The regulation of symbiotic N2 fixation: a conceptual model of N feedback from the ecosystem to the gene expression level

Symbiotic nitrogen (N₂) fixation in legumes may give the host plant a distinct competitive advantage; at the same time it is mainly responsible for introducing N into terrestrial ecosystems which may ultimately benefit all organisms. Depending on environmental conditions, symbiotic N₂ fixation may be tuned to the plant's N demand or specifically inhibited (a disadvantage for plants which depend mainly on symbiotic N₂ fixation), or even prevented. Thus, the ecological range for symbiotic N₂ fixation can be narrower than that of the host plants. A shortage of mineral N is the only case in which adverse environmental conditions clearly favour symbiotic N₂ fixation. Variations in number or mass of nodules or nodule morphology are persistent features, that may represent one kind of regulation of N₂ fixation. In addition, varying O₂ permeability of nodules functions as a rapid and reversible control of N₂ fixation which may compensate partially or fully for poor nodulation. The plant's demand for symbiotically fixed N is thought to play a central role in modulating both nodulation and N₂ fixation activity; an N feedback mechanism is assumed. The control of symbiotic N₂ fixation operates through a series of ecophysiological triggers which are also influenced by complex interactions between legume plants and other organisms in the ecosystem. The proportion of legume biomass and the performance of symbiotic N₂ fixation in each individual legume are the main parameters which determine the amount of symbiotically fixed N introduced into a terrestrial ecosystem. The various triggers and N feedback mechanisms from the whole ecosystem to the gene expression level which regulate symbiotic N₂ fixation in terrestrial ecosystems are reviewed and discussed in terms of a conceptual model. Although the presented model is based primarily on our knowledge about the physiology of a few leguminous crop species and of ecosystem processes in managed, perennial grassland in temperate climatic conditions, it may stimulate thinking about functional relationships between symbiotic N₂ fixation and terrestrial ecosystems at various system levels.     

异养微生物固定CO2的合成生物学研究进展

人类活动造成大气二氧化碳（CO₂）浓度不断升高,使当今世界面临着气候变化的重大危机。微生物CO₂固定为实现地球“碳中和”提供了一条有前景的绿色发展路线。与自养微生物相比,异养微生物具有更快的生长速度和更先进的遗传工具,但是其固定CO₂的能力还很有限。近年来,基于合成生物学技术强化异养微生物CO₂固定受到诸多关注,主要包括优化能量供给、改造羧化途径以及基于异养微生物间接固定CO₂。本综述将围绕上述3个方面重点讨论异养微生物CO₂固定的研究进展,为将来更好地利用微生物CO₂固定技术实现“碳达峰、碳中和”提供参考。     

Dark microbial CO2 fixation in temperate forest soils increases with CO2 concentration

Dark, that is, nonphototrophic, microbial CO₂ fixation occurs in a large range of soils. However, it is still not known whether dark microbial CO₂ fixation substantially contributes to the C balance of soils and what factors control this process. Therefore, the objective of this study was to quantitate dark microbial CO₂ fixation in temperate forest soils, to determine the relationship between the soil CO₂ concentration and dark microbial CO₂ fixation, and to estimate the relative contribution of different microbial groups to dark CO₂ fixation. For this purpose, we conducted a ¹³C‐CO₂ labeling experiment. We found that the rates of dark microbial CO₂ fixation were positively correlated with the CO₂ concentration in all soils. Dark microbial CO₂ fixation amounted to up to 320 µg C kg^?1 soil day^?1 in the Ah horizon. The fixation rates were 2.8–8.9 times higher in the Ah horizon than in the Bw1 horizon. Although the rates of dark microbial fixation were small compared to the respiration rate (1.2%–3.9% of the respiration rate), our findings suggest that organic matter formed by microorganisms from CO₂ contributes to the soil organic matter pool, especially given that microbial detritus is more stable in soil than plant detritus. Phospholipid fatty acid analyses indicated that CO₂ was mostly fixed by gram‐positive bacteria, and not by fungi. In conclusion, our study shows that the dark microbial CO₂ fixation rate in temperate forest soils increases in periods of high CO₂ concentrations, that dark microbial CO₂ fixation is mostly accomplished by gram‐positive bacteria, and that dark microbial CO₂ fixation contributes to the formation of soil organic matter.     

