Interrelationship Between Plant Developmental Stage, Plant Growth Rate, Nitrate Utilization and Nitrogen Fixation in Hydroponically Grown Soybean

The growth rate of the indeterminate soybean plant [Glycinemax (L.)Merr.] slows as it proceeds from vegetative phase intoreproductive growth. Yet, the well-nodulated plant acquiresmost of its nitrogen during reproductive growth. Thus, the interrelationshipbetween plant developmental stage and nitrogen fixation wasexamined. It is shown that, regardless of the age of the hydroponicallygrown soybean plant at the time of its inoculation with Bradyrhizobiumjaponicum, the highest rate of nitrogen fixation occurs duringthe pod-filling stage (R5). Nevertheless, maximum total nitrogenfixation is generally achieved when inoculation occurs at thefull-bloom stage (R2). It is shown, however, that flower budsand flowering are not responsible for the enhanced nodulationand nitrogen fixation. Rather, the data suggest that the onsetof rapid nodulation occurs soon after the initiation of thedevelopmentally programmed drop in foliar nitrate reductaseactivity. The ensuing increase in nitrogen fixation providesthe plant with much of its needed nitrogen and hence stimulatesplant mass accumulation during pod-fill. It is suggested thatnitrogen fixation enhances growth of the soybean plant by increasingits net photosynthetic efficiency during reproductive growthand by providing the needed nitrogen at the appropriate timefor maximum seed growth. Key words: Glycine max, nitrate, nitrogen fixation, nodulation     

Enhanced Nitrogen Fixation Increases Net Photosynthetic Output and Seed Yield of Hydroponically Grown Soybean

Ten to 20% of the net photosynthetic output of a tropical grainlegume may be consumed by the nodulation-nitrogen-fixation process.If plant growth activities during the reproductive phase werelimited by photosynthetic output, enhanced nitrogen fixationwould seemingly lower total plant mass and seed yield. To testthis possibility, soybean [Glycine max (L.) Merr.] plants weregrown hydroponically on nutrient medium supplemented with minimalurea or with an excess of either nitrate or nitrate plus urea.Acetylene reduction activities (i.e. nitrogen fixation rates)and transpiration rates were measured twice weekly on theseplants through pod fill. Of the plants inoculated, those grownon minimal urea revealed significantly greater acetylene reductionactivities and transpiration rates. At maturity, plants thathad fixed nitrogen at a rapid rate during pod fill had a significantlygreater seed size, total plant mass (i.e. net photosyntheticrate) and nitrogen content than uninoculated or poorly nodulatedplants grown on an excess of nitrate. It is concluded, therefore,that a rapid rate of nitrogen fixation during pod fill enhancesboth transpiration and net photosynthetic output. The increasedavailability of usable nitrogen (i.e. ureides), coupled withenhanced transpiration and photosynthetic output, significantlyincreases total plant mass and seed yield. Thus, enhanced nitrogenfixation seems to be an inexpensive means of increasing seedyield of soybean and perhaps of other tropical grain legumes. Key words: Glycine max, nodulation, nitrate, urea     

Nitrate-Ammonium Ratio Required for pH Homeostasis in Hydroponically Grown Soybean

Imsande, J. 1986. Nitrate-ammonium ratio required for pH homeostasisin hydroponically grown soybean.—J. exp. Bot. 37: 341–347. Plant acid-base homeostasis is achieved when the mmoles of hydroxylions produced in the plant equal the mmoles of protons. Reductionof nitrate to ammonia is the major source of hydroxyl ions whereasammonium uptake-assimilation and the metabolism of neutral sugarsto organic acids are the primary sources of protons. Soybean[Glycine max (L.) Merr plants were grown hydroponically on mediumsupplemented with 3·0 mol m^–3 nitrogen providedas various combinations of KNO₃ and NH₄NO₃ Plant growth consumedessentially all available nitrogen in each case; however, onlyin flasks supplemented with approximately 1·8 minolesof KNO₃ plus 0·6 mmole of NH₄NO₃ was the pH of the mediumunchanged. Thus, for every mmole of nitrogen assimilated, approximately0·6 mmole of dissociable protons must have been producedby the conversion of neutral sugars to carboxylic acids. Also,it was shown that a plant obtaining all of its nitrogen fromnitrate must neutralize or excrete approximately 0·5mmole of hydroxyl ion d^–1. Conversely, the plant derivingall of its nitrogen from dinitrogen must excrete or neutralizeat least 0·8 mmole of hydrogen ion d^–1 whereasthe plant deriving all of its nitrogen from aminonium must excreteor neutralize approximately 2·1 mmoles of hydrogen iond^–1. Nevertheless, plants grown on medium supplementedwith 2·4 mol m^–1 nitrate plus 0·6 mol m^–3ammonium did not achieve a higher growth rate than plants grownon 3·0 mol m^–3 nitrate. Key words: Glycine max, nitrogen fixation, nitrate utilization     

