Competition Between Cucumber Mosaic Virus Subgroup I and II Isolates in Tobacco

Historical reports indicate that Cucumber mosaic virus (CMV) strain subgroup I is more prevalent than subgroup II in most parts of the world, but recent reports suggest that subgroup II isolates may be far more abundant than previously found in China. In order to evaluate the dominance of CMV subgroup I and subgroup II strains in co-infected tobacco plants, four isolates, NX and YQ in subgroup I, and ZL and AG in subgroup II, were tested in competition experiments. In these comparisons, the frequency of infection was assessed, and ratios between singly and doubly infected plants were calculated based on ELISA tests of tobacco leaves. In contrast to previous reports suggesting that subgroup I strains are usually more competitive than subgroup II strains in the field, the results from the present study indicate that the subgroup II ZL isolate was more competitive than the subgroup I YQ isolate, even though the ZL isolate caused milder symptoms than YQ in singly infected tobacco. In contrast, the subgroup I strains NX and YQ were more competitive than subgroup II AG. This information provides evidence for variation in the competitive abilities of subgroup II strains in tests with subgroup I strains, and suggests that direct competition during mixed infections may account in part for the recent spread of some subgroup II strains in China and elsewhere.     

不同CMV分离物侵染寄主的超微结构变化

应用电镜观察了黄瓜花叶病毒CMV不同分离物侵染寄主的细胞 超微结构变化。来自一串红(Salvia splendens)的不含卫星RNA分离物M-22侵染心叶烟,病毒粒子散布于细胞质,在液泡中形成大片病毒粒子结晶,液泡膜边缘产生小泡结构,完整的病毒粒子穿过胞间连丝在细胞间运转,胞间连丝中央部分有扩张现象。自然感染三生烟的含坏死卫星RNA分离物8-S1侵染普通烟,病毒粒子分散于细胞质,在液泡中未观察到结晶体,叶绿体产生囊泡结构,部分病毒粒子处在叶绿体空泡中。田间寄主上受8-S1侵染的三生烟细胞质中分布着大量球形病毒粒子,叶绿体也产生含有病毒粒子的囊泡结构。表明含有卫星RNA和不含卫星RNA的CMV分离物引起的细胞病变特征存在差别,可能是CMV卫星RNA参与病理变化的依据之一。     

不同CMV分离物侵染寄主的超微结构变化

应用电镜观察了黄瓜花叶病毒ＣＭＶ不同分离物侵染寄主的细胞超微结构变化。来自一患红（Ｓａｌｖｉａｓｐｌｅｎｄｅｎｓ）的不含卫星ＲＮＡ分离物Ｍ－２２侵染心叶烟,病毒粒子散布于细胞质,在液泡中形成大片病毒粒子结果,液泡膜边缘产生小泡结构,完整的病毒粒子穿过胞间连丝在细胞间运转,胞间连丝中央部分有扩张现象。     

黄瓜花叶病毒及抗病转基因烟草研究进展

黄瓜花叶病毒(Cucumber mosaic virus,CMV)是世界上分布最广的植物病毒之一,其株系繁多,给烟草生产造成很大的损失。近些年来,各国学者对CMV开展了大量研究。该文主要介绍了黄瓜花叶病毒的基因组结构、亚组以及抗CMV的烟草基因工程的研究进展。     

Differentiation of Bulgarian Isolates from Cucumber Mosaic Virus by Serological Methods

Cucumber mosaic virus (CMV) is of great importance to the Bulgarian economy and hence a detailed knowledge of its diversity under local geographic and climatic conditions is required. An extended study was carried out on CMV strains the currently occur in Bulgaria. Fifty-one isolates and strains found in different regions and various crops were biologically characterized and serologically differentiated into subgroups I and II using different variants of enzyme-linked immunosorbent assay (ELISA) [double antibody sandwich (DAS)-, antigen-coated plate (ACP)-, triple antibody sandwich (TAS)- with poly and monoclonal antibodies] and immunodiffusion tests. The ELISA modifications with monoclonal antibodies individually (ACP) or in combination with polyclonal antibodies (TAS-ELISA) are suitable for mass screening of CMV isolates. The hyperimmune sera against strains from CMV subgroups I and II were very efficient for use in isolate differentiation via gel double immunodiffusion. The results obtained correlated with the polymerase chain reaction and restriction fragment length polymorphism data reported by other authors. The majority of the isolates belonged to subgroup I, whereas 10, mainly from tomato and pepper, belonged to subgroup II. Most of the subgroup II isolates came from the north of Bulgaria. The results of the present study will help to clarify the virus epidemiology and to develop specific control measures.     

