Assessing the symbiotic dependency of grain and tree legumes on N2 fixation for their N nutrition in five agro-ecological zones of Botswana

To assess the symbiotic dependency of grain and shrub/tree legumes within five agro-ecological zones of Botswana, fully expanded leaves of the test species were sampled from about 26 study sites within Ngwaketse, Gaborone, Central, Ghanzi and Kalahari agro-ecological zones. Isotopic analysis revealed significant differences in δ¹⁵N values of the grain legumes [cowpea (Vigna unguiculata L. Walp), Bambara groundnut (Vigna subterranea L. Verde.), and groundnut (Arachis hypogaea L.)] from the 26 farming areas in both 2005 and 2006. Estimates of %Ndfa of leaves also showed significant differences between farming areas, with cowpea deriving more than 50% of its N nutrition from symbiotic fixation. In terms of distribution, many more symbiotic shrub/tree species were found in the wetter Ngwaketse agro-zone compared to the fewer numbers in the drier Kalahari region. Acacias were the more dominant species at all sites. Leaf δ¹⁵N values of shrub/tree species also varied strongly across Botswana, with 11 out of 18 of these legumes deriving about 50%, or more, of their N from symbiotic N₂ fixation.Acacia caffra, in particular, obtained as much as 93.6% of its N nutrition from symbiotic fixation in the wetter Ngwaketse agro-zone. This study has shown that grain legumes sampled from farmer’s fields in Botswana obtained considerable amounts of their N from symbiotic fixation. We have also shown that shrub and tree legumes probably play an important role in the N economy of the savanna ecosystems in Botswana. However, the decline in the number of functional N₂-fixing shrub/tree legumes along an aridity gradient suggests that soil moisture is a major constraint to N₂ fixation in the tree legumes of Botswana.     

Nitrogen fixation by groundnut and soyabean and residual nitrogen benefits to rice in farmers' fields in Northeast Thailand

Nitrogen fixation in groundnut and soyabean and the residual benefits of incoporated legume stover to subsequent rice crops were estimated in farmers' fields using¹⁵N-isotope methods. Three field experiments were conducted, two which examined N₂-fixation in groundnut by¹⁵N-isotope dilution using a non-nodulating groundnut as a reference crop and one in which N₂-fixation in two soyabean genotypes was compared using maize as the non-fixing reference crop. Groundnut fixed 72–77% of its N amounting to 150–200 kg N ha^-1 in 106–119 days and soyabean derived 66–68% of its N from N₂-fixation which amounted to 108–152 kg N ha^-1 under similar conditions. When legume stover was returned to the soil, there was a net contribution of N from N₂-fixing varieties of groundnut in all cases ranging from 13–100 kg N ha^-1, whilst due to the high % N harvest index in soyabean (87–88%) there was a net removal of N of 37–46 kg N ha^-1. In all cases if the legume stover was removed there was a net removal of N in the legume crop which ranged between 54 and 74 kg N ha^-1 in N₂-fixing varieties of groundnut and from 58 to 73 kg N ha^-1 in soyabean, whilst maize removed 66 kg N ha^-1 if its stover was returned and 101 kg N ha^-1 when the stover was removed. Growth of rice was improved in all cases where groundnut stover was returned resulting in increases in grain yield of 12–26% and increases in total dry matter production of 26–31%. Soyabean residues gave no increases in rice grain yield but increased total dry matter production by 12–20%. Rice accumulated more N in all cases where legume stover was returned to the soil, and N yields were larger in all cases after the N₂-fixing legumes than after the non-fixing reference crops. N difference estimates of the total residual N benefits from the N₂-fixing legumes ranged from 11–19 kg N ha^-1 after groundnut and 15–16 kg N ha^-1 after soyabean. The amounts of N estimated directly by application of¹⁵N-labelled stover amounted to 7.2–20.5 kg N ha^-1 with groundnut which represented recovery of 8–22% of the N added in the stover. In soyabean only 3.0–5.8 kg N ha^-1 was estimated to be recovered by¹⁵N-labelling which was 15–23% of the added N, whilst only 1.3 kg N ha^-1 (4% of the N added) was recovered by rice from the maize stover. An indirect¹⁵N-method based on addition of unlabelled stover to microplots where the soil had previously been labelled with¹⁵N gave extremely variable and often negative estimates of residual N benefits. Estimates of residual N from the added stover made by N difference calculations did not correspond with the estimates by direct¹⁵N-labelling in all cases and possible reasons for this are discussed.     

