植物寄生线虫效应子研究进展

植物寄生线虫是一类专性活体营养寄生物,在植物根部维管束细胞附近诱导形成取食位点,与植物形成稳定的寄生关系,严重影响植物的生长发育,最终导致植物大幅度减产,甚至绝收。为长期、有效地防控植物寄生线虫,对线虫寄生、致病机制及其与植物的相互作用模式进行研究显得尤为重要。效应子在线虫整个寄生阶段中起着关键作用。近些年,线虫效应子的鉴定、功能、线虫效应子与植物相互作用模式等方面的研究取得了重要进展。本文主要针对病原物与寄主植物的相互作用模型、植物寄生线虫效应子鉴定、功能及线虫效应子与寄主植物的相互作用等方面的研究进展进行简单概述。     

禾谷炭疽菌CFEM效应子的生物信息学鉴定与转录分析

植物病原真菌效应子是指可以改变寄主植物细胞结构或者细胞功能的分泌蛋白或其他小分子物质。效应子对病原真菌的侵入、扩展以及致病发挥着至关重要的作用,是植物病原真菌与寄主的互作不断演化的必然结果。真菌特有的CFEM(common in several fungal extracellular membrane protein)蛋白对于病原真菌的致病性起重要作用,一些能够被分泌到胞外的CFEM蛋白被证明是病原真菌效应子。由禾谷炭疽菌(Colletotrichum graminicola)引起的玉米炭疽病是玉米上的重要病害,常年造成严重损失。本研究运用生物信息学工具对禾谷炭疽菌中的CFEM蛋白进行信号肽分析和亚细胞定位分析,进而通过转录分析明确禾谷炭疽菌CFEM蛋白的表达时期。分析结果表明,该病原真菌编码32个CFEM蛋白,其中22个具有信号肽并可分泌至胞外,为分泌蛋白。转录分析表明,10个CFEM分泌蛋白于病菌侵染时附着胞形成期表达,2个CFEM分泌蛋白于侵染后的活体寄生阶段表达,1个于死体寄生阶段表达,其余9个CFEM分泌蛋白在病菌侵染时期的3个阶段均稳定表达。结合生物信息学和转录分析结果,我们预测这22个CFEM分泌蛋白为禾谷炭疽菌致病相关的效应子(简称CFEM效应子)。明确禾谷炭疽菌中CFEM蛋白数量,预测病菌致病相关的CFEM效应子组成,可为开展病原真菌CFEM蛋白介导的病菌—寄主互作研究奠定基础,并为玉米炭疽病的防治和抗性育种研究提供参考。     

植物寄生线虫分支酸变位酶基因的研究进展

 植物寄生线虫食道腺中表达的寄生基因编码的分泌蛋白在线虫侵入寄主植物、建立取食位点和抑制寄主的防御反应过程中起重要作用。利用植物寄生线虫食道腺削减cDNA文库及基于同源克隆等方法,鉴定了这些过程中起作用的分支酸变位酶(CM)基因。带有氨基酸末端信号肽的根结线虫CM、孢囊线虫CM与细菌CM的蛋白质序列非常相似。mRNA原位杂交表明,CM基因专门存在于植物寄生线虫的亚腹食道腺中。RT-PCR分析表明,它们的转录丰度在线虫寄生的早期丰度较高,在后期较低或者难以检测到。Southern杂交表明,这些CM基因为多基因家族。CM的蛋白质在专性内寄生线虫中广泛存在,表明这种多功能的酶在控制线虫侵染植物的过程中起重要作用。     

试论病原物的致病基因

 致病基因(pathogenicity genes)是病原物中决定对植物致病性的有关基因。在侵染植物过程中,病原物致病基因决定了与植物建立寄生关系和破坏植物正常生理功能,调控了对植物趋性、吸附、侵入、定殖、扩展、破坏寄主和显症等病理学过程。     

植物病原真菌的血清学研究进展

植物病原真菌的血清学研究进展华中农业大学生命科学院（４３００７０）陈福生罗信昌湖北省植保总站（４３００７０）周文科血清学技术最早是用于医学病原菌的分类鉴定和诊断。７０年代初期Ｃｌａｒｋ和Ａｄａｍｓ将酶联免疫吸附检测（Ｅｎｚｙｍｅ－ｌｉｎｋｅｄＩｍ－ｍ...     

