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Paeonia ostii is known for its excellent medicinal values as Chinese traditional plant. To date, the diversity of culturable endophytes associated with P. ostii is in its initial phase of exploration. In this study, 56 endophytic bacteria and 51 endophytic fungi were isolated from P. ostii roots in China. Subsequent characterization of 56 bacterial strains by 16S rDNA gene sequence analysis revealed that nine families and 13 different genera were represented. All the fungal strains were classed into six families and 12 genera based on ITS gene sequence. The biosynthetic potential of all the endophytes was further investigated by the detection of putative polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) genes. The PCR screens were successful in targeting thirteen bacterial PKS, five bacterial NRPS, ten fungal PKS and nine fungal NRPS gene fragments. Bioinformatic analysis of these detected endophyte gene fragments facilitated inference of the potential bioactivity of endophyte bioactive products, suggesting that the isolated endophytes are capable of producing a plethora of secondary metabolites. These results suggest that endophytes isolated from P. ostii had abundant population diversity and biosynthetic potential, which further proved that endophytes are valuable reservoirs of novel bioactive compounds.
The aim of this study was to isolate and identify endophytic bacteria from sterns of Chelidonium mqjus L. (greater celandine) and to evaluate their antifungal properties. In total, 34 bacterial endophyte strains were isolated. The fungistatic effects of these bacteria on the growth of five moulds (Alternaría altérnala, Chaetonium sp., Paecilomyces variotti, Byssochlamys fitlva, Aureobasidium pullulans) and one species of black yeast (Exophiala mesophila) were tested. The majority of the bacterial isolates were found to inhibit the growth of fungi and those with the strongest antifungal properties were further characterized. Of the twelve isolates examined, 11 were species of Bacillus thuringiensis and one was Bacillus amyloliquefaciens.
A search for endophytes in Castanea sativa Miller (Fagales: Fagaceae) grafting scions showed that a latent pathogenic fungus Gnomoniopsis smithogilvyi (Diaporthales: Gnomoniaceae) was present as the major component of the endophytic flora. Initially, the goal of this study was to develop a biological control method of Cryphonectria parasitica (Diaporthales: Valsaceae), the chestnut blight agent, by soaking chestnut scions before grafting in antagonists suspension. However, the healthy chestnut material used in in vitro and glasshouse experiments turned out to be naturally infected by a pathogen. At first view, the symptoms looked very similar to those caused by C. parasitica but some differences were noticed. DNA sequencing and application of Koch’s postulates revealed that G. smithogilvyi was the agent responsible of those symptoms. Preventive biocontrol experiments were carried out with chestnut tree scions soaked overnight in a liquid suspension of Bacillus amyloliquefaciens (Bacillales: Bacillaceae). This bacterium was then frequently found in the lower parts of scions (CF of 100% between 3.1 and 6 cm) and up to a height of 18 cm. It was observed that when B. amyloliquefaciens was present, the endophytic and opportunistic pathogenic fungus G. smithogilvyi was not present. Conversely, the parts not colonized by the bacteria were always naturally infected by the endophytic fungus. This would indicate that the endophytic behavior of B. amyloliquefaciens inhibited the growth of G. smithogilvyi and reduced its presence in scions. A similar experiment, carried out with the Trichoderma atroviride (Hypocreales: Hypocreaceae), led to similar observations. Trichoderma atroviride was frequently isolated in the lower parts of scions (CF of 100% until 6 cm) and up to a height of 27 cm. Inoculating B. amyloliquefaciens and T. atroviride as part of a preventive biocontrol treatment would allow these biological control agents to colonize the plant as endophytes and prevent the development of G. smithogilvyi.
