稻纹枯菌酯酶同工酶、可溶性蛋白及致病力的研究初报

 对稻纹枯菌25个菌株分别进行了致病力测定、酯酶同工酶和可溶性蛋白分析.结果表明:不同菌株存在明显的致病力分化,而酯酶同工酶图谱却有较强的一致性,它们的主酶带基本相同,但在副酶带上反映出一定的同工酶表型异质性,表现为带的数目及着色深浅的不同;可溶性蛋白图谱比酯酶同工酶图谱表现出更多的多样性,大多数致病力较强的菌株都有l~2条特征谱带,而且它们的谱带数目比弱毒力菌株的多出3~5条.     

稻瘟菌不同小种间可溶性蛋白及酯酶同工酶的多态性

采用聚丙烯酰胺凝胶泳对来自太湖稻区的１０个水稻稻瘟病菌株和２个禾本科杂草瘟菌的可溶性蛋白及酯酶同工酶进行了分析比较。结果表明：供试菌株菌体可溶性蛋白可呈现１６～２５条电泳谱带，其中在Ｒｆ值为０．２３，０．３３，０．４４的３条谱带为１２２个菌株所共有，没有发现生理小种特异性谱带。对１２个菌株α－酯酶同工酶的分析表明，在所有菌株同工酶谱带中可分辩出具有不同迁移率的１２条谱带，据此可将菌株分为８个不同的同工酶谱带类型。同时表明，α－酯酶同工酶在太湖稻区稻瘟病菌中存在丰富的多态性，但其谱带类型与病菌生理小种无相关性。     

稻瘟病菌生理小种酯酶同工酶特性的研究

 运用聚丙烯酰胺凝胶等电聚焦法对来自江、浙两省经全国统一鉴别寄主鉴定的稻瘟病菌7群12个生理小种的16个菌株菌体酯酶同工酶进行了分析比较,结果表明:稻瘟菌菌体酯酶同工酶带有13-18条,等电点分布范围为3.00-7.20。在等电点4.15附近有各生理小种共同的酶带。同一生理小种群内不同生理小种间酯酶同工酶的相似系数一般大于0.50,不同生理小种群的生理小种间酶谱的相似系数一般为0.30-0.40,也存在少数不是同一生理小种群的小种间酶谱相似系数大于0.50的情况。供试各生理小种间酶谱均存在不同程度的差异,表现为酶带数目的多少和等电点分布的不同。来自不同省份经鉴别寄主鉴定为同一生理小种的菌株间酶谱的相似系数亦仅0.05-0.60。研究结果初步反映出供试大多数稻瘟菌菌株间菌体酯酶同工酶的相似性与致病性有关,但也受菌株来源地生态环境影响。本文还讨论了稻瘟病菌致病性容易变异但在许多情况下又具有相对稳定性的可能机制,探讨在一定范围内聚丙烯酰胺凝胶等电聚焦技术用于稻瘟菌生理分化和致病性变异监测的可能性及尚待进一步研究解决的穗题等。     

西藏高寒植物垫状雪灵芝酯酶同工酶变异研究

以色季拉山的高寒植物垫状雪灵芝为材料,选取分布在海拔4500~4650m范围内的垫状雪灵芝3个居群为研究对象,采用聚丙烯酰胺琼脂电泳技术,对垫状雪灵芝3个居群的48个个体功能叶片的酯酶同工酶谱带进行测定并编码,依据酶谱带测定结果进行聚类分析;同时结合酯酶同工酶的3个遗传指标进行相关分析。结果表明:来自同一居群的个体,同工酶谱带相似性较大,居群内的遗传杂合度较小(HeA=0.735,HeB=0.765,HeC=0.726),这也说明居群内个体存在一定程度的遗传分化;酯酶在不同居群间的相似性系数的平均值相对较小,遗传距离相对较大(IAB=0.392,IAC=0.542,IBC=0.610;DAB=0.937,DAC=0.613,DBC=0.495)。研究数据显示垫状雪灵芝酯酶同工酶在不同的居群内与居群间存在不同程度的遗传分化,缺少有效的基因交流。     

