Biological dinitrogen fixation (acetylene reduction) associated with Florida mangroves

Biological dinitrogen fixation in mangrove communities of the Tampa Bay region of South Florida was investigated using the acetylene reduction technique. Low rates of acetylene reduction (0.01 to 1.84 nmol of C(2)H(4)/g [wet weight] per h) were associated with plant-free sediments, while plant-associated sediments gave rise to slightly higher rates. Activity in sediments increased greatly upon the addition of various carbon sources, indicating an energy limitation for nitrogenase (C(2)H(2)) activity. In situ determinations of dinitrogen fixation in sediments also indicated low rates and exhibited a similar response to glucose amendment. Litter from the green macroalga, Ulva spp., mangrove leaves, and sea grass also gave rise to significant rates of acetylene reduction.Higher rates of nitrogenase activity (15 to 53 nmol of C(2)H(4)/g [wet weight] per h were associated with washed excised roots of three Florida mangrove species [Rhizophora mangle L., Avicennia germinans (L) Stern, and Laguncularia racemosa Gaertn.] as well as with isolated root systems of intact plants (11 to 58 mug of N/g [dry weight] per h). Following a short lag period, root-associated activity was linear and did not exhibit a marked response to glucose amendment. It appears that dinitrogen-fixing bacteria in the mangrove rhizoplane are able to use root exudates and/or sloughed cell debris as energy sources for dinitrogen fixation.     

N2 Fixation Associated with Decaying Leaves of the Red Mangrove (Rhizophora mangle)

N₂ (C₂H₂) fixation was associated with decaying leaves of Rhizophora mangle. The process was predominantly anaerobic, with about two-thirds of the nitrogenase activity being light dependent. Average N₂ fixation rates in the light were 11 μg of N per g (dry weight) per h for leaves that had decayed for 2 to 3 weeks. This nitrogen input is probably significant in the estuarine, detrital food chains linked to R. mangle.     

Photosynthesis in leaves and siliques of winter oilseed rape (Brassica napus L.)

The rate of photosynthesis and its relation to tissue nitrogen content was studied in leaves and siliques of winter oilseed rape (Brassica napus L.) growing under field conditions including three rates of nitrogen application (0, 100 or 200 kg N ha^-1) and two levels of irrigation (rainfed or irrigated at a deficit of 20 mm). The predominant effect of increasing N application under conditions without water deficiency was enhanced expansion of photosynthetically active leaf and silique surfaces, while the rate of photosynthesis per unit leaf or silique surface area was similar in the different N treatments. Thus, oilseed rape did not increase N investment in leaf area expansion before a decline in photosynthetic rate per unit leaf area due to N deficiency could be avoided. Much less photosynthetically active radiation penetrated into high-N canopies than into low-N canopies. The specific leaf area increased markedly in low light conditions, causing leaves in shade to be less dense than leaves exposed to ample light. In both leaves and siliques the photosynthetic rate per unit surface area responded linearly to increasing N content up to about 2 g m^-2, thus showing a constant rate of net CO₂ assimilation per unit increment in N (constant photosynthetic N use efficiency). At higher tissue N contents, photosynthetic rate responded less to changes in N status. Expressed per unit N, light saturated photosynthetic rate was three times higher in leaves than in silique valves, indicating a more efficient photosynthetic N utilization in leaves than in siliques. Nevertheless, from about two weeks after completion of flowering and onwards total net CO₂ fixation in silique valves exceeded that in leaves because siliques received much higher radiation intensities than leaves and because the leaf area declined rapidly during the reproductive phase of growth. Water deficiency in late vegetative and early reproductive growth stages reduced the photosynthetic rate in leaves and, in particular, siliques of medium- and high-N plants, but not of low-N plants.     

