Symbiotic nitrogen fixation in eight Acacia senegal provenances in dryland clays of the Blue Nile Sudan estimated by the 15N natural abundance method

The symbiotic biological N₂fixation by Acacia senegal was estimated using the ¹⁵N natural abundance (δ ¹⁵N) procedure on eight provenances collected from different environments and soil types grown in a clay soil in the Blue Nile region, Sudan. Balanites aegyptiaca (a non-legume) was used as a non-N₂-fixing reference plant to allow ¹⁵N-based estimates of the proportion of the Acacia N derived from atmospheric N₂ (N_dfa) to be calculated. Results show variation in leaf δ ¹⁵N between A. senegal and the reference plant and among years. The relative δ ¹⁵N values (‰) were higher in B. aegyptiaca than in the N₂-fixing acacia provenances. Provenances originally collected from clay soils fixed little N in the first year, but the amount fixed increased as the trees aged. All provenances showed a decrease in δ ¹⁵N with age. The N_dfa varied between 24% (Mazmoom provenance) and 61% (Rahad provenance) 4 years after planting. There was no significant difference in δ ¹⁵N between provenance groups based on soil type or rainfall at original growing site. The amount of N_dfa increased significantly with age in all provenances. The above-ground contribution of fixed N to foliage growth in a 4-year-old A. senegal was highest in the Rahad sand–soil provenance (46.7 kg N ha⁻¹) and lowest in the Mazmoom clay-soil provenance (28.7 kg N ha⁻¹). Our study represents the first use of the δ ¹⁵N method for estimating the N input by A. senegal to the clay plain soils of the gum belt in the Sudan.     

Alder and lupine enhance nitrogen cycling in a degraded forest soil in Northern Sweden

Positive effects of legumes and actinorhizal plants on N-poor soils have been observed in many studies but few have been done at high latitudes, which was the location of our study. We measured N₂ fixation and several indices of soil N at a site near the Arctic Circle in northern Sweden. More than 20 years ago lupine (Lupinus nootkatensis Donn) and gray alder (Alnus incana L. Moench) were planted on this degraded forest site. We measured total soil N, net N mineralization and nitrification with a buried bag technique, and fluxes of NH⁺ ₄ and NO^– ₃ as collected on ion exchange membranes. We also estimated N₂ fixation activity of the N₂-fixing plants by the natural abundance of ¹⁵N of leaves with Betula pendula Roth. as reference species. Foliar nitrogen in the N₂-fixing plants was almost totally derived from N₂ fixation. Plots containing N₂-fixing species generally had significantly higher soil N and N availability than a control plot without N₂-fixing plants. Taken together, all measurements indicated that N₂-fixing plants can be used to effectively improve soil fertility at high latitudes in northern Sweden.     

N2 fixation in Thai soybeans: Effect of tillage and inoculation on15N-determined N2 fixation in recommended cultivars and advanced breeding lines

The practice of seeding soybeans following paddy rice in Thailand has encountered difficulties in seedling germination, nodulation and crop establishment. This research project evaluated the choice of a non-fixing control to quantify N₂ fixation by¹⁵N isotope dilution, and the effect of tillage regime, soybean cultivar, strain ofBradyrhizobium japonicum and P fertilization on yield and N₂ fixation after paddy rice in northern and central Thailand.Japanese non-nodulating lines Tol-0 and A62-2 were the most appropriatecontrol plants for¹⁵N isotope dilution for Thai soybeans in these soils which contained indigenous rhizobia. Cereals such as maize, sorghum and barley were also appropriate controls at some sites. The choice of the appropriate non-fixing control plant for the¹⁵N isotope dilution technique remains a dilemma and no alternative exists other than to use several possible controls with each experiment. Acetylene reduction assay (ARA) proved of little value for screening varieties on their N₂ fixing capacity.The recommended Thai soybean cultivars (SJ1, 2, 4, 5) and an advanced line 16–4 differed little in their ability to support N₂ fixation or yield, possibly due to similar breeding ancestry. The ten AVRDC (ASET) lines showed considerable genotypic control in their ability to utilize their three available N sources (soil, fertilizer, atmosphere) and to translate them into yields. None of these lines were consistently superior to Thai cultivars SJ4 or SJ5 although ASET lines 129, 209 and 217 showed considerable promise.Neither recommended Thai or ASET cultivars were affected by tillage regime. Zero tillage resulted in superior N₂ fixation and yield at two sites but conventional tillage was superior at another site. Soybean cultivars grown in Thailand were well adapted to zero tillage. Levels of N₂ fixation were similar to world figures, averaging more than 100 kg N ha^–1 and supplying over 50% of the plant's N yield. However, seed yields seldom exceeded 2 t ha^–1, well below yields for temperately-grown soybeans. It is not clear why Thai soybeans support N₂ fixation, but do not translate this into higher seed yields.     

