Nutrient Solution pH Control using Dipolar Buffers in Studies of Trifolium repens L. Nitrogen Nutrition

In studies of Trifolium repens nitrogen nutrition, the controlof nutrient solution pH using dipolar buffers, was evaluatedin tube culture under sterile conditions. Five buffers; MES,ADA, ACES, BES and MOPS with pK₂s (20 °C) of 6.15, 6.60,6.90, 7.15 and 7.20 respectively, at a concentration of 2.0mol m^–3, were provided to inoculated Trifolium repensgrowing in nutrient solution containing 7.13 mol m^–3 nitrogenas (NH₄)₂SO₄. Initial pH of each solution was adjusted to theappropriate buffer pK₂ Two buffers, ADA and ACES completelyinhibited plant growth. The remaining buffers had little effectin limiting pH change, although plant dry matter was higherand nodule numbers lower in the presence of these buffers. MESand MOPS were supplied to nutrient solutions with and without7.13 mol m^–3 (NH₄)₂SO₄, at concentrations ranging from0–12 mol m^–3. MES at 9 mol m^–3 and 12 molm^–3 reduced growth of plants reliant on the symbiosisfor providing nitrogen. The provision of MES to plants providedwith NH₄⁺ significantly increased plant yield and reduced nodulenumber at all concentrations. MOPS did not affect plant yieldor nodule number. The use of dipolar buffers in legume nitrogennutrition studies is considered in terms of buffering capacity,and the side effects on plant growth and symbiotic development. Key words: Ammonium, Dipolar buffer, Nitrogen nutrition, pH control, Symbiosis, Trifolium repens     

A comparison of the effect of combined nitrogen on nodulation in non-legumes and legumes

Summary 1. The effect of the presence of combined (ammonium) nitrogen in the rooting medium on nodule development has been investigated inAlnus glutinosa andMyrica gale, the plants being grown by a water culture technique with provision for the control of the level of combined nitrogen and of pH. For purposes of comparison a similar experiment with a legume (Ulex europaeus) has been included.2. In some instances the differential treatment with combined nitrogen was commenced and inoculation effected at an early stage in plant development, while in others the plants were grown on to a larger size before these treatments were applied.3. In the two non-legumes the presence of combined nitrogen led to an increase in the weight of nodulus formed per plant, at least at the lower levels of nitrogen. Relative to the enhanced growth of the plant as a whole, nodule development was continuously depressed as the level of combined nitrogen was increased.4. InUlex when the nitrogen status of the plants at the time of inoculation was similar to that in the corresponding non-legume experiment, the above responses were also shown by the legume. It is concluded thatAlnus andMyrica resemble legumes closely in their responses to combined nitrogen.5. The data show that inAlnus the effect of a given level of combined nitrogen on nodulation depends on the nitrogen status of the plant at the time of inoculation, the adverse effect tending to be stronger if the plant is initially relatively high in nitrogen. This suggests that as in legumes the effect of the combined nitrogen is exerted internally in the plant tissues.     

The Effects of Aluminium on the Growth of White Clover dependent upon Fixation of Atmospheric Nitrogen

Jarvis, S. C. and Hatch, D. J. 1985. The effects of aluminiumon the growth of white clover dependent upon fixation of atmosphericnitrogen.–J. exp. Bot. 36: 1075–1086. The effects of aluminium (Al^{3 +}) on the growth of white cloverdependent upon symbiotic fixation of atmospheric nitrogen wereexamined at concentrations that may be encountered in solutionsfrom soils of low pH. Well established plants were grown inflowing solution culture with carefully maintained concentrationsof Al and of P and with pH held constant at a value (4.5) atwhich insoluble precipitates are unlikely to form. After 3 weeksof treatment, there were major visual differences between treatmentsin both shoots and roots. Although added Al at 25, 50 and 100mmol m^–3 reduced dry weight, the differences between theplants were not significant. There were, however, some considerabledifferences in P and Ca contents between the treatments. Muchof the Al held by the roots was displaced when plants were transferredto solutions containing either scandium or gallium. As wellas the effects on the plant, Al had a considerable influenceon the fixation of atmospheric nitrogen. At concentrations of50 and 100 mmol m^–3 Al³⁺ nodule initiation was reducedand there was a much reduced nodule nitrogenase activity perunit of root. Even at 25 mmol m^–3 Al³⁺, when nodule numberswere not reduced, nitrogenase activity was adversely affected. Key words: —Aluminium, Trifolium repens, nitrogen fixation     

