稻区杂草上的梨孢菌与稻瘟病发生的关系

 新发现的稻曲病菌白化菌株其分生孢子座白色,厚垣孢子表面光滑,大小为3.5-6.0×3.5-6.8μm。在麦芽糖-稻芽浸渍液中可产生较多的分生孢子。pH值为6时生长最好。普通培养基中加入几种供试碳源和氮源均有利于菌落生长,以蔗糖为碳源,浓度为2%或以L-天门冬酰胺为氮源浓度为0.2%~0.3%时时其菌落生长有促进作用。培养过程中可产生对稻种萌发有抑制作用的毒素,而固体培养时产毒能力高于液体培养。白化菌株回接到水稻上可重新产生白色稻曲球。该菌和稻曲病菌的酯酶同功酶和RAPD扩增谱带明显不同。可见白化菌株具有独立于稻曲病菌的新的分类地位。     

解淀粉芽孢杆菌(Bacillus amyloliquefaciens)HAB-7的培养基优化及抑菌活性

研究了解解淀粉芽孢杆菌(Bacillus amyloliquefaciens)HAB-7菌株在不同培养基下生长速率及产生的活性物质对几种植物病原菌的抑菌活性。采用摇瓶法测定了HAB-7耐盐性及生长速率;平板对峙法测定了其抑菌活性;以NA培养基为基础培养基,芒果炭疽菌(Colletotrichum gloeosporioides)为靶标菌,研究了不同碳、氮源组合培养基对HAB-7菌株生长影响,以及对芒果炭疽菌的抑制率;用单因子正交试验,进一步优化碳源、氮源组合培养基。结果表明,HAB-7菌株在0~20%NaCl(g/L)含盐量上均能生长;最佳接种时间应在培养24~28 h;在NA、LB、BPY、AYDA、BPA及牛肉膏蛋白胨培养基上培养的HAB-7,对8种植物病原真菌平均抑制率分别为53.23%、52.03%、55.52%、52.77%、56.58%及53.82%。优化后的最佳碳源为蔗糖(10.0 g/L)和葡萄糖(10.0 g/L)复合碳源;最佳的氮源为酵母粉(20.0 g/L)和胰蛋白胨(20.0 g/L)复合氮源。单因子正交试验结果显示的碳、氮源为蔗糖30.0 g/L、葡萄糖10.0 g/L、酵母粉30.0 g/L、胰蛋白胨40.0 g/L;对芒果炭疽菌抑菌率为78.3%,较BPA培养的HAB-7提高了10.80%,活菌数达到1.97×10~(10)cfu/m L。     

稻曲病菌无性孢子萌发及在不同培养基上生长特性

采用不同的培养基研究稻曲病菌孢子萌发及病菌生长特性.结果表明,稻曲病菌厚垣孢子和分生孢子在蒸馏水和2%蔗糖溶液中3h不能萌发;6h时分生孢子、厚垣孢子分别萌发产生次生分生孢子和分生孢子;新采集的黄色和黑色稻曲球上的厚垣孢子在2%蔗糖溶液中的萌发率均高于其在蒸馏水中的萌发率.供试的9种固体培养基中,以马铃薯蔗糖琼脂培养基(PSA)最适于病原菌生长.生长速率为2.54 mm/天;病菌在7种培养液中振荡培养7天后,在马铃薯葡萄糖培养液(PDB)中菌丝干重最大,达24.5mg/mL,在大米汁和马铃薯蔗糖培养液(PSB)中产生的分生孢子量最多,孢子浓度分别为2.63×107和2.28×107个/mL;而固、液态的Czapek、稻秆汁和胁本哲氏培养基均不适宜于稻曲病菌的培养.     