Influence of carbon availability on the production of NO, N2O, N2 and CO2 by soil cores during anaerobic incubation

Net productions of permanent soil atmosphere gases (N₂, CO₂, O₂) and temporary gases (N₂O, NO) were monitored in soil cores using a non-interfering, fully automated measuring technique allowing highly time resolved measurements over prolonged periods. The influence of changes in available organic carbon on CO₂, N₂O, NO and N₂ production was studied by changing the soil carbon content through aerobic preincubations of different length, up to 21 days.The aerobic preincubation caused an increase in NO₃ ^- concentration and a decrease in available carbon content. Available carbon content dominated both CO₂ and total N gas (N₂+N₂O+NO) production during anaerobiosis. Both CO₂ and total N gas production rates decreased with increasing length of the previous aerobic preincubation, this in spite of the higher initial NO₃ ^- concentration.Total denitrification rates were closely related to the anaerobic CO₂ production rates. No relation was found between water soluble carbon content and total denitrification. The N₂O/N₂ ratio could be explained by an interaction of carbon availability, NO₃ ^- concentration and enzyme status. Net N₂O consumption was monitored. The balance between cumulative total N gas production and NO₃ ^- consumption varied according to the different treatments. Cumulative N₂O production exceeded cumulative N₂ production for 0 up to 5 days.     

Symbiotic N2-fixation by the cover crop Pueraria phaseoloides as influenced by litter mineralization

The perennial legume Pueraria phaseoloides is widely used as a cover crop in rubber and oil palm plantations. However, very little knowledge exists on the effect of litter mineralization from P. phaseoloides on its symbiotic N₂-fixation. The contribution from symbiotic N₂-fixation (Ndfa) and litter N (Ndfl) to total plant N in P. phaseoloides was determined in a pot experiment using a ¹⁵N cross-labeling technique. For determination of N₂-fixation the non-fixing plant Axonopus compressus was used as a reference. The experiment was carried out in a growth chamber during 9 weeks with a sandy soil and 4 rates of ground litter (C/N=16,2.8% N). P. phaseoloides plants supplied with the highest amount of litter produced 26% more dry matter and fixed 23% more N than plants grown in soil with no litter application, but the percentage of Ndfa decreased slightly, but significantly, from 87 to 84%. The litter N uptake was directly proportional to the rate of application and constituted 10% of total plant N at the highest application rate. Additionally, a positive correlation was found between litter N uptake and the amount of fixed N₂. The total recovery of litter N in plants averaged 26% at harvest (shoot + root) and was not affected by the quantity added. A parallel incubation experiment also showed that, as an average of all litter levels, 26% of the litter N was present in the inorganic N pool. The amounts of fertilizer and soil N taken up by plants decreased with litter application, probably due to microbial immobilization and denitrification. It is concluded that, within the litter levels studied, litter mineralization will result in a higher amount of N₂-fixed by P. phaseoloides.     

