Data profile chemoautotrophic assimilation of inorganic carbon by the microbial communities through the water column of the Eastern Tropical North Pacific’s Oxygen Minimum Zone. Samples were collected on various dates in February and March of 2023 from two stations. Niskin bottle-collected whole water samples were collected in 1L bottles, dispensed into incubation septa test tubes and spiked with 14C-bicarbonate. One depth per sample set was selected for stimulation experiments. At these selected depths, replicates were additionally spiked with alternative electron donors. The unstimulated replicates from the same depth were size fractionated into ‘free living’ and ‘particle associated’ size fractions. Samples were transported to the home laboratory, suspended in scintillation cocktail and radioassayed. Samples were collected and prepared by Natalie Butkevich, Dr. Gordon Taylor and Dr. Elena Yakubovskaya. Samples were processed by Natalie Butkevich and Dr. Gordon Taylor.

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

1L whole water samples were collected on deck under nitrogen pressure at the rosette. A three-volume overflow was performed for each sample to minimize oxygen contamination. Bottles were sealed with deoxygenated butyl rubber septa after confirming all bubbles were excluded. A He gas headspace was added in the shipboard lab no more than 15 minutes after collection. To quantify community chemoautotrophic activity, triplicate incubations with ¹⁴C-bicarbonate were prepared in 16 mL Hungate tubes with degassed butyl rubber septa under nitrogen pressure. Chilled N2-purged ¹⁴C-bicarbonate in an alkaline brine (pH 9.5; S = 60 on the practical salinity scale) was injected (0.92 µCi Mex1, 0.54 µCi Mex2) into the bottom before sealing (Tuttle and Jannasch 1973a). One depth per cast was selected for stimulation experiments with alternative electron donors. Selected incubations were spiked with ¹⁴C-bicarbonate and one of the following: NH₄⁺, DMSP, H₂S or S₂O₃. Incubations were conducted in a dark ice chest for 18-24 hours and terminated by filtration onto 25 mm nitrocellulose 0.22 µm filters (25 mm diameter Osmonics). Selected non-amended incubations were size fractionated into a “particle-associated” fraction (>2.7 µm) and a free-living size fraction (0.22 – 2.7 µm) by sequential filtration through stacked 25 mm Swinnex filter holders. Unassimilated ¹⁴C-bicarbonate was purged from the filters in a saturated HCl atmosphere enclosure for >1h in a fume hood. Filters were then dried, suspended in scintillation cocktail (Hionic-Fluor) and radioassayed. Table 1 summarizes sample types. Data were corrected for isotopic fractionation (multiplied by 1.06) and for nonbiological sorption by use of control samples processed immediately after introduction of the radiotracer (T₀ samples). If blanks were higher than T₀ samples, blanks were used instead. Three T₀ sample sets were made, one for standard+stimulation experiments, one for the free-living size fraction and one for the particle-associated size fraction. Rates of dark ¹⁴C-assimilation were normalized to total nanomoles of carbon per day via equation detailed under 'Data Processing' section.

Sample code	Tracer/Amendments	Experiment type	Notes
standard	14C-bicarbonate	Standard
S2O3	14C-bicarbonate and S2O3	Alternative electron donor amendment	Target stimulant concentration: 10uM. For all samples prepared Feb.19 or later, target concentration is 5uM
DMSP	14C-bicarbonate and DMSP	Alternative electron donor amendment	Target stimulant concentration: 100nM. For all samples prepared Feb.19 or later, target concentration is 50nM
NH4+	14C-bicarbonate and NH4+	Alternative electron donor amendment	Target stimulant concentration: 10uM. For all samples prepared Feb.19 or later, target concentration is 5uM
H2S	14C-bicarbonate and H2S	Alternative electron donor amendment	Target stimulant concentration: 10uM. For all samples prepared Feb.19 or later, target concentration is 5uM
0.2_SF	14C-bicarbonate	Size fractionation
2.7_SF	14C-bicarbonate	Size fractionation

Assimilation equation: 

rate (nanomole carbon per liter per day) = (((2*10^6) * 1.06 * max(DPM - blank or t0, 0))/specific activity value)*(24/incubation time)

 

Constants: 

2*10^6 = dissolved inorganic carbon in seawater (from Feely et al. 1995)

1.06 = isotopic fractionation factor

29 = standard blank in DPM

30 = small size fraction blank in DPM

37 = large size fraction blank in DPM

2032708 = specific activity for mex1 in DPM, 0.916 microCurie injected per sample

1190572 = specific activity for mex2 in DPM, 0.536 microCurie injected per sample

 

 

Notes: 

-separate specific activities were used for Mex1 and Mex2 samples because separate stocks were prepared. The same blank values were used across all samples.

-max(DPM-blank or t0,0) implies that if the blank/t0 subtraction resulted in a negative value, 0 was substituted instead. 

