Sampling and analytical procedures:

Capture and husbandry: Antarctic krill (Euphausia superba) were captured during the austral summer of 2014/2015. Krill were collected by net tow (2 m diameter, 1000 m mesh, non-filtering cod end) off the R/V Laurence M. Gould near the Western Antarctic Peninsula and transported directly to the Palmer Station biological laboratory. One to two thousand krill were housed in one 4'w x 3'h circular holding tank and two 5’ x 2’ x 1’ rectangular tanks provided with aeration and flow-through seawater. Water was non-filtered and individuals were able to feed on plankton ad libitum throughout the season.

Experimental treatments: Four experimental treatments were targeted in this study, (1) ambient temperature (0 degrees C) and ambient pCO2/pH (400ppm/8.10), (2) ambient temperature (0 degrees C) and elevated pCO2 (800ppm)/reduced pH (7.7), (3) elevated temperature (3 degrees C) and ambient pCO2 (400ppm/8.10), and (4) elevated temperature (3 degrees C) and elevated pCO2 (800ppm)/reduced pH (7.7). Temperature treatments were obtained using two separate recirculating systems. Two 800 L cylindrical polycarbonate carboys were attached to temperature controlled chillers (Delta Star) and inline pumps. The carboys were filled with non-filtered seawater acquired from the Palmer Station intake line, placed in a flow-through water bath, and maintained at 0 degrees C. Two other 800 L carboys were set up without a chiller and placed in an environmental chamber at 3 degrees C. The systems were replaced with new water daily and allowed to acclimate to temperature for a minimum of 24 hours before the start of a trial or water change. High CO2 conditions were obtained using a peristaltic pump to inject straight CO2 into the propeller of a pump submerged in seawater. Treated water was then gently siphoned with minimal disturbance into treatment bottles.

Krill of comparable size (31-38 mm) were picked and placed in individual 4 L wide-mouth polycarbonate bottles with airtight lids (n = 30 per experimental treatment). Bottles were filled with one of the four pre-acclimated treatment waters as described above. Bottles were immediately closed and placed in environmental chambers at either 0 or 3 degrees C. Every 24-48 hours, 80% of the water was siphoned out of each experimental bottle and replaced with pre-acclimated water from the 800 L carboys to minimize excretory and respiratory effect of the animals on treatment conditions. During the 23-day experiment, bottles were checked for molts twice per day. If a molt was observed, that bottle was pulled from the experiment and the molt and live krill processed. The experiment ended after the last krill molted.

Analyses: Once a krill molted, both the krill and molt were gently removed from the bottle. Both the molt and krill were measured for uropod and total length (mm) with digital calipers (Fowler), and the krill wet weight (mg) was measured.

pH was measured in each experimental bottle at the start of the experiment. Salinity, pH, and total alkalinity were measured during water changes. The water changes occurred every 24-48 hours, but we were unable to collect salinity, pH, and total alkalinity samples at every water change due to time constraints. Salinity was measured with a bench top conductivity meter (YSI 3100) calibrated daily with a conductivity standard (50,000 uS/cm; Ricca Chemical Company). pH was determined spectrophotometrically using the indicator dye thymol blue (Dickson et al. 2007; Zhang and Byrne 1996). Total alkalinity was determined on 100 ml subsamples with an open-cell, potentiometric titration of seawater (Metrohm 888 Titrando) with 0.1 M HCl following the potential of a pH electrode (Dickson et al. 2007). Tiamo software (version 2.3) was used to process the alkalinity data. Measurements of pH and TA were quality controlled using certified reference materials (CRMs) obtained from Andrew Dickson at UCSD Scripps Institute of Oceanography.

2015_krill_growth1.csv