The effect of culture medium composition and incubation time on synthesis of gibberellin-like substances by bacteria isolated from the roots of pine seedlings (Pinus silvestris L.)

It was found that synthesis of gibberellin-like substances by ten strains of Coryneform bacteria isolated from the roots of pine seedlings depended on both the composition of the medium and incubation time. More of these substances were produced in mineral medium with glucose in complex medium with casamino acids and yeast extract. Most gibberellin-like substances were found in 7 or 14-day old cultures. Culture supernatant fluids of most of the bacteria tested contained several gibberellin-like substances which on chromatograms run with the solvent system benzene, acetic acid (10:3, v/v) were located at Rf 0.0-0.3; 0.4-0.6 and 0.8-1.0.     

Synthesis of auxins by fungi isolated from the roots of pine seedings (Pinus silvestris L.) and from soil.

Synthesis of auxins by fungi grown with and without tryptophan has been studied. 26 out of 30 fungal strains produced detectable amounts of auxins in tryptophan contain media. 18 strains produced but very small amounts of auxins in media without this amino acid. By means of paper chromatography, chromogenic reagents and biotest three active substances could be distinguished. They were found on the chromatograms run with isopropanol, ammonia, water (10:1:1 v/v) at Rf 0.05--0.2, 0.3--0.5 and 0.8--1.0. Most strains produced active substances with Rf 0.3--0.5.     

Production of auxins by bacteria isolated from the roots of pine seedlings (Pinus silvestris L.).

Qualitative and quantitative studies were carried out on the production of auxins by Coryneform bacteria, the only bacterial types isolated from roots of pine seedlings. Almost all isolates were capable of producing auxins in tryptophan containing media. In media without this amino acid only trace or no auxins were produced. Most of the bacteria studied synthesized auxins located on the chromatograms run with isopropanol, ammonia, water (10:1:1 v/v) at Rf 0.3--0.5. Moreover substances with Rf values 0.05--0.2 and 0.8--1.0 were produced by some strains. No plant growth inhibitors detected with the Avena coleoptiles biotest were produced by the bacteria studied.     

Decomposition of organic matter by the ericoid mycorrhizal endophytes of Formosan rhododendron (Rhododendron formosanum Hemsl.)

Ericoid mycorrhizas are associated with a number of host plants in the Ericaceae in high-elevation regions of Taiwan. The ability of these microorganisms to thrive in harsh environmental conditions in the regions implies their capability of decomposing plant organic matter (raw humus). The objective of this study was to investigate the decomposition characteristics of three ericoid mycorrhizal endophytes isolated from the roots of Formosan rhododendron (Rhododendron formosanum Hemsl.). Molecular analysis indicated that strains Rf9 and Rf32 belong to the genus Cryptosporiopsis while strain Rf28 is a member of the genus Phialocephala. Mycorrhizal synthesis experiment showed that the roots of synthesized seedlings produced hyphal coils, a characteristic of ericoid mycorrhiza. Decomposition ability analysis revealed that strains Rf28 and Rf32 had the highest rates of decomposition of organic matter (up to 10.4% after 70 days) while the value for strain Rf9 was about 6.8%. Consistently, these strains secreted extracellular oxidases when cultured on tannic acid medium. Enzyme assay revealed that strains Rf28 and Rf32 secreted peroxidase, laccase, tyrosinase, and cellulase, but strain Rf9 secreted mainly peroxidase and tyrosinase. Apparently, the differences in secreted hydrolytic enzymes among the three endophytes are related to their ability to decompose organic matter. In the mycorrhizal synthesis experiment, all inoculated seedlings survived in the organic matter substrate for 70 days and exhibited a stronger vigor than the control. This study demonstrated that these three isolated endophytes, Rf9, Rf28, and Rf32, are ericoid mycorrhizal fungi, capable of forming ericoid mycorrhiza with Formosan rhododendron. Meanwhile, all three endophytes can secrete hydrolytic enzymes to decompose organic matter for growth, presumably a prerequisite for the adaptation of Formosan rhododendron to the harsh environments of high elevation.     