Nitrogen fixation (acetylene reduction) associated with duckweed (lemnaceae) mats

Duckweed (Lemnaceae) mats in Texas and Florida were investigated, using the acetylene reduction assay, to determine whether nitrogen fixation occurred in these floating aquatic macrophyte communities. N(2)-fixing microorganisms were enumerated by plating or most-probable-number techniques, using appropriate N-free media. Results of the investigations indicated that substantial N(2)-fixation (C(2)H(2)) was associated with duckweed mats in Texas and Florida. Acetylene reduction values ranged from 1 to 18 mumol of C(2)H(4) g (dry weight) day for samples incubated aerobically in light. Dark N(2) fixation was always two- to fivefold lower. 3-(3,4-Dichlorophenyl)-1,1-dimethylurea (7 to 10 muM) reduced acetylene reduction to levels intermediate between light and dark incubation. Acetylene reduction was generally greatest for samples incubated anaerobically in the light. It was estimated that 15 to 20% of the N requirement of the duckweed could be supplied through biological nitrogen fixation. N(2)-fixing heterotrophic bacteria (10 cells g [wet weight] and cyanobacteria (10 propagules g [wet weight] were associated with the duckweed mats. Azotobacter sp. was not detected in these investigations. One diazotrophic isolate was classified as Klebsiella.     

Photosynthesis-Nitrogen Relationships in Pioneer Plants of Disturbed Tropical Montane Forest Sites

Tropical forest disturbances lead to the establishment of secondary successional plant communities constituted by light demanding species with high relative growth rate that conduct to rapid canopy closure. Two main strategies for N nutrition are: (a) mineral N acquisition in the form of NH4 and NO3, and (b) symbiotic atmospheric N2 fixation. Given the high N requirement for maximization of leaf area and radiant energy absorption, we hypothesize that contrasting strategies of N nutrition in these environments are reflected in leaf photosynthetic characteristics. We compared the N-photosynthesis relationships and carbon balance parameters per unit leaf area as they vary with age in two species with contrasting N acquisition strategies: a N2-fixer Crotalaria anagyroides HBK (Papilionoideae), and a mineral-N user Verbesina turbacensis HBK (Asteraceae). N2 fixation capacity was associated to higher specific leaf area (SLA), higher photosynthetic capacity (Pmax) per unit leaf area and leaf mass, and higher N content per unit leaf mass. The N2-fixer species showed higher slope in the relationship Pmax-N per unit leaf mass and area when compared to the leaves of non-fixer species. Moreover, the intrinsic photosynthetic N use efficiency (Pmax/N) was higher in the N2 fixer than in leaves of the non-fixer species. Changes in N due to leaf age resulted in larger changes in CO2 flux density at the leaf level in the N2-fixer species. The higher photosynthetic capacity of the N2-fixer species was mechanistically related to higher stomatal conductance, internal CO2 concentration (ci) values closer to atmospheric CO2 concentration (ca), and lower intrinsic water use efficiency than the mineral N-user species. Despite their higher Pmax per unit leaf area, total non-structural saccharides concentration was lower in mature leaves of the N2-fixer plant as compared to the non-fixer counterpart. This might be caused by the presence of a larger root sink (symbionts) stimulating saccharides export and higher diurnal respiration rates.     

Partitioning of leaf nitrogen with respect to within canopy light exposure and nitrogen availability in peach (Prunus persica)

Summary Relationships between leaf nitrogen content and within canopy light exposure were studied in mature nectarine peach trees (Prunus persica cv. Fantasia) that had received 0, 112, 196, 280 or 364 kg of fertilizer nitrogen per hectare per year for the previous 3 years. The relationships between light saturated leaf CO₂ assimilation rates and leaf nitrogen concentration were also determined on trees in the highest and lowest nitrogen fertilization treatments. The slope of the linear relationship between leaf N content per unit leaf area and light exposure was similar for all nitrogen treatments but the y-intercept of the relationship increased with increasing N status. The slope of the relationship between leaf N content per unit leaf area and light saturated CO₂ assimilation rates was greater for the high N trees than the low N trees, but maximum measured leaf CO₂ assimilation rates were similar for both the high and low N treatments. A diagrammatic model of the partitioning of leaf photosynthetic capacity with respect to leaf light exposure for high and low nitrogen trees suggests that the major influence of increased N availability is an increase in the photosynthetic capacity of partially shaded leaves but not of the maximum capacity of highly exposed leaves.     

