A numerical model of nitrogen flxation and its application to Lake Valencia,Venezuela*

SUMMARY. 1. A numerical model for calculation of daily and annual nitrogen fixation in lakes is presented. The model is based on empirically-derived equations for the rates of nitrogen fixation by heterocysts (nitrogen-fixing cells) in relation to light and on functions for the vertical and tetnporal distributions of heterocysts and light in a lake. 2. Applications of the model to Lake Valencia, Venezuela, between December 1980 and December 1981 indicated that nitrogen fixation is largely a surface phenomenon in this lake: 80% of diurnal fixation occurred within 1m of the water surface. 3. Nitrogen fixation is largely restricted to periods of lake stratification, when the phytoplankton have sufficient light for growth, but dissolved inorganic nitrogen is scarce. Nitrogen fixation was maximal late in the stratification period of 1981: 85 % of fixation occurred within the last 3 months of the 9-month period. 4. The annual nitrogen fixation in Lake Valencia is 26 kg ha^?1, which is comparable to the nitrogen fixation in temperate eutrophic lakes with seasonal blue-green algal blooms. However, nitrogen fixation accounted for only 23% of the total nitrogen supply to Lake Valencia in 1981.     

Physiological studies on nitrogen fixation in the blue-green alga anabaena cylindrica

Summary Short-term manometric experiments with bacteria-free cultures of Anabaena cylindrica showed that the close dependency of nitrogen fixation upon photosynthesis could be temporarily eliminated in nitrogen-starved cells. Initial rates of nitrogen uptake by these cells in the absence of carbon dioxide were equally rapid in the light and dark, decreasing and finally ceasing after two hours. Continued steady nitrogen uptake was only maintained for long periods in the presence of carbon dioxide in the light. In the dark, nitrogen uptake was accompanied by carbon dioxide evolution.More oxygen was evolved in the light by cells fixing nitrogen than by those incubated under argon. This additional oxygen evolution could be accounted for by extra carbon dioxide fixation in the presence of nitrogen.Of a number of organic compounds tested, only sodium pyruvate stimulated nitrogen fixation. This stimulation was achieved both in the light and dark and in the presence and absence of carbon dioxide, showing that the role of pyruvate was other than acting as a carbon skeleton.Three metabolic inhibitors, cyanide and chlorpromazine (chiefly respiratory) and phenylurethane (photosynthetic) differentially inhibited photosynthesis and nitrogen fixation. The latter inhibitor had a more marked effect on photosynthesis while the two chiefly respiratory inhibitors had a stronger effect on nitrogen fixation.     

Resolution of Conflicting Signals at the Single-Cell Level in the Regulation of Cyanobacterial Photosynthesis and Nitrogen Fixation

Unicellular, diazotrophic cyanobacteria temporally separate dinitrogen (N₂) fixation and photosynthesis to prevent inactivation of the nitrogenase by oxygen. This temporal segregation is regulated by a circadian clock with oscillating activities of N₂ fixation in the dark and photosynthesis in the light. On the population level, this separation is not always complete, since the two processes can overlap during transitions from dark to light. How do single cells avoid inactivation of nitrogenase during these periods? One possibility is that phenotypic heterogeneity in populations leads to segregation of the two processes. Here, we measured N₂ fixation and photosynthesis of individual cells using nanometer-scale secondary ion mass spectrometry (nanoSIMS) to assess both processes in a culture of the unicellular, diazotrophic cyanobacterium Crocosphaera watsonii during a dark-light and a continuous light phase. We compared single-cell rates with bulk rates and gene expression profiles. During the regular dark and light phases, C. watsonii exhibited the temporal segregation of N₂ fixation and photosynthesis commonly observed. However, N₂ fixation and photosynthesis were concurrently measurable at the population level during the subjective dark phase in which cells were kept in the light rather than returned to the expected dark phase. At the single-cell level, though, cells discriminated against either one of the two processes. Cells that showed high levels of photosynthesis had low nitrogen fixing activities, and vice versa. These results suggest that, under ambiguous environmental signals, single cells discriminate against either photosynthesis or nitrogen fixation, and thereby might reduce costs associated with running incompatible processes in the same cell.     

