国内一种果树新病害--石榴枯萎病

2001年在云南省蒙自县相继发生石榴枯萎病，经作者分离鉴定以及致病性测定，证明病害是由甘薯长喙壳Ceratocystis fimbriata Ellis and Halsted引起的，为国内果树新病害。对病害的症状、鉴定方法、病原菌形态及发生危害情况进行了描述。     

紫茎泽兰对石榴枯萎病菌和枯草芽孢杆菌生长的影响

在室内条件下,利用20%的MS营养液（MS0.2）和灭菌蒸馏水培养的紫茎泽兰根系分泌物对石榴枯萎病和生防细菌枯草芽孢杆菌生长进行测定。结果表明,2种方式培养的紫茎泽兰根系分泌物均不能有效抑制枯萎病菌的生长,但对枯草芽孢杆菌的生长具有极强的促进作用。其中30%MS0.2对枯草芽孢杆菌的促进率达29.85%,而40%蒸馏水的促进率可达87.46%。恢复石榴园紫茎泽兰可能提高利用生防细菌枯草芽孢杆菌防治石榴枯萎病的效果。     

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。     

香蕉枯萎病菌生理小种鉴定及其SCAR标记

 通过室内人工接种蕉类鉴别寄主,对采集于广东蕉区的18个蕉类枯萎病菌菌株进行鉴定,KP021、KP022、GZ981和JL021 4个菌株属Racel,其余14个菌株属Race4,说明广东蕉区同时存在尖孢镰刀菌古巴专化型Race1和Race4。用RAPD技术对上述18个菌株进行分析,从200条随机引物中筛选出8条引物可产生生理小种RAPD标记12个,其中标记Racel的8个,标记Race4的4个。对这些RAPD标记带分别进行回收、克隆、测序,根据这些特异片段序列分别设计相应的SCAR引物,通过对18个菌株的PCR扩增检验,有4个RAPD标记成功地转化为SCAR标记,其中Race1-SCAR标记1个、Race4-SCAR标记2个、同时能鉴定出2个小种的SCAR标记1个。应用这4个SCAR标记同时对采自田间的9个病菌分离物进行检测,能够准确地鉴定出广东蕉区的尖孢镰刀菌古巴专化型Racel和Race4,这为下一步开展香蕉枯萎病菌生理小种的分子鉴定及各生理小种田间流行动态监测奠定了基础。     

香蕉枯萎病菌致病力分化与ISSR遗传多样性分析

香蕉枯萎病是一种破坏香蕉维管束的全株性土传病害。本研究旨在探讨香蕉枯萎病菌致病力分化和遗传多样性。以30株采自我国广西的香蕉枯萎病菌,16株分别来自澳大利亚和我国广东、海南、福建、云南等地的香蕉枯萎病菌为对象,采用伤根灌淋法测定香蕉枯萎病菌的致病力,然后用筛选到的ISSR引物对46个香蕉枯萎病菌菌株和4个对照菌株(3个非致病性尖孢镰刀菌和1个茄腐镰刀菌)进行ISSR遗传多样性分析。结果显示,分离到广西香蕉枯萎病菌1号生理小种(FOC1)8株,致病力强、中、弱类型比例分别为62.5%、12.5%和25%;广西香蕉枯萎病菌4号生理小种(FOC4)22株,致病力强、中、弱类型比例分别为18.18%、63.64%和18.18%。14条ISSR引物扩增出237个条带,多态性条带161个,多态性比例为67.93%,遗传相似系数0.76～0.96。聚类分析显示,以遗传距离0.80为阈值时菌株被分为8个类群,所占比例分别为4%、10%、60%、16%、4%、2%、2%、2%。第三类群全部为香蕉枯萎病菌4号生理小种。第一、二、四和五类群总量的70.59%为香蕉枯萎病菌1号生理小种。第八类群为香蕉枯萎病菌3号生理小种。结果表明,在香蕉枯萎病菌与寄主协同进化中,广西的FOC1和FOC4出现明显致病力分化。1号生理小种的遗传多样性比4号生理小种丰富。广西香蕉枯萎病菌4号生理小种与海南、广东的FOC4遗传相似性较高。香蕉枯萎病菌生理小种类型与遗传多样性相关。致病力变异与遗传多样性无相关性。研究结果对香蕉枯萎病菌种群扩张机制探讨、遗传动态分析以及有效防控措施的制定具有一定的指导意义。     

