Microbial Ecology

, Volume 55, Issue 1, pp 152–161 | Cite as

Changes in Morphology and Elemental Composition of Vibrio splendidus Along a Gradient from Carbon-limited to Phosphate-limited Growth

  • Trond Løvdal
  • Evy F. Skjoldal
  • Mikal Heldal
  • Svein Norland
  • T. Frede Thingstad


We examined morphology, elemental composition (C, N, P), and orthophosphate-uptake efficiency in the marine heterotrophic bacterium Vibrio splendidus grown in continuous cultures. Eight chemostats were arranged along a gradient of increasing glucose concentrations in the reservoirs, shifting the limiting factor from glucose to phosphate. The content of carbon, nitrogen, and phosphorus was measured in individual cells by x-ray microanalysis using a transmission electron microscope (TEM). Cell volumes (V) were estimated from length and width measurements of unfixed, air-dried cells in TEM. There was a transition from coccoid cells in C-limited cultures toward rod-shaped cells in P-limited cultures. Cells in P-limited cultures with free glucose in the media were significantly larger than cells in glucose-depleted cultures (P < 0.0001). We found functional allometry between cellular C-, N-, and P content (in femtograms) and V (in cubic micrometers) in V. splendidus (C = 224 × V 0.89, N = 52.5 × V 0.80, P = 2 × V 0.65); i.e., larger bacteria had less elemental C, N, and P per V than smaller cells, and also less P relative to C. Biomass-specific affinity for orthophosphate uptake in large P-limited V. splendidus approached theoretical maxima predicted for uptake limited by molecular diffusion toward the cells. Comparing these theoretical values to respective values for the smaller, coccoid, C-limited V. splendidus indicated, contrary to the traditional view, that large size did not represent a trade-off when competing for the non-C-limiting nutrients.


Vibrio Soluble Reactive Phosphorus Free Glucose Cell Quota Late Logarithmic Phase 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



This work was supported by the Strategic Institution Programme “Patterns in Biodiversity” Contract 158936/S40 from the Research Council of Norway. The FACSCalibur flow cytometer was in part funded by a grant from The Knut and Alice Wallenberg Foundation to the Virtue program. We gratefully thank Philippe Lebaron for providing the V. splendidus culture, Tsuneo Tanaka for advice and assistance, George Jackson for providing information on diffusion literature, and Egil S. Erichsen for his assistance at the Laboratory for Electron Microscopy (LEM), Science Faculty, University of Bergen.


