Trends in biological nitrogen fixation research and application

In the world each year 17.2×10⁷ tons of N are biologically fixed. Biological nitrogen fixation (BNF) contributes to plant production in arable lands and in natural ecosystems. Research to improve BNF is progressing through the breeding of efficient N-fixing organisms and host plants, selection of the best combinations of host plant and microsymbiont, and by the improvement of inoculation techniques and field management. Biotechnology is useful for the creation of promising N₂-fixing organisms. However, to increase plant production through enhanced BNF the constraints in establishing effective N₂-fixing systems in the field should be understood and eliminated.     

Fertilizers and biological nitrogen fixation as sources of plant nutrients: Perspectives for future agriculture

Biological nitrogen fixation (BNF) has an assured place in agriculture, mainly as a source of nitrogen for legumes. Legumes are currently grown mostly as a source of vegetable oil and as food for humans and animals, but not as nitrogen source.Other crops with BNF capability may be eventually be developed eventually. Such crops will also need mineral fertilizers to maintain a good status of soil nutrients, but their possible effects to the environment is also a concern. Fertilizers, however, will remain a necessary and sustainable input to agriculture to feed the present and increasing human population. It is not a case of whether BNF is better or worse than mineral fertilizers because both plays an important role in agriculture.     

Phosphorus availability and elevated CO2 affect biological nitrogen fixation and nutrient fluxes in a clover-dominated sward

The response of biological nitrogen fixation (BNF) to elevated CO(2) was examined in white clover (Trifolium repens)-dominated swards under both high and low phosphorus availability. Mixed swards of clover and buffalo grass (Stenotaphrum secundatum) were grown for 15 months in 0.2 m2 sand-filled mesocosms under two CO2 treatments (ambient and twice ambient) and three nutrient treatments [no N, and either low or high P (5 or 134 kg P ha(-1)); the third nutrient treatment was supplied with high P and N (240 kg N ha(-1))]. Under ambient CO2, high P increased BNF from 410 to 900 kg ha(-1). Elevated CO2 further increased BNF to 1180 kg ha(-1) with high P, but there was no effect of CO2 on BNF with low P. Allocation of N belowground increased by approx. 50% under elevated CO2 irrespective of supplied P. The results suggest that where soil P availability is low, elevated CO2 will not increase BNF, and pasture quality could decrease because of a reduction in aboveground N.     

Biological nitrogen fixation: rates,patterns and ecological controls in terrestrial ecosystems

New techniques have identified a wide range of organisms with the capacity to carry out biological nitrogen fixation (BNF)—greatly expanding our appreciation of the diversity and ubiquity of N fixers—but our understanding of the rates and controls of BNF at ecosystem and global scales has not advanced at the same pace. Nevertheless, determining rates and controls of BNF is crucial to placing anthropogenic changes to the N cycle in context, and to understanding, predicting and managing many aspects of global environmental change. Here, we estimate terrestrial BNF for a pre-industrial world by combining information on N fluxes with ¹⁵N relative abundance data for terrestrial ecosystems. Our estimate is that pre-industrial N fixation was 58 (range of 40–100) Tg N fixed yr⁻¹; adding conservative assumptions for geological N reduces our best estimate to 44 Tg N yr⁻¹. This approach yields substantially lower estimates than most recent calculations; it suggests that the magnitude of human alternation of the N cycle is substantially larger than has been assumed.     

