Production of Cellulolytic Enzymes by Botryodiplodia theobromae, Pat.

Carboxymethylcellulase (CM-cellulase) and ß-glucosidaseactivities were induced in cultures of Botryodiplodia theobromae,Pat. Both growth of the fungus and CM-cellulase production werebetter with sodium nitrate as nitrogen source than with eitherammonium nitrate or ammonium chloride. Growth, per unit nitrogensupplied, was greater with glutamic acid and aspartic acid asnitrogen sources than with sodium nitrate; this was probablybecause the fungus utilized the carbon of these amino-acids.Most ß-glucosidase activity was formed with ammoniumchloride or ammonium nitrate as nitrogen source. Glucose inhibited the activity of ß-glucosidase markedly.Low glucose concentration (c. 0.003 M) stimulated CM-cellulaseactivity but concentrations above 0.05 M inhibited. The formationof both enzymic activities was repressed in the presence ofglucose. ß-glucosidase activity was more thermolabilethan that of CM-cellulase.     

Effect of nitrate and ammonium concentration on nitrate reductase activity in five species of mycorrhizal fungi

The nitrate reducing capacity of pure cultures of Cenococcum geophilum (Sow.) Ferd. & Winge, Paxillus involutes (Batsch: Fr.) Fr. (strains 1 and 2), Piloderma croceum Erikss. & Hjortst., Suillus variegatus (Fr.) O. Kuntze (strains 1 and 2) and an ectendomycorrhizal (E-strain) fungus was measured using an in vivo nitrate reductase (EC 1.6.6.3) assay. Differences between species and strains were established. The nitrate concentration of the culture medium influenced the nitrate reductase activities of the E-strain fungus and one strain of S. variegatus. The nitrate reductase activity of certain species and strains was a function of nitrate concentration. Addition of ammonium to the growth medium did not have any significant effect on the in vivo or in vitro nitrate reductase activity. The in vivo nitrate reductase activity in the mycelia of C. geophilum and the E-strain fungus decreased during 28 day growth in modified Melin-Norkrans medium. For mycelia of Paxillus involutus, Piloderma croceum and S. variegatus grown on agar the in vitro assays showed higher nitrate reductase activity than the in vivo assays.     

The Effects of Changes in the Inorganic Nitrogen Supply on the Growth and Development of Marsilea in Aseptic Culture

The effects of changes in the inorganic nitrogen source of themedium on the growth and development of Marsilea in asepticculture are described. A change in nitrate concentration to0·2 or 5 x that of the normal medium had initially noeffect on growth or heteroblastic development, but in the 0·2nitrogen medium there was finally a reversion to juvenile stagesfollowed by early cessation of growth. Growth and heteroblastic development were stimulated by thesubstitution of the nitrate of the medium by ammonium saltsor urea. Nitrite was little inferior to nitrate as a nitrogensource, but hydroxylamine was toxic even at low concentrations.Addition of sodium chloride led to improved growth and developmentin nitrate media, an effect referred to beneficial effects ofthe chloride ion. Symptoms of toxicity and eventual death of the cultures whichoccurred after prolonged growth on ammonium media were due toan acid drift of the media rather than to a specific effectof the ammonium ion.     

Carbon and nitrogen requirements of Fusarium oxysporum causing cotton wilt

Summary The present work was carried out to study the nutritional requirements of the cotton wilt-inducing fungus, i.e.Fusarium oxysporum on a synthetic liquid medium with regard to the carbon and nitrogen sources at varying concentrations in terms of the average mycelial dry weights.The optimum carbon requirements of the fungus ranged from 7000–8000 p.p.m. irrespective of the carbon source used in experiment. Carbon utilization was best on sucrose followed by maltose, starch, glucose, fructose and cellulose successively.The optimum nitrogen requirements of the fungus were 300 p.p.m. of nitrogen in the medium; nitrogen utilization was best on using nitrate-nitrogen followed by glycine, glutamic acid, ammonium nitrate, asparagine and ammonium sulphate.Maximum growth of the fungus took place on media containing a C/N ratio ranging between 22.8 and 25.7.Colour formation is correlated with varying either source or concentration of nitrogen and not carbon.     

