利用15N2直接标记法研究水稻种植对稻田固氮量和固氮活性的影响

稻田固氮对土壤维持肥力有着重要的作用,但水稻种植与固氮菌及其活性之间的关系尚不清楚。本试验利用15N2直接标记法测定了下位砂姜土发育的简育水耕人为土在种水稻和不种水稻条件下的生物固氮量,及其在土壤不同层次(0~1、1~5、5~15 cm)和水稻中的分配,并通过实时荧光定量PCR技术测定了土壤中固氮菌nifH DNA及RNA基因数量。结果表明:种水稻处理显著提高了土壤各层固氮量,尤其提高了1~5 cm和5~15 cm土层土壤固氮量对总固氮量的贡献;种水稻处理的总固氮量是不种水稻处理的10.3倍;水稻植株中生物固定的氮占总固氮量的31.48%;在0~1 cm土层,种水稻处理显著提高了nifH RNA基因数量,而对nifH DNA基因数量的增加不显著。可见,水稻种植没有增加固氮菌的数量,稻田固氮量的增加是因为水稻种植极大地促进了固氮菌nifH基因的表达,提高了固氮菌的固氮活性。     

Growth and Nitrogen Uptake in Sorghum Plants Manured with Leucaena leucocephala Leaves as Affected by Nitrogen Rate and Time of Application

A pot experiment was conducted to determine the effect of four rates of nitrogen (N) in the form of leucaena leaves and the time of application on the performance of sorghum plants using the ¹⁵N isotopic dilution technique. Results showed that leucaena green manure (LGM) increased dry matter and N yield of sorghum. Nitrogen recoveries of LGM ranged between 23 and 47%. An additional beneficial effect of LGM was attributed to the enhancement of soil N uptake. The best timing of LGM incorporation for obtaining more N derived from LGM, less soil N uptake, and greater dry matter and N in sorghum leaves seemed to be at planting. However, the appropriate timing and rate of LGM to obtain greater dry matter and N yield in panicles, as well as in the whole plant of sorghum, appeared to be at 30 days before planting, particularly a rate of 120 kg N ha^?1.     

Determining Symbiotic Nitrogen Fixation in Dry Bean Cultivars Using Ureide Method and Isotope Dilution Techniques

Symbiotic nitrogen fixation (SNF) is an environmentally safe source of nitrogen (N) to the crop plants. In total, 12 dry bean (Phaseolus vulgaris L.) cultivars from pinto, navy, black, and kidney market classes were inoculated with rhizobia and grown in a greenhouse. SNF was estimated using isotope dilution technique and ‘ureide’ method. The amount of SNF ranged between 33 and 68 mg N plant^–1 when determined using ¹⁵N isotope dilution and followed the order: pinto > navy > black > kidney. Percent N derived from atmosphere (%Ndfa) significantly varied between 49% and 90% at V₃ and between 71% and 98% at R₂ stages. The outcomes of the experiment suggested that dry bean cultivars from different market classes have variable N₂ fixation ability, and fertilizer N required should be calculated according to their SNF potentials and N need of a specific market class or cultivar. Stable isotope dilution should be used as the standard procedure to estimate the SNF in dry bean.     

Performance of Sorghum Grown on a Salt Affected Soil Manured with Dhaincha Plant Residues Using a 15N Isotopic Dilution Technique

A field experiment was conducted on a salt-affected soil to determine the effect of application of three types of Dhaincha (Sesbania aculeata Pers.) residues (R, roots; L, shoots; L+R, shoots plus roots) on the performance of sorghum (Sorghum bicolor L.) using the indirect ¹⁵N isotopic dilution technique. Results indicated that sesbania residues (L and L+R), used as green manures, significantly increased grain yield, dry matter production, N uptake, and water use efficiency of sorghum. Percentages of nitrogen (N) derived from residues (%Ndfr) in sorghum ranged from 6.4% to 28%. The N recoveries in sorghum were 52%, 19.6% and 19.7% of the total amount contained in sesbania roots, shoots and roots plus shoots, respectively. The beneficial effects of sesbania residues are attributed not only to the additional N availability to the plants, but also to effects on the enhancement of soil N uptake, particularly in the L+R treatment. The findings suggest that the use of Sesbania aculeata residues, as a green manure, can provide a substantial portion of total N in sorghum. In addition, the use of sesbania green manure in salt-affected soils, as a bio-reclaiming material, can be a promising approach for enhancing plant growth on a sustainable basis.     