Managing native and legume-fixed nitrogen in lowland rice-based cropping systems

Lowlands comprise 87% of the 145 M ha of world rice area. Lowland rice-based cropping systems are characterized by soil flooding during most of the rice growing season. Rainfall distribution, availability of irrigation water and prevailing temperatures determine when rice or other crops are grown. Nitrogen is the most required nutrient in lowland rice-based cropping systems. Reducing fertilizer N use in these cropping systems, while maintaining or enhancing crop output, is desirable from both environmental and economic perspectives. This may be possible by producing N on the land through legume biological nitrogen fixation (BNF), minimizing soil N losses, and by improved recycling of N through plant residues. At the end of a flooded rice crop, organic- and NH₄-N dominate in the soil, with negligible amounts of NO₃. Subsequent drying of the soil favors aerobic N transformations. Organic N mineralizes to NH₄, which is rapidly nitrified into NO₃. As a result, NO₃ accumulates in soil during the aerobic phase. Recent evidence indicates that large amounts of accumulated soil NO₃ may be lost from rice lowlands upon the flooding of aerobic soil for rice production. Plant uptake during the aerobic phase can conserve soil NO₃ from potential loss. Legumes grown during the aerobic phase additionally capture atmospheric N through BNF. The length of the nonflooded season, water availability, soil properties, and prevailing temperatures determine when and where legumes are, or can be, grown. The amount of N derived by legumes through BNF depends on the interaction of microbial, plant, and environmental determinants. Suitable legumes for lowland rice soils are those that can deplete soil NO₃ while deriving large amounts of N through BNF. Reducing soil N supply to the legume by suitable soil and crop management can increase BNF. Much of the N in legume biomass might be removed from the land in an economic crop produce. As biomass is removed, the likelihood of obtaining a positive soil N balance diminishes. Nonetheless, use of legumes rather than non-legumes is likely to contribute higher quantities of N to a subsequent rice crop. A whole-system approach to N management will be necessary to capture and effectively use soil and atmospheric sources of N in the lowland rice ecosystem.IRRI-NifTAL-IFDC joint contribution.     

Estimation of nitrogen fixation and transfer from alfalfa to associated grasses in mixed swards under field conditions

Fixation and transfer of nitrogen (N) from alfalfa (Medicago sativa L.) to different grass species including timothy (Phleum pratense L.) and bromegrass (Bromus inermis Leyss) were studied under field conditions, using the¹⁵N dilution technique.The percentage of alfalfa N derived from fixation (%NF) increased throughout the growing seasons and ranged from 62 to 83%. Nitrogen transfer (NT) from alfalfa to associated grasses was evident and contributed 26,46 and 38% of the total annual N yield of associated grasses or represented absolute amounts of 5, 20 and 19 kg N ha^–1 during the first, second and third year, respectively. The gradual and consistent percentage of NT that occurred before first harvest indicated that this transfer is a result of a direct excretion of N compounds from alfalfa root systems. Decomposition of root and nodule debris seems to contribute to the NT from alfalfa to associated grasses in the later cuts. All grass species benefitted similarly from alfalfa, although earlier maturing species with greater competitive ability were slightly more responsive.Contribution No. 1159 from the Plant Research Centre     

Biological nitrogen fixation in trees in agro-ecosystems

The integration of trees, especially nitrogen fixing trees (NFTs), into agroforestry and silvo-pastoral systems can make a major contribution to sustainable agriculture by restoring and maintaining soil fertility, and in combating erosion and desertification as well as providing fuelwood. The particular advantage of NFTs is their biological nitrogen fixation (BNF), their ability to establish in nitrogen-deficient soils and the benefits of the nitrogen fixed (and extra organic matter) to succeeding or associated crops.The importance of NFTs leads to the question of how we can maximise or optimize their effects and how we can manage BNF and the transfer of nitrogen to associated or succeeding plantings. To be able to achieve these goals, suitable methods of measuring BNF in trees are necessary. The total nitrogen difference (TND) method is simple, but is better suited for low than high soil N conditions. The acetylene reduction assay (ARA), although sensitive and simple, has many technical limitations especially for NFTs, and the estimates of BNF have generally been very low, compared to other methods. For NFTs, the ¹⁵N techniques are still under development, but have already given some promising results (e.g., has been used to measure large genetic variability in BNF within different NFTs).Various factors affect BNF in trees. They include the age of trees, the microbial component, soil moisture, temperature, salinity, pH, soil N level and plant nutrient deficiencies. Some of the factors, e.g. temperature, affect the symbiosis more than plant growth, and differences in the effects of these factors on BNF in different NFT genotypes have been reported. These factors and research needs for improving BNF in trees are discussed.     