新月弯孢菌及其近似菌菌落形态和可溶性蛋白电泳图谱比较分析

通过对分离自玉米及其他寄主的 12个新月弯孢菌菌株和 5个近似菌株菌落形态及可溶性蛋白电泳谱比较分析 ,表明菌落形态在不同种之间存在明显的差异 ,同种之间也存在一定差异 ,但不稳定和存在变异。 17个菌株可溶性蛋白SDS聚丙烯酰胺凝胶电泳显示 ,Curvularia属在Rf值为 0.177处有一条该属的特征蛋白带 ,新月弯孢菌在Rf值为0.225处有一条该种的特征蛋白带 ,但新月弯孢菌蛋白带存在多样性 ,聚类分析将未能确定的 13号菌株定为不等弯孢菌     

鱼类肝脏酯酶的同工酶构成及其对磷酸三苯酯和马拉氧磷的敏感性

用聚丙烯酰胺凝胶电泳法分离了麦穗鱼 (Pseudorasbora p arva)、金鱼 (Carassius auratus)、尼罗罗非鱼 (Tilapja nilotica)、食蚊鱼 (Gambusia affinis)、虹鳟 (Salmo gairdneri)等五种鱼的肝脏酯酶的同工酶 ,以乙酸α-萘酯为底物测定了它们的活性。在麦穗鱼、金鱼、尼罗罗非鱼体内发现两条酶带 ,在食蚊鱼和虹鳟体内发现一条酶带。离体抑制率实验表明 ,五种鱼肝脏内均含有对磷酸三苯酶敏感的酶带 ,而对马拉氧磷敏感的酶带只存在于麦穗鱼、金鱼、尼罗罗非鱼体内。酯酶同工酶分布型的意义及与马拉硫磷选择毒性的关系在本文中进行了讨论。     

转sck/cry1Ac基因稻谷对米蛾幼虫解毒酶系的影响

采用酶活力测定及酯酶同工酶电泳方法研究米蛾(Corcyra cephalonica)幼虫取食表达SCK/Cry1Ac毒蛋白的转基因稻谷后体内重要解毒酶活力的变化,明确其代谢毒蛋白的主要酶类。结果表明:取食转Bt基因稻谷米蛾幼虫体内的α-乙酸萘酯酶、碱性磷酸酯酶的活力在24～72h显著低于对照;酸性磷酸酯酶活力在12、48h和72h时也显著低于对照;而α-乙酸萘酯羧酸酯酶和乙酰胆碱酯酶的活力显著高于对照。谷胱甘肽-S-转移酶的活性除饲喂24h的处理外,其他处理与对照无显著差异。此外处理72h后米蛾幼虫体内酯酶同工酶谱有4条酯酶酶带,4种酯酶Est1、Est2、Est3、Est4的活性均受到不同程度的抑制,酯酶Est4的抑制效果最明显。这些解毒酶和酯酶同工酶直接参与Bt毒蛋白的代谢作用。     

贝莱斯芽孢杆菌Bacillus velezensis YB15β-葡聚糖酶的抑菌作用与基因克隆

本文在对峙法验证贝莱斯芽孢杆菌B. velezensis YB15具抑菌作用的基础上,用透明圈法、DNS法研究其产β-葡聚糖酶特性,利用对峙法验证该酶抑菌作用,通过PCR法获得目的基因,分析克隆序列并预测其蛋白质结构与功能。结果表明,该菌株对多种病原真菌有拮抗作用,杨树紫纹羽病菌拮抗带达11.0 mm,该菌提取的葡聚糖酶粗酶液对杨树紫纹羽病菌抑菌带为10.6 mm,说明葡聚糖酶在菌株YB15抑菌中有重要作用。不同接种方法影响菌株YB15葡聚糖酶水解透明圈形成,点种法水解圈与菌落直径之比在72 h可达14.1,效果最好。克隆所得菌株YB15葡聚糖酶基因命名为Bglu1,该基因序列长732 bp,编码243个氨基酸,此酶蛋白氨基酸序列与解淀粉芽孢杆菌TB2β-1,3-1,4-葡聚糖酶同源性较高,属糖基水解酶16家族,N端疏水区存在信号肽并具跨膜区域,推测其为分泌蛋白。     

余庆县茶叶斑病致病菌Pestalotiopsis trachicarpicola鉴定及其生物学研究

为有效控制贵州省余庆县茶叶斑病的发生及为害,从该区域茶树叶部病害样品中分离纯化多个菌株,并对其中多个形态相似的分离物进行菌落形态、子实体、产孢细胞、分生孢子的形态观察.测定了核糖体转录间隔区(ITS)、微管蛋白(TUB2)和翻译延长因子1-α(TEF-1α)等核酸序列,采用PAUP软件及最大简约法,进行了多基因系统发育...     

