小麦赤霉病镰孢菌毒素的生物合成及其分子调控

禾谷镰刀菌是小麦赤霉病的主要致病菌,其真菌次生代谢产生的单端孢霉烯类B型毒素,如雪腐镰刀菌烯醇(nivalenol,NIV)、脱氧雪腐镰刀菌烯醇(deoxynivalenol,DON)和其它乙酰化衍生物等污染小麦籽粒后对人畜健康构成威胁。综述了近年来国内外对小麦赤霉病镰孢菌单端孢霉烯类B型毒素生物合成的主要途径及分子调控研究进展,对毒素合成过程中的重要调控基因如TRI5、TRI7和TRI13在农业中的应用进行了阐述。     

DON生物合成的亚细胞定位和精准外排研究进展

单端孢霉烯B族毒素脱氧雪腐镰刀烯醇（deoxinivalenol, DON）是产毒镰刀菌在侵染小麦等作物过程中的一类重要的致病因子,可以帮助产毒镰刀菌在麦穗间扩展。DON会抑制蛋白质合成,对动物、微生物和寄主具有毒性（cytotoxicity and phytotoxicity）,然而产毒镰刀菌自身借助何种保护机制免受DON毒害目前研究甚少。DON毒害机制的研究对于镰刀菌毒素的持续防控和粮食安全、人民生命健康保障具有重要意义。综述了产毒镰刀菌DON合成解毒机制的最新研究进展,主要包括DON合成的亚细胞定位、合成基因簇内的外排蛋白和解毒基因作用方式,以期为有针对性地破解其解毒机制,设计研发高效靶向控毒技术的相关研究提供参考。     

麦类赤霉病菌毒素的研究II．禾谷镰刀菌的固体、液体培养产毒研究

自然病麦中分离、鉴定的禾谷镰刀菌(Fusarium gramincarum)FG l接种在固体、液体培养基中,培养物的粗捉取掖不但引起试验鸽子的强烈呕咀,还能抑制婉豆种了的山荣。了度温条件下固体培养,菌株的产毒能力增加。液体培养则以在S·P·M·中的培养物显示的生物毒性最蛆。从液体培养物中提取的粗制毒素是一个多组分化合物,柱层层析得到的6个组分中以0,值0．53部分生物毒性最强,与已知的赤霉病麦毒紊I(脱氧雪腐镰刀菌烯醇)、11(3～乳酰氧基脱氧雪腐镰刀菌烯醇)和赤霉烯酮比较,显示不同的层析现象。根据毒素的生物特性,它属于单端孢霉烯族毒素中的一个未知成员。     

中国小麦赤霉病菌优势种—禾谷镰刀菌产毒素能力的研究

试验测定了分离自中国小麦赤霉病常发生地区病麦穗上的４７个禾谷镰刀菌（Ｆｕｓａｒｉｕｍ ｇｒａｍｉｎｅａｒｕｍ）菌株的产毒素能力。结果表明,它们可以产生２５种包括 单端孢霉烯族化合物（Ｔｒｉｃｈｏｔｈｅｃｅｎｅｓ）、倍半萜类化合物（Ｓｅｓｑｕｉｔｅｒｐｅｎｅｓ）、赤霉烯酮（Ｚｅａｒａｌｅｎｏｎｅ）和丁烯羟酸内酯（Ｂｕｔｅｎｏｌｉｄｅ）等类的已知次生代谢物。这些菌株属于化学型Ｉ,其中,来自我国温暖麦区的     

禾谷镰孢菌高产毒菌株的构建

为研究禾谷镰孢菌Fusarium graminearum Schw.单端孢霉烯族毒素生物合成基因（产毒基因）在寄主体内的表达,作者构建了带报告基因GUS（β-葡糖苷酸酶基因）的质粒pGUSTRI6P5,并通过对野生型菌株的转化获得禾谷镰孢高产毒菌株,该质粒含有由TRI5（禾谷镰隐单端孢霉的二烯合酶基因）启动子（TRI5 Prom)驱动的GUS基因编码区、潮霉素B抗性基因和拟枝孢镰孢F.sporotrichioides的产毒调控基因TRI6（FSTR16）。用pGUSTRI6P5转化野生型菌株GZ3639后,在含潮霉素B的培养基上选取抗性菌落。单孢分离获单孢菌株（转化子）。在GYEP（葡萄糖-酵母粉-蛋白胨）液体培养基上,转化子B4-1和B16-1的GUS比活力强,15-AcDON(15-乙酰脱氧雪腐镰鼠菌烯醇）产量高,且两者呈正相关（相关系数（r)分别为0.9839和0.9523)。B41-1和B16-1两个转化子可作为研究禾谷镰孢与其寄主相互作用的工具菌株。     