  1. 1.
    Bratbak, G (1985) Bacterial biovolume and biomass estimations. Appl Environ Microbiol 49: 1488–1493PubMedGoogle Scholar
  2. 2.
    Clift, R, Grace, JR, Weber, ME (1978) Bubbles, drops, and particles. Academic Press, New YorkGoogle Scholar
  3. 3.
    Currie, DJ, Kalff, J (1984) A comparison of the abilities of freshwater algae and bacteria to acquire and retain phosphorus. Limnol Oceanogr 29: 298–310Google Scholar
  4. 4.
    Droop, MR (1974) The nutrient status of algal cells in continuous culture. J Mar Biol Assoc UK 54: 825–855CrossRefGoogle Scholar
  5. 5.
    Fagerbakke, KM, Heldal, M, Norland, S (1996) Content of carbon, nitrogen, oxygen, sulfur and phosphorus in native aquatic and cultured bacteria. Aquat Microb Ecol 10: 15–27CrossRefGoogle Scholar
  6. 6.
    Flaten, GAF (2004) Phosphate limitation in marine osmotrophs: affinity and competition. DSc thesis, University of Bergen, BergenGoogle Scholar
  7. 7.
    Goldman, JC, Caron, DA, Dennet, MR (1987) Regulation of gross growth efficiency and ammonium regeneration in bacteria by substrate C:N ratio. Limnol Oceanogr 32: 1239–1252Google Scholar
  8. 8.
    Gundersen, K, Heldal, M, Norland, S, Purdie, DA, Knap, AH (2002) Elemental C, N, and P cell content of individual bacteria collected at the Bermuda Atlantic time-series study (BATS) site. Limnol Oceanogr 47: 1525–1530CrossRefGoogle Scholar
  9. 9.
    Jumars, PA (1993) Concepts in biological oceanography: an interdisciplinary primer. Oxford University Press, OxfordGoogle Scholar
  10. 10.
    Jumars, PA, Deming, JW, Hill, PS, Karp-Boss, L, Yager, PL, Dade, WB (1993) Physical constraints on marine osmotrophy in an optimal foraging context. Mar Microb Food Webs 7: 121–159Google Scholar
  11. 11.
    Karp-Boss, L, Boss, E, Jumars, PA (1996) Nutrient fluxes to planktonic osmotrophs in the presence of fluid motion. Oceanogr Mar Biol Ann Rev 34: 71–107Google Scholar
  12. 12.
    Klausmeier, CA, Litchman, E, Levin, SA (2004) Phytoplankton growth and stoichiometry under multiple nutrient limitation. Limnol Oceanogr 49: 1463–1470CrossRefGoogle Scholar
  13. 13.
    Koch, AL (1996) What size should a bacterium be? a question of scale. Annu Rev Microbiol 50: 317–348PubMedCrossRefGoogle Scholar
  14. 14.
    Koroleff, F (1983) Determination of phosphorus. In: Grasshoff, K, Erhardt, M and Kremling, K (Eds.) Methods in seawater analysis. Verlag Chemie, pp 125–131Google Scholar
  15. 15.
    La Ferla, R, Leonardi, M (2005) Ecological implications of biomass and morphotype variations of bacterioplankton: an example in a coastal zone of the Northern Adriatic Sea (Mediterranean). Mar Ecol 26: 82–88CrossRefGoogle Scholar
  16. 16.
    Lee, S, Fuhrman, JA (1987) Relationships between biovolume and biomass of naturally derived marine bacterioplankton. Appl Environ Microbiol 53: 1298–1303PubMedGoogle Scholar
  17. 17.
    Loferer-Krössbacher, M, Klima, J, Psenner, R (1998) Determination of bacterial cell dry mass by transmission electron microscopy and densitometric image analysis. Appl Environ Microbiol 64: 688–694PubMedGoogle Scholar
  18. 18.
    Makino, W, Cotner, JB, Sterner, RW, Elser, JJ (2003) Are bacteria more like plants or animals? growth rate and resource dependence of bacterial C : N : P stoichiometry. Funct Ecol 17: 121–130CrossRefGoogle Scholar
  19. 19.
    Marie, D, Partensky, F, Jacquet, S, Vaulot, D (1997) Enumeration and cell cycle analysis of natural populations of marine picoplankton by flow cytometry using the nucleic acid stain SYBR Green I. Appl Environ Microbiol 63: 186–193PubMedGoogle Scholar
  20. 20.
    Martinussen, I, Thingstad, TF (1987) Utilization of N, P and organic C by heterotrophic bacteria. II. Comparison of experiments and a mathematical model. Mar Ecol Prog Ser 37: 285-293CrossRefGoogle Scholar
  21. 21.
    Morita, RY (1982) Starvation-survival of heterotrophs in the marine environment. Adv Microb Ecol 6: 171–198Google Scholar
  22. 22.
    Moutin, T, Thingstad, TF, van Wambeke, F, Marie, D, Slawyk, G, Raimbault, P, Claustre, H (2002) Does competition for nanomolar phosphate supply explain the predominance of the cyanobacterium Synechococcus? Limnol Oceanogr 47: 1562–1567CrossRefGoogle Scholar
  23. 23.
    Nissen, H, Heldal, M, Norland, S (1987) Growth, elemental composition, and formation of polyphosphate bodies in Vibrio natriegens cultures shifted from phosphate-limited to phosphate pulsed media. Can J Microbiol 33: 583–588CrossRefGoogle Scholar
  24. 24.
    Norland, S, Fagerbakke, KM, Heldal, M (1995) Light element analysis of individual bacteria by X-ray microanalysis. Appl Environ Microbiol 61: 1357–1362PubMedGoogle Scholar
