Time-course of dinitrogen fixation of promiscuous soybean cultivars measured by the isotope dilution method

 Soybean cultivars capable of nodulating with indigenous Bradyrhizobium spp. have been developed by the International Institute of Tropical Agriculture (IITA) and national programs in Africa in order to avoid artificial inoculation by resource-poor farmers in Africa. The current selection procedure for enhanced N₂ fixation is based on an assessment of nodule formation which does not directly quantify the proportions of crop N derived from the atmosphere. We have monitored N accumulation patterns and N₂ fixation in nine promiscuous soybean cultivars with different maturity periods, using the ¹⁵N dilution technique. Nodule development generally peaked at the early podfill stage for all cultivars except Tgx 1519-1D and Tgx 1447-2D in which it continued to increase. The proportion of crop N derived from fixation (%NDFA) ranged between 51% and 67%, 77% and 84%, and 66% and 73% at full bloom, early podfill, and physiological maturity stages, respectively. Total N accumulation increased in all soybean genotypes with increasing plant age. Significant correlations (P<0.001) were established between nodule weight and %NDFA, even though this did not explain the relationship between nodule development and N₂ fixation in cultivars such as Tgx 1519-1D. Promiscuous soybean cultivars retained between 10% and 19% of total N accumulated at the final harvest, in belowground biomass. Our results indicated that these soybean cultivars can derive substantial proportions of plant N from N₂ fixation in soils where compatible indigenous bradyrhizobia populations are adequate and effective. Also, we have substantiated the claims that qualitative nodulation parameters currently used to select varieties with a high N₂ fixation capacity need to be validated with other measurements of N₂ fixation. Received: 5 November 1998     

Nodulation and nitrogen fixation of Lupinus species with Bradyrhizobium (lupin) strains in iron-deficient soil

 The effect of six Bradyrhizobium sp. (lupin) strains (WPBS 3201D, WPBS 3211D, USDA 3040, USDA 3041, USDA 3042 and CB 2272) and Fe supply on nodulation, N₂-fixation and growth of three lupin species (Lupinus termis, L. albus and L. triticale) grown under Fe deficiency in an alkaline soil, were examined in sterilized and non-sterilized pot experiments. When inoculated with USDA 3040, 3041, 3042 and CB2272 without Fe addition, the three lupin species had a very low nodule number and mass, low shoot and root dry matter accumulation and lower N yield. However, inoculation with WPBS 3201D and 3211D without Fe treatments increased all these parameters substantially. The ability of WPBS 3201D and 3211D to form nodules on the three lupin species under conditions of Fe stress could be attributed to their ability to scavenge Fe from Fe-deficient environments through their siderophore production. Addition of Fe to the other four strains significantly increased nodulation and N₂-fixation of the three lupin species, indicating that the poorer nodulation and N₂-fixation of these strains in the absence of Fe, resulted from a low ability to obtain Fe from alkaline soils. Bradyrhizobium strains WPBS 3201D and 3211D were superior to the other four strains in terms of promoting greater nodulation, N₂-fixation, plant growth and N accumulation of L. termis and L. albus. However, the other four strains were more efficient in symbiotic association with L. triticale. The greater variations in nodule efficiencies (specific nitrogenase activity) under different levels of Fe supply could be attributed to the quantities of bacteroid protein and leghaemoglobin in the nodules. The results suggested that Bradyrhizobium (lupin) strains differ greatly in their ability to obtain Fe from alkaline soils, and that the selection of bradyrhizobial strains which are tolerant of Fe deficient soils could complement plant breeding for the selection of legume crops for Fe-deficient soils. Received: 5 January 1998     

