Turnip mosaic virus induces expression of the LRR II subfamily genes and regulates the salicylic acid signaling pathway in non-heading Chinese cabbage

In order to study the defense response to turnip mosaic virus (TuMV) infection in non-heading Chinese cabbage (Brassica campestris ssp. chinensis Makino), we cloned the LRR II subfamily genes which comprises six members. They were high homologous to the function-known LRR II genes of Arabidopsis. We investigated their expression through quantitative real-time PCR analysis. TuMV infection induced the expression of these genes locally and systematically, and regulated the endogenous accumulation of salicylic acid (SA). Exogenous SA spraying was able to induce resistance to the susceptibility of the TuMV-infected plants, which might function via inhibiting the viral duplication. Though TuMV-induced SA accumulation was not the determinant in regulating gene expression, it mediated the reaction oxygen species (ROS) burst as a channel of defense.     

大豆与疫霉菌非亲和互作早期差异显示基因的表达分析

大豆疫霉根腐病是大豆的毁灭性病害。为了深入了解大豆对疫霉菌的分子抗病机制,以大豆疫霉菌1号生理小种游动孢子接种抗性品种绥农10的根部及下胚轴,通过反转录差异显示技术分离到疫霉菌侵染0、0.5、1、2和4h后大豆下胚轴和茎部的差异表达基因,其中至少有8个基因与抗病相关。接种后0.5 h开始上调表达的有肉桂酸-4-羟化酶基因、ATP合成酶β亚基基因,以及类花生泛素结合酶基因;接种后1h和2h依次开始上调表达的有尿苷二磷酸-N-乙酰基-α-D-氨基半乳糖基因和豌豆蓝铜蛋白基因;接种后4 h才上调表达的有TGA型碱性亮氨酸拉链基因、大豆环孢素基因和14-3-3蛋白基因。这8个基因中有1个基因与信号传导有关、4个基因与抗病和防御有关、2个基因与转录调控有关、1个基因与能量代谢有关。研究表明,以上8个基因在疫霉菌游动孢子萌发、侵入大豆和在大豆体内扩展过程中起着重要作用。     

Serotypic variation in turnip mosaic virus

A panel of 30 monoclonal antibodies (MAbs) was produced against four isolates of turnip mosaic virus (TuMV). The panel was tested in plate-trapped antigen ELISA tests against 41 TuMV isolates (with different host and geographical origins and of differing pathotypes). The antibodies were also tested against four other potyviruses (bean common mosaic virus, bean common mosaic necrosis virus, lettuce mosaic virus and zucchini yellow mosaic virus). The reactions were assessed quantitatively (using multivariate analysis) and qualitatively (using the standard deviation obtained against healthy leaf material). The MAbs recognized 16–17 TuMV epitopes that were not present in the other potyviruses and a further two potyvirus epitopes. The isolates were grouped into three serotypes. Only one isolate did not fit this grouping. The classification of seven isolates in coat protein amino acid sequence homology groups correlated with serotypes. There was no correlation between serotype and pathotype, or between reactions to individual MAbs and single lines. There was therefore no evidence that the epitopes recognized by the MAbs are elicitors for the resistance genes present in the Brassica napus lines. However, the sensitivity and specificity of the MAbs will be useful for both routine detection of TuMV and fundamental studies on plant–virus interactions.     

大白菜抗芜菁花叶病毒的QTL分析

 以大白菜高抗TuMV-C₃株系的高代自交系A52-2和感病自交系GCⅣ杂交的F₂代255个单株作为构建遗传图谱的作图群体,通过EST-PCR-RFLP和AFLP分子标记的遗传分析,构建了包含12个连锁群,由124个遗传标记组成的大白菜分子遗传图谱。其中包括6个EST-PCR-RFLP多态性位点和118个AFLP多态性位点,该图谱覆盖长度为683.9 cM,平均图距5.52 cM。利用Windows QTL CartographerV 2.0软件和复合区间作图法进行分析,共检测到4个抗TuMV-C₃株系的QTLs位点。     

Purification and characterization of turnip mosaic virus helper component protein

ABSTRACT The helper component (HC) protein of turnip mosaic virus (TuMV) was concentrated by differential centrifugation followed by ammonium sulfate precipitation. The partially purified HC was then loaded onto a Ni(2+)-resin column that bound the HC; a histidine tag was not required for binding. The HC eluted from the column migrated as a band of about 50 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In its native state, the HC did not pass through an ultrafiltration membrane with a molecular mass cutoff of 100 kDa, which suggested that the HC is in a multimeric form when it is biologically active. The molecular mass of the multimeric form was determined by gel filtration to be approximately 145 kDa. Purified HC retained its activity for several months at -20 degrees C. Using a protein blotting-overlay protocol, purified HC interacted in vitro with an aphid-transmissible TuMV isolate, but not with a non-aphid-transmissible isolate.     

