Estimation of symbiotic N2 fixation in an Amazon floodplain forest

Sustainable management for existing Amazonian forests requires an extensive knowledge about the limits of ecosystem nutrient cycles. Therefore, symbiotic nitrogen (N₂) fixation of legumes was investigated in a periodically flooded forest of the central Amazon floodplain (Várzea) over two hydrological cycles (20 months) using the ¹⁵N natural abundance method. No seasonal variation in ¹⁵N abundance (δ ¹⁵N values) in trees which would suggest differences in N₂ fixation rates between the terrestrial and the aquatic phase was found. Estimations of the percentage of N derived from atmosphere (%Ndfa) for the nodulated legumes with Neptunia oleracea on the one side and Teramnus volubilis on the other resulted in mean %Ndfa values between 9 and 66%, respectively. More than half of the nodulated legume species had %Ndfa values above 45%. These relatively high N gains are important for the nodulated legumes during the whole hydrological cycle. With a %Ndfa of 4–5% for the entire Várzea forest, N₂ fixation is important for the ecosystem and therefore, has to be taken into consideration for new sustainable land-use strategies in this area.     

Alder and lupine enhance nitrogen cycling in a degraded forest soil in Northern Sweden

Positive effects of legumes and actinorhizal plants on N-poor soils have been observed in many studies but few have been done at high latitudes, which was the location of our study. We measured N₂ fixation and several indices of soil N at a site near the Arctic Circle in northern Sweden. More than 20 years ago lupine (Lupinus nootkatensis Donn) and gray alder (Alnus incana L. Moench) were planted on this degraded forest site. We measured total soil N, net N mineralization and nitrification with a buried bag technique, and fluxes of NH⁺ ₄ and NO^– ₃ as collected on ion exchange membranes. We also estimated N₂ fixation activity of the N₂-fixing plants by the natural abundance of ¹⁵N of leaves with Betula pendula Roth. as reference species. Foliar nitrogen in the N₂-fixing plants was almost totally derived from N₂ fixation. Plots containing N₂-fixing species generally had significantly higher soil N and N availability than a control plot without N₂-fixing plants. Taken together, all measurements indicated that N₂-fixing plants can be used to effectively improve soil fertility at high latitudes in northern Sweden.     

Biological nitrogen fixation: Investments,expectations and actual contributions to agriculture

Inputs of biologically fixed N into agricultural systems may be derived from symbiotic relationships involving legumes and Rhizobium spp., partnerships between plants and Frankia spp. or cyanobacteria, or from non-symbiotic associations between free-living diazotrophs and plant roots. It is assumed that these N₂-fixing systems will satisfy a large portion of their own N requirements from atmospheric N₂, and that additional fixed N will be contributed to soil reserves for the benefit of other crops or forage species. This paper reviews the actual levels of N₂ fixation attained by legume and non-legume associations and assesses their role as a source of N in tropical and sub-tropical agriculture. We discuss factors influencing N₂ fixation and identify possible strategies for improving the amount of N₂ fixed.     

Interactions among nitrogen fixation and soil phosphorus acquisition strategies in lowland tropical rain forests

Paradoxically, symbiotic dinitrogen (N₂) fixers are abundant in nitrogen (N)‐rich, phosphorus (P)‐poor lowland tropical rain forests. One hypothesis to explain this pattern states that N₂ fixers have an advantage in acquiring soil P by producing more N‐rich enzymes (phosphatases) that mineralise organic P than non‐N₂ fixers. We assessed soil and root phosphatase activity between fixers and non‐fixers in two lowland tropical rain forest sites, but also addressed the hypothesis that arbuscular mycorrhizal (AM) colonisation (another P acquisition strategy) is greater on fixers than non‐fixers. Root phosphatase activity and AM colonisation were higher for fixers than non‐fixers, and strong correlations between AM colonisation and N₂ fixation at both sites suggest that the N–P interactions mediated by fixers may generally apply across tropical forests. We suggest that phosphatase enzymes and AM fungi enhance the capacity of N₂ fixers to acquire soil P, thus contributing to their high abundance in tropical forests.     