棉蚜不同抗性品系羧酸酯酶比较

用分光光度计终点测定法和酶标仪动力学测定法对3个抗性水平不同的棉蚜品系和1个敏感品系的羧酸酯酶进行了研究。棉蚜的不同抗性品系羧酸酯酶对α-乙酸萘酯(α-NA)和β-乙酸萘酯(β-NA)的水解活性具有极显著的相关性(r=0.95,n=400)。两种测定法均显示出R1、R2和R3品系的羧酸酯酶比活力明显高于S品系。以α-NA为底物时,用酶标仪动力学测定法研究表明,S、R1、R2和R3棉蚜品系羧酸酯酶活性分别为38、85、198和762mOD·min~(-1)·aphid~(-1);与4个品系的抗性程度比较,酶动力学方法的测定结果更可靠。     

Evidence of asexual recombination in Rhynchosporium secalis

Three single-spore isolates of Rhynchosporium secalis that differed in their α-esterase and β-glucosidase isozyme patterns were inoculated as two mixtures, each of two isolates, on to seedlings of the susceptible barley cultivar Maris Mink. Approximately 100 single-spore isolates were taken from mature lesions produced by each mixture. These were subjected to polyacrylamide gel electrophoresis and the gels were stained for α-esterase and β-glucosidase. Parental types only were produced by one of the isolate mixtures. However, one of the 10 lesions examined for the second mixture produced nine isolates with a novel combination of isozymes, indicating that some form of asexual recombination had occurred. The use of isozymes as a natural marker system for the detection in vivo of asexual recombination in pathogenic fungi is discussed.     

大豆疫霉菌单孢分离物生物学性状的遗传变异研究

 本文研究了大豆疫霉菌单游动孢子无性分离物和自交单卵孢子分离物的菌丝生长速率、菌落形态、同宗配合性状、产孢量以及对甲霜灵敏感性的遗传变异。结果表明:菌落形态、生长速率和同宗配合性状在单游动孢子后代和自交后代可稳定遗传,控制上述性状的遗传因子是纯合的;大豆疫霉菌的游动孢子产生能力和对甲霜灵的敏感性在单游动孢子后代和自交后代中均发生连续性变异,表明这两种性状可能是数量遗传性状,也可能控制这两种性状的基因为杂合基因或细胞质遗传因子。     

Characteristics of strains of Septoria nodorum adapted to wheat or to barley

Sixty-four strains of Septoria which had been isolated from a range of hosts at different locations and in different years were characterized for their adaptation to wheat or barley, growth at near-maximum temperature, fluorescence, colony morphology, conidial (pycnidiospore) length and hexokinase and alkaline phosphatase isozymes. For each character except conidial length, the strains could be divided into two or three discrete groups. The variation in these six characters was strongly associated, such that 60 strains could be classified into two groups, designated W-type and B-type. W-type strains are adapted to wheat, produce large colonies at 31 C, fluoresce, produce brown-pigmented colonies, and have fast isozymes. B-type strains are adapted to barley, produce small colonies at 31 C, do not fluoresce, produce pink-pigmented colonies, and have slow isozymes. A few strains differed from these norms in one of the six characters, but only one showed an atypical host adaptation. The four unclassified strains differed from W- or B-type in two or more characters. The many differences between the W- and B-types suggest they are genetically distinct populations within Septoria nodorum.     

