Arbuscular mycorrhizae affect the N and C economy of nodulated Phaseolus vulgaris (L.) during NH4 nutrition

In the symbiosis between nodulated legume roots and arbuscular mycorrhizal (AM) fungi, the C and N economy can be influenced by the source of N-supply from either AM-derived NH₄⁺ uptake or nodule-derived biological nitrogen fixation (BNF). This relationship was investigated in terms of NH₄⁺ supply and BNF by the two symbionts. Nodulated Phaseolus vulgaris seedlings with and without AM, were hydroponically grown with either 0 N or 1 mM NH₄⁺ supply. Plants were harvested at 30 days after emergence and measurements were taken for biomass, N₂ fixation, photosynthesis, CO₂ and O₂ root respiration, calculated C and N economy. AM roots had higher NH₄⁺ uptake and this was associated with the suppression of BNF and nodule growth. The higher NH₄⁺ uptake in AM roots occurred with lower root maintenance respiration, compared to when N was derived from BNF. There was also an increase in the below-ground sink strength of NH₄⁺ fed AM roots compared to NH₄⁺ fed non-AM roots, as evidenced by the increases in root CO₂ and O₂ respiration and photosynthetic stimulation. These results indicate that although the AM root had higher total below-ground respiratory costs during NH₄⁺ nutrition, there were lower respiratory C costs associated with N derived from AM symbionts in comparison to N from BNF.     

Transfer of N fixed by a legume tree to the associated grass in a tropical silvopastoral system

Below-ground transfer of nitrogen (N) fixed by legume trees to associated non-N₂-fixing crops has received little attention in agroforestry, although the importance of below-ground interactions is shown in other ecosystems. We used ¹⁵N natural abundance to estimate N transfer from the legume tree Gliricidia sepium (Jacq.) Kunth ex Walp. to C₄ grass Dichanthium aristatum (Poir.) C.E. Hubb. in a silvopastoral system, where N was recycled exclusively by below-ground processes and N₂ fixation by G. sepium was the sole N input to the system. Finding a suitable reference plant, a grass without contact with tree roots or litter, was problematic because tree roots invaded adjacent grass monocrop plots and soil isotopic signature in soil below distant grass monocrops differed significantly from the agroforestry plots. Thus, we used grass cultivated under greenhouse conditions in pots filled with agroforestry soil as the reference. A model of soil ¹⁵N fractionation during N mineralization was developed for testing the reliability of that estimate. Experimental and theoretical results indicated that 9 months after greenhouse transplanting, the percentage of fixed N in the grass decreased from 35% to <1%, due to N export in cut grass and dilution of fixed N with N taken up from the soil. The effect of soil ¹⁵N fractionation on the estimate of the reference value was negligible. This indicates that potted grass is a suitable reference N transfer studies using ¹⁵N natural abundance. About one third of N in field-grown grass was of atmospheric origin in agroforestry plots and in adjacent D. aristatum grassland invaded by G. sepium roots. The concentration of fixed N was correlated with fine root density of G. sepium but not with soil isotopic signature. This suggests a direct N transfer from trees to grass, e.g. via root exudates or common mycorrhizal networks.     

N natural abundance of biologically fixed N2 in soybean is controlled more by the Bradyrhizobium strain than by the variety of the host plant

The ¹⁵N natural abundance technique is one of those most easily applied ‘on farm’ to evaluate the contribution of biological N₂ fixation (BNF) to legume crops. When proportional BNF inputs are high, the accuracy of this technique is highly dependent on an accurate estimate of the ¹⁵N abundance of the N derived from N₂ fixation (the ‘B’ value). The objective of this study was to determine the influence of soybean variety on ‘B’ value. Plants of five soybean varieties were inoculated separately with two Bradyrhizobium strains (one Bradyrhizobium japonicum and one Bradyrhizobium elkanii) grown in pots of soil virtually free of bradyrhizobia capable of nodulating soybean. The proportion of N derived from BNF (%Ndfa) was estimated in separate pots where a small quantity of enriched ¹⁵N ammonium sulphate was added. The %Ndfa was then used with the ¹⁵N natural abundance data of the nodulated soybean and non-N₂-fixing reference plants, to determine the ‘B’ value for each soybean variety/Bradyrhizobium association. The varieties nodulated by the B. japonicum strain showed significantly greater N content and %Ndfa than those nodulated by the B. elkanii strain, and in all cases the ‘B’ value of the shoot tissue (‘B_s’) was higher. The differences in ‘B_s’ values between varieties nodulated by the same Bradyrhizobium strain were insignificant, indicating that this parameter is influenced much more by the Bradyrhizobium strain than by the variety of the host plant.     