Light-dark (12:12) cycle of carbon and nitrogen metabolism in Crocosphaera watsonii WH8501: relation to the cell cycle

This study provides with original data sets on the physiology of the unicellular diazotrophic cyanobacterium Crocosphaera watsonii WH8501, maintained in continuous culture in conditions of obligate diazotrophy. Cultures were exposed to a 12:12 light-dark regime, representative of what they experience in nature and where growth is expected to be balanced. Nitrogen and carbon metabolism were monitored at high frequency and their dynamics was compared with the cell cycle. Results reveal a daily cycle in the physiological and biochemical parameters, tightly constrained by the timely decoupled processes of N(2) fixation and carbon acquisition. The cell division rate increased concomitantly to carbon accumulation and peaked 6 h into the light. The carbon content reached a maximum at the end of the light phase. N(2) fixation occurred mostly during the dark period and peaked between 9 and 10 h into the night, while DNA synthesis, reflected by DNA fluorescence, increased until the end of the night. Consequently, cells in G1- and S-phases present a marked decrease in their C:N ratio. Nitrogen acquisition through N(2) fixation exceeded 1.3- to 3-fold the nitrogen requirements for growth, suggesting that important amounts of nitrogen are excreted even under conditions supposed to favour balanced, carbon and nitrogen acquisitions.     

Traffic lights in trichodesmium. Regulation of photosynthesis for nitrogen fixation studied by chlorophyll fluorescence kinetic microscopy

We investigated interactions between photosynthesis and nitrogen fixation in the non-heterocystous marine cyanobacterium Trichodesmium IMS101 at the single-cell level by two-dimensional (imaging) microscopic measurements of chlorophyll fluorescence kinetics. Nitrogen fixation was closely associated with the appearance of cells with high basic fluorescence yield (F(0)), termed bright cells. In cultures aerated with normal air, both nitrogen fixation and bright cells appeared in the middle of the light phase. In cultures aerated with 5% oxygen, both processes occurred at a low level throughout most of the day. Under 50% oxygen, nitrogen fixation commenced at the beginning of the light phase but declined soon afterwards. Rapid reversible switches between fluorescence levels were observed, which indicated that the elevated F(0) of the bright cells originates from reversible uncoupling of the photosystem II (PSII) antenna from the PSII reaction center. Two physiologically distinct types of bright cells were observed. Type I had about double F(0) compared to the normal F(0) in the dark phase and a PSII activity, measured as variable fluorescence (F(v) = F(m) - F(0)), similar to normal non-diazotrophic cells. Correlation of type I cells with nitrogen fixation, oxygen concentration, and light suggests that this physiological state is connected to an up-regulation of the Mehler reaction, resulting in oxygen consumption despite functional PSII. Type II cells had more than three times the normal F(0) and hardly any PSII activity measurable by variable fluorescence. They did not occur under low-oxygen concentrations, but appeared under high-oxygen levels outside the diazotrophic period, suggesting that this state represents a reaction to oxidative stress not necessarily connected to nitrogen fixation. In addition to the two high-fluorescence states, cells were observed to reversibly enter a low-fluorescence state. This occurred mainly after a cell went through its bright phase and may represent a fluorescence-quenching recovery phase.     

Comparative adaptations of Aphanizomenon and Anabaena for nitrogen fixation under weak irradiance

1. In situ measurements of nitrogen fixation rates for Aphanizomenon in fertile Colorado lakes with low inorganic nitrogen concentrations demonstrated high efficiency of nitrogen fixation at low irradiance. 2. For study populations, rates of N₂ fixation in darkness and with alternating exposure to light and darkness were a higher percentage of light‐saturated rates for Aphanizomenon than for Anabaena, suggesting storage of reduced metabolites at high irradiance that are used subsequently by Aphanizomenon when cells are forced by mixing into zones of low irradiance. Also, saturation of N₂ fixation occurred over a lower range of irradiance for Aphanizomenon than for Anabaena. 3. High efficiency of N₂ fixation in Aphanizomenon at low or fluctuating irradiance is complementary to its previously demonstrated high efficiency of photosynthesis at low irradiance. Nitrogen fixation rate was also strongly related to DIN concentration; fixation was highest at low DIN (maximum < 5 μg L^?1) but was also most vulnerable to photoinhibition under such conditions. 4. The fixation capabilities of Aphanizomenon under weak or varying irradiance could explain its commonly observed domination over Anabaena when transparency is low and available nitrogen is scarce.     

