厚皮甜瓜（cucumis melo L.）主要采后病害的潜伏侵染及生物学控制
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摘要
本文以厚皮甜瓜为试材，研究了厚皮甜瓜果实主要采后病害的潜伏侵染。同时，利用诱抗剂BTH和Harpin，芽孢杆菌B_1、B_2对厚皮甜瓜采后病害的控制及机理进行了初步研究。
黑斑病和白斑病是哈密瓜低温贮藏期间发生的主要病害，它们分别由链格孢(Alternaria alternata(Fr.)keissl)和镰刀菌(Fusarium spp.)潜伏侵染引起。其中，链格孢在采前可通过两种途径侵入甜瓜果实：一是自花期开始侵入，经由花柱进入子房，形成花期潜伏侵染；二是在果实生长发育过程中，通过果实表皮的自然孔口及表皮网纹侵入，潜伏于皮下组织。镰刀菌在网纹形成期才开始侵染甜瓜果实。网纹形成期是病原真菌潜伏侵染哈密瓜果实的关键时期。
利用柱层析法分离不同发育期哈密瓜果实果皮甲醇-二氯甲烷提取物，并以链格孢为指示菌进行生物活性测定，发现果皮中存在抗菌物质，经色谱-质谱联用(GC／MS)分析，邻苯二甲酸二-(2-乙基)-己酯、邻苯二甲酸二辛酯、邻苯二甲酸二丁酯、棕榈酸甲酯、亚油酸甲酯、亚麻酸甲酯等可能为哈密瓜果实果皮中的预先合成抗菌物质。In vitro试验表明，邻苯二甲酸二甲酯具有直接抑菌作用。在果实生长发育过程中，预合成抗菌物质的种类和含量都呈下降趋势，果实膨大期种类最多，含量最高，至成熟时很难检测到。预合成抗菌物质是果实潜伏侵染机制之一。
采后化学诱抗剂BTH和Harpin处理能够诱导哈密瓜果实产生对采后黑斑病的抗性。诱抗作用随诱抗剂浓度的升高而增强，100mg／L的BTH效果最佳，处理后48小时接种黑斑病菌，贮藏7天时的病斑直径显著小于对照。
BTH和Harpin诱导果实采后抗病性的机理是产生病程相关蛋白(PRs)。采后100mg／L BTH和90mg／LHarpin处理均能诱导哈密瓜果实过氧化物酶和几丁质酶的活性升高。黑斑病(Alternaria alternata)和粉霉病(Trichothecium roseum)是“黄河蜜”甜瓜采后贮藏运输过程中的主要病害，芽孢杆菌B_1、B_2对“黄河蜜”甜瓜采后黑斑病和粉霉病具有显著的抑制效果。In vitro条件下，B_1、B_2活菌液对黑斑病的抑制率分别达到100％和94％，而对粉霉病的抑制率均达到100％。In vivo条件下，B_1、B_2活菌液能显著抑制“黄河蜜”甜瓜采后黑斑病和粉霉病的侵染和扩展。B_1、B_2代谢液显著抑制粉霉病的扩展，但对黑斑病的扩展无抑制效果。
Latent infection and biological control of main postharvest diseases on muskmelon were studied during 2001 and 2002.
Black spot caused by Alternaria alternata and white mould caused by Fusarium spp. occurred frequently after harvest on 'Hami' melons. Both of two pathogens could infect fruits at preharvest. There were two principal periods to infect melons for A.alternata in the field. One was at flowering period. The fungi initially infected the styles and settled in ovary, then kept quiescence till after harvest. Another happened during development of fruit. The pathogen penetrate the peel through epidermal lenticels and net in the surface of fruit, and remained latent in the underlying epidermal cells. For Fusarium spp., the infection occurred mainly during developing of fruit, and the nets formed in peel were important pathway. Net forming stage was the key period when latent fungi infected muskmelon.
