Possible reasons for relative salt stress tolerance in nodules of white lupin cv. Multolupa

The effects of different NaCl concentrations on the growth andnitrogen fixation activity of white lupin (Lupinus albus [L.])was studied over a 6 d period. Plant growth parameters, photosynthesisand shoot respiration were unaffected by NaCl concentrationsup to 150 mol m^–3. However, nitrogenase activity decreasedwith increased NaCl concentration up to 100 mol m_–3, whilstthe O₂ diffusion resistance increased with 100 mol m^–3NaCl, but showed no further change when 150 mol m^–3 NaClwas applied for 6 d. Increases in NaCl concentration decreasednodular starch content while increasing sucrose content, suggestingan osmotic regulation. These changes were associated with a77% decrease in sucrose synthase activity. The effect on theO₂ diffusion resistance was paralleled by changes in glycoproteincontent of the nodules, as determined by immunogold localizationand ELISA. X-ray microanalysis studies of nodules showed that,following a 6 d exposure to 150 mol m^–3 NaCl, Na⁺ ionswere largely excluded from the infected zone, whilst only lowlevels of Cl^- ions penetrated into this region. Na⁺ entry intoroots and leaves was also at a low level. Leghaemoglobin contentdecreased with saline stress, as did superoxide dismutase; whichdecreased by 36% following exposure to 100 mol m^–3 saltfor 6 d. These results are discussed in relation to the relativesalt tolerance of the Multolupa/ISLU-16 symbiosis. Key words: Salt stress, nodules, nitrogen fixation, oxygen diffusion, carbohydrates, Lupinus albus     

Nodule-specific host proteins in effective and ineffective root nodules of Pisum sativum

Nodule-specific root proteins – so called nodulins – were identified in root nodules of pea plants by an immunological assay. Nodulin patterns were examined at different stages of nodule development. About 30 nodulins were detectable during development. Some were preferentially synthesized before nitrogen fixation started, whereas the majority were synthesized concomitantly with leghaemoglobin. Some of the nodulins were located within the peribacteroid membrane. Ineffective Rhizobium strains (a natural nod⁺fix^- and a pop ^-fix^-) appeared to be useful in studying the expression of nodulin genes. Synthesis of some nodulins was repressed in ineffective root nodules, indicating that nodulins are essential for the establishment of nitrogen fixation. In both types of ineffective root nodules, leghaemoglobin synthesis was not completely repressed. Low amounts of leghaemoglobin were always detected in young ineffective root nodules whereas in old nodules no leghaemoglobin was present.     

Biological and Cultural Characteristics of the Effective Frankia Strain HFPCcl3 (Actinomycetales) from Casuarina cunninghamiana (Casuarinaceae)

From homogenates prepared from surface-sterilized nodules ofseedlings of Casuarina cunninghamiana grown aeroponically, astrain of Frankia designated HFPCc13 was isolated and has beengrown in pure filamentous culture in a defined synthetic nutrientmedium. Vesicle and sporangium formation can be induced by removalof combined nitrogen from the medium.Frankia strain HFPCc13nodulates young seedlings of C. cunninghamiana and C. equisetifoliawithin three weeks of inoculation with an optimum root mediumpH of 6–7 for nodulation and optimum temperature of 30–35^°C. The presence of combined nitrogen in the root mediuminhibits nodulation with NH₄⁺ more inhibitory than NO₃^–.Frankia HFPCc13 does not nodulate Allocasuarina species withinthe same family nor several other possible actinorhizal plantstested. Thus this strain is quite precise in its host specificity.The rate of acetylene reduction was greater in C. cunninghamianathan the closely related species C. equisetifolia. In neitherof these host species were vesicles observed to occur withinthe infected root nodules which had been demonstrated to beactively fixing dinitrogen. Root nodules were shown to be activein acetylene reduction over a range of O₂ concentration in thegaseous environment with an optimum at about 20 per cent O₂,the ambient P_O2 of the air. The mechanism(s) for oxygen protectionof nitrogenase within the filamentous form of Frankia withinthese nodules remains to be explained. Casuarina, Frankia, nodulation, nitrogen fixation     

