Does the white clover response to sulphur availability correspond to phenotypic or ontogenetic plasticity?

White clover (Trifolium repens L.) is a key species in grasslands. Its performance in grassland communities is strongly linked to nitrogen (N) availability. A decrease in soil sulphur (S) content has appeared in the last few decades in grasslands in Northern Europe and this could change the behaviour of white clover. S is essential for plants and particularly for legumes through its effect upon nitrogen fixation. The aim of this study was to determine the effects of S deficiency on white clover fitness, analysing its plasticity in a time course of growth.Three concentrations of SO₄^2?, “Low S” (0.009 mM), “Medium S” (0.384 mM) and “High S” (1.509 mM), were used to grow plants in a hydroponic system. S availability modified biomasses significantly only at the end of the experiment (11 weeks). Medium S appeared optimal while Low S induced a lower aboveground dry mass. An appropriate S availability (Medium S) not only increased S content but also increased N content by stimulating N₂ fixation. Plant growth analysis using growth fitted curves and the calculation of RGR revealed that S effects on biomass corresponded to the production of different phenotypes and not to a growth delay. This work shows that the acceleration growth phase (49–56 days) is a key period for the nutritional needs of white clover and should be the best period for a sulphur fertilisation regime that aims to enhance white clover fitness.     

Nitrogen fixation by white clover when competing with grasses at moderately low temperatures

It was the aim of this study to determine the way in which low temperature modifies the effect of a competing grass on nitrogen fixation of a forage legume. White clover (Trifolium repens L.) was grown in monoculture or in different planting ratios with timothy (Phleum pratense L.) or perennial ryegress (Lolium perenne L.) in growth chambers at either 7.5/5°C (LoT) or 15/10°C (HiT) average day/night temperatures, and with 2.5 or 7.5 mM ¹⁵N-labelled nitrate in the nutrient solution.Competition with grass led to a marked increase in the proportion of clover nitrogen derived from symbiosis (% N_sym). This increase was slower at LoT where % N_sym was reduced considerably; it was closely related to the reduction in the amount of available nitrate as a result of its being utilized by the grass.Nitrogen concentration in white clover herbage and dry matter yield per clover plant were reduced, for the most part, when a competing grass was present. The amount of nitrogen fixed per plant of white clover decreased markedly with temperature. Low temperature consequently accentuated competition for nitrate. The capacity of white clover to compete successfully was limited by its slower growth and nitrogen accumulation.     

Influence of dissimilarities in temporal and spatial N-uptake patterns on15N-based estimates of fixation and transfer of N in ryegrass-clover mixtures

The temporal N-uptake patterns of white clover (Trifolium repens L.) mixed with perennial ryegrass (Lolium perenne L.) and of red clover (Trifolium pratense L.) mixed with Italian ryegrass (Lolium multiflorum Lam.) were determined in successive harvests of herbage within the growth cycles of a ley established near Zürich (Switzerland). Rooting patterns were examined by injecting¹⁵N-fertilizer at soil depths ranging from 10 to 40 cm. The results were analyzed to determine the effect of variations in time and depth of N-uptake on the¹⁵N-based measurement of N from symbiosis (N_sym) and N from transfer (N_trans).Grasses in mixture appeared to have deeper rooting systems than grass monocultures, which led to an overestimation of N transfer from white clover to perennial ryegrass if¹⁵N was spread on the soil surface.White clover generally lagged behind grass in soil N- uptake. Soil N-uptake of red clover slowed down before that of the grass because % N_sym almost reached 100% during the second half of each growth cycle. However, the effect of these dissimilarities on the seasonal average of %N_sym did not exceed 2%.It is concluded that at the observed high levels of N₂ fixation, failure to account for the N-uptake patterns of the test and reference crops only slightly affected the estimates of % N_sym and % N_trans, and did not invalidate the observed differences between species.     

