Diversity of Endophytes in Panax quinquefolium Root / 中国实验方剂学杂志

ObjectiveTo analyze the community structure of endophytes in Panax quinquefolium root and explore the dominant endophytic bacteria and fungi， to provide scientific basis for the establishment of endophytic microbial bank in P. quinquefolium root. MethodInternal Transcribed Spacer （ITS） sequencing and 16S sequencing were performed on six P. quinquefolium root samples collected from Wendeng， Shandong province on PacBio Sequel Ⅱ. ResultA total of 8 phyla， 11 classes， 23 orders， 27 families and 53 genera of endophytic bacteria were identified in P. quinquefolium root， among which an unidentified Burkholderiaceae and an unidentified Rhizobiaceae were dominant. A total of 9 phyla， 23 classes， 35 orders， 43 families and 48 genera of endophytic fungi were identified in P. quinquefolium root， among which an unclassified Helotiales and Pseudogymnoascus were dominant. The community structure of endophytic bacteria revealed that the roots were selectively enriched with nitrogen-fixing bacteria such as unidentified Rhizobiaceae， Bradyrhizobium and Herbaspirillum， which suggested that nitrogen is important for the growth of P. quinquefolium root. The community structure of endophytic fungi indicated that P. quinquefolium in Shandong province might be infected by unclassified Helotiales. ConclusionThere is a rich diversity of endophytic bacteria and fungi in P. quinquefolium root， which provides scientific basis for studying the interaction of the plant with endophytic microorganisms and screening the endophytes to promote the growth of P. quinquefolium root.

Secondary metabolites of endophyte Alternaria sp. W-1 from Laminaria japonica / 药学学报

Six compounds were isolated from the crude extract of the liquid culture of Alternaria sp. W-1 by silica gel column chromatography, Sephadex LH-20 gel column chromatography, and HPLC. They were identified as 6-iso-tricycloalternarene 6a (1), tricycloalternarene 6a (2), tricycloalternarene B (3), uracil (4), 5-methyluracil (5), and lumichrome (6) through HR-MS, NMR and literature comparison. 6-iso-Tricycloalternarene 6a (1) is a new compound which has never been reported in the literature. In cytotoxicity assay, compounds 1-3 showed weak inhibition activity to human hepatoma cell line SMMC-7721 and human gastric cell line SGC-7901.

A new cytochalasan compound from endophyte Chaetomium globosum WQ in Imperata cylindrical / 药学学报

italic>Chaetomium globosum WQ, an endophyte derived from Imperata cylindrical, can produce abundant cytochalasan compounds through solid state fermentation. Based on previous research and guided by 1H NMR spectrum and TLC, a new cytochalasan compound was isolated from the ethyl acetate extract of a solid culture of C. globosum WQ using silica gel column chromatography, gel filtration over Sephadex LH-20 and HPLC. The new compound was characterized as 20-iso-chaetoglobosin E (1) by a combination of spectroscopic (HR-MS, 1D and 2D NMR) analyses.

Promoting tanshinone synthesis of Salvia miltiorrhiza root by a seed endophytic fungus, Phoma herbarum D603 / 中国中药杂志

The interaction of endophytes and host plant is an effective mean to regulate the growth and secondary metabolism of medicinal plants. Here we want to elucidate the effects and mechanism of Phoma herbarum D603 on the root development and tanshinone synthesis in root of Salvia miltiorrhiza by endophyte-plant coculture system. The mycelium of P. herbarum D603 was colonized in the root tissue space, and formed a stable symbiotic relationship with host plant. The in vitro activities analysis showed that the concentration of IAA produced by D603 can reach(6.45±0.23) μg·mL~(-1), and this strain had some abilities of phosphorus solubilization and siderophore production activities. The coculture experiment showed that strain D603 can significantly promote the synthesis and accumulation of tanshinones in the root of S. miltiorrhiza, in which after 8 weeks of treatment with D603, the content of tanshinone Ⅱ_A in the roots reached up to(1.42±0.59) mg·g~(-1). By the qRT-PCR analysis results, we found that D603 could improve the expression levels of some key genes(DXR, DXS, GGPP, HMGR, CPS) of tanshinone biosynthesis pathway in host plant S. miltiorrhiza, but the promoting effect mainly occurred in the early stage of the interaction, and the enzyme activity level decreased in varying degrees of the later stage. In summary, seed-associated endophyte P. herbarum D603 can promote the growth and root development of S. miltiorrhiza by producing hormones, promoting nutrient absorption and siderophore production, and promote the synthesis and accumulation of tanshinones by regulating the expression level of key genes in the synthetic pathway in S. miltiorrhiza.