侵染番茄的黄瓜花叶病毒（CMV）株系特性的比较研究

从山东省主要番茄种植区病毒样本中分离到111个黄瓜花叶病毒(CMV)分离物,根据病毒的寄主范围和症状,初步鉴定为三个株系:番茄蕨叶株系(CMV-ToF)、番茄花叶株系(CMV-ToM)和番茄轻花叶株系(CMV-ToL)。三株系除在生物学特性存在明显差异外,其病毒粒体形态大小,电泳迁移率,病毒外壳蛋白亚基分子量、核酸组分以及病毒粒体血清学特性亦存在差别。     

Problems Encountered with the Selection of Cucumber Mosaic Virus (CMV) Isolates for Resistance Breeding Programs

Among the Chili breeding lines from the Asian Vegetable Research Center, two were chosen for the screening of a larger selection of Cucumber mosaic virus (CMV) isolates, mainly from Asian countries. The chili line (VC246) showed a resistance against several CMV‐isolates and was compared with chili line VC27a that was susceptible to CMV infection. Among the 28 CMV isolates, five were identified as resistance breaking (AN‐like) and non‐resistance breaking (P3613‐like) for the line VC246, whereas all isolates could establish a systemic infection on VC27a. However, further testing revealed that resistance in VC246 was also dependent on the way of inoculation and the inoculums itself. Graft inoculation could overcome the resistance, and the inoculation with isolated viral RNA resulted in no infection at all on the resistant chili line, independent of the virus isolate. Using a pseudo‐recombinant approach, we identified RNA2 of resistance breaking isolates as responsible for systemic infection and confined the area within RNA2 to the 3′ terminal part including the ORF 2b. Sequence alignments of that area revealed eight distinct mutations on amino acid level, which was present either in resistance or non‐resistance breaking isolates. A reversion from the P3613‐like to the AN‐like sequence of two of these mutations induced no effect on Capsicum sp., but induced symptoms on several tobacco species distinct from those induced by the wild‐type virus. However, pseudorecombinants, each generated from sets of two different AN‐like isolates, which were expected to infect VC246 systemically, did not indicating that probably RNA2 must be in a specific context to have the effect. In this case, a generalized attribution of functions to single amino acid exchanges might be impossible or at least extremely difficult.     

Transmission of Zucchini yellow mosaic virus by Colonizing and Non-colonizing Aphids in Greece and New Aphid Species Vectors of the Virus

Nineteen aphid species were tested for their ability to transmit Zucchini yellow mosaic virus (ZYMV) from and to zucchini under laboratory conditions. Sixteen species were found to be new vectors of ZYMV (i.e. Aphis craccae, Aphis fabae, Aphis nerii, Aulacorthum solani, Brachycaudus cardui, Brevicoryne brassicae, Hyalopterus pruni complex, Hyperomyzus lactucae, Macrosiphoniella sanborni, Macrosiphum rosae, Metopolophium dirhodum, Myzus cerasi, Rhopalosiphum maidis, R. padi, Semiaphis dauci and Sipha maydis). Their transmission efficiency by a single aphid was low (0.1–4.2%). Myzus persicae was used as a control and was the most efficient vector (41.1%, one aphid per plant). Hayhurstia atriplicis, Myzus ascalonicus and Sitobion avenae did not transmit the virus. In four out of six new vectors assayed in arena tests for propensity estimation, propensity was higher than efficiency. Data from an experimental zucchini field in northern Greece revealed a high correlation between ZYMV spread and alatae of the vector species. The most abundant aphid vectors during 2 years experimentation were M. persicae, Aphis gossypii and Aphis spiraecola. The possible role of the 16 new and the previously known aphid vectors in the epidemiology of ZYMV was investigated using data of transmission efficiency combined with the captures of their alatae in the Greek net of a Rothamsted type suction trap.     

Aphid Transmission of Potato aucuba mosaic virus Strains Mediated by Different Strains of Potato virus Y

Three British strains of potato aucuba mosaic virus (PAMV) were tested for transmissibility by the aphid Myzus persicae. None was aphid transmissible on its own but all three were transmitted in the nonpersistent manner by aphids that had previously been fed on a source of the potyvirus potato virus Y (PVY). Different PVY strains mediated PAMV transmission from Nicotiana clevelandii to Capsicum annuum to different degrees, and different PAMV strains were transmitted at different frequencies when assisted by the same PVY strain. These results are compatible with the idea that subtle differences in the PAMV coat protein and in the PVY helper component are responsible for diffrences in frequencies of transmission of PAMV, without however, excluding the possibility of effects of other undefined factors. Transmission of PAMV was no less frequent when mediated by a PVY strain that was unable to infect C. annuum than when a C. annuum‐infecting PVY strain was used.     