Short-term effects of a dung pat on N2 fixation and total N uptake in a perennial ryegrass/white clover mixture

The short-term effects of a simulated cattle dung pat on N₂ fixation and total uptake of N in a perennial ryegrass/white clover mixture was studied in a container experiment using sheep faeces mixed with water to a DM content of 13%. We used a new ¹⁵N cross-labelling technique to determine the influence of dung-pat N on N₂ fixation in a grass/clover mixture and the uptake of dung N in grass and clover. The proportion of N in clover derived from N₂ fixation (%Ndfa) varied between 88–99% during the 16 weeks following application of the dung. There was no effect of dung on the %Ndfa in clover grown in mixture, whereas the %Ndfa in clover grown in pure stand decreased (nominal 2–3%) after dung application. Dung did not influence the amount of N₂ fixed, and the uptake of dung N in grass and clover proceeded at an almost constant rate. After 16 weeks, 10% of the applied dung N was taken up by grass and clover, 57% had been incorporated in the soil by faunal activity and 27% remained in residual dung on the soil surface. The dung N unaccounted for (7%) was probably lost by ammonia volatilisation and denitrification. The uptake of dung N in grass/clover mixtures in the field was similarly followed by using simulated ¹⁵N-labelled dung pats. The total dry matter production and N yields increased in the 0–30 cm distance from the edge of the dung patch, but the proportion of clover decreased. Thirteen months after application of the dung 4% of the applied dung N was recovered in the harvested herbage, 78% was recovered from the soil and the residual dung, and 18% was not accounted for. It is concluded that N₂ fixation in the dung patch border area in grass/clover mixtures is not influenced directly by the release of N from dung pats in the short term. However the amount of N₂ fixed may be reduced, if the growth of clover is reduced in the patch border area.     

Improving N2 fixation from the plant down: Compatibility of Trifolium subterraneum L. cultivars with soil rhizobia can influence symbiotic performance

Plant genotypes of Trifolium subterraneum L. (subterranean clover) were evaluated for differences in symbiotic N₂ fixation with soil rhizobia, with the long-term aim of using plant selection to overcome sub-optimal N₂ fixation associated with poorly effective soil rhizobia. Symbiotic performance (SP) was assessed for 49 genotypes of subterranean clover with each of four pure Rhizobium strains isolated from soil. Plants were grown in N free media in the greenhouse and their shoot dry weights measured and expressed as a percentage of dry weight with R. leguminosarm bv. trifolii WSM1325, the recommended commercial inoculant. Average SP with two Rhizobium strains (H and J) ranged from completely ineffective to 80% of potential for the subterranean clover genotypes. Two clover cultivars with high (cv. Campeda) and low (cv. Clare) SP values were investigated in more detail. Campeda typically fixed more N₂ than Clare when inoculated with 30 soil extracts (4.2 vs 2.4 mg N₂ fixed/shoot) and with 14 pure strains isolated from those soils (4.2 vs 2.2 mg N₂ fixed/shoot). The poor performance of Clare could be attributed to interruptions at multiple stages of the symbiotic association, from nodule initiation (less nodules), nodule development (small, white nodules), through to reduced nodule function (N₂ fixed/mg nodule) depending on the inoculation treatment. Through the careful use of subterranean clover genotypes by plant breeders it should be possible to make significant gains in the SP of future subterranean clover cultivars.     

Nitrogen fixation by white clover in pastures grazed by dairy cows: Temporal variation and effects of nitrogen fertilization

Effects of rate of nitrogen (N) fertilizer and stocking rate on production and N₂ fixation by white clover (Trifolium repens L.) grown with perennial ryegrass (Lolium perenne L.) were determined over 5 years in farmlets near Hamilton, New Zealand. Three farmlets carried 3.3 dairy cows ha^–1 and received urea at 0, 200 or 400 kg N ha^–1 yr^–1 in 8–10 split applications. A fourth farmlet received 400 kg N ha^–1 yr^–1 and had 4.4 cows ha^–1.There was large variation in annual clover production and total N₂ fixation, which in the 0 N treatment ranged from 9 to 20% clover content in pasture and from 79 to 212 kg N fixed ha^–1 yr^–1. Despite this variation, total pasture production in the 0 N treatment remained at 75–85% of that in the 400 N treatments in all years, due in part to the moderating effect of carry-over of fixed N between years.Fertilizer N application decreased the average proportion of clover N derived from N₂ fixation (P_N; estimated by ¹⁵N dilution) from 77% in the 0 N treatment to 43–48% in the 400 N treatments. The corresponding average total N₂ fixation decreased from 154 kg N ha^–1 yr^–1 to 39–53 kg N ha^–1 yr^–1. This includes N₂ fixation in clover tissue below grazing height estimated at 70% of N₂ fixation in above grazing height tissue, based on associated measurements, and confirmed by field N balance calculations. Effects of N fertilizer on clover growth and N₂ fixation were greatest in spring and summer. In autumn, the 200 N treatment grew more clover than the 0 N treatment and N₂ fixation was the same. This was attributed to more severe grazing during summer in the 0 N treatment, resulting in higher surface soil temperatures and a deleterious effect on clover stolons.In the 400 N treatments, a 33% increase in cow stocking rate tended to decrease P_N from 48 to 43% due to more N cycling in excreta, but resulted in up to 2-fold more clover dry matter and N₂ fixation because lower pasture mass reduced grass competition, particularly during spring.     