三隔镰刀菌大米培养物产生二苯甲醇的研究

<正> T-2毒素是镰刀菌属三隔镰刀菌Fusarium tricinctum(Corda)Sacc.的一种次生代谢产物,隶属单端孢霉烯簇毒素。发霉大米中毒症及豆荚中毒症等均与单端孢霉烯簇毒素有关。张树荣等从真菌培养物M-20中提取T-2毒素成功,我们对其     

禾谷镰孢菌高产毒菌株的构建*

为研究禾谷镰孢菌Fusarium graminearum Schw.单端孢霉烯族毒素生物合成基因（产毒基因）在寄主体内的表达,作者构建了带报告基因GUS（(-葡糖苷酸酶基因）的质粒pGUSTRI6P5,并通过对野生型菌株的转化获得禾谷镰孢高产毒菌株。该质粒含有由TRI5（禾谷镰孢单端孢霉二烯合酶基因）启动子（TRI5 Prom）驱动的GUS基因编码区、潮霉素B抗性基因和拟枝孢镰孢F. sporotrichioides的产毒调控基因TRI6（FSTRI6）。用pGUSTRI6P5转化野生型菌株GZ3639后,在含潮霉素 B的培养基上选取抗性菌落,单孢分离获单孢菌株（转化子）。在GYEP（葡萄糖-酵母粉-蛋白胨）液体培养基上,转化子B4-1和B16-1的GUS比活力强,15-AcDON（15-乙酰脱氧雪腐镰刀菌烯醇）产量高,且两者呈正相关（相关系数（r）分别为0.9839和0.9523）。B4-1和B16-1两个转化子可作为研究禾谷镰孢与其寄主相互作用的工具菌株。     

镰刀菌毒素的研究 Ⅰ.T-2毒素产生菌株的筛选

<正> 镰刀菌属(Fusarium)是危害性最大的真菌之一,不仅侵染田间作物和库贮谷物,造成经济损失,而且产生毒素,引起人畜镰刀菌毒素中毒。黄曲霉毒素的致癌作用被证实后,更引起世界各国对镰刀菌毒素及其中毒现象的重视(孟昭赫等,1979)。镰刀菌产生的毒素主要是单端孢霉烯族化合物,其中 T-2毒素的毒性最强(Ronald,1981),它能引起兔子皮肤反应和抑制植物种子发芽等生物毒性效应(Burme-istes,1970;Ueno,1971)。国内尚未见人工培养产生 T-2毒素的报告。为给农业、食品卫生和环境保护等方面研究提供科学依据,本文报告 T-2毒素产生菌株的筛选。     

禾谷镰刀菌蛋白激酶研究进展

由禾谷镰刀菌引起的小麦赤霉病是小麦生产最重要的真菌病害之一,除了造成严重的产量损失外,其病原菌还会产生多种真菌毒素危害人畜健康。蛋白激酶在禾谷镰刀菌生长发育、植物侵染和胁迫应答等方面具有重要作用。综述了禾谷镰刀菌主要蛋白激酶在生物学功能和分子作用机制等方面的研究进展,并对未来禾谷镰刀菌蛋白激酶的研究趋势进行了展望,以期为今后禾谷镰刀菌蛋白激酶的研究与小麦赤霉病的防治提供理论参考。     

一种新的霉菌毒素——CBD2的分离和结构研究

从田间感染禾谷镰刀菌(Fusarium graminearum Schw.)的赤霉病大麦中分离得到一种新的霉菌毒素——赤霉病麦毒素II(CBD2)。通过光谱分析和化学反应研究了它的结构,初步确定该毒素为3一乳酰氧基一7,1 5一二羟基一12,13一环氧单端孢霉素一9一烯一8一酮。该毒素可抑制豌豆种子发芽并引起兔子皮肤发炎。     

