致病疫霉超级生理小种遗传多样性分析

通过交配型和甲霜灵抗性以及线粒体DNA单倍型、SSR和AFLP基因型分析对40个超级生理小种菌株进行了遗传多样性分析。在被测菌株中发现了A1、A2和自育3种不同类型的交配型。其中,A1和自育型菌株数量多,分别为21株和14株,而A2交配型仅5株。甲霜灵抗性测定检测出高抗菌株26株,敏感菌株14株。线粒体DNA单倍型测定出Ia型和IIa型两种,比例接近1:1。基于5个基因座被测40个超级生理小种菌株共鉴定出了7种SSR基因型。利用6对荧光引物共检测到258条AFLP谱带,其中多态性谱带204条,多态性为79.1%。将供试的40个菌株划分为38个基因型,几乎每个菌株都为1个特有基因型。而且,我国南方和北方超级生理小种群体存在着明显的遗传差异。结果表明我国致病疫霉超级生理小种具有丰富的遗传多样性,可以推断致病疫霉中的任何小种都可在多个抗病基因的强大选择压力下,在短时间内通过与之对应的无毒基因快速突变而成为超级生理小种。当前对致病疫霉生理小种的鉴定及监测对生产上利用抗病品种防控晚疫病的指导意义不大。     

河北省番茄上致病疫霉Phytophthora infestans群体结构的分析

番茄晚疫病是河北省番茄生产上最具毁灭性的病害之一,对引起该病害的致病疫霉群体结构进行分析有利于病害的防治。利用对峙培养法和菌落直径法对2007-2008年采自河北省保定、沧州和唐山分离自番茄的49个致病疫霉菌株进行了交配型和甲霜灵抗性的表型测定,结果表明该群体所有菌株均为A1交配型,以甲霜灵敏感菌株为主,抗性菌株仅7株。利用聚合酶链式反应-限制性片段长度多态性(PCR-RFLP)、简单序列重复(SSR)和扩增片段长度多态性(AFLP)等分子技术对该群体的基因型进行了分析,结果表明供试菌株线粒体基因型均为Ia型,共鉴定出了Ⅰ、Ⅱ和Ⅲ3种SSR基因型,AFLP聚类分析在相似系数0.87时可以形成α、β和γ等3个不同的分支。河北省所有番茄上致病疫霉菌株均分布在α分支上,该分支又可进一步分为7个亚分支。AFLP亚分支与甲霜灵抗性和地理来源均无明显相关性,但Ⅱ型SSR与甲霜灵抗性和地理来源有明显的相关性。综合表型和基因型数据说明河北省番茄上致病疫霉群体结构比较单一,遗传多样性程度较低。     

四川省马铃薯晚疫病菌群体表型和遗传变异的分析

致病疫霉Phytophthora infestans引起的晚疫病是马铃薯的一种毁灭性病害。为了对四川省近年马铃薯晚疫病菌进行系统多样性分析,本研究从表现型和基因型两个方面鉴定了四川省马铃薯晚疫病菌的群体多态性。交配型、甲霜灵敏感性和生理小种的鉴定结果发现A1交配型菌株只有2份,A2型有29份,自育型12份;甲霜灵敏感菌株3株,中抗菌株22株,高抗菌株18株;共测定出11个生理小种,其中全毒力小种发生频率最高,占供试菌株的25.58%。基因型鉴定结果表明,43份材料共发现了10种SSR基因型,5个SSR标记在该种群上共产生了16个SSR位点,每个标记平均有3.2个位点,多态性信息含量平均值为0.46,SSR4是多态性最丰富的标记。本试验中A2菌株和自育型的基因型SSR相同,其中全毒力小种是当地的优势致病菌,该研究为今后四川马铃薯晚疫病的有效防控提供理论依据。     

