竹红菌甲素对番茄灰霉病菌的抑制作用
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摘要
为研究竹红菌甲素(HA)对番茄灰霉病菌的抑制作用,分别采用菌丝生长速率法、电导率法、紫外分光光度法和电镜观察法研究了HA对番茄灰霉病菌抑制率以及其对电导率、蛋白质和核酸相对泄漏度、菌丝形态与超微结构的影响。结果表明:在黑暗条件下,HA对番茄灰霉病菌无明显抑制作用;但在12 000 lx光照条件下,抑菌作用显著,EC50值为17.16 mg/L,且随着HA浓度上升,抑菌率呈极显著增加趋势,40 mg/L下抑菌率达73.9%;番茄灰霉病菌细胞相对电导率、蛋白质与核酸相对泄漏度随HA浓度上升而明显升高;电镜观察可见,经34.32 mg/L的HA处理后,大多数菌丝出现畸形、膨大或缢缩等现象,细胞器几乎全部消失,细胞内出现大面积空腔,细胞遭到严重破坏。研究结果可为HA开发成绿色高效生物农药提供理论依据。
In order to study the inhibitory effect of Hypocrellin A( HA) on Botrytis cinerea,the inhibition rate, conductivity, leakage of protein and nucleic acid, mycelial morphology and ultrastructure of B. cinerea that being treated with HA were determined by methods of mycelium growth rate,conductivity,UV spectrophotometry and electron microscope,respectively. The results showed that HA had weak or even no antifungal activity to B. cinerea in the dark; but under luminous intensity of 12 000 lx,the activity was obvious,with an EC50 value of 17. 16 mg / L,and the inhibition rate increased significantly with the increase of HA concentration,the inhibition rate could reach 73. 9% at40 mg / L HA; The conductivity, leakage of protein and nucleic acid rose obviously as the concentration of HA increased. According to the results of electron microscope,most of hyphae malformed with swollen or constriction after being treated by 34. 32 mg / L HA; M eanwhile,almost allthe organelles disappeared and cavities with large area appeared in the cells,suggesting the cells were damaged severely. The results of present study provided theoretical basis for developing HA as a novel green biological pesticide.
In order to study the inhibitory effect of Hypocrellin A( HA) on Botrytis cinerea,the inhibition rate, conductivity, leakage of protein and nucleic acid, mycelial morphology and ultrastructure of B. cinerea that being treated with HA were determined by methods of mycelium growth rate,conductivity,UV spectrophotometry and electron microscope,respectively. The results showed that HA had weak or even no antifungal activity to B. cinerea in the dark; but under luminous intensity of 12 000 lx,the activity was obvious,with an EC50 value of 17. 16 mg / L,and the inhibition rate increased significantly with the increase of HA concentration,the inhibition rate could reach 73. 9% at40 mg / L HA; The conductivity, leakage of protein and nucleic acid rose obviously as the concentration of HA increased. According to the results of electron microscope,most of hyphae malformed with swollen or constriction after being treated by 34. 32 mg / L HA; M eanwhile,almost allthe organelles disappeared and cavities with large area appeared in the cells,suggesting the cells were damaged severely. The results of present study provided theoretical basis for developing HA as a novel green biological pesticide.
引文
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[2]徐大勇,李峰.番茄灰霉病拮抗内生细菌的筛选、鉴定及其活性[J].生态学杂志,2012,31(4):994-999.Xu Dayong,Li Feng.Screening,identification,and bioactivity of an endophytic bacterium from tomato against Botrytis cinerea[J].Chin J Ecol,2012,31(4):994-999.(in Chinese)
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[5]Jiang Yuan,Leung Albert Wingnang,Wang Xinna,et al.Inactivation of Staphylococcus aureus by photodynamic action of hypocrellin B[J].Photodiagnosis Photodyn Ther,2013,10(4):600-606.
