15N isotope dilution techniques to study soil nitrogen transformations and plant uptake

The use of¹⁵N as a tracer in soil/plant research is examined. The limitations of the so-called Ndff approach are discussed to show the need to consider not just the fate of the added label but also the path that was followed and the rate of the transformation. The development of¹⁵N isotope dilution techniques to determine gross rates of nitrogen transformation in soil is reviewed with some indications as to the further development of the approach.     

Estimation of the contribution of biological N2 fixation to twoPhaseolus vulgaris genotypes using simulation of plant nitrogen uptake from15N-labelled soil

A technique for the application of the¹⁵N isotope dilution technique for the quantification of plant associated biological nitrogen fixation (BNF) was tested and applied to quantify the BNF contribution to two genotypes ofPhaseolus vulgaris. The technique makes use of sequential measurements of the¹⁵N enrichment of soil mineral N, and the uptake of labelled N by the N₂-fixing plant, to simulate its uptake of soil N (the soil to plant simulation technique). The test was made with two non-N₂-fixing crops (non-nodulating beans and wheat) and two bean genotypes (PR 923450 and Puebla 152), at two levels of N fertilizer addition (10 and 40 kg N ha^–1), to compare the actual N uptake with that simulated from the soil and crop¹⁵N data. The simulation of the soil N uptake by the non-nod bean crop using this soil to plant simulation technique underestimated by 20 to 30% the true N uptake, suggesting that the mineral N extracted from soil samples taken from the 0–15cm layer had a higher¹⁵N enrichment than that N sampled by the roots of this crop. In the case of the wheat crop the simulation resulted in a much greater underestimation of actual N uptake. In general the results using this technique suggested that BNF inputs to the bean cultivars was higher than would be expected from the nodulation and acetylene reduction data, except for the early PR beans in the 40 kg N ha^–1 treatment. In this case the total N and simulated soil N accumulation were well matched suggesting no BNF inputs. An allied technique (the plant to plant simulation technique) was proposed where the¹⁵N enrichrnent of soil mineral N was simulated from the data for total N and labelled N accumulation taken from sequential harvests of either of the non-N₂ -fixing control crops. This was then utilized in combination with the labelled N uptake data of the other crop to simulate its soil N uptake. However, the results using either technique indicated that the wheat and non-nod or nodulating beans exploited pools of N in the soil with completely different¹⁵N enrichments probably due to differences in exploitation of the soil N with depth.     

The influence of rate and time of nitrate supply on nitrogen fixation and yield in pea (Pisum sativum L.)

The influence of nitrate N supply on dry matter production, N content and symbiotic nitrogen fixation in soil-grown pea (Pisum sativum L.) was studied in a pot experiment by means of¹⁵N fertilizer dilution. In pea receiving no fertilizer N symbiotic nitrogen fixation, soil and seed-borne N contributed with 82, 13 and 5% of total plant N, respectively. The supply of low rates of nitrate fertilizer at sowing (starter N) increased the vegetative dry matter production, but not the seed yield significantly. Nitrogen fixation was not significantly decreased by the lower rates of nitrate but higher rates supplied at sowing reduced the nitrogen fixation considerably. Applying nitrate N at the flat pod growth stage increased the yield of seed dry matter and N about 30% compared to pea receiving no nitrate fertilizer. Symbiotic nitrogen fixation was reduced only about 11%, compared with unfertilized pea, by the lowest rate of nitrate at this application time. The pea very efficiently took up and assimilated the nitrate N supplied. The average fertilizer N recovery was 82%. The later the N was supplied the more efficiently it was recovered. When nitrate was supplied at the flat pod growth stage 88% was recovered, and 90% of this N was located in the seeds.     

