Variability in nitrogen fixation of common bean (Phaseolus vulgaris L.) intercropped with maize

Thirty one selected bean lines were evaluated in the field for ability to support N₂ fixation when intercropped with maize which received 0, 30 and 60 kg N ha^–1 as ammonium sulphate. The amount of fixed N₂ was estimated using the natural variation of ¹⁵N and wheat as the standard non-fixing crop. Nitrogen as low as 15 kg N ha^–1 at sowing suppressed nodule weight and activity (acetylene reduction activity) but not nodule number, suggesting that the main effect of mineral N was on nodule development and function. ¹⁵N data revealed a high potential of the bean genotypes to fix N₂, with the most promising ones averaging between 50–60% of seed N coming from fixation. Bean lines CNF-480, Puebla-152, Mexico-309, Negro Argel, CNF-178, Venezuela-350 and WBR22-3, WBR22-50 and WBR22-55 were ranked as good fixers.     

Biotic and Abiotic Induction of a Fungitoxic Compound in Rice Bean (Vigna umbellata [Thunb.] Ohwi and Ohashi)

Young leaves of rice bean produced relatively more fungitoxic compound(s) than the older leaves. Epicotyls, roots, mature leaves and pods, but not cotyledon and cotyledonary leaves, synthesized the antifungal compound(s) following inoculation with anon-pathogen of this plant. Epicotyl segments when chopped or pinpricked produced the phytoalexin. Treatment of epicotyls with a dilute concentration of various chemicals caused induction of the fungitoxic compound(s). Induction by inoculation, injury and chemicals suggest a phytoalexin nature of the compound. Presence of phaseollin in the extracts of inoculated, injured or chemical-treated epicotyls has been demonstrated.     

Phosphorus requirements and nitrogen accumulation by three mungbean (Vigna radiata (L) Welzek) cultivars

A promising approach for overcoming poor crop yields in phosphorus (P)-deficient soils is to exploit the genetic variation among plants to grow under low P conditions. We examined the P requirements of three mungbean cultivars, T-77, MI-5 and E-72, using four P rates, 0, 30, 60 and 90 mg P kg^-1 soil (designated P₀, P₁, P₂ and P₃, respectively). Nodulation was highest in T-77, and unlike the other cultivars, nodule numbers were not increased by P application. Similarly, growth of T-77 was the highest, and was not influenced by P rates. In contrast shoot yields of MI-5 and E-72 at P₀ were only 76 and 65%, respectively, of the maximum obtained under P application. Nodule dry weight and the amount of N fixed (Ndfa) in each cultivar was enhanced by P application, with T-77 generally giving the lowest response, and accumulating the highest Ndfa. The data suggest a higher P requirement for N₂ fixation (especially for T-77) than for growth. All plants increased their P uptake as P rates increased, with T-77 accumulating the highest amount of P at each P level. Differences in the physiological P use efficiency, PPUE (g shoot mg^-1 P) among genotypes were generally not significant, neither were there any consistent trends as P rates changed. The ability to absorb P therefore appeared to be more important than PPUE in enhancing growth. We conclude from our data that it is possible by selection to obtain plants capable of good growth and high N₂ fixation in soils of low P; cultivar T-77 is a good example.     

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

There remains conflicting evidence on the relationship between P supply and biological N₂-fixation rates, particularly N₂-fixing plant adaptive strategies under P limitation. This is important, as edaphic conditions inherent to many economically and ecologically important semi-arid leguminous tree species, such as Acacia senegal, are P deficient. Our research objective was to verify N acquisition strategies under phosphorus limitations using isotopic techniques. Acacia senegal var. senegal was cultivated in sand culture with three levels of exponentially supplied phosphorus [low (200 μmol of P seedling⁻¹ over 12 weeks), mid (400 μmol) and high (600 μmol)] to achieve steady-state nutrition over the growth period. Uniform additions of N were also supplied. Plant growth and nutrition were evaluated. Seedlings exhibited significantly greater total biomass under high P supply compared to low P supply. Both P and N content significantly increased with increasing P supply. Similarly, N derived from solution increased with elevated P availability. However, both the number of nodules and the N derived from atmosphere, determined by the ¹⁵N natural abundance method, did not increase along the P gradient. Phosphorus stimulated growth and increased mineral N uptake from solution without affecting the amount of N derived from the atmosphere. We conclude that, under non-limiting N conditions, A. senegal N acquisition strategies change with P supply, with less reliance on N₂-fixation when the rhizosphere achieves a sufficient N uptake zone.     

