Micro- and mesozooplankton respiratory metabolism: comparison of simultaneous incubation experiments and ETS activity

A Microand mesozooplankton respiratory metabolism: comparison of simultaneous incubation experiments and ETS activity. By Miguel Alcaraz1*, Theodor T. Packard1, Enric Saiz1, Albert Calbet1, Isabel Trepat1 and Alf Skovgaard2 1 Group of Zooplankton Ecology. Institut de Ciències del Mar, CSIC. P. Marítim de la Barceloneta 37-49, 08003 Barcelona (Spain). 2 University of Copenhagen, Department of Biology. Øster Farimagsgade 2D, DK-1353 Copenhagen K, Denmark * e-mail: miq[email protected]c.es 1. Introduction º The contribution of the different plankton components to the Ocean respiratory carbon losses is of prime importance for the improvement of models dealing with global climate changes and plankton production. However, the obtention of plankton respiratory losses for the separate groups by classical incubation methods is extremely timeand labour consuming. Enzymatic methods like the activity of the respiratory electron transport system (ETS) overcome most of the experimental inconveniences, but gives potential, not actual respiration rates. In order to use ETS as valid estimator of respiration rates, it is necessary to determine wheter there exist robust relationships between ETS activity and directly measured O2 consumption. 2. Aims Alcaraz M., et al. (1996). J. Mar Systems, 17: 347-359. Alcaraz M, and Packard, T.T. (1989). Sci. Mar., 53: 247-250. Bamstedt, U (1980). J. exp. mar. Biol. Ecol. 42: 267-283. Kin, F.D., and Packard, T.T. (1975). Limnol. Oceanogr. 20: 849-857. Controls Experiments Electrode Syringe Experimental organisms The study included three groups of experimental zooplankton organisms (Fig. 1): >200 µm mixed zooplankton (mesoplankton) from 3 stations in the NW Mediterranean in March 1999 (Alcaraz et al. (2007). Obtained by 100 – 0 m depth vertical net hauls made with a 200 µm-mesh WP-2. 3. Methods Fig. 3.- Electrode graphs of O2 concentration inIncubation experiments Fig. 2.- Incubation flasks showing the electrode and the pressure control The study was based in the comparison of O2 consumption rates obtained by classical incubation methods with those obtained from ETS activity in the incubated organisms. Direct measurements of respiration rates: O2 consumption rates were measured by 24 h-duration incubation experiments at “in situ” temperature as described in Alcaraz et al. (1998). In most of the experiments, semi-continuous analysis of dissolved O2 concentration were obtained with pulsed polarographic O2 electrodes (ENDECO , Fig. 2). Time - O2 concentration relationships were fitted by linear regression (Fig. 3). In few cases, the incubation was made in BOD bottles and the O2 concentration measured by Winkler titration (Fig. 4). ETS activity: After the incubations, the experimental organisms were transferred by filtration into GF/A glass fibre filters, immediately frozen in liquid N2 and maintained at -80 oC until the analysis of the ETS activity. These were performed according to Packard et al. (1996). Fig. 4.- BOD bottles for respiratory incubation experiments controls Experiments O2 electrode <200 >50 µm mixed zooplankton (metazoan microzooplankton) from a station 1 nM offshore Barcelona. Samples obtained from november 2003 to january 2004. 40 – 0 m depth vertical hauls made with a 50 µm-mesh net.. Copepod nauplii. (nauplii II-IV) obtained from a laboratory culture of the cyclopoid Oithona davisae maintained by the Zooplankton Ecology Group in the Institute of Marine Sciences. A B C Fig. 1.- The three experrimental zooplankton groups: A: Mixed mesozooplankton; B: Mixed microzooplankton; C Oithona davisae nauplii. 1.- The obtention of robust relationships between directly measured O2 consumption rates and ETS activity for different zooplankton components. 2.- To ascertain whether the relationships between respiration rates and ETS for different zooplankton size-fractions are also different and can be used as a proxy for respiration rates. 1.- The obtention of robust relationships between directly measured O2 consumption rates and ETS activity for different zooplankton components. 2.