Host range and some properties of Physalis mosaic virus,a new virus of the turnip yellow mosaic virus group

An apparently undescribed virus was isolated fromPhysalis subglabrata in Illinois, USA, and its properties were studied. The virus was namedPhysalis mosaic virus (PMV). It was readily transmitted by sap inoculation to 23 out of 34 Solanaceae tested, toChenopodium foetidum andSonchus oleraceus but not to 28 other non-solanaceous species inoculated. Purified preparations of PMV contained isometric particles of 27 nm in diameter, which sedimented as two components with sedimentation coefficients of 50 and 112 S. The 112 S component was infectious, the 52 S component was not. The virus contained 38% ribonucleic acid with a molar base content of G 14.4%, A 22.9%, C 37,2% and U 25.5%.Purified preparations were highly infectious; a concentration of about 6000 particles per ml was infectious on plants.PMV is a member of the Andean potato latent virus subgroup of the turnip yellow mosaic virus group. The virus was closely related to the viruses: Andean potato latent, belladonna mottle, dulcamara mottle and egg-plant mosaic.Samenvatting Een nog niet eerder beschreven virus, dat in de staat Illinois (V.S. van Amerika) opPhysalis subglabrata was gevonden, werd in Wageningen bestudeerd. Het virus dat Physalis mosaic virus (PMV) (in het Nederlands:Physalis-mozaïekvirus) werd genoemd, kon met sap worden overgebracht.BehalveChenopodium foetidum enSonchus oleraceus bleken ook 23 van de 34 getoetste soorten uit de familie Solanaceae vatbaar voor dit virus te zijn. Gezuiverde virus preparaten bevatten isometrische deeltjes met een diameter van 27 nm (Fig. 2) Het virus bestaat uit twee deeltjes met sedimentatie-coëfficiënten van 112 en 50 S. Het 112 S deeltje bleek infectieus te zijn, het andere niet. Op grond van de sedimentatiecoëfficiënten kan worden berekend dat het 112 S deeltje 38% nucleïnezuur bevat. Voor de basenverhouding in het nucleïnezuur werd 22,9% adenine, 14,4% guanine, 37,2% cytosine en 25,5% uracil gevonden (Tabel 1). Het hoge gehalte van cytosine kwam ook tot uiting in de U.V. absorptiekromme van het virus en het nucleïnezuur (Fig. 1). Het gezuiverde virus bleek zeer infectieus te zijn; 6000 deeltjes/ml waren in staat een plant van de soortNicotiana clevelandii ziek te maken.Op grond van serologisch onderzoek kon het virus tot de turnip yellow mosaic virus groep worden gerekend. Het vertoonde serologische verwantschap met de Andean potato latent virus (APLV) subgroep (Tabel 2). In premunitieproeven bood het slechts een geringe bescherming tegen APLV en dulcamara mottle virus. Het omgekeerde werd eveneens geconstateerd. De leden van de APLV-subgroep kunnen op grond van hun waardplantenreeks van elkaar onderscheiden worden (Tabel 3).     

