Rhizobium japonicum USDA191 is a member of a new group of Rhizobium japonicum strains found in China. This strain is one of several strains shown to be salt-tolerant and fast-growing; it is unique in being the only strain of this group that effectively nodulates American soybean cultivars. For these reasons strain USDA 191 was chosen for further study and comparison to the common American Rhizobium japonicum isolate USDA 1 10. Strain USDA 19 1 has a doubling time of 3.2 h in complex medium and grows in concentrations of up to 0.4 M-NaCl, while strain USDAl 10, which has a doubling time of 12 h, is severely inhibited in media containing 0.1 MNaCl. Under salt stress conditions, intracellular levels of K+ and glutamate were shown to increase. A comparison based on carbohydrate metabolism, DNA homology and protein patterns on polyacrylamide gels reveals that strain USDA191 is more closely related to the fastgrowing rhizobia than to Rhizobium japonicum. However, the strain retains capacity to nodulate American soybean and cowpea cultivars effectively.

Ammonium suppresses nitrogenase activity in Anabaena flos-aquae (Lyng) Breb. at all pH values tested. L-Methionine-DL-sulfoximine at 1 millimolar totally inhibited glutamine synthetase, and 10 micromolar partially inhibited. Both concentrations protected nitrogenase activity from ammonium-induced suppression at pH 7.1 and 8.1. At pH 93 and 10.2, methionine sulfoximine did not alleviate the suppression of nitrogenase by ammonium. This pH-dependent protection of nitrogenase activity is a result of the noncompetitive inhibition of the ammonium transporter by methionine sulfoximine. At pH 7.1 and 8.2, ammonium is protonated and methionine sulfoximine inhibits its entry into the cell. At pH 93 and 10.2, unprotonated ammonia is abundant and may enter the cell independent of the transport system. The effects of ammonium are closely mimicked by the ammonium analog methylamine. These results suggest that ammoniumperse is an important in vivo regulator ofnitrogen fixation and its function can be mimicked by methylamine. Previous studies employing methionine sulfoximine may have to be re-evaluated in light of the inhibitory effects of methionine sulfoximine on the ammonium transporter.In the absence ofcombined nitrogen, heterocystous blue-green algae fix atmospheric N2 and produce NH3 which is subsequently assimilated by the GS2/GOGAT pathway (2,8,20,21). Exogenous NH3 inhibits nitrogenase activity and heterocyst differentiation in nitrogen fixing algae (3). Stewart and Rowell (20) have shown that this effect can be alleviated by the presence ofthe GS inhibitor L-methionine-DL-sulfoximine (MSX). This result prompted Stewart and Rowell (20) to suggest that the inhibition of nitrogen fixation was not caused by ammonium directly but rather by GS, GOGAT, or a product ofammonium assimilation. Subsequent work (14) suggested that the size of the glutamine pool was an important factor in nitrogenase regulation. Recent Cultures were maintained at 30°C in 400-ml culture bottles bubbled with filter sterilized laboratory air which had been passed through an activated charcoal filter in series with two gas scrubbers containing 1 N H2SO4 and distilled H20, respectively. This treatment removed particulate matter and gaseous ammonia while at the same time saturating the air with water vapor. Continuous illumination was provided by two Gro-lux fluorescent tubes. The quantum flux density was attenuated to 40 gE m-2 s-' with neutral density screening. Experiments were carried out using 20-ml portions of algal cultures in mid-logarithmic phase of growth (1-2 ,ug Chl a ml-'). These portions were incubated in 40-ml air-bubbled test tubes. Culture pH was adjusted to pH 7.1, 8.1, and 10.2 with phosphate (20 mM), bicine (20 mM), and CHES (20 mM), respectively.Nitrogenase Activity. Nitrogenase activity was determined by acetylene reduction. Acetylene reduction was measured using 2-ml aliquots placed in 1 5-ml test tubes sealed with rubber septa. The tubes were preincubated for 10 min in a shaking water bath and illuminated with a quantum...

