Partial Characterization of the Antimicrobial Activity of Streptomyces antimycoticus FZB53

The paper describes experiments aimed at evaluating the sensitivity of different fungi, most of them plant pathogens and bacteria towards Streptomyces antimycoticus FZB53, a biocontrol agent that, when applied as a seed treatment, in previous studies has shown good activity against different seed‐borne fungal diseases. When incorporated into agar media, the filtrate from shake cultures of S. antimycoticus FZB53 inhibited the mycelial growth or spore germination, respectively, of a broad spectrum of fungi. The most sensitive of the fungi tested was Fusarium culmorum. The inhibitory activity could be removed from the culture filtrate by extraction with ethyl acetate. When ethyl acetate extracts of the pellet and supernatant obtained by centrifugation of the shake culture were added to the agar medium, inhibition of mycelial growth of F. culmorum was restored, especially with the extracts of the pelleted biomass. Autoclaving of the culture filtrate reduced the inhibition of F. culmorum but completely eliminated the inhibitory activity against Fusarium graminearum. Among the bunt fungi tested, spore germination of Tilletia tritici was more sensitive to the culture filtrate of S. antimycoticus FZB53 than spore germination of Ustilago avenae and U. tritici. Separation by thin layer chromatography (tlc) and spraying with different reagents showed that ethyl acetate extracts from shake cultures or biomass scraped from agar media contained several hydrophobic metabolites. When eluted from the tlc‐plates, the material from one of the spots had strong antifungal activity against spore germination of T. tritici and mycelial growth of F. culmorum, respectively. Ethyl acetate extracts from biomass of S. antimycoticus FZB53 prevented the growth of the tested Gram‐positive bacteria, namely Clavibacter michiganensis and different species of Bacillus. The results indicated that these bacteria were at least as sensitive towards the metabolites of S. antimycoticus FZB53 as F. culmorum. The tested Gram‐negative bacteria were not affected.     

飞机草地上部分化学成分及其抑菌活性研究

为探究飞机草地上部分化学成分及其抑菌作用.采用正相硅胶、Sephadex LH-20等色谱方法,从飞机草乙酸乙酯提取物中得到10个化合物,根据1H NMR、13C NMR、MS波谱数据,分别鉴定为:5,6,7,4'-四甲氧基黄烷酮(1)、圣草素-7,4'-二甲醚(2)、5,7-二羟基-6,4'-二甲氧基黄酮(3)、二氢...     

Tumor cell biotransformation products of prostaglandin A1 with growth inhibitory activity

The growth inhibitory effect and the fate of prostaglandin A1 (10(-6) M) were followed in cultures of rat B104 neuroblastoma and C6 glioma cells. More than 40% and 85% of the drug were neither recognized by a prostaglandin A1 antiserum nor extracted from the acidified medium with ethyl acetate, after 6 h and 24 h-incubation, respectively. When the supernatant of cells cultured in the presence of prostaglandin A1 during 24 hours was transferred to other cells and used as culture medium, the same growth inhibitory effect as with prostaglandin A1 was observed even when no prostaglandin A1 was added. After extensive purification and reverse phase HPLC of supernatant, four peaks more polar than prostaglandin A1 were shown; two of them were still active as growth inhibitors. This biotransformation was not observed with normal cells like L 929 or chick embryo fibroblasts, for which prostaglandin A1 had no inhibitory effect. The identification of these metabolites will allow the study of the structure-activity relationship.     

Stoffwechselprodukte von Mikroorganismen

Aspergillus microcysticus Sappa TÜ 502 produces a formerly unknown antibiotic named asposterol. Extractions of culture filtrates from fermentations with ethyl acetate and subsequent column chromatography of the extracts on silica gel using benzene/ethyl acetate for the elution yielded up to 35 mg/l of practically pure preparations of the antibiotic. Asposterol is lipophilic and instable. The elemental composition was determined as C₂₄H₃₃NO₄, molecular weight 399. The antibiotic exists in two isomeric forms. A steran skeleton is attributed to the compound. There is no evidence for other similarities with known steroid antibiotics. Asposterol inhibits both bacteria and fungi. The minimal inhibitory concentrations are 0.3 μg/ml for Bacillus subtilis and 1.2 μg/ml for Schizosaccharomyces pombe. The inhibitory effect of asposterol is compensated by α-tocopherol. It is assumed that the cytoplasmic membrane is the site of the primary antibiotic action. Linolenic acid, undecylenic acid and other unsaturated fatty acids show similarities with respect to the type of their antimicrobial effects.     