Global inputs of biological nitrogen fixation in agricultural systems

Biological dinitrogen (N₂) fixation is a natural process of significant importance in world agriculture. The demand for accurate determinations of global inputs of biologically-fixed nitrogen (N) is strong and will continue to be fuelled by the need to understand and effectively manage the global N cycle. In this paper we review and update long-standing and more recent estimates of biological N₂ fixation for the different agricultural systems, including the extensive, uncultivated tropical savannas used for grazing. Our methodology was to combine data on the areas and yields of legumes and cereals from the Food and Agriculture Organization (FAO) database on world agricultural production (FAOSTAT) with published and unpublished data on N₂ fixation. As the FAO lists grain legumes only, and not forage, fodder and green manure legumes, other literature was accessed to obtain approximate estimates in these cases. Below-ground plant N was factored into the estimations. The most important N₂-fixing agents in agricultural systems are the symbiotic associations between crop and forage/fodder legumes and rhizobia. Annual inputs of fixed N are calculated to be 2.95 Tg for the pulses and 18.5 Tg for the oilseed legumes. Soybean (Glycine max) is the dominant crop legume, representing 50% of the global crop legume area and 68% of global production. We calculate soybean to fix 16.4 Tg N annually, representing 77% of the N fixed by the crop legumes. Annual N₂ fixation by soybean in the U.S., Brazil and Argentina is calculated at 5.7, 4.6 and 3.4 Tg, respectively. Accurately estimating global N₂ fixation for the symbioses of the forage and fodder legumes is challenging because statistics on the areas and productivity of these legumes are almost impossible to obtain. The uncertainty increases as we move to the other agricultural-production systems—rice (Oryza sativa), sugar cane (Saccharum spp.), cereal and oilseed (non-legume) crop lands and extensive, grazed savannas. Nonetheless, the estimates of annual N₂ fixation inputs are 12–25 Tg (pasture and fodder legumes), 5 Tg (rice), 0.5 Tg (sugar cane), <4 Tg (non-legume crop lands) and <14 Tg (extensive savannas). Aggregating these individual estimates provides an overall estimate of 50–70 Tg N fixed biologically in agricultural systems. The uncertainty of this range would be reduced with the publication of more accurate statistics on areas and productivity of forage and fodder legumes and the publication of many more estimates of N₂ fixation, particularly in the cereal, oilseed and non-legume crop lands and extensive tropical savannas used for grazing.     

Taxonomic identity determines N2 fixation by canopy trees across lowland tropical forests

Legumes capable of fixing atmospheric N₂ are abundant and diverse in many tropical forests, but the factors determining ecological patterns in fixation are unresolved. A long‐standing idea is that fixation depends on soil nutrients (N, P or Mo), but recent evidence shows that fixation may also differ among N₂‐fixing species. We sampled canopy‐height trees across five species and one species group of N₂‐fixers along a landscape P gradient, and manipulated P and Mo to seedlings in a shadehouse. Our results identify taxonomy as the major determinant of fixation, with P (and possibly Mo) only influencing fixation following tree‐fall disturbances. While 44% of trees did not fix N₂, other trees fixed at high rates, with two species functioning as superfixers across the landscape. Our results raise the possibility that fixation is determined by biodiversity, evolutionary history and species–specific traits (tree growth rate, canopy stature and response to disturbance) in the tropical biome.     

Natural abundance of 15N in tropical plants with emphasis on tree legumes

Natural abundance of ¹⁵N ( ¹⁵N) of leaves harvested from tropical plants in Brazil and Thailand was analyzed. The ¹⁵N values of non-N₂-fixing trees in Brazil were +4.5±1.9, which is lower than those of soil nitrogen (+8.0±2.2). In contrast, mimosa and kudzu had very low ¹⁵N values (–1.4+0.5). The ¹⁵N values of Panicum maximum and leguminous trees, except Leucaena leucocephala, were similar to those of non-N₂-fixing trees, suggesting that the contribution of fixed N in these plants is negligible. The ¹⁵N values of non-N₂-fixing trees in Thailand were +4.9±2.0. Leucaena leucocephala, Sesbania grandiflora, Casuarina spp. and Cycas spp. had low ¹⁵N values, close to the value of atmospheric N₂ (0), pointing to a major contribution of N₂ fixation in these plants. Cassia spp. and Tamarindus indica had high ¹⁵N values, which confirms that these species are non-nodulating legumes. The ¹⁵N values of Acacia spp. and Gliricidia sepium and other potentially nodulating tree legumes were, on average, slightly lower than those of non-N₂-fixing trees, indicating a small contribution of N₂ fixation in these legumes.     

