Table of Contents

• Coverage
• Dataset Description
  • Methods & Sampling
  • Data Processing Description
• Parameters
• Instruments
• Deployments
• Project Information
• Funding

Seawater samples (10 × 100 L) were collected during R/V Atlantis cruise AT50-08 (Feb–Mar 2023) at two stations in the Oxygen Minimum Zone of the Eastern Tropical North Pacific (10–600 m depth). Samples were collected by CTD rosette, pre-filtered through 0.22 µm filters to remove cells and debris, and concentrated ~500× by tangential flow filtration (100 kDa cutoff).

The resulting concentrates contained both viruses and extracellular vesicles (EVs). EVs were enriched on board using a lectin-based binding column. Transmission electron microscopy (JEOL JEM 1400, Stony Brook University CryoEM facility) confirmed EV enrichment after column purification.

Subsequent EV analyses were performed at the University of Pennsylvania School of Veterinary Medicine EV Core Facility. Methods included size-based separation (SEC-HPLC, gravity-driven iZon column, density-gradient ultracentrifugation), particle quantification (resistive pulse sensing, nCS1; nanoparticle tracking analysis, NTA), and immunophenotyping (nanoscale flow cytometry; ExoView™ chip array). See related datasets.

CTD data were processed following standard Sea-Bird CTD procedures. Raw measurements were converted to engineering units using manufacturer calibration coefficients, then time-aligned across sensors (e.g., conductivity and oxygen aligned to pressure/temperature) and corrected for conductivity cell thermal mass.

Data were screened to remove periods of unstable flow and package motion (surface soak, pump start-up, winch stops/pressure reversals/loops), and automated despiking (“wild edit”) was applied to remove dropouts and isolated spikes.

Final profiles were generated from the downcast and used to compute derived variables including practical salinity, depth (from pressure using latitude), density, sound velocity (Chen–Millero), and dissolved oxygen products (µmol/kg, mL/L, % saturation); some oxygen fields include a smoothed/windowed version indicated in the headers (e.g., “WS=2”).

NSF Award Abstract:
This EAGER project is a proof-of-concept study on the composition, origins, and dynamics of extracellular vesicles (EVs) produced by marine microalgae in response to various environmental and biotic stressors. EVs are microscopic lipid-encased particles that are released naturally from almost all cell types and are vehicles for a variety of cargo, including genetic material (RNA, DNA), proteins, and lipids. EVs have been variously postulated to serve as a defense against viral attack, a waste disposal mechanism, a stress response, or a means of cell-to-cell communication. Marine microalgae are pivotal players in the global carbon cycle. By better understanding processes that govern their population dynamics and responses to environmental changes, we can develop better predictive models of responses to global climate change. The need to understand these mechanisms is becoming increasingly urgent as climate change becomes more manifest. Very recent findings suggest that EVs play a key role in marine phytoplankton population regulation, but our understanding of their function(s) in planktonic systems is severely limited and fragmentary. This project addresses significant knowledge gaps and explores the potential complexities of marine planktonic EV production. This project provides support and training to a female graduate and undergraduate marine sciences students, who are receiving unique opportunities to master new experimental approaches and state-of-the-art research tools that are extremely rare in marine sciences programs. The project supports high school students in marine sciences studies as a part of the summer science camp (www.sigmacamp.org). A female postdoc is also being trained on the project.

Using the cosmopolitan and geochemically-important microalga E. huxleyi as a model system, this project tests three major hypotheses to enhance our understanding of the purpose(s) of microalgal EV production. (1) Microalgae produce distinctive types of EVs (ectosomes or exosomes) in response to different environmental conditions, and EV types have definitive functions (stress response, viral defense, intercellular communication, waste disposal). (2) EVs’ cargo is diverse, so their production and release reflect a complex intercellular communication mechanism. (3) Exosome genesis is a multistage process, and its stages are separated in time. Therefore, algal cells may contain a pool of pre-formed EVs loaded with different cargo that are stored internally, and when induced by a sudden change in external conditions are released through the outer membrane. To adequately test these hypotheses requires using single particle analytical methods in addition to ensemble measurements. The investigators are using an assortment of recently developed methods and original experimental approaches developed by our group to investigate EV compositional variability under selected stress conditions. They use single particle Raman microspectroscopy, pulse-chase Stable Isotope Probing, and LC-MSMS for compositional analysis of EVs, and Cryo-EM and AFM for morphological analyses. If experimental data confirm our suspicions, then phytoplankton EVs represent a novel and essentially overlooked mechanism of extracellular interactions that potentially govern a wide range of globally-important processes.

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.