Biogeosciences www.biogeosciences.net 1726-4170 1726-4189 5 2 2008 10.5194/bg-5-371-2008 http://www.biogeosciences.net/5/371/2008/ http://www.biogeosciences.net/5/371/2008/bg-5-371-2008.html http://www.biogeosciences.net/5/371/2008/bg-5-371-2008.pdf 371 383 2008-03-12 Competition for inorganic and organic forms of nitrogen and phosphorous between phytoplankton and bacteria during an Emiliania huxleyi spring bloom T. Løvdal trond.lovdal@uis.no C. Eichner H.-P. Grossart V. Carbonnel L. Chou V. Martin-Jézéquel T. F. Thingstad Department of Biology, University of Bergen, Jahnebakken 5, PO Box 7800, 5020 Bergen, Norway Leibniz Institute of Freshwater Ecology and Inland Fisheries, Department of Limnology of Stratified Lakes, Alte Fischehuette 2, 16775 Stechlin, Germany Laboratoire d'Océanographie Chimique et Géochimie des Eaux, Université Libre de Bruxelles, Campus Plaine – CP 208, 1050 Brussels, Belgium Laboratoire 'Mer, Molécule, Santé', EA 2160, EA2663, Université de Nantes, BP 92208, 44322 Nantes, France present address: Department of Mathematics and Natural Sciences, Faculty of Science and Technology, University of Stavanger, 4036 Stavanger, Norway present address: Institute of Marine Research, PO Box 1870 Nordnes, 5817 Bergen, Norway Using 15N and 33P, we measured the turnover of organic and inorganic nitrogen (N) and phosphorus (P) substrates, and the partitioning of N and P from these sources into two size fractions of marine osmotrophs during the course of a phytoplankton bloom in a nutrient manipulated mesocosm. The larger size fraction (>0.8 μm), mainly consisting of the coccolithophorid Emiliania huxleyi, but also including an increasing amount of large particle-associated bacteria as the bloom proceeded, dominated uptake of the inorganic forms NH4+, NO3−, and PO43−. The uptake of N from leucine, and P from ATP and dissolved DNA, was initially dominated by the 0.8–0.2 μm size fraction, but shifted towards dominance by the >0.8 μm size fraction as the system turned to an increasing degree of N-deficiency. Normalizing uptake to biomass of phytoplankton and heterotrophic bacteria revealed that organisms in the 0.8–0.2 μm size fraction had higher specific affinity for leucine-N than those in the >0.8 μm size fraction when N was deficient, whereas the opposite was the case for NH4+. There was no such difference regarding the specific affinity for P substrates. Since heterotrophic bacteria seem to acquire N from organic compounds like leucine more efficiently than phytoplankton, our results suggest different structuring of the microbial food chain in N-limited relative to P-limited environments. Antia, N. J., Harrison, P. J., and Oliveira, L.: The role of dissolved organic nitrogen in phytoplankton nutrition, cell biology and ecology, Phycologia 30, 1–89, 1991. Becquevort, S., Rousseau, V., and Lancelot, C.: Major and comparable roles for free-living and attached bacteria in the degradation of \textitPhaeocystis-derived organic matter in Belgian coastal waters of the North Sea, Aquat. Microb. Ecol., 14, 39–48, 1998. Benitez-Nelson, C. R. and Buesseler, K. O.: Variability of inorganic and organic phosphorus turnover rates in the coastal ocean, Nature, 398, 502–505, 1999. Berges, J. A. and Falkowski, P. G.: Cell-associated proteolytic enzymes from marine phytoplankton, J. Phycol., 32, 566–574, 1996. Berman, T. and Bronk, D. A.: Dissolved organic nitrogen: a dynamic participant in aquatic systems, Aquat. Microb. Ecol., 31, 279–305, 2003. Billen, G.: Heterotrophic utilization and regeneration of nitrogen, in: Heterotrophic activity in the sea, edited by: