一株侵染黄桃果实的葡萄座腔菌菌株鉴定  

Identification of A Strain in Botryosphaeria Infecting Fruit of Yellow Peach Variety“Jinxiu”

作　　者：虞凡霜 强遥 秦双林 张凯东 刘冰[1] 熊桂红 蒋军喜[1] YU Fanshuang;QIANG Yao;QIN Shuanglin;ZHANG Kaidong;LIU Bing;XIONG Guihong;JIANG Junxi(College of Agronomy,Jiangxi Agricultural University,Nanchang 330045,China;Agricultural Technology Service Center of Changxing Town,Chongming District,Shanghai 201913,China)

机构地区：[1]江西农业大学农学院,江西南昌330045 [2]上海市崇明区长兴镇农业技术服务中心,上海201913

出　　处：《江西农业大学学报》2021年第3期566-573,共8页Acta Agriculturae Universitatis Jiangxiensis

基　　金：国家自然科学基金项目(31460452);江西省科技计划项目(20181ACF60017)。

摘　　要：【目的】明确从江西省井冈山市“锦绣”黄桃病果上分离到的1株葡萄座腔菌的种类归属及其致病性。【方法】对黄桃病果进行症状描述,使用PDA平板按照常规真菌分离法从病果中分离病菌并进行菌种纯化,观察该菌的培养性状和形态特征,测量其分生孢子器和分生孢子大小,根据观测值,依据相关文献对其进行形态学鉴定。提取该菌株基因组DNA,对rDNA-ITS、EF-1和β-tubulin基因分别进行PCR扩增和测序,将测定序列输入到NCBI中利用BLAST程序搜索其同源序列并构建系统发育树,根据序列同源性大小和亲缘发生关系,对该菌株进行分子生物学鉴定;采用菌饼接种法分别对健康的桃一年生枝条和果实进行刺伤接种,测定该菌株的致病性。【结果】该黄桃病果病斑褐色、圆形或不规则形,病健交界不清楚,病斑果皮较完好,内部果肉变软腐烂。从病果中分离获得形态特征一致的真菌菌落,取其中一个菌株进行观察,其菌落初呈白色,后从中央开始变为灰绿色,菌落背面为墨绿色,菌落蓬松。菌落生长迅速,生长速率达35.5 mm/d,其培养性状与已报道的葡萄座腔菌(Botryosphaeria dothidea)培养性状一致。分生孢子器球形或扁球形,大小221.2(214.2~244.8)μm×225.9(204~255)μm。分生孢子梭形、无色、单胞,大小22.6(18.7~28.6)μm×5.1(3.9~5.2)μm,其形态大小与B.dothidea的无性态七叶树壳梭孢(Fusicoccum aesculi)的分生孢子器和分生孢子形态大小相吻合;测定的rDNA-ITS、EF-1和β-tubulin 3个基因的序列长度分别为583,297,681 bp,其在GenBank中的序列登录号分别为MW202368、MW202401和MW202404,此3个序列与GenBank中葡萄座腔菌(B.dothidea)对应序列具有最高的同源性,均为100%。将本菌株的3个序列按rDNA-ITS、EF-1、β-tubulin顺序连接成一个大的串联序列,与从GenBank中下载连接的对应序列一起构建系统发育树,结果显示,该菌株与B.dothidea的各菌株聚[Objective]The objective of this study is to identify the species and pathogenicity of a strain in Botryosphaeria isolated from the diseased fruit of yellow peach variety"Jinxiu"in Jinggangshan City,Jiangxi Province.[Method]Symptoms of the diseased fruits of yellow peach were described.The pathogen of the diseased fruit was isolated and purified on PDA plates using the conventional fungal isolation method.The cultural characteristics and morphological features of the strain were observed,and the sizes of its pycnidia and conidia were measured.The strain was identified based on comparing its morphology and size with the corresponding description in the literature.Genomic DNA of the strain was extracted and used as a template to amplify rDNAITS,EF-1 and β-tubulin genes by PCR.The PCR products were sequenced and the determined sequences were sent to NCBI to search for their homologous sequences by using BLAST program.A phylogenetic tree was constructed based on the alignment of sequences here and those homologous sequences.Molecular identification of the strain was performed based on the sequence homology and phylogenetic relationship.To determine pathogenicity of the strain,healthy annual branches and fruits of peach were inoculated with stab wounds using mycelium plug method.[Results]The lesions on the peach fruits were brown,round or irregular,and the boundary between disease and health was unclear.The skin on the lesion was intact,and the flesh became soft and rotten.Fungal colonies with the same morphological characteristics were isolated from the diseased fruits.One of the strains was randomly selected for identification.The colony of the strain was initially white,and then turned grey green from the center,while the back of the colony was dark green.It was fluffy and grew rapidly,and the colony growth rate reached 35.5 mm·d-1.This character is consistent with that of B. dothidea in literature.Its pycnidia were spherical or oblate with a size of 221.2(214.2~244.8)μm×225.9(204~255)μm,and the conidia were s

关 键 词：黄桃果实腐烂病 病原鉴定 葡萄座腔菌 七叶树壳梭孢 

分 类 号：S436.621.12[农业科学—农业昆虫与害虫防治]