Genetic resources for improving nitrogen fixation in legume-rhizobia symbiosis

Leguminous crops are genetically polymorphous for the balance between symbiotrophic and combined types of nitrogen nutrition. In pea, polebean, alfalfa and fenugreek the wild-growing populations and local varieties exceed the agronomically advanced cultivars in the activity of N₂ fixation that occurs in symbiosis with nodule bacteria (rhizobia). Combined nitrogen nutrition ensures higher productivity than symbiotrophic one in the old leguminous crops (pea, alfalfa, common vetch, polebean, soybean), while the symbiotrophic type dominates in some young crops (hairy vetch, kura clover, goat's rue). An importance is emphasized of using the symbiotically active wild-growing genotypes as the initial material for breeding the legume cultivars. The data on high heritability (broad sense, narrow sense, realized) of the legume symbiotic activity demonstrate that the plant selection for this activity may be highly effective. A range of methods to select the legumes for an improved symbiotic activity is available including plant growth in N-depleted substrates, analysis of nodulation scores, direct (isotopic) and indirect (acetylene reduction) estimation of nitrogenase activity. Analysis of the specificity of interactions between different plant genotypes and bacterial strains (via two-factor analysis of variance) demonstrates the strain-specific plant polygenes are of a special importance in controlling the intensity of nitrogen fixation. Therefore, a coordinated plant-bacteria breeding is required to create the optimal combinations of partners' genotypes. Selection and genetic construction of the commercially attractive rhizobia strains should involve improvement of nitrogen fixing, nodulation and competitive abilities expressed in combination with the symbiotically active plant genotypes, Breeding of the leguminous crops for the preferential nodulation by highly active rhizobia strains, for the ability to support N₂ fixation under moderate N fertilization levels and to ensure a sufficient energy supply of symbiotrophic nitrogen nutrition is required     

Determination of competitive abilities of Bradyrhizobium japonicum strains in soils from soybean production regions in South Africa

Bradyrhizobium japonicum strain CB 1809 was recently chosen to replace strain WB 1 in commercial soybean [Glycine max (L.) Merr.] inoculants in South Africa, the selection criterion being N₂-fixing effectiveness. Nodulation competitiveness is an additional characteristic required of inoculants and was determined for CB 1809 and WB 1 as well as two other strains, USDA 110 and a Brazilian strain 965, using the gusA marker gene to identify strains. Initial experiments with plants grown in sterile sand showed that the competitive index of strain WB 1 was less than that of the other strains. Further comparisons used plants grown in five soils containing established populations of B. japonicum. When strains were applied in peat inoculum to seed at a rate of 1,000 cells per seed in a soil containing 300 rhizobia g^–1, significant differences in nodule occupancy were detected and strains ranked in the order 965>CB 1809>USDA 110>WB 1. The remaining four soils each contained about 10⁶ rhizobia g^–1 and 5×10⁶ cells were applied per seed. Nodule occupancy by inoculant strains ranged from 22% to 81% between soils. In this experiment, WB 1 was consistently the poorest performer and its competitiveness was significantly less than CB 1809. The competition results supported the recent decision to replace WB 1 with CB 1809 in commercial inoculants. Although WB 1 had been used in inoculants over a period of 19 years, this strain was detected in only one soil, where it comprised 8% of isolates. In contrast, a substantial proportion (32–78%) of isolates from the soils corresponded serologically to a former inoculant strain WB 66, which had been discontinued in 1966. This illustrates the difficulty of replacing a resident population with an introduced strain. The effect of naturalized populations on the establishment of CB 1809 in South African soils will need monitoring Received: 23 November 1999     

Effect of organic matter on the growth,nodulation, and nitrogen fixation of soybean grown under acid and saline conditions

Abstract

A field experiment was conducted to examine the effects of two organic materials (Bark and Tenporon composts) on the growth, nodulation and nitrogen fixation (measured as acetylene-reducing activity) of soybean plants (Glycine max L.) under acid and saline conditions. These organic materials were incorporated into acid and saline (induced by irrigating the soil with a NaCl solution) soils at 4% rate of application.

These composts tended to improve nodule formation in soybean under acid and saline conditions, especially in the plants treated with Bark compost. Acetylene-reducing activity was significantly (P<0.05) increased by these composts under acid condition.

These composts also tended to improve shoot growth under acid and saline conditions, presumably due to the improvement of the soil moisture status of the soils and nutrient uptake. These results suggest that the growth of soybean could be improved by the application of organic matter under acid and saline conditions.     