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.     

Niche-based assessment of contributions of legumes to the nitrogen economy of Western Kenya smallholder farms

Nitrogen (N) deficiency is a major constraint to the productivity of the African smallholder farming systems. Grain, green manure and forage legumes have the potential to improve the soil N fertility of smallholder farming systems through biological N₂-fixation. The N₂-fixation of bean (Phaseolus vulgaris), soyabean (Glycine max), groundnut (Arachis hypogaea), Lima bean (Phaseolus lunatus), lablab (Lablab purpureus), velvet bean (Mucuna pruriens), crotalaria (Crotalaria ochroleuca), jackbean (Canavalia ensiformis), desmodium (Desmodium uncinatum), stylo (Stylosanthes guianensis) and siratro (Macroptilium atropurpureum) was assessed using the ¹⁵N natural abundance method. The experiments were conducted at three sites in western Kenya, selected on an agro-ecological zone (AEZ) gradient defined by rainfall. On a relative scale, Museno represents high potential AEZ 1, Majengo medium potential AEZ 2 and Ndori low potential AEZ 3. Rainfall in the year of experimentation was highest in AEZ 2, followed by AEZ 1 and AEZ 3. Experimental fields were classified into high, medium and low fertility classes, to assess the influence of soil fertility on N₂-fixation performance. The legumes were planted with triple super phosphate (TSP) at 30 kg P ha^?1, with an extra soyabean plot planted without TSP (soyabean-P), to assess response to P, and no artificial inoculation was done. Legume grain yield, shoot N accumulation, %N derived from N₂-fixation, N₂-fixation and net N inputs differed significantly (P<0.01) with rainfall and soil fertility. Mean grain yield ranged from 0.86 Mg ha^?1, in AEZ 2, to 0.30 Mg ha^?1, in AEZ 3, and from 0.78 Mg ha^?1, in the high fertility field, to 0.48 Mg ha^?1, in the low fertility field. Shoot N accumulation ranged from a maximum of 486 kg N ha^?1 in AEZ 2, to a minimum of 10 kg N ha^?1 in AEZ 3. Based on shoot biomass estimates, the species fixed 25–90% of their N requirements in AEZ 2, 23–90% in AEZ 1, and 7–77% in AEZ 3. Mean N₂-fixation by green manure legumes ranged from 319 kg ha^?1 (velvet bean) in AEZ 2 to 29 kg ha^?1 (jackbean) in AEZ 3. For the forage legumes, mean N₂-fixation ranged from 97 kg N ha^?1 for desmodium in AEZ 2 to 39 kg N ha^?1 for siratro in AEZ 3, while for the grain legumes, the range was from 172 kg N ha^?1 for lablab in AEZ 1 to 3 kg N ha^?1 for soyabean-P in AEZ 3. Lablab and groundnut showed consistently greater N₂-fixation and net N inputs across agro-ecological and soil fertility gradients. The use of maize as reference crop resulted in lower N₂-fixation values than when broad-leaved weed plants were used. The results demonstrate differential contributions of the green manure, forage and grain legume species to soil fertility improvement in different biophysical niches in smallholder farming systems and suggest that appropriate selection is needed to match species with the niches and farmers’ needs.     

