Field Application of Calcium to Reduce Phytophthora Stem Rot of Soybean, and Calcium Distribution in Plants.

The effect of calcium compounds [Ca(HCOO)2-A and Ca(NO3)2] on the incidence of Phytophthora stem rot of soybean (Glycine max) cv. Tanbakuro was investigated in the field. Disease incidence in control plants in three fields naturally infested with Phytophthora sojae ranged from 11.7 to 52.0% at 140 days after transplanting. Independent of the pathotype diversity, 4 and 10 mM of the calcium compounds applied twice (prior to transplanting and 14 days after transplanting) significantly suppressed disease incidence and delayed onset. Ca(HCOO)2-A (Suicaru) was more effective than calcium nitrate for reducing disease incidence. In most cases, the calcium amendments increased plant height, number of nodes and pods, and seed yields, and reduced low-quality seeds. Scanning electron microscopy with fresh samples showed increased accumulation of calcium crystals around the cambium and xylem elements of soybean plants treated with 10-mM Ca(HCOO)2-A and Ca(NO3)2. Mycelial penetration was inhibited at these sites. These results indicated that calcium-rich areas may be more resistant to invasion by P. sojae, and the calcium crystals may play an important role in calcium ion storage and its availability for those tissues to maintain long-term field resistance.

Boron toxicity in rice (Oryza sativa L.). I. Quantitative trait locus (QTL) analysis of tolerance to boron toxicity.

Boron toxicity tolerance of rice plants was studied. Modern japonica subspecies such as Koshihikari, Nipponbare, and Sasanishiki were tolerant, whereas indica subspecies such as Kasalath and IR36 were intolerant to excessive application of boron (B), even though their shoot B contents under B toxicity were not significantly different. Recombinant inbred lines (RILs) of japonica Nekken-1 and indica IR36 were used for quantitative trait locus (QTL) analysis to identify the gene responsible for B toxicity tolerance. A major QTL that could explain 45% of the phenotypic variation was detected in chromosome 4. The QTL was confirmed using a population derived from a recombinant inbred line which is heterogenic at the QTL region. The QTL was also confirmed in other chromosome segment substitution lines (CSSLs).

Select Calcium Compounds Reduce the Severity of Phytophthora Stem Rot of Soybean.

This study investigated the effects of several calcium compounds on Phytophthora stem rot of soybean (Glycine max) and fungal growth and zoospore release of a Phytophthora sojae isolate in vitro. All concentrations of five formulated calcium products [Ca(COOH)2-A, Ca(COOH)2-B, Ca(COOH)2-C, CaSO4-A, and CaCl2-A] and two chemical compounds [CaCl2 and Ca(NO3)2] applied prior to inoculation significantly suppressed disease incidence. Among all the products and chemicals, Ca(COOH)2-A was the most effective in suppressing the incidence of disease. In most cases, no significant relationship was observed between inhibition of growth rate in vitro and disease reduction in growth chamber tests. Therefore, disease suppression recorded in laboratory experiments using pathogen mycelium was likely due to the responses of plant tissues rather than the direct inhibition of pathogen fungal growth by the calcium compounds. The extent of disease reduction was related to increased calcium uptake by plants, suggesting that calcium was the effective element in reducing Phytophthora stem rot. Seedling tray experiments using zoospores indicated that the application of 10 mM Ca(COOH)2-A was more effective for reducing incidence of disease under growth chamber conditions, compared to other concentrations. The presence of 4 to 20 mM of all seven calcium solutions decreased the release of zoospores, although 0.4 mM of all compounds significantly increased zoospore release. Therefore, disease reduction in the growth-chamber experiments was due to the multiple effects of direct suppression on zoospore release and fungal growth in combination with the response of the host plant tissue to Ca(COOH)2-A.

Boron nutrition of cultured tobacco BY-2 cells. III. Characterization of the boron-rhamnogalacturonan II complex in cells acclimated to low levels of boron.

Cultured cells of tobacco (Nicotiana tabacum L.) BY-2 which could propagate at the same rate as the parent cells (1 mg B liter(-1)) under a lower level of boron (0.01 mg B liter(-1)) were obtained. The selected cells had swollen cell walls. In the parent cells, all the RG-II occurred as a B-RG-II complex, however, two-thirds of the RG-II occurred in a monomeric form in the selected cells.