香蕉枯萎病菌RAPD分析及4号生理小种的快速检测

 用随机扩增多态性DNA(RAPD)技术,对采自广东、广西的香蕉和粉蕉上的30个香蕉枯萎病菌(Fusarium oxysporum f.sp.cubense)菌株和3个其它尖孢镰刀菌专化型的菌株进行比较及聚类分析。在遗传相似系数0.67时,可将供试菌株划分为3个RAPD群(RGs),其中香蕉枯萎病菌4号生理小种(FOC4)共15个菌株属于RGⅠ,1号生理小种(FOC1)共15个菌株属于RGⅡ,供试的其它尖孢镰刀菌专化型的3个菌株则属于RGⅢ。这说明香蕉枯萎病菌和供试3个其它专化型菌株与致病性间存在明显的相关性。1号生理小种内菌株间的遗传分化大于4号生理小种内菌株间的遗传分化。从90条RAPD随机引物中筛选出2条引物可产生4号生理小种的RAPD标记2个。将这2个RAPD标记电泳切胶回收、克隆及测序,并根据这2个特异片段序列设计SCAR上下游特异引物,通过对30个菌株的PCR扩增检验,其中一个RAPD标记成功地转化为SCAR标记,初步建立了以此为基础的4号生理小种快速检测技术,其检测灵敏度为2 ng新鲜菌丝。对采自不同地区的显症样品、吸芽、室内接种未显症的香蕉苗以及发病的香蕉植株不同部位进行检测,能够准确灵敏地鉴定出4号生理小种,从而为香蕉枯萎病菌的快速检测及防治奠定了基础。同时,快速检测结果发现,田间发病植株果柄的各部位及果实内并没有枯萎病菌的存在。     

石榴枯萎病菌拮抗放线菌对南方根结线虫的毒力

从云南蒙自万亩石榴园、建水石榴园采集的土样中分离筛选得到对石榴枯萎病菌有较强拮抗活性的放线菌,其中菌株JS1对石榴枯萎病菌的室内抑菌半径为11mm。测定了菌株JS1发酵液对南方根结线虫的抑杀活性,结果表明,50%JS1菌株发酵液处理南方根结线虫卵6d,卵孵化率与对照相比降低了51.67%;50%JS1菌株发酵液处理南方根结线虫2龄幼虫72h,2龄幼虫的校正死亡率增加了88.10%;菌株JS1兼有拮抗石榴枯萎病菌及抑杀南方根结线虫的活性。结合菌株JS1在鉴定培养基上的培养特征、形态学观察及16SrDNA序列分析,将JS1初步鉴定为壮观链霉菌(Streptomyces spectabilis)。     

黄瓜枯萎病菌遗传多样性的AFLP分析

黄瓜枯萎病是由半知菌亚门尖孢镰刀菌黄瓜专化型(Fusarium oxysporumf.sp.cucumainum Owen)侵染引起的一种土传病害,是影响黄瓜生产的最主要病害之一[1].近年来随着分子生物学技术的迅速发展,国内外学者对于病原真菌的遗传多样性做了大量的研究,Wang等[2]对影响黄瓜枯萎病菌AFLP技术体系的多种因素作了探讨,得到了1种适合于黄瓜枯萎病菌AFLP分析的优化体系;Duan等[3]应用RAPD、ISSR和AFLP标记揭示出了西瓜枯萎病菌株在分子水平上的遗传多样性.     