Monovalent cation transporters; establishing a link between bioinformatics and physiology

Toxic aluminum (Al) ion is a major constraint to plant growth in acid soils. Aluminum tolerance in wheat (Triticum aestivum L.) is strongly related to the Al-triggered efflux of malate from root apices. A role of the secreted malate has been postulated to be in chelating Al and thus excluding it from root apices (malate hypothesis), but the actual process has yet to be fully elucidated. We measured Al content and root growth during and after Al exposure using seedlings of near-isogenic lines [ET8 (Al tolerant) and ES8 (Al sensitive)] differing in the capacity to induce Al-triggered malate efflux. Aluminum doses that caused 50% root growth inhibition during 24-h exposure to Al in calcium (Ca) solution (0.5 mM CaCl₂, pH 4.5) were 50 μM in ET8 and 5 μM in ES8. Under such conditions, the amount of Al accumulated in root apices was approximately 2-fold higher in ET8 than ES8. Al-treated seedlings were then transferred to the Al-free Ca solution for 24 h. Compared to control roots (no Al pretreatment), root regrowth of Al-treated roots was about 100% in ET8 and about 25% in ES8. The impaired regrowth in ES8 was observed even after 24-h exposure to 2.5 μM Al which had caused only 20% root growth inhibition. The addition of malate (100 μM) during exposure to 50 μM Al in ES8 enhanced root growth 1.6 times and regrowth in Al-free solution 7 times, resulting in similar root growth and regrowth as in ET8. Short-term Al treatments of ES8 for up to 5 h indicated that the Al-caused inhibition of root regrowth started after 1-h exposure to Al. The stimulating effect of malate on root regrowth was observed when malate was present during Al exposure, but not when roots previously exposed to Al were rinsed with malate, although Al accumulation in root apices was similar under these malate treatments. We conclude that the malate secreted from root apices under Al exposure is essential for the apices to commence regrowth in Al-free medium, the trait that is not related to the exclusion of Al from the apices.     

Combating food insecurity on sandy soils in Zimbabwe: The legume challenge

The continued rise in mineral fertilizer costs has demanded cheaper alternative N sources for resource-constrained smallholder farmers, with N₂-fixing legumes presenting a viable option to maintain crop productivity. A study was conducted over two years on a coarse sandy soil (Lixisol with <80 g clay kg^?1 soil) to determine the productivity of (i) five grain legumes, (ii) a green manure legume, and (iii) maize on smallholder farmers’ fields, identified as SOFECSA Leaming Centres, in Chinyika, north-east Zimbabwe. The objective of the study was to promote appropriate targeting of soil fertility technologies to different farmer resource groups. Emphasis was put on establishing the scope for improving nutrient resource allocation efficiency and crop yields in relation to different management practices as dictated by resource endowment. Both biomass and grain yield results indicated a general conformity to farmer resource group as follows: Resource-endowed farmers (RG1) > Intermediate farmers (RG2) > Resource-constrained farmers (RG3). Although overall biomass productivity for the grain legumes was generally low, <2.8 Mg ha^?1 across all Learning Centres, soyabean grain yields increased by between 30% (RG1) and >500% (RG3) over the two seasons. However, there was a general preference for bambara nut by RG3 farmers who cited low cash demands in terms of seed and external inputs, and pest-resistance compared with other grain legumes. Increased maize grain yields following legumes, and which exceeded 7 Mg ha^?1 for RG1 under green-manure, was apparently due to an increase in soil available N. The results showed scope for enhancing the contribution of legumes to both soil fertility and household nutrition within smallholder farming systems if targeted according to farmers’ resource endowment. The challenge is availing the minimum level of external inputs to RG3 farmers to achieve significant yield benefits on poor soils. The paper presents three main scenarios constituting major challenges for integrating legumes into the current farming systems.     

