蒜头果中植物内生真菌烟曲霉的抗菌活性研究
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摘要
植物内生真菌是一类生活在植物体内的微生物,能够产生化学结构新颖、生物活性多样的次生代谢产物。本研究用纸片法检测从我国特有植物蒜头果中分离得到的植物内生真菌烟曲霉的抗菌活性,并采用单菌多产物(OSMAC)策略寻找植物内生真菌烟曲霉产生抗菌化合物的最佳条件。结果表明:该烟曲霉液体发酵提取物对供试的6种人体致病细菌均有抑菌活性(蜡样芽孢杆菌、副溶血性弧菌、无乳链球菌、福氏志贺氏菌、金黄色葡萄球菌、藤黄微球菌),其中对副溶血性弧菌、藤黄微球菌最低抑制浓度达到6.25mg/mL。以马铃薯粉为氮源,麦芽糖或葡萄糖为碳源,提取物得率较大且抑菌活性最好,接种量为60mg/100mL时提取物收获量最高。此外,改变培养温度能提高提取物对除无乳链球菌及藤黄微球菌以外的4种致病菌的抗菌活性。与细菌共培养后对蜡样芽孢杆菌、金黄色葡萄球菌、藤黄微球菌的抗菌活性有所增强,但对副溶血性弧菌、无乳链球菌、福氏志贺氏菌的抗菌活性下降。本研究为植物内生真菌抗菌化合物的开发利用奠定基础。
Endophytic fungi are microorganisms living in the body of plants,and it can produce secondary metabolites with novel chemical structure and biological activities. In this study,the antibacterial activity of Aspergillus fumigatus that isolated from Malania oleifera Chun et Lee was detected by paper method for the first time. And one strain-many compounds(OSMAC) strategy was used to find the best conditions for the production of antibacterial compounds of A. fumigatus. The results showed that the extracts of A. fumigatus had antibacterial activity on six kinds of human pathogenic bacteria(Bacillus cereus,Vibrio parahaemolyticus,Streptcococcus aglactiae,Shigella flexneri,Staphylococcus aureus,Micrococcus luteus),the lowest inhibition concentration of Vibrio parahaemolyticus and Micrococcus luteus was 6. 25 mg/mL. OSMAC strategy was adopted to optimize the training conditions and the results showed that extracted rate of extract and antimicrobial activity was both greater with potato powder as nitrogen source,maltose or glucose as the source of carbon and the rate of extract was highest when the inoculation was60 mg/mL. In addition,the change of culture temperature can improve the antibacterial activity of 4 pathogenic bacteria except Streptcococcus agalactiae and Micrococcus luteus. Co-culture with bacteria increased the antibacterial activity to Bacillus cereus,Staphylococcus aureus and Micrococcus luteus but reduced the antibacterial activity to Vibrio parahaemolyticus,Streptcococcus agalactiae and Shigella flexneri. The study could provide help for the development and utilization of plant endophytic fungi antibacterial compounds.
Endophytic fungi are microorganisms living in the body of plants,and it can produce secondary metabolites with novel chemical structure and biological activities. In this study,the antibacterial activity of Aspergillus fumigatus that isolated from Malania oleifera Chun et Lee was detected by paper method for the first time. And one strain-many compounds(OSMAC) strategy was used to find the best conditions for the production of antibacterial compounds of A. fumigatus. The results showed that the extracts of A. fumigatus had antibacterial activity on six kinds of human pathogenic bacteria(Bacillus cereus,Vibrio parahaemolyticus,Streptcococcus aglactiae,Shigella flexneri,Staphylococcus aureus,Micrococcus luteus),the lowest inhibition concentration of Vibrio parahaemolyticus and Micrococcus luteus was 6. 25 mg/mL. OSMAC strategy was adopted to optimize the training conditions and the results showed that extracted rate of extract and antimicrobial activity was both greater with potato powder as nitrogen source,maltose or glucose as the source of carbon and the rate of extract was highest when the inoculation was60 mg/mL. In addition,the change of culture temperature can improve the antibacterial activity of 4 pathogenic bacteria except Streptcococcus agalactiae and Micrococcus luteus. Co-culture with bacteria increased the antibacterial activity to Bacillus cereus,Staphylococcus aureus and Micrococcus luteus but reduced the antibacterial activity to Vibrio parahaemolyticus,Streptcococcus agalactiae and Shigella flexneri. The study could provide help for the development and utilization of plant endophytic fungi antibacterial compounds.
引文
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[14]国家环境保护局.珍稀濒危植物保护与研究[M].北京:中国环境科学出版社,1991.
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[17]林淑婷,李从发,胡晓苹,等.不同发酵方式对海南粗榧内生真菌CH1307c合成次级代谢产物的影响[J].热带作物学报,2016,37(7):1407-1412.
[18]王胤.烟曲霉次级代谢产物抗菌与抗肿瘤活性研究[J].化学与生物工程,2011,28(4):70-72.