Seasonal variation in nitrogen fixation and effects of climate change in a subarctic heath

Background and aims

Nitrogen fixation associated with cryptogams is potentially very important in arctic and subarctic terrestrial ecosystems, as it is a source of new nitrogen (N) into these highly N limited systems. Moss-, lichen- and legume-associated N₂ fixation was studied with high frequency (every second week) during spring, summer, autumn and early winter to uncover the seasonal variation in input of atmospheric N₂ to a subarctic heath with an altered climate.

Methods

We estimated N₂ fixation from ethylene production by acetylene reduction assay in situ in a field experiment with the treatments: long- vs. short-term summer warming using plastic tents and litter addition (simulating expansion of the birch forest).

Results

N₂ fixation activity was measured from late April to mid November and 33 % of all N₂ was fixed outside the vascular plant growing season (Jun–Aug). This substantial amount underlines the importance of N₂ fixation in the cold period. Warming increased N₂ fixation two- to fivefold during late spring. However, long-term summer warming tended to decrease N₂ fixation outside the treatment (tents present) period. Litter alone did not alter N₂ fixation but in combination with warming N₂ fixation increased, probably because N₂ fixation became phosphorus limited under higher temperatures, which was alleviated by the P supply from the litter.

Conclusion

In subarctic heath, the current N₂ fixation period extends far beyond the vascular plant growing season. Climate warming and indirect effects such as vegetation changes affect the process of N₂ fixation in different directions and thereby complicate predictions of future N cycling.     

Nitrogen fixation and assimilation efficiency in Ethiopian and German pea varieties

Dinitrogen (N₂) fixation and assimilation efficiency in a German and two Ethiopian varieties of Pisum sativum L. was studied in a pot experiment during vegetative and reproductive growth. The objective of the study was to assess whether genotypes having contrasting growth habits showed differences in physiological processes that affect the efficiency of N₂ fixation and assimilation. Dry matter formation, nodulation and nitrogen assimilation were compared between two treatments where one depended solely on N₂ fixation while the other was nourished with nitrate. Moreover, carbon (C) costs of N₂ fixation and the capacity of different respiratory chains in roots and nodules were determined at vegetative and reproductive growth. As compared to the Ethiopian cultivars, the German variety displayed a more rapid vegetative growth with intensive N₂ fixation and assimilation and highly efficient individual nodules. However, during reproductive growth, N₂ fixation in the German variety declined sharply, while continuing in the Ethiopian varieties. Lowest C costs of N₂ fixation coincided with most efficient individual nodules in both growth intervals. C costs of N₂ fixation were lower during reproductive growth in all varieties which was accompanied by a shift in root/nodule respiratory capacity towards more C efficient respiratory pathways. The results provide further evidence that unreliable nitrogen fixation rates during reproductive growth of pea can be connected with restricted C supply to nodules. One strategy of pea plants to adapt to critical C availability is an increase in capacity of more C efficient root/nodule respiration.     

The role of pH in the regulation of carbon fixation in the chloroplast stroma. Studies on CO2 fixation in the light and dark

