C2H2 and Ar induce rapid declines in nitrogenase activity and CO2 evolution in nodules of Datisca glomerata

Preliminary studies have indicated that after addition of C₂H₂ there is a rapid decline in nitrogenase activity in the nodules of Datisca glomerata . The present work was undertaken to determine whether (1) there is also a decline in respiration and (2) the decline is associated with the cessation of ammonia production. The rates of C₂H₄ and CO₂ evolution by nodulated root systems of Datisca were measured as a function of time after exposure to C₂H₂. The peak rate of C₂H₄ evolution occurred at 30 s after C₂H₂ exposure, while the rate of CO₂ evolution started to decline at 60 s after exposure to C₂H₂. Incubation of nodules in a gas mixture containing Ar also caused a decline in CO₂ evolution. Further, pretreatment with Ar eliminated most of the C₂H₂-induced decline in nitrogenase activity and CO₂ evolution. These C₂H₂- and Ar-induced declines in Datisca nodules are more rapid than those reported in any other nodules. They are evidence that continued ammonia formation is essential for maintenance of normal nitrogenase activity in Datisca nodules.     

The Glycine-Glomus-Bradyrhizobiun symbiosis. XI. Nodule gas exchange and efficiency as a function of soil and root water status in mycorrhizal soybean

Soybean [ Glycine max (L.) Merr. cv. Hobbit] plants were inoculated with a HUP− strain of Bradyrhizobium japonicum (Nitragin 61A118) and either colonized by the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus mosseae (Nicol & Gerd.) Gerd. and Trappe or fertilized with KH₂PO₄ (nonVAM). They were grown for 50 days in a growth chamber and harvested over a 4-day drought period during which available soil water decreased to 0. Nodule P concentrations and P-use efficiency declined linearly with soil and root water content during the harvest period in both VAM and nonVAM plants. Nitrogenase activity, estimated from H₂ evolution and C₂H₂ reduction data, was also a linear function of declining nodule P concentrations and CO₂-exchange rates and showed simular patterns in both treatments. Hydrogen evolution and the relative efficiency of N₂ fixation, on the other hand, reacted differently to increasing drought in VAM and nonVAM plants. Differences in the responses of nodule activity in VAM and nonVAM plants to drought are interpreted in terms of demand for nodule P and carbohydrates and of the effects of dehydration on O₂ diffusion through nodule tissue.     

The Glycine-Glomus-Bradyrhizobium symbiosis. IX. Nutritional, morphological and physiological responses of nodulated soybean to geographic isolates of the mycorrhizal fungus Glomus mosseae.

The objective of the work was to determine differences in plant response to geographic isolates of a vesicular-arbuscular mycorrhizal (VAM) fungus, and to demonstrate the need for such determinations in the selection of desirable host-endophyte combinations for practical applications. Soybean ( Glycine max (L.) Merr.) plants were inoculated with Bradyrhizobium japonicum and isolates of the VAM-fungal morphospecies Glomus mosseae (Nicol. & Gerd.) Gerd. and Trappe, collected from an arid (AR), semiarid (SA) or mesic (ME) area. Inoculum potentials of the VAM-fungal isolates were determined and the inocula equalized, achieving the same level of root colonization (41%, P >0.05) at harvest (50 days). Plants of the three VAM treatments (AR, SA and ME) were evaluated against von VAM controls. Significant differences in plant response to colonization were found in dry mass, leaf K, N and P concentrations, and in root/shoot, nodule/root, root length/leaf area and root length/root mass ratios. The differences were most pronounced and consistent between the AR and all other treatments. Photosynthesis and nodule activity were higher ( P <0.05) in all VAM treatments, but only the AR plants had higher ( P <0.05) photosynthetic water-use efficiency than the controls. Nodule activity, evaluated by H₂ evolution and C₂H₂ reduction, differed significantly between treatments. The results are discussed in terms of nutritional and non-nutritional effects of VAM colonization on the development and physiology of the tripartite soybean association in the light of intraspecific variability within the fungal endophyte.     