[6]薛东运,李华,滕学洲.竹红菌素加用黑光灯照射治疗增生性疤痕127例[J].皮肤病与性病,2006,28(2):35-36.Xue Dongyun,Li Hua,Teng Xuezhou.The 127 cases of therapy for hypertrophic scar by use of Hypocrellins w ith black light[J].J Dermatol Venereol,2006,28(2):35-36.(in Chinese)
[7]吴瑞芳,张菊英.聚焦超声联合竹红菌素软膏加光疗治疗外阴白色病变疗效观察[J].中华实用诊断与治疗杂志,2010,24(1):64-65.Wu Ruifang,Zhang Juying.The curative effect observation to vulvar w hite lesion by use focal ultrasonic treatment combine photodynamic therapy of Hypocrellin ointment[J].J Pract Diagn Ther,2010,24(1):64-65.(in Chinese)
[8]王瑛.聚焦超声合并竹红菌素治疗外阴白色病变[J].当代医学,2010,16(25):54.Wang Ying.Focal ultrasonic treatment combine Hypocrellin cure vulvar w hite lesion[J].Contemporary Med,2010,16(25):54.(in Chinese)
[9]赵菊芬,李茜,杨柳风,等.聚焦超声联合竹红菌素软膏治疗外阴白色病变164例疗效观察[J].中国计划生育和妇产科,2012,4(1):61-63.Zhao Jufen,Li Qian,Yang Liufeng,et al.Clinical observation of curative effect on focused-ultrasound combined w ith hypocrellin-ointment to treat leukoplakia vulvae in 164 cases[J].Chin J Fam Plann G ynecotokol,2012,4(1):61-63.(in Chinese)
[10]Wang Fang,Zhou Lin,Zhou Jiahong,et al.Characterization of anticancer hypocrellin A encapsulated w ith silica nanoparticles[J].J Therm Anal Calorim,2010,102(1):69-74.
[11]Jin Shan,Zhou Liangjun,Gu Zhanjun,et al.A new near infrared photosensitizing nanoplatform containing blue-emitting up-conversion nanoparticles and hypocrellin A for photodynamic therapy of cancer cells[J].Nanoscale,2013,5(23):11910-11918.
[12]Chio-Srichan S,Réfrégiers M,Jamme F,et al.Photosensitizer effects on cancerous cells:a combined study using synchrotron infrared and fluorescence microscopies[J].Biochim Biophys Acta(BBA)-G eneral Subjects,2008,1780(5):854-860.
[13]Hudson J B,Zhou J,Chen J,et al.Hypocrellin,form Hypcrella bambusae,is phototoxic to human immunodeficiency virus[J].J Photochem Photobiol B:Biol,1994,60(3):253-255.
[14]Junichi H,Kenji I,Hideki A,et al.Photoinactivation of virus infectivity by hypocrellin A[J].Photochem Photobiol,1997,66(5):697-700.
[15]Sirinart C S,Noufissa O,Annelise B G,et al.Toxicity and phototoxicity of Hypocrellin A on malignant human cell lines,evidence of a synergistic action of photodynamic therapy w ith Imatinib mesylate[J].J Photochem Photobiol B:Biol,2010,99(2):100-104.
[16]Fei X F,Chen J,Zheng K Y,et al.Apoptotic effects of hypocrellin A on He La cells[J].Chem Res Chin Univ,2006,22(6):772-775.
[17]陈毅坚,钟文武,杨松艳.滇西竹黄抗菌活性研究[J].云南民族大学学报:自然科学版,2010,19(2):154-156.Chen Yijian,Zhong Wenw u,Yang Songyan.Study on the antibacterial activity of Shiraia bambusicola Henn[J].J Yunnan Univ Natl:Nat Sci Ed,2010,19(2):154-156.(in Chinese)
[18]高波,刘卫,张举成,等.竹红菌素光敏反应抑菌效果研究[J].安徽农业科学,2010,38(15):7839-7840.Gao Bo,Liu Wei,Zhang Jucheng,et al.Study about photosensitive bacteriostasis effects of hypocrellins[J].J Anhui Agric Sci,2010,38(15):7839-7840.(in Chinese)
[19]Su Yujie,Sun Jun,Rao Shengqi,et al.Photodynamic antimicrobial activity of hypocrellin A[J].J Photochem Photobiol B:Biol,2011,103(14):29-34.