Direct Test for Altered Gas Exchange Rates in Water stressed Soybean Nodules

Water stress usually lowers the nitrogenase activity of soybeanroot nodules. This loss in activity might result from an increasedbarrier to nodular gas exchange, from a general reduction inbiochemical function, or both. To test for the possibility ofan increased barrier to gas diffusion, we measured the apparentlag time for initiation of acetylene reduction by intact soybeanplants, both before and after water stress. Mild nodular waterloss (i.e. 10% of fresh weight or less) seldom altered the apparentlag time, whereas severe water stress (20–40% f. wt loss)frequently produced a small increase in apparent lag time. Severewater stress also produced a large decrease (24%) in the externaldiameter of the nodules and a loss of the white lenticel traces.Water stress usually caused a decrease in the apparent K_m foracetylene. The data do not suggest a large change in the diffusiveresistance to acetylene of nodules subjected to water stress.Thus, the observed decrease in nitrogenase activity may resultprimarily from biochemical, rather than physical, changes. However,because of the relatively greater importance of gas-phase diffusionfor oxygen entry, we cannot exclude the possibility of a largechange in a small gas pathway that affects oxygen influx morethan acetylene influx. Diffusion, Glycine max, nitrogen fixation, water stress     

Entry of Oxygen and Nitrogen into Intact Soybean Nodules

The apparent nitrogenase activity of soybean root nodules isreduced approximately 70% upon nodul detachment. By increasingthe partial pressure of oxygen in the assay vessel from approximate0.2 to 0.5 atm, the nitrogenase activity of detached nodulesis restored to the original attached level. Oi the other hand,maximal assayable nitrogenase activity of nodules attached tothe intact soybean plan is achieved in the presence of 0.2–0.3atm of oxygen. Exposure of attached nodules to 0.5 atm oxygeicauses a decrease in nitrogenase activity seemingly due to inactivation of nitrogenase. These and othe data suggest that,upon detachment, the entry of oxygen into the nodules is impaired.A model for th regulation of airflow into attached and detachednodules is presented. This model is also consisten with responseof nodule activity to water stress.     

Inhibition of Nodule Development in Soybean by Nitrate or Reduced Nitrogen

Imsande, J. 1986. Inhibition of nodule development in soybeanby nitrate or reduced nitrogen.—J. exp. Bot. 37: 348–355. Nodulation of hydroponically grown soybean plants [Glycine max(L.) Merr.] is inhibited by continuous growth in the presenceof 4· mol m^–3 KNO₃ The presence of 4·0 molm^–3 ‘starter nitrate’ for 3-6 d during noduledevelopment, however, subsequently stimulates nodule dry weightaccumulation and nitrogenase activity. These stimulations occureven though 4· mol m^–3 nitrate temporarily delaysnodule development, i.e. the late steps of nodule developmentare reversibly inhibited by a short-term exposure to 4·0mol m^–3 nitrate. On the other hand, treatment with 4·0mol m^–3 nitrate in excess of 14 d significantly reducesnodule dry weight Thus, extended growth in the presence of 4·0mol m^–3 KNO₃ seems to block both early and late stepsof nodule development. Nodulation of hydroponically grown soybeansis also inhibited by continuous growth in the presence of 2·0mol m^–3 (NH₄)₂SO₄ This inhibition is not caused by acidityof the growth medium. On the other hand, nodule development6 d after inoculation with Rhizoblum japonicum is not delayedby a 7-d exposure to 2·0 mol m^–3 (NH₄)₂SO₄ butis partially inhibited by a prolonged exposure to (NH₄)₂SO₄Because repression of nodulation by 4·0 mol m^–3KNO₃ is more severe than that by 2·0 mol m^–3 (NH₄)₂SO₄and because ammonium taken up by the soybean plant is not activelyoxidized to nitrate, it is suggested that there are at leasttwo mechanisms by which nitrate utilization represses noduleformation in soybean. Key words: Glycine max, nitrogen, nitrogen fixation, nodulation     

Regulation of Nodule Efficiency by the Undisturbed Soybean Plant1

Acetylene reduction activity (ARA) has been widely used to estimatecomparative rates of nitrogen fixation in soybean [Glycine max(L.) Merr.]. To use the non-destructive closed-container ARAassay to study plant-mediated regulation of N₂ fixation, itwas first necessary to determine the validity of the ARA procedure.Thus, well-nodulated, hydroponically-grown soybean plants wereindividually assayed for ARA twice weekly for 5 weeks. Plantswere subsequently analysed for total nitrogen by the Kjeldahlprocedure and the values obtained were compared with the N-accumulationvalues estimated by ARA. For the 168 plants examined, ARA measurementsunderestimated N₂ fixation by approximately 50%. However, N₂fixation was underestimated by approximately 80% in plants witha very low ARA whereas N₂ fixation was underestimated by only20% in plants with a high ARA. Because of the accuracy and reliabilityof the Kjeldahl measurements, these results show that nodulatedsoybean plants can exhibit different efficiencies in acetylenereduction. Thus, vigorous, well-nodulated, hydroponically-grownsoybean plants readily control nodule efficiency and the N fixationprocess.     

Nodulation of Hydroponically Grown Soybean Plants and Inhibition of Nodule Development by Nitrate

A procedure is described for obtaining a large number of nodules(approximately 1000) on a hydroponically grown soybean plantafter a brief exposure to Rhizobium japonicum. This procedurewas used to study the effect of medium nitrate upon nodulationof the soybean plant. It was found that nitrate, at concentrationsequal to or greater than 2.0 mM, restricted nodule developmentbut did not inhibit appreciably the activity or synthesis ofnitrogenase in those few nodules that were formed. Key words: Soybean, Rhizobium japonicum, Nodulation