Growth and Nitrogen Fixation in Soybean as Affected by Phosphorus Fertilizer and Sheep Manure using 15N Isotopic Dilution

A field experiment was conducted to study the effect of adding different phosphorus (P) fertilizer levels [0, 40, and 80 kg phosphorus pentoxide (P₂O₅) ha^?1 (abbreviated as P0, P1, and P2, respectively)] and rates of sheep manure (M) [0, 20, and 40 ton ha^?1 (abbreviated as M0, M1, and M2, respectively)] on growth and nitrogen (N₂) fixation of soybean (Glycine max L.). Sorghum bicolor L. was employed as a reference crop to evaluate N₂ fixation using the ¹⁵N-isotpic dilution technique. Results showed that addition of P fertilizer or sheep manure had positive effects on dry-matter production, N accumulation, and seed yield. Such effects were more pronounced when adding sheep manure and P together than adding separately. Solely P fertilizer had a small impact on N₂ fixation. A tangible increase in the amounts of N₂ fixed due to manure addition occurred. The efficient use of N fertilizer (%NUE) increased significantly as the result of adding a high level of P fertilizer. However, a drastic decrease in %NUE was observed when sheep manure was added solely or in combination with P fertilizer. From productivity and ecological standpoints, P2M1 and P2M2 surpassed the other treatments in showing greater grain yield and greater N₂ fixation. However, considering the high cost of sheep manure, P2M1 was the optimal treatment for improving growth and N₂ fixation in soybean plants with minimal manure consumption. In conclusion, the integrated use of manure and P fertilizer could be considered a useful agricultural practice for improving the performance of soybean plants grown in an Aridisol. Their beneficial effects were mainly attributed to the enhancement of N₂ fixation through root growth and soil property improvements besides being a source of P and other nutrients that are essential for N₂-fixation process.     

Natural 15N and 13C abundance in Andisols influenced by long-term fertilization management in Japan

Two field experiments were conducted on Andisols in Japan to evaluate the changes in the natural ¹⁵N and ¹³C abundance in the soil profile and to determine whether the values of δ¹⁵N could be used as an indicator of fertilizer sources or fertilizer fate. The 6-year experiment conducted at the National Agricultural Research Center (NARC) consisted of the following treatments: application of swine compost (COMPOST), slow-release nitrogen fertilizer (SRNF), readily available nitrogen fertilizer (RANF), and absence of fertilization (CONTROL). Experimental plots located at the Nippon Agricultural Research Institute (NARI) received cattle compost at different rates for 12 years; a forest soil at this site was sampled for comparison. Swine compost application led to a considerable change in the δ¹⁵N distribution pattern in the soil profile, with the highest δ¹⁵N values recorded in the top 20 cm layers of the COMPOST plot, decreasing in the sequence of CONTROL >- RANF > SRNF, mainly due to the relatively high δ¹⁵N value of swine compost and its subsequent decomposition. In contrast, SRNF application resulted in the lowest δ¹⁵N values in soil, indicating the presence of negligible nitrogen losses relative to input and low nitrogen cycling rates. Values of δ¹⁵N increased with compost application rates at NARI. In the leachate collected at 1-m depth, the δ¹⁵N values decreased in the sequence of COMPOST > RANF ≥ CONTROL > SRNF. The δ¹³C values in soil peaked in the 40–60 cm layers for all the fertilizers. The δ¹³C value was lowest in forest soil due to the presence of plant residues in soil organic matter. These results indicated that the δ¹⁵N values in the upper soil layers or leachate may enable to detect pollution sources of organic or inorganic nitrogen qualitatively in Andisols.     

