南方水稻黑条矮缩病毒dot-ELISA检测试剂盒的应用性能验证

以制备的抗南方水稻黑条矮缩病毒(Southern rice black-streaked dwarf virus,SRBSDV)单抗2C2为核心,建立了检测水稻叶片和白背飞虱虫中SRBSDV的dot-ELISA试剂盒。试剂盒的灵敏度分析表明,当SRBSDV感染病叶稀释到1∶10 240倍(w/v,g/m L)、单头携毒白背飞虱稀释到1∶51 200倍(头/μL)时仍能检测到SRBSDV。建立的试剂盒检测感染SRBSDV的水稻和携毒白背飞虱呈阳性反应,而检测感染水稻黑条矮缩病毒、水稻矮缩病毒、水稻条纹病毒、水稻瘤矮病毒、水稻条纹花叶病毒、水稻锯齿矮缩病毒的病叶和健康水稻及非携毒白背飞虱呈阴性反应。试剂盒的田间样品检测结果与RT-PCR方法的检测结果的符合率达到100%,核酸测序和序列比对结果发现RT-PCR检测阳性的样品确实感染SRBSDV。试验结果表明,建立的检测试剂盒能准确、有效地检测田间白背飞虱及水稻样品中的SRBSDV,可为我国南方水稻黑条矮缩病的检测和诊断、预测预警及科学防控提供技术服务。     

南方水稻黑条矮缩病毒和水稻黑条矮缩病毒的单抗制备及其检测应用

 用与牛血清白蛋白偶联的南方水稻黑条矮缩病毒（Southern rice black-streaked dwarf virus,SRBSDV）衣壳蛋白的C端12个氨基酸多肽为抗原免疫BALB/c小鼠,经细胞融合、筛选、克隆,获得2株能稳定传代并分泌抗SRBSDV和水稻黑条矮缩病毒（Rice black-streaked dwarf virus,RBSDV）单克隆抗体（MAb）的杂交瘤细胞株3F1、5G1。3F1、5G1单克隆抗体腹水间接ELISA效价达10^-6,抗体类型及亚类均为IgG1, kappa链。 Western blot分析表明,2株单克隆抗体均与SRBSDV和RBSDV的外壳蛋白亚基有特异反应。利用单克隆抗体3F1建立的dot-ELISA检测方法能准确、特异、灵敏地检测田间稻飞虱及水稻样品中的SRBSDV和RBSDV。SRBSDV和RBSDV单克隆抗体的制备及检测方法的建立为水稻黑条矮缩病的诊断、预测预报及科学防控提供了技术支撑。     

公安县南方水稻黑条矮缩病的发生规律和防控技术初探

南方水稻黑条矮缩病是近年来发现并命名的一种新的水稻病毒病。该病由呼肠孤病毒科斐济病毒属（Fijivirus）的一个建议新种——南方水稻黑条矮缩病毒（Southern rice black streaked dwarf virus,SRBSDV）引起。SRBSDV由迁飞性昆虫白背飞虱传播,所引发的南方水稻黑条矮缩病已经在我国华南、华中、华东、西南等广大稻区以及越南、日本等国家的部分稻区发生。2009年,该病害在我国南部及越南北部晚季稻上暴发成灾,我国的受害面积30万hm2以上,     

南方水稻黑条矮缩病毒检测方法的比较

为明确南方水稻黑条矮缩病毒（Southern rice black-streaked dwarf virus, SRBSDV）检测方法的最佳适用范围,对其现有检测方法Real time RT-PCR、RT-LAMP、RT-PCR的灵敏性及特异性进行了比较,并分析了依据SRBSDV单克隆抗体3F1建立的斑点免疫结合印迹（dot immunobinding assay, DIBA）方法对检测植物寄主和白背飞虱Sogatella furcifera Horvth的特异性。结果表明,灵敏性以Real time RT-PCR方法最高,其次为RT-LAMP方法,而普通RT-PCR方法相对较低。这3种方法均可特异性检测SRBSDV植物寄主和白背飞虱;DIBA方法可以满足SRBSDV和水稻黑条矮缩病毒（Rice black-streaked dwarf virus, RBSDV）植物寄主和白背飞虱大量样品的检测,但不能区分SRBSDV和RBSDV。Real time RT-PCR方法实现了短时间内对SRBSDV RNA拷贝数的相对定量;RT-LAMP方法全程恒温反应,无需热循环仪。     

南方水稻黑条矮缩病的发生规律及防治措施

南方水稻黑条矮缩病(Southern rice black-streaked dwarf virus,SRBSDV,Genus Fijivirus)在关岭县水稻种植过程中常年发生,造成水稻矮缩不抽穗,极大地影响水稻产量和品质,给农业带来极大的经济损失.本文详细介绍了贵州省关岭县南方水稻黑条矮缩病发病症状和发病规律,并根...     