Seasonal N uptake and N2fixation by common and adzuki bean at various spacings

The adzuki bean (Vigna angularis (Wild.) Ohwi and Ohashi) and common bean (Phaseolus vulgaris L.) have a high physiological demand for N. A 2-year field study was conducted to investigate the seasonal change of available soil N and symbiotic N₂ fixation usage. The beans were seeded at two densities, 22.2 plants m^–2 with a row spacing of 0.3 m and 11.1 plants m^–2 with a row spacing of 0.6 m. The amount of fixed N₂ in the shoot was calculated using the ¹⁵N natural abundance method. The common bean demonstrated low N₂ fixation and the ability to accumulate high levels of soil N. Soil nitrate under the common bean was continually absorbed. The adzuki bean, on the other hand, had a remarkable peak of N accumulation in the early reproductive stage. This was mainly due to N₂ fixation, though the soil nitrate level was high. Narrowing the plant row spacing increased the dry matter yield of both species, but the origin of the increased N differed between the species. For the first 77 DAP in 1999 (73 DAP in 2000) the N increase for both beans was due to both soil and atmospheric N₂. At harvest, though, the increase of N in common bean was mainly due to soil N, while that in adzuki bean was mainly due to atmospheric N₂. It can be concluded that the low symbiotic N₂ fixation ability of common bean was due to its high soil N uptake ability and constant N accumulation, which enabled an efficient soil N absorption. Adzuki bean absorbed N mainly for a short period and depended more on symbiotically fixed N₂ and, in contrast to common bean, left a high level of NO₃-N remaining in the soil after cropping.     

Crop residue application increases nitrogen fixation and dry matter production in groundnut (Arachis hypogaea L.) grown on an acid sandy soil in Niger,West Africa

Field experiments were conducted during the rainy reasons of 1989, 1990 and 1991 on an acid sandy soil in Niger, West Africa, to assess the effect of millet straw application (+CR) on growth and N₂ fixation of groundnut (Arachis hypogaea L.).Three years of +CR (4 t ha^–1 yr^–1) increased symbiotic N₂ fixation, total dry matter production (haulm plus pods) by 83% and total nitrogen (N) accumulation by 100%. Concentration of N in the shoot dry matter and total N in the soil were only slightly affected by the +CR treatment.Crop residue application increased the concentration of potassium (K) and molybdenum (Mo) and decreased the concentrations of aluminium (Al) and manganese (Mn) distinctly, both in the plant (shoot and nodule dry matter) and in the soil.The increase in dry matter production and N uptake was mainly due to improved N₂ fixation reflected by enhanced formation and growth of nodules as well as nitrogenase activity. This was attributed to improved chemical soil conditions, particularly to the higher availability of Mo and the lowered content of available Al and Mn.Although with the application of 4 t CR ha^–1, 60 kg K were supplied, increased growth could not be attributed to the additional supply of K.ICRISAT Journal Article No. 1229.ICRISAT Journal Article No. 1229.     

Influence of pruning management on P and N distribution and use efficiency by N2 fixing and non-N2 fixing trees used in alley cropping systems

Pruning of hedgerow trees is an important management practice for the successful establishment of an alley cropping system. Although pruning affects biomass production, only meager evidence of this management on distribution of nutrients among the different plant organs after tree regrowth is available. This study examined the effect of pruning on the distribution and use efficiency of N and P in a N₂ fixing leguminous tree species, Gliricidia sepium, and two non-N₂ fixing leguminous tree species, Senna siamea and S. spectabilis, grown in a field on an Alfisol (low in P) at Fashola (Guinea Savanna Zone), Southwestern Nigeria. Four P rates, 0, 20, 40 and 80 kg P ha^–1 as single superphosphate were used and management treatments included pruned versus unpruned plants. The ¹⁵N isotope dilution technique was used to measure N₂ fixation in G. sepium. Partitioning of total P among different plant organs was influenced by plant species and pruning management, but was not affected by P application rates. The distribution of total P in the various plant organs followed that of dry matter yield while N partitioning had a different pattern. Pruned plants distributed about 118% more total P to branches and had a higher physiological P use efficiency (PPUE) than unpruned plants. Leaves were the biggest sink for total N and N allocation in the other plant organs was influenced by plant species and pruning management, G. sepium had relatively more of its total N and P partitioned into roots (about double that of the non-N₂ fixing trees) but had a lower PPUE. Unpruned and pruned G. sepium derived 35 and 54% respectively of their total N from atmospheric N2, with about 54% of the fixed N2 being allocated to leaves and roots. Results showed that N and P pools turned over in the branches during plant regrowth after pruning but the causative factors associated with this phenomenon were not clear.     