Leaf ureide degradation and N(2) fixation tolerance to water deficit in soybean

Accumulation of ureides in leaves is associated with the sensitivity of N(2) fixation in soybean to soil water deficit. Consequently, ureide degradation in leaves may be a key to increasing soybean tolerance to dry soils. Previous research indicated that allantoic acid degradation is catalysed by different enzymes in cultivars Maple Arrow and Williams. The enzyme found in Williams requires manganese as a cofactor. The first objective of this study was to determine if the two degradation pathways were associated with differences in N(2) sensitivity to soil water deficits. N(2) fixation of Williams grown on low-Mn soil was sensitive to stress, but it was relatively tolerant when grown on soil amended with Mn. N(2) fixation in Maple Arrow was relatively tolerant of soil drying regardless of the Mn treatment. The second objective of this study was to expand the study of the degradation pathway to nine additional genotypes. Based on ureide degradation in the presence and absence of Mn, these genotypes also segregated for the two degradation pathways. Those genotypes with the Mn-dependent pathway tended to have drought-sensitive N(2) fixation, but there was one exception. The genotypes not requiring Mn for ureide degradation were drought-tolerant except for one genotype. These results demonstrated the possibility for increasing N(2) fixation tolerance to soil water deficits in soybean by selection of lines with high ureide degradation rates, which were commonly associated with the Mn-independent pathway.     

Inhibition of Dark CO(2) Fixation and Photosynthesis in Leaf Discs of Corn Susceptible to the Host-specific Toxin Produced by Helminthosporium maydis, Race T

The host-specific toxin produced by Helminthosporium maydis, race T, causes 50% inhibition of dark fixation of ¹⁴CO₂ by leaf discs of susceptible (Texas male sterile) corn when it is diluted to approximately 1/10,000 of the volume of the original fungus culture filtrate. Dilutions of 1/10 or less are required for equivalent inhibition of discs prepared from resistant (N) corn. Root growth and photosynthesis were considerably less sensitive (dilution values 1/3000 and 1/1200, respectively), as was leakage of ¹⁴C induced by toxin from preloaded discs. Based on literature values for dilutions causing ion leakage or inhibition of mitochondrial oxidation, toxin dilutions several orders of magnitude greater bring about inhibition of dark CO₂ fixation. Preincubation of discs in light increased sensitivity of dark fixation to toxin and an effect of light on symptom development was shown. Phosphoenolypruvate carboxylase activity in extracts of roots or leaves was not affected by toxin nor was the enzyme level altered in excised leaves treated with toxin. Inhibition of dark fixation of CO₂ provides a bioassaay for race T toxin which is both reliable and rapid.     

Complex adjustments of photosynthetic potentials and internal diffusion conductance to current and previous light availabilities and leaf age in Mediterranean evergreen species Quercus ilex

Mature non-senescent leaves of evergreen species become gradually shaded as new foliage develops and canopy expands, but the interactive effects of integrated light during leaf formation (Q(int)G), current light (Q(int)C) and leaf age on foliage photosynthetic competence are poorly understood. In Quercus ilex L., we measured the responses of leaf structural and physiological variables to Q(int)C and Q(int)G for four leaf age classes. Leaf aging resulted in increases in leaf dry mass per unit area (M(A)), and leaf dry to fresh mass ratio (D(F)) and decreases in N content per dry mass (N(M)). N content per area (N(A)) was independent of age, indicating that decreases in N(M) reflected dilution of leaf N because of accumulation of dry mass (NA = N(M) M(A)). M(A), D(F) and N(A) scaled positively with irradiance, whereas these age-specific correlations were stronger with leaf growth light than with current leaf light. Area-based maximum ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) carboxylase activity (V(cmax)A), capacity for photosynthetic electron transport (J(max)A) and the rate of non-photorespiratory respiration in light (R(d)A) were also positively associated with irradiance. Differently from leaf structural characteristics, for all data pooled, these relationships were stronger with current light with little differences among leaves of different age. Acclimation to current leaf light environment was achieved by light-dependent partitioning of N in rate-limiting proteins. Mass-based physiological activities decreased with increasing leaf age, reflecting dilution of leaf N and a larger fraction of non-photosynthetic N in older leaves. This resulted in age-dependent modification of leaf photosynthetic potentials versus N relationships. Internal diffusion conductance (g(m)) per unit area (g(m)A) increased curvilinearly with increasing irradiance for two youngest leaf age classes and was independent of light for older leaves. In contrast, g(m) per dry mass (g(m)M) was negatively associated with light in current-year leaves. Greater photosynthetic potentials and moderate changes in diffusion conductance resulted in greater internal diffusion limitations of photosynthesis in higher light. Both area- and mass-based g(m) decreased with increasing leaf age. The decrease in diffusion conductance was larger than changes in photosynthetic potentials, leading to larger CO2 drawdown from leaf internal air space to chloroplasts (delta(c)) in older leaves. The increases in diffusion limitations in older leaves and at higher light scaled with age- and light-dependent increases in MA and D(F). Overall, our study demonstrates a large potential of foliage photosynthetic acclimation to changes in leaf light environment, but also highlights enhanced structural diffusion limitations in older leaves that result from leaf structural acclimation to previous rather than to current light environment and accumulation of structural compounds with leaf age.     