The15N methodology in estimating N2 fixation by vetch and pea grown in pure stand or in mixtures with oat

Cereal-legume mixtures are frequently the best management decision for forage production instead of growing crops in pure stands. Nitrogen fertilization of cereal-legume mixtures is questionable since combined nitrogen could depress N₂ fixation by legumes. The objectives of this study were (1) to examine the effect of N fertilization on N₂ fixation by vetch and field peas in pure and in mixed stands with oats, and (2) to examine if there is any transfer of N from legumes to associated cereals. The field experiment was conducted for two growing seasons. The treatments were pure stands of vetch, pea and oats, and the mixtures of the two legumes with oats at the seeding ratios 90:10 and 75:25, fertilized with labelled¹⁵N at the rates of 15 and 90 kg N ha⁻¹. Nitrogen fertilization of 90 kg N ha⁻¹ suppressed N₂ fixation in both legumes grown in pure and in mixed stands. Crops grown in mixtures in many instances had lower atom %¹⁵N excess. Whether this was due to high N₂ fixation in the case of legume and transfer in the case of oat or the differences were due to practical problems of the¹⁵N technique is not clearly shown by the results, so based on the literature the aspect is discussed as well as the precautions which should be considered in using the¹⁵N technique in such studies.     

Australian mulga ecosystems –13C and 15N natural abundances of biota components and their ecophysiological significance

Samples of recently produced shoot material collected in winter/spring from common plant species of mulga vegetation in eastern and Western Australia were assayed for ¹³C and ¹⁵N natural abundance. ¹³C analyses showed only three of the 88 test species to exhibit C₄ metabolism and only one of seven succulent species to be in CAM mode. Non-succulent winter ephemeral C₃ species showed significantly lower mean δ¹³C values (– 28·0‰) than corresponding C₃-type herbaceous perennials, woody shrubs or trees (– 26·9, – 25·7 and – 26·2‰, respectively), suggesting lower water stress and poorer water use efficiency in carbon acquisition by the former than latter groups of taxa. Corresponding values for δ¹⁵N of the above growth and life forms lay within the range 7·5–15·5‰. δ¹⁵N of soil NH₄⁺ (mean 19·6‰) at a soft mulga site in Western Australia was considerably higher than that of NO₃^– (4·3‰). Shoot dry matter of Acacia spp. exhibited mean δ¹⁵N values (9·10 ± 0·6‰) identical to those of 37 companion non-N₂-fixing woody shrubs and trees (9·06 ± 0·5‰). These data, with no evidence of nodulation, suggested little or no input of fixed N₂ by the legumes in question. However, two acacias and two papilionoid legumes from a dune of wind-blown, heavily leached sand bordering a lake in mulga in Western Australia recorded δ¹⁵N values in the range 2·0–3·0‰ versus 6·4–10·7‰ for associated non-N₂-fixing taxa. These differences in δ¹⁵N, and prolific nodulation of the legumes, indicated symbiotic inputs of fixed N in this unusual situation. δ¹⁵N signals of lichens, termites, ants and grasshoppers from mulga of Western Australia provided evidence of N₂ fixation in certain termite colonies and by a cyanobacteria-containing species of lichen. Data are discussed in relation to earlier evidence of nitrophily and water availability constraints on nitrate utilization by mulga vegetation.     

N<Subscript>2</Subscript> fixation in three perennial <Emphasis Type="Italic">Trifolium</Emphasis> species in experimental grasslands of varied plant species richness and composition

This study is the first to investigate quantitative effects of plant community composition and diversity on N₂ fixation in legumes. N₂ fixation in three perennial Trifolium species grown in field plots with varied number of neighbouring species was evaluated with the ¹⁵N natural abundance method (two field sites, several growing seasons, no N addition) and the isotope dilution method (one site, one growing season, 5 g N m⁻²). The proportion of plant N derived from N₂ fixation, pNdfa, was generally high, but the N addition decreased pNdfa, especially in species-poor communities. Also following N addition, the presence of grasses in species-rich communities increased pNdfa in T. hybridum and T. repens L., while legume abundance had the opposite effect. In T. repens, competition for light from grasses appeared to limit growth and thereby the amount of N₂ fixed at the plant level, expressed as mg N₂ fixed per sown seed. We conclude that the occurrence of diversity effects seems to be largely context dependent, with soil N availability being a major determinant, and that species composition and functional traits are more important than species richness regarding how neighbouring plant species influence N₂ fixation in legumes.     

Does nitrogen transfer between plants confound 15N-based quantifications of N2 fixation?