The nodulation of micro-propagated plants of Parasponia andersonii by tropical legume rhizobia

A simple clonal micro-propagation system for Parasponia andersoniiwas employed to study the nodulation response of this non-legumeto inoculation by the broad host range Rhizobium sp. NGR234,isolated from Lablab purpureus, and also to tropical legumerhizobia isolated from Aeschynomene species. Partially effectivenodules, assayed by acetylene reduction and ¹⁵N dilution procedures,were induced with strain NGR234 and its spontaneous streptomycinresistantmutant ANU240. Effective nodules were produced by one of theAeschynomene strains (ORS302) tested, with rates of acetylenereduction comparable to those of root nodules produced by Bradyrhizobiumstrain CP279, originally isolated from P. andersonii. Lightand transmission electron microscopy showed that there was acorrelation between the nitrogen fixing capability of the symbiosisbetween NGR234 and Parasponia and the number of persistent infection(fixation) threads within the nodule cells. Key words: Parasponia, Bradyrhizobium, Rhizobium, Aeschynomene, micro-propagation, root nodules, nitrogen fixation     

A Host-specific, Nitrogen Fixation Mutant of Bradyrhizobium: Physiology on Three Host Plants

NC92 #284 is a transposon mutant of Bradyrhizobium sp. (Arachis)strain NC92 and has a host-specific fixation phenotype. It appearsto be ineffective on the host pigeonpea (90% reduction in shootN compared to that of the wild type), but partially effectiveon two other host plants, groundnut and siratro (50% and 20%reduction in shoot N compared to the wild type, respectively).To understand this phenomenon of host-specific fixation, thephysiological basis of the phenotypes was investigated. Host-dependentdifferences in symbiotic effectiveness were largely explainedby the degree to which nitrogenase activity was impaired inthe various #284 symbioses. Nodulation and the onset of nitrogenfixation were found to be delayed on all three hosts, but tothe greatest degree on pigeonpea. The specific activity of nitrogenaseper gram nodule was also reduced on all three hosts, again tothe greatest extent on pigeonpea. By contrast, the carbon costsand relative efficiencies of each symbiosis were similar tothe wild type. The results indicate that the host-specific fixationphenotype of #284 is reflected in a quantitative reduction inthe amount of N₂ fixed. Thus the phenotypes reflect the differentability of the three host plants to tolerate or support the#284 mutation, rather than a defect in a specific interactionbetween #284 and a particular host plant. Key words: Bradyrhizobium, nodules, nitrogenase activity, relative efficiency, Arachis hypogaea, Cajanus cajan, Macroptilium atropurpureum     

Xylem fluxes of fixed N through nodules of the legume Acacia littorea and haustoria of an associated N-dependent root hemiparasite Olax phyllanthi

Nodulated 1-1.5-year-old plants of Acacia littorea grown inminus nitrogen culture were each partnered with a single seedlingof the root hemiparasite Olax phyllanthi. Partitioning of fixedN between plant organs of the host and parasite was studiedfor the period 4–8 months after introducing the parasite.N fluxes through nodules of Acacia and xylem-tapping haustoriaof Olax were compared using measured xylem flows of fixed Nand anatomical information for the two organs. N₂ fixation duringthe study interval (635 µg N g FW nodules^–1 d^–1)corresponded to a xylem loading flux of 0.20 µg N mm^–2d^–1 across the secretory membranes of the pencycle parenchymaof the nodule vascular strands. A much higher flux of N (4891µg mm^–2 d^–1) exited through xylem at the junctionof nodule and root. The corresponding flux of N from host xylemacross absorptive membranes of the endophyte parenchyma of Olaxhaustorium was 1.15 µg N mm^–1 d^–1, six timesthe loading flux in nodules. The exit flux from haustorium toparasite rootlet was 20.0 pg N mm^–1 d^–1, 200-foldless than that passing through xylem elements of the nodule.Fluxes of individual amino compounds in xylem of nodule andhaustorium were assessed on a molar and N basis. N flux valuesare related to data for transpiration and partitioning of Cand N of the association recorded in a companion paper. Key words: Olax phyllanthi, host-parasite relationships, N flux, Acacia, N₂ fixation     