Pathotypic variation in turnip mosaic virus with special reference to European isolates

A collection of 124 isolates of turnip mosaic virus was gathered from around the world, principally from European countries, and characterized by inoculation to four differential lines of Brassica napus (oilseed rape and swede). Three symptom phenotypes were induced—apparent immunity, local infection only, or systemic infection. Twelve distinct patterns, i.e. pathotypes, were observed. Three pathotypes were predominant in the collection; pathotype 1 isolates, which were the most common, did not overcome any of the most extreme sources of resistance in the differential lines. Of the other two, pathotype 3 isolates overcame one of the major sources of resistance and pathotype 4 isolates overcame all sources of resistance. The distribution of pathotypes within Europe was examined. No pathotype was confined to any geographical area, although pathotype 4 isolates were not found in southern Europe or Asia. Most isolates (90) originated from Brassica hosts, while others were from other cruciferae genera (19) or non-crucifers (5). The species of plant that the isolates originated from was not clearly related to the pathotype of the isolates. Resistance to pathotype 1 isolates is controlled by a dominant allele in one of the differential lines, and resistance sources are being examined in the other lines. Isolates belonging to pathotype 1 appeared to be able to mutate readily to overcome the resistance in one of the rape differential lines, but no isolates appeared to mutate to overcome the other major source of resistance in the differentials. The implications of the results for disease control strategies are discussed.     

大白菜抗芜菁花叶病毒基因EST-PCR-RFLP分子标记的研究

 本试验以高抗芜菁花叶病毒C₃株系(TuMV-C₃)的高代自交系A₁56-2和感病自交系P9805杂交后代的F₂代为群体,根据大白菜的抗性相关的表达序列标签(EST)设计引物,利用分离群体分群分析法(BSA),筛选出2个与TuMV-C₃株系抗病基因紧密连锁的EST-PCR-RFLP分子标记BS300及BS160,遗传距离均为6.5 cM,为大白菜分子辅助育种、抗病基因克隆以及研究抗病基因编码特性等奠定基础。     

香蕉苞片花叶病毒CP基因的原核表达及抗血清制备

本试验成功构建了香蕉苞片花叶病毒(Banana bract mosaic virus,BBrMV)外壳蛋白(coat pro-tein,CP)基因的原核表达载体,并诱导表达了34 kDa的融合蛋白His.CP。对该原核表达蛋白的可溶性分析表明,该融合蛋白以包涵体形式存在。利用组氨酸标签纯化试剂盒对目的蛋白进行了纯化,获得了高纯度的融合蛋白。以纯化的蛋白为抗原免疫健康家兔,成功制备了抗BBrMV CP基因编码蛋白的兔抗血清。Western-blotting结果表明这种抗血清有很强的特异性。血清效价测定的效价在51 200倍以上,对植物材料的合适检测浓度为1:800～1:3 200。     

雀麦花叶病毒外壳蛋白基因原核表达及抗血清制备

正雀麦花叶病毒(brome mosaic virus,BMV)为正义单链RNA病毒。BMV寄主范围广泛,在波兰等地均有发现(Trzmiel et al.,2015),大多危害禾本科、豆科、茄科等植物,感病植株常表现出褪绿、轻微花叶症状。因此,为减轻BMV造成的经济损失和丰富BMV的检测方式,本研究拟构建BMV的外壳蛋白(coat protein,CP)基因的原核表达载体,在大肠杆菌Escherichia coli中表达BMV的外壳蛋白,制备高特异性和高灵敏度的抗血清,以期用于BMV的有     

小麦FtsH2蛋白与WYMV CP互作研究

 小麦黄花叶病毒(Wheat yellow mosaic virus, WYMV)是马铃薯Y病毒科大麦黄花叶病毒属成员,主要危害冬小麦。实验室前期以WYMV外壳蛋白(coat protein,CP)为诱饵,通过酵母双杂交筛选小麦cDNA文库,发现FtsH2蛋白部分片段与WYMV CP互作。FtsH2蛋白是AAA蛋白酶家族成员,参与植物叶绿体光损伤修复和类囊体发育进程。本研究利用酵母双杂交和双分子荧光互补技术进一步对FtsH2蛋白全长与WYMV CP进行互作验证,并鉴定互作结构域;利用荧光蛋白标记技术研究FtsH2蛋白与CP的亚细胞定位。实验结果表明FtsH2蛋白全长与CP互作;两者互作的关键结构域包含FtsH2蛋白低复杂区、跨膜区及AAA结构域和WYMV CP的中段区域(61-293 aa)。FtsH2蛋白单独表达时定位在细胞质、细胞核和叶绿体;CP单独表达时定位在细胞质;两者共同表达时亚细胞定位均没有发生明显变化,且主要共定位于细胞质中。WYMV CP与小麦FtsH2蛋白的互作可能会干扰植物绿叶体的发育和功能。本研究对了解WYMV的症状形成机制具有一定意义。     