Effectiveness of Lotus Root Nodules: III. EFFECT OF COMBINED NITROGEN ON NODULE EFFECTIVENESS AND FLAVOLAN SYNTHESIS IN PLANT ROOTS

When grown in a nutrient solution containing combined nitrogen(NH₄NO₃), Lotus pedunculatus and L. tenuis seedlings inoculatedwith a fast-growing strain of Rhizoblum (NZP2037) did neitherdevelop root nodules nor develop flavolans in their roots. Incontrast, the roots of nodulated seedlings growing in a nitrogen-freenutrient solution contained flavolans. Flavolan synthesis coincidedwith root nodule development on these plants. When added as a single dose, high concentrations of NH₄NO₃ (5and 10 mg N per plant) stimulated the growth of L. pedunculatusplants but suppressed nodulation and nitrogen fixation. In contrastthe continued supply of a low concentration of NH₄NO₃ (1?0 mgN d^–1 per plant) stimulated nitrogen fixation by up to500%. This large increase in nitrogen fixation was associatedwith a large increase in nodule fresh weight per plant, a doublingof nodule nitrogenase activity, and a lowering of the flavolancontent of the plant roots. The close relationship between nitrogendeficiency, nodule development, and flavolan synthesis in L.pedunculatus meant that it was not possible (by nitrogen pretreatmentof plants) to alter the ineffective nodule response of a Rhizobiumstrain (NZP2213) sensitive to the flavolan present in the rootsof this plant.     

Cinnamate toxicity expression on phenylalanine ammonia-lyase activity,germination and growth of cucumber (Cucumis sativis) seedlings

Low pH (5.2) decreased nodule number and acetylene reduction. Aluminium further depressed those parameters in theRhizobium leguminosarum-Pisum sativum associations examined. In the Al-treated plants nodule formation by strains 128C53 and 128C30 was not affected by 3 or 15 and 30 or 60 μM Al, respectively, as compared with the number of nodules on plants grown at pH 5.2 in the absence of Al. However, improved nodulation rates by those strains did not enhance plant dry weight or reduced nitrogen content. No differences in nitrogenase activity were found among strains of nodulating plants grown at the same aluminium level. These results suggest that Al-ions affected specifically nitrogenase activity and that this effect was primarily responsible for the reduction in plant growth.     

Effects of solution pH and bicarbonate on the growth and nodulation of a range of grain legume species

Defining plant adaptation to soil conditions is critical for the successful introduction of grain legume species into farming systems. This nutrient solution study examined the effects of pH (4, 5, 6, 7 and 8) and bicarbonate (5 mM KHCO₃) on the growth and nodulation of 14 grain legume species supplied with N or reliant on N₂-fixation. Species includedPisum sativum L.,Cicer arietinum L.,Lens culinaris Med., and a range ofLupinus, Vicia andLathyrus species. Species differed greatly in response to solution pH. For both N-fertilized and N₂-fixing plants, shoot growth ofL. culinaris was very sensitive to low pH (pH < 7), whereas shoot growth ofLupinus angustifolius L. andLupinus albus L. was sensitive to higher pH (pH ≥ 6). Other species had a broader optimal pH range for growth when supplied with N, but were generally sensitive to low pH (pH < 7 forC. arietinum andVicia sativa L., pH < 6 forP. sativum, Vicia faba L.,Lathyrus sativus L. andLathyrus cicera L., and pH < 5 forVicia benghalensis L. andVicia narbonensis L.) when reliant on N₂-fixation. For these other species, symbiotic N₂-fixation appeared to be more sensitive than host plant growth to low pH. This finding was supported by lower nodule numbers and mass, and lower N concentrations in shoots of sensitive species at low pH relative to higher pH. ForL. culinaris, nodule numbers and mass were relatively unaffected by pH 5–8, N concentrations in shoots were high at low pH and plants developed symptoms relating to H⁺ toxicity at pH as high as 7. These results indicate that host plant growth ofL. culinaris is more sensitive to low pH than theRhizobium symbiosis. ForL. albus andL. angustifolius, both host plant growth and symbiotic N₂-fixation appeared to be equally sensitive to pH ≥ 6.Lupinus pilosus Murr. was more tolerant of high pH than the otherLupinus species. At pH 4, two genotypes ofC. arietinum had better early nodulation than other species.Vicia ervilia L. nodulated poorly at all levels of solution pH, indicating that the commercial Group E inoculum (Rhizobium leguminosurum bv.viceae SU303) may not be effective for this species in solution culture. Addition of bicarbonate decreased shoot growth, nodulation and N concentrations in shoots of most species. Early nodulation (nodule number) ofLathyrus ochrus (L.) DC was not affected by the bicarbonate treatment.     