甘薯苗期黑斑病对薯苗生长的影响及药剂浸种对黑斑病的防治效果

为明确甘薯苗期黑斑病对薯苗生长的影响及25%肟菌·异菌脲悬浮剂浸种的防治效果, 通过人工接菌和药液浸种试验, 对各处理的种薯发芽、生长、产苗量及薯苗带菌率进行了调查。结果表明, 种薯或基质带菌育苗, 种薯发芽数与未接菌处理相比没有显著差异;不同浓度25%肟菌·异菌脲悬浮剂浸种对种薯发芽数也未见显著影响。与未接菌处理相比, 种薯带菌和育苗基质带菌处理未药剂浸种时产苗量分别减少43.97%和59.82%。使用25%肟菌·异菌脲悬浮剂500 mg/L浸种时, 种薯和基质带菌试验中千克种薯产苗分别为74.12株和69.56株, 苗床接菌试验中产苗分别为126.92株和85.31株, 均显著高于未药剂浸种的处理。调查发现, 种薯或苗床带菌会使萌发的嫩芽受黑斑病菌感染, 引起死苗、生长停滞和薯苗带菌, 是产苗量减少的主要原因。25%肟菌·异菌脲悬浮剂浸种可减少薯苗发病, 增加产苗量。     

嘧菌酯对石榴干腐病菌的生物学活性

为探究石榴干腐病的病原菌种类及嘧菌酯对石榴干腐病菌的生物学活性,采用单孢分离法对石榴病果中的病原菌进行分离纯化,然后进行分子鉴定和致病性测定,并通过室内生测法测定了嘧菌酯对其菌丝生长和孢子萌发的影响,且连续2年开展了大田防治试验。经形态学、分子生物学鉴定及致病性测定结果表明,石榴干腐病的病原菌为石榴壳座月孢Pilidiella granati。嘧菌酯在水杨肟酸(SHAM)的协同作用下,对石榴干腐病菌的菌丝生长和孢子萌发具有强烈的抑制作用,对其菌丝生长和孢子萌发的平均EC50分别为0.202μg/m L和0.007μg/m L(含100μg/m L SHAM);在大田防治试验中,嘧菌酯对石榴干腐病具有良好的防效,其中1 000倍25%嘧菌酯SC稀释液在2013和2014年的防效分别可达90.85%和81.91%,显著高于其它处理。表明嘧菌酯可作为防治石榴干腐病的候选药剂。     

9种生物制剂对甘薯长喙壳菌的抑制作用及防治效果

为筛选防治甘薯黑斑病的生物农药,测定了9种生物制剂对甘薯长喙壳菌孢子萌发和菌丝生长的抑制作用,并进行了防治效果验证。结果表明,四霉素对分生孢子萌发以及菌丝生长均表现出最强的抑制作用,EC50分别为0.10和0.15 mg/L;丁子香酚次之,EC50分别为0.24和0.93 mg/L。四霉素对薯苗黑斑病具有较好的防治效果,当接种甘薯长喙壳菌分生孢子浓度为5×104孢子/m L时,四霉素有效用量6与9 mg/L处理的防治效果分别为66.67%和78.57%;当接种浓度为5×105孢子/m L时,四霉素有效用量6与9 mg/L处理的防治效果分别为77.44%和90.23%。本研究为生物农药应用于甘薯黑斑病的防治提供了依据。     

Genetic diversity and interfertility among highly differentiated populations of Ceratocystis fimbriata in Brazil

Mating studies showed that isolates of the insect‐associated wilt pathogen Ceratocystis fimbriata from Eucalyptus spp., mango, fig, inhame (Colocasia esculenta), Gmelina arborea and sweet potato were interfertile, and progeny from those crosses showed normal segregation for microsatellite markers. Genetic diversity was compared among 13 populations of C. fimbriata collected from six states in Brazil using 15 highly polymorphic microsatellite markers. The gene diversity values of most eucalyptus and mango populations from Minas Gerais, Bahia, Rio de Janeiro and São Paulo states were similar to putatively native populations of Ceratocystis platani and C. cacaofunesta, two other species in the C. fimbriata complex that are homothallic. Index of association values indicated substantial asexual reproduction or selfing in populations on mango and eucalyptus. Most of these eucalyptus and mango populations were not highly differentiated from each other, and these populations and genotypes appeared to be more closely related to each other than to other populations by upgma analyses. By contrast, the G. arborea population from Pará and the fig and inhame populations from São Paulo had relatively low levels of diversity and were highly differentiated from each other and all other studied populations, suggesting that they were from different origins and had gone through genetic bottlenecks. One of the eucalyptus populations in Bahia consisted of a single genotype and may have been introduced to the site in infected cuttings from another Bahia location. Similarly, a mango population from Mato Grosso do Sul consisted of a single genotype, which was identical to one of the genotypes found on mango in São Paulo. Aside from introductions by humans, mating studies and genetic analyses suggest that limited dispersal distance and a high degree of selfing or asexual reproduction lead to local populations of C. fimbriata that have limited diversity but are highly differentiated from other populations.     