Nitrate removal from drained and reflooded fen soils affected by soil N transformation processes and plant uptake

Knowledge about nitrate transformation processes and how they are affected by different plants is essential in order to reduce the loss of valuable N fertiliser as well as to prevent environmental pollution due to nitrate leaching or N₂O emission after fertilisation or the reflooding of degraded fens with nitrate-containing municipal sewage. Therefore four microcosm ¹⁵N tracer experiments were performed to evaluate the effect of common wetland plants (Phalaris arundinacea, Phragmites australis) combined with different soil moisture conditions (from dry to reflooded) on nitrate turnover processes. At the end of experiment, the total formation of gaseous N compounds was calculated using the ¹⁵N balance method. In two experiments (wet and reflooded soil conditions) the N₂O and N₂ emissions were also directly determined.Our results show that in degraded fen soils, which process mainly takes place—denitrification or transformation into organic N compounds—is determined by the soil moisture conditions. Under dry soil moisture conditions (water filled pore space: 31%) up to 80% of the ¹⁵N nitrate added was transformed into organic N compounds. This transformation process is not affected by plant growth. Under reflooded conditions (water filled pore space: 100%), the total gaseous N losses were highest (77-95% of the ¹⁵N-nitrate added) and the transformation into organic N compounds was very low (1.8% of ¹⁵N nitrate added). Under almost all soil conditions plant growth reduced the N losses by 20-25% of the ¹⁵N nitrate added due to plant uptake. The N₂ emissions exceeded the N₂O emissions by a factor of 10-20 in planted soil, and as much as 30 in unplanted soil. In the treatments planted with Phragmites australis, N₂O emission was about two times higher than in the corresponding unplanted treatment. 15% of the N₂O and N₂ formed was transported via the Phragmites shoots from the soil into the atmosphere. By contrast, Phalaris arundinacea did not affect N₂O emissions and no emission via the shoots was observed.     

N supply by groundnuts to maize in a maize plus groundnut intercropping system,as affected by the genotype

Summary Identification of legume genotypes with high N-supplying ability is important in improving and sustaining the productivity of low-input cropping systems. Hence, ¹⁵N-aided studies were made to ascertain the relative N-supplying ability of some cultivars of groundnut, a widely grown tropical legume. The study was conducted outdoors in 1991 at Kamburupitiya, Sri Lanka, in tanks filled with 64 kg soil which had been tagged by incorporating ¹⁵N-labelled plant material. Maize cv. Badra was grown as a monocrop and as an intercrop with five genotypes of groundnut, X-14, MI-1, Red Spanish, ICGV 87127, and a non-nodulating line. All the nodulating genotypes derived over 90% of their N from the atmosphere. Significant genotypic differences in N₂ fixation were observed. X-14 fixed the highest amount (1.95 g plant^-1), while Red Spanish the lowest (0.88 g plant^-1). Intercropping of maize with nodulating groundnut significantly decreased the ¹⁵N atom excess of maize, depending on the genotype. However, this decrease did not appear to be related to the amount of N₂ fixed, based on aboveground material. The per cent N derived by maize from the intercropped groundnuts varied from 17% (X-14) to 39% (Red Spanish), indicating a marked genotypic variability in N-suppling ability. X-14, which fixed the largest amounts of N₂, grew most vigorously compared to other genotypes, causing a growth depression in the maize. The genotype that fixes the most N₂ may therefore not necessarily have the greatest N-supplying ability. The transfer of N from the legume and the consequent improvement of N nutrition in the associated cereal in low-fertility situations is therefore expected to be high when the growth of the legume is intermediate and does not suppress the growth of the cereal.     