1. The pH in the stroma and in the thylakoid space has been measured in a number of chloroplast preparations in the dark and in the light at 20 °C. Illumination causes a decrease of the pH in the thylakoid space by 1.5 and an increase of the pH in the stroma by almost 1 pH unit.2. CO₂ fixation is shown to be strongly dependent on the pH in the stroma. The pH optimum was 8.1, with almost zero activity below pH 7.3. Phosphoglycerate reduction, which is a partial reaction of CO₂ fixation, shows very little pH dependency.3. Low concentrations of the uncoupler m-chlorocarbonylcyanide phenylhydrazone (CCCP) inhibit CO₂ fixation without affecting phosophoglycerate reduction. This inhibition of CO₂ fixation appears to be caused by reversal of light induced alkalisation in the stroma by CCCP.4. Methylamine has a very different effect compared to CCCP. Increasing concentrations of methylamine inhibit CO₂ fixation and phosphoglycerate reduction to the same extent. The light induced alkalisation of the stroma appears not to be significantly inhibited by methylamine, but the protons in the thylakoid space are neutralized. The inhibition of CO₂ fixation by higher concentrations of methylamine is explained by an inhibition of photophosphorylation. It appears that methylamine does not abolish proton transport.5. It is shown that intact chloroplasts are able to fix CO₂ in the dark, yielding 3-phosphoglycerate. This requires the addition of dihydroxyacetone phosphate as precursor of ribulosemonophosphate and also to supply ATP, and the addition of oxaloacetate for reoxidation of the NADPH in the stroma.6. Dark CO₂ fixation in the presence of dihydroxyacetone phosphate and oxaloacetate has the same pH dependency as CO₂ fixation in the light. This demonstrates that CO₂ fixation in the dark is not possible, unless the pH in the medium is artificially raised to pH 8.8.7. It is shown that pH changes occurring in the stroma after illumination are sufficient to switch CO₂ fixation from zero to maximal activity. This offers a mechanism for light control of CO₂ fixation, avoiding wasteful CO₂ fixation in the dark.     

Contemporaneous N2 Fixation and Oxygenic Photosynthesis in the Nonheterocystous Mat-Forming Cyanobacterium Lyngbya aestuarii

The nonheterocystous filamentous cyanobacterial genus Lyngbya is a widespread and frequently dominant component of marine microbial mats. It is suspected of contributing to relatively high rates of N₂ fixation associated with mats. The ability to contemporaneously conduct O₂-sensitive N₂ fixation and oxygenic photosynthesis was investigated in Lyngbya aestuarii isolates from a North Carolina intertidal mat. Short-term (<4-h) additions of the photosystem II (O₂ evolution) inhibitor 3(3,4-dichlorophenyl)-1,1-dimethylurea stimulated light-mediated N₂ fixation (nitrogenase activity), indicating potential inhibition of N₂ fixation by O₂ production. However, some degree of light-mediated N₂ fixation in the absence of 3(3,4-dichlorophenyl)-1,1-dimethylurea was observed. Electron microscopic immunocytochemical localization of nitrogenase, coupled to microautoradiographic studies of ¹⁴CO₂ fixation and cellular deposition of the tetrazolium salt 2,4,5-triphenyltetrazolium chloride, revealed that (i) nitrogenase was widely distributed throughout individual filaments during illuminated and dark periods, (ii) ¹⁴CO₂ fixation was most active in intercalary regions, and (iii) daylight 2,4,5-triphenyltetrazolium chloride reduction (formazan deposition) was most intense in terminal regions. Results suggest lateral partitioning of photosynthesis and N₂ fixation during illumination, with N₂ fixation being confined to terminal regions. During darkness, a larger share of the filament appears capable of N₂ fixation.     

Evaluation of the Relative Ureide Content of Xylem Sap as an Indicator of N(2) Fixation in Soybeans: GREENHOUSE STUDIES

The use of the relative ureide content of xylem sap [(ureide-N/total N) × 100] as an indicator of N₂ fixation in soybeans (Merr.) was examined under greenhouse conditions. Acetylene treatments to inhibit N₂ fixation were imposed upon the root systems of plants totally dependent upon N₂ fixation as their source of N and of plants dependent upon both N₂ fixation and uptake of exogenous nitrate. Significant decreases in the total N concentration of xylem sap from plants of the former type were observed, but no significant decrease was observed in the total N concentration of sap from the latter type of plants. In both types of plants, acetylene treatment caused significant decreases in the relative ureide content of xylem sap. The results provided further support for a link between the presence of ureides in the xylem and the occurrence of N₂ fixation in soybeans. The relative ureide content of xylem sap from plants totally dependent upon N₂ fixation was shown to be insensitive to changes in the exudation rate and total N concentration of xylem sap brought about by diurnal changes in environmental factors. There was little evidence of soybean cultivars or nodulating strains affecting the relative ureide content of xylem sap. `Ransom' soybeans nodulated with Rhizobium japonicum strain USDA 110 were grown under conditions to obtain plants exhibiting a wide range of dependency upon N₂ fixation. The relative ureide content of xylem sap was shown to indicate reliably the N₂ fixation of these plants during vegetative growth using a ¹⁵N method to measure N₂ fixation activity. The use of the relative ureide content of xylem sap for quantification of N₂ fixation in soybeans should be evaluated further.     