Factors influencing dark nitrogen fixation in a blue-green alga.

Nitrogen-fixing activity declines first rapidly and then more gradually when Anabaenopsis circularis is transferred from light into dark conditions. The rate and duration of dark acetylene reduction (nitrogen fixation) depend upon conditions prevailing during the preceding light period. Factors (such as light intensity, CO2 concentration, and supply of glucose), which in the light affect photosynthesis and the accumulation of reserve carbon, have a profound effect on dark nitrogen fixation. Glucose greatly promotes nitrogen fixation in the light and supports prolonged nitrogenase activity in the dark. The results suggest that heterotrophic nitrogen fixation by blue-green algae in the field may be important both under light and dark conditions.     

Inhibition of cell division blocks the synthesis of the second nitrogenase (Nif2) in the cyanobacterium Anabaena variabilis

Anabaena variabilis ATCC 29413 belongs to the cyanobacteria that use a specific cell type, heterocysts, for fixation of atmospheric nitrogen under aerobic conditions. Nitrogen fixation under anaerobic conditions is catalyzed by a Mo-dependent nitrogenase (Nif2) that is expressed in the vegetative cells. We demonstrate here using immunolocalization/light microscopy (LM) that the synthesis of NifH2 is mainly initiated in dividing vegetative cells along the trichomes. Blocking cell division by cephalexin abolished nitrogenase synthesis under anaerobic conditions.     

In situ nitrogen fixation associated with seagrasses in the Gulf of Elat (Red Sea)

In situ nitrogen fixation associated with the seagrass Halophila stipulacea, at the northern Gulf of Elat (Red Sea), is eight to ten times higher than that of nearby plant-free areas. A daily cycle of nitrogen fixation is evident, with rates during the day being seven times greater than during the night. Removal of seagrass leaves only from a patch within a seagrass bed gradually decreases nitrogen fixation activity, reaching the rates of plant-free areas after ten hours. A method devised for the in situ measurement of nitrogen fixation rates using belljars is described in detail. Nitrogen fixation rates in situ are higher than in the laboratory and lack the lag period typical to laboratory measurements. In laboratory experiments using intact plant samples, glucose enhances nitrogen fixation rates both in light and dark. Photosystem II inhibitor (3-3,4-dichloro-phenyl-1,1-dimethylurea) doubles nitrogen fixation rates in light. Both field and laboratory results indicate that light is essential for nitrogen fixation activity in the H. stipulacea bed possibly through its effect on cyanobacterial population that occupy the aerobic niches of the phyllosphere and on photosynthetic Rhodospirillacean bacteria that inhabit the anaerobic ones. Nitrogen fixation rates evident in H. stipulacea beds in situ account for a considerable portion of the biomass production by the seagrasses. The dependence of high nitrogenase activity by the diazotrophs on the presence of the seagrasses indicates the great importance of the seagrass community to the nitrogen cycle in its highly oligotrophic surroundings of the Gulf of Elat.     

Effect of lectins on auxin-induced elongation and wall loosening in oat coleoptile and azuki bean epicotyl segments

A marine unicellular aerobic nitrogen-fixing cyanobacterium Synechococcus sp. strain Miarni BG 043511 was pretreated with different light and dark regimes in order to induce higher growth synchrony. A pretreatment of two dark and light cycles of 16 h each yielded good synchrony for 3 cell division cycles. Longer dark treatments decreased the degree of synchrony and shorter dark treatments caused irregular cell division. Once synchronous culture was established, distinct phases of cellular carbohydrate accumulation and cellular carbohydrate degradation were observed even under continuous illumination. Changes in carbohydrate content were repeated in a cyclic manner with approximately 20 h intervals, the same as the cell division cycle. This change in carbohydrate metabolism provided a good index of growth synchrony under nitrogen-fixing conditions.
Photosynthetic oxygen evolution and nitrogen fixation capabilities and their activities in near, in situ, culture conditions were measured in well synchronized cultures of this strain under continuous illumination. Distinct oscillations of both photosynthetic oxygen evolution and nitrogen fixation capabilities with ca 20-h intervals, similar to the interval of the cell division cycle, were observed for three cycles. However, the activities of photosynthetic oxygen evolution were inversely correlated with those of nitrogen fixation. During the nitrogen fixation period, net oxygen consumption was observed even in the light under conditions approximating in situ culture conditions. The phase of temporal appearance of nitrogenase activity during the cell division cycle coincided with the phase of carbohydrate net degradation. These data indicate that this unicellular cyanobacterium can grow diazotrophically under conditions of continuous illumination by the segregation of photosynthesis and nitrogen fixation within a cell division cycle.     