Stimulation of Nodulation in the Clover-Rhizobium trifolii 0403 System by Penicillin and Mecillinam

The number of nodules produced per clover seedling inoculated with Rhizobium trifolii 0403 can be increased almost 2-fold by the addition of penicillin or mecillinam. Two-day-old dutch white clover seedlings grown in 250 milliliter boston round jars containing agar-solidified plant growth medium were inoculated with exponentially growing Rhizobium trifolii 0403 cells. Penicillin or mecillinam (100 micrograms per milliliter) were added immediately or after 24 hours. Following 42 days growth, 10 replicate sets of 5 plants for each treatment were assayed for nodule number, plant dry weight, and Kjeldahl nitrogen. Both antibiotics increased nodule number, plant dry weight, and Kjeldahl nitrogen. Increases in nodule number and dry weight were statistically significant. The range of values in Kjeldahl nitrogen was so extensive as to make the data insignificant at the P < 0.05 level, however nodule number, plant dry weight, and Kjeldahl nitrogen displayed a significant correlation with each other. There were no significant differences in treatment with either antibiotic or with time of treatment. Nodule number increased by about 85%, and plant dry weight and nitrogen increased by about 30%.     

Effect of NO3 on N2 fixation and nitrogenous solutes of xylem in two nodulated West African geocarpic legumes,Kersting's bean (Macrotyloma geocarpum L.) and Bambara groundnut (Vigna subterranea L.)

Nodulation, nitrogen (N₂) fixation and xylem sap composition were examined in sand cultured plants of Bambara groundnut (Vigna subterranea L.) and Kersting's bean (Macrotyloma geocarpum L.) inoculated with Bradyrhizobium strain CB756 and supplied via the roots for a 4 week period from the third week onwards with different levels of (¹⁵N)-nitrate (0–15 mM). The separate contributions of nitrate and N₂ to plant nitrogen were measured by isotope dilution. Increasing levels of nitrate inhibited nodule growth (measured as dry matter or nodule N) of both species parallel with decreased dependence on symbiotically-fixed N. Specific nodule activity (N₂ fixed g nodule dry⁻¹ d⁻¹ of nodules) was reduced progressively with time in V. subterranea at higher (5 or 15 mM) levels of NO₃, but this was not so for M. geocarpum. Root xylem bleeding sap of both species showed ureides (allantoin and allantoic acid) as predominant (>90%) solutes of nitrogen when plants were relying solely on atmospheric N. Levels of ureide and glutamine decreased and those of asparagine and nitrate in xylem increased with increasing level of applied nitrate. Relative levels of xylem ureide-N were positively correlated (R²=0.842 for M. geocarpum and 0.556 for V. subterranea), and the ratio of asparagine to glutamine in xylem exudate negatively correlated (R²=0.955 for M. geocarpum and 0.736 for V. subterranea) with plant reliance on nitrogen fixation. The data indicate that xylem sap analyses might be useful for indirect field assays of nitrogen fixation by the species and that Kersting's bean might offer some potential as a symbiosis in which N₂ fixation is relatively tolerant of soil N.     

