Translocation of Nitrogenous Compounds in Symbiotic and Nitrate-Fed Amide-Exporting Legumes

Peoples, M. B., Sudin, M. N. and Herridge, D. F. 1987. Translocationof nitrogenous compounds insymbiotic and nitrate-fed amide-exportinglegumes.–J. exp. Bot. 38: 567–579. The transport of nitrogen from the roots and nodules of chickpea(Cicer anetinum L.), lentil (Lens culinaris Medic), faba bean(Vicia faba L.) and pea (Pisum sativum L.) was examined in glasshouse-grownplants supplied either with nitrate-free nutrients or with nutrientssupplemented with 1,2,4 or 8 mol m^-3153N-nitrate. A sixth treatmentcomprised uninoculated plants supplied with 8–0 mol m^-31513N-nitrate. For each species, more than 75% of the nitrogenwas exported from the nodules as the amides, asparagine andglutamine. In fully symbiotic plants, the amides also dominatednitrogen transport to the shoot When N2 fixation activity wasdecreased by the addition of nitrate to the rooting medium,the N-composition of xylem exudate and stem solutes changedconsiderably. The relative concentrations of asparagine tendedto increase in the xylem whilst those of glutamine were reduced;the levels of nitrate increased in both xylem exudate and thesoluble nitrogen pool of the stem with a rise in nitrate supply.The changes in relative nitrate contents reflected generallythe contributions of root and shoot to overall nitrate reductaseactivity at the different levels of nitrate used. The relationshipsbetween the relative contents of xylary or stem nitrate andamino nitrogen and the plants' reliance on N2 fixation (determinedby the ¹⁵N isotope dilution procedure) were examined. Data suggestthat compositional relationships based on nitrate may be reasonableindicators of symbiotic dependence for all species under studyexcept faba bean when greater than 25% of plant nitrogen wasderived from N2 fixation. Key words: Nitrogen, translocation, legumes     

Development of the Xylem Ureide Assay for the Measurement of Nitrogen Fixation by Pigeonpea (Cajanus cajan (L.) Millsp.)

Quantification of N₂ fixation by pigeonpea (Cajanus cajan (L.)Millsp.) in the field has proved difficult using techniquessuch as ¹⁵N isotope dilution, acetylene reduction and N difference.We report experiments to develop the ureide assay of N₂ fixationbased on extraction and analysis of xylem exudate. Plants ofpigeonpea cv. Quantum, inoculated with effective Rhizobium spp.CB756, were grown in a temperature-controlled glasshouse inlarge pots filled with a sand: vermiculite mixture, in waterculture and in a slightly acidic, red-brown earth in replicatedfield plots. Xylem exudate was collected as bleeding sap fromboth nodulated and unnodulated roots, and from detached nodules.Exudate was extracted also from detached shoots and stems ofpigeonpea using a mild vacuum (60–70 kN m^–2). Largedifferences in the composition of N solutes exported from rootsof N₂-dependent and nitrate-dependent plants suggested thatshifts in plant dependence on N₂ fixation may be reflected byconcomitant changes in N solutes. Thus, nodulated plants weresupplied throughout growth with either N-free nutrients or nutrientssupplemented with 1, 2, 5, 5, 10, or 20 mol m^{–3 15}. Plants were harvested at regular intervals fordry matter and vacuum-extracted exudate. The relative abundanceof ureides ([ureide-N/ureide-N + nitrate-N + -amino-N] ? 100)in the exudate was highly correlated with the proportion ofplant N (calculated using a ¹⁵N isotope dilution technique)derived from N₂ fixation. Two distinct phases of plant growthwere recognized and standard curves were prepared for each.The relationship between proportional dependence of plants onN₂ and xylem relative ureides was unaffected by mineral-N source,i.e. nitrate or ammonium. This result is discussed in relationto interpretation of material from field-grown plants. The effectsof plant genotype, strain of rhizobia, section of stem extracted,removal of leaves, time delay between shoot detachment and extraction,and diurnal characteristics were examined in order to identifypotential sources of error and to optimize sampling procedures. Key words: Ureides, allantoin, allantoic acid, N₂ fixation, pigeonpea, Cajanus cajan     

