Economy of Photosynthate Use in Nitrogen-fixing Legume Nodules: Observations on Two Contrasting Symbioses

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 ¹⁴CO₂ 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.     

Metabolism of carbon and nitrogen by soybean seedlings in response to vegetative apex removal

Short-term (31-hour diurnal) growth-chamber studies were conducted to determine the effects of removing the vegetative apex (meristem and developing trifoliolate leaves) on net photosynthesis (changes in plant dry weight), on distribution of metabolites among plant parts, and on nitrate metabolism and reduced-N accumulation by soybean [Glycine max (L.) Merr.] seedlings. Roots and stems served as alternate sinks for dry matter accumulation in the absence of the vegetative apex. Sugar concentration in roots increased (42%) within 4 hours of vegetative apex removal, and remained higher than for the controls during the 31-hour experimental period. Nitrate assimilation (nitrate reductase activity and total accumulation of reduced-N) was also enhanced in response to vegetative apex removal. Although dry matter accumulation was similar between treated and control plants (113 versus 116 milligrams per plant) over the 31-hour sampling period, more nitrate (1.31 versus 0.79 milligrams per plant) and more reduced-N (3.96 versus 3.45 milligrams per plant) accumulated in treated plants during the same interval. It was concluded that vegetative apex removal had little effect on overall net photosynthesis of soybean seedlings during the 31-hour treatment period, but did alter partitioning of photosynthate and enhanced uptake, transport, and reduction of nitrate. Implications are that uptake and metabolism of nitrate by soybeans may be limited by flux of carbohydrate to the roots, although hormonal effects due to vegetative apex removal cannot be ruled out.     

Effects of nitrogen dioxide and nitrate nutrition on nodulation, nitrogenase activity, growth, and nitrogen content of bean

The influence of nutrient nitrate level (0-20 millimolar) on the effects of NO₂ (0-0.5 parts per million) on nodulation and in vivo acetylene reduction activity of the roots and on growth and nitrate and Kjeldahl N concentration in shoots was studied in bean (Phaseolus vulgaris L. cv Kinghorn Wax) plants. Exposing 8-day old seedlings for 6 hours each day, for 15 days, to 0.02 to 0.5 parts per million NO₂ decreased total nodule weight at 0 and 1 millimolar nitrate, and nitrogenase (acetylene reduction) activity at all concentrations of nitrate. The pollutant had little effect on root fresh or dry weights. Shoot growth was inhibited by NO₂. The NO₂ exposure increased nitrate concentration in roots only at 20 millimolar nutrient nitrate. Exposure to NO₂ markedly increased Kjeldahl N concentration in roots but generally decreased that in shoots. The experiments demonstrated that nutrient N level and NO₂ concentration act jointly in affecting nodulation and N fixing capability, plant growth and composition, and root/shoot relationships of bean plants.     

Effect of phosphorus and zinc nutrition on soybean seed phytic Acid and zinc

The relationships between nutrient P and Zn levels and the phytic acid, P, and Zn concentrations in soybean (Glycine max L. Merr. cv `Williams 79') seed were studied. Phytic acid increased linearly from 4.2 to 19.2 milligrams per gram as nutrient P treatment was varied from 2.0 to 50 milligrams per liter and Zn was held constant at 0.05 milligrams per liter. Leaf P concentration during seed development was found to be closely related to the concentrations of seed P and phytic acid. Leaf and seed Zn concentrations both responded positively to increasing nutrient Zn treatment. The effects of P treatment on plant and seed P and phytic acid were largely independent of the effects of Zn treatment on leaf and seed Zn. Phytic acid to Zn molar ratios ranging from 3.6 to 33.8 were observed.