江西省南方水稻黑条矮缩病发生情况及防治建议

由斐济病毒属(Fijivirus)暂定新种南方水稻黑条矮缩病毒(southern rice black-streaked dwarf virus,SRBSDV)引起的水稻矮缩病,自2001年在广东省阳西县被华南农业大学周国辉教授等人鉴定发现以来,已迅速扩散至我国南方广大稻区。2009年,该病害在我国南部及越南晚季稻上暴发成灾,造成严重的产量损失。通过对南方水稻黑条矮缩病毒的生物学特性、近几年在江西的发生、预测预报情况进行综述,提出了该病的综合防治技术措施,以期为江西省南方水稻黑条矮缩病的发生及防治提供参考。     

南方水稻黑条矮缩病发生现状及防控对策

<正>南方水稻黑条矮缩病毒(Southern rice black-streaked dwarf virus,SRBSDV)是由我国科学工作者首先发现,并经华南农业大学周国辉教授等人鉴定和命名的为害农作物的病毒新种,属呼肠孤病毒科     

关于南方水稻黑条矮缩病防控工作的几点思考

南方水稻黑条矮缩病是近年来传人湖北省的新发生病害,它是由南方水稻黑条矮缩病毒South—ern rice black-streaked dwarf virus,SRBSDV引起的,2009年在公安县晚稻田首次见病,近年来扩散蔓延迅速,由于该病害可经迁飞性害虫白背飞虱进行远距离传播,扩展快,隐蔽性强,危害大,已对我省水稻生产安全构成了严重威胁。为科学、有效防控南方水稻黑条矮缩病,全国和我省均制定了防控技术方案,但由于该病研究时间较短,一些研究还不够深入,故在防治策略和防治实践上还存在不足,值得进一步探讨。     

南方水稻黑条矮缩病毒的快速检测

南方水稻黑条矮缩病毒(Southern rice black-streaked dwarf virus,SRBSDV)病原鉴定和介体带毒率检测是病害防治的前提条件。本研究分别建立了病株dsRNA基因组鉴定法和介体带毒率测定的斑点杂交法。结果显示:dsRNA基因组鉴定法可以从0.5 g水稻病株组织中快速检测到SRBSDV,不需要经过RT-PCR过程,具有快速简便等特点;斑点杂交法检测介体昆虫白背飞虱带毒率的灵敏度较高,单头带毒介体的总RNA稀释1 000倍后依然可以检测到病毒,并且可以大批量处理样品,用于病害流行研究和测报。     

抑制南方水稻黑条矮缩病毒非结构蛋白P5-1的表达阻碍病毒在昆虫体内的增殖

为明确南方水稻黑条矮缩病毒(Southern rice black-streaked dwarf virus,SRBSDV)编码的非结构蛋白P5-1参于SRBSDV在介体白背飞虱体内侵染过程中的作用机制,通过原核表达蛋白制备SRBSDV编码的非结构蛋白P5-1的多克隆抗体,并应用Western blot和免疫荧光标记法检测抗体的特异性,以注射法将来源于P5-1基因的dsRNA(ds P5-1)注入获毒1 d的白背飞虱体内,5 d后通过免疫荧光标记法检测ds P5-1对SRBSDV在白背飞虱体内增殖的影响,同时以注射来源于GFP基因的dsRNA(ds GFP)为对照。结果显示,Western blot和免疫荧光标记分别检测到SRBSDV侵染水稻和白背飞虱表达的P5-1蛋白,表明所制备的P5-1抗体具有特异性。ds GFP处理的对照组白背飞虱带毒率高达81%,而ds P5-1处理的白背飞虱带毒率仅为21%,且P5-1蛋白的表达和SRBSDV在昆虫体内的增殖均受到抑制。表明P5-1蛋白是病毒在昆虫体内增殖的关键因子,可作为阻断病毒在昆虫体内增殖的理想靶标。     

