山药漆腐叶斑病病原菌的鉴定及其生物学特性研究

Identification and Biological Characteristics of the Pathogen Causing Myrothecium Leaf Spot on Chinese Yam

在河南山药(Dioscorea opposita)主产品种‘铁棍山药’生产中出现了一种新的叶部病害,田间发病株率为6.6%。该病害主要发生在植株下部叶片,病斑圆形褐色,具同心轮纹,后期病斑上产生大量黑色孢子堆。对该病害的病原菌进行了组织分离培养和柯赫氏法则验证。其分生孢子座碗状或浅盘状,分生孢子梗呈笤帚状分枝,分生孢子梗顶端着生棒状产孢细胞;分生孢子长椭圆形、棒形、长卵形或梨形,透明无色,不分隔,两端钝圆,大小为1.7~2.5μm×4.9~6.8μm。病原菌菌落白色绒毛状,产生大量墨绿色胶质状分生孢子团,呈环状排列。用ITS通用引物对病原菌的基因组DNA进行扩增并测序(序列在GenBank的登录号为KY369166),同源性比对结果显示其与露湿漆斑菌(Myrothecium roridum)的一致性达到99%,结合形态学鉴定结果,确定该病害的病原菌为露湿漆斑菌。病原菌的最适生长温度为30℃,分生孢子致死温度为52℃,pH 5~10之间该菌生长良好,光暗(12 h/12 h)交替最有利于病原菌产孢,菌丝生长的最适碳氮源为乳糖和丙氨酸,产孢的最适碳氮源为果糖和酵母。

A new foliar disease on Chinese yam （Dioscorea opposita） was observed in Jiaozuo city, Henan province. The spots with a size ranged from 3 - 18 mm in diameter show circular and pale to dark brown concentric rings on the leaves. There were black spore masses on older lesions. It was isolated and identified according Koch＇s Postulates. The single spore （with a isolation number of 0901 ） was isolated from the symptomatic portions of the leaves. The pathogenicity test has shown that the pure isolate 0901 was the pathogen causing the disease. Morphological characteristics of 0901 were consistent with the descriptions of Myrothecium roridum. Genomic DNA was extracted from mycelia and which was amplified as a template with the universal primer pair of fungal rDNA ITS 1 and ITS4, then the fragment was sequenced. The sequence showed highest identity （99%） to M. roridum. It was concluded that M. roridum was a new pathogen in Chinese yam in Henan province, China. The biological characteristics of the pathogen were tested. The colony spread fastest under 30℃, 12 h light/dark condition. The pathogen could grow well under pH 5 - 10, conidia lethal temperature was 52 ℃. The most suitable carbon and nitrogen sources for mycelia growth were lactose and alanine, and the optimum carbon and nitrogen sources were fructose and yeast for spore production.