The effects of nutrient P treatments on the concentrations of phytic acid, seed P, and leaf P were also studied in the P-sensitive (gene np) cultivars `Harosoy' and `Clark' and their respective P-tolerant (gene Np) near-isogenic lines L66-704 and L63-1677. In general, the positive relationships observed among nutrient P, leaf P, seed P, and phytic acid concentrations were similar to those observed in the studies with Williams 79. When fertilized with low or moderate nutrient P (2.5 and 25.0 milligrams P per liter, respectively) no significant differences in any parameter were observed between Harosoy or Clark and their respective P-tolerant isolines. When fertilized with high nutrient P (100 milligrams P per liter), Harosoy seed had a significantly higher concentration of phytic acid (30 milligrams per gram) than did seed of its P-tolerant near-isogenic line L66-704 (24.2 milligrams per gram phytic acid), whereas no significant difference was observed between Clark and its P-tolerant near-isogenic line L63-1677 (22.8 and 21.6 milligrams per gram, respectively). Variation in the phytic acid concentrations in the mature seed of the cultivars and isolines more closely paralleled leaf P concentrations observed during seed development (49 days after flowering), than those observed at the onset of seed development (14 days after flowering). Electrophoresis and ion-exchange chromatography revealed that partially phosphorylated intermediates do not appear when phytic acid accumulation is greatly reduced by limiting the nutrient P or when accumulation is greatly accelerated by excess P.

     

Increases in Phosphorus Requirements for CO(2)-Enriched Pine Species

Pinus radiata D. Don (half-sib families 20010 and 20062) and Pinus caribaea var hondurensis (an open-pollinated family) were grown for 49 weeks at seven levels of phosphorus and at CO₂ concentrations of either 340 or 660 microliters per liter, to establish if the phosphorus requirements differed between the CO₂ concentrations and if mycorrhizal associations were affected. When soil phosphorus availability was low, phosphorus uptake was increased by elevated CO₂. This may have been related to changes in mycorrhizal competition. When the phosphorus concentration in the youngest fully expanded needles was above 600 milligrams per kilogram the shoot weight of all pine families was greater at high CO₂ due to increases in rates of photosynthesis. More dry weight was partitioned to the stems of P. radiata family 20010 and P. caribaea. At foliar phosphorus concentrations above 1000 milligrams per kilogram (P. radiata) and 700 milligrams per kilogram (P. caribaea), growth did not increase at 340 microliters of CO₂ per liter. Soluble sugar levels in the same needles mirrored the growth response, but the starch concentration declined with increasing phosphorus. At 660 microliters of CO₂ per liter, shoot weight and soluble sugar concentrations were still increasing up to a foliar P concentration of 1800 milligrams per kilogram for P. radiata and 1600 milligrams per kilogram for P. caribaea. The starch concentrations did not decline. These results indicate that higher foliar phosphorus concentrations are required to realize the maximum growth potential of pines at elevated CO₂.     

Effect of Contrasting Patterns of Nitrate Application on the Nitrate Uptake, N2-Fixation, Nodulation and Growth of White Clover

White clover (Trifolium repens L.) plants were grown from seedin perlite, inoculated with effective rhizobia and exposed tothe same ‘concentration x days’ of ¹⁵N-labellednitrate in four contrasting patterns of doses. Acetylene reductionwas measured at intervals using an open, continuous-flow sytem.Mean dry weight per nodule and rates of acetylene reductionfell rapidly (2–3 d) during periods of exposure to highnitrate concentrations (> 7 mM N) and rose again, equallyrapidly, when nitrate was withdrawn or substantially reduced.The fall in mean dry weight per nodule (50–66 per cent)was almost certainly too large to be accounted for by loss ofsoluble or storage carbohydrate only. No new nodules were formedduring periods of high nitrate availability. When nitrate wassupplied continuously at a moderate concentration (5.7 mM N)nodule numbers stabilised although existing nodules increasedin dry weight by almost four-fold over the 30 d measurementperiod. Treatment had no effect on the percentage nitrogen in planttissues although there were large differences in the proportionsderived from nitrate and N₂-fixation. Plants exposed continuouslyor frequently to small doses of nitrate took up more nitrate,and hence relied less heavily on N₂-fixation, than those exposedto larger doses less often. Increased reliance on nitrate broughtwith it increased total dry weight and shoot: root ratios. Possiblemechanisms involved in bringing about these differences in nitrogennutrition and growth are discussed. White clover, Trifolium repens, nitrate, N₂-fixation, nodule, acetylene reduction, ¹⁵N     

Low concentrations of nitrate and ammonium stimulate nodulation and N2 fixation while inhibiting specific nodulation (nodule DW g−1 root dry weight) and specific N2 fixation (N2 fixed g−1 root dry weight) in soybean