福建省部分地区马铃薯致病疫霉群体遗传多样性分析

致病疫霉(Phytophthora infestans)引起的晚疫病是马铃薯的一种毁灭性病害。有效控制马铃薯晚疫病需要明确致病疫霉的群体遗传结构特征。采用8对SSR引物对采自福建省福州、长乐、漳州2010年分离的95株马铃薯致病疫霉进行遗传多样性分析。结果共检测出21个等位基因和26个基因型。三个地点致病疫霉菌群体间的平均遗传分化系数FST为0.22,在8个位点中有5个位点的等位基因频率分布差异显著。三个群体的观测纯合度小于期望纯合度,观测杂合度大于期望杂合度,以无性生殖为主。结果表明福建群体的遗传多样性高,群体间的存在较高的遗传分化度。     

安徽和黑龙江省大豆疫霉群体遗传结构的SSR分析

采用简单重复序列(SSR)的分析方法,对来自安徽和黑龙江省的大豆疫霉群体进行了遗传多样性分析。通过使用20对SSR引物对供试的83株大豆疫霉菌株进行PCR扩增,共得到109个SSR标记,全部为多态性标记,平均每对引物扩增出5.5条带。遗传变异与相似性分析表明,安徽群体具有更高的遗传变异度,安徽群体与黑龙江群体间遗传相似性较低;聚类分析显示,供试菌株在80%的相似性水平上可被区分为7个类群,且安徽群体分布于更多的聚类组中;Shannon-Wiener多样性指数也表明安徽群体的遗传多样性较黑龙江群体丰富。综合分析表明,本研究的结果不支持关于安徽的大豆疫霉可能来源于黑龙江的推测。     

致病疫霉有性生殖诱导及F1代遗传多样性

致病疫霉Phytophthora infestans为马铃薯晚疫病的重要病原菌。通过从昆明市寻甸县采集110P和H-6两株致病疫霉,明确其染色体倍性、交配型、线粒体单倍型、毒性和甲霜灵敏感性,经对峙培养,利用改良的卵孢子萌发方法获得有性生殖F1代群体POP1（60株）,并对POP1进行表型和基因型测定。结果表明：冷冻处理24h为最佳条件,卵孢子萌发率达5.09%±0.15%;POP1的交配型、毒性和甲霜灵敏感性均发生了分离,其中交配型分离比为A1:A2:A1A2:自育型（SF）=16:5:17:22,毒性分离比为抗性（R）:敏感性（S）=11:49,甲霜灵敏感性分离比为抗性（R）:敏感性（S）=2:58;3个表型的分离均偏离孟德尔单基因显性遗传特点。基于8对SSR多态性引物对POP1基因型分析表明,遗传相似系数为0.98时,可将所有菌株分为14个基因型;遗传相似系数为0.95时,可将POP1分为6个分支,其中优势群体为S1,占分离群体的61.67%。关联分析进一步表明,8对SSR所代表的基因型和几个重要表型有显著相关性（R2=0.6667）。本研究建立了高效的致病疫霉卵孢子萌发体系,解析了有性生殖后代群体遗传结构特点,为深入探索致病疫霉的变异规律及病害流行趋势提供了理论基础。     

一个马铃薯种质资源圃致病疫霉群体的分析

由致病疫霉Phytophthora infestans引起的晚疫病是马铃薯生产上最严重的病害之一,认识其群体结构特征,可为晚疫病防控策略的制定以及抗病品种的合理布局提供指导。对2009年采自宁夏一个种植有93个品种(品系)的马铃薯种质资源圃的致病疫霉进行了交配型、致病型和线粒体DNA单倍型分析,结果表明,116个致病疫霉菌株中存在A1、A2和自育型3种交配型,发生频率分别为24.1%、57.8%和18.1%,A2交配型为优势类型;对其中43个菌株的致病型进行测试,检测到两种致病类型:1.2.3.4.5.6.7.8.9.10.11和3.4.10,发生频率分别为95.3%和4.7%,可克服所有11个抗病基因的1.2.3.4.5.6.7.8.9.10.11类型占绝对优势;对62个菌株的线粒体DNA单倍型进行分析,检测到Ia和IIa两种类型,发生频率分别为74.2%和25.8%。综合表型和基因型数据分析发现,该马铃薯种质资源圃中致病疫霉群体致病型单一,但致病型毒力因子高度复合;线粒体DNA分析表明,该马铃薯种质资源圃引入了遗传背景较为复杂的致病疫霉"新"群体。     