- To ascertain whether the relationships between respiration rates and ETS for different zooplankton size-fractions are also different. The relationships between ETS values and respiration rates (RR) for the different groups were statistically significant (Fig. 5 and Table I). The regression coefficients for > 200 µm zooplankton were not statistically different from those of other groups (p < 0.05), even after a single point from the mesozooplankton (*) was removed. For the pooled data the differences with respect the remaining groups were also non significant (p < 0.05). A A 4. Results and Discussion A Fig. 5.- Relationships betwee RR and ETS for the different grups and for all the data pooled. For > 200 µm zooplankton and pooled data, a line with an asterisc means the corresponding equation after the high value point has been removed. Group Equation R2n > 200 µm mixed zooplankton RR = 0.28 ETS + 1.05 0.93 18 > 200 µm mixed zooplankton *RR = 0.36 ETS + 0.29 0.88 17 < 200, > 50 µm mixed zooplankton RR = o.441 ETS + 0.65 0.91 17 Nauplius II-IV Oithona davisae RR = 0.45 ETS + 0.44 0.92 10 Pooled dataRR = 0.31 ETS + 1.46 0.87 45 Pooled data *RR = 0.41 ETS + 0.51 0.88 44 * Same as the above with the largest value point removed Table I.- Regression eaquations, determination coefficients (r2), and number of data (n) for the different zooplankton groups and for all grouped. > 200 µm zooplankton Oithona nauplii II-IV > 50 µm, < 200 µm zooplankton * * Pooled data Groups ETS/RR SD N Authors Mixed copepods 0.46 0.09 6 1 Ctenophores0.69 0.19 15 1 Medusae 1.19 1.13 36 1 Mixed zooplankton 0.65 0.25 7 1 Mixed zooplankton 0.39 0.11 6 2 Acartia tonsa1.25 0.42 5 3 Acartia tonsa1.75 1.45 5 3 Acartia tonsa* 2.27 0.23 5 3 Mixed zooplankton 2.46 1.35 18 This study Mixed microzooplankton 1.75 0.66 17 This study Oithona Nauplii II-IV 2.07 0.59 10 This study Total (pooled data) 2.09 1.75 45 This study * Winter values ETS/RR ratios (Table II) were similar to those obtained by Bamstedt (1980) for Acartia tonsa. The values ranged from 1.75 to 2.46, with an average value for pooled data of 2.09. These values contrast with previous ones in which ETS gave potential respiration rates lower than those directly measured (seeTable II). The reason could be the improvement of the analysis of ETS activity by the two substrates method (Packard et al. 1996). The relationship between calculated and measured respiration rates, using the regression equation for the pooled data, and the average ETS/RR ratio explain the same percentage of the variance, 88 % in both cases (Fig. 6). Table II.- Table II.- Average ETS/RR ratios; standard deviation (SD), number of data (N) and authors: 1) King and Packard (1975); 2) Alcaraz and Packard (1989); 3) Bamstedt (1980). The numbers in red are potential respiration rates (ETS) lower than those measured. measured calculated (equations) calculated (ratios) This relatively good agreement obtained between observed and calculated respiration rates (Fig. 6), both using the regression equation for pooled data (A) and the average ETS/RR ratio (B), could be due to the relatively homogeneous environmental conditions (mainly temperature and food) experienced by all the zooplankton groups before catching them and during the incubation experiments. Fig. 6.- Relationships between calculated (abscissae) and measured (ordinates) respiration rates for the threegroups of zooplankton (pooled data). A: Equations for pooled data as in Table I. B: ETS/RR ratios (Table II). A B 0 5 10 15 20 25 30 35 0 20 40 60 80 100 120 measured µmol O 2 d -1 0 5 10 15 20 0 5 10 15 20 25 30 35 0 5 10 15 20 0 5 10 15 20 25 30 35 ETS µmol O 2 0 5 10 15 20 25 30 35 020 40 60 80 100 120 d -1 0 5 10 15 20 25 0 5 10 15 20 25 y = 0.0386 + 0.996x R 2= 0.877 0 5 10 15 20 25 0 5 10 15 20 25 y = 0.547 + 0.847x R 2= 0.877 •The ETS-RR regression equations and ETS/RR ratios for the different zooplankton components appear to be relatively constant. • At least when the experimental conditions (mainly temperature and food) are similar, both regression equations and ETS/RR ratios are robust predictors of actual respiration rates (RR). The variance explained is in both cases up to 88 %. • The factors affecting the variability of the relationships between ETS-derived and directly measured RR must be ascertained in order to efficiently use ETS activity as a proxy of respiration rates. 6. References 5. Conclusions