Nitrogen fixation rates in algal turf communities of a degraded versus less degraded coral reef

Algal turf communities are ubiquitous on coral reefs in the Caribbean and are often dominated by N₂-fixing cyanobacteria. However, it is largely unknown (1) how much N₂ is actually fixed by turf communities and (2) which factors affect their N₂ fixation rates. Therefore, we compared N₂ fixation activity by turf communities at different depths and during day and night-time on a degraded versus a less degraded coral reef site on the island of Curaçao. N₂ fixation rates measured with the acetylene reduction assay were slightly higher in shallow (5–10-m depth) than in deep turf communities (30-m depth), and N₂ fixation rates during the daytime significantly exceeded those during the night. N₂ fixation rates by the turf communities did not differ between the degraded and less degraded reef. Both our study and a literature survey of earlier studies indicated that turf communities tend to have lower N₂ fixation rates than cyanobacterial mats. However, at least in our study area, turf communities were more abundant than cyanobacterial mats. Our results therefore suggest that turf communities play an important role in the nitrogen cycle of coral reefs. N₂ fixation by turfs may contribute to an undesirable positive feedback that promotes the proliferation of algal turf communities while accelerating coral reef degradation.     

STUDIES ON THE METABOLISM OF AUTOTROPHIC BACTERIA : II. THE NATURE OF THE CHEMOSYNTHETIC REACTION

In a study of chemosynthesis (the fixation of CO₂ by autotrophic bacteria in the dark) in Thiobacillus thiooxidans, the data obtained support the following conclusions: 1. CO₂ can be fixed by "resting cells" of Thiobacillus thiooxidans; the fixation is not "growth bound." 2. The physiological condition of the cell is of considerable importance in determining CO₂ fixation. 3. CO₂ fixation can occur in the absence of oxidizable sulfur in "young" cells. The extent of this fixation appears to be dependent upon the pCO₂. 4. CO₂ fixation can also occur under anaerobic conditions and the presence of sulfur does not influence such fixation. 5. However, in the CO₂ fixation by cells in the absence of sulfur, only a limited amount of CO₂ can be fixed. This amount is approximately 40 µl. CO₂ per 100 micrograms bacterial nitrogen. After a culture has utilized this amount of CO₂ it no longer has the ability to fix CO₂ but releases it during its respiration. 6. Relatively short periods of sulfur oxidation can restore the ability of cells to fix CO₂ under conditions where sulfur oxidation is prevented. 7. It is possible to oxidize sulfur in the absence of CO₂ and to store the energy thus formed within the cell. It is then possible to use this energy at a later time for the fixation of CO₂ in the entire absence of sulfur oxidation. 8. Cultures of Thiobacillus thiooxidans respiring on sulfur utilize CO₂ in a reaction which proceeds to a zero concentration of CO₂ in the atmosphere. 9. CO₂ may act as an oxidizing agent for sulfur. 10. Hydrogen is not utilized by the organism. 11. It is possible to selectively inhibit sulfur oxidation and CO₂ fixation.     

Non-phototrophic CO 2 fixation by soil microorganisms

Although soils are generally known to be a net source of CO₂ due to microbial respiration, CO₂ fixation may also be an important process. The non-phototrophic fixation of CO₂ was investigated in a tracer experiment with ¹⁴CO₂ in order to obtain information about the extent and the mechanisms of this process. Soils were incubated for up to 91 days in the dark. In three independent incubation experiments, a significant transfer of radioactivity from ¹⁴CO₂ to soil organic matter was observed. The process was related to microbial activity and could be enhanced by the addition of readily available substrates such as acetate. CO₂ fixation exhibited biphasic kinetics and was linearly related to respiration during the first phase of incubation (about 20–40 days). The fixation amounted to 3–5% of the net respiration. After this phase, the CO₂ fixation decreased to 1–2% of the respiration. The amount of carbon fixed by an agricultural soil corresponded to 0.05% of the organic carbon present in the soil at the beginning of the experiment, and virtually all of the fixed CO₂ was converted to organic compounds. Many autotrophic and heterotrophic biochemical processes result in the fixation of CO₂. However, the enhancement of the fixation by addition of readily available substrates and the linear correlation with respiration suggested that the process is mainly driven by aerobic heterotrophic microorganisms. We conclude that heterotrophic CO₂ fixation represents a significant factor of microbial activity in soils.     

