湖北省葡萄主产区灰葡萄孢菌多样性分析

Diversity of Botrytis cinerea in major grape production regions of Hubei Province

【目的】了解湖北省葡萄主产区灰葡萄孢菌(Botrytis cinerea)多样性,为葡萄灰霉病防治提供科学依据。【方法】采用常规微生物分离法对从湖北省8个葡萄主产区采集的葡萄灰霉病样本进行灰葡萄孢菌分离,在PDA培养基上观察分离物菌落培养形态并测定菌丝生长速率;利用特异引物对各分离物的Flipper和Boty转座子片段进行扩增,区分转座子类型;扩增Bc-hch基因并用Hha I酶切检测分离物的多态性以区分组群;依据菌丝生长速率、菌落形态及转座子类型挑选4株典型分离物在4个葡萄品种果实上进行致病力测定。【结果】从病害样本中共分离获得51株分离物,菌落培养形态表明30株分离物为菌核型,20株为孢子型,仅1株为菌丝型,出现频率分别为58.82%、39.22%和1.96%。所有分离物菌丝生长速率均较高,在10.86~13.94 mm/d。分离物只存在2种转座子类型,其中50株为Transposa型,仅1株为Flipper型。Bc-hch基因Hha I酶切多态性鉴定结果表明,所有菌株均为Group II,为狭义的灰葡萄孢菌。分离物致病力测定结果表明,菌丝生长速率最低的WH3在供试葡萄品种上的致病力均最强,其次是Flipper型分离物SX1,菌丝生长速率最高的XN2和菌丝型WH6分离物致病力均较弱,且不同分离物在不同品种上致病力趋势不同。【结论】湖北省葡萄主产区灰葡萄孢菌菌落培养形态较丰富,所有分离物均为狭义灰葡萄孢菌,只存在Transposa和Flipper 2种转座子类型,前者占绝对优势,典型分离物间致病力差异明显。

【Objective】To assess the diversity of Botrytis cinerea in major grape production regions of Hubei Province,so as to provide scientific support for grape gray mold control.【Method】Isolates of B.cinerea from 8 main grape production regions in Hubei Province were isolated by conventional microbial separation method.All the isolates were cultured on PDA medium to observe the morphological characteristics and measure the mycelium growth rates.Transposon segments Flipper and Boty were amplified using specific primers to detect the transposon types.Bc-hch gene was amplified and detected by Hha I digestion to identify the group.Four representative isolates were used for pathogenicity analysis by inoculating on fruits of 4 grape varieties.【Result】Totally,51 isolate were obtained.Results of the morphological analysis showed that:Among them,30 isolates were the sclerotia type with the frequency of 58.82%,which was the most prevalent type;twenty isolates were the conidial type with the frequency of 39.22%;only 1 isolate was the mycelium type with the frequency of 1.96%.Mycelium growth rates of all isolates were high and ranged from 10.86 mm/d to 13.94 mm/d.Only two transposon types were found in isolates:50 were Transposa and only 1 was Flipper.All the isolates were classified into group II according to the restriction polymorphism analysis of enzyme Hha I of Bc-hch gene.Pathogencity analysis revealed that,isolate WH3 which had the lowest mycelium growth rate had the strongest pathogenicity.SX1,the only Flipper type isolate,had the second strongest pathogenicity.Pathogenicity of XN2 which had the highest growth rate and WH6 which was the only mycelium type were weaker.Pathogenicity regulations of isolates on different varieties were different.【Conclusion】Morphological characteristics of B.cinerea in the major grape production regions of Hubei Province is abundant.All the isolates are identified as B.cinerea in a narrow sense.There are two genotypes of transposable elements:Transposa and Flipper types.The Transposa type is predominant.The differentiation of pathogenicity is obvious between representative isolates.