辣椒疫霉致病毒素

辣椒疫霉在培养液中振荡培养时可分泌毒素,这种纱能经志辣椒叶片形成水渍状褐腐斑,类似病原菌侵染后形成的症状,Ｐｌｉｃｈ’ｓ和Ｖ８Ｃ２液适于Ｐ．ｃａｐｓｉｃｉ的生长和产毒;生长适宜温度和ＰＨ范围分别为２０－３０℃和ＰＨ６－７,在２５－３０℃、Ｐ５－８的条件下２滤液毒性最强;培养１５天产毒达到最高值,液先后经８５％硫酸铵沉淀,ＤＥ５２、Ｃ几ＳｅｐｈａｄｅｘＧ－７５柱层将毒素纯化。经鉴定该纯毒素为３９．８     

辣椒疫霉致病毒素

辣椒疫霉(Phytophthora capsici Leonian)在培养液中振荡培养时可分泌毒素,这种毒素能引起辣椒叶片形成水渍状褐腐斑,类似病原菌侵染后形成的症状。Hich’s和V₈培养液适于P capsici的生长和产毒;生长适宜温度和pH范围分别为20～30℃与pH6～7,在25～30℃、pH5～8的条件下培养滤液毒性最强;培养15天产毒达到最高值。滤液先后经85％硫酸铵沉淀、DE52、CM32和SephadexG-75柱层析将毒索纯化。经鉴定该纯毒索为39．8kD酸性糖蛋白。并对80℃以上高温或蛋白酶敏感,而β-D-葡萄糖苷酶处理不影响其毒性,蛋白亚基为毒紊的活性中心。毒素处理寄主叶片引致病理性坏死,这一作用与辣椒品种抗病性有关。     

辣椒疫霉产毒共分离RAPD标记的研究

辣椒疫霉（Ｐｈｙｔｏｐｈｔｈｏｒａ　ｃｏｐｓｉｃｉ）毒素的产生是由一个不完全显性基因所控制,通过ＲＡＰＤ／ＢＳＡ分析证明,用ＯＰＷ１２扩增得到一条约１３００ｂｐ的ＲＡＰＤ标记带,该带与辣椒疫霉产毒共分离。纯化回收ＯＰＷ１２　１３００ＤＮＡ,共转化感受态Ｅ．ｃｏｌｉ　ＤＨ５ａ,筛选出３个白色阳性克隆,序列分析发现该标记ＤＮＡ为１２９１ｂｐ。这一标记ＤＮＡ序列的阐明为进一分析产毒遗传机理提供了新的信息。     

辣椒疫霉产毒共分离RAPD标记的研究

辣椒疫毒（Ｐｈｙｔｏｐｈｔｈｏｒａ ｃａｐｓｉｃｉ）毒素的产生是由一个不完全显性基因所控制,通过ＲＡＰＤ／ＢＳＡ分析证明,用ＯＰＷ１２扩增得到一条约１３００ｂｐ的ＲＡＰＤ标记带,该带与辣椒疫霉产毒共分离。纯化回收ＯＰＷ１２１３００ＤＮＡ,共转化感受态Ｅ．ｃｏｌｉ ＤＨ５α,筛选出３个白色阳性克隆,序列分析发现该标记ＤＮＡ为１２９１ｂｐ。这一标记ＤＮＡ序列的阐明为进一步分析产毒遗传机理提供了新的信息     

辣椒疫霉产毒共分离RAPD标记的研究*

辣椒疫霉（Ｐｈｙｔｏｐｈｔｈｏｒａ　ｃｏｐｓｉｃｉ）毒素的产生是由一个不完全显性基因所控制,通过ＲＡＰＤ／ＢＳＡ分析证明,用ＯＰＷ１２扩增得到一条约１３００ｂｐ的ＲＡＰＤ标记带,该带与辣椒疫霉产毒共分离。纯化回收ＯＰＷ１２　１３００ＤＮＡ,共转化感受态Ｅ．ｃｏｌｉ　ＤＨ５ａ,筛选出３个白色阳性克隆,序列分析发现该标记ＤＮＡ为１２９１ｂｐ。这一标记ＤＮＡ序列的阐明为进一分析产毒遗传机理提供了新的信息。     