The effects of elevated CO2 on symbiotic N2 fixation: a link between the carbon and nitrogen cycles in grassland ecosystems

The response of plants to elevated CO₂ is dependent on the availability of nutrients, especially nitrogen. It is generally accepted that an increase in the atmospheric CO₂ concentration increases the C:N ratio of plant residues and exudates. This promotes temporary N-immobilization which might, in turn, reduce the availability of soil nitrogen. In addition, both a CO₂ stimulated increase in plant growth (thus requiring more nitrogen) and an increased N demand for the decomposition of soil residues with a large C:N will result under elevated CO₂ in a larger N-sink of the whole grassland ecosystem. One way to maintain the balance between the C and N cycles in elevated CO₂ would be to increase N-import to the grassland ecosystem through symbiotic N₂ fixation. Whether this might happen in the context of temperate ecosystems is discussed, by assessing the following hypothesis: i) symbiotic N₂ fixation in legumes will be enhanced under elevated CO₂, ii) this enhancement of N₂ fixation will result in a larger N-input to the grassland ecosystem, and iii) a larger N-input will allow the sequestration of additional carbon, either above or below-ground, into the ecosystem. Data from long-term experiments with model grassland ecosystems, consisting of monocultures or mixtures of perennial ryegrass and white clover, grown under elevated CO₂ under free-air or field-like conditions, supports the first two hypothesis, since: i) both the percentage and the amount of fixed N increases in white clover grown under elevated CO₂, ii) the contribution of fixed N to the nitrogen nutrition of the mixed grass also increases in elevated CO₂. Concerning the third hypothesis, an increased nitrogen input to the grassland ecosystem from N₂ fixation usually promotes shoot growth (above-ground C storage) in elevated CO₂. However, the consequences of this larger N input under elevated CO₂ on the below-ground carbon fluxes are not fully understood. On one hand, the positive effect of elevated CO₂ on the quantity of plant residues might be overwhelming and lead to an increased long-term below-ground C storage; on the other hand, the enhancement of the decomposition process by the N-rich legume material might favour carbon turn-over and, hence, limit the storage of below-ground carbon.     

Interactions between white clover and ryegrass under contrasting nitrogen availability: N2 fixation,N fertilizer recovery,N transfer and water use efficiency

Seasonal variation in N₂ fixation, N transfer from clover to ryegrass, and soil N absorption in white clover–ryegrass swards were investigated under field conditions over three consecutive years. The plots were established with different seeding ratios of clover and ryegrass and contrasting fertilizer N ranging from 3 to 72 kg ha^-1 year^-1.An initially poor clover population needed at least one growing season to reach the same yield output as an initially well established clover population. The clover content of the sward decreased by the annual application of 72 kg N ha^-1 but not by smaller N dressings.The total amount of atmospherically derived N in clover growing in mixture with ryegrass was, on average over the three years equal to 83, 71, 68 and 60 kg N ha^-1 for the treatments of 3, 24, 48 and 72 kg N ha^-1, respectively. The proportion of atmospherically derived N declined with increasing N application, but never became smaller than 80% of total clover N. The proportion of atmospherically derived N in a pure stand white clover amounted to 60–80% of the total N content, equivalent to 109, 110, 103 and 90 kg N ha^-1 for the treatments of 3, 24, 48 and 72 kg N ha^-1, respectively.Only small amounts of atmospherically derived N was transferred to the associated ryegrass during the first production year, while in each of the following years up to 21 kg ha^-1 was transferred. The average amount of N transferred from clover to ryegrass was equivalent to 3, 16 and 31% of the N accumulated in ryegrass in the first, second and third production year, respectively. Expressed relative to the total amount of fixed N₂ in the clover–ryegrass mixture, the transfer amounted to 3, 17 and 22% in the first, second and third production year, respectively. Thus transfer of atmospherically derived N from clover contributed significantly to the N economy of the associated ryegrass.The clover–ryegrass mixture absorbed constantly higher amount of soil derived N than the pure stands of the two species. Only 11% of the total accumulated fertilizer N and soil derived N in the mixture was contained within the clover component. Lower water use efficiencies for the plants grown in mixture compared to pure stands were mainly related to the increased N uptake in the mixture, with the subsequent increase in growth compared to the pure stands.It is concluded that positive interactions between clover and ryegrass growing in mixture ensure a more efficient fixation of atmospheric N₂ and absorption of fertilizer N and soil derived N than pure stands of the same species.     

