柬埔寨龙血树褐斑病的病原鉴定及其培养条件分析

Identification of the pathogen causing brown leaf spot in Dracaena cambodiana and analysis of its cultivation conditions

[目的]柬埔寨龙血树为我国各省市广泛栽培的室内观叶植物,近年来普遍发生叶部病害影响其观赏价值。2011年8月至2016年12月,在云南省蒙自市、新疆乌鲁木齐的柬埔寨龙血树发现一种叶部初期褪绿逐渐变为灰褐色凹陷最后暗褐色枯死的褐斑病。为了确定其病原菌种类和发病条件,进行病原菌鉴定及培养条件测试。[方法]采用在云南蒙自、新疆乌鲁木齐采集的12份柬埔寨龙血树褐斑病病叶,开展了组织分离、致病性测试、形态学观察、分子鉴定与培养条件测试。[结果]从12份病叶分离到15株真菌,菌落中央产生黑色孢子,边缘白色,顶囊球形或近球形,直径38~45μm,分生孢子球形暗褐色,直径2~5μm,代表性菌株LZB-6、LZB-12经CaM或β-tubulin系统发育分析确定为塔宾曲霉;刺伤法接种14 d的龙血树叶片均发病,症状与田间一致,并能分离到菌落形态、分生孢子形态与原分离菌株一致的真菌,完成科赫法则验证确定为病原菌。培养条件测试表明,病原菌菌丝生长的适宜温度为22~34℃,最适宜温度为28℃,光照抑制菌丝生长,菌丝生长偏爱碱性环境,pH为7~10时,随着pH值升高生长速率仍在增加,最适宜碳源和氮源分别为甘露醇、明胶,菌丝致死温度为55℃10 min;产孢适宜温度为28~40℃,最适宜温度为32℃,光照抑制产孢,适宜产孢pH值为6~8,最适宜pH值为8,最适宜碳源和氮源分别为蔗糖、明胶,孢子的致死温度为60℃10 min。[结论]导致云南蒙自、新疆乌鲁木齐的柬埔寨龙血树褐斑病菌均为塔宾曲霉,除了光照和氮源之外,温度、pH值、碳源对菌丝生长和产孢的影响均不同,22~34℃时病原菌能较好的生长与产孢,应加强对柬埔寨龙血树褐斑病的防控。

[Objective]Dracaena cambodiana is a widely cultivated indoor ornamental plant in various provinces of China.In recent years,a common leaf disease has affected its ornamental value.Between August 2011 and December 2016,a brown spot disease was discovered in the leaves of D.cambodiana in Mengzi City,Yunnan Province and Urumqi,Xinjiang.The disease started with the leaves turning chlorotic,gradually becoming grayish-brown,then developing sunken areas,and eventually turning dark brown and dying.In order to determine the type of pathogen and the conditions under which the disease developed,we conducted pathogen identification and cultivation condition tests.[Methods]A total of 12 samples of Cambodian dragon tree leaves with brown spot disease were collected from Mengzi,Yunnan,and Urumqi,Xinjiang.These samples were used for tissue isolation,pathogenicity testing,morphological observation,molecular identification,and cultivation condition testing.[Results]Fifteen fungal strains were isolated from the 12 diseased leaf samples.The colonies had black spores at the center,with white edges.The spore bodies were spherical or nearly spherical,with a diameter of 38~45 μm.The conidia were dark brown,spherical,and 2~5 μm in diameter.Phylogenetic analysis of representative strains using CaM or β-tubulin confirmed that the pathogen Aspergillus tubingensis;Inoculation of the leaves of the dragon tree with the pathogen caused disease within 14 days,and the symptoms matched those observed in the field.Fungi with colony and conidia morphology consistent with the original isolates were re-isolated,confirming the pathogen through Koch's postulates.Cultivation condition tests showed that the optimal temperature for mycelial growth was 22~34 ℃,with the most suitable temperature being 28 ℃.Light inhibited mycelial growth,and the fungus preferred an alkaline environment.The growth rate increased with a pH range of 7~10,with the best growth at pH 8.The most suitable carbon and nitrogen sources were mannitol and gelatin,respectively.The lethal temperature for the mycelium was 55 ℃ for 10 minutes.The optimal temperature for spore production was 28~40 ℃,with the most suitable temperature being 32 ℃.Light inhibited spore production,and the optimal pH for spore production was 6~8,with the most suitable pH being 8.The most suitable carbon and nitrogen sources for spore production were sucrose and gelatin,respectively.The lethal temperature for spores was 60 ℃ for 10 minutes.[Conclusion]A.tubingensis was causal agent of brown leaf spot on D.cambodiana in Mengzi,Yunnan and Urumqi,Xinjiang.Except for light and nitrogen sources,temperature,pH value,and carbon sources all had varying effects on mycelial growth and spore production.The pathogen grows and sporulates best between 22~34 ℃.It is recommended to strengthen control measures for brown leaf spot in the D.cambodiana.