Study on the Resistance to Dicarboximide and Benzamide Fungicides in Botrytis Cinerea from Tomato and Strawberry in Hubei Province,China

摘要

(厌)葡萄孢（Botrytis cinerea）是包括番茄和草莓在内的超过200种寄主植物的病原菌，其引起的(厌)霉病是世界范围的严重病害。本研究调查了采自番茄和草莓的(厌)葡萄孢对二甲酰亚胺类和苯甲酰胺类杀菌剂的抗性发生情况，同时对抗性菌株进行了抗性分子检测和适合度测定。主要结果如下:
　　1)草莓和番茄是世界上种植的重要经济作物。中国作为重要的草莓种植国和最大的番茄生产国，深受(厌)霉病的困扰。本研究测定了从湖北省不同地区番茄和草莓保护地采集的(厌)葡萄孢对杀菌剂腐霉利的敏感性。腐霉利敏感和抗性菌株的平均EC50值（抑制50％生长时的有效浓度）分别为0.25μg/ml和3.02μg/ml。抗性菌株(ProR)的频率为16％，最高的频率发生在荆门。腐霉利抗性菌株对其他二甲酰亚胺类杀菌剂存在正交互抗性，但对苯吡咯类杀菌剂没有交互抗性。敏感菌株和抗性菌株的适合度，比如菌丝生长、渗透压敏感性和致病力，两者之间存在显著差异。此对BcOs1基因的氨基酸序列发现腐霉利抗性菌株在第365位密码子(舍)有点突变(I365S)，或者在369位密码子和373位密码子存在一对点突变(Q369P，N373S)。
　　2)本研究还测定了以上(厌)葡萄孢菌株对苯甲酰胺类杀菌剂苯酰菌胺的敏感性。经菌丝生长分析，苯酰菌胺敏感(ZoxS)和抗性菌株(ZoxR)的平均EC50值分别为0.27μg/ml和4.98μg/ml。来自八个地点的菌株约有8.5％对苯酰菌胺有抗性。在所检测的样品中，最高的抗性频率发生在荆门。苯酰菌胺抗性菌株对多菌灵存在正交互抗性。敏感菌株和抗性菌株的适合度之间不存在显著差异，表明抗性发生风险较高。对(厌)葡萄孢的TUB2基因进行序列分析发现三个之前报道的点突变(E198A，E198K，F200Y)和一个新点突变(T351I)。这个新突变总出现在舍有E198K点突变的菌株中。在高抗菌株中则检测到点突变F200Y。新点突变T351I和所有其他突变的发现表明抗性发生的风险与日俱增。本研究较好地阐明了中国湖北省(厌)葡萄孢对常用杀菌剂的抗性发生特点，结果有利于提高杀菌剂抗性的治理策略。

目录

声明

Table of Contents

ABSTRACT

摘要

List of Abbreviations

CHAPTER 1．REVIEW OF LITERATURE

1．1．1．Nomenclature and systematics

1．1．2．Biology，host range，and distribution

1．1．3．Adaptability and diversity

1．1．4．Epidemiology of B．cinema

1．2．Gray mold diseases

1．2．1 Gray mold on strawberries

1．2．2．Gray mold on tomatoes

1．3．Management and Control

1．3．1 Integrated management

1．3．2．Chemical control of B．cinerea

1．3．3 Fungicide resistance of B．cinerea

1．4．Objectives of study

CHAPTER 2．DETECTION AND MOLECULAR CHARACTERIZATION OF RESISTANCE TO THE PROCYMIDONE IN B．CINEREA FROM TOMATO AND STRAWBERRY IN HUBEI PROVINCE，CHINA

2．1．Introduction

2．2．Materials and methods

2．2．1．Fungicides

2．2．2．Source of B．cinerea isolates

2．2．3．Screening for resistance to fungicides

2．2．4．Sensitivity of B．cinerea mycelial growth to procymidone

2．2．5．Cross-resistance

2．2．6．Stability of resistant isolates

2．2．7．Fitness of resistant isolates

2．2．8．Efficacy of fungicides for control of B．cinerea

2．2．9．DNA extraction

2．2．10．Amplification and sequencing of BcOs1 gene

2．3．Results

2．3．1．Frequency distribution of resistant isolates

2．3．2．Sensitivity of B．cinerea to procymidone

2．3．3．Cross-resistance

2．3．4．Stability of resistant isolates

2．3．5．Fitness of resistant isolates

2．3．6．Control efficacy for resistant isolates

2．3．7．Molecular characterization of BcOs1

2．4．Discussion

CHAPTER 3．CHARACTERIZALTION OF ZOXAMIDE RESISTANCE IN BOTRYTIS CINEREA FROM TOMATO AND STRAWBERRY

3．1．Introduction

3．2．Materials and methods

3．2．1．Fungicides

3．2．2．Collection of B．cinerea isolates

3．2．3．Screening of B．cinerea isolates to zoxamide

3．2．3．Sensitivity of B．cinema to zoxamide

3．2．4．Cross-resistance evaluation

3．2．5．Evaluation of stability for zoxamide resistance

3．2．6．Fitness of zoxamide resistant isolates

3．2．7．Evaluation of control efficacy for zoxamide resistant isolates

3．2．8．Amplification and sequencing of TUB2 gene

3．3．Results

3．3．1．Frequency distribution of resistant isolates

3．3．2．Sensitivity of B．cinema to zoxamide

3．3．3．Cross-resistance of ZoxR isolates to carbendazim

3．3．4．Evaluation of resistance stability

3．3．5．Fitness of resistant isolates

3．3．6．Evaluation of control efficacy for ZoxR isolates

3．3．7．Molecular characterization of TUB2 gene

3．4．Discussion

CHAPTER 4．SUMMARY AND PERSPECTIVES

REFERENCES

APPENDIX

PUBLICATIONS

ACKNOWLEDGMENT