Kinetic characterization of reduced pyridine nucleotide dehydrogenases (duroquinone-dependent) in cucurbita microsomes

Stomatal conductances of normally oriented and inverted leaves were measured as light levels (photosynthetic photon flux densities) were increased to determine whether abaxial stomata of Vicia faba leaves were more sensitive to light than adaxial stomata. Light levels were increased over uniform populations of leaves of plants grown in an environmental chamber. Adaxial stomata of inverted leaves reached maximum water vapor conductances at a light level of 60 micromoles per square meter per second, the same light level at which abaxial stomata of normally oriented leaves reached maximum conductances. Abaxial stomata of inverted leaves reached maximum conductances at a light level of 500 micromoles per square meter per second, the same light level at which adaxial stomata of normally oriented leaves reached maximum conductances. Maximum conductances in both normally oriented and inverted leaves were about 200 millimoles per square meter per second for adaxial stomata and 330 millimoles per square meter per second for abaxial stomata. Regardless of whether leaves were normally oriented or inverted, when light levels were increased to values high enough that upper leaf surfaces reached maximum conductances (about 500 micromoles per square meter per second), light levels incident on lower, shaded leaf surfaces were just sufficient (about 60 micromoles per square meter per second) for stomata of those surfaces to reach maximum conductances. This `coordinated' stomatal opening on the separate epidermes resulted in total leaf conductances for normally oriented and inverted leaves that were the same at any given light level. We conclude that stomata in abaxial epidermes of intact Vicia leaves are not more sensitive to light than those in adaxial epidermes, and that stomata in leaves of this plant do not respond to light alone. Additional factors in bulk leaf tissue probably produce coordinated stomatal opening on upper and lower leaf epidermes to optimally meet photosynthetic requirements of the whole leaf for CO₂.     

Gas exchange in the filamentous cyanobacterium Nostoc punctiforme strain ATCC 29133 and Its hydrogenase-deficient mutant strain NHM5

Nostoc punctiforme ATCC 29133 is a nitrogen-fixing, heterocystous cyanobacterium of symbiotic origin. During nitrogen fixation, it produces molecular hydrogen (H(2)), which is recaptured by an uptake hydrogenase. Gas exchange in cultures of N. punctiforme ATCC 29133 and its hydrogenase-free mutant strain NHM5 was studied. Exchange of O(2), CO(2), N(2), and H(2) was followed simultaneously with a mass spectrometer in cultures grown under nitrogen-fixing conditions. Isotopic tracing was used to separate evolution and uptake of CO(2) and O(2). The amount of H(2) produced per molecule of N(2) fixed was found to vary with light conditions, high light giving a greater increase in H(2) production than N(2) fixation. The ratio under low light and high light was approximately 1.4 and 6.1 molecules of H(2) produced per molecule of N(2) fixed, respectively. Incubation under high light for a longer time, until the culture was depleted of CO(2), caused a decrease in the nitrogen fixation rate. At the same time, hydrogen production in the hydrogenase-deficient strain was increased from an initial rate of approximately 6 micro mol (mg of chlorophyll a)(-1) h(-1) to 9 micro mol (mg of chlorophyll a)(-1) h(-1) after about 50 min. A light-stimulated hydrogen-deuterium exchange activity stemming from the nitrogenase was observed in the two strains. The present findings are important for understanding this nitrogenase-based system, aiming at photobiological hydrogen production, as we have identified the conditions under which the energy flow through the nitrogenase can be directed towards hydrogen production rather than nitrogen fixation.     