Production of auxins and gibberellin-like substances by mycorrhizal fungi,bacteria and actinomycetes isolated from soil and the mycorrhizosphere of pine (Pinus silvestris L.)

Summary Production of auxins and gibberellin-like substances by mycorrhizal fungi, bacteria and actinomycetes isolated from the mycorrhizosphere of Scots pine was studied. Chromatography and biossays were used.Most of the organisms required tryptophan for auxins production. The highest biological activity exhibited substances located at R_f 0.2–0.4.The organisms produced minute amounts of gibberellin-like substances which appeared at different R_f values. It was stated that auxins production is much more common among the root zone organisms of pine than the production of gibberellin-like substances.This research was carried out under problem MR.II.16 coordinated by the Institute of Dendrology, Polish Academy of Sciences.     

Synthesis of auxins from tryptophan and tryptophan-precursors by fungi isolated from mycorrhizae of pine (Pinus silvestris L.).

Fungi isolated from mycorrhizae of pine required tryptophan for auxin synthesis. More auxins were found in culture grown with pyrogallol than in those without this compound. The fungi studied produced also auxins from other than tryptophan compounds. Indole employed with serine was more suitable for the production of auxins than indole or anthranilic acid used separately. The active compounds showing auxin activity were located on the chromatograms at Rf 0.2--0.4 and 0.3--0.5 with the solvent system isopropanol, ammonia, water (10:1:1 v/v).     

The Gibberellins of Developing Bean Seeds

Properties of gibberellin-like substances extracted from Frenchbean seeds during their development were studied. Two zonesof gibberellin-like activity were detected on paper chromatograms. Changes in activity of one of the zones correlated with changesin growth-rate of the seed. From considerations of Rf on paperchromatograms, and differential activity on dwarf peas, dwarf-1and dwarf-5 corn, it was deduced that activity of this zonewas due to substances resembling gibberellin A₁ and gibberellinA₅. Gibberellin A₅-like activity was highest in young seedsand disappeared after cell division in the cotyledons had ceased.Gibberellin A₁-like activity rose to its highest level duringthe period of rapid cell expansion in the cotyledons, afterthe disappearance of gibberellin A₅-like activity. The second zone of gibberellin-like activity was due mainlyto a non-acidic substance, which disappeared at the time GAj-likeactivity was rising to its highest level. A non-acidic substance that stimulated lateral bud growth ofdwarf peas also was detected in the extracts. It is presumedto be a cytokinin.     

Induced defence responses limit Hartig net formation in ectomycorrhizal birch roots

Roots of clonal birches ( Betula pendula ) were inoculated with the ectomycorrhizal fungi Paxillus involutus (isolates P0 and Mi) and Hebeloma cylindrosporum (strains D1 and D105). These fungi showed different rates of mycorrhiza formation in vitro . Mature mycorrhizas were obtained after only 2–4 d with H. cylindrosporum , whereas 6–8 d were necessary with P. involutus isolate P0, and P. involutus isolate Mi was not able to form mature mycorrhiza during the 10 d of the experiment. Temporal changes in PAL activity and the expression of genes encoding intracellular pathogenesis-related proteins were followed after inoculating birch roots with these fungi. Transient increase of PAL activity, and transient induction of expression of the wound-inducible Bet v1-SC1 gene, were observed in roots challenged with both H. cylindrosporum strains and the P. involutus isolate P0. These changes were found to coincide with hyphal penetration between root cells during Hartig net formation, and were never observed in roots inoculated with the poorly aggressive P. involutus isolate Mi. Examination of mycorrhizal root sections under u.v. light indicated the presence of phenolic compounds in the host cell walls at the vicinity of the Hartig net. These results strongly suggest that hyphal penetration between the root cells triggers a transient defence response which, in turn, could limit Hartig net formation to the outer layer of the root cortex.     