Nitrate N is a major inhibitor of the soybean/Bradyrhizobium symbiosis in legumes and although this inhibition has been studied for many years, as yet no consensus has been reached on the specific and quantitative interactions between nitrate and ammonium supply and N₂ fixation. The effect of nitrate and ammonium supply on plant growth, nodulation and N₂ fixation capacity during the full growth cycle was investigated in both greenhouse and growth chamber experiments with three soybean genotypes. The results show that a high concentration of mineral N (10 mM), either as nitrate or ammonium or ammonium nitrate significantly suppressed nodule number, nodule dry weight and total N₂ fixed per plant of nodulated soybeans. However, lower mineral N concentrations, either 1 mM or 3.75 mM significantly enhanced nodule number, nodule dry weight and total N₂ fixed per plant, while specific nodulation (nodule dry weight g^–1 root DW, SNOD) and specific N₂ fixation (total N₂ fixed g^–1 root DW, SNF) were significantly reduced, particularly at the early vegetative growth stage V4, compared to the treatment with N₂ fixation as the only N source, in both growth chamber and greenhouse experiments. Therefore, we suggest that SNOD or SNF might be better indicators to express the suppressing effect of mineral N addition on nodule performance and N₂ fixed. Our studies also showed that ammonium alone was the more efficient N source than either ammonium nitrate or nitrate for soybean, as it resulted in higher biomass accumulation, nodule dry weight, total N accumulation and total N₂ fixed by 23, 20, 18 and 44%, respectively, compared to NO₃ ^– as the N source.     

Nitrate inhibition of legume nodule growth and activity : I. Long term studies with a continuous supply of nitrate

The synthesis and accumulation of nitrite has been suggested as a causative factor in the inhibition of legume nodules supplied with nitrate. Plants were grown in sand culture with a moderate level of nitrate (2.1 to 6.4 millimolar) supplied continuously from seed germination to 30 to 50 days after planting. In a comparison of nitrate treatments, a highly significant negative correlation between nitrite concentration in soybean (Glycine max [L.] Merr.) nodules and nodule fresh weight per shoot dry weight was found even when bacteroids lacked nitrate reductase (NR). However, in a comparison of two Rhizobium japonicum strains, there was only 12% as much nitrite in nodules formed by NR⁻R. japonicum as in nodules formed by NR⁺R. japonicum, and growth and acetylene reduction activity of both types of nodules was about equally inhibited. In a comparison of eight other NR⁺ and NR⁻R. japonicum strains, and a comparison of G. max, Phaseolus vulgaris, and Pisum sativum, the concentration of nitrite in nodules was unrelated to nodule weight per plant or to specific acetylene reduction activity. The very small concentration of nitrite found in P. vulgaris nodules (0.05 micrograms NO₂⁻-N per gram fresh weight) was probably below that required for the inhibition of nitrogenase based on published in vitro experiments, and yet the specific acetylene reduction activity was inhibited 83% by nitrate. The overall results do not support the idea that nitrite plays a role in the inhibition of nodule growth and nitrogenase activity by nitrate.     

Nitrogen Nutrition of Nodules in Relation to ;N-Hunger' in Cowpea (Vigna unguiculata L. Walp)

Early growth, nodule development, and nitrogen fixation by two cultivars of cowpea (Vigna unguiculata L. Walp), one large-seeded (Vita 3; 146.0 ± 0.9 milligrams seed dry weight, 4.1 ± 0.2 milligrams seed N), the other small-seeded (Caloona; 57.5 ± 2.5 milligrams seed dry weight, 1.8 ± 0.1 milligrams seed N), were compared under conditions of sand culture with nutrient solution free of combined N. The seed stocks used had been obtained from plants uniformly labeled with ¹⁵N, thus enabling changes with time in distribution of cotyledon and fixed N among plant parts to be measured by isotope dilution. Caloona, but not Vita 3, showed physiological symptoms of `N hunger,' i.e. transient loss of chlorophyll (visible yellowing) and N from the first-formed unifoliolate leaves at or around the onset of symbiotic functioning and N<sub>2</sub> fixation. The smaller-seeded Caloona showed higher early nitrogenase activity than the larger-seeded Vita 3 and by 28 days had fixed 6.6 milligrams of N per milligram of seed N [mg N · (mg seed N)<sup>−1</sup>] versus only 3.5 mg N · (mg seed N)<sup>−1</sup> in Vita 3. Both cultivars lost around 30% of their initial seed N at germination, mostly as fallen cotyledons. Abscised cotyledons of Caloona contained 1.21 ± 0.17% N; those of Vita 3 contained 2.61 ± 0.37% N. When compared on the basis of cotyledon N available for seedling growth, Caloona was shown to have fixed 10.6 mg N · (mg seed N)<sup>−1</sup> and Vita 3 only 5.3 mg N · (mg seed N)<sup>−1</sup>. Most of the cotyledon N withdrawn from the unifoliolate leaf pair of Caloona during `N-hunger' was committed to early nodule growth and, in total, 20 to 25% of the cotyledon N resource of this cultivar was ultimately invested in establishment of symbiosis compared with only 7% in Vita 3.     

