一株水稻纹枯菌拮抗细菌的分离与鉴定

【目的】从土壤中分离并鉴定水稻纹枯菌拮抗细菌,测定其体外抑菌和温室防治效果。【方法】采用系列稀释法和平板对峙法筛选拮抗细菌,基于形态、生理特征及16S rDNA序列鉴定其分类地位,采用种子细菌化温室试验测定其防效。【结果】从蔬菜根际土壤中筛选出一株纹枯菌拮抗细菌,命名为kwkjT4。菌株具有明显的体外抑菌活性,对水稻纹枯病的温室防效与井冈霉素相当,初步鉴定为假紫色色杆菌(Chromobacterium pseudoviolaceum)。最适生长条件为pH 7.0,温度32°C,培养时间为36 h;抑菌活性物质产生的最适培养条件为pH 6.0,温度28°C,培养时间为48 h;表明两者并不一致。【结论】kwkjT4菌株在水稻纹枯病的生物防治中具有潜在的应用价值。这是C.pseudoviolaceum拮抗纹枯菌的首次报道。     

Influence of organic amendments on arbuscular mycorrhizal fungi in relation to rice sheath blight disease

 The effect of various organic soil amendments on arbuscular myorrhizal (AM) fungal activity on rice plants was tested under greenhouse and field conditions with reference to sheath blight (ShB) disease caused by Rhizoctonia solani. AM spore density, per cent infection, and intensity of infection were increased by organic amendments, whilst ShB disease was decreased. Certain amendments, especially green leaf manure, stimulated arbuscule development in rice plants. Mycorrhiza formation and sporulation were higher with healthy rice plants than with rice plants infected with R. solani. Our results indicate the possibility of using selective organic amendments to enhance development of native AM fungi and thus reduce disease incidence. Accepted: 9 November 1995     

Concentric leaf spot of yam (Dioscorea spp) in South-western Nigeria

The etiology and epidemiology of concentric leaf spot and seedling blight diseases of yam (Dioscorea spp) was investigated at Jbadan, south western Nigeria, in the low land humid tropics. Sclerotium rolfsii was associated with the concentric leaf spot and seedling blight of yam. The pathogen was also harbored by weeds namely Mucuna pruriensis, Commelina erecta, ipomea triloba, I. involucrataand Ipomoea sp found growing within the yam plots. A high mean inoculum densities of 3.0 ± 0.5 × 10⁶ colony forming unit/g of soil was estimated in the soil samples obtained from the yam plots This revised version was published online in June 2006 with corrections to the Cover Date.     

Detoxification of oxalic acid by pseudomonas fluorescens strain pfMDU2: implications for the biological control of rice sheath blight caused by Rhizoctonia solani

Rhizoctonia solani isolates varying in their virulence were tested for their ability to produce oxalic acid (OA) in vitro. The results indicated that the virulent isolates produced more OA than the less virulent isolates. In order to isolate OA-detoxifying strains of Pseudomonas fluorescens, rhizosphere soil of rice was drenched with 100 mM OA and fluorescent pseudomonads were isolated from the OA-amended soil by using King's medium B. These isolates were tested for their antagonistic effect towards growth of R. solani in vitro. Among them P. fluorescens PfMDU2 was the most effective in inhibiting the mycelial growth of R. solani. P. fluorescens PfMDU2 was capable of detoxifying OA and several proteins were detected in the culture filtrate of PfMDU2 when it was grown in medium containing OA. To investigate whether the gene(s) involved in OA-detoxification resides on the plasmids in P. fluorescens PfMDU2, a plasmid-deficient strain of P. fluorescens was generated by plasmid curing. The plasmid-deficient strain (PfMDU2P-) failed to grow in medium containing OA and did not inhibit the growth of R. solani. Both PfMDU2 and PfMDU2P- were tested for their efficacy in controlling sheath blight of rice under greenhouse conditions. Seed treatment followed by soil application of rice with P. fluorescens strain, PfMDU2, reduced the severity of sheath blight by 75% compared with the control, whereas PfMDU2P- failed to control sheath blight disease.     

