A phosphoenol pyruvate phosphatase transcript is induced in the root nodule cortex of Phaseolus vulgaris under conditions of phosphorus deficiency.

Although previous studies on N2-fixing legumes have demonstrated the contribution of acid phosphatases to their phosphorus (P) use efficiency under P-deficient growth conditions, localization of these enzymes in bean nodules has not been demonstrated. In this study, phosphoenol pyruvate phosphatase (PEPase) gene transcripts were localized within the nodule tissues of two recombinant inbred lines, RIL115 (P-deficiency tolerant) and RIL147 (P-deficiency sensitive), of Phaseolus vulgaris. Nodules were induced by Rhizobium tropici CIAT899 under hydroaeroponic conditions with a sufficient versus a deficient P supply. The results indicated that PEPase transcripts were particularly abundant in the nodule infected zone and cortex of both RILs. Analysis of fluorescence intensity indicated that nodule PEPase was induced under conditions of P deficiency to a significantly higher extent in RIL147 than in RIL115, and more in the inner cortex (91%) than in the outer cortex (71%) or the infected zone (79%). In addition, a significant increase (39%) in PEPase enzyme activity in the P-deficient RIL147 correlated with an increase (58%) in the efficiency of use in rhizobial symbiosis. It was concluded that nodule PEPase is upregulated under conditions of P deficiency in the P-deficiency-sensitive RIL147, and that this gene may contribute to adaptation of rhizobial symbiosis to low-P environments.

Phosphorous deficiency decreases nitrogenase activity but increases proton efflux in N2-fixing Medicago truncatula.

Effects of Sinorhizobium strain and P nutrition on N(2)-dependent growth, nitrogenase activity and proton efflux by nodulated roots were investigated in the model legume Medicago truncatula cultivar Jemalong grown in hydroaeroponic culture in symbioses with Sinorhizobium meliloti strains 102F51 and 2011. Sinorhizobium strain had strong effects on nitrogenase activity and N(2)-dependent growth, with S. meliloti 102F51 being the more efficient strain. Apparent and total nitrogenase activities, measured by hydrogen evolution in air and argon, respectively, were drastically reduced in plants supplied with 5 µmol P plant(-1) week(-1) as compared with 15 µmol P plant(-1) week(-1). There was a net proton efflux as soon as 2 weeks after inoculation and, in contrast to the effect of P nutrition on nitrogenase activity, P deficiency increased total and specific proton effluxes, irrespective of Sinorhizobium strain.

Growth and nitrogen acquisition strategies of Acacia senegal seedlings under exponential phosphorus additions.

There remains conflicting evidence on the relationship between P supply and biological N(2)-fixation rates, particularly N(2)-fixing plant adaptive strategies under P limitation. This is important, as edaphic conditions inherent to many economically and ecologically important semi-arid leguminous tree species, such as Acacia senegal, are P deficient. Our research objective was to verify N acquisition strategies under phosphorus limitations using isotopic techniques. Acacia senegal var. senegal was cultivated in sand culture with three levels of exponentially supplied phosphorus [low (200 µmol of P seedling(-1) over 12 weeks), mid (400 µmol) and high (600 µmol)] to achieve steady-state nutrition over the growth period. Uniform additions of N were also supplied. Plant growth and nutrition were evaluated. Seedlings exhibited significantly greater total biomass under high P supply compared to low P supply. Both P and N content significantly increased with increasing P supply. Similarly, N derived from solution increased with elevated P availability. However, both the number of nodules and the N derived from atmosphere, determined by the (15)N natural abundance method, did not increase along the P gradient. Phosphorus stimulated growth and increased mineral N uptake from solution without affecting the amount of N derived from the atmosphere. We conclude that, under non-limiting N conditions, A. senegal N acquisition strategies change with P supply, with less reliance on N(2)-fixation when the rhizosphere achieves a sufficient N uptake zone.

