辽宁省稻曲病菌遗传多样性和致病力分析

为有效防治辽宁省稻曲病菌Ustilaginoidea virens,利用重复序列PCR(repetitive elementbased PCR,rep-PCR)分子指纹技术,对2017年自辽宁省8个市8个主产稻区采集的51株稻曲病菌菌株进行遗传多样性和致病力分析。结果显示,在3对引物中,以BOX1/BOX2和ERIC1/ERIC2为引物扩增的DNA指纹图谱的遗传多样性值分别为0.764、0.707,均大于0.7,故选择这2种引物扩增的DNA指纹图谱进行遗传多样性分析;当DNA指纹相似系数为0.78时,以BOX1/BOX2为引物和以ERIC1/ERIC2为引物扩增的DNA指纹图谱分别将供试菌株划分为12个和10个遗传类群;供试菌株致病力可划分为弱致病型、中等致病型和强致病型3个致病型,所占比例分别为33.33%、58.82%和7.85%,强致病型菌株仅在沈阳市、鞍山市和大连市出现;所有优势类群均包含3种致病型菌株。表明辽宁省稻曲病菌遗传结构复杂,不同地理来源的稻曲病菌菌株致病力存在一定差异,相同致病型的稻曲病菌菌株分属于不同的遗传类群,同一遗传类群中包含不同的致病型菌株。     

四川省籼稻区水稻稻曲病菌遗传多样性分析

应用ERIC-PCR指纹技术,对采自四川省6个籼稻自然生态区的60个稻曲病菌菌株进行了DNA分子水平上的遗传多样性研究,并进行UPGMA聚类分析和相似性分析.结果表明,所采用的ERIC引物在不同供试菌株中分别扩增出5～20条不等的带谱;在0.75相似水平上,供试菌株被划分为11个遗传型群,其中L1、L4、L5为优势群,并存在次要小型群和特异性型群;来自同一地区的稻曲病菌菌株具有较高的遗传相似性,而不同地区的稻曲病菌则表现出程度不同的变异,且寄主品种与稻曲病菌遗传差异之间的相关性较小.     

辽宁稻区稻曲病菌生物学特性及遗传多样性分析

为明确采自辽宁省12个稻区稻曲病菌Ustilaginoidea virens的生物学特性、群体遗传多样性与地理区域的关系。本研究采用生物学方法测定稻曲病菌菌株的生长速率和产孢能力,并采用SPSS 20.0分析软件对稻曲病菌菌株的菌丝生长速率和产孢能力进行相关性分析。提取稻曲病菌基因组DNA,采用特异性引物US、交配型引物MAT、遗传多样性引物ERIC对其进行PCR扩增,通过聚类分析进行群体遗传多样性研究。结果显示:157个稻曲病菌菌株的菌丝生长速率与产孢能力相关系数为0.19。采用稻曲病菌特异性引物US进行扩增,157个菌株均为稻曲病菌株;采用交配型引物MAT进行扩增,53个菌株为MATⅠ型,104个菌株为MATⅡ型。采用ERIC引物可扩增出2~7条不等的条带,157个稻曲病菌株被划分为10个基因类群,其中第1类群为优势类型,有44个菌株,占总数的28.0%;第2类群有20个菌株,占总数的12.7%;第3类群1个菌株,占总数的0.6%;第4类群1个菌株,占总数的0.6%;第5类群有21个菌株,占总数的13.4%;第6类群有17个菌株,占总数的10.8%;第7类群有2个菌株,占总数的1.2%;第8类群有5个菌株,占总数的3.2%;第9类群有30个菌株,占总数的19.1%;第10类群有16个菌株,占总数的10.2%。来自辽宁省12个稻区的157个稻曲病菌株菌丝生长速率与菌株产孢量之间没有相关性,产孢量与地域之间有相关性。基于ERIC-PCR扩增的稻曲病菌株基因组DNA指纹图谱的多态性进行划分的基因类群与地理区域之间有相关性,基因类群与生物学特性之间没有相关性。     