新疆甜瓜疫霉菌毒素的分离及其致病效应（简报）

从瓜类疫霉菌液体培养液中分离到一种对新疆甜瓜幼苗有明显致萎效应的毒素，用ＤＥＡＥ－５２柱纯化得到两个组分，其中组分Ⅰ蛋白质含量低于组分Ⅱ，而糖含量则高于组分Ⅱ，组分Ⅱ的致萎效应大于部分Ⅰ。     

贵州地区木霉菌分离鉴定及对辣椒疫霉的拮抗作用

【背景】辣椒疫霉是一种毁灭性的土传病害,当前主要使用化学合成杀菌剂防治,但容易导致环境污染和食品安全等问题。【目的】筛选可拮抗辣椒疫霉的候选菌株,探究分离菌株拮抗辣椒疫霉的生理生化作用机制。【方法】综合应用形态学、核糖体RNA (rRNA)基因非转录区ITS序列相似性方法鉴定分离菌株,通过对峙实验筛选抑菌效果较高的拮抗菌株,基于比色法测定分离菌株发酵液粗提物对辣椒疫霉菌丝脂质过氧化、纤维素酶、β-葡萄糖苷酶(β-GC)和多聚半乳糖醛酸酶(PG)活性的影响。【结果】从腐木和土壤样品中分离得到11株木霉,分属于绿色木霉(Trichodermavirens)、哈茨木霉(Trichoderma harzianum)、钩状木霉(Trichoderma hamatum)和棘孢木霉(Trichoderma asperellum) 4个种。11株木霉对辣椒疫霉均有一定的抑制作用,抑制率达到90%以上的菌株包括:绿色木霉Tv-1(92.68%)、Tv-2 (95.12%),哈茨木霉Thz-2 (92.68%),钩状木霉Tha-1 (90.24%)。以4株高效木霉的发酵液粗提物处理辣椒疫霉菌丝5 d后,因脂质过氧化产生的丙二醛含量显著增加,分别达到1.20、1.48、2.69和3.16 nmol/g,显著高于对照处理的0.77 nmol/g;与对照组相比,β-GC、PG酶活性显著下降,分别降低了12.28%-64.91%、7.2%-15.5%;同时纤维素酶活性呈上升趋势,最显著组为2.647 U/mL,相对于对照组增加了0.831U/mL。【结论】分离得到4株明显抑制辣椒疫霉菌生长的高效木霉菌,主要通过破坏细胞壁结构、降低致病因子酶活力和增强脂质过氧化等方式起拮抗作用,可为辣椒疫病的生物防治提供理论依据和技术支持。     

中国橡胶树疫霉种的研究

疫病是我国植胶区的主要病害。近年来,作者从云南西双版纳和广东海南岛的橡胶树和胶园土共分离出57株疫霉菌种。通过分类研究,共鉴定出4个种:恶疫霉 Phytophthoracactorum(Leb.＆ Cohn)Schroeter,辣椒疫霉 P.capsici Leoman,柑桔褐腐疫霉 P.citrophthora(Sm.＆ Sm.)Leonian,和棕榈疫霉 P.palmivora(Butl.)Butler。其中辣椒疫霉是首次在橡胶树上发现。我国橡胶树疫霉的种群结构与东南亚和南亚的有所不同,除棕榈疫霉外,其余3种在东南亚和南亚均未发现。而东南亚常见种:簇囊疫霉(P.botryosa)、橡胶疫霉(P.heveae)和蜜色疫霉(P.meadii),在我国却迄今尚未发现或有待证实。以前报道分离自胶园土壤中的芋疫霉(P.colocasiae),可能系柑桔褐腐疫霉之误。绝大多数分离物经配对培养均可产生性器官:辣椒疫霉的A~1交配型和A~2交配型大致相等;柑桔褐腐疫霉和棕榈疫霉的A~2交配型则明显多于A~1交配型。     