Measurement of N2 fixation in maize (Zea mays L.)—ricebean (Vigna umbellata [Thunb.] Ohwi and Ohashi) intercrops

The yield of N in maize (Zea mays L.) and ricebean (Vigna umbellata [Thumb.] Ohwi and Ohashi) were compared on a Tropoqualf soil in North Thailand in 1984 and 1985. Both species were grown in field plots in monoculture or as intercrops at a constant planting density equivalent to 8 maize or 16 ricebean plants per m². The contribution of symbiotic N₂ fixation to ricebean growth was estimated from measurements of the natural abundance of¹⁵N (δ¹⁵N) in shoot nitrogen and from analysis of ureides in xylem sap vacuumextracted from detached stems. The natural abundance of¹⁵N in the intercropped ricebean was found to be considerably less than that in monoculture in both growing seasons. Using maize and a weed (Ageratum conyzoides L.) as non-fixing¹⁵N reference plants the proportions (P ¹⁵N) of ricebean shoot N derived from N₂ fixation ranged from 0.27 to 0.36 in monoculture ricebean up to 0.86 when grown in a 75% maize: 25% ricebean intercrop. When glasshouse-derived calibration curves were used to calculate plant proportional N₂ fixation (Pur) from the relative ureide contents of field collected xylem exudates, the contribution of N₂ fixation to ricebean N yields throughout the 1985 growing season were greater in intercrop than in monocrop even at the lowest maize:legume ratio (25∶75). Seasonal patterns of sap ureide abundance indicated that N₂ fixation was greatest at the time of ricebean podset. The averagePur andP ¹⁵N in ricebean during the first 90 days of growth showed identical rankings of monocrop and intercrop treatments in terms of N₂ fixation, although the two sets ofP values were different. Nonetheless, seasonal estimates of N₂ fixation during the entire 147 days of legume growth determined from ureide analyses indicated that equivalent amounts of N could be fixed by ricebean in a 75∶25 intercrop and in monoculture despite the former being planted at one-quarter the density.     

Influence of dissimilarities in temporal and spatial N-uptake patterns on15N-based estimates of fixation and transfer of N in ryegrass-clover mixtures

The temporal N-uptake patterns of white clover (Trifolium repens L.) mixed with perennial ryegrass (Lolium perenne L.) and of red clover (Trifolium pratense L.) mixed with Italian ryegrass (Lolium multiflorum Lam.) were determined in successive harvests of herbage within the growth cycles of a ley established near Zürich (Switzerland). Rooting patterns were examined by injecting¹⁵N-fertilizer at soil depths ranging from 10 to 40 cm. The results were analyzed to determine the effect of variations in time and depth of N-uptake on the¹⁵N-based measurement of N from symbiosis (N_sym) and N from transfer (N_trans).Grasses in mixture appeared to have deeper rooting systems than grass monocultures, which led to an overestimation of N transfer from white clover to perennial ryegrass if¹⁵N was spread on the soil surface.White clover generally lagged behind grass in soil N- uptake. Soil N-uptake of red clover slowed down before that of the grass because % N_sym almost reached 100% during the second half of each growth cycle. However, the effect of these dissimilarities on the seasonal average of %N_sym did not exceed 2%.It is concluded that at the observed high levels of N₂ fixation, failure to account for the N-uptake patterns of the test and reference crops only slightly affected the estimates of % N_sym and % N_trans, and did not invalidate the observed differences between species.     

Contrasting growth response of an N2-fixing and non-fixing forb to elevated CO2: dependence on soil N supply