Hobbie, J. E., Williams, P. J. L. B., Plenum Press, New York, London, 313–355, 1984. Bratbak, G. and Thingstad, T. F.: Phytoplankton-bacteria interactions: an apparent paradox? analysis of a model system with both competition and commensalism, Mar. Ecol. Prog. Ser., 25, 23–30, 1985. Chróst, R. J.: Microbial ectoenzymes in aquatic environments, in: Aquatic microbial ecology: Biochemical and molecular approaches, edited by: Overbeck, J., Chróst, R. J., Springer Verlag, 47–78, 1990. DeFlaun, M. F., Paul, J. H., and Jeffrey, W. H.: Distribution and molecular weight of dissolved DNA in subtropical estuarine and oceanic environments, Mar. Ecol. Prog. Ser., 38, 65–73, 1987. Diehl, H., Ihlefeld, H., and Schwegler, H.: Physik für Biologen, Springer-Verlag, 391 pp., 1991. Engel, A., Zondervan, I., Aerts, K., Beaufort, L., Benthien, A., Chou, L., Delille, B., Gattuso, J.-P., Harlay, J., Heemann, C., Hoffmann, L., Jacquet, S., Nejstgaard, J., Pizay, M.-D., Rochelle-Newall, E., Schneider, U., Terbrueggen, A., and Riebesell, U.: Testing the direct effect of CO2 concentration on a bloom of the coccolithophorid \textitEmiliania huxleyi in mesocosm experiment, Limnol. Oceanogr., 50, 493–507, 2005. Fernández, E., Marañón, E., Harbour, D. S., Kristiansen, S., and Heimdal, B. R.: Patterns of carbon and nitrogen uptake during blooms of \textitEmiliania huxleyi in two Norwegian fjords, J. Plankton Res., 18, 2349–2366, 1996. Grasshoff, K.: Determination of nitrate, in: Methods of seawater analysis, edited by: Grasshoff, K., Erhardt, M., and Kremling, K., Verlag Chemie, Weinheim, 143–150, 1983. Grossart, H.-P., Allgaier, M., Passow, U., and Riebesell, U.: Testing the effect of CO2 concentration on the dynamics of marine heterotrophic bacterioplankton, Limnol. Oceanogr., 51, 1–11, 2006a. Grossart, H.-P., Czub, G., and Simon, M.: Specific interactions of planktonic algae and bacteria: Implications for aggregation and organic matter cycling in the sea, Environ. Microbiol., 8, 1074–1084, 2006b. Havskum, H., Thingstad, T. F., Scharek, R., Peters, F., Berdalet, E., Sala, M. M., Alcaraz, M., Bangsholt, J. C., Zweifel, U. L., Hagström, Å., Perez, M., and Dolan, J. R.: Silicate and labile DOC interfere in structuring the microbial food web via algal-bacterial competiton for mineral nutrients: Results from a mesocosm experiment, Limnol. Oceanogr., 48, 129–140, 2003. Hillebrand, H., Dürselen, C. D., Kirschtel, D., Pollingher, U., and Zohary, T.: Biovolume calculation for pelagic and benthic microalgae, J. Phycol., 35, 403–424, 1999. Hollibaugh, J. T. and Azam, F.: Microbial degradation of dissolved proteins in seawater, Limnol. Oceanogr., 28, 1104–1116, 1983. Ietswaart, T., Schneider, P. J., and Prins, R. A.: Utilization of organic nitrogen sources by two phytoplankton species and a bacterial isolate in pure and mixed cultures, Appl. Environ. Microbiol., 60, 1554–1560, 1994. Jacobsen, A., Egge, J. K., and Heimdal, B. R.: Effects of increased concentration of nitrate and phosphate during a springbloom experiment in mesocosm, J. Exp. Mar. Biol. Ecol., 187, 239–251, 1995. Joint, I., Henriksen, P., Fonnes, G. A., Bourne, D., Thingstad, T. F., and Riemann, B.: Competition for inorganic nutrients between phytoplankton and bacterioplankton in nutrient manipulated mesocosms, Aquat. Microb. Ecol., 29, 145–159, 2002. Junk, G. and Svec, H. J.: The absolute abundance of the nitrogen isotopes in the atmosphere and compressed gas from various sources, Geochim. Cosmochim. Acta, 14, 234–243, 1958. Jørgensen, N. O. G. and Jacobsen, C. S.: Bacterial uptake and utilization of dissolved DNA, Aquat. Microb. Ecol., 11, 263–270, 1996. Jørgensen, N. O. G., Kroer, N., Coffin, R. B., Yang, X.