Production of Plant Growth-Regulating Substances by the Vesicular-Arbuscular Mycorrhizal Fungus Glomus mosseae

Glomus mosseae, a representative species of Endogonaceae (Phycomycetes) able to form vesicular-arbuscular mycorrhiza, was investigated for phytohormone production. Spores of G. mosseae were axenically germinated in water, and the resultant mycelial growth was assayed by standard procedures for extracting plant hormones from microbial cultures. Paper partition chromatography and specific bioassays were used to separate and identify plant growth-regulating substances. The microorganism synthesized at least two gibberellin-like substances, one with R_f corresponding in position to authentic gibberellic acid, and four substances with the properties of cytokinins.     

Changes in Levels of Naturally Occurring Gibberellin-like Substances during Germination of Seed of Lycopersicon esculentum Mill.

Naturally occurring gibberellin-like substances possessing acidic,basic, and neutral properties were detected, by paper partitionchromatography, in ethanolic extracts of tomato seed and ofetiolated seedlings after 72 and 116 hours' growth. Dwarf maizemutants of the d-1 and d-5 types, ‘Meteor’ pea seedlingsand young ‘Potentate’ tomato plants were used asbioassay material. Hydrolysis of seed and seedling proteinsby ficin in phosphate buffer, pH 6.2, after removal of ethanol-solublesubstances, liberated more and different ‘bound’gibberellin-like substances. It is suggested that protein hydrolysisduring germination is an important means of liberating thesesubstances at different stages of seedling development. Acidic substances were present in all the extracts prepared,but in general two with Rfs 0.25 and 0.55 in iso-propanol: ammonia:water : : 10:1:1 v/v were differentiated on d-2 and d-5 maizerespectively. Neutral substances in dry seed extracts chromatographedin the same solvent, had Rfs of 0.05, 0.35, and 0.95 and thesewere found only in the ethanolic (‘free’) extracts.They were active on d-1 and d-2 maize and ‘Meteor’pea. Basic gibberellin-like substances with Rfs of 0.05 and0.35 were found in ‘free’ extracts of both dry seedand etiolated seedlings after 116 hours' growth which were activeon d-2 maize only. Two others with Rfs 0.45 and 0.95 were extractedfrom seedlings after 72 hours' growth and these were activeon young ‘Potentate’ tomato plants. It is suggested that certain gibberellin-like substances, capableof reversing dwarfism in test plants, may be responsible formorphogenetic or other responses not involving stem extensionin the parent species. Changes were found in the levels of gibberellin-likesubstances but there was no evidence of changes in levels ofseed inhibitors relative to seed growth substances.     

Endogenous Gibberellins in Floating Plants and Turions of Wolffiella floridana

Endogenous gibberellins from floating plants and in vitro induced turions of Wolffiella floridana (Lemnaceae) were extracted and partially purified. Gibberellin-like activity was detected in two zones of the chromatogram corresponding to Rf 0-0.1 and Rf 0.4-0.5 by dwarf pea bioassay. The active compounds in the two zones have been referred to in this paper as factor I and factor II respectively. There were quantitative differences in the gibberellin-like substances of floating plants and turions. The floating plants contained more of factor I but less of factor II as compared to the turions. The chromatographic behavior of factor I was similar to either gibberellin A₁ and A₃ while the identity of factor II was uncertain.     

Production of fungal and bacterial growth modulating secondary metabolites is widespread among mycorrhiza-associated streptomycetes

ABSTRACT: BACKGROUND: Studies on mycorrhiza associated bacteria suggest that bacterial-fungal interactions play important roles during mycorrhiza formation and affect plant health. We surveyed Streptomyces Actinobacteria, known as antibiotic producers and antagonists of fungi, from Norway spruce mycorrhizas with predominantly Piloderma species as the fungal partner. RESULTS: None of the fifteen Streptomyces isolates inhibited all seven tested mycorrhizal and plant pathogenic fungi (Amanita muscaria, Fusarium oxysporum, Hebeloma cylindrosporum, Heterobasidion abietinum, Heterobasidion annosum, Laccaria bicolor, Piloderma croceum). The growth of only one of the tested fungi, the mycorrhiza-forming fungus Laccaria bicolor, was stimulated by the streptomycetes, and Piloderma croceum was only moderately affected. Bacteria responded to the streptomycetes differently than the fungi. For instance the strain Streptomyces sp. AcM11, which inhibited most tested fungi, was less inhibitory to bacteria than other tested streptomycetes. The determined patterns of Streptomyces-microbe interactions were associated with distinct patterns of secondary metabolite production. Notably, potentially novel metabolites were produced by strains that were less antagonistic to fungi. Most of the identified metabolites were antibiotics (e.g. cycloheximide, actiphenol) and siderophores (e.g. ferulic acid, desferroxiamines). Plant disease resistance was activated by a single streptomycete strain only. CONCLUSIONS: Our results show that the primary characteristic of mycorrhiza associated streptomycetes is to inhibit the growth of fungi and bacteria. In parallel, our study indicates that Streptomyces strains which are not general antagonists may produce previously un-described metabolites.     