Biological nitrogen fixation by two tropical forage legumes assessed from the relative ureide abundance of stem solutes: 15N calibration of the technique in sand culture

The use of the relative ureide abundance (RUA) in the sap of mainly tropical ureide-producing legumes as a means to estimate the contribution of biological nitrogen fixation (BNF) is potentially an useful technique as it does not require the use of reference plants or additions of ¹⁵N-labelled fertilizer, and the analyses necessitate only relatively simple equipment. However, one problem in the application of the technique arises from the difficulty of obtaining sap samples from such legumes, especially small-stemmed forage legumes under field conditions. This study was conducted to investigate the possibility of using RUA in hot-water extracts of the stems of two forage legumes, Desmodium ovalifolium and a Centrosema hybrid, to estimate the contribution of BNF. In this case only ureide and nitrate are analysed to calculate RUA (100 × ureide-N/(ureide-N + nitrate-N)). The technique was calibrated with the ¹⁵N isotope dilution technique in sand culture where the plants were fed with 5 different levels of nitrate (0, 12.5, 25, 50 and 100 mg N pot^-1). Despite the fact that in many stem extracts more than 90% of the N was neither nitrate or ureide, the colorimetric techniques utilised proved reliable and relatively immune to interference from other solutes in the extracts. One problem with the use of the ¹⁵N dilution technique to calibrate the RUA technique is that the former gives an integrated estimate of the BNF contribution since planting (or between harvests) and the latter is a point estimate at the time of sampling. This was overcome by using a `plant to plant simulation technique' where estimates of BNF are calculated from the daily accumulation of total N and the labelled N derived from the growth medium by the legumes using a curve-fitting strategy. These estimates of BNF for the days when stem extracts were analysed for nitrate and ureide showed linear correlations (r ² = 0.82 and 0.90 for the D. ovalifoliumand Centrosema hybrid, respectively). This indicated that RUA of stem extracts of these two legumes was a reliable indicator of the BNF contribution, at least under controlled conditions.     

Use of the 15N natural abundance technique to quantify biological nitrogen fixation by woody perennials

Biological nitrogen fixation (BNF) associated with trees and shrubs plays a major role in the functioning of many ecosystems, from natural woodlands to plantations and agroforestry systems, but it is surprisingly difficult to quantify the amounts of N₂ fixed. Some of the problems involved in measuring N₂ fixation by woody perennials include: (a) diversity in occurrence, and large plant-to-plant variation in growth and nodulation status of N₂-fixing species, especially in natural ecosystems; (b) long-term, perennial nature of growth and the seasonal or year-to-year changes in patterns of N assimilation; and (c) logistical limitations of working with mature trees which are generally impossible to harvest in their entirety. The methodology which holds most promise to quantify the contributions of N₂ fixation to trees is the so-called `¹⁵N natural abundance' technique which exploits naturally occurring differences in ¹⁵N composition between plant-available N sources in the soil and that of atmospheric N₂. In this review we discuss probable explanations for the origin of the small differences in ¹⁵N abundance found in different N pools in both natural and man-made ecosystems and utilise previously published information and unpublished data to examine the potential advantages and limitations inherent in the application of the technique to study N₂ fixation by woody perennials. Calculation of the proportion of the plant N derived from atmospheric N₂ (%Ndfa) using the natural abundance procedure requires that both the ¹⁵N natural abundance of the N derived from BNF and that derived from the soil by the target N₂-fixing species be determined. It is then assumed that the ¹⁵N abundance of the N₂-fixing species reflects the relative contributions of the N derived from these two sources. The ¹⁵N abundance of the N derived from BNF (B) can vary with micro-symbiont, plant species/provenance and growth stage, all of which create considerable difficulties for its precise evaluation. If the%Ndfa is large and the ¹⁵N abundance of the N acquired from other sources is not several ¹⁵N units higher or lower than B, then this can be a major source of error. Further difficulties can arise in determining the ¹⁵N abundance of the N derived from soil (and plant litter, etc.) by the target plant as it is usually impossible to predict which, if any, non-N₂-fixing reference species will obtain N from the same N sources in the same proportions with the same temporal and spatial patterns as the N₂-fixing perennial. The compromise solution is to evaluate the ¹⁵N abundance of a diverse range of neighbouring non-N₂-fixing plants and to compare these values with that of the N₂-fixing species and the estimate of B. Only then can it be determined whether the contribution of BNF to the target species can be quantified with any degree of confidence. This review of the literature suggests that while the natural abundance technique appears to provide quantitative measures of BNF in tree plantation and agroforestry systems, particular difficulties may arise which can often limit its application in natural ecosystems.     