With the ability to symbiotically fix atmospheric N₂, legumes may lack the N-limitations thought to constrain plant response to elevated concentrations of atmospheric CO₂. The growth and photosynthetic responses of two perennial grassland species were compared to test the hypotheses that (1) the CO₂ response of wild species is limited at low N availability, (2) legumes respond to a greater extent than non-fixing forbs to elevated CO₂, and (3) elevated CO₂ stimulates symbiotic N₂ fixation, resulting in an increased amount of N derived from the atmosphere. This study investigated the effects of atmospheric CO₂ concentration (365 and 700 mol mol^–1) and N addition on whole plant growth and C and N acquisition in an N₂-fixing legume (Lupinus perennis) and a non-fixing forb (Achillea millefolium) in controlled-chamber environments. To evaluate the effects of a wide range of N availability on the CO₂ response, we incorporated six levels of soil N addition starting with native field soil inherently low in N (field soil + 0, 4, 8, 12, 16, or 20 g N m^–2 yr^–1). Whole plant growth, leaf net photosynthetic rates (A), and the proportion of N derived from N₂ fixation were determined in plants grown from seed over one growing season. Both species increased growth with CO₂enrichment, but this response was mediated by N supply only for the non-fixer, Achillea. Its response depended on mineral N supply as growth enhancements under elevated CO₂ increased from 0% in low N soil to +25% at the higher levels of N addition. In contrast, Lupinus plants had 80% greater biomass under elevated CO₂ regardless of N treatment. Although partial photosynthetic acclimation to CO₂ enrichment occurred, both species maintained comparably higher A in elevated compared to ambient CO₂ (+38%). N addition facilitated increased A in Achillea, however, in neither species did additional N availability affect the acclimation response of A to CO₂. Elevated CO₂ increased plant total N yield by 57% in Lupinus but had no effect on Achillea. The increased N in Lupinus came from symbiotic N₂ fixation, which resulted in a 47% greater proportion of N derived from fixation relative to other sources of N. These results suggest that compared to non-fixing forbs, N₂-fixers exhibit positive photosynthetic and growth responses to increased atmospheric CO₂ that are independent of soil N supply. The enhanced amount of N derived from N₂ fixation under elevated CO₂ presumably helps meet the increased N demand in N₂-fixing species. This response may lead to modified roles of N₂-fixers and N₂-fixer/non-fixer species interactions in grassland communities, especially those that are inherently N-poor, under projected rising atmospheric CO₂.     

Nitrogen use efficiency of sugarcane in relation to its BNF potential and population of endophytic diazotrophs at different N levels

The nitrogen fixing bacterial endophytes Gluconacetobacter diazotrophicus and Herbaspirillum spp. have been proposed to benefit sugarcane (Saccaharum spp. hybrids) growth. Variable populations of these endophytes exist depending upon ontogenic and climatic variations as well. This study investigates the effect of variable chemical nitrogen application in soil on the population of endophytic diazotrophs, acetylene reduction ability of excised roots, plant N-nutrient use efficiency and probable interactions among different parameters in eight commercial sugarcane varieties of subtropical India. Recovery efficiency (RE), agronomic efficiency (AE), partial factor productivity (PFP) and physiologic efficiency (PE) indicators were used for accounting N-nutrient use efficiency. The population of G. diazotrophicus was more at N₇₅ compared to N₀ and N₁₅₀, whereas Herbaspirillum population increased from N₀ to N₁₅₀. ARA was positively correlated with Gluconacetobacter population in rhizosphere and root, whereas it had poor correlation with Herbaspirillum population. Positive correlation of RE and AE with ARA of roots, Gluconacetobacter and Herbaspirillum populations in roots and stems indicate their positive contribution in total nitrogen uptake by the plant per kg of N applied. Average PFP was 808.9 at N₇₅ compared to 408.7 at N₁₅₀ indicating that N was utilized efficiently at low N input status in sugarcane. Strong positive correlations of AE₇₅ (agronomic efficiency from 75 kg N ha⁻¹ to 150 kg N ha⁻¹) with N-uptake (r ² = 0.615), cane yield (r ² = 0.758) and PFP (r ² = 0.758) and other parameters compared to AE (agronomic efficiency from 0 kg N ha⁻¹ to 75 kg N ha⁻¹ or 150 kg N ha⁻¹) correlations with N-uptake (r ² = 0.111), cane yield (r ² = 0.368) and PFP (r ² = 0.190) indicated that the AE of sugarcane was strongly directed towards producing more cane yield per unit of N fertilizer once the sugarcane plant has established using initial dose of nitrogen and thus AE₇₅ seems to be a more appropriate indicator for accounting N-nutrient use efficiency in sugarcane.     

Application of 15N natural abundance technique for evaluating biological nitrogen fixation in oil palm ecotypes at nursery stage in pot experiments and at mature plantation sites

A range of different species of diazotrophic bacteria has been found in tissues and the rhizosphere of oil palm plants, suggesting a potential to benefit from biological nitrogen fixation (BNF). A few studies have confirmed that plantlets at nursery stage can benefit significantly from BNF after inoculation with Azospirillum spp. but no data are available regarding the benefit from naturally-occurring diazotrophic bacteria in oil palm. The results described here were derived from two pot trials laid out under controlled conditions with plantlets from two important regions for palm oil production in Brazil, as well as from different field sites of mature oil palm plantations. The ¹⁵N natural abundance technique was employed to estimate plant dependence on BNF (%Ndfa) by the different ecotypes grown in soil and previously characterized as hosting diazotrophic bacteria. From both pot trials it was possible to identify some ecotypes of high potential for N₂-fixation that reached in some cases approximately 50%Ndfa. However, the accuracy of measurement still needs to be improved using more suitable reference plants for pot experiments. Values of δ ¹⁵N signals from oil palm and reference plants in the field were inconclusive concerning any benefit from BNF to oil palm, owing to apparently high temporal and spatial variability of δ ¹⁵N of the plant-available N in the heterogeneous soil matrix for the different palm and reference plant tested.     