生物固氮在我国农业生产中的研究现状及发展对策

简述了生物固氦在我国农业生产中3个热点的研究现状:提高现有固氦生物的固氦能力、化学模拟固氦酶、固氦微生物的生物工程,同时提出了生物固氦在令后生态农业中的发展对策。     

Effect of nitrogen fertilization and cropping system of the reference crop on estimation of N2 fixation by vetch using15N methodology

This study was conducted to examine the effects of varying N rates and cropping systems (mixedversus pure stand) on the suitability of oats (Avena sativa L.) for estimating N₂ fixed in sequentially harvested vetch (Vicia sativa L.) over two growing seasons (1984–85 and 1985–86). The N rates were, 20 and 100 kg N ha^–1 in 1984–85 and 15 and 60 kg N ha^–1 in 1985–86. In the 1984–85 season, vetch at maturity derived 76 and 63% N from fixation at the high and low N rates respectively. The corresponding values for the second season were 66 and 42%. Except in the 1985–86 season when some significantly higher values of % N₂ fixed were estimated by using the reference crop grown at the higher (A-value approach) than at the lower N rate (isotope-dilution approach), both approaches resulted in similar measurements of N₂ fixed. In the 1984–85 season, similar values of N₂ fixed were obtained using either the pure or mixed stand oats reference crops. Although in the 1985–86 season, the mixed reference crop occasionally estimated lower % N₂ fixed than pure oats, total N₂ fixed estimates were always similar (P<0.05). Thus, in general, N fertilization and cropping system of the reference crop did not significantly influence estimates of N₂ fixation.     

Crop rotation and residue management effects on carbon sequestration,nitrogen cycling and productivity of irrigated rice systems

The effects of soil aeration, N fertilizer, and crop residue management on crop performance, soil N supply, organic carbon (C) and nitrogen (N) content were evaluated in two annual double-crop systems for a 2-year period (1994–1995). In the maize-rice (M-R) rotation, maize (Zea mays, L.) was grown in aerated soil in the dry season (DS) followed by rice (Oriza sativa, L.) grown in flooded soil in the wet season (WS). In the continuous rice system (R-R), rice was grown in flooded soil in both the DS and WS. Subplot treatments within cropping-system main plots were N fertilizer rates, including a control without applied N. In the second year, sub-subplot treatments with early or late crop residue incorporation were initiated after the 1995 DS maize or rice crop. Soil N supply and plant N uptake of 1995 WS rice were sensitive to the timing of residue incorporation. Early residue corporation improved the congruence between soil N supply and crop demand although the size of this effect was influenced by the amount and quality of incorporated residue. Grain yields were 13-20% greater with early compared to late residue incorporation in R-R treatments without applied N or with moderate rates of applied N. Although substitution of maize for rice in the DS greatly reduced the amount of time soils remained submerged, the direct effects of crop rotation on plant growth and N uptake in the WS rice crops were small. However, replacement of DS rice by maize caused a reduction in soil C and N sequestration due to a 33–41% increase in the estimated amount of mineralized C and less N input from biological N fixation during the DS maize crop. As a result, there was 11–12% more C sequestration and 5–12% more N accumulation in soils continuously cropped with rice than in the M-R rotation with the greater amounts sequestered in N-fertilized treatments. These results document the capacity of continuous, irrigated rice systems to sequester C and N during relatively short time periods. This revised version was published online in June 2006 with corrections to the Cover Date.     

Photosynthesis and nitrogen fixation in a cyanobacterial bloom in the Baltic Sea

Daily integrals of photosynthesis by a cyanobacterial bloom in the Baltic Sea, during the summer of 1993, were calculated from the vertical distributions of light, temperature and the organisms in the water column and from photosynthesis/irradiance curves of picoplanktonic and diazotrophic cyanobacteria isolated from the community. The distribution of chlorophyll a in size-classes <20?µm and >20?µm was monitored over 9 days that included a deep mixing event followed by calm. Picocyanobacteria formed 70% of the cyanobacterial biomass and contributed 56% of the total primary production. Of the filamentous diazotrophs that formed the other 30%, Aphanizomenon contributed 28% and a Nodularia-containing fraction 16% of the primary production. For the whole population there was little change in standardized photosynthetic O₂ production, which remained at about 31?mmol?m^?2 before and after the mixing event. There were differences, however, between the classes of cyanobacteria: in picocyanobacteria primary production hardly changed, while in Aphanizomenon it increased by 2.6 and in Nodularia it fell below zero. Total phytoplankton photosynthesis was strongly dependent on total daily insolation with the compensation point at a photon insolation of 22.7?mol?m^?2?d^?1. Similar analyses of N₂ fixation showed much less dependence on depth distribution of light and biomass: Aphanizomenon fixed about twice as much N₂ as Nodularia their; their fixation exceeded their own N demand by about 12%. Together, these species contributed 49% of the total N demand of the phytoplankton population. Computer models based on the measured light attenuation and photosynthetic coefficients indicate that growth of the cyanobacterial population could occur only in the summer months when the critical depth of the cyanobacteria exceeds the depth of mixing.     