Interactions between Nitrogen Fixation, Mycorrhizal Colonization, and Host-Plant Growth in the Phaseolus-Rhizobium-Glomus Symbiosis

Bean (Phaseolus vulgaris L. cv. Dwarf) roots were inoculated with Rhizobium phaseoli and colonized by the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus fasciculatum Gerd. and Trappe or left uncolonized as controls. The symbiotic associations were grown in an inert substrate using 0, 25, 50, 100, or 200 milligrams hydroxyapatite (HAP) (Ca₁₀[PO₄]₆[OH]₂) per pot as a P amendment. Plant and nodule dry weights and nodule activity increased for both VAM and control plants with increasing P availability, but values for VAM plants were significantly lower in all parameters than for controls. Inhibition of growth and of N₂ fixation in VAM plants was greatest at the lowest and highest P regimes. It was smallest at 50 milligrams HAP, where available P at harvest (7 weeks after planting) was 5 micrograms P per gram substrate. At this level of P availability, the association apparently benefited from increased P uptake by the fungal endophyte. Percent P values for shoots, roots, and nodules did not differ significantly (p > 0.05) between VAM and control plants. The extent of colonization, fungal biomass, and the fungus/association dry weight ratio increased several fold as HAP was increased from 0 to 200 milligrams. It is concluded that intersymbiont competition for P and photosynthate was the primary cause for the inhibition of growth, nodulation, and nodule activity in VAM plants. Impaired N₂ fixation resulted in N stress which contributed to inhibition of host plant growth at all levels of P availability.     

Effect of Ammonium Ions on Egg Hatching and Second-Stage Juveniles of Meloidogyne incognita in Axenic Tomato Root Culture

Eggs, either dispersed or in masses, and second-stage juveniles (J2) of Meloidogyne incognita were exposed to different concentrations of ammonium ions in a nutrient agar medium upon which excised tomato roots were growing. Egg hatch and J2 penetration of the roots was slowed or inhibited at high (54 and 324 mg/liter) but not at low (1.5 and 9 mg/liter) concentrations of ammonium nitrate. The effect of ammonium on J2 appeared to be temporary and reversible. High potassium nitrate concentration (1,116 mg/liter) did not modify egg hatch or J2 penetration. There was no adverse effect from the high ammonium nitrate concentrations or an equivalent potassium nitrate concentration on root dry weight. Ammonium ions influence nematodes both directly and via plant roots and do so independently of microbial organisms.     

Citrus tissue culture: stimulation of fruit explant cultures with orange juice

In vitro growth of explant (juice vesicle or albedo tissues) cultures from citron (Citrus medica), lemon (C. limon), grapefruit (C. paradisi), sweet orange (C. sinensis), and mandarin (C. reticulata) fruits was stimulated by addition of orange juice (10% v/v optimum) to a basal medium containing Murashige and Skoog salts, 50 grams per liter sucrose, 100 milligrams per liter myo-inositol, 5 milligrams per liter thiamine·HCl, 2 milligrams per liter 2,4-dichlorophenoxyacetic acid and 0.5 milligrams per liter kinetin. In analyzing this effect of orange juice on citron explant cultures, we failed to obtain increased yields by addition of appropriate concentrations of citric acid to the basal medium but obtained growth stimulation when the medium was supplemented with juice from an “acidless” orange variety (cv. Lima). These facts suggest that some component(s) other than citric acid is involved. Addition of the inorganic ash corresponding to 10% (v/v) orange juice to the basal medium had no effect on yields. Similarly, the stimulatory effect of orange juice could not be explained based on its content of sucrose or of organic growth factors already present in the basal medium.     