The contribution of associative and symbiotic nitrogen fixation to the nitrogen nutrition of non-legumes

During the past 10 years estimates of N₂ fixation associated with sugar cane, forage grasses, cereals and actinorhizal plants grown in soil with and without addition of inoculum have been obtained using the ¹⁵N isotope dilution technique. These experiments are reviewed in this paper with the aim of determining the proportional and absolute contribution of N₂ fixation to the N nutrition of non-legumes, and its role as a source of N in agriculture. The review also identifies deficiencies in both the totality of data which are currently available and the experimental approaches used to quantify N₂ fixation associated with non-legumes.Field data indicate that associative N₂ fixation can potentially contribute agronomically-significant amounts of N (>30–40 kg N ha^-1 y^-1) to the N nutrition of plants of importance in tropical agriculture, including sugar cane (Saccharum sp.) and forage grasses (Panicum maximum, Brachiaria sp. and Leptochloa fusca) when grown in uninoculated, N-deficient soils. Marked variations in proportions of plant N derived from the atmosphere have been measured between species or cultivars within species.Limited pot-culture data indicate that rice can benefit naturally from associative N₂ fixation, and that inoculation responses due to N₂ fixation can occur. Wheat can also respond to inoculation but responses do not appear to be due to associative N₂ fixation. ¹⁵N dilution studies confirm that substantial amounts of N₂ can be fixed by actinorhizal plants.     

Natural 15N abundances of maize and soil amended with urea and composted pig manure

To investigate the effect of inorganic fertilizer and composted manure amendments on the N isotope composition (delta ¹⁵N) of crop and soil, maize (Zea mays L.) was cultivated under greenhouse conditions for 30, 40, 50, 60, and 70 days. Composted pig manure (delta ¹⁵N= +13.9) and urea (-2.3) were applied at 0 and 0 kg N ha^–1 (C0U0), 0 and 150 kg N ha^–1 (C0U2), 150 and 0 kg N ha^–1 (C2U0), and 75 and 75 kg N ha^–1 (C1U1), respectively. The delta ¹⁵N of total soil-N was not affected by both amendments, but delta ¹⁵N of NH⁺ ₄ and NO^– ₃ provided some information on the N isotope fractionation in soil. During the early growth stage, significant differences (P < 0.05) in delta ¹⁵N among maize subjected to different treatments were observed. After 30 days of growth, the delta ¹⁵N values of maize were +6.6 for C0U0, +1.1 for C0U2, +7.7 for C2U0, and +4.5 for C1U1. However, effects of urea and composted manure application on maize delta ¹⁵N progressively decreased with increasing growth period, probably due to isotope fractionation accompanying N losses and increased uptake of soil-derived N by maize. After 70 days of growth, delta ¹⁵N of leaves and grains of maize amended with composted pig manure were significantly (P < 0.05) higher than those with urea. The temporal variations in delta ¹⁵N of maize amended with urea and composted manure indicate that plant delta ¹⁵N is generally not a good tracer for N sources applied to field. Our data can be used in validation of delta ¹⁵N fractionation models in relation to N source inputs.     

Growing vetches (Vicia villosa Roth) in irrigated cotton systems: inputs of fixed N, N fertiliser savings and cotton productivity

