Explaining mutualistic cooperation between species remains one of the greatest problems for evolutionary biology. Why do symbionts provide costly services to a host, indirectly benefiting competitors sharing the same individual host? Host monitoring of symbiont performance and the imposition of sanctions on 'cheats' could stabilize mutualism. Here we show that soybeans penalize rhizobia that fail to fix N(2) inside their root nodules. We prevented a normally mutualistic rhizobium strain from cooperating (fixing N(2)) by replacing air with an N(2)-free atmosphere (Ar:O(2)). A series of experiments at three spatial scales (whole plants, half root systems and individual nodules) demonstrated that forcing non-cooperation (analogous to cheating) decreased the reproductive success of rhizobia by about 50%. Non-invasive monitoring implicated decreased O(2) supply as a possible mechanism for sanctions against cheating rhizobia. More generally, such sanctions by one or both partners may be important in stabilizing a wide range of mutualistic symbioses.

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Host sanctions and the legume–rhizobium mutualism

Publisher Summary This chapter describes the various aspects of cereal-legume intercropping systems Intercropping is the growing of two or more crop species simultaneously in the same field during a growing season The intercropping of legumes with cereals offers scope for developing energy-efficient and sustainable agriculture The main types of intercropping include mixed intercropping, row intercropping, and strip intercropping Crop combinations differ with geographical location and there may be intercropping of tree crops, intercropping of tree and field crops, or intercropping of field crops Combinations of crops are determined primarily by the length of the growing season and the adaptation of crops to particular environments Different indices have been suggested for evaluating productivity and efficiency per unit area of land of cereal-legume intercrop systems These include comparisons of absolute yields, protein yields, caloric equivalent, and in economic terms, gross returns from intercrops and sole crops Differences in competitive ability affect the relative performance of component crops and thus the land equivalent ratio values of different cereal-legume intercrop systems It is found that phosphorus is a major nutrient that determines the production potential of most grain legumes usually intercropped with cereals

Cereal–Legume Intercropping Systems

INTRODUCfION Techniques for Studying the Identity of Transported Solutes of Nitrogen . Nitrogen in Phloem and Xylem Fluids . Uptake. Assimilation. and Transport of Nitrogen by Roots . Transport of Nitrogen from Nodulated Roots of Nr/ixing Plants .. Initial Distribution of Xylem-Borne Nitrogen in Shoots .. Export of Nitrogen from Leaves in Phloem . Export of Nitrogen from Stem Tissue in the Phloem . RelationshipS Between Transport and Storage of Nitrogen .. Transport of Nitrogen to Deve/oping Fruits and Seeds Loading of Nitrogenous Solutes onto Xylem and Phloem .. Relationships Between Nitrogen Transport and the Production and Partitioning of Photosynthate .. CONCLUSIONS ..

Transport and Partitioning of Nitrogenous Solutes

INTRODUCTION 539 THE EXCHANGE OF REDUCED CARBON AND NITROGEN: AN OVERVIEW. ... 540 MAJOR PRODUCTS OF RECENTLY FIXED N2 EXPORTED FROM NODULES.... 542 SYNTHESIS OF AMIDES AND UREIDES 546 Ammonium Assimilation .... 546 Asparagine Synthesis . 547 Synthesis of 4-Methyleneglutamine 548 Synthesis of Citrulline . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 549 Biogenesis of Allantoin and Allantoic Acid 551 METABOLISM OF AMIDES AND UREIDES . ......... . . ... . . ........ .... . . .. ..... . 558 ENERGETICS OF SYNTHESIS AND METABOLISM OF THE PRODUCTS OF Nz FIXATION 564 SUMMARy......... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 567

Products of Biological Nitrogen Fixation in Higher Plants: Synthesis, Transport, and Metabolism

Inputs of biologically fixed N into agricultural systems may be derived from symbiotic relationships involving legumes and Rhizobium spp., partnerships between plants and Frankia spp. or cyanobacteria, or from non-symbiotic associations between free-living diazotrophs and plant roots. It is assumed that these N2-fixing systems will satisfy a large portion of their own N requirements from atmospheric N2, and that additional fixed N will be contributed to soil reserves for the benefit of other crops or forage species. This paper reviews the actual levels of N2 fixation attained by legume and non-legume associations and assesses their role as a source of N in tropical and sub-tropical agriculture. We discuss factors influencing N2 fixation and identify possible strategies for improving the amount of N2 fixed.