Potential of fungal antagonists for biocontrol of Fusarium spp. in wheat and maize through competition in crop debris

Pathogenic Fusarium spp. cause head blight in wheat or ear rot in maize leading to yield losses and also a reduction in quality due to mycotoxin contamination of the grain. Infected crop residues are the main inoculum source for epidemics. Saprophytic fungi, obtained from cereal tissues or necrotic tissues of other crops, were screened for their ability to colonise wheat straw and maize stalks and to suppress sporulation of pathogenic Fusarium spp. Results of bio-assays conducted under controlled conditions were variable among Fusarium spp. and host substrates for most antagonists tested, such as yeasts, Trichoderma spp. and non-pathogenic Fusarium spp. Isolates of Clonostachys rosea consistently suppressed sporulation of F. culmorum and F. graminearum on wheat straw, and of F. culmorum, F. graminearum, F. proliferatum and F. verticillioides on maize stalks. Isolates of C. rosea, C. cladosporioides and F. equiseti were applied to pieces of maize stalks or flowering ears in preliminary experiments conducted under field conditions. The colonisation of stalk pieces by pathogenic Fusarium spp. was assessed after 9 months. Colonisation of stalk pieces by pathogenic Fusarium spp. was significantly reduced at several sampling dates. However, results obtained with the antagonists were not consistent for all sampling dates and between experiments.     

小麦抗赤霉病利器——他山之石

赤霉病是我国乃至世界小麦(Triticum aestivum)产区的重要病害, 给农业生产和人畜健康造成重大威胁。分离鉴定优质抗病基因、培育抗病品种, 是控制我国麦区赤霉病的重要手段。最近, 山东农业大学孔令让团队完成了二倍体长穗偃麦草(Thinopyrum elongatum)基因组的组装, 并在此基础上通过精细定位和图位克隆分离得到来自长穗偃麦草的抗赤霉病基因Fhb7。他们发现Fhb7编码1个谷胱甘肽转移酶, 对禾谷镰孢菌(Fusarium graminearum)分泌的包括呕吐毒素等在内的多种毒素具有解毒作用, 是1个广谱持久抗病基因。他们还发现Fhb7很可能最初源于内生真菌, 经过基因水平转移进入到偃麦草基因组中。此外, Fhb7不影响其它农艺性状, 且其抗性不受小麦遗传背景影响。这一系列工作揭示了作物抗病演化中的全新机制, 对小麦抗赤霉病育种以及更好地利用长穗偃麦草的丰富基因资源都具有重要意义。     

蛋白激酶FgBud32参与禾谷镰刀菌的生长发育、致病和胁迫应答

由禾谷镰刀菌引起的小麦赤霉病是一种毁灭性的小麦真菌病害,在世界范围内造成小麦产量和质量的巨大损失。实验室前期在禾谷镰刀菌中共鉴定到116个蛋白激酶,其中FgBUD32基因的缺失会造成营养生长和有性生殖方面的严重缺陷,但其在禾谷镰刀菌中的详细功能尚未报道。本研究通过系统比较Fgbud32突变体与野生型PH-1及互补菌株的表型差异,对FgBud32在禾谷镰刀菌中的生物学功能进行了解析。研究结果显示Fgbud32突变体在多个表型方面存在缺陷,与野生型菌株以及互补菌株相比,其生长速率急剧下降,菌丝弯曲且分支减少;分生孢子的产量显著降低,形态变短,隔膜减少,萌发率降低且萌发速率延迟;在有性生殖时期不能产生子囊壳或子囊壳前体;对小麦穗和胚芽鞘的致病力以及DON毒素的合成能力均显著下降。进一步胁迫试验表明,FgBUD32基因的缺失导致禾谷镰刀菌对氧化胁迫(H₂O₂)以及DNA损伤胁迫(羟基脲和甲磺甲酯)的敏感性增加。此外,我们还发现FgBud32在细胞核和细胞质中均有定位,且在一定时期或条件下会从细胞质向细胞核内聚集。综上所述,FgBUD32基因参与了禾谷镰刀菌的营养生长、极性生长、无性/有性生殖、DON毒素合成、致病以及对氧化胁迫和DNA损伤胁迫的应答等多种生命活动,但其具体的作用机制还有待深入研究。     