-H., and Lee, C.: Dissolved free amino acids, combined amino acids, and DNA as sources of carbon and nitrogen to marine bacteria, Mar. Ecol. Prog. Ser., 98, 135–148, 1993. Karner, M. and Herndl, G. F.: Extracellular enzymatic activity and secondary production in free-living and marine-snow-associated bacteria, Mar. Biol., 113, 341–347, 1992. Koroleff, F.: Direct determination of ammonia in natural waters as indophenol blue, Council Meeting Document – International Council for the Exploration of the Sea, 19–22, 1969. Koroleff, F.: Determination of phosphorus, in: Methods in seawater analysis, edited by: Grasshoff, K., Erhardt, M., and Kremling, K., Verlag Chemie, 125–131, 1983. Kristiansen, S., Thingstad, T. F., van der Wal, P., Farbrot, T., and Skjoldal, E. F.: An \textitEmiliania huxleyi dominated subsurface bloom in Samnangerfjorden, western Norway. Importance of hydrography and nutrients, Sarsia, 79, 357–368, 1994. Kuenzler, E. J. and Perras, J. P.: Phosphatases of marine algae, Biol. Bull., 128, 271–284, 1965. Laws, E. A.: Photosynthetic quotients, new production and net community production in the open ocean, Deep-Sea Res., 38, 143–167, 1991. Lessard, E. J., Merico, A., and Tyrrell, T.: Nitrate: phosphate ratios and \textitEmiliania huxleyi blooms, Limnol. Oceanogr., 50, 1020–1024, 2005. Loferer-Krößbacher, M., Klima, J., and Psenner, R.: Determination of bacterial cell dry mass by transmission electron microscopy and densitometric image analysis, Appl. Environ. Microbiol., 64, 688–694, 1998. Løvdal, T., Skjoldal, E. F., Heldal, M., Norland, S., and Thingstad, T. F.: Changes in morphology and elemental composition of \textitVibrio splendidus along a gradient from carbon-limited to phosphate-limited growth, Microb. Ecol., 55, 152–161, 2008. Løvdal, T., Tanaka, T., and Thingstad, T. F.: Algal-bacterial competition for phosphorus from dissolved DNA, ATP, and orthophosphate in a mesocosm experiment, Limnol. Oceanogr. 52, 1407–1419, 2007. Lukacs, G. L., Haggie, P., Seksek, O., Lechardeur, D., Freedman, N., and Verkman, A. S.: Size-dependent DNA mobility in cytoplasm and nucleus, J. Biol. Chem., 275, 1625–1629, 2000. Martinez, J., and Azam, F.: Aminopeptidase activity in marine chroococcoid cyanobacteria, Appl. Environ. Microbiol., 59, 3701–3707, 1993. Menden-Deuer, S. and Lessard, E. J.: Carbon to volume relationships for dinoflagellates, diatoms, and other protist plankton, Limnol. Oceanogr., 45, 569–579, 2000. Middelboe, M., Søndergaard, M., Letarte, Y., and Borch, N. H.: Attached and free-living bacteria: Production and polymer hydrolysis during a diatom bloom, Microb. Ecol., 29, 231–248, 1995. Moal, J., Martin-Jézéquel, V., Harris, R. P., Samain, J. F., and Poulet, S. A.: Interspecific and intraspecific variability of the chemical composition of marine phytoplankton, Oceanologica Acta, 10, 339–346, 1987. Montagnes, D. J. S., Berges, J. A., Harrison, P. J., and Taylor, F. J. R.: Estimating carbon, nitrogen, protein, and chlorophyll $a$ from volume in marine phytoplankton, Limnol. Oceanogr., 39, 1044–1060, 1994. Montoya, J. P., Voss, M., Kähler, P., and Capone, D. G.: A simple, high-precision, high-sensitivity tracer assay for N2 fixation, Appl. Environ. Microbiol., 62, 986–993, 1996. Nimer, E., Schneiderman, R., and Maroudas, A.: Diffusion and partition of solutes in cartilage under static load, Biophys. Chem., 106, 125–146, 2003. Palenik, B. and Henson, S. E.: The use of amides and other organic nitrogen sources by the phytoplankton \textitEmiliania huxleyi, Limnol. Oceanogr., 42, 1544–1551, 1997. Pantoja, S. and