Effects of 62 isolates (of 41 taxa) of fungi on growth of 1-year-old oak (Quercus robur) seedlings were studied in an inoculation experiment. The fungi were isolated from roots of 80–96-year-old Q. robur that had been subjected to periodic flooding and had symptoms of oak decline. The fungal genera included Alternaria, Aspergillus, Calonectria, Chaetomium, Cladosporium, Clonostachys, Corynespora, Cylindrocarpon, Dicyma, Geotrichum, Ilyonectria, Isaria, Metarhizium, Oidiodendron, Ophiostoma, Pezicula, Phialocephala, Phialophora, Pyrenochaeta, Sporendocladia, Sporothrix, Thelonectria, Trichoderma and Trimmatostroma. Mycelial colonies of fungi growing in potato-dextrose broth were used for soil inoculation. Plant growth was assessed 2 years after inoculation, when the plants were 3 years old. Stem lengths, and dry weights of stems, roots and leaves were measured. Stem growth was inhibited by 31 isolates (50%) and root growth by 12 isolates (19%). Stem growth was stimulated by two isolates (3%) and root growth by 17 isolates (27%). The overall ratio of inhibitors to stimulants was 2.1. The proportion of taxa that inhibited stem growth was 16 times greater than that which promoted stem growth. The proportion of taxa that promoted root growth was only 1.5 times greater than that which inhibited root growth. The structure of the fungal communities in periodically flooded oak forests suggests that they are more likely to inhibit than to promote vigour in oaks.
Eighty four different fungal endophytes isolated from sea grasses (5), marine algae (36) and leaves or barks of forest trees (43) were grown in vitro and the secondary metabolites secreted by them were harvested by immobilizing them on XAD beads. These metabolites were eluted with methanol and screened using SYBR Green I assay for their antiplasmodial activity against blood stage Plasmodium falciparum in human red blood cell culture. Our results revealed that fungal endophytes belonging to diverse genera elaborate antiplasmodial metabolites. A Fusarium sp. (580, IC50: 1.94 μg ml−1) endophytic in a marine alga and a Nigrospora sp. (151, IC50: 2.88 μg ml−1) endophytic in a tree species were subjected to antiplasmodial activity-guided reversed phase high performance liquid chromatography separation. Purification led to potentiation as reflected in IC50 values of 0.12 μg ml-1 and 0.15 μg ml−1 for two of the fractions obtained from 580. Our study adds further credence to the notion that fungal endophytes are a potential storehouse for a variety of novel secondary metabolites vested with different bioactivities including some that can stall the growth of the malaria parasite.
The present investigation was carried out to analyse the presence of endophytes in the above-ground parts of Hipericum perforatum and to analyse the biodiversity and enumeration of epiphytes. Plant material was collected in Poland three times during the growing season. Phenotypic and genotypic diversity of all the endophytes and the most abundant epiphytes were researched. We analysed fungistatic activity of this isolates. From the endosphere of tested plant Alcaligenes faecalis and Bacillus licheniformis were isolated. The most numerous epiphytes were the copiotrophs and a bit less numerous were oligotrophs, bacteria cultivated on Bunt and Rovir’s medium and fungi. The least numerous bacteria were Azotobacter sp. Among all the molds dominant were: Cladosporium herbarum, C. cladosporioides and Alternaria consortialis, A. alternata, Clonostachys rosea f. catenulata (Gliocladium catenulatum), Scopulariopsis brevicaulis and Penicillium terrestre. Among phyllobacteria there were found mostly the following species: Burkholderia cepacia, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas fluorescens, P. putida, Pantoea agglomerans, Paenibacillus polymyxa, Bacillus cereus, Rhodococcus sp., R. erythropolis and Cellulosimicrobium cellulans. The broadest spectrum of antifungal activity was examined for the following species: Paenibacillus polymyxa, Pseudomonas putida and Pantoea agglomerans. P. polymyxa limited the growth of over 82% tested molds, so did the other two strains: P. agglomerans over 77% and P. putida over 73%.
