大豆枯萎病菌尖孢镰孢遗传多样性及大豆品种抗性

 了解大豆枯萎病菌的群体遗传特征及明确大豆种质对大豆枯萎病的抗性,对抗病育种、抗性品种的合理布局以及制定更有效的病害防治策略具有重要的参考价值。本研究利用随机扩增多态性DNA(random amplified polymorphic DNA,RAPD),对采自我国不同地区的大豆枯萎病菌—尖孢镰孢菌(Fusarium oxysporum)进行遗传多样性分析,筛选到10个多态性随机引物,共扩增出75条RAPD条带,其中55条为多态性条带,占73.3%。利用UPGMA法对DNA扩增图谱进行聚类分析,以相似系数0.68为阈值,55个分离物可分为9个遗传聚类组,表明我国大豆枯萎病菌具有丰富的种内遗传多样性,所划分的群体与分离物来源地不相关。同时,对上述分离物进行致病性分析,发现我国的大豆枯萎病菌具有明显的致病力分化现象。进一步利用3个代表性分离物对来自我国不同大豆产区的180个大豆品种(资源)进行抗大豆枯萎病鉴定,发现皖豆28、中黄13、中黄51、中作X08076和5D034等5个品种对大豆枯萎病具有良好抗性,占供试材料的2.8%,表明不同大豆品种对枯萎病的抗性存在一定的差异。     

尖孢镰刀菌磷脂脂肪酸的遗传多样性分析

为明确尖孢镰刀菌Fusarium oxysporum不同专化型菌株的遗传多样性,利用MIDI微生物脂肪酸鉴定系统分析了来源于5个专化型的27株菌株的磷脂脂肪酸种类和含量,并进行了磷脂脂肪酸聚类分析。结果表明,供试尖孢镰刀菌中共检测到9种脂肪酸,在兰氏距离为25.98时分为3个类群,即主要脂肪酸、微量脂肪酸和偶现脂肪酸,其中主要脂肪酸有16:0、18:0、18:1cis9(ω9)和18:2cis9,12/18:0a,含量共占总磷脂脂肪酸的96.41%;微量脂肪酸有14:0、15:0、16:1cis9(ω7)和20:0,偶现脂肪酸仅有17:1(ω11)。不同专化型菌株的主要脂肪酸仅18:2cis9,12/18:0a存在显著差异,微量脂肪酸除15:0外其余3种均有显著差异。聚类分析结果表明,以供试的全部27株菌株聚类时,其磷脂脂肪酸与寄主专化型间无明显相关性,以寄主来源为同科(葫芦科或茄科)的菌株聚类时,其磷脂脂肪酸与寄主专化型间存在着一定的对应关系。     

拟轮枝镰孢菌EST-SSR信息分析与标记开发

为开发可用于拟轮枝镰孢菌Fusarium verticillioides及其近缘种遗传多样性分析的SSR引物,利用生物信息学方法和PCR技术,通过对从NCBI下载的87 086条拟轮枝镰孢菌的EST序列信息进行分析,设计EST-SSR引物,检测其在拟轮枝镰孢菌及其近缘种中的扩增情况,并用筛选出的多态性引物对15株拟轮枝镰孢菌进行SSR遗传多样性分析。结果表明,在EST序列中,共查找到11 952个SSR位点,592种重复基元,SSR出现频率为1.09%,重复基元出现数量最多的为三核苷酸(54.00%),其中(CAA/TTG)n基元出现频率最高。设计的25对EST-SSR引物在拟轮枝镰孢菌种内的有效扩增率和多态率分别为80.00%与32.00%,对5种近缘镰孢菌种的通用率和多态率分别为40.00%和8.00%。遗传多样性分析结果表明,在相似系数为0.664水平下,供试菌株可划分为4个SSR类群,但类群的划分与菌株的地理来源无关;不同菌株间存在明显的遗传分化。表明基于拟轮枝镰孢菌EST序列开发的SSR引物可用于拟轮枝镰孢菌及其近缘种的遗传多样性分析。     