Relationship between C2H2 reduction, H2 evolution and 15N2 fixation in root nodules of pea (Pisum sativum)

The quantitative relationship between C₂H₂ reduction, H₂ evolution and ¹⁵N₂ fixation was investigated in excised root nodules from pea plants ( Pisum sativum L. cv. Bodil) grown under controlled conditions. The C₂H₂/N₂ conversion factor varied from 3.31 to 5.12 between the 32nd and the 67th day after planting. After correction for H₂ evolution in air, the factor (C₂H₂-H₂)/N₂ decreased to values near the theoretical value 3, or in one case to a value significantly ( P < 0.05) below 3. The proportion of the total electron flow through nitrogenase, which is not wasted in H₂ production but used for N₂ reduction, is often stated as the relative efficiency (1-H₂/C₂H₂). This factor varied significantly ( P < 0.05) during the growth period. The actual allocation of electrons to H₂ and N₂, expressed as the H₂/N₂ ratio, was independent of plant age, however. This discrepancy and the observation that the (C₂H₂-H₂)/N₂ conversion factor tended to be lower than 3, suggests that the C₂H₂reduction assay underestimates the total electron flow through nitrogenase.     

Effect of H2 evolution on 15N2 fixation, C2H2 reduction and relative efficiency of leguminous symbionts

The (C₂H₄+ H_2(C2H₂))/¹⁵N₂ ratios of 15 clover- Rhizobium symbionts. soybean, and black medick symbionts were measured. Relative efficiency based on the C2H4 production and on ¹⁵N₂ incorporation were compared, and in most symbionts there was little difference between the two measures of relative efficiency. Total measurable electron flux through nitrogenase during acetylene reduction and ¹⁵N₂ incorporation were nearly equal for most symbionts studied. The relative efficiency and the (C₂H₄+ H_2(C2H₂))/¹⁵N₂ ratio showed an inverse correlation. Use of this ratio appears preferable to use of the ratio of C2H2 reduction/N₂ reduction. Some evolution of H₂ was observed in the presence of C₂H₂.     

Relationship between photosystem II activity and CO2 fixation in leaves

There is now potential to estimate photosystem II (PSII) activity in vivo from chlorophyll fluorescence measurements and thus gauge PSII activity per CO₂ fixed. A measure of the quantum yield of photosystem II, Φ_II (electron/photon absorbed by PSII), can be obtained in leaves under steady-state conditions in the light using a modulated fluorescence system. The rate of electron transport from PSII equals Φ_II times incident light intensity times the fraction of incident light absorbed by PSII. In C₄ plants, there is a linear relationship between PSII activity and CO₂ fixation, since there are no other major sinks for electrons; thus measurements of quantum yield of PSII may be used to estimate rates of photosynthesis in C₄ species. In C₃ plants, both CO₂ fixation and photorespiration are major sinks for electrons from PSII (a minimum of 4 electrons are required per CO₂, or per O₂ reacting with RuBP). The rates of PSII activity associated with photosynthesis in C₃ plants, based on estimates of the rates of carboxylation (v_o) and oxygenation (v_o) at various levels of CO₂ and O₂, largely account for the PSII activity determined from fluorescence measurements. Thus, in C₃ plants, the partitioning of electron flow between photosynthesis and photorespiration can be evaluated from analysis of fluorescence and CO₂ fixation.     

Interaction of elevated CO2 and O3 on growth, photosynthesis and respiration of three perennial species grown in low and high nitrogen

Seedlings of three species native to central North America, a C₃ tree, Populus tremuloides Michx., a C₃ grass, Agropyron smithii Rybd., and a C₄ grass, Bouteloua curtipendula Michx., were grown in all eight combinations of two levels each of CO₂, O₃ and nitrogen (N) for 58 days in a controlled environment. Treatment levels consisted of 360 or 674 μmol mol^-1 CO₂, 3 or 92 nmol mol^-1 O₃, and 0.5 or 6.0 m M N. In situ photosynthesis and relative growth rate (RGR) and its determinants were obtained at each of three sequential harvests, and leaf dark respiration was measured at the second and third harvests. In all three species, plants grown in high N had significantly greater whole-plant mass, RGR and photosynthesis than plants grown in low N. Within a N treatment, elevated CO₂ did not significantly enhance any of these parameters nor did it affect leaf respiration. However, plants of all three species grown in elevated CO₂ had lower stomatal conductance compared to ambient CO₂-exposed plants. Seedlings of P. tremuloides (in both N treatments) and B. curtipendula (in high N) had significant ozone-induced reductions in whole-plant mass and RGR in ambient but not under elevated CO_2. This negative O₃ impact on RGR in ambient CO₂ was related to increased leaf dark respiration, decreased photosynthesis and/or decreased leaf area ratio, none of which were noted in high O₃ treatments in the elevated CO₂ environment. In contrast, A. smithii was marginally negatively affected by high O_3.     