Tumours of Begonia and some other ornamentals,induced by Corynebacterium fascians

In 1975 many tumours were observed in plants ofBegonia Schwabenland grown in Aalsmeer. Submersion of the roots ofNicotiana megalosiphon seedlings in a homogenate of tumorous tissue, induced tumours after two weeks. Short periods of submergence yielded results similar to those obtained after longer periods. Tumour homogenates lost their infectivity after ten min at 50°C. Aphids transmitted the infectious agent.Treatment with propylene oxide did not inhibit infectivity completely. Filtration through a 450 nm filter removed the infectious agent.Tobacco tumor virus or a viroid could not be isolated. Cultures ofCorynebacterium fascians, isolated from tumours ofN. megalosiphon were highly infectious and induced tumours in healthyN. megalosiphon andBegonia. Tumorous tissue homogenates ofPelargonium zonale, Dahlia sp.,Gladiolus sp., andLilium sp. also caused tumours inN. megalosiphon, from whichC. fascians was isolated. It was not possible to produce tumours inN. megalosiphon with homogenates from roses with symptoms of bud proliferation.Samenvatting In 1975 werden vele tumoren waargenomen inBegonia Schwabenland op Aalsmeerse bedrijven (Fig. 1). De infectiositeit van tumorweefsel kon goed en snel worden vastgesteld door de wortels van zaailingen vanNicotiana megalosiphon in een homogenaat van tumorweefsel te dompelen. Tumoren ontstonden na twee weken, de eindbeoordeling geschiedde na een maand (Fig. 2). Ook verschillende andereNicotiana spp.,Melilotus officinalis (Fig. 3) enPisum odoratum (Fig. 4) werden aangetast.Bij de infectiositeitstoets gaven zeer korte dompeltijden even goede resultaten als langere (Tabel 1). Infectieus sap verloor zijn infectievermogen na 10 min verhitting bij 50°C. Bladluizen brachten de smetstof over. Propyleenoxide verminderde de infectiositeit wel, doch onderdrukte deze niet totaal. Bij filtratie door een 450 nm filter bleef het infectieuse agens op het filter achter. Het tumor-inducerende agens was ook aanwezig in die delen van planten met tumoren welke gezond leken en het ging voor een gering deel over met zaad (Tabel 2).Uit tumoren konden wij geen tabakstumorvirus of een viroïde isoleren. Culturen vanCorynebacterium fascians, geïsoleerd uit tumoren vanN. megalosiphon bleken zeer infectieus en veroorzaakten tumoren inN. megalosiphon enBegonia. Homogenaten van tumorweefsel vanPelargonium zonale, dahlia (Fig. 5), gladiool (Fig. 6) enLilium Mid Century Hybrid Enchantment (Fig. 7) veroorzaakten ook tumoren opN. megalosiphon, waaruitC. fascians werd geïsoleerd. Met sap van kroeskopzieke rozen konden wijN. megalosiphon niet besmetten.     

Plant Viruses Transmitted by Whiteflies

One-hundred and fourteen virus species are transmitted by whiteflies (family Aleyrodidae). Bemisia tabaci transmits 111 of these species while Trialeurodes vaporariorum and T. abutilonia transmit three species each. B. tabaci and T. vaporariorum are present in the European–Mediterranean region, though the former is restricted in its distribution. Of the whitefly-transmitted virus species, 90% belong to the Begomovirus genus, 6% to the Crinivirus genus and the remaining 4% are in the Closterovirus, Ipomovirus or Carlavirus genera. Other named, whitefly-transmitted viruses that have not yet been ranked as species are also documented. The names, abbreviations and synonyms of the whitefly-transmitted viruses are presented in tabulated form together with details of their whitefly vectors, natural hosts and distribution. Entries are also annotated with references. Whitefly-transmitted viruses affecting plants in the European–Mediterranean region have been highlighted in the text.     