Water availability moderates N2 fixation benefit from elevated [CO2]: A 2‐year free‐air CO2 enrichment study on lentil (Lens culinaris MEDIK.) in a water limited agroecosystem

Increased biomass and yield of plants grown under elevated [CO₂] often corresponds to decreased grain N concentration ([N]), diminishing nutritional quality of crops. Legumes through their symbiotic N₂ fixation may be better able to maintain biomass [N] and grain [N] under elevated [CO₂], provided N₂ fixation is stimulated by elevated [CO₂] in line with growth and yield. In Mediterranean‐type agroecosystems, N₂ fixation may be impaired by drought, and it is unclear whether elevated [CO₂] stimulation of N₂ fixation can overcome this impact in dry years. To address this question, we grew lentil under two [CO₂] (ambient ~400 ppm and elevated ~550 ppm) levels in a free‐air CO₂ enrichment facility over two growing seasons sharply contrasting in rainfall. Elevated [CO₂] stimulated N₂ fixation through greater nodule number (+27%), mass (+18%), and specific fixation activity (+17%), and this stimulation was greater in the high than in the low rainfall/dry season. Elevated [CO₂] depressed grain [N] (?4%) in the dry season. In contrast, grain [N] increased (+3%) in the high rainfall season under elevated [CO₂], as a consequence of greater post‐flowering N₂ fixation. Our results suggest that the benefit for N₂ fixation from elevated [CO₂] is high as long as there is enough soil water to continue N₂ fixation during grain filling.     

C(4) Photosynthesis: Light-dependent CO(2) Fixation by Mesophyll Cells, Protoplasts, and Protoplast Extracts of Digitaria sanguinalis

Mesophyll cells, protoplasts, and protoplast extracts of Digitaria sanguinalis were used for comparative studies of light-dependent CO₂ fixation. CO₂ fixation was low without the addition of organic substrates. Pyruvate, oxaloacetate, and 3-phosphoglycerate induced relatively low rates (10 to 90 μmoles/mg chlorophyll·hr) of CO₂ fixation when added separately. However, a highly synergistic relationship was found between pyruvate + oxaloacetate and pyruvate + 3-phosphoglycerate for inducing light-dependent CO₂ fixation in the mesophyll preparations. Highest rates of CO₂ fixation were obtained with protoplast extracts. Pyruvate, in combination with oxaloacetate or 3-phosphoglycerate induced light-dependent rates from 150 to 380 μmoles of CO₂ fixed/mg chlorophyll·hr which are equivalent to or exceed reported rates of whole leaf photosynthesis in C₄ species. Concentrations of various substrates required to give half-maximum velocities of CO₂ fixation were determined, with the protoplast extracts generally saturating at the lowest substrate concentrations. Chloroplasts separated from protoplast extracts showed little capacity for CO₂ fixation. The results suggest that CO₂ fixation in C₄ mesophyll cells is dependent on chloroplasts and extrachloroplastic phosphoenolpyruvate carboxylase.     

Dynamics of periphyton nitrogen fixation in short-hydroperiod wetlands revealed by high-resolution seasonal sampling

Periphyton N₂ 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 N₂ 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 N₂ 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. N₂ 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 N₂ 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.     