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

Hansen, A. P., Pate, J. S. and Atkins, C. A. 1987. Relationshipsbetween acetylene reduction activity, hydrogen evolution andnitrogen fixation in nodules of Acacia spp.: Experimental backgroundto assaying fixation by acetylene reduction under field conditions.—J.exp. Bot. 38: 1–12 Glasshouse grown, symbiotically-dependent seedlings of Acaciaalata R.Br., .A. extensa Lindl., and A. pulchella R.Br. wereexamined for acetylene reduction in closed assay systems usingundisturbed potted plants, excavated whole plants, nodulatedroots or detached nodules. Nitrogenase activity declined sharplyover 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 periodin assays involving attached nodules. Using detached nodules,rates of acetylene reduction, nitrogen (¹⁵N₂) fixation, andhydrogen evolution in air (¹⁵N₂) and acetylene-containing atmosphereswere measured in comparable 30 min assays. Total electron flowthrough nitrogenase in air was determined from rates of nitrogen(¹⁵N₂) fixation ( ? 3) plus hydrogen evolution, that in thepresence of acetylene from rates of acetylene reduction andhydrogen evolution in air: acetylene. Values for the ratio ofelectron flow in air: acetylene to that in air ranged from 0?43to 0?83 in A. pulcheila, from 0?44 to 0?66 in A. alala and from0?37 to 0?70 in A. extensa, indicating substantial inhibitionof electron flow through nitrogenase of detached nodules byacetylene. Relative efficiencies of nitrogenase functioningbased on hydrogen evolution and acetylene reduction were from0?15 to 0?79, those based on nitrogen (¹⁵N₂) fixation and hydrogenevolution from 0?53 to 0?87. Molar ratios of acetylene reducedto nitrogen (¹⁵N₂) fixed were 2?82 ? 0?24, 201 ? 0?15, and 1?91? 0?11 (?s.e.; n = 7) for A. pulcheila,A. extensa and A. alata respectively A standard 5–10 min acetylene reduction assay, conductedon freshly detached unwashed nodules in daytime (12.00–14.00h), was calibrated for field use by comparing total N accumulationof seedlings with estimated cumulative acetylene reduction overa 7-week period of glasshouse culture. Molar ratios for acetylenereduced: nitrogen fixed using this arbitrary method were 3?58for 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     

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     

Effects of NaCl on Growth, Nitrogen Incorporation and Chemical Composition of Inoculated and NH4NO3 Fertilized Vicia faba (L. ) Plants

Vicia faba cv. Maris Bead was grown either on fixed nitrogenor on ammonium nitrate. After 4 weeks growth, nutrient solutionswere supplemented with 50, 75 and 100 mol m^–3 NaCl for15 d. Five harvests were made at weekly intervals, beginningat 4 weeks. Effects of salinity were directly related to dose,plant growth (fresh and dry weight) being depressed in bothN-fixing and N-fertilized plants. The number of nodules perplant and the proportion of those formed which developed intothe active nitrogen fixing state were depressed by salt stress.Increased size of nodules in salt-stressed plants only partlycompensated for the lower specific nitrogenase activity. Theeffects of salinity on plant nitrogen content were more pronouncedon N-flxing than on N-fertilized plants. The former took upmore Na⁺ and Cl^– than the latter: the implications ofthis and of ionic imbalance are discussed. Key words: Vicia faba, Growth, Salt stress, Nodulation     

The Growth and Photosynthesis of Seedling Plants of White Clover at Low Temperature

