福建省一种蓝莓枝枯病病原鉴定

张小艳; 陈细红; 谢丽雪; 张立杰; 郑姗; 李韬

doi:10.19303/j.issn.1008-0384.2023.10.010

福建省一种蓝莓枝枯病病原鉴定

doi: 10.19303/j.issn.1008-0384.2023.10.010

1.
福建省农业科学院果树研究所，福建　福州　350013
2.
福州海关技术中心，福建　福州　350001

基金项目: 福建省科技计划公益类专项（2020R10280011）

详细信息

作者简介:
张小艳（1982 —），女，硕士，助理研究员，主要从事果树病虫害综合治理研究，E-mail：zxyan1204@163.com

通讯作者:
李韬（1969 —），男，学士，副研究员，主要从事果树遗传育种与病虫害研究，E-mail: leetao06@163.com

中图分类号: S436.63
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出版历程
- 收稿日期: 2023-06-30
- 修回日期: 2023-07-26
- 网络出版日期: 2023-10-25
- 刊出日期: 2023-10-28

Pathogen of Branch Blight on Blueberry Plants in Fujian

1.
Fruit Research Institute, Fujian Academy of Agricultural Sciences, Fuzhou, Fujian　350013, China
2.
Fujian Key Laboratory for Technology Research of Inspection and Quarantine, Technology Center of Fuzhou Customs District, Fuzhou, Fujian　350001, China

摘要

摘要: 目的明确福建省三明地区蓝莓枝枯病的病原菌，为该病害的有效防治提供理论依据。方法采集受害蓝莓枝条样品，通过常规组织分离法进行病原菌分离纯化，采用针刺法进行致病性测定完成柯赫氏法则验证，利用形态学特征并联合分子生物学，明确致病菌的分类地位。结果从病组织中分离获得菌株KW1-4，将其回接至健康蓝莓枝条后，接种部位出现干枯病斑，再次分离获得的菌株与接种菌株一致，表明该菌株是引起蓝莓枝枯病的病原菌。菌株KW1-4菌落圆形，白色至灰色，有2种类型分生孢子，α型分生孢子长椭圆形至梭形，具2个明显油球，（4.77～7.63）µm×（1.55～2.71）µm，β型分生孢子线形，无油球，（11.95～19.65）µm×（1.05～1.94）µm。采用ITS，TEF1-ɑ，β-tubulin3段基因联合构建系统发育树，供试菌株KW1-4与Diaporthe australiana聚在同一分支。对比形态学特征，确定菌株KW1-4为间座壳属真菌Diaporthe australiana。结论引起福建省三明市蓝莓枝枯病的病原菌是Diaporthe australiana，本研究首次报道Diaporthe australiana能够侵染蓝莓枝条。
- 蓝莓 /
- 枝枯病 /
- 间座壳菌 /
- 鉴定
Abstract: Objective The pathogen that caused the blueberry branch blight in Sanming, Fujian was identified. Method Microbes were isolated and purified from the blueberry branches showing anthracnose symptoms collected from the infected area. Pathogenicity of the isolates was tested by needle injection on the plant and verified by the Koch's method. Pathogen taxonomy was determined by morphological observations and molecular technology. Result Strain KW1-4 was isolated from the diseased tissue and inoculated onto healthy blueberry branches to show on the inoculation site dry spots same as what produced by the strain from the infected plants. The white to gray, round colonies of KW1-4 had two types of conidia. The alpha conidia were elliptical or spindle in shape and 4.77-7.63 μm × 1.55-2.71 μm in size with two distinct oil globules, while the linear beta conidia sized 11.95-19.65 μm × 1.05-1.94 μm with no oil globule. The 3 fragments of the genes, ITS, TEF1-ɑ, and β-tubulin, were used to construct the phylogenetic tree that showed KW1-4 to cluster with Diaporthe australiana in a same branch, which agreed with the morphological characteristics. Conclusion It was determinedD. australiana to be the pathogen that caused the blight on blueberry bushes in Sanming. The identification paved the way for the disease control.
- Blueberry /
- branch blight /
- Diaporthe australiana /
- identification

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图 1 蓝莓枝枯病田间症状

Figure 1. Symptoms of blueberry branch blight in the field

下载: 全尺寸图片幻灯片

图 2 KW1-4的接种症状

Figure 2. Pathogenicity of KW1-4

下载: 全尺寸图片幻灯片

图 3 KW1-4 菌落形态及其分生孢子

A：3 d ；B：9 d ；C：25 d ；D：α型分生孢子； E：β型分生孢子。

Figure 3. Colony morphology and conidia of KW1-4

A: 3 d; B: 9 d; C: 25 d; D: alpha conidia; E: beta conidia.

下载: 全尺寸图片幻灯片

图 4 基于rDNA-ITS、TEF1-ɑ和β-tubulin基因序列构建的间座壳菌系统发育树

Figure 4. Phylogenetic tree of Diaporthe species constructed based on concatenated sequences of ITS, TEF1-ɑ, and β-tubulin

下载: 全尺寸图片幻灯片

表 1 PCR扩增ITS、TEF1-ɑ和β-tubulin片段的引物

Table 1. Primers for PCR amplification of fragments of ITS, TEF1-ɑ, and β-tubulin

基因 Gene	引物 Primer	序列（5′-3′） Sequence（5′-3′）	参考文献 Reference
ITS	ITS1	TCCGTAGGTGAACCTGCGG	[21]
ITS	ITS4	TCCTCCGCTTATTGATATGC	[21]
TEF1-ɑ	EF-728F	CATYGAGAAGTTCGAGAAGG	[22]
TEF1-ɑ	EF2	GGARGTACCAGTSATCATGTT	[23]
β-tubulin	Bt2a	GGTAACCAAATCGGTGCTGCTTTC	[24]
β-tubulin	Bt2b	ACCCTCAGTGTAGTGACCCTTGGC	[24]

下载: 导出CSV

表 2 ITS、TEF1-ɑ和β-tubulin引物的PCR反应条件

Table 2. PCR reaction conditions for fragments of ITS, TEF1-ɑ, and β-tubulin

扩增片段 Amplified fragment	预变性 Pre-denaturation	变性 Denaturation	退火 Annealing	延伸 Extension	循环次数 Cycle number	延伸 Extension
ITS	94 ℃，3 min	94 ℃，45 s	52 ℃，45 s	72 ℃，60 s	30	72 ℃，7 min
TEF1-ɑ	95 ℃，5 min	95 ℃，60 s	55 ℃，80 s	72 ℃，90 s	35	72 ℃，5 min
β-tubulin	95 ℃，5 min	95 ℃，60 s	45 ℃，80 s	72 ℃，90 s	35	72 ℃，5 min

下载: 导出CSV

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