Isotopic fractionation during N2 fixation by four tropical legumes

To quantify the contribution of biological nitrogen fixation (BNF) to legume crops using the ¹⁵N natural abundance technique, it is necessary to determine the ¹⁵N abundance of the N derived from BNF—the B value. In this study, we used a technique to determine B whereby both legume and non-N₂-fixing reference plants were grown under the same conditions in two similar soils, one artificially labelled with ¹⁵N, and the other not. The proportion of N derived from BNF (%Ndfa) was determined from the plants grown in the ¹⁵N-labelled soil and it was assumed that the %Ndfa values of the legumes grown in the two soils were the same, hence the B value of the legumes could be calculated. The legumes used were velvet bean (Mucuna pruriens), sunnhemp (Crotalaria juncea), groundnut (Arachis hypogaea) and soybean (Glycine max) inoculated, or not, with different strains of rhizobium. The values of %Ndfa were all over 89%, and all the legumes grown in unlabelled soil showed negative δ¹⁵N values even though the plant-available N in this soil was found to be approximately +6.0‰. The B values for the shoot tissue (B_s) were calculated and ranged from approximately −1.4‰ for inoculated sunnhemp and groundnut to −2.4 and −4.5‰ for soybean inoculated with Bradyrhizobium japonicum strain CPAC 7 and Bradyrhizobium elkanii strain 29W, respectively. The B (B_wp) values for the whole plants including roots, nodules and the original seed N were still significantly different between the soybean plants inoculated with CPAC 7 (−1.33‰) and 29W (−2.25‰). In a parallel experiment conducted in monoxenic culture using the same soybean variety and Bradyrhizobium strains, the plants accumulated less N from BNF and the values were less negative, but still significantly different for soybean inoculated with the two different Bradyrhizobium strains. The results suggest that the technique utilized in this study to determine B with legume plants grown in soil in the open air, yields B values that are more appropriate for use under field conditions.     

Assessment of the relative uptake of added and indigenous soil nitrogen by nodulated legumes and reference plants in the 15N dilution measurement of N2 fixation: Glasshouse application of method

A method for calculating the relative uptake (R) of added N and indigenous soil N by a legume (Trifolium subterraneum) and non-legume (Lolium rigidum), growing together, was investigated in two pot experiments. In the first experiment, ¹⁵N-labelled sodium nitrate was applied to the soil surface at rates equivalent to 0.3 or 1.0kg N ha^?1. Twenty one days later, the legume had fixed about 95% of its total N and this was unaffected by N addition. There was no difference in R values between legume and non-legume at both N rates.In the second experiment using a soil of higher total N, sodium nitrate or ammonium sulphate were surface-applied at a rate equivalent to 1 kg N ha^?1 and harvests were made at 3, 6, 12 and 27 days after N addition. Fixation of atmospheric N₂ by the legume did not begin until day 12 but accounted for about 40% of the total N assimilated by the legume by day 27. There was no difference in R values between legume and non-legume throughout the growth period when sodium nitrate was applied. However, when ammonium sulphate was added to label to soil N, the uptake of added N relative to indigenous soil N was greater for the non-legume than the legume. This caused an overestimation (51 vs 43%) of the proportion on N fixed by the legume when compared with that for the control or sodium nitrate treatments.     

Growth promotion of plants by plant growth-promoting rhizobacteria under greenhouse and two different field soil conditions

This study was conducted with sugar beet in greenhouse and field at two soil type with different organic matter (containing 2.4 and 15.9% OM, referred as the low- and high-OM soil) conditions in order to investigate seed inoculation of sugar beet, with five N₂-fixing and two phosphate solubilizing bacteria in comparison to control and mineral fertilizers (N and P) application. Three bacterial strains dissolved P; all bacterial strains fixed N₂ and significantly increased growth of sugar beet. In the greenhouse, inoculations with PGPR increased sugar beet root weight by 2.8-46.7% depending on the species. Leaf, root and sugar yield were increased by the bacterial inoculation by 15.5-20.8, 12.3-16.1, and 9.8-14.7%, respectively, in the experiment of low- and high-OM soil. Plant growth responses were variable and dependent on the inoculants strain, soil organic matter content, growing stage, harvest date and growth parameter evaluated. The effect of PGPR was greater at early growth stages than at the later. Effective Bacillus species, such as OSU-142, RC07 and M-13, Paenibacillus polymyxa RC05, Pseudomonas putida RC06 and Rhodobacter capsulatus RC04 may be used in organic and sustainable agriculture.     

Nitrogen fixation and beneficial effects of some grain legumes and green-manure crops on rice

Studies were conducted on paddy soils to ascertain N₂ fixation, growth, and N supplying ability of some green-manure crops and grain legumes. In a 60-day pot trial, sunhemp (Crotalaria juncia) produced a significantly higher dry matter content and N yield than Sesbania sesban, S. rostrata, cowpeas (Vigna unguiculata), and blackgram (V. mungo), deriving 91% of its N content from the atmosphere. Dry matter production and N yield by the legumes were significantly correlated with the quantity of N₂ fixed. In a lowland field study involving sunhemp, blackgram, cowpeas, and mungbean, the former produced the highest stover yield and the stover N content, accumulating 160–250 kg N ha^-1 in 60 days, and showed great promise as a biofertilizer for rice. The grain legumes showed good adaptability to rice-based cropping systems and produced a seed yield of 1125–2080 kg ha^-1, depending on the location, species, and cultivar. Significant inter- and intraspecific differences in the stover N content were evident among the grain legumes, with blackgram having the highest N (104–155 kg N ha^-1). In a trial on sequential cropping, the groundnut (Arachis hypogaea) showed a significantly higher N₂ fixation and residual N effect on the succeeding rice crop than cowpeas, blackgram, mungbeans (V. radiata), and pigeonpeas (Cajanus cajan). The growth and N yield of the rice crop were positively correlated with the quantity of N₂ fixed by the preceding legume crop.     

