Original Article | Published:

Microbial Ecology and Functional Diversity of Natural Habitats

Diazotrophic bacterioplankton in a coral reef lagoon: phylogeny, diel nitrogenase expression and response to phosphate enrichment

The ISME Journal volume 1, pages 7891 (2007) | Download Citation

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Abstract

We investigated diazotrophic bacterioplankton assemblage composition in the Heron Reef lagoon (Great Barrier Reef, Australia) using culture-independent techniques targeting the nifH fragment of the nitrogenase gene. Seawater was collected at 3 h intervals over a period of 72 h (i.e. over diel as well as tidal cycles). An incubation experiment was also conducted to assess the impact of phosphate (PO43−) availability on nifH expression patterns. DNA-based nifH libraries contained primarily sequences that were most similar to nifH from sediment, microbial mat and surface-associated microorganisms, with a few sequences that clustered with typical open ocean phylotypes. In contrast to genomic DNA sequences, libraries prepared from gene transcripts (mRNA amplified by reverse transcription-polymerase chain reaction) were entirely cyanobacterial and contained phylotypes similar to those observed in open ocean plankton. The abundance of Trichodesmium and two uncultured cyanobacterial phylotypes from previous studies (group A and group B) were studied by quantitative-polymerase chain reaction in the lagoon samples. These were detected as transcripts, but were not detected in genomic DNA. The gene transcript abundance of these phylotypes demonstrated variability over several diel cycles. The PO43− enrichment experiment had a clearer pattern of gene expression over diel cycles than the lagoon sampling, however PO43− additions did not result in enhanced transcript abundance relative to control incubations. The results suggest that a number of diazotrophs in bacterioplankton of the reef lagoon may originate from sediment, coral or beachrock surfaces, sloughing into plankton with the flooding tide. The presence of typical open ocean phylotype transcripts in lagoon bacterioplankton may indicate that they are an important component of the N cycle of the coral reef.

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References

  1. , , , , , et al. (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucl Acid Res 25: 3389–3402.

  2. . (1998). Microbial control of oceanic carbon flux: the plot thickens. Science 280: 694.

  3. , , , . (2004). The demise of the marine cyanobacterium, Trichodesmium spp. via an autocatalyzed cell death pathway. Limnol Oceanogr 49: 997–1005.

  4. , . (1996). Effects of nitrogen limitation and starvation on Chroococcidiopsis sp. (Chroococcales). New Phytol 133: 563–571.

  5. , , , , . (2000). Ionizing-radiation resistance in the desiccation-tolerant cyanobacterium Chroococcidiopsis. Appl Environ Microbiol 66: 1489–1492.

  6. , , , . (2005). Spatial distribution and transcriptional activity of an uncultured clade of planktonic diazotrophic gamma-proteobacteria in the Arabian Sea. Appl Environ Microbiol 71: 2079–2085.

  7. , , , . (2004). Bacterial life and dinitrogen fixation at a gypsum rock. Appl Environ Microbiol 70: 7070–7077.

  8. . (1996). Coral reef ecosystems in the context of the marine nitrogen cycle. In: Bjork M, Semesi AK, Pedersen M, Bergman B (eds). Current Trends in Marine Botanical Research in the East African Region. SIDA, University of Mauritius: Mahe, pp 61–76.

  9. , , , . (1992). Microbial nitrogen transformations in unconsolidated coral reef sediments. Mar Ecol Progr Ser 80: 75–88.

  10. , . (1991). Major role of the cyanobacterium Trichodesmium in nutrient cycling in the North Atlantic Ocean. Science 254: 1356–1358.

  11. , . (2005). Dinitrogen fixation by exposed communities on the rum of Tikehau atoll (Tuamotu Archipelago, French Polynesia). Coral Reefs 24: 622–628.

  12. , , . (2001). Atmospheric dinitrogen fixation by benthic communities of Tikehau Lagoon (Tuamotu Archipelago, French Polynesia) and its contribution to benthic primary production. Mar Biol 139: 991–997.

  13. , , . (1990). Nutrient budget of the lagoonal waters in an open central South Pacific atoll (Tikehau, Tuamotu, French Polynesia). Mar Biol 107: 67–73.

  14. . (2001). Phosphorus supply for atoll biological productivity. Coral Reefs 20: 357–360.

  15. . (2005). Importance of photosynthetic picoplankton in coral reef ecosystems. Vie et Milieu Life and Environment 55: 217–223.

  16. , , , . (2005a). Vertical distributions of nitrogen-fixing phylotypes at Stn ALOHA in the oligotrophic North Pacific Ocean. Aquat Microb Ecol 38: 3–14.

  17. , , , , . (2005b). Temporal patterns of nitrogenase gene (nifH) expression in the oligotrophic North Pacific Ocean. Appl Environ Microbiol 71: 5362–5370.

  18. , , , , . (2002). Characterisation of the bacterial consortium associated with black band disease in coral using molecular microbiological techniques. Environ Microbiol 4: 401–413.

