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1.
The acetylene reduction assay for the measurement of N 2 fixation in a water-saturated paddy soil is limited by the slow diffusion of acetylene and ethylene. In laboratory incubation tests, vigorous shaking after the assay period is needed to release ethylene into the gas within the assay vials. Shaking prior to the incubation is also effective for dissolving acetylene in the water-saturated soil. However, a water-saturated soil depth of less than 10 mm during incubation is recommended. In field assays, some amounts of ethylene remain in the water-saturated soil phase of the acetylene reduction assay chamber, but stirring the water-saturated soil before sampling reduces the amount of ethylene remaining in soil. Evidence of a downward movement of acetylene and an upward movement of ethylene through rice plants was obtained. Because of the rapid transfer of acetylene to rice plant roots, an in situ acetylene reduction assay covering a rice hill is likely to detect nitrogen fixation in the proximity of roots where acetylene is easily accessible. Acetylene introduction to the water-saturated soil phase prior to assay did not greatly increase the acetylene reduction rate. Carbon dioxide enrichment in the assay chamber did not enhance nitrogen fixation in a paddy including rice and algae during a 1-day cycle. 相似文献
2.
An apparatus was designed that permitted acetylene reduction (N 2 fixation) by root nodules to be measured in situ simultaneously with net photosynthesis, dark respiration, and transpiration of the shoot in soybean plants ( Glycine max [L.] Merr. var. Beeson). Tests showed that acetylene reduction was linear with time for at least 5 hours, except for the first 30 to 60 minutes. Endogenous ethylene production did not affect the measurements. Successive determinations of acetylene reduction could be made without apparent aftereffects on the plant. 相似文献
3.
The relationship between ureide N and N 2 fixation was evaluated in greenhouse-grown soybean ( Glycine max L. Merr.) and lima bean ( Phaseolus lunatus L.) and in field studies with soybean. In the greenhouse, plant N accumulation from N 2 fixation in soybean and lima bean correlated with ureide N. In soybean, N 2 fixation, ureide N, acetylene reduction, and nodule mass were correlated when N 2 fixation was inhibited by applying KNO 3 solutions to the plants. The ureide-N concentrations of different plant tissues and of total plant ureide N varied according to the effectiveness of the strain of Bradyrhizobium japonicum used to inoculate plants. The ureide-N concentrations in the different plant tissues correlated with N 2 fixation. Ureide N determinations in field studies with soybean correlated with N 2 fixation, aboveground N accumulation, nodule weight, and acetylene reduction. N 2 fixation was estimated by 15N isotope dilution with nine and ten soybean genotypes in 1979 and 1980, respectively, at the V9, R2, and R5 growth stages. In 1981, we investigated the relationship between ureide N, aboveground N accumulation, acetylene reduction, and nodule mass using four soybean genotypes harvested at the V4, V6, R2, R4, R5, and R6 growth stages. Ureide N concentrations of young stem tissues or plants or aboveground ureide N content of the four soybean genotypes varied throughout growth correlating with acetylene reduction, nodule mass, and aboveground N accumulation. The ureide-N concentrations of young stem tissues or plants or aboveground ureide-N content in three soybean genotypes varied across inoculation treatments of 14 and 13 strains of Bradyrhizobium japonicum in 1981 and 1982, respectively, and correlated with nodule mass and acetylene reduction. In the greenhouse, results correlating nodule mass with N 2 fixation and ureide N across strains were variable. Acetylene reduction in soybean across host-strain combinations did not correlate with N 2 fixation and ureide N. N 2 fixation, ureide N, acetylene reduction, and nodule mass correlated across inoculation treatments with strains of Bradyrhizobium spp. varying in effectiveness on lima beans. Our data indicate that ureide-N determinations may be used as an additional method to acetylene reduction in studies of the physiology of N 2 fixation in soybean. Ureide-N measurements also may be useful to rank strains of B. japonicum for effectiveness of N 2 fixation. 相似文献
4.