The preformed antifungal compounds were extracted by ethanol-dichlormethane and column-chromatography from the peel of 'Hami' melon at different mature stage, and detected by bioassay on PDA. The possible anti-fungal substances were identified with gas chromatography and mass spectroscopy (GC/MS) as bis(2-ethylhexyl) pathalate, 1,2-benzenedicarboxylic acid,dioctyl ester, butyl phthalate, methyl palmitate, methyl linolelaidate, methyl linolenate. In vitro test indicated that diisoocytl adipates inhibited growth of Alternaria alternata directly. The species and content of anti-fungal substances decreased with the development and maturity of the muskmelon, little compounds were detected at harvest time. The results indicated that duration of latent infection could be determined by these compounds.
The resistance of 'Hami' melon to postharvest Alternaria rot could be induced by postharvest treatment of chemical inducers BTH and Harpin. BTH at 100mg/L showed more
effective. Fruits were treated with the chemical, then inoculated with the conidia of Alternaria alternata after 48 hours, the diameter of black spot were measured after stored 7 days at 20癈. The diameter of black spot of l00mg/L BTH treated fruit was smaller than the control significantly.
The mechanism of induced resistance by elicitors on melons was due to production of pathogenesis related proteins (PRs). Activities of Chitinase and Peroxidase (POD) increased in treated fruit at lOOmg/L BTH and 90mg/L Harpin.
Alternaria rot and Pink rot (caused by Trichothecium roseum} were the main postharvest diseases on 'Huanghemi' melon. Two isolates of Bacillus (B1 and B2) were tested as antagonists for postharvest biological control of Alternaria rot and Pink rot on 'Huanghemi' melon. The inhibition rate of B1 and B2 cell culture to Alternaria alternata were 100% and 94% respectively on PDA, both inhibition rates to Trichothecium roseum were 100%. The cell-free filtrates of B1 and B2 by means of centrifugation and filtration also inhibited A.alternata and T. roseum in vitro compared with control significantly. The results of in vivo test showed that the cell cultures of B1 and B2 inhibited the infection rate and lesion development of A.alternata and T. roseum significantly. The cell-free filtrates of B1 and B2 inhibited T. roseum in vivo significantly.
黑斑病和白斑病是哈密瓜低温贮藏期间发生的主要病害，它们分别由链格孢(Alternaria alternata(Fr.)keissl)和镰刀菌(Fusarium spp.)潜伏侵染引起。其中，链格孢在采前可通过两种途径侵入甜瓜果实：一是自花期开始侵入，经由花柱进入子房，形成花期潜伏侵染；二是在果实生长发育过程中，通过果实表皮的自然孔口及表皮网纹侵入，潜伏于皮下组织。镰刀菌在网纹形成期才开始侵染甜瓜果实。网纹形成期是病原真菌潜伏侵染哈密瓜果实的关键时期。
利用柱层析法分离不同发育期哈密瓜果实果皮甲醇-二氯甲烷提取物，并以链格孢为指示菌进行生物活性测定，发现果皮中存在抗菌物质，经色谱-质谱联用(GC／MS)分析，邻苯二甲酸二-(2-乙基)-己酯、邻苯二甲酸二辛酯、邻苯二甲酸二丁酯、棕榈酸甲酯、亚油酸甲酯、亚麻酸甲酯等可能为哈密瓜果实果皮中的预先合成抗菌物质。In vitro试验表明，邻苯二甲酸二甲酯具有直接抑菌作用。在果实生长发育过程中，预合成抗菌物质的种类和含量都呈下降趋势，果实膨大期种类最多，含量最高，至成熟时很难检测到。预合成抗菌物质是果实潜伏侵染机制之一。
采后化学诱抗剂BTH和Harpin处理能够诱导哈密瓜果实产生对采后黑斑病的抗性。诱抗作用随诱抗剂浓度的升高而增强，100mg／L的BTH效果最佳，处理后48小时接种黑斑病菌，贮藏7天时的病斑直径显著小于对照。
BTH和Harpin诱导果实采后抗病性的机理是产生病程相关蛋白(PRs)。采后100mg／L BTH和90mg／LHarpin处理均能诱导哈密瓜果实过氧化物酶和几丁质酶的活性升高。黑斑病(Alternaria alternata)和粉霉病(Trichothecium roseum)是“黄河蜜”甜瓜采后贮藏运输过程中的主要病害，芽孢杆菌B_1、B_2对“黄河蜜”甜瓜采后黑斑病和粉霉病具有显著的抑制效果。In vitro条件下，B_1、B_2活菌液对黑斑病的抑制率分别达到100％和94％，而对粉霉病的抑制率均达到100％。In vivo条件下，B_1、B_2活菌液能显著抑制“黄河蜜”甜瓜采后黑斑病和粉霉病的侵染和扩展。B_1、B_2代谢液显著抑制粉霉病的扩展，但对黑斑病的扩展无抑制效果。
Latent infection and biological control of main postharvest diseases on muskmelon were studied during 2001 and 2002.