芝麻菜品种对尖镰孢菌枯萎病的抗性评价

尖镰孢菌枯萎病是意大利西北部芝麻菜栽培品种(Eruca vesicaria)和野生品种(Diplotaxis spp．)上的重要病害之一,常引起严重的损失。为评价芝麻菜品种对尖镰孢菌枯萎病的抗性,2004年6～11月,选用了44个栽培及野生品种资源在玻璃温室进行抗性试验,使用浓度为1×10~6cfu/ mL的孢子悬浮液蘸根法人工接种30天苗龄的芝麻菜幼苗,以分离自芝麻菜两个不同野生品种的菌株Fus．Ruc．9A/02和Fus．Ruc．13/03,以及分离自栽培和野生品种的32个菌株混合(MIX),分别作为抗病试验接种体,每次接种试验设A、B、C三个重复。整个试验重复进行2次。调查病害引起的死苗率(%)和病情指数(DI),作为评价芝麻菜品种抗性差异的两项指标。研究结果显示,这两项指标间的相关性极显著(P＜0．01),即表明可采用其中一项指标评价芝麻菜品种对尖镰孢菌枯萎病的抗性。供试的44个栽培及野生芝麻菜品种中,多数品种抗尖镰孢茵茵株Fus．Ruc．13/ 03,而对菌株Fus．Ruc．9A/02及菌株混合MIX表现为易感性,而且这些易感品种在菌株Fus．Ruc．9A/02及菌株混合MIX两试验中的抗性表现较为一致,Fus．Ruc．9A/02和MIX的毒性也更强于Fus．Ruc．13/03菌株。芝麻菜品种12/03和品种20/03高抗菌株混合MIX,品种6/03和品种7/04高抗菌株Fus．Ruc．9A/02,而品种9/02、2/03、5/03、6/03、7/03、9/03、11/03、12/03、20/03、21/03、24/03、3/04、6/04、7/04和11/04高抗菌株Fus．Ruc．13/03。     

西安地区番茄枯萎病尖镰孢菌的生理型鉴定

 西安地区番茄枯萎病早在60年代初就发现在东郊的分布和危害。70年代中后期推广满丝、特罗皮克等抗枯萎品种后,病情得到控制。     

木霉对尖镰孢菌的拮抗机制及生防效果研究

将土壤中分离出的木霉菌株(T₄₁) ,通过对峙法对尖镰孢菌进行了体外拮抗作用测定和评价 ,并利用制备的木霉生防菌剂对番茄枯萎病进行了室内盆栽防治试验。结果表明 :木霉T₄₁对番茄枯萎病的防效达86% ,并具有防效长久和刺激作物生长的作用     

尖孢镰刀菌的遗传多态性

 尖孢镰刀菌是重要的植物维管束病原真菌。近年来,有关该菌的遗传多态性研究报道很多,本文着重综述了利用营养体亲和群、DNA多态性技术研究尖孢镰刀菌专化型、小种及其相互关系等方面的进展,同时介绍了对棉花枯萎病菌、瓜类枯萎病菌及尖孢镰刀菌小种起源等的研究概况。     

京津冀设施大棚根际土壤中筛选出的防治根腐病的生防镰孢菌

 由致病性尖孢镰孢菌(Fusarium oxysporum)引起的根腐病严重危害果蔬生产,但非致病性镰孢菌可作为潜在的生防菌。为筛选防治根腐病的非致病生防镰孢菌,从京津冀设施大棚采集茄科、葫芦科果蔬78份根际土样中分离2 402株真菌,筛选出对致病性尖孢镰孢菌(F. oxysporum)具有拮抗效果的真菌173株。利用镰孢菌通用引物进行PCR扩增,从中筛选出28株候选镰孢菌;通过镰孢菌发酵液泡根进行安全性测试,筛选出对寄主黄瓜幼苗安全无害的镰孢菌菌株4株(1418、1441、1436和1473)。进一步通过镰孢菌测序通用引物TEF1αF/TEF1αR结合菌落和分生孢子的形态学特征,1418菌株和1441菌株被鉴定为尖孢镰孢菌(F. oxysporum)、1436菌株被鉴定为茄病镰孢菌(F. solani)。盆栽测试发现,除1441菌株外,1418菌株、1436菌株和1473菌株对黄瓜根腐病的防效均在50%以上,其中1418菌株的防效为70%,与杀菌剂咪酰胺的防效相当,具有很好的应用潜力。本研究筛选获得的具有生防潜力的镰孢菌不仅为镰孢菌致病力分化的研究提供了实验材料,也为新型生防产品的研制奠定了基础。     