培养基碳源和氮源对中国被毛孢菌落生长的影响

中国被毛孢(Hirsutella sinensis)是冬虫夏草真正的无性型,其生长需要有碳源的存在,没有碳源存在菌落生长缓慢甚至不生长;其菌丝生长对氮源具有选择性,不适合的氮源也会抑制菌丝的生长。本研究通过改变培养基碳源和氮源,得到了最适合中国被毛孢生长的碳、氮源。结果表明,在蔗糖为碳源的培养基上生长最佳,且菌落生长状况较接近菌落的典型特征。水解蛋白胨是较理想的氮源,菌落生长最快,且生长状况最接近菌落的典型特征。不同碳、氮源培养基上,中国被毛孢菌落生长状况差异较大,对蝙蝠蛾幼虫的侵染能力的差异还有待于研究。     

青海省樱桃叶斑病菌生物学特性及室内药剂毒力测定

为明确青海省樱桃叶斑病菌链格孢Alternaria alternata的生物学特性及筛选出对其防效最佳的药剂,本研究采用菌丝生长速率法和孢子计数法研究了不同培养条件对该病原菌菌丝生长和产孢量的影响,并用菌丝生长抑制率法对9种杀菌剂进行室内毒力测定。结果表明,菌丝生长和产孢的最适培养基分别为马铃薯蔗糖琼脂(PSA)培养基和马铃薯葡萄糖琼脂(PDA)培养基,最适温度为25℃, 最适氮源为蛋白胨,最适碳源分别为肌醇和乳糖,最适pH分别为6~8和6~7,24 h持续黑暗处理更有利于菌丝生长和产孢。9种化学药剂对病原菌均有不同程度的抑制作用,其中250 g/L嘧菌酯悬浮剂抑菌效果最好,EC50为1.258 mg/L。该研究结果可为青海省樱桃叶斑病的防治提供理论依据。     

高粱链格孢叶斑病菌生物学特性研究

为明确高粱链格孢叶斑病菌Alternaria alternata的生物学特性, 本文采用生长速率法研究了不同营养及环境条件对该病菌生长的影响。试验结果表明:不同培养基上菌丝生长有明显差异, 其中马铃薯蔗糖琼脂培养基(PSA)最适合菌丝生长和孢子产生; 不同碳、氮源条件下菌丝的生长有显著差异, 其中最适宜菌丝生长碳源为葡萄糖, 氮源是蛋白胨; 该病原菌最适宜营养生长的温度为28℃, pH为7; 菌丝的致死温度为47℃; 光照时间对高粱叶斑病菌丝的营养生长没有影响。     

甜瓜黑点根腐病菌生物学特性及其对药剂敏感性测定

为明确甜瓜黑点根腐病的发生和流行规律及有效防治方法,在不同培养基、碳源、氮源、温度、pH等培养条件下,测定甜瓜黑点根腐病菌的生物学特性及其对多种瓜类作物致病性,并采用菌丝生长速率法测定菌株对不同药剂的敏感性。结果表明,甜瓜黑点根腐病菌MC8菌丝生长的适宜培养基为马铃薯蔗糖培养基、燕麦粉琼脂和马铃薯葡萄糖琼脂培养基,马铃薯蔗糖培养基、燕麦粉琼脂和玉米琼脂粉这3种培养基可产生子囊壳;碳源和氮源对菌丝生长无明显促进作用,但乳糖可显著促进子囊壳产生,该发现为国内外首次报道;MC8菌丝生长的适宜pH为5～7,菌丝和子囊壳生长最适温度为30℃。致病力试验结果显示MC8菌株可侵染西瓜、甜瓜、南瓜和葫芦多种瓜类作物。咪酰胺和嘧菌酯在PDA上对瓜类黑点根腐病菌的菌丝生长有较好的抑制效果。     

寒地水稻秆腐菌核病菌的生物学研究

研究结果表明,水稻秆腐菌核病菌可利用多种碳源和氮源,碳源以可溶性淀粉最好,氮源以酒石酸铵最好,混合碳源(蔗糖+鼠李糖)对病菌菌丝生长有促进作用,比单独蔗糖和鼠李糖生长的好;病菌能在多种天然培养基上生长并形成菌核;菌丝在碳氮比为60∶1的培养基上生长最好;生长物质VB₆+肌醇、VB₆+谷氨酰胺等11个组合对菌丝生长有促进作用;微量元素中亚铁、锰和硼对菌丝生长有抑制作用;液体培养中以Czapek培养基生长最好,但不利于菌核的形成;菌核在15～35℃均可萌发,最适温度为30℃;菌核在0.05%葡萄糖溶液、蒸馏水、水琼脂上均可萌发,以0.05%葡萄糖溶液萌发最好。     

