豌豆茎基腐病病原菌分离鉴定及其对杀菌剂的敏感性测定

doi:10.11983/CBB22169

植物学报 ›› 2023, Vol. 58 ›› Issue (1): 132-139.DOI: 10.11983/CBB22169

所属专题：杂粮生物学专辑 (2023年58卷1期)

豌豆茎基腐病病原菌分离鉴定及其对杀菌剂的敏感性测定

宋雅静¹^,², 欧晋稳²^,³, 张古文², 冯志娟², 卜远鹏², 王斌², 龚亚明², 徐建强¹^,^*(), 刘娜²^,^*()

¹河南科技大学园艺与植物保护学院, 洛阳 471003
²浙江省农业科学院蔬菜研究所, 农产品质量安全危害因子与风险防控国家重点实验室, 农业农村部南方豆类蔬菜种质创新与分子育种重点实验室, 杭州 310021
³浙江农林大学园艺科学学院, 杭州 311300

收稿日期:2022-07-27 接受日期:2022-11-14 出版日期:2023-01-01 发布日期:2023-01-05
通讯作者: *E-mail: xujqhust@126.com;ln200811@163.com
基金资助:
浙江省基础公益研究计划(LGN20C150006);浙江省“十四五”蔬菜新品种选育重大科技专项(2021C02065-6-2);国家自然科学基金(31872114)

Pathogen Identification of Pea Crown Rot and Its Sensitivity to Fungicides

Yajing Song¹^,², Jinwen Ou²^,³, Guwen Zhang², Zhijuan Feng², Yuanpeng Bu², Bin Wang², Yaming Gong², Jianqiang Xu¹^,^*(), Na Liu²^,^*()

¹College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471003, China
²Key Laboratory of Vegetable Legumes Germplasm Enhancement and Molecular Breeding in Southern China, Ministry of Agriculture and Rural Affairs, State Key Laboratory for Managing Biotic and Chemical Threats to the Quality and Safety of Agro-products, Institute of Vegetables, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
³College of Horticulture Science, Zhejiang Agriculture and Forestry University, Hangzhou 311300, China

Received:2022-07-27 Accepted:2022-11-14 Online:2023-01-01 Published:2023-01-05
Contact: *E-mail: xujqhust@126.com;ln200811@163.com

摘要/Abstract

摘要： 豌豆(Pisum sativum)是我国重要的豆类经济作物, 病害对豌豆生产造成重大经济损失。通过形态学观察、分子鉴定以及致病性测定, 最终确定引起豌豆茎基腐病的3种病原菌分别为尖孢镰刀菌(Fusarium oxysporum)、芸苔链格孢菌(Alternaria brassicae)和格氏镰刀菌(F. grosmichelii), 优势菌株为尖孢镰刀菌, 分离率为53.6%。室内毒力测定结果表明, 5种供试杀菌剂对3种病原菌的菌丝生长均有抑制作用, 其中咯菌腈和戊唑醇的抑菌效果最好。研究结果为豌豆茎基腐病的防治提供了科学依据。

关键词: 豌豆, 茎基腐病, 分离与鉴定, 杀菌剂, 敏感性测定

Abstract: Pea (Pisum sativum) is one of the most important economic crops of beans in China. The occurrence of diseases will cause great economic losses in pea production. Through morphological observation, molecular identification and pathogenicity determination, the results showed that the three kinds of pathogenic fungi causing pea crown rot were Fusarium oxysporum, Alternaria brassicae and F. grosmichelii, among which F. oxysporum was the most pathogenic, with the isolation frequency of 53.6%. The results of indoor toxicity measurement showed that five fungicides had inhibitory effects on the mycelial growth of three kinds of pathogenic bacteria, among which, the inhibition effect of fludioxonil and tebuconazole have top results. The results provide a scientific basis for the prevention and control of pea crown rot.

Key words: pea, crown rot, isolation and identification, fungicides, sensitivity determination

宋雅静, 欧晋稳, 张古文, 冯志娟, 卜远鹏, 王斌, 龚亚明, 徐建强, 刘娜. 豌豆茎基腐病病原菌分离鉴定及其对杀菌剂的敏感性测定. 植物学报, 2023, 58(1): 132-139.

Yajing Song, Jinwen Ou, Guwen Zhang, Zhijuan Feng, Yuanpeng Bu, Bin Wang, Yaming Gong, Jianqiang Xu, Na Liu. Pathogen Identification of Pea Crown Rot and Its Sensitivity to Fungicides. Chinese Bulletin of Botany, 2023, 58(1): 132-139.