Nitrogen fixation of nodulation mutants of soybean as affected by nitrate

It was previously reported that three soybean (Glycine max [L.] Merr.) nodulation mutants (NOD1-3, NOD2-4, and NOD3-7) were partially tolerant to nitrate when nitrate was supplied simultaneously with inoculation at the time of transplanting. The current study evaluated the effect of short-term nitrate treatment on nitrogenase activity (C₂H₂ reduction per plant and per nodule weight) and on relative abundance of ureides when nitrate application was delayed until plants were 3 weeks old and nodules were fully developed. Nitrogenase activity of the mutants was similar to that of Williams after an initial 3-week growth period, prior to nitrate treatment. Application of 5 millimolar nitrate resulted in greater inhibition of nitrogenase activity in Williams than in the three mutants. NOD1-3 was most tolerant of nitrate among the mutants tested and showed the highest relative abundance of ureides. Although C₂H₂ reduction activity per plant for NOD1-3 was higher than for Williams in the presence of nitrate, C₂H₂ reduction activity per gram of nodules was lower for NOD1-3 than for Williams in the presence and absence of nitrate. Compared to Williams, NOD1-3 had higher nodule ureide concentration and had similar glutamine synthetase activity in nodule tissue, indicating its nodules have normal nitrogen assimilation pathways. Nitrate application resulted in ureide accumulation in nodule tissue as well as in all plant parts assayed. Unexpectedly, nitrate treatment also increased the rate of ureide degradative capacity of leaves in both NOD1-3 and Williams. The data confirmed that nitrogenase activity of the selected nodulation mutants was more, but still only partially, tolerant of nitrate compared with the Williams parent.     

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.     

Endogenous Abscisic Acid and Indole-3-Acetic Acid and Somatic Embryogenesis in Cultured Leaf Explants of Pennisetum purpureum Schum. : Effects in Vivo and in Vitro of Glyphosate, Fluridone, and Paclobutrazol

Effects of application in vivo of glyphosate, fluridone, and paclobutrazol to glasshouse-grown donor plants of Pennisetum purpureum Schum. on endogenous levels of abscisic acid (ABA) and indole-3-acetic acid (IAA) in young leaves and on somatic embryogenesis in cultured leaf explants were studied. Treatment of plants with glyphosate (100 milligrams per liter) resulted in elevated levels of endogenous ABA and IAA in young leaves. In contrast, paclobutrazol (50% active ingredient; 200 milligrams per liter) did not alter the endogenous levels of ABA and IAA. Fluridone (100 milligrams per liter) markedly inhibited synthesis of ABA and leaf explants from fluridone-treated plants lost the capacity for somatic embryogenesis. Explants from glyphosate- or paclobutrazol-treated plants did not show any reduction in embryogenic capacity when compared with untreated control plants. Glyphosate and fluridone were also incorporated into the culture media at various concentrations (0 to 20 milligrams per liter) to study their effects in vitro on somatic embryogenesis in leaf explants from untreated, field-grown plants. Glyphosate was inhibitory to somatic embryogenesis but only at concentrations above 5 milligrams per liter. Fluridone inhibited somatic embryogenesis at all concentrations tested. Inhibition of somatic embryogenesis by fluridone, by either in vivo or in vitro application, could be overcome partially by (±)-ABA added to the culture medium. Exogenous application of (±)-ABA enhanced somatic embryogenesis and reduced the formation of nonembryogenic callus. Application of IAA or gibberellic acid (GA₃; >5 milligrams per liter) was inhibitory to somatic embryogenesis. These results indicate that endogenous ABA is one of the important factors controlling the embryogenic capacity of leaf explants in Napier grass.     

Mineral nutrient requirements of cucumber seedlings

Mineral nutrient requirements for maximum growth rate of cucumber (Cucumis sativus L.) seedlings are estimated on the basis of three criteria. 1. The optimal weight proportions among the nutrients present in the seedlings. In relation to N = 100, close to 75 K, 13 P, 9 Ca, and 9 Mg are required. 2. The optimal ratio between the nitrogen sources NH₄ and NO₃ in the solution. The best growth is recorded with about equivalent amounts. Nitrate alone is also highly productive. Cucumber prefers nitrate and is sensitive to high ammonium concentrations. 3. The optimal total concentration in the solution corresponds to 200 to 300 milligrams of nitrogen per liter, with the proportions of the nutrients according to criterion 1. Simple growth methods are suggested in which the three criteria are fulfilled.     

