This is an experiment of a series investigating the mechanisms driving the incorporation of oil into diatom aggregates using a water accommodated fraction (WAF) of oil. The diatom Thalassiosira CCMP strain #1335 was used, and WAF was prepared from Macondo oil (Deepwater Horizon oil spill in the Gulf of Mexico April 2010) and from Refugio oil (pipeline oil spill off California May 2015). Roller tank experiments mimic in situ conditions of the ocean in that continuous sinking of particles is possible (infinity water column). Once aggregates>1mm (operationally defined) have formed, they are harvested separately from the surrounding seawater, which contains non-aggregates cells or small aggregates. Both fractions are analyzed separately, so that the fraction of material incorporated within aggregates may be calculated. Samples were analyzed for cell abundance, particulate organic carbon (POC) and nitrogen (PON), the isotopes Carbon-13 and Nitrogen-15, estimated oil equivalent (EOE), and transparent exopolymer particles (TEP). Sinking velocity of aggregates was determined.
Uta Passow. 2018. EPS mechanisms in aggregation and dispersion of oil: roller tank experiment using Thalassiosira pseudonana culture and natural bacterial inoculum. Distributed by: Gulf of Mexico Research Initiative Information and Data Cooperative (GRIIDC), Harte Research Institute, Texas A&M University–Corpus Christi. doi:10.7266/N78W3BP3
To test the effect of different oils and natural microbial populations on marine oil snow formation: 3 treatments (Control and RefugioB-WAF, MacondoWAF) with Thalassiosira pseudonana (CCMP 1335 strain #1335) culture and natural bacterial inoculum. Marine snow and surrounding seawater (SSW) sampled separately.
Data Parameters and Units:
GRIIDC-WAF-Agg-Summary-R4x263-000-0006.csv: Experiment: exp. Tank number = tank # Sampling Date (MM/DD/YYYY) Sampling Day = number of experiment days Treatment: Control, Refugio, or Macondo Replicate # = number of replicates Type of Sample: Slurry or SSW Slurry = the fraction of aggregates >1 mm with some surrounding seawater SSW = surrounding seawater fraction Maximum tank volume (5773 mL)/Slurry volume = Tank Vol/Slurry volume (mL) Avg POC = Average particulate organic carbon (ug/tank) Std POC = Standard deviation particulate organic carbon (ug/tank) Avg PON = Average particulate organic nitrogen (ug/tank) Std PON = Standard deviation particulate organic nitrogen (ug/tank) Avg d13CNorm = Average delta Carbon-13 (unitless) standard expressed as per mille:(δ13C = (Rsample/Rstandard -1) X 1000, where R = 13C/12C) Std d13CNorm = Standard deviation delta Carbon-13 (unitless) Avg d15NNorm = Average delta Nitrogen-15 (unitless) Std d15NNorm = Standard deviation delta Nitrogen-15 (unitless) Avg EOE corrected = Average Estimate Oil Equivalent (µg/tank) Avg TEP = Average transparent exopolymer particles (µgGXeq/Tank) Std TEP = Average transparent exopolymer particles (µgGXeq/Tank) Gum Xanthan equivalent = GXeq Avg cell numbers per tank Sinking velocity measured: yes or blank GRIIDC-WAF-Agg-Sinking Velocity-R4x263-000-0006.csv: Experiment = exp. Treatment: Control, WAFRef, or WAFMac Water accommodated fraction of Macondo oil = WAFMAC Water accommodated fraction of Refugio oil – WAFRef Control = no oil Aggregate ID number = Agg # Equivalent spherical diameter = ESD (mm) Sinking velocity = w (m/d)
Replicate acrylic roller tanks were filled bubble free. Experiments were incubated in the dark, at 13˚C at an rpm of 1.93 Aggregate appearance and numbers were monitored during the experiments. When sufficient aggregates had formed, all treatments were harvested. Aggregates >1mm (visually discernable) were manually collected and analyzed. After all aggregates >1 mm were removed the remainder of the material, termed surrounding seawater (SSW), was subsampled. Both fractions were analyzed for POC/PON, cell abundance, TEP, CSP. Relative oil concentration was estimated from fluorescence using a Trilogy Turner Fluorometer with the crude oil module 7200-63, which measures at excitation wavelength of 365 nm and emission wavelength of 410-600nm. Duplicate filters (GF/F) prepared for POC/PON analysis were measured in a CEC44OHA elemental analyzer (Control equipment). Diatom cells were counted (Olympus CX41) using a hemocytometer; at least 6 subsamples and 200 cells each were counted per sample. Counts were at times only conducted on 1 of the two replicate treatments. TEP and CSP concentrations were determined in triplicate each using the respective colorimetric methods and are expressed in Gum Xanthan equivalents (GXeq.) and Bovine serum albumin equivalent (BSA eq.) respectively. All biomass results (cell abundance, POC, TEP concentration) are normalized per tank to allow budgets and make aggregate and SSW fractions directly comparable. Make WAF directly before use. Make 3L filtered seawater (0.2 µm filter), and pasteurize it for at least 2 hours at 65 degrees C. Add into 4+ L glass bottle with bottom spigot and glass stopper. Leave 20-25% headspace. Add a sterile stir bar to the bottle and add 3 mL (3000 µL) oil. Immediately cap, shake. Place on a stir table at room temperature for about 24 hours at a fairly high stirring speed (not enough to generate a vortex, but enough to keep things homogenized). Keep dark during mixing (aluminum foil or brown glass). Let the bottles stand for a couple hours. Then, harvest discarding the upper oily layer. Sinking velocity measurements were conducted using the column/ setcol method and the non-destructive orbit method of calculating sinking velocities based on tank rotation speed and the orbital path of aggregates within the turning tanks. Ploug, H., Terbrüggen, A., Kaufmann, A., Wolf-Gladrow, D. and Passow, U. (2010) A novel method to measure particle sinking velocity in vitro, and its comparison to three other in vitro methods. Limnology and Oceanography: Methods 8, 386-393.