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     

Plant-endophyte symbiosis in non-leguminous plants

Summary A wide taxonomic range of non-leguminous dicotyledonous plants bear root nodules and are able to fix atmospheric nitrogen. These plants belong to the orders Casuarinales, Myricales, Fagales, Rhamnales, Coriariales, and Rosales. Actinomycetes are involved in the root-nodule symbiosis. Nitrogen fixation is inhibited by hydrogen and carbon monoxide. Combined nitrogen depress nodule formation, but nitrogen fixation still occurs in the presence of combined nitrogen in the medium. In nitrogen-free medium Alnus plants fix in one season of 48 weeks 500 mg N per plant and Ceanothus plants 760 mg N per plant. Fixation by the other plant species is about of the same order. Field estimates showed that the nitrogen increase of the soil was about 61.5–157 kg N per ha per annum, depending on the age of the trees, under Alnus, 58.5 kg N per ha per annum under Casuarina, about 60 kg N per ha per annum under Ceanothus, 27–179 kg N per ha per annum underHippopha? rhamnoides, and about 61.5 kg N per ha per annum underDryas drummondii with someShepherdia spp. Non-leguminous root nodules belong to two types: coralloid root nodules and root nodules where the apex of each nodule lobe produces a negatively geotropic root. The primary infection occurs through the root hairs where a curling effect is observed. In the host cells the endophyte presents itself in three forms: hyphae, vesicles and bacteria-like cells. Vesicles are probably associated with nitrogen fixation, whereas the bacteria-like cells function in the endophyte's survival and dispersal. The endophyte is an obligate symbiont. TheAlnus glutinosa endophyte has been isolated and grownin vitro in root-nodule callus tissue. However, the isolated endophyte produces only ineffective root nodules in re-inoculation tests.     

Nodulation and symbiotic nitrogen fixation of cowpea (Vigna unguiculata (L.) Walp)

Summary Rhizobium strains CIAT 301, CIAT 79 and SLM 602 were tested and found effective in the nodulation and nitrogen fixation of cowpea cv. MI-35 (Vigna unguiculata (L.) Walp) plants in growth chamber experiments. Fresh weight of nodules increased with plant age initially and stabilized in 20–30 days from planting, followed by a secondary flush of nodule growth after 30 days. Apparent nitrogen fixation per gram nodule fresh weight reached a maximum in 20–30 days after planting and then decreased, even though a flush of new nodules was produced.     

Preincubation of B. japonicum cells with genistein reduces the inhibitory effects of mineral nitrogen on soybean nodulation and nitrogen fixation under field conditions

Genistein is the major root produced isoflavonoid inducer of nod genes in the symbiosis between B. japonicum and soybean plants. Reduction in the isoflavonoid content of the host plants has recently been suggested as a possible explanation for the inhibition of mineral nitrogen (N) on the establishment of the symbiosis. In order to determine whether genistein addition could overcome this inhibition, we incubated B. japonicum cells (strain 532C) with genistein. Mineral N (in the form of NH₄NO₃) was applied at 0, 20 and 100 kg ha^-1. The experiments were conducted on both a sandy-loam soil and a clay-loam soil. Preincubation of B. japonicum cells with genistein increased soybean nodule number and nodule weight, especially in the low-N-containing sandy-loam soil and the low N fertilizer treatment. Plant growth and yield were less affected by genistein preincubation treatments than nitrogen assimilation. Total plant nitrogen content was increased by the two genistein preincubation treatments at the early flowering stage. At maturity, shoot and total plant nitrogen contents were increased by the 40 μM genistein preincubation treatment at the sandy-loam soil site. Total nitrogen contents were increased by the 20 μM genistein preincubation treatment only at the 0 and 20 kg ha^-1 nitrate levels in clay-loam soil. Forty μM genistein preincubation treatment increased soybean yield on the sandy-loam soil. There was no difference among treatments for 100-seed weight. The results suggest that preincubation of B. japonicum cells with genistein could improve soybean nodulation and nitrogen fixation, and at least partially overcome the inhibition of mineral nitrogen on soybean nodulation and nitrogen fixation. This revised version was published online in June 2006 with corrections to the Cover Date.     