重要植物寄生线虫内切葡聚糖酶基因研究进展

 在线虫与植物互作的过程中,降解寄主细胞壁是植物线虫成功建立寄生关系的关键环节。β-1,4-内切葡聚糖酶(β-1,4-endoglucanase,ENG)是由线虫食道腺细胞产生并由口针分泌、对细胞壁降解起关键作用的酶类之一。本文对近年来植物寄生线虫eng基因的克隆、组织定位、表达分析、基因、编码蛋白的结构功能以及ENG来源、进化及在线虫与植物互作中的潜在作用等进行了概述。     

中草药水剂LS-1对十种植物病原真菌的抑菌测定

目前关于植物源杀虫剂的研究已很深入,但植物源杀菌剂的研究尚处初级阶段。本课题从上百种植物中筛选出一些具有抑菌效果的植物,然后根据中药配伍原理进行合理配比,研制成植物源杀菌剂ＬＳ 1。本文报道ＬＳ 1对10种病原真菌的室内抑菌效果。1　材料与方法1.1　供试菌　禾生腐霉菌(Ｐｙｔｈｉｕｍｇｒａｍｉｎｉｃｏｌａ)、立枯丝核菌(Ｒｈｉｚｏｃｔｏｎｉａｓｏｌａｎｉ)、禾谷镰刀菌(Ｆｕｓａｒｉｕｍｇｒａｍｉｎｅａｒｕｍ)和小麦纹枯病菌(Ｒ.ｓｏｌａｎｉ)ＡＧ 4由中国科学院微生物所提供;南瓜疫病菌(Ｐｈｙｔｏｐｈｔｈｏｒａｃａｐｓ…     

草莓胶孢炭疽菌CFEM候选效应子的生物信息学鉴定及其侵染过程中的转录分析

病原真菌通常分泌效应子到寄主组织中调控寄主的生理过程,从而有利于其侵染。CFEM(common in several fungal extracellular membrane)蛋白是真菌所独有的,且与致病性密切相关。本研究利用Pfam数据库对草莓胶孢炭疽菌全基因组进行搜索,鉴定获得22个CFEM蛋白。对CFEM蛋白的信号肽、跨膜结构域和亚细胞定位进行分析,结果表明仅有8个CFEM蛋白为分泌蛋白。对CFEM分泌蛋白在不同侵染阶段的转录情况进行转录组学及RT-PCR分析,结果显示8个CFEM蛋白在侵染后不同时期均有表达。其中,1个CFEM分泌蛋白于附着胞形成期特异表达,2个于活体寄生阶段特异表达,2个于死体寄生阶段特异表达。综合上述分析结果,预测这8个分泌蛋白可能为草莓胶孢炭疽菌的效应子。本研究为深入解析植物病原真菌CFEM效应子提供了理论依据。     

真菌源蛋白类激发子及其转基因植物研究进展

真菌源蛋白类激发子是广泛存在于植物病原真菌的信号传导分子,不仅在植物与病原菌互作中起重要作用,而且还具有广谱诱导植物抗性和促进植物生长的功能。本文就真菌源蛋白类激发子的种类、功能及蛋白激发子基因转化植物等研究进行了综述。     

植物抗病基因工程研究进展（Ⅰ）──植物抗病毒基因工程

植物病原真菌、细菌、病毒常年危害农作物，给农业生产带来巨大损失，而防治病害的最有效措施是抗病育种。生物技术的发展给抗病育种开辟了新途径。本文分三大部分综述了国内外抗病毒病害、抗细菌病害、抗真菌病害基因工程的研究进展，包括转病毒基因成分（ＣＰ基因、复制酶基因、反义ＲＮＡ、ｓａｔＲＮＡ），转植物抗生物质编码基因和转核酶基因的抗病毒病植株；转溶菌酶基因、转抗菌肽基因、转解除细菌毒素基因的抗细菌病害植株；转芪合成酶基因、转植保素和木质素的关键合成酶基因、转几丁质酶基因、转葡聚糖基因、转ＲＩＰ基因的抗真菌病害植株等研究所取得的成果。     