Potential for control of seedling blight of wheat caused by Fusarium graminearum and related species using the bacterial endophyte Bacillus mojavensis

Fusarium-infected wheat seed decreases germination, seedling emergence, and causes post emergence seedling death, and can contribute to wheat scab and ear rot of maize, with consequent production of mycotoxins such as deoxynivalenol and zearalenone. Current seed treatments have proved ineffective in controlling seedling blight and scab. A patented endophytic bacterial strain, Bacillus mojavensis RRC 101, and several other strains of this species were studied to determine in vitro antagonism to some Fusarium species and to assess the potential of this bacterium to serve as an endophytic biocontrol for seedling blight of wheat produced by species within the F. graminearum complex, as well as other species of Fusarium. Seedling emergence and seed germination were two tests used as indicators of seedling blight. These tests were conducted in growth rooms with two wheat cultivars highly susceptible to scab, Norm and Pioneer 2552, and other cultivars with varying resistance to scab. The results indicated that all strains of this bacterium were antagonistic in vitro to the strains of F. graminearum and its seven related species, as well as four strains of F. pseudograminearum and the two strains of F. verticillioides. Germination of the highly scab susceptible cultivar 2552 was increased from 77 to 97% when planted in soil containing a mixed inoculum of F. graminearum and related species. Seedling emergence in the very susceptible wheat cultivar Norm increased from 20 to 82% when treated with the bacterium. The data indicated that inoculating wheat kernels with B. mojavensis reduced seedling blight of wheat produced by F. graminearum and related Fusarium species indicating the potential for this bacterium as a biocontrol under field condition.     

小麦响应禾谷镰刀菌侵染的转录组学研究进展

由禾谷镰刀菌引起的小麦赤霉病直接为害作物穗部,不仅严重影响小麦产量,还可因为毒素污染问题威胁人畜健康。近年来对小麦与禾谷镰刀菌互作的转录组学研究带来了很多新见解,概述了小麦响应禾谷镰刀菌侵染的转录组学研究进展,主要比较了不同抗性品种、不同器官、不同籽粒发育时期的小麦穗部在禾谷镰刀菌侵染时的基因表达特征,总结了赤霉病感染时小麦的激素响应、信号传导、转录调控和防卫相关基因的表达规律,以期促进研究者对小麦响应禾谷镰刀菌侵染规律的理解。     

禾谷镰孢菌β2微管蛋白与苯并咪唑类杀菌剂互作研究

小麦赤霉病是小麦上的主要病害之一,在全世界范围内引起该病害的致病菌主要是禾谷镰孢菌Fusarium graminearum。目前,使用杀菌剂是生产上防治小麦赤霉病发生和危害的主要手段,常用的杀菌剂主要有苯并咪唑类杀菌剂(benzimidazoles)等,苯并咪唑类杀菌剂的作用靶标是β2微管蛋白。本研究旨在探究小麦赤霉病菌中β2微管蛋白与苯并咪唑类杀菌剂的互作机制,通过同源建模的方法获得了禾谷镰孢菌β2微管蛋白的三维结构,并在此基础上将β2微管蛋白与4种苯并咪唑类杀菌剂(多菌灵、苯菌灵、噻菌灵、甲基硫菌灵)进行分子对接。分子对接结果显示β2微管蛋白第198位苯丙氨酸和第236位缬氨酸与4种苯并咪唑类杀菌剂直接互作形成氢键,第50、134、165、167、198、200、236、237、239、240、250、253、257、314位氨基酸形成药剂结合口袋。通过比较β2微管蛋白与4种苯并咪唑类杀菌剂结合自由能,发现与其他3种杀菌剂相比,β2微管蛋白与多菌灵的结合自由能最小(-5.72 kcal/mol),说明其与多菌灵互作亲和力更强。采用菌丝生长速率法测定了禾谷镰孢菌对4种苯并咪唑类杀菌剂的EC₅₀值,禾谷镰孢菌对多菌灵、苯菌灵、噻菌灵、甲基硫菌灵的EC₅₀值分别为0.772、0.862、1.088、13.266 mg/L,该结果表明禾谷镰孢菌对多菌灵的敏感性强于其他3种杀菌剂,与分子对接结果相吻合。     

