栗疫病致病性研究和栗疫菌混合体系探讨

经过１０余年栗疫病发生特点和致病性的大量调查研究，结果表明栗疫菌在板栗分布区普遍存在，是弱寄生菌，在本地只寄生那些生长十分衰弱，即将死亡的板栗树干，苗木等，但栗疫菌的寄生不是这些部位死亡的主要和直接原因，而且它所寄生的部位局部死亡后对整株栗树的影响不大，因此，栗疫病在一般情况下非毁灭性病害，本文首次提出了栗疫菌混合体系概念，并用此说明中国栗疫病普遍分布，危害很轻的原因。     

栗疫病菌培养性状及毒力的研究

将采自湖南、重庆两省的56个板栗疫病菌(Cryphonectria parasitica (Murr．)Barr．)的培养性状划分为两类：一种为正常培养性状即：菌落桔黄色或黄色、生长快、形成分生孢子器的菌株，其占菌株总数的58．93％；其它为非正常培养性状，菌落颜色表现出黄褐色、深褐色或白色，其生长速率与正常菌株均不同。致病性测定结果表明，各个菌株可分为强毒力菌株、中等毒力菌株、中低毒力菌株、低毒力菌株。     

栗疫菌弱毒力特征的传递及其影响因素

栗疫菌 (Cryphonectriaparasitica)弱毒力 (hypovirulence)的本质是菌体中含有一种弱毒性病毒 (cryphonectriahypovirulenceviruses ,CHV)dsRNA(Choietal,1992 ;Morrisetal,1979)。栗疫菌的弱毒力菌株 (H菌株 )一般都具有将强毒力 (virulent)菌株 (V菌株 )转化为弱毒力菌株的能     

栗疫病菌分生孢子后代培养性状分化的观察

通过对6个含dsRNA及4个不含dsRNA的菌株进行培养性状的观察，结果表明，含dsRNA的菌株其分生孢子后代发生明显的分化，表现为白色和桔黄色两种类型菌落；而不含dsRNA的菌株后代均不发生分化；并对各菌株的后代进行了dsRNA检测。     

香味全缘孔菌培养条件的研究

香味全缘孔菌是一种可以提取天然香料的高等真菌。本项研究对分离于野外的香味全缘孔菌从不同的温度、pH值以及不同的C源和N源的营养成分进行室内培养。其结果显示香味全缘孔菌营养菌丝生长的相对最适温度为33℃;最佳N源为酵母汁,其次为牛肉膏;相对在培养基pH值为6.0时菌丝生长较快,较好的C源为葡萄糖。这些结果为大规模人工培养香味全缘孔菌提供了很好的指导作用。     

湘渝地区栗疫菌低毒力菌株dsRNA的转化研究

通过对采自湘渝两地区板栗上的溃疡斑进行分离，共得到56个菌株，各菌株经检测，有11个中低、低毒力菌株和1个中等毒力菌株含dsRNA，所有供试强毒菌株均不形成电泳带。通过弱毒菌株对毒菌株的转化，结果表明，弱毒力菌株对毒力菌株的转化率表现出较大差异，其中，弱毒力菌株185、618、293对所试的10个毒力菌株转化情况较好，转化率均在90％以上。弱毒力菌株729、518、392对毒菌株的转化差异较大，转化率在20—80％之间不等。     

栗疫菌营养体亲和型多样性比较及野生栗栗疫菌交配型的分子检测

在北京周边县区、湖北大老岭山区以及陕西安康地区分别调查栗疫病发生情况,采集病原栗疫菌(Cryphonectria parasitica)菌株并进行营养体亲和性试验.结果表明:北京板栗栽培区和陕西安康地区野生栗栗疫病发病率较高,局部地区危害严重;湖北大老岭山区栗疫病发病率低,危害轻微.北京地区菌株群体营养体亲和型多样性指数(Shannon-Wiener's diversity index)极显著低于湖北菌株群体和陕西菌株群体,而陕西与湖北菌株群体营养体亲和型多样性指数差异不显著.随机选取湖北、陕西野生栗栗疫菌部分菌株,利用特异性引物,通过普通PCR和巢氏PCR,对其交配型进行测定,发现陕西与湖北野生栗菌株群体中均存在MAT-1与MAT-2两种交配型的菌株.通过PCR扩增,在两地的野生菌株群体里均发现同时具有两种交配型基因的菌株,其中,陕西群体此类菌株比例较大,湖北菌株比例较小     