侵染菊花的黄瓜花叶病毒的初步鉴定和血清学检测

侵染菊花的黄瓜花叶病毒的初步鉴定和血清学检测张海保,朱西儒,张云开（中国科学院华南植物研究所广州５１０６５０）关键词菊花,黄瓜花叶病毒,间接ＥＬＩＳＡ菊花（Ｃｈｒａｙｓａｎｔｈｅｍｕｍｍｏｒｉｆｏｌｉｕｍ）病毒病是危害菊花的一类主要病害。国外文献报道...     

Biological and Molecular Characterization of Taiwanese Isolates of Cucumber mosaic virus Associated with Banana Mosaic Disease

Banana mosaic disease (BMD) caused by Cucumber mosaic virus (CMV) has become an important threat to the banana industry. We collected and characterized 10 CMV isolates associated with BMD in Taiwan and compared their biological characteristics and coat protein sequences. The isolates fell into four pathotypes on the basis of the symptoms they induce on banana, Nicotiana glutinosa and Vigna unguiculata (cowpea). Double-stranded RNA analysis revealed that the different pathotypes are not related to the presence of CMV satellite RNA. Phylogenetic analysis of worldwide CMV coat protein sequences revealed that among the currently known CMV subgroups IA, IB and II, subgroup IB is phylogenetically unresolved. Our CMV isolates form a new subgroup, IT, within subgroup I. In addition, we resolved another new CMV subgroup, IS, within subgroup I. The analysis also revealed that isolates within different subgroups can infect the banana.     

黄瓜花叶病毒（CMV）运动蛋白基因介导的抗病性

利用Ｆｎｙ＿ＣＭＶ株系ＲＮＡ３ｃＤＮＡ克隆,构建了含有全长和编码区缺失５０１个核苷酸的运动蛋白（ＭＰ）基因植物表达载体ｐＢＭＰＲ和ｐＢＭＰＫ。在土壤农杆菌（Ａｇｒｏｂａｃｔｅｒｉｕｍｔｕｍｅｆａｃｉｅｎｓ（ＳｍｉｔｈｅｔＴｏｗｎｓｅｎｄ）Ｃｏｎｎ）ＬＢＡ４４０４介导下转化烟草（ＮｉｃｏｔｉａｎａｔａｂａｃｕｍＬ．）品种“ＮＣ８９”,分别经Ｓｏｕｔｈｅｒｎｂｌｏｔｉｎｇ、ＲＴ＿ＰＣＲ或Ｗｅｓｔｅｒｎｂｌｏｔｉｎｇ分析,外源基因已整合到再生植株中并得到表达。抗病性分析表明,含有缺失型ＭＰ基因的Ｒ０代转基因植株抗性较好,接种５０ｄ后,１０株转化植株中仍有５株不表现症状。在自然发病条件下,这５个含有缺失型ＭＰ基因转基因株系在Ｒ１代都表现了一定的抗病性。抗性主要表现为症状出现推迟,严重度减轻。利用ＰＣＲ筛选、种子卡那霉素抗性试验和温室抗病性测定等方法,初步认为Ｒ２代转基因烟草Ｋ＿６＿５株系为转基因抗病纯合系。而含有全长ＭＰ基因的Ｒ０代转化植株,前期没有表现明显的抗病性,但在接种４０ｄ后部分发病植株有恢复健康的趋势。     

Virus Movement Protein Gene Mediated Resistance Against Cucumber Mosaic Virus Infection

Tobacco (Nicotiana tabactum L. ) "NC89" plants were transformed with deletion mutant of cucumber mosaic virus (CMV) movement protein (MP) gene and full-length CMV MP gene, respectively. The transformed plants were analyzed with polymerase chain reaction (PCR), PCR-Southem, Southern and Western blots. R0 generation of the transgenic plants were inoculated with CMV. Five out of 10 lines of tobacco plants (BMPK) transformed with CMV MP deletion mutant gene showed high resistance to CMV infection and remained symptomless for up to 50 days post-inoculation. In contrast, tobacco plants (BMPR) transformed with full-length CMV MP gene did not show resistance to CMV infection. However, most of the infected full-length CMV MP gene transgenic plants recovered by showing none or very mild mosaic symptoms in 40 days post-inoculation. The results of R1 generation of the BMPK transgenic plants tested under field conditions showed that all 5 lines of transgenic plants could delay the virus disease development.     

