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。     

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.     

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.     

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。     

Transmission Comparisons of Cucumber Mosaic Virus Subgroup I and II Isolates by Different Aphid Species

To investigate the transmission differences between Cucumber mosaic virus (CMV) subgroup isolates, we carried out a comparative study with five aphid species Myzus persicae, Aphis gossypii, Lipaphis erysimi, Aphis craccivora and Megoura viciae in laboratory and field experiments to evaluate spread of CMV Subgroup I NX and subgroup II AG isolates in tobacco. Both NX and AG varied in transmission efficiency by the five aphids, and our transmission results revealed important differences in transmission efficiency of two isolates by Myzus persicae and Aphis gossypii. In contrast, significant transmission differences were not detected with Lipaphis erysimi, Aphis craccivora or Megoura viciae. Interestingly, the overall transmission efficiencies of the two different subgroup strains were almost equal when field transmissions were tested with mixed populations of the five aphid species. Our results together with our previously reported experiments on competition of CMV subgroup isolates in tobacco suggest that variations in aphid vector populations contribute substantially to the epidemic potential of CMV subgroup isolates.     

用黄瓜花叶病毒卫星RNA生防制剂大面积防治麦茬辣椒病毒病

用黄瓜花叶病毒（ＣＭＶ）卫星ＲＮＡ生防制剂Ｓ＿（５２）免疫辣椒,能防治辣椒由ＣＭＶ引起的病毒病。用Ｓ＿（５２）免疫接种辣椒的大田对比试验,防效达５９．５－７１．０％,产量比对照增加２６．１－３２．２％,产值增加３３．５－４５．２％。小区对比试验,用Ｓ＿（５２）免疫接种５０天的辣椒,防效达７１．３－９２．９％,免疫接种８０天,防效达５５．９－７８．５％,产量比对照增加３６．０－６５．３％,产值增加４９．３－９３．５％,还有刺激生长,促进早熟和增强对真菌、细菌病害抵抗作用。     

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。     

黄瓜花叶病毒衣壳蛋白基因转化辣椒研究

The plant expression vector of the coat protein(CP) gene of cucumber mosaic virus (CMV) BS strain was used to transform three kinds of pepper (Capsicum annuum) tissues (cotyledon, stem and root) by agrobacterium-mediated co-cultivation. 53%-68.4% of the total tissues (639) can be induced to be calli, but only cotyledon calli can be further regenerated to form shoots (regenerated efficiency 39.7%). 70%(42/60) of the putative transformed plants were confirmed to have CP gene in their genomes by Southern blot. The mRNAs and the CP were respectively found in 80% of transgenic plants by Northern blot and DAS-ELISA. 24 of the transgenic plants expressing CP gene of BS strain showed three kinds of resistant level (severe symptom, delay of symptomatic development, no symptom) to infection of CMV-BS and of CMV-P. However, there was distinctly higher resistance to inoculation of CMV-BS than that with CMV-P in these transgenic plants.     

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...     

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.     

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     

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。     

CMV抑制PVYN CP基因沉默及其介导的抗病性

以前曾报道用RNA介导的抗病毒策略,获得了高度抗病的表达马铃薯Y病毒坏死株系外壳蛋白基因(PVY^N CP)的转基因烟草,并对T1、T2代转基因植株进行了遗传和抗病性分析。此次以T,代转基因植株为试验材料,在筛选高度抗病植株并证明其抗病性是基于转基因沉默的基础上,采用Northern杂交的方法,证明CMV侵染抑制了转基因植株中PVY^N CP基因的沉默,而且CMV对PVY^N CP基因沉默的抑制部位是发生在接种后的新生叶上,接种叶及其下部叶片中PVY^N CP基因沉默则未受到影响。采用ELISA方法对CMV PVY^N复合接种的转基因植株进行PVY^N检测,结果表明,接种叶及下部叶没有检测到PVY^N,植株叶片对PVY^N表现为抗病。而在CMV接种后植株新生叶中则检测出了高滴度的PVY^N,植株叶片对PVY^N表现为感病。该文报道了在表达PVY^N CP基因的RNA介导抗性转基因植株中,异源病毒侵染抑制了转基因的沉默,并导致转基因植株的抗病性丧失。     

黄瓜绿斑驳花叶病毒辽宁分离物全基因组序列测定

以感病组织总RNA为模板,采用RT-PCR方法扩增并测定黄瓜绿斑驳花叶病毒(Cucumber green mottle mosaic virus,CGMMV)辽宁分离物(CGMMV-LN)的基因组全序列。CGMMV-LN基因组全长6 422 nt,5'非编码区(noncoding region,NCR)和3'NCR分别为59 nt和175 nt。CGMMV-LN编码的4个蛋白依次是186 kD和129kD的复制酶,29 kD的移动蛋白和17.4 kD的外壳蛋白。CGMMV-LN与其他4个CGMMV分离物基因组核苷酸序列同源性为97.6%～99.3%,与同属其他3种病毒基因组核苷酸序列同源性仅为61.7%～62.8%。基于186kD复制酶和外壳蛋白氨基酸序列的同源树显示:侵染葫芦科作物的烟草花叶病毒属病毒可分为2个亚组,亚组I包括所有CGMMV分离物,亚组II包括Kyuri绿斑驳花叶病毒(Kyuri green mottle mosaic virus,KGMMV)、黄瓜果实斑驳花叶病毒(Cucumber fruit mottle mosaic virus,CFMMV)和小西葫芦绿斑驳花叶病毒(Zucchini ...     

小麦梭条花叶病抗性遗传和育种研究进展

由土壤真菌禾谷多粘菌(Polymyxa graminis)传播的小麦梭条花叶病在我国长江流域和黄淮平原麦区已成为危害小麦生产的一种严重病害。本文系统地综述了我国在小麦梭条花叶病种质资源筛选、抗性机理、抗性遗传、抗病基因的分子标记以及抗病育种方面的进展。研究表明,在我国地方品种和改良品种中存在着较丰富的抗病资源;植物体内某些酶的活性和可溶性糖含量与抗性有密切的联系;抗病性表现为数量性状的遗传特征,可能受1~3对显性基因控制。通过抗感品种间杂交,可以育成抗病丰产的新品种。文中还对今后开展小麦抗梭条花叶病育种提出了几点建议。