Estimates of seasonal nitrogen fixation of annual subterranean clover-based pastures using the15N natural abundance technique

Annual pasture legumes play a key role in ley farming systems of southern Australia, providing biologically fixed nitrogen (N) to drive the production of the pastures as well as subsequent crops grown in rotation. Seasonal inputs of biologically fixed N in shoot biomass of the subterranean clover (Trifolium subterraneum) component of grazed annual pastures were assessed using the¹⁵N natural abundance technique and appropriately timed sampling of herbage dry matter (DM) for N accumulation. At three study sites spanning a gradient across the Western Australian wheatbelt from 300 to 600 mm annual rainfall the performance of the clover and non-legume herbs and grasses was examined as paired comparisons involving two management treatments expected to give contrasting effects on pasture productivity, botanical composition and N₂ fixation. The proportion of clover N derived from atmospheric N₂ fixation (%Ndfa) ranged from 65 to 95% across sites, treatments and sampling times. Amounts of fixed N accumulated in clover shoot biomass ranged from 50 to 125 kg ha⁻¹, and paralleled trends in clover production. Substantial increases in pasture production in high yielding treatments generally occurred without decrease in %Ndfa, suggesting that N₂ fixation was essentially non-limiting to performance of the clover component. Seasonal profiles for accumulation of fixed N were skewed towards the late winter and spring period, particularly in low plant density pastures following a cereal crop. There were seasonal, site and treatment-specific effects on the proportion of clover and non-legume pasture components and consequently clover yield and N₂ fixation were variably affected by competition from non-legume species.     

Effect of light and atmospheric carbon dioxide concentration on nitrogen fixation by herbage legumes

Summary Lucerne, red clover and white clover were grown at two atmospheric concentrations of CO₂ (300 and 1000 μl l⁻¹) and the effects on N₂ fixation, nodule mass/number and root/shoot dry matter production determined. Pea plants were similarly evaluated as a comparison with grain legumes. CO₂ enrichment increased N₂ fixation activity in all cases but activity/unit nodule mass was significantly increased only in the pea. The enhancement of N₂ fixation in herbage legumes by CO₂ enrichment reflected an increase in nodule mass which in turn was attributed to increased nodule number, and results show that under the experimental conditions obtaining here photosynthate supply did not limit nodule N₂ fixation in these plants though it was limiting in the case of peas. White clover growing in a 6 and 14 hour photoperiod was studied for response of the N₂ fixing system to light. Long photoperiod (14 hour) plants assayed at constant temperature (20°C) did not show a significant response to light at the end of the dark period either in terms of fixation per plant or per unit nodule mass, in contrast with short photoperiod (6 hour) plants which showed significant responses. Short photoperiod plants compensated for reduced photosynthates by maintaining only half the root nodule mass and fixation activity of 14 hour photoperiod plants though plants in both systems supported similar rates of N₂ fixation per unit mass of nodule during the photoperiod. Comparison of N₂ fixation activities in whole and decapitated plant systems indicates the importance of shoot reserves for sustaining nitrogenase activity in white clover during short-term interruption of photosynthesis. These results support the conclusion of the CO₂ enrichment studies, that herbage legumes have the potential for supplying their nodule photosynthate requirements for sustaining optimum rates of N₂ fixation and excess carbon supply is used solely to promote further nodulation. Nodules of short photoperiod white clover plants were less efficient in N₂ fixation in that they evolved more H₂ relative to N₂ (C₂H₂) reduced than did long photoperiod plants.     