Relationship Between Biological Control, Incidence of Hypovirulence, and Diversity of Vegetative Compatibility Types of Cryphonectria parasitica in France

ABSTRACT In France, chestnut blight, caused by Cryphonectria parasitica, has been controlled since 1974 in orchards, but never in coppice forests, by releasing hypovirulent strains infected with CHV1 hypovirus. We tested the hypothesis that this biological control (BC) has lead to a decrease in blight severity, spread of hypovirulence, and change in C. parasitica populations. The low severity of chestnut blight was confirmed in the six regions studied (subdivided into zones). The remission of cankers was associated with the presence of white isolates presumed to be hypovirulent. These two parameters were also correlated, at the zonal level, to the frequency of sites where BC was used. However, the estimates of the natural background level of hypovirulence, independent of BC, ranged from 4% in forests in Dordogne to 60% in orchards in Lozère. Differences in the rate of hypovirulent isolates among regions were consistent with the diversity of vegetative compatibility (VC) types in populations of C. parasitica. The highest VC-type diversity and mean allelic diversity for known vegetative incompatibility (vic) genes were observed in Dordogne. We showed that the current diversity of VC types in populations of C. parasitica was lower than in 1981. We found 30 VC types among 1,113 isolates of C. parasitica. Ten VC types were incompatible with known EU testers, suggesting that one additional vic gene or allele at one of the six vic loci known should be present in Europe.     

Phytophthora boehmeriae boll rot: a new threat to cotton cultivation in the mediterranean region

A boll rot of cotton (Gossypium hirsutum L.) was observed for the first time in Greece in August 1993 in Larissa and Volos counties, and in August and September 1995 in Trikala and Phthiotis counties. Fungi of the genusPhytophthora were isolated from diseased plants. Morphological characteristics of the pathogen were recorded on mounts made directly from the infected tissues or after growth of the isolated fungus on corn meal agar or sterile distilled water. Colony morphology, growth rates, features of asexual and sexual structures and maximum growth temperatures were examined. APhytophthora species new to Europe,Phytophthora boehmeriae Sawada, attacking cotton bolls, was identified. The pathogenicity of the isolates was confirmed by artificial inoculations of detached cotton bolls. Analysis of α-esterase isozymes revealed unique banding patterns for isolates ofP. boehmeriae compared with those ofP. cactorum andP. parasitica, which arePhytophthora species with similar morphology.     

Phylogenetic history of Phytophthora cryptogea and P. drechsleri isolates from floriculture crops in North Carolina greenhouses

The evolutionary history of Phytophthora cryptogea and P. drechsleri isolates previously collected from floriculture crops in North Carolina commercial greenhouses was explored with coalescent- and parsimony-based analyses. Initially, 68 isolates representing 13 location-host groups were sequenced at multiple loci. Sequences of all isolates within a group were identical. A subset of isolates were selected, cloned to resolve heterozygous sites, and analyzed with SNAP Workbench. The internal transcribed spacer (ITS) region of the ribosomal DNA and cytochrome oxidase II gene genealogies were congruent and indicated that P. cryptogea and P. drechsleri are sister species diverged from a common ancestor with no evidence of gene flow. In contrast, genealogies inferred from β-tubulin (β-tub) and translation elongation factor 1α (EF-1α) genes were in conflict with these loci. Coalescent analysis based on a nonrecombining partition in β-tub and EF-1α showed an initial (older) split between P. cryptogea and P. drechsleri, with a later (recent) event separating the remaining P. cryptogea haplotypes from P. drechsleri. A parsimony-based minimal ancestral recombination graph inferred recombination between P. cryptogea and P. drechsleri isolates in the ITS region and β-tub, suggesting genetic exchange between species. Also, putative recombination between A1 and A2 mating types of P. cryptogea suggests that sexual reproduction has occurred in the history of these P. cryptogea isolates.     