Soybean nodulation and nitrogen fixation on soil previously amended with sewage sludge

Summary A greenhouse study was conducted to examine the residual effects of sewage sludge on soybean Glycine max (L.) Merr., nodulation, and N fixation. Nodulating and nonnodulating isolines of Clark soybean were grown to the R2 stage in soils (Typic Paleudults) obtained from plots where heat-treated sludge had been applied in 1976 at rates equal to 0, 56,112, and 224 Mg ha^–1 high (7.0) and low (6.2) soil pH regimes were established by CaCO₃ additions. Sludge and soil pH treatments resulted in clearly defined differences in metal uptake by soybean shoots. Plant Zn, Cd, and Ni concentrations were greater on pH 6.2, sludge-amended soil than on the pH 7.0, amended soil. At low soil pH, soybean Zn and Cd concentrations, respectively, increased from 41 and 0.19 mg kg^–1 (control) to 120 and 0.58 mg kg^–1 at the 224 Mg hat sludge rate. At the high soil pH and 224 hg hat sludge rate, Zn and Cd concentrations were 45 and 0.15 mg kg^–1, respectively.Symbiotic N fixation provided 90% of the total N accumulation. Total N accumulation, shoot N concentration, dry matter, and N fixation by nodulating soybeans exhibited a significant linear increase with sludge rate. Total N accumulation, dry matter, and N fixation were significantly greater at high soil pH. For high and low soil pH, respectively, N fixation increased from 422 and 382 mg N per plant (control) to 614 and 518 mg N per plant at the 224 Mg ha^–1 sludge rate. While soybean nodulation also increased linearly on sludge-amended soil, a significant rate times pH interaction for nodule number indicated that nodulation was less strongly enhanced by sludge at low soil pH.     

Cobalt reduces nitrate inhibition of nodulation in mungbean (Vigna radiata)

 A cobalt-mediated decrease in ethylene production reduced the inhibition of nodulation by nitrate in Vigna radiata (mungbean). Nitrate increased the ethylene production in 5-day-old seedlings, while it caused a reduction in the nodulation status (nodule number and nodule weight) and nodule efficiency (acetylene reduction activity) in mungbean plants. The application of cobalt chloride inhibited nitrate-affected ethylene production and also decreased the inhibitory effect of nitrate on nodulation. The effect of cobalt was most marked on nodule number. Received: 6 August 1999     

Glyphosate effects on photosynthesis,nutrient accumulation,and nodulation in glyphosate‐resistant soybean

Previous greenhouse studies have demonstrated that photosynthesis in some cultivars of first‐ (GR1) and second‐generation (GR2) glyphosate‐resistant soybean was reduced by glyphosate. The reduction in photosynthesis that resulted from glyphosate might affect nutrient uptake and lead to lower plant biomass production and ultimately reduced grain yield. Therefore, a field study was conducted to determine if glyphosate‐induced damage to soybean (Glycine max L. Merr. cv. Asgrow AG3539) plants observed under controlled greenhouse conditions might occur in the field environment. The present study evaluated photosynthetic rate, nutrient accumulation, nodulation, and biomass production of GR2 soybean receiving different rates of glyphosate (0, 800, 1200, 2400 g a.e. ha^–1) applied at V2, V4, and V6 growth stages. In general, plant damage observed in the field study was similar to that in previous greenhouse studies. Increasing glyphosate rates and applications at later growth stages decreased nutrient accumulation, nodulation, leaf area, and shoot biomass production. Thus, to reduce potential undesirable effects of glyphosate on plant growth, application of the lowest glyphosate rate for weed‐control efficacy at early growth stages (V2 to V4) is suggested as an advantageous practice within current weed control in GR soybean for optimal crop productivity.     

Genetic variability in dinitrogen fixation between cowpea [Vigna unguiculata (L.) Walp] cultivars determined using the nitrogen-15 isotope dilution technique

 N fixed in 16 cultivars of cowpea [Vigna unguiculata (L.) Walp] inoculated with effective Bradyrhizobium strains collected from the West African MIRCEN culture collection was measured by ¹⁵N isotope dilution technique. In all plant parts, significant differences in the percentage of N derived from the atmosphere (%Ndfa) and the amount of Ndfa occurred between the cultivars. Ndoute variety exhibited the highest %Ndfa (74.33% in shoots; 60.90% in roots) and accumulated more fixed N (960 mg N plant^–1 and 38 mg N plant^–1 in shoots and roots, respectively). Therefore this cultivar should be selected as the highest N-fixing cowpea cultivar. It also should be used in a breeding programme to contribute to the development of cultivars that could stimulate an intensive use of cowpea in many different cropping systems in Africa with a view to maintaining soil fertility. Received: 14 June 1999     