The effect of a single application of cow urine on annual N2 fixation under varying simulated grazing intensity, as measured by four 15N isotope techniques

The effects of dairy cow urine and defoliation severity on biological nitrogen fixation and pasture production of a mixed ryegrass-white clover sward were investigated over 12 months using mowing for defoliation. A single application of urine (equivalent to 746 kg N ha^–1), was applied in late spring to plots immediately after light and moderately-severe defoliation (35 mm and 85 mm cutting heights, respectively) treatments were imposed. Estimates of percentage clover N derived from N₂ fixation (%Ndfa) were compared by labelling the soil with ¹⁵N either by applying a low rate of ¹⁵N-labelled ammonium sulphate, immobilising ¹⁵N in soil organic matter, adding ¹⁵N to applied urine, or by utilising the small differences in natural abundance of ¹⁵N in soil. Urine application increased annual grass production by 85%, but had little effect on annual clover production. However, urine caused a marked decline in %Ndfa (using an average of all ¹⁵N methods) from 84% to a low of 22% by 108 days, with recovery to control levels taking almost a year. As a result, total N fixed (in above ground clover herbage) was reduced from 232 to 145 kg N ha^–1 yr^–1. Moderately–severe defoliation had no immediate effect on N₂ fixation, but after 108 days the %Ndfa was consistently higher than light defoliation during summer and autumn, and increased by up to 18%, coinciding with an increase in growth of weeds and summer-grass species. Annual N₂ fixation was 218 kg N ha^–1 yr^–1 under moderately-severe defoliation compared to 160 kg N ha^–1 yr^–1 under light defoliation. Estimates of %Ndfa were generally similar when ¹⁵N-labelled or immobilised ¹⁵N were used to label soil regardless of urine and defoliation severity. The natural abundance technique gave highly variable estimates of %Ndfa (–56 to 24%) during the first 23 days after urine application but, thereafter, estimates of %Ndfa were similar to those using ¹⁵N-labelling methods. In contrast, in urine treated plots the use of ¹⁵N-labelled urine gave estimates of %Ndfa that were 20–30% below values calculated using conventional ¹⁵N-labelling during the first 161 days. These differences were probably due to differences in the rooting depth between ryegrass and white clover in conjunction with treatment differences in ¹⁵N distribution with depth. This study shows that urine has a prolonged effect on reducing N₂ fixation in pasture. In addition, defoliation severity is a potential pasture management tool for strategically enhancing N₂ fixation.     

Dinitrogen fixation in white clover grown in pure stand and mixture with ryegrass estimated by the immobilized 15N isotope dilution method

Dinitrogen fixation in white clover (Trifolium repens L.) grown in pure stand and mixture with perennial ryegrass (Lolium perenne L.) was determined in the field using ¹⁵N isotope dilution and harvest of the shoots. The apparent transfer of clover N to perennial ryegrass was simultaneously assessed. The soil was labelled either by immobilizing ¹⁵N in organic matter prior to establishment of the sward or by using the conventional labelling procedure in which ¹⁵N fertilizer is added after sward establishment. Immobilization of ¹⁵N in the soil organic matter has not previously been used in studies of N₂ fixation in grass/clover pastures. However, this approach was a successful means of labelling, since the ¹⁵N enrichment only declined at a very slow rate during the experiment. After the second production year only 10–16% of the applied ¹⁵N was recovered in the harvested herbage. The two labelling methods gave, nonetheless, a similar estimate of the percentage of clover N derived from N₂ fixation. In pure stand clover, 75–94% of the N was derived from N₂ fixation and in the mixture 85–97%. The dry matter yield of the clover in mixture as percentage of total dry matter yield was relatively high and increased from 59% in the first to 65% in the second production year. The average daily N₂ fixation rate in the mixture-grown clover varied from less than 0.5 kg N ha⁻¹ day⁻¹ in autumn to more than 2.6 kg N ha⁻¹ day⁻¹ in June. For clover in pure stand the average N₂ fixation rate was greater and varied between 0.5 and 3.3 kg N ha⁻¹ day⁻¹, but with the same seasonal pattern as for clover in mixture. The amount of N fixed in the mixture was 23, 187 and 177 kg N ha⁻¹ in the seeding, first and second production year, respectively, whereas pure stand clover fixed 28, 262 and 211 kg N ha⁻¹ in the three years. The apparent transfer of clover N to grass was negligible in the seeding year, but clover N deposited in the rhizosphere or released by turnover of stolons, roots and nodules, contributed 19 and 28 kg N ha⁻¹ to the grass in the first and second production year, respectively. This revised version was published online in June 2006 with corrections to the Cover Date.     