We compared symbiotic N₂ fixation by winter forage legumes (clovers, medics and vetches) using the ¹⁵N natural abundance technique in three experiments. Vetches (Vicia spp.) were the most productive legumes, and woollypod vetch fixed (shoot+root) up to 265 kg N ha^–1 (mean 227 kg N ha^–1) during a 4–5 months period over winter and early spring. Balansa and Berseem clovers, and Gama medic were highly productive in the first experiment, but fixed significantly less N than woollypod vetch in the second experiment. A 6-year study (1997–2003) compared cotton (Gossypium hirsutum L.) systems with and without vetch, or with faba beans (Vicia faba L.) to assess the effects of these crops on cotton production. Woollypod vetch was grown either between annual cotton crops, or between wheat (Triticum aestivumL.) and cotton crops. Vetch added 230 kg N ha^–1 (174 kg fixed N ha^–1) to the soil when incorporated as a green manure. Faba bean shoot residues and nodulated roots contributed 108 kg fixed N ha^–1 to the soil, following the removal of 80 kg N ha^–1 in the harvested seed (meaned over three crops). Lablab (Lablab purpureus L. – summer-growing and irrigated) added 277 kg N ha^–1 (244 kg fixed N ha^–1) before incorporation as a green manure in the first year of the experiment. The economic optimum N fertiliser rate for each cropping system was determined every second year when all systems were sown to cotton. Cotton following cotton required 105 kg fertiliser N ha^–1, but only 40 kg N ha^–1 when vetch was grown between each cotton crop. Cotton following wheat required 83 kg fertiliser N ha^–1 but no N fertiliser was needed when vetch was grown after wheat (the highest yielding system). Cotton following faba beans also required no N fertiliser. The vetch-based systems became more N fertile over the course of the experiment and produced greater lint yields than the comparative non-legume systems, and required less N fertiliser. While no cash flow was derived from growing vetch, economic benefits accrued from enhanced cotton yields, reduced N fertiliser requirements and improved soil fertility. These findings help explain the rotational benefits of vetches observed in other regions of the world.     

Measurement of N2-fixation in field-grown pigeonpea [Cajanus cajan (L.) Millsp.] using15N-labelled fertilizer

Two experiments were carried out from 1981 to 1983 in Vertisol field at ICRISAT Center, Patancheru, India to measure N₂-fixation of pigeonpea [Cajanus cajan (L.) Millsp.] using the¹⁵N isotope dilution technique. One experiment examined the effect of control of a nodule-eating insect on fixation while another in vestigated the effect of intercroping with cereals on fixation and the residual effect of pigeonpea on a succeeding cereal crop. Although both experiments indicated that at least 88% of the N in pigeonpea was fixed from the atmosphere, one result is considered fortuitous in view of the differential rates of growth of the legume and the control, sorghum [Sorghum bicolor (L.) Moench]. The difference method of calculation in dieated negative fixation and the results emphasized the problem of finding a suitable nonfixing control. In a second experiment, when all plants were confined to a known volume of soil to which¹⁵N fertilizer was added in the field, these problems were overcome, and isotope dilution and difference methods gave similar results of N₂-fixation of about 90%. In intercropped pigeonpea 96% of the total N was derived from the atmosphere. This estimate might be an artifact. There was no evidence of benefit from N fixed by pigeonpea to intercropped sorghum plants. Plant tissue¹⁵N enrichments of cereal crops grown after pigeonpea indicated that the cereal derived some N fixed by the previous pigeonpea. Thus residual benefits to cereals are not only an effect of ‘sparing’ of soil N.     

Nitrogen fixation in legumes and actinorhizal plants in natural ecosystems: values obtained using 15N natural abundance

Background: Nitrogen fixation has been quantified for a range of crop legumes and actinorhizal plants under different agricultural/agroforestry conditions, but much less is known of legume and actinorhizal plant N₂ fixation in natural ecosystems.

Aims: To assess the proportion of total plant N derived from the atmosphere via the process of N₂ fixation (%Ndfa) by actinorhizal and legume plants in natural ecosystems and their N input into these ecosystems as indicated by their ¹⁵N natural abundance.

Methods: A comprehensive collation of published values of %Ndfa for legumes and actinorhizal plants in natural ecosystems and their N input into these ecosystems as estimated by their ¹⁵N natural abundance was carried out by searching the ISI Web of Science database using relevant key words.

Results: The %Ndfa was consistently large for actinorhizal plants but very variable for legumes in natural ecosystems, and the average value for %Ndfa was substantially greater for actinorhizal plants. High soil N, in particular, but also low soil P and water content were correlated with low legume N₂ fixation. N input into ecosystems from N₂ fixation was very variable for actinorhizal and legume plants and greatly dependent on their biomass within the system.

Conclusions: Measurement of ¹⁵N natural abundance has given greater understanding of where legume and actinorhizal plant N₂ fixation is important in natural ecosystems. Across studies, the average value for %Ndfa was substantially greater for actinorhizal plants than for legumes, and the relative abilities of the two groups of plants to utilise mineral N requires further study.     

