Nitrate and nitrite reductase negative mutants of N2-fixingAzospirillum spp.

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Abstract
Chlorate resistant spontaneous mutants ofAzospirillum spp. (syn.Spirillum lipoferum) were selected in oxygen limited, deep agar tubes with chlorate. Among 20 mutants fromA. brasilense and 13 fromA. lipoferum all retained their functional nitrogenase and 11 from each species were nitrate reductase negative (nr−). Most of the mutants were also nitrite reductase negative (nir−), only 3 remaining nir+. Two mutants from nr+ nir+ parent strains lost only nir and became like the nr+ nir− parent strain ofA. brasilense. No parent strain or nr+ mutant showed any nitrogenase activity with 10 mM NO 3 − . In all nr− mutants, nitrogenase was unaffected by 10 mM NO 3 − . Nitrite inhibited nitrogenase activity of all parent strains and mutants including those which were nir−. It seems therefore, that inhibition of nitrogenase by nitrate is dependent on nitrate reduction. Under aerobic conditions, where nitrogenase activity is inhibited by oxygen, nitrate could be used as sole nitrogen source for growth of the parent strains and one mutant (nr− nir−) and nitritite of the parent strains and 10 mutants (all types). This indicates the loss of both assimilatory and dissimilatory nitrate reduction but only dissimilatory nitrite reduction in the mutants selected with chlorate.