Effect of enhanced atmospheric CO2 on mycorrhizal colonization by Glomus mosseae in Plantago lanceolata and Trifolium repens

Plantago lanceolata L. and Trifolium repens L. were grown for 16 wk in ambient (360 μmol mol⁻¹) and elevated (610 μmol mol⁻¹) atmospheric CO₂. Plants were inoculated with the arbuscular mycorrhizal (AM) fungus Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe and given a phosphorus supply in the form of bonemeal, which would not be immediately available to the plants. Seven sequential harvests were taken to determine whether the effect of elevated CO₂ on mycorrhizal colonization was independent of the effect of CO₂ on plant growth. Plant growth analysis showed that both species grew faster in elevated CO₂ and that P. lanceolata had increased carbon allocation towards the roots. Elevated CO₂ did not affect the percentage of root length colonized (RLC); although total colonized root length was greater, when plant size was taken into account this effect disappeared. This finding was also true for root length colonized by arbuscules. No CO₂ effect was found on hyphal density (colonization intensity) in roots. The P content of plants was increased at elevated CO₂, although both shoot and root tissue P concentration were unchanged. This was again as a result of bigger plants at elevated CO₂. Phosphorus inflow was unaffected by CO₂ concentrations. It is concluded that there is no direct permanent effect of elevated CO₂ on mycorrhizal functioning, as internal mycorrhizal development and the mycorrhizal P uptake mechanism are unaffected. The importance of sequential harvests in experiments is discussed. The direction for future research is highlighted, especially in relation to C storage in the soil.     

Nitrogen fixation (C2H2 reduction) by Medicago nodules and bacteroids under sodium chloride stress

Medicago ciliaris (L.) All., a salt-tolerant legume, was not nodulated by Rhizobium meliloti (2011), a strain commonly used for field inoculation of alfalfas. A strain of Rhizobium meliloti (ABS7) was isolated from saline Algerian soils. It is generally more salt-resistant than strain 2011, exhibits a higher rate of growth and induces the formation of nodules on M. ciliaris . C₂H₂ reduction activity of M. ciliaris nodules was inhibited by 50% in the presence of 200 m M NaCl in the culture medium. whereas 100 m M NaCl was sufficient to inhibit the activity of nodules of M. sativa (L. cv. Europe). C₂H₂ reduction by bacteroids, isolated from nodules of the two species of alfalfa, was directly inhibited by the presence of NaCl in the incubation medium. In both cases, glucose could support bacteroid nitrogen fixation, but only in a narrow range of O₂ tensions. Bacteriods from M. ciliaris were more tolerant to salt than M. sativa ones. The salt resistance of bacteroids from nodules of plants watered with NaCl solutions was not improved in either species. Salt directly added to the incubation mixture of bacteroids or to the culture medium of plants inhibited O₂ uptake of bacteroids isolated from nodules of both M. ciliaris and M. sativa . The depressive effect of NaCl on bacteroid C₂H₂ reduction could be directly related to the drop in bacteroid respiration. The nitrogen fixation capacity of the M. ciliaris-Rhizobium meliloti (ABS7) symbiosis under saline conditions leads us to recommend the introduction of this association in salt-troubled areas.     

