Table of Contents

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

This dataset has been deprecated and replaced. Please see dataset https://www.bco-dmo.org/dataset/779368.

Thalassiosira pseudonanana were grown in nitrate replete culture at eight temperatures, 300 umol photons m–2 s­–1, and 400 ppm CO2. Growth rates, photosynthetic rates, respirations rates, C:N ratio, C:Chlorophyll-a ratio, maximum quantum yield, and the range of pCO2 values are reported.

The culture was grown in a nutrient-replete continuous culture system on a 14:10 L:D cycle of illumination at a temperatures of 10, 15, 20, 25, 30, 31, and 32°C. The irradiance during the photoperiod was 300 micro-mol photons m^–2 s^–1. Photosynthetically active radiation (400–700 nm) was measured with a Biospherical Instruments model QSL 2100 quantum sensor. Temperature was controlled to within 0.1°C by circulating water from a Haake model DC10 temperature-controlled water bath through the outer jacket of the reaction chamber. The dilution rate of the growth chamber was controlled with a peristaltic pump (Masterflex Model 77200-60) to within ± 0.002 per day. The CO₂ concentration in the laboratory was monitored with a CO2METER model AZ-004 meter calibrated at 0 and 400 ppm CO₂ with a standard gas mixture.

The system was judged to be in steady state when cell counts, measured with a Beckman Coulter model Z1 particle counter, had been reproducible to within ± 2% for at least 4 doubling times. Chlorophyll a concentrations were determined from samples collected on glass fiber filters and extracted in methanol. The absorbances were measured at 664 and 750 nm with a Cary Model 50 spectrophotometer. Concentrations of particulate organic carbon (POC) and particulate nitrogen (PN) were determined by filtering replicate 50-mL samples from the growth chamber onto GF/F glass fiber filters followed by analysis with an Exeter Analytical model CE-440 elemental analyzer. pH was measured with a Thermo Spectronic Heios spectrophotometer, as described in SOP 6B by Dickson, et al 2007 with minor modifications, and with a Hach SensION model PH31 pH meter calibrated with standards on the total pH scale, prepared as per Millero, F.J., et al. "The use of buffers to measure the pH of seawater." Marine Chemistry 44.2 (1993): 143-152, with minor modifications.

The growth medium consisted of artificial seawater with a total alkalinity of 2365 meq L^–1. Nutrient concentrations corresponded to f/2 medium, with the exception of trace metals, which were added at the concentrations specified by Sunda and Hardison (Limnology & Oceanography 52[6]:  2496–2506 [2007]). The medium was sterile filtered (0.2 micron) into a 40-liter glass carboy that had been previously autoclaved. The growth chamber was an autoclaved glass reaction flask with a working volume of 2183 mL. The cells in the growth chamber were uniformly labeled with C-14 by adding 20 microcuries of C-14 bicarbonate to the nutrient reservoir. Five-milliliter samples for C-14 activity in the organic carbon were withdrawn in triplicate from the growth chamber at two-hour intervals during the photoperiod. The samples were acidified with 1 mL of 1 N HCl to drive off inorganic carbon.

The activity of C-14 in the samples was then determined by counting on a Packard Tri-Carb model 3100 TR liquid scintillation counter. Short-term (5-minute) photosynthesis versus irradiance curves were measured at the start, middle, and end of the photoperiod. For these experiments, triplicate 5-mL aliquots from the growth chamber were added to liquid scintillation vials pre-inoculated with 0.85 microcuries of C-14 bicarbonate. The vials were incubated at irradiances of 5, 10, 20, 30, 55, 80, 120, 150, 200, 250, 300, and 350 micro-mol photons m^–2 s^–1 for 5 minutes. Fixation was stopped by adding 0.5 mL of 1 N HCl to the vials. Total alkalinity was determined using the open cell titration method described as SOP 3B by Dickson, et al 2007 DIC concentrations were then calculated from temperature, salinity, total alkalinity, and pH using the equations in Zeebe and Wolf-Gladrow, CO₂ in Seawater: Equilibrium, Kinetics, Isotopes.

Photosynthetic rates as a function of irradiance were found to be best described by Hill reaction kinetics with n = 2 (Hill, A. V., J. Physiol. 40, iv-vii [1910]). For n = 2, the Hill equation takes the form

P = P_maxI²/(K_I^² + I^²)

The light-saturated photosynthetic rate (Pmax) with units of grams carbon per gram chlorophyll a per hour and the parameter KI (Hill coefficient) with units of micro-mol photons m^–2 s^–1 were determined by least squares. Dark-adapted photosynthetic quantum yield (QY) was measured in triplicate for each continuous culture in steady state at mid-photoperiod. QY measurements were made with a PSI AquaPen C100 with manufacturer’s supplied plastic cuvettes containing 4 mL of culture each. Dark-adaptation of the culture samples was achieved by wrapping each of three cuvettes in aluminum foil and incubating at room temperature for 30 minutes, after which QY was measured in a darkened room. 

References:

Dickson, A.G., Sabine, C.L. and Christian, J.R. (Eds.) 2007. Guide to best practices for ocean CO₂ measurements. PICES Special Publication 3, 191 pp.

Hill, A. V. 1910. The possible effects of the aggregation of the molecules of haemoglobin on its dissociation curves. J. Physiol. 40: iv–vii.

Sunda, W. G., and D. R. Hardison. 2007. Ammonium uptake and growth limitation in marine phytoplankton. Limnol. Oceanogr. 52(6): 2496–2506.

