胶孢炭疽菌G蛋白信号调控因子CgRGS3的生物学功能

G蛋白信号调控因子(regulators of G-protein signaling,RGS)是G蛋白的一类负调控因子,在植物病原真菌生长发育及致病过程中起着重要作用。为探究胶孢炭疽病菌1个RGS基因CgRGS3的生物学功能,利用PCR技术扩增CgRGS3基因并进行生物信息学分析,通过同源重组的方法获得该基因的敲除突变体,并在突变体的基础上获得互补株,通过表型分析确定CgRGS3的生物学功能。结果表明:CgRGS3编码1个含有367个氨基酸的蛋白,包含1个RGS功能域和3个跨膜区域,表型分析发现与野生型菌株相比,敲除突变体营养生长缓慢,分生孢子产量附着胞形成率显著降低,对Cu~(2+)、Fe~(2+)、Zn~(2+)离子更加敏感,细胞壁完整性发生改变,黑色素产量降低及致病力减弱等。由此可见,CgRGS3参与调控胶孢炭疽菌的营养生长,分生孢子产生及附着胞的分化、细胞壁完整性、黑色素产生及致病性等。     

稻瘟病菌CAAX蛋白酶MoRce1的功能研究

 蛋白质的翻译后异戊烯化修饰(CAAX修饰)能够介导真核生物中许多重要蛋白质的亚细胞定位以及蛋白与蛋白间的相互作用。稻瘟病菌(Magnaporthe oryzae)引致的稻瘟病是水稻最重要病害之一,造成全球水稻严重减产。为深入了解稻瘟病菌致病机理,更好地防控稻瘟病,我们研究了异戊烯修饰是否影响稻瘟病菌的生长发育和致病性。首先从稻瘟病菌基因组数据库中鉴定到一个稻瘟病菌的异戊烯蛋白酶MoRce1,同源比对发现MoRce1保守结构域在各物种之间变化较大,猜测在不同物种中该蛋白可能出现了功能分化。经同源重组方法敲除MoRCE1基因,发现MoRCE1缺失突变体在胁迫培养条件下细胞壁完整性明显缺陷,但是对营养生长、产孢、萌发以及致病性没有明显影响,说明MoRce1蛋白可能通过参与稻瘟病菌细胞壁合成相关蛋白的异戊烯化修饰进而影响该菌细胞壁的完整性,其具体机制还有待深入研究。     

禾谷镰刀菌中磷脂酰肌醇转运蛋白功能分析

磷脂酰肌醇转运蛋白(phosphatidylinositol transfer protein,PITP)保守存在于真核生物中,负责膜系统中磷脂酰肌醇和磷脂酰胆碱的转运,参与胞内的脂类信号调控过程。由禾谷镰刀菌(Fusarium graminearum)引起的小麦赤霉病严重威胁粮食产量和食品安全,而在禾谷镰刀菌的研究中发现磷脂信号网络中的关键元件参与病原菌致病过程的调控。我们利用基因敲除的方法对禾谷镰刀菌中PITP家族的功能进行分析,在禾谷镰刀菌中鉴定了7个PITP蛋白的编码基因,并成功获得了其单敲除突变体。通过表型分析发现,这7个基因的单敲除突变体在营养生长、无性及有性生殖和致病性方面与野生型菌株并无明显差异。研究结果表明:1)禾谷镰刀菌中的PITP基因并不是看家基因;2)不同PITP基因之间可能存在功能冗余;3)丝状真菌中PITP的生理功能与酵母中的PITP存在显著差异。     