Effects of Light and Nutrients on Leaf Size, CO(2) Exchange, and Anatomy in Wild Strawberry (Fragaria virginiana)

Plants of a single genotype of wild strawberry, Fragaria virginiana Duchesne, were grown with or without fertilizer in high (406 microeinsteins per square meter per second) and low (80 microeinsteins per square meter per second) light. High-light leaves were thicker than low-light leaves and had greater development of the mesophyll. Within a light level, high-nutrient leaves were thicker, but the proportions of leaf tissues did not change with nutrient level. Maximum net CO₂ exchange rate and leaf size were greatest in high-light, high-nutrient leaves and lowest in high-light, low-nutrient leaves. Changes in mesophyll cell volume largely accounted for differences in CO₂ exchange rate in low-light leaves, but not in high-light leaves.

Leaf size in these experiments was apparently determined by nutrient and carbon supply. This may explain the observation that the largest leaves produced by wild strawberries in the field occur in high-light, mesic habitats, rather than in shady habitats.

     

Acetylene reduction (nitrogen fixation) associated with corn inoculated with Spirillum.

Sorghum and corn breeding lines were grown in soil in field and greenhouse experiments with and without an inoculum of N2-fixing in Spirillum strains from Brazil. Estimated rates of N2 fixation associated with field-grown corn and sorghum plants were less than 4 g of N2/ha per day. The mean estimated N2-fixation rates determined on segments of roots from corn inoculated with Spirillum and grown in the greenhouse at 24 to 27 degrees C were 15 g of N2/ha per day (16 inbreds), 25 g of N2/ha per day (six hybrids), and 165 g of N2/ha per day for one hybird which was heavily inoculated. The corresponding mean rates determined from measurements of in situ cultures of the same series of corn plants (i.e., 16 inbreds, six hybrids, and one heavily inoculated hybrid) were 0.4, 2.3, and 1.1 g of N2/ha per day, respectively. Lower rates of C2H2 reduction were associated with control corn cultures which had been treated with autoclaved Spirillum than with cultures inoculated with live Spirillum. No C2H2 reduction was detected in plant cultures treated with ammonium nitrate. Numbers of nitrogen-fixing bacteria on excised roots of corn plants increased an average of about 30-fold during an overnight preincubation period, and as a result acetylene reduction assays of root samples after preincubation failed to serve as a valid basis for estimating N2 fixation by corn in pot cultures. Plants grown without added nitrogen either with or without inoculum exhibited severe symptoms of nitrogen deficiency and in most cases produced significantly less dry weight than those supplied with fixed nitrogen. Although substantial rates of C2H2 reduction by excised corn roots were observed after preincubation under limited oxygen, the yield and nitrogen content of inoculated plants and the C2H2-reduction rates by inoculated pot cultures of corn, in situ, provided no evidence of appreciable N2 fixation.     

Root respiration associated with nitrogenase activity (c(2)h(2)) of soybean, and a comparison of estimates

Root respiration associated with symbiotic fixation in soybean (Glycine max [L.] Merr.) was estimated by four methods.

Averaged over the life of the plant, the root respires 5.8 milligrams C per milligram N accumulated from fixation. When nitrogenase (C₂H₂) activity and root respiration were decreased by treating roots briefly with 1.0 atmosphere O₂, the respiration associated with nitrogenase was estimated as 2.10 micromoles CO₂ per micromole C₂H₄.

When nitrogenase activity and respiration were decreased by addition of nitrate, the respiration associated with fixation was calculated as 2.90 micromoles CO₂ per micromole C₂H₄. Removing nodules from roots decreased fixation and root respiration, and the ratio was 4.08 micromoles CO₂ per micromole C₂H₄. When soybean plants were kept in prolonged darkness, then returned to light, the associated drop and recovery of respiration and nitrogenase activity had a ratio of 4.36 micromoles CO₂ per micromole C₂H₂.