Nitrogen fixation in soybean as influenced by cultivar and Rhizobium strain

Summary The¹⁵N-substratum labeling technique and other indirect methods were used to compare nitrogen (N₂) fixation in soybean varieties grown in the field in Greece and Romania. Significant variation in the amount (Ndfa) and proportion of N derived from fixation (% Ndfa) was found in different varieties. With 20 kg N/ha applied to soil, N₂ fixed ranged from 22 to 236 kg N/ha in Greece and from 17 to 132 kg N/ha in Romania. In general, varieties or treatments with higher dry matter yield supported greater fixation. Also, varieties with high Ndfa had high % Ndfa andvice versa. Breeding N₂-fixing legumes for high yields at low soil N levels therefore appears to be a reasonable strategy for enhancing N₂ fixation. Heavy applications of inorganic N fertilizer severely depressed N₂ fixation in two out of the three varieties used in Romania. One variety, F 74–412, however, derived slightly higher amounts of N₂ from fixation at 100 kg N/ha rate than when fertilized with 20 kg N/ha. In Greece, Chippewa, Williams and Amsoy-71 inoculated with a Nitragin inoculant fixed similar amounts of N₂ at both 20 and 100 kg N/ha fertilizer rates. However, when Chippewa and Williams were inoculated with amother, locally-isolated Rhizobium strain, N₂ fixation was substantially depressed at the higher N rate.     

Significance of nitrogen-fixing actinorhizal symbioses for restoration of depleted,degraded, and contaminated soil

Atmospheric nitrogen (N₂)-fixing legume trees are frequently used for the restoration of depleted, degraded, and contaminated soils. However, biological N₂ fixation (BNF) can also be performed by so-called actinorhizal plants. Actinorhizal plants include a high diversity of woody species and therefore can be applied in a broad spectrum of environments. In contrast to N₂-fixing legumes, the potential of actinorhizal plants for soil restoration remains largely unexplored. In this Opinion, we propose related basic research requirements for the characterization of environmental stress responses that determine the restoration potential of actinorhizal plants for depleted, degraded, and contaminated soils. We identify advantages and unexplored processes of actinorhizal plants and describe a mainly uncharted avenue of future research for this important group of plant species.     

Bi-directional transfer of nitrogen between alfalfa and bromegrass: Short and long term evidence

Transfer of N from legumes to associated non-legumes has been demonstrated under a wide range of conditions. Because legumes are able to derive their N requirements from N₂ fixation, legumes can serve, through the transfer of N, as a source of N for accompanying non-legumes. Studies, therefore, are often limited to the transfer of N from the legume to the non-legume. However, legumes preferentially rely on available soil N as their source of N. To determine whether N can be transferred from a non-legume to a legume, two greenhouse experiments were conducted. In the short-term N-transfer experiment, a portion of the foliage of meadow bromegrass (Bromus riparius Rhem.) or alfalfa (Medicago sativa L.) was immersed in a highly labelled ¹⁵N-solution and following a 64 h incubation, the roots and leaves of the associated alfalfa and bromegrass were analyzed for ¹⁵N. In the long-term N transfer experiment, alfalfa and bromegrass were grown in an ¹⁵N-labelled nutrient solution and transplanted in pots with unlabelled bromegrass and alfalfa plants. Plants were harvested at 50 and 79 d after transplanting and analyzed for ¹⁵N content. Whether alfalfa or bromegrass were the donor plants in the short-term experiment, roots and leaves of all neighbouring alfalfa and bromegrass plants were enriched with ¹⁵N. Similarly, when alfalfa or bromegrass was labelled in the long-term experiment, the roots and shoots of neighbouring alfalfa and bromegrass plants became enriched with ¹⁵N. These two studies conclusively show that within a short period of time, N is transferred from both the N₂-fixing legume to the associated non-legume and also from the non-legume to the N₂-fixing legume. The occurrence of a bi-directional N transfer between N₂-fixing and non-N₂-fixing plants should be taken into consideration when the intensity of N cycling and the directional flow of N in pastures and natural ecosystems are investigated.     