An automated method was developed for continuous, in situ determination of acetylene reduction (N 2 fixation) by intact soybean plants ( Glycine max [L.]). The culture vessel containing the roots of intact plants grown in sand culture is sealed at the surface and an air-acetylene mixture continuously injected into the root chamber. The effluent gas is automatically sampled and injected into a gas chromatograph. Continuous acetylene assay at intervals as short as 3.5 min may be made over a period of several days, without attention, except for plant watering. Adverse effects of prolonged exposure of the root system to acetylene were mitigated by pulse injection of acetylene for 20 min followed by 40 min of acetylene-free air. Bare root systems can be suspended in a reaction chamber and sprayed with water or nutrient solution; this permits periodic removal of the root system for sampling nodules. 相似文献
5.
The role of lateral root nodules in N 2 fixation and the relationships between total shoot N and several traits which influence or control N 2 fixation in common bean ( Phaseolus vulgaris L.) i.e., acetylene reduction value, specific nodule activity, leghemoglobin concentration, total leghemoglobin and nodule mass, were investigated in field studies. Significant variation among bean lines was observed for all the traits measured. Lines varied for the proportion of total N accumulated up to the R3 growth state, thus measurements of total shoot N near maturity ( e.g., R7) provided a better estimate of total N 2 fixation than measurements taken at an early growth stage. Nodule mass was correlated with acetylene reduction and total leghemoglobin, and total leghemoglobin was correlated with acetylene reduction value. Total shoot N at R7 was correlated with seasonal means of nodule mass and number, acetylene reduction value and total leghemoglobin. For all traits except total leghemoglobin, values for lateral roots were more highly correlated with total shoot N than were values for either crown roots or the whole root system. Seed yield was most highly correlated with nodule mass of the lateral roots. These results will be useful in devising breeding strategies for improved N 2 fixation of the host plant. 相似文献
6.
The two commonly applied methods to assess dinitrogen (N 2) fixation rates are the 15N 2-tracer addition and the acetylene reduction assay (ARA). Discrepancies between the two methods as well as inconsistencies between N 2 fixation rates and biomass/growth rates in culture experiments have been attributed to variable excretion of recently fixed N 2. Here we demonstrate that the 15N 2-tracer addition method underestimates N 2 fixation rates significantly when the 15N 2 tracer is introduced as a gas bubble. The injected 15N 2 gas bubble does not attain equilibrium with the surrounding water leading to a 15N 2 concentration lower than assumed by the method used to calculate 15N 2-fixation rates. The resulting magnitude of underestimation varies with the incubation time, to a lesser extent on the amount of injected gas and is sensitive to the timing of the bubble injection relative to diel N 2 fixation patterns. Here, we propose and test a modified 15N 2 tracer method based on the addition of 15N 2-enriched seawater that provides an instantaneous, constant enrichment and allows more accurate calculation of N 2 fixation rates for both field and laboratory studies. We hypothesise that application of N 2 fixation measurements using this modified method will significantly reduce the apparent imbalances in the oceanic fixed-nitrogen budget. 相似文献
7.
It is well established that nitrate is a potent inhibitor of nodulation and nitrogen fixation in legumes. The objective of
this study was to demonstrate the relative insensitivity of these processes to nitrate with Calopogonium mucunoides, a tropical South American perennial legume, native to the cerrado (savannah) region. It was found that nodule number was
reduced by about half in the presence of high levels of nitrate (15 mM) but nodule growth (total nodule mass per plant) and
nitrogen fixation (acetylene reduction activity and xylem sap ureide levels) were not affected. Other sources of N (ammonium
and urea) were also without effect at these concentrations. At even higher concentrations (30 mM), nitrate did promote significant
inhibition (ca. 50%) of acetylene reduction activity, but no significant reduction in xylem sap ureides was found. The extraordinary
insensitivity of nodulation and N 2 fixation of C. mucunoides to nitrate suggests that this species should be useful in studies aimed at elucidating the mechanisms of nitrate inhibition
of these processes. 相似文献
8.