Grassland species show similar strategies for sulphur and nitrogen acquisition

Backgrounds and aims

Plant nutrition strategies play a crucial role in community structure and ecosystem functioning. However, these strategies have been established only for nitrogen (N) acquisition, and it is not known whether similar strategies hold for other macronutrients such as sulphur (S). The aim of our study was to determine whether strategies for S acquisition of some grassland species were similar to those observed for N acquisition, and to analyse the relationships between these plant strategies and the soil microbial activity involved in soil organic S mineralisation.

Methods

We used three exploitative and three conservative grass species grown with and without S fertilisation. We measured a set of plant traits, namely root and shoot biomass, leaf area, root length, N and S content, leaf nutrient use efficiency, and sulphate uptake rates in plants, and one microbial trait linked to S mineralisation, namely soil arylsulphatase activity.

Results

The set of plant traits differentiated exploitative from conservative species. Close relationships were found between traits associated with strategies for N acquisition, namely total N content and Leaf N Use Efficiency (LNUE), and traits associated with strategies for S acquisition, namely total S content and Leaf S Use Efficiency (LSUE). Exploitative species exhibited similar or lower sulphate uptake capacities per unit of biomass than conservative species, but acquired more S through their larger root systems. Greater arylsulphatase activity was observed in the rhizosphere of the most exploitative species.

Conclusion

Overall, our results show that nutrient strategies defined in grassland species for N acquisition can be extended to S.     

Effects of white clover (Trifolium repens L.) on plant and soil nitrogen and soil organic matter in mixtures with perennial ryegrass (Lolium perenne L.)

To increase our insight into the above- and belowground N flows in grass and grass-clover swards relations between crop and soil parameters were studied in a cutting trial with perennial ryegrass (Lolium perenne) monocultures and ryegrass–white clover (Trifolium repens) mixtures. The effects of clover cultivar on herbage yield, the amount of clover-derived nitrogen, apparent N transfer to companion grass, dynamics of N and organic matter in the soil were estimated.The grass monocultures had very low DM yields (<2.1 t ha^-1) and a low N concentration in the harvested herbage. During 1992–1995 the annual herbage DM yield in the mixtures ranged from 7.0 to 14.3 t ha^-1, the white clover DM yield from 2.4 to 11.2 t ha^-1 and the mean annual clover content in the herbage DM harvested from 34 to 78%. Mixtures with the large-leaved clover cv. Alice yielded significantly more herbage and clover DM and had a higher clover content than mixtures with small/medium-leaved cvs. Gwenda and Retor. Grass cultivar did not consistently affect yield, botanical composition or soil characteristics.The apparent N₂ fixation was very high, ranging from 150 to 545 kg N ha^-1 in the different mixtures. For each tonne of clover DM in the harvested herbage 49 to 63 kg N was harvested, while the apparent N transfer from clover to grass varied between 55 and 113 kg N ha^-1 year^-1.The net N mineralization rate was lower under monocultures than under mixtures. The C mineralization and the amounts of C and N in active soil organic matter fractions were similar for monocultures and mixtures, but the C:N ratio of the active soil organic matter fractions were higher under grass than under mixtures. This explains the lower N mineralization under grass.     

The effect of source,concentration and time of application of nitrogen on the growth,nodulation and nitrogen fixation ofLotus pedunculatus andTrifolium repens

