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Applied and Environmental Microbiology, February 2003, p. 960-970, Vol. 69, No. 2
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.2.960-970.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Phylogenetic Diversity of Nitrogenase (nifH) Genes in Deep-Sea and Hydrothermal Vent Environments of the Juan de Fuca Ridge

Mausmi P. Mehta,1* David A. Butterfield,2 and John A. Baross1

School of Oceanography,1 Joint Institute for the Study of Atmosphere and Ocean, University of Washington, Seattle, Washington 981952

Received 27 August 2002/ Accepted 14 November 2002

The subseafloor microbial habitat associated with typical unsedimented mid-ocean-ridge hydrothermal vent ecosystems may be limited by the availability of fixed nitrogen, inferred by the low ammonium and nitrate concentrations measured in diffuse hydrothermal fluid. Dissolved N2 gas, the largest reservoir of nitrogen in the ocean, is abundant in deep-sea and hydrothermal vent fluid. In order to test the hypothesis that biological nitrogen fixation plays an important role in nitrogen cycling in the subseafloor associated with unsedimented hydrothermal vents, degenerate PCR primers were designed to amplify the nitrogenase iron protein gene nifH from hydrothermal vent fluid. A total of 120 nifH sequences were obtained from four samples: a nitrogen-poor diffuse vent named marker 33 on Axial Volcano, sampled twice over a period of 1 year as its temperature decreased; a nitrogen-rich diffuse vent near Puffer on Endeavour Segment; and deep seawater with no detectable hydrothermal plume signal. Subseafloor nifH genes from marker 33 and Puffer are related to anaerobic clostridia and sulfate reducers. Other nifH genes unique to the vent samples include proteobacteria and divergent Archaea. All of the nifH genes from the deep-seawater sample are most closely related to the thermophilic, anaerobic archaeon Methanococcus thermolithotrophicus (77 to 83% amino acid similarity). These results provide the first genetic evidence of potential nitrogen fixers in hydrothermal vent environments and indicate that at least two sources contribute to the diverse assemblage of nifH genes detected in hydrothermal vent fluid: nifH genes from an anaerobic, hot subseafloor and nifH genes from cold, oxygenated deep seawater.

* Corresponding author. Mailing address: University of Washington, School of Oceanography, Box 357940, Seattle, WA 98195. Phone: (206) 543-4911. Fax: (206) 543-0275. E-mail: mausmi{at}ocean.washington.edu.

Applied and Environmental Microbiology, February 2003, p. 960-970, Vol. 69, No. 2
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.2.960-970.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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