Growth and N2 fixation in mixed cropping of Medicago arborea and Atriplex halimus grown on a salt-affected soil using a 15N tracer technique

Abstract

The objective of this study was to evaluate dry matter (DM), nitrogen yield, N₂ fixation (Ndfa) and soil N uptake (Ndfs) in the shrubby medic (Medicago arborea) and saltbush (Atriplex halimus) grown in pots either solely or in a mixture on a salt-affected soil, using ¹⁵N dilution method. The combined DM of both species was considerably higher than that of solely grown shrubs. The inclusion of saltbush in the mixed cropping system decreased Ndfs by shrubby medic and enhanced % Ndfa without affecting amounts of N₂ fixed. It can be concluded that the use of mixed cropping system of shrubby medic and saltbush could be a promising bio-saline agricultural approach to utilize salt affected soils in terms of forage yield and N₂ fixation.     

Role of biological nitrogen fixation in legume based cropping systems; a case study of West Africa farming systems

Nitrogen (N) has been gradually depleted from West African soils and now poses serious threats to food production. Many ways of increasing N supply (e.g. judicious use of inorganic fertilizers and nitrogen-fixing plants) have been tried in West African farming systems. Herbaceous and woody legumes commonly contribute 40–70 kg N ha^–1 season. This represents about 30% of the total N applied as residues. Nevertheless and despite repeated demonstrations of the usefulness of green manures in enhancing soil fertility, their practices and adoption are still limited. Promiscuous soyabeans are being used to develop sustainable cropping systems in the moist savannah. Reliable estimates of N₂ fixed by soyabeans and their residual N benefits to subsequent cereal crops in the savannah zone of southern Guinea have only infrequently been made. The actual amounts measured varied between 38 and 126 kg N ha^–1 assuming that only seeds of soyabeans are removed from the plots, the net N accrual of soil nitrogen ranges between minus 8 kg N ha^–1 and plus 47 kg N ha^–1 depending on the soyabean cultivar. Residual soyabean N values of 10–24 kg N ha^–1 (14–36% of the total N in maize) were obtained in a soyabean-maize rotation. Although cereal yields following legume cultivation have been attributed to greater N accumulation, our data show that the relative increase in maize N was smaller than the relative increase in dry-matter yield. Hence, the increased yields of maize following soybeans are not entirely due to the carry-over of N from soyabean residues (as well as to conservation of soil N) but to other rotational effects as well. It is thus clear that the Nbenefit of grain legumes to non-legumes is small compared to the level of N fertilizer use in more intensive cereal production systems but is nevertheless significant in the context of the low amounts of input in subsistence farming.     