川木香对实验性胃溃疡形成的抑制作用研究

目的:研究川木香对实验性胃溃疡形成的抑制作用.方法:采用利血平型小鼠胃溃疡模型、醋酸型大鼠胃溃疡模型,以雷尼替丁为对照药物,观察动物溃疡指数和溃疡抑制率.结果:川木香单体提取物(去氢木香内酯)、醋酸乙酯提取物、乙醇提取物,抑制利血平型溃疡存在统计学差异(与模型对照组比较,P＜0.01);醋酸乙酯提取物,抑制醋酸型溃疡存在统计学差异(与模型对照组比较,P＜0.05),其中高剂量组作用存在统计学差异(与模型对照组比较,P＜0.01).结论:川木香具有抑制实验性胃溃疡的形成作用,醋酸乙酯提取物可以作为川木香抑制胃溃疡形成的有效部位.     

Siderophore containing 2,3-dihydroxybenzoic acid and threonine formed by Rhizobium trifolli

An iron-binding compound was isolated from ethyl acetate extract of culture supernatant fluid of Rhizobium trifolii AR6 and was purified by iron-exchange chromatography. The compound was characterized by UV and IR. It contained 2,3-dihydroxy-benzoic acid and threonine and was accumulated during stationary phase of growth in iron-deficient media. Synthesis of the siderophore was repressed by FeCl3. In iron limited medium the compound promoted growth of R. trifolii strains.     

Mycorrhizal growth in pure cultures in the presence of pesticides

The effects of pesticides on 64 ectomycorrhizal fungi of boreal forest trees were studied in vitro. The pesticides (fungicides: benomyl, chlorothalonil, copper oxychloride, maneb and propiconazole; herbicides: chlorthiamid, glyphosate, hexazinone, linuron and terbuthylazine; insecticide: cypermethrin) were selected as those commonly used in Nordic forest nurseries and afforestation sites. In general, the fungicides proved to be more toxic to ectomycorrhizal fungi than the herbicides and cypermethrin. The fungicides, chlorothalonil and propiconazole, had the clearest inhibitory effect on growth of mycorrhizal fungi. Conversely, maneb, glyphosate and terbuthylazine stimulated the growth of some mycorrhizal fungi. Leccinum versipelle and L. scabrum, Paxillus involutus and Cenococcum geophilum were the most sensitive ectomycorrhizal fungi to the various pesticides.     

Tolerance and biodegradation of m-toluate by Scots pine, a mycorrhizal fungus and fluorescent pseudomonads individually and under associative conditions

The tolerance to, and degradation of m-toluate by Scots pine (Pinus sylvestris), a symbiotic mycorrhizal fungus (Suillus bovinus) and Pseudomonas fluorescens strains, with or without m-toluate-degrading capacity, was determined individually and in all symbiotic/associative plant-microbe combinations. Fungal survival on medium with m-toluate was increased in co-culture with the degradative bacterial strains on agar plates (up to 0.02%, w/v). When fungi were grown in mycorrhizal association with Scots pine seedlings in test-tube microcosms containing expanded clay pellets and growth media, the fungus was able to withstand m-toluate concentrations up to 2.0%, w/v in all treatments. The seedling tolerance remained unaltered regardless of the presence or absence of mycorrhizal fungi or biodegradative bacteria. Reduction in m-toluate levels was only detected in treatments inoculated with bacterial strains harbouring TOL catabolic plasmids. The plant and fungus, alone or in mycorrhizal symbiosis, were unable to cleave m-toluate. The presence of easily available plant-derived carbon sources did not impede m-toluate degradation by the bacteria in the mycorrhizosphere.     

Investigations of aneuploidy-inducing chemical combinations in Saccharomyces cerevisiae

For several years we have been investigating combinations of chemicals for their ability to induce aneuploidy. Earlier published results indicated that combinations of certain chemicals showed a potentiation effect while other combinations did not. We have continued to explore this phenomenon and report additional findings in this communication. Combinations of ethyl acetate and methyl ethyl ketone showed a potentiation effect as did 1-methyl-2-pyrrolidinone-nocodazole combinations. Combinations that did not show a potentiation effect were 2-pyrrolidinone-nocodazole and 1-methyl-2-pyrrolidinone-ethyl acetate. We also found that nocodazole, which is a potent inducer of aneuploidy in yeast extract-peptone-dextrose (YEPD) medium but not in synthetic complete (SC) medium, showed a potentiation effect with ethyl acetate in SC medium. This effect in SC medium is similar to that previously reported for nocodazole with ethyl acetate in YEPD medium. When nocodazole was dissolved in 1-methyl-2-pyrrolidinone as a concentrated stock solution, a potentiation effect occurred even at low concentrations of the solvent.     