抗甲霜灵辣椒疫霉菌的环境适合度

摘要:【目的】研究抗甲霜灵辣椒疫霉菌的环境适合度,对于评估甲霜灵防治辣椒疫霉的抗药性风险具有重要意义。【方法】以室内药剂驯化的抗甲霜灵辣椒疫霉菌株Pc2-3为研究对象,分析比较其与原始敏感菌株Pc2的主要生物学特性、生长竞争力、致病力及土壤适合度等环境适合度指标。【结果】Pc2-3孢子囊产生量(3 d后)、孢子囊释放率(24 h后)和游动孢子萌发率(8 h后)分别是Pc2的0.44、0.09和0.54倍。Pc2-3可生长温度和pH范围及最适生长温度与Pc2基本一致,但菌丝生长速率低于Pc2。竞争力测定显示,在胡萝卜(CA)平板培养条件下,Pc2-3生长极显著弱于Pc2。盆栽致病试验显示,Pc2-3对辣椒植株的致病率为14.3%,明显低于Pc2(88.6%)。两者等量混合后接种,辣椒植株的发病率为75.7%,接近单独接种Pc2时的发病率,且所有病株分离出的辣椒疫霉菌均为甲霜灵敏感型。分别将Pc2-3和Pc2游动孢子加入自然土壤培养20 d后,实时定量PCR检测显示Pc2-3数量是Pc2的0.28倍,当土壤中含有300 mg/kg干土的甲霜灵,则前者为后者的0.42倍。此外,2个菌株最适存活土壤温度和湿度基本一致,当土壤温度和湿度利于辣椒疫霉存活时,Pc2-3土壤适合度显著低于Pc2,不利于辣椒疫霉存活时,Pc2-3土壤适合度略低于Pc2。【结论】抗甲霜灵菌株Pc2-3环境适合度弱于原始敏感菌株Pc2。     

辣椒炭疽病菌毒素

辣椒炭疽病菌（辣椒刺盘孢Ｃｏｌｌｅｔｏｔｒｉｃｈｕｍ ｃａｐｓｉｃｉ）在２５－２８℃的Ｃｚａｐｅｋ－Ｄｏｘ培养液中振荡培养时分泌出毒素,这种毒素能引起辣椒叶片形成坏死斑,类似于病原菌侵染形成的症状。辣椒叶煎汁培养液和Ｃｚａｐｅｋ－Ｄｏｘ培养液适于Ｃ．ｃａｐｓｉｃｉ的生长和产毒,生长适宜温度和范围分别为２５℃和ｐＨ６－７,在２５－２８℃,ｐＨ６－７的条件下培养滤液毒性最强;光线和通气条件可促进Ｃ．ｃ     

辣椒炭疽病菌毒素

辣椒炭疽病菌(辣椒刺盘孢Colletotrichum capsici)在25—28℃的Czapek-Dox培养液中振荡培养时分泌出毒素,这种毒素能引起辣椒叶片形成坏死斑,类似于病原菌侵染形成的症状。辣椒叶煎汁培养液和Czapek-Dox培养液适于C.capsici的生长和产毒,生长适宜温度和范围分别为25℃和pH6—7,在25—28℃,pH6—7的条件下培养滤液毒性最强;光线和通气条件可促进C.capsici的生长和产毒;培养14天的培养滤液毒性最强。培养滤液经丙酮沉淀及离子交换树脂柱和Sephadex G-50柱层析将毒素纯化。实验结果表明该毒素为多聚糖类物质。生物检测结果表明:培养滤液和C.capsici毒素溶液能抑制辣椒、绿豆、豌豆、豇豆种子胚根生长;并能使辣椒幼苗发生萎蔫,这一作用与辣椒品种抗性有关。     

