Relative Abundance Of Ureides Research Articles

Seed inoculation with Azospirillum brasilense in the U.S. soybean systems

Symbiotic nitrogen (N) fixation (SNF) is critical to satisfying the nutritional need of soybean (Glycine max (L.) Merr.) and maintaining productivity and high seed protein concentration. Due to its low environmental impact, a key factor for increasing the sustainability of soybean systems is to enhance SNF. Seed inoculation with the free-living Azospirillum brasilense alone or with Bradyrhizobium japonicum (herein called co-inoculation) are plausible strategies that have been explored in tropical environments but lack information in temperate regions. Following this rationale, this study aimed to evaluate the impact of seed inoculation with Azospirillum brasilense (herein called Azospirillum) alone or combined with Bradyrhizobium japonicum (herein called Bradyrhizobium) in a range of environments in the United States (US) for: (i) seed yield, (ii) relative abundance of ureides (RAU) as a proxy of SNF, and (iii) seed protein concentration. Twenty-five field studies across the US states with the same experimental design were performed during the 2019 and 2020 growing seasons. The primary outcomes of this research were: (i) yield responses to co-inoculation were considered significant in only 2 out of 25 site-years, (ii) RAU was not increased by Azospirillum inoculation or co-inoculation, and lastly, (iii) seed protein concentration was marginally associated with the inoculation strategies. Although Azospirillum did not impose remarkable gain in any observed plant traits, future studies should focus on mechanistically understanding whether Azospirillum can naturalize in temperate region soils. Still, strategies for enhancing SNF are required for sustainably improving productivity and quality for US soybean systems.

Soybean Seed Yield Productivity and Biological Nitrogen Fixation in Kansas

Soybean [Glycine max (L.) Merr.] productivity (seed yield) and biological nitrogen fixation (BNF) were evaluated in response to different fertilization strategies. The study comprised four different locations in Kansas during the 2021 growing season, two irrigated (Topeka and Scandia) and two dryland (Kiro and Ashland Bottoms) sites. Greater seed yields were recorded in Topeka and Kiro (80 bu/a) relative to Scandia (55 bu/a) and Ashland Bottoms (51 bu/a), without observing fertilizer effects on yields. Overall, the relative abundance of ureides (% RAU), an indicator of the level of BNF, increased as the crop matured and showed a negative association with the soil N level. The main objective of this study was to identify how different levels of nitrogen (N) and sulfur (S) fertilization affect the seed yield and the biological nitrogen fixation (BNF) in soybean.

Environmental Factors Associated With Nitrogen Fixation Prediction in Soybean.

Biological nitrogen (N)-fixation is the most important source of N for soybean [Glycine max (L.) Merr.], with considerable implications for sustainable intensification. Therefore, this study aimed to investigate the relevance of environmental factors driving N-fixation and to develop predictive models defining the role of N-fixation for improved productivity and increased seed protein concentration. Using the elastic net regularization of multiple linear regression, we analyzed 40 environmental factors related to weather, soil, and crop management. We selected the most important factors associated with the relative abundance of ureides (RAU) as an indicator of the fraction of N derived from N-fixation. The most relevant RAU predictors were N fertilization, atmospheric vapor pressure deficit (VPD) and precipitation during early reproductive growth (R1–R4 stages), sowing date, drought stress during seed filling (R5–R6), soil cation exchange capacity (CEC), and soil sulfate concentration before sowing. Soybean N-fixation ranged from 60 to 98% across locations and years (n = 95). The predictive model for RAU showed relative mean square error (RRMSE) of 4.5% and an R2 value of 0.69, estimated via cross-validation. In addition, we built similar predictive models of yield and seed protein to assess the association of RAU and these plant traits. The variable RAU was selected as a covariable for the models predicting yield and seed protein, but with a small magnitude relative to the sowing date for yield or soil sulfate for protein. The early-reproductive period VPD affected all independent variables, namely RAU, yield, and seed protein. The elastic net algorithm successfully depicted some otherwise challenging empirical relationships to assess with bivariate associations in observational data. This approach provides inference about environmental variables while predicting N-fixation. The outcomes of this study will provide a foundation for improving the understanding of N-fixation within the context of sustainable intensification of soybean production.