GTP酶激活蛋白MoGcs1参与调控稻瘟病菌的无性繁殖,附着胞分化及对外界胁迫的应答

稻瘟病菌(Magnaporthe oryzae)引起的稻瘟病是水稻上的一种毁灭性真菌病害,每年导致的稻谷产量损失可养活6千万人口。从分子水平了解该病菌的致病机理,对新型药剂靶标的挖掘和病害防控策略的制定具有重要的理论和实践指导意义。生物体存在两类GTP结合蛋白,一类是异三聚体G蛋白,另一类是小G蛋白。小G蛋白分子量为20~30 KD,具有GTP酶活性,在多种细胞反应中具有分子开关作用。小G蛋白结合GTP时被激活,可作用于下游分子使之活化。当GTP水解成为GDP时,则恢复为非活化状态。细胞中存在一些专门控制小G蛋白活性的调节因子,如鸟嘌呤核苷酸交换因子(guanine nucleotide exchange factor,GEF)和GTP酶激活蛋白(GTPase activating protein,GAP)。GEF能够催化GTP的转换,而GAP的作用是加速GTP的水解,使小G蛋白向失活的状态转变。本研究在稻瘟病菌中鉴定到一个GAP蛋白MoGcsl,并对该蛋白编码基因的生物学功能进行了研究。对MoGCS1基因进行敲除突变发现,△Mogcsl突变体产孢量显著下降,分生孢子形态异常且附着胞形成加快,但突变体的营养生长和致病能力与野生型比较无明显变化。进一步研究发现,突变体对外界盐胁迫敏感性升高,对细胞壁胁迫敏感性降低。上述结果表明,MoGcsl是稻瘟病菌生长发育过程中一个重要的蛋白,参与调控病菌的无性繁殖、附着胞分化及对外界胁迫的应答。     

MoHRD3基因参与调控稻瘟病菌的生长发育和致病力

 稻瘟病菌(Magnaporthe oryzae)作为主要的农业病原微生物,其引起的稻瘟病严重威胁着水稻等谷类作物的生产安全。内质网相关蛋白质降解途径(Endoplasmic reticulum-associated protein degradation, ERAD)是生物体应答内质网压力的主要方式之一,其在机体生长发育过程中具有重要作用。而HRD(HMG-CoA reductase degradation)复合物作为ERAD的关键组分,主要由Hrd1、Hrd3、以及凝集素Yos9等蛋白组成,负责内质网中错误折叠蛋白的识别、转运以及泛素化过程,最终由蛋白酶体降解,从而有效缓解内质网压力,保证细胞的正常生理活动。有研究表明,Hrd3属于单次跨膜蛋白,在内质网腔中与Hrd1、Yos9相结合,负责底物的识别并起着稳定Hrd1的作用。目前Hrd3在稻瘟病菌中的生物学功能尚不清楚。本研究通过基因敲除及互补试验获得了稻瘟病菌的ΔMohrd3突变体和ΔMohrd3-C回补菌株,并以野生型为对照,对突变体的生物学表型进行了分析。结果显示,ΔMohrd3突变体的生长速率、产孢量明显下降;对大麦和水稻的致病力显著减弱。进一步胁迫试验表明,MoHRD3的缺失导致稻瘟病菌对外界盐胁迫、渗透压胁迫的耐受性增强,对内质网胁迫耐受性减弱,而对细胞壁胁迫无明显变化。同时,MoHRD3基因的缺失激活了未折叠蛋白响应途径(Unfolded protein response, UPR)。上述结果表明,MoHRD3参与调控稻瘟病菌的营养生长、无性繁殖、致病及对不同环境胁迫的响应过程。     

MoHRD3基因参与调控稻瘟病菌的生长发育和致病力

 稻瘟病菌(Magnaporthe oryzae)作为主要的农业病原微生物,其引起的稻瘟病严重威胁着水稻等谷类作物的生产安全。内质网相关蛋白质降解途径(Endoplasmic reticulum-associated protein degradation, ERAD)是生物体应答内质网压力的主要方式之一,其在机体生长发育过程中具有重要作用。而HRD(HMG-CoA reductase degradation)复合物作为ERAD的关键组分,主要由Hrd1、Hrd3、以及凝集素Yos9等蛋白组成,负责内质网中错误折叠蛋白的识别、转运以及泛素化过程,最终由蛋白酶体降解,从而有效缓解内质网压力,保证细胞的正常生理活动。有研究表明,Hrd3属于单次跨膜蛋白,在内质网腔中与Hrd1、Yos9相结合,负责底物的识别并起着稳定Hrd1的作用。目前Hrd3在稻瘟病菌中的生物学功能尚不清楚。本研究通过基因敲除及互补试验获得了稻瘟病菌的ΔMohrd3突变体和ΔMohrd3-C回补菌株,并以野生型为对照,对突变体的生物学表型进行了分析。结果显示,ΔMohrd3突变体的生长速率、产孢量明显下降;对大麦和水稻的致病力显著减弱。进一步胁迫试验表明,MoHRD3的缺失导致稻瘟病菌对外界盐胁迫、渗透压胁迫的耐受性增强,对内质网胁迫耐受性减弱,而对细胞壁胁迫无明显变化。同时,MoHRD3基因的缺失激活了未折叠蛋白响应途径(Unfolded protein response, UPR)。上述结果表明,MoHRD3参与调控稻瘟病菌的营养生长、无性繁殖、致病及对不同环境胁迫的响应过程。     