主要环境及营养因素对西瓜枯萎病菌生长的影响

采用菌丝生长速率法和产孢量测定研究了不同温度、光照、pH及碳氮源对来源于海南、河北和上海的3株西瓜枯萎病菌Fo-HN-46、Fo-HB-12和Fo-SH-1生长速率和产孢量的影响。结果表明:西瓜枯萎病菌菌株的适宜生长温度为20~30℃,Fo-HN-46最适生长温度为28℃,Fo-HB-12和Fo-SH-1最适生长温度为25℃,产孢的最适温度为28~30℃;光照对西瓜枯萎病菌生长速率无显著影响,对产孢影响显著,Fo-HN-46在黑暗条件下单位面积产孢量最高,Fo-HB-12和Fo-SH-1在半光照条件下产孢量最高;pH对西瓜枯萎病菌菌株的生长速率和产孢量有显著影响,pH 7~9时菌丝生长速率快,pH 8~11时产孢量高;葡萄糖、淀粉和乳糖作为碳源能够显著增加西瓜枯萎病菌的产孢量;蛋白胨作为氮源能显著促进菌丝生长,硝酸钠、酵母粉和蛋白胨均能显著增加西瓜枯萎病菌的产孢量。用最适碳源、氮源培养,Fo-HN-46生长速率大于Fo-HB-12和Fo-SH-1。Fo-SH-1产孢量高于Fo-HN-46和Fo-HB-12。     

飞机草菌绒孢菌生物学性状研究

由飞机草菌绒孢菌引起的紫茎泽兰叶斑病是紫茎泽兰的重要病害。病原菌孢子萌发和病原菌生长的最适温度为25℃、pH为5.0。饱和湿度下孢子萌发率最高。紫外光照射对菌丝生长发育有抑制作用,而日光灯与室内自然光照射菌丝生长最好,连续黑暗最利于孢子产生。紫茎泽兰鲜叶片加入培养基有促进病原菌生殖生长的作用。病原菌能有效地利用各种碳源,以蔗糖最佳,氮源以蛋白胨最好。     

Simple and rapid detection of rice false smut pathogen Ustilaginoidea virens in rice seeds

A method for simple and rapid detection of Ustilaginoidea virens in rice seeds was developed based on specific polymerase chain reaction (PCR). To design the specific primers for detection of U. virens, the comparison was made on 5.8S rDNA intra- and inter-specific variations in nucleotide sequences of U. virens and other pathogens: Fusarium verticillioides, Sclerotinia sclerotiorum, Tilletia barclayana, Fusarium graminearum and Magnaporthe oryzae. A 346-bp fragment could be amplified by using the specific primers uvr-F and uvr-R in U. virens, but not in other test fungi, indicating that the designed primers were specific for the detection of U. virens. U. virens was detected in nine of the 24 tested rice samples, indicating that 37.5% of the rice samples were contaminated with the false smut pathogen. The development of the simple and rapid detection technique using specific primers will be applicable for direct identification of U. virens in rice seeds to screen large numbers of samples.     

