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蛋白毒素AP-Toxin诱导撑绿杂交竹抗梢枯病及其分泌物响应
何倩倩, 彭麒, 宗绡卓, 朱天辉, 李姝江
南京林业大学学报（自然科学版） ›› 2020, Vol. 44 ›› Issue (5) : 199-208.
PDF(1975 KB)
PDF(1975 KB)
蛋白毒素AP-Toxin诱导撑绿杂交竹抗梢枯病及其分泌物响应
Induced resistance of Bambusa pervariabilis× Dendrocalamopsis grandis to Arthrinium phaeospermum
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【目的】以撑绿杂交竹梢枯病病原菌(Arthrinium phaeospermum)灭活的蛋白毒素AP-Toxin为诱导因子,对在其诱导下3个杂交竹品种分泌物特性与病情指数、诱抗效果相关性进行分析,为诱导抗病性在林木病害治理中的应用奠定理论基础。【方法】选择健康的1年生撑绿杂交竹3号(No.3)、6号(No.6)、8号(No.8),对其上部嫩枝按4种处理方式进行针刺接种,接种后在4个不同时期分别采样。采用修订后的索姆基(Sormgy)糖含量测定法、双缩脲法、碱液滴定法分别测定分泌物中糖、蛋白质和脂类含量,分析糖、蛋白质和脂类分泌物对灭活毒素诱导的响应。【结果】撑绿杂交竹在引入灭活蛋白毒素作为诱导因子后,其分泌物(蛋白质、可溶性多糖、总糖、脂类)含量增加,病情指数显著下降、诱抗效果显著增加,撑绿杂交竹抵抗病原菌侵染的能力增强。在3个品种中,诱导后病情指数No.6最低、动态变化最小,其次是No.3,No.8病情指数最高。相关性分析表明,蛋白质、可溶性糖、总糖和脂类含量与病情指数和抗性显著正相关,其中还原糖含量与抗性呈显著负相关。【结论】暗孢节菱孢菌灭活蛋白毒素诱导后能使撑绿杂交竹抗性增强,抵抗病原菌的侵染;且抗性越强的品种,诱抗效果越好。
【Objective】 The inactivated protein toxin from Arthrinium phaeospermum(AP-Toxin) was used as the induction factor for the resistance of three Bambusa pervariabilis × Dendrocalamopsis grandis varieties, and the correlations of secretory response with the disease index and resistance were analyzed.【Method】 The upper twigs of healthy one-year-old B. pervariabilis× D. grandis varieties of No. 3, 6, and 8 were inoculated with AP-Toxin by puncturing. The twig samples were collected at four different time points after inoculation. Secretion contents including sugar, proteins, and lipids were determined using the sormy sugar content determination method, biuret method, and alkali droplet method, respectively, with slight modifications. Responses of sugar, protein, and lipid contents after induction with AP-Toxin were assessed.【Result】 After induction with Ap-Toxin, protein, soluble sugar, total sugar, and lipids contents were increased in secretions of all B. pervariabilis × D. grandis varieties. Moreover, the disease index of all varieties increased significantly. The resistance of B. pervariabilis× D. grandis to A. phaeospermum infestation was enhanced. Among the three varieties, the disease index of No.6 was the lowest and its secretory response was the weakest, whereas the disease index of No. 8 was the highest and its secretory response was the strongest. Correlation analyses showed that protein, soluble sugar, total sugar, and lipid contents were significantly and positively correlated with the disease index and resistance, while the reducing sugar content was significantly and negatively correlated with disease resistance.【Conclusion】The resistance of B. pervariabilis × D. grandis varieties to A. phaeospermum could be enhanced using AP-Toxin as the induction factor. The stronger the resistance of the variety, the better the induction effect. Our results lay the theoretical foundation for the application of induced disease resistance for the management of forest pests.
蛋白毒素 / 诱导因子 / 撑绿杂交竹梢枯病 / 分泌物响应 / 暗孢节菱孢菌
AP-Toxin / induced factor / Bambusa pervariabilis × Dendrocalamopsis grandis / secretory response / Arthrinium phaeospermum
| [1] |
李姝江 . 撑绿杂交竹梢枯病菌蛋白毒素及其精确作用机制研究[D]. 雅安: 四川农业大学, 2013.
|
| [2] |
朱天辉, 黄宗超, 高强章 , 等. 撑×绿杂交竹梢枯病调查及空间分布型研究[J]. 四川林业科技, 2009,30(1):29-31, 37.