Sensitivity of bacterial and fungal plant pathogens to the lytic peptides,MSI-99, magainin II,and cecropin B

In vitro and leaf disk assays of bacterial and fungal plant pathogens were conducted using three cationic lytic peptides, MSI-99, magainin II (MII), and cecropin B (CB). Growth of bacterial organisms was retarded or completely inhibited by low concentrations of these lytic peptides. The peptides also significantly reduced germination of fungal spores and growth of mycelia; however, higher concentrations of peptides were needed to inhibit fungal growth compared with those needed to inhibit bacteria. The relative efficacy of the peptides depended on the microorganism tested, but CB was the most inhibitory to the majority of the bacteria and fungi assayed. MSI-99, a synthetic derivative of MII with increased positive charge, showed equal or two- to fivefold higher antibacterial activity compared to MII in the in vitro assays. MSI-99 was also superior to MII against the oomycete, Phytophthora infestans but was slightly inferior to MII in assays with the true fungi, Penicillium digitatum and Alternaria solani. In the leaf disk assays, pretreating spores of Alternaria solani and Phytophthora infestans with the peptides at concentrations as low as 10 microg per ml led to significant reductions in the size of early blight lesions and prevented development of any late blight lesions on tomato leaf disks. Our results from in vitro and leaf disk assays suggest that MSI-99 can be used as a transgene to generate tomato lines with enhanced resistance to bacterial and fungal diseases of this crop.     

施硅增强水稻对纹枯病的抗性

采用水培的方法,从细胞学和生理生化方面研究了硅增强水稻对纹枯病的抗性作用。结果表明：加硅处理的水稻叶片硅化细胞和叶片表面的硅元素含量均显著高于缺硅处理（对照）：接种纹枯病菌后,加硅处理的MDA含量总体上低于缺硅处理,峰值尤为显著;加硅处理的SOD活性始终高于缺硅处理,接种后第4天加硅处理SOD活性较低时,其POD活性较高,而缺硅处理的POD活性较低,表明硅增强了SOD和POD之间的协调性;接种后硅对CAT和PAL活性没有产生明显影响,但降低了PP0活性;加硅能显著降低水稻植株的纹枯病病情指数。     

Involvement of secondary metabolites and extracellular lytic enzymes produced by Pseudomonas fluorescens in inhibition of Rhizoctonia solani, the rice sheath blight pathogen

Fourteen strains of Pseudomonas fluorescens isolated from rhizosphere soil of rice were tested for their antagonistic effect towards Rhizoctonia solani, the rice sheath blight fungus. Among them, PfMDU2 was the most effective in inhibiting mycelial growth of R. solani in vitro. Production of chitinase, beta-1,3-glucanase, siderophores, salicylic acid (SA) and hydrogen cyanide (HCN) by P. fluorescens strains was evaluated. The highest beta-1,3-glucanase activity, siderophore production, SA production and HCN production were recorded with PfMDU2. A significant relationship between the antagonistic potential of P. fluorescens against R. solani and its level of beta-1,3-glucanase, SA and HCN was observed.     

水稻株形对纹枯病抗性的影响

利用240份源于珍汕97B/明恢63的重组自交系水稻(Oryza sativa L.)群体,连续2年调查纹枯病病级与水稻生育期、株高和叶片长宽等18个株形性状的关系.对株形性状与纹枯病病级进行了偏相关分析.实验结果,只有植株松紧度与病级表型偏相关两年中都达到了显著或极显著水平,倒2叶基角、穗层整齐度等8个性状与病级之间的偏相关只有一年达显著或极显著水平.结合构建的分子标记遗传连锁图谱,对各性状进行QTL定位.在抗纹枯病QTL相近区间仅检测到控制分蘖角、植株松紧度和倒2叶基角的QTLS,未发现其余株形性状QTLs与抗纹枯病QTLs分布在同一染色体上.结果表明,水稻对纹枯病的抗性主要是由本身抗性基因控制,株形对纹枯病抗性表达的影响主要是间接影响,即通过改变田间小气候而影响发病程度.抗纹枯病育种在累加主效抗纹枯病QTLs的同时,也要注重选择不利于纹枯病发展的株形性状.     

Cloning and overexpression of antifungal barley chitinase gene in Escherichia coli

Plant chitinases are pathogenesis-related proteins, which are believed to be involved in plant defense responses to pathogen infection. In this study, chitinase gene from barley was cloned and overexpressed in Escherichia coli. Chitinase (35 kDa) was isolated and purified. Since the protein was produced as insoluble inclusion bodies, the protein was solubilized and refolded. Purified chitinase exerted broad-spectrum antifungal activity against Botrytis cinerea (blight of tobacco), Pestalotia theae (leaf spot of tea), Bipolaris oryzae (brown spot of rice), Alternaria sp. (grain discoloration of rice), Curvularia lunata (leaf spot of clover) and Rhizoctonia solani (sheath blight of rice). Due to the potential of broad-spectrum antifungal activity barley chitinase gene can be used to enhance fungal-resistance in crop plants such as rice, tobacco, tea and clover.     