Relative abundance of ureides differs among plant fractions in soybean

The present study aims to compare the relative abundance of ureides (RAU) among soybean [Glycine max (L.) Merr.] plant fractions in contrasting trial settings and under three nitrogen (N) rates, designed to create variability on symbiotic N fixation. A greenhouse and a field experiment evaluated the RAU in plant fractions (base of the main stem, petiole and leaflets of the uppermost fully developed leaf) at varying growth stages and under absent, intermediate, or high N fertilization rates. The RAU was determined by measuring dry tissue nitrate and ureides and considered the entire main stem as benchmarking plant fraction, previously calibrated based on 15N isotope methods. The main outcomes of this study were: 1) RAU in the base of the stem accurately reflected the one obtained in the main stem; 2) upper leaflets RAU poorly correlate with the main stem values; and 3) the RAU from the upper petiole presented similar trends as the base of the stem but did not provide as reliable RAU estimation. The base of the stem is a reliable fraction to quantify soybean RAU while simplifying dry mass collection. The upper petiole fraction did not estimate RAU accurately, but it could differentiate contrasting N fixation levels.

Soybean yield, biological N2 fixation and seed composition responses to additional inoculation in the United States

It is unclear if additional inoculation with Bradyrhizobia at varying soybean [Glycine max (L.) Merr.] growth stages can impact biological nitrogen fixation (BNF), increase yield and improve seed composition [protein, oil, and amino acid (AA) concentrations]. The objectives of this study were to evaluate the effect of different soybean inoculation strategies (seed coating and additional soil inoculation at V4 or R1) on: (i) seed yield, (ii) seed composition, and (iii) BNF traits [nodule number and relative abundance of ureides (RAU)]. Soybean field trials were conducted in 11 environments (four states of the US) to evaluate four treatments: (i) control without inoculation, (ii) seed inoculation, (iii) seed inoculation + soil inoculation at V4, and (iv) seed inoculation + soil inoculation at R1. Results demonstrated no effect of seed or additional soil inoculation at V4 or R1 on either soybean seed yield or composition. Also, inoculation strategies produced similar values to the non-inoculated control in terms of nodule number and RAU, a reflection of BNF. Therefore, we conclude that in soils with previous history of soybean and under non-severe stress conditions (e.g. high early-season temperature and/or saturated soils), there is no benefit to implementing additional inoculation on soybean yield and seed composition.

Abstracts of Nippon Dojo-Hiryogaku Zasshi

Abstracts of Nippon Dojo-Hiryogaku Zasshi

Soybean N2 Fixation Estimates, Ureide Concentration, and Yield Responses to Drought

Increasing N2 fixation tolerance to drought has been hindered by the labor and costs of quantifying N2 fixation using 15N methodologies. The relative abundance of ureides (RAU) in plant tissues has been used for estimating N2 fixation in soybean [Glycine max (L.) Merr.] grown under well‐watered conditions, but it has not been evaluated for drought conditions. The present research evaluated the response of N accumulation to N fertilization, the ability of the RAU technique to predict N2 fixation under drought conditions, and the response of yield to N fertilization under well‐watered and drought conditions. Under drought, shoot N accumulation rate during vegetative growth approximately doubled as the amount of N fertilizer was increased from 10 to 200 kg N ha−1, indicating a greater sensitivity of N2 fixation to drought than uptake and assimilation of inorganic N. Under well‐watered conditions, the relationship between estimates of N2 fixation made by 15N‐dilution and RAU agreed within 15% of published reports. Under drought conditions, however, this relationship was greatly different (13 to 43%) from published reports. Fertilization with inorganic N in 1 yr increased grain yield 15 to 25% for the drought treatment and 12 to 15% for the well‐watered treatment. In a second year, N fertilization increased yield of both drought and well‐watered treatments approximately 9%. This research indicates that the RAU technique for estimating N2 fixation under drought conditions may be invalid without further refinement, that N2 fixation is more sensitive to drought than the uptake and assimilation of inorganic soil N, and that increasing the tolerance of N2 fixation to drought would likely result in yield increases.