     

Nitrogen from senescing lower leaves of common bean is re-translocated to nodules and might be involved in a N-feedback regulation of nitrogen fixation

The objective of the present study was to elucidate whether remobilized N from lower leaves is involved in causing the drop in N(2) fixation during pod-filling in common bean (Phaseolus vulgaris L). Moreover, we addressed the question of whether remobilized N from lower leaves would reach the nodules. Nodulated common bean plants were grown in a growth chamber in quartz sand. During a 2-week period, at vegetative and at reproductive growth, 50% of the leaves (lower part) were either excised or individually darkened, thereby removing the same photosynthetic capacity yet allowing N to be remobilized from the darkened leaves. Moreover, at the vegetative growth period, three lower leaves per plants were (15)N labelled by applying (15)NH(4)NO(3) prior to imposing the darkening treatment. Leaf darkening at vegetative growth induced N remobilization as well as reduced N(2)-fixation rates and growth. Leaf excision at reproductive growth enhanced N(2) fixation. Changes in N(2)-fixation rates were in all cases the result of altered growth rates, while the % N in the whole plant and in various plant parts remained conserved. Directly after leaf labelling, but also at the end of the vegetative growth period, substantial amounts of (15)N from the leaves could be recovered in nodules in the control, and in higher amounts in the leaf-darkening treatment. It is proposed that nitrogen from leaves circulates within the plant via nodules, and that the strength or composition of this circular flow may be the signal for a putative N-feedback effect.     

Why abaxial illumination limits photosynthetic carbon fixation in spinach leaves

Limitations of carbon fixation within spinach leaves due to light and CO2 were investigated. Under equivalent photon fluxes, carbon fixation was higher when leaves were irradiated adaxially compared to abaxially. Maximal carbon fixation occurred in the middle of the palisade mesophyll under adaxial illumination, whereas, maximal carbon fixation occurred in the spongy mesophyll under abaxial illumination. Total carbon fixation and the pattern of carbon fixation across leaves were similar, when leaves were irradiated with 800 micromol quanta m(-2) s(-1) either adaxially alone or adaxially plus abaxially (1,600 micromol quanta m(-2) s(-1)). In contrast, when both leaf surfaces were irradiated simultaneously with 200 micromol quanta m(-2) s(-1), total carbon fixation increased and carbon fixation in the middle of the leaf was higher compared to leaves irradiated unilaterally with the low light. Feeding 14CO2 through either the adaxial or abaxial leaf surface did not change the pattern of carbon fixation across the leaf. Increasing 14CO2 pulse-feeding times from 5 s to 60 s allowed more 14CO2 to be fixed but did not change the pattern of 14CO2 fixation across the leaf. We concluded that in spinach, the distribution of both light and Rubisco activity within leaves has significant effects on the patterns of carbon fixation across leaves; whereas CO2 diffusion does not appear to affect the carbon fixation pattern within spinach leaves.     

N(2) Fixation by Azospirillum brasilense and Its Incorporation into Host Setaria italica

Growth and nitrogen fixation were followed during the life cycle of Setaria italica (foxtail millet) inoculated with Azospirillum brasilense in controlled-environment growth chambers. The plants were fertilized at seeding with a limiting amount of combined nitrogen and maintained with an N-free mineral solution. During maturation of the plants, substantial nitrogenase activity, measured by acetylene reduction, developed in the rhizosphere, with total fixation estimated to be equivalent to 20% of the N in the inoculated plants. The peak of this activity coincided with depletion of soluble nitrogen from the system, which in turn was reflected by a sharp decrease in the nitrate reductase activity of the leaves. A. brasilense was found in association with the root populations at 8 x 10 cells per gram of dry weight. An increase in shoot growth occurred at this time, but no significant increase in total plant nitrogen could be demonstrated. N(2) enrichment experiments confirmed that fixation was occurring, but only about 5% of the nitrogen fixed by A. brasilense was incorporated into the plants within 3 weeks. There was thus no evidence of direct bacterium-to-plant transport of fixed nitrogen, but rather a slow transfer suggesting the gradual death of bacteria and subsequent mineralization of their nitrogen, at least under growth-room conditions.     