Levels of Daily Light Doses Under Changed Day-Night Cycles Regulate Temporal Segregation of Photosynthesis and N2 Fixation in the Cyanobacterium Trichodesmium erythraeum IMS101

While the diazotrophic cyanobacterium Trichodesmium is known to display inverse diurnal performances of photosynthesis and N₂ fixation, such a phenomenon has not been well documented under different day-night (L-D) cycles and different levels of light dose exposed to the cells. Here, we show differences in growth, N₂ fixation and photosynthetic carbon fixation as well as photochemical performances of Trichodesmium IMS101 grown under 12L:12D, 8L:16D and 16L:8D L-D cycles at 70 μmol photons m^-2 s^-1 PAR (LL) and 350 μmol photons m^-2 s^-1 PAR (HL). The specific growth rate was the highest under LL and the lowest under HL under 16L:8D, and it increased under LL and decreased under HL with increased levels of daytime light doses exposed under the different light regimes, respectively. N₂ fixation and photosynthetic carbon fixation were affected differentially by changes in the day-night regimes, with the former increasing directly under LL with increased daytime light doses and decreased under HL over growth-saturating light levels. Temporal segregation of N₂ fixation from photosynthetic carbon fixation was evidenced under all day-night regimes, showing a time lag between the peak in N₂ fixation and dip in carbon fixation. Elongation of light period led to higher N₂ fixation rate under LL than under HL, while shortening the light exposure to 8 h delayed the N₂ fixation peaking time (at the end of light period) and extended it to night period. Photosynthetic carbon fixation rates and transfer of light photons were always higher under HL than LL, regardless of the day-night cycles. Conclusively, diel performance of N₂ fixation possesses functional plasticity, which was regulated by levels of light energy supplies either via changing light levels or length of light exposure.     

Interaction of nitrate uptake and nitrogen fixation in faba beans

An experiment was carried out to determine the relationship between nitrate uptake and nitrogen fixation of faba beans. Therefore inoculated and uninoculated faba beans were grown in nutrient solution with different nitrate concentrations. Nitrate uptake was measured every two days during the growing period. At the end of the experiment the nitrate uptake kinetics were determined with a short time depletion technique and nitrogen fixation was measured with the acetylene reduction method. A limitation of nitrate uptake due to nitrogen fixation was relatively small. Nitrate concentrations of approximately 1 mol m^–3 and 5 mol m^–3 decreased nitrogen fixation to values of 16% and 1% of the control plants which received no nitrate nitrogen. A reduction of nitrogen fixation was mainly due to a decrease of specific nitrogen fixation per unit nodule weight and to a lesser extent due to a reduction of nodule growth. Only the maximum nitrate influx (I_max) seemed to be influenced by nitrogen fixation. Michaelis-Menten constants (K_m) and minimum NO _inf3 ^– -concentrations (C_min) were not significantly influenced by nitrogen fixation.     

AMMONIUM STIMULATION OF DARK CARBON FIXATION AS AN INDICATOR OF NITROGEN DEFICIENCY IN PHYTOPLANKTON: POTENTIAL ERRORS CAUSED BY AMMONIUM-OXIDIZING BACTERIA1

Stimulation of dark fixation of carbon by NH₄⁺ is often used as an indicator of phytoplankton N deficiency. This assay is based on the influence of available NH₄⁺ on anaplerotic CO₂ fixation by algae. However, carbon fixation by chemoautotrophic NH₄⁺-oxidizing bacteria may also be stimulated by NH₄⁺ enrichment, a process that can mask the algal response in natural communities. NH₄⁺ addition enhanced dark carbon fixation up to 300%, relative to unamended controls, in organisms collected on a 0.7-μm retention filter in oligotrophic Flathead Lake, Montana, but the effect was not detectable in the presence of nitrapyrin, an inhibitor of NH₄⁺-oxidizing bacteria. Dark carbon fixation was enhanced with addition of NH₄⁺ in organisms retained on 2-μm filters (which should allow passage of most bacteria). NH₄⁺ stimulated dark carbon fixation in N-deficient axenic cultures of Chlamydomonas reinhardtii Dang but not in N-replete cultures in both the presence and absence of nitrapyrin. Application of nitrapyrin or size fractionation treatments, to separate the processes of dark carbon fixation by nitrifiers and phytoplankton, may improve the efficacy of assays using NH₄⁺ stimulation of dark carbon fixation to specifically indicate N deficiency in natural algal communities.     