The growth of white clover (Trifolium repens L.) in conditionstypical of April in Southern England (8 °C day/4 °Cnight, 12 h photoperiod of 90 J m^–2 s^–1 visibleradiation) was extremely slow, whether the plants were dependentfor nitrogen on fixation by their root nodules or were suppliedwith abundant nitrate; although growth was slower in the nodulatedplants. The reasons for slow growth were a large root: shootratio and a small leaf area, particularly in the nodulated plants,and a low photosynthetic rate in all plants. The probable effectsof these characteristics on the growth of white clover withgrasses in mixed pastures are discussed. Trifolium repens L, white clover, low temperature, leaf area, photosynthetic rate, nitrogen supply, growth     

Anatomy, physiology and biochemistry of root nodules of Sprint-2 Fix-, a symbiotically defective mutant of pea (Pisum sativum L.)

A plant-determined pea mutant Sprint-2 Fix^– and the parentalline Sprint-2 were compared for selected physiological and biochemicalparameters. The Fix^– mutation prevented differentiationof Rhizobium leguminosarum bacteria into bacteroids and producedlarge, white, non-fixing nodules. These lacked nitrogenase-linkedrespiration, but had a background rate of CO₂ evolution similarto the normal Fix⁺ nodules. The cortical structure of the ineffectivenodules suggests the existence of an oxygen diffusion barrierand this was supported by a low oxygen concentration in thecentral region (0.5–3.0%), measured using an O₂ sensitivemicro-electrode. Sucrose and starch contents were similar innormal and ineffective nodules while ononitol content was about15 times lower in the Fix^– nodules. The distribution ofstarch was also different in the two nodule types. The activitiesof glutamine synthetase (GS), sucrose synthase (SS), phosphoenolpyruvatecarboxylase (PEPC) and alanine pyruvate aminotransferase (APAT)were markedly higher in Fix⁺ nodules while the activities ofpyruvate decarboxylase (PDC), alcohol dehydrogenase (ADH) andglutamate dehydrogenase (GDH) were higher in Fix^– nodules.The data from immunodetection of host nodule proteins confirmedthe reduced levels of sucrose synthase and the almost completeabsence of glutamine synthetase and leghaemoglobin in mutantnodules. There was no significant difference in the amount ofnitrogenase component 1 extracted from the microsymbiont ofnormal and ineffective nodules, but component 2 was hardly detectablein the Fix^– mutant. Key words: Pisum sativum, Fix^– mutant, nodules     

Hydrogen Effect on Nitrogenase (C2H2 Reduction) Response to Oxygen in Bradyrhizobium japonicum-Phaseoleae Symbioses

The influence of hydrogenase in Bradyrizobum-Phaseoleae symbioseswas studied ex-planta and in-planra in soybean (Glycine max)and cowpea (Vigna unguiculata). The hydrogenase was activatedby the addition of hydrogen in the incubation gas phase whichmodified the response of nitrogenase activity of Hup⁺ (hydrogenuptake positive) symbiosis to the external oxygen partial pressure.For bacteroids the hydrogenase expression increased nitrogenaseactivity at supraoptimal pO₂, acting possibly as a respiratoryprotection of nitrogenase. However, at suboptimal pO₂, nitrogenaseactivity of Hup⁺ bacteroids decreased with hydrogen, a phenomenonattributed to the lower efficiency of ATP synthesis from hydrogenthan from carbon substrates oxidation. For undisturbed nodules,the hydrogenase expression in soybean increased the optimalpO₂ for ARA (COP), from 35.3 to 40.3 kPa O₂, and the ARA atsupraoptimal pO₂; at suboptimal PO₂ there was a negative effectof hydrogenase on ARA, although this inhibition was less thanon bacteroids and was not detected if plants were grown at 15°C rather than 20 °C root temperature. No H₂ effectwas detected on cowpea nodules. The results on soybean nodulesare consistent with the concept that symbiotic nitrogen fixationis oxygen-limited and that hydrogenase activity has no beneficialeffect on nitrogen fixation in O₂ limitation. Key words: Glycine max, hydrogenase, nitrogenase, nitrogen fixation, nodules, Vigna unguiculata     