Pigeon peas [ Cajanus cajan (L.) Millsp.] were grown in soil columns containing 15N-enriched organic matter. Seasonal N 2 fixation activity was determined by periodically assaying plants for reduction of C 2H 2. N 2 fixation rose sharply from the first assay period at 51 days after planting to a peak of activity between floral initiation and fruit set. N 2 fixation (acetylene reduction) activity dropped concomitantly with pod maturation but recovered after pod harvests. Analysis of 15N content of plant shoots revealed that approximately 91 to 94% of plant N was derived from N 2 fixation. The effect of inoculation with hydrogenase-positive and hydrogenase-negative rhizobia was examined. Pigeon peas inoculated with strain P132 (hydrogenase-positive) yielded significantly more total shoot N than other inoculated or uninoculated treatments. However, two other hydrogenase-positive strains did not yield significantly more total shoot N than a hydrogenase-negative strain. The extent of nodulation by inoculum strains compared to indigenous rhizobia was determined by typing nodules according to intrinsic antibiotic resistance of the inoculum strains. The inoculum strains were detected in almost all typed nodules of inoculated plants. Gas samples were taken from soil columns several times during the growth cycle of the plants. H2 was never detected, even in columns containing pigeon peas inoculated with hydrogenase-negative rhizobia. This was attributed to H2 consumption by soil bacteria. Estimation of N2 fixation by acetylene reduction activity was closest to the direct 15N method when ethylene concentrations in the gas headspace (between the column lid and soil surface) were extrapolated to include the soil pore space as opposed solely to measurement in the headspace. There was an 8-fold difference between the two acetylene reduction assay methods of estimation. Based on a planting density of 15,000 plants per hectare, the direct 15N fixation rates ranged from 67 (noninoculated) to 134 kilograms per hectare, while grain yields ranged from 540 to 825 kilograms per hectare. Grain yields were not increased with N fertilizer. 相似文献
9.
The effect of nitrate on N 2 fixation and the assimilation of fixed N 2 in legume nodules was investigated by supplying nitrate to well established soybean ( Glycine max L. Merr. cv Bragg)- Rhizobium japonicum (strain 3I1b110) symbioses. Three different techniques, acetylene reduction, 15N 2 fixation and relative abundance of ureides ([ureides/(ureides + nitrate + α-amino nitrogen)] × 100) in xylem exudate, gave similar results for the effect of nitrate on N 2 fixation by nodulated roots. After 2 days of treatment with 10 millimolar nitrate, acetylene reduction by nodulated roots was inhibited by 48% but there was no effect on either acetylene reduction by isolated bacteroids or in vitro activity of nodule cytoplasmic glutamine synthetase, glutamine oxoglutarate aminotransferase, xanthine dehydrogenase, uricase, or allantoinase. After 7 days, acetylene reduction by isolated bacteroids was almost completely inhibited but, except for glutamine oxoglutarate aminotransferase, there was still no effect on the nodule cytoplasmic enzymes. It was concluded that, when nitrate is supplied to an established symbiosis, inhibition of nodulated root N 2 fixation precedes the loss of the potential of bacteroids to fix N 2. This in turn precedes the loss of the potential of nodules to assimilate fixed N 2. 相似文献
10.