Photosynthetic rates during two-hour intervals during the photoperiod were calculated by solving the differential equation

d(POC)/dt = P – m x POC                                                                                           (1)

where P is the rate of production of POC in the growth chamber, m is the dilution rate of the growth chamber and d(POC)/dt is the rate of change of POC in the growth chamber. The solution of equation (1) between two points in time is

P = m(POC_t – POC₀ e^^–mt) /(1 – e^^–mt )                                                                            (2)

where POC₀  and POC_t are the concentrations of POC at the beginning and end of the time interval, respectively, and t is the duration of the time interval, which in this experiment was 2 hours. Values of P were calculated for each two-hour time interval during the photoperiod, normalized to the chlorophyll a concentration during each time interval, and then averaged to determine the photosynthetic rate per unit chlorophyll (productivity index or PI) during the photoperiod. Results are reported as grams of carbon per gram of chlorophyll a per hour averaged over the 14-h photoperiod.

Dark respiration rates were calculated from the natural logarithm of the ratio of the total organic carbon ¹⁴C activity at the end of the photoperiod and the beginning of the subsequent photoperiod. All TO¹⁴C activities were corrected for blank counts, which were typically 24 count per minute. The natural logarithm of the ratio of the TO¹⁴C counts was equated to (m + m_r)10/24, where m_r is the dark respiration rate (d^–1) and m is the dilution rate (d^–1). Division by 24 converts these rates to h^–1, and multiplication by 10 corrects for the fact that the duration of the dark period was 10 hours. Thus

  m_r = (24/10)ln (TO¹⁴C_e/ TO¹⁴C_b) – m                                                                     (3)

where TO¹⁴C_e and TO¹⁴C_b are the ¹⁴C counts in the TOC at the end of one photoperiod and the beginning of the next photoperiod, respectively.

The minimum quantum requirement (i.e., smallest number of photons required to fix one carbon atom) was estimated at the beginning, midpoint, and end of the 14-h photoperiod based on the 5-minute uptake of inorganic carbon (gram C per gram chlorophyll per hour) versus irradiance. Hill kinetics with n = 2 was assumed  (Hill, A. V., J. Physiol. 40, iv-vii [1910]). For n = 2, the Hill equation takes the form

P = P_maxI^²/(K_I^² + I^²)

For such an uptake curve, the minimum quantum requirement equals 2K_IA_abs/P_max, where A_abs is the chlorophyll-specific absorption coefficient of light. Because our experiments were conducted with white light (daylight fluorescent lamps), we assumed an A_abs value of 14 m² per gram of chlorophyll based on Atlas and Bannister, Limnology & Oceanography 25(1): 157–159 (1980). If K_I has units of micro-mol photons m^–2 s^–1 and P_max has units of g C g^–1 chl a h^–1, the minimum quantum requirement based on the Hill equation with n = 2 is 1.2096K_I/P_max. Minimum quantum requirements were calculated at the beginning, middle, and end of the 14-h photoperiod and are reported as the averages of those three estimates.

BCO-DMO Processing Notes:
- added conventional header with dataset name, PI name, version date
- modified parameter names to conform with BCO-DMO naming conventions
- replaced 'no data' and empty cells with nd

The overarching goal of this project is to develop a framework for understanding the response of phytoplankton to multiple environmental stresses. Marine phytoplankton, which are tiny algae, produce as much oxygen as terrestrial plants and provide food, directly or indirectly, to all marine animals. Their productivity is thus important both for global elemental cycles of oxygen and carbon, as well as for the productivity of the ocean. Globally the productivity of marine phytoplankton appears to be changing, but while we have some understanding of the response of phytoplankton to shifts in one environmental parameter at a time, like temperature, there is very little knowledge of their response to simultaneous changes in several parameters. Increased atmospheric carbon dioxide concentrations result in both ocean acidification and increased surface water temperatures. The latter in turn leads to greater ocean stratification and associated changes in light exposure and nutrient availability for the plankton. Recently it has become apparent that the response of phytoplankton to simultaneous changes in these growth parameters is not additive. For example, the effect of ocean acidification may be severe at one temperature-light combination and negligible at another. The researchers of this project will carry out experiments that will provide a theoretical understanding of the relevant interactions so that the impact of climate change on marine phytoplankton can be predicted in an informed way. This project will engage high schools students through training of a teacher and the development of a teaching unit. Undergraduate and graduate students will work directly on the research. A cartoon journalist will create a cartoon story on the research results to translate the findings to a broader general public audience.

Each phytoplankton species has the capability to acclimatize to changes in temperature, light, pCO2, and nutrient availability - at least within a finite range. However, the response of phytoplankton to multiple simultaneous stressors is frequently complex, because the effects on physiological responses are interactive. To date, no datasets exist for even a single species that could fully test the assumptions and implications of existing models of phytoplankton acclimation to multiple environmental stressors. The investigators will combine modeling analysis with laboratory experiments to investigate the combined influences of changes in pCO2, temperature, light, and nitrate availability on phytoplankton growth using cultures of open ocean and coastal diatom strains (Thalassiosira pseudonana) and an open ocean cyanobacteria species (Synechococcus sp.). The planned experiments represent ideal case studies of the complex and interactive effects of environmental conditions on organisms, and results will provide the basis for predictive modeling of the response of phytoplankton taxa to multiple environmental stresses.