Skip to main content Accessibility help

Growth, nitrogen fixation, respiration, and a nitrogen budget for cultures of a cosmopolitan diazotrophic endosymbiont (Teredinibacter turnerae) of shipworms

  • Rachel E.A. Horak (a1) and Joseph P. Montoya (a1)


Wood-boring bivalves (Bivalvia, family Teredinidae), also known as shipworms, host dinitrogen-fixing and cellulolytic symbiotic bacteria in gill bacteriocytes, which may be a necessary adaptation to a wooden diet. Although oxygen (O2) inhibits nitrogenase in other species, symbionts are able to fix nitrogen (N) within the gill tissue and provide newly fixed N to the host shipworm. The recent direct evidence of new N incorporation into the host tissue indicates that there are potentially complex nutrient cycles in this symbiosis and uninvestigated controls upon these cycles.To elucidate the mechanisms of this unique N2-fixing symbiosis and determine whether symbionts can excrete newly fixed N, we measured rates of growth, N2-fixation, respiration, and inorganic N content for the cultivated symbiont Teredinibacter turnerae (γ-proteobacteria, strain T7901) under a range of headspace O2 conditions. In all conditions, headspace O2 did not affect maximum specific N2-fixation and respiration activity, but did influence the rate and timing of growth. These results are consistent with the development of microaerobic conditions through an oxygen gradient in the culture medium, which facilitates N2-fixation and growth. The medium accumulated a small amount of NH4+, which represented 0.5–2.5% of the total N fixed by the culture. We constructed a simple N budget for T. turnerae to assess the role of the major known N sources and sinks. The N budget was not closed, indicating that new N is allocated to currently unidentified sinks, which may include excreted dissolved organic nitrogen.


Corresponding author

Correspondence should be addressed to: R.E.A. Horak, University of Washington, School of Oceanography, Box 357940, Seattle, WA 98195, USA email:


Hide All
Beshay, U. (2003) Production of alkaline protease by Teredinobacter turnirae cells immobilized in Ca-alginate beads. African Journal of Biotechnology 2, 6065.
Bradford, M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principal of protein-dye binding. Analytical Biochemistry 72, 248254.
Brune, A. and Friedrich, M. (2000) Microecology of the termite gut: structure and function on a microscale. Current Opinion in Microbiology 3, 263269.
Capone, D. (1994) Amino acid cycling in colonies of the planktonic marine cyanobacterium Trichdesmium thiebautii. Applied and Environmental Microbiology 60, 39893995.
Capone, D. and Montoya, J.P. (2001) Nitrogen fixation and denitrification. Methods in Microbiology 30, 501515.
Delatolla, R., Berk, D. and Tufenkji, N. (2008) Rapid and reliable quantification of biofilm weight and nitrogen content of biofilm attached to polystyrene beads. Water Research 42, 30823088.
Distel, D. (2003) The biology of marine wood-boring bivalves and their bacterial endosymbionts. In Goodell, B., Nicholas, D. and Schultz, T. (eds) Wood deterioration and preservation. Washington, DC: American Chemical Society, pp. 253271.
Distel, D., DeLong, E. and Waterbury, J. (1991) Phylogenetic characterization and in situ localization of the bacterial symbiont of shipworms (Teredinidae: Bivalvia) by using 16S rRNA sequence analysis and oligodeoxynucleotide probe hybridization. Applied and Environmental Microbiology 57, 23762382.
Distel, D., Morrill, N., MacLaren-Toussaint, N., Franks, D. and Waterbury, J. (2002) Teredinibacter turnerae gen. nov., sp. nov., a dinitrogen-fixing, cellulolytic, endosymbiotic gamma-proteobacterium isolated from the gills of wood-boring molluscs (Bivalvia: Tereinidae). International Journal of Systematics and Evolutionary Microbiology 52, 22612269.
Dixon, R. and Kahn, D. (2004) Genetic regulation of biological nitrogen fixation. Nature Reviews Microbiology 2, 621631.
Ferreira, G., Ahuja, S., Sierks, M. and Moreira, A. (2001) Pleomorphism of the marine bacterium Teredinobacter turnirae. Letters in Applied Microbiology 33, 5660.
Foster, R., Kuypers, M., Vagner, T., Paerl, R., Musat, N. and Zehr, J. (2011) Nitrogen fixation and transfer in open ocean diatom-cyanobacterial symbioses. ISME Journal 5, 14841493.
Gallager, S., Turner, R. and Berg, C.J. Jr (1981) Physiological aspects of wood consumption, growth, and reproduction in the shipworm Lyrodus pedicellatus (Bivalvia: Teredinidae). Journal of Experimental Marine Biology and Ecology 52, 6377.
Garcia, H. and Gordon, L. (1992) Oxygen solubility in seawater: better fitting equations. Limnology and Oceanography 37, 13071312.
Glibert, P. and Bronk, D. (1994) Release of dissolved organic nitrogen by marine diazotrophic cyanobacteria, Trichodesmium spp. Applied and Environmental Microbiology 60, 39964000.
Greene, R. (1994) Challenges from the sea: marine shipworms and their symbiotic bacterium. Society of Industrial Microbiology News 44, 5159.
Greene, R., Cotta, M. and Griffin, H. (1989) A novel, symbiotic bacteria isolated from marine shipworm secretes proteolytic activity. Current Microbiology 19, 353356.
Greene, R. and Freer, S. (1986) Growth characteristics of a novel nitrogen-fixing cellulolytic bacterium. Applied and Environmental Microbiology 52, 982986.
Griffin, H., Greene, R. and Cotta, M. (1992) Isolation and characterization of an alkaline protease from the marine shipworm bacterium. Current Microbiology 24, 111117.
Großkopf, T. and LaRoche, J. (2012) Direct and indirect costs of nitrogen fixation in Crocosphaera watsonii WH8501 and possible implications for the nitrogen cycle. Frontiers in Microbiology 3, article 236.
Hill, S. (1988) How is nitrogenase regulated by oxygen? FEMS Microbiology Reviews 54, 111130.
Holl, C. and Montoya, J. (2008) Diazotrophic growth of the marine cyanobacterium Trichodesmium IMS101 in continuous culture: effects of growth rate on N2-fixation rate, biomass, and C:N:P stoichiometry. Journal of Phycology 44, 929937.
Kuhla, J., Dingler, C. and Oelze, J. (1985) Production of extracellular nitrogen-containing components by Azotobacter vinelandii fixing dinitrogen in oxygen-controlled continuous culture. Archives of Microbiology 141, 297302.
Lechene, C., Luyten, Y., McMahon, G. and Distel, D. (2007) Quantitative imaging of nitrogen fixation by individual bacteria within animal cells. Science 317, 15631566.