[20]朱齐超,林海萍,李朋举,等.竹红菌甲素对18种植物病原菌的抑制作用[J].植物病理学报,2014,44(1):107-109.Zhu Qichao,Lin Haiping,Li Pengju,et al.Inhibition of Hypocrellin A against 18 pathogenic fungi[J].Acta Phytopathologica Sinica,2014,44(1):107-109.(in Chinese)
[21]黄小波.药用真菌竹黄的液态发酵工艺优化及竹红菌素的提取研究[D].杭州:浙江农林大学,2010.Huang Xiaobo.Study on the optimization of liquid fermentation process of Shiraia bambusicola and extraction of hypocrellin[D].Hangzhou:Zhejiang A&F University,2010.(in Chinese)
[22]朱齐超,张媚,李朋举,等.竹红菌甲素Triton X-100微乳剂的制备、光热稳定性及抑菌活性[J].农药学学报,2013,15(5):578-592.Zhu Qichao,Zhang M ei,Li Pengju,et al.Preperation light&thermal stability and antimicrobial activity of hypocrellin A Triton X-100 micro-emulsion[J].Chin J Pestic Sci,2013,15(5):578-592.(in Chinese)
[23]方中达.植病研究方法[M].北京:中国农业出版社,1998:152.Fang Zhongda.The Research M ethod of Plant Disease[M].Beijing:China Agriculture Press,1998:152.(in Chinese)
[24]毛胜凤,林海萍,陈安良,等.喜树碱对番茄灰霉病菌的病理反应[J].中国生物防治学报,2014,30(1):143-148.M ao Shengfeng,Lin Haiping,Chen Anliang,et al.Antifungal reaction of Botrytis cinerea on camptothecin[J].Chin J Biolog Contr,2014,30(1):143-148.(in Chinese)
[25]宋风平,王树桐,胡同乐,等.芒果苷对马铃薯晚疫病菌的抑菌作用机制初探[J].农药学学报,2009,11(2):213-218.Song Fengping,Wang Shutong,Hu Tongle,et al.Preliminary studies on the mode of action of mangiferin against Phytophthora infestans[J].Chin J Pest Sci,2009,11(2):213-218.(in Chinese)
[26]金林红,陈红军,杨松,等.新型砜类化合物对辣椒枯萎病菌的抑菌作用初探[J].农药学学报,2010,12(2):233-236.Jin Linhong,Chen Hongjun,Yang Song,et al.Preliminary study on antifungal activity of new derivative of sulfone against Fusarium oxysporum[J].Chin J Pestic Sci,2010,12(2):233-236.(in Chinese)
[27]刘淑宇,于新,陈发河,等.绿色木霉菌发酵液对杧果炭疽菌的生长抑制及细胞损伤作用[J].果树学报,2012,29(6):1097-1102.Liu Shuyu,Yu Xin,Chen Fahe,et al.Effect of Trichoderma uiride fermentation broth on Colletorichum gloeosporioides grow th inhibition and cell damages[J].J Fruit Sci,2012,29(6):1097-1102.(in Chinese)
[28]Xie J Y,Dong G J,Liu Z Y.Method of preparing specimens of microorganism for SEM[J].J Chin Electron Microsc Soc,2005,43(7):975-980.
[29]陈安良,冯美杰,冯俊涛,等.丙烷脒对灰霉病菌菌丝形态和超微结构的影响[J].中国农业科学,2007,40(3):633-637.Chen Anliang,Feng M eijie,Feng Juntao,et al.Effects of propamidine on mycelial morphology and ultrastructure of Botrytis cirerea[J].Scientia Agricultura Sinica,2007,40(3):633-637.(in Chinese)
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