Summary Studies under growth cabinet conditions investigated the effect of source and concentration of nitrogen and timing of nitrogen application on the growth and nitrogen fixation byLotus pedunculatus cv. Maku andTrifolium repens cv. S184. KNO₃, NaNO₃ and NH₄NO₃ were added at transplanting at the following rates: 3.33, 7.78 and 13.33 mg N/plant. KNO₃ was added at 3.33 and 7.78 mg N/plant at 0, 6, 12, 18, 24 or 30 days after transplanting.Lotus shoot weight increased with all increasing nitrogen sources but clover only responded to KNO₃ and NaNO₃. The root weight of both species increased with increasing KNO₃ and NH₄NO₃. The percentage increase in lotus and clover shoot growth was greater than that of root growth when KNO₃ was added within a week of transplanting. Increases in growth by both species resulted from added nitrogen except with lotus when NaNO₃ was applied where increased nitrogen fixation also contributed to increased growth.Weight and number of effective nodules on both species were increased with 3.33 mg N per plant as KNO₃ but nitrogen fixation was not affected. Addition of 13.33 mg N as NaNO₃ reduced weight and number of effective nodules in both species and also nitrogen fixation by lotus.KNO₃ increased growth and nodulation of both species when applied within one week after transplanting. Nodulated lotus plants responded to KNO₃ by increasing growth but not nodulation.KNO₃ appeared to affect infection and development of nodules on lotus and may affect the growth of existing nodules on clover.     

Interactions between perennial ryegrass ( Lolium perenne L.) and white clover (Trifolium repens L.) under contrasting nitrogen availability: productivity,seasonal patterns of species composition,N2 fixation,N transfer and N recovery

Nitrogen (N) fertiliser and clover cultivar choice affect competition and productivity in grass-clover mixtures. Pure stands and mixtures of perennial ryegrass and white clover cultivars with contrasting growth habits were examined. The aim of this work was to study the effect of repetitive nitrogen (N) application and cultivar combination on competition and productivity, N yield in the harvested herbage, N₂ fixation in mixtures and pure stands, and transfer of N from clover to the companion grass. Large-leaved white clover cultivar Alice and small-leaved cv. Gwenda and perennial ryegrass cvs. Barlet (erect) and Heraut (prostrate) were sown in pure stands and as four binary grass-clover mixtures on a sandy soil in 1995. In the mixtures, two levels of N fertiliser were applied: 0 (-N) and 150 and 180 kg ha^-1 y^-1 N (+N) in 1996 and 1997, respectively, while the grass monocultures received three N levels (0, 140/180 and 280/360 kg ha^-1) in 1996 and 1997, respectively. No N was applied to pure clover. The plots were cut five times during 1996 and six times during 1997. Fertiliser N was applied in early spring and after every harvest. The treatments were continued until the summer of 1999. In pure grass, the applied N was effectively recovered. In mixtures, N application affected competition by enhancing grass growth and the overall effect of N application was 17 kg DM per kg N applied in 1996. However, there was no yield response to N fertilizer in 1997, because this was compensated for by a higher clover production in unfertilised mixtures. In 1997, -N mixtures yielded more N than +N mixtures, owing to the higher clover content and N₂ fixation. Large-leaved clover cv. Alice was better able to withstand the negative effect of repetitive N application on clover production in mixtures and increased its proportion during the growing season of the second harvest year. In 1997, mixtures with Alice yielded more N than mixtures with Gwenda, but in pure clover swards, there was no cultivar effect on N yield. Also, during the autumn of 1998 and the spring of 1999, the clover content was highest in mixtures with Alice. Harvested N and apparent N₂ fixation were almost twice as high in 1997 as in 1996. N yield and apparent N₂ fixation were higher in pure clover than in mixtures. In mixtures, the apparent N₂ fixation in 1996 was 142 kg N ha^-1, irrespective of cultivar or N treatment. In 1997, it was on average 337 kg N ha^-1, and higher in -N mixtures and in mixtures with Alice. For each tonne of clover DM in the harvested herbage, 65 and 57 kg N was harvested in 1996 and 1997 in -N mixtures, respectively. The apparent transfer of clover-derived N to grass was on average 29 and 70 kg N ha^-1 yr^-1 in 1996 and 1997, respectively. It was highest in +N mixtures and highest in mixtures with Gwenda in 1997. In contrast to clover, the grass cultivars were very similar in their productivity and seasonal patterns, despite their contrasting growth habits. Seasonal trends in N yield, N transfer and N recovery are discussed in relation to fertilizer application regimes and variation in production patterns in mixtures and pure stands. This revised version was published online in June 2006 with corrections to the Cover Date.     