Seasonal accumulation and partitioning of nitrogen by lentil

Although wheat (Triticum aestivum L.) is the dominant crop of the semi-arid plains of Canada and the western United States, lentil (Lens culinaris Medik.) has become an important alternative crop. Sources and seasonal accumulation of N must be understood in order to identify parameters that can lead to increased N₂-fixing activity and yield. Inoculated lentil was grown in a sandy-loam soil at an irrigated site in Saskatchewan, Canada. Wheat was used as the reference crop to estimate N₂ fixation by the A-value approach. Lentil and wheat received 10 and 100 kg N ha⁻¹ of ammonium nitrate, respectively. Crops were harvested six times during the growing season and plant components analyzed. During the first 71 days after planting the wheat had a higher daily dry matter and N accumulation compared to lentil. However, during the latter part of the growing season, daily dry matter and N accumulation were greater for lentil. The maximum total N accumulation for lentil at maturity was 149 kg ha⁻¹. In contrast, wheat had a maximum N accumulation of 98 kg ha⁻¹ in the Feekes 11.1 stage, or 86 days after planting. The maximum daily rates of N accumulation were 3.82 kg N ha⁻¹ day⁻¹ for lentil and 2.21 kg N ha⁻¹ day⁻¹ for wheat. The percentage of N derived from N₂ fixation (% Ndfa) ranged from 0 at the first harvest to 92 % at final harvest. Generative plant components had higher values for % Ndfa than the vegetative components which indicates that N in the reproductive plant parts was derived largely from current N₂ fixation and lentil continued to fix N until the end of the pod fill stage. At final harvest, lentil had derived 129 kg N ha⁻¹ from N₂ fixation with maximum N₂-fixing activity (4.4 kg N ha⁻¹ day⁻¹) occurring during the early stages of pod fill. Higher maximum rates of N₂-fixing activity than net N accumulation (3.82 kg N ha⁻¹ day⁻¹) may have been caused by N losses like volatilization. In addition, lentil provided a net N contribution to the soil of 59 kg ha⁻¹ following the removal of the grain.     

Inoculation of forage grasses with N2-fixing Enterobacteriaceae

Summary Previous investigations indicated some forage grass roots in Texas are heavily colonized with N₂-fixing bacteria. The most numerous N₂-fixing bacteria were in the genera Klebsiella and Enterobacter. In the present investigation inoculation experiments were conducted using 18 isolates of these bacteria to determine if a N₂-fixing association could be established between the bacteria and the grassesCynodon dactylon andPanicum coloratum. Plants were grown in soil for approximately 5 months in a greenhouse and were measured periodically for dry matter, nitrogen accumulation, and acetylene reduction activity. Results of the investigation indicated that 25% of the plant-soil systems were active in acetylene reduction and the activity was high enough to indicate agronomically significant quantities of N₂ were being fixed (>8kg N ha⁻¹). However, plant systems extrapolated to fix>8 kg N ha⁻¹ contained less nitrogen and accumulated less dry matter than plants less active in acetylene reduction. Inocula could not be re-isolated from healthy grass roots indicating that the N₂-fixing activity may have not have been closely assiciated with plant roots. Future research is needed to determine factors limiting colonization of grass roots.     

