Dataset: BioLog Eco-plates experiment - Metabolic potential

BioLog Eco-plates contained 96-wells, prefilled with: (1) a colorless tetrazolium dye that reduces to a violet formazin if the substrate is oxidized, (2) triplicates of 31 different organic compounds in equimolar quantities, and (3) triplicate water blanks as a control for airborne bacterial contamination. We used a single coccolithophore strain in each plate. We started each experiment by inoculating a 96-well plate with 100 μL of log-phase cell suspension from each log phase culture. We carried out all inoculations under subdued light and all incubations in complete darkness. At time-zero (T0), 24 h, 48 h, and 72 h, we measured the reduced violet tetrazolium dye absorption on a FilterMax F5 multimode plate reader (Molecular Devices, LLC, San Jose, CA, U.S.A.) for absorption at 595 nm. We interpreted the increasing optical density at 595 nm as heterotrophic metabolism of the organic compound by the coccolithophore cultures. (Note: For two strains, CCMP298 and CCMP 3337, T72 was not performed, but the measurement was instead taken at T120.)

Several limitations to the BioLog Eco-plates should be noted (Stefanowicz 2006). One limitation is that the technique assumes that the uptake of each substrate is independent from any other (i.e., two substrates are not used in a synergistic fashion). The technique also assumes the concentrations of the substrates in each well of the microtiter plate are optimal for the organism in question, rather than too high (i.e., inhibitory) or low (limiting) based on the organism's specific uptake kinetic parameters. Therefore, we could not make any inferences on the uptake kinetics of DOC utilization using the microtiter plates. Furthermore, ions in seawater (specifically Ca++) can cause false positives in BioLog Eco-plates (Pierce et al. 2014). It is critical to lower the Ca++ concentration from 10 mmol L⁻¹ (normal concentration in seawater) to ~ 2.5 mmol L⁻¹ in order to eliminate false positives (Pierce et al. 2014). The BioLog Company suggests doing this with chelators. An alternative method was suggested by Tuchman et al. (2006) in which the cultures were centrifuged and pelleted to separate them from the nutrient-rich media, which might contain other growth-inducing substrates. Then the pellet should be resuspended in nutrient-free saline solution. We followed these latter recommendations and resuspended the coccolithophores in ASW with 2.5 mmol L⁻¹ calcium.