Background and aims

Transfer of fixed N from legumes to non-legume reference plants may alter the ¹⁵N signature of the reference plant as compared to the soil N available to the legume. This study investigates how N transfer influences the result of ¹⁵N-based N₂ fixation measurements.

Methods

We labelled either legumes or non-legumes with ¹⁵N and performed detailed analyses of ¹⁵N enrichment in mixed plant communities in the field. The results were used in a conceptual model comparing how different N transfer scenarios influenced the ¹⁵N signatures of legumes and reference plants, and how the resulting N₂ fixation estimate was influenced by using reference plants in pure stand or in mixture with the legume.

Results

Based on isotopic signatures, N transfer was detected in all directions: from legume to legume, from legume to non-legume, from non-legume to legume, from non-legume to non-legume. In the scenario of multidirectional N transfer, N₂ fixation was overestimated by using a reference plant in pure stand.

Conclusions

Fixed N transferred to neighbouring reference plants modifies the ¹⁵N signature of the soil N available both to the reference plant and the N₂-fixing legume. This provides strong support for using reference plants growing in mixture with the legumes for reliable quantifications of N₂ fixation.     

Optimising biological N2 fixation by legumes in farming systems

Whether grown as pulses for grain, as green manure, as pastures or as the tree components of agro-forestry systems, the value of leguminous crops lies in their ability to fix atmospheric N₂, so reducing the use of expensive fertiliser-N and enhancing soil fertility. N₂ fixing legumes provide the basis for developing sustainable farming systems that incorporate integrated nutrient management. By exploiting the stable nitrogen isotope ¹⁵N, it has been possible to reliably measure rates of N₂ fixation in a wide range of agro-ecological field situations involving many leguminous species. The accumulated data demonstrate that there is a wealth of genetic diversity among legumes and their Rhizobium symbionts which can be used to enhance N₂ fixation. Practical agronomic and microbiological means to maximise N inputs by legumes have also been identified.     

Factors regulating the contributions of fixed nitrogen by pasture and crop legumes to different farming systems of eastern Australia

On-farm and experimental measures of the proportion (%Ndfa) and amounts of N₂ fixed were undertaken for 158 pastures either based on annual legume species (annual medics, clovers or vetch), or lucerne (alfalfa), and 170 winter pulse crops (chickpea, faba bean, field pea, lentil, lupin) over a 1200 km north-south transect of eastern Australia. The average annual amounts of N₂ fixed ranged from 30 to 160 kg shoot N fixed ha^–1 yr^–1 for annual pasture species, 37–128 kg N ha^–1 yr^–1 for lucerne, and 14 to 160 kg N ha^–1 yr^–1 by pulses. These data have provided new insights into differences in factors controlling N₂ fixation in the main agricultural systems. Mean levels of %Ndfa were uniformly high (65–94%) for legumes growing at different locations under dryland (rainfed) conditions in the winter-dominant rainfall areas of the cereal-livestock belt of Victoria and southern New South Wales, and under irrigation in the main cotton-growing areas of northern New South Wales. Consequently N₂ fixation was primarily regulated by biomass production in these areas and both pasture and crop legumes fixed between 20 and 25 kg shoot N for every tonne of shoot dry matter (DM) produced. Nitrogen fixation by legumes in the dryland systems of the summer-dominant rainfall regions of central and northern New South Wales on the other hand was greatly influenced by large variations in %Ndfa (0–81%) caused by yearly fluctuations in growing season (April–October) rainfall and common farmer practice which resulted in a build up of soil mineral-N prior to sowing. The net result was a lower average reliance of legumes upon N₂ fixation for growth (19–74%) and more variable relationships between N₂ fixation and DM accumulation (9–16 kg shoot N fixed/t legume DM). Although pulses often fixed more N than pastures, legume-dominant pastures provided greater net inputs of fixed N, since a much larger fraction of the total plant N was removed when pulses were harvested for grain than was estimated to be removed or lost from grazed pastures. Conclusions about the relative size of the contributions of fixed N to the N-economies of the different farming systems depended upon the inclusion or omission of an estimate of fixed N associated with the nodulated roots. The net amounts of fixed N remaining after each year of either legume-based pasture or pulse crop were calculated to be sufficient to balance the N removed by at least one subsequent non-legume crop only when below-ground N components were included. This has important implications for the interpretation of the results of previous N₂ fixation studies undertaken in Australia and elsewhere in the world, which have either ignored or underestimated the N present in the nodulated root when evaluating the contributions of fixed N to rotations.     