Nodulation and nitrogenase activity in nitrogen-fixing woody plants stimulated by CO2 enrichment of the atmosphere

The responses of three species of nitrogen-fixing trees to CO₂ enrichment of the atmosphere were investigated under nutrient-poor conditions. Seedlings of the legume, Robinia pseudoacacia L. and the actinorhizal species, Alnus glutinosa (L.) Gaertn. and Elaeagnus angustifolia L. were grown in an infertile forest soil in controlled-environment chambers with atmospheric CO₂ concentrations of 350 μl ⁻¹ (ambient) or 700 μl ⁻¹. In R. pseudoacacia and A. glutinosa , total nitrogenase (N₂ reduction) activity per plant, assayed by the acetylene reduction method, was significantly higher in elevated CO₂, because the plants were larger and had more nodule mass than did plants in ambient CO₂. The specific nitrogenase activity of the nodules, however, was not consistently or significantly affected by CO₂ enrichment. Substantial increases in plant growth occurred with CO₂ enrichment despite probable nitrogen and phosphorus deficiencies. These results support the premises that nutrient limitations will not preclude growth responses of woody plants to elevated CO₂ and that stimulation of symbiotic activity by CO₂ enrichment of the atmosphere could increase nutrient availability in infertile habitats.     

Defoliation effects on mycorrhizal colonization, nitrogen fixation and photosynthesis in the Glycine-Glomus-Rhizobium symbiosis

Soybean [ Glycine max (L.) Merr. cv. Wells] plants grown in a greenhouse were inoculated with Rhizobium japonicum strain 61A118 and the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus fasciculatum (Thaxt. sensu Gerd.) Gerd. & Trappe. Plants were defoliated (26, 48 and 66%) throughout the growth period and evaluated for VAM colonization, N₂, fixation and photosynthesis at harvest (six weeks). Photosynthate stress as a result of defoliation affected nodulation and nodule activity most severely. Colonization of the roots by the VAM fungus was little affected in comparison, and the intensity of colonization increased with increasing stress. The CO₂-exchange rate decreased less with defoliation than did leaf mass, and photosynthetic efficiency increased with the severity of defoliation. The increase in photosynthetic efficiency was significantly correlated with increases in leaf P (r = 0.91) and N (r = 0.97) concentrations. The results suggest that the VAM fungus should not be regarded as a simple P source and C sink in the tripartite legume association. Threeway source/sink relationships (VAM-P, Rhizobium-N, and host leaf-C) are discussed.     

N2(C2H2)-fixation in early stages of a primary succession on a reclaimed salt marsh

Nitrogen fixation activity was determined for Lotus tenuis. Medicago lupulina and Trifolium pratense . The three species grew in clones in grassland in an area reclaimed from brackish water in the 1940s. The N₂[C₂H₂]-fixation was measured in soil cores throughout 1974 and 1975. From cores taken in dense and uniform stands of the species, the yearly N₂[C₂H₂]-fixation at maximum cover was estimated. L. tenuis fixed about 4 g N m⁻² yr⁻¹ (area with max. cover 130%), i.e. 30–56% of its requirement. Both M. lupulina and T. pratense fixed about 7 g N m⁻² yr⁻¹ (maximum cover 37% and 80%) i.e. 67% of their N-requirement. Average N₂[C₂H₂]-fixation for the whole area was 0.4 g N m⁻² yr⁻¹, considerably less than the N-addition through rainfall.     

Effects of age, supra-ambient oxygen and repeated assays on acetylene reduction and root respiration in pea

An open flow-through gas system was used to investigate the effect of plant age on nitrogenase activity in relation to root respiration (measured as CO₂ release) and supra-ambient O₂ levels in 24- to 51-day-old, nodulated Pisum sativum L. cv. Bodil. The effect of assaying plants repeatedly was also studied. The respiratory efficiency of nitrogenase [mol CO₂ (mol C₂H₄)⁻¹] and the relative decline in nitrogenase (EC 1.7.99.2) activity in response to introduction of C₂H₂ in the gas stream were unaffected by plant age. In contrast, the nitrogenase-linked respiration as a proportion of total root respiration increased with time. Accordingly, the specific respiration linked-to growth and maintenace of the noduled root system decreased with time. C₂H₂ reduction and root respiration were increased by supra-ambient O₂ levels, but the tolerance to high O₂ concentrations seemed to decrease with plant age. Repeated C₂H₂ assays on the same plants decreased their rate of growth and N accumulation: in addition, nitrogenase activity and root respiration were somewhat negatively affected. The results indicate that results from experiments with plants of different ages cannot always be directly compared, and that repeated C₂H₂ assays on the same plants should be applied with caution in physiological work.     

