树木溃疡病病原菌Botryosphaeria dothidea代谢产物的研究
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摘要
树木溃疡病主要由病原真菌引起,其中Botryosphaeria属是最为重要的病原菌之一。世界各地屡有该属真菌导致树木溃疡病害的报道。其危害单子叶植物、双子叶植物和裸子植物的枝干、果实等。我国早在上个世纪就已经开始了对树木溃疡病的研究,并在培养中发现该菌在生长发育过程中能产生诱使寄主树木发生病害的代谢物质。本文对Botryosphaeria属真菌在致病过程中产生的代谢产物成分进行了研究,结果如下。
粗毒素萃取后生测试验表明,丙沉中所含成分比丙浮具较强的致病力。方差分析显示,丙浮、丙沉、CK接种北京杨、毛白杨的感病指数差异显著,LSD和Duncan测验均值大小排序为:丙沉>丙浮>CK。毒素透析后生测试验表明,IN液体接种北京杨、毛白杨的感病指数明显高于OUT的接种处理,方差分析二者与CK接种北京杨、毛白杨后的感病面积差异显著,LSD和Duncan测验均值大小排序为:IN>OUT>CK,随着时间的变化IN和OUT处理后的感病面积呈上升趋势,并在15d时IN和OUT的感病指数均达到最大值。其中OUT的致病成分为分子量小于3500的有毒物质。
通过HPLC示差面积归一法鉴定透析袋内液体成分,主要成分出现时间为4 min左右,分子量为5000的多糖类化合物。经过Benidict和Seliwanoff试验,初步认为该代谢物成分内含有分子量为5000具致病能力的酮糖类化合物。
从试验中得到溶解性较小的白色透明晶体,可以溶于丙酮有机分析纯溶液,部分溶解于乙醇溶液,在水中的溶解性较小,在NaCl水溶液中呈现较淡的茶红色。从物理和化学的一般性质以及相关文献的整理,判断该晶体应为鹿蹄草素或类鹿蹄草素化合物。
Canker diseases of woody plants were caused by disease fungus, Botryosphaeria genus was one of the most important genuses. There were many infection reports of this genus around the world. They could harm the trunk and fruits of monocotyledon, dicotyledon and gymnosperm. The research started from last century at this diseases in our country. And there were some metabolites could cause trees canker in experiments. This paper was researched the metabolites of Botryosphaeria dothidea, and results were followed.
The bioassay results of crude phytotoxin after extraction with 100% acetone, the ingredient of sediment caused this disease was more strength than the suspended solid.The infected index were significantly different of the inoculated the suspended solid, sediment acetone extraction and CK through anova. And the mean of LSD and Duncan were order as sediment acetone extraction, suspended solid and CK. In the phytotoxin bioassary after dialysis, the infection indexes of the inner solution were obviously highly than OUT. The infected areas were significantly different of the inoculated IN, OUT and CK through anova. The infect areas were significantly different and rising step by step by the time, and the 15d was biggest. The molecular of OUT were less than 3500.
Through the area normalization of HPLC evaluated the main composition of inner bag filter that come out at 4min, it was considered as polyoses that molecular was about 5000. After the qualitation test of Benidict and Seliwanoff , it was considered as ketose-like chemical compound that molecular was about 5000 and had pathogenecity.
The small white transparent crystal with solubleness was gained in experiment, and soluble in acetone, part soluble in ethanol and solubleness in water. They could become congo red when put them in NaCl solution. From the repot and its general character of physical and chemical, it was considered as pyrolin or pyrolin-like.
粗毒素萃取后生测试验表明,丙沉中所含成分比丙浮具较强的致病力。方差分析显示,丙浮、丙沉、CK接种北京杨、毛白杨的感病指数差异显著,LSD和Duncan测验均值大小排序为:丙沉>丙浮>CK。毒素透析后生测试验表明,IN液体接种北京杨、毛白杨的感病指数明显高于OUT的接种处理,方差分析二者与CK接种北京杨、毛白杨后的感病面积差异显著,LSD和Duncan测验均值大小排序为:IN>OUT>CK,随着时间的变化IN和OUT处理后的感病面积呈上升趋势,并在15d时IN和OUT的感病指数均达到最大值。其中OUT的致病成分为分子量小于3500的有毒物质。
通过HPLC示差面积归一法鉴定透析袋内液体成分,主要成分出现时间为4 min左右,分子量为5000的多糖类化合物。经过Benidict和Seliwanoff试验,初步认为该代谢物成分内含有分子量为5000具致病能力的酮糖类化合物。
从试验中得到溶解性较小的白色透明晶体,可以溶于丙酮有机分析纯溶液,部分溶解于乙醇溶液,在水中的溶解性较小,在NaCl水溶液中呈现较淡的茶红色。从物理和化学的一般性质以及相关文献的整理,判断该晶体应为鹿蹄草素或类鹿蹄草素化合物。
Canker diseases of woody plants were caused by disease fungus, Botryosphaeria genus was one of the most important genuses. There were many infection reports of this genus around the world. They could harm the trunk and fruits of monocotyledon, dicotyledon and gymnosperm. The research started from last century at this diseases in our country. And there were some metabolites could cause trees canker in experiments. This paper was researched the metabolites of Botryosphaeria dothidea, and results were followed.