葡萄座腔菌(Botryosphaeria dothidea)全基因组分泌蛋白的预测及功能分析

 葡萄座腔菌(Botryosphaeria dothidea)是引起多种果树和林木枝干溃疡病和果实腐烂病的重要病原菌。为明确葡萄座腔菌的致病机理,采用SignalP、WoLF PSORT、TMHMM、GPI-SOM等软件,对葡萄座腔菌全基因组14 998条蛋白序列进行了分泌蛋白预测和功能分析。结果表明B.dothidea全基因组编码蛋白中有851条序列具有典型的分泌蛋白特征,占蛋白总数的5.67%,其长度主要分布在100~700个氨基酸范围内。信号肽的分析结果表明,以19个氨基酸为信号肽的分泌蛋白数目最多,非极性氨基酸丙氨酸(A)在信号肽中的使用频率最高,而有带电侧链的天冬氨酸(D)和谷氨酸(E)使用频率最低,信号肽-3和-1位置的氨基酸相对保守,其切割位点属于典型的A-X-A型。对分泌蛋白功能预测结果表明,578个分泌蛋白获得了功能注释,其功能主要涉及碳水化合物的运输、代谢过程,蛋白翻译后修饰和氨基酸代谢、运输过程。分泌蛋白中效应蛋白(effector)预测结果表明,B. dothidea分泌蛋白中共有119个潜在的效应蛋白,其中11个可被PHI数据库注释到,其与引起其他植物病原菌致病力变化的致病效应蛋白具有较高的相似性。碳水化合物活性酶类(CAZymes)的预测结果表明,B. dothidea分泌蛋白组中共有279个CAZymes,其中GHs家族最多。这些结果的获得为今后进一步筛选B. dothidea的效应蛋白,明确B. dothidea的致病机理,以及筛选寄主新的抗性基因提供了必要的基础。     

The secreted proteome of necrotrophic Ciborinia camelliae causes nonhost-specific virulence

Ciborinia camelliae (Sclerotiniaceae) is a host- and organ-specific fungal pathogen that causes rapid browning and flower drop on ornamental plants of the genus Camellia. To determine the nature of its necrotrophic factors, we tested whether proteins secreted by C. camelliae can damage host-plant tissues. Fungal culture filtrate caused necrogenic activity, abolished by heat or protease treatments, thus indicating that the secreted necrogenic agents are probably proteinaceous in nature. Mass spectrometry-based proteomics was used to detect and identify secreted proteins of C. camelliae. Proteins secreted in culture media (in vitro) and in petal apoplast (in planta) had similar functional distributions, and key identified proteins included homologs to known virulence factors of the closely related Sclerotiniaceae fungus Botrytis cinerea, including endopolygalacturonases, cerato-platanin family proteins, and necrosis- and ethylene-inducing peptides. The main class of secreted proteins were carbohydrate-active enzymes, a characteristic signature of necrotrophic plant pathogens. Both fungal culture filtrate and apoplastic washes of infected petals induced necrosis when infiltrated into host and nonhost plants. This suggests that while some of the secreted proteins might contribute to virulence of C. camelliae, and can cause necrosis similar to secreted proteins of broad-host Sclerotiniaceae pathogens, they do not have a role in determining its host specificity.     

Localized hemibiotrophy in Colletotrichum: cytological and molecular taxonomic similarities among C. destructivum, C. linicola and C. truncatum