The diffusion of gibberellins into agar and water during early germination of Pharbitis nil Choisy

Agar diffusion of imbibed seeds yielded significant amounts of diffusible Gibberellin-like substances. An analysis of the extractable and diffusible gibberellin-like substance, including an analysis of the remaining imbibition water of the seeds, indicated that a significant part of these gibberellin-like substances could be attributed to a net biosynthesis of these substances in the imbibing seeds. At the same time it was found that water diffusion yielded considerably more gibberellin-like activities than comparable agar diffusions i.e. 10 to 12 fold in general.Agar as well as water diffusion showed a temperature effect with regard to the yield of gibberellin-like substances particularly during the first 6 h of diffusion. The yield of these substances is lower at 10°C, and remains lower as shown with consecutive diffusions, in comparison with the yields at 20°C or 30°C.With both agar and water diffusion the sum of activities obtained with consecutive diffusions is always higher, often considerably higher, than equal periods of continuous diffusion which is probably due to inactivation and/or interference of inhibitory substances with the bioassay responses. Finally, water diffusates of both seeds and seedlings of the normal growing cv. Violet of Japanese morning glory contained considerably more gibberellin-like activities than those of the dwarf cv. Kidachi which indicated that normals synthesize more gibberellins than dwarfs.     

Auxin and gibberellin-like substances synthesis by Fusarium isolates pathogenic to corn seedlings

The four isolates studied produced considerable amounts of auxin in media with tryptophan and two of them produced also considerable amounts in tryptophan-free media. The quantity of auxin synthesized increased with decreasing pathogenicity of the fungi. All of them produced also considerable amounts of gibberellin-like substances. There was no clear reaction between the production of the above substances and the pathogenicity.     

Phytochrome controlled gibberellin metabolism in etioplast envelopes

Summary Disruption of preparations of intact etioplasts isolated from wheat (Triticum aestivum L.) leaves in the dark leads to an increase in the level of extractable gibberellin-like (GA-like) activity. Irradiation of disrupted preparations with red light causes an alteration in the Rf of part of the GA-like activity. A study of sub-etioplast fractions prepared by gradient centrifugation indicates that the phytochrome and the GA-like substances are associated principally with the etioplast envelope.Abbreviations GA gibberellin - MOPS N-morpholino-3-propanesulphonic acid - EDTA ethylene diamine tetra-acetic acid - ATP adenosine triphosphate - R red light     

Physiological effects of gibberellic Acid. X. The release of gibberellin-like substances by germinating barley embryos

Barley embryos, completely free from endosperm, were excised from germinating grain at various times and allowed to diffuse into an aqueous medium for varying lengths of time. At the end of this time, the embryos and ambient solutions were separately extracted. Gibberellin-like activity in the extracts was determined with the barley endosperm bioassay using seed from the same variety, harvest and treatment schedule as was employed for the embryo diffusion experiments. Gibberellin-like substances were released by embryos throughout the 60 hour germination period, though at no time during this period could sufficient activity be extracted from the embryos themselves to account for the observed release. Solvent partitioning and chromatography identified at least one major acidic component migrating at an Rf similar to that of GA₃.