致病疫霉在中国云南的马铃薯田间形成卵孢子*

由致病疫霉Phytophthora infestans (Mont.)de Bary引起的马铃薯和番茄晚疫病是世界性的作物病害,每年均造成巨大的经济损失和社会影响。致病疫霉是异宗配合的卵菌,有两个已知交配型A1和A2,两个相对交配型互作时可进行有性生殖产生卵孢子( Gallegly & Galindo,1958)。过去许多年一直认为致病疫霉在除墨西哥以外的国家中只存在A1交配型,通过产生孢子囊进行无性繁殖。近年来,由于致病疫霉新致病群体的产生以及全球范围的迁移和替代,A2交配型菌株先后在欧洲、美洲、亚洲和非洲的许多国家被发现。A1、A2两种交配型菌株的同时存在,增…     

华南瓜类疫霉的RAPD遗传多样性分析

为了解来自广东和广西的瓜类疫霉的遗传多样性,利用从180条RAPD引物中所筛选出的多态扩增性强、重复性好的12条引物,对分离自两省区的96株瓜类疫霉进行了全基因组DNA遗传多样性分析和指纹图谱构建。通过对供试菌株的RAPD-PCR扩增,共获得135条DNA标记谱带,其中124条为多态性谱带,多态检测率为91.9%。利用NTSYSpc Version2.1软件对供试菌株间的遗传距离进行聚类分析并构建系统树,以遗传相似系数0.81为阈值,将96个供试菌株划分为12个RAPD群,多数分离物之间遗传相似性较低,在DNA水平上存在显著的遗传变异,具有较丰富的遗传多样性。不同地区间菌株的遗传分化程度不同,分离自黄瓜的菌株遗传分化明显高于分离自冬瓜的菌株。RAPD群与菌株地理来源、分离寄主、致病力、交配型及甲霜灵抗性均无明显的相关性。     

致病疫霉组蛋白H2A变异体序列与表达

在真核生物中,DNA缠绕在组蛋白上形成核小体,一个组蛋白分子包括H2A、H2B、H3和H4各2个核心组蛋白亚基。在这4种核心组蛋白中,H2A富含多样化,且在细胞的生物途径中起重要作用的变异体,因此,H2A家族一直是研究热点。致病疫霉是重要的病原菌也是研究卵菌的模式物种之一,目前关于卵菌表观遗传的研究还未见报道。本研究针对致病疫霉组蛋白H2A变异体,利用基因组信息和基因芯片数据,通过序列比对、系统发育分析以及基因表达水平检测,发现在致病疫霉基因组中存在组蛋白H2A变异体H2A.X.1、H2A.X.2和H2A.Z,它们在不同生长发育阶段和侵染过程呈现特异的表达谱。研究结果为进一步研究致病疫霉表观遗传机制奠定了基础。     

Phytophthora infestans isolates from Northern China show high virulence diversity but low genotypic diversity

Phenotypic and genotypic characteristics of 48 Phytophthora infestans isolates , collected in five provinces in Northern China between 1997 and 2003, were determined and compared with reference isolates. Characterisation included mating type, virulence, mitochondrial DNA (mtDNA) haplotype and DNA fingerprinting patterns based on simple sequence repeats (SSR) and amplified fragment length polymorphisms (AFLP). All isolates had the A1 mating type, mtDNA haplotype IIa and an identical SSR genotype (designated as SG-01-01) that differed from SSR genotypes found in the reference isolates, including those representing the 'old' US-1 lineage that dominated the P. infestans population worldwide prior to 1980. In contrast, the virulence spectra were highly variable and virulence to all resistance genes present in the standard differential set ( R1 to R11 ) was found. AFLP analysis revealed some diversity; eight different AFLP genotypes were found that could be grouped into two major clusters. This study shows that there is very little genotypic diversity in the P. infestans population in Northern China. The occurrence of many different races within this rather uniform population is discussed in the framework of recent insights into the molecular determinants of avirulence in potato– P. infestans 'gene-for-gene' interactions.     

