Niche specificity and functional diversity of the bacterial communities associated with Ginkgo biloba and Panax quinquefolius.

Plant-associated bacteria can establish mutualistic relationships with plants to support plant health. Plant tissues represent heterogeneous niches with distinct characteristics and may thus host distinct microbial populations. The objectives of this study are to investigate the bacterial communities associated with two medicinally and commercially important plant species; Ginkgo biloba and Panax quinquefolius using high Throughput Sequencing (HTS) of 16S rRNA gene, and to evaluate the extent of heterogeneity in bacterial communities associated with different plant niches. Alpha diversity showed that number of operational taxonomic units (OTUs) varied significantly by tissue type. Beta diversity revealed that the composition of bacterial communities varied between tissue types. In Ginkgo biloba and Panax quinquefolius, 13% and 49% of OTUs, respectively, were ubiquitous in leaf, stem and root. Proteobacteria, Bacteroidetes, Actinobacteria and Acidobacteria were the most abundant phyla in Ginkgo biloba while Proteobacteria, Bacteroidetes, Actinobacteria, Plantomycetes and Acidobacteria were the most abundant phyla in Panax quinquefolius. Functional prediction of these bacterial communities using MicrobiomeAnalyst revealed 5843 and 6251 KEGG orthologs in Ginkgo biloba and Panax quinquefolius, respectively. A number of these KEGG pathways were predicted at significantly different levels between tissues. These findings demonstrate the heterogeneity, niche specificity and functional diversity of plant-associated bacteria.

Changes of volatile organic compounds and bioactivity of Alternaria brassicae GL07 in different ages.

Plant endophytes are rich in secondary metabolites and are widely used in medicine, chemical, food, agriculture, and other fields. Here, an endophytic fungus is isolated from Ginkgo biloba L. leaves and identified as Alternaria brassicae GL07 through genotypic characterizations. It can produce fruity scented volatiles. The analysis of volatile organic compounds (VOCs) was done by gas chromatography-mass spectrometry. A total of 32 components were identified; and at different culture times, the composition of VOCs was different. It had more components in the first two weeks, but a fewer components on the 21st day. More olefins, ketone, aldehyde, and alcohol were found in the growth period and more amines and esters were found in the decline period. Also, 2,5-dihydroxyacetophenone, ß-ionone, and nonanal were found. They were the same ingredients in Ginkgo essential oils and some of them were isolated for the first time in the volatile constituents of endophytes. The antioxidant activity and whitening activities of all extracts were also evaluated. When cultured for 10 days, it had the strongest 2,2-diphenyl-2-picrylhydrazyl radical (IC50 , 0.56 g/L), hydroxyl radical scavenging ability (IC50 , 0.47 g/L), reducing ability, and tyrosinase inhibition ability (IC50 , 5.18 g/L), which may be due to a large amount of ketones and alcohols produced during the log phase. This demonstrates the potential of A. brassicae GL07 to produce bioactive compounds and to be used for perfume and cosmetic industries.

An endophytic Fungi of Ginkgo biloba L. produces antimicrobial metabolites as potential inhibitors of FtsZ of Staphylococcus aureus.

A total of 58 fungal isolates, belonging to 24 genera, were obtained from the leaves, stems and roots of Ginkgo biloba L.. Among them, one endophytic fungal strain, Penicillium cataractum SYPF 7131, displayed the strongest antibacterial activity. Four new compounds (1-4) were isolated from the strain fermentation broth together with four known compounds (5-8). These structures were determined on the basis of 1D and 2D NMR and [Rh2(OCOCF3)4]-induced electronic circular dichroism (ECD) spectroscopic analyses. All the isolated compounds were screened for their in vitro antimicrobial activities. Compound 3 and 4 showed moderate inhibitory activity against Staphylococcus aureus. Compound 7 exhibited significant inhibitory activity against S. aureus with MIC value of 10 µg/mL. Further, the in silico molecular docking studies of the active compounds was used to explore the binding interactions with the active site of filamentous temperature-sensitive protein Z (FtsZ) from Staphylococcus aureus. The docking results revealed that compounds 3, 4 and 7 showed high binding energies, strong H-bond interactions and hydrophobic interactions with FtsZ from S. aureus validating the observed antimicrobial activity. Based on antimicrobial activities and docking studies, compounds 3, 4 and 7 were identified as promising antimicrobial lead molecules.

A Bilobalide-Producing Endophytic Fungus, Pestalotiopsis uvicola from Medicinal Plant Ginkgo biloba.

For screening bilobalide (BB)-producing endophytic fungi from medicinal plant Ginkgo biloba, a total of 57 fungal isolates were isolated from the internal stem, root, leaf, and bark of the plant G. biloba. Fermentation processes using BB-producing fungi other than G. biloba may become a novel way to produce BB, which is a terpene trilactones exhibiting neuroprotective effects. In this study, a BB-producing endophytic fungal strain GZUYX13 was isolated from the leaves of G. biloba grown in the campus of Guizhou University, Guiyang city, Guizhou province, China. The strain produced BB when grown in potato dextrose liquid medium. The amount of BB produced by this endophytic fungus was quantified to be 106 µg/L via high-performance liquid chromatography (HPLC), substantially lower than that produced by the host tissue. The fungal BB which was analyzed by thin layer chromatography (TLC) and HPLC was proven to be identical to authentic BB. The strain GZUYX13 was identified as Pestalotiopsis uvicola via morphology and ITS rDNA phylogeny. To the best of our knowledge, this is the first report concerning the isolation and identification of endophytic BB-producing Pestalotiopsis spp. from the host plant, which further proved that endophytic fungi have the potential to produce bioactive compounds.