Effects of Irrigation Water Quality and Nitrogen Rate on the Recovery of 15N Fertilizer by Sorghum in Field Study

The effects of ¹⁵N-labeled ammonium nitrate on yield, uptake of nitrogen (N) by sorghum (Sorghum sudanense, Piper), and on N remaining in the soil were studied in a field experiment with different N rates (0, 50, and 100 kg N ha^?1) and with two irrigation water qualities, well water (WW) and treated wastewater (TWW). Treated wastewater irrigation increased dry matter and N yield compared to WW. At equal N rates, recovery of ¹⁵N-labeled fertilizer by plants increased with TWW irrigation compared to WW (36% versus 23%). Neither fertilizer rate nor water quality had an effect on the ¹⁵N-labeled fertilizer remaining in the 0- to 60-cm layer of soil. On average 41% in the TWW treatment (49–33%) and 38% in WW treatment was mostly present in the surface 20-cm layer. Losses of ¹⁵N-labeled fertilizer were unaffected by irrigation water quality (35%) and increased with N application rate in TWW (4% versus 31%).     

Nitrogen accumulation in three legumes and two cereals with emphasis on estimation of N2 fixation in the legumes by the natural 15N-abundance technique

Summary N accumulation and natural ¹⁵N abundance in three legumes (groundnuts, cowpeas, and soybeans) and in two cereals (sorghum and maize) were investigated over two seasons in Alfisols with and without N fertilization. Using the N uptake and natural ¹⁵N abundance of non-nodulating plants as the indication of N derived from soil and fertilizer, the per cent N derived from atmospheric N₂ was calculated for nodulated plants. In the first experiment, the groundnut genotype contained 85% atmosphere-derived N, but the percentage decreased with N application. Estimates of atmosphere-derived N by the N-difference and ¹⁵N-abundance techniques gave identical results. The percentages of atmosphere-derived N estimated by the two methods at different stages of groundnut growth were also similar. In the second experiment, atmosphere-derived N was estimated in plants grown with 0–200 kg ha^-1 applied N. The estimated atmosphere-derived N ranged from 42% to 61% for groundnuts from 33% to 77% for cowpeas, and from 24% to 48% for soybeans, depending on the amount of N applied. Inoculation with a Bradyrhizobium strain increased the percentage of atmospherederived N in soybean plants grown without any fertilizer N. The natural ¹⁵N abundance of sorghum and maize was very close to that of the non-nodulating groundnut, suggesting that these cereals can be used as reference plants in the estimation of atmosphere-derived N by the natural ¹⁵N-abundance method.ICRISAT Journal Article No. 876     

Soil N Losses by Denitrification Evaluated Using the 15N Tracer Method

Molecular nitrogen (N₂) and nitrous oxide (N₂O) generated by denitrification increase N losses in the soil–plant system. This study aimed to quantify N₂ and N₂O from potassium nitrate (K¹⁵NO₃) applied to soils with different textures and moisture contents in the absence and presence of a source of carbon (C) using the ¹⁵N tracer method. In the three soils used (sandy texture (ST), sandy clay loam texture (SCLT), and clayey texture (CT)), three moisture contents were evaluated (40%, 60%, and 80% of the water holding capacity (WHC)) with (D⁺) and without (D^?) dextrose added. The treatments received 100 mg N kg^?1 (KNO₃ with 23.24 atom% ¹⁵N). N₂ emissions occurred in all of the treatments, but N₂O emissions only occurred in the D⁺ treatment, showing increases with increasing moisture content. SCLT with 80% WHC in the D⁺ treatment exhibited the highest accumulated N emission (48.26 mg kg^?1). The ¹⁵N balance suggested trapping of the gases in the soil.     