小麦FtsH2蛋白与WYMV CP互作研究

 小麦黄花叶病毒(Wheat yellow mosaic virus, WYMV)是马铃薯Y病毒科大麦黄花叶病毒属成员,主要危害冬小麦。实验室前期以WYMV外壳蛋白(coat protein,CP)为诱饵,通过酵母双杂交筛选小麦cDNA文库,发现FtsH2蛋白部分片段与WYMV CP互作。FtsH2蛋白是AAA蛋白酶家族成员,参与植物叶绿体光损伤修复和类囊体发育进程。本研究利用酵母双杂交和双分子荧光互补技术进一步对FtsH2蛋白全长与WYMV CP进行互作验证,并鉴定互作结构域;利用荧光蛋白标记技术研究FtsH2蛋白与CP的亚细胞定位。实验结果表明FtsH2蛋白全长与CP互作;两者互作的关键结构域包含FtsH2蛋白低复杂区、跨膜区及AAA结构域和WYMV CP的中段区域(61-293 aa)。FtsH2蛋白单独表达时定位在细胞质、细胞核和叶绿体;CP单独表达时定位在细胞质;两者共同表达时亚细胞定位均没有发生明显变化,且主要共定位于细胞质中。WYMV CP与小麦FtsH2蛋白的互作可能会干扰植物绿叶体的发育和功能。本研究对了解WYMV的症状形成机制具有一定意义。     

Changes in the glucosinolate content of oilseed rape plants following infection with turnip mosaic virus

Low and high glucosinolate cultivars of oilseed rape were analysed for variation in both individual and total glucosinolate content up to 27 days after inoculation with turnip mosaic virus. Both infected and control plants of the two cultivars differed in the concentration of aliphatic, aromatic and indole glucosinolates. The biggest differences in glucosinolate content between the cultivars were found in the aliphatic and aromatic glucosinolate groups. Infected plants of the high glucosinolate cultivar, Solida, possessed lower amounts of glucosinolates, than did healthy control plants. Similar results were obtained with a low glucosinolate cultivar, S1. No significant difference was found between the two cultivars in resistance to turnip mosaic virus, using qualitative and quantitative ELISA to determine virus content in their tissues.     

Barriers to gene flow between emerging populations of Watermelon mosaic virus in Southeastern France

Since 1999, "emerging" (EM) strains of Watermelon mosaic virus (WMV) have been detected in cucurbit crops of southeastern France, probably as a result of recent introductions. Population genetic approaches were used to study the structure of EM isolates in southeastern France and to identify factors involved in their spatial distribution. A population clustering method (SAMOVA) and a maximum-difference algorithm (Monmonier's algorithm) were combined to visualize and quantify barriers to gene flow between populations. Both methods yielded similar results and two main barriers were identified. A North/South oriented barrier may be related to physical obstacles to gene flow (Rh?ne River, presence of an area with few cucurbit crops). Although the barrier was very strong, some "crossing" events were detected. A second barrier, oriented Northwest to Southeast, was not correlated with obvious geographical features. The two methods used here are complementary and confirm the limited spread of WMV-EM isolates. This approach can be useful in epidemiology studies to characterize the structure of viral populations and identify barriers to gene flow.     

甘蔗线条花叶病毒P1蛋白基因原核表达及抗血清制备

甘蔗线条花叶病毒Sugarcane streak mosaic virus(SCSMV)是引起甘蔗花叶病的主要病原之一,在世界各大蔗区普遍发生,严重威胁甘蔗产业的发展.建立快速有效的检测方法对于SCSMV的防控有着重要意义.本研究依据SCSMV P1基因序列合成一对引物,扩增获得1 074 bp的目的基因,将目的基因与...     