Improving flavonoid extraction from Ginkgo biloba leaves by prefermentation processing.

This paper presents a prefermentation treatment method involving fungi to improve flavonoid extraction from the leaves of Ginkgo biloba . The fungi employed for this treatment were screened from the soil present under an ancient ginkgo tree. Seventy-six strains belonging to 23 genera were isolated and identified by a molecular identification method employing 18S rDNA sequences. Thirty-three strains grew well using ginkgo leaves as the growth medium. One strain, Gyx086, with higher extracted yield of flavonoids and more similar to the control, was finally selected for prefermentation processing. The major fermentation factors were optimized by response surface methodology. The optimal conditions for the highest total falvonoid yield were 27.8 °C for temperature, 64.2% for moisture content, and 61 h for fermentation time. Under the optimal condition, a actual total flavonoid yield of 27.59 ± 0.52 mg/g dry weight culture sample was obtained, which was about 70% higher than that of unfermented gingko leaf samples.

New flavonol and diterpenoids from the endophytic fungus Aspergillus sp. YXf3.

One new flavonol, chlorflavonin A (1), four new diterpenoids, aspergiloids E-H (3, 5-7), together with eight known compounds (2, 4, 8-13) were isolated from solid fermentation of Aspergillus sp. (strain no. YXf3), an endophytic fungus from Ginkgo biloba. Their structures were determined through detailed spectroscopic analysis combined with comparison of NMR spectra data with reported ones. All of them were screened on cytotoxicity against KB, SGC-7901, SW1116, and A549 cell lines; compounds 4, 9-11 exhibited moderate activities with IC50 values ranging from 6.74 to 46.64 µM.

Ginkgolide B produced endophytic fungus (Fusarium oxysporum) isolated from Ginkgo biloba.

To screen the presence of ginkgolide B-producing endophytic fungi from the root bark of Ginkgo biloba, a total of 27 fungal isolates, belonging to 6 different genus, were isolated from the internal root bark of the plant Ginkgo biloba. The fungal isolates were fermented on solid media and their metabolites were analyzed by TLC. The obtained potential ginkgolides-producing fungus, the isolate SYP0056 which was identified as Fusarium oxysporum, was successively cultured in the liquid fermentation media, and its metabolite was analyzed by HPLC. The ginkgolide B was successfully isolated from the metabolite and identified by HPLC/ESI-MS and (13)C-NMR. The current research provides a new method to produce ginkgolide B by fungal fermentation, which could overcome the natural resource limitation of isolating from the leaves and barks of the plant Ginkgo biloba.

Two compounds from the endophytic Colletotrichum sp. of Ginkgo biloba.

Two compounds, apigenin-8-C-beta-D-glucopyranoside and 2-(hydroxymethylthio)ethanol, were extracted from the fermentation products of a strain of endophytic fungus, Colletotrichum sp. NTB-2, isolated from the leafstalk of Ginkgo biloba. The structures of the two compounds were determined on the basis of extensive spectroscopic analysis, including 1D and 2D NMR spectral data. The compounds wereobtained from microorganisms for the first time.

[Endophytic fungi from Ginkgo biloba and their biological activities].

OBJECTIVE: To research the isolation method, identification and screen for bioactivities endophytic fungi from ginkgo. METHOD: Endophytic fungi from ginkgo were separated. By means of microdilution method, activities of endophytes against pathogenic fungi were tested. Then, using DPPH, the antioxidant activities were measured. RESULT: Nine strains (16.1%) showed antifungal activities against Candida albicans, Cryptococcus neoformans, Trichophyton rubrum and Aspergillus fumigatus. Among these bioactive strains, the growth of T. rubrum was strongly inhibited by T-1-2-1, as the MIC80 was equal to fluconazole, the positive control. Five strains (8.9%) showed antioxidant activities. Among them sample T-3-2-2 and T-6-5-7 showed the strongest antioxidant activities. CONCLUSION: Endophytic fungi of ginkgo would be potential and rich resources for drug development.

Cytotoxic metabolites produced by Alternaria no.28, an endophytic fungus isolated from Ginkgo biloba.

From the medicinal plant Ginkgo biloba the fungal endophyte Alternaria no.28 was isolated. Extract of the fungus grown in liquid culture media exhibited marked cytotoxic activity when tested in vitro against brine shrimp (Artemia salina). Eight compounds were isolated from the extract of cultures of this endophytic fungus and were elucidated as alterperylenol (1), altertoxin I (2), alternariol (3), alternariol monomethyl ether (4), tenuazonic acid (5) and its derivative (6), together with ergosterol and ergosta-4, 6, 8, 22-tetraen-3-one by means of spectroscopic analysis. Among them, both 5 and 6 showed significant cytotoxic effects in the brine shrimp bioassy, with mortality rates of 73.6% and 68.9%, respectively, at a concentration of 10 microg x mL(-1), and they were first isolated from endophytic fungi.

Bioactive metabolites produced by Chaetomium globosum, an endophytic fungus isolated from Ginkgo biloba.

A novel cytotoxic chlorinated azaphilone derivative named chaetomugilin D (1), together with three known metabolites, chaetomugilin A (2), chaetoglobosins A (3) and C (4), has been isolated by a bioassay-guided fractionation from the EtOAc extract of the cultures of Chaetomium globosum, an endophytic fungus found in the leaves of Ginkgo biloba. Structure of 1 was established by analyses of spectroscopic methods, including 2D-NMR experiments (COSY, NOESY, HMQC, and HMBC). Compounds 1-4 displayed significant growth inhibitory activity against the brine shrimp (Artemia salina) and Mucor miehei.