Effect of Nitrate on Bradyrhizobium — Mungbean (Vigna radiata) Symbiosis in presence of Syringaldehyde

Bradyrhizoblum — mungbean (Vigna radiata) symbiosis was influenced by the application of nitrate in combination with.yrlngaldehyde. Application of nitrate alone at lower concentrations (2, 5 mM) caused a reduction at initial stage (10 DAS) while at later stage (35 DAS–45 DAS) enhanced the nodulation status (nodule number, nodule weight) and nodule efficiency (acetylene reduction activity). Higher concentration of nitrate (10 mM) caused a reduction in nodule number, nodule weight and acetylene reduction activity. Syringaldehyde alone improved the nodulation status and nodule efficiency while it acted synergistically when applied in combination with nitrate. These effects were also reflected in the biomass of plants.     

Oxygen-Nitrogen Relationships in Autotrophic Nitrification

Oxygen utilization by the autotrophic nitrifiers Nitrosomonas and Nitrobacter was studied. Experimental evidence is presented which reflects the effect of carbon dioxide fixation on overall oxygen utilization in autotrophic nitrification. Measurement of dissolved oxygen and inorganic nitrogen changes indicates that oxygen-nitrogen ratios in inorganic nitrogen oxidation are equal to 3.22 parts (expressed in milligrams per liter) of oxygen per part of ammonia nitrogen oxidized to nitrite nitrogen and 1.11 parts of oxygen per part of nitrite nitrogen oxidized to nitrate nitrogen. These values rather than the stoichiometric ratios should be used in nitrogenous oxygen demand calculations.     

Effect of nitrate on acetylene reduction activity and carbohydrate composition of Phaseolus vulgaris nodules

When Phaseolus vulgaris L. cv. Kentucky Wonder plants were supplied with various levels of nitrate for 34 days, nodule weight (plant)⁻¹, acetylene reduction activity (g nodule)⁻¹, and sugar concentration in nodules were depressed >60% (7.5 m M nitrate vs nil nitrate). Starch concentration in nodules was more than double the sugar concentration and declined only slightly in response to nitrate level. At the highest level of nitrate, sugar concentration in nodules was 50% greater than that in roots and nodule starch was about 6-fold greater than root starch on a fresh weight basis. When plants were grown with 1 m M nitrate and then supplied with 12 m M nitrate for 7 days, the rapid decline in acetylene reduction activity coincided with a decline in sucrose concentration. However, glucose and fructose concentrations declined only after the largest decrease in acetylene reduction had occurred, and the quantitative decrease in glucose and fructose in nodules was small relative to sucrose. Other results showed that the magnitude of the effect of nitrate on some nodule carbohydrate compounds depends on Rhizobium phaseoli strain and on whether plants were grown with or without nitrate prior to experimental treatments. Some of the results are consistent with the carbohydrate-deprivation hypothesis for inhibition of legume nodules by nitrate. However, there are several complications involved in the interpretation of results of this type, and other possible explanations for the results are suggested.     

Xylem and Phloem transport and the functional economy of carbon and nitrogen of a legume leaf

Exchanges of CO₂ and changes in content of C and N were studied over the life of a leaf of Lupinus albus L. These data were combined with measurements of C:N weight ratios of xylem (upper stem tracheal) and phloem (petiole) sap to determine net fluxes of C and N between leaf and plant. Phase 1 of leaf development (first 11 days, leaf to one-third area) showed increasing net import of C and N, with phloem contributing 61% of the imported C and 18% of the N. ¹⁴C feeding studies suggested the potential for simultaneous import and export through phloem over the period 9 to 12 days. Phase 2 (11-20 days, leaf attaining maximum area and net photosynthesis rate) exhibited net import through xylem and increasing export through phloem. Eighty-two% of xylem-delivered N was consumed in leaf growth, the remainder exported in phloem. Phase 3 (20-38 days) showed high but declining rates of photosynthesis, translocation, and net export of N. Phase 4 (38-66 days) exhibited substantial losses of N and declining photosynthesis and translocation of C. C:N ratio of xylem sap remained constant (2.3-2.6) during leaf life; petiole phloem sap C:N ratio varied from 25 to 135 over leaf development. The relationships between net photosynthesis and N import in xylem were: phase 1, 4.8 milligrams C per milligram N; phase 2, 24.7 milligrams C per milligram N; phase 3, 91.9 milligrams C per milligram N; and phase 4, 47.7 milligrams C per milligram N.