Effects of arbuscular mycorrhizal inoculation on growth, yield, nitrogen, and phosphorus nutrition of nodulating bean varieties in two soil substrates of contrasting fertility

A pot experiment was conducted to study the response of nodulated bean (Phaseolus vulgaris L.), vars. Ceca and Montalbán, to inoculation with the arbuscular‐mycorrhizae (AM) Glomus spurcum strain IES‐3 and G. mosseae strain IES‐8 in two non‐sterilized non‐amended soil substrates (S‐l and S‐2) of contrasting fertility. S‐l was an acid soil (pH 5.0) with 77% exchangeable aluminum (Al), low available phosphorus (P) (3.5 ug g‐1 dry soil) and 1.8% organic matter. S‐2 was a soil with pH 7.0, 7.9 μg P g‐1 dry soil, 9.6% organic matter and no exchangeable Al. Plants growing in S‐1 (S‐1 plants) and S‐2 (S‐2 plants) soils were sampled at the beginning of the flowering stage. S‐1 and S‐2 plants were nodulated by the rhizobial populations native of each soil. S‐1 plants of both varieties did not respond to AM inoculation in term of shoot and root mass, P content, relative abundance of ureides (RAU) and seed yields. Only the total chlorophyll, chlorophyll‐a, chlrophyll‐a/b ratio and nodule mass were significantly enhanced in AM‐inoculated plants. Increased total reducing sugars concentration was detected in roots of S‐1 plants inoculated with G. mosseae. S‐2 plants displayed significantly higher shoot and nodule mass as well as increased total chlorophyll, chlorophyll‐a, chlorophyll‐a/b ratio and P content than S‐1 plants, regardless of the variety and AM inoculation. S‐2 plants of the var Ceca showed a two fold increase in seed yields but similar RAU values (>60%) when compared to S‐1 plants of the same variety. In contrast, the significant increase in RAU detected in S‐2 plants of the var Montalban was not translated into higher seed yields. In S‐2, the productivity of plants of the var Ceca doubled that of the var Montalban. For both bean varieties the highest significant P content and seed yield were observed exclusively in S‐2 plants inoculated with G. mosseae. This Glomelean strain enhanced the sink‐source ratio of the S‐2 plants as evidenced by the higher total reducing sugar concentration in the root mass. Arbuscular‐mycorrhizae inoculation significantly decreased the acid phosphatase activity in the rhizosphere of S‐1 and S‐2 plants, respectively, pointing toward a negative effect of foreign AM on the native microbial biomass. The effectiveness of the rhizobial populations native of each soil type and the weak response elicited by G. mosseae in S‐2 plants do not justify, at present, the inclusion of foreign inocula in the bean crops carried out at S‐1 and S‐2 soils of the Sucre State of Venezuela. Results also indicated the higher adaptability of var Ceca to conditions prevailing in S‐1 and S‐2.

Xylem-solute technique to measure N2 fixation by Phaseolus vulgaris L.: Calibration and sources of error

Utilising the 15N dilution technique the relationship between the proportion of N derived from N2 fixation and relative abundance of ureides in xylem sap was evaluated for Phaseolus vulgaris L. cv. Mokcham during vegetative and reproductive development. In order to establish calibration curves for time integrated estimates of N2 fixation, plants were raised in sand culture during the dry season in northern Thailand and continuously supplied with a N-free nutrient solution or the same solution amended with 0, 3, 6 or 9 mol m−3 nitrate. Large changes in plant dependence on N2 fixation were concomitantly reflected by corresponding alterations in N solutes in xylem sap. Regression analyses of the data suggested high correlations between relative ureide content and N2 fixation, but different slopes and line intercepts indicated the requirement for the use of calibration curves established for different phases of the development of the plant. Largest age related differences were noted between vegetative and reproductive development. Judging from 95% confidence limits, utilisation of appropriate calibrations can reduce errors of the technique to close to ±5%.

Physiological and Biochemical Aspects of Symbiotic Nitrogen Fixation in Cowpea (Vigna unguiculata(L.) Walp. var. Tuy) Plants Infected by Cowpea Mosaic Virus

