Table of Contents

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

One pair of 1m² MOCNESS (Multiple Open Closing Nets with Environmental Sensor System) tows were performed during each cruise- one during the day, and one at night (MOCNESS, Wiebe et al, 1985). Nets with 150um mesh were used to better capture the smaller midwater zooplankton community in the region. Eight nets were fired in sequence along the upcast to capture spatially discrete zooplankton samples between 600m and the surface. While nets one, two, and three consistently targeted depths of 600-500m, 500m-400m, and 400-300m, depths for nets four through eight varied based on hydrographic features including the thermocline, deep chlorophyll maximum, and oxygen minimum zone (Maas et al, 2014, Steinberg et al, 2008).  Once onboard, samples were split in two using a Motoda splitter (Motoda, 1959)  with half preserved with sodium tetraborate buffered 4% formalin in seawater to be scanned with a ZooSCAN (Gorsky et al, 2010) and half placed in 95% undenatured ethanol for metabarcoding. 

A representative subsample of the formalin-preserved zooplankton community from each net were imaged using a ZooSCAN ver. 4 at either 4,800 dpi or 2,400 dpi (following the methods in: Gorsky et al., 2010, Vandromme et al., 2012 as detailed in Maas et al. 2021). The change in resolution partway through the project was a result of recommendations from Hydroptic and loss of software support for 4800dpi imaging. In order to better represent all size classes in the images, the original sample was divided into three size categories. All individuals larger than 2 cm were selected by eye and scanned separately from all the others (fraction "d1"). The remainder of the sample was sieved through a 1-mm mesh sieve, and both size fractions were individually scanned ("d2" >1000um, "d3" 153-1000um). From these smaller size fractions, at least 1500 particles were scanned after subsampling using a Motoda splitter (Motoda, 1959), requiring generation of two separate scans for both size classes. This resulted in a total of five images per net.

Image names:

Image names include:
cruise#_mocnessID_net#_sizefraction_ and _a|b if a replicate and end in _raw_1.tif

Multiple images of the same size fraction were sometimes taken to obtain a sufficient number of particles. These replicates are named a or b. If there is no replicate they don’t have a letter in the image name. An a and b scan were always done for size classes d2 and d3.  This was important because the split size is for the sum of a+b (e.g. if a is ¼ and b is ¼, the acq_sub_part will be 0.5).

Example of image names:

ae2112_m22_n4_d3_a_raw_1.tif  [a replicate]
ae2112_m22_n4_d3_b_raw_1.tif  [b replicate]
ae2204_m27_n5_d1_raw_1.tif      [no replicate]

Related Datasets may contain the "object_id" (the particle/organism id) which is constructed the same way as the image name except it as an additional _# at the end.  This additional number in the object_id is added by the ZooProcess software (Hydroptic, 2016).
e.g.
object_id:       ae1614_m3_n1_d2_a_1_100
image_name: ae1614_m3_n1_d2_a_1.tif

Scans were processed using ZooProcess (version 8.22, ImageJ macro suite). The "Convert and process from RAW" function was used to separate particles into individual vignettes and generate a suite of measurements for each particle. "Doubles" (vignettes containing more than one particle) were manually separated in the software and reprocessed. 

Processed scans and their corresponding metadata were then uploaded to Ecotaxa (Picheral et al, https://ecotaxa.obs-vlfr.fr/), where a training set was created using manually classified images from this project as well as existing validated images from other projects in the Sargasso Sea. Classification categories were chosen based on taxon of interest, identification level in previous projects, and known limitations of the software. Generally, broader level taxonomic groups are used. Identification of all particles was predicted, then manually validated. 

Version 1:
* All raw scan images were bundled into zip files named by _.zip
* The filesize and md5 checksum was added to the image metadata table.
** After images were transferred to BCO-DMO, a file inventory was made containing the filename, filesize in bytes, and checksum (md5sum). This file inventory table was joined with the provided metadata table to verify the file collection is complete without any missing files.
* an additional supplemental file was added containing metadata for each file (including zips) which includes file access links to aid in programmatic download of all image bundles in the dataset once the dataset is released.
* two imagenames in the metadata table were changed to match the actual filenames. The data submitter was consulted and advised this was the correct course of action. The 3 was omitted in d3 of the imagename.
image_name ae2124_m24_n1_d_a_raw_1.tif -> ae2124_m24_n1_d3_a_raw_1.tif
image_name ae2124_m24_n1_d_b_raw_1.tif -> ae2124_m24_n1_d3_b_raw_1.tif

NSF Award Abstract:
The purpose of this collaborative project is to advance understanding of the role of marine planktonic animals (or zooplankton) in the biological pump, or transport of carbon from surface to deeper ocean waters. This movement of carbon from surface to deep ocean water can ultimately affect carbon dioxide in the atmosphere, with implications for global climate. Many marine zooplankton, including species of copepods and krill, play a direct role in the biological pump both because they are abundant and because they can migrate from surface waters at night, where they feed, to depths of more than 500 m at night. At the same time, some organisms called flux feeders will remain at depth and do not migrate. Instead, they rely on particles produced by other zooplankton feeding in surface waters. In this project, the investigators are focusing on populations of flux feeders in the deeper ocean waters of the Sargasso Sea. They are leveraging an ongoing long-term research program, conducting field collections using specialized nets and particle traps, as well lab experiments, as a way to understand how these organisms modify the particles around them. This project is supporting a postdoctoral scientist and providing research experiences for undergraduates at two institutions. An education specialist is creating lesson plans for an award-winning Ask-A-Biologist website, designed for public and K-12 audiences. Images of zooplankton will be disseminated to the public and scientific community via EcoTaxa (a web platform devoted to plankton biodiversity, with images and taxonomic annotation) and physical samples will be archived as part of a teaching library.

The oceanic biological carbon pump refers to the export of dissolved and particulate organic carbon to the deep ocean, and it is a significant driver of atmospheric carbon uptake by the oceans. Evidence from long-term research carried out at the Bermuda Atlantic Time-series Study (BATS) site suggests that the spectrum of particles collected by gel-traps below the euphotic zone changes drastically below 150 m, which is attributed to resident populations of zooplankton that feed on vertically migrating zooplankton as well as sinking particles. The goals of this study are to investigate the role of different zooplankton taxa on both particle aggregate formation and in particle transformation, and to compare and characterize the particles generated by the zooplankton communities with those collected by particle traps. The investigators are combining field collections with experiments onboard ship and in environmental chambers. They are collecting samples over two years, with three cruises a year to capture distinct seasons. They are assessing high-resolution vertical distribution of zooplankton in the upper 600 m using Multiple Opening-Closing Net and Environmental Sensing System (MOCNESS) tows during day- and night-time, to distinguish diel vertical migrators from resident populations and to quantify contributions to particulate organic carbon flux via fecal pellet production. On each cruise, sinking particles are being collected using gel trap tubes attached to the particle traps deployed monthly at BATS. In addition, roller tank experiments are determining how individual zooplankton mediate aggregate formation. Particle types and fecal pellets are being characterized using image analysis and DNA-based analysis of microbial communities. Finally, ongoing data collection from the long-term BATS program is providing invaluable environmental context and will ensure results from this study contribute to ongoing community efforts to observe and predict the fate of carbon in our global system.

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.