Effects of growth light and nitrogen nutrition on the organization of the photosynthetic apparatus in leaves of a C4 plant, Amaranthus cruentus

Properties of C4 photosynthesis were examined in Amaranthus cruentus L. (NAD-malic enzyme (ME) subtype, dicot) grown under different light and nitrogen (N) conditions, from the viewpoint of N investment into their photosynthetic components. In low-light (LL) leaves, chlorophyll content per leaf area was greater and chlorophyll alb ratio was lower than in high-light (HL) leaves. These indicate that LL leaves invest more N into their light-harvesting systems. However, this N investment did not contribute to the increase in the quantum yield of photosynthesis on the incident photon flux density (PFD) basis (Qi) in LL leaves. N allocation to ribulose 1,5-bisphosphate carboxylasel oxygenase (Rubisco) was significantly higher in HL-high N (HN) leaves than in other leaves. On the other hand, N allocation to C4 enzymes [phosphoenolpyruvate carboxylase (PEPC) and pyruvate Pi dikinase (PPDK)] was unaffected by the growth conditions. Maximum photosynthetic rates (Pmax) per Rubisco content were similar irrespective of the growth light treatments. Carbon isotope ratios (delta13 C) in the leaf dry matter were more negative in LL leaves than in HL leaves (LL = -19.3% per hundred, HL = -16.0% per hundred) and independent of leaf N. Vein density was highest in HL-HN leaves, and leaf thickness was unaffected by the growth light treatments. From these results, we conclude that A. cruentus leaves would not acclimate efficiently to low growth light.     

Evidence that P deficiency induces N feedback regulation of symbiotic N2 fixation in white clover (Trifolium repens L.)

Trifolium repens L. was grown to test the following hypotheses: when P is deficient (i) N2 fixation decreases as a result of the plant's adaptation to the low N demand, regulated by an N feedback mechanism, and (ii) the decrease in the photosynthetic capacity of the leaves does not limit N2 fixation. Severe P deficiency prevented nodulation or stopped nodule growth when the P deficiency occurred after the plants had formed nodules. At low P, the proportion of whole-plant-N derived from symbiotic N2 fixation decreased, whereas specific N2 fixation increased and compensated partially for poor nodulation. Leaf photosynthesis was reduced under P deficiency due to low Vc,max and Jmax. Poor growth or poor performance of the nodules was not due to C limitation, because (i) the improved photosynthetic performance at elevated pCO2 had no effect on the growth and functioning of the nodules, (ii) starch accumulated in the leaves, particularly under elevated pCO2, and (iii) the concentration of WSC in the nodules was highest under P deficiency. Under severe P deficiency, the concentrations of whole-plant-N and leaf-N were the highest, indicating that the assimilation of N exceeded the amount of N required by the plant for growth. This was clearly demonstrated by a strong increase in asparagine concentrations in the roots and nodules under low P supply. This indicates that nodulation and the proportion of N derived from symbiotic N2 fixation are down-regulated by an N feedback mechanism.     

Growth and N2 fixation in an Alnus hirsuta (Turcz.) var. sibirica stand in Japan

To estimate the N₂ fixation ability of the alder (Alnus hirsuta (Turcz.) var. sibirica), we examined the seasonal variation in nitrogenase activity of nodules using the acetylene reduction method in an 18-year-old stand naturally regenerated after disturbance by road construction in Japan. To evaluate the contribution of N₂ fixation to the nitrogen (N) economy in this alder stand, we also measured the phenology of the alder, the litterfall, the decomposition rate of the leaf litter, and N accumulation in the soil. The acetylene reduction activity per unit nodule mass (ARA) under field conditions appeared after bud break, peaked the maximum in midsummer after full expansion of the leaves, and disappeared after all leaves had fallen. There was no consistent correlation between ARA and tree size (dbh). The amount of N₂ fixed in this alder stand was estimated at 56.4 kg ha^?1 year^?1 when a theoretical molar ratio of 3 was used to convert the amount of reduced acetylene to the amount of fixed N₂. This amount of N₂ fixation corresponded to the 66.4% of N in the leaf litter produced in a year. These results suggested that N₂ fixation still contributed to the large portion of N economy in this alder stand.