Three-day-old cowpea seedlings were inoculated with the severe strain of Cowpea Mosaic Virus (CpMV) and 24 h later with Bradyrhizobium sp. cowpea, strain 1-125, when virus translocation to roots still had not taken place. Plants were harvested at 22, 30, 45 and 59 d after germination. Active virus replication was associated with increased protein content, detected in the leaves of 22-d-old plants. CpMV infection reduced the total leaf area, dry weight of shoots and the chlorophyll content of the first trifoliolate leaf at all experimental times. Low sugar content was recorded in leaves of 22- and 30-d-old CpMV-infected plants and in nodules and roots of 30- and 45-d-old CpMV-infected plants. Up to 45 d, the nodule mass of CpMV-infected plants was lower than in controls, but reached control values at the 4th harvest. In CpMV-infected nodules, massive agglomeration of virus particles, crystalline virus inclusions and the proliferation of the endoplasmic reticulum were observed only in those cells containing bacteroids. In 30- and 45-d-old plants, CpMV infection decreased the content of ureides in nodules, roots, and petioles. Virus infection did not alter the a-amino-N content of roots and nodules but induced a transient 74% reduction in the level of a-amino-N in petioles of 45-d-old plants. At the 1st and 2nd harvests the activity of uricase (EC 1.7.3.3) in the nodules and of pyruvate kinase (EC 2.7.1.40) in the nodules and leaves were decreased severely by virus infection. CpMV did not hinder the allantoinase (EC 3.5.2.5) activity in the leaves but caused a 9% transitory decrease in the activity of this enzyme in nodules of 45-d-old plants. Measurements of NAD-malic enzyme (EC 1.1.1.38) in nodules also showed the non-effect of CpMV on this enzyme, except for a temporary 16% reduction at the 2nd harvest. As compared to controls, the relative abundance of ureides in 30-d-old CpMV-infected plants indicated a 15%, 10%, and 51% reduction in the nodules, roots, and petioles, respectively. Results indicate that at the time of the 4th harvest the symbiotic process, measured in terms of ureide content and enzymatic activities, was functioning at a near normal level despite nodule infection by CpMV.

Nitrogen Fixation of Nodulation Mutants of Soybean as Affected by Nitrate

It was previously reported that three soybean (Glycine max [L.] Merr.) nodulation mutants (NOD1-3, NOD2-4, and NOD3-7) were partially tolerant to nitrate when nitrate was supplied simultaneously with inoculation at the time of transplanting. The current study evaluated the effect of short-term nitrate treatment on nitrogenase activity (C(2)H(2) reduction per plant and per nodule weight) and on relative abundance of ureides when nitrate application was delayed until plants were 3 weeks old and nodules were fully developed. Nitrogenase activity of the mutants was similar to that of Williams after an initial 3-week growth period, prior to nitrate treatment. Application of 5 millimolar nitrate resulted in greater inhibition of nitrogenase activity in Williams than in the three mutants. NOD1-3 was most tolerant of nitrate among the mutants tested and showed the highest relative abundance of ureides. Although C(2)H(2) reduction activity per plant for NOD1-3 was higher than for Williams in the presence of nitrate, C(2)H(2) reduction activity per gram of nodules was lower for NOD1-3 than for Williams in the presence and absence of nitrate. Compared to Williams, NOD1-3 had higher nodule ureide concentration and had similar glutamine synthetase activity in nodule tissue, indicating its nodules have normal nitrogen assimilation pathways. Nitrate application resulted in ureide accumulation in nodule tissue as well as in all plant parts assayed. Unexpectedly, nitrate treatment also increased the rate of ureide degradative capacity of leaves in both NOD1-3 and Williams. The data confirmed that nitrogenase activity of the selected nodulation mutants was more, but still only partially, tolerant of nitrate compared with the Williams parent.

Development of the Xylem Ureide Assay for the Measurement of Nitrogen Fixation by Pigeonpea (Cajanus cajan(L.) Millsp.)

Quantification of N2 fixation by pigeonpea (Cajanus cajan (L.) Millsp.) in the field has proved difficult using techniques such as 15N isotope dilution, acetylene reduction and N difference. We report experiments to develop the ureide assay of N2 fixation based on extraction and analysis of xylem exudate. Plants of pigeonpea cv. Quantum, inoculated with effective Rhizobium spp. CB756, were grown in a temperature-controlled glasshouse in large pots filled with a sand: vermiculite mixture, in water culture and in a slightly acidic, red-brown earth in replicated field plots. Xylem exudate was collected as bleeding sap from both nodulated and unnodulated roots, and from detached nodules. Exudate was extracted also from detached shoots and stems of pigeonpea using a mild vacuum (60–70 kN m−2). Large differences in the composition of N solutes exported from roots of N2-dependent and nitrate-dependent plants suggested that shifts in plant dependence on N2 fixation may be reflected by concomitant changes in N solutes. Thus, nodulated plants were supplied throughout growth with either N-free nutrients or nutrients supplemented with 1, 2, 5, 5, 10, or 20 mol m−315NO3-⁠. Plants were harvested at regular intervals for dry matter and vacuum-extracted exudate. The relative abundance of ureides ([ureide-N/ureide-N + nitrate-N + α-amino-N] × 100) in the exudate was highly correlated with the proportion of plant N (calculated using a 15N isotope dilution technique) derived from N2 fixation. Two distinct phases of plant growth were recognized and standard curves were prepared for each. The relationship between proportional dependence of plants on N2 and xylem relative ureides was unaffected by mineral-N source, i.e. nitrate or ammonium. This result is discussed in relation to interpretation of material from field-grown plants. The effects of plant genotype, strain of rhizobia, section of stem extracted, removal of leaves, time delay between shoot detachment and extraction, and diurnal characteristics were examined in order to identify potential sources of error and to optimize sampling procedures.