黄瓜花叶病毒CP基因原核表达及抗血清的制备

把黄瓜花叶病毒 (CMV)西番莲分离物的外壳蛋白 (CP)基因 ,通过BamHI SacI位点定向插入pET 2 2b( )载体 ,转化大肠杆菌BL2 1 (DE3)中 ,经IPTG诱导下 37℃培养 6h ,SDS PAGE电泳示表达蛋白分子质量为 31 8kDa ,表达量占菌体总蛋白的 2 8 9%,表明该蛋白得到了高效表达。用冰冷的氯化钾溶液显色 ,用表达的特异蛋白质条带制备抗原 ,免疫家兔制备出病毒特异抗血清。采用ID ELISA测定抗血清效价为 1 0 -6;抗血清和CMV几个分离物均有特异反应 ,和TMV、菌体蛋白不发生非特异性反应 ;检测病毒灵敏度达 30ng ml,能够从稀释 31 2 5倍的感病植物汁液中检测出病毒     

侵染香蕉的黄瓜花叶病毒分离物衣壳蛋白的肽链分析

香蕉花叶病为华南香蕉生产的重要限制因素之一［‘1。特别是随着组培苗应用面积的扩大,发生越来越严重,一般地块发病率为20—40％,个别重病地块高达90％,损失严重。此病病原虽在1930年【’1就被鉴定为黄瓜花叶病毒（Cucumbermosaicvirus,CMV）,但至今对其研究不多。从有限的文献看,大多数作者【’·’嘟认为其病原为CMV的一个株系,即香蕉株系。但也有认为为二个株系［‘］、或三个株系的［’］。因此有关病原的调查鉴定就成为当前迫切需要解决的一个重要问题。1991～1993年我｛I’J在广东省广州市天河和黄埔区,以及顺德、番禹…     

Structural Analysis of A-Protein of Cucumber Green Mottle Mosaic Virus and Tobacco Mosaic Virus by Synchrotron Small-Angle X-Ray Scattering

The size and shape of A-protein of tobacco mosaic virus coat protein (TMVP) and cucumber green mottle mosaic virus coat protein (CGMMVP) were evaluated by means of small-angle X-ray scattering (SAXS) using a synchrotron radiation source, complemeted by electron microscopic observations. The results imply that TMV and CGMMV A-proteins are composed of three and two subunits, respectively, stacked in the shape of an isosceles triangular prism at lower ionic strength. Considering the difference of the A-protein structure at higher and lower ionic strength, the globular core structure was proposed as a subunit which might be modeled as a thin isosceles triangular prism composed of four globular cores joined by rather flexible segments. These cores correspond probably to four helical regions in a subunit, and rearrange their relative positions according to the external conditions. A slight rearrangement of core positions in a subunit may result in the formation of A-proteins of various shapes.     

核酸酶保护试验在黄瓜花叶病毒株系鉴定中的初步应用

采用黄瓜花叶病毒（ＣＭＶ）亚组Ⅰ株系Ｆｎｙ－ＣＭＶＲＮＡ＿２的１２０９～１６２６核苷酸片段和亚组Ⅱ株系Ｌｓ－ＣＭＶＲＮＡ＿２的２００２～２４３３核苷酸片段的ｃＤＮＡ克隆,体外转录,同时掺入￣（３２）Ｐ获得负链ＲＮＡ探针,与纯化的番茄和甜椒上的ＣＭＶ中国分离物的ＲＮＡ杂交,结果表明：ＣＭＶ番茄和甜椒中国分离物与Ｆｎｙ－ＣＭＶ的核苷酸有高度同源性,隶属于Ｆｎｙ－ＣＭＶ为代表的亚组Ⅰ株系。并利用Ｋ－ＣＭＶ株系（亚组Ⅰ,源于中国）的ＲＮＡ＿２全长ｃＤＮＡ克隆的两个ＥｃｏＲＩ位点间的核苷酸序列（１６５７～２１２５ｎｔ）作探针,与上述两种ＣＭＶ中国分离物的ＲＮＡ杂交,进一步比较分析了这两个分离物和Ｋ－ＣＭＶ株系的关系。讨论了核酸酶保护法在ＣＭＶ株系鉴定中的作用。