Significant nonsymbiotic nitrogen fixation in Patagonian ombrotrophic bogs

Nitrogen (N) nutrition in pristine peatlands relies on the natural input of inorganic N through atmospheric deposition or biological dinitrogen (N₂) fixation. However, N₂ fixation and its significance for N cycling, plant productivity, and peat buildup are mostly associated with the presence of Sphagnum mosses. Here, we report high nonsymbiotic N₂‐fixation rates in two pristine Patagonian bogs with diversified vegetation and natural N deposition. Nonsymbiotic N₂ fixation was measured in samples from 0 to 10, 10 to 20, and 40 to 50 cm depth using the ¹⁵N₂ assay as well as the acetylene reduction assay (ARA). The ARA considerably underestimated N₂ fixation and can thus not be recommended for peatland studies. Based on the ¹⁵N₂ assay, high nonsymbiotic N₂‐fixation rates of 0.3–1.4 μmol N₂ g^?1 day^?1 were found down to 50 cm under micro‐oxic conditions (2 vol.%) in samples from plots covered by Sphagnum magellanicum or by vascular cushion plants, latter characterized by dense and deep aerenchyma roots. Peat N concentrations point to greater potential of nonsymbiotic N₂ fixation under cushion plants, likely because of the availability of easily decomposable organic compounds and oxic conditions in the rhizosphere. In the Sphagnum plots, high N₂ fixation below 10 cm depth rather reflects the potential during dry periods or low water level when oxygen penetrates the top peat layer and triggers peat mineralization. Natural abundance of the ¹⁵N isotope of live Sphagnum (5.6 δ‰) from 0 to 10 cm points to solely N uptake from atmospheric deposition and nonsymbiotic N₂ fixation. A mean ¹⁵N signature of ?0.7 δ‰ of peat from the cushion plant plots indicates additional N supply from N mineralization. Our findings suggest that nonsymbiotic N₂ fixation overcomes N deficiency in different vegetation communities and has great significance for N cycling and peat accumulation in pristine peatlands.     

River-wetland interaction and carbon cycling in a semi-arid riverine system: the Okavango Delta, Botswana

The Okavango River, in semi-arid northwestern Botswana, flows for over 400 km in a pristine wetland developed on a large (>22,000 km²) alluvial fan (Okavango Delta). An annual flood pulse inundates the floodplains of the wetlands and travels across the Delta in 4–6 months. In this study, we assess the effects of long hydraulic residence time, variable hydrologic interaction between river–floodplain–wetland and evapotranspiration on carbon cycling. We measured dissolved inorganic carbon (DIC) concentrations and stable carbon isotopes of DIC (δ¹³C_DIC) from river water when the Delta was not flooded (low water) and during flooding (high water). During low water, the average DIC concentration was 31 % higher and the δ¹³C_DIC 2.1 ‰ more enriched compared to high water. In the lower Delta with seasonally flooded wetlands, the average DIC concentration increased by 70 % during low water and by 331 % during high water compared to the Panhandle with permanently flooded wetlands. The increasing DIC concentration downriver is mostly due to evapoconcentration from transpiration and evaporation with increased transit time. The average δ¹³C_DIC between low and high water decreased by 3.7 ‰ in the permanently flooded reaches compared to an increase of 1.6 ‰ in the seasonally flooded reaches. The lower δ¹³C_DIC during high water in the permanently flooded reaches suggest that DIC influx from the floodplain-wetland affects river’s DIC cycling. In contrast, higher river channel elevations relative to the wetlands along seasonal flooded reaches limit hydrologic interaction and DIC cycling occurs mostly by water column processes and river-atmospheric exchange. We conclude that river-wetlands interaction and evapoconcentration are important factors controlling carbon cycling in the Okavango Delta.     

