Denitrification in lucerne nodules is not involved in nitrite detoxification

Dissimilatory reduction of ionic nitrogen oxides to gaseous forms such as nitrous oxide or nitrogen can be carried out by free living or symbiotic forms of some strains of Rhizobium meliloti. In this paper we investigate whether bacteroid denitrification plays a role in the alleviation of the inhibitory effects of nitrate on nitrogen fixation both in bacteroid incubations as in whole nodules. The presence of a constitutive nitrate reductase (NR) activity in isolated bacteroids caused nitrite accumulation in the incubation medium, and acetylene reduction activity in these bacteroids was progressively inhibited, since nitrite reductase (NiR) activity was unable to reduce all the nitrite produced by NR and denitrification occurred slowly. Even nodules infiltrated with nitrate and nitrite failed to increase gaseous forms of nitrogen substantially, indicating that nitrite availability was not limiting denitrification by bacteroids. In spite of the low rates of bacteroidal denitrification, the effect of nodule denitrification on the inhibition of nitrogen fixation by nitrate in whole plants was tested. For that purpose, lucerne plants (Medicago sativa L. cv. Aragon) were inoculated with two Rhizobium meliloti strains: 102-F-65 (non denitrifying) and 102-F-51 (a highly denitrifying strain). After a seven days nitrate treatment, both strains showed the same pattern of inhibition, and it occurred before any nitrate or nitrite accumulation within the nodules could be detected. This observation, together with the lack of alleviation of the ARA inhibition in the denitrifying strain, and the limited activity of dissimilatory nitrogen reduction present in these bacteroids, indicate a role other than nitrite detoxification for denitrification in nodules under natural conditions.