Quantification of the contribution of biological nitrogen fixation to Cratylia mollis using the N natural abundance technique in the semi-arid Caatinga region of Brazil

This study was performed to investigate the capacity of the woody perennial Cratylia mollis, a legume endemic to the semi-arid region of the North-East of Brazil, to nodulate, and obtain N from BNF using the ¹⁵N natural abundance technique. To estimate the ¹⁵N abundance of the N-derived from soil, the leaves of several (4-6) non-legume and non-nodulating legume species growing in close proximity to the Cratylia were analysed for δ¹⁵N. The δ¹⁵N values of these reference plants were high (from +9 to +16‰) and showed relatively small differences between species at each site/sampling time. At the irrigated site at both samplings, and at the non-irrigated site sampled in the rainy season, the ¹⁵N abundance of the Cratylia mollis leaves was far lower (+1 to +5‰) strongly suggesting that the legume obtained large proportional contributions from BNF. As was to be expected, no nodules were found on the Cratylia plants at the non-irrigated site in the dry season, and the ¹⁵N abundance of the Cratylia mollis plants were very similar to that of the reference plants, consistent and nodulation and BNF being limited by water deficit at this time.     

Size, symbiotic effectiveness and genetic diversity of field pea rhizobia (Rhizobium leguminosarum bv. viciae) populations in South Australian soils

Field pea (Pisum sativum L.) is widely grown in South Australia (SA), often without inoculation with commercial rhizobia. To establish if symbiotic factors are limiting the growth of field pea we examined the size, symbiotic effectiveness and diversity of populations of field pea rhizobia (Rhizobium leguminosarum bv. viciae) that have become naturalised in South Australian soils and nodulate many pea crops. Most probable number plant infection tests on 33 soils showed that R. l. bv. viciae populations ranged from undetectable (six soils) to 32×10³ rhizobia g⁻¹ of dry soil. Twenty-four of the 33 soils contained more than 100 rhizobia g⁻¹ soil. Three of the six soils in which no R. l. bv. viciae were detected had not grown a host legume (field pea, faba bean, vetch or lentil). For soils that had grown a host legume, there was no correlation between the size of R. l. bv. viciae populations and either the time since a host legume had been grown or any measured soil factor (pH, inorganic N and organic C). In glasshouse experiments, inoculation of the field pea cultivar Parafield with the commercial Rhizobium strain SU303 resulted in a highly effective symbiosis. The SU303 treatment produced as much shoot dry weight as the mineral N treatment and more than 2.9 times the shoot dry weight of the uninoculated treatment. Twenty-two of the 33 naturalised populations of rhizobia (applied to pea plants as soil suspensions) produced prompt and abundant nodulation. These symbioses were generally effective at N₂ fixation, with shoot dry weight ranging from 98% (soil 21) down to 61% (soil 30) of the SU303 treatment, the least effective population of rhizobia still producing nearly double the growth of the uninoculated treatment. Low shoot dry weights resulting from most of the remaining soil treatments were associated with delayed or erratic nodulation caused by low numbers of rhizobia. Random amplified polymorphic DNA (RAPD) polymerase chain reaction (PCR) fingerprinting of 70 rhizobial isolates recovered from five of the 33 soils (14 isolates from each soil) showed that naturalised populations were composed of multiple (5-9) strain types. There was little evidence of strain dominance, with a single strain type occupying more than 30% of trap host nodules in only two of the five populations. Cluster analysis of RAPD PCR banding patterns showed that strain types in naturalised populations were not closely related to the current commercial inoculant strain for field pea (SU303, ≥75% dissimilarity), six previous field pea inoculant strains (≥55% dissimilarity) or a former commercial inoculant strain for faba bean (WSM1274, ≥66% dissimilarity). Two of the most closely related strain types (≤15% dissimilarity) were found at widely separate locations in SA and may have potential as commercial inoculant strains. Given the size and diversity of the naturalised pea rhizobia populations in SA soils and their relative effectiveness, it is unlikely that inoculation with a commercial strain of rhizobia will improve N₂ fixation in field pea crops, unless the number of rhizobia in the soil is very low or absent (e.g. where a legume host has not been previously grown and for three soils from western Eyre Peninsula). The general effectiveness of the pea rhizobia populations also indicates that reduced N₂ fixation is unlikely to be the major cause of the declining field pea yields observed in recent times.     