Deleterious effects of fungi isolated from paddy soils on seminal root of rice

Soil samples were collected from rice paddies at 22 locations in northeastern Honshu, Japan. In 20 of the samples, seedling growth of rice was improved by soil pasteurization (aerated steaming at 60°C for 30 min), although no typical disease symptoms were observed in the seedlings grown in the untreated soil samples. In most locations, rice seedlings grew better in a potting medium containing root material from plants grown in the pasteurized portion of a soil sample than in a medium with root material from plants grown in the unpasteurized portion of the same sample. The results suggest that microorganisms that restrained rice seedling growth may be common in the soils of rice paddies. Approximately 800 isolates were obtained from seedling roots grown in unpasteurized soils and grouped by cultural and microscopic morphologies. The deleterious effects of 79 isolates from 21 major groups, most of which were soil-dwelling taxa, were examined after direct inoculation of the seminal roots. Isolates of Curvularia sp., Cirrenalia sp., Eppicoccum nigrum, Fusarium graminearum, F. oxysporum, Gliocladium virens, Humicola sp., Penicillium sp., Rhizoctonia oryzae-sativae, Sclerotium hydrophilum, Trichoderma aureoviride, and T. harzianum inhibited root growth, suggesting that deleterious root-infecting fungi were more common in paddy soil than previously thought. These fungi may be involved in the restraint of rice seedling growth.     

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。     

Bacillomycin D发酵与纯化工艺优化及其抑菌活性初步研究

为建立高纯度bacillomycin D（杆菌霉素D）脂肽的可放大制备工艺,促进其在农用抗生素领域应用价值的开发,经分离纯化及分子结构鉴定,确定了海洋源解淀粉芽孢杆菌Bacillus amyloliquefaciens TYg 3-2所产活性化合物为bacillomycin D脂肽类抗生素;选择精制过的天然培养基（酵母提取物和蛋白胨）,与组分明确的速效碳源、氮源（葡萄糖和硫酸铵）组成复合培养基,结合响应面优化调整培养基配方,建立了bacillomycin D发酵水平高达990 mg/L的5 L罐发酵工艺,其20.62 mg/（L·h）的比生产速率远高于文献报道的10.23 mg/（L·h）;在高品质原料基础上,发酵液经菌液分离、酸化沉淀、甲醇浸提、乙醚沉淀、大孔树脂吸附及硅胶柱层析工艺,可高效制备纯度达94.72%的样品。针对所制备的高纯度bacillomycin D样品进行了抑菌活性初步研究,证实其可离体拮抗德氏霉菌、玉米弯孢霉、稻绿核菌和长柄链格孢,并通过盆栽试验证实了bacillomycin D能提升同源芽孢杆菌TYg 3-2的防效,500 mg/L的bacillomycin D与4×106 CFU/mL的TYg 3-2菌体混合施用,对黄瓜炭疽病的防效可达95.3%,优于单独施用bacillomycin D（78.4%）和菌体（0）的防治效果。研究初步探索了bacillomycin D在农用抗生素领域的开发价值,同时其高纯度脂肽可放大制备工艺的建立至关重要,不仅能在保全生产菌（其芽孢为生防菌剂原料）的同时从其发酵液中分离和纯化目标脂肽,以促成对农用抗生素价值的深入研究,也能够满足中国农药登记相关毒理学试验对样品的要求（纯度>85%的克量级样品）,其结果可为芽孢杆菌源脂肽类抗生素的研究与开发提供参考。     

Biological Control of Sclerotinia pseudotuberosa and Other Fungi During Moist Storage of Quercus robur Seeds

The fungal pathogen Sclerotinia pseudotuberosa is a major cause of deterioration during storage of Quercus robur seeds (acorns) and along with other, mainly saprotrophic fungi, contributes to the decline of viability and vigour in the acorn population. Hot-water thermotherapy (HWT; 41 °C for 2.5 h) killed the fungal pathogen S. pseudotuberosa and prolonged the storage life of acorns. The addition of the systemic fungicide benomyl to the HWT and/or the broad-spectrum fungicide thiram as a seed dressing further enhanced the storage life of acorns. Three fungal antagonists, Coniothyrium minitans, Trichoderma sp. (KW3) and Trichoderma virens (G20), were also applied as a film-coating to acorns using a polyvinylacetate sticker achieving ca. 10⁷–10⁸ viable conidia per acorn. The biological treatments provided protection against infection and the spread of infection of S. pseudotuberosa and other fungi on the acorns during storage over several months. All treated and stored acorns grew normally following sowing in seedbeds. The Trichoderma species were more effective than C. minitans, with T. virens being the most effective. T. virens reduced pathogen spread from acorns infected with S. pseudotuberosa to `clean' acorns when T. virens coated infected and 'clean' acorns were mixed together. However, T. virens was less effective than HWT at preventing the proliferation of this pathogen within individual acorns that were infected before coating. A combination of HWT and subsequent coating with T. virens provided a more effective control against both S. pseudotuberosa and saprotrophic fungi than when either treatment was applied alone.     