The infection process of hemibiotrophic isolates of Colletotrichum linicola (from flax, Linum usitatissimum ) and C . truncatum (from broad bean, Vicia faba and lentil, Lens culinaris ) was studied by light microscopy. Host surfaces were penetrated directly leading to a symptomless, biotrophic phase characterized by the elaboration of large multilobed, multiseptate, vesicular primary hyphae that were restricted to the initially infected epidermal cells. Biotrophy lasted for the first 48 h of the host-pathogen interaction and was rapidly succeeded by a necrotrophic phase during which narrow, secondary hyphae invaded the surrounding leaf tissues and water-soaked spreading lesions with sporulating, monosetate acervuli were produced on infected host surfaces. Molecular taxonomic analysis of the nucleotide sequences of the amplified D2 and ITS-2 regions of rDNA revealed very close similarities (97–99%) between these isolates and those of C . destructivum obtained from cowpea ( Vigna unguiculata ) and lucerne ( Medicago sativa ), and also of C . truncatum obtained from pea ( Pisum sativum ). This association was consistent with results from a comparative assessment of some in-planta and in-vitro morphological and growth characteristics of these hemibiotrophic fungi. It was concluded that localized hemibiotrophy is an infection strategy utilized predominantly by a closely-related group of pathogens comprising C . destructivum , C . linicola and C. truncatum , and the formation of multilobed primary hyphae restricted to the first penetrated cell might therefore be a key taxonomic character which correlates consistently with ITS sequence data.     

Dynamic changes in pod and fungal physiology associated with the shift from biotrophy to necrotrophy during the infection of Theobroma cacao by Moniliophthora roreri

Theobroma cacao pods were inoculated with meiospores of Moniliophthora roreri (Mr), a hemibiotrophic basidiomycete causing frosty pod rot. Pods were malformed 30 days after inoculation (DAI) and sporulation was observed 60 DAI. Glucose and asparagine concentrations decreased and mannitol and malonate increased in infected pods 30 DAI. By 60 DAI, most carbohydrates, amino acids, and organic acids were drastically reduced by infection. Mannitol and succinic acid levels increased 60 DAI and likely originated from Mr. RT-qPCR analysis of cacao ESTs indicated a strong response to infection 30 DAI in malformed pod. Evidence indicated that biotrophic hyphae colonized pods and a shift to necrotrophic growth occurred later (during the end stages of infection). Expression of cacao ESTs associated with plant hormone biosynthesis and action was altered. Changes in the expression of Mr ESTs in response to nutrient deficiency in pure culture were small. Changes in Mr gene expression patterns and levels of specific metabolites in necrotic sporulating pods 60 DAI compared to malformed pods 30 DAI indicated that the glyoxylate cycle of Mr was up regulated during the shift from biotrophic to necrotrophic phases of the disease cycle.     

Effects of the saprophytic leaf mycoflora on growth and productivity of winter wheat

The effects of the saprophytic mycoflora and its interference with cereal aphids on growth and yield of winter and spring wheat was studied in field experiments in 1980, 1981 and 1982.Yields varied between 5000 and 8000 kg dry matter of kernels per ha. The effect of the saprophytic mycoflora on yield was determined in different treatments: A) no control measures against cereal aphids and saprophytic and necrotrophic fungi, B) no control of cereal aphids, control of saprophytic and necrotrophic fungi, C) control of cereal aphids and control of saprophytic and necrotrophic fungi, D) control of cereal aphids and stimulation of saprophytic mycoflora and E) control of cereal aphids, no control of saprophytic and necrotrophic fungi nor stimulation of saprophytic mycoflora.Considerable differences in top densities of saprophytic mycoflora (10 times as large in A and D as in B and C) were determined. The consequences of these differences for the growth and productivity of wheat were minor. A negative effect of saprophytic mycoflora on the yield could not be detected in 1981 and 1982, whereas a small positive significant effect was found in 1980. This stimulation may have been due to competition between necrotrophic fungal pathogens and saprophytic mycoflora. As a result of favourable weather conditions necrotrophic fungal pathogens were very numerous in 1980 and could form an important yield reducing factor. Yield levels may effect the importance of the necrotrophic and saprophytic mycoflora as yield reducing factors. Additionally, in the presence of aphid honeydew captafol was found to be relatively ineffective against saprophytic fungi.     