Biological nitrogen fixation: investments, expectations and actual contributions to agriculture

The economy of C use by root nodules was examined in two symbioses, Vigna unguiculata (L.) Walp. (cv. Caloona):Rhizobium CB756 and Lupinus albus L. (cv. Ultra):Rhizobium WU425 over a 2-week period in early vegetative growth. Plants were grown in minus N water culture with cuvettes attached to the nodulated zone of their primary roots for collection of evolved CO2 and H2. Increments in total plant N and in C and N of nodules, and C:N weight ratios of xylem and phloem exudates were studied by periodic sampling from the plant populations. Itemized budgets were constructed for the partitioning and utilization of C in the two species. For each milligram N fixed and assimilated by the cowpea association, 1.54 ? 0.26 (standard error) milligrams C as CO2 and negligible H2 were evolved and 3.11 milligrams of translocated C utilized by the nodules. Comparable values for nodules of the lupin association were 3.64 ? 0.28 milligrams C as CO2, 0.22 ? 0.05 milligrams H2, and 6.58 milligrams C. More efficient use of C by cowpea nodules was due to a lesser requirement of C for synthesis of exported N compounds, a smaller allocation of C to nodule dry matter, and a lower evolution of CO2. The activity of phosphoenolpyruvate carboxylase in nodule extracts and the rate of 14C02 fixation by detached nodules were greater for the cowpea symbiosis (0.56 ? 0.06 and 0.22 milligrams C as CO2 fixed per gram fresh weight per hour, respectively) than for the lupin 0.06 ? 0.02 and 0.01 milligrams C as CO2 fixed per gram fresh weight per hour. The significance of the data was discussed in relation to current information on theoretical costs of nitrogenase functioning and associated nodule processes. Several investigations have considered the energy relationships of N fixation by nodulated legumes. Some of these have concerned the thermodynamics of the reactions (3), the energetics of nitrogenase functioning (5, 6), and the effects of H2 evolution by nitrogenase on the efficiency of N fixation in specific symbioses (7, 20-23). Other investigations have assessed nodule functioning in terms of carbohydrate intake from the parent plant (9, 12, 15), CO2 loss by the below-ground organs of nodulated plants (9, 11, 14) or by a detailed C economy of nodules (1). This paper describes an experimentally based approach to determine the cost of symbiosis involving simultaneous measurements of N2 fixation, and CO2 and H2 evolution by attached nodules of intact legumes. These data are used to construct itemized budgets for the utilization of photosynthetically fixed C in two contrasting legume:

Economy of Photosynthate Use in Nitrogen-fixing Legume

Mobilization of Nitrogen in Fruiting Plants of a Cultivar of Cowpea

Growth and symbiotic performance of soybean (Glycine max (L.) Merrill) cv. Bragg and three of its induced nodulation mutants (nod49, non-nodulating; ntsl 116, intermediate supernodulator; nts1007, extreme supernodulator) were compared throughout development under different nitrogen regimes (0, 2, 5 and 10 mol nitrate m−3). Nitrogen fixation was assessed using 15N-isotope dilution and xylem sap analysis for ureide content. Both techniques confirmed a complete lack of N2 fixation activity in nod49. Plant reliance on nitrogen fixation by the other genotypes was dependent on the nitrate regime and the developmental stage. The ntsl007 and ntsl 116 mutants fixed more nitrogen than the parent cultivar in the presence of 10 mol m−3 nitrate in the nutrient solution, but higher input of symbiotically derived nitrogen was still insufficient to offset the amount of nitrogen removed in the harvested seed. However, the mutants utilized less nitrate for growth than Bragg. Comparison of estimates of N2 fixation derived from the 15N-dilution technique with those based on relative ureide content of xylem sap indicated that the latter offered a simple and reliable procedure for evaluating the symbiotic performance of supernodulating plants.