中华羊茅内生真菌Epichlo? sp.挥发性物质的抑菌活性及成分分析

为测定中华羊茅内生真菌Epichloe sp.挥发性物质的抑菌活性和成分,依次采用挥发油提取器和乙酸乙酯萃取Epichloe sp.菌株的发酵产物,再采用琼脂扩散法、悬滴法和滤纸片扩散法测定其抑菌活性;基于顶空固相微萃取和气相质谱联用法分析挥发性物质成分。结果表明:挥发性物质、乙酸乙酯提取物、水相部分和发酵液可以抑制小麦根腐离蠕孢(Bipolaris sorokiniana)、新月弯孢(Curvularia lunata)和黑曲霉(Aspergillus niger)菌丝生长。高浓度的挥发性物质对四种真菌孢子萌发和芽管长度抑制效果显著高于低浓度(P<0.05)。结果表明,随着挥发性物质浓度的增加对金黄色葡萄球菌(Staphylococcus aureus)和大肠杆菌(Escherichia coli)的抑菌作用增强,同一挥发性物质浓度对金黄色葡萄球菌的抑制作用强于大肠杆菌。此外,从挥发性物质中鉴定出32种具有较高质谱匹配度的化合物,其中主要成分包括异山梨醇(16.26%)、甲基叔丁基醚(8.18%)、二氢甘露醇(3.75%)、羟基丙酮(2.99%)、乙酸(1.22%)、己醇(1.20%)、3-糠醛(0.92%)、5-羟甲基二氢呋喃-2-酮(0.86%)、5-羟基戊酸-2,4-二叔丁基苯酯(0.86%)和棕榈酸甲酯(0.85%)。总之,中华羊茅内生真菌Epichlosp.挥发性物质具有拮抗细菌和真菌的作用。     

Influence of glucose and oxygen on the production of ethyl acetate and isoamyl acetate by a <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> strain during alcoholic fermentation

The effect of glucose and dissolved oxygen in a synthetic medium simulating the standard composition of grape juice on the production of ethyl acetate and isoamyl acetate by a Saccharomyces cerevisiae strain during alcoholic fermentation was studied. The specific in vitro activity of alcohol acetyltransferase (AATase, EC 2.3.1.84) and esterases (ESase, EC 3.1.1.1; hydrolysis and synthesis of esters) in cell-free extracts was also examined. The specific activity of AATase for ethyl acetate was found to peak at the beginning of the exponential growth phase and that for isoamyl acetate at its end. While the glucose concentration only affected the maximum specific activity of AATase, and only slightly, oxygen inhibited such activity, to a greater extent for isoamyl acetate than for ethyl acetate. On the other hand, esterases were found to catalyse the synthesis of ethyl acetate only at a low or medium glucose concentration (50 or 100 g l^-1, respectively), and to reach their maximum hydrolytic activity on isoamyl acetate during the stationary growth phase. The highest ethyl acetate and isoamyl acetate concentrations in the medium were obtained with a glucose concentration of 250 g l^-1 and semianaerobic conditions.     

Regulation of chlorophyll synthesis in the green alga golenkinia

Chlorophyll synthesis in Golenkinia is inhibited 10-fold by growth in darkness on acetate or by growth on elevated concentrations of acetate in the light, particularly if the growth medium contains low levels of nitrogen. Glucose has no such inhibitory effect. delta-Aminolevulinic acid, with a maximal effect at 0.01 m, but not its precursors, overrides the inhibitory effect of acetate and darkness, restoring chlorophyll synthesis. Glycine, succinate, and alpha-ketoglutarate, the precursors tested, all enter the cell. Cells forming chlorophyll produce significantly more aminolevulinic acid than do cells becoming bleached, further indicating the important regulatory role of this compound. Cyclic AMP has no effect on chlorophyll synthesis. These results are compared with those obtained studying other algae, and a mechanism relating light and acetate to chlorophyll formation is proposed.     

Stimulation of some litter-decomposing basidiomycetes by shikimic acid

Shikimic acid, which constitutes 1.5-2.5% of the dry matter in needles of Scots pine, was found to stimulate the growth of various litter-decomposing basidiomycetes of the genera Marasmius, Mycena and Xeromphalina in a synthetic nutrient medium. Out of eighteen litter-decomposing species, ten were stimulated by shikimic acid, wheras the eight mycorrhizal and four wood-rotting species tested were not affected. Maximal effect was obtained at a concentration of ca 2 m M . Growth experiments at varying pH-values indicated active uptake of shikimic acid. Even in the presence of aromatic amino acids, shikimic acid stimulated the growth of the fungi. In certain species the strong inhibiting effect of phenylalanine, tyrosine and tryptophan, when added simultaneously, was reversed in the presence of shikimic acid. Fungi which were stimulated by shikimic acid were also able to use this compound as their sole carbon source. Maximal stimulating effect of shikimic acid occurred when glucose had been added at optimal concentration.     

