| Endophytic fungi of medicinal plants,as special microorganisms,are important sources of bioactivity compounds.However,the diversity and bioactivity metabolites of endophytic fungi from Pinellia Tenore have not been systematically studied.In this thesis,P.ternata and P.pedatisecta were used as experimental materials to isolate endophytic fungi using various isolation media.The species of endophytic fungi were identified using molecular biology methods,and their diversity was analyzed.The antibacterial and herbicidal activities of the endophytic fungi were screened,and the active components of the target strains were further isolated and purified.The structures of the compounds were further identified by a variety of spectral methods.The main results are as follows:1.Endophytic fungi of Pinellia Tenore were isolated and purified using three kinds of isolation media(Czapek–Dox Medium,MEA,and PDA).A total of 53 and 24 strains of endophytic fungi were isolated from P.ternata and P.pedatisecta,respectively.The identification results showed that the endophytic fungi of P.ternata were distributed in 19 different genera of 5 classes(Agaricomycetes,Pezizomycetes,Orbiliomycetes,Saccharomycetes,and Coprinellomycetes),and the main genera identified were Fusarium,Aspergillus,Penicillium,Talaromyces,Bjerkandera,Lecanicillium,with isolation frequencies of 16.98%,16.98%,11.32%,9.43%,5.66%,and 5.66%,respectively.The endophytic fungi of P.pedatisecta were distributed in 11 different genera of 4 classes(Agaricomycetes,Pezizomycetes,Orbiliomycetes,and Coprinellomycetes),and the main genera were Fusarium,Aspergillus,Cladosporium,Bjerkandera,Colletotrichum,with isolation frequencies of 33.33%,16.67%,8.33%,8.33%,and 8.33%,respectively.The dominant endophytic fungi in both Pinellia Tenore were Fusarium,and they were distributed in all tissues of P.ternata and P.pedatisecta.The analysis of diversity indices indicated that the endophytic fungi in the tubers of P.ternata exhibited high species richness and diversity.The species richness index,Margalef index,and Shannon-Wiener index were 12,3.3011,and 2.2299,respectively.In addition,the endophytic fungi in the roots of P.ternata and P.pedatisecta showed high species evenness,with Pielou’s evenness index of 0.9697 and1.0000,respectively.2.The results of antibacterial activity test showed that the crude extract of ethyl acetate from 19 strains(24.7%)of endophytic fungi showed antibacterial activity against at least one pathogenic bacterium(Staphylococcus aureus,Micrococcus tetragenus,Escherichia coli,and Pseudomonas syringae pv.actinidiae),with the inhibition zone diameter of 7.0 –20.0 mm under the concentration of 30 μg/6 mm.In addition,the results of herbicidal activity test showed that the fermentation broth of 52 strains(67.5%)of endophytic fungi showed good herbicidal activity against Echinochloa crusgalli,with inhibition rates of more than50%.The endophytic fungi Penicillium citrinum PT58 and Alternaria angustiovoidea PT09 of P.ternata and Aspergillus floccosus PP39 of P.pedatisecta exhibited inhibitory activity against at least two pathogenic bacteria and potent herbicidal activity with inhibition rates of more than 91%.Therefore,they were selected as target strains for further experiments.3.Four compounds were isolated and identified from the endophytic fungus A.angustiovoidea PT09 in P.ternata: alternariol monomethyl ether(1),alternariol(2),dehydroaltenusin(3),alternarotoxin II(4).Five compounds were isolated and identified from the endophytic fungus P.citrinum PT58 in P.ternata: citrinin(5),frangula emodin(6),citreorosein(7),sclerotinin B(8),dihydrocitrinone(9).Three compounds were isolated and identified from the endophytic fungus A.floccosus PP39 in P.pedatisecta: citrinin(5),terreic acid(10),terremutin(11).The antibacterial activity test results showed that compound 10 exhibited strong antibacterial activities against E.coli,M.tetragenus,S.aureus,and P.syringae pv.actinidiae with the IZD of 36.0,31.0,33.7,40.2 mm under the concentration of30 μg/6 mm and minimum inhibitory concentration(MIC)values of 1.56,3.13,1.56,1.56μg/m L respectively,which were better than or equal to those of positive gentamicin sulfate(IZD: 21.7,25.7,26.7,and 26.0 mm;MIC: 1.56,3.13,3.13,and 6.25 μg/m L);compound 5also exhibited strong inhibitory activity against the P.syringae pv.actinidiae with the IZD of 26.0 mm and MIC value of 6.25 μg/m L,which were equal to those of positive gentamicin sulfate.In addition,the compound 5 had potent phytotoxic activity against E.crusgalli and Abutilon theophrasti with the inhibition rate of 73.4% and 41.7% under the concentration of100 μg/m L,respectively.In conclusion,this study showed endophytic fungi from P.ternata and P.pedatisecta were a resource pool of bioactive natural products,with broad prospects for development of new antibiotics and biological herbicides. |
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