Lee, C.: Cell-surface oxidation of amino acids in seawater, Limnol. Oceanogr., 39, 1718–1726, 1994. Paul, J. H.: Uptake of organic nitrogen, in: Nitrogen in the marine environment, edited by: Carpenter, E. J., and Capone, D. G., Academic Press, New York, 275–308, 1983. Pengerud, B., Skjoldal, E. F., and Thingstad, T. F.: The reciprocal interactions between degradation of glucose and ecosystem structure. Studies in mixed chemostat cultures of marine bacteria, algae, and bacterivorous nanoflagellates, Mar. Ecol. Prog. Ser., 35, 111–117, 1987. Perry, M. J.: Alkaline phosphatase activity in subtropical Central North Pacific waters using a sensitive fluorometric method, Mar. Biol., 15, 113–119, 1972. Paasche, E.: A review of the coccolithophorid \textitEmiliania huxleyi (Prymnesiophyceae), with particular reference to growth, coccolith formation, and calcification-photosynthesis interactions, Phycologia, 40, 503–529, 2002. Rees, A. P., Woodward, E. M. S., Robinson, C., Cummings, D. G., Tarran, G. A., and Joint, I.: Size-fractionated nitrogen uptake and carbon fixation during a developing coccolithophore bloom in the North Sea during June 1999, Deep-Sea Res. II, 49, 2905–2927, 2002. Riegman, R., Noordeloos, A. A. M. and Cadée, G. C.: \textitPhaeocystis blooms and eutrophication of the continental coastal zones of the North Sea, Mar. Biol., 112, 479–484, 1992. Riegman, R., Stolte, W., Noordeloos, A. A. M., and Slezak, D.: Nutrient uptake and alkaline phosphatase (EC 3:1:3:1) activity of \textitEmiliania huxleyi (Prymnesiophyceae) during growth under N and P limitation in continuous cultures, J. Phycol., 36, 87–96, 2000. Sambrook, J. and Russell, D. W.: Molecular cloning: A laboratory journal, Cold Spring Harbour Laboratory Press, 2001. Samuelsson, K., Berglund, J., Haecky, P., and Andersson, A.: Structural changes in an aquatic microbial food web caused by inorganic nutrient addition, Aquat. Microb. Ecol., 29, 29–38, 2002. Simon, M., Grossart, H.-P., Schweitzer, B., and Ploug, H.: Microbial ecology of organic aggregates in aquatic ecosystems, Aquat. Microb. Ecol., 28, 175–211, 2002. Smith, D. C., Simon, M., Alldredge, A. L., and Azam, F.: Intense hydrolytic enzyme activity on marine aggregates and implications for rapid particle dissolution, Nature, 359, 139–142, 1992. Sokal, R. R. and Rohlf, F. J.: Biometry, 3rd Ed., W. H. Freeman, 207–271, 1995. Tanaka, T., Henriksen, P., Lignell, R., Olli, K., Seppälä, J., Tamminen, T., and Thingstad, T. F.: Specific affinity for phosphate uptake and specific alkaline phosphatase activity as diagnostic tools for detecting P-limited phytoplankton and bacteria, Estuaries and Coasts, 29, 1226–1241, 2006. Thingstad, T. F.: Control of bacterial growth in idealized food webs, in: Microbial ecology of the oceans, edited by: Kirchman, D. L., Wiley-Liss, 229–260, 2000. Thingstad, T. F., Øvreås, L., Egge, J. K., Løvdal, T., and Heldal, M.: Use of non-limiting substrates to increase size; a generic strategy to simultaneosly optimize uptake and minimize predation in pelagic osmotrophs?, Ecol. Lett., 8, 675–682, 2005. Thingstad, T. F. and Rassoulzadegan, F.: Conceptual models for the biogeochemical role of the photic zone microbial food web, with particular reference to the Mediterranean Sea, Prog. Oceanogr., 44, 271–286, 1999. Thingstad, T. F., Skjoldal, E. F., and Bohne, R. A.: Phosphorus cycling and algal-bacterial competition in Sandsfjord, western Norway, Mar. Ecol. Prog. Ser., 99, 239–259, 1993. Utermöhl, H.: Zur vervolkommung der quantitativen Phytoplankton-methodik, Mitteilungen – Internationale Vereinigung für Limnologie, 9, 1–38, 1958.