Heterotrophic Nitrification in an Acid Forest Soil and by an Acid-Tolerant Fungus

Nitrate was formed from ammonium at pH 3.2 to 6.1 in suspensions of a naturally acid forest soil; the maximum rates of formation occurred at ca. pH 4 to 5. Nitrate was also formed from soil nitrogen in suspensions incubated at 50°C. Autotrophic nitrifying bacteria could not be isolated from this soil. Enrichment cultures produced nitrate in a medium with β-alanine if much soil was added to the medium, and nitrite but not nitrate was formed in the presence of small amounts of soil. Nitrification by these enrichments was abolished by eucaryotic but not procaryotic inhibitors. A strain of Absidia cylindrospora isolated from this soil was found to produce nitrate and nitrite in a medium with β-alanine at pH values ranging from 4.0 to 4.8. Nitrate production by A. cylindrospora required the presence of sterile soil. Free and bound hydroxylamine, hydroxamic acids, and primary aliphatic nitro compounds did not accumulate during the conversion of β-alanine to nitrite by the fungus. The organism also formed nitrite from ammonium in a medium containing acetate. We suggest that nitrification in this soil is a heterotrophic process catalyzed by acid-tolerant fungi and not by autotrophs or heterotrophs in nonacid microsites.     

Spodoptera littoralis detoxifies neurotoxic 3-nitropropanoic acid by conjugation with amino acids

Spodoptera littoralis is a phytophagous generalist. Its host range includes more than 40 plant species, some of which produce 3-nitropropanoic acid (3-NPA), an irreversible inhibitor of mitochondrial succinate dehydrogenase. Growth in larvae fed an artificial diet with a sublethal admixture of 3-NPA (4.2 μmol per g) was slowed significantly, but larvae experienced no increase in mortality. In contrast, larvae injected with 25.2 μmol/g (bodyweight) 3-NPA experienced acute toxicity and death. To study the detoxification mechanism of 3-NPA in S. littoralis, the insect frass was analyzed by HPLC-MS. Comparative analysis of 3-NPA-treated and -untreated control samples using HR-MS² revealed a group of differential signals that were identified as amino acid amides of 3-NPA with glycine, alanine, serine, and threonine. When sublethal amounts of stable isotope-labeled 3-NPA were injected into a larva's hemolymph, 3-NPA amino acid conjugates were identified as putative detoxification products. Bioassays with synthetic standards confirmed that the toxicity of the amides was negligible in comparison to the toxicity of free 3-NPA, demonstrating that amino acid conjugation in S. littoralis represents an efficient way to detoxify 3-NPA. Furthermore, biosynthetic studies using crude fractions of the gut tissue indicated that conjugation of 3-NPA with amino acids occurs in epithelial cells of the insect's gut. Taken together, these results suggest that the detoxification of 3-NPA in S. littoralis proceeds via conjugation to specific amino acids within the epithelial cells followed by export of the nontoxic amino acid conjugates to the hemolymph via as yet uncharacterized mechanisms, most likely involving the Malpighian tubules.     

Interaction between Streptococcus lactis and Aspergillus flavus on production of aflatoxin

The inoculation of Aspergillus flavus spores into a culture of Streptococcus lactis in Lablemco tryptone broth medium resulted in little or no aflatoxin accumulation even though the growth of the fungus was not hindered. The drop in pH and reduced nutrient levels in the medium as a result of the S. lactis growth were not the cause of the observed inhibition. The inhibition was not eliminated by the addition of carbohydrate equal to the amount used by the bacterium before the inoculation with the fungus. Aflatoxin levels were also markedly reduced when S. lactis was inoculated into a growing A. flavus culture. In addition to inhibiting the synthesis of aflatoxin, S. lactis also degraded preformed toxin. A. flavus, on the other hand, not only reduced the growth of S. lactis but also affected the morphology of the bacterial cell; the cells became elongated and formed long chains. S. lactis produced and excreted the inhibitor into the medium late in its growth phase. The inhibitor was a heat-stable low-molecular-weight compound. Chloroform extracts of A. flavus grown in the presence of S. lactis were toxic to Bacillus megaterium but did not exhibit mutagenic or carcinogenic activity in the Salmonella/mammalian microsome mutagenicity test.     

Interaction between Streptococcus lactis and Aspergillus flavus on production of aflatoxin.