Segregation of non‐target‐site resistance to herbicides in multiple‐resistant Alopecurus myosuroides plants

Non‐target‐site resistance (NTSR) comprises a set of mechanisms conferring resistance to multiple modes of action. Investigation of the number of loci involved in NTSR will aid in the understanding of these resistance mechanisms. Therefore, six different multiple herbicide‐resistant Alopecurus myosuroides plants with different herbicide history were crossed in two generations with a susceptible wild type. Seeds from the backcrossing generation were studied for their segregation rate for resistance to five herbicides with four different modes of action (HRAC groups C₂, A, B and K₃). Taking into account that NTSR is a set of quantitative traits, the numbers of loci controlling NTSR were estimated using a normal mixture model fitted by the NLMIXED procedure of SAS. Each herbicide was controlled by a different number of loci comparing the six plants. In most of the cases, chlorotoluron resistance was controlled by one locus, whereas resistance to fenoxaprop‐P‐ethyl needed one or two loci. Resistance to pinoxaden was in all plants conferred by two loci. Cross‐resistance of fenoxaprop‐P‐ethyl and pinoxaden was found in all backcrossings, indicating that at least one of the two loci is responsible for both resistances. Resistance to mesosulfuron + iodosulfuron was conferred by a minimum of two loci. Results indicated that a minimum of five different loci can be involved in a multiple NTSR plant. Furthermore, the plant‐specific accumulation of NTSR loci was demonstrated. Such behaviour should be taken into account when evaluating the development and further spread of herbicide resistance.     

A Single Gene Cluster Controls Incompatibility and Partial Resistance to Various Melampsora larici-populina Races in Hybrid Poplars

ABSTRACT Complete cosegregation for race-specific incompatibility with three Melampsora larici-populina rust races was observed in five F(1) hybrid progenies of Populus, with different patterns among the various progenies. A single gene cluster could explain these segregations: one locus with multiple alleles or two tightly linked loci controlling complete resistance to E1 and E3, and two tightly linked loci for E2. The random amplified polymorphic DNA marker OPM03/04_480 was linked to that cluster in all families (<1 cM). This marker accounted for more than 70% of the genetic variation for field resistance in each family (heritability approximately 0.40). The same marker accounted for up to 64% of the clonal variation for growth in the nursery under natural inoculum pressure; the weak tolerance to rust of F(1) interspecific hybrids was attributed to a genetic background effect. Partial resistance was split into epidemiological components (heritability ranged from 0.35 to 0.87). Genotypic correlations among resistance traits for the different races were high (0.73 to 0.90). However, correlations among different resistance components for a single race were not all significant. A major quantitative trait locus for all components of partial resistance to E2 was associated to the cluster controlling incompatibility to E1 and E3 and marked by OPM03/04_480 (R(2)from 48 to 68%).     

Genetic control of the response of Erysiphe graminis f.sp. hordei to ethirimol and triadimenol

A study was conducted on the genetic control of the response of Erysiphe graminis f.sp. hordei , the causal agent of barley powdery mildew, to two fungicides: the hydroxypyrimidine ethirimol, and the triazole, sterol demethylation inhibitor triadimenol. In tests of responses to both fungicides, sets of progeny of various crosses were classified by principal components analysis into discrete resistant and sensitive classes. A single allele controlled the response to ethirimol of the resistant isolate DH14 in crosses with the sensitive isolates CC52 and CC138. The ethirimol-resistance alleles of DH14 and another resistant isolate. CC107, are at the same locus or are closely linked. Alleles at single loci controlled resistance and sensitivity to triadimenol in crosses of DH14 (sensitive) with CC107 (moderately resistant) and CC138 (highly resistant). There was no evidence for polygenic control of response to either fungicide. The ethirimol response locus and the two putative triadimenol response loci are designated Eth1 and Tdl1 and Tdl2 , respectively. There was no evidence for linkage of Eth1 and Tdl2 in the cross CC138 × DH14, in which responses to both fungicides segregated.     

在叶锈菌侵染过程中小麦过氧化物酶同工酶的变化

本文研究了抗叶锈性不同的小麦品种和毒力不同的叶锈菌小种相互作用过程中过氧化物酶(POD)同工酶谱的变化。结果表明:(1)不亲和组合在接种后48小时出现新酶带3,酶带4活性明显增强,120小时后又出现新酶带9。但亲和组合没有新酶带出现,而在接种后96小时酶带4活性也明显增强;(2)慢锈品种在接种后120小时以前,酶带4活性稍有增强,在120小时后其活性骤然增强。这可能是慢锈品种的特点。如果这些特点在更多的品种上得到验证,则POD同工酶谱分析有可能作为鉴定品种抗叶锈性和区分感病或慢锈品种的一种生理生化指标。