GENETIC EVIDENCE FOR ZYGOTIC MEIOSIS IN PHYTOPHTHORA CAPSICI

Variation of several traits among single-oospore isolates of the heterothallic oomycete, Phytophthora capsici, was studied to determine genetically the place of meiosis in the life cycle. Low percentages of met^– offspring were recovered from two crosses of met^– X prototrophic isolates. Arg^– cultures were also recovered from the crosses indicating the possible presence of a suppressed arg^– gene in one of the parental cultures. Three arg^– X met^– crosses yielded all four of the possible classes of offspring. Thus meiosis was either zygotic or, if gametangial, the original parental cultures must have been heterozygous at both the arg and met loci. However, when these parental cultures were crossed, only prototrophic offspring were recovered. A ratio of 3 prototrophs: 1 met^– was obtained from cross R in which two prototrophic offspring from a previous arg^– X met^– cross were mated. This indicated the presence of a suppressor of met^– and may explain the low recovery of met^– offspring from earlier crosses. From backcrosses of met^– offspring from this cross to the A² prototrophic parent, ratios close to the expected 1:1 were obtained. When the A¹ prototrophic parent of cross R (K-10) was crossed to an A² prototrophic isolate, met^– offspring were again recovered, which indicated that K-10 carried a suppressed met deficiency. In two crosses of streptomycin-resistant (S^r) X sensitive (S^s) isolates, ratio of 108 S^s: 14 S^r and 84 S^s:14 S^r were obtained. Although 1:1 ratios would have been expected if meiosis were zygotic, segregation of S^s and S^r in the F₁ generation indicated that gametangial meiosis was unlikely. Segregation of the A¹ and A² mating types in all crosses also indicated that meiosis was zygotic in P. capsici. Possible control of mating type by a single pair of alleles, by plasmids, and by more complex mechanisms is discussed in relation to the mating type ratios obtained.     

木霉对辣椒疫霉菌抑制作用的超微结构与细胞化学

哈茨木霉菌株NF9和TC3对辣椒疫霉病菌具有强烈的抑制作用。超微结构与细胞化学研究表明,木霉菌丝能够缠绕并寄生疫霉菌菌丝,且重寄生作用主要发生在培养基内的菌丝上。但在绝大多数情况下,木霉菌可直接水解和破坏疫霉菌菌丝,说明木霉抑菌作用的主要机制是直接向外分泌水解酶分解疫霉的细胞壁及细胞质,而不是重寄生作用。不同培养基对木霉的重寄生作用有重要影响,减少培养基中葡萄糖含量可以增强木霉的重寄生作用和抑菌效果。此外,在菌落生长后期,疫霉的新生菌丝均可侵染自身的老菌丝。     

疫霉属菌种的液氮超低温保藏

对疫霉属的15个种68株菌和霜疫霉2株菌进行了液氮超低温的保藏试验并得到成功。比较了冷冻速度、保护剂和解冻速度对菌存活的影响。严格控制每分钟降温摄氏1度直到-40℃后再放入液氮罐中,对疫霉和霜疫霉来说都是非常必要的。这种降温程序可通过简单设备人工操作达到。而直接由室温降到-150℃以下会损伤菌种以致死亡。在所用的保护剂中,不论10％甘油还是5-15％二甲基亚砜都能起到保护作用。尚看不出对那种保护剂有特别的要求,似可任意选用。至于解冻条件,由液氮中取出放置在38℃水浴中快速融化与在20℃水浴中中等速度融化效果相等,对菌的成活都没有太大影响。我们的试验肯定了在有保护剂存在下,用慢速冷冻可以在液氮中保存疫霉和霜疫霉。     

几种真菌发酵液对致病疫霉的抑制作用

测定了８种真菌发酵液在５种不同浓度下对致病疫霉菌丝生长、游动孢子静止、静止胞萌发、附着胞形成和侵入丝形成等不同阶段的影响。结果表明,供试真菌不同浓度的发酵液,对致病疫霉上述各个阶段均有一定程度的抑制作用,并均随发酵液浓度增加,抑制作用逐渐增强,浓度为１００％时,抑制作用均达到最高。其中,立枯丝核菌发酵液的抑制作用最强,浓度为１００％时,对致病疫霉菌丝生长的抑制率达到９０．４％,而静止胞萌发率仅为２．４％,附着胞及侵入丝均未见形成。     