[19]李梓木.刺五加内生真菌CJ7活性代谢物研究[D].哈尔滨:黑龙江大学,2010.
[20]Liu J Y,Song Y C,Zhang Z,et al.Aspergillus fumigatus CY018,an endophytic fungus in Cynodon dactylon as a versatile producer of new and bioactive metabolites[J].Journal of Biotechnology,2004,114(3):279-287.
[21]谢海平,黄晖,黄登峰,等.海洋枯草芽孢杆菌Bs-1产生多种抗真菌活性物质[J].中山大学学报(自然科学版),2003,42(3):122-123.
[22]Lee S Y,Kinoshita H,Ihara F,et al.Identification of novel derivative of helvolic acid from Metarhizium anisopliae,grown in medium with insect component[J].Journal of Bioscience&Bioengineering,2008,105(5):476-480.
[2]郭顺星.药用植物内生真菌研究现状和发展趋势[J].菌物学报,2018,37(1):1-13.
[3]Zhang X,Li C,Nan Z.Effects of cadmium stress on seed germination and seedling growth of Elymus dahuricus infected with the Neotyphodium endophyte[J].Science China-Life Sciences,2012,55(9):793-799.
[4]Oberhofer M,Gusewell S,Leuchtmann A.Effects of natural hybrid and non-hybrid Epichloe endophytes on the response of Hordelymus europaeus to drought stress[J].New Phytologist,2014,201(1):242-253.
[5]Rodriguez R J,White J F J,Arnold A E,et al.Fungal endophytes:diversity and functional roles[J].New Phytologist,2009,182(2):314-330.
[6]Xia C,Zhang X,Christensen M J,et al.Epichloe endophyte affects the ability of powdery mildew(Blumeria graminis)to colonise drunkenhorse grass(Achnatherum inebrians)[J].Fungal Ecology,2015,16:26-33.
[7]黄敬瑜,张楚军,姚瑜龙,等.植物内生菌生物抗菌活性物质研究进展[J].生物工程学报,2017,33(2):178-186.
[8]Stierle A,Stierle D.Taxol and taxane production by Taxomyces andreanae,an endophytic fungus of Pacific yew[J].Science,1993,260(5105):214-216.
[9]Cacho R A,Tang Y,Chooi Y H.Next-generation sequencing approach for connecting secondary metabolites to biosynthetic gene clusters in fungi[J].Frontiers in Microbiology,2015,5(774):1-16.
[10]胡付品,朱德妹,汪复,等.CHINET监测2010年碳青霉烯类抗生素耐药肠杆菌科细菌的分布特点和药物敏感性[J].中国感染与化疗杂志,2013,13(1):1-7.
[11]崔海滨,梅文莉,缪承杜,等.红树植物卤蕨内生真菌Penicillium sp.0935030中的抗菌活性成分研究[J].中国抗生素杂志,2008,33(7):407-410.
[12]Weber D,Sterner O,Anke T.Phomol,a new antiinflammatory metabolite from an endophyte of the medicinal plant erythrina crista-galli[J].Journal of Antibiotics,2004,57(9):559-563.
[13]卢轩,陈刚,裴月湖.单菌多产物策略及其在药物先导化合物研发中的应用[J].沈阳药科大学学报,2011,28(8):640-645.
[14]国家环境保护局.珍稀濒危植物保护与研究[M].北京:中国环境科学出版社,1991.
[15]Strobel G,Daisy B,Castillo U,et al.Natural products from endophytic microorganisms[J].Journal of Natural Products,2004,67(2):257-268.
[16]马养民,赵洁.植物内生真菌抗菌活性物质的研究进展[J].有机化学,2010,30(2):220-232.
[17]林淑婷,李从发,胡晓苹,等.不同发酵方式对海南粗榧内生真菌CH1307c合成次级代谢产物的影响[J].热带作物学报,2016,37(7):1407-1412.
[18]王胤.烟曲霉次级代谢产物抗菌与抗肿瘤活性研究[J].化学与生物工程,2011,28(4):70-72.
[19]李梓木.刺五加内生真菌CJ7活性代谢物研究[D].哈尔滨:黑龙江大学,2010.
[20]Liu J Y,Song Y C,Zhang Z,et al.Aspergillus fumigatus CY018,an endophytic fungus in Cynodon dactylon as a versatile producer of new and bioactive metabolites[J].Journal of Biotechnology,2004,114(3):279-287.
[21]谢海平,黄晖,黄登峰,等.海洋枯草芽孢杆菌Bs-1产生多种抗真菌活性物质[J].中山大学学报(自然科学版),2003,42(3):122-123.
[22]Lee S Y,Kinoshita H,Ihara F,et al.Identification of novel derivative of helvolic acid from Metarhizium anisopliae,grown in medium with insect component[J].Journal of Bioscience&Bioengineering,2008,105(5):476-480.