Fusarium mycotoxins: a review of global implications for animal health, welfare and productivity

Trichothecenes, zearalenone (ZEN) and fumonisins are the major Fusarium mycotoxins occurring on a worldwide basis in cereal grains, animal feeds and forages. Other important Fusarium mycotoxins include moniliformin and fusaric acid. Spontaneous outbreaks of Fusarium mycotoxicoses have been recorded in Europe, Asia, New Zealand and South America and, in addition, chronic exposure occurs on a regular and more widespread scale. The metabolism and adverse effects of the Fusarium mycotoxins are considered in this review with particular reference to recent data on specific and proposed syndromes and to interactions among co-occurring mycotoxins. Within the trichothecene group, deoxynivalenol (DON) is associated with emesis, feed refusal and depressed feed intake in pigs, while T-2 toxin and diacetoxyscirpenol (DAS) are now clearly linked with oral lesions in poultry. The gut microflora of farm livestock are able to transform DON to a de-epoxy derivative. In contrast, the ovine metabolism of ZEN results in the production of five metabolites and relatively high levels of these forms may be excreted in the urine as glucuronides. There is now undisputed evidence that ZEN and its metabolites possess estrogenic activity in pigs, cattle and sheep, but T-2 toxin has also been implicated in reproductive disorders in farm livestock. Fumonisins are positively linked with pulmonary edema in pigs, leukoencephalomalacia in equines and with deranged sphingolipid metabolism in these animals. Fusarium mycotoxins have also been provisionally implicated in ovine ill-thrift, acute mortality of poultry and in duodenitis/proximal jejunitis of horses. Several Fusarium mycotoxins may co-occur in a particular feed ingredient or in compound feedingstuffs. In general, combinations of Fusarium mycotoxins result in additive effects, but synergistic and/or potentiating interactions have been observed and are of greater concern in livestock health and productivity. Synergistic effects have been reported between DON and fusaric acid; DON and fumonisin B₁ (FB₁); and DAS and the Aspergillus-derived aflatoxins. Limited evidence of potentiation between FB₁ and DON or T-2 toxin has also emerged recently. Additive and synergistic effects between known and unidentified mycotoxins may account for enhanced adverse effects observed on feeding Fusarium-contaminated diets. The potential for transmission of DON into eggs and of ZEN into porcine kidney and liver has been demonstrated. However, lactational carry-over of FB₁ appears not to occur, at least in cows and sows. It is concluded that livestock health, welfare and productivity may be severely compromised by consumption of DON, T-2 toxin, DAS, ZEN and fumonisins and by interactions among these mycotoxins. Safety of some animal products may also be at risk. Furthermore, in view of the limited options available for remediation, it is concluded that exploitation of crops resistant to Fusarium infection offers the most viable strategy for reducing mycotoxin contamination of grain and animal feed.     

PCR-based strategy to detect contamination with mycotoxigenic Fusarium species in maize

Contamination of cereals with mycotoxigenic species of Fusarium is an important source of trichothecenes, fumonisins and other mycotoxins which cause serious diseases in human and animals. In addition, these species are phytopathogenic and produce severe losses in cereal yield. Methods for early detection of these Fusarium species are crucial to prevent toxins entering the food chain and are a useful tool in disease management practices. We have developed an integrated protocol for diagnosis of mycotoxigenic Fusarium contamination in maize which can also be used for other cereals. The protocol consisted in an easy and rapid DNA extraction from maize samples (grain and germ), and subsequent group-specific polymerase chain reaction (PCR) assays for genus Fusarium, Gibberella fujikuroi complex, and trichothecene-producing species of Fusarium, that orientate the search of the critical species. We have additionally developed a PCR assay for the identification of F. proliferatum. The primers were designed on the basis of IGS sequence (Intergenic Spacer of rDNA), a multi-copy region in the genome that permits to enhance the sensitivity of the assay in comparison with PCR assays based on single-copy sequences. The suitability of the protocol and the relative efficacy of single and multi-copy sequence-based PCR assays have been tested in a wide range of fumonisin-contaminated maize samples.