  25. 25.
    Norland, S, Heldal, M, Tumyr, O (1987) On the relation between dry matter and volume of bacteria. Microb Ecol 13: 95–101CrossRefGoogle Scholar
  26. 26.
    Novitsky, JA, Morita, RY (1976) Morphological characterization of small cells resulting from nutrient starvation of a psychrophilic marine vibrio. Appl Environ Microbiol 32: 617–622PubMedGoogle Scholar
  27. 27.
    Nyström, T, Olsson, RM, Kjelleberg, S (1992) Survival, stress resistance, and alterations in protein expression in the marine Vibrio sp. strain S14 during starvation for different individual nutrients. Appl Environ Microbiol 58: 55–65PubMedGoogle Scholar
  28. 28.
    Pengerud, B, Skjoldal, EF, Thingstad, TF (1987) 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: 111-117CrossRefGoogle Scholar
  29. 29.
    Pernthaler, J, Sattler, B, Simek, K, Schwarzenbacher, A, Psenner, R (1996) Top–down effects on the size-biomass distribution of a freshwater bacterioplankton community. Aquat Microb Ecol 10: 255–263CrossRefGoogle Scholar
  30. 30.
    Perry, MJ (1972) Alkaline phosphatase activity in subtropical Central North Pacific waters using a sensitive fluorometric method. Mar Biol 15: 113–119CrossRefGoogle Scholar
  31. 31.
    Schulz, HN, Jørgensen, BB (2001) Big bacteria. Annu Rev Microbiol 55: 105–137PubMedCrossRefGoogle Scholar
  32. 32.
    Shannon, SP, Chrzanowski, TH, Grover, JP (2007) Prey food quality affects flagellate ingestion rates. Microb Ecol 53: 66–73PubMedCrossRefGoogle Scholar
  33. 33.
    Sokal, RR, Rohlf, FJ (1995) Biometry, 3rd ed. W.H. Freeman, New YorkGoogle Scholar
  34. 34.
    Sterner, RW, Elser, JJ (2002) Ecological stoichiometry. The biology of elements from molecules to the biosphere. Princeton University Press, Princeton, NJGoogle Scholar
  35. 35.
    Takeuchi, M, Sawada, H, Oyaizu, H, Yokota, A (1994) Phylogenetic evidence for Sphingomonas and Rhizomonas as nonphotosynthetic members of the alpha-4 subclass of the Proteobacteria. Int J Syst Bacteriol 44: 308–314PubMedCrossRefGoogle Scholar
  36. 36.
    Tezuka, Y (1990) Bacterial regeneration of ammonium and phosphate as affected by the carbon:nitrogen:phosphorus ratio of organic substrates. Microb Ecol 19: 227–238CrossRefGoogle Scholar
  37. 37.
    Thingstad, TF (1987) Utilization of N, P, and organic C by heterotrophic bacteria. I. Outline of a chemostat theory with a consistent concept of ‘maintenance' metabolism. Mar Ecol Prog Ser 35: 99–109CrossRefGoogle Scholar
  38. 38.
    Thingstad, TF, Rassoulzadegan, F (1999) Conceptual models for the biogeochemical role of the photic zone microbial food web, with particular reference to the Mediterranean Sea. Prog Oceanogr 44: 271–286CrossRefGoogle Scholar
  39. 39.
    Thingstad, TF, Skjoldal, EF, Bohne, RA (1993) Phosphorus cycling and algal-bacterial competition in Sandsfjord, western Norway. Mar Ecol Prog Ser 99: 239–259CrossRefGoogle Scholar
  40. 40.
    Thingstad, TF, Øvreås, L, Egge, JK, Løvdal, T, Heldal, M (2005) Use of non-limiting substrates to increase size; a generic strategy to simultaneously optimize uptake and minimize predation in pelagic osmotrophs? Ecol Lett 8: 675–682CrossRefGoogle Scholar
  41. 41.
    Thompson, JR, Randa, MA, Marcelino, LA, Tomita-Mitchell, A, Lim, E, Polz, MF (2004) Diversity and dynamics of a North Atlantic coastal Vibrio community. Appl Environ Microbiol 70: 4103–4110PubMedCrossRefGoogle Scholar
  42. 42.
    Vadstein, O (1998) Evaluation of competitive ability of two heterotrophic planktonic bacteria under phosphorus limitation. Aquat Microb Ecol 14: 119–127CrossRefGoogle Scholar
  43. 43.
    Vrede, K, Heldal, M, Norland, S, Bratbak, G (2002) Elemental composition (C, N, P) and cell volume of exponentially growing and nutrient-limited bacterioplankton. Appl Environ Microbiol 68: 2965–2971PubMedCrossRefGoogle Scholar
  44. 44.
    Wyman, M, Gregory, RPF, Carr, NG (1985) Novel role of phycoerythrin in a marine cyanobacterium, Synechococcus strain Dc2. Science 230: 818–820PubMedCrossRefGoogle Scholar
  45. 45.
    Øvreås, L, Bourne, D, Sandaa, R-A, Casamayor, EO, Benlloch, S, Goddard, V, Smerdon, G, Heldal, M, Thingstad, TF (2003) Response of bacterial and viral communities to nutrient manipulations in seawater mesocosms. Aquat Microb Ecol 31: 109–121CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Trond Løvdal
    • 1
  • Evy F. Skjoldal
    • 1
  • Mikal Heldal
    • 1
  • Svein Norland
    • 1
  • T. Frede Thingstad
    • 1
  1. 1.Department of BiologyUniversity of BergenBergenNorway

Personalised recommendations