Estimation of symbiotically fixed nitrogen in field grown soybeans: An application of natural15N/14N abundance and a low level15N-tracer technique

Summary Plants from agricultural and natural upland ecosystem were investigated for¹⁵N content to evaluate the role of symbiotic N₂-fixation in the nitrogen nutrition of soybean. Increased yields and lower δ¹⁵N values of nodulating soybeansvs, non-nodulating isolines gave semi-quantitative estimates of N₂ fixation. A fairly large discrepancy was found between estimations by δ¹⁵N and by N yield at 0 kg N/ha of fertilizer. More precise estimates were made by following changes in plant δ¹⁵N when fertilizer δ¹⁵N was varied near¹⁵N natural abundance level. Clearcut linear relationships between δ¹⁵N values of whole plants and of fertilizer were obtained at 30 kg N/ha of fertilizer for three kinds of soils. In experimental field plots, nodulating soybeans obtained 13±1% of their nitrogen from fertilizer, 66±8% from N₂ fixation and 21±10% from soil nitrogen in Andosol brown soil; 30%, 16% and 54% in Andosol black soil; 7%, 77% and 16% in Alluvial soil, respectively. These values for N₂ fixation coincided with each corresponding estimation by N yield method. Other results include: 1)¹⁵N content in upland soils and plants was variable, and may reflect differences in the mode of mineralization of soil organics, and 2) nitrogen isotopic discrimination during fertilizer uptake (δ¹⁵N of plant minus fertilizer) ranged from −2.2 to +4.9‰ at 0–30 kg N/ha of fertilizer, depending on soil type and plant species. The proposed method can accurately and relatively simply establish the importance of symbiotic nitrogen fixation for soybeans growing in agricultural settings.     

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.     

Early seedling and seasonal N2-fixing symbiotic activity of two soybean [Glycine max (L.) Merr.] cultivars inoculated with Bradyrhizobium strains of diverse origin

In areas with a short growing season the poor adaptability of soybean [Glycine max Meer. (L.)] to cool soil conditions is considered the primary yield limiting factor. Soybean requires temperatures in the 25 to 30°C range for optimum N₂-fixation and yield. Field studies were conducted in 1990 and 1991 at Montreal, Quebec to determine whether adaptability to cool soil conditions, with respect to earlier symbiosis establishment and function, existed among either Bradyrhizobium strains or soybean genotypes. An early maturing isoline of the soybean cultivar Evans and the cultivar Maple Arrow were inoculated with one of four strains isolated from the cold soils of Hakkaido, northern Japan, or the commercially used strains 532C or USDA110, at two planting dates. Plot biomass and nodulation were assessed at seedling (V2), and flowering(R2) growth stages and harvest maturity. Soybean genotypes did not differ for pre-flowering nodulation or N₂-fixation in the cool spring conditions of the first year. Seasonal N₂-fixation rates were also determined at the final harvest by the N-balance and ¹⁵N-isotope dilution methods. Significantly higher symbiotic activity was found for two of the four Hakkaido strains and was reflected in higher final soybean seed yield and total N₂-fixation for the growing season, as compared to the two commercial strains. Planting 14 days earlier resulted in greater early vegetative and total seasonal N₂ fixation and yield in the second year when soil temperatures were warmer, emphasizing the need for the development of soybean-Bradyrhizobium combinations superior in nodule development and function under cool soil conditions.     

An indirect method for estimating 15N isotope fractionation during nitrogen fixation by a legume under field conditions

A new technique is proposed for measuring ¹⁵N isotope fractionation during N fixation that obviates some of the possible disadvantages of existing methods. Accurate calculation of N fixation by legumes using the ¹⁵N natural abundance technique requires a value for the isotopic composition of fixed N as an input. Isotopic fractionation in fixed N in legumes has usually been measured using N- free solution culture but results can vary with Rhizobium strain and growth conditions. The proposed method avoids these problems and can be used as an integral part of a field experiment for evaluating N fixation.The technique is essentially a process of adjusting values of ¹⁵ N for fixed N until % N fixation calculated by the ¹⁵N natural abundance method best matches % N fixation estimated by the ¹⁵N enrichment method. The use of high % N fixation values improves the sensitivity and reliability of the method.A field evaluation of this comparison technique using chickpea (Cicer arietinum L.) provided a ¹⁵N isotope fractionation factor (–2.37) for fixed N close to that obtained by N-free solution culture methods (–2.10). The availability of these two independent techniques allowed mutual corroboration of estimates of ¹⁵N isotope fractionation during N fixation.