苎麻疫霉雄器侧生性状的遗传研究

在来自江苏、江西棉花、芒麻和构树的12个些麻疫霉（PhytophthoraboehmeriaeSawada）菌株中均观察到侧生雄器,其比率为4．0％～16．5％。在以菌株JS－5和PM－8（雄器侧生比率分别为16．5％和9．5％）为亲本所建立的连续2～3代单游动孢子无性系后代中,雄器侧生性状可以遗传,但单孢株间雄器侧生比率有一定差异,其分布范围分别为9．0％～34．0％和2．5％～15．5％。进一步诱导菌株JS－5的单游动孢子株的卵孢子萌发,分别对具侧生雄器和具围生雄器的藏卵器（卵孢子）进行单孢分离,检测雄器位置性状在由上述不同来源卵孢子萌发所形成的单卵孢株后代的遗传与变异。结果表明,无论是由具侧生雄器的藏卵器中的卵孢子萌发形成的单卵孢株,还是由具围生雄器的藏卵器中卵孢子萌发形成的单卵孢株,其有性器官均具有雄器侧生与雄器围生两种类型,但侧生比率在各代菌株间有很大差异。S1代单卵孢株的雄器侧生比率分布范围为1．0％～79．0％,其中具侧生雄器的藏卵器中卵孢子萌发形成的单卵孢株雄器侧生比率较高（分布范围7．0％～79．0％,平均33．6％）,而来自具围生雄器的藏卵器中的卵孢子萌发形成的单卵孢株雄器侧生比率较低（分布范围1．0％～32．0％,平均11．52％）。S2代单卵孢株的雄器侧生比率为10％～9     

Biological Control of Oomycete Soilborne Diseases Caused by Phytophthora capsici,Phytophthora infestans,and Phytophthora nicotianae in Solanaceous Crops

Oomycete pathogens that belong to the genus Phytophthora cause devastating diseases in solanaceous crops such as pepper, potato, and tobacco, resulting in crop production losses worldwide. Although the application of fungicides efficiently controls these diseases, it has been shown to trigger negative side effects such as environmental pollution, phytotoxicity, and fungicide resistance in plant pathogens. Therefore, biological control of Phytophthora-induced diseases was proposed as an environmentally sound alternative to conventional chemical control. In this review, progress on biological control of the soilborne oomycete plant pathogens, Phytophthora capsici, Phytophthora infestans, and Phytophthora nicotianae, infecting pepper, potato, and tobacco is described. Bacterial (e.g., Acinetobacter, Bacillus, Chryseobacterium, Paenibacillus, Pseudomonas, and Streptomyces) and fungal (e.g., Trichoderma and arbuscular mycorrhizal fungi) agents, and yeasts (e.g., Aureobasidium, Curvibasidium, and Metschnikowia) have been reported as successful biocontrol agents of Phytophthora pathogens. These microorganisms antagonize Phytophthora spp. via antimicrobial compounds with inhibitory activities against mycelial growth, sporulation, and zoospore germination. They also trigger plant immunity-inducing systemic resistance via several pathways, resulting in enhanced defense responses in their hosts. Along with plant protection, some of the microorganisms promote plant growth, thereby enhancing their beneficial relations with host plants. Although the beneficial effects of the biocontrol microorganisms are acceptable, single applications of antagonistic microorganisms tend to lack consistent efficacy compared with chemical analogues. Therefore, strategies to improve the biocontrol performance of these prominent antagonists are also discussed in this review.     

A Survey of Phytophthora Species on Hainan Island of South China

During the period 1997–2007, a comprehensive study of the occurrence and distribution of Phytophthora species was conducted on Hainan Island of South China. To date, 14 species of Phytophthora have been recovered and their distribution determined. Phytophthora nicotianae ( =P. parasitica ) is the most important species attacking a wide variety of crops, followed by Phytophthora capsici and Phytophthora citrophthora . In contrast to Phytophthora colocasiae attacking taro leaves throughout the entire island, Phytophthora cyperi was found only once on Digitaria ciliaris in Danzhou. It is of interest to note that Phytophthora heveae, Phytophthora katsurae and Phytophthora insolita are commonly found in forest soil/water of protected mountains without causing any plant diseases. Although Phytophthora species are usually terrestrial or found in fresh water, one isolate of Phytophthora resembling closely the asexual isolates of P. insolita in Hainan was obtained from decaying Rhizophora leaves submerged in seawater. An unidentified Phytophthora species producing non-papillate; internally proliferating sporangia was isolated from the soil in which Ceriops tagel and Bruguiera serangula were growing in a salt water shrimp farm.