Theory of a Crop Enclosure System for Measuring Nitrogen Fixation, Photosynthesis, Respiration and Biological Processes in the Soil

This paper contains a theory for the analysis of gas exchangemeasurements obtained using a combined shoot and soil enclosure.Particular attention is given to the non-steady state ratesof nitrogen fixation and initial patterns of carbon dioxideevolution. Under steady state conditions virtually all of themathematical complexity disappears leaving equations very similarin form to those traditionally used by physiologists to estimaterates of gas exchange. A theory for measuring hydrogen evolutionfrom legume crops in the field is also presented. Nitrogen fixation, hydrogen, respiration, photosynthesis, soil     

Perspectives in biological nitrogen fixation research

Nitrogen fixation, along with photosynthesis is the basis of all life on earth. Current understanding suggests that no plant fixes its own nitrogen. Some plants (mainly legumes) fix nitrogen via symbiotic anaerobic microorganisms (mainly rhizobia). The nature of biological nitrogen fixation is that the dinitrogenase catalyzes the reaction-splitting triple-bond inert atmospheric nitrogen (N2) into organic ammonia molecule (NH3). All known nitrogenases are found to be prokaryotic,multi.complex and normally oxygen liable. Not surprisingly, the engineering of autonomous nitrogen-fixing plants would be a long-term effort because it requires the assembly of a complex enzyme and provision of anaerobic conditions. However,in the light of evolving protein catalysts, the anaerobic enzyme has almost certainly been replaced in many reactions by the more efficient and irreversible aerobic version that uses O2. On the other hand, nature has shown numerous examples of evolutionary convergence where an enzyme catalyzing a highly specific, O2-requiring reaction has an oxygen-independent counterpart, able to carry out the same reaction under anoxic conditions. In this review, I attempt to take the reader on a simplified journey from conventional nitrogenase complex to a possible simplified version of a yet to be discovered Ilght-utilizing nitrogenase.     

植物叶片氮分配及其影响因子研究进展

氮是植物生长的基本限制性因子,它的有效利用可以增加植物的适应性。叶片氮分配是指氮在植物叶片细胞各细胞结构以及游离化合物中所分配的比例。叶片氮的分配方式决定了叶片光合作用的强弱,影响叶片的坚韧程度以及化学防御强度,因此研究氮在植物叶片内的分配方式具有重要意义。阐述了叶片氮分配的方式,分析了影响叶片氮分配的生物和非生物因子(CO2,光,土壤养分),介绍了常用的叶片氮分配的研究方法,并对未来的研究进行了展望。     

Nitrogen use efficiency in instantaneous and daily photosynthesis of leaves in the canopy of a Solidago altissima stand

Photosynthetic capacity was measured on detached leaves sampled in a canopy of Solidago altissima L. Non-rectangular hyperbola fitted the light response curve of photosynthesis and significant correlations were observed between leaf nitrogen per unit area and four parameters which characterize the light-response curve. Using regressions of the parameters on leaf nitrogen, a model of leaf photosynthesis was constructed which gave the relationships between leaf nitrogen, photon flux density (PFD) and photosynthesis. Curvilinear relations were obtained between leaf nitrogen and photosynthetic rate on both an instantaneous and a daily basis. Nitrogen use efficiency (NUE, photosynthesis per unit leaf nitrogen) was calculated against leaf nitrogen under varying PFDs. The optimum nitrogen content per unit leaf area that maximizes NUE shifted to higher values with increasing PFD. Field measurements of PFD showed high positive correlations between the distribution of leaf nitrogen in the canopy and relative PFD. The predicted optimum leaf nitrogen content for each level in the canopy, to achieve maximized NUE during a clear day, was close to the actual nitrogen distribution as found through sampling.     