Epidemiology and simulation of population development of Sitobion avenae in winter wheat

The epidemiology ofSitobion avenae and its natural enemies in winter wheat was studied in 1975, 1976 and 1977. Immigration was important until the end of flowering. The alate immigrants had apterous offspring. These became the driving force in population growth. Their offspring were mostly alatae which usually left the field. A model of the epidemic was developed. Quantitative relations between the aphids and their environment were obtained from literature or established in laboratory trials. The model simulated population development and population composition from the beginning of June till the population peak at the end of June or early in July. Because quantitative data on relations between aphids and their natural enemies and pathogens are scarce, and since the knowledge on wing formation is still limited, the population collapse could not be predicted. In the future, prognosis over a period of three weeks seems possible.Samenvatting De toenemende betekenis van graanbladluizen (vooralSitobion avenae) gepaard gaande met een sterke toename van het gebruik van insecticiden op granen maakte verbetering van de prognose over het schadelijk optreden wenselijk. Door gedetailleerde tellingen in het veld (Fig. 1–7) werden gegevens verkregen over het verloop van de epidemie en het optreden van natuurlijke vijanden in 1975, 1976 en 1977.Een immigratieperiode tot in de bloei kon worden vastgesteld. Daarna lijkt de aantrekkelijkheid van het gewas voor alate luizen te verminderen. De alate immigranten krijgen aptere nakomelingen. Deze vormen de stuwende kracht van de populatiegroei. De nakomelingen van apteren zijn merendeels alaat. Zij verlaten het gewas.Een model van de populatieontwikkeling gedurende de epidemie werd opgesteld. De relatiediagrammen Fig. 9 en 10 laten groei en ontwikkeling vanS. avenae en een predator (Syrphus corollae) zien. Kwantificering van de betrekkingen werd mogelijk door literatuurgegevens en laboratoriumexperimenten.Met het model kon de populatieontwikkeling vanS. avenae vanaf begin juni tot aan de populatiepiek in 1975, 1976 en 1977 vrij goed worden gesimuleerd (Fig. 12). Ook de populatieopbouw kon worden gesimuleerd (Fig. 14). De teruggang van de populatiedichtheid blijkt moeilijker te voorspellen door het ontbreken van gegevens over natuurlijke vijanden.Het lijkt waarschijnlijk dat in de toekomst met het model een prognose over de piek van de bladluispopulatie circa 3 weken tevoren mogelijk zal zijn.     

Broad bean mottle virus in Morocco; curculionid vectors,and natural occurrence in food legumes other than faba bean (Vicia faba)

Broad bean mottle virus (BBMV) was transmitted from infected to healthy faba-bean plants by the curculionid weevilsApion radiolus Kirby,Hypera variabilis Herbst,Pachytychius strumarius Gyll,Smicronyx cyaneus Gyll, andSitona lineatus L. The latter appeared to be an efficient vector: acquisition and inoculation occurred at the first bite, the rate of transmission was c. 41%, and virus retention lasted for at least seven days.S. lineatus transmitted the virus from faba bean to lentil and pea, but not to the three genotypes of chickpea tested. This is the first report on the generaHypera, Pachytychius, andSmicronyx as virus vectors, and onA. radiolus, H. variabilis, P. strumarius, andS. cyaneus as vectors of BBMV.Out of 351 samples of food legumes with symptoms suggestive of virus infection, 16, 11, 19, and 17% of the samples of chickpea, lentil, pea, and common bean, respectively, were found infected when tested for BBMV in DAS-ELISA. This is the first report on the natural occurrence of BBMV in chickpea, lentil, pea, and common bean. The virus should be regarded as a food-legume virus rather than a faba-bean virus solely, and is considered an actual threat to food legume improvement programmes.     

Relationships of <Emphasis Type="Italic">Barley yellow dwarf virus</Emphasis>-PAV and <Emphasis Type="Italic">Cereal yellow dwarf virus</Emphasis>-RPV from Iran with viruses of the family <Emphasis Type="Italic">Luteoviridae</Emphasis>