This study is the first to investigate quantitative effects of plant community composition and diversity on N 2 fixation in legumes. N 2 fixation in three perennial Trifolium species grown in field plots with varied number of neighbouring species was evaluated with the 15N natural abundance method (two field sites, several growing seasons, no N addition) and the isotope dilution method (one
site, one growing season, 5 g N m −2). The proportion of plant N derived from N 2 fixation, pNdfa, was generally high, but the N addition decreased pNdfa, especially in species-poor communities. Also following
N addition, the presence of grasses in species-rich communities increased pNdfa in T. hybridum and T. repens L., while legume abundance had the opposite effect. In T. repens, competition for light from grasses appeared to limit growth and thereby the amount of N 2 fixed at the plant level, expressed as mg N 2 fixed per sown seed . We conclude that the occurrence of diversity effects seems to be largely context dependent, with soil N availability being
a major determinant, and that species composition and functional traits are more important than species richness regarding
how neighbouring plant species influence N 2 fixation in legumes. 相似文献
11.
The role of nitrogen (N 2) fixation in balancing N supply to wetland metaphyton was assessed by comparing primary production with enzymatic, isotopic,
and elemental correlates. Primary production, N 2 fixation (acetylene reduction, AR), phosphatase activity, C:N:P ratio, and N isotopic composition of metaphyton were measured
along a nutrient gradient in a freshwater marsh during May through September 2004. N 2 fixation and phosphatase activity in metaphyton were negatively correlated with inorganic N and P concentrations, respectively.
Although metaphyton N 2 fixation demonstrated a clear spatial pattern along the nutrient gradient, N 2 fixation rates varied monthly and decreased sharply in September. However, the percent contribution of N 2 fixation to N uptake by metaphyton consistently decreased throughout the summer. Furthermore, the decreased contribution
of N 2 fixation to N uptake corresponded with an increase in metaphyton N content during the growing season. Nitrogen isotopic data
suggested the sustained importance of an atmospheric N 2 source through September at the most downstream (nutrient poor) site even though the percent contribution of N 2 fixition to N uptake was lowest in that month. This suggests that metaphyton were efficiently accumulating and recycling
fixed N 2 in support of primary production. Over the course of the summer, metaphyton primary production showed a weak inverse correlation
with metaphyton phosphatase activity ( r
2 = 0.58). The largest residuals in this regression corresponded to the largest vartiation in metaphyton N content. When metaphyton
primary production was normalized to metaphyton N content, production rates for the entire growing season were more strongly
inversely correlated with metaphyton phosphatase activity ( r
2 = 0.78). Results of the study suggest that N 2 fixation in N poor areas may adequately supplement community N requirements in metaphyton, thereby causing limitation by
other elemental resources such as P. 相似文献
12.
Summary Non-symbiotic heterotrophic N 2 fixation in coniferous bark litter was investigated with the acetylene reduction assay under aerobic and anaerobic conditions. The litter studied was composed essentially of bark, of pH 5 and a C/N ratio of 101; the ratio of available C to available N, which governs N 2 fixation, was considerably higher. The rate of N 2 fixation was estimated as 2.5–4.4 g N. g –1 dry wt. day –1. Nitrogenase activity was still evident after seven months of incubation under aerobic conditions. The N 2-ase activity was O 2 dependent: under anaerobic conditions no N 2-ase activity was found unless a fermentable C source was added. The importance of N 2 fixation in N-poor litter for the maintenance of soil fertility is emphasized. 相似文献
13.
It has previously been reported that endophytic diazotrophic bacteria contribute significantly to the nitrogen budgets of
some graminaceous species. In this study the contribution of biological nitrogen fixation to the N-budget of a South African
sugarcane cultivar was evaluated using 15N natural abundance, acetylene reduction and 15N incorporation. Plants were also screened for the presence of endophytic diazotrophic bacteria using acetylene reduction
and nifH-gene targeted PCR with the pure bacterial strains. 15N natural abundance studies on field-grown sugarcane indicated that the plants did not rely extensively on biological nitrogen
fixation. Furthermore, no evidence was found for significant N 2-fixation or nitrogenase activity in field-grown or glasshouse-grown plants using 15N incorporation measurements and acetylene reduction assays. Seven endophytic bacterial strains were isolated from glasshouse-grown
and field-grown plants and cultured on N-free medium. The diazotrophic character of these seven strains could not be confirmed
using acetylene reduction and PCR screening for nifH. Thus, although biological nitrogen fixation may occur in South African sugarcane varieties, the contribution of this N-source
in the tested cultivar was not significant. 相似文献
14.