An interdisciplinary research strategy to improve symbiotic nitrogen fixation and yield of common bean (Phaseolus vulgaris) in salinised areas of the Mediterranean basin.

The main findings of a cooperative research group of agronomists, plant breeders, microbiologists, physiologists and molecularists to improve the symbiotic nitrogen fixation (SNF) and N2-dependent yield of common bean under moderate salinity in the Mediterranean basin are summarised. Agronomic surveys in reference production areas show large spatial and temporal variations in plant nodulation and growth, and in efficiency of utilisation of the rhizobial symbiosis. The latter was associated with a large rhizobial diversity, including new bean nodulating species. Macrosymbiont diversity in SNF and adaptation to NaCl was found. However, contrasts between plant genotypes could be altered by specific interactions with some native rhizobia. Therefore, variations in soil rhizobial population, in addition to agronomic practices and environmental constraints, may have contributed to erratic results observed in field inoculations. At the mechanistic level, nodule C and N metabolisms, and abcissic acid content, were related to SNF potential and tolerance to NaCl. Their relation with nodule conductance to O2 diffusion was addressed by in situ hybridisation of candidate carbonic anhydrase and aquaporin genes in nodule cortex. The limits and prospects of the cooperative strategy are discussed.

The phosphorus requirement of N2 -fixing and urea-fed Acacia mangium.

The fast-growing leguminous tree Acacia mangium Willd, was grown for at least 22 wk in aerated solution culture either under N2 -fixing conditions or with 2 mmol urea per plant per wk. Inorganic phosphorus was supplied at between 1 and 100 µmol P(1) per plant per wk: the latter was determined to be the optimum P supply for growth. The external P requirement for growth and the efficiency of utilization of internal P were similar for both N sources. However, shoot growth and the concentrations of N and P in leaves were decreased by P deficiency to a slightly higher extent in N2 -fixing plants than in urea-fed plants. Even though P deficiency limited nodule growth more drastically than it did shoot growth, the fraction of P allocated to both nodules and nodule nitrogenase activity (acetylene reduction) increased with P deficiency. It is concluded that this increase in nitrogenase activity reflects a higher N requirement per unit nodule mass in P-deficient plants and that, once nodules are fully functional the efficiency of utilization of internal P in N2 -fixing A. mangium is high compared with that of other N2 -fixing legumes.

A Short-Term Decrease in Nitrogenase Activity (C2H2 Reduction) Is Induced by Exposure of Soybean Shoots to Their CO2 Compensation Point.

Photosynthesis and nitrogenase acetylene-reducing activity (ARA) were measured in soybeans (Glycine max [L.] Merr.) in which the shoots were exposed for 48 h to 60 [mu]L L-1 CO2, a value corresponding to their CO2 compensation point. Six hours after the beginning of the light period at low CO2, the ARA started to decrease, reaching a rate of 50% of the control rate in 14 to 24 h and 20% of the control rate in 34 to 38 h after the beginning of the CO2 treatment. At these times, there was no net photosynthesis, and the transpiration rate was 20% lower than that in the control plants. An increase in the partial pressure of O2 around the nodules alleviated this inhibition of ARA. The maximal ARA achieved at 40 kPaO2 was 3 times higher than that at 20 kPa O2 and similar to the maximal ARA of the control plants. It was argued that the decrease in ARA of soybean exposed to the CO2 compensation point was due to a decrease in the nodule's permeability to O2 diffusion.

Effect of Oxygen and Malate on NO(3) Inhibition of Nitrogenase in Soybean Nodules.