Concurrent Rates of N2 Fixation, Nitrate and Ammonium Uptake by White Clover in Response to Growth at Different Root Temperatures

Nodulated white clover plants (Trifolium repens L. cv. Huia)were grown for 71 d in flowing nutrient solutions containingN as 10 mmol m_–3 NH₄NO₃, under artificial illumination,with shoots at 20/15°C day/night temperatures and root temperaturereduced decrementally from 20 to 5°C. Root temperatureswere then changed to 3, 7, 9, 11, 13, 17 or 25°C, and theacquisition of N by N₂ fixation, NH₄+ and NO₃^– uptakewas measured over 14 d. Shoot specific growth rates (d. wt)doubled with increasing temperature between 7 and 17°C,whilst root specific growth rates showed little response; shoot:root ratios increased with root temperature, and over time at11°C. Net uptake of total N per plant (N₂ fixation + NH₄++ NO₃^–) over 14 d increased three-fold between 3 and 17°C.The proportion contributed by N₂ fixation decreased with increasingtemperature from 51% at 5°C to 18% at 25°C. Uptake ofNH₄+ as a proportion of NH₄+ + NO₃^– uptake over 14 d variedlittle (55–62%) with root temperature between 3 and 25°C,although it increased with time at most temperatures. Mean ratesof total N uptake per unit shoot f. wt over 14 d changed littlebetween 9 and 25°C, but decreased progressively with temperaturebelow 9°C, due to the decline in the rates of NH₄+ and NO₃^–uptake, even though N₂ fixation increased. The results suggestthat N₂ fixation in the presence of sustained low concentrationsof NH₄+ and NO₄^– is less sensitive to low root temperaturethan are either NH₄+ or NO₃^– uptake systems. White clover, Trifolium repens L. cv. Huia, root temperature, nitrogen fixation, ammonium, nitrate     

Comparisons of the Respiratory Effluxes of Nodules and Roots in Six Temperate Legumes

The specific respiration rates of nodulated root systems, ofnodules and of roots were determined during active nitrogenfixation in soya bean, navy bean, pea, lucerne, red clover andwhite clover, by measurements on whole plants before and afterthe removal of nodule populations. Similar measurements weremade on comparable populations of the six legumes, lacking nodulesbut receiving abundant nitrate-nitrogen, to determine the specificrespiration of their roots. All plants were grown in a controlled-environmentclimate which fostered rapid growth. The specific respiration rates of nodulated root systems ofthe three grain and three forage legumes during a 7–14-dayperiod of vegetative growth varied between 10 and 17 mg CO₂g^–1 (dry weight) h^–1. This mean value consistedof two components: a specific root respiration rate of 6–9mg CO₂ g^–1 h^–1 and a specific nodule respirationrate of 22–46 mg CO₂ g^–1 h^–1. Nodule respirationaccounted for 42–70 per cent of nodulated root respiration;nodule weight accounted for 12–40 per cent of nodulatedroot weight. The specific respiration rates of roots lackingnodules and utilizing nitrate nitrogen were generally 20–30per cent greater than the equivalent rates of roots from nodulatedplants. The measured respiratory effluxes are discussed in thecontext of nitrogen nitrogen fixation, nitrate assimilation. Glycine max, Phaseolus vulgaris, Pisum sativum, Medicago sativa, Trifolium pratense, Trifolium repens, soya bean, navy bean, pea, lucerne, red clover, white clover, nodule respiration, root respiration, fixation, nitrate assimilation     

Effects of Temperature on Growth and Nitrogen Fixation by Swards of Subterranean Clover