Nitrogen fixation in perennial forage legumes in the field

Nitrogen acquisition is one of the most important factors for plant production, and N contribution from biological N₂ fixation can reduce the need for industrial N fertilizers. Perennial forages are widespread in temperate and boreal areas, where much of the agriculture is based on livestock production. Due to the symbiosis with N₂-fixing rhizobia, perennial forage legumes have great potential to increase sustainability in such grassland farming systems. The present work is a summary of a large number of studies investigating N₂ fixation in three perennial forage legumes primarily relating to ungrazed northern temperate/boreal areas. Reported rates of N₂ fixation in above-ground plant tissues were in the range of up to 373 kg N ha^–1 year^–1 in red clover (Trifolium pratense L.), 545 kg N ha^–1 year^–1 in white clover (T. repens L.) and 350 kg N ha^–1 year^–1 in alfalfa (Medicago sativa L.). When grown in mixtures with grasses, these species took a large fraction of their nitrogen from N₂ fixation (average around 80%), regardless of management, dry matter yield and location. There was a large variation in N₂ fixation data and part of this variation was ascribed to differences in plant production between years. Studies with experiments at more than one site showed that also geographic location was an important source of variation. On the other hand, when all data were plotted against latitude, there was no simple correlation. Climatic conditions seem therefore to give as high N₂ fixation per ha and year in northern areas (around 60°N) as in areas with a milder climate (around 40°N). Analyzing whole plants or just above-ground plant parts influenced the estimate of N₂ fixation, and most reported values were underestimated since roots were not included. Despite large differences in environmental conditions, such as N fertilization and geographic location, N₂ fixation (Nfix; kg N per ha and year) was significantly (P<0.001) correlated to legume dry matter yield (DM; kg per ha and year). Very rough, but nevertheless valuable estimations of Nfix in legume/grass mixtures (roots not considered) are given by Nfix = 0.026DM + 7 for T. pratense, Nfix = 0.031DM + 24 for T. repens, and Nfix = 0.021DM + 17 for M. sativa.     

Nitrogen and carbon partitioning in soybean under variable nitrogen supplies and acclimation to the prolonged action of elevated CO2

This study was conducted to determine reciprocal effects of low to high doses of nitrogenous fertilizer (N₃₀, N₄₀, N₅₀, N₆₀ and N₇₀ — 30, 40, 50, 60 and 70 kg ha⁻¹ respectively) and CO₂ enriched environment on C and N partitioning in soybean (Glycine max (L.) Merril cv JS-335). Plants were grown from seedling emergence to maturity inside open top chambers under ambient, AC (350±50 mol mol⁻¹) and elevated, EC (600±50 mol mol⁻¹) CO₂ and analyzed at seedling, vegetative, flowering, pod setting and maturity stages. Soybean responded to both CO₂ enrichment and N supply. Leaves, stem and root reserves at different growth stages were analyzed for total C and N contents. Consistent increase in the C contents of the leaf, stem and root was observed under EC than in AC. N contents in the different plant parts were found to be decreased under EC-grown plants specially at seedling and vegetative stage despite providing N doses to the soil. Significant increase observed for C to N dry mass ratio under EC in the root, stems and leaves at seedling and vegetative stage was decreased in the middle and later growth stages possibly due to combined impact of N doses to the soil and increased N₂ fixing activities due to EC conditions. Critical analysis of our findings reveals that the composition and partitioning of C and N of soybean under variable rates of N supply and CO₂ enrichment alter according to need under altered metabolic process. These changes eventually may lead to alteration in uptake of not only N but other essential nutrients also under changing atmosphere.     

Assessment of genetic variability for N2 fixation between and within provenances of Leucaena leucocephala and Acacia albida estimated by 15N labelling techniques

Nitrogen fixed in 13 provenances of Acacia albida and 11 isolines of Leucaena leucocephala inoculated with effective Rhizobium strains was measured by ¹⁵N techniques and the total N difference method. In the test soil, on the average, L. leucocephala derived about 65% of its total N from atmospheric N₂ fixation compared to about 20% by A. albida. Significant differences in the percentage of N derived from atmospheric N₂ (% Ndfa) occurred, between provenances or isolines within species. The % Ndfa ranged from 37 to 74% within L. leucocephala and from 6 to 37 within A. albida; (equivalent to 20–50 mg N plant^–1 and 4–37 mg N plant^–1 for the two species over three months, respectively) and was correlated with the nodule mass (r=0.91). The time course of N₂ fixation of three selected provenances (low, intermediate and good fixers) was followed at 12 weekly intervals over a 36 week period. The % Ndfa of all provenances and isolines increased with time; and except for one of the L. leucocephala provenances, % Ndfa was similar within species at the 36 weeks harvest. There was a significant correlation between % Ndfa and the amount of N₂ fixed (r=0.96). Significant interactions occurred between provenances and N treatments and often growth of uninoculated but N fertilized plants was less variable than for inoculated unfertilized plants.     