Curation Actions Performed on Data
- Loaded data from "Chemoautotrophic assimilation of inorganic carbon_FINAL.xlsx" (sheet 1), using xlsx format with formatting preservation and floating point error adjustment enabled, headers on row 1, blank headers ignored, empty rows removed, and empty string and "nd" values treated as missing; table named 1005988_v1_ETNP_OMZ_AT50_Chemoautotrophic_Assimilation_of_Inorganic_Carbon
- Created new column ISO_Cast_Start_DateTime_UTC by combining Cast Date (UTC) and Cast Time (UTC) columns, parsing with formats %Y-%m-%d and %H:%M:%S respectively, treating input as UTC, and outputting as datetime in format %Y-%m-%dT%H:%M:%SZ, also in UTC
- Renamed columns: "Sample Type" to Sample_Type, "Cast Date (UTC)" to Cast_Date_UTC, "Cast Time (UTC)" to Cast_Time_UTC, "CTD Cast Filename" to CTD_Cast_Filename, "Incubation Duration" to Incubation_Duration, "Raw DPM" to Raw_DPM, "Assimilation_nMC_d^-1" to Assimilation_nMC, "Assimilation_sd_nMC_d^-1" to Assimilation_sd_nMC, "Replicate ID" to Replicate_ID
- Reordered columns to: Depth, Sample_Type, Station, Lat, Long, ISO_Cast_Start_DateTime_UTC, Cast_Date_UTC, Cast_Time_UTC, CTD_Cast_Filename, Incubation Start (local ship time), Incubation Stop (local ship time), Incubation_Duration, Raw_DPM, Assimilation_nMC, Assimilation_sd_nMC, Replicate_ID
- Set column types and formats: Depth and Raw_DPM as integer; Sample_Type, CTD_Cast_Filename, Replicate_ID, Station as string; Lat, Long, Incubation_Duration, Assimilation_nMC, Assimilation_sd_nMC as number; Cast_Date_UTC as date (format %Y-%m-%d); Cast_Time_UTC as time (format %H:%M:%S); ISO_Cast_Start_DateTime_UTC as datetime (format %Y-%m-%dT%H:%M:%SZ); Incubation Start (local ship time) and Incubation Stop (local ship time) as datetime (format %Y-%m-%d %H:%M)
- Updated column metadata with standard name IDs, descriptions, and supplied units for all columns, including marking Depth, ISO_Cast_Start_DateTime_UTC, Lat, and Long as primary parameters
- Dumped final table to CSV output named 1005988_v1_ETNP_OMZ_AT50_Chemoautotrophic_Assimilation_of_Inorganic_Carbon.csv

Curation Actions Performed on Metadata
- Standard metadata entry and text formatting steps were performed.

No problems or issues have been reported by the authors for this dataset.

Coverage: East Tropical North Pacific Ocean

NSF Award Abstract:

Oxygen availability shapes the distributions and activities of marine organisms. Ongoing human activities and climate change are expected to lead to expansion and intensification of already large oxygen-stressed areas of the coastal and open ocean. Decreases in ocean oxygen have significant ecological consequences, including habitat loss for migratory and bottom-dwelling organisms, modification of the marine food web, and production of trace gases with pronounced feedbacks on climate, such as methane and nitrous oxide. Intense chemical cycling by microorganisms occurs in oxygen-depleted marine habitats. However, a full understanding of the consequences for marine ecosystems is hampered by limited knowledge of actual rates of key microbiological processes and dynamics of the microorganisms mediating them. This study combines novel methods and sampling techniques to understand how these processes are influenced by changes in oxygen concentration to inform predictions of important chemical exchanges within a changing ocean and its production of climate-active gases. This deeply collaborative project trains undergraduates (four of whom participate on the cruise), a graduate student and a postdoctoral fellow. Outreach takes place in middle and high schools and through social media. Data and samples from the cruise are integrated in coursework.

Oxygen depletion alters cycling of major elements (especially carbon, nitrogen, and sulfur) as well as food web functionality. This project addresses major gaps in our knowledge of oxygen minimum zone (OMZ) processes by applying in situ approaches to more accurately measure rates of several key microbial processes (chemoautotrophy, denitrification, anammox, sulfate reduction and sulfide oxidation) central to marine biogeochemical cycling. This work studies the Eastern Tropical North Pacific OMZ, the largest open ocean oxygen-depleted system, to 1) determine the in situ rates of microbial processes involved in carbon, nitrogen, and sulfur cycling, 2) reveal the genomic blueprint of active single cells involved in these processes, and 3) obtain estimates of the relative contributions of the dominant chemoautotrophic and heterotrophic groups to the measured rates. This work include applies cutting-edge equipment for in situ sampling and incubations that minimize artifacts associated with traditional water sampling approaches, allowing more accurate estimates of rates of important biogeochemical processes. Additionally, rate measurements of relatively undisturbed bulk and fractionated water samples make it easier to distinguish the potential role of particle-associated microorganisms in these OMZ processes. Single cell sorting of microorganisms using a fluorescent dye indicative of cell activity together with metatranscriptomics informs on metabolic pathways used for key processes by active microbial community members, as well as the potential coupling of chemoautotrophy and nitrogen or/and sulfur cycling. By combining stable isotope probing, fluorescence in situ hybridization and single cell Raman microspectrometry the relative activity levels of different microbial phylotypes involved in chemoautotrophic and heterotrophic elemental cycling are assessed.

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.