The use of GUS-reporter gene to study the effect of Azospirillum-Rhizobium coinoculation on nodulation of white clover

 The gusA-marked Azospirillum lipoferum T1371, constructed by inserting transposon mTn5SSgusA20 from Escherichia coli S17-1 λ-pir into the genome of Azospirillum lipoferum 137, was used to evaluate its effect on nodulation of white clover with and without Rhizobium inoculation. When inoculated alone, Azospirillum colonized the tap roots, secondary roots and root hairs. The combined inoculation of white clover with Rhizobium leguminosarum bv trifolii and A. lipoferum enhanced the number of nodules by 2–3 times from 5 to 20 days after inoculation (DAI). The combined inoculation also enhanced acetylene reduction activity by 2.3–2.7 times at 20 DAI. Moreover, Azospirillum was observed colonizing the tap root, root hairs and sites near or on the nodules. These results suggest that the formation of additional infection sites by A. lipoferum, with a combined inoculation, may be the mechanism that will enhance nodulation and nitrogen fixation of white clover. Received: 23 July 1997     

Nodulation pattern and acetylene reduction (nitrogen fixation) activity of some highland and lowland Acacia species of Ethiopia

 Endosymbionts from the Ethiopian highland acacia species Acacia abyssinica, A. negrii and A. etbaica, and the lowland species A. nilotica, A. prasinata, A.senegal, A. seyal, A. tortilis and Faidherbia (Acacia) albida were isolated and characterized. Seven tree species were found to be nodulated by species of both Rhizobium and Bradyrhizobium. F. (Acacia) albida and A. senegal were nodulated by only Bradyrhizobium or Rhizobium, respectively. In A. abyssinica, both genera were isolated from the same nodule, whereas in A. nilotica and A. tortilis, both strains were isolated from different nodules of the same plant. The nitrogen fixation (acetylene reduction) activities varied considerably and showed no correlation with the nitrogen content of the plant. Highland species were as effective as lowland plants, thus demonstrating good potential for soil reclamation. The endosymbionts isolated proved rather promiscuous, efficiently nodulating other Acacia spp. and some tropical grain legumes, but did not nodulate temperate legumes. Received: 7 August 1997     

Soybean nodulation and nitrogen fixation on soil amended with plant residues

Residues from some tree species may contain allelopathic chemicals that have the potential to inhibit plant growth and symbiotic N₂-fixing microorganisms. Soybean [Glycine max (L.) Merr] was grown in pots to compare nodulation and N₂-fixation responses of the following soil amendments: control soil, leaf compost, red oak (Quercus rubra L.) leaves, sugar maple (Acer saccharum Marsh) leaves, sycamore (Platanus occidentalis L.) leaves, black walnut (Juglans nigra L.) leaves, rye (Secale cereale L.) straw, and corn (Zea mays L.) stover. Freshly fallen leaves were collected from urban shade trees. Soil was amended with 20 g kg^-1 air-dried, ground plant materials. Nodulating and nonnodulating isolines of Clark soybean were grown to the R2 stage to determine N₂-fixation by the difference method. Although nodulation was not adversely affected, soybean grown on leaf-amended soil exhibited temporary N deficiency until nodulation. Nodule number was increased by more than 40% for soybean grown on amended soil, but nodule dry matter per plant generally was not changed compared with control soil. Nonnodulating plants were severely N deficient and stunted as a consequence of N immobilization. Nodulating soybean plants grown on leaf or crop residue amended soil were more dependent on symbiotically fixed N and had lower dry matter yields than the controls. When leaves were composted, the problem of N immobilization was avoided and dry matter yield was not reduced. No indication of an allelopathic inhibition on nodulation or N₂-fixation from heavy application of oak, maple, sycamore, or walnut leaves to soil was observed.     