Black spot caused by Alternaria alternata and white mould caused by Fusarium spp. occurred frequently after harvest on 'Hami' melons. Both of two pathogens could infect fruits at preharvest. There were two principal periods to infect melons for A.alternata in the field. One was at flowering period. The fungi initially infected the styles and settled in ovary, then kept quiescence till after harvest. Another happened during development of fruit. The pathogen penetrate the peel through epidermal lenticels and net in the surface of fruit, and remained latent in the underlying epidermal cells. For Fusarium spp., the infection occurred mainly during developing of fruit, and the nets formed in peel were important pathway. Net forming stage was the key period when latent fungi infected muskmelon.
The preformed antifungal compounds were extracted by ethanol-dichlormethane and column-chromatography from the peel of 'Hami' melon at different mature stage, and detected by bioassay on PDA. The possible anti-fungal substances were identified with gas chromatography and mass spectroscopy (GC/MS) as bis(2-ethylhexyl) pathalate, 1,2-benzenedicarboxylic acid,dioctyl ester, butyl phthalate, methyl palmitate, methyl linolelaidate, methyl linolenate. In vitro test indicated that diisoocytl adipates inhibited growth of Alternaria alternata directly. The species and content of anti-fungal substances decreased with the development and maturity of the muskmelon, little compounds were detected at harvest time. The results indicated that duration of latent infection could be determined by these compounds.
The resistance of 'Hami' melon to postharvest Alternaria rot could be induced by postharvest treatment of chemical inducers BTH and Harpin. BTH at 100mg/L showed more
effective. Fruits were treated with the chemical, then inoculated with the conidia of Alternaria alternata after 48 hours, the diameter of black spot were measured after stored 7 days at 20癈. The diameter of black spot of l00mg/L BTH treated fruit was smaller than the control significantly.
The mechanism of induced resistance by elicitors on melons was due to production of pathogenesis related proteins (PRs). Activities of Chitinase and Peroxidase (POD) increased in treated fruit at lOOmg/L BTH and 90mg/L Harpin.
Alternaria rot and Pink rot (caused by Trichothecium roseum} were the main postharvest diseases on 'Huanghemi' melon. Two isolates of Bacillus (B1 and B2) were tested as antagonists for postharvest biological control of Alternaria rot and Pink rot on 'Huanghemi' melon. The inhibition rate of B1 and B2 cell culture to Alternaria alternata were 100% and 94% respectively on PDA, both inhibition rates to Trichothecium roseum were 100%. The cell-free filtrates of B1 and B2 by means of centrifugation and filtration also inhibited A.alternata and T. roseum in vitro compared with control significantly. The results of in vivo test showed that the cell cultures of B1 and B2 inhibited the infection rate and lesion development of A.alternata and T. roseum significantly. The cell-free filtrates of B1 and B2 inhibited T. roseum in vivo significantly.
引文
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