Subterranean clover (Trifolium subterraneum L.) cv. ‘Woogenellup’ swards were grown at 10, 15, 20 and 25 Cwith a 12 h photoperiod of 500 or 1000 µmol m^–2s^–1 [low and high photosynthetic photon flux density (PPFD)].Nitrogen-fixing swards received nutrient solution lacking combinednitrogen while control swards received a complete nutrient solution.Growth was measured by infra-red analysis of carbon dioxideexchange and by accumulation of dry matter. Swards were harvestedat intervals between 95 and 570 g d. wt m^–2 for estimationof nitrogenase activity by acetylene reduction and hydrogenevolution assays. Nitrogen fixation was also measured by increasein organic nitrogen. The growth rate was highest at 10 C at low PPFD, and at 10–15C at high PPFD. Nitrogen-fixing swards grew slower than thosereceiving combined nitrogen. Nitrogen fixation measured by increasein organic nitrogen responded similarly to the growth rate,as did acetylene reduction between 10 and 20 C. At 25 C therelationship between acetylene reduction and nitrogen fixationwas distrupted. The difference between the rates of acetylenereduction and hydrogen evolution, theoretically proportionalto nitrogen fixation, was not a reliable indicator of nitrogenfixation because hydrogen uptake developed. Trifolium subterraneum L, subterranean clover, growth, nitrogen fixation, temperature, acetylene reduction     

The Effect of Irradiance on the Recovery of Soybean Nodules from Sodium Chloride-Induced Senescence

Soybean (Glycine max L. Merr) cv. Clarke plants inoculated withBradyrhizobium japonicum strain RCR3407 were grown either ina greenhouse with a low irradiance (200–400)µmolm^–2 s^–1) or in a controlled-environment growth cabinetwith a higher irradiance (600 µimol m^–2 s^–1).At 42 d plants were given a nitrogen-free nutrient solutioncontaining 50 mol m^–3 sodium chloride for 2 weeks andthen allowed to recover from salt-stress for a further 2 weeks. Salt treatment reduced plant growth by at least half in bothgrowth regimes, however, the controlled environment-grown (CEG)plants were five times larger than the greenhouse-grown (GG)plants in terms of dry weight and number/weight of nodules perplant, regardless of treatment. The structure of nodules, from both growth regimes, harvestedat the end of the 2 week salt-stress was similar to unstressedcontrol nodules. However, nodules harvested 1 week later fromboth CEG and GG plants had structural changes including degradationof bacteria in vacuoles around host cell nuclei, particularlyin the outer cell layers of the infected tissue. In addition,meristematic activity was seen in the cortex of some nodulesfrom GG plants. Young cells here contained infection threadsand newly-released bacteria. Nodules harvested 2 weeks after removal of the salt-stress fromCEG plants showed an apparent recovery from the stress. However,there was a very marked increase in the amount of starch inthe cortex which was not seen in equivalent GG nodules. In contrast,nodules from GG plants contained many vacuolate infected cellsand, consequently, a lowered bacteroid population. Further,meristematic activity was seen in a zone concentric to the infectedzone, newly-formed cells contained many large infection threadsand were interspersed with intercellular bacteria. The meristematicactivity increased the relative volume of cortical to infectedcells in these nodules. Growth conditions did not affect control nodule specific nitrogenaseactivity or oxygen diffusion resistance (R) and these parameterswere also not altered in CEG nodules exposed to salt plus the14 d recovery period. However, nitrogenase activity was greatlyreduced, and R increased by more than eight times in equivalentGG nodules exposed to salt plus recovery. It is hypothesized that the gross morphological changes werean attempt to counter salt toxicity and/or oxygen damage underconditions of reduced photosynthate supply to the nodules dueto the poor light levels in the greenhouse. However, soybeannodules supplied with adequate photosynthate were able to withstandand recover from long-term salt-stress with little alterationto their structural integrity. Key words: Soybean, sodium chloride, nitrogen fixation, light intensity, oxygen diffusion resistance     

Effect of Nitrogen on Nitrate Reductase Activity in the Nodules and Leaves of Summer Moong (Vigna radiata)