Estimating N2 fixation by field-grown lupins (Lupinus angustifolius L.) using soil and plant 15N enrichment

Summary Biological N₂ fixation was estimated in a field experiment following the addition of NH₄Cl or KNO₃ to unconfined microplots (1.5 m²) at 2.5 g N m^-2 (10 atom% ¹⁵N). A model of total N and ¹⁵N accumulation in lupins and decreasing ¹⁵N enrichment in the KCl-extractable soil-N pool (0–0.15 m depth) was used to estimate the proportion of N in lupins derived from biological N₂ fixation. Estimates of N₂ fixation derived from the model were compared with ¹⁵N isotope-dilution estimates obtained using canola, annual ryegrass, and wheat as nonfixing reference plants. Biomass, total N accumulation, or ¹⁵N enrichment in the lupin and reference crops did not differ whether NH _inf4 ^sup+ or NO _inf3 ^sup- was added as the labelled inorganic-N source. The decrease in soil ¹⁵N enrichment was described by first-order kinetics, whereas total N and ¹⁵N accumulation in the lupins were described by logistical equations. Using these equations, the uptake of soil N by lupins was estimated and was then used to calculate fixed N₂. Estimates of N₂ fixation derived from the model increased from 0 at 50 days after sowing to a maximum of 0.79 at 190 days after sowing. Those based on the ¹⁵N enrichment of the NO _inf3 ^sup- pool were 10% higher than those based on the mineral-N pool. ¹⁵N isotope-dilution estimates of N₂ fixation ranged from 0.37 to 0.55 at 68 days after sowing and from 0.71 to 0.77 at 190 days after sowing. Reference plant-derived values of N₂ fixation were all higher than modelled estimates during the early states of growth, but were similar to modelled estimates at physiological maturity. The use of the model to estimate N₂ derived from the atmosphere has the intrinsic advantage that the need for a non-fixing reference plant is avoided.     

Interactions between residue placement and earthworm ecological strategy affect aggregate turnover and N2O dynamics in agricultural soil

Previous laboratory studies using epigeic and anecic earthworms have shown that earthworm activity can considerably increase nitrous oxide (N₂O) emissions from crop residues in soils. However, the universality of this effect across earthworm functional groups and its underlying mechanisms remain unclear. The aims of this study were (i) to determine whether earthworms with an endogeic strategy also affect N₂O emissions; (ii) to quantify possible interactions with epigeic earthworms; and (iii) to link these effects to earthworm-induced differences in selected soil properties. We initiated a 90-day ¹⁵N-tracer mesocosm study with the endogeic earthworm species Aporrectodea caliginosa (Savigny) and the epigeic species Lumbricus rubellus (Hoffmeister). ¹⁵N-labeled radish (Raphanus sativus cv. Adagio L.) residue was placed on top or incorporated into the loamy (Fluvaquent) soil. When residue was incorporated, only A. caliginosa significantly (p < 0.01) increased cumulative N₂O emissions from 1350 to 2223 μg N₂O-N kg⁻¹ soil, with a corresponding increase in the turnover rate of macroaggregates. When residue was applied on top, L. rubellus significantly (p < 0.001) increased emissions from 524 to 929 μg N₂O-N kg⁻¹, and a significant (p < 0.05) interaction between the two earthworm species increased emissions to 1397 μg N₂O-N kg⁻¹. These effects coincided with an 84% increase in incorporation of residue ¹⁵N into the microaggregate fraction by A. caliginosa (p = 0.003) and an 85% increase in incorporation into the macroaggregate fraction by L. rubellus (p = 0.018). Cumulative CO₂ fluxes were only significantly increased by earthworm activity (from 473.9 to 593.6 mg CO₂-C kg⁻¹ soil; p = 0.037) in the presence of L. rubellus when residue was applied on top. We conclude that earthworm-induced N₂O emissions reflect earthworm feeding strategies: epigeic earthworms can increase N₂O emissions when residue is applied on top; endogeic earthworms when residue is incorporated into the soil by humans (tillage) or by other earthworm species. The effects of residue placement and earthworm addition are accompanied by changes in aggregate and SOM turnover, possibly controlling carbon, nitrogen and oxygen availability and therefore denitrification. Our results contribute to understanding the important but intricate relations between (functional) soil biodiversity and the soil greenhouse gas balance. Further research should focus on elucidating the links between the observed changes in soil aggregation and controls on denitrification, including the microbial community.     