Class I hydrophobin BcHpb1 is important for adhesion but not for later infection of <Emphasis Type="Italic">Botrytis cinerea</Emphasis>

Hydrophobins are small secreted proteins unique to filamentous fungi. In this study, we cloned and characterized the class I hydrophobin gene BcHpb1 in the necrotrophic pathogen Botrytis cinerea. The BcHpb1 protein consisted of 117 amino acids. Similar to class I hydrophobins from other fungi, BcHpb1 contains eight conserved cysteine residues. The hydropathy plot of the BcHpb1 amino acid sequence was characteristic of a class I hydrophobin. These results indicated that the BcHpb1 gene encodes a class I hydrophobin. Vegetative growth of ΔBcHpb1 strains, null mutants of BcHpb1, was similar to that of the wild-type strain as were the conidiophores, conidia, appressoria and virulence on host plants. However, adherence of ΔBcHpb1 strains to hydrophobic surfaces was greatly reduced, implying that BcHpb1 is important for the hydrophobicity of conidia and that BcHpb1 may be required to adhere to plant surfaces under certain environmental conditions.     

Development of the infection strategy of the hemibiotrophic plant pathogen,Colletotrichum orbiculare,and plant immunity

The hemibiotrophic fungus Colletotrichum orbiculare forms appressoria as infection structures and primarily establishes biotrophic infection in cucumber epidermal cells. Subsequently, it develops necrotrophic infection. In the pre-invasion stage, morphogenesis of appressoria of C. orbiculare is triggered by signals from the plant surface. We found that C. orbiculare PAG1 (Perish-in-the-Absence-of-GYP1), a component of MOR [morphogenesis-related NDR (nuclear Dbf2-related) kinase network] plays an essential role as a key component of the plant-specific signaling pathway and that hydrolysis of cutin by a spore surface esterase creates a cutin monomer that constitutes a key plant-derived signal. Development of the infection structure of C. orbiculare is strictly regulated by the cell cycle and we found that proper regulation of G1/S progression via two-component GAP genes, consisting of BUB2 (Budding-Uninhibited-by-Benomyl-2) and BFA1 (Byr-Four-Alike-1) is essential for the establishment of successful infection. In the post-invasion stage, the establishment of the biotrophic phase of hemibiotrophic fungi is crucial for successful infection. We found that C. orbiculare WHI2 (WHIsky-2), an Saccharomyces cerevisiae stress regulator homolog, is involved in the phase transition from biotrophy to necrotrophy through TOR (Target of Rapamycin) signaling, and is thus essential for full pathogenesis.     

灰霉菌(Botrytis cinerea)采后致病性研究

 灰霉菌(Botrytis cinerea)是引起葡萄采后病害的主要病原真菌之一。葡萄灰霉菌可在田间潜伏侵染,采后由健康果实携带进入销售市场,该菌的显著致病症状为果实软腐和脱落。灰霉菌与葡萄的其它采后致病菌,如链格孢菌(Alternaria alternata)、镰刀菌(Fusarium sp.)、芽枝霉(Cladosporium sp.)、青霉菌(Penicillium sp.)、黑曲霉(Aspergillus nigar)和粉红单端孢菌(Trichothecium roseum)相比,不仅表现出明显的潜伏侵染优势,而且具有较强的低温(4℃)条件下的致病优势。4℃低温下灰霉菌在寄主葡萄体外和体内分泌多聚半乳糖醛酸酶(PG)活力均显著高于以上各菌,而在25℃下无显著差异,这一结果与该2种温度下灰霉菌接种果实后的症状表现一致。     

高温处理西花蓟马若虫对其雌成虫寿命、繁殖力及后代发育的影响

温度是影响昆虫生长发育的重要生态因素之一,高温逆境作为一种绿色防控手段可以有效控制温室害虫种群发展。为了探究高温处理西花蓟马若虫对其种群发展的影响,本试验用41℃和43℃两个温度分别处理西花蓟马初孵若虫2、6、12、24和36h后,观察并记录其雌成虫寿命、繁殖力及后代发育指标的变化情况。结果表明,随着处理时间的延长,两个高温处理后西花蓟马雌成虫寿命,子代若虫总数、成虫总数和总存活率(若虫发育到成虫的存活率)均显著下降,后代雌雄性比呈总体下降趋势。41℃处理后,性比从2.30∶1降低到2.13∶1;43℃处理后从2.25∶1降低到2.07∶1,均明显低于对照的2.69∶1。另外,两性生殖种群相比于孤雌生殖种群更易遭受高温处理的影响。