Application of 15N and xylem ureide methods for assessing N2 fixation of three shrub legumes periodically pruned for forage

The apparently diminished capacity for N₂ fixation by the shrub legume Calliandra calothyrsus (Calliandra) relative to other woody perennial legumes was investigated in a field experiment in northern Queensland, Australia. In this trial, (i) the proportion of plant nitrogen (N) derived from symbiotic N₂ fixation (%Pfix) and the amounts of N₂ fixed were compared in Calliandra, Gliricidia sepium (Gliricidia) and Codariocalyx gyroides (Codariocalyx), (ii) variations in N₂ fixation due to season or tree age were determined, (iii) estimates of Pfix derived with the ¹⁵N natural abundance technique were compared with values obtained from ¹⁵N enrichment or xylem sap ureide procedures to determine whether the previous conclusions about Calliandra's ability to fix N had resulted from specific problems with the natural abundance methodology used in the earlier studies.Inoculated seedlings of each of the three shrub legume species were planted in dense stands (1.5 m rows, 0.5 m between trees) in two randomised blocks. The northern block was used solely for natural abundance measurements, while ¹⁵N-enriched KNO₃ (10 atom % ¹⁵N excess) was applied four times over a 52 week period to plots in the southern block. The non-nodulating tree legume Senna spectabilis (formally Cassia spectabilis) was used as a non-N₂-fixing reference for the ¹⁵N-based procedures, with Guinea grass (Panicum maximum) included as an additional non-fixing check. Growth by the trees above 75 cm was first cut and removed after 22 weeks and regrowth was subsequently pruned periodically for another 95 weeks. Sampling for dry matter production, N yield and estimates of Pfix were restricted to the central four of the 32 plants which constituted each replicate plot. Information generated during the 117 week study indicated that estimates of Pfix by ¹⁵N natural abundance were closely similar to values derived with ¹⁵N-enrichment or sap ureides. The data indicated that Calliandra had a reduced reliance upon N₂ fixation relative to Gliricidia and Codariocalyx for the first 65 weeks after establishment. This appeared to be due to more prolifc root growth by Calliandra than either of the other N₂-fixing species and an ability to extract a greater proportion of its N requirements from soil mineral N. However, after week 65 and for the remainder of the experiment, estimates of Pfix for Calliandra were similar to the other shrub legumes. Over 117 weeks, prunings from Calliandra and Gliricidia had removed 52–58 t dry matter ha^-1, and between 1471 and 1678 kg N ha^-1, of which 1026–1063 kg N ha^-1 was estimated to have been derived from N₂ fixation. At the time of final harvest, 65–73% of the fixed N was present in shoot regrowth of the N₂ fixing shrubs, 9–18% in the roots, 15% in the trunk, and 2–6% in fallen leaves.     

Nitrogenase activity and N2 fixation are stimulated by elevated CO2 in a tropical N2-fixing tree