Influence of mycorrhiza vs. soluble phosphate on growth,nodulation, and N2 fixation (15N) in alfalfa under different levels of water potential

Summary The legume Medicago sativa (+Rhizobium melilott) was grown under controlled conditions to study the interactions between soluble P in soil (four levels), or a mycorrhizal inoculum, and the degree of water potential (four levels) in relation to plant development and N₂ fixation. ¹⁵N-labelled ammonium sulphate was added to each pot for a qualitative estimate of N₂ fixation, in order to rank the effects of the different treatments.Dry-matter yield, nutrient content and nodulation increased with the amount of plant-available P in the soil, and decreased as the water stress increased, for each P-level. The mycorrhizal effect on dry matter, N yield, and on nodulation was little affected by the water potential. Since P uptake was affected by the water content in mycorrhizal plants, additional mechanisms, other than those mediated by P, must be involved in the mycorrhizal activity.There was a positive correlation between N yield and nodulation for the different P levels and the mycorrhizal treatment at all water levels. A high correlation between plant unlabelled N content and atom% ¹⁵N excess was also found for all levels of P. In mycorrhizal plants, however, the correlation between unlabelled N yield and ¹⁵N was lower. This suggests that mycorrhiza supply plants with other N sources in addition to those derived from the improvement on N₂ fixation.     

Natural N abundances of inorganic nitrogen in soil treated with fertilizer and compost under changing soil moisture regimes

This study was conducted to examine whether the applications of N-inputs (compost and fertilizer) having different N isotopic compositions (δ¹⁵N) produce isotopically different inorganic-N and to investigate the effect of soil moisture regimes on the temporal variations in the δ¹⁵N of inorganic-N in soils. To do so, the temporal variations in the concentrations and the δ¹⁵N of NH₄⁺ and NO₃⁻ in soils treated with two levels (0 and 150 mg N kg⁻¹) of ammonium sulfate (δ¹⁵N=−2.3‰) and compost (+13.9‰) during a 10-week incubation were compared by changing soil moisture regime after 6 weeks either from saturated to unsaturated conditions or vice versa. Another incubation study using ¹⁵N-labeled ammonium sulfate (3.05 ¹⁵N atom%) was conducted to estimate the rates of nitrification and denitrification with a numerical model FLUAZ. The δ¹⁵N values of NH₄⁺ and NO₃⁻ were greatly affected by the availability of substrate for each of the nitrification and denitrification processes and the soil moisture status that affects the relative predominance between the two processes. Under saturated conditions for 6 weeks, the δ¹⁵N of NH₄⁺ in soils treated with fertilizer progressively increased from +2.9‰ at 0.5 week to +18.9‰ at 6 weeks due to nitrification. During the same period, NO₃⁻ concentrations were consistently low and the corresponding δ¹⁵N increased from +16.3 to +39.2‰ through denitrification. Under subsequent water-unsaturated conditions, the NO₃⁻ concentrations increased through nitrification, which resulted in the decrease in the δ¹⁵N of NO₃⁻. In soils, which were unsaturated for the first 6-weeks incubation, the δ¹⁵N of NH₄⁺ increased sharply at 0.5 week due to fast nitrification. On the other hand, the δ¹⁵N of NO₃⁻ showed the lowest value at 0.5 week due to incomplete nitrification, but after a subsequence increase, they remained stable while nitrification and denitrification were negligible between 1 and 6 weeks. Changing to saturated conditions after the initial 6-weeks incubation, however, increased the δ¹⁵N of NO₃⁻ progressively with a concurrent decrease in NO₃⁻ concentration through denitrification. The differences in δ¹⁵N of NO⁻₃ between compost and fertilizer treatments were consistent throughout the incubation period. The δ¹⁵N of NO₃⁻ increased with the addition of compost (range: +13.0 to +35.4‰), but decreased with the addition of fertilizer (−10.8 to +11.4‰), thus resulting in intermediate values in soils receiving both fertilizer and compost (−3.5 to +20.3‰). Therefore, such differences in δ¹⁵N of NO₃⁻ observed in this study suggest a possibility that the δ¹⁵N of upland-grown plants receiving compost would be higher than those treated with fertilizer because NO₃⁻ is the most abundant N for plant uptake in upland soils.     