The relation of the in vivo nitrate reductase (NR) activityto growth period was studied in the nodules and the leaves ofthe summer moong (Vigna radiata). The maximum NR activity wasobserved 31 days after sowing (DAS) in the leaves and 28 DASin the case of the nodules. In a pot experiment, the effectof the various nitrogen concentrations, namely 0, 3, 6, 9 and12 mg kg^–1 was studied on NR activity at three growthstages. The maximum NR activity was observed at 6 mg kg^–1N during the pre-flowering stage (26 DAS). Though the noduleshave higher NR activity, its expression was limited by substrateavailability. The NR activity in the leaf could be used as anindex of NR activity in the nodules. Nitrate reductase, nitrogen, nitrate, moong, Vigna radiata     

Rates of Hydrogen Ion Efflux by Nodulated Legumes Grown in Flowing Solution Culture with Continuous pH Monitoring and Adjustment

The net efflux of H⁺ from lucerne (Medicago saliva L.), redclover (Trifolium pratense L.) and white clover (Trifolium repensL.) growing in flowing solution culture and dependent upon symbioticfixation of atmospheric N, was measured over a 75 d experimentalperiod. Considerable and rapid increases in acidity of the nutrientsolution of up to 1.45 pH units were recorded when the pH wasriot held constant over a 30 h period. There was little differencein H⁺ efflux when solution pH was held constant at 4.75, 5.75or 6.75, but there was an immediate cessation when it was adjustedto 3.75. Differences in the daily net efflux of H⁺ closely followedthe pattern of daily differences in incoming radiation, andthere was also evidence of a diurnal pattern of H⁺ efflux. Althoughthere were initially distinct differences between the speciesin the calculated rate of net H⁺ efflux (µg H⁺ g^–1dry shoot d^–), by day 75 these had diminished. In allspecies, however, the maximum rate of efflux per unit of shootsoccurred during the earlier rapid phases of growth. The measuredefflux of H⁺ was well equated with the plant content of excesscations (as measured by ash alkalinity) and, on average, theratio of acidity produced to N assimilated (expressed as anequivalent) was 0-24. Medicago sativa L., Trifolium pratense L., Trifolium repens L., lucerne, red clover, white clover, acidification, cation/anion balance, flowing solution culture, H⁺ efflux, nitrogen fixation     

Oxygen Diffusion in Lupin Nodules: I.VISUALIZATION OF DIFFUSION BARRIER OPERATION

Root nodules of Lupinus albus (L.) cv. Multolupa were subjectedto short- and medium-term stresses by lowering rhizosphere temperaturefrom 25 to 16°C (2 h), detopping plants (3 h), darkeningplants (21 h) or exposing roots to 20 mol m^–3 KNO₃ for4 d. All experimental treatments produced increases in oxygendiffusion resistance, compared with control plants. These correlatedwith structural changes in the nodule cortex, which is describedin detail for the first time. The most noticeable change isthe occlusion of intercellular spaces by a glycoprotein whichwas identified using the monoclonal antibody MAC236. This glycoproteinwas also found surrounding bacteria in intercellular spacesof the cortex of control nodules. Key words: Oxygen diffusion resistance, glycoprotein, nodules, nitrogen fixation, Lupinus albus     

Directed transposon Tn5 mutagenesis and complementation in slow-growing, broad host range cowpea Rhizobium

Summary Site-directed Tn5 mutagenesis by gene replacement method, via homologous recombination, was used to identify symbiotically essential regions in the genome of cowpea Rhizobium spp. IRc78. Transposon insertions with-in the nifK hybridizing region or in the regions spanning 10 kb downstream of the nifK have revealed the presence of functional genes required for nitrogen fixation. Six single Tn5 insertions resulted in nod⁺ fix^– phenotypes and one in nod⁺ but reduced fix⁺ phenotype. All seven Tn5 insertions were stable before, during and after plant passage. However, IRc78 transconjugants containing duplicated nif copies, (a normal and a Tn5 inserted copy separated by vector sequences) were unstable. In five IRc78::Tn5 strains, the mutant phenotypes were corrected by an extrachromosomally stable vector containing wild type nif alleles. Our experiments suggest that the correction to nod⁺ fix⁺ phenotype is by complementation although correction by recombination cannot be completely excluded.