It is concluded that the endogenous gibberellin-like substance(s) originates within the embryo during germination, and that the release of this substance(s) is temporally consistent with, and quantitatively sufficient to account for the in vivo endosperm mobilization response syndrome. A gibberellin-like substance is undoubtedly the endosperm mobilizing hormone.

     

Gibberellin-like Substances from Norway Spruce (Picea abies)

By use of methanol extraction, two different consecutive partition procedures, repeated polyvinylpyrrolidone column chromatography, silicic acid partition column chromatography and the dwarf rice, lettuce, and barley half-seed bioassays, several gibberellin-like substances were detected in elongating shoots of Picea abies (L.) Karst. No significant differences in the content of gibberellin-like substances could be detected between juvenile and mature trees. The shoot axes were found to contain fewer gibberellin-like substances than the needles.     

Inhibitor and Auxin Activity in the Avocado Fruit

The wheat coleoptile elongation bioassay was used for determination of growth-promoting and growth-inhibiting substances in avocado (Persea americana Mill.) fruit tissues, during development and maturation. Growth-promoting activity was found in two zones on paper chromatograms developed with isopropanol: ammonia : water (10:1:1 v/v): Rf 0.30–0.50 and Rf 0.8–0.9. Growth inhibiting activity was found in three different zones: “A” Rf 0.0–0.2, “B” (abscisic acid) Rf 0.6–0.8, and “C” (l-acetoxy-2,4-dihyroxy-n-hepta-deca-16-ene) Rf 0.85–1.0. Higher levels of auxins were found in seed tissues than in the mesocarp. No correlation was found between fruit growth rate and level of extractable auxins in the mesocarp. The amount of abscisic acid (ABA) in the mesocarp was constant during fruit growth. A gradual and consistent increase in 1-acetoxy-2,4-dihydroxy-n-hepta-deca-16-ene was found during fruit growth, reaching a maximum when the fruit attained maturity.     

Intracellular colonization of Rhododendron and Vaccinium roots by Cenococcum geophilum, Geomyces pannorum and Meliniomyces variabilis

Four in vitro experiments were set up to verify the colonization potential of ectomycorrhizal (EcM) Cenococcum geophilum FR. (strain CGE-4), saprotrophic Geomyces pannorum (LINK) SIGLER & CARMICHAEL (GPA-1) and a frequent root-associated, potentially ericoid mycorrhiza (ErM)-forming Meliniomyces variabilis Hambleton & Sigler (MVA-1) in roots of Rhododendron and Vaccinium. A typical ErM fungus, Rhizoscyphus ericae (Read) Zhuang & Korf (RER-1), was included for comparison. All fungal strains intracellularly colonized rooted Vaccinium microcuttings: GPA-1 occasionally produced hyphal loops similar to ErM, MVA-1 and RER-1 exhibited a typical ErM colonization pattern. CGE-4 hyphae grew vigorously on and around newly formed roots and rarely penetrated turgescent rhizodermal cells forming intracellular loose loops. Rooting of Rhododendron sp. microcuttings was not promoted by any fungal strain except CGE-4, which also promoted the most vigorous growth of Rhododendron ponticum L. seedlings. The widespread EcM fungus C. geophilum has a potential to colonize non-EcM roots and support their development which may influence overall growth of ericaceous plants. As shown for G. pannorum, structures resembling ErM may be formed by fungi that are to date not regarded as ericoid mycorrhizal.     

Phase Change in Hedera helix L: I. GIBBERELLIN-LIKE SUBSTANCES IN THE TWO GROWTH PHASES

Gibberellin-like substances were extracted from all parts ofthe juvenile and adult growth phases of Hedera helix. A numberof extraction techniques were employed and estimatos of theamount of gibberellin-like activity present were made usingthree bioassays. Gibberellin like substances were present inapproximately equal amounts in leaves at the two growth phasesand appeared to be localized, in the main, in chloroplasts.Apical ‘buds’ of the juvenile phase were shown tocontain higher levels of gibberellin-like substances than theiradult counterparts. Roots, which under normal circumstancesare associated only with the juvenile phase, contained highlevels of extractable gibberellin-like substances.