The bioassay results of crude phytotoxin after extraction with 100% acetone, the ingredient of sediment caused this disease was more strength than the suspended solid.The infected index were significantly different of the inoculated the suspended solid, sediment acetone extraction and CK through anova. And the mean of LSD and Duncan were order as sediment acetone extraction, suspended solid and CK. In the phytotoxin bioassary after dialysis, the infection indexes of the inner solution were obviously highly than OUT. The infected areas were significantly different of the inoculated IN, OUT and CK through anova. The infect areas were significantly different and rising step by step by the time, and the 15d was biggest. The molecular of OUT were less than 3500.
Through the area normalization of HPLC evaluated the main composition of inner bag filter that come out at 4min, it was considered as polyoses that molecular was about 5000. After the qualitation test of Benidict and Seliwanoff , it was considered as ketose-like chemical compound that molecular was about 5000 and had pathogenecity.
The small white transparent crystal with solubleness was gained in experiment, and soluble in acetone, part soluble in ethanol and solubleness in water. They could become congo red when put them in NaCl solution. From the repot and its general character of physical and chemical, it was considered as pyrolin or pyrolin-like.
引文
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2 Bernard slippers, Micheal J Wingfield. Botryosphaeriaceae as endophytes and latent pathogens of woody plants: diversity, ecology and impact[J]. Fungal biology reviews, 2007, 21: 90-106.
3 E. Turco, L. Marianelli, et al. First Report of Botryosphaeria dothidea on Sycamore, Red Oak, and English Oak in northwestern Italy[J]. Plant disease. 2006, 90(8): 1106.
4 D. Jurc, N. Ogris. First report of Botryosphaeria dothidea causing bark dieback of European hop hornbeam in Slovenia[J]. Plant Pathology. 2006. 55, 299.
5 J.Moral, M.Lovera, et al. First report of Botryosphaeria obtusa causing fruit rot of quince (Cydonia oblonga) in Spain[J]. New Disease Reports. 2006.13,33.
6 J.R.úrbez Torres, W.D.Gubler, et al. First report of Botryosphaeria iberica and B. viticola associated with grapevine decline in California[J]. Plant disease.2007, 91(6):772.
7 B. T. Linaldeddu, A. Franceschini, et al. First report of canker disease caused by Botryosphaeria parva on cork oak trees in Italy[J]. Plant disease. 2007, 91(3): 324.
8 Ulrike Damm, Pedro W.Crous, et al. Botryosphaeriaceae as potential pathogens of Prunus species in South Africa, with descriptions of Diplodia africana and Lasiodiplodia plurivora sp.Nov[J]. Mycologia, 2007, 99(5): 664-680.
9 J.Moral, F.Luque, et al. First report of Diplodia seriata, the anamorph of“Botryosphaeria”obtusa, causing fruit rot of olive in Spain[J].Plant disease .2008, 92(2): 311.
10 Macedo, Davi Mesquita de, et al. First record of Botryosphaeria ribis associated with leaf spots on Magnolia aff. candollei in Brazil[J]. Brazilian Journal of Microbiology. 2008. 39(2): 321-324.
11 N.T.Amponsah, E.E.Jones, et al. First report of Neofusicoccum australe [Botryosphaeria australis],a cause of grapevine dieback in New Zealand[J]. Australasian Plant Disease Notes, 2009,4: 6–8.
12 Qiu Y, Savocchia S, et al. Botryosphaeria dothidea associated with grapevine trunk disease in southeastern Australia[J]. Australasian Plant Pathology, 2008, 37: 482-485.
13 Jules DésiréDjoukeng, Suzanna Polli, et al. Identification of phytotoxins from Botryosphaeria obtusa, a pathogen of black dead arm disease of grapevine[J]. Eur J Plant Pathol,2009,124: 303-308.
14 Annalisa Cabras, Maria A Mannoni, et al. Occurrence,isolation and biological activity ofphytotoxic metabolites produced in vitro by Sphaeropsis sapinea ,pathogenic fungus of Pinus radiata[J].European Journal of Plant Pathology,2006, 115: 187-193.
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