Application of 15N and xylem ureide methods for assessing N2 fixation of three shrub legumes periodically pruned for forage

The apparently diminished capacity for N₂ fixation by the shrub legume Calliandra calothyrsus (Calliandra) relative to other woody perennial legumes was investigated in a field experiment in northern Queensland, Australia. In this trial, (i) the proportion of plant nitrogen (N) derived from symbiotic N₂ fixation (%Pfix) and the amounts of N₂ fixed were compared in Calliandra, Gliricidia sepium (Gliricidia) and Codariocalyx gyroides (Codariocalyx), (ii) variations in N₂ fixation due to season or tree age were determined, (iii) estimates of Pfix derived with the ¹⁵N natural abundance technique were compared with values obtained from ¹⁵N enrichment or xylem sap ureide procedures to determine whether the previous conclusions about Calliandra's ability to fix N had resulted from specific problems with the natural abundance methodology used in the earlier studies.Inoculated seedlings of each of the three shrub legume species were planted in dense stands (1.5 m rows, 0.5 m between trees) in two randomised blocks. The northern block was used solely for natural abundance measurements, while ¹⁵N-enriched KNO₃ (10 atom % ¹⁵N excess) was applied four times over a 52 week period to plots in the southern block. The non-nodulating tree legume Senna spectabilis (formally Cassia spectabilis) was used as a non-N₂-fixing reference for the ¹⁵N-based procedures, with Guinea grass (Panicum maximum) included as an additional non-fixing check. Growth by the trees above 75 cm was first cut and removed after 22 weeks and regrowth was subsequently pruned periodically for another 95 weeks. Sampling for dry matter production, N yield and estimates of Pfix were restricted to the central four of the 32 plants which constituted each replicate plot. Information generated during the 117 week study indicated that estimates of Pfix by ¹⁵N natural abundance were closely similar to values derived with ¹⁵N-enrichment or sap ureides. The data indicated that Calliandra had a reduced reliance upon N₂ fixation relative to Gliricidia and Codariocalyx for the first 65 weeks after establishment. This appeared to be due to more prolifc root growth by Calliandra than either of the other N₂-fixing species and an ability to extract a greater proportion of its N requirements from soil mineral N. However, after week 65 and for the remainder of the experiment, estimates of Pfix for Calliandra were similar to the other shrub legumes. Over 117 weeks, prunings from Calliandra and Gliricidia had removed 52–58 t dry matter ha^-1, and between 1471 and 1678 kg N ha^-1, of which 1026–1063 kg N ha^-1 was estimated to have been derived from N₂ fixation. At the time of final harvest, 65–73% of the fixed N was present in shoot regrowth of the N₂ fixing shrubs, 9–18% in the roots, 15% in the trunk, and 2–6% in fallen leaves.     

Carbon isotope composition of N2-fixing and N-fertilized legumes along an elevational gradient

Dinitrogen-fixing legumes are frequently assumed to be less water-use efficient than plants utilizing soil mineral N, because of the high respiratory requirements for driving N₂ fixation. However, since respiration is assumed not to discriminate against ¹³C, any differences in water-use efficiency exclusively due to respiration should not be apparent in carbon isotope discrimination () values. Our objective was to determine if the source of N (N₂ fixation versus soil N) had any effect on of field-grown grain legumes grown at different elevations. Four legume species, Glycine max, Phaseolus lunatus, P. vulgaris, and Vigna unguiculata, were grown on five field sites spanning a 633 m elevational gradient on the island of Maui, Hawaii. The legumes were either inoculated with a mixture of three effective strains of rhizobia or fertilized weekly with urea at 100 kg N ha^-1 in an attempt to completely suppress symbiotic N₂-fixing activity. In 14 of 20 analyses of stover and 12 of 15 analyses of seed values were significantly higher (p=0.10) in the inoculated plants than the N-fertilized plants. Nitrogen concentrations were generally higher in the fertilized treatments than the inoculated treatments. The different values obtained depending on N-source may have implications in using as an indicator of water-use efficiency or yield potential of legumes.