MoRAD26参与稻瘟病菌的胁迫应答和致病过程

 稻瘟病菌引起的稻瘟病是水稻三大病害之一。DNA修复对维持基因组的完整性具有重要作用,为了解析DNA修复基因在稻瘟病菌致病过程中的功能,本试验对稻瘟病菌DNA损伤检控点蛋白MoRad26进行了功能分析。首先我们利用同源重组的方法获得了MoRAD26基因的稻瘟病菌敲除突变体。表型观察发现MoRAD26缺失突变体的致病力下降,对DNA损伤胁迫和H₂O₂胁迫更加敏感,但是对稻瘟病菌的生长、产孢以及分生孢子萌发和附着胞的形成没有明显影响。进一步分析表明,敲除MoRAD26基因降低了稻瘟病菌侵染菌丝的扩展能力进而影响稻瘟病菌的致病力,但其具体机制还有待深入研究。     

苹果树腐烂病菌异柠檬酸裂解酶基因VmICL的功能分析

由黑腐皮壳菌(Valsa mali Miyabe et Yamada)引起的苹果树腐烂病严重影响了苹果树的健康生长。异柠檬酸裂解酶是真菌乙醛酸循环中的关键酶,异柠檬酸裂解酶基因(ICL)在真菌生长发育及致病过程中发挥重要作用。本研究在苹果树腐烂病菌基因VmICL生物信息学分析的基础上,应用PEG介导原生质体转化法获得了基因VmICL的敲除突变体及回补菌株,解析了VmICL对病菌生长发育、细胞壁完整性、渗透胁迫、碳氮源利用及致病力的影响。研究表明,基因VmICL位于9号染色体,VmICL蛋白在第23-545位氨基酸区域具TIM super family结构域,系统发育分析显示与水稻稻瘟病菌(Magnaporthe grisea)亲缘关系最近。基因VmICL缺失突变体生长速率减慢,分生孢子器产生数量减少,分生孢子萌发延迟;基因VmICL参与维持细胞壁的完整性,但不参与渗透胁迫;基因VmICL正调控碳源吸收,负调控氮源吸收,正调控致病力。总之,基因VmICL参与调控苹果树腐烂病菌的生长发育、细胞壁完整性的维持、碳氮源利用及致病力的发挥。     

OsDCL1基因沉默突变体诱导稻瘟病抗性的转录组分析

稻瘟病是严重危害水稻生长的真菌病害,每年给水稻生产带来重大经济损失。与野生型日本晴NPB相比,OsDCL1基因沉默突变体增强了水稻对稻瘟菌的广谱抗病性。为了解水稻OsDCL1-RNAi突变体的抗病机制,我们分析和比较了它和野生型日本晴NPB在稻瘟菌侵染前后转录组水平的变化。响应稻瘟菌侵染的7 129个差异表达基因(DEGs)中,NPB和OsDCL1-RNAi突变体分别有5 382和5 180个基因表达发生了变化,参与生物胁迫反应、信号通路、蛋白代谢和RNA调控4个通路的基因明显被富集。以野生型未受稻瘟菌侵染的基因表达为对照,在OsDCL1-RNAi突变体中共发现1 318个DEGs,且超过70%是上调的,其中很多基因参与了生物胁迫反应(26.9%)和信号通路(11%),上调表达基因中与生物胁迫反应相关的主要为PR基因和细胞壁相关基因。另外,还发现10个茉莉酸相关基因和11个水杨酸相关基因分别在OsDCL1-RNAi突变体中显著下调和上调表达。在OsDCL1-RNAi突变体中还发现WRKY和MYB等一些转录因子家族以及部分受体类激酶被诱导表达。用qRT-PCR方法验证部分差异表达的基因,其结果与DEGs基本一致。OsDCL1-RNAi突变体中转录组变化的分析,为深入了解该突变体抗瘟性增强的机制奠定了基础。     

稻瘟病菌效应蛋白MoIEEP1功能初探

由稻瘟病菌引起的稻瘟病是全球最严重的植物真菌病害之一, 严重威胁水稻生产安全?效应蛋白是病原菌与植物对抗过程中分泌产生的一种蛋白质, 可作为毒力因子促进侵染或作为无毒因子触发防御反应?本研究对实验室前期筛选到的在稻瘟病菌侵染早期诱导表达的效应蛋白(infection-induced expression effector protein 1, MoIEEP1)进行了功能验证与分析?结果表明: MoIeep1 在稻瘟病菌侵染初期8 h表达量最高; 信号肽和亚细胞定位分析验证该蛋白N端含有17个氨基酸的信号肽且在水稻原生质体中呈明亮点状的定位信号, 初步推测该定位信号为过氧化物酶体或线粒体等细胞器; 与野生型菌株Guy11相比, MoIeep1 基因敲除突变体菌丝生长无明显差异, 但其致病性受到影响?以上研究结果为进一步探究该蛋白质的作用机理打下良好基础, 也为揭示其他效应蛋白在稻瘟病菌侵染水稻过程中的功能提供新的理论依据?     