Assessment of biological nitrogen fixation

The four commonly used methods for measuring biological nitrogen fixation (BNF) in plants are: the total nitrogen difference (TND) method, acetylene reduction assay (ARA) technique, xylem-solute (or ureide production) method and the use of¹⁵N labelled compounds.The TND method relies on a control non-N₂-fixing plant to estimate the amount of N absorbed by the fixing plant from soil. It is one of the simplest and least expensive methods, but works best under low soil N conditions. The ARA technique measures the rate of acetylene conversion to ethylene by the N₂-fixing enzyme, nitrogenase. The ethylene produced can then be converted into N₂ fixed, using a conversion ratio, originally recommended as 3. Although the method is inexpensive and highly sensitive, its major disadvantages are, the short-term nature of the assays, the doubtful validity of always using a conversion ratio of 3 and the auto-inhibition of acetylene conversion to ethylene. The ARA technique is therefore not a method of choice for measuring BNF.The xylem-solute technique can be used to measure BNF for those species that produce significant quantities of ureide as product of BNF. Although simple and relatively inexpensive, it is an instantaneous assay and also needs to be calibrated against a known method. The most serious limitation is, that only a small proportion of N₂-fixing plants examined are ureide exporters, and the method is therefore not widely applicable.The¹⁵N methods, classified into the isotope dilution and A-value methods, appear to be the most accurate, but also the most expensive. They involve labelling soil with¹⁵N fertilizer and using a non-N₂-fixing reference plant to measure the¹⁵N/¹⁴N ratio in the soil. The¹⁵N isotope dilution approach is both operationally and mathematically simpler than the A-value approach. To limit potential errors in the selection of reference crops, it is recommended to use¹⁵N labelled compounds or soil labelling methods that result in the slow release of¹⁵N or the slow decline of¹⁵N/¹⁴N ratio in the soil. Additionally, the use of several reference plants rather than a single one can improve the accuracy of the results.     

Influence of delaying flood and preflood nitrogen application on dry-seeded rice

In the southern U.S. rice belt it is recommended that rice (Oryza sativa L.) grown in the dry-seeded, delayed flood cultural system have the preflood N fertilizer applied and the field flooded at the fourth to fifth leaf stage of plant development. The objective of this field study was to determine if delaying the flood and preflood N application past the fifth leaf stage was detrimental to rice total N and fertilizer¹⁵N uptake, total dry matter, and grain yield. This study was conducted on a Crowley silt loam (Typic Albaqualfs) and a Perry clay (Vertic Haplaquepts). The preflood N fertilizer and flood were delayed 0, 7, 14, or 21 d past the fourth to fifth leaf stage, after which time a permanent flood was established and maintained until maturity. All treatments received 20.5 g N m^–2 as¹⁵N-labeled urea in three topdress applications. All plant and soil samples were taken at maturity. Harvest index increased as the preflood N and flood were delayed past the 4 to 5 leaf stage. Total N in the grain + straw either decreased or showed a decreasing trend as the N and flood were delayed. Similarly, uptake of native soil N decreased as flood was delayed. Conversely, percent recovery of fertilizer N in the rice plant and the plant-soil system increased as the preflood N and flood were delayed. Rice grain yield was not significantly affected by delaying the preflood N and flood up to 21 d.Received....... . Published with permission of the Director of the Arkansas Agric. Exp. Stn. Project ARK01386. Supported in part by the Tennessee Valley Authority National Fertilizer and Environmental Research Center and the Arkansas Rice Research and Promotion Board.     