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链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB22169

https://www.chinbullbotany.com/CN/Y2023/V58/I1/132

图/表 8

表1 5种杀菌剂的浓度设置

Table 1 Concentration settings of 5 fungicides

Fungicides	Concentration 2 (μg?mL^-1)	Concentration 3 (μg?mL^-1)	Concentration 4 (μg?mL^-1)	Concentration 5 (μg?mL^-1)
Tebuconazole	0.4	0.6	0.8	1.0
Carbendazim	1.0	1.5	2.0	2.5
Fludioxonil	0.05	0.1	0.2	0.4
Iprodione	1.5	2.0	2.5	3.0
Kresoximmethyl	0.2	0.5	0.8	1.0

图1 豌豆茎基腐病发病植株 (A) 豌豆茎基腐病发病症状(bar=5 cm); (B) 茎基部病状(bar= 2 cm)

Figure 1 Plants infected with pea crown rot (A) Symptoms of pea crown rot (bar=5 cm); (B) Symptoms of crown rot (bar=2 cm)

图2 豌豆茎基腐病病原菌YD-7、YD-21和YD-28的形态特征 (A1)-(A3) YD-7的菌落及孢子形态; (B1)-(B3) YD-21的菌落及孢子形态; (C1)-(C3) YD-28的菌落及孢子形态。(A1), (A2), (B1), (B2), (C1), (C2) Bars=2 cm; (A3), (B3), (C3) Bars=25 μm

Figure 2 Morphological characteristics of pathogenic fungus YD-7, YD-21 and YD-28 of pea crown rot (A1)-(A3) Colony and spore morphology of YD-7; (B1)-(B3) Colony and spore morphology of YD-21; (C1)-(C3) Colony and spore morphology of YD-28. (A1), (A2), (B1), (B2), (C1), (C2) Bars=2 cm; (A3), (B3), (C3) Bars=25 μm

图3 豌豆茎基腐病病原菌PCR扩增产物凝胶电泳及相关近缘种的系统发育树 (A) 分离菌株YD-7、YD-21和YD-28的PCR扩增产物凝胶电泳; (B) 基于ITS及其同源序列构建的系统发育树

Figure 3 Agarose gel electrophoresis of PCR products of pea crown rot pathogen and phylogenetic tree of related sibling species (A) Agarose gel electrophoresis of PCR products of isolated strains YD-7, YD-21 and YD-28; (B) Phylogenetic tree based on ITS and its homologous sequences

表2 菌株ITS序列长度

Table 2 ITS sequence length of the strains

Species	Isolates	Length of ITS (bp)	GenBank number	Species	Isolates	Length of ITS (bp)	GenBank number
Fusarium equiseti	BMS4	570	OM876892.1	A. solani	43	545	MN871610.1
F. falciforme	M232	530	MN907530.1	A. infectoria	F16	775	MW301214.1
F. solani	CFE-95	569	MW045606.1	Alternaria sp.	PYCC 8684	917	OK094892.1
F. oxysporum	S17F3	488	MK968009.1	Fusarium sp.	160	545	OM262322.1
F. grosmichelii	JXN4-10	514	OL744454.1	A. brassicae	W1 18S	589	JF439448.1
Alternaria brassicae	ALE-53	534	MF380676.1	F. nirenbergiae	9245	510	ON365710.1

图4 分离菌株YD-7、YD-21和YD-28与其它相似菌株核苷酸序列多重比对

Figure 4 Multiple alignment of nucleotide sequences of isolated strains YD-7, YD-21 and YD-28 with other similar strains

图5 分离菌株YD-7、YD-21和YD-28对浙豌2号的致病性 (A) 以接种PDA为空白对照; (B) YD-7对浙豌2号的致病性; (C) YD-21对浙豌2号的致病性; (D) YD-28对浙豌2号的致病性。Bars=5 cm

Figure 5 Pathogenicity of isolated strains YD-7, YD-21 and YD-28 to Zhewan 2 (A) Inoculate PDA as control; (B) Pathogenicity of YD-7 to Zhewan 2; (C) Pathogenicity of YD-21 to Zhewan 2; (D) Pathogenicity of YD-28 to Zhewan 2. Bars=5 cm

表3 不同杀菌剂对YD-7、YD-21和YD-28的毒力测定

Table 3 Toxicity measurement of different fungicides to YD-7, YD-21 and YD-28

Fungicides/Classification	Strain	Regression equation	EC₅₀ (μg?mL^-1)	Means±SD	r
Tebuconazole/Triazoles	YD-7	Y=0.8566X+5.1100	0.74	1.17±1.10	0.99
	YD-21	Y=2.0295X+4.1263	2.69		0.99
	YD-28	Y=0.7605X+5.8069	0.09		0.99
Carbendazim/Benzimidazoles	YD-7	Y=2.8425X+6.5769	0.28	8.28±10.96	0.97
	YD-21	Y=1.4663X+2.9820	23.78		0.99
	YD-28	Y=9.3029X+6.0223	0.78		0.96
Fludioxonil/Pyrroles	YD-7	Y=0.1054X+4.9459	3.26	1.12±1.52	0.99
	YD-21	Y=0.6536X+5.7413	0.07		0.99
	YD-28	Y=0.6242X+6.1209	0.02		0.99
Iprodione/Dicarbonamides	YD-7	Y=4.9338X+0.9405	6.65	3.44±2.46	0.97
	YD-21	Y=1.6099X+5.2905	0.66		0.99
	YD-28	Y=3.2891X+3.4191	3.02		0.98
Kresoximmethyl/Methoxy acrylates	YD-7	Y=0.9294X+4.9657	1.09	5.03±5.50	0.99
	YD-21	Y=1.0512X+3.8358	12.81		0.99
	YD-28	Y=1.0638X+4.9143	1.20		0.99

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豌豆茎基腐病病原菌分离鉴定及其对杀菌剂的敏感性测定

Pathogen Identification of Pea Crown Rot and Its Sensitivity to Fungicides

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