Barley yellow dwarf disease is one of the most important problems confronting cereal production in Iran. Barley yellow dwarf virus-PAV (BYDV-PAV) and Cereal yellow dwarf virus-RPV (CYDV-RPV) are the predominant viruses associated with the disease. One isolate of BYDV-PAV from wheat (PAV-IR) and one isolate of CYDV-RPV from barley (RPV-IR) were selected for molecular characterisations. A genome segment of each isolate was amplified by PCR. The PAV-IR fragment (1264 nt) covered a region containing partial genes for coat protein (CP), read through protein (RTP) and movement protein (MP). PAV-IR showed a high sequence identity to PAV isolates from USA, France and Japan (96–97%). In a phylogenetic analysis it was placed into PAV group I together with PAV isolates from barley and oats. The fragment of RPV-IR (719 nt) contained partial genes for CP, RTP and MP. The sequence information confirmed its identity as CYDV. However, RPV-IR showed 90–91% identity with both RPV and Cereal yellow dwarf virus-RPS (CYDV-RPS). Phylogenetic analyses suggested that it was more closely related to RPS. These data comprise the first attempt to characterise BYD-causing viruses in Iran and southwest Asia. The nucleotide sequence data reported appear in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under the accession numbers AY450425 and AY450454     

Identification,incidence and characterization of <Emphasis Type="Italic">Fusarium proliferatum</Emphasis>on ornamental palms in Spain

During a survey conducted from 1998 to 2002, Fusarium proliferatum was found associated with young and adult palms belonging to the genera Chamaerops, Phoenix, Trachycarpus and Washingtonia showing symptoms of wilt and dieback. The fertility and toxicological profile of 36 strains representing different locations and hosts were studied. All of them except two, which were infertile, belonged to mating population D. Both mating types (MATD-1 and MATD-2) were isolated from the same host species, showing a high potential of genetic recombination in the field. Additionally, eight strains were fertile once crossed as female. Toxin analysis showed differences in the ability of strains to produce fumonisin B₁, moniliformin, beauvericin, fusaric acid and fusaproliferin. Only 17 of them produced all the toxins analyzed. Pathogenicity tests were conducted on Phoenix dactylifera and P. canariensis using nine F. proliferatum Spanish strains and two reference strains from Saudi Arabia. Eight months after inoculation all strains caused disease, with palms showing lesions on the bases of leaves and development of wilt symptoms similar to those originally observed in affected plants. This is the first report on the occurrence of F. proliferatum on P. dactylifera in Spain and also the first report of this pathogen on C. humilis, P. canariensis, P. reclinata, T. fortunei, W. filifera and W. robusta.     

A new <Emphasis Type="Italic">Rhizoctonia</Emphasis> sp. closely related to <Emphasis Type="Italic">Waitea circinata</Emphasis> causes a new disease of creeping bentgrass

Isolates of an unidentified Rhizoctonia sp. (UR isolates) were obtained from creeping bentgrass and Kentucky bluegrass with reddish brown sheath and foliar rots. Because the UR isolates anastomosed with isolates of three varieties of Waitea circinata (var. oryzae, var. zeae, and var. circinata), colony morphology, hyphal growth rate at different temperatures, pathogenicity, sequence analysis of the internal transcribed spacers (ITS) region of ribosomal RNA genes (rDNA) were compared. The colony color of mature UR isolates was distinct from isolates of the other three varieties of W. circinata. In pathogenicity tests on creeping bentgrass, the severity of the disease caused by UR isolates was significantly higher than that caused by the three varieties of W. circinata. Sequence similarities of the rDNA-ITS region between UR isolates and between isolates within each variety were high (97–100%), but they were lower among isolates from UR and the varieties of W. circinata (88–94%). In a phylogenetic tree based on the rDNA-ITS sequences, UR isolates formed a cluster separate from each of the clusters formed by the three varieties of W. circinata. These results indicate that the UR isolates clearly differ from the three varieties of W. circinata. We therefore propose that the UR isolates be classified as new Rhizoctonia sp. that are closely related to W. circinata and that the disease on creeping bentgrass should be called Waitea reddish-brown patch disease (Sekikasshoku-hagusare-byo in Japanese).     