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Natural15N abundance as a method of estimating the contribution of biologically fixed nitrogen to N2-fixing systems: Potential for non-legumes

The¹⁵N abundance of plants usually closely reflects the¹⁵N abundance of their major immediate N source(s); plant-available soil N in the case of non-N₂-fixing plants and atmospheric N₂ in the case of N₂ fixing plants. The¹⁵N abundance values of these sources are usually sufficiently different from each other that a significant and systematic difference in the¹⁵N abundance between the two kinds of plants can be detected. This difference provides the basis for the natural¹⁵N abundance method of estimating the relative contribution of atmospheric N₂ to N₂-fixing plants growing in natural and agricultural settings. The natural¹⁵N abundance method has certain advantages over more conventional methods, particularly in natural ecosystems, since disturbance of the system is not required and the measurements may be made on samples dried in the field. This method has been tested mainly with legumes in agricultural settings. The tests have demonstrated the validity of this method of arriving at semi-quantitative estimates of biological N₂-fixation in these settings. More limited tests and applications have been made for legumes in natural ecosystems. An understanding of the limits and utility of this method in these systems is beginning to emerge. Examples of systematic measurements of differences in¹⁵N abundance between non-legume N₂-fixing systems and neighbouring non-fixing systems are more unusual. In principle, application of the method to estimate N₂-fixation by nodulated non-legumes, using the natural¹⁵N abundance method, is as feasible as estimating N₂-fixation by legumes. Most of the studies involving N₂-fixing non-legumes are with this type of system (e.g., Ceanothus, Chamabatia, Eleagnus, Alnus, Myrica, and so forth). Resuls of these studies are described. Applicability for associative N₂-fixation is an empirical question, the answer to which probably depends upon the degree to which fixed N goes predominantly to the plant rather than to the soil N pool. The natural¹⁵N abundance method is probably not well suited to assessing the contribution of N₂-fixation by free-living microorganisms in their natural habitat, particularly soil microorganisms.This work was supported in part by subcontracts under grants from the US National Science Foundation (DEB79-21971 and BSR821618)     

Effect of NO3 on N2 fixation and nitrogenous solutes of xylem in two nodulated West African geocarpic legumes,Kersting's bean (Macrotyloma geocarpum L.) and Bambara groundnut (Vigna subterranea L.)

Nodulation, nitrogen (N₂) fixation and xylem sap composition were examined in sand cultured plants of Bambara groundnut (Vigna subterranea L.) and Kersting's bean (Macrotyloma geocarpum L.) inoculated with Bradyrhizobium strain CB756 and supplied via the roots for a 4 week period from the third week onwards with different levels of (¹⁵N)-nitrate (0–15 mM). The separate contributions of nitrate and N₂ to plant nitrogen were measured by isotope dilution. Increasing levels of nitrate inhibited nodule growth (measured as dry matter or nodule N) of both species parallel with decreased dependence on symbiotically-fixed N. Specific nodule activity (N₂ fixed g nodule dry⁻¹ d⁻¹ of nodules) was reduced progressively with time in V. subterranea at higher (5 or 15 mM) levels of NO₃, but this was not so for M. geocarpum. Root xylem bleeding sap of both species showed ureides (allantoin and allantoic acid) as predominant (>90%) solutes of nitrogen when plants were relying solely on atmospheric N. Levels of ureide and glutamine decreased and those of asparagine and nitrate in xylem increased with increasing level of applied nitrate. Relative levels of xylem ureide-N were positively correlated (R²=0.842 for M. geocarpum and 0.556 for V. subterranea), and the ratio of asparagine to glutamine in xylem exudate negatively correlated (R²=0.955 for M. geocarpum and 0.736 for V. subterranea) with plant reliance on nitrogen fixation. The data indicate that xylem sap analyses might be useful for indirect field assays of nitrogen fixation by the species and that Kersting's bean might offer some potential as a symbiosis in which N₂ fixation is relatively tolerant of soil N.     

