Carbon Catabolite Repressor UvCreA is Required for Development and Pathogenicity in Ustilaginoidea virens

The rice false smut disease, caused by Ustilaginoidea virens, has emerged as a significantglobal threat to rice production. The mechanism of carbon catabolite repression plays a crucial role in theefficient utilization of carbon nutrients and enzyme regulation in the presence of complex nutritionalconditions. Although significant progress has been made in understanding carbon catabolite repression infungi such as Aspergillus nidulans and Magnaporthe oryzae, its role in U. virens remains unclear. Toaddress this knowledge gap, we identified UvCreA, a pivotal component of carbon catabolite repression,in U. virens. Our investigation revealed that UvCreA localized to the nucleus. Deletion of UvCreA resultedin decreased growth and pathogenicity in U. virens. Through RNA-seq analysis, it was found that theknockout of UvCreA led to the up-regulation of 514 genes and down-regulation of 640 genes. Moreover,UvCreA was found to be involved in the transcriptional regulation of pathogenic genes and genesassociated with carbon metabolism in U. virens. In summary, our findings indicated that UvCreA isimportant in fungal development, virulence, and the utilization of carbon sources through transcriptionalregulation, thus making it a critical element of carbon catabolite repression.

Survey and examination of the potential alternative hosts of Villosiclava virens, the pathogen of rice false smut, in China

Rice false smut is caused by ascomycete Villosiclava virens, whose potential alternative hosts have been assumed previ- ously. Here its potential alternative hosts were surveyed and identified from 2008 to 2013 in the main rice-growing regions in China. Two common weeds in paddy fields, Digitaria sanguinalis Scop. and Echinochloa crusgalli （L.） Beauv., were found to present the similar symptoms to smut diseases in a few individuals in 2012 and 2013 in Zhejiang and Sichuan provinces of China, respectively. After the examinations of the spore morphology, their infection and extension mode in hosts, pathogen cell wall components, and molecular identification, the two pathogens were identified to be the Basidiomycetes, Ustilago syntherismae and Ustilago trichophora, respectively. So far there has been no alternative host of V. virens to be identified in China. These suggest that the alternative hosts of V. virens, if they do exist, are not possible to play an important role in the pathogen life cycle and the disease epidemics.

Rapid detection of the rice false smut fungus Ustilaginoidea virens by lateral flow strip-based recombinase polymerase amplification assay

Rice false smut,caused by Ustilaginoidea virens,is a devastating disease that greatly reduces rice yield and quality.However,controlling rice false smut disease is challenging due to the unique infection mode of U.virens.Therefore,there is a need for early diagnosis and monitoring techniques to prevent the spread of this disease.Lateral flow strip-based recombinase polymerase amplification(LF-RPA)overcomes the limitations of current U.virens detection technologies,which are time-consuming,require delicate equipment,and have a high false-positive rate.In this study,we used a comparative genomics approach to identify Uv_3611,a specific gene of U.virens,as the target for the LF-RPA assay.The designed primers and probe efffectively detected the genomic DNA(gDNA)of U.virens and demonstrated no cross-reactivity with related pathogens.Under optimal conditions,the LF-RPA assay demonstrated a sensitivity of 10 pg of U.virens gDNA.Additionally,by incorporating a simplified PEG-NaOH method for plant DNA extraction,the LF-RPA assay enabled the detection of U.virens in rice spikelets within 30 min,without the need for specialized equipment.Furthermore,the LF-RPA assay successfully detected U.virens in naturally infected rice and seed samples in the field.Therefore,the LF-RPA assay is sensitive,efficient,and convenient,and could be developed as a kit for monitoring rice false smut disease in the field.

Ustilaginoidea virens: A Fungus Infects Rice Flower and Threats World Rice Production

Rice false smut disease, which is caused by the fungus Ustilaginoidea virens, is currently one of the most devastating rice fungal diseases in the world. Rice false smut disease not only causes severe yield loss and grain quality reduction, but also threatens food safety due to its production of mycotoxins. In this review, the most recent progresses regarding the life cycle, infection processes, genome and genetic diversity, pathogenic gene and disease resistance in rice were summarized in order to provide theoretical basis for the control of U. virens. We also proposed some future directions and key questions that need to be addressed for a better understanding of the molecular mechanism that leads to rice false smut disease and the prospects for sustainable control of rice false smut.

