果树学报2017，Vol.34Issue(10)：1323-1329,7.DOI:10.13925/j.cnki.gsxb.20170121
苯醚甲环唑对苹果树腐烂病菌茵丝形态和超微结构的影响
Effects of difenoconazole on the mycelial morphology and ultrastructure of Valsa mali
摘要
Abstract
[Objective] Apple tree Valsa canker is primarily caused by the ascomycetous Valsa species.Valsa mali is a worldwide and destructive disease in apple production areas,which can cause the bark rot of apple trees and then lead to serious economic losses.At present,the prevention to this disease mainly depends on the chemical agents,difenoconazole is a type of triazole fungicide,which could effectively prevent and cure Valsa canker,by inhibiting spore germination and mycelial growth.In this study,our aim is to determine the operation of difenoconazole on Valsa mali,and then to provide an important theoretical basis for the prevention and control of Valsa canker.[Methods] A 5 mm diameter disc from the V.mail culture was inoculated in the center of a PDA plate,and cultured at 25 ℃.One day after being cultured,the petri dishes were used for fungicide treament.Concentration of the fungicide was diluted from a stock solution 10 g·L-1 to 40 mg·L-1 with germ-free double-distilled water.Sepsis filter paper strips (0.5 cm×5 cm) were immersed in the fungicide and then placed in a square (5 cm×5 cm) around the fungal colony.For control,filter paper strips were immersed in 0.4％ acetone.After treatment,the petri dishes were continually cultured at 25 ℃ 12,24,48 h after fungicide treatment,and then small pieces of PDA from the colony edge were taken for mycelial morphology and ultrastructure studies.[Results]Observations from scanning with an electron microscope discovered that the growth of apical hyphae was inhibited and the top expanded into a sphere with a rough surface at 12 h after fungicide treatment compared with the control.Then 24 h after fungicide treatment,fungal colonies grew slowly and appeared to show a serious abnormality:hyphae were irregularly swelled and excessively branched,the hyphal cell walls gradually ruptured,and even showed the phenomenon of protoplasm extravasation.After 48 h with fungicide treament,colonies stopped their growth,and the hyphae collapsed,and the majority of the hyphal cell walls seriously ruptured.The part of the hyphae was swollen,and the hyphae were badly shriveled and even necrotic.Observations using a transmission electron microscope discovered that cell structure changed significantly at 12 h after treatment.The important change was the hyphal cell walls,which were irregularly thickened accompanied by an internal extending cytoplasm.On the other hand,the number of the mitochondria in the cytoplasm increased and its morphology was abnormally thickened.Moreover,it was also found that the nucleus of the cells changed to a certain extent:the number of nuclei increased and the nucleolus gradually spreaded and the area of nucleolar also decreased at the same time.When the hyphae were treated for 48 h with fungicide,the structure of the hyphea cell became worse:the whole cell walls became thickened and some of their exosporium disintegrated,and the membrane of the mitochondria showed irregularly thickened and malformed,and there was extracellular deep staining which maybe caused by cytoplasm leakage material.At this moment,the cell had some septum and part of the septum became irregularly thickened and malformed,and they could not even develop into a complete diaphragm.In addition,the vacuolization of the mycelial cell was very serious,some intracellular organelles disorganized even they disappeared completely,and the cytoplasms became necrosis.It is worthy of being mentioned that there was a new discovery:a daughter hypha could form in the original degenerated hypha.The cell walls became irregularly thickened,and the cytoplasms of the daughter hyphae with disorganized organelles were necrosis.However,the original degenerated hypha cell walls disappeared.[Conclusion] As a type of fungicide difenoconazole can affect the structure of the cell wall and the normal growth of the whole cell by affecting the morphology of the cell wall of Valsa mali,resulting in the hyphal malformation and cell necrosis.Specifically:the hyphae irregularly swelled and excessively branched and contorted,protoplasm extravasation occurred,cell walls irregularly thickened and disintegrated until completely obiterated,the membranes of the mitochondria irregularly thickened and malformed,the number of nuclei increased and the nucleolus gradually spread,the area of nucleolar decreased,a daughter hypha formed within the original degenerated hypha and the cell walls disappeared.关键词
苹果树腐烂病菌/苯醚甲环唑/形态/超微结构Key words
Valsa mali/Difenoconazole/Morphology/Ultrastructure分类
农业科技引用本文复制引用
高双,高小宁,冯浩,韩青梅,黄丽丽..苯醚甲环唑对苹果树腐烂病菌茵丝形态和超微结构的影响[J].果树学报,2017,34(10):1323-1329,7.基金项目
国家重点基础研究发展计划(2016YFD0201100) （2016YFD0201100）