Field evaluation of soybean genotypes selected for enhanced capacity to nodulate and fix nitrogen in the presence of nitrate

The development of cultivars of soybean (Glycine max L. Merr.) which are capable of near-maximum levels of N2 fixation in high-NO 3 − soils remains a high priority in N2 fixation research. We report a field study to evaluate nodulation and N2 fixation by 32 genotypes of soybean, selected after two years of glasshouse screening for superior symbiotic activity in the presence of 2.5 mM NO 3 − . The 32 “NO 3 − -tolerant” genotypes and eight others (three commercial “check” cultivars and five “non-fixing” lines) were inoculated withBradyrhizobium japonicum CB 1809 and sown into a black earth soil (fine, montmorillonitic, thermic Udic Pellusterts) which contained high levels of soil NO 3 − (260 kg N ha−1; 0 to 120 cm depth) and which was free of soybean rhizobia. Nitrogen fixation activity was assessed at 89 days after sowing using the relative abundance of ureides in xylem exudate [(ureide-N/ureide-N+NO 3 − -N+α amino-N)×100] as an index of fixation. Plant growth and nodulation were assessed 11 days later. Genotypes 466, 468, 469 and 464, all of Korean origin, showed the highest levels of symbiotic activity. Many of the remaining 28 “tolerant” genotypes nodulated poorly in the field and displayed levels of N2 fixation (relative ureides) which were equivalent to two of the commercial “check” cultivars, Bragg and Elf. Correlation matrices of the measured parameters revealed highly significant correlations among the indices of nodulation and N2 fixation and poor correlation between those measurements and plant growth-seed yield. The levels of NO 3 − tolerance, displayed by the four Korean lines, may prove useful in breeding programs which aim to enhance N2 fixation by soybean in high-NO 3 − soils.

Ureide production by N 2-fixing and non-N 2-fixing leguminous trees

Xylem-sap and stem and leaf extracts from 35 species, comprising 14 genera, of leguminous trees were analyzed for ureides, nitrate and α-amino acids. Trees were either inoculated with Rhizobium or fertilized with NH 4NO 3. The dominant form of soluble N in stem and leaf extracts and xylem sap was 2-amino acids. Certain non-N 2-fixing species, i.e. Tamarindus indica and Adenantherapavonina, produced significant amounts of ureides. Several N 2-fixing species. Mimosa scabrella, Sesbania grandiflora. Acacia mearnsii and Gliricida sepium, grown on mineral-N had higher absolute amounts of ureides in both extracts and exudates than did most nodulated species. Nodulated A. meamsii and S. grandiflora, had the highest amounts of ureides in xylem sap. The relative abundance of ureides in stem and leaf extracts was lower than in xylem sap, but was correlated. Results indicated that the presence of ureides, per se, was not a reliable indicator of N 2-fixing activity. Moreover, the relative abundance of ureides in most of the species tested was too low to use as a presumptive test for, or as a means of, estimating N 2 fixation.