Effects of Plant Contact, Inoculation Pattern, Leaf Wetness Regime, and Nitrogen Supply on Inoculum Efficiency in Rice Sheath Blight

An experimental design was developed which allowed the measurement of inoculum efficiency (IE) represented by lesions of a leaf-borne disease such as rice sheath blight. In this design. IE is measured as the ratio of newly established lesions on trap plants relative to the inoculum present in a canopy, i.e. lesions artificially established on source plants, IE of the rice sheath blight pathogen, Rhizoctonia solani , was studied under semi-controlled conditions in which the effects of the following factors were measured; contact frequency among host tissues, amount and location of inoculum in the canopy, leaf wetness regime, and nitrogen content of the host plant. The conduciveness of these factors to sheath blight infection was measured in terms of IE which was expressed as the ratio of the density of daughter lesions on trap rice hills to the density of mother lesions on the inoculated quadrat hills. IE generally declined with the three successive batches of trap hills used in the experiments. It was significantly higher at closer plant spacings and under interrupted leaf wetness regimes. IE was not affected by the amount, nor by the location, of inoculum (mother lesions) in the canopy, but was significantly lower in hills with high total nitrogen content.     

Evaluation of a Powder Formulation of Pseudomonas fluorescens Pf1 for Control of Rice Sheath Blight

Pseudomonas fluorescens strain Pf1, inhibitory to the growth of the rice sheath blight pathogen Rhizoctonia solani in vitro , was developed as a talc-based formulation. W hen the formulation was applied as a seed treatment, the bacteria established well in the rice rhizosphere. Root treatment or soil application of the formulation was less effective in establishing the bacteria into the rhizosphere. Effective control of rice sheath blight was obtained when the powder formulation was applied to seed, roots, soil and foliage, and these methods of application increased crop grain yield in four field trials. The possible commercial exploitation of the powder formulation of the bacteria is discussed.     

致病疫霉拮抗菌株B25-I-3的鉴定及其次级代谢产物

【背景】粘细菌是一类具有社会性行为的高等原核生物,能产生丰富、多样、新颖的具有生物活性的次级代谢产物,具有很大的研究开发价值。【目的】从土壤样品中筛选出对致病疫霉(Phytophthora infestans)具有拮抗作用的粘细菌,并对其次级代谢产物进行研究。【方法】对分离到的一株拮抗P. infestans菌株,结合形态观察和16S rRNA基因序列分析确定其分类地位,并通过单因素分析与正交优化相结合的方法对菌株的发酵参数进行研究。采用滤纸片法对其浓缩发酵液中活性物质的稳定性及抗菌活性进行检测,并通过薄层色谱法(thin layer chromatography,TLC)与高效液相色谱法(high performance liquid chromatography,HPLC)等初步分离后,将具有抗P. infestans活性的组分利用液相色谱-质谱联用技术(HPLC-MS)进行检测,最后通过离体叶片法测定活性物质对马铃薯晚疫病的防病作用。【结果】从土壤样品中分离得到的菌株B25-I-3对P. infestans表现出较强的拮抗活性,经鉴定为橙色粘球菌(Myxococcus fulvus)。其拮抗P. infestans的活性物质主要存在于胞外,产活性物质的最优发酵条件为：摇床转速180 r/min,接种量10%,培养温度30 °C,发酵周期7 d。该菌株产生的活性物质耐受蛋白酶K以及紫外线与自然光照射,对温度耐受性极强,而且易于在低温下保存,对枯草芽孢杆菌(Bacillus subtilis)、金黄色葡萄球菌(Staphylococcus aureus)、大肠杆菌(Escherichia coli)、酿酒酵母(Saccharomyces cerevisiae)、立枯丝核菌(Rhizoctonia solani)、尖孢镰刀菌(Fusarium oxysporum)、向日葵核盘菌(Sclerotinia sclerotiorum)均有不同程度的抑制作用。其对P. infestans的拮抗组分中含有N-(3-氨基-2-羟丙基)-N-甲基硫酸二酰胺[N-(3-amino-2-hydroxypropyl)-N-methylsulfuric diamide]与甲基(2R)-2-叠氮基-3-羟基-2-甲基丙酸酯[methyl(2R)-2-azido-3-hydroxyl-2-methylpropanoate],该活性物质能明显抑制P. infestans侵染马铃薯叶片且对不同品种的叶片均无损伤作用。【结论】研究为M. fulvus B25-I-3活性物质的分离鉴定及抗马铃薯晚疫病生物农药的研发提供了基础数据。     

Effect of carbendazim and mancozeb combinationon Alternaria leaf blight and seed yield in sunflower (Helianthus annus L.)