Recent observations on leafy gall in Liliaceae and some other families

Corynebacterium fascians, which causes leafy gall, has been shown to be responsible for the unusual symptoms recently found in lilies. The symptoms are described and the different bacterial isolates compared by biochemical, serological and pathogenicity experiments. No evidence could be found for the existence of specialized strains even though considerable variation in virulence could be demonstrated. Although the role of variation in susceptibility of the different lily cultivars should not be underestimated, it would appear that high inoculum levels ofC. fascians in the soil may be largely responsible for these outbreaks.Also included in this study are the results of biochemical, serological and pathogenicity experiments ofC. fascians which has been isolated fromKalanchoe, Euphorbia, Brodiaea, Hebe andVerbascum.Samenvatting Reeds een aantal jaren was in lelies een woekering in stengelbollen bekend, die echter sporadisch voorkwam en economisch van weinig betekenis was. Tijdens de zomer van 1977 echter trad de ziekte op één enkel perceel zo ernstig op dat zeer aanzienlijke oogstderving het gevolg was. Behalve de stengelbol kan ook de hoofdbol misvormd zijn. Bovendien kunnen misvormde stengelbollen lange spruiten vormen, zgn. sabeltandvorming. Soms is het wortelstelsel gereduceerd; in de kas zijn verdikte stengels waargenomen. De ziekte staat nu bekend onder de naam woekerziekte.Door de PD te Wageningen en het LBO te Lisse werd de bacterieCorynebacterium fascians (Tilford) Dows. uit het zieke materiaal geïsoleerd. In gezamenlijk onderzoek werd nagegaan of de bacterie de veroorzaker is van bovengenoemde symptomen en of hier sprake is van een op lelie gespecialiseerde stam.Biochemische en serologische vergelijkingen tussen isolaten vanC. fascians uit lelie,Kalanchoe, Euphorbia, Brodiaeea, Hebe enVerbascum laten zien dat er zeer weinig verschillen bestaan. Alle isolaten reageren op een standaard antiserum met titers variërend tussen 1: 640–1: 5120.Een oriënterende inoculatieproef werd uitgevoerd op het LBO in 1977. Daarna werden op de PD 10 van de 25 isolaten uit lelie en 8 uit andere waardplanten getoetst op lelie, chrysant en lathyrus. Uit de inoculatieproeven blijkt, dat de bacterie de veroorzaker is van de waargenomen symptomen. Tussen de isolaten blijkt een groot verschil in pathogeniteit te bestaan (12–100% aantasting), zonder dat dit aan herkomst gebonden is. Het voorkomen van op lelie of andere waardplanten gespecialiseerde stammen is daarom niet aan te nemen.In hoeverre de hevige aantasting een gevolg is van een opbouw van de bacterie-populatie door een te nauwe vruchtwisseling en mechanische rooimethoden moet worden nagegaan.     

Pathogenic fungi involved in root rot of peas in the Netherlands and their physiological specialization

Research on root rot pathogens of peas in the Netherlands has confirmed the prevalence ofFusarium solani, F. oxysporum, Pythium spp.,Mycosphaerella pinodes andPhoma medicaginis var.pinodella. Aphanomyces euteiches andThielaviopsis basicola were identified for the first time as pea pathogens in the Netherlands. Other pathogens such asRhizoctonia solani andCylindrocarpon destructans were also found on diseased parts of roots. F. solani existed in different degrees of pathogenicity, and was sometimes highly specific to pea, dwarf bean of field bean, depending on the cropping history of the field.A. euteiches was specific to peas, whereasT. basicola showed some degree of physiological specialization.     

Nematicidal Activity of Chrysanthemum coronarium

Organic amendments and green manure are potential alternatives to the harmful chemical control means currently used against plant-parasitic nematodes. In this work, Chrysanthemum coronarium was applied to the soil as a green manure to control the root-knot nematodes Meloidogyne incognita and M. javanica. Chrysanthemum coronarium significantly reduced nematode infection of tomato roots and improved plant-top fresh weight, both in the greenhouse and in microplots. Other green manures, derived from Anthemis pseudocotula, wild chickpea (Cicer pinnatifidum), Geranium spp. and wheat, were not as effective as C. coronarium. Chrysanthemum coronarium, retained its nematicidal activity even when applied as a dried material. Only mature C. coronarium plants, in their flowering stage, exhibited nematode control activity, but the green plant parts were more effective than the flowers. An aqueous extract of C. coronarium exhibited in vitro, nematostatic activity towards M. incognita and M. javanica second-stage juveniles and inhibited their hatching from eggs and egg-masses; its nematostatic activity was expressed also against other phytonematode species such as Heterodera avenae and Pratylenchus mediterraneus, but did not affect the beneficial entomopathogenic nematode Steinernema feltiae.