The Xylem Ureide Assay of Nitrogen Fixation: Sampling Procedures and Sources of Error

Herridge, D. F., O'Connell, P. and Donnelly, K. 1988. The xylemureide assay of nitrogen fixation: sampling procedures and sourcesof error.—J. exp. BoL 39: 12–22. The relative abundance of ureides [(ureide-N/ureide-N + NO₃^––N+ amino-N)x 100] in vacuum-extracted xylem exudate of soyabean(Glycine max [L.] Merr.) can be used to quantify N₂ fixationactivity. We report experiments designed to (1) develop appropriateprocedures for sampling and storage of xylem exudate extractedfrom plants in the field and (2) develop a single plant, non-destructivesampling procedure, useful for evaluation of segregating breedingmaterial. Soyabean cultivars Bragg, Essex, Davis, Hardee andDodds were grown at three sites at Brecza, New South Wales,on a deep, alkaline vertisol. All plots were inoculated witheffective Bradyrhizobium japonicum CB 1809. Vacuum strengthsof between 50•5 and 80•8 kN m^–2 did not affectthe relative abundance of ureides of xylem exudate. Delays betweenplant sampling and extraction of exudate, however, resultedin time-dependent increases in the relative ureide contents.Extraction should immediately follow shoot detachment to allowvalid comparison of treatments and accurate interpretation ofdata. The relative ureide contents of exudate showed only minordiurnal fluctuations. Exudate, once collected, could be stabilizedby dilution (1:1) in absolute ethanoL Both diluted and undilutedsamples were stable for at least 7 d at 4°C. Differencesduring early flowering, but not subsequently, in the compositionsof N solutes of exudate extracted from either the whole shoot,the top half or the lower half indicated the possibility ofnon-destructive sampling where seed, as well as xylem exudate,could be harvested. Best results in terms of volume of exudateextracted, treatment separation and seed production were obtainedwhen single plants were sampled during late flowering by detachmentof shoots at internode 5. Key words: Ureides, nitrogen fixation, soyabean     

Variation in Colony Characteristics and Symbiotic Effectiveness of Rhizobium

Seventeen cultures of Rhizobium CB756 varied in symbiotic effectiveness and contained a number of different colony types. An examination of single colony isolates from one culture of CB756 indicated that colony characteristics of most isolates were unstable and did not breed true. There was a relationship between symbiotic effectiveness and colony type of the original isolate. The 3 most ineffective sub-strains were all isolated from large, gummy colonies whereas the most effective were isolated from pinpoint, dry colonies.     

Partitioning and Utilization of Net Photosynthate in a Nodulated Annual Legume

The economy of carbon in nodulated white lupin (Lupinus albusL.) was studied in terms of consumption of net photosynthatein nitrogen fixation, in maintenance of respiration, and inthe production of dry matter and protein. Net photosynthesisrose to a maximum in early fruiting and then fell abruptly dueto shedding of leaves. Nodulated roots acquired translocateequivalent to 51% of the plant's net photosynthate, 78% of thecarbon of this translocate being respired, 10% entering drymatter, and 12% returning to the shoot attached to productsof nitrogen fixation. Nodules utilized 4?0–6?5 g C infixing 1 g nitrogen. Photosynthate was utilized most effectivelyfor nitrogen fixation in late vegetative growth. Fruits sequestered16% of the plant's net photosynthate, shoot night respiration17%, and dry matter formation in shoot vegetative parts 22%.Averaged over growth, 9?9 g net photosynthate was required toproduce 1 g seed dry matter and 31 g net photosynthate to produce1 g seed protein. Budgets for utilization of the carbon of netphotosynthate were constructed for 10 d intervals of the plant'sgrowth cycle. Feeding of shoots with ¹⁴CO₂ resulted in radiocarbonbecoming partitioned approximately as predicted by these budgets.The dependence of root respiration on recent photosynthate wasassessed by following the time course of release of ¹⁴CO₂ tothe rooting medium of the ¹⁴CO-labelled plants.