稻曲病菌生物学特性与致病力相关性研究

 测定了100个稻曲病菌(Ustilaginoidea virens)菌株的产孢能力、生长速率、色泽、胞外物质对麦种萌发抑制率和致病力,用SPSS软件分析了其产孢能力、生长速率、色泽、对麦种萌发抑制率与致病力的相关性。结果表明：菌株的产孢能力、色泽与致病力呈正相关;对麦种萌发抑制率与致病力的相关系数无统计学意义;菌株的生长速率与致病力呈负相关。其中,菌株的产孢能力和致病力呈中度相关;色泽、生长速率与致病力呈低度相关。     

稻曲病菌致病机制研究进展

 稻曲病是一种水稻穗部病害,在世界各水稻主产区均有发生,已成为世界性病害,在我国也属于水稻主要病害之一。稻曲病的发生严重影响水稻产量和稻米品质。研究其病原菌致病机制,对找寻防治药物的特异性靶标具有重要作用,可为抗病分子育种提供新的思路,为稻曲病菌防治药剂研发提供新的靶标。本文综述了稻曲病菌侵染过程、致病相关基因功能研究、水稻响应稻曲病菌侵染等方面的研究进展,提出进一步研究的策略。现有研究表明,稻曲病菌能成功侵染水稻孕穗期雄蕊的花丝引起发病。病原菌可通过抑制多个水稻免疫途径实现成功侵染;还可模仿胚珠受精激活水稻灌浆、糖代谢等途径为稻曲球的形成提供营养物质;全基因组测序完成和基因编辑技术在基因敲除中的成功应用,为稻曲病菌功能基因研究提供了很好的基础。稻曲病菌独特的侵染过程和营养利用机制是未来研究的重要方向。     

稻曲病菌致病机制研究进展

 稻曲病是一种水稻穗部病害,在世界各水稻主产区均有发生,已成为世界性病害,在我国也属于水稻主要病害之一。稻曲病的发生严重影响水稻产量和稻米品质。研究其病原菌致病机制,对找寻防治药物的特异性靶标具有重要作用,可为抗病分子育种提供新的思路,为稻曲病菌防治药剂研发提供新的靶标。本文综述了稻曲病菌侵染过程、致病相关基因功能研究、水稻响应稻曲病菌侵染等方面的研究进展,提出进一步研究的策略。现有研究表明,稻曲病菌能成功侵染水稻孕穗期雄蕊的花丝引起发病。病原菌可通过抑制多个水稻免疫途径实现成功侵染;还可模仿胚珠受精激活水稻灌浆、糖代谢等途径为稻曲球的形成提供营养物质;全基因组测序完成和基因编辑技术在基因敲除中的成功应用,为稻曲病菌功能基因研究提供了很好的基础。稻曲病菌独特的侵染过程和营养利用机制是未来研究的重要方向。     

中国部分地区稻曲病菌培养特性及其遗传多样性分析

采用生物学方法和RAPD-PCR技术,对来自11个省(市)84个菌株的菌丝生长速率、分生孢子数量、孢子萌发率及其遗传多样性进行分析,以明确中国部分地区稻曲病菌株的培养特性和遗传多样性。依据菌丝生长速率,菌株可被划分为快和慢2种类型,分别占58.33%和41.67%。依据产孢力和分生孢子萌发力,菌株可划分为强、中和弱3种类型。采用12条RAPD引物共扩增出323条带,多态性条带比率为98.14%,遗传距离变化范围为0.02～1.00。在遗传距离0.725水平上,所有菌株被划分成7个遗传聚类组,聚类组R4和R5为优势聚类组,并存在一些亚组。不同地区之间和同一地区内的菌株表现出不同程度的变异,内陆地区的菌株群体变异程度明显高于沿海地区。从采用相同接种体接种的水稻品种上分离的稻曲病菌具有紧密的亲缘关系。     