HMG-CoA Reductase Inhibitory Activity of Leaf-associated Fungi

Background@#Cholesterol is an important biomolecule for cells. However, because of certain lifestyle, the amount of cholesterol can increase beyond what our bodies can consume. Increased intake may result in the accumulation of cholesterol in the blood leading to atherosclerosis that can lead to congestive heart failure. With the use of statins, cholesterol levels are reduced therefore lowering the risk for this disease. It does so by inhibiting the hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase, the first committed enzyme in cholesterol biosynthesis. In light of the increasing cases of hypercholesterolemia, there is a need to discover novel classes of HMG-CoA inhibitors. @*Objective@#The primary objective of this study was to screen extracts of leaf-associated fungi for their capacity to inhibit hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase, the first committed enzyme in cholesterol biosynthesis. @* Methods@#Eight (8) plants were sampled for the fungal epiphytes and endophytes on their leaves. The fungal isolates were identified by sequencing the internal transcribed region (ITS) of the 18S rDNA. All fungal isolates were cultured in 20 flasks containing 300 mL potato dextrose broth at 30°C for 14 days. The culture broths were then subjected to reduction in volume using a rotary evaporator. HMG-CoA reductase inhibition assay was carried out using the ethyl acetate extracts from the culture broths. Reagent grade pravastatin was used as the positive control. @* Results@#There were a total of sixty-six (66) fungal taxa that were tested for their capacity to inhibit the enzyme HMG-CoA reductase. Of the 50 that tested positive, there were 25 families represented and 5 isolates were of uncertain status taxonomically (incertae sedis). There were also nine classes that were represented and the class with the most number of isolates was Sordariomycetes. The highest percentage inhibition was observed in Pestalotiopsis lespedezae with 65.01 % inhibition. The other fungi with notable inhibitory activities were Colletotrichum cymbidiicola (64.16%) and Schizophyllum commune (57.85%). @*Conclusions@#This study may be the first report of production of HMG-CoA reductase inhibitors in the genus Colletotrichum and in the other leaf-associated fungal taxa that tested positive for HMG-CoA reductase inhibition. It is therefore very promising that a novel statin compound, or even a new class of substances with antihypercholesterolemic bioactivity may be discovered from these leaf-associated fungal isolates. The crude extracts of these fungi are prime candidates for downstream fractionation for the subsequent isolation and structure elucidation of the bioactive compound.

Root-fungal associations in plants from home gardens of Tripura, Northeast India

Mycorrhizal association is an integral part of terrestrial ecosystems. The present work was focused to examinearbuscular mycorrhiza (AM), dark septate endophyte (DSE) fungal colonization, and the composition of AMfungi from two home gardens of Tripura in Northeast India. The results reveal eight plants commonly occurringin two sites belonging to seven families. Of the eight plants, dual colonization of AM and DSE fungi wasobserved in seven plants from two sites. A total of 18 AM fungal species were recovered from both the sites.The study reveals a robust composition of AM fungi in the home garden ecosystem. AM fungi isolated fromthese ecosystems confirm their occurrence and these fungi may be beneficial in improving the cultivationpractices in the home garden systems of the region.

Bacterial endophytes from mangrove leaves with antibacterial and enzymatic activities

Aims@#The aim of this study was to screen bacterial endophytes with antibacterial and enzyme activity from mangrove leaves of Indonesia. @*Methodology and results@#Bacterial endophytes were isolated and evaluated for antibacterial activity against five strains of pathogenic bacteria. Enzymatic Index (EI) was measured to evaluate the production of protease, amylase and cellulase. Hemolysin test was performed on Blood Agar and the sensitivity to antibiotic was performed. Bacterial endophyte Strain 1-1 isolated from Bruguiera gymnorrhiza showed strong inhibition against Escherichia coli, Salmonella sp., Staphylococcus aureus and Pseudomonas aeruginosa with inhibition zone of 12.6 ± 1.4, 8.8 ± 4.1, 12.5 ± 2.3 and 8.4 ± 0.9 mm respectively. Isolate 1-16 which isolated from B. gymnorrhiza exhibited antibacterial activity against E. coli and P. aeruginosa, while Isolate 6-10 isolated from Avicennia lanata exhibited strong inhibition on Salmonella sp. (13.1 ± 3.3 mm). All of those three isolates produced protease, non-haemolysin-producing strain and sensitive to gentamicin or kanamycin but resistant to ampicillin, tetracycline and chloramphenicol. Those three isolates were identified based on homology of 16S rDNA sequence. Strain 1-1 and 1-16 were identified as P. aeruginosa, while strain 6-10 identified as S. marcescens. @*Conclusion, significance and impact of study@#This finding was showed the potential endophytic bacteria from Indonesian mangrove plants with antibacterial and enzyme production.