Estimation of N2 fixation in Desmodium ovalifolium from the relative ureide abundance of stem solutes: Comparison with the 15N-dilution and an in situ soil core technique

Many, but not all, legumes of tropical origin, transport fixed N from the nodules to the shoot tissue in the form of ureides, and the mineral N absorbed from the soil is principally transported in the form of nitrate. The analysis of stem xylem sap, or hot-water extracts of stem tissue, for ureide and nitrate has been used successfully to quantify BNF contributions to several grain legumes and more recently to some shrub and forage legumes. The objective of this study was to investigate the application of this technique to the quantification of the contribution of BNF to the forage legume Desmodium ovalifolium by comparing the relative ureide abundance (RUA) of stem extracts of this plant with simultaneous estimates of BNF obtained using the ¹⁵N isotope dilution technique. The first experiment was performed in pots of soil, taken from a grazing study, amended with ¹⁵N-labelled organic matter at four different application rates. The ureide concentration in the stem extracts reflected the changes in BNF activity during plant growth and the RUA was closely correlated with the proportion of N derived from BNF as determined from the ¹⁵N technique (r ² = 0.86 and 0.88 for inoculated and non-inoculated plants, respectively). The use of a calibration curve derived from a previous study where the same legume was fed increasing concentrations of ¹⁵N labelled nitrate in sand/vermiculite culture, resulted in an over-estimation of the BNF contribution which may have been due to a significant uptake of ammonium from this acidic soil. The second experiment was performed in field plots and a good agreement was found between the estimates of BNF derived from using the ureide and ¹⁵N dilution techniques at two harvests six months apart. The uptake of soil N by the D. ovalifoliumand two forage grasses (Brachiaria humidicola and Panicum maximum) was estimated using an in situ soil core technique, and, while the uptake of N by the grasses was successfully estimated, this technique underestimated the N derived from the soil by the legume as determined by the ureide and ¹⁵N dilution techniques.     

Effect of climate on the recovery in crop and soil of15N-labelled fertilizer applied to wheat

Data was assembled from experiments on the fate of¹⁵N-labelled fertilizer applied to wheat (Triticum spp.) grown in different parts of the world. These data were then ranked according to the annual precipitation-evaporation quotient for each experimental location calculated from the average long-term values of precipitation and potential evaporation. Percentage recovery of¹⁵N fertilizer in crop and soil varied with location in accordance with the precipitation-evaporation quotient. In humid environments more¹⁵N fertilizer was recovered in the crop than in the soil, while in dry environments more¹⁵N fertilizer was recovered in the soil than in the crop. Irrespective of climatic differences between locations 20% (on average) of the¹⁵N fertilizer applied to wheat crops was unaccounted for at harvest. Most of the¹⁵N fertilizer remaining in the soil was found in the 0–30 cm layer. The most likely explanation of these differences is that wheat grown in dry environments has a greater root:shoot ratio than wheat grown in humid environments and, further, that the residue of dryland crops have higher C/N ratios. Both factors could contribute to the greater recovery of¹⁵N fertilizer in the soil in dry environments than in humid ones.     

1. The chemistry and biology of flooded soils in relation to the nitrogen economy in rice fields

Response of lowland rice to sources and methods of nitrogen fertilizer application were summarized for more than 100 experiments. In about 2/3 of the experiments, the yield increase per kg of fertilizer N was judged to be relatively poor with best split applications of urea. Based on frequency distribution, sulfur coated urea and urea briquets or urea supergranules deep placed more often produced satisfactory yield increases than best split urea, but even with these sources/methods the yield increases were judged to be relatively poor in about 1/2 of the experiments. There is an enormous potential to increase rice production with no further increases in inputs of fertilizer N if we could learn to match the best method/source of fertilizer with the soil-crop management complex.About 60% of the yields with no fertilizer N were in the range of 2 to 4 t/ha. Based on the average yield response to urea, this is equivalent to about 100 kg of urea N. It would appear worthwhile to study ways to improve utilization of soil nitrogen since it is already in place on the land and apparently in fairly abundant amounts in many soils.About 50 experiments with¹⁵N fertilizers were summarized. In almost all cases, the uptake of tagged fertilizer was less than the net increase in N in the above ground matter. In about 2/3 of the experiments, the addition of fertilizer N increased soil N uptake more than 20% and in 1/3 of the experiments the uptake of soil N was increased more than 40%. These results lead to much uncertainty about practical interpretation and use of¹⁵N data.     