Nitrogen nutrition of C3 plants at elevated atmospheric CO2 concentrations

The atmospheric CO₂ concentration has risen from the preindustrial level of approximately 290 μl l⁻¹ to more than 350 μl l⁻¹ in 1993. The current rate of rise is such that concentrations of 420 μl l⁻¹ are expected in the next 20 years. For C₃ plants, higher CO₂ levels favour the photosynthetic carbon reduction cycle over the photorespiratory cycle, resulting in higher rates of carbohydrate production and plant productivity. The change in balance between the two photosynthetic cycles appears to alter nitrogen and carbon metabolism in the leaf, possibly causing decreases in nitrogen concentrations in the leaf. This may result from increases in the concentration of storage carbohydrates of high molecular weight (soluble or insoluble) and/or changes in distribution of protein or other nitrogen containing compounds. Uptake of nitrogen may also be reduced at high CO₂ due to lower transpiration rates. Decreases in foliar nitrogen levels have important implications for production of crops such as wheat, because fertilizer management is often based on leaf chemical analysis, using standards estimated when the CO₂ levels were considerably lower. These standards will need to be re-evaluated as the CO₂ concentration continues to rise. Lower levels of leaf nitrogen will also have implications for the quality of wheat grain produced, because it is likely that less nitrogen would be retranslocated during grain filling.     

Canopy photosynthesis of crops and native plant communities exposed to long-term elevated CO2

Abstract. There have been seven studies of canopy photosynthesis of plants grown in elevated atmospheric CO₂: three of seed crops, two of forage crops and two of native plant ecosystems. Growth in elevated CO₂ increased canopy photosynthesis in all cases. The relative effect of CO₂ was correlated with increasing temperature: the least stimulation occurred in tundra vegetation grown at an average temperature near 10°C and the greatest in rice grown at 43°C. In soybean, effects of CO₂ were greater during leaf expansion and pod fill than at other stages of crop maturation. In the longest running experiment with elevated CO₂ treatment to date, monospecific stands of a C₃ sedge, Scirpus olneyi (Grey), and a C₄ grass, Spartina patens (Ait.) Muhl., have been exposed to twice normal ambient CO₂ concentrations for four growing seasons, in open top chambers on a Chesapeake Bay salt marsh. Net ecosystem CO₂ exchange per unit green biomass (NCE_b) increased by an average of 48% throughout the growing season of 1988, the second year of treatment. Elevated CO₂ increased net ecosystem carbon assimilation by 88% in the Scirpus olneyi community and 40% in the Spartina patens community.     

Elevated pyruvate,orthophosphate dikinase (PPDK) activity alters carbon metabolism in C3 transgenic potatoes with a C4 maize PPDK gene

Changes in carbon metabolism and δ¹³C value of transgenic potato plants with a maize pyruvate,orthophosphate dikinase (PPDK; EC 2.7.9.1) gene are reported. PPDK catalyzes the formation of phospho enol pyruvate (PEP), the initial acceptor of CO₂ in the C₄ photosynthetic pathway. PPDK activities in the leases of transgenic potatoes were up to 5.4‐fold higher than those of control potato plants (wild‐type and treated control plants). In the transgenic potato plants, PPDK activity in leaves was negatively correlated with pyruvate content (r²= 0.81), and was positively correlated with malate content (r²= 0.88). A significant increase in the δ¹³C value was observed in the transgenic potato plants, suggesting a certain contribution of PEP carboxylase as the initial acceptor of atmospheric CO₂. These data suggest that elevated PPDK activity may alter carbon metabolism and lead to a partial operation of C₄‐type carbon metabolism. However, since parameters associated with CO₂ gas exchange were not affected, the altered carbon metabolism had only a small effect on the total photosynthetic characteristics of the transgenic plants.     