Comparison of populations of the wilt pathogen Ceratocystis albifundus in South Africa and Uganda

Ceratocystis albifundus is an important fungal pathogen of Acacia mearnsii trees in South Africa. In a previous study, a high level of gene diversity was demonstrated in a South African population of C. albifundus . This, together with the occurrence of the pathogen on native Protea species and its exclusive occurrence in South Africa, led to the hypothesis that C. albifundus is probably native to that country. More recently, C. albifundus has been reported from A. mearnsii in south-western Uganda. The aim of this study was to compare the populations of C. albifundus from Uganda and South Africa based on genetic diversity, population structure and possible gene flow. This was achieved using codominant microsatellite markers developed for the closely related species Ceratocystis fimbriata . Available isolates for comparison were from six different areas of South Africa and six jungle stands in Uganda. Eight of the 11 available markers amplified loci in C. albifundus . Gene diversity was higher in the Ugandan population, but genotypic diversity was greater for the South African isolates. There were no common genotypes between the two populations and they shared only 22% of the total alleles. The populations were genetically isolated from each other and highly substructured within. There was no association between isolates collected from the same geographic locations, and gene flow between the two populations was low. Results suggest that C. albifundus was probably not introduced into Uganda from South Africa but rather that an ancestral population, yet to be discovered, is the source of both populations.     

Genetic variation in populations of the cacao wilt pathogen, Ceratocystis cacaofunesta

Ceratocystis cacaofunesta (=  Ceratocystis fimbriata ) causes a lethal wilt disease of cacao ( Theobroma cacao ) in Latin America. Polymorphic microsatellite markers, (CAT)₅ nuclear DNA fingerprints and Hae III mitochondrial DNA fingerprints were used to compare genetic diversity among isolates of C. cacaofunesta collected from populations in western Ecuador, Costa Rica, Colombia, and Rondônia and Bahia in Brazil. Microsatellite markers and nuclear DNA fingerprints separated Ecuadorian isolates from isolates of the other four populations, and these two major groups correspond to genetic lineages already identified from ITS-rDNA sequences and intersterility groupings. Mitochondrial DNA fingerprints also demonstrated substantial diversity and split the Ecuadorian isolates into two groups. All marker types showed limited variation in the Colombian, Costa Rican and Bahian populations, as might be expected for introduced populations that have gone through recent genetic bottlenecks. In contrast, the Rondonian and western Ecuadorian populations showed gene diversity values similar to natural populations of other Ceratocystis species. The Rondonian population was the only sampled population in the native range of T. cacao (the Upper Amazon), and the putatively introduced populations were more closely related to the Rondonian population than to the western Ecuadorian population. The Ecuadorian population is in an area with other native Theobroma species, which may serve as natural hosts.     

中国发现甘薯长喙壳菌引起枣子果实腐烂

 2013年8月,在中国昆明发现一种枣子果实的腐烂病。通过致病性测试,证明从发病组织分离出的病菌能引起枣子果实腐烂。经过形态学和分子鉴定,确定该病原菌为甘薯长喙壳菌。这是在中国首次报道由甘薯长喙壳菌引起的枣子果实腐烂病。     

Histopathological aspects of mango resistance to the infection process of Ceratocystis fimbriata