Fate of nitrogen (15N) from oxamide and urea applied to turf grass: A lysimeter study

Slow release N fertilizers are receiving increasing attention for use on turf grass, but their fate in the plant-soil system is still poorly understood. We aimed to quantify the uptake and recovery of N by a mixture of grasses when applied as either urea or oxamide in different diameter granules using a tracer technique (¹⁵N). The effects of the N source on soil biomass, root density and amount of readily available organic C in soil were also evaluated.In a first experiment oxamide in 4–5 mm diameter granules was compared with urea. The initial N absorption, 40 days after fertilization (d.a.f.), was higher for urea (23.5%) than for oxamide (12.1%), but after 64 days absorption efficiencies were about the same (11%) for both fertilizers. Fertilizer-derived N lost by leaching was much greater from the urea-fertilized soil (1.57 g), compared with losses from oxamide-fertilized soil (0.05 g). The total residual fertilizer N remaining in the system at the end of the experiment was 26.7% of applied urea N and 39.6% of applied oxamide N. Cumulated absorption efficiencies, calculated after dismantling the lysimeters, were 43.1% for urea and 54.8% for oxamide (roots included). A priming effect caused by a larger uptake of soil N because of the better root development was found in the oxamide-treated lysimeter. Fertilization with oxamide also caused an increase in the amount of soil microbial biomass.In a second experiment, the efficiencies and fertilizer N uptake rates from oxamide applied at two different granule sizes (1–2 mm and 5–10 mm) were evaluated. The amount of soil N taken up by the grass was linearly related to root density (r = 0.92).     

Effect of the timing and rate of nitrogen fertilization on the growth and recovery of fertilizer nitrogen within the potato (Solanum tuberosum L.) crop

The effect of the timing of N fertilizer application on the uptake and partitioning of N within the crop and the yield of tubers has been studied in two experiments. In 1985 either none, 8 or 12 g N m^–2 was applied and in 1986 none, 12 or 18 g N m^–2. Fertilizer N was applied either at planting, around the time of tuber initiation or half at planting and the remainder in four foliar sprays of urea during tuber bulking.¹⁵N-labelled fertilizer was applied to measure the recovery of fertilizer N in the crops.There was an apparent pre-emergence loss of nitrate from the soil when N was applied at planting in 1986, thereby reducing the efficiency of fertilizer use. Applying the N at tuber initiation delayed and reduced the accumulation of N in the canopy compared with crops receiving all their fertilizer at planting. Foliar sprays of urea slightly increased both tuber yields and tuber N contents when compared to a single application at planting. The proportion of the fertilizer N recovered in the crop was little affected by the rate of N application, but a greater proportion of foliar-applied N was recovered than N broadcast at planting, due partly to pre-emergence losses of nitrate in 1986. It is suggested that late applications of N was foliar sprays can be of benefit to crops with a long growing season and reduce environmental losses of N.     

Effect of lime nitrogen on the efficiency of urea and other ammonium nitrogen fertilizers

Five pot experiments were conducted with wheat and rice in a net house to study the effect of lime nitrogen (LN, contains about 55% calcium cyanamide) amendment rates on the efficiency of urea, the recovery urea-¹⁵N, the efficiency of the three nitrogen fertilizers(NF), on the efficiency of urea in the three soils, and on NO ₃ ^- -N leaching from a flooded soil. A rate of LN-N of 5–8% of applied fertilizer N increased the recovery of labeled urea-N by 9.42%. The effect of LN on the efficiency of NF was urea > ammonium sulfate > ammonium chloride. Under flooded conditions, LN decreased NO ₃ ^- formation and leaching.Responses of several crops to LN amended fertilizers were also studied in field experiments. At equal NPK applications, the efficiency of basal applications to rice, wheat, corn, potatoes, soybean, peanut, grapes, peaches, melon and watermelon were bette r with LN than without. Efficiency with a basal fertilizer for rice or wheat with LN were the same as with the same fertilizer without LN applied in split applications.     

