Cloning of Potato POTHR-1 Gene and Its Expression in Response to Infection by Phytophthora infestans and Other Abiotic Stimuli

A complete cDNA of potato Phytophthora infestans-induced hypersensitive response-related protein gene (POTHR-1) was cloned using rapid amplification of cDNA ends (RACE) strategy according to a fragment sequence which we had cloned using suppression subtractive hybridization (SSH) technique. The potato POTHR-1 gene encodes a protein of 225 amino acids, which shares 81% identity with tobacco hin1 gene-enoded protein (harpin-induced protein). Southern blot revealed that there are two to three copies of POTHR-1 in potato genome. The POTHR-1 gene expression in potato leaves showed that its transcripts accumulated remarkably in leaves after 36 h inoculation with P. infestans. Mechanical wounding and jasmonic acid (JA) could induce the POTHR-1 gene expression and osmotic stress just induce a slight accumulation of POTHR-1 gene mRNA, while salicylic acid (SA) had no detectable function on the induction accumulation of POTHR-1 gene transcripts. The potato POTHR-1 gene may preferentially associate with hypersensitive response (HR) or biotic cell death during interaction between host and pathogen.

影响大豆疫霉菌Phytophthora sojae卵孢子生活力和萌发的因素

本研究的目的是寻找一种能够促使大豆疫霉菌卵孢子在短时间内大量萌发的方法。结果表明,卵孢子菌龄、预处理温度、化学物质及其浓度在某种程度上均影响大豆疫霉菌卵孢子的生活力。胡萝卜琼脂平板上24℃密封培养条件下,30d菌龄的卵孢子中只有10.56%处于萌动状态,但经0.4%的KMnO4处理20min或35℃处理5d后,萌动率分别提高到58.49%和83.55%,但并不萌发。以3mL大豆感病品种Sloan的根系分泌物为培养液,26℃黑暗或光照培养7d即可获得80%以上的卵孢子萌发率。

Phylogenetic Analysis of the Sequences of rDNA Internal Transcribed Spacer (ITS) of Phytophthora sojae

The internal transcribed spacer （ITS） region （ITS1, ITS2 and 5.8S rDNA） of the nuclear ribosomal DNA （nrDNA） was amplified via the PCR method in seventeen different isolates of Phytophthora sojae using the common primers of the ITS of fungi. Around 800 bp- 1,000 bp fragments were obtained based on the DL2000 marker and the sequences of the PCR products were tested. Taking isolate USA as outgroup, the phylogenetic tree was constructed by means of maximum parsimony analysis, and the genetic evolution among isolates was analyzed. The results showed that there is a great difference between the base constitution of ITS 1 and ITS2 among various isolates. The seventeen isolates are classified into three groups, and the isolates from the same region belong to the same group, which shows the variation in geography.

Screening of Antagonistic Bacteria against Phytophthora infestans and Its Inhibition Effect

[ Objective ] The paper was to screen bacterial strain with significant antagonistic effect against Phytophthora infestans, so as to provide basis for further development and utilization of antagonistic bacteria to inhibit P. infestans and control potato late bright. [ Method] Plate dual culture and filter paper method were used to determine the inhibition effect of strains in vivo, fermentation broth and bacterial liquid of 61 strains against P. infestans and the resistance-induction effect of SR13-2 strain. [ Result] The inhibition rate of 24 strains among 61 tested strains against mycelial growth of P. infestans was greater than 60%, and the inhibi- tion effect of HT-6 strain was the strongest with the inhibition rate of 89.92%. However, fermentation broth of all tested strains had no significant inhibition effect against P. infestans, while the inhibition effect of bacterial liquid of most strains was significantly higher than strain in vivo; the inhibition effect of $34-1 strain was the strongest with inhibition rate of 91.50%. The bacterial liquid of SR13-2 strain was found to have significant resistance-induction effect with protective rate of 60%. [ Conclusion] The inhibition effect of strains in vivo and fermentation broth of antagonistic strains S34-1 and SR13-2 had no relationship with each other, while bacterial liquid had great application potential in controlling potato late bright.