The inoculation of Aspergillus flavus spores into a culture of Streptococcus lactis in Lablemco tryptone broth medium resulted in little or no aflatoxin accumulation even though the growth of the fungus was not hindered. The drop in pH and reduced nutrient levels in the medium as a result of the S. lactis growth were not the cause of the observed inhibition. The inhibition was not eliminated by the addition of carbohydrate equal to the amount used by the bacterium before the inoculation with the fungus. Aflatoxin levels were also markedly reduced when S. lactis was inoculated into a growing A. flavus culture. In addition to inhibiting the synthesis of aflatoxin, S. lactis also degraded preformed toxin. A. flavus, on the other hand, not only reduced the growth of S. lactis but also affected the morphology of the bacterial cell; the cells became elongated and formed long chains. S. lactis produced and excreted the inhibitor into the medium late in its growth phase. The inhibitor was a heat-stable low-molecular-weight compound. Chloroform extracts of A. flavus grown in the presence of S. lactis were toxic to Bacillus megaterium but did not exhibit mutagenic or carcinogenic activity in the Salmonella/mammalian microsome mutagenicity test.     

NUTRITIONAL REQUIREMENTS FOR OVULE FORMATION IN EXCISED PISTILS OF NIGELLA

The nutritional requirements for ovule formation in Nigella saliva L. were investigated by growing excised pistils on defined media. Pistils grown on a medium containing the minerals of Murashige and Skoog produced significantly more ovules than on a medium containing the minerals of Bilderback. When the nitrogen, sulfate, and phosphate of the Bilderback medium were adjusted to levels comparable to those of the Murashige and Skoog medium, a similar number of ovules was formed. The effect of different forms and concentrations of nitrogen on ovule formation and pistil growth was investigated. High concentrations of nitrate (40 mil) favored pistil growth and ovule formation, but comparable levels of ammonium were toxic. When ammonium at concentrations above 10 mM was added to nitrate media, ovule formation was inhibited. A medium containing low concentrations of ammonium (10 mM) and nitrate (5 mM) supported more ovule formation and pistil growth in young pistils than a low-nitrate (5 mM) medium without ammonium. However, ovule formation on a medium containing 10 mM ammonium and 5 mM nitrate was significantly less than on a medium containing only 15 mM nitrate. Low concentrations of organic nitrogen in the form of α-alanine (1 mM) and γ-aminobutyric acid (5 mM) supported ovule formation and pistil growth similar to a high nitrate medium.     

Nitrogen acquisition in four co-existing species from an upland acidic grassland

The concentration of both nitrate and ammonium nitrogen was measured in soil taken from an upland acidic (pH 4.5) grassland habitat, containing four co-existing species, Deschampsia flexuosa (L.) Trin., Festuca ovina L., Juncus squarrosus L. and Nardus stricta L. Both nitrate and ammonium nitrogen were found to be present in the soil, in similarly small quantities. The effect of both sources of nitrogen on relative growth rate was studied, and an attempt was made to determine whether nitrate or ammonium nitrogen is the immediate source of nitrogen for these plants using assays of nitrate reductase (EC 1.6.6.2) and ammonium uptake. All four species showed larger growth rates on the same concentration of ammonium nitrogen compared to nitrate nitrogen. All species showed low activities of leaf nitrate reductase, even in plants grown on 18 mol nitrate m⁻³. Ammonium uptake activity appeared to be higher in species which showed the lowest nitrate reductase activity and least response to increasing nitrate concentration in the growth medium.     

Absorption of Copper by Lemna as Influenced by Some Factors Which Nullify the Copper Effect on Flowering and Growth

The effect of copper on flowering and growth of Lemna paucicostata6746 and Lemna gibba G3 in a copper-containing medium is nullifiedby the addition of EDTA, ammonium ions or salicylic acid tothe medium or a decrease in its nitrate concentration. Thesefactors were examined for their effects on the absorption ofcopper by the plants. The addition of EDTA to the medium completelyinhibited the absorption of copper in both species, thus eliminatingthe copper effect. Ammonium ions also inhibited copper absorption,their effectiveness rising with their concentration. Loweringthe nitrate concentration in the medium nullified the coppereffect on flowering in L. paucicostata 6746, and the additionof salicylic acid to the medium also nullified the copper effectin L. gibba G3, both without affecting the absorption of copper. (Received June 7, 1982; Accepted August 27, 1982)     