The effect of autumn applied 15N-labelled fertilizer on nitrate leaching in a cultivated soil during winter

Nitrogen (N) fertilizer applied in autumn to arable farm land raises concerns over affects on ground water quality. The contribution of autumn ¹⁵N-labelled fertilizer (50 kg N ha^-1) to nitrate leaching losses from a cultivated soil (silt loam on sandy loam; Udic Ustochrept) was measured using undisturbed monolith lysimeters (500 mm diameter, 700 mm long) during consecutive winters in Canterbury, New Zealand. The addition of ¹⁵N-labelled fertilizer at 50 kg N ha^-1 did not significantly increase nitrate leaching losses. Soil-derived-N contributed 78 and 88% (1996 and 1997, respectively) of the nitrate leached beneath fertilized lysimeters. Warmer weather and wetter soil conditions at cultivation and fertilizer application during 1997, compared with 1996, resulted in an increased release of soil-derived-N in 1997. Nitrate leaching and average nitrate concentrations were therefore 41% and 56% higher, respectively, during the winter of 1997 than the winter of 1996. However, fertilizer leaching losses were relatively consistent between years (7.8 and 8.6%). Although not statistically significant, total N leaching losses and average nitrate concentration were 24 to 30% higher below fertilized lysimeters as compared with unfertilized lysimeters, indicating a priming effect of fertilizer on soil N release. During both late winter periods, leachate nitrate concentrations from fertilized and unfertilized lysimeters exceeded World Health Organisation (WHO) limits for drinking water. Higher release of soil-derived-N in 1997 also meant WHO limits were exceeded for 6 weeks longer than in 1996. In conclusion, the application of ¹⁵N fertilizer in autumn directly contributed only a small proportion of the total amount of N leached in this cultivated soil. However, the apparent priming effect of autumn applied-N fertilizer has importance on the overall environmental impact of this production system, as the amount of N leached, and extent to which health limits were exceeded, was largely determined by the factors which controlled the release of soil-derived-N.     

Nitrogen fertilizer in leucaena alley cropping. I. Maize response to nitrogen fertilizer and fate of fertilizer-15N

Field microplot experiments were conducted in the semi-arid tropics of northern Australia to evaluate the response of maize (Zea mays L.) growth to addition of N fertilizer and plant residues and to examine the fate of fertilizer¹⁵N in a leucaena (Leucaena leucocephala) alley cropping system, in which supplemental irrigation was used. Leucaena prunings, maize residues and N fertilizer were applied to alley-cropped maize grown in microplots which were installed in the alleys formed by leucaena hedgerows spaced 4.5 metres apart. The¹⁵N-labelled fertilizer was used to examine the fate of fertilizer N applied in the presence of mulched leucaena prunings and maize residues.Application of leucaena prunings increased maize yield while addition of N fertilizer in the presence of the prunings produced a further increase in maize production. There was a significant positive interaction between N fertilizer and leucaena prunings in increasing maize production. The addition of maize residues in the presence of N fertilizer and leucaena prunings decreased maize yield and N uptake and increased fertilizer¹⁵N loss from 38% to 47%. Maize recovered 24–79% of fertilizer¹⁵N in one cropping season, depending on application rate of N fertilizer and field management of plant residues. About 20–34% of fertilizer¹⁵N remained in the soil. More than 37% of fertilizer¹⁵N was apparently lost from the soil and plant system largely through denitrification when N fertilizer was applied at 40 kg N ha^–1 or more in the presence or absence of plant residues. Application of N fertilizer improved maize yield and increased the contribution of mulched leucaena prunings to crop production in the alley cropping system.     