Competition between inoculated and indigenous Rhizobium/Bradyrhizobium spp. strains for nodulation of grain and fodder legumes in Pakistan

Summary The competitive ability of inoculated and indigenous Rhizobium/Bradyrhizobium spp. to nodulate and fix N₂ in grain legumes (Glycine max, Vigna unguiculata, Phaseolus vulgaris) and fodder legumes (Vicia sativa, Medicago sativa, and Trifolium subterraneum) was studied in pots with two local soils collected from two different fields on the basis of cropping history. The native population was estimated by a most-probable-number plant infectivity test in growth pouches and culture tubes. The indigenous rhizobial/bradyrhizobial population ranged from 3 to 2×10⁴ and 0 to 4.4×10³ cells g^-1 in the two soils (the first with, the second without a history of legume cropping). Inoculated G. max, P. vulgaris, and T. subterraneum plants had significantly more nodules with a greater nodule mass than uninoculated plants, but N₂ fixation was increased only in G. max and P. vulgaris. A significant response to inoculation was observed in the grain legume P. vulgaris in the soil not previously used to grow legumes, even in the presence of higher indigenous population (>10³ cells g^-1 soil of Rhizobium leguminosarum bv phaseoli). No difference in yield was observed with the fodder legumes in response to inoculation, even with the indigenous Rhizobium sp. as low as <14 cells g^-1 soil and although the number and weight of nodules were significantly increased by the inoculation in T. subterraneum. Overall recovery of the inoculated strains was 38–100%, as determined by a fluorescent antibody technique. In general, the inoculation increased N₂ fixation only in 3 out of 12 legume species-soil combinations in the presence of an indigenous population of rhizobial/bradyrhizobial strains.     

Sampling effects on the assessment of genetic diversity of rhizobia associated with soybean and common bean

Biological nitrogen fixation plays a key role in agriculture sustainability, and assessment of rhizobial diversity contributes to worldwide knowledge of biodiversity of soil microorganisms, to the usefulness of rhizobial collections and to the establishment of long-term strategies aimed at increasing contributions of legume-fixed N to agriculture. Although in recent decades the use of molecular techniques has contributed greatly to enhancing knowledge of rhizobial diversity, concerns remain over simple issues such as the effects of sampling on estimates of diversity. In this study, rhizobia were isolated from nodules of plants grown under field conditions, in pots containing soil, or in Leonard jars receiving a 10⁻² or a 10⁻⁴ serially-diluted soil inoculum, using one exotic (soybean, Glycine max) and one indigenous (common bean, Phaseolus vulgaris) legume species. The experiments were performed using an oxisol with a high population (10⁵ cells g⁻¹ soil) of both soybean rhizobia, composed of naturalized strains introduced in inoculants and of indigenous common-bean rhizobia. BOX-PCR was used to evaluate strain diversity, while RFLP-PCR of the ITS (internally transcribed spacer) region with five restriction enzymes aimed at discriminating rhizobial species. In both analyses the genetic diversity of common-bean rhizobia was greater than that of soybean. For the common bean, diversity was greatly enhanced at the 10⁻⁴ dilution, while for the soybean dilution decreased diversity. Qualitative differences were also observed, as the DNA profiles differed for each treatment in both host plants. Differences obtained can be attributed to dissimilarity in the history of the introduction of both the host plant and the rhizobia (exotic vs. indigenous), to host-plant specificity, rhizobial competitiveness, and population structure, including ease with which some types are released from microcolonies in soil. Therefore, sampling method should be considered both in the interpretation and comparison of the results obtained in different studies, and in the setting of the goals of any study, e.g. selection of competitive strains, or collection of a larger spectrum of rhizobia. Furthermore, effects of sampling should be investigated for each symbiosis.     

Turnover of grain legume N rhizodeposits and effect of rhizodeposition on the turnover of crop residues