Isolation and characterization of <Emphasis Type="Italic">Ustilaginoidea virens</Emphasis> and survey of false smut disease of rice in India

The intensity of rice false smut disease in selected states of northwest and south India was studied. In northern Indian states as a whole, disease incidence (percentage of false smut-infected tillers) varied from 2% to 75%. In the state of Haryana, maximum infection was recorded on hybrids like PA 6444 and PA 6129 while in Punjab state, 10–20% disease incidence was recorded in popular inbred rice varieties like PR 114, PA 116 and PAU 201. In the southern state of Tamil Nadu, the disease incidence varied from 5% to 85%. A heavy incidence of the disease was noticed in variety BPT 5204 and due to this, the air above the infected field gave a black smoky appearance from a distance as a result of release of spore mass in the atmosphere. In severe cases the number of infected grains reached even more than 100 per panicle. The pathogen Ustilaginoidea virens was isolated in potato dextrose agar medium and was characterized by both pathogenicity test and molecular analysis. Under glasshouse conditions, when a conidial suspension of the pathogen was injected during boot leaf stage of the rice variety TN1, typical smut balls were observed. The identity of the pathogen was further confirmed through polymerase chain reaction (PCR) analysis using U. virens-specific internal transcribed spacer (ITS) primers. The primer pair US 1-5/US3-3 and US2-5/US4-3 amplified 380 bp and 232 bp product, respectively, which are typical for the U. virens fungus.     

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.     

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.     

Frequency distribution of sensitivity of Ustilaginoidea virens to four EBI fungicides, prochloraz, difenoconazole, propiconazole and tebuconazole, and their efficacy in controlling rice false smut in Anhui Province of China

False smut, caused by Ustilaginoidea virens, is an important emerging disease of rice (Oryza sativa L.) in China. Up to now, as most varieties with high yielding and good quality are susceptible or even highly susceptible to false smut in most rice-growing ecological regions, especially in Anhui Province, chemical control with fungicides would be an important measure for the control of this disease. The ergosterol biosynthesis inhibitor (EBI) fungicides, such as prochloraz, difenoconazole, propiconazole and tebuconazole, are extensively used in China for the control of rice diseases, such as rice sheath blight and rice blast. In this study, a total of 102 U. virens isolates (from Anhui Province of China) were tested for their sensitivity to these four EBI fungicides during the stage of mycelial growth. The EC₅₀ ranges of values for prochloraz, difenoconazole, propiconazole and tebuconazole inhibiting mycelial growth of the 102 U. virens isolates were 0.04–0.75, 0.04–1.08, 0.04–0.38 and 0.03–0.57 μg?ml^?1, with the average EC₅₀ values of 0.32?±?0.08, 0.45?±?0.08, 0.19?±?0.03 and 0.21?±?0.06 μg?ml^?1, respectively. These values suggested that the tested U. virens isolates were very sensitive to these four EBI fungicides. Results of field trials showed that two sprays of three of the fungicides exhibited greater control efficacy than a single spray for the control of rice false smut. Two sprays of each was better than a single spray for the control of rice sheath blight. Two sprays of 50% propiconazole EC at 300 g a.i. ha^?1 gave the best control of rice false smut at both two sites during the two consecutive years, 2010 and 2011, with the control efficacy ranging from 71.5 to 74.3%. Sensitivity of the field U. virens isolates to EBI fungicides should be monitored. Mixtures, as well as alternation with other fungicides with different modes of action, should be tested.