稻曲病菌分生孢子的生物学研究

 本文对稻曲病菌分生孢子的一些生物学特性进行了研究,结果表明,基质养分对分生孢子萌发影响较大,纯水不利于孢子萌发,PSA最适于孢子萌发,葡萄糖则强烈抑制孢子萌发,马铃薯煮汁既可抵消葡萄糖的抑制作用,又可刺激孢子萌发。分生孢子在琼脂面上比在液滴中萌发率高。分生孢子萌发的适宜温度为22~31℃,以28℃最好。分生孢子萌发对pH值敏感,以pH 6~7最适宜。用振荡培养法获取分生孢子,培养10 d后,孢子的萌发力开始下降。分生孢子的存活对水的依赖性强,在水中保存8 d萌发力不变,在100% RH中8 d萌发力略有降低,而在25% RH中5 h萌发力即迅速下降。根据这些特性,作者对分生孢子在田间的动态作了一些推测。     

稻曲病菌的病原学

 稻曲病菌产生的黄色和黑色厚垣孢子表面都有疣状突起,但黑色厚垣孢子的疣状突起明显。常规萌发试验,黄色厚垣孢子萌发,黑色不萌发。黄色厚垣孢子在4℃和25℃下贮存分别保持萌发能力1年和80d左右,萌发的最适pH值和最适温度分别为pH5~pH8和25℃~30℃。光照可诱导产生厚垣孢子。病菌在PD和PS培养液中可产生大量分生孢子。田间采集的稻曲病菌菌核在砂培条件下可以萌发并可产生成熟的子囊孢子。该菌可产生毒素,在一定的浓度下,对稻种萌发和胚芽生长有明显抑制作用。     

Genetic diversity and population structure analysis of isolates of the rice false smut pathogen Ustilaginoidea virens in India

Genetic diversity assessment and population structure analysis are essential for characterization of pathogens and their isolates. Markers are essential tools for exploring genetic variation among the isolates. False smut of rice caused by Ustilaginoidea virens, formerly Villosiclava virens, is a major emerging disease of rice in India. A high level of variability is observed at the field level, but no information is available from India on genetic diversity and population structure. This is the first report of genetic diversity and population structure of U. virens from India that included 63 isolates distributed across the vast geographical area of eastern and north-eastern India (18.9 to 26.7°N and 82.6 to 94.2°E). Seventeen RAPDs and 14 SSRs were identified as polymorphic and a total of 140 alleles were detected across the populations. The average number of alleles per locus for each primer was 4.5. All the isolates were grouped into two major clusters, with partial geographical segregation that was supported by principal coordinate analysis. Mantel test suggested genetic distance within the isolates increased with increasing geographical distance. Analysis of molecular variation showed more genetic variation within populations and less among populations. This outcome will help in understanding genetic diversity of U. virens from eastern and north-eastern India and in planning effective management strategies.     

稻曲病菌遗传多样性与群体结构的初步分析

 利用随机扩增多态性DNA (random amplified polymorphic DNA,RAPD)初步分析了稻曲病菌(Ustilaginoidea virens)的群体遗传结构。从1 60个随机引物中筛选32个扩增带型清晰、重复性好的引物,对不同年份采自辽宁、云南、湖北和浙江等水稻种植区的5 6个菌株进行扩增。32个引物扩增出2 2 3条带,绝大多数引物对不同年度采自不同稻区的菌株扩增的DNA谱型相同,大多数菌株间相似性系数达0.80以上。根据扩增DNA片段的多态性,从空间分布来看,来源于北方、长江流域和南方的菌株难以划分出明显的地理宗谱;不同年度的菌株DNA多态性也无明显的差异。上述结果初步表明稻曲病菌遗传稳定,寄主选择作用(寄主的基因型及其时空分布)对稻曲病菌变异的影响较小。但是尚需采用其它的分子技术测试更多的菌系,才能较系统地分析我国稻曲病菌系的遗传变异及群体结构特点。     

Epiphytic colonization of Ustilaginoidea virens on biotic and abiotic surfaces implies the widespread presence of primary inoculum for rice false smut disease