Biological Activity of Several Fungicides against Ustilaginoidea virens

[Objective] This study aims to screen for the high effective fungicides which could significantly decrease the disease incidence and disease index of rice false smut. [Method] The inhibitory activities of the fungicide against mycelial growth of Ustilaginoidea virens were measured to in vitro evaluate the ECho values. And 17 fungicides were sprayed to evaluate the efficacy and effect of the fungicides tested in the field trials on the rice characters, [Result] The results showed that epoxicona- zole, difenoconazole, propiconazole and procloraz exhibited high inhibitory activity against mycelial growth of Ustilaginoidea virens with the ECso values 0.04, 0.07, 0.12 and 0.11 pg/ml, respectively. The results of field trials showed that the efficacy of Wen- quning, and fungicides such as difenoconazole, prochloraz, propiconazole, epoxi- conazole and their mixtures in controlling rice false smut were all 70% or more. [Conclusion] The 17 tested fungicides behaved efficacy in controlling rice false smut and did not cause drug injury on leaves and grains of rice plants, sprayed when flag leaves of rice fully expanded.

UvWhi2 Is Required for Stress Response and Pathogenicity in Ustilaginoidea virens

Some stress response-related genes have been identified in Ustilaginoidea virens,but it is not clear whether and how defects of stress responses affect the pathogenesis processes of U.virens.In this study,we identified a general stress response factor UvWHI2 as a homolog of Saccharomyces cerevisiae Whi2 in U.virens.The relative expression level of Uv Whi2 was significantly up-regulated during infection,suggesting that UvWHI2 may be involved in pathogenesis.Furthermore,knockout of Uv Whi2 showed decreased mycelial growth,increased conidiation in the potato sucrose medium and a defect in pathogenicity.In addition,the RNA-Seq and phenotypic analysis showed that UvWHI2 was involved in response to oxidative,hyperosmotic,cell wall stress and nutrient limitation.Further studies revealed that the defects of stress responses of the?Uvwhi2 mutant affected the formation of secondary spores on the nutrient limited surface and the rice surface,resulting in a significant reduction of pathogenicity of U.virens.Our results suggest that UvWHI2 is necessary for fungal growth,stress responses and the formation of secondary spores in U.virens.In addition,the defects of stress responses can affect the formation of secondary spores on the rice surface,and then compromise the pathogenicity of U.virens.

Effect of temperature on the development of sclerotia in Villosiclava virens

The sclerotia of Villosiclava virens were found commonly in high altitude and the temperate regions, where the temperatures are relatively low in rice filling stage. To make sure if low temperature induce the sclerotial formation in V. virens, the inoculated rice panicles in laboratory and the diseased rice panicles cut from paddy fields were treated under different temperatures. The results showed that 3 days of night temperature at 15°C were enough to induce the sclerotial formation. The low temperature was much more effective for young balls with intact membranes. After appearance of chlamydospores on the ball surfaces, the sclerotium could not differentiate anymore. The sclerotia began to differentiate below the chlamydospore layer and gradually grew onto the ball surfaces. This suggests that low temperature in the early development stage of false smut balls is an important factor to induce the sclerotial differentiation, and rice cultivars with long growth periods are able to produce more sclerotium-bearing balls, which will produce mass of spores in paddy field in the coming year.

Putative Phosphatase UvPsr1 Is Required for Mycelial Growth, Conidiation, Stress Response and Pathogenicity in Ustilaginonidea virens

Ustilaginoidea virens is the causal agent of rice false smut,which can be a highly destructive disease of rice.The plasma membrane phosphatase Psr1 proteins,which act as a regulator of the salinity stress response in yeast,are widely distributed across fungi,but their functional characterization is sketchy.In this study,we characterized the functions of Psr1 protein,UvPsr1,in U.virens.Analyses of the AUvpsr1 and its complementation strain showed that UvPsr1 is required for normal mycelial growth,conidiation and tolerance to oxidative,osmotic and cell wall stresses.When rice panicles were inoculated with the AUvpsr1 strains,no symptoms of false smut disease developed,showing that UvPSR1 also contributes to the pathogenicity of the fungus.The deletion mutant of UvPSR1 also appeared to produce a smaller titer of toxic compounds able to inhibit elongation of the germinated seeds.In conclusion,our results indicated that UvPsr1 is a new pathogenic factor of U.virens.

