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Appl Environ Microbiol. 1985 August; 50(2): 350-355
Copyright © 1985, American Society for Microbiology. All Rights Reserved.

Enumeration, Isolation, and Characterization of N₂-Fixing Bacteria from Seawater

Mary Lou Guerinot and Rita R. Colwell^*

Department of Microbiology, University of Maryland, College Park, Maryland 20742

ABSTRACT

Marine pelagic N₂-fixing bacteria have not, in general, been identified or quantified, since low or negligible rates of N₂ fixation have been recorded for seawater when blue-green algae (cyanobacteria) are absent. In the study reported here, marine N₂-fixing bacteria were found in all samples of seawater collected and were analyzed by using a most-probable-number (MPN) method. Two different media were used which allowed growth of microaerophiles, as well as that of aerobes and facultative anaerobes. MPN values obtained for N₂-fixing bacteria ranged from 0.4 to 1 x 10³ per liter for water collected off the coast of Puerto Rico and from 2 to 5.5 x 10² per liter for Chesapeake Bay water. Over 100 strains of N₂-fixing bacteria were isolated from the MPN tubes and classified, yielding four major groups of NaCl-requiring bacteria based on biochemical characteristics. Results of differential filtration studies indicate that N₂-fixing bacteria may be associated with phytoplankton. In addition, when N₂-fixing bacteria were inoculated into unfiltered seawater and incubated in situ, nitrogenase activity could be detected within 1 h. However, no nitrogenase activity was detected in uninoculated seawater or when bacteria were incubated in 0.2-µm-filtered (phytoplankton-free) seawater. The ability of these isolates to fix N₂ at ambient conditions in seawater and the large variety of N₂-fixing bacteria isolated and identified lead to the conclusion that N₂ fixation in the ocean may occur to a greater degree than previously believed.

^* Corresponding author.

Present address: MSU/DOE Plant Research Laboratory, Michigan State University, East Lansing, MI 48824.

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