  19. , , , . (2001). Atoll morphology controls lagoon nutrient regime. Limnol Oceanogr 46: 456–461.

  20. . (1998). Profile hidden Markov models. Bioinformatics 14: 755–763.

  21. , , , , . (2004). N2 fixation by unicellular bacterioplankton from the Atlantic and Pacific Oceans: phylogeny and in situ rates. Appl Environ Microbiol 70: 765–770.

  22. , , , . (2002). Diversity of diazotrophic unicellular cyanobacteria in the tropical North Atlantic Ocean. Appl Environ Microbiol 68: 5760–5764.

  23. , . (1998). Biomass, production and grazing rates of pico- and nanoplankton in coral reef waters (Miyako Island, Japan). Microb Ecol 35: 46–57.

  24. , , , . (1988). Extraction of DNA suitable for molecular biological studies from natural planktonic microorganisms. Appl Environ Microbiol 54: 1426–1429.

  25. . (1981). The oxygen sensitivity of nitrogenase – a problem for biochemists and microorganisms. Trend Biochem Sci 6: 19–23.

  26. , , , , . (2003). Long-term region-wide declines in carribean corals. Science 301: 958–960.

  27. . (1976). Some physical and biological determinants of coral community structure in the Eastern Pacific. Ecol Monogr 46: 431–456.

  28. . (1991). Coral reef bleaching in the 1980s and possible connections with global warming. Trend Ecol Evol 6: 175–179.

  29. , . (1992). Effect of oxygen and temperature on the induction of a circadian nitrogenase activity rhythm in Synechococcus RF-1. J Plant Physiol 140: 391–394.

  30. , . (1999). Reef framework diagenesis across wave-flushed oxic-suboxic-anoxic transition zones. Coral Reefs 18: 229–240.

  31. . (1999). BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl Acid Symp Ser 41: 95–98.

  32. , . (2004). Bacterioplankton species richness and diversity along an estuarine gradient in Moreton Bay, Australia. Appl Environ Microbiol 70: 3425–3433.

  33. , . (2006). Spatial and vertical biogeography of coral reef sediment bacterial and diazotroph communities. Mar Ecol Progr Ser 306: 79–86.

  34. , , , , . (2004). Evidence for Trichodesmium viral lysis and potential significance for biogeochemical cycling in the oligotrophic ocean. Aquat Microb Ecol 36: 1–8.

  35. . (1992). Physiology of nitrogen fixation in free-living heterotrophs. In: Stacey G, Burris RH, Evans HJ (eds). Biological Nitrogen Fixation. Chapman & Hall: New York, pp 87–134.

  36. , , , , , et al. (2003). Climate Change, human impacts, and the resilience of coral reefs. Science 301: 929–933.

  37. . (2004). Tropical Archaea: diversity associated with the surface microlayer of corals. Mar Ecol Progr Ser 273: 81–88.

  38. , . (1984). Inhibition of nitrogenase activity by ammonium chloride in Azotobacter vinelandii cells. J Bacteriol 157: 148–151.

  39. , , , , , et al. (2001). ENCORE: The effect of nutrient enrichment on coral reefs. Synthesis of results and conclusions. Mar Poll Bull 42: 91–120.

  40. , , . (2005). Diazotrophic diversity and distribution in the tropical and subtropical Atlantic Ocean. Appl Environ Microbiol 71: 7910–7919.

  41. , , , , . (2004). PCR analysis of the distribution of unicellular cyanobacterial diazotrophs in the Arabian Sea. Appl Environ Microbiol 70: 7355–7364.

  42. . (1992). Regulation of nitrogen fixation genes in free-living and symbiotic bacteria. In: Stacey G, Burris RH, Evans HJ (eds). Biological Nitrogen Fixation. Chapman & Hall: New York, pp 835–876.

  43. , , , , , . (2006). Application of a nifH oligonucleotide microarray for profiling diversity of N2-fixing microorganisms in marine microbial mats. Environ Microbiol 8: 1721–1735.

  44. , , , , , . (2004). High rates of N2 fixation by unicellular diazotrophs in the oligotrophic Pacific Ocean. Nature 430: 1027–1031.

  45. , . (1990). Productivity and growth rates of coral reef bacteria on hard calcareous substrated and in sandy sediments in summer. Aust J Mar Fresh Res 41: 785–794.

  46. , , , , . (1985). Productivity of bacteria and microalgae and the effect of grazing by holothurians in sediments on a coral reef flat. Mar Biol 85: 293–300.

  47. , . (1994). Ingestion of the cyanobacterium Trichodesmium spp. by pelagic harpacticoid copepods Macrosetella, Miracia and Oculosetella. Hydrobiol 292/293: 235–240.

  48. . (1999). A possible role of temperate phage in the regulation of Trichodesmium biomass. Bull d l'Inst Oceanogr Monaco 19: 287–292.