水稻稻曲病研究进展

稻曲病是水稻后期发生的一种真菌性病害,在中国及世界各地均有发生,现已成为水稻的主要病害之一。近年来有大量学者致力于水稻抗稻曲病研究,并取得了一定的进展。对水稻稻曲病的分布及危害、稻曲菌的生物学特征、稻曲菌的侵染机制、水稻与稻曲菌之间的相互作用以及水稻稻曲病的分子生物学研究等方面进行了综述,为育种家培育抗稻曲病水稻品种提供借鉴。

丁香酚和香芹酚对稻曲病菌的抑菌作用及田间防效评价

基于天然成分开发防治水稻稻曲病的新型抑菌剂,本文选取了丁香酚和香芹酚两种两种功能丰富、来源广泛的天然植物源萜类化合物,通过浓度梯度试验测定发现,两者均能显著抑制稻曲病菌菌丝生长、分生孢子萌发,并呈现剂量依赖效应。0.2mmol/L的丁香酚和香芹酚对稻曲病菌的菌丝生长抑制率分别为12.53%和44.39%,分生孢子萌发抑制率分别为94.02%和72.24%。扫描电子显微镜观察结果表明,两者均能破坏稻曲病菌菌丝的完整性。田间药效试验中两者均有效抑制水稻稻曲病发生,减少稻谷中稻曲菌素A积累。本研究应用丁香酚和香芹酚防治稻曲病和防控稻曲菌素A,初步解析了两者抑菌防毒机理,展现了两者具有的广阔应用前景。

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 2 2～ 31℃ ,以 2 8℃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 10d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8d萌发力不变 ,在 10 0 %RH中 8d萌发力略有降低 ,而在 2 5 %RH中 5h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 ( Ustilaginoideavirens)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 。

采用AFLP技术分析稻曲病菌的遗传多样性Ⅰ:同一块稻田中稻曲病菌的遗传结构

采用AFLP(Am plified fragm ent length po lym orphism)技术分析了来自北京昌平同一块稻田中不同水稻品种和育种中间材料上稻曲病菌(U stilag ino idea virens)的遗传多样性。从256对E coRⅠ和M seⅠ引物中选择30对扩增40个菌株。结果表明,同一块稻田中稻曲病菌菌株间的相似性系数达0.72以上,来自同一小区的多数菌株能聚成亚类;发现从同一水稻品种分离的菌株没有特异性的AFLP谱带。初步推断稻曲病菌与水稻品种不存在明显的专化性互作。

水稻稻曲病的药效试验和防治适期的选定

于水稻破口前3 d施用8种药剂防治稻曲病,结果表明,爱苗、万兴、好力克、井冈霉素、HB20-1、纹曲宁、三唑酮对稻曲病的防效分别达67.53%、62.15%、61.75%、57.52%、52.11%、48.35%和29.68%,其每公顷理论和实际增产幅度分别达1.71%-19.62%和3.85%-15.03%。于水稻孕穗期、破口前3 d、齐穗期、扬花期施用爱苗和井冈霉素防治稻曲病,结果表明,随着生育期推迟,对稻曲病的防治效果及产量均呈下降趋势,且以孕穗期施药防效最佳,增产最高。爱苗、井冈霉素孕穗期施药防效分别达到67.03%、58.54%,每公顷理论产量分别比不施药处理增产11.89%和10.68%,每公顷实际产量分别比不施药处理增产了11.31%和9.34%。

水稻对稻曲病抗性的分级及相应级别的产量损失

对24个水稻品种(系)与稻曲病抗性有关的3个主要组分(病穗率、平均每穗病粒数、每千粒含病粒数)进行主成分分析;用线性回归法分析衡阳具代表性的14个水稻品种产量损失率与平均每穗病粒数的关系;以中选组分用欧氏距离和最长聚类法对101个品种进行聚类。结果表明:平均每穗病粒数为主要抗性组分;14个品种的产量损失率与平均每穗病粒数的回归方程为y=2.8624x+0.5427(P=0.0001);用平均每穗病粒数组分,当阈值T=0.4000,将101个水稻品种聚为6类,依6类品种间的平均每穗病粒数值,并参照国际水稻病害分级标准,将水稻品种对稻曲病的抗性划分为6级,并将各级平均每穗病粒数值代入回归方程得出其产量损失率,即各级平均每穗病粒数和产量损失率为:0级,0.000粒,产量损失率0,高度抗病;1级,0.001～0.100粒,产量损失率0.546%～0.829%,抗病;3级,0.101～0.410粒,产量损失率0.830%～1.716%,中度抗病;5级,0.411～1.010粒,产量损失率1.717%～3.434%,中度感病;7级,1.011～1.760粒,产量损失率3.435%～5.581%,感病;9级,每穗病粒数≥1.761粒,产量损失率≥5.582%,高度感病。

