal Investigator(s)
<p><em><strong>Designations for experiments and blanks</strong></em></p>
<ul>
<li>P designates incorporation of radiolabeled compound into cellular organic matter</li>
<li>C designates incorporation of radiolabeled compound into particulate inorganic carbon (calcium carbonate)</li>
<li>TOT designates the total radioactivity of the compound added to the incubation at time-zero (usually sampled in a 100 microliter volume)</li>
<li>TOP/BOT designate radioactive counts in 1) the upper bucket (TOP) with PEA CO2 trap or 2) bottom&nbsp; (BOT) of the scintillation vial (as organic carbon)</li>
<li>PEA designates the radioactive counts associated with a 0.2 mL addition of phenethylamine (placed in the upper hanging bucket as a CO2 trap). This is done as a type of procedural blank.</li>
<li>PEA TIP designates the radioactive counts associated with the tip of the PEA dispenser, typically measured at the end of the experiment.&nbsp; This was simply a check to make sure that <sup>14</sup>C-CO2 within the fume hood had not been adsorbed onto the tip of the PEA dispenser, thereby becoming a source of potential contamination in the low-level scintillation counts.</li>
<li>BLANK designates a blank filter</li>
</ul>
<p><strong>BCO-DMO processing</strong><br />
- Data is concatenated from the 'raw counts and calc' tabs on each of the four separate files of EN616 isotope results.<br />
-&nbsp;Modified parameter (column) names to conform with BCO-DMO naming conventions.&nbsp;<br />
- Added latitudes and longitudes to match Stations 1, 3, 5, and 9.<br />
- Added sampling datetimes for the four stations from 'reduced' tab on the isotope results files which indicated times of inoculation, filtration, and sorting of the incubation samples<br />
- Converted datetimes to UTC ISO8601 format.<br />
-&nbsp;Added columns for&nbsp;identification numbers and processes captured&nbsp;within the&nbsp;Sample ID<br />
- Separated statistics portion from the scintillation counts and served as a Supplemental document</p>
Specified by the Principal Investigator(s)
asNeeded
7.x-1.1
Biological and Chemical Oceanography Data Management Office (BCO-DMO)
Unavailable
508-289-2009
WHOI MS#36
Woods Hole
MA
02543
USA
info@bco-dmo.org
http://www.bco-dmo.org
Monday - Friday 8:00am - 5:00pm
For questions regarding this resource, please contact BCO-DMO via the email address provided.
pointOfContact
Tri-Carb 3110TR time-resolved liquid scintillation counter
Tri-Carb 3110TR time-resolved liquid scintillation counter
PI Supplied Instrument Name: Tri-Carb 3110TR time-resolved liquid scintillation counter PI Supplied Instrument Description:The radioactivity of the filters was measured using a Tri-Carb 3110TR time-resolved liquid scintillation counter (PerkinElmer), set for transformed spectral index of external standards. Instrument Name: Liquid Scintillation Counter Instrument Short Name:LSC 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 to 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. Community Standard Description: http://vocab.nerc.ac.uk/collection/L05/current/LAB21/
shaker table
shaker table
PI Supplied Instrument Name: shaker table PI Supplied Instrument Description:The sealed scintillation vials were gently shaken on a shaker table for 24 hours Instrument Name: Shaker Instrument Short Name: Instrument Description: A Shaker is a piece of lab equipment used to mix, blend, or to agitate substances in tube(s) or flask(s) by shaking them, which is mainly used in the fields of chemistry and biology. A shaker contains an oscillating board which is used to place the flasks, beakers, test tubes, etc.
Cruise: EN616
EN616
R/V Endeavor
Community Standard Description
International Council for the Exploration of the Sea
R/V Endeavor
vessel
EN616
William M. Balch
Bigelow Laboratory for Ocean Sciences
R/V Endeavor
Community Standard Description
International Council for the Exploration of the Sea
R/V Endeavor
vessel