不同浓度臭氧水对致病疫霉菌Phytophthora infestans孢子萌发抑制作用初报

采用不同浓度臭氧水对致病疫霉菌Phytophthora infestans(Mont.)de Bary孢子囊萌发进行了试验,结果表明:臭氧水处理对孢子囊萌发的影响与臭氧水浓度密切相关,以9 mg/kg臭氧水处理1 min可以抑制95%以上孢子囊萌发,以18 mg/kg臭氧水处理1 min则100%抑制孢子囊萌发。无论在高浓度还是在低浓度条件下,处理时间1 min或2 min对孢子囊萌发没有产生显著影响,杀菌作用几乎是瞬间完成的。臭氧水处理孢子囊后导致孢子囊形态发生变化,这种变化随时间延长变得更加明显。鉴于高浓度臭氧水对植物本身不产生毒害作用,因此,高浓度臭氧水在控制晚疫病方面显示出广阔的应用前景。

Screening and Identification of Antagonistic Strain against Phytophthora sojae

[ Objective] The paper was to screen the antagonistic strain against Phytophthora sojae with biocontrol potential, and provide basis for searching control measures and designing new control strategies against P. sojae. [ Method] The rhizosphere soil of soybean was collected from three different places in Heilongjiang Province, and various soil microorganisms were isolated. Dual culture method was used to screen the microorganism with antagonistic effect against P. sojae. On this basis, the growth inhibition rate of the microorganism with stronger antagonistic effect against P. sojae was determined, and its control effect against P. sojae was also measured. [ Result] A strain of bacterium with relatively good antagonistic effect was isolated from soil, and named as strain B048. Dual test showed that the growth inhibition rate of antagonistic bacterium 11048 against P. sojac reached 97.5%. Antagonistic endurance tests showed that the width of inhibition zone was still 20.0 mm after dual culture with P. sojac for21 d. In potting experiment, the control effect of B048 against P. sojae was 100%. The antagonistic bacterium was primarily identified to be Bacillus pumilus through morphology and 16S rDNA sequence analysis. [Condusion] The antagonistic bacterium B048 had good prospect to be developed as the biocontrol bacterium against P. sojae.

Improvement of Polyacrylamide Gel Electrophorus in Phytophthora infeatans SSR Marker

[Objective] The paper was to improve polyacrylamide gel electrophorus in Phytophthora infeatans SSR Marker.[Method] With the disease sample of P.infeatans collected from Guyuan in Ningxia Province in 2009 as test material,its DNA was extracted and amplified with PCR,and its products were carried out polyacrylamide gel electrophoresis.[Result] 12% polyacrylamide gel electrophoresis was used to detect primers D13,G11 and PI02,and 8% polyacrylamide gel electrophoresis was used to detect primers PI4B,PI63,SSR4,SSR8 and SSR11,then 0.1% silver nitrate was used to stain,and an ideal electrophoresis and staining effect was obtained.[Conclusion] The electrophoresis and staining method suitable for P.infeatans SSR Marker established in the study had the characteristics of high sensitivity,simple operation and clear bands,which was an effective,simple and quick detection method.

Distribution and Virulence Diversity of Phytophthora sojae in China

By investigating occurrence of Phytophthora root rot in fields and isolating P.sojae fromdiseased plants and soils, the distribution of P.sojae in China was surveyed. In addition tonortheast region, P.sojae existed in Huanghe-Huaihe basin and Yangtze basin too. Eighty- threeisolates of P.sojae isolated from different areas were identified on virulence using 13differential soybean cultivars, abundant virulence diversity was found in P.sojae. The greaterdiversity in virulence of P.sojae was in isolates from soil than from plants. And the greatestvirulence diversity of P.sojae was found in Yangtze basin.

烟草黑胫病原菌(Phytophthora parasitica Var·nicotianae)生理生化特性的研究

本文主要介绍了用人工合成培养基测定烟草黑胫病原菌的致死温度、最适生长的pH值和能够生长的pH值范围的方法及结果。对黄河等“适宜于马铃薯晚疫病菌的一种液体合成培养基”,在配方和实验方法上做了一些改进。通过实验,进一步证实了该菌在营养生长过程中钙素的重要性和三羧酸循环(TCA)中有机酸的重要性;该菌的营养生长并不需要较复杂的营养条件。

Effect of Different Culture Conditions on Sporulation Quantity of Phytophthora capsici in Capsicum annuum L. var. dactylus M. in Xunhua

[Objective] The paper was to explore the effect of different culture conditions on sporulation quantity of Phytophthora capsici in Capsicum annuum L.var.dactylus M.in Xunhua.[Method] The effects of light hours,pH,medium and temperature on sporulation quantity during isolation and culture process of P.capsici were studied.[Result] The sporulation quantity of P.capsici under the conditions of 24 h/day light,pH 7.0,potato medium（PSA） and 30 ℃ was the largest,and pH,basal medium and temperature had greater impact on sporulation quantity.[Conclusion]The study laid foundation for the study on natural incidence condition of C.annuum in Xunhua.