On three separate occasions we investigated morphological and physiological aspects of bacterial associations with planktonic aggregates of the ubiquitous marine N 2 fixing cyanobacterium Trichodesmium sp. Close associations generally characterized Trichodesmium blooms; associations were present during day- and night-time. Colonization by both rod-shaped and filamentous heterotrophic bacteria occurred on Trichodesmiun aggregates actively fixing N 2 (acetylene reduction). Scanning electron and optical microscopy showed bacteria located both around and within aggregates. Microautoradiography demonstrated that associated bacteria largely mediated utilization of trace additions of 3H-labeled carbohydrates (fructose, glucose, mannitol) and amino acids, whereas Trichodesmium utilized amino acids only. Oxygen measurements using microelectrodes revealed high localized oxygen consumption among aggregates, with rapid (within a minute) changes from supersaturated to subsaturated oxygen following the transition from photosynthetic illuminated to dark periods. Stab culturing techniques confirmed the presence of heterotrophic N 2 fixers among aggregate-associated bacteria. Parallel deployment of oxygen microelectrodes, the tetrazolium salt 2,3,5 triphenyl tetrazolium chloride (TTC) and acetylene reduction assays demonstrated microaerophilic requirements for expression of nitrogenase activity among cultured bacteria. Trichodesmium aggregates are characterized by dynamic nutrient and oxygen regimes, which promote and maintain simultaneous and contiguous oxygenic photosynthesis and N 2 fixation. In part, the above-mentioned consortial interactions with a variety of heterotrophic bacteria facilitate Trichodesmium biomass production and bloom formation in nitrogen depleted, oligotrophic tropical/subtropical waters. 相似文献
15.
Measurements of CO 2 and H 2O fluxes were carried out using two different techniques—eddy-covariance (EC) and open system gas exchange chamber (OC)—during
two-years’ period (2003–2004) at three different grassland sites. OC measurements were made during fourteen measurement campaigns.
We found good agreement between the OC and EC CO 2 flux values ( n = 63, r
2 = 0.5323, OC F CO2 = −0.6408+0.9508 EC F CO2). The OC F H2O values were consistently lower than those measured by the EC technique, probably caused by the air stream difference inside
and outside the chamber. Adjusting flow rate within the chamber to the natural conditions would be necessary in future OC
measurements. In comparison with EC, the OC proved to be a good tool for gas exchange measurements in grassland ecosystems. 相似文献
16.
Specimens of Chamaebatia foliolosa Benth. with nodule structures on their roots fix atmospheric nitrogen. The nodules are similar to those of other non-legumes in gross morphology and structure, containing hyphal strands, some with club-shaped vesicles at their ends. A fixation rate of 130 nmoles N 2 per g fresh weight per hr is reported by using 15N 2 as a tracer. Equivalent rates of acetylene reduction were observed. 相似文献
17.
Periphyton N 2 fixation plays a key role in N cycling of aquatic systems, but temporal studies of this process are often lacking, especially in systems with only seasonal flooding. We used seven samplings to characterize nitrogenase activity (acetylene reduction method) of periphyton in short-hydroperiod marl prairies and two wetlands restored from agricultural disturbance in Everglades National Park, USA. We hypothesized that the seasonal drying and rewetting would increase the temporal dynamics of the process. All sites showed significant periphyton N 2 fixation, but in restored areas highest rates were observed only in the early wet season (July), while in reference sites fixation was spread throughout the summer. Most N 2 fixation in the restored areas was confined to a 3-month-period resulting in large underestimates of annual fixation in previous studies with few seasonal measurements. N 2 fixation rates correlated with total P, N and TN:TP ratio, and periphyton moisture content in the dry season. N stable isotopic signature was a good indicator of N 2 fixation rates between sites, but did not correctly indicate seasonal patterns. These findings improve our understanding of N cycling in wetlands like the Everglades and indicate a need for more detailed measurements of processes in seasonally flooded systems. 相似文献
18.
