蓝莓叶斑病病原菌鉴定及其生物学特性

Identification and biological characteristics of the pathogen Pestalotiopsis clavispora causing blueberry leaf spot

为明确山东省蓝莓叶斑病病原菌的种类及其生物学特性,采用组织分离法获得菌株B1,通过形态学特征并结合r DNA-ITS序列分析对该菌株B1进行鉴定,以离体和活体叶片接种法测定菌株B1的致病性,并对其生物学特性进行研究。结果表明,菌株B1菌落呈白色绒毛状,背面淡黄色,分生孢子纺锤形,5个细胞,中间3个细胞为褐色,具有2～4根顶端附属丝;菌株B1的ITS序列与GenBank中棒状拟盘多毛孢Pestalotiopsis clavispora的相似性达99%以上,结合形态特征与rDNAITS序列分析将病原菌鉴定为棒状拟盘多毛孢,GenBank登录号为MG009201。菌株B1接种蓝莓叶片后产生褐色病斑,后期密生黑色分生孢子盘,与蓝莓田间自然发病症状一致。该菌株在5～35℃、pH 5～11范围内均可生长,最适温度范围为25～30℃,最适pH范围为5～9;但仅在25～30℃时产生分生孢子,最适产孢温度为25℃;光照条件对菌丝生长无明显影响,但连续黑暗有利于产孢;以葡萄糖为碳源时利用率最高,对乳糖利用效果最差;最适氮源为硝酸钠、硫酸铵和蛋白胨,几乎不能利用尿素。

In order to clarify the pathogen and the biological characteristics of blueberry leaf spot disease occurring in Shandong Province, the strain B1 was obtained by using tissue isolation method. The strain was identified based on morphological characteristics and rDNA-internal transcribed spacer(ITS)analysis;its pathogenicity was determined by inoculation in vitro and in vivo, and the optimum conditions for mycelium growth and sporulation were studied. The results showed that colonies of strain B1 were white villous, and the back was pale yellow. The conidia consisting of five cells were spindles with two to four hyaline appendages and three middle brown cells. The ITS sequence of strain B1 had over 99% homology with that of Pestalotiopsis clavispora isolates in GenBank, and the pathogen was identified as P. clavispora(accession No. MG009201). After inoculation of strain B1 on blueberry leaves, brown spots were observed and abundant black acervuli were produced in the later stage, which was similar to the diseased symptoms in the field. The strain B1 could grow under the conditions of 5-35℃ and pH 5-11, with the optimal growth temperature ranging from 25 to 30℃ and the optimal pH value at 5 to 9. However, the conidia were produced only at 25 to 30℃, and the optimal sporulation temperature was 25℃. The light conditions had no significant effect on mycelial growth of strain B1,but continuous darkness was beneficial to sporulation. For mycelial growth of strain B1, glucose was the optimum carbon source and lactose utilization was the worst. The optimum nitrogen sources were sodium nitrate, ammonium sulfate and peptone;however, urea could hardly be utilized.