与盾叶薯蓣细胞共生培养的内生真菌分离及其生物学特性

从盾叶薯蓣根状茎中分离到一株与盾叶薯蓣细胞共生培养的内生真菌（RE0105）。测定了该菌ITS1-5．8S rDNA—ITS2序列并构建了系统发育树,发现RE0105与Libertella属形成一个类群,具有较近亲缘关系。该真菌能产生与宿主植物相同的代谢产物——薯蓣皂苷元。对RE0105培养特性研究表明：富含有机营养的培养基有利于其生长;盾叶薯蓣细胞提取物可明显刺激其生长;20μmol／L生长素NAA和2,4-D能够促进RE0105的生长。200μmol／L细胞分裂素KT和BA对RE0105生长具有一定的促进作用,但与生长素相比,细胞分裂素对RE0105生长的刺激作用较小。蔗糖、葡萄糖、果糖、乳糖、木糖和NH4NO3是RE0105生长的良好C源和无机N源,但NH4Cl和（NH4）2SO4对该菌生长具有明显抑制作用,并且显著影响其形态特征。     

Effects of high ammonium concentrations on growth and anthocyanin formation in grape (Vitis vinifera L.) cell suspension cultured in a production medium

Cultivating Vitis vinifera cell suspensions in a production medium which is characterized by high sucrose and low nitrate concentrations (132 mM and 6.25 mM respectively) repressed growth but enhanced the intracellular accumulation of anthocyanins, especially peonidin 3-glucoside. Increasing the ammonium concentration of the production medium from 2 to 8–16 mM increased growth and decreased the accumulation of anthocyanins and peonidin 3-glucoside specifically. Instead, peonidin 3-p-coumaroylglucoside accumulated. At 24 mM ammonium concentration, growth was inhibited and accumulation of peonidin 3-p-coumaroylglucoside was significant (p<0.05) and represented 42% of total anthocyanins after 12 days of culture compared with 19% in the production medium with 2 mM ammonium.Contribution Number 217.     

Heterotrophic growth and nutritional aspects of the diatom Cyclotella cryptica (Bacillariophyceae): effect of nitrogen source and concentration

To investigate the nutritional value of the diatom Cyclotella cryptica Reimann, Lewin, and Guillard as an alternative feed for the use in the aquaculture industry, the heterotrophic growth characteristics, total fatty acids, and the resultant fatty acid profile of the microalga were studied when cultivated with sodium nitrate, ammonium chloride, or urea. All three nitrogen sources supported growth under heterotrophic conditions, and their uptake affected the pH of the cultivation medium, even when buffered. The use of sodium nitrate or urea resulted in a significant increase in the pH of the cultivation media, whereas the use of ammonium chloride caused a minor decrease in the pH of the cultivation media. The maximum specific growth rate was highest when urea and ammonium chloride were supplied at a low concentration; however, the total fatty acid content was not significantly affected (P = 0.101) by the nitrogen source when supplied at 10.7 mM nitrogen. The total fatty acid content and fatty acid profile of C. cryptica was more affected by the growth phase (predominately influenced by the initial nitrogen concentration) than by the source of nitrogen.     

Formation of Nitrate from 3-Nitropropionate by Aspergillus flavus

Extracts of the hyphae of a nitrifying strain of Aspergillus flavus formed nitrite and nitrate from 3-nitropropionate. Nicotinamide adenine dinucleotide phosphate and nicotinamide adenine dinucleotide enhanced the production of nitrate but not nitrite, whereas cysteine and diethyldithiocarbamate increased nitrite but diminished nitrate synthesis. Quinacrine reduced the extent of conversion of the nitro compound to nitrite and nitrate, but only the inhibition of nitrite formation was completely reversed by flavine coenzymes. Molecular oxygen was essential for this part of the nitrification sequence. 3-Chloropropionate stimulated the oxidation of nitrite by hyphae or enzyme preparations. Although the fungus contained a noncytochrome-linked nitrite-oxidizing enzyme, partially purified preparations free of this enzyme formed both nitrite and nitrate from 3-nitropropionate. Possible mechanisms of this latter stage of heterotrophic nitrification are discussed.     