固态13C和15N核磁共振法研究15N标记土壤的腐殖质组分

Five humic fractions were obtained from a uniformly ^15N-labelled soil by extraction with 0.1 mol L^-1 Na4P2O7,0.1mol L^-1 NaOH ,and HF/HCl-0.1 mol L^-1 NaOH,consecutively,and analyzed by ^13C and ^15N CPMAS NMR (cross polarization and magic angle spinning nuclear magnetic resonace).Compared with those of native soils humic fractions studied as a whole contained more alkyls ,methoxyls and O-alkyls,being 27%-36%,17%-21%and 36%-40%,respectively,but fewer aromatics and carboxyls(bein 14%-20% and 13%-90%,respectively),Among those humic fractions ,the humic acid(HA)and fulvic acid(FA) extracted by 0.1 mol L^-1 Na4P2O7 contained slightly more carboxyls than corresponding humic fractions extracted by 0.1 mol L^-1 NaOH ,and the HA extacted by 0.1 mol L^-1 NaOH after treatment with HF/HCl contained the least aromatics and carboxyls.The distribution of nitrogen functional groups of soil humic fractions studied was quite similar to each other and also quite similar to that of humic fraction from native soils.More than 75% of total N in each fraction was in amide from,with 9%-13% present as aromatic and /or aliphatic amines and the remainder as heerocyclic N.     

Evaluation of N2 fixation by applying 15N labeled plant material and ammonium sulfate

Effect of different ¹⁵N labeled sources on the estimation of N₂ fixation was investigated. The combination of ¹⁵N labeled ammonium sulfate, ¹⁵N labeled plant material, and ¹⁵N labeled ammonium sulfate with unlabeled plant material, was examined in pot experiments. Two cultivars of soybean (Glycine max) and one of mungbean (Vigna radiata) were used. No significant difference was observed among the treatments for the estimation of N₂ fixation. This was due to the homogeneity and stability of the ¹⁵N abundance in soil which resulted in a similar N uptake from the soil by the N² fixing and reference crops. The plant yield, total N uptake and amount of N₂ fixed were higher in the Yellow Soil than in the Andosol. The amount of N₂ fixed was strongly influenced by the plant growth and consequently it affected the plant yield. The slow decomposition of plant material in the Andosol resulted in a low yield in both the N₂ fixing and reference crops. Thus, the artificial decrease of the available N content in soil, by application of plant material, did not stimulate N, fixation but suppressed plant growth and N₂ fixation.     

施氮量对嘎啦幼苗~(15)N、~(13)C分配利用特性影响

【目的】采用15N、13C同位素示踪技术,通过对不同施氮量下嘎啦幼苗生长状况及氮、碳分配、利用特性等的研究,以期为苹果生产合理施肥提供依据。【方法】将2年生盆栽嘎啦幼苗进行低、中、高三个氮水平处理,同时进行15N标记。在新梢旺长初始期、新梢旺长期、新梢缓长期分别进行整株13C标记,72小时后,整株解析为叶、梢、根三部分,进行15N、13C测定。样品全氮用凯氏定氮法测定,15N丰度用ZHT-03质谱计测定。13C丰度用DELTA V Advantage同位素比率质谱仪测定。【结果】1)中、高氮水平的施肥处理可在不同程度上提高整株及叶片干物质量和新梢长度。新梢旺长初始期和新梢缓长期嘎啦幼苗整株干物质量、新梢旺长期叶片干物质分配比率在中、高氮水平处理间差异不显著,中氮水平经济有效。新梢旺长期以后新梢长度以中氮高氮低氮,三者间差异性显著,中氮处理有利于新梢生长。2)在新梢旺长初始期,低氮处理植株叶片15N分配率达50%,比其他处理高出13个百分点左右,表明低氮处理更多的氮被叶片所利用,中氮和高氮处理间差异不显著,说明在本试验施氮条件下中氮供应水平已能满足氮素营养需求。3)新梢旺长期和新梢缓长期幼苗13C固定量均以中氮处理最高,新梢旺长初始期3个处理间根系13C分配率中氮高氮低氮,表明中氮处理有利于碳同化物在嘎啦幼苗中的分配。4)不同施氮量处理的嘎啦幼苗,15N利用率随施氮水平提高而降低,高氮处理对碳同化物分配没有显著贡献。【结论】低、中、高氮不同处理新梢缓长期碳同化物在各器官间的分配比较均衡,氮素水平不能影响碳同化物的分配。盆栽试验表明,中氮水平在保证营养供应的同时,能够促进新梢生长和树势健壮。     