三种桉树菌根菌培养条件的研究

多根硬皮马勃9215、彩色豆马勃9216及西澳粘滑菇E4070菌株是广东地区应用较多、效果也较好的菌根真菌,为摸清其培养条件,分别进行了适温范围、最适pH以及C源、N源需求的测定。其结果表明:菌株9216适温范围最广,为10～38C,菌株E4070适温范围最窄,为15～30C;最适pH范围以9215菌株最广,为3～7,菌株E4070最窄,仅为5～7;除菌株9216不能利用有机N源中的牛肉膏及酵母浸膏外,其它8种N源均有不同程度的利用,9215菌株对有机N利用较好,而9216菌株对氨态N利用较好;除9215菌株对果糖的利用较差外,其它C源差异不大,菌株9216对麦芽糖及葡萄糖利用较好,而E4070对6种C源利用效果差异不大。     

培养条件对紫丁香花粉离体萌发的影响

以紫丁香盛花期的新鲜花粉为材料,采用单因素试验,研究了培养基、pH值、培养温度、培养时间对紫丁香花粉离体萌发和花粉管生长的影响。结果表明:花粉萌发和花粉管生长的最适培养基pH值为6.0,其花粉萌发率为32.45%,花粉管长度为0.350mm,最佳培养温度为25℃,最佳培养时间为1.5～2h。     

Variations in Disease Resistance of Chestnut Cultivars against Chestnut Blight Fungus,Cryphonectria parasitica

INTRODUCTION Chestnut trees cultivated worldwide are planted as fruit tree in Korea and the area is increasing annually. In the 1970s, chestnut trees were enormously planted in many provinces in Korea. However,chestnut blight occurred and increased due to lack of disease management and the aging of the trees. Blight caused by Cryphonectria parasitica is a kind of important disease and occurred in oak and maple. American chestnut is more susceptible to this pathogen than Korean or Japane…     

细菌嗜铁素、DAPG和PCA对板栗疫病菌的抑制作用

为了探讨生防细菌防治板栗疫病的潜力,测定了恶臭假单胞杆菌(Pseudomonas putida)WCS358r菌株及其嗜铁素缺失突变体JM218,转基因菌株WCS358::phl和WCS358::phz对板栗疫病菌生长和分生孢子萌发的抑制作用。在KB和板栗枝条韧皮部抽提液培养基(BA)中,WCS358r对病菌生长和分生孢子萌发具有较强的抑制作用,而JM218几乎没有抑制效果;在培养基中加入200μmol/L FeCl3后,WCS358r对病菌的抑制能力显著降低,证明WCS358r产生的嗜铁素是抑制菌丝生长和分生孢子萌发的重要因子。在PDA培养基中,WCS358r对病菌无抑制效果。在上述三种培养基中,产生抗生素2,4-二乙酰间苯三酚(DAPG)的转基因菌株WCS358::phl与原始菌株WCS358r相比,抑菌效果明显增强,而产生酚嗪-1-羧酸(PCA)的WCS358::phz的抑菌能力无明显提高。培养基中添加铁对WCS358::phl的抑菌活性无影响,但能降低WCS358::phz对板栗疫病菌的抑制效果。由此说明,嗜铁素和DAPG对抑制板栗疫病菌生长和孢子萌发具有重要作用,而PCA的抑菌效果不明显。     

Rindenkrebs der Eßkastanie (Cryphonectria parasitica) in Südwestdeutschland

Chestnut blight (Cryphonectria parasitica) in South-west Germany . In 1992 and 1993, chestnut blight (Cryphonectria parasitica) was found in three stands in the south-west of Germany. Each of the three stands has only one compatibility group (vc-group). Two of them (97 and 75 Isolates) belong to the vc-group I. They are compatible with the Swiss vc-group I. The 43 Isolates from the third stand were not compatible with the most common Swiss vc-groups (vc-group I-V). In the German vc-group I, one hypovirulent strain was found.     

板栗疫病发生原因调查及对策

通过对板栗疫病发生现状调查，认为板栗疫病发生与管理粗放、栽植不合理、蛀干害虫危害、人为造成大量伤口等有一定关系，提出今后应加强检疫、抚育、伤口保护、重病区改造、药剂防治等工作 。     

Spread of Cryphonectria hypovirus1 into 45 vegetative compatibility types of Cryphonectria parasitica on grafted American chestnut trees