Mycorrhizal fungi of Vanilla: diversity, specificity and effects on seed germination and plant growth

Mycorrhizal fungi are essential for the germination of orchid seeds. However, the specificity of orchids for their mycorrhizal fungi and the effects of the fungi on orchid growth are controversial. Mycorrhizal fungi have been studied in some temperate and tropical, epiphytic orchids, but the symbionts of tropical, terrestrial orchids are still unknown. Here we study diversity, specificity and function of mycorrhizal fungi in Vanilla, a pantropical genus that is both terrestrial and epiphytic. Mycorrhizal roots were collected from four Vanilla species in Puerto Rico, Costa Rica and Cuba. Cultured and uncultured mycorrhizal fungi were identified by sequencing the internal transcribed spacer region of nuclear rDNA (nrITS) and part of the mitochondrial ribosomal large subunit (mtLSU), and by counting number of nuclei in hyphae. Vanilla spp. were associated with a wide range of mycorrhizal fungi: Ceratobasidium, Thanatephorus and Tulasnella. Related fungi were found in different species of Vanilla, although at different relative frequencies. Ceratobasidium was more common in roots in soil and Tulasnella was more common in roots on tree bark, but several clades of fungi included strains from both substrates. Relative frequencies of genera of mycorrhizal fungi differed significantly between cultured fungi and those detected by direct amplification. Ceratobasidium and Tulasnella were tested for effects on seed germination of Vanilla and effects on growth of Vanilla and Dendrobium plants. We found significant differences among fungi in effects on seed germination and plant growth. Effects of mycorrhizal fungi on Vanilla and Dendrobium were similar: a clade of Ceratobasidium had a consistently positive effect on plant growth and seed germination. This clade has potential use in germination and propagation of orchids. Results confirmed that a single orchid species can be associated with several mycorrhizal fungi with different functional consequences for the plant.     

Plant-plant interactions vary with different mycorrhizal fungus species

Because different species of mycorrhizal fungi have different effects on the growth of particular plant species, variation in mycorrhizal fungus species composition could cause changes in the strength of plant-plant interactions. Results are presented from a growth chamber experiment that compared the strength of interactions among seedlings of ponderosa pine (Pinus ponderosa) when the pines were colonized by two different groups of ectomycorrhizal fungi in the genus Rhizopogon. Plant density effects differed between the two groups of mycorrhizal fungi: plant growth was low regardless of density when plants were colonized with pine-specific Rhizopogon species, while plant growth declined with plant density when plants were colonized by Rhizopogon species having a broader host range. This result parallels results from previous studies showing that plant interactions are more antagonistic with mycorrhizal fungi than without, implying that plant responsiveness to beneficial mycorrhizal fungi declines with increasing plant density. If such effects are prevalent in plant communities, then variation in mycorrhizal fungus community composition is predicted to have a density-dependent effect on plants.     

Evidence for Naturally Occurring Inhibitors of Archegonial Differentiation in Lygodium japonicum

Archegonial differentiation in prothallia of Lygodium japonicum was inhibited when the filtrate of conditioned medium or the extracts of prothallia with organic solvents were added to the medium. By varying the timing of treatment with the methanol extract, archegonial differentiation was shown to start at least 4 days before microscopically detectable change. The inhibitory effect of methanol extract was nullified by transferring the treated plants to a fresh medium omitting the methanol extract, so that the archegonial formation became discernible 6 days after the transfer. The inhibitory activity was stable in both acidic and basic solutions at room temperature, and was partially lost by boiling at pH 3 or 11 for 30 min. The inhibitor, which could be retrieved from the filtrate and the methanol extract, was fractionated into the neutral ethyl acetate fraction, but was not found in the acidic ethyl acetate fraction and in the aqueous residue. At least two active zones were separated on thin layer chromatograms of the ethyl acetate extracts from the filtrate and the methanol extract, and the relative flow-rates of each active zone from these two sources were very similar. The evidence described above indicates that specific inhibitors of archegonial differentiation may be produced in the tissue of prothallia of Lygodium and eventually be secreted to the medium.     