Abstract

Leaf blight caused by Alternaria helianthi (Hansf.) Tubaki & Nishihara, is the major disease of sunflower affecting the successful cultivation across India. Five individual fungicides and two combination fungicides were evaluated against this pathogen in laboratory and in field experiments. Among them, the combination of carbendazim + mancozeb completely (100%) inhibited the mycelial growth of A. helianthi, irrespective of the concentrations tested followed by carbendazim alone and metalaxyl + mancozeb under in vitro condition. In field conditions, the combination of carbendazim + mancozeb was found to be highly effective in reducing the leaf blight disease of sunflower in all the three experiments as compared to other fungicides and unsprayed control. The reduction of Alternaria leaf blight was also directly associated with an increase in seed yield. The economics of the fungicides spray has been worked out and the benefit cost ratio for the combination of carbendazim + mancozeb at 2.0 g/l was 7.1 as compared to unsprayed control. The overall analysis of the results revealed that the combination of carbendazim + mancozeb at 2.0 g/l can be used for the management of foliar diseases such as Alternaria leaf spot/blight in agricultural crops.     

钾素营养对防治番木瓜叶枯病的影响

对番木瓜叶枯病与植株营养、土壤相关养分的关系等进行研究,结果表明,番木瓜叶枯病是由于土壤有效钾缺乏而导致的一种生理性缺钾症;施用钾肥能有效控制该症的发生并获得较高产量。在发生缺钾叶枯病的番木瓜根、茎、叶片、叶柄等器官中镁含量均明显升高,说明番木瓜植株钾镁间存在拮抗作用。     

Synthesis and fungicidal activity of tubulin polymerisation promoters. Part 2: pyridazines

Special tetrasubstituted pyridazines are potent fungicides by promoting the tubulin polymerisation, hereby disrupting the microtubule dynamics in the fungus. They are monocyclic analogs of similar substituted triazolopyrimidines and pyridopyrazines with the same mode of action. The fungicidal activity of these pyridazines was evaluated against the plant pathogens Botrytis cinerea (grey mould), Mycosphaerella graminicola (wheat leaf blotch) and Alternaria solani (potato and tomato early blight). Structure-activity relationship studies revealed the importance of a methyl and a chlorine substituent next to both ring nitrogen atoms and two aryl or heteroaryl groups in the other two pyridazine positions.     

The beginning of the end? Extensive dieback of an open-grown Amur honeysuckle stand in northern Kentucky,USA

Amur honeysuckle (Lonicera maackii) is recognized as one of the most important invasive species in the Ohio River Valley. In 2012–2013, an outbreak of honeysuckle leaf blight on this species was observed in the region around Cincinnati, OH, USA. Dieback of open-grown honeysuckle stands was then noted in 2013, along with reduced physiological performance. One of these stands with signs of blight and dieback was surveyed in late summer 2013. The honeysuckle dead/total stem density fraction was 61.8 %, compared to 3.2 % reported in the 1980s. The dead/total basal area fraction was 36.9 %. The stem population size structure may also indicate a decline. Future work is needed to determine how widespread the decline is and if the leaf blight is the causative agent or an opportunistic infection.     

Screening for fire blight resistance in apple (Malus pumila) using excised leaf assays from in vitro and in vivo grown material

The apple cultivars Greensleeves and Novole and somaclones derived from Greensleeves were tested for resistance to fire blight (Erwinia amylovora) using excised leaf bioassays. Cv. Greensleeves was consistently susceptible to fire blight in in vitro, and in glasshouse and excised leaf bioassays whilst cv. Novole was resistant. Greensleeves somaclones, identified as having intermediate levels of fire blight resistance in glasshouse and in vitro screening tests, also showed intermediate levels of resistance in excised leaf bioassays. The length of leaves and the inoculum concentration employed affected the severity of symptoms observed. Excised leaf bioassays were unsuitable as tests for fire blight resistance using field grown material.     

Survival of Rhizoctonia solani AG‐1 IA,the Causal Agent of Rice Sheath Blight,under Different Environmental Conditions

The ability of Rhizoctonia solani AG‐1 IA, the causal agent of rice sheath blight, to survive in diseased rice straw and as sclerotia and mycelia was investigated. After storage for 10 months at 4°C, 25°C and non‐air‐conditioned natural room temperature (NRT, temperature range from 6°C to 35°C), sclerotia placed inside a desiccator, soaked in sterile water or immersed in wet paddy soil were viable. In contrast, only 15% of sclerotia in dry paddy soil survived. Survival of mycelia was severely affected by temperature and humidity. After 10 months in a desiccator at 4°C, 55% of mycelia samples could survive, whereas at 25°C and NRT, mycelial samples survived for only 7 and 5 months, respectively. However, mycelia stored in sterile water at constant temperatures (4°C or 25°C) survived for 10 months. A certain amount of UV radiation had no obvious effect on the survival of sclerotia or mycelia. The survival rate of the fungus in diseased rice straw stored for 16 months could reach 100% at 4°C, 50% at 25°C and 35% at NRT. The survival rates of the pathogen in diseased rice straw buried in dry, wet and flooded paddy soils after 10‐month storage at NRT were 75, 100 and 100%, respectively, indicating that soil humidity is a crucial factor for the survival of this fungus.