 Seeds of Gliricidia sepium, a fast-growing woody legume native to seasonal tropical forests of Central America, were inoculated with N₂-fixing Rhizobium bacteria and grown in environmentally controlled glasshouses for 67–71 days under ambient CO₂ (35 Pa) and elevated CO₂ (70 Pa) conditions. Seedlings were watered with an N-free, but otherwise complete, nutrient solution such that bacterial N₂ fixation was the only source of N available to the plant. The primary objective of our study was to quantify the effect of CO₂ enrichment on the kinetics of photosynthate transport to nodules and determine its subsequent effect on N₂ fixation. Photosynthetic rates and carbon storage in leaves were higher in elevated CO₂ plants indicating that more carbon was available for transport to nodules. A ¹⁴CO₂ pulse-chase experiment demonstrated that photosynthetically fixed carbon was supplied by leaves to nodules at a faster rate when plants were grown in elevated CO₂. Greater rates of carbon supply to nodules did not affect nodule mass per plant, but did increase specific nitrogenase activity (SNA) and total nitrogenase activity (TNA) resulting in greater N₂ fixation. In fact, a 23% increase in the rate of carbon supplied to nodules coincided with a 23% increase in SNA for plants grown in elevated CO₂, suggesting a direct correlation between carbon supply and nitrogenase activity. The improvement in plant N status produced much larger plants when grown in elevated CO₂. These results suggest that Gliricidia, and possibly other N₂-fixing trees, may show an early and positive growth response to elevated CO₂, even in severely N-deficient soils, due to increased nitrogenase activity. Received: 27 February 1996 / Accepted: 19 June 1996     

Emissions of nitrous oxide from three tropical forests in Southern China in response to simulated nitrogen deposition

Emissions of nitrous oxide (N₂O) from the soil following simulated nitrogen (N) deposition in a disturbed (pine), a rehabilitated (pine and broadleaf mixed) and a mature (monsoon evergreen broadleaf) tropical forest in southern China were studied. The following hypotheses were tested: (1) addition of N will increase soil N₂O emission in tropical forests; and (2) any observed increase will be more pronounced in the mature forest than in the disturbed or rehabilitated forest due to the relatively high initial soil N concentration in the mature forest. The experiment was designed with four N treatment levels (three replicates; 0, 50, 100, 150 kg N ha⁻¹ year⁻¹ for C (Control), LN (Low-N), MN (Medium-N), and HN (High-N) treatment, respectively) in the mature forest, but only three levels in the disturbed and rehabilitated forests (C, LN and MN). Between October 2005 to September 2006, soil N₂O flux was measured using static chamber and gas chromatography methodology. Nitrogen had been applied previously to the plots since July 2003 and continued during soil N₂O flux measurement period. The annual mean rates of soil N₂O emission in the C plots were 24.1 ± 1.5, 26.2 ± 1.4, and 29.3 ± 1.6 μg N₂O–N m⁻² h⁻¹ in the disturbed, rehabilitated and mature forest, respectively. There was a significant increase in soil N₂O emission following N additions in the mature forest (38%, 41%, and 58% when compared to the C plots for the LN, MN, and HN plots, respectively). In the disturbed forest a significant increase (35%) was observed in the MN plots, but not in the LN plots. The rehabilitated forest showed no significant response to N additions. Increases in soil N₂O emission occurred primarily in the cool-dry season (November, December and January). Our results suggest that the response of soil N₂O emission to N deposition in tropical forests in southern China may vary depending on the soil N status and land-use history of the forest.     

Biological Nitrogen Fixation in Two Tropical Forests: Ecosystem-Level Patterns and Effects of Nitrogen Fertilization

Humid tropical forests are often characterized by large nitrogen (N) pools, and are known to have large potential N losses. Although rarely measured, tropical forests likely maintain considerable biological N fixation (BNF) to balance N losses. We estimated inputs of N via BNF by free-living microbes for two tropical forests in Puerto Rico, and assessed the response to increased N availability using an on-going N fertilization experiment. Nitrogenase activity was measured across forest strata, including the soil, forest floor, mosses, canopy epiphylls, and lichens using acetylene (C₂H₂) reduction assays. BNF varied significantly among ecosystem compartments in both forests. Mosses had the highest rates of nitrogenase activity per gram of sample, with 11 ± 6 nmol C₂H₂ reduced/g dry weight/h (mean ± SE) in a lower elevation forest, and 6 ± 1 nmol C₂H₂/g/h in an upper elevation forest. We calculated potential N fluxes via BNF to each forest compartment using surveys of standing stocks. Soils and mosses provided the largest potential inputs of N via BNF to these ecosystems. Summing all components, total background BNF inputs were 120 ± 29 μg N/m²/h in the lower elevation forest, and 95 ± 15 μg N/m²/h in the upper elevation forest, with added N significantly suppressing BNF in soils and forest floor. Moisture content was significantly positively correlated with BNF rates for soils and the forest floor. We conclude that BNF is an active biological process across forest strata for these tropical forests, and is likely to be sensitive to increases in N deposition in tropical regions.     

