Uptake Hydrogenase Activity Determined by Plasmid pRL6JI in Rhizobium leguminosarum Does Not Increase Symbiotic Nitrogen Fixation

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Six mutants of Rhizobium leguminosarum 3855 lacking uptake hydrogenase activity (Hup− phenotype) as a result of Tn5-mob mutagenesis of the hup-containing plasmid pRL6JI were tested for symbiotic performance on Pisum sativum L. and Vicia benghalensis L. Three pea cultivars and one vetch line, which induce four different levels of Hup activity in strain 3855, were grown to flowering under microbiologically controlled conditions in the absence of combined N. Direct Kjeldahl N measurements showed that in every case at least one Hup− mutant fixed as much N2 as the isogenic Hup+ strain. Measures of C2H2 reduction, H2 evolution, 3H2 incorporation, and plant dry weight were consistent with the interpretation that the oxidation of H2 produced by the nitrogenase enzyme complex was not necessarily associated with increased N2 fixation in these symbiotic associations. Tests with a smaller subset of the Hup− strains under four different root environments ranging from pH 5.0 to 8.2 likewise showed no significant advantage for the isogenic Hup+ strain. It was concluded that the improvements in symbiotic N2 fixation produced by pRL6JI are associated with some trait other than the Hup+ phenotype.

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