Biological nitrogen fixation in four Gliricidia sepium genotypes

A field experiment was conducted at the Coconut Research Institute in Sri Lanka to examine the biological nitrogen fixation potential of three Gliricidia sepium provenances (OFI 14/84, 17/84, 12/86) and a local landrace (designated LL), using the ¹⁵N isotope dilution method. There was marked variation in dry matter, total N, nodulation and ¹⁵N enrichment among the Gliricidia genotypes (=0.001), and the dry matter yield of Cassia siamea (syn. Senna siamea), the non-N₂ fixing reference plant was higher than for G. sepium. In all cases, highest biomass and total N were aboveground, with roots on average accounting for < 20 % of total dry matter or the total N in plants. Atom % ¹⁵N excess was highest in C. siamea, and lowest in OFI 14/84. Although atom % ¹⁵N excess was lower in Gliricidia leaves than in the other organs (all of which had similar ¹⁵N enrichments), values of % N derived from atmospheric N₂ fixation (% Ndfa) calculated for any individual organ or for the whole plant were similar. This was because the relative distribution of ¹⁵N in the different parts of the fixing plant followed the same trend as in the reference plant. There were significant differences (p=0.01) in N₂ fixation between the Gliricidia genotypes. The values ranged from 17.8 g N tree^-1 (equivalent to 86 kg N ha^-1 at 5000 trees ha^-1) in OFI 12/86 to 61.7g N tree^-1 (equivalent to 309 kg N ha^-1) in OFI 14/84. Although most of this variability was due to differences in both % Ndfa and total N in plant, amount of N fixed was more correlated with total N in plant (r=0.935) than with % Ndfa (r=0.707). On average, % Ndfa in all three G. sepium provenances and LL was about 55 % or 34.6 g N tree^-1 (equivalent to some 166 kg N ha^-1) in the 9 months within which N₂ fixation was measured. This represents a substantial contribution of N into the soil-plant system.     

The effects of elevated CO2 on symbiotic N2 fixation: a link between the carbon and nitrogen cycles in grassland ecosystems

The response of plants to elevated CO₂ is dependent on the availability of nutrients, especially nitrogen. It is generally accepted that an increase in the atmospheric CO₂ concentration increases the C:N ratio of plant residues and exudates. This promotes temporary N-immobilization which might, in turn, reduce the availability of soil nitrogen. In addition, both a CO₂ stimulated increase in plant growth (thus requiring more nitrogen) and an increased N demand for the decomposition of soil residues with a large C:N will result under elevated CO₂ in a larger N-sink of the whole grassland ecosystem. One way to maintain the balance between the C and N cycles in elevated CO₂ would be to increase N-import to the grassland ecosystem through symbiotic N₂ fixation. Whether this might happen in the context of temperate ecosystems is discussed, by assessing the following hypothesis: i) symbiotic N₂ fixation in legumes will be enhanced under elevated CO₂, ii) this enhancement of N₂ fixation will result in a larger N-input to the grassland ecosystem, and iii) a larger N-input will allow the sequestration of additional carbon, either above or below-ground, into the ecosystem. Data from long-term experiments with model grassland ecosystems, consisting of monocultures or mixtures of perennial ryegrass and white clover, grown under elevated CO₂ under free-air or field-like conditions, supports the first two hypothesis, since: i) both the percentage and the amount of fixed N increases in white clover grown under elevated CO₂, ii) the contribution of fixed N to the nitrogen nutrition of the mixed grass also increases in elevated CO₂. Concerning the third hypothesis, an increased nitrogen input to the grassland ecosystem from N₂ fixation usually promotes shoot growth (above-ground C storage) in elevated CO₂. However, the consequences of this larger N input under elevated CO₂ on the below-ground carbon fluxes are not fully understood. On one hand, the positive effect of elevated CO₂ on the quantity of plant residues might be overwhelming and lead to an increased long-term below-ground C storage; on the other hand, the enhancement of the decomposition process by the N-rich legume material might favour carbon turn-over and, hence, limit the storage of below-ground carbon.     

Interactions between white clover and ryegrass under contrasting nitrogen availability: N2 fixation,N fertilizer recovery,N transfer and water use efficiency

