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Symbiotic and Free-living Denitrification by Bradyrhizobium japonicum¹

ABSTRACT

Denitrification by Bradyrhizobium japonicum (formerly Rhizobium japonicum) was characterized in two strains, USDA-110 and USDA-23, to determine the extent of denitrifying enzyme expression during symbiotic growth with soybean [Glycine max (L.) Merr.]. The rhizobia grew in anaerobic batch cultures, depleting NO₃^- and producing N₂O in the presence of C₂H₂. Specific denitrifying activities of resting cells of rhizobia were compared with those of the common soil denitrifier Pseudomonas fluorescens, and were found to range from 14 to 18% of the P. fluorescens activity. Bacteroid specific activities were less than activities of free-living rhizobial cells, though the ratio depended on the growth stage of the cells. Denitrification was observed in detached whole nodules containing either of the strains, but not in nodules formed by a NO₃^- reductase mutant of B. japonicum 110. Denitrification rates of nodules were unaffected by the presence of external O₂, though the rates were limited by the external NO₃^- concentration. Denitrification by intact, nodulated soybean plants under aerobic conditions was observed. In the presence of NO₃^- denitrification and N₂ fixation (C₂H₂ reduction) occurred simultaneously. The estimated ratio of N denitrified to N fixed ranged from 0 to 0.3, depending on assay conditions. The observations suggest that denitrification in soybean nodules in the field can be a continuous process, independent of soil aeration. However, it is likely to be slow relative to N₂ fixation under most conditions.

¹ Published with the approval of the Director of the Kentucky Agricultural Experiment Station as Journal Article no. 85-3-188.

² Graduate Research Assistant and Associate Professor of Soil Microbiology, Univ. of Kentucky, Lexington, KY 40546.

Received for publication May 17, 1985.