Tight coupling of root-associated nitrogen fixation and plant photosynthesis in the salt marsh grass Spartina alterniflora and carbon dioxide enhancement of nitrogenase activity.

The coupling of root-associated nitrogen fixation and plant photosynthesis was examined in the salt marsh grass Spartina alterniflora. In both field experiments and hydroponic assay chambers, nitrogen fixation associated with the roots was rapidly enhanced by stimulating plant photosynthesis. A kinetic analysis of acetylene reduction activity (ARA) showed that a five-to sixfold stimulation occurred within 10 to 60 min after the plant leaves were exposed to light or increased CO2 concentrations (with the light held constant). In field experiments, CO2 enrichment increased plant-associated ARA by 27%. Further evidence of the dependence of ARA on plant photosynthate was obtained when activity in excised roots was shown to decrease after young greenhouse plants were placed in the dark. Seasonal variation in the ARA of excised plant roots from field cores appears to be related to the annual cycle of net photosynthesis in S. alterniflora.     

Tight coupling of root-associated nitrogen fixation and plant photosynthesis in the salt marsh grass Spartina alterniflora and carbon dioxide enhancement of nitrogenase activity

The coupling of root-associated nitrogen fixation and plant photosynthesis was examined in the salt marsh grass Spartina alterniflora. In both field experiments and hydroponic assay chambers, nitrogen fixation associated with the roots was rapidly enhanced by stimulating plant photosynthesis. A kinetic analysis of acetylene reduction activity (ARA) showed that a five-to sixfold stimulation occurred within 10 to 60 min after the plant leaves were exposed to light or increased CO2 concentrations (with the light held constant). In field experiments, CO2 enrichment increased plant-associated ARA by 27%. Further evidence of the dependence of ARA on plant photosynthate was obtained when activity in excised roots was shown to decrease after young greenhouse plants were placed in the dark. Seasonal variation in the ARA of excised plant roots from field cores appears to be related to the annual cycle of net photosynthesis in S. alterniflora.     

Leaf canopy as a dynamic system: ecophysiology and optimality in leaf turnover

BACKGROUND AND AIMS: In a leaf canopy, there is a turnover of leaves; i.e. they are produced, senesce and fall. These processes determine the amount of leaf area in the canopy, which in turn determines canopy photosynthesis. The turnover rate of leaves is affected by environmental factors and is different among species. This mini-review discusses factors responsible for leaf dynamics in plant canopies, focusing on the role of nitrogen. SCOPE: Leaf production is supported by canopy photosynthesis that is determined by distribution of light and leaf nitrogen. Leaf nitrogen determines photosynthetic capacity. Nitrogen taken up from roots is allocated to new leaves. When leaves age or their light availability is lowered, part of the leaf nitrogen is resorbed. Resorbed nitrogen is re-utilized in new organs and the rest is lost with dead leaves. The sink-source balance is important in the regulation of leaf senescence. Several models have been proposed to predict response to environmental changes. A mathematical model that incorporated nitrogen use for photosynthesis explained well the variations in leaf lifespan within and between species. CONCLUSION: When leaf turnover is at a steady state, the ratio of biomass production to nitrogen uptake is equal to the ratio of litter fall to nitrogen loss, which is an inverse of the nitrogen concentration in dead leaves. Thus nitrogen concentration in dead leaves (nitrogen resorption proficiency) and nitrogen availability in the soil determine the rate of photosynthesis in the canopy. Dynamics of leaves are regulated so as to maximize carbon gain and resource-use efficiency of the plant.     