The Xylem Ureide Assay of Nitrogen Fixation: Sampling Procedures and Sources of Error

Herridge, D. F., O'Connell, P. and Donnelly, K. 1988. The xylem ureide assay of nitrogen fixation: sampling procedures and sources of error.—J. exp. Bot. 39: 12-22. The relative abundance of ureides [(ureide-N/ureide-N + NOj-N + a amino-N)x 100] in vacuum extracted xylem exudate of soyabean (Glycine max [L.] Merr.) can be used to quantify N2 fixation activity. We report experiments designed to (1) develop appropriate procedures for sampling and storage of xylem exudate extracted from plants in the field and (2) develop a single plant, non-destructive sampling procedure, useful for evaluation of segregating breeding material. Soyabean cultivars Bragg, Essex, Davis, Hardee and Dodds were grown at three sites at Breeza, New South

Contributions of fixed nitrogen and soil nitrate to the nitrogen economy of irrigated soybean

Effects of soil nitrate and numbers of Bradyrhizobium japonicum on the development and functioning of a soybean symbiosis and on crop production were studied in a field experiment at Breeza, New South Wales, Bragg soybean was grown with irrigation on soil, initially free of B. japonicum, with four rates of fertilizer-N (0, 100, 200, 300 kg N ha −1 as ammonium nitrate applied 6 weeks before sowing to provide four concentrations of soil nitrate) and four rates of inoculation [nil, normal (n). 100n, 1000n]. The inoculant strain was B. japonicum CB1809. Observations were made on nodulation, the relative abundance of ureidcs in xylem exudates as an index of N 2 fixation, dry matter and seed yield, and total nitrogen in shoots and seed. Results showed clearly that soil nitrate repressed nodulation, that the effect was magnified as soil nitrate concentrations increased, but that inhibition was substantially ameliorated by increased numbers of rhizobia. The relative abundance of ureides in xylem exudates responded similiarly. The highest yields of dry matter and of N in shoots and in seed occurred at the highest rates of inoculation (100n, 1000n) at intermediate and high soil nitrate (N100, N200, N300); at low soil nitrate (N0), yields were increased by inoculation per se but not by the rate used. Uninoculated plants did not nodulate and yields in these plots reflected concentrations of soil nitrate. Data suggested that soil nitrate and N 2 fixation were not always complementary in meeting the N requirements of the growing crop. Absence of rhizobia. except at the highest rate of nitrate, and repression of nodulation at the normal rate of inoculation by intermediate concentrations of nitrate resulted in reduced N yields because of insufficient N supply to the crop during the final stages of growth.

Isolation of Soybean Lines Capable of Nodulation and Nitrogen Fixation under High Levels of Nitrate Supply1

Results from numerous studies indicate that the proportion of soybean [Glycine max (L.) Men.] crop N derived from N2 fixation ranges between 25 and 50% and rarely exceeds 75%. The remainder is taken up as NO−3 from the soil and the net effect of soybean cropping is depletion of the soil N pool. Therefore, a program was commenced to identify genotypes of soybean which were capable of high levels of nodulation and high rates of N2 fixation in the presence of NO−3. In the first year of screening, 489 genotypes of diverse origin were grown in sand, free of Bradyrhizobium japonicum prior to inoculation with effective strain CB 1809 (USDA136), in unreplicated, 14‐L free‐draining pots in a glasshouse and supplied with nutrients containing either zero or 2.5 mMNO−3. Plants were sampled at R2 for plant growth (shoot mass), for nodulation, and for the proportion of plant N derived from N2 fixation (relative abundance of ureides in xylem exudate and extracts of shoot axes and nodulated roots). From the original 489 genotypes, 66 “NO−3‐tolerant”, 9 “NO−3‐sensitive” and 12 others were selected for a second year of screening on the basis of an overall ranking index which related nodulation and N2 fixation (relative ureides) activity at 2.5 mMNO−3 to activity at zero NO−3. In year two, treatments were replicated twice; plant culture and sampling procedures were otherwise identical to those used in the first year. Variation in symbiotic activity at high NO−3 supply was again evident with between 5‐ and 13‐fold differences in the nodulation and ureide indices recorded. In both screenings, genotypes of Korean origin ranked first overall and provided three of the top 10 lines. Of the original 19 Korean genotypes, 15 (80%) were screened in year 2 and nine (47%) were identified for future evaluation. Only 5% of the other 470 genotypes were selected as “NO−3‐tolerant” after two years of screening. Isolation of genotypes, superior in nodulation and N2 fixation in the presence of NO−3 under both glasshouse and field conditions, should be achieved with further screening in a high‐NO−3 field soil.