  49. , , , . (2004a). Comparison of diazotroph community structure in Lyngbya sp and Microcoleus chthonoplastes dominated microbial mats from Guerrero Negro, Baja, Mexico. FEMS Microbiol Ecol 47: 305–318.

  50. , , , . (2004b). Determination of nitrogen-fixing phylotypes in Lyngbya sp and Microcoleus chthonoplastes cyanobacterial mats from Guerrero Negro, Baja California, Mexico. Appl Environ Microbiol 70: 2119–2128.

  51. . (1996). TreeView: an application to display phylogenetic trees on personal computers. Computer Appl Biosci 12: 357–358.

  52. , , , , , et al. (2003). Global trajectories of the long-term decline of coral reef ecosystems. Science 301: 955–958.

  53. , , . (1993). Growth, nitrogen fixation and spectral attenuation in cultivated Trichodesmium species. Appl Environ Microbiol 59: 1367–1375.

  54. , , , . (2006). Modeling the dynamic regulation of nitrogen fixation in the cyanobacterium Trichodesmium sp. Appl Environ Microbiol 72: 3217–3227.

  55. , , , . (2002). Diversity and distribution of coral-associated bacteria. Mar Ecol Progr Ser 243: 1–10.

  56. , , . (2003). Photosynthetic responses of the coral Montipora digitata to cold temperature stress. Mar Ecol Progr Ser 248: 85–97.

  57. , , . (2004). Spatial and temporal distribution of two diazotrophic bacteria in the Chesapeake Bay. Appl Environ Microbiol 70: 2186–2192.

  58. . (1994). Role of plankton in the turnover of organic matter on the Great Barrier Reef, Australia. Hydrobiologia 308: 35–44.

  59. , , , . (2004). Development and testing of a DNA macroarray to assess nitrogenase (nifH) gene diversity. Appl Environ Microbiol 70: 1455–1465.

  60. , , , , . (1997). The ClustalX windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucl Acid Res 25: 4876–4882.

  61. , . (2000). Xanthogenate nucleic acid isolation from cultured and environmental cyanobacteria. J Phycol 36: 251–258.

  62. , , , . (2006). The effect of water exchange on bacterioplankton depletion and inorganic nutrient dynamics in coral reef cavities. Coral Reefs 25: 23–36.

  63. . (1991). Nitrogen fixation by the cyanobacterial symbiont of the diatom genus Hemiaulus. Mar Ecol Progr Ser 76: 201–204.

  64. , , , , , . (2004). Coral-associated archaea. Mar Ecol Progr Ser 273: 89–96.

  65. , , , , , . (2004a). Coral mucus functions as an energy carrier and particle trap in the reef ecosystem. Nature 428: 66–70.

  66. , , , , , . (2004b). Degradation and mineralization of coral mucus in reef environments. Mar Ecol Progr Ser 267: 159–171.

  67. , , . (2005). Influence of coral mucus on nutrient fluxes in carbonate sands. Mar Ecol Progr Ser 287: 87–98.

  68. , , , . (2000). Expression of nifH genes in natural microbial assemblages in Lake George, New York, detected by reverse transcriptase PCR. Appl Environ Microbiol 66: 3119–3124.

  69. , , , , , et al. (2001). Unicellular cyanobacteria fix N-2 in the subtropical North Pacific Ocean. Nature 412: 635–638.

  70. , , , . (2003). Nitrogenase gene diversity and microbial community structure: a cross-system comparison. Environ Microbiol 5: 539–554.

  71. , . (1989). Use of degenerate oligonucleotides for amplification of the nifH gene from the marine cyanobacterium Trichodesmium thiebautii. Appl Environ Microbiol 55: 2522–2526.

  72. , , . (1998). New nitrogen-fixing microorganisms detected in oligotrophic oceans by the amplification of nitrogenase (nifH) genes. Appl Environ Microbiol 64: 3444–3450.

  73. , , , , , et al. (2007). Nitrogenase gene expression and N2 fixation in the North Pacific Subtropical Gyre. Limnol Oceanogr 52: 169–183.

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Acknowledgements

We acknowledge the assistance of C Bagnato, P Hallam and K Grieg from the Heron Island Research Station, University of Queensland staff for field assistance; and R Foster, R Paerl and K London for assistance with lab work. This study was supported by award OCE0425363 from the National Science Foundation, and an award from the Gordon and Betty Moore Foundation Marine Microbiology Initiative. Sampling for this study was conducted under permit G05/15688.1 from the Great Barrier Reef Marine Parks Authority (Commonwealth of Australia).

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  1. Department of Ocean Sciences, University of California Santa Cruz, Santa Cruz, CA, USA

    • Ian Hewson
    • , Pia H Moisander
    • , Amanda E Morrison
    •  & Jonathan P Zehr

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Correspondence to Ian Hewson.

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https://doi.org/10.1038/ismej.2007.5

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