Hymenopteran Parasitoids on Fruit-infesting Tephritidae (Diptera) in Latin America and the Southern United States: Diversity, Distribution, Taxonomic Status and their use in Fruit Fly Biological Control

We first discuss the diversity of fruit fly (Diptera: Tephritidae) parasitoids (Hymenoptera) of the Neotropics. Even though the emphasis is on Anastrepha parasitoids, we also review all the information available on parasitoids attacking flies in the genera Ceratitis, Rhagoletis, Rhagoletotrypeta, Toxotrypana and Zonosemata. We center our analysis in parasitoid guilds, parasitoid assemblage size and fly host profiles. We also discuss distribution patterns and the taxonomic status of all known Anastrepha parasitoids. We follow by providing a historical overview of biological control of pestiferous tephritids in Latin American and Florida (U.S.A.) and by analyzing the success or failure of classical and augmentative biological control programs implemented to date in these regions. We also discuss the lack of success of introductions of exotic fruit fly parasitoids in various Latin American countries. We finish by discussing the most pressing needs related to fruit fly biological control (classical, augmentative, and conservation modalities) in areas of the Neotropics where fruit fly populations severely restrict the development of commercial fruit growing. We also address the need for much more intensive research on the bioecology of native fruit fly parasitoids.     

Brief Guide for Authors

正Journal of Arid Land(JAL)is an international journal(ISSN 1674-6767;CN 65-1278/K)for the natural sciences,sponsored by the Xinjiang Institute of Ecology and Geography,Chinese Academy of Sciences and Science Press.It is published by Science Press and Springer-Verlag Berlin Heidelberg bimonthly.JAL publishes original,innovative,and integrative research from arid and semiarid regions,ad     

The parasitoid complex ofLiriomyza cicerina on chickpea (Cicer arietinum)

Liriomyza cicerina (Diptera: Agromyzidae) is an important pest on chickpea in Turkey. The objective of this study was to determine the parasitoids and rates of parasitism ofL. cicerina on chickpea (Cicer arietinum L.) during the 2005 and 2006 seasons in ?anl?urfa province, Turkey. Leaves with mines were sampled weekly and kept in the laboratory to observe and count emerging leafminer and parasitoid adults. Eight parasitoid species were collected: the braconidsOpius monilicornis Fischer andOpius tersus Foerster and the eulophidsDiaulinopsis arenaria (Erdös) andNeochrysocharis formosa (Westwood), which occurred in both the winter and summer seasons;Diglyphus crassinervis Erdös,Neochrysocharis ambitiosa Hansson,Neochrysocharis sericea (Erdös) andPediobius metallicus (Nees), which occurred only in the summer growing areas.Diaulinopsis arenaria was the predominant parasitoid with 4–7.7% parasitism rate whileN. ambitiosa andO. monilicornis were the second and third most predominant species. The results of these trials show that sinceDia. arenaria occurred throughout every season, it could potentially be used for control of the leafminerL. cicerina.     

Plant–herbivore asynchrony necessitates augmentative releases of the exotic beetle,Zygogramma bicolorata,to enhance the biological control of P arthenium hysterophorus

Systematic information on the quantitative impact of Z ygogramma bicolorata on the biology of P arthenium hysterophorus is crucial as the seeds of this weed continue to germinate from the accumulated soil seed bank throughout the year in the form of different germinating flushes, while the activity of the beetle ceases during winter as it enters diapause. Therefore, plant–herbivore interactions need to be explored to develop predictions of the overall impact of the introduced beetle on the weed. The findings revealed that defoliation by Z . bicolorata had a significant impact on the plant height, density and flower production in flushes F ₃, F ₄ and F ₅, but not in F ₁ and F ₂ that exhibited longer periodicity, profuse branching, a longer flowering period and maximum flower production and contributed mostly to the existing seed soil bank. Therefore, total depletion of the existing soil seed bank was not possible. Consequently, the effect of augmentative field releases of laboratory‐reared beetles was explored on F ₁ and F ₂ in February for three consecutive years (2011–2013). Before initiating the trial, random soil samples were taken from the plots that were assigned to the paired treatments (i.e. with the beetle and without the beetle [insecticide‐treated]) and it was found that the seed bank in those samples did not differ. The single release of Z . bicolorata adults at five per plant at the six‐leaf stage significantly reduced the soil seed bank, compared to without the biocontrol agent, irrespective of the flushes at the end of the season.     