Enzymes of Ammonia Assimilation and Ureide Biosynthesis in Soybean Nodules: Effect of Nitrate

The effect of nitrate on N(2) fixation and the assimilation of fixed N(2) in legume nodules was investigated by supplying nitrate to well established soybean (Glycine max L. Merr. cv Bragg)-Rhizobium japonicum (strain 3I1b110) symbioses. Three different techniques, acetylene reduction, (15)N(2) fixation and relative abundance of ureides ([ureides/(ureides + nitrate + alpha-amino nitrogen)] x 100) in xylem exudate, gave similar results for the effect of nitrate on N(2) fixation by nodulated roots. After 2 days of treatment with 10 millimolar nitrate, acetylene reduction by nodulated roots was inhibited by 48% but there was no effect on either acetylene reduction by isolated bacteroids or in vitro activity of nodule cytoplasmic glutamine synthetase, glutamine oxoglutarate aminotransferase, xanthine dehydrogenase, uricase, or allantoinase. After 7 days, acetylene reduction by isolated bacteroids was almost completely inhibited but, except for glutamine oxoglutarate aminotransferase, there was still no effect on the nodule cytoplasmic enzymes. It was concluded that, when nitrate is supplied to an established symbiosis, inhibition of nodulated root N(2) fixation precedes the loss of the potential of bacteroids to fix N(2). This in turn precedes the loss of the potential of nodules to assimilate fixed N(2).

N2 Fixation (C2H2) and Ureide Content of Soybeans: Ureides as an Index of Fixation1

The ureide contents of soybeans [Glycine max (L.) Merr.] were measured to determine if ureide concentration is a useful indicator of N2 fixation. Acetylene reduction activity (ARA) excised roots and the concentrations of ureides in stem segments were measured in field grown soybeans at weekly intervals. Ureides occurred in every soybean cultivar tested. Their concentration in the shoot varied over the growing season. The concentration droppeduring early vegetative growth, then rose and fell during reproductive growth. The plant age when maximum ureide concentration occurred was similar in three seasons. Within maturity groups there was little difference in ureide concentration. Stem nitrate levels decreased continuously over the season and were negatively correlated with ARA and ureides. A relative abundance of ureides was calculated as the fraction of ureide‐N in the ureide‐N + NO3‐N pool. The relative abundance correlated well with ARA, and gave a correction for the effect of soil nitrate on ureide concentration. Ureide concentration and ARA, measured at the same time, were significantly correlated. Dinitrogen fixation over the season was estimated by integrating ARA. The concentration of ureides in the young stem at the R3 stage showed the highest significant correlation with integrated seasonal ARA. The single ureide sampling was not as closely correlated with the estimated seasonal fixation as a single ARA determination at the R2 or R3 stage.

Relative Abundance of Ureides and Nitrate in Plant Tissues of Soybean as a Quantitative Assay of Nitrogen Fixation

The relationship between the relative abundance of ureides ([ureide-N/ureide-N plus nitrate-N] x 100) in the shoot axis (stems plus petioles), nodulated roots and leaflets of "Bragg" soybean (Glycine max [L.] Merrill) and the symbiotic dependence of these plants was examined under glass-house conditions. Plants, inoculated with effective Rhizobium japonicum CB1809, were grown with their roots exposed continuously to a nutrient solution containing either 0, 1.5, 3.0, 6.0 or 12.0 millimolar NO(3)-N per liter. Nodulation and N(2)-acetylene fixation were correlated inversely with the level of nitrate. Seasonal acetylene reduction profiles for each of the nitrate treatments were integrated and the symbiotic dependence ([N(2) fixed per total plant N] x 100) determined using a conversion ratio of 1.5:1 (acetylene reduced:N(2) fixed), calculated from the zero NO(3) treatment. Examination of the nitrogenous solutes of the shoot axis and nodulated roots showed linear relationships between the relative abundance of ureides and the symbiotic dependence of the plants. Two standard curves, depicting these relationships during vegetative and reproductive growth, were drawn for each plant part. The overriding effect of plant age invalidated any attempt to develop a standard relationship for leaflets. Data from two diurnal studies suggested that relative ureides were insensitive to diurnal fluctuations, thus simplifying sampling procedures. Plant material could be stored at ambient temperatures (20-30 degrees C) for up to 24 h without affecting the relative concentration of ureides and nitrate. It is suggested that the shoot axis provides the most suitable target organ when using this technique as a quantitative assay for N(2) fixation because of ease of sampling of these tissues, especially with field-grown plants.