Adaptation of methods for evaluating N2 fixation in food legumes and legume cover crops

Methods for partitioning the nitrogen assimilated by nodulated legumes, between nitrogen derived from soil sources and from N₂ fixation, are described as applied in peninsular Malaysia. The analysis of nitrogenous components translocated from the roots to the shoots of nodulated plants in the xylem sap is outlined, with some precautions to be observed for applications in the tropics. Some examples of the use of the technique in surverying apparent N₂ fixation by tropical legumes, in studying interrow cropping in plantation systems and in assessing effects of experimental treatments on N₂ fixation by food legumes, are described. Techniques for assesing N₂ fixation by means of¹⁵N abundance have been used to show that applications of nitrogenous fertilizers commonly used in Malaysia for soybeans depress N₂ fixation, that similar results are obtained with natural abundance and¹⁵N-enrichment methods and that, in at least two locations in Malaysia, differences between the natural abundance of¹⁵N in plant-available soil nitrogen and in atmospheric N₂ are great enough to permit application to measurement of N₂ fixation by leguminous crops.     

Enhancing legume N2 fixation through plant and soil management

Atmospheric N₂ fixed symbiotically by associations between Rhizobium spp. and legumes represents a renewable source of N for agriculture. Contribution of legume N₂ fixation to the N-economy of any ecosystem is mediated by: (i) legume reliance upon N₂ fixation for growth, and (ii) the total amount of legume-N accumulated. Strategies that change the numbers of effective rhizobia present in soil, reduce the inhibitory effects of soil nitrate, or influence legume biomass all have potential to alter net inputs of fixed N. A range of management options can be applied to legumes growing in farming systems to manipulate N₂ fixation and improve the N benefits to agriculture and agroforestry.     

Natural 15N abundance of vegetation and soil in the Kapalga savanna,Australia

Abstract The natural abundance of the stable isotope ¹⁵N was measured in different vegetation components and in the soil of a northern Australian savanna. Most of the vegetation was found to be ¹⁵N-depleted compared to atmospheric N₂. Herbaceous legumes, perennial grasses, tree legumes, non-legume trees and annual grasses exhibited mean δ¹⁵N of ? 1.7, ? 0.8, ? 0.7, 0.0 and + 0.3‰, respectively. These results are in good agreement with previous studies. Legumes exhibit slightly negative values, indicating that they are likely to be nitrogen-fixing plants. Non-legume plants have a δ¹⁵N close to zero, which could equally result from non-symbiotic fixation, soil organic matter mineralization, or fresh root litter mineralization. In contrast, soil organic matter was ¹⁵N-enriched. Values of δ¹⁵N increased with depth and were + 2.5, + 5.2 and +6.1‰ in the 0–10, 10–20 and 20–40cm layers, respectively. Soil organic matter δ¹⁵N shows a typical profile of mature soils.     

Effect of NPK on growth and nitrogen fixation of Sesbania rostrata as a green manure for lowland rice (Oryza sativa L.)

The stem-nodulating tropical legume Sesbania rostrata is a promising green manure species for low input rice-farming systems in lowland areas. However, its success as biofertilizer depends on its biomass production and N₂ fixation. Nutrient imbalances and soils low in available nutrients can considerably affect biofertilizer production. Use of mineral N, P, and K fertilizers in growing S. rostrata as biofertilizer for lowland rice was therefore evaluated in pot experiments, and in the fields in Central Luzon, Philippines. Two soils low in Olsen P (3–7.3 mg kg^–1) and exchangeable K (0.05–0.08 meq 100g^-1) were used. Increasing amounts of N (0, 10, 20, 30, and 40 mg kg^-1), P (0, 50, and 100 mg kg^-1), and K (0, 100, 200, and 300 mg kg^-1) were applied to S. rostrata grown in the greenhouse, whereas small amounts of N, P, and K fertilizers (30, 15, and 33 kg ha^-1, respectively) were applied in the field.Mineral N application depressed nodulation and N₂ fixation in roots. It however, stimulated nodulation and N₂ fixation in stems. Applying 30 kg N ha^-1 as urea increased total N accumulation by S. rostrata and yield of the subsequent rice crop (IR64). Applied P and K both stimulated growth, nodulation, and N₂ fixation of S. rostrata. Nitrogen accumulation in P- and K-fertilized S. rostrata was about 40% higher than that in nonfertilized green manure. Thus integration of mineral N, P, and K fertilizers in a green manure-based rice-farming system can considerably improve biofertilizer production and increase rice grain yield.     

