Survey area
The U.S. AMLR Program conducted five winter surveys (August and September 2012–2016) around the northern Antarctic Peninsula aboard the U.S. National Science Foundation research vessel/ice breaker (RVIB) Nathaniel B. Palmer (Fig. 1)19. We surveyed a historical grid of 110 fixed stations located 20–40 km apart around the northern Antarctic Peninsula and South Shetland Islands, which was divided into four sampling areas: the Elephant Island Area (EI; 43,865 km2), the South Area (the Bransfield Strait, SA; 24,479 km2), the West Area (the west shelf immediately north of Livingston and King George Islands, WA; 38,524 km2), and the Joinville Island Area (JI; 18,151 km2). In 2016, we also surveyed the Gerlache Strait (GS; 24,479 km2).
At-sea sampling
Detailed methods for all at-sea sampling by the U.S. AMLR Program have been previously reported19. At each sampling station, we performed a Conductivity-Temperature-Depth (CTD) cast to 750 m or to within 10 m of the bottom in shallower areas (SBE9/11; Sea-Bird Electronics). The CTD rosette was equipped with 24 10 l Niskin bottles triggered to close on the upcast at 750, 200, 100, 75, 50, 40, 30, 20, 15, and 5 m. We defined the Upper Mixed Layer (UML) depth (m) as the depth at which the density of the water changed by 0.05 kg m−3 relative to the mean density of the upper 10 m of the water column20. UML temperature and salinities were defined to be means over the depth range of the UML. We defined daylight conditions according to three categories. Day (D) was defined as one hour after local sunrise to one hour before local sunset; night (N) was defined as one hour after sunset to one hour before sunrise; and Twilight (T) was defined as one hour before and after sunrise and sunset.
Chlorophyll-a (hereafter chl-a) was determined from water samples collected between 5 and 200 m21,22. Samples (285 ml) were filtered at a pressure differential of < 150 mm Hg through 25.4 mm glass fiber filters (Whatman, Inc.), and chl-a was extracted from filters in 7 ml methanol over 24 hrs. Samples were then centrifuged and chl-a was determined from the recovered supernatant using a Trilogy fluorometer (Turner Designs). Samples were then acidified using 2 drops 1.0 N HCl and read again to determine phaeopigment concentration. We calculated integrated chl-a (to 100 m, mg chl-a m−2) using concentrations at the discrete depths of each sample from 5 m to 100 m and averaging between depths to calculate the intermediate depth concentrations. Concentrations were then summed over the top 100 m for each CTD cast as an indicator of water-column food availability.
At each station, we towed an Isaacs-Kidd Midwater Trawl (IKMT) net with 505 µm mesh to 170 m or to within 10 m of the bottom in shallower areas. The volume of seawater filtered by each tow was determined using a calibrated flow meter mounted on the net frame (Model 2030 R, General Oceanics, Inc.). Large zooplankton (identifiable without a microscope) were enumerated and identified to species. Small zooplankton (requiring a microscope for identification) were subsampled by rinsing the cod end of the net into a 505 µm sieve and suspending the recovered zooplankton in a known volume of ambient seawater. The water was thoroughly mixed and zooplankton in 1–2 ml subsamples were placed on Petri dishes and identified under a microscope. At least five subsamples were counted per tow, and total counts of individuals were scaled up to the volume of the tow. Abundance of each species was calculated by dividing the total number of individuals by the tow volume and multiplying by the tow depth.
Sea ice
Beginning in 2013, we determined percent sea-ice cover and sea-ice type at each sampling station by observing the ice conditions in a 200 m arc around the stern of the ship during each net tow. Percent sea-ice cover was characterized as 0% (open water), 1–25% (recorded as 25%), 26–50% (recorded as 50%), 51–75% (recorded as 75%), and 75–100% (recorded as 100%). Sea-ice type was classified according to a modification of the standardized visual approach from the Scientific Committee on Antarctic Research Antarctic Sea Ice Processes and Climate Program (ASPeCT)23. We classified ice type as slush (frazil, shuga, and grease ice; less than 100 mm thick), thin (nilas, pancake, young grey, and young grey-white ice; 100–300 mm thick), first-year (thicker and more solid ice; 300–1200 mm thick), and multi-year (more solid; any ice thicker than 1200 mm).
Lipid analysis
Individuals of each species collected from the same station were frozen together at −20 °C until lipid analysis (usually no later than 48 hrs after collection). At stations where too few individuals were collected to achieve a mass suitable for lipid analysis (0.5–1.5 g), individuals from multiple stations within the same sampling area were pooled for analysis.
Lipid was extracted from aliquots of 0.5–1.5 g of homogenized animals24,25. Samples were refrigerated overnight in a 2:1 chloroform:methanol solution with 0.01% butylated hydroxytoluene (BHT) as a preservative. The following day, samples were filtered to collect the solution, and the lipid-extracted zooplankton were retained in glass vials for stable isotope analysis. The solution was centrifuged for 20 min and the methanol layer was removed and discarded. Lipid dissolved in the chloroform layer was filtered through anhydrous sodium sulfate to remove all traces of remaining methanol and water. The chloroform solvent was then evaporated under nitrogen and lipid was weighed to the nearest 0.001 g (Model PA214, S/N: 8331270072, Ohaus) to determine percent lipid in each sample.
Percent lipid of both wet mass and dry mass are reported in the full dataset. Dry mass of samples was determined by freeze-drying all 2016 lipid-extracted samples in glass vials for 16 hrs (VirTis Benchtop K Lyophilizer, SP Scientific), adding back the mass of the removed lipid, and then subtracting the mass of the vial. The mean percent mass loss for each species was calculated and applied to the wet mass of each sample to determine dry mass.
Stable isotope analysis
The same samples used for lipid analysis were used for stable isotope analysis and prepared according to the analytical laboratory’s guidelines26. Vials of lipid-free zooplankton were freeze-dried for 14–16 hrs. Dried samples were pulverized and 0.8–1.2 mg aliquots were weighed into tin capsules (5 × 9 mm, Costech Analytical Technologies) and analyzed for δ13C and δ15N at the University of California Davis Stable Isotope Facility (UCD SIF) using a PDZ Europa ANCA-GSL elemental analyzer interfaced to a PDZ Europa 20–20 isotope ratio mass spectrometer (Sercon). Isotope values are reported as ratios of heavy to light isotopes (δ13C and δ15N) in parts per thousand (‰) compared to standards (PeeDee limestone and atmospheric N2, respectively).
Because the zooplankton species analyzed contain few, if any, calcified structures, samples were not acid treated prior to stable isotope analysis to eliminate the risk of skewing δ15N values27. Lipid removal prior to stable isotope analysis may also skew δ15N values. Lipid is often removed because it is more depleted in 13C than protein or carbohydrate and may result in inaccurate characterizations of δ13C if not removed, particularly in organisms with high lipid content28. However, chemical lipid extraction following the same technique we used24 may only skew δ15N by approximately 0.25‰28, which is close to typical analytical error. To address the issue of skewed δ15N resulting from lipid extraction, we collected as many extra samples of all zooplankton species as possible in 2015 and 2016 for stable isotope analysis of non-lipid-extracted animals. Those data are included in the dataset and denoted as “WAN” (whole-animal), whereas lipid-extracted samples are denoted as “LFR” (lipid-free). The δ13C in whole-animal samples was not normalized for lipid content.
Source: Ecology - nature.com