Summary Isotopic as well as non-isotopic methods were used to assess symbiotic nitrogen fixation within eight soybean [ Glycine max (L.) Merr.] cultivars grown at 20 and 100 kg N/ha levels of nitrogen fertilizer under field conditions.The 15N methodology revealed large differences between soybean cultivars in their abilities to support nitrogen fixation. In almost all cases, the application of 100 kg N/ha resulted in lower N 2 fixed in soybean than at 20 kg N/ha in the first year of the study. However, N 2 fixed in one cultivar, Dunadja, was not significantly affected by the higher rate of N fertilizer application. These results were confirmed by measurements of acetylene reduction activity, nodule dry weight and N 2 fixed as measured by the difference method. Further proof of differences in N 2 fixed within soybean cultivars and the ability of Dunadja to fix similar amounts of N 2 at 20 and 100 kg N/ha was obtained during a second year experiment. Dunadja yield was affected by N fertilizer and produced larger yield at 100 kg N/ha than at 20 kg N/ha. This type of cultivar could be particularly useful in situations where soil N levels are high or where there is need to apply high amounts of N fertilizer.The present study reveals the great variability between legume germplasms in the ability to fix N 2 at different inorganic N levels, and also the potential that exists in breeding for nitrogen fixation associative traits. The 15N methodology offers a unique tool to evaluate germplasms directly in the field for their N 2 fixation abilities at different N fertilizer levels. 相似文献
19.
Summary A system for employing open-ended root chambers to measure in situ acetylene reduction rates under field conditions is described. Gas mixtures containing about 2 mbar acetylene were continuously
flowed through the chambers providing a continuous record of acetylene reduction. These chambers have been used to measure
acetylene reduction rates of soybeans during three growing seasons. The system has proved to be reliable with a high degree
of precision. The large amount of plant-to-plant variability observed in N 2 fixation research has been confirmed by the data collected with this system. However, such variability in physiological studies
can be reduced by using a non-destructive system to compare the response of an individual plant with its rates before treatment. 相似文献
20.
Microcoleus chthonoplastes dominated microbial mats are conspicuous along the shallow littoral zone in Lake Chiprana, a hypersaline lake located in the Ebro river basin in north-eastern Spain. Pigment data show that these mats included diatom species and anoxygenic phototrophs, Chloroflexus-type bacteria and purple bacteria. In situ, these mats showed low rates of dinitrogen fixation (acetylene reduction). Acetylene reduction was stimulated about 30-fold in excised mats after moderate phosphate fertilisation during 2 weeks incubation in a mesocosm. Pigment analyses showed that this treatment had little impact on the phototrophic community structure, except that it induced a decrease of Chloroflexus-type bacteria. The use of metabolic inhibitors indicated that methanogenic archaea and aerobic heterotrophic bacteria were the major dinitrogen fixers in this system. This is in agreement with the fact that the mat-building cyanobacterium M. chthonoplastes lacks the dinitrogenase reductase nifH gene and with the fact that acetylene reduction rates were strongly stimulated by additions of H 2/CO 2, methanol, fructose and sucrose, but not by lactate, acetate, formate and glucose. No significant differences where found for acetylene reduction rates when comparing light and dark incubations of these microbial mats. However, acetylene reduction rates were enhanced in the light when the near infrared (NIR) light was filtered out, which arrested anoxygenic photosynthesis. We suggest, therefore, that the chemoheterotrophic dinitrogen fixing bacteria were in competition with anoxygenic phototrophic bacteria for organic substrates, while the latter did not contribute to dinitrogen fixation in the mat. 相似文献
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