A conceptual assessment of the importance of denitrification as a source of soil nitrogen loss in tropical agro-ecosystems

This paper attempts to answer the question: is denitrification a major route of N loss from tropical agro-ecosystems? This question turns out to be very difficult to answer due to a severe shortage of data on this process for tropical agro-ecosystems other than rice. Given this lack of data, I approach this question by analyzing data on denitrification and nitrous oxide flux in tropical native forest and pasture soils and attempt to make some conclusions and pose some hypotheses about the significance of denitrification in tropical agricultural soils. I also briefly review methods for measuring denitrification. The data analysis suggests that denitrification in tropical forest soils is strongly influenced by the nature and amount of soil C and N turnover. Studies to examine differences in denitrification in different tropical agricultural systems should focus on the effects of system management on C and N turnover. The data analysis also suggests that, just as in temperate regions, denitrification may not be a significant route of N loss from most tropical agricultural systems. However, field studies are necessary to determine if this is actually the case.     

Efficiency of urea and ammonium sulfate for wetland rice grown on an acid sulfate soil as affected by rate and time of application

A pot experiment was conducted in a greenhouse to assess the effect of rate and time of N application on yield and N uptake of wetland rice grown on a Rangsit acid sulfate soil (Sulfic Tropaquepts). Response of rice at N rates of 800, 1600 and 2400 mg N/pot (5 kg of soil) was compared between urea and ammonium sulfate when applied at two times: (i) full-rate basal at transplanting and (ii) one half at transplanting and one half at the PI stage. In addition, labelled¹⁵N sources were applied either at transplanting or at the PI stage to determine the nitrogen balance sheet in the soil/plant system.No significant difference in grain and straw yields between urea and ammonium sulfate at low rate was observed. At the higher N rates, urea produced higher yields than did ammonium sulfate regardless of timing. The highest yields were obtained when urea at the high N rate was applied either in a single dose or a split dose while lowest yields were observed particularly when ammonium sulfate at the same rate was applied. Split application of N fertilizer was shown to be no better than a single basal application. The occurrence of nutritional disorder, a symptom likely reflected by high concentration of Fe (II) in combination with soluble Al, was induced with high rate of ammonium sulfate.In terms of fertilizer N recovery by using¹⁵N-labelling, ammonium sulfate was more efficient than urea when both were applied at transplanting. In contrast, application at the PI stage resulted in higher utilization of urea than of ammonium sulfate. The recovery of labelled N in the soil was higher with urea than with ammonium sulfate when the two sources were applied at transplanting, while the opposite result was obtained when the same fertilizers were applied at the PI stage. The losses from urea and ammonium sulfate were not different when these fertilizers were applied at transplanting but loss from urea was higher than that from ammonium sulfate when both were applied at the PI stage.     

The application of isotopic (32P and 15N) dilution techniques to evaluate the interactive effect of phosphate-solubilizing rhizobacteria, mycorrhizal fungi and Rhizobium to improve the agronomic efficiency of rock phosphate for legume crops

A pot experiment was designed to evaluate the interactive effects of multifunctional microbial inoculation treatments and rock phosphate (RP) application on N and P uptake by alfalfa through the use of ¹⁵N and ³²P isotopic dilution approaches. The microbial inocula consisted of a wild type (WT) Rhizobium meliloti strain, the arbuscular mycorrhizal (AM) fungus Glomus mosseae (Nicol. and Gerd.) Gerd. and Trappe, and a phosphate solubilizing rhizobacterium (Enterobacter sp.). Inoculated microorganisms were established in the root tissues and/or in the rhizosphere soil of alfalfa plants (Medicago sativa L.). Improvements in N and P accumulation in alfalfa corroborate beneficial effects of Rhizobium and AM interactions. Inoculation with selected rhizobacteria improved the AM effect on N or P accumulation in both the RP-added soil and in the non RP-amended controls. Measurements of the ¹⁵N/¹⁴N ratio in plant shoots indicate an enhancement of the N₂ fixation rates in Rhizobium-inoculated AM-plants, over that achieved by Rhizobium in non-mycorrhizal plants. Whether or not RP was added, AM-inoculated plants showed a lower specific activity (³²P/³¹P) than did their comparable non-mycorrhizal controls, suggesting that the plant was using otherwise unavailable P sources. The phosphate-solubilizing, AM-associated, microbiota could in fact release phosphate ions, either from the added RP or from the indigenous ``less-available' soil phosphate. A low Ca concentrations in the test soil may have benefited P solubilization. Under field conditions, the inoculation with AM fungi significantly increased plant biomass and N and P accumulation in plant tissues. Phosphate-solubilizing rhizobacteria improved mycorrhizal responses in soil dually receiving RP and organic matter amendments. Organic matter addition favoured RP solubilization. This, together with a tailored microbial inoculation, increased the agronomic efficiency of RP in the test soil that was Ca deficient at neutral pH.     