|
| [3] |
Arthrinium phaeospermum (Corda) M.B. Ellis is a globally distributed pathogenic fungus with a wide host range; its hosts include not only plants, but also humans and animals. This study aimed to develop genomic resources for A. phaeospermum to provide solid data and a theoretical basis for further studies of its pathogenesis, transcriptomics, proteomics, metabolomics and RNA genomics. The genome was obtained from the mycelia of the strain AP-Z13 using a combination of analyses with the high-throughput Illumina HiSeq 4000 system and PacBio RSII LongRead sequencing platform. Functional annotation was performed by BLASTing protein sequences against those in different publicly available databases to obtain their corresponding annotations. The genome is 48.45Mb in size, with an N90 scaffold size of 1,931,147bp, and encodes 19,836 putative predicted genes. This is the first report of the genome-scale assembly and annotation for A. phaeospermum, the first species in the genus Arthrinium to be subjected to whole genome sequencing.
|
| [4] |
马桂莲, 胡国良, 俞彩珠 , 等. 高节竹梢枯病病原菌及其生物学特性[J]. 浙江林学院学报, 2003,20(1):44-48.
|
| [5] |
朱天辉, 黄宗超, 高强章 , 等. 撑×绿杂交竹梢枯病病原及发生规律研究[J]. 中国森林病虫, 2009,28(2):10-12.
|
| [6] |
杨佐忠, 叶建仁 . 杂交竹枯梢病的病原鉴定[J]. 四川农业大学学报, 2004,22(3):225-227.
|
| [7] |
李姝江, 朱天辉, 李芳莲 , 等. 撑绿杂交竹梢枯病菌拮抗细菌筛选及其抗菌物质研究[J]. 中国生物防治, 2010,26(S1):55-61.
|
| [8] |
李姝江, 梁漫, 朱天辉 , 等. 撑绿杂交竹梢枯病拮抗菌的筛选及抗菌蛋白分析[J]. 南京林业大学学报(自然科学版), 2013,37(6):27-32.
|
| [9] |
黄宗超, 朱天辉 . 撑绿杂交竹枯梢病菌的室内杀菌剂筛选[J]. 中国森林病虫, 2007,26(3):35-38.
|
| [10] |
Bamboo blight in hybrid Bambusa pervariabilisxDendrocalamopsis grandis is caused by the fungus Arthrinium phaeospermum. One component designated as P-C with pathogenic activity and a molecular weight of about 27kDa was obtained from this fungus by ammonium sulphate precipitation, polyethylene glycol concentration, Sephadex G-100 gel chromatography and Q-Sepharose Fast Flow anion exchange resin. The toxin P-C was shown to cause the same typical symptoms caused by the pathogen itself, although the reaction of the toxin was quicker than the pathogen in all four hybrid bamboo varieties. A threshold toxic dose of 10-20g/ml was observed for two resistant varieties, while only 5-10g/ml was observed for two susceptible varieties. Moreover, effects of the purified toxic protein on the activities of superoxide dismutase, peroxidase, phenylalanine amonnialyase, polyphenol oxidase, chitinase and -1,3 glucanase in the different bamboo varieties were analysed using the impregnation and leaf disc methods. The results showed a positive correlation between the resistance of bamboo and the activities of all six enzymes. The activities of all enzymes except -1,3 glucanase decreased over the time course of investigation. These results proved the important role played by toxic proteins in the pathogenicity of A.phaeospermum and their potential for selecting resistant varieties or identify the resistance of this bamboo.
|
| [11] |
Bambusa pervariabilis x Dendrocalamopisis grandis blight is caused by a toxin produced by the fungus Arthrinium phaeospermum. In this study, a toxin fraction (P1-2-2) with an estimated molecular mass of 31 kDa was purified from a culture filtrate of this fungus by ammonium sulfate precipitation, Sephadex G-50 gel chromatography, Q Sepharose Fast Flow anion exchange resin, and Sephadex G-75 chromatography. The N-terminal amino acid sequence (i.e., H(2)N-Gln-Val-Arg-Asp-Arg-Leu-Glu-Ser-Thr) determined by Edman degradation showed homology to known serine alkaline proteases. The purified protein was named AP-toxin. Effects of the purified protein toxin on total phenol, flavonoid, total nucleic acid, DNA, RNA, soluble protein, and soluble sugar content, as well as DNase and RNase activities and disease index, were analyzed in different bamboo varieties by the impregnation method. The toxin had a significant effect on each parameter tested. In addition, a significant correlation was observed among the metabolic index, treatment time, bamboo resistance, and disease index. These data suggest that AP-toxin plays an important role in mediating the phytotoxic activities of A. phaeospermum. This study also indicates that metabolic indices could reflect the resistance indices of hybrid bamboo to blight.