Secondary metabolites from Corynespora cassiicola J9, an endophytic fungus associated with Blumea balsamifera (L.) DC / 药学学报

Chemical investigation on the rice culture of Corynespora cassiicola J9, an endophyte inhabiting in Blumea balsamifera (L.) DC. resulted in isolation of eight compounds, including a new depsidone derivative, corynether C (1), and seven analogues, corynether B (2), corynetherlactone A (3), corynether A (4), diaryl ether (5), corynesidone C (6), corynesidone D (7), and corynesidone A (8). Their structures were deduced based on 1D and 2D NMR spectroscopy, and HR-ESI-MS data. All of the isolated compounds were evaluated for inhibitory activities against Lissorhoptrus oryzophilus Kuschel by the leaf spray assay. Unfortunately, none of them showed inhibitory effects.

Biotransformation and antitumor activity of triterpenoid derivatives from Ardisia gigantifolia / 中草药

Objective: To study the chemical constituents of transformed products by Sphingomonas yabuuchiae GTC 868T (AB071955) and Pectinex Ultra AFP from the saponin of Ardisia gigantifolia. Methods: Transformation products separated by the process of silica gel column, compounds were identified and elucidated by spectral and chemical methods. Their cytotoxicity activities were tested by Cell Counting Kit 8 colorimetric assay. Results: Five triterpenoid saponins were obtained, including 3β-O-{α-L-rhamnopyranosyl-(1→3)-[β-D-xylopyranosyl-(1→2)]-β-D-glucopyranosyl-(1→4)-α-L-arabinopyranosyl}-cyclamiretin A (1), 3β-O-{β-D-glucopyranosyl-(1→4)-[β-D-glucopyranosyl-(1→2)]-α-L-arabinopyranoside}-cyclamiretin A (2), 3β-O-{β-D-glucopyranosyl-(1→2)-α-L-arabinopyranoside}-cyclamiretin A (3), 3-O-α-L-arabinopyranosyl cyclamiretin A (4), and cyclamiretin A (5). Conclusion: Compounds 2-5 are obtained by biotransformation for the first time. Some of the compounds showed certain antitumor activity, among them, compound 2 shows more cytotoxicity activity than Ag3 and positive control.

Investigations on secondary metabolites of endophyte Diaporthe sp. hosted in Tylophora ovata / 中国中药杂志

One new compound ()-2,3-dihydroxybutyl 2-hydroxy-3,5,6-trimethylbenzoate (1) and six known compounds xylariphthalide A (2), convolvulol (3), cis-4-hydroxy-6-deoxytalone (4), phomoxydienes B (5), 5,6-dihydroxy-2,3,6-trimethylcyclohex-2-enone (6), trans-cyclo-(D-tryptophanyl-L-tyrosyl) (7) were isolated from Diaporthe sp., an endophytic fungus hosted in the leaves of the toxic Chinese folk medicine Tylophora ouata, using the combination methods of silica gel column chromatography, medium-pressure ODS column chromatography and RP-preparative HPLC. The structure of compound 1 was elucidated by NMR and MS data analyses. The absolute configurations were established according to the ¹H-NMR data and exciton chirality method. Compound 1 inhibited the activation of human lung fibroblasts MRC-5 cells by 64.0% at 10 μmol·L⁻¹. The MTT assay showed that compounds 2 and 4 displayed cytotoxic activity against human tumor cell lines BGC-823 cells with IC₅₀ values of 1.5 and 8.6 μmol·L⁻¹, respectively.

Antimicrobial activity of Aspergillus sp. IBRL MP15 CCL, an endophytic fungus isolated from Swietenia macrophylla leaf against human pathogens

@#Aims: Endophytes are microorganisms residing in the living tissues of the host plant and may contribute to their hostplant by producing a plethora of bioactive compounds that provide survival value to the plant. This study aimed toevaluate the antimicrobial activity of Aspergillus sp. IBRL MP15 CCL, an endophytic fungus isolated from Swieteniamacrophylla leaf.Methodology and results: The antimicrobial activity was evaluated with disc diffusion and a colorimetric brothmicrodilution test against 15 organisms comprising of 4 Gram-positive bacteria and 4 Gram-negative bacteria, 4 fungiand 3 yeast. On disc diffusion assay, the fungal extract was shown to inhibit the growth of 7 test bacteria and 3 testyeast. The antibacterial activity was more pronounced with extract from fungal culture with host plant extractsupplementation with significantly larger inhibition zones on all susceptible test microorganisms. The minimal inhibitoryconcentration of the extract ranged from 250 to 4000 μg/mL indicating different level of susceptibility of the testedpathogens against the fungal extract. The killing kinetic study shows that antimicrobial activity of the fungal extract isconcentration dependent and it can act as bactericidal at higher concentration.Conclusion, significance and impact of study: The findings of this study suggest that Aspergillus sp. IBRL MP15CCL can be a promising source of antimicrobial agent to be further studied and developed