Borate-rhamnogalacturonan II bonding reinforced by Ca2+ retains pectic polysaccharides in higher-plant cell walls

The extent of in vitro formation of the borate-dimeric-rhamnogalacturonan II (RG-II) complex was stimulated by Ca2+. The complex formed in the presence of Ca2+ was more stable than that without Ca2+. A naturally occurring boron (B)-RG-II complex isolated from radish (Raphanus sativus L. cv Aokubi-daikon) root contained equimolar amounts of Ca2+ and B. Removal of the Ca2+ by trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid induced cleavage of the complex into monomeric RG-II. These data suggest that Ca2+ is a normal component of the B-RG-II complex. Washing the crude cell walls of radish roots with a 1.5% (w/v) sodium dodecyl sulfate solution, pH 6.5, released 98% of the tissue Ca2+ but only 13% of the B and 22% of the pectic polysaccharides. The remaining Ca2+ was associated with RG-II. Extraction of the sodium dodecyl sulfate-washed cell walls with 50 mM trans-1,2-diaminocyclohexane-N, N,N',N'-tetraacetic acid, pH 6.5, removed the remaining Ca2+, 78% of B, and 49% of pectic polysaccharides. These results suggest that not only Ca2+ but also borate and Ca2+ cross-linking in the RG-II region retain so-called chelator-soluble pectic polysaccharides in cell walls.

Two Chains of Rhamnogalacturonan II Are Cross-Linked by Borate-Diol Ester Bonds in Higher Plant Cell Walls.

Polysaccharide moiety of the boron-polysaccharide complex (T. Matoh, K. Ishigaki, K. Ohno, J. Azuma [1993] Plant Cell Physiol 34: 639-642) isolated from radish (Raphanus sativus) roots has been shown to be rhamnogalacturonan II by glycosyl-linkage analysis and the presence of diagnostic monosaccharides, including apiose, aceric acid, 2-O-methylfucose, and 3-deoxy-D-manno-2-octulosonic acid. Removal of boron from the complex reduced the molecular weight by one-half without causing a significant increase in the number of reducing end groups, indicating that boron, as boric acid, links two rhamnogalacturonan II chains together to form the boron-polysaccharide complex.

Sodium Stimulates Growth of Panicum coloratum through Enhanced Photosynthesis.

A sodium-requiring C(4) plant, Panicum coloratum Walt. cv Kabulabula, was grown with and without sodium. Rate of nitrogen uptake and photosynthesis were measured during the recovery from sodium deficiency. The beneficial effect of sodium on growth was apparent irrespective of nitrogen source, ammonium- or nitrate-nitrogen. The leaf photosynthetic rate ((14)CO(2) fixation) doubled by sodium within 1 hour of the application.

Sodium Stimulates Growth of Amaranthus tricolor L. Plants through Enhanced Nitrate Assimilation.

Effects of Na application on the capacity of NO(3) (-) assimilation were studied in Na-deficient Amaranthus tricolor L. cv Tricolor plants. On day 30 after germination, Na-deficient A. tricolor plants received either 0.5 millimolar NaCl or KCl. The level of nitrate reductase activity doubled within 24 hours by the addition of Na and the enhanced level was maintained thereafter. When the plants were exposed to 2 millimolar (15)NO(3) (-), total (15)N taken up by the plants was greater in the Na-treated plants than in the K-treated plants within 24 hours of the Na treatment. Incorporation of (15)N into the 80% ethanol-insoluble nitrogen fraction of the Na-treated plants in the light period was about 260% of those of the K-treated plants indicating greater capacity of NO(3) (-) assimilation in the Na-treated plants. From these results, it was demonstrated that Na application to the Na-deficient A. tricolor plants promoted NO(3) (-) reduction and its subsequent assimilation into protein, resulting in growth enhancement.

Collapse of ATP-Induced pH Gradient by Sodium Ions in Microsomal Membrane Vesicles Prepared from Atriplex gmelini Leaves: Possibility of Na/H Antiport.

Sealed microsomal membrane vesicles were prepared from leaves of a 250 millimolar NaCl-grown halophyte (Atriplex gmelini C. A. Mey). The vesicles exhibited ATP-dependent proton-transporting activity which was inhibited 60% by NO(3) (-) (50 millimolar) but not by vanadate (100 micromolar) and 23% by oligomycin (10 micrograms per milliliter), suggesting that tonoplast-derived vesicles were the major constituents of the preparation. The pH gradient established by the vesicles by ATP in the presence of oligomycin collapsed upon the addition of Na(+) salts. The vesicles took up Na(+) ions in the presence of ATP and this activity was canceled by gramicidin. These results suggest that Na(+) ions were taken up by the vesicles via a Na(+)-specific uptake system, possibly a Na(+)/H(+) antiport.

Sodium-Stimulated NO(3) Uptake in Amaranthus tricolor L. Plants.

Nitrate uptake of Na(+) -deficient Amaranthus tricolor L. cv Tricolor seedlings from complete culture solution was stimulated by about 210% within 5 hours by application of 0.5 millimolar NaCl. From a Na(+) -preloading experiment, intracellular Na(+) was shown to be responsible for the stimulation of NO(3) (-) uptake. The results suggest a possible role of Na(+) in NO(3) (-) uptake in C(4) plants.

Early Responses of Sodium-Deficient Amaranthus tricolor L. Plants to Sodium Application.