Influence of 5-Methyltryptophan-Resistant Bradyrhizobium japonicum on Soybean Root Nodule Indole-3-Acetic Acid Content

Bradyrhizobium japonicum mutants resistant to 5-methyltryptophan were isolated. Some of these mutants were found to accumulate indole-3-acetic acid (IAA) and tryptophan in culture. In greenhouse studies, nodules from control plants inoculated with wild-type bradyrhizobia contained 0.04, 0.10, and 0.58 μg of free, ester-linked, and peptidyl IAA g (fresh weight) of nodules⁻¹, respectively. Nodules from plants inoculated with 5-methyltryptophan-resistant bradyrhizobia contained 0.94, 1.30, and 10.6 μg of free, ester-linked, and peptidyl IAA g (fresh weight) of nodules⁻¹, respectively. This manyfold increase in nodule IAA content indicates that the Bradyrhizobium inoculum can have a considerable influence on the endogenous IAA level of the nodule. Further, these data imply that much of the IAA that accumulated in the high-IAA-containing nodules was of bacterial rather than plant origin. These high-IAA-producing 5-methyltryptophan-resistant bacteria were poor symbiotic nitrogen fixers. Plants inoculated with these bacteria had a lower nodule mass and fixed less nitrogen per gram of nodule than did plants inoculated with wild-type bacteria.     

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     

Quantification of N2-fixation and survival of inoculated diazotrophs associated with roots of Kallar grass

White clover plants were grown for 97 days under two temperature regimes (20/15°C and 8/5°C day/night temperatures) and were supplied with either small amounts (a total of 80 mg N pot^–1) of ammonium (NH ₄ ⁺ ) or nitrate (NO ₃ ^– ) nitrogen, or received no mineral N and relied on N₂ fixation. Greatest growth and total leaf area of clover plants occurred in N₂ fixing and NO ₃ ^– -fed plants grown at 20/15°C and poorest growth occurred in NH ₄ ⁺ -fed plants grown at 8/5°C. Nodule mass per plant was greater at 8/5°C due to increased nodule numbers rather than increased dry weight per nodule. This compensated to some extent for the reduced N₂-fixing activity per unit dry weight of nodule tissue found at the low growth temperature up to 116 d after sowing, but thereafter both activity per nodule dry weight and activity per plant were greater at the low temperature. Highest nitrate reductase activity (NRA) per g fresh weight and total activity per leaf, petiole or root occurred in NO ₃ ^– -fed plants at 8/5°C. Low growth temperature resulted in a greater partitioning of total plant NRA to the roots of NO ₃ ^– -fed plants. The results are considered in relation to the use of N fertiliser in the spring under field conditions.     

Symbiotic dinitrogen fixation and host-plant growth during development of and recovery from phosphorus deficiency

Characterization of nodule growth and function, phosphorus and nitrogen status of plant tissues and host-plant growth of nodulated soybean ( Glycine max L. Merr.) plants developing and recovering from phosphorus deficiency was used to evaluate the role of phosphorus in symbiotic dinitrogen fixation. The sequence of physiological responses during recovery from phosphorus deficiency was; (1) rapid uptake of phosphorus, (2) rapid increases in the phosphorus concentration of leaves and nodules, (3) enhanced growth and function of nodules, (4) increased nitrogen concentrations in all plant organs and (5) enhanced plant growth. The sequence of physiological responses to onset of phosphorus deficiency was; (1) decreased phosphorus uptake, (2) decreased phosphorus concentrations in leaves and nodules, (3) decreased nodule function, (4) decreased nitrogen concentration in plant organs and (5) decreased plant growth. These results, in conjunction with previously published data (Sa and Israel, Plant Physiol. 97: 928–935, 1991), support an interpretation that the total response of symbiotic dinitrogen fixation in soybean plants to altered phosphorus supply is a function of both indirect effects on host-plant growth and more direct effects on the metabolic function of nodules.     