Progress in antimicrobial substances of endophytes / 生物工程学报

Isolation and characterization of metabolites produced by endophytes are significant ways to search for novel natural active substances, proving that the endophytes are the unique resources of newer and more effective compounds. Many new compounds with antimicrobial activity from different endophytes have been isolated so far. These new compounds provide alternatives to fight against multi-drug resistance of microorganisms. This review outlined the major achievements and latest developments of endophytes, including the diversity of endophytes and antimicrobial activity of endophytes, as well as its development in China.

Distribution, Characterization, and Diversity of the Endophytic Fungal Communities on Korean Seacoasts Showing Contrasting Geographic Conditions

This study analyzed the distribution of endophytic fungi in 3 coastal environments with different climatic, geographical, and geological characteristics: the volcanic islands of Dokdo, the East Sea, and the West Sea of Korea. The isolated fungal endophytes were characterized and analyzed with respect to the characteristics of their host environments. For this purpose, we selected common native coastal halophyte communities from three regions. Molecular identification of the fungal endophytes showed clear differences among the sampling sites and halophyte host species. Isolates were also characterized by growth at specific salinities or pH gradients, with reference to previous geographical, geological, and climate studies. Unlike the East Sea or West Sea isolates, some Dokdo Islands isolates showed endurable traits with growth in high salinity, and many showed growth under extremely alkaline conditions. A smaller proportion of West Sea coast isolates tolerate compared to the East Sea or Dokdo Islands isolates. These results suggest that these unique fungal biota developed through a close interaction between the host halophyte and their environment, even within the same halophyte species. Therefore, this study proposes the application of specific fungal resources for restoring sand dunes and salt-damaged agricultural lands and industrialization of halophytic plants.

Bioactive compounds fractionated from endophyte Streptomyces SUK 08 with promising ex-vivo antimalarial activity

Objective:To determine ex vivo antimalarial activity and cytotoxicity of endophytic Streptomyces SUK 08 as well as the main core structure fractionated from its crude extract.Methods:The activities of SUK 08 crude extract were evaluated by using the Plasmodium lactate dehydrogenase assay and synchronization test against rodent malaria parasite Plasmodium berghei,instead of human malarial parasite Plasmodium falciparum.The cytotoxicity of the crude extract was determined by MTT assay.The crude extract was analyzed by thin-layer chromatography and gas chromatography-mass spectrophotometry.Results:The ethyl acetate crude extract showed very promising antimalarial activity with IC50 of 1.25 mg/mL.The synchronization tests showed that ethyl acetate extraction could inhibit all stages of the Plasmodium life cycle,but it was most effective at the Plasmodium ring stage.On the basis of a MTT assay on Chang Liver cells,ethyl acetate and ethanol demonstrated IC50 values of ＞ 1.0 mg/mL.The IC50 of parasitemia at 5％ and 30％ for this extract was lower than chloroquine.Thin-layer chromatography,with 1∶9ratio of ethyl acetate:hexane,was used to isolate several distinct compounds.Based on gas chromatography-mass spectrophotometry analysis,three core structures were identiffed as cyclohexane,butyl propyl ester,and 2,3-heptanedione.Structurally,these compounds were similar to currently available antimalarial drugs.Conclusions:The results suggest that compounds isolated from Streptomyces SUK 08 are viable antimalarial drug candidates that require further investigations.

Bioactive compounds fractionated from endophyte Streptomyces SUK 08 with promising ex-vivo antimalarial activity

Objective To determine ex vivo antimalarial activity and cytotoxicity of endophytic Streptomyces SUK 08 as well as the main core structure fractionated from its crude extract. Methods The activities of SUK 08 crude extract were evaluated by using the Plasmodium lactate dehydrogenase assay and synchronization test against rodent malaria parasite Plasmodium berghei, instead of human malarial parasite Plasmodium falciparum. The cytotoxicity of the crude extract was determined by MTT assay. The crude extract was analyzed by thin-layer chromatography and gas chromatography–mass spectrophotometry. Results The ethyl acetate crude extract showed very promising antimalarial activity with IC

Isolation and identification of endophytic fungi from five medicinal plants species of orchidaceae / 中国药学杂志