Mineral nitrogen impairs the biological nitrogen fixation in soybean of determinate and indeterminate growth types

Soybean (Glycine max Merrill) crop production in Brazil relies mainly on biological nitrogen fixation (BNF) for nitrogen (N) supply. Recent adoption of indeterminate growth-type genotypes has raised doubts on the need for supplemental mineral N that might negatively affect the BNF. We assessed the effects of mineral N on BNF attributes of soybean genotypes grown in central and southern Brazil. Genotypes were inoculated with Bradyrhizobium sp. and/or received mineral N in three sets of experiments. In the first set, two genotypes received increasing rates of mineral N in nutrient solution, which consistently reduced the BNF. In the second set, mineral N applied at sowing and/or topdressing reduced nodulation and ureides-N in determinate and indeterminate growth-type genotypes. In the third set, mineral N applied at R5.3 stage, foliar or as topdressing, did not increase grain yield in four field experiments. Mineral N impaired BNF irrespective of the growth type and had no effect on grain yield.     

Soil test levels of potassium,yields, and seed size in soybean cultivars

Abstract

The effect of soil test levels of K (1 N, NH₄OAc) on yields of soybean (Glycine max Merr) reproductive growth, seed size and numbers were evaluated for early, medium, and late maturing cultivars on a Maury silt loam (Typic Paleudalf). Relative yields were near maximum when extractable K was approximately 210 pp2m. Concentrations of K in uppermost, fully expanded leaves at flower initiation at the high production level were approximately 2.0 to 2.15%.

Neither dates of flower initiation nor pod maturation of the three cultivars, Bonus, Cutler 71, and Williams, were altered by K nutrition. Either seed size or seed number were approximately equally capable of serving as a mechanism for increasing yield in response to improved K nutrition. Among cultivars, Bonus tended to have more smaller seeds, whereas Cutler 71 tended to have larger seeds with fewer seeds being produced.     

Effects of CO2 concentration in rhizosphere on nodulation and N2 fixation of soybean and cowpea

Soybean (Glycine max L. Merr.) cvs. Akisengoku and Peking, and cowpea (Vigna unguiculata Walp.) cv. Kegonnotaki were inoculated with Bradyrhizobium japonicum AlO17, Shinorhizobium fredii USDAI93, and B. sp. Vigna MAFF03-03063, respectively and were cultured hydroponically with supply of CO₂-free air, 3dm³ m^-3 CO₂ air, or 25 dm³ m^-3 CO₂ air to study the effects of the CO₂ concentration in the rhizosphere on plant growth, nodulation, and nitrogen fixation. Increase of the CO₂ concentration in the rhizosphere led to the increase of the plant dry weight in the symbiosis between Peking and USDAI93, and that between Kegonnotaki and MAFF03-03063. On the other hand, dry matter accumulation in the symbiosis between Akisengoku and AI017 decreased under the supply of 25 dm³ m^-3 CO₂ air aimed at increasing the CO₂ concentration in the rhizosphere beyond the optimum CO₂ concentration for growth. Nodule mass and nodule number per plant were highest in Akisengoku, followed by Kegonnotaki and lowest in Peking. Also the increase of the CO₂ concentration in the rhizosphere led to the increase of the nodule mass and number in Kegonnotaki, while no changes were observed in Akisengoku and Peking. Biological nitrogen fixation (BNF) was highest in Akisengoku, followed by Kegonnotaki, and lowest or near zero in Peking. BNF in Akisengoku and Kegonnotaki showed a similar tendency to that of dry matter accumulation. BNF of Peking was especially low under the supply of CO₂-free air, and it increased with the increase of the CO₂ concentration in the rhizosphere. For the symbiosis of Bradyrhizobium strains with soybean and cowpea, the most suitable CO₂ concentration for N₂ fixation and plant growth was estimated to be about 10 dm³ m^-3, while for the symbiosis of S. fredii with soybean, the value was estimated to be above 30 dm³ m^-3.     