Effects of some fungicides used against cereal pathogens on the growth of Rhizobium trifolii and its capacity to fix nitrogen in white clover

Fungicide residues in soils may affect nitrogen fixation by legumes. Effects of nine systemic fungicides, used against cereal pathogens, on Rhizobium trifolii and white clover were measured. Fenarimol and oxycarboxin inhibited growth of R. trifolii in vitro. The weight of clover plants after 10 weeks' growth in soil containing carboxin, oxycarboxin, benodanil, tridemorph and pyracarbolid was reduced. Symbiotic nitrogen fixation was decreased by carboxin, oxycarboxin and tridemorph present in soil at concentrations somewhat greater than that likely to result from several applications of the fungicide. Only tridemorph reduced nodulation. None of the compounds seems likely to affect nitrogen fixation in the field if applied at the recommended rate.     

Assimilate Partitioning in Red and White Clover Either Dependent on N2 Fixation in Root Nodules or Utilizing Nitrate Nitrogen

During vegetative growth in controlled environments, the patternof distribution of ¹⁴C-labelled assimilates to shoot and root,and to the meristems of the shoot, was measured in red and whiteclover plants either wholly dependent on N₂ fixation in rootnodules or receiving abundant nitrate nitrogen but lacking nodules. In experiments where single leaves on the primary shoot wereexposed to ¹⁴CO₂, nodulated plants of both clovers generallyexported more of their labelled assimilates to root (+nodules),than equivalent plants utilizing nitrate nitrogen, and thiswas offset by reduced export to branches (red clover) or stolons(white clover). The intensity of these effects varied with experiment.The export of labelled assimilate to growing leaves at the terminalmeristem of the donor shoot was not influenced by source ofnitrogen. Internode elongation in the donor shoot utilized nolabelled assimilate. Whole plants of white clover exposed to ¹⁴CO₂ on seven occasionsover 32 days exhibited the same effect on export to root (+nodules),which increased slightly in intensity with increasing plantage. Nodulated plants had larger root: shoot ratios than theirequivalents utilizing nitrate nitrogen. Trifolium repens, Trifolium pratense, red clover, white clover, nitrogen fixation, nitrate utilization, assimilate partitioning     

Dynamic phenotypic plasticity for root growth in Polygonum: a comparative study

Species differences in patterns of phenotypic plasticity may be an important aspect of adaptive diversity. Plasticity for functionally important root traits was studied in inbred field lineages of Polygonum persicaria and P. cespitosum (Polygonaceae). Replicate seedlings were grown in plexiglass rhizotrons under a range of constant and temporally variable moisture treatments. Plasticity was determined for final whole-plant biomass, root biomass allocation, and absolute and proportional root length. The dynamic aspect of root plasticity was examined by digitizing weekly tracings of the proportional deployment of each plant's root system to different vertical soil layers. Plants of both species expressed significant functionally adaptive phenotypic plasticity in the relative allocation, length, and vertical deployment of root systems in response to contrasting moisture conditions. Plasticity patterns in these closely related species were in general qualitatively similar, but for most traits differed in the magnitude and/or the timing of the plastic response. Dynamic changes in root deployment were more marked as well as faster in P. persicaria. Species differences in patterns of individual plasticity were generally consistent with the broader ecological distribution of P. persicaria in diverse as well as temporally variable moisture habitats.     

Nitrogen fixation,transfer and turnover in upland and lowland grass-clover swards,using15N isotope dilution

The stable isotope¹⁵N is particularly valuable in the field for measuring N fixation by isotope dilution. At the same time other soil-plant processes can be studied, including¹⁵N recovery, and nitrogen transfer between clover and grass. Three contrasting sites and soils were used in the present work: a lowland soil, an upland soil, and an upland peat. Nitrogen fixation varied from 12 gm^–2 on lowland soil to 2.7 gm^–2 on upland peat. Most N transfer occurred on upland soil (4.2 gm^–2) which, added to nitrogen fixed, made a total of 8.7 gm² input during summer 1985.¹⁵N recovery for the whole experiment was small, around 25%.Measurement of dead and dying leaves, stubble and roots, suggests that plant organ death is the first stage in N transfer from white clover to ryegrass, through the decomposer cycle. Decomposition was fastest on lowland soils, slowest on peat. On lowland soil this decomposer nitrogen is apparently subverted before transfer, probably by soil microbes.Variations in natural abundance of¹⁵N in plants were found in the two species on the different soils. These might be used to measure nitrogen fixation without adding isotope, but the need for many replicates and repeat samples would limit throughput.