Lechene, C., Hillion, F., McMahon, G., Benson, D., Kleinfeld, A., Kampf, J., Distel, D., Luyten, Y., Bonventre, J., Hentschel, D., Park, K., Ito, S., Schwart, M., Begichou, G. and Slodzian, G. (2006) High-resolution quantitative imaging of mammalian and bacterial cells using stable isotope mass spectrometry. Journal of Biology 5, 20.
Loladze, I. and Elser, J. (2011) The origins of the Redfield nitrogen-to-phosphorus ratio are in a homeostatic protein-to-rRNA ratio. Ecology Letters 14, 244250.
Marchal, K. and Vanderleyden, J. (2000) The ‘oxygen paradox’ of dinitrogen-fixing bacteria. Biology and Fertility of Soils 30, 363373.
Meeks, J., Enderlin, C., Joseph, C., Chapman, J. and Lollar, M. (1985) Fixation of [13N]N2 and transfer of fixed nitrogen in the Anthoceros–Nostoc symbiotic association. Planta 164, 406414.
Mulholland, M. and Bernhardt, P. (2005) The effect of growth rate, phosphorus concentration, and temperature on N2 fixation, carbon fixation, and nitrogen release in continuous cultures of Trichodesmium IMS101. Limnology and Oceanography 50, 839849.
Mulholland, M. and Capone, D. (1999) Nitrogen fixation, uptake, and metabolism in natural and cultured populations of Trichodesmium spp. Marine Ecology Progress Series 188, 3349.
Neidhardt, F.C. and Umbarger, H. (1996) Chemical composition of Escherichia coli. In Neidhardt, F.C. (ed) Escherichia coli and salmonella: cellular and molecular biology, Volume 1. 2nd edition. Washington, DC: American Society of Microbiology Press, pp. 13–16.
Ohkuma, M. (2001) Symbiosis within the gut microbial community of termites. RIKEN Review 41, 6972.
Peters, G. (1977) The Azolla-Anabaena azollae symbiosis. In Hollaender, A., Burns, R., Day, P., Hardy, R., Helinski, D., Lamborg, M., Owens, L. and Volentine, R. (eds) Genetic engineering for nitrogen fixation. New York: Plenum Press, pp. 231257.
Silvester, W., Parsons, R. and Watt, P. (1996) Direct measurement of release and assimilation of ammonia in the Gunnera–Nostoc symbiosis. New Phytologist 132, 617625.
Sterner, R. and Elser, J. (2002) Ecological stoichiometry: the biology of elements from molecules to the biosphere. Princeton, NJ: Princeton University Press.
Streams, M., Fisher, C. and Fiala-Medioni, A. (1997) Methanotrophic symbiont location and fate of carbon incorportated from methane in a hydrocarbon seep mussel. Marine Biology 129, 465476.
Strickland, J. and Parsons, T. (1972) A practical handbook of seawater analysis. 2nd edition. Ottawa: Bulletin of Fisheries Research Board of Canada.
Trindade-Silva, A., Machado-Forrera, E., Senra, M., Vizzoni, V., Ypanaguine, L., Leoncini, O. and Soares, C. (2009) Physiological traits of the symbiotic bacterium Teredinibacter turnerae isolated from the mangrove shipworm Neoteredo reynei. Genetics and Molecular Biology 32, 572581.
Trytek, R. and Allen, W. (1980) Synthesis of essential amino acids by bacterial symbionts in the gills of the shipworm Bankia setacea (Tryon). Comparative Biochemistry and Physiology 67A, 419427.
Turner, R. (1984) An overview of research on marine borers: past progress and future direction. In Costlow, J. and Tipper, R. (eds) Marine biodeterioration: an interdisciplinary study. Annapolis, MD: Naval Institute Press, pp. 316.
Van Dover, C. (2000) The ecology of deep-sea hydrothermal vents. Princeton, NJ: Princeton University Press.
Waterbury, J., Calloway, C. and Turner, R. (1983) A cellulolytic nitrogen-fixing bacterium cultured from the gland of Deshayes in shipworms (Bivalvia: Teredinidae). Science 221, 14011403.
Waters, J., Hughes II, B., Purcell, L., Gerhardt, K., Mawhinney, T. and Emerich, D. (1998) Alanine, not ammonia, is excreted from N2-fixing soybean nodule bacteroids. Proceedings of the National Academy of Sciences of the United States of America 95, 1203812042.


Growth, nitrogen fixation, respiration, and a nitrogen budget for cultures of a cosmopolitan diazotrophic endosymbiont (Teredinibacter turnerae) of shipworms

  • Rachel E.A. Horak (a1) and Joseph P. Montoya (a1)


Full text views

Total number of HTML views: 0
Total number of PDF views: 0 *
Loading metrics...

Abstract views

Total abstract views: 0 *
Loading metrics...

* Views captured on Cambridge Core between <date>. This data will be updated every 24 hours.

Usage data cannot currently be displayed