The turnover of N derived from rhizodeposition of faba bean (Vicia faba L.), pea (Pisum sativum L.) and white lupin (Lupinus albus L.) and the effects of the rhizodeposition on the subsequent C and N turnover of its crop residues were investigated in an incubation experiment (168 days, 15 °C). A sandy loam soil for the experiment was either stored at 6 °C or planted with the respective grain legume in pots. Legumes were in situ ¹⁵N stem labelled during growth and visible roots were removed at maturity. The remaining plant-derived N in soil was defined as N rhizodeposition. In the experiment the turnover of C and N was compared in soils with and without previous growth of three legumes and with and without incorporation of crop residues. After 168 days, 21% (lupin), 26% (faba bean) and 27% (pea) of rhizodeposition N was mineralised in the treatments without crop residues. A smaller amount of 15–17% was present as microbial biomass and between 30 and 55% of mineralised rhizodeposition N was present as microbial residue pool, which consists of microbial exoenzymes, mucous substances and dead microbial biomass. The effect of rhizodeposition on the C and N turnover of crop residues was inconsistent. Rhizodeposition increased the crop residue C mineralisation only in the lupin treatment; a similar pattern was found for microbial C, whereas the microbial N was increased by rhizodeposition in all treatments. The recovery of residual ¹⁵N in the microbial and mineral N pool was similar between the treatments containing only labelled crop residues and labelled crop residues + labelled rhizodeposits. This indicates a similar decomposability of both rhizodeposition N and crop residue N and may be attributable to an immobilisation of both N sources (rhizodeposits and crop residues) as microbial residues and a subsequent remineralisation mainly from this pool.Abbreviations C or N_dec C or N decomposed from residues - C or N_mic microbial C or N - C or N_micres microbial residue C or N - C or N_min mineralised C or N - C or N_input added C or N as crop residues and/or rhizodeposits - dfr derived from residues - dfR derived from rhizodeposition - Ndfr N derived from residues - NdfR N derived from rhizodeposition - N_loss losses of N derived from residues - SOM soil organic matter - WHC water holding capacity     

BIOMASS ALLOCATION AND NITROGEN DISTRIBUTION IN RYEGRASS UNDER WATER AND NITROGEN SUPPLIES

Ryegrass (Lolium perenne L.) in grassland is known to sustain with water and nitrogen (N). This study investigates biomass and N partitioning in plant organs (roots, main and the youngest tillers) under water-nitrogen interactions. Nitrogen was applied at the rates of 50 and 100 mg N kg^?1 as N₁ (low N) and N₂ (high N) treatments, respectively, with uniform irrigation until 440 growing degree-days (GDD). Thereafter, the water supply was restricted to 50 mL on a weekly basis (W₁) against 50 mL on a daily basis (W₂) and concurrently, N enriched with 1 atom% ¹⁵N isotopes. Cumulative tillers’ biomass increased linearly from 1st to 8th order, but thereafter reached a plateau with further increases in number of negligible weights. Initially tiller mass and number per plan did not differ (P < 0.05) with water and/or N applications but changed at 788 GDD with clear differences at 911 GDD with the highest under N₂W₂ and lowest under N₁W₁. Nitrogen concentration sharply decreased from 530 to 700 GDD and then levelled off with age. The decline was more pronounced in tillers than roots. The high N treatment showed elevated N-concentration under both water treatments. Watering on a daily basis promoted vegetative growth. High water and N levels significantly (P < 0.05) influenced concentration of N absorbed during ¹⁵N labeling (N_L) in all organs with relatively pronounced N_L under N₂. The additive positive effect of W₂ and N₂ was obvious on N_L as compared to N_T, which showed that plants discriminate N-uptake on mass basis. Nitrogen (mobile) was higher in young and ¹⁵N (heavier) was low in young tillers and vice versa. Accumulation of N absorbed during ¹⁵N labeling (¹⁵N_A) was significant knowing that water is a strong determining factor of N concentration in ryegrass organs.     

Natural N abundances in a tallgrass prairie ecosystem exposed to 8-y of elevated atmospheric CO2

After 8-y of elevated CO₂, we previously detected greater amounts of total soil nitrogen, suggesting that rates of ecosystem N flux into or out of tallgrass prairie had been altered. Denitrification and associative N fixation rates are the two primary biological processes that are known to control N loss and accumulation in tallgrass prairie soil. Therefore, our objective was to assess the natural abundance of plant and soil ¹⁵N isotopes as a cumulative index of potential change in efflux or influx of N into and out of the tallgrass prairie after 8-y of exposure to elevated CO₂. Aboveground plant delta ¹⁵N values of Andropogon gerardii were close to zero and more positive as a result of elevated CO₂, but whole-soil values at the 5-30 cm depth were significantly reduced (6.8 vs 7.3; P<0.05) under elevated CO₂-chamber (EC) relative to ambient CO₂- chamber (AC). Total, aboveground plant biomass, root-in-growth, extractable N, microbial biomass N, and soil pools collectively exhibited a range of delta ¹⁵N values from −2.8 to 7.3. Measurements of surface soil ¹⁵N indicate that a change in N inputs and outputs has occurred as a result of elevated atmospheric CO₂. In addition to possible changes in denitrification and N₂ fixation, other sources of N such as the re-translocation of N to the surface from deeper soil layers are needed to explain how soil N accrues in surface soils as a consequence of elevated CO₂. Our results support the notion that C accrual may promote N accrual, possibly driven by high plant and microbial N demand amplified by soil N limitation.