Effects of estrone and 17 β-estradiol on vegetative growth of Medicago sativa

Alfalfa ( Medicago sativa L.) plants were grown in the absence or presence of the steroidal estrogens, estrone and 17β-estradiol, under varying conditions. Plants were analysed for the following parameters: plant weight, estrogen content, phytoestrogen content, degree of nodulation and nitrogen fixation activity. It was found that under controlled greenhouse conditions: (1) Treatment with estrogens in the range of 0.005 to 0.5 μg 1⁻¹ increases both shoot and root dry weitht. In contrast, estrogen in concentrations of 50 to 500 μg 1⁻¹ decreases plant growth. (2) The effect of estrogen of growth is most marked in the absence of nitrogen. (3) Estrone is more effective in increasing growth than 17 β-estradiol. (4) In the plants where estrogen induced growth there was no significant increase in nitrogen fixation activity and nodule number. (5) Endogenous estrogen content of the plant did not increase at concentrations (0.005-0.5 μg 1⁻¹) which increased vegetative growth. (6) Endogenous estrogen content of the plant did increase at concentrations of estrogen (50-500 μg 1⁻¹ which inhibited vegetative growth and nodule weight. It can be concluded that estrogen in concentrations found in sewage water (0.3 μg estrogen 1⁻¹) can affect the vegetative growth of alfalfa plants.     

Diurnal Functioning of the Legume Root Nodule

Diurnal changes in plant and nodule performance were studiedin 28–9 d plants of Pisum sativum L. in two environments,both with a 12 h (27 000 lx):12 h::light:dark cycle, but one(A) with a fluctuating temperature-humidity regime (photoperiod18 ?C, 60 per cent relative humidity:night 12 ?C, 85 per cent),the other (B) with constant temperature (18 ?C) and humidity(75 per cent). Fixation rate (C₂H₂ reduction), respiratory output of the nodulatedroot, and nodule sugar level increased throughout the photoperiod,whereas nodule soluble nitrogen level declined steadily. Reversalof these trends in the night period led, at its end, to minimain fixation rate, sugar level and respiration, but a maximumin soluble nitrogen. The A environment produced the greaterday:night fluctuations in transpiration and nodule soluble nitrogen,but B, with its higher night temperature, induced the more pronounceddecrease in fixation at night. Slightly less nitrogen was fixed during the photoperiod thanduring the night in the A environment, yet since some fixationproducts were retained in the nodules at night and not releaseduntil the next photoperiod, the day: night difference in nitrogenexport from nodules was 1.8:1. The photoperiod of A was alsoa time of higher nodule respiration and replenishment of nodulesugar and starch, so that the nodules' requirement for translocatedcarbohydrate was more than twice that at night. Humidity decrease in the photoperiod (of A) elicited higherrates of transpiration and a more rapid than normal emptyingof soluble nitrogen from the nodules: elevation of humidityhad the opposite effects. Shoot removal (A-grown plants) causednodule sugar levels to fall rapidly below those normally encounteredin intact plants.     

The Influence of the Plant Growth-regulator, 2-Methyl-4-Chlorophenoxyacetic Acid, on the Metabolism of Carbohydrate, Nitrogen and Minerals in Solanum lycopersicum (Tomato)