Soybean (Glycine max cv Hodgson) nitrogenase activity (C(2)H(2) reduction) in the presence or absence of nitrate was studied at various external O(2) tensions. Nitrogenase activity increased with oxygen partial pressure up to 30 kilopascals, which appeared to be the optimum. A parallel increase in ATP/ADP ratios indicated a limitation of respiration rate by low O(2) tensions in the nodule, and the values found for adenine nucleotide ratios suggested that the nitrogenase activity was limited by the rate of ATP regeneration. In the presence of nitrate, the nitrogenase activity was low and less stimulated by increased pO(2), although the nitrite content per gram of nodules decreased from 0.05 to 0.02 micromole when pO(2) increased from 10 to 30 kilopascals. Therefore, the accumulation of nitrite inside the nodule was probably not the major cause of the inhibition. Instead, inhibition by nitrate could be due to competition for reducing power between nitrate reduction and bacteroid or mitochondrial respiration inside the nodule. This is supported by the observation of decrease in ATP/ADP ratios from 1.65, in absence of nitrate, to 0.93 in the presence of this anion at 30 kilopascals O(2). Furthermore, the inhibition was suppressed by the addition, to the plant nutrient solution, of 15 millimolar l-malate, a carbon substrate that is considered to be the major source of reductant for the bacteroids in the symbiosis.

Influence of the Bradyrhizobium japonicum Hydrogenase on the Growth of Glycine and Vigna Species.

The effect of the Bradyrhizobium japonicum hydrogenase on nitrogen fixation was evaluated by comparing the growth of Vigna and Glycine species inoculated with a Hup mutant and its Hup revertant. In all experiments, the growth of plants inoculated with the strain without hydrogenase was at least equal to the growth of the strain with hydrogenase. For Glycine usuriensis and Glycine max cv. Hodgson in liquid culture, the growth was higher with the Hup strain. It is possible that reduced rates of nitrogen fixation in the presence of hydrogenase are due to O(2) depletion caused by the hydrogen oxidizing, since the oxygen pressure in the air appears to be a limiting factor of symbiotic nitrogen fixation in the soybean.

Effects of periodic fluctuations of photon flux density on anatomical and photosynthetic characteristics of soybean leaves.

The development of soybean leaves grown at fluctuating photon flux density between 100 and 1500µM m(-2)s(-1) with a period of 160 sec were compared to leaves developed under continuous light with the same mean photon flux density. Number of epidermal cells and stomata, leaf area and specific leaf weight were not affected by the periodic fluctuation of photon flux density. Chloroplastic pigment concentration and chlorophyll fluorescence reveal some photoinhibitory effects of the high photon flux density phase. Stomatal and internal CO2 conductance and the quantum yield were not affected by the light regime. In contrast ribulose 1.5 bisphosphate carboxylase/oxygenase activity before in vitro activation by CO2 and Mg(++) was stimulated by the periodic illumination whereas the total amount of the enzyme and the internal leaf CO2 conductance remained steady. In conclusion, there was no major difference between leaves of plant grown either under a steady or under a periodic fluctuation of the photon flux density except some photoinhibitory symptoms under fluctuating illumination, and a higher in vivo level of activation of the Rubisco.

Respiratory and Nitrogenase Activities of Soybean Nodules Formed by Hydrogen Uptake Negative (Hup) Mutant and Revertant Strains of Rhizobium japonicum Characterized by Protein Patterns.

Rates of respiratory CO(2) loss and nitrogenase activities of H(2) uptake-negative mutant strains and H(2) uptake-positive revertant strains of Rhizobium japonicum have been investigated. Two-dimensional gel protein patterns of bacteroids formed by inoculation of soybeans (Glycine max L.) with these two strains show that they are closely related and revealed only one obvious difference between them. On the basis of molecular weight standards, it was concluded that the missing protein spot in the H(2) uptake-negative mutant strain could be caused by a failure of the mutant to synthesize hydrogenase. Nodules formed by the H(2) uptake-negative mutant strain evolved respiratory CO(2) at a rate of about 10% higher than that of nodules formed by the H(2) uptake-positive revertant strain. During short-term experiments employed, rates of both C(2)H(2) reduction and (15)N(2) fixation varied considerably among replicate samples and no statistically significant differences between mutant and revertant strains were observed. It was observed that increasing the partial pressure of O(2) over nodules significantly decreased the proportion of nitrogenase electrons allocated to H(+).