Resistance Evaluation of Soybean Germplasm from Huanghuai Region to Phytophthora Root Rot

The aim of the study was to establish a set of differential strains and to identify soybean resistant genes to Phytophthora root rot and then to apply those strains for analysis of the resistant genes Rps1a,Rps1c,and Rps1k that soybean cultivars or lines may carry.Virulence formula of 125 Phytophthora sojae isolates were determined using the hypocotyls inoculation technique,the strains,which includ 6 isolates with different virulence formulas,were applied to identify the resistance of 55 soybean cultivars or lines and resistant genes were analyzed using the gene postulating procedure.Eighteen reaction types occurred in 55 cultivars or lines and results of gene postulation indicated that 2 cultivars or lines probably carried gene Rps1c and no cultivar may carry genes Rps1a or Rps1k.A few of soybean cultivars or lines from Huanghuai Region carry Rps genes Rps1a,Rps1c and Rps1k and tend to infect by P.sojae,so resistant cultivars or lines need to be bred and popularized actively.

Combined Toxicity Test of Complex Agents of Metalaxyl and Other Fungicides against Phytophthora parasitica var. nicotianae

[ Objective] To overcome the resistance of Phytophthora parasitica var. nicotianae against metalaxyl and effectively control its damage, new efficient complex agent of metalaxyl was studied and developed. [ Method] The toxieities of nine fungicides against P. parasitica were measured using growth rate method. On this basis, the fungicides with good effects were selected to compound with metalaxyl, and the optimum complex ratio was confirmed. [Result] The toxicity of metalaxyl was the strongest with EC50 of 2. 130 5μg/ml; followed by carbendazim, mancezeb and dimethomorph with EC50 of 2.357 9, 2.639 8 and 2. 778 8 μg/ml. The effect of cyazofamid was the poorest with EC50 of 6. 278 8 μg/ml. The optimum complex ratios of dimethomorph, carbendazim and mancezeb with metalaxyl were 40： 60, 30：70 and 20： 80, and their co-toxicity coefficients were 138.80,124.25 and 115.00, respectively. [ Conclusion] The complex agents had application and promotion value, which could be used to carry out further field trials.

大豆疫霉菌(Phytophthora sojae)增强型绿色荧光蛋白遗传转化载体的构建

载体的构建是建立遗传转化体系的基础。以真核表达载体pcDNA3.1(-)/hygro为基本骨架,构建大豆疫霉菌遗传转化载体,通过限制性内切酶酶切、去磷酸化、连接等基因重组技术,将增强型绿色荧光蛋白(Enhanced Green Fluorescent Protein,EGFP)基因和来自莴苣霜霉菌(Bremia lactucae)的启动子(ham34)、终止子重组到真核表达载体pcDNA3.1(-)/hygro中,经大肠杆菌转化后对转化子进行了酶切验证,为大豆疫霉菌遗传转化体系的建立提供载体。

Phytophthora α1性信息素C(9)～C(16)片段的合成

以甾体皂甙元氧化降解废弃物（R）-4-甲基-5-羟基戊酸甲酯为原料，经包括酯的格氏加成反应、烯烃的臭氧化反应和环氧化合物的亲核加成等六步反应，以60．5％的总收率合成得到了Phytophthora α1性信息素的C（9）-C（16）片段4。

柑桔脚腐病菌Phytophthora parasitica杀菌剂的室内筛选

柑桔脚腐病是为害柑桔主干基部的病害。采用菌丝生长速率法对柑桔脚腐病菌寄生疫霉Phytophthoraparasitica进行了21种农药的室内毒力测定。结果表明，各药剂对寄生疫霉菌丝的生长均有抑制作用，并且其抑制率与药剂浓度呈正相关。其中，毒力作用最强的是50％烯酰吗啉水分散粒剂，EC50为0．2598μg／mL；其次是60％唑醚·代森联水分散剂、2％春雷霉素水乳剂、70％丙森锌可湿性粉剂、10％多抗霉素B可湿性粉剂、50％多·锰锌可湿性粉剂和50％醚菌酯水分散粒剂，EC5。在1．0～7．7μg／mL之间；再次是65％代森锌可湿性粉剂、10％井冈霉素水乳剂、10％苯醚甲环唑水分散粒剂、40％腈菌唑水分散粒剂、43％戊唑醇悬浮剂、25％丙环唑乳油、3％中生菌素可湿性粉剂、68％碱式硫酸铜水分散粒剂、40％氟硅唑乳油和12．5％烯唑醇可湿性粉剂，EC50在18．0～69．0μg／mL之间；较差的是50％敌克松可湿性粉剂、50％异茵脲可湿性粉剂、70％甲基硫菌灵可湿性粉剂和50％甲基硫菌灵悬浮剂，EC50在329．0μg／mL以上，尤其甲基硫茵灵70％可湿性粉剂和50％悬浮剂的EC50分别高达979．1665和1713．8330μg／mL，不宜推荐继续用于柑桔脚腐病的防治。