Light affects competition for inorganic and organic nitrogen between maize and rhizosphere microorganisms

Effects of light on the short term competition for organic and inorganic nitrogen between maize and rhizosphere microorganisms were investigated using a mixture of amino acid, ammonium and nitrate under controlled conditions. The amount and forms of N added in the three treatments was identical, but only one of the three N forms was labeled with ¹⁵N. Glycine was additionally labeled with ¹⁴C to prove its uptake by maize and incorporation into microbial biomass in an intact form. Maize out-competed microorganisms for during the whole experiment under low and high light intensity. Microbial uptake of ¹⁵N and ¹⁴C was not directly influenced by the light intensity, but was indirectly related to the impact the light intensity had on the plant. More was recovered in microbial biomass than in plants in the initial 4 h under the two light intensities, although more ¹⁵N-glycine was incorporated into microbial biomass than in plants in the initial 4 h under low light intensity. Light had a significant effect on uptake by maize, but no significant effects on the uptake of or ¹⁵N-glycine. High light intensity significantly increased plant uptake of and glycine ¹⁴C. Based on ¹⁴C to ¹⁵N recovery ratios of plants, intact glycine contributed at least 13% to glycine-derived nitrogen 4 h after tracer additions, but it contributed only 0.5% to total nitrogen uptake. These findings suggest that light intensity alters the competitive relationship between maize roots and rhizosphere microorganisms and that C4 cereals such as maize are able to access small amounts of intact glycine. We conclude that roots were stronger competitor than microorganisms for inorganic N, but microorganisms out competed plants during a short period for organic N, which was mineralized into inorganic N within a few hours of application to the soil and was thereafter available for root uptake.     

The nitrogen mineralization rate of legume residues in soil as influenced by their polyphenol,lignin, and nitrogen contents

A 12-week greenhouse experiment was conducted to determine the effect of the polyphenol, lignin and N contents of six legumes on their N mineralization rate in soil and to compare estimates of legume-N release by the difference and ¹⁵N-recovery methods. Mature tops of alfalfa (Medicago sativa L.), round leaf cassia (Cassia rotundifolia Pers., var. Wynn), leucaena (Leucaena leucocephala Lam., deWit), Fitzroy stylo (Stylosanthes scabra Vog., var Fitzroy), snail medic (Medicago scutellata L.), and vigna (Vigna trilobata L., var verde) were incorporated in soil at the rate of 100 mg legume N kg^-1 soil. The medic and vigna were labeled with ¹⁵N. Sorghum-sudan hybrid (Sorghum bicolor, L. Moench) was used as the test crop. A non-amended treatment was used as a control. Net N mineralization after 12 weeks ranged from 11% of added N with cassia to 47% of added N for alfalfa. With the two legumes that contained less than 20 g kg^-1 of N, stylo and cassia, there was net N immobilization for the first 6 weeks of the experiment. The legume (lignin + polyphenol):N ratio was significantly correlated with N mineralization at all sampling dates at the 0.05 level and at the 0.01 level at 6 weeks (r²=0.866). Legume N, lignin, or polyphenol concentrations or the lignin:N ratio were not significantly correlated with N mineralization at any time. The polyphenol:N ratio was only significantly correlated with N mineralization after 9 weeks (r²=0.692). The (lignin + polyphenol):N ratio appears to be a good predictor of N mineralization rates of incorporated legumes, but the method for analyzing plant polyphenol needs to be standardized. Estimates of legume-N mineralization by the difference and ¹⁵N recovery methods were significantly different at all sampling dates for both ¹⁵N-labeled legumes. After 12 weeks, estimates of legume-N mineralization averaged 20% more with the difference method than with the ¹⁵N recovery method. This finding suggests that estimates of legume N available to subsequent crops should not be based solely on results from ¹⁵N recovery experiments.     

Physiological changes in soybean (Glycine max) Wuyin9 in response to N and P nutrition