Contrasting growth response of an N2-fixing and non-fixing forb to elevated CO2: dependence on soil N supply

With the ability to symbiotically fix atmospheric N₂, legumes may lack the N-limitations thought to constrain plant response to elevated concentrations of atmospheric CO₂. The growth and photosynthetic responses of two perennial grassland species were compared to test the hypotheses that (1) the CO₂ response of wild species is limited at low N availability, (2) legumes respond to a greater extent than non-fixing forbs to elevated CO₂, and (3) elevated CO₂ stimulates symbiotic N₂ fixation, resulting in an increased amount of N derived from the atmosphere. This study investigated the effects of atmospheric CO₂ concentration (365 and 700 mol mol^–1) and N addition on whole plant growth and C and N acquisition in an N₂-fixing legume (Lupinus perennis) and a non-fixing forb (Achillea millefolium) in controlled-chamber environments. To evaluate the effects of a wide range of N availability on the CO₂ response, we incorporated six levels of soil N addition starting with native field soil inherently low in N (field soil + 0, 4, 8, 12, 16, or 20 g N m^–2 yr^–1). Whole plant growth, leaf net photosynthetic rates (A), and the proportion of N derived from N₂ fixation were determined in plants grown from seed over one growing season. Both species increased growth with CO₂enrichment, but this response was mediated by N supply only for the non-fixer, Achillea. Its response depended on mineral N supply as growth enhancements under elevated CO₂ increased from 0% in low N soil to +25% at the higher levels of N addition. In contrast, Lupinus plants had 80% greater biomass under elevated CO₂ regardless of N treatment. Although partial photosynthetic acclimation to CO₂ enrichment occurred, both species maintained comparably higher A in elevated compared to ambient CO₂ (+38%). N addition facilitated increased A in Achillea, however, in neither species did additional N availability affect the acclimation response of A to CO₂. Elevated CO₂ increased plant total N yield by 57% in Lupinus but had no effect on Achillea. The increased N in Lupinus came from symbiotic N₂ fixation, which resulted in a 47% greater proportion of N derived from fixation relative to other sources of N. These results suggest that compared to non-fixing forbs, N₂-fixers exhibit positive photosynthetic and growth responses to increased atmospheric CO₂ that are independent of soil N supply. The enhanced amount of N derived from N₂ fixation under elevated CO₂ presumably helps meet the increased N demand in N₂-fixing species. This response may lead to modified roles of N₂-fixers and N₂-fixer/non-fixer species interactions in grassland communities, especially those that are inherently N-poor, under projected rising atmospheric CO₂.     

Nitrogen Oxide Fluxes and Nitrogen Cycling during Postagricultural Succession and Forest Fertilization in the Humid Tropics

The effects of changes in tropical land use on soil emissions of nitrous oxide (N₂O) and nitric oxide (NO) are not well understood. We examined emissions of N₂O and NO and their relationships to land use and forest composition, litterfall, soil nitrogen (N) pools and turnover, soil moisture, and patterns of carbon (C) cycling in a lower montane, subtropical wet region of Puerto Rico. Fluxes of N₂O and NO were measured monthly for over 1 year in old (more than 60 years old) pastures, early- and mid-successional forests previously in pasture, and late-successional forests not known to have been in pasture within the tabonuco (Dacryodes excelsa) forest zone. Additional, though less frequent, measures were also made in an experimentally fertilized tabonuco forest. N₂O fluxes exceeded NO fluxes at all sites, reflecting the consistently wet environment. The fertilized forest had the highest N oxide emissions (22.0 kg N · ha⁻¹· y⁻¹). Among the unfertilized sites, the expected pattern of increasing emissions with stand age did not occur in all cases. The mid-successional forest most dominated by leguminous trees had the highest emissions (9.0 kg N · ha⁻¹· y⁻¹), whereas the mid-successional forest lacking legumes had the lowest emissions (0.09 kg N · ha⁻¹· y⁻¹). N oxide fluxes from late-successional forests were higher than fluxes from pastures. Annual N oxide fluxes correlated positively to leaf litter N, net nitrification, potential nitrification, soil nitrate, and net N mineralization and negatively to leaf litter C:N ratio. Soil ammonium was not related to N oxide emissions. Forests with lower fluxes of N oxides had higher rates of C mineralization than sites with higher N oxide emissions. We conclude that (a) N oxide fluxes were substantial where the availability of inorganic N exceeded the requirements of competing biota; (b) species composition resulting from historical land use or varying successional dynamics played an important role in determining N availability; and (c) the established ecosystem models that predict N oxide loss from positive relationships with soil ammonium may need to be modified. Received 22 February 2000; accepted 6 September 2000.     