基于稻曲球结构分析稻曲菌的侵染部位和侵染时期

对田间采集的稻曲病病粒进行解剖分析,结果显示厚垣孢子层未形成的稻曲病病粒可分离出完整雌雄蕊;典型的稻曲球均可解剖出完整子房,且子房绿色未见明显膨大。雄性不育的水稻品系受稻曲病为害产生典型症状,其解剖结果与正常水稻品系相同。由此推测稻曲菌侵染水稻雌蕊发生在水稻开花之前,并直接从子房获得营养形成稻曲球。

稻曲病对水稻产量的影响及水稻新品种抗病性测定

在田间调查稻曲病病粒数对水稻秕谷率和产量损失率的影响,同时采用自然诱发方法测定62个水稻新品种对稻曲病的抗性。结果表明,单穗稻曲球病粒数的增加能引起水稻秕谷率和产量损失率的增加;当单穗病粒数在5粒以上时,引起产量损失率可达36%以上;62个水稻品种中表现中抗稻曲病的品种有7个,占供试品种数量的11.29%,表现中感稻曲病的品种有22个,感稻曲病的品种有33个,分别占35.48%和53.23%。

绿木霉菌株TY009防治纹枯病等水稻主要真菌病害的潜力

【目的】研究绿木霉(Trichoderma virens)菌株TY009对水稻纹枯病、稻瘟病和稻曲病菌作用效果,以及其代谢产物胶霉毒素在不同温度和pH条件下的活性稳定性,评价菌株TY009防治水稻纹枯病和其它水稻主要真菌病害的生防潜力。【方法】用色谱法从绿木霉菌株TY009培养液中分离、纯化出胶霉毒素;采用对峙培养方法测定菌株TY009对水稻纹枯病菌的拮抗作用;采用常规抑制方法测定其代谢产物胶霉毒素对水稻纹枯病菌菌丝生长、菌核萌发的抑制作用,以及对稻瘟病菌孢子萌发、附着胞形成和稻曲病菌孢子萌发和次生孢子形成的影响;在不同温度、pH条件下测定胶霉毒素生物活性的变化。【结果】菌株TY009对水稻纹枯病菌有显著的拮抗作用。150μg·mL-1胶霉毒素完全抑制水稻纹枯病菌的菌丝生长;160μg·mL-1胶霉毒素处理菌核4h后,完全抑制了水稻纹枯病菌菌核的萌发;50和10μg·mL-1胶霉毒素分别完全抑制稻瘟病菌分生孢子萌发和附着胞的形成;1.0μg·mL-1胶霉毒素完全抑制稻曲病菌分生孢子萌发和次生孢子的形成。胶霉毒素在100℃处理60min后,其对水稻纹枯病菌菌丝的生物活性没有改变;在pH值大于6.0时,0.5μg·mL-1胶霉毒素对稻曲病分生孢子没有明显的生物活性,但低的pH孢子悬浮液中,胶霉毒素显著抑制分生孢子萌发和次生孢子的形成。【结论】绿木霉(T.virens)菌株TY009拥有多种生防机制,是一个潜在的、有效的生防菌株,具有应用防治水稻纹枯病并兼防稻瘟病和稻曲病的潜力。

福建省水稻新品种对稻曲病和穗瘟病的抗性评价

采用室内人工注射接种法和田间自然诱发法评价福建省15个水稻新品种对稻曲病和穗瘟病的抗性。水稻品种对稻曲病的室内抗性评价结果表明,有3个水稻品种对稻曲病表现为中抗,如FJZD10-5、FJZD10-8,有4个品种表现为中感,8个品种表现为感病。水稻品种对稻曲病的田间抗性评价结果表明,有11个品种表现为中抗,如FJZD10-2、FJZD10-3,有4个品种表现为中感。水稻品种对穗瘟病的田间抗性评价结果表明,有7个品种表现为中抗,如FJZD10-6、FJZD10-7,4个品种表现为中感,4个品种表现为感病。在建阳、宁化和福安3个田间试验点,同时发生稻曲病和穗瘟病的水稻品种出现的频率分别为100%、73.33%和73.33%。未出现对稻曲病和稻瘟病均表现为抗病的水稻品种。

抗稻曲病水稻品种与材料的筛选与评价

通过田间分期播种自然诱发和温室人工接种试验,对湖北省主栽品种及高代育种材料进行了抗稻曲病鉴定,共筛选出对稻曲病免疫的材料4份,为稻曲病的抗病育种奠定了基础。