Effects of sodium application on sodium-deficient Amaranthus tricolor L. cv Tricolor seedlings were studied. Thirty-day-old A. tricolor seedlings grown without sodium received either 0.5 millimolar of NaCl or KCl, and the changes in the growth rate, chlorophyll concentration, photosynthetic oxygen evolution, and dark-oxygen consumption, and some enzyme activities were compared. Following the sodium treatment, the sodium concentration in the leaves increased from the initial value of 0.4 millimolar to 2 to 3 millimolar within 24 hours, and also the relative growth rate and O(2) evolution were enhanced within 24 hours. The stimulation of O(2) evolution was greater in the upper leaves than in the lower leaves. Although total chlorophyll concentration did not increase significantly, the increase in the chlorophyll a/b ratio was apparent within 24 hours. There were not significant increases in the C(4) photosynthetic enzyme activities; however, nitrate reductase activity increased by 350% by the sodium treatment within 24 hours, and this increase is considered not to be one of the consequences of the improved photosynthesis. Results suggest that the sodium treatment promoted CO(2) and nitrate assimilation resulting in the growth enhancement, and that sodium can be involved in some other functions than C(4) photosynthesis in A. tricolor plants.

Sodium, Potassium, Chloride, and Betaine Concentrations in Isolated Vacuoles from Salt-Grown Atriplex gmelini Leaves.

Vacuoles were isolated via protoplasts from the leaves of a halophyte Atriplex gmelini C.A.Mey., grown in culture solution supplemented with 250 millimolar NaCl. Lysis of the protoplasts was induced by lowering the medium osmolarity (1.2 to 1.0 molar sorbitol) and adding a detergent, a synthesized cholate derivative, 3-([3-cholamidopropyl] dimethylammonio)-1-propanesulfonate at a concentration of 0.5 millimolar and the resulting vacuoles were purified by successive dilution and floatation. Isolated vacuoles contained almost the same concentration of sodium (569 millimolar) and chloride (260 millimolar) as recorded in protoplasts (582 and 254 millimolar, respectively), suggesting that the vacuoles are the major sequestration site of NaCl in leaves of halophytes. Betaine concentration in the protoplasts was about 16 millimolar, while that in vacuoles was only about 0.24 millimolar, indicating that betaine is accumulated in the cytoplasm as a compatible solute.

Changes in the activities of ferredoxin- and NADH-glutamate synthase during seedling development of peas.

Ferredoxin-glutamate synthase (EC 1.4.7.1) and NADH-glutamate synthase (EC 1.4.1.14) activities in pea seedlings (Pisum sativum L., cv. Alaska) were measured during germination and the early stages of growth. Both enzymes were detected at all stages in the developing roots and shoots, but their relative activities varied according to the growth stages. In shoots of 5-d-old seedlings, the ratio of the NADH-enzyme to the ferredoxin-enzyme was 72:28. However, the ferredoxin-activity increased rapidly and in shoots of 17-d-old seedlings, the activities were in the ratio of 3:97. Similar trends in these ratio changes were observed in the roots. In both tissues, the NADH-enzyme was shown to be predominant in the immature parts. When chloroplasts prepared from mature pea leaves were incubated with [(14)C]glutamine and 2-oxoglutarate, the production of [(14)C]glutamate was found to be light dependent and was inhibited by azaserine (3 mM) and 3-(3,4-dichlorophenyl)-1-1-dimethylurea (20 µM). In contrast, considerable amounts of [(14)C]glutamate were formed by chloroplasts from young leaves even in the dark. Addition of malate or dihydroxyacetone phosphate to the reaction mixture resulted in a twofold increase of this dark- and azaserine-sensitive [(14)C]glutamate formation.

Isolation and characterization of NADH-glutamate synthase from pea (Pisum sativum L.).

Both ferredoxin-glutamate synthase (EC 1.4.7.1) and NADH-glutamate synthase (EC 1.4.1.14) were isolated separately on DEAE-cellulose chromatography from etiolated pea shoots. The latter enzyme was purified 1,400-fold by ammonium sulfate fractionation and column chromatographies of DEAE-cellulose, Sephadex G-200 and blue-Sepharose. The enzyme had a molecular weight of 220,000 and an isoelectric point of 4.3. The optimum pH was 7.6. Apparent Km values for l-glutamine, 2-oxoglutarate and NADH were 400, 37 and 4 µm, respectively. The enzyme had its absorption maxima at 275, 375 and 440 nm, suggesting that pea NADH-glutamate synthase is a flavoprotein. It showed NADH-diaphorase activity toward ferricyanide and 2,6-dichlorophenol indophenol as the electron acceptor. Sulfhydryl reagents, metal-chelating reagents, phthalein acids and azaserine were strong inhibitors. Ammonium and phosphate ions enhanced the enzyme activity.