Richtlinien der Verbände des ökologischen Landbaus zum Vorratsschutz

Zusammenfassung Verbände des ökologischen Landbaus wie z. B. Bioland, Gäa, Demeter; Naturland, Ernte für das Leben oder Bio Suisse beschränken ihre Mitglieder bei der Wahl von Vorratsschutzmaßnahmen. Vorrang besitzen Maßnahmen zur Vermeidung von Schädlingen gegenüber Bekämpfungsmaßnahmen. Fallen müssen zur Befallsüberwachung eingesetzt werden, um einen Befall durch Vorratsschädlinge frühzeitig zu erkennen. Diese Maßnahmen sollen den weitgehenden Verzicht auf chemisch-synthetische Mittel ermöglichen. In diesem Beitrag werden die Empfehlungen der Verbände mit den derzeit verfügbaren chemischen Mitteln für den Vorratsschutz abgeglichen. Erfahrung in der praktischen Umsetzung von physikalischen und biologischen Verfahren werden diskutiert und Defizite bei der Befallsüberwachung und Bekämpfung beschrieben.     

Development of protocols for detection of Colletotrichum acutatum and monitoring of strawberry anthracnose using real-time PCR

Real-time PCR (TaqMan®) assays were developed for the specific detection and discrimination of Colletotrichum spp., C. acutatum and C. gloeosporioides causing anthracnose in strawberry using the most divergent area of the internal transcribed spacers (ITS1 and ITS2) and 5·8S ribosomal RNA (rRNA) gene region. The specificity of the new assays was tested using DNA from six species of Colletotrichum and nine fungal species commonly found associated with strawberry material, and additionally by comparing the sequences with those from databases using a blast search. The sequences only showed identity with homologous sequences from the desired target organisms. The new assays were 10–100 times more sensitive than conventional PCR methods previously published for the diagnosis of strawberry anthracnose. When real-time PCR was compared with ELISA methods, PCR improved the sensitivity of the identification by obtaining positive results for samples of strawberry plant material that tested negative with ELISA. The development of C. acutatum was monitored using artificially infected strawberry crowns from two strawberry cultivars (Camarosa and Ventana) and a real-time PCR assay specific for this species between January and June 2006. The amount of C. acutatum detected using real-time PCR varied significantly by month ( P  < 0·001), but not by cultivar ( P  = 0·394). The new assays were shown to be useful tools for rapid detection and identification of these pathogens and to allow rapid and accurate assessment of the casual agents of anthracnose in strawberry.     

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.     

Relationship Between Production of the Phytotoxin Prehelminthosporol and Virulence in Isolates of the Plant Pathogenic Fungus Bipolaris sorokiniana

A gas chromatographic method was developed to quantify the phytotoxin prehelminthosporol, which is a sesquiterpene metabolite of the plant pathogen Bipolaris sorokiniana. The toxin was extracted from mycelium or culture filtrates, pre-cleaned using solid phase extraction, and analyzed by gas chromatography as a trimethylsilyl-derivative. The detection limit of the method was 5ngl^–1 (signal to noise ratio 4:1) which corresponds to ca. 15ng prehelminthosporol per mg dry weight of mycelium or 15ng prehelminthosporol per ml culture filtrate. The total amount of prehelminthosporol (mycelium plus culture filtrate) increased with cultivation time when examined in six isolates of B. sorokiniana after 6, 9, 12 and 15 days of incubation. The screening experiment of 17 isolates for prehelminthosporol production after 8 days of incubation revealed significant differences in the toxin production between the isolates. The isolates with low toxin production had lower virulence towards barley roots compared to those with higher production of the toxin. However, the virulence did not increase with prehelminthosporol level among the high producing isolates. Prehelminthosporol was also analyzed in a number of related Bipolaris and Drechslera species. In addition to B. sorokiniana, three out of six Bipolaris species (B. setariae, B. zeicola, B. victoriae) produced prehelminthosporol, which indicates that ability to produce prehelminthosporol is conserved among closely-related Bipolaris species.