Symbiotic Performance of Supernoclulating Soybean (Glycine max (L.) Merrill) Mutants during Development on Different Nitrogen Regimes

Budgets for transfer of carbon from individual leaves and other source organs to fruits and nodulated roots were constructed for stages of the post-flowering development of symbiotically-dependent cowpea ( Vigna unguiculata L. Walp. cv. Vita 3-Rhizobium strain CB756). Exportable surpluses of carbon from sources, assessed from net exchanges of C02 and changes in carbon content, were allocated to sink organs in proportion to carbon consumption (growth and respiration) and the ability of each sink organ to attract assimilates from the sources, as demonstrated by 14C-feeding. The first 10 d after flowering showed high sink activity by roots, stem and petioles, low consumption by fruits, with the upper three trifoliate blossom leaves providing the bulk of the required assimilates. The next 10 d showed a sharp decline in photosynthesis of the leaf subtending the oldest fruit followed by similar declines in leaves at the other fruiting nodes. All leaflets at fruiting nodes abscised during the final 10 d period, while the two lower leaves, not subtending fruits, remained green and supplied most of the carbon required by developing fruits and roots. Throughout fruiting all currently-active sources supplied all sinks, with only slight evidence of blossom leaves specializing in nourishing their subtended fruits. Of the carbon translocated from leaves during fruiting 32% came from the topmost leaf, 28% from the leaf below this, 16% from the next leaf, and the remaining 24% from the lowest three leaves. Some 80% of the fruit's total intake of carbon came from leaves, the rest from mobilization of stored carbon (partly sugars and starch) from other vegetative parts.

Post-Anthesis Economy of Carbon in a Cultivar of Cowpea

Hansen, A. P., Pate, J. S. and Atkins, C. A. 1987. Relationships between acetylene reduction activity, hydrogen evolution and nitrogen fixation in nodules of Acacia spp.: Experimental background to assaying fixation by acetylene reduction under field conditions.—J. exp. Bot. 38: 1-12. Glasshouse grown, symbiotically-dependent seedlings of Acacia alata R.Br., A. extensa Lindl., and A. pulchella R.Br, were examined for acetylene reduction in closed assay systems using undisturbed potted plants, excavated whole plants, nodulated roots or detached nodules. Nitrogenase activity declined sharply over the first hour after exposure of detached nodules to acetylene (10% v/v in air), less steeply or not at all over a 3 h period in assays involving attached nodules. Using detached nodules, rates of acetylene reduction, nitrogen (15N2) fixation, and hydrogen evolution in air (15N2) and acetylene-containing atmospheres were measured in comparable 30 min assays. Total electron flow through nitrogenase in air was determined from rates of nitrogen (15N2) fixation (x3) plus hydrogen evolution, that in the presence of acetylene from rates of acetylene reduction and hydrogen evolution in air : acetylene. Values for the ratio of electron flow in air : acetylene to that in air ranged from 0-43 to 0-83 in A. pulchella, from 0-44 to 0-66 in A. alata and from 0-37 to 0-70 in A. extensa, indicating substantial inhibition of electron flow through nitrogenase of detached nodules by acetylene. Relative efficiencies of nitrogenase functioning based on hydrogen evolution and acetylene reduction were from 015 to 0-79, those based on nitrogen (15N2) fixation and hydrogen evolution from 0-53 to 0-87. Molar ratios of acetylene reduced to nitrogen (15N2) fixed were 2-82 ±0-24, 2-01 ±0-15, and 1-91 +0-11 (x + s.e.; n = 7) for A. pulchella, A. extensa and A. alata respectively. A standard 5-10 min acetylene reduction assay, conducted on freshly detached unwashed nodules in daytime (12.00-14.00 h), was calibrated for field use by comparing total N accumulation of seedlings with estimated cumulative acetylene reduction over a 7-week period of glasshouse culture. Molar ratios for acetylene reduced : nitrogen fixed using this arbitrary method were 3-58 for A. alata, 4-82 for A. extensa and 1-60 for A. pulchella. The significance of the data is discussed. Key words—Acacia spp., nitrogenase functioning.

Relationships between Acetylene Reduction Activity, Hydrogen Evolution and Nitrogen Fixation in Nodules of Acacia spp.: Experimental Background to Assaying Fixation by Acetylene Reduction under Field Conditions

Craig A. Atkins

Papers

Economy of Photosynthate Use in Nitrogen-fixing Legume

Mobilization of Nitrogen in Fruiting Plants of a Cultivar of Cowpea

Symbiotic Performance of Supernoclulating Soybean (Glycine max (L.) Merrill) Mutants during Development on Different Nitrogen Regimes

Post-Anthesis Economy of Carbon in a Cultivar of Cowpea

Relationships between Acetylene Reduction Activity, Hydrogen Evolution and Nitrogen Fixation in Nodules of Acacia spp.: Experimental Background to Assaying Fixation by Acetylene Reduction under Field Conditions