Comparison of 15N natural abundance and difference methods for estimating N2 fixation of soybeans grown in a tropical soil

Abstract

A study was carried out to compare the difference or N-yield method with the ¹⁵N natural abundance method for the estimation of the fractional contribution of biological N₂ fixation in the different plant parts of nodulating and non-nodulating isolines of soybeans. The results indicated that the δ¹⁵N values of most plant parts of soybeans were significantly lower (p<0.05) in the nodulating than in the non-nodulating isoline. However, in the case of the root+nodule component, the δ¹⁵N value was higher in the nodulating than in the non-nodulating isoline possibly due to isotopic discrimination of ¹⁵N over ¹⁴N which may have occurred in the nodules. Inoculation of soybeans with the Bradyrhizobium japonicum strain CB 1809 increased significantly (p<0.05) the δ¹⁵N value of the root+nodule component implying that the effectiveness of the soybean-rhizobium symbiosis had increased by inoculation.

Percentage of plant N derived from atmospheric N₂ fixation (%Ndfa) estimated by the ¹⁵N natural abundance method was highly correlated (r=0.762, p<0.01) with that by the difference or N-yield method and the differences between the two methods were not statistically significant. The agreement between the two methods was closer at maturity than at the early reproductive stage.

The %Ndfa obtained by the difference method ranged from 48.4 to 92.6% whereas the %Ndfa obtained by the ¹⁵N natural abundance method ranged from 43.2 to 92.4% in the different plant parts. Based on the ¹⁵N natural abundance method, approximately 15% of the N in pod, shoot, grain, and shell was derived from the soil but in the case of stover, this fraction was about 55%.     

应用15N稀释法测定土壤氮素初级转化速率的一些关键技术

本文综合评述了应用~(15)N库稀释法测定土壤氮素初级转化速率的一些关键技术,即~(15)N标记土壤氮库的方法、~(15)N的加入量、丰度和标记物种类的选择,以及初始取样时间的确定。只有合理地运用这些关键技术,才能更准确地测定土壤氮素初级转化速率,进而更真实地表征土壤氮素的实际周转状况。     

15N标记的土壤估测某些豆科植物N2固定的温室实验

Pot experiments were carried out to estimate N2 fixation by vetch,milk vetch,sickle alfalfa and broadbean in pure stand using a ^15N-labelled soil.Winter wheat was used as the non-fixing control.The 15N-labelled soil used was prepared by growing corn-wheat-corn successively on a nearly organic-matter-free Xiashu loess supplemented with adequate amounts of (15NH4)2SO4,P,K and micronutrients,then incorporating these 15N-labelled plant materials into the soil after each havest,and allowing the plant materials to be decomposed aerobically for 410d after incorporation of the plant material of the thire crop.The 15N enrichment of wheat plant-N varied slightly with organs,with a maximum difference of 9.8%,Based on 15N enrichment of soil N inferred from the mean value of the 15N enrichment in different organs of wheat 79%-91% of total N in the tops and 67%-74% of total N in the roots of legumes studied were derived from atmosphere .Estimate by isotope dilution method was in good agreement with that by the conventional difference method provided values obtained by the latter were corrected for seed N,and also with that from the measurement of N accumulated in the tops of the legumes.     

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.     