Plant recovery of 15N-labelled nitrogen applied to reed canary grass grown for biomass

Reed canary grass (Phalaris arundinacea L.) is apotential crop for production of bioenergy and biomass in northern Europe. In this study labelled ¹⁵N was used to follow the fate of applied N in roots and shoots of reed canary grass during a year. Two rates of¹⁵N fertiliser were applied in spring 1995 and 1996 to a clay (50 kg ha⁻¹ and 100 kg ha⁻¹) and an organic soil (30 kg ha⁻¹ and 60 kg ha⁻¹). The data did not indicate significant differences between recoveries of nitrogen following application of fertiliser at recommended and half of the recommended rates. The recovery of added N in shoots was highest at midsummer. The median values were 68% and 58% inorganic soil and 42% and 65% in clay soil, in 1995 and 1996respectively. Some of the N utilised by shoots was remobilised to the roots during autumn. The highest median recovery of applied N in roots was 19%in clay soil in October 1996, corresponding to a 13 percentage unit increase in recovery during autumn. In contrast, the lowest remobilisation was recorded after a rainy spring in clay soil, being only 3 percentage units. During winter the loss of N and fertiliser N from the shoots continued, and consequently the total N content in shoots was about half of that for autumn. In spring, one year after N application, the shoots contained 9–20% of applied N. The data suggest both intensive uptake and remobilisation of fertiliser N during over a year, following delayed harvest, and indicate the importance of the rhizome system in N turnover. This revised version was published online in August 2006 with corrections to the Cover Date.     

Effect of K on nitrogen fixation of intercrop groundnut and the competition between intercrop groundnut and maize

Application of adequate level of K has shown to improve the competitive ability of the legume in legume/grass mixtures. However, the effect of K on the competitive ability of grain legumes in legume/cereal intercropping systems has not been adequately studied. Hence, studies were made to ascertain if the effects of K could be exploited in improving the performance of groundnut (Arachis hypogaea L.) cv. No. 45 when intercropped with maize (Zea mays L.) cv. Badra. The study was conducted at the Faculty of Agriculture, University of Ruhuna, Kamburupitiya, Sri Lanka in 1988 in basins filled with 36 kg of soil. It involved establishing maize and groundnut as monocrops and as intercrops at three K levels viz. 0, 20 and 40 mg of K kg^–1 of soil. Monocrop maize and groundnut had 2 and 5 plants/basin, respectively while the intercrop had 1 maize plant and 3 groundnut plants/basin. The soil used was Red Yellow Podzolic which was tagged by incorporating¹⁵N-labelled plant material. When grown as a monocrop, K had no effect on the percent N derived from atmosphere, amount of N₂ fixed, dry matter production, pod yield and total N content of groundnut. However, when intercropped with maize lack of K application affected the above parameters significantly which was overcome by improving K level. Thus, the optimum level of K for groundnut was greater when intercropped than monocropped. A significant interaction between K level and cropping system was evident with regard to N₂ fixation, pod yield and total dry matter production of groundnut. Intercrop maize derived 30–35% of its N content from the associated groundnut plants which amounted to 13–22 mg N/plant. The amount of N supplied by groundnut to associated maize plant was not affected by K level. It appears that there is scope for alleviating growth depression of the legume component in legume/cereal intercropping systems by developing appropriate K fertilizer practices.