春兰叶斑病病原的鉴定和分析  

Identification and analysis of pathogens caused Cymbidium goeringii leaf spot

作　　者：罗维宇 梁嘉莉 张荣[1] 罗易华 冯淑杰[1] LUOWei-yu;LIANG Jia-li;ZHANG Rong;LUO Yi-hua;FENG Shu-jie(College of Horticulture,South China Agricultural University/Integrative Microbiology Research Centre of Guangdong Province,South China Agricultural University/College of Forestry and Landscape Architecture,South China Agricultural University,Guangzhou,Guangdong 510642,China)

机构地区：[1]华南农业大学园艺学院/华南农业大学广东省群体微生物研究中心/华南农业大学林学与风景学院,广东广州510642

出　　处：《南方农业学报》2023年第1期158-168,共11页Journal of Southern Agriculture

基　　金：广东省现代农业产业技术体系花卉创新团队项目(2021KJ121)。

摘　　要：【目的】明确野生春兰引种驯化过程中引发叶斑病的病原菌种类,为春兰叶斑病的诊断和防治提供科学依据。【方法】采集广东省广州市和贵州省遵义市典型春兰叶斑病的病叶,利用组织分离法对病原菌进行分离纯化;依据柯赫氏法则,通过刺伤接种和摩擦接种验证其致病性;根据病原菌形态学特征,结合ITS、ACT、CAL、EF-1a、GAPDH、GS和TUB2部分序列的多基因分子系统学分析等方法进行病原菌分类鉴定。【结果】从采集的3种不同类型症状的叶斑病病叶中共检测到4株致病菌,分别命名为GZH1、GZH2、GZH3和GZH4,其中GZH1和GZH4分离自广东广州采集的叶斑病病叶,GZH2和GZH3分离自贵州遵义采集的叶斑病病叶。致病性测定结果显示,菌株GZH1表现出较强的致病性,离体针刺接种发病明显;菌株GZH2、GZH3和GZH4离体针刺接种发病不明显,通过摩擦接种可在接种部位形成典型的坏死斑。菌株GZH1在PSA培养基上的菌落特征,大、小型分生孢子,小型分生孢子梗、厚垣孢子的形态及大小均符合尖镰孢(Fusarium oxysporum)的特征,其ITS和EF-1a部分序列与F. oxysporum菌株UACH-217相应序列的一致性分别达98.23%和97.24%,结合其多基因序列构建系统发育进化树分析,菌株GZH1被鉴定为尖镰孢(F. oxysporum)。菌株GZH2、GZH3和GZH4在PDA培养基上的菌落形态,分生孢子及其附着胞的形态和大小等,分别与兰花炭疽菌(Colletotrichum cymbidiicola)、果生炭疽菌(C. fructicola)和江西炭疽菌(C. jiangxiense)的特征相似;供试菌株GZH2的ITS、ACT、CAL、GAPDH和TUB2部分序列与C. cymbidiicola菌株CBS:123757对应序列的一致性分别达99.00%、100.00%、99.00%、99.00%、99.60%;供试菌株GZH3的ITS、ACT、CAL、GAPDH和TUB2部分序列与C. fructicola菌株ICMP:18613对应序列的一致性分别达99.00%、99.00%、99.00%、100.00%、99.70%;供试菌株GZH4的ITS、ACT、CAL、GAPDH、TUB2和GS部分序列与C. jiangxiense菌�【Objective】To identify pathogens causing the leaf spot disease during introduction and domestication of wild Cymbidium goeringii,and to provide scientific basis for diagnosis,prevention and control of C.goeringii leaf spot disease.【Method】C.goeringii leaves with typical leaf spot disease were collected from Guangzhou City,Guangdong Province and Zunyi City,Guizhou Province,and the pathogens were isolated and purified by tissue separation;according to Kochʼs rule,their pathogenicity was tested by acupuncture inoculation and friction inoculation in vitro.Based on their morphological characteristics,pathogen classification and identification were carried out through multi-gene molecular systematic analysis of partial sequences of ITS,ACT,CAL,EF-1a,GAPDH,GS and TUB2.【Result】In the collected leaves with 3 typical leaf spot diseases Symptoms,4 strains of pathogens were detected,and were named as GZH1,GZH2,GZH3 and GZH4.Among them,GZH1 and GZH4 were isolated from diseased leaves collected in Guangzhou City,Guangdong Province;GZH2 and GZH3 were isolated from diseased leaves collected in Zunyi City,Guizhou Province.The results of pathogenicity tests showed that strain GZH1 showed strong pathogenicity through acupuncture inoculation in vitro,while strains GZH2,GZH3 and GZH4 could cause typical necrotic spots at the inoculation site by friction inoculation.On PSA medium,strain GZH1 showed colony morphology,morphology and size of macroconidia,microconidia,microconidia conidiophore and chlamydospores were consistent with those of Fusarium oxysporum,and partial sequences of ITS and EF-1A of strain GZH1 had a consistency of 98.23%and 97.24%with F.oxysporum strain UACH-217;combined with phylogenetic tree analysis by multi-gene sequences,strain GZH1 was identified as F.oxysporum.On PDA medium,colony morphology,conidia morphology,conidia size,appressoria morphology and appressoria size of strains GZH2,GZH3 and GZH4 were highly consistent with those of Colletotrichum cymbidiicola,C.fructicola and C.jiangxiense,respectiv

关 键 词：春兰 叶斑病 鉴定 Fusarium oxysporum Colletotrichum spp 

分 类 号：S436.81[农业科学—农业昆虫与害虫防治] S682.31[农业科学—植物保护]