Divergent selection for dinitrogen fixation and yield in soybean

Summary Nitrogen fixation is generally considered to be a major parameter of productivity in soybean (Glycine max). The aim of the investigations reported here was to analyse the genetic behaviour of this trait in view of its possible use as an indirect criterion of selection for productivity. Divergent selection for nitrogen fixation rate was carried out on F₂ populations obtained from crosses between high-yielding cultivars that are well adapted to French climatic conditions. The genetic component of nitrogen fixation and yield was isolated through the analysis of (1) the nitrogen fixation potentials of the genotypes under controlled conditions and (2) the field yields under favourable conditions. Divergent selection resulted in two groups of genotypes whose nitrogen fixation abilities are significantly different. The F₆ filial progeny obtained by single seed descent from the two groups displayed significantly different abilities for nitrogen fixation and for field productivity. The gain achieved for the nitrogen fixation activity with respect to the mean value of the parents ranged from 20% to 33% for the positive selection, depending on the crosses. The occurrence of positive and significant correlations between the level of nitrogen fixation activity in F₂ plants and N₂ fixation or yield in the F₆ generation corroborates the relatively high heritability of this trait and suggests its possible use as an indirect selection criterion for yield.     

Isotopic analysis of cyanobacterial nitrogen fixation associated with subarctic lichen and bryophyte species

Dinitrogen fixation by cyanobacteria is of particular importance for the nutrient economy of cold biomes, constituting the main pathway for new N supplies to tundra ecosystems. It is prevalent in cyanobacterial colonies on bryophytes and in obligate associations within cyanolichens. Recent studies, applying interspecific variation in plant functional traits to upscale species effects on ecosystems, have all but neglected cryptogams and their association with cyanobacteria. Here we looked for species-specific patterns that determine cryptogam-mediated rates of N₂ fixation in the Subarctic. We hypothesised a contrast in N₂ fixation rates (1) between the structurally and physiologically different lichens and bryophytes, and (2) within bryophytes based on their respective plant functional types. Throughout the survey we supplied ¹⁵N-labelled N₂ gas to quantify fixation rates for monospecific moss, liverwort and lichen turfs. We sampled fifteen species in a design that captures spatial and temporal variations during the growing season in Abisko region, Sweden. We measured N₂ fixation potential of each turf in a common environment and in its field sampling site, in order to embrace both comparativeness and realism. Cyanolichens and bryophytes differed significantly in their cyanobacterial N₂ fixation capacity, which was not driven by microhabitat characteristics, but rather by morphology and physiology. Cyanolichens were much more prominent fixers than bryophytes per unit dry weight, but not per unit area due to their low specific thallus weight. Mosses did not exhibit consistent differences in N₂ fixation rates across species and functional types. Liverworts did not fix detectable amounts of N₂. Despite the very high rates of N₂ fixation associated with cyanolichens, large cover of mosses per unit area at the landscape scale compensates for their lower fixation rates, thereby probably making them the primary regional atmospheric nitrogen sink.     

The response and recovery of nitrogen fixation activity in soybean to water deficit at different reproductive developmental stages

Soybean (Glycine max [L.] Merr.) N₂ fixation is a primary plant mechanism responsible for meeting plant-N demand during seed development. Nitrogen fixation is recognized as a drought-sensitive mechanism; however, N₂ fixation response to water deficit and N₂ fixation recovery at different reproductive stages are not well documented. We tested the hypothesis that water deficit during late reproductive stages would inhibit N₂ fixation and lead to the breakdown of essential leaf proteins and an inability to recover N₂ fixation. Acetylene reduction activity (ARA) and N redistribution response to a 5-d drought period at flowering (R2), early seed fill (R5), and late seed fill (R6) were evaluated in one genotype (Hendricks, maturity group 0). Control plants maintained high rates of nodule activity until late seed fill. Plants drought stressed at R2 and R5 recovered ARA after rewatering and in some cases had higher nitrogenase activity than control plants during mid-seed fill. Recovery of ARA on plants stressed at R2 and R5 was associated with higher shoot N concentration than control plants at maturity. Drought stress at R6 reduced ARA, and the inability to recover ARA after stress alleviation at R6 resulted in decreased individual seed mass, which was likely caused by an acceleration of leaf N redistribution and a shorter seed-fill period. Results emphasized the importance of soybean N₂ fixation during late seed development on seed yield and that the ability to recover N₂ fixation following drought is dependent upon crop developmental stage.