Dominance of legume trees alters nutrient relations in mixed species forest restoration plantings within seven years

Failures in reforestation are often attributed to nutrient limitation for tree growth. We compared tree performance and nitrogen and phosphorus relations in adjacent mixed-species plantings of contrasting composition, established for forest restoration on Ultisol soil, originally covered by tropical semi-deciduous Atlantic Forest in Southeast Brazil. Nutrient relations of four tree species occurring in both planting mixtures were compared between a legume-dominated, species-poor direct seeding mixture of early-successional species (“legume mixture”), and a species-diverse, legume-poor mixture of all successional groups (“diverse mixture”). After 7 years, the legume mixture had 6-fold higher abundance of N₂-fixing trees, 177% higher total tree basal area, 22% lower litter C/N, six-fold higher in situ soil resin-nitrate, and 40% lower in situ soil resin-P, compared to the diverse mixture. In the legume mixture, non-N₂-fixing legume Schizolobium parahyba (Fabaceae-Caesalpinioideae) had significantly lower proportional N resorption, and both naturally regenerating non-legume trees had significantly higher leaf N concentrations, and higher proportional P resorption, than in the diverse mixture. This demonstrate forms of plastic adjustment in all three non-N₂-fixing species to diverged nutrient relations between mixtures. By contrast, leaf nutrient relations in N₂-fixing Enterolobium contortisiliquum (Fabaceae-Mimosoideae) did not respond to planting mixtures. Rapid N accumulation in the legume mixture caused excess soil nitrification over nitrate immobilization and tighter P recycling compared with the diverse mixture. The legume mixture succeeded in accelerating tree growth and canopy closure, but may imply periods of N losses and possibly P limitation. Incorporation of species with efficient nitrate uptake and P mobilization from resistant soil pools offers potential to optimize these tradeoffs.     

Successional changes in soil nitrogen availability,non-symbiotic nitrogen fixation and carbon/nitrogen ratios in southern Chilean forest ecosystems

Vast areas of southern Chile are now covered by second-growth forests because of fire and logging. To study successional patterns after moderate-intensity, anthropogenic fire disturbance, we assessed differences in soil properties and N fluxes across a chronosequence of seven successional stands (2–130 years old). We examined current predictions of successional theory concerning changes in the N cycle in forest ecosystems. Seasonal fluctuations of net N mineralization (N_min) in surface soil and N availability (N_a; N_a=NH ₄ ⁺ –N+NO ₃ ^– –N) in upper and deep soil horizons were positively correlated with monthly precipitation. In accordance with theoretical predictions, stand age was positively, but weakly related to both N_a (r ²=0.282, P<0.001) and total N (N_tot; r ²=0.192, P<0.01), and negatively related to soil C/N ratios (r ²=0.187, P<0.01) in surface soils. A weak linear increase in soil N_min (upper plus deep soil horizons) was found across the chronosequence (r ²=0.124, P<0.022). N_min occurred at modest rates in early successional stands, suggesting that soil disturbance did not impair microbial processes. The relationship between N fixation (N_fix) in the litter layer and stand age best fitted a quadratic model (r ²=0.228, P<0.01). In contrast to documented successional trends for most temperate, tropical and Mediterranean forests, non-symbiotic N_fix in the litter layer is a steady N input to unpolluted southern temperate forests during mid and late succession, which may compensate for hydrological losses of organic N from old-growth ecosystems.     