对马铃薯晚疫病菌(Phytophthora infestans)有杀菌毒性的中草药的筛选(英文)

将来源于 44科的 88种中草药等植物材料的提取物稀释 5 0 0倍后 ,与马铃薯晚疫病菌 (Phytophthorainfestans)的孢子囊悬浮液混合。有 31种材料的提取物在该浓度下表现了对马铃薯晚疫病菌的游动孢子囊间接萌发的完全抑制作用 ,有 46种提取物也表现了不同程度的抑制作用 ,另有 11种提取物在该浓度下没有表现抑制作用。在抑制菌丝生长作用的试验中 ,植物提取物用黑麦培养基稀释 2 0 0倍 ,然后倒皿 ,在 8.5cm直径的培养皿中央放一马铃薯晚疫病菌菌片 ,16℃下培养 ,15d后测量菌落直径大小。结果表明 ,在供试的 83种提取物中 ,在稀释 2 0 0倍浓度下 ,有 32种提取物表现了对菌丝生长的完全抑制作用 ,2 4种表现了比较强的抑制作用 ,而有 2 7种提取物没有表现对菌丝生长的抑制作用 ,甚至还表现了对菌丝生长的促进作用。同时 ,还有

云南省发现马铃薯晚疫病菌(Phytophthora infestans)A2交配型

将一批分离自楚雄、大理、昆明、曲靖、宣威等马铃薯产区的晚疫病菌在利马豆 Ｖ８ 琼脂培养基上与 Ａ１ 交配型标准菌株进行交配型测定， ４７ 个菌株中出现了１０ 个 Ａ２ 交配型菌株。亲和性菌株交配时产生大量具有典型特征的有性结构。对卵孢子、穿雄生的藏卵器以及雄器的形态特征和大小进行了描述。这是云南马铃薯晚疫病菌中存在 Ａ２ 交配型菌株的首次报道。

A Preliminary Study on the Resistance to Different Fungicides in Phytophthora parasitica var. nicotianae

[Objective] This study aimed to investigate the resistance to different fungicides in Phytophthora parasitica var. nicotianae. [Method] Under indoor incubation conditions, the resistance to dimethomorph, metalaxyl-mancozeb, propamocarb and ovraclostrobin.dimethomorph in P. parasitica strain isolated from Zhenyuan County in Qiandongnan State was analyzed with colony growth measurement method. [Result] P. parasitica exhibited different levels of sensitivity to four fungicides. To be specific, P. parasitica exhibited the highest resistance to dimethomorph, and ECho reached 1.19 μg/ml. [Conclusion] In Zhenyuan tobacco-growing area, long-term single use of dimethomorph possesses certain resistance risk in prevention and control of black shank disease.

影响大豆疫霉菌(Phytophthora sojae)游动孢子产生的条件

间歇性地给菌丝块换水可以诱导菌丝产生孢子囊 ,进而释放游动孢子。挑取 5～ 8块直径为 8mm于利马豆或 V8汁平板上培养 4～ 8d的菌块 ,置于直径 7cm的培养皿内 ,加入蒸馏水正好没过菌块表面 ,每 30 min换水一次 ,换水 4次后加入 15 ml Petri培养液 ,2 5℃、黑暗条件下培养 18～ 2 0 h,可诱发菌块大量产孢。

致病疫霉(Phytophthora infestans)对杀菌剂抗药性研究进展

综述了致病疫霉 (Phytophthorainfestans)对内吸性杀菌剂甲霜灵、霜脲氰和保护性杀菌剂代森锰锌和百菌清的抗性研究进展 ,总结了当前用于防治晚疫病的药剂种类并提出了抗性治理策略。对于致病疫霉抗性遗传研究和抗性治理及药剂防治都具有一定的参考价值。