Nitrogen Transfer from Four Nitrogen-Fixer Associations to Plants and Soils

Nitrogen (N) fixation is the main source of ‘new’ N for N-limited ecosystems like subarctic and arctic tundra. This crucial ecosystem function is performed by a wide range of N₂ fixer (diazotroph) associations that could differ fundamentally in their timing and amount of N release to the soil. To assess the importance of different associative N₂ fixers for ecosystem N cycling, we tracked ¹⁵N-N₂ into four N₂-fixer associations (with a legume, lichen, free-living, moss) and into soil, microbial biomass and non-diazotroph-associated plants 3 days and 5 weeks after in situ labelling. In addition, we tracked ¹³C from ¹³CO₂ labelling to assess if N and C fixation are linked. Three days after labelling, half of the fixed ¹⁵N was recovered in the legume soils, indicating a fast release of fixed N₂. Within 5 weeks, the free-living N₂ fixers released two-thirds of the fixed ¹⁵N into the soil, whereas the lichen and moss retained the fixed ¹⁵N. Carbon and N₂ fixation were linked in the lichen shortly after labelling, in free-living N₂ fixers 5 weeks after labelling, and in the moss at both sampling times. The four investigated N₂-fixer associations released fixed N₂ at different rates into the soil, and non-diazotroph-associated plants have no access to ‘new’ N within several weeks after N₂ fixation. Although legumes and free-living N₂ fixers are immediate sources of ‘new’ N for N-limited tundra ecosystems, lichens and especially mosses, do not contribute to increase the N pool via N₂ fixation in the short term.     

Contrasting δ<Superscript>15</Superscript>N Values of Atmospheric Deposition and <Emphasis Type="Italic">Sphagnum</Emphasis> Peat Bogs: N Fixation as a Possible Cause

Nitrogen (N) isotope systematics were investigated at two high-elevation ombrotrophic peat bogs polluted by farming and heavy industry. Our objective was to identify N sources and sinks for isotope mass balance considerations. For the first time, we present a time-series of δ¹⁵Ν values of atmospheric input at the same locations as δ¹⁵Ν values of living Sphagnum and peat. The mean δ¹⁵Ν values systematically increased in the order: input NH₄ ⁺ (?10.0‰) < input NO₃ ^? (?7.9‰) < peat porewater (?5.6‰) < Sphagnum (?5.0‰) < shallow peat (?4.2‰) < deep peat (?2.2‰) < runoff (?1.4‰) < porewater N₂O (1.4‰). Surprisingly, N of Sphagnum was isotopically heavier than N of the atmospheric input (P < 0.001). If partial incorporation of reactive N from the atmosphere into Sphagnum was isotopically selective, the residual N would have to be isotopically extremely light. Such N, however, was not identified anywhere in the ecosystem. Alternatively, Sphagnum may have contained an admixture of isotopically heavier N. Ambient air contains such N in the form of N₂ (δ¹⁵Ν_N2 = 0‰). Because high energy is required to break the triple bond, microbial N fixation is likely to proceed only under limited availability of pollutant N. Also for the first time, a δ¹⁵Ν comparison is presented between anoxic deeper peat and porewater N₂O. Isotopically light N is removed from anoxic substrate by denitrification, whose final product, N₂, escapes into the atmosphere. Porewater N₂O is an isotopically heavy residuum following partial N₂O reduction to N₂.