Mango wilt, caused by Ceratocystis fimbriata, is one of the most important diseases affecting mango yields in Brazil. Information regarding the infection process of C. fimbriata in the stem tissues of mango from different cultivars and the basis of host resistance to the pathogen is rare in the literature. Thus, the objective of the study was to investigate how infection by two isolates of C. fimbriata can be affected by mango cultivar‐specific mechanisms of resistance. Disease progress on the inoculated stem tissues of the mango cultivars was evaluated and stem sections were obtained from the site of inoculation and prepared for histopathological observations using light microscopy. The factors mango cultivars and C. fimbriata isolates and their interaction were significant for all measures of disease development. Plants from the cultivars Espada, Haden and Palmer inoculated with isolates of C. fimbriata were more susceptible, whereas plants from the cultivars Tommy and Ubá were moderately resistant and resistant, respectively. Histopathologically, fungal isolates apparently massively colonized the stem tissues of plants from the susceptible cultivars Espada, Haden and Palmer, starting from the collenchyma and moving in the direction of the cortical parenchyma, xylem vessels and pith parenchyma. By contrast, on stem tissues of plants from the resistant cultivars Tommy Atkins and Ubá, most of the cells reacted to C. fimbriata infection by accumulating amorphous material. The results from the present study strongly indicated the importance of phenolic compounds for mango cultivar resistance against infection by Brazilian C. fimbriata isolates.     

土壤样品中长喙壳属真菌诱导分离方法的研究

 长喙壳属真菌(Ceratocystis spp.)是一类重要的土传植物病原真菌,可危害多种作物。由于技术方法的限制,对于土壤样品中该病原菌的分离比较困难。本研究通过比较以胡萝卜、马铃薯、甘薯作为诱导富集载体对土壤中长喙壳属真菌的富集分离效果,确定了胡萝卜为最优诱导富集载体。进一步通过对胡萝卜片厚度、土壤相对湿度、培养温度、光照条件、保湿方式等培养条件的优化,建立了从土壤中分离长喙壳属真菌的有效方法,具体为:以厚度为3~5 mm的胡萝卜片作为诱导富集载体;土壤样品相对含水量为10%~30%;12 h光暗交替或24 h黑暗;在22℃~26℃条件下不保湿进行培养,8 d后可从携带最低数量为10个孢子/g的土壤样品中分离到靶标菌。采用该方法,对采自新疆、河南、湖南、云南等地共计108份自然或耕作土壤样品进行了分离,结果表明从采自云南蒙自市、临沧市的45份土壤样品中均分离获得了长喙壳属真菌。该方法具有操作简便、所需仪器设备简单、灵敏度较高、成本低和高通量等特点,适用于田间调查,可对样本量较大的土壤样品进行快速及时分离,以获得不同土壤生境中的长喙壳属真菌分离物。     

Populations of Ceratocystis fimbriata on Colocasia esculenta and other hosts in the Mata Atlântica region in Brazil

Ceratocystis fimbriata is native to Brazil, where it is able to cause serious diseases on numerous hosts, especially on non‐native plants. Because C. fimbriata is soilborne and not wind dispersed, highly differentiated populations are found in different regions of Brazil. The present study compared populations of C. fimbriata on taro, mango, eucalyptus and kiwifruit from the coastal Mata Atlântica region with native populations of the fungus from the Cerrado‐transition region in Brazil by using 14 SSR markers and DNA sequences of ITS and mating type genes. Microsatellite and phylogenetic analyses were performed to test the hypothesis that populations on different hosts from the Mata Atlântica region are related to each other and are native to the region. The ITS sequences varied greatly among the taro isolates, with six sequences identified, from which two had not been previously reported. For mating type genes, four sequences were identified among the isolates on taro, mango, eucalyptus and kiwifruit. Phylogenetic analyses showed that Mata Atlântica populations formed a monophyletic group distinct from Cerrado‐transition region populations, although earlier studies had shown that isolates from the two regions are interfertile and are considered as a single biological species. Microsatellite analysis revealed low gene diversity for each of the three Mata Atlântica populations on taro, mango and kiwifruit, suggesting that these populations had gone through genetic bottlenecks, probably by dispersal of select genotypes in vegetative propagation material. Also, microsatellite markers showed that two microsatellite genotypes from taro are widely spread in Brazil, probably by infected corms.