不同施氮方式对嘎啦苹果碳氮利用和产量品质的影响

以15年生嘎啦苹果/平邑甜茶为试材,采用C、N双标记技术,研究果实发育期不同施氮方式(传统一次性施氮、分次施氮和渗灌施氮,分别用ON、TN和IN表示)对苹果植株碳氮营养吸收、利用、分配、损失及果实产量和品质的影响。结果表明:至果实成熟期,苹果各器官Ndff值均为INTNON,新生器官间(果实、叶片和1年生枝)差异显著。植株全氮量和~(15)N吸收量均以IN处理最大,ON处理最低。与ON处理相比,TN和IN处理~(15)N利用率分别提高了41.63%和68.60%,而~(15)N损失率分别降低了10.60%和18.63%。各处理不同土层~(15)N残留量差异显著,0—40 cm土层~(15)N残留量为INTNON,60—120 cm土层趋势相反。TN和IN处理果实和贮藏器官(多年生枝、中心干和粗根)的~(13)C分配率均显著高于ON处理,而营养器官(叶片和1年生枝)的~(13)C分配率则以ON处理最高,IN处理最低。同时,在IN处理下,苹果产量、硬度、可溶性糖和糖酸比等品质指标均达到最高水平。综上,渗灌施氮通过降低氮素损失,显著促进植株对氮素的吸收利用,并优化光合产物在各器官间分配,从而最有利于苹果产量和品质的提高。     

利用15N揭示滴灌区盐旱胁迫对土壤氮素分布与棉花生长的影响

为了探究盐旱胁迫对土壤中氮素分布和棉花生长的影响,通过测坑试验研究滴灌区不同盐分、干旱条件下土壤全氮、硝氮、氨氮的分布和棉花生长情况。试验设置3种盐分梯度的土壤（电导率,EC）：3,6,9 dS/m,分别用T1、T2、T3表示;3个灌水量：2 700,3 600,4 500 m³/hm²,分别用W1、W2、W3表示（4 500 m³/hm²为当地推荐灌水量）。结果表明：当土壤盐分梯度> 3 dS/m时土壤全氮累积量显著高于低盐土壤（P<0.05）,且土壤盐分对棉花花期生长影响较大。土壤的氨氮挥发量和土壤盐分梯度成正比。土壤硝态氮的淋失与灌水量呈正比,与正常灌水量的硝态氮淋失相比,水分胁迫对棉花产量的影响更为严重（P<0.01）。随土层深度的增加,土壤碱解氮以每20 cm土层8%的速度减少。各处理土壤¹⁵N残留率为11%~40%,随土壤盐度增加而增加,随灌水量增加而减少,与土壤全氮含量呈正比,与棉花产量呈反比。综上所述,T1W3处理更有利于棉花对氮肥的利用和产量的提高,推荐滴灌区棉花土壤盐度<3 dS/m,灌水量4 500 m³/hm²,可在花期适当提高施肥量以稳定产量。     

NATURAL ABUNDANCES OF 15NITROGEN AND 13CARBON INDICATIVE OF GROWTH AND N2 FIXATION IN POTASSIUM FED LENTIL GROWN UNDER WATER STRESS

Dual natural abundance analysis of ¹⁵Nitrogen (N) and ¹³Carbon (C) isotopes in lentil plants subjected to different soil moisture levels and rates of potassium (K) fertilizer were determined to assess crop performance variability in terms of growth and N₂-fixation (Ndfa). The δ¹⁵N values in lentils ranged from +0.67 to +1.36‰; whereas, those of the N₂-fixed and reference plant were ?0.45 and +2.94‰, respectively. Consequently, the Ndfa% ranged from 45 and 65% of total plant N uptake. Water stress reduced Δ¹³C values. However, K fertilization enhanced whole plant Δ¹³C along with dry matter yield and N₂-fixation. The water stressed plants amended with K fertilizer seemed to be the best treatment because of its highest pod yield, high N balance, and N₂-fixation with low consumption of irrigation water. This illustrates the ecological and economical importance of K fertilizer in alleviating water stress occurring during the post-flowering period of lentil.