Phenotypic and Genotypic Characterization of Phytophthora infestans Isolates from China

A total of 288 (202 from potato and 86 from tomato) isolates of Phytophthora infestans were collected from 1998 to 2007 in China. The isolates were characterized based on mating type, in vitro metalaxyl sensitivity, virulence on potato differentials, allozymes of glucose-6-phosphate isomerase ( Gpi ), peptidase ( Pep ), and mitochondrial DNA (mtDNA) haplotype and examined by DNA-based simple sequence repeat (SSR) and random amplified polymorphic DNA (RAPD) fingerprinting. The majority (283 of 288) of the isolates were of the A1 mating type, the other three were the A2 mating type and two were the A1A2 mating type. Resistance to metalaxyl was frequently observed, with 248 (86.1%) resistant, 21 (7.3%) intermediate and 19 (6.6%) sensitive isolates identified. Virulence was assessed for 125 isolates on a set of 11 potato differentials and 61 races were detected. Most isolates were virulent on the differential genotype with gene R3, and all known virulence genes were found, with race 3.4.7.11 being the most common. This pattern did not appear to be associated with geographic origin, sample type, mating type or metalaxyl sensitivity. The dominant banding patterns for Gpi were 100/100/111 (176 isolates) and 100/100 (109 isolates), but genotypes 86/100 and 100/111 were also identified. All isolates tested were homozygous (100/100) at the Pep locus. The majority (205 of 288) of isolates tested was of mtDNA haplotype IIb, 76 were haplotype IIa and seven were the rare Ib haplotype. The genetic diversity of 60 representative isolates from China was assayed by two types of molecular markers, RAPD and SSR. A high level of polymorphism was found. The results demonstrated the diverse phenotypic and genotypic structure of the current populations of P. infestans in China.     

Genetic analysis of Phytophthora infestans populations in the Nordic European countries reveals high genetic variability

Late blight, caused by the oomycete Phytophthora infestans, is the most important disease of potato (Solanum tuberosum). The pathogen is highly adaptable and to get an overview of the genetic variation in the Nordic countries, Denmark, Finland, Norway and Sweden we have analyzed 200 isolates from different fields using nine simple-sequence repeat (SSR) markers. Forty-nine alleles were detected among the nine SSR loci and isolates from all four Nordic countries shared the most common alleles across the loci. In total 169 multilocus genotypes (based on seven loci) were identified among 191 isolates. The genotypic diversities, quantified by a normalized Shannon's diversity index (H(s)), were 0.95 for the four Nordic countries. The low F(ST) value of 0.04 indicates that the majority of variation is found within the four Nordic countries. The large number of genotypes and the frequency distribution of mating types (60% A1) support the hypothesis that sexual reproduction is contributing notably to the genetic variation of P. infestans in the Nordic countries.     

中国马铃薯晚疫病菌AFLP遗传多样性分析

应用AFLP分子标记检测了我国部分马铃薯主要产区马铃薯晚疫病菌的遗传多样性及不同地区菌株间的亲缘关系。在200对引物组合中,利用6个菌株筛选出12对多态性好、带型清晰的引物组合。利用这12对引物组合对1997-2002年间采自我国黑龙江、河北、四川和云南4省的50株菌株进行了PCR扩增,共扩增出922条谱带,其中多态性标记530条,占57.5%。利用NTSYSpc软件中UPGMA算法构建了我国马铃薯晚疫病菌的亲缘关系树状图,聚类分析结果表明我国马铃薯晚疫病菌的遗传多样性与病原菌的地理来源有一定的相关性,而与交配型、生理小种和对甲霜灵的抗性无明显的相关性。用POPGENE软件计算了各群体间的遗传多样性参数,结果表明我国马铃薯晚疫病菌的遗传多样性程度不高,不同地区种群间分化不明显。     