Assessing the symbiotic dependency of grain and tree legumes on N2 fixation for their N nutrition in five agro-ecological zones of Botswana

To assess the symbiotic dependency of grain and shrub/tree legumes within five agro-ecological zones of Botswana, fully expanded leaves of the test species were sampled from about 26 study sites within Ngwaketse, Gaborone, Central, Ghanzi and Kalahari agro-ecological zones. Isotopic analysis revealed significant differences in δ¹⁵N values of the grain legumes [cowpea (Vigna unguiculata L. Walp), Bambara groundnut (Vigna subterranea L. Verde.), and groundnut (Arachis hypogaea L.)] from the 26 farming areas in both 2005 and 2006. Estimates of %Ndfa of leaves also showed significant differences between farming areas, with cowpea deriving more than 50% of its N nutrition from symbiotic fixation. In terms of distribution, many more symbiotic shrub/tree species were found in the wetter Ngwaketse agro-zone compared to the fewer numbers in the drier Kalahari region. Acacias were the more dominant species at all sites. Leaf δ¹⁵N values of shrub/tree species also varied strongly across Botswana, with 11 out of 18 of these legumes deriving about 50%, or more, of their N from symbiotic N₂ fixation.Acacia caffra, in particular, obtained as much as 93.6% of its N nutrition from symbiotic fixation in the wetter Ngwaketse agro-zone. This study has shown that grain legumes sampled from farmer’s fields in Botswana obtained considerable amounts of their N from symbiotic fixation. We have also shown that shrub and tree legumes probably play an important role in the N economy of the savanna ecosystems in Botswana. However, the decline in the number of functional N₂-fixing shrub/tree legumes along an aridity gradient suggests that soil moisture is a major constraint to N₂ fixation in the tree legumes of Botswana.     

Dinitrogen-fixation by three neotropical agroforestry tree species under semi-controlled field conditions

Cultivating dinitrogen-fixing legume trees with crops in agroforestry is a relatively common N management practice in the Neotropics. The objective of this study was to assess the N₂ fixation potential of three important Neotropical agroforestry tree species, Erythrina poeppigiana, Erythrina fusca, and Inga edulis, under semi-controlled field conditions. The study was conducted in the humid tropical climate of the Caribbean coastal plain of Costa Rica. In 2002, seedlings of I. edulis and Vochysia guatemalensis were planted in one-meter-deep open-ended plastic cylinders buried in soil within hedgerows of the same species. Overall tree spacing was 1 × 4 m to simulate a typical alley-cropping design. The ¹⁵N was applied as (NH₄)₂SO₄ at 10% ¹⁵N atom excess 15 days after planting at the rate of 20 kg [N] ha⁻¹. In 2003, seedlings of E. poeppigiana, E. fusca, and V. guatemalensis were planted in the same field using the existing cylinders. The ¹⁵N application was repeated at the rate of 20 kg [N] ha⁻¹ 15 days after planting and 10 kg [N] ha⁻¹ was added three months after planting. Trees were harvested 9 months after planting in both years. The ¹⁵N content of leaves, branches, stems, and roots was determined by mass spectrometry. The percentage of atmospheric N fixed out of total N (%N_f) was calculated based on ¹⁵N atom excess in leaves or total biomass. The difference between the two calculation methods was insignificant for all species. Sixty percent of I. edulis trees fixed N₂; %N_f was 57% for the N₂-fixing trees. Biomass production and N yield were similar in N₂-fixing and non-N₂-fixing I. edulis. No obvious cause was found for why not all I. edulis trees fixed N₂. All E. poeppigiana and E. fusca trees fixed N₂; %N_f was ca. 59% and 64%, respectively. These data were extrapolated to typical agroforestry systems using published data on N recycling by the studied species. Inga edulis may recycle ca. 100 kg ha⁻¹ a⁻¹ of N fixed from atmosphere to soil if only 60% of trees fix N₂, E. poeppigiana 60–160 kg ha⁻¹ a⁻¹, and E. fusca ca. 80 kg ha⁻¹ a⁻¹.