|
| [12] |
|
| [13] |
Tomato plants resistant to the fungal pathogen, Fusarium oxysporum f. sp. lycopersici, race 2, were obtained using in vitro selection against fusaric acid, a non-specific toxin, as well as non-challenged cells. Protoplasts were isolated from cotyledonary tissue of tomato cv. 'UC-82', which is susceptible to Fusarium race 2. Protoplasts were challenged with the toxin, and the resistant calli were further subjected to the toxin. Plants regenerated from toxin-resistant calli were screened for resistance to the pathogen by using the Fusarium slurry inoculation technique. Seeds were collected from the surviving individuals, germinated and rescreened for resistance to the pathogen. Data obtained from this test showed a ratio of three resistant to one susceptible among R1 progenies. Further analysis of the R2 progenies confirmed that the fusarium-resistant plants were either homozygous or heterozygous dominant for the gene conferring the resistance. Similar results were recorded for tomato plants regenerated from cells that received no selection pressure. The nature of this single dominant gene-type of resistance is under investigation.
|
| [14] |
|
| [15] |
杨艳丽, 肖浪涛, 胡先奇 . 马铃薯晚疫病菌毒素与寄主品种抗性关系研究[J]. 中国农业科学, 2009,42(6):2202-2210.
|
| [16] |
何倩倩, 刘雨欣, 方馨玫 , 等. 撑绿杂交竹抗梢枯病诱导因子筛选及其持续期研究[J]. 植物保护, 2019,45(2):81-89.
|
| [17] |
|
| [18] |
方中达 . 植病研究方法[M]. 3版.北京: 中国农业出版社, 1998: 8-12.
|
| [19] |
王俊刚, 张树珍, 杨本鹏 , 等. 3,5-二硝基水杨酸(DNS)法测定甘蔗茎节总糖和还原糖含量[J]. 甘蔗糖业, 2008,27(5):45-48.
|
| [20] |
钟岩, 潘浦群, 王艳红 , 等. 苯酚-硫酸法测定鲜人参中多糖含量[J]. 时珍国医国药, 2008,19(8):1957-1958.
|
| [21] |
王晓丽, 苟琳 . 生物化学实验教程 [M]. 成都: 四川科学技术出版社, 2005: 53-54, 63, 72-73.
|
| [22] |
|
| [23] |
杨莉 . 暗孢节菱孢菌对撑×绿撑绿杂交竹致病机理的研究[D]. 雅安: 四川农业大学, 2009.
|
| [24] |
李姝江, 朱天辉, 杨莉 , 等. 暗孢节菱孢菌非蛋白类毒素对撑绿杂交竹生理代谢的影响[J]. 植物病理学报, 2011,41(6):587-595.
|
| [25] |
景康康 . 牛角花齿蓟马(Odontothrips loti)为害对苜蓿同化产物及矿质元素分配的影响[D]. 兰州: 甘肃农业大学, 2013.
|
| [26] |
敖雪, 邵世勤, 张少英 , 等. 甜菜抗(感)丛根病品种苗期某些生理生化特性差异的研究[J]. 中国甜菜糖业, 2005, ( 1):15-17.
|
| [27] |
|
| [28] |
|
| [29] |
程洁 . 白菜、甘蓝根肿病相关生理生化指标的分析[D]. 重庆: 西南大学, 2013.
|
| [30] |
|
| [31] |
王金生, 张学君, 方中达 . 马铃薯品种对细菌软腐病抗性生理指标的研究Ⅱ.品种还原糖含量及其与软腐病抗性的关系[J]. 南京农业大学学报, 1990,13(2):69-75.
|
| [32] |
龙书生, 李亚玲, 张宇宏 , 等. 糖分含量作为抗镰刀菌茎腐病玉米品种的育种指标研究[J]. 山东农业大学学报(自然科学版), 1999,30(4):372-376, 380.
|
| [33] |
丁九敏, 高洪斌, 刘玉石 , 等. 黄瓜霜霉病抗性与叶片中生理生化物质含量关系的研究[J]. 辽宁农业科学, 2005,5(1):11-13.
|
| [34] |
周博如, 李永镐, 刘太国 , 等. 不同抗性的大豆品种接种大豆细菌性疫病菌后可溶性蛋白、总糖含量变化的研究[J]. 大豆科学, 2000,19(2):111-114.
|
| [35] |
|
| [36] |
|
| [37] |
Two Bradyrhizobium japonicum, Tn5-induced, mutant strains, ML126 and ML150, were studied. Both induce host cell division to form normal-sized nodules that do not fix nitrogen and whose cells have very few bacteroids (Bar-). Early-infection (15 days post infection) cells have much endoplasmic reticulum (ER), numerous Golgi bodies, and large vacuoles that are probably secondary lysosomes. Later the cytoplasm of the host cells of both are dominated by hundreds of vesicles containing only finely fibrous material and that appear to originate by the degradation of the cell walls of the infection threads; they have been named
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