Phosphorus deficiency is a very common problem in the acid soil of central China. Previous research has shown that starter N and N topdressing at the flowering stage (Rl) increased soybean (Glycine max) yield and N₂ fixation (Gan et al, 1997, 2000). However, there is little information available concerning soybean response to P‐fertiliser in soybean production in central China (Gan, 1999). A field experiment was conducted to investigate the response to P (0 kg P ha^?1, 22 kg P ha^?1, 44 kg P ha^?1 before sowing) and N fertiliser application (N1: 0 kg N ha^?1, N2: 25 kg N ha^?1 before sowing, N3: N2 + 50 kg N ha^?1 at the V2 stage and N4: N2 + 50 kg N ha^?1 at the R1 stage) on growth, yield and N₂ fixation of soybean. Both N and P fertiliser increased growth and seed yield of soybean (P < 0.01). Application of basal P fertiliser at 22 kg P ha^?1 or 44 kg P ha^?1 increased total N accumulation by 11% and 10% (P < 0.01) and seed yield by 12% and 13% (P < 0.01), respectively, compared to the zero P treatment. Although application of starter N at 25 kg N ha^?1 had no positive effect on seed yield at any P level (P > 0.05), an application of a topdressing of 50 kg N ha^?1 at the V2 or R1 stage increased total N accumulation by 11% and 14% (P < 0.01) and seed yield by 16% and 21% (P < 0.01), respectively, compared to the zero N treatment. Soybean plants were grown on sterilised Perlite in the greenhouse experiment to study the physiological response to different concentrations of phosphate (P1: 0 mM; P2: 0.05 mM; P3: 0.5 mM; P4:1.0 mN) and nitrate (N1: 0 mM with inoculation, N2: 20 mM with inoculation). The result confirmed that N and P nutrients both had positive effects on growth, nodulation and yield (P < 0.01). The relative importance of growth parameters that contributed to the larger biomass with N and P fertilisation was in decreasing order: (i) total leaf area, (ii) individual leaf area, (iii) shoot/root ratio, (iv) leaf area ratio and (v) specific leaf area. The yield increase at N and P supply was mainly associated with more seeds and a larger pod number per plant, which confirmed the result from the field experiment.     

A field evaluation using the 15N isotope dilution method of lines of Phaseolus vulgaris L. bred for increased nitrogen fixation

N₂ fixation in lines of Phaseolus vulgaris was measured by ¹⁵N-isotope dilution to determine whether a programme of crossing and recurrent selection had resulted in enhanced nitrogen fixation. In field experiments on an isohyperthermic Aquic Hapludoll soil the amount of N₂ fixed by the different lines ranged from 18 to 36 kg ha^–1 (32 to 47% of plant N) in 56 days. The quantity of N₂ fixed and the proportion of plant N derived from fixation was not significantly greater in the lines selected for N₂ fixation (RIZ lines) than parental lines. Total shoot N ranged from 53 to 77 kg ha^–1 and partitioning of N to pods differed from 28 to 52% among the lines which all had similar growth habit and duration. Nodulation patterns were also distinct. Nodules formed early (10 to 15 plant^–1 at 13 days) in many lines, and smallest amounts of fixation were observed in those lines which nodulated slowly and did not form substantial nodule mass until after 40 days. The screening criteria used in the selection of the RIZ lines had been largely indirect with other factors such as disease resistance also being included. Progress for increasing N₂ fixation over good-fixing parental lines such as BAT76 was not significant and it is recommended that more attention be paid to early nodulation, to the use of soils with lower available N and to inter-crossing of lines having different good N₂ fixation traits in order to further enhance the potential for N₂ fixation in beans.     

Nitrogen fixation efficiency,natural15N abundance,and morphology of mesquite (Prosopis glandulosa) root nodules

Summary The relative nitrogen fixation efficiencies (RE 1-[H₂ evolved÷C₂H₂ reduced]·100) of four mesquite (Prosopis glandulosa var.torreyana) rhizobia (Strains WR 1001, WR 1002, L5, L9) and a cowpea rhizobia (Strain 176A32) on mesquite were evaluated in a glasshouse experiment. Plant yield, shoot N accumulation, and the natural¹⁵N abundance (¹⁵N) of nodule tissue were determined. Strain WR 1002 failed to nodulate mesquite and strain L5 produced ineffective nodules. Among the three effective strains (WR 1001, L9, 176A32) the cowpea strain (176A32) and strain L9 had significantly higher RE than strain WR 1001. Differences in RE, however, were not accompanied by significantly higher plant yield and shoot N accumulation. The difference in¹⁵N abundance between foliar tissue and nodules (nodules minus leaves) was 0.47 ¹⁵N for the ineffective L5 nodules, while for the effective WR 1001, L9, and 176A32 nodules, respectively, this difference was 8.35, 7.81, and 8.35 ¹⁵N. This indicates a similar relationship between N₂-fixing effectiveness and natural¹⁵N enrichment of nodules that was previously observed in soybeans (Glycine max, L. Merr.). Strains WR 1001 and L9 produced elongate, indeterminate nodules typical for mesquite. The ineffective L5 nodules had few infected cells and an abundance of cortical amyloplasts. Mesquite nodules produced by the cowpea strain were spherical and were somewhat more similar in internal morphology to determinate nodules typical of cowpea than indeterminate nodules normally associated with mesquite.