Effect of darkness and CO2 starvation on NH4+ and NO3− assimilation in the unicellular alga Cyanidium caldarium

Cyanidium caldarium (Tilden) Geitler, a non-vacuolate unicellular alga, resuspended in medium flushed with air enriched with 5% CO₂, assimilated NH₄⁺ at high rates both in the light and in the dark. The assimilation of NO₃⁻, by contrast, was inhibited by 63% in the dark. In cell suspensions flushed with CO₂-free air, NH₄⁺ assimilation decreased with time both in the light and in the dark and ceased almost completely after 90 min. The addition of CO₂ completely restored the capacity of the alga to assimilate NH₄⁺. NO₃⁻ assimilation, by contrast, was 33% higher in the absence of CO₂ and was linear with time. It is suggested that NO₃⁻ and NH₄⁺ metabolism in C. caldarium are differently controlled in response to the light and carbon conditions of the cell.     

Respiration in a future, higher-CO2 world

Abstract. Apart from its impact on global warming, the annually increasing atmospheric [CO₂] is of interest to plant scientists primarily because of its direct influence on photosynthesis and photorespiration in C₃ species. But in addition, 'dark' respiration, another major component of the carbon budget of higher plants, may be affected by a change in [CO₂] independent of an increase in temperature. Literature pertaining to an impact of [CO₂] on respiration rate is reviewed. With an increase in [CO₂], respiration rate is increased in some cases, but decreased in others. The effects of [CO₂] on respiration rate may be direct or indirect. Mechanisms responsible for various observations are proposed. These proposed mechanisms relate to changes in: (1) levels of nonstructural carbohydrates, (2) growth rate and structural phytomass accumulation, (3) composition of phytomass, (4) direct chemical interactions between CO₂ and respiratory enzymes, (5) direct chemical interactions between CO₂ and other cellular components, (6) dark CO₂ fixation rate, and (7) ethylene biosynthesis rate. Because a range-of (possibly interactive) effects exist, and present knowledge is limited, the impact of future [CO₂] on respiration rate cannot be predicted. Theoretical considerations and types of experiments that can lead to an increase in the understanding of this issue are outlined.     

The effects of increasing CO2 on crop photosynthesis and productivity: a review of field studies

Abstract. Only a small proportion of elevated CO₂ studies on crops have taken place in the field. They generally confirm results obtained in controlled environments: CO₂ increases photosynthesis, dry matter production and yield, substantially in C₃ species, but less in C₄, it decreases stomatal conductance and transpiration in C₃ and C₄ species and greatly improves water-use efficiency in all plants. The increased productivity of crops with CO₂ enrichment is also related to the greater leaf area produced. Stimulation of yield is due more to an increase in the number of yield-forming structures than in their size. There is little evidence of a consistent effect of CO₂ on partitioning of dry matter between organs or on their chemical composition, except for tubers. Work has concentrated on a few crops (largely soybean) and more is needed on crops for which there are few data (e.g. rice). Field studies on the effects of elevated CO₂ in combination with temperature, water and nutrition are essential; they should be related to the development and improvement of mechanistic crop models, and designed to test their predictions.     

Effects of long-term elevated [CO2] from natural CO2 springs on Nardus stricta: photosynthesis, biochemistry, growth and phenology

Plants of Nardus stricta growing near a cold, naturally emitting CO₂ spring in Iceland were used to investigate the long-term (> 100 years) effects of elevated [CO₂] on photosynthesis, biochemistry, growth and phenology in a northern grassland ecosystem. Comparisons were made between plants growing in an atmosphere naturally enriched with CO₂ (≈ 790 μ mol mol^–1) near the CO₂ spring and plants of the same species growing in adjacent areas exposed to ambient CO₂ concentrations (≈360 μ mol mol^–1). Nardus stricta growing near the spring exhibited earlier senescence and reductions in photosynthetic capacity (≈25%), Rubisco content (≈26%), Rubisco activity (≈40%), Rubisco activation state (≈23%), chlorophyll content (≈33%) and leaf area index (≈22%) compared with plants growing away from the spring. The potential positive effects of elevated [CO₂] on grassland ecosystems in Iceland are likely to be reduced by strong down-regulation in the photosynthetic apparatus of the abundant N. stricta species.