Seasonal variation in rates of heterotrophic nitrogen fixation (acetylene reduction) in Zostera noltii meadows and uncolonised sediments of the Bassin d'Arcachon, south-west France

Nitrogen fixation (acetylene reduction) rates were measured over an annual cycle in meadows of the seagrass Z. noltii and uncolonised sediments of the Bassin d'Arcachon, south-west France, using both slurry and whole core techniques. Measured rates using the slurry technique in Z. noltii colonised sediments were consistently higher than those determined in isolated cores. This was probably due to the release of labile organic carbon sources during preparation of the slurries. Thus, in colonised sediments the whole core technique may provide a more accurate estimate of in situ activity. Acetylene reduction rates measured by the whole core technique in colonised sediments were 1.8 to 4-fold greater, dependent upon the season, in the light compared with those measured in the dark, indicating that organic carbon released by the plant roots during photosynthesis was an important factor regulating nitrogen fixation. In contrast acetylene reduction rates in uncolonised sediments were independent of light.Addition of sodium molybdate, a specific inhibitor of sulphate reduction inhibited acetylene reduction activity in Z. noltii colonised sediments by > 80% as measured by both slurry and whole core techniques irrespective of the light regime, throughout the year inferring that sulphate reducing bacteria (SRB) were the dominant component of the nitrogen fixing microflora. A mutualistic relationship between Z. noltii and nitrogen fixing SRB in the rhizosphere, based on the exchange of organic carbon and fixed nitrogen is proposed. In uncolonised sediments sodium molybdate initially severely inhibited acetylene reduction rates, but the level of this inhibition declined over the course of the year. These data indicate that the nitrogen fixing SRB associated with the Zostera roots and rhizomes were progressively replaced by an aerobic population of nitrogen fixers associated with the decomposition of this recalcitrant high C:N ratio organic matter.Acetylene and sulphate reduction rates in the seagrass beds showed distinct summer maxima which correlated with a reduced availability of NH ₄ ⁺ in the sediment and the growth cycle of Z. noltii in the Bassin. Overall, these data indicate that acetylene reduction (nitrogen fixation) activity in the rhizosphere of Z. noltii was regulated both by release of organic carbon from the plant roots and maintenance of low ammonium concentrations in the root zone due to efficient ammonium assimilation.Nitrogen fixation rates determined from acetylene reduction rates measured by the whole core technique ranged from 0.1 to 7.3 mg N m^–2 d^–1 in the Z. noltii beds and between 0.02 and 3.7 mg N m^–2 d^–1 in uncolonised sediments, dependent upon the season. Nitrogen fixation in the rhizosphere of Z. noltii was calculated to contribute between 0.4 and 1.1 g N m^–2 y^–1 or between 6.3 and 12% of the annual fixed nitrogen requirement of the plants. Heterotrophic nitrogen fixation therefore represents a substantial local input of fixed nitrogen to the sediments of this shallow coastal lagoon and contributes to the overall productivity of Z. noltii in this ecosystem.     

Dirunal leaf movement, chlorophyll fluorescence and carbon assimilation in soybean grown under different nitrogen and water availabilities

Diurnal heliotropic leaf movements, photosynthetic gas exchange, and the ratio of variable fluorescence to maximum fluorescence (Fv/Fm) of unrestrained and horizontally restrained leaves from soybean (Glycine max cv. Cumberland) plants grown in two different water and two different nitrogen treatments were measured. Leaves of plants grown in low water or low nitrogen availability treatments displayed more pronounced diaheliotropism (solar tracking) in the afternoon and a longer period of paraheliotropism (light avoiding) at midday relative to those of well-watered, high-nitrogen-grown plants. Photosaturated photosynthetic rates and the photon flux required to saturate photosynthesis were reduced by water stress and nitrogen deficiency. Compared to horizontal leaves, irradiance on orienting leaves was nearer to the breakpoint of the photosynthetic light response curve, where photosynthesis is co-limited by ribulose biphosphate regeneration and carboxylation. This would increase the carbon return on investments of nitrogen into photosynthesis. A positive linear relationship between Fv/Fm and quantum yield of photosynthesis was measured. Leaves of low-nitrogen-grown plants had earlier and more prolonged reductions in Fv/Fm at midday compared to leaves of high nitrogen grown plants of the same water treatment. Within the same water and nitrogen treatment, horizontally restrained leaves had lower midday Fv/Fm in relation to orienting leaves. Nitrogen deficiency and water stress enhanced this difference such that horizontally restrained leaves of low water and low nitrogen grown plants had earlier and longer midday depressions in Fv/Fm.