Importance of seed Zn content for wheat growth on Zn-deficient soil

Eggplants (Solanum melongena L. cv. Bonica) were grown in a glasshouse during summer under natural light with one unbranched shoot or one shoot with 3 to 4 branches and with or without fruit in quartz sand buffered and not buffered with 0.5% CaCO₃ (w : v), respectively. Nutrient solutions supplied contained nitrate or ammonium as the sole nitrogen source. Compared with nutrient solutions containing nitrate (10 mM), solutions containing ammonium (10 mM) caused a decrease in net photosynthesis of eggplants during early stages of vegetative growth when grown in quartz sand not buffered with CaCO₃. The decrease was not observed before leaves showed interveinal chlorosis. In contrast, net photosynthesis after bloom at first increased more rapidly in eggplants supplied with ammonium than with nitrate nitrogen. However, even in this case, net photosynthesis decreased four weeks later when ammonium nutrition was continued. The decrease was accompanied by epinasty and interveinal chlorosis on the lower leaves and later by severe wilting, leaf drop, stem lesions, and hampered growth of stems, roots, and fruits. These symptoms appeared later on plants not bearing fruits than on plants bearing fruits. If nutrient solutions containing increasing concentrations of ammonium (0.5–30 mM) were supplied after the time of first fruit ripening, shoot growth and set of later flowers and fruits were promoted. In contrast, vegetative growth and reproduction was only slightly affected by increasing the concentration of nitrate in the nutrient solutions. In quartz sand buffered with CaCO₃ ammonium nutrition caused deleterious effects only under low light conditions (shade) and on young plants during rapid fruit growth. If eggplants were supplied with ammonium nitrogen before bloom, vegetative growth was promoted, and set of flowers and fruit occurred earlier than on plants supplied with nitrate. Furthermore, the number of flowers and fruit yield increased. These effects of ammonium nutrition were more pronounced when plants were grown with branched shoots than with unbranched shoots. The results indicate that vegetative and reproductive growth of eggplants may be manipulated without causing injury to the plants by supplying ammonium nitrogen as long as the age of the plants, carbohydrate reserves of the roots, quantity of ammonium nitrogen supplied, and pH of the growth medium are favourable. T W Rufty Section editor     

Effect of nitrogen form on the growth and nitrate concentration of lettuce

Summary A pot experiment with lettuce involving three N forms each at six application levels, showed that lettuce can be grown satisfactorily with a very low nitrate content when supplied with ammonium sulphate and a nitrification inhibitor. For plants growing on nitrate N, the optimum midrib sap nitrate concentration as maturity approached was about 1400 mg/1 NO₃-N. Large losses of mineral N were observed from the peat medium, even in the absence of plants. A relationship is presented which would enable a lettuce grower to estimate whole-shoot nitrate concentration from a quick test of midrib sapi.e. NO₃-N (mg/kg in fresh shoot) =0.14×NO₃-N (mg/l in sap). Tipburn was worst at intermediate levels of applied N, and was less serious with pure ammonium nutrition than with nitrate.     

Induction,maintenance, and differentiation of rice callus cultures on ammonium as sole nitrogen source

Rice callus was initiated from cultured anthers and was maintained on medium containing ammonium as sole nitrogen source and supplemented with either a buffer or an organic acid. Plant regeneration also was obtained on this medium, but the morphogenetic capacity of the callus culture declined as rapidly as on a medium containing a mixture of ammonium and nitrate. The results of the studies suggest that organic acids provide an additional function beyond that of buffering the medium during growth on ammonium.     

Effect of specific amino acids on growth and aflatoxin production by Aspergillus parasiticus and Aspergillus flavus in defined media.

Four amino acids were used as sole nitrogen sources or as supplements to ammonium sulfate, and casein and ammonium sulfate were used as sole nitrogen sources to examine their effects on aflatoxin production by Aspergillus parasiticus NRRL 2999 and Aspergillus flavus 3357 grown on synthetic liquid media. In general, when proline, asparagine, casein, and ammonium sulfate were used as sole nitrogen sources, they supported more growth and toxin production than tryptophan or methionine. However, proline stimulated more toxin production per gram of mycelium in stationary cultures than the other nitrogen sources, including the amino acid asparagine, which is generally recognized as supporting good aflatoxin production. The exact responses to individual nitrogen sources were influenced by the species of fungus and whether cultures were stationary or shaken. In shake cultures, but not in stationary cultures, increased growth was generally associated with increased toxin production.