Research on soil microorganisms associated with plant roots (endophytes) and their outside habitats (rhizosphere, soil) under exposure to high concentrations of heavy metals in the environment may contribute to an increased knowledge about the effects of unfavorable pollutants in the natural ecosystem and improve the efficiency of remediation processes of degraded soils. The main goal of our study was to evaluate properties of microbial populations inhabiting different but spatially closed habitats: interior of the tree roots – endophytes (Betula pendula L. and Alnus glutinosa L.), rhizosphere and soil not influenced by plant roots at two heavy metal polluted areas in southern Poland where the content of cadmium, lead and zinc ranged 9.2–14.9, 26.2–75.1 and 1080–1559 mg∙kg–1 of dried soil, respectively. In our experiment, we analyzed their density as well as determined the total metabolic activity and ability for utilization of different carbon sources. The density of endophytes (5.76–7.16 log10 c.f.u. × g dw–1) was higher than the density of bacteria inhabiting outside habitats – the rhizosphere and soil (5.43–5.91 and 5.08–5.57 log10 c.f.u. × g dw–1, respectively) and was not affected by heavy metal content. However, the total metabolic activity of endophytic populations was strongly influenced by the physicochemical properties of the soil and was significantly lower at more contaminated site. In general, the metabolic activity of endophytes was ~15 times lower than the activity of rhizobacteria and at the same level or lower than the activity of soil microorganisms. Discriminant analysis revealed a higher utilization of α-Cyclodextrin and D-Xylose by microorganisms isolated from more polluted test site. The use of an artificial neural network (ANN) followed by principal component analysis (PCA) allowed the distinguishing of preferences in the utilization of carbon sources by microorganisms due to tree species but not the zone of isolation – endophytes, rhizosphere, soil. Our study revealed no significant impact of the metal concentrations present in the soil on the density of microorganisms (bacteria and fungi) inhabiting three analyzed zones (interior of the roots, rhziosphere, soil); however, their total metabolic activity was decreased in more contaminated site. Since the density of endophytes was higher compared to the number of bacteria present in the outside habitats (rhizosphere and soil), and their total metabolic activity was in general lower, we suggest a high specialization of microorganisms associated with the cellular spaces of plant roots compared to other microbial populations.
Molecular diversity studies of 19 rhizobia isolates from chickpea were conducted using simple sequence repeats (SSR) and 16S rDNA-RFLP markers. Phenotypic characterization with special reference to salinity and pH tolerance was performed. These isolates were identified as different strains of Mesorhizobium, Rhizobium, Bradyrhizobium, and Agrobacterium. Twenty SSR loci of Mesorhizobium ciceri, distributed across the other rhizobial genome, clearly differentiated 19 rhizobial isolates. Analogous clustering supported the results of 16S rDNA sequence-based phylogeny. Analysis of the 16S rDNA sequences from M. ciceri strains revealed that nucleotide variables (signature sites) were located at 20 different positions; most of them were present in the first 820 bp region from 5’ terminal. Interestingly, 14 signature sites were located in two main regions, the variable region V1 (nt 527–584), and variable region V2 (nt 754–813). The secondary structure and minimal free energy were determined in these two regions. These results will be useful in characterizing the micro-evolutionary mechanisms of species formation and increase understanding of the symbiotic relationship.
Studies were carried out on the effect of inoculation with Azospirillum brasilense on development and yielding of winter wheat and oat. Plants were grown in the field conditions. In the experiments conducted in randomized block design, three factors were applied: inoculation with an active strain of Azospirillum brasilense, two levels of nitrogen fertilizer and seed treatment with fungicidal seed dressing. It was found, that inoculation of cereals with active strain of Azospirillum brasilense may be a factor increasing their vigour and yielding. Under climatic and soil conditions of Wielkopolska region and despite the fact, that no dinitrogen fixation was observed, cereal inoculation with Azospirillum brasilense contributed to the yield increase up to 27% in wheat and to 6% in oat.
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This is a first part of work devoted highland rush endophytes: Penicillium expansum, Cladosporium oxysporum, Arthrinium state of Apiospora montagnei and Aureobasidium pullulans. The basidiomycete strain, possibly Lagarobasidium detriticum was also isolated.
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Współżycie endofitów bakteryjnych z roślinami

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W pracy przeanalizowano wpływ bakterii endofitycznych na różne gatunki roślin uprawnych. Przedstawiono sposoby zasiedlania tkanek gospodarza przez zespoły endofitów oraz mechanizmy, za pomocą których oddziałują na rośliny. Oddziaływanie to może mieć charakter bezpośredni – poprzez wytwarzanie różnorodnych stymulatorów wzrostu oraz udostępnianie związanego azotu – lub pośredni, kiedy bakterie odgrywają rolę czynników kontroli biologicznej.
The influence of inoculation with Azospirillum brasilense strain, two levels of nitrogen fertilizer, and seed treatment with fungicidal seed dressing on the development acid yield of maize were determined in field experiments. In these studies nitrogenase activity, chlorophyll content in plants, yield size and quality were used as control parameters. It has been found that inoculation of maize crops with an active strain of Azospirillum brasilense has a beneficial effect on maize vigour and yield under the climatic and soil conditions of the Wielkopolska region.
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