植物与黄瓜花叶病毒互作的研究

植物对病原物的抗性包括由单基因控制的质量抗病性和由多基因控制的数量抗病性两种。作物对黄瓜花叶病毒病(CMV)的抗病性属于数量抗病性,即有多个微效抗性基因同时起作用控制黄瓜花叶病毒病的感染,其中研究最精细的是其数量主基因RCY1,该基因位于拟南芥5号染色体,编码一套具有卷曲螺旋-核苷酸结合位点-富含亮氨酸重复序列(CCNBLRR)结构的抗性蛋白;黄瓜花叶病毒病的外壳蛋白基因与RCY1基因共同作用决定作物对黄瓜花叶病毒病的抗性反应。但是在其它的互作体系中,可能存在另外的诱发子和抗性基因,到目前为止尚无定论。     

The relationships between pea necrosis virus and bean yellow mosaic virus

A standard pea necrosis virus isolate (PNV-E178) and two isolates resembling PNV (Kow 14 and E242) were fully compared with bean yellow mosaic virus type strain B25 (BYMV-B25). PNV-E178 and PNV-like isolates Kow 14 and E242 resembled each other and the earlier described pea necrosis strain of BYMV in their reaction on pea, but differed from BYMV strains studied so far in inclusion bodies, and in their reaction in cucumber.Serologically, PNV isolates E178 and E242 were closely related to each other and both showed a more distant relationship to BYMV-B25. PNV isolate Kow14 was serologically intermediate between PNV and BYMV-B25, but was hardly infectious toPhaseolus beans.E242 and, to a lesser extent, also Kow14 were considered strains of the pea necrosis virus, which is closely related to BYMV, but apparently not more so than bean common mosaic virus, pea seedborne mosaic virus, clover yellow vein virus and some other members of the potyvirus group.The lack of well-definable borderlines between the different taxonomic entities unavoidably leads to problems in diagnosing (identifying) intermediate isolates.Samenvatting Een aantal virussen kan in erwten ernstige aftervingsverschijnselen doen ontstaan, zoals een eerder beschreven ertwtenecroserivus (PNV-E178) en een erwtenecrosestam van het bonescherpmozaïekvirus (BYMV). Twee nieuwe erwtenecrose-isolaten (Kow14 ene E242) werden bestudeerd en hun verwantschap met het erwtenecroserirus (PNV) en het bonescherpmozaïekvirus (BYMV) gaf aanleiding tot een nieuw onderzoek naar de relatie tussen de twee laatstgenoemde virussen.In hun reactie op erwt en boon leken PNV-E178, Kow14 en E242 veel op elkaar en op de erwtenecrosestam van BYMV. Ze verschilden echter van de mozaïekstam (B25) van dit virus in de geproduceerde celinsluitsels, vooral wat betreft de sterke vergroting der nucleodi en bij E178 bovendien door de opvallende uitstaande kristalnaalden in de celkern. Ook zijn de eerste drie isolaten duidelijk minder pathogeen voorPhaseolus-bonen (Fig. 1) dan de bonemozaïkstam en de erwtegeelmozaïekstam van het BYMV. Op deze plantesoort doet Kow14 slecht een gering aanal lokale vlekken ontstaan. Verder wordenNicotiana-soorten gemakkelijker door de drie isolaten aangetast (Fig. 2 en Tabel 1). Deze laatste worden ook niet veroorzaakt door de erwtenecrosestam van het BYMV, hoewel ook deze vrij gemakkelijkNicotiana-soorten aantast (o.a. het nu in de serologieproef gebruikte isolaat E221).Serologisch zijn PNV-E178 en E242 nauw aan elkaar verwant en zijn beide minder nauw verwant aan B25 (Tabel 2). Kow14 neemt serologisch een tussenpositie in, hoewel dit isolaat in biologisch opzicht meer een uiterste is omdat het nauwelijks infectieus is voorPhaseolus-bonen.E242, en hoewel minder verwant, ook Kow14, worden nu beschouwd als stammen van het erwtenecrosevirus (Tabel 3). Dit virus is weliswaar vrij nauw verwant aan het bonescherpmozaïekvirus, maar niet nauwer dan het bonerolmozaïekvirus, het erwterolmozaïkvirus, het nerfvergelingsvirus van klaver en enkele andere leden van de aardappelvirus-Y-groep.Het klaarblijkelijk on tbreken van shcerpe grenzen tussen de verschillende taxonomische eenheden leidt onvermijdelijk tot moeilijkheden bij de beschrijving en herkenning van intermediatire isolaten(tussenvormen).Guestworker from February through May 1, as afellow of the Netherlands Ministry of Education, research plant virologist of the Institute for Plant Protection, Budaperst, Hungary.