Formation of ethyl acetate by Kluyveromyces marxianus on whey: Influence of aeration and inhibition of yeast growth by ethyl acetate

The ability of the yeast Kluyveromyces marxianus to convert lactose into ethyl acetate offers good opportunities for the economical reuse of whey. The formation of ethyl acetate as a bulk product depends on aerobic conditions. Aeration of the bioreactor results in discharge of the volatile ester with the exhaust gas that allows its process‐integrated recovery. The influence of aeration (varied from 10 to 50 L/h) was investigated during batch cultivation of K. marxianus DSM 5422 in 0.6 L whey‐borne medium using a stirred reactor. With lower aeration rates, the ester accumulated in the bioreactor and reached higher concentrations in the culture medium and the off gas. A high ester concentration in the gas phase is considered beneficial for ester recovery from the gas, while a high ester concentration in the medium inhibited yeast growth and slowed down the process. To further investigate this effect, the inhibition of growth by ethyl acetate was studied in a sealed cultivation system. Here, increasing ester concentrations caused a nearly linear decrease of the growth rate with complete inhibition at concentrations greater than 17 g/L ethyl acetate. Both the cultivation process and the growth rate depending on ethyl acetate were described by mathematical models. The simulated processes agreed well with the measured data.     

Effect of onion (Allium cepa) root exudates on the hyphal growth of Gigaspora margarita

 The effect of root exudates from onions differing in P status on spore germination and hyphal growth of arbuscular mycorrhizal fungi was investigated. Onion (Allium cepa) was grown in solution culture at different phosphorus concentrations (0, 0.1, 1.0, 8.0 and 24.0 mg P l^–1) and root exudates were collected. When spores of the arbuscular mycorrhizal fungus, Gigaspora margarita were incubated with these root exudates, spore germination was only slightly affected but hyphal growth was greatly affected, particularly with exudates from P-deficient plants. This suggests that the P nutrition of host plants influences the composition of root exudates and thereby the hyphal growth of arbuscular mycorrhizal fungi. Accepted: 25 June 1995     

Effect of the activity of primary proton pumps on the growth of Escherichia coli in the presence of acetate

Escherichia coli batch cultures were grown under aerobic and anaerobic conditions on glucose with the substrate addition at pH 7.0. The cultures accumulated acetate in the medium at concentrations sufficient to inhibit the growth. This inhibitory effect of acetate was mediated apparently via its action on the intracellular pH. The inhibition of E. coli growth by acetate increased when the redox proton pump was switched off in the course of transition from aerobiosis to anaerobiosis and when the regulation of K+ fluxes was disordered in the presence of valinomycin. H+-ATPase was not essentially involved in maintaining the high rate of E. coli growth in the presence of acetate under aerobic conditions. If the activity of H+-ATPase was inhibited under anaerobic conditions at pH 7.0, the growth ceased after the dissipation of ionic gradients on the membrane. When CCCP was added under aerobic conditions, the growth did not stop at once if the medium had a pH of 7.6, but ceased immediately at pHout 7.0 in the glucose-salt medium.     

Inhibition of the synthesis of cell wall polysaccharides in oat coleoptile segments by jasmonic acid: Relevance to its growth inhibition

The inhibitory mode of action of jasmonic acid (JA) on the growth of etiolated oat (Avena sativa L. cv. Victory) coleoptile segments was studied in relation to the synthesis of cell wall polysaccharides using [¹⁴C]glucose. Exogenously applied JA significantly inhibited indoleacetic acid (IAA)-induced elongation of oat coleoptile segments and prevented the increase of the total amounts of cell wall polysaccharides in both the noncellulosic and cellulosic fractions during coleoptile growth. JA had no effect on neutral sugar compositions of hemicellulosic polysaccharides but substantially inhibited the IAA-stimulated incorporation of [¹⁴C]glucose into noncellulosic and cellulosic polysaccharides. JA-induced inhibition of growth was completely prevented by pretreating segments with 30 mm sucrose for 4 h before the addition of IAA. The endogenous levels of UDP-sugars, which are key intermediates for the synthesis of cell wall polysaccharides, were not reduced significantly by JA. Although these observations suggest that the inhibitory mode of action of JA associated with the growth of oat coleoptile segments is relevant to sugar metabolism during cell wall polysaccharide synthesis, the precise site of inhibition remains to be investigated.Abbreviations JA jasmonic acid - ABA abscisic acid - IAA indoleacetic acid - T ₀ minimum stress relaxation time - TFA trifluoroacetic acid - TCA trichloroacetic acid - HPLC high-performance liquid chromatography - EtOAc ethyl acetate - TLC thin-layer chromatography - JA-Me methyl jasmonate - GLC-SIM gas-liquid chromatography-selected ion monitoring