Seasonal variation in N₂ fixation, N transfer from clover to ryegrass, and soil N absorption in white clover–ryegrass swards were investigated under field conditions over three consecutive years. The plots were established with different seeding ratios of clover and ryegrass and contrasting fertilizer N ranging from 3 to 72 kg ha^-1 year^-1.An initially poor clover population needed at least one growing season to reach the same yield output as an initially well established clover population. The clover content of the sward decreased by the annual application of 72 kg N ha^-1 but not by smaller N dressings.The total amount of atmospherically derived N in clover growing in mixture with ryegrass was, on average over the three years equal to 83, 71, 68 and 60 kg N ha^-1 for the treatments of 3, 24, 48 and 72 kg N ha^-1, respectively. The proportion of atmospherically derived N declined with increasing N application, but never became smaller than 80% of total clover N. The proportion of atmospherically derived N in a pure stand white clover amounted to 60–80% of the total N content, equivalent to 109, 110, 103 and 90 kg N ha^-1 for the treatments of 3, 24, 48 and 72 kg N ha^-1, respectively.Only small amounts of atmospherically derived N was transferred to the associated ryegrass during the first production year, while in each of the following years up to 21 kg ha^-1 was transferred. The average amount of N transferred from clover to ryegrass was equivalent to 3, 16 and 31% of the N accumulated in ryegrass in the first, second and third production year, respectively. Expressed relative to the total amount of fixed N₂ in the clover–ryegrass mixture, the transfer amounted to 3, 17 and 22% in the first, second and third production year, respectively. Thus transfer of atmospherically derived N from clover contributed significantly to the N economy of the associated ryegrass.The clover–ryegrass mixture absorbed constantly higher amount of soil derived N than the pure stands of the two species. Only 11% of the total accumulated fertilizer N and soil derived N in the mixture was contained within the clover component. Lower water use efficiencies for the plants grown in mixture compared to pure stands were mainly related to the increased N uptake in the mixture, with the subsequent increase in growth compared to the pure stands.It is concluded that positive interactions between clover and ryegrass growing in mixture ensure a more efficient fixation of atmospheric N₂ and absorption of fertilizer N and soil derived N than pure stands of the same species.     

Effects of white clover (Trifolium repens L.) on plant and soil nitrogen and soil organic matter in mixtures with perennial ryegrass (Lolium perenne L.)

To increase our insight into the above- and belowground N flows in grass and grass-clover swards relations between crop and soil parameters were studied in a cutting trial with perennial ryegrass (Lolium perenne) monocultures and ryegrass–white clover (Trifolium repens) mixtures. The effects of clover cultivar on herbage yield, the amount of clover-derived nitrogen, apparent N transfer to companion grass, dynamics of N and organic matter in the soil were estimated.The grass monocultures had very low DM yields (<2.1 t ha^-1) and a low N concentration in the harvested herbage. During 1992–1995 the annual herbage DM yield in the mixtures ranged from 7.0 to 14.3 t ha^-1, the white clover DM yield from 2.4 to 11.2 t ha^-1 and the mean annual clover content in the herbage DM harvested from 34 to 78%. Mixtures with the large-leaved clover cv. Alice yielded significantly more herbage and clover DM and had a higher clover content than mixtures with small/medium-leaved cvs. Gwenda and Retor. Grass cultivar did not consistently affect yield, botanical composition or soil characteristics.The apparent N₂ fixation was very high, ranging from 150 to 545 kg N ha^-1 in the different mixtures. For each tonne of clover DM in the harvested herbage 49 to 63 kg N was harvested, while the apparent N transfer from clover to grass varied between 55 and 113 kg N ha^-1 year^-1.The net N mineralization rate was lower under monocultures than under mixtures. The C mineralization and the amounts of C and N in active soil organic matter fractions were similar for monocultures and mixtures, but the C:N ratio of the active soil organic matter fractions were higher under grass than under mixtures. This explains the lower N mineralization under grass.     

Symbiotic nitrogen fixation in a tropical rainforest: 15N natural abundance measurements supported by experimental isotopic enrichment

* Leguminous trees are very common in the tropical rainforests of Guyana. Here, species-specific differences in N(2) fixation capability among nodulating legumes growing on different soils and a possible limitation of N(2) fixation by a relatively high nitrogen (N) and low phosphorus (P) availability in the forest were investigated. * Leaves of 17 nodulating species and 17 non-nodulating reference trees were sampled and their delta(15)N values measured. Estimates of N(2) fixation rates were calculated using the (15)N natural abundance method. Pot experiments were conducted on the effect of N and P availability on N(2) fixation using the (15)N-enriched isotope dilution method. * Nine species showed estimates of > 33% leaf N derived from N(2) fixation, while the others had low or undetectable N(2) fixation rates. High N and low P availability reduced N(2) fixation substantially. * The results suggest that a high N and low P availability in the forest limit N(2) fixation. At the forest ecosystem level, N(2) fixation was estimated at c. 6% of total N uptake by the tree community. We conclude that symbiotic N(2) fixation plays an important role in maintaining high amounts of soil available N in undisturbed forest.