The effects of the foliar application of phytocidal concentrationsof 2-methyl-4-chlorophenoxyacetic acid (MCPA) on change in totaldry weight, and in ‘available carbohydrate’ (starch,‘total’ and ‘reducing’ sugars), totalnitrogen, phosphorus, potassium, calcium, and magnesium of ‘tops’and roots of tomato plants have been followed over a periodof 14 days following spraying. There were two main treatments—‘nutrient’(nutrient supply to roots continued after spraying) and ‘water’(distilled water only supplied to roots after spraying) and‘water’ (distilled water only supplied to rootsafter spraying)—the sub-treatments consisting of ‘MCPA’versus ‘no-MCPA’ for each of the main treatments.Twelve different times of sampling were used. In analysing the present data, the quantity ‘residualdry weight’ (total dry weight less ‘available carbohydrate’),which was originally introduced by Mason and Maskell as a basisof reference for analyses of plant organs in short-period experimentsnot involving appreciable growth, has been used as an estimateof the permanent structure of plant growth. This new use ofthe ‘residual dry weight’ basis has brought outimportant features which were obscured when the data were leftin their primary form (as percentages of total dry weight oramounts per plant). Growth, as measured by increase in ‘residual dry weight’,was greatly inhibited by 2-methyl-4-chlorophenoxyacetic acidshortly after spraying, in both the presence and the absenceof nutrient. In the presence of 2-methyl-4-chlorophenoxyacetic acid, netassimilation rate (estimated as rate of increase in total dryweight per gram ‘residual dry weight’ of the ‘tops’)was greatly diminished while uptake of total nitrogen and ofP₂O₅ (estimated as increase in total nitrogen or of P₂O₅ ofthe whole plant per day per 1 g. ‘residual dry weight’of the roots) appeared to undergo a similar but much smallerdiminution. It seemed probable, however, that in the presenceof MCPA a larger proportion of the carbohydrate actually formedwas utilized for synthesis of aminoacids and protein. In the plant as a whole there was no evidence of actual depletionof ‘available carbohydrate’ as a result of MCPAtreatment, this fraction showing a steady increase in all treatmentsthroughout the experiment. The rate of increase was, however,much reduced by MCPA treatment. The ‘tops’ presentedmuch the same picture as the whole plant, but for the rootsthe situation was quite different. While the roots of the ‘no-MCPA’plants and also of the ‘MCPA-water’ plants showeda steady increase in available carbohydrate, those of the ‘MCPA-nutrient’plants rose only very slightly (from the initial value of 8mg. per plant to about 10 mg.) during the first 2 days, andthen in the next 2 days declined to a value (about 6 mg.) belowthe initial and remained at this low level for the rest of theexperiment. It is suggested that the phytocidal effect of 2-methyl-4-chlorophenoxyaceticacid in the presence of nutrient may be due to depletion ofthe ‘available carbohydrate’ supplies in the roots,which is shown to be brought about, in part, by reduced transportfrom the tops, and partly by the relatively greater utilizationof the carbohydrate present. These results offer an explanationfor the facts that plants showing vigorous growth are more easilykilled by MCPA and that perennial plants, particularly thosewith storage tissues in their roots, are more resistant. Further,they suggest the useful practical application that MCPA treatmentshould be given when the carbohydrate reserves of the rootsare at a minimum. For perennial plants, conditions might beexpected to be optimal for the application of MCPA in late spring,at a time when the first ‘flush’ of growth is slowingdown and before any appreciable new reserves of carbohydratehave been accumulated. It was also shown that 2-methyl-4-chlorophenoxyacetic acid preventedthe net synthesis of starch, but still permitted an appreciablenet formation of sucrose. 2-methyl-4-chlorophenoxyacetic acid appeared to have no effecton the uptake of potassium, calcium, or of magnesium. The lackof effect on potassium is contrasted with the previous observationby Rhodes, Templeman, and Thruston (1950) that sub-lethal concentrationsof MCPA, applied over a relatively long period to the rootsof tomato plants, specifically depressed the uptake of potassium.     

Alleviation of Salt Stress in Common Bean (Phaseolus vulgaris) by Exogenous Abscisic Acid Supply

In this work the effect of abscisic acid (ABA) and 100 mM NaCl on common bean (Phaseolus vulgaris var. Coco) growth, nitrogenase activity, and nodule metabolism was studied. Experiments were carried out in a controlled environmental chamber and plants, at the vegetative growth stage (16 days old), were treated with ABA (1 μM and 10 μM) and 48 h later were exposed to saline treatment. Results revealed that plant dry weight, nodule dry weight, nitrogen fixation (acetylene reduction activity and ureides content), and most enzymes of ammonium and ureides metabolism were affected by both ABA and NaCl. The addition of 1 μM ABA to the nutrient solution before the exposure to salt stress reduced the negative effect of NaCl. Based on our results, we suggest that ABA application improves the response of Phaseolus vulgaris symbiosis under saline stress conditions, including the nitrogen fixation process and enzymes of ammonium assimilation and purine catabolism.