OBJECTIVE: To investigate extensively the cultural endophytic fungal community colonized mainly in the roots of five orchid medicinal plants species. METHODS: Cultural endophytic fungi were identified using the morphological methods and molecular sequences. RESULTS: A total of 241 strains of fungi were isolated and identified into at least 52 genera. Among them, Fusarium (14.9%), Colletotrichum(12.9%), Xylaria(5.4%) and Alternaria(3.7%) were dominated. Ten interested fungal strains were identified, including Dark septate endophyte (DSE) and Orchid mycorrhizal fungi(e. g. Tulasnella and Sebacina). CONCLUSION: Fungal taxa and richness are related to host plant species and its geological distribution. This study provides important fungal resources for cultivation of orchid medicinal plants and screening of bioactive compounds in the future.

Interaction between endophytes and host plant and the role of endophytes in genuineness analysis of medicinal plant / 药学学报

Research of plant development and metabolism has drawn lots of attention with the fast development of science of mycorrhizal biology, molecular biology and metabonomics technology. It has become one of hot fields in the study of endophytes and plant, which would affect plant's metabolite composition. This would provide opportunity for appraising and modifying traits to medicinal plant, and would also perfect the tranditional standpoint on forming reason of medicinal plant genuineness. Here we provide a review of theory and mechanism, research and application of interaction between plant and endophyte. This review may enhance understanding of medicinal plant, and evaluating the quality of herbs in production.

Isolation, identification and inhibition activity of endophytes in Fengdan / 中国中药杂志

Endophytes were isolated and purified from the roots of medicinal plant Fengdan also known as Paeonia suffruticosa from Tongling region, Anhui province, China. Morphology and molecular biology methods were applied to indentify the endophyte strains. And methods of growth rate and filtering paper were also used for studying antibacterial/antifungal effects of the strains. As a result, 129 endophyte strains were isolated. Fifty-eight endophytic fungi strains were identified as 6 species in 4 genera and the dominant genus was Fusarium. Seventy-one endophytic bacteria strains were identified as 9 species in 3 genera and the dominant genus was Bacillus. Inhibitory diameter with endophytic fermenting liquid of Pseudomonas chlororaphis, F. nematophilum and B. megaterium from P. suffruticosa against Staphyloccocus aureus, Escherichia coli and B. subtilis reached 25.0,20.2,24.0 mm respectively. The inhibition rate of endophytic fermenting liquid from F. nematophilum against Penicillium sp. and Colletotrichum dematium reached 90.6% and 83.3%, respectively. The inhibition effect of P. chlororaphis against F. oxysporum f. sp. niveum and Mucor sp. was good and the antifungal rate reached 80.0% and 84.9%, respectively. P. suffruticosa in Tongling region contains abundant endophytes. P. chlororaphis and F. nematophilum are valuable species as starting strain about microbicide.

Research Progress in Endophyte in Rhizoma Paridis / 中国药师

The endogenous microorganism in paridis rhizoma mainly includes bacteria and fungi,and shows different physiological activities,such as microorganism inhibition and anti-tumor activity. The principal active substance steroid saponin in paridis rhizoma can be generated from the fermentation liquor of some endogenous bacteria,and the chemical structure of vancomycin and progesterone can be transformed by the endophyte. It is supposed that paridis rhizoma may benefit from the attributions of the endogenous bacteria against illness and environment changes. It would also benefit in productivity, transformation and modification of the other active sub-metabolites and compounds in paridis rhizoma. Thus,it is valuable in drug development and source insufficiency alleviation of Chinese medicines through separation,identification,screening and clone of the endogenous microorganism in paridis rhizoma.

Broad spectrum antibacterial activity of granaticinic acid, isolated from Streptomyces thermoviolaceus NT1; an endophyte in Catharanthus roseus (L.) G. Don.

Searching for endophytic actinomycetes, strain NT1 was isolated from surface sterilized stem of Catharanthus roseus (L.) G. Don collected from Paschim Medinipur, India. The strain was identified as Streptomyces thermoviolaceus NT1 on the basis of morphological, biochemical and 16s rDNA based phylogenetic analysis. It showed potential antagonism against several Gram positive and Gram negative bacterial pathogens along with drug resistant Staphylococcus aureus. Maximum antibacterial production was obtained in ISP2 media at pH 7.2, 35 °C for 10 days. The active antibacterial substance was purified by Silica gel column chromatography and activity guided TLC. IR and NMR analysis identified the active compound as granaticinic acid of m/z 463.26 [M+H]. These results suggest that the antimicrobial produced by the isolated endophytic strain will be useful in near future.