Potato late blight in Morocco: characterization of Phytophthora infestans populations (virulence and mating type)

Late blight caused by Phytophthora infestans, is the most important disease of potato in Morocco. Use of partially resistant cultivars should be an essential component of a sustainable management strategy of potato late blight, provided the durability of this form of resistance. It is therefore important to determine the nature of P. infestans Moroccan populations. Mating types were determined for 91 strains of P. infestans collected in the northern (Larache-northern plain), north western (Kénitra) and north eastern (Méknès, Middle Atlas) potato cropping areas of Morocco in 1999-2000, 2000-2001 and 2003-2004. They showed a clear regional structure of these populations, with the presence of both mating types (A1 and A2). Of all isolates collected since 1999, A2 mating type constituted 56% (54 isolates), following by A1 mating type (40.7%, 31 isolates) and A1-A2 (self-fertile) mating type (3.30%, 3 isolates). Populations from Méknès and Kénitra consisted mainly of A2 mating type, whereas populations from Larache predominantly included A1 mating type. Physiological race study revealed the presence of 19 races of P. infestans in the first collection of 25 isolates tested between 1999 and 2001. All known virulence genes were detected in western and northern Moroccan isolates, except virulence for resistance genes R2, R5, and R6 which were absent. All isolates were able to overcome two or more R genes except one isolate (5-1) corresponding to race 1.     

Diversity of the Phytophthora infestans population in Flanders, Belgium

A total of 94 isolates of Phytophthora infestans were collected from disease outbreaks in commercial potato crops and private gardens in 2002 and 2003. The isolates were recovered successfully from single lesions of diseased potato foliage. Not from all isolates pure cultures were obtained due to contaminations with Fusarium species and bacteria. The structure of the population was analysed phenotypically. Characteristics of the isolates included in vitro growth rate, mating type, in vitro sensitivity to the phenylamide fungicide metalaxyl-M and allozyme genotype at glucose-6-phosphate isomerase (Gpi) and peptidase (Pep) loci. Significant differences in in vitro growth rate were observed among the 52 isolates by comparing the main radial growth of the isolates after 7 days. Forty seven from the isolates tested were the Al mating type. Only one isolate was characterized as A2 mating type. Isolates with sensitive, intermediate and resistant responses to metalaxyl-M were detected in the populations. Forty isolates had a growth of less then 40 % at 5 ppm metalaxyl-M. Three isolates had a growth of less then 40 % at 100 ppm metalaxyl-M. Eight isolates had a growth of more then 40 % at 5 and 100 ppm metalaxyl-M. Cellulose acetate electrophoresis was used to examine Gpi and Pep banding pattern of the population of P. infestans attacking potato in Flanders. All the isolates tested produced the 100/100 Gpi isozyme electromorph. Five different allozyme genotypes of the Pep loci were identified: 92/92, 96/96, 100/100, 92/100, 83/100.     

Monitoring of Virulence in Populations of Pbytophthora infestans Using a Schwarzbach Spore Trap

A modified Sehwarzbach jet spore trap was used to sample Phytophthora infestans populations in small crops of potato within polyethylene tunnels and in the field. Airborne sporangia were eaptured in the trap and deposited upon potato leaf discs, of a cultivar lacking race-specific resistance genes, contained within the trap. Virulence characters of isolates captured in the spore trap were determined by subculturing the isolates on leaf discs of a range of potato genotypes with race-specific resistance (R-genes), incubating them and then recording compatible and mcompatible reactions, reactions. For four isolates of P. infestans the levels of sporulation on the differential leaf discs were noted and although cultivar × isolate interactions were inconsistent, variable light and temperature conditions had no significant effect on the reactions of the cultivars.