A mixture of hypovirulent strains of Cryphonectria parasitica, including four white (European) strains infected with Cryphonectria hypovirus1 (CHV1), was used in 1982 and 1983 to inoculate natural blight cankers located within a zone ranging from the ground to 183 cm on grafted American chestnut trees. These four white strains belonged to three vegetative compatibility (vc) types. Using pigmented, single‐spore colonies from white isolates, 48 vc types were identified among 110 white isolates recovered in 1996, 1998, and 1999 from cankers located outside the inoculated zone. Twenty‐five of the 48 white vc types consisted of two or more isolates. The 25 major white vc types were vegetatively incompatible with all four of the original white hypovirulent strains, providing evidence for spread of CHV1 but not for spread of the original inoculated strains. Forty‐five vc types represent the minimum number of `new' vc types into which CHV1 had spread. The ratio of white vc types to white isolates tested (S/N) and Shannon diversity index were 0.436 and 3.64, respectively. The spatial pattern of white vc types on the grafts was found to be non‐random (p=0.019). White single‐spore colonies of white isolates were placed into four cultural morphology (CM) groups. The two largest groups contained 37 (CM group 3) and 33 (CM group 1) isolates. Single‐spore colonies from the original, white inoculated strain, Ep 49, were classified into CM groups 3 and 1, and colonies of Ep 51 W were classified into CM group 1.     

韩国、美国和中国栗疫病菌的遗传变异分析

用RAPD方法,对来自韩国、美国和中国的26个栗疫病菌菌株进行了遗传变异分析.使用筛选的12个随机引物,共扩增了115个0.19～3.1 kb大小的扩增片段,其中多态性片段占61.7%.聚类分析结果,相似系数为0.92时,26个菌株分为两大组,一组由21个菌株组成,包括大部分韩国菌株和美国菌株;另一组包括部分韩国菌株和中国菌株.表明美国菌株和大部分韩国菌株的遗传相似性很高,部分韩国菌株有较大的变异,而中国菌株则表现出了遗传上的远缘关系.     

Incomplete movement of Cryphonectria hypovirus 1 within a vegetative compatibility type of Cryphonectria parasitica in natural cankers on grafted American chestnut trees

American chestnut trees, grafted in 1980 from large survivors, were inoculated in 1982 and 1983 with four white (European) hypovirulent strains of Cryphonectria parasitica, infected with C. hypovirus 1 (CHV1); this hypovirus has been shown to be capable of moving rapidly within the mycelium of a vegetative compatibility (vc) type of C. parasitica in blight cankers. Using a 49‐cell lattice plot, 17.8×17.8 cm, the spatial patterns and frequencies of white and pigmented isolates and white and pigmented vc types were investigated within superficial cankers on the grafts located outside the hypovirulent‐strain‐inoculated zone. Four of six cankers assayed contained white isolates, and three of the four had random spatial patterns of white isolates, based on join‐count statistics. Vc tests, using pigmented isolates and pigmented single‐spore colonies of white isolates, indicated that the majority of white and pigmented isolates recovered from each of two cankers assayed were in one vc type. White and pigmented lattice‐plot cells of the same vc type were frequently in contact with each other, indicating incomplete movement of CHV1 within a vc type. Nine and 10 vc types were found in the two cankers; it is hypothesized that small, white vc type areas in each canker may be a source of CHV1 transmission to the major vc types. Based on join‐count statistics, the spatial pattern of the single, major vc type in one canker was non‐random (aggregated), whereas the other canker had a random major vc type pattern. White and pigmented in vitro variants (sectors) of C. parasitica, that resemble white and pigmented in vivo variants in spatial contact and vc compatibility, were intermediate hypovirulent and virulent on forest American chestnuts, and dsRNA positive and negative, respectively. Incomplete movement of CHV1 within a vc type could be a major cause of the prevalence of pigmented isolates in superficial cankers on chestnut trees.     

木霉菌和毛壳菌对板栗疫病的抗生作用

哈茨木霉Trichoderma harzianumT88菌株、深绿木霉T.atrovorideT95菌株和螺旋毛壳Chaetomium spiraleND35菌株与板栗疫病菌Cryphonectria parasiticaLY菌株的平板对峙培养结果表明,木霉菌对营养和空间有强烈的竞争作用,表现在可使病原菌菌丝消解,对病菌菌丝的生长有明显的抑制作用;毛壳菌生长速度较慢,但可形成抑菌带,使病菌不扩展。室内水培枝条接种试验结果表明,接种拮抗菌的拮抗效果好,而且先接种拮抗菌后接种病原菌的处理比拮抗菌和病原菌同时接种的处理效果还好,毛壳菌没有木霉菌的作用明显。接种枝条的木霉菌、毛壳菌和病原菌的再分离结果表明,T95和ND35的定殖能力强。