The effect of host plant cultivar on H2 evolution by root nodules was examined in symbioses between Pisum sativum L. and selected strains of Rhizobium leguminosarum. Hydrogen evolution from root nodules containing Rhizobium represents the sum of H2 produced by the nitrogenase enzyme complex and H2 oxidized by any uptake hydrogenase present in those bacterial cells. Relative efficiency (RE) calculated as RE -1 -(H2 evolved in air/C2H2 reduced) did not vary significantly among 'Feltham First,"Alaska,' and 'JI1205' peas inoculated with R. kguminosarum strain 300, which lacks uptake hydrogenase activity (Hup-). That observation suggests that the three host cultivars had no effect on H2 production by nitrogenase. However, RE of strain 128C53 was significantly (P c (25). That work was confirmed by Dixon (11), who localized uptake hydrogenase activity in pea root nodule bacteroids (12) and found it to be similar to Azotobacter hydrogenase in all respects examined (13 Many data suggest that at least 25% of the reductant used by nitrogenase is allocated to protons for H2 formation, while the remaining fraction of reductant is used to convert N2 to NH3 (6). The EAC4 is a convenient expression that reflects the partitioning of reductant among protons and alternative substrates such as N2 or C2H2 (6): EAC = (exogenous substrate reduced/H30 reduced + exogenous substrate reduced). In the presence of saturating levels of C2H2, all reductant is used to produce C2H4 and no H2 formation is detected. Schubert and Evans (28) defined the relative efficiency of N2 fixation as RE = 1 -(H2 evolved in air/C2H2 reduced) and showed that RE varied greatly among different Rhizobium-legume symbioses. Such RE values depend on the EAC of nitrogenase and on the capacity of the rhizobial cells to recover H2 by a separate uptake hydrogenase. Thus, in the absence ofuptake hydrogenase activity (Hup-phenotype), RE presumably is a measure of apparent EAC.Previous reports indicate that both plant and bacterial factors can affect H2 evolution from leguminous root nodules. Dixon (13) showed that R leguminosarum strain ONA 311 expressed strongly Hup+, slightly Hup+, or Hup-phenotypes on Pisum sativum, Vicia bengalensis, and V. faba, respectively. Similar reversals in uptake hydrogenase phenotype have been observed for two R. japonicum strains associated with Vigna unguiculata and Glycine max (20). Both strains were Hup+ on three cowpea cultivars and Hup-on three soybean cultivars. Host plant effects on apparent rhizobial EAC have been indicated by the observation that lengthening the normal dark period for P. sativum and Trifolium subterraneum increased RE of Hup-Rhizobium symbionts (15). Host plant age and long-term environmental factors such as availability of combined nitrogen, irradiance, and CO2 level also can affect apparent EAC (16). These latter findings are consistent with a previous study which concluded that both EAC and uptake hydrogenase activity of a Hup+ rhizobial strain were affected by irradiance treatment of the host pea pl...

Leaves of commercial soybeans [Glycine max (L.) Merr.] typically senesce and abscise during seed development, but certain genetic lines produce mature seeds and show a delayed leaf senescence (DLS) phenotype in which leaves remain green until killed by frost. Field studies with such DLS lines provide information on the physiological and agronomic traits associated with such plants. Acetylene reduction and carbon exchange rates of DLS plants declined greatly after seeds matured, but positive values were measured until frost. Leaves formed before flowering which remained on DLS plants at the R8 stage had significantly (P ≤ 0.01) higher concentrations of reduced N and starch than those measured in leaves of normal plants just prior to abscission. In addition, a new population of axillary leaves containing substantial N and starch often developed during pod filling in DLS lines. The highest seed yield measured in DLS materials was 2370 kg/ha, which was 75 and 86% of the yields recorded in plots of ‘Elf’ and ‘Clark’, respectively. The highest yielding DLS lines accumulated as much N as Elf under conditions where both entries obtained about 75% of their N from N2. Progeny row studies of the DLS trait in two different genetic backgrounds showed significant (P ≤ 0.001) negative correlations between seed yield and the DLS phenotype. Reciprocal shoot/root grafts between Clark and DLS plants established that the DLS trait was controlled by the shoot genotype. It is concluded that expression of the DLS trait does not require severe inhibitions of reproductive development, such as is associated with male sterility. However, in the materials used in this study, the DLS phenotype was associated with a decrease in seed yield.

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