Ustilaginoidea virens (Uv), the causative agent of rice false smut disease, infects developing rice spikelets at the booting stage, and transforms individual grains of the panicle into smut balls. Epidemics of the disease occur when the rice booting and heading stages coincide with rainy days. Using a green fluorescent protein (GFP)‐labelled Uv isolate that can form false smut balls on rice panicles, it was found that under high humidity and free water conditions the Uv isolate could colonize leaves of plants belonging to various families including the Poaceae (Oryza sativa, Echinochloa crusgalli, Digitaria sanguinalis and Leptochloa chinensis), the Brassicaceae (Arabidopsis thaliana) and the Solanaceae (Nicotiana benthamiana) without symptoms. Over several days, some conidia could germinate on the leaves of these plants and in water on the surface of Parafilm and cellophane, form hyphae and differentiate conidiophores to generate a large number of secondary conidia, while other conidia were able to directly produce secondary conidia. Conversely, in the absence of water some conidia could either bud to form new conidia or were converted into chlamydospores. These data indicate that Uv is one of a few fungal pathogens reported to have epiphytic characteristics. The rapid generation of a large number of spores on biotic and abiotic surfaces greatly increases the inoculum that can infect rice spikelets, resulting in the occurrence of rice false smut disease epidemics. These findings are important in the development of disease control strategies.     

水稻品种构成对稻曲病菌遗传结构影响的初步研究

由稻曲病菌（Ustilaginoidea virens）引起的病害是当前限制我国水稻优质高产的重要病害之一。家畜食用被稻曲病菌厚垣孢子污染的谷物可导致中毒。近20年来，稻曲病防治主要以喷施化学农药为主。研究品种布局对稻曲病菌群体结构的影响，有助于了解病害的发生规律及提出有效的病害防治策略。本文初步分析了稻曲病菌群体结构与水稻品种构成的关系。     

3种防御酶在水稻抗稻曲病中的活性变化

不同水稻品种对稻曲病抗性存在明显差异。本研究选用12个不同抗、感病水稻品种,在抽穗前7~10 d,用注射法接种稻曲病菌分生孢子悬浮液。结果表明,在抗病品种中苯丙氨酸解氨酶（PAL）、过氧化物酶（POD）和多酚氧化酶（PPO）3种酶活性接种后迅速提高,变化幅度较大;而感病品种的酶活性在接种后缓慢增加,变化幅度小。由此说明,这3种防御酶与水稻抗稻曲病有一定的关系。     

稻曲病菌不同颜色厚垣孢子超微结构比较

 The surface and internal structure difference of yellow and black dormant chlamydospores of Ustilaginoidea virens on mid-and late-season rice were examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). SEM revealed that the spherical yellow and black chlamydospores had prominent spines, with the diameters from 3 to 6 μm and 4 to 6 μm, respectively. The spines were 200-700 nm and 400-1 100 nm long in yellow and black chlamydospores, respectively. Variance analysis showed that there was no difference in diameter of the yellow chlamydospores but significant difference among the black ones collected from both rice cropping types. Similarly, there was no difference in length of the spines on the same color chlamydospores but extremely different between the yellow and black chlamydospores collected from the two cropping types. TEM revealed that the cell wall of both color chlamydospores were composed of two layers, the thick endosporium and thin exosporium, and the thickness of the former layer of the black chlamydospores was nearly two times as that of the yellow ones. The organelles in yellow chlamydospores such as the nucleus were obviously visible, but invisible in black ones. The cytoplasm of black chlamydospores was occupied by a big lipid globule. This study suggests that the formation of dormant chlamydospores of U. virens is related with the structural changes in the cell.     

中国不同地理来源稻曲病菌rDNA-IGS的初步分析

 The rDNA-IGS of 11 Ustilaginoidea virens isolates from different geographical regions in China were amplified by PCR and sequenced. The results showed that the rDNA-IGS fragments consisted of one cen-tral variable region and two lateral conservative regions. The two conservative sequences among the11 isolates shared the sequence similarity of 99.44%. The variable region consisted of 2, 4, or 6 of 77 bp-length-repeats in the isolates and the sequences of the repeat units were high conservative. Primers UIGS Ⅲ and UIGS IV were designed for amplification of the variable region in the rDNA-IGS.