尖镰孢胁迫下黄瓜茎部蛋白质差异表达分析

为揭示尖镰孢胁迫下黄瓜茎部蛋白质差异,以黄瓜枯萎病高抗品系‘D0327’和感病品系‘649’为试验材料,分别接种2个毒力不同的尖镰孢C9和S1菌株后提取黄瓜茎部蛋白,运用双向电泳技术研究了尖镰孢侵染后茎部蛋白质组的变化。结果表明,2个尖镰孢菌胁迫下,抗感材料茎部蛋白表达发生了变化。茎部蛋白质双向电泳图谱与对照图谱进行比较分析,成功鉴定出8个差异蛋白点。强毒力菌株C9胁迫下,感病品系中鉴定出3个差异蛋白,分别是烯醇化酶及亚基、肌动蛋白和放氧增强蛋白,抗病品系中差异蛋白为烯醇化酶及亚基、磷酸丙糖异构酶和核苷酸二磷酸激酶。弱毒力菌株S1胁迫下,感病品系中差异蛋白为Csf-2蛋白,抗病品系中差异蛋白为放氧增强蛋白。这些蛋白分别参与能量代谢、光合作用和胁迫反应。     

尖孢镰刀菌及芬芳镰刀菌遗传多样性的ISSR分析

 为了明确镰刀菌属(Fusarium)美丽组（Section Elegans）中尖孢镰刀菌(F. oxysporum)和芬芳镰刀菌（F. redolens）2种菌的遗传差异性和亲缘关系,利用ISSR分子标记技术对这2种菌的35个菌株进行了遗传多样性分析。结果表明,用筛选的15条引物对35个供试菌株共扩增出231条条带,其中多态性条带220条,平均多态性比率为95.2%,平均每条引物产生条带为14.7条。聚类结果和遗传相似系数分析显示,35个菌株间的遗传相似系数范围为0.506~0.935,平均为0.661。在遗传相似系数为0.593时,供试的35株镰刀菌可明显的分成2个ISSR类群（IG）,其中IGⅠ包括1～23号菌株,全部为F. oxysporum;IGⅡ包括24～35号菌株,全部为F. redolens。ISSR类群划分与菌种分类之间存在一定相关性 (IGⅠ中23株F. oxysporum间的平均相似系数为0.720,IGⅡ中12株F. redolens间的平均相似系数为0.717),但与菌株的地理来源不存在相关性。而同一类群中,菌株之间的遗传相似性与菌株的地理来源存在一定的相关性。     

检疫性杂草毒莴苣EST-SSR标记的开发及应用

为明确毒莴苣EST序列中SSR的特点,并开发EST-SSR分子标记用于毒莴苣及其近似种的分类鉴定及遗传多样性分析,以筛选的7 438条毒莴苣EST序列为基础,利用SSRIT软件结合人工搜索查找SSR位点,并采用Primer Premier 5.0软件设计EST-SSR引物,以毒莴苣及其近似种为材料对所合成引物进行多态性检测,应用UPGMA法构建11个毒莴苣及其近似种的系统发育树。结果表明,共搜索到SSR位点1 465个,发现重复基元598种,其中二、三、六核苷酸重复是主要的重复类型。合成的24对引物中有9对表现出多态性,利用这9对EST-SSR引物分析毒莴苣等11份材料的亲缘关系,其相似性系数在0.432~1.000之间;于0.513处可被划分为3大类:Ⅰ类包括翅果菊和北山莴苣,Ⅱ类只有宿根莴苣1种,Ⅲ类包括毒莴苣和莴苣。     

Cultural characteristics, pathogenicity and genetic diversity of Fusarium oxysporum isolates from tobacco fields in Spain

Several formae speciales of Fusarium oxysporum are capable to produce disease in tobacco plants. Different authors have classified those isolates as a forma specialis or a race within on the basis of the severity of disease and host specificity. Fusarium wilt of tobacco plant in Extremadura (central Spain) tobacco fields have been recorded in the last years and F. oxysporum was isolated from symptomatic plants. The aim of our study was to characterize these F. oxysporum populations. For this purpose, the in vitro spore production and growth and the virulence (severity of disease) have been tested. Although all isolates behaved as pathogen, the virulence of isolates was different. The differences in growth could not be correlated with other characteristics but the two isolates with scarce spore production have also behaved as the weakest pathogen. We have analyzed intergenic spacer (IGS) region polymorphism of ribosomal DNA and random amplified polymorphic DNA (RAPD) markers to assess the genetic diversity within F. oxysporum isolates. These molecular analyses showed two major groups with different physiological capabilities that could reflect two different lineages. One group was characterized by medium–high sporulation, high virulence and the same IGS-RFLP pattern. The other group was more heterogeneous featuring low–medium sporulation and variable virulence and growth. This first experimental approach to pathogen population could be a good starting point for further studies including non-pathogenic isolates and a larger number of pathogen that could clarify if there are two or more genetic lineages.     