Effectiveness of foreign bradyrhizobia strains in enhancing nodulation, dry matter and seed yield of soybean (Glycine max L.) cultivars in Nigeria

 Field experiments were conducted to investigate the performance of three soybean cultivars with five foreign bradyrhizobia strains in different regions. The experiments at the two sites were designed with soybean (Glycine max L.) cultivars as the main factor and bradyrhizobia strains (USDA 136, TAL 122, USDA 6, TAL 377 and TAL 102) as the sub-factor. The experiments were arranged in randomised complete block design with four replications. Results show that nodule number, nodule dry weight and shoot dry weight, total N and seed yield were significantly increased when soybean cultivars were inoculated with foreign bradyrhizobia in two locations in the south east of Nigeria. At 63 days after planting the percentage increase in nodule number and dry weight after inoculation of soybean cultivars with bradyrhizobia strains ranged from 71 to 486% and from 0 to 200%, respectively. The percentage increase in shoot dry matter, %N and total N after bradyrhizobia inoculation ranged between 2–130%, 18–62% and 35–191%, respectively at Awka, and at the Igbariam site the percentage increase in shoot dry weight, %N and total N ranged between 3–76%, 0–43% and 19–125%, respectively. Seed yields after bradyrhizobia inoculation of soybean cultivar TGX 1485–1D at Igbariam ranged between 1.20 and 2.18 t ha^–1 against the uninoculated plants, which had seed yields of 1.05 t ha^–1. The poorest yield response after inoculation with bradyrhizobia strains was observed in soybean cultivar M-351, with a seed yield ranging from 0.60 to 0.98 t ha^–1. The fact that foreign bradyrhizobia strains were more effective than the indigenous strains for all the parameters studied suggests that there is a need to use bradyrhizobia inoculants for increased soybean production in Nigeria. The variations in the strain performance with the different soybean cultivars at the two sites, emphasises the need for careful Bradyrhizobium spp. strain selection. The fact that inoculation response was cultivar- and site-specific suggests that strategies for improving inoculation response in soybean cultivars should also consider the soil environment where the soybean is to be produced. Received: 25 May 1999     

Phosphorus uptake by mycorrhizal and nonmycorrhizal roots of soybean

Kinetics of phosphorus absorption were investigated with mycorrhizal and nonmycorrhizal soybean (Glycine max Merr.) plants which were grown in a greenhouse and growth chamber and with nutrient solutions. Mycorrhizal plants depleted phosphorus from a dilute (<30uM) nutrient solution three times faster and had lower apparent efflux of phosphorus than the uninfected plants. The affinity (K_m) of roots for phosphorus from the solution was not increased by mycorrhizal infection. The results indicate that enhanced phosphorus absorption by mycorrhizal roots of soybean is due to an increase in the number of uptake sites per unit area of root (^v max).     

玉米/大豆、玉米/花生间作对作物氮素吸收及结瘤固氮的影响

禾本科与豆科作物间作具有显著的增氮作用。为探明玉米/大豆、玉米/花生间作模式的氮素吸收、氮营养竞争能力及豆科结瘤特性的变化,解释玉米与豆科间作体系的增氮效应,通过田间试验,设置玉米单作（MM）、大豆单作（SS）、玉米/大豆间作（MS）、花生单作（PP）、玉米/花生间作（MP）等5种种植模式,研究不同种植模式对作物氮素积累、氮营养竞争强弱及豆科结瘤固氮特性的调控作用。结果表明,与单作相比,间作显著降低玉米和大豆的氮素积累量,对花生的氮素积累量影响不显著。5种模式系统氮素积累总量表现为MS > SS > MP,PP和MM处理最低且差异不显著,MS处理比MP处理显著高21.8%。与MM处理相比,MS和MP处理的玉米氮素积累量分别降低20.5%和11.7%,其中MP处理籽粒、叶片和茎秆氮素积累量比MS处理高8.9%、21.2%和14.3%。与SS处理相比,MS处理的大豆氮素积累量降低28.5%,其中,中行、边行分别降低10.1%、15.4%。玉米相对大豆氮营养竞争比率表现为强（CR_ms>1）,相对花生则表现为弱（CR_mp<1）。与SS处理相比,五叶期MS处理的大豆根瘤数量显著增加,根瘤鲜重无显著差异,盛花期后根瘤数量和鲜重均显著降低;MS处理的大豆根瘤固氮酶活性均降低,且中行降低幅度更大。与PP处理相比,开花期MP处理的花生根瘤数量和鲜重均显著增加,下针期后均显著降低;MP处理的花生根瘤固氮酶活性均降低,且边行降低幅度更大。各间作模式作物的氮素积累量虽然降低,但间作模式的系统氮素积累量却显著高于各单作模式,两种间作模式中MS处理的氮素积累总量最高。