土壤强还原消毒过程中产生气体对土传病原菌的抑制作用

土壤强还原(reductive soil disinfestation,RSD)是一种高效、环保的土壤消毒方法,为明确RSD杀菌机理,采用气相色谱和荧光定量PCR等技术研究RSD过程中产生的气体对土传病原菌的杀菌作用。结果表明:RSD过程中土壤能够产生H_2S和NH_3;当在装有25 g病土的三角瓶中分别加入25 m L的H_2S和NH_3时,土壤中尖孢镰刀菌的数量均显著下降,其杀菌效果分别为90.3%和100.0%;当在病土中加入0.125%(w/w)氨水时,尖孢镰刀菌、茄劳尔氏菌、辣椒疫霉和立枯丝核菌的数量分别下降至对照土壤的1.0%、0.3%、0.1%和0.9%;此外,加入10μL氨水即可显著抑制尖孢镰刀菌菌丝生长和孢子萌发,抑制率分别为52.8%和100.0%。表明RSD过程中产生的H_2S和NH_3对RSD过程的杀菌效果起着一定的作用。     

Ultrastructural and cell wall modifications during infection of <Emphasis Type="Italic">Eucalyptus viminalis</Emphasis> roots by a pathogenic <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> strain

Fusarium species are soil-borne fungal pathogens that produce a variety of disease symptoms when attacking crop plants. The mode of root colonization of Eucalyptus viminalis seedlings by a pathogenic F. oxyporum strain (Foeu1) at the ultrastructural level and changes in cell wall pectin during host pathogen interactions are described. Root systems of E. viminalis plants were inoculated with F. oxysporum in an in vitro model system. Hyphae of F. oxysporum adhered to the outer epidermal cell walls through fibrillar material, and after penetration they spread into the internal tissues. They developed intercellularly and intracellularly in the root cortex and invaded vascular tissues. Papillae were induced, and the host plasma membrane ruptured in colonized cells, causing rapid host tissue and cell damage. Changes in distribution and occurrence of nonesterified and methyl-esterified pectins were evaluated after root colonization by F. oxysporum using two monoclonal antibodies, JIM 5 and JIM 7, respectively. Nonesterified pectin in control roots was mainly localized in the epidermal cell walls and middle lamellae in parenchymal cortex, whereas methyl-esterified pectin accumulated more in primary cell walls of the cortex and phloem. Decreases in immunodetected nonesterified and methyl-esterified pectins were associated with extensive plant tissue degradation after root colonization by the pathogenic fungus.     

太子参根腐病病原菌的鉴定及防治药剂筛选

为有效防控太子参根腐病,本研究于2017年3月-2019年6月自福建省宁德市柘荣县太子参主产区采集具有典型根腐病症状的植株和块根病样,分离纯化获得病原菌,利用形态学特征、分子生物学特性及致病性测定对其进行鉴定;同时通过测定平板抑制率和扫描电镜观察分析6种杀菌剂对太子参根腐病病原菌菌丝生长的抑制效果.结果表明:太子参根腐...     

Cell interactions between a nonpathogenic <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> strain and root tissues of <Emphasis Type="Italic">Eucalyptus viminalis</Emphasis>

Nonpathogenic isolates of Fusarium oxysporum can be successful antagonists of pathogenic forms of the same fungal species that commonly attacks crop plants. The characteristics that distinguish nonpathogenic from pathogenic forms are not well understood. In this study, the mode of root colonization of Eucalyptus viminalis seedlings by a nonpathogenic F. oxysporum strain is described at the ultrastructural level. Root systems of E. viminalis plants were inoculated with nonpathogenic F. oxysporum strain Fo47 in an in vitro model system. Changes in the occurrence of nonesterified and methyl-esterified pectins in colonized E. viminalis roots were evaluated by in situ immunolabeling using two monoclonal antibodies, JIM 5 and JIM 7. Modes of penetration and root colonization patterns in E. viminalis seedlings by the nonpathogenic fungus were similar to those described for pathogenic forms of F. oxysporum. However, root interactions differed in that the nonpathogenic fungus did not induce host tissue damage. No papilla-like appositions were observed in host cells in response to invading hyphae, which did not disrupt the host plasma membrane in many cases, suggesting that a biotrophic relationship was established. Root colonization by the nonpathogenic strain did not induce alteration in JIM 7 labeling of methyl-esterified pectin in E. viminalis cell walls, whereas nonesterified pectin was detected to a significantly greater extent in cell walls of roots colonized by the fungus. Pectin components decreased slightly only at points of hyphal contact with host cells. Because nonpathogenic strains utilize pectin in pure culture, host control over enzyme activity or production by the fungi may at least partly explain their compatible interactions with host tissues.