我国部分地区常见农作物上黄瓜花叶病毒分离物核酸多样性分析

为明确我国黄瓜花叶病毒株系分化及系统进化基本情况,从湖南、新疆、青海和海南4省区采集1 367个样品对其进行酶联免疫和RT-PCR检测,并对分离获得的15个黄瓜花叶病毒(Cucumber mosaic virus,CMV)纯化分离物CP、MP、2b核苷酸序列进行相似性和进化树分析及生物学性状比较。结果表明,辣椒、龙葵和黄瓜的CMV阳性检出率较高,分别为54.13%、29.19%和18.46%。进化树分析显示CMV-Q5与CMV亚组II的亲缘性较高;CMV-N7为新发现的重组株系,其CP、2b基因属于CMV亚组IB,MP基因却属于CMV亚组II;其余13个分离物均属于CMV亚组IB。CMV-N7和CMV-Q5在系统寄主心叶烟和枯斑寄主苋色藜上引发的症状相似,但比对照株系CMV-P3613(IB)的发病时间要晚1~2 d,系统花叶较温和,枯斑较小。表明在以上4省区常见农作物上广泛流行的CMV存在分子变异。     

植物病毒卫星RNA生防制剂CMV-S52防治番茄病毒病

 1976年Kaper等发现,有些黄瓜花叶病毒(CMV)中除含有CMV基因组RNA1-4外,还含有1种可伴随CMV复制的RNA5,称CMV卫星RNA,它的存在能够改变CMV在各种寄主上的症状及其严重程度,还能干扰CMV-RNA1-4的复制,降低植株内病毒浓度。     

引起番茄植株坏死病的病毒研究

 2004~2005年,在上海郊区夏番茄上连续发生严重的植株坏死病。表现为番茄植株矮化、顶端坏死、果实畸形和整株枯萎症状,并在叶柄和茎干上出现不规则坏死条斑,具有典型的病毒病特征。通过采集典型病叶、病果,进行病毒分离、dsRNA分析、寄主生物学研究、病毒纯化、病毒蛋白分析、组织病理学与形态学观察和ELISA检测,初步确定是由番茄花叶病毒(Tomato mosaic virus,ToMV)和黄瓜花叶病毒(Cucumber mosaic virus,CMV)侵染。经5次心叶烟单斑分离,获得N5分离物,对CP基因克隆测序的结果确定该分离物为ToMV。寄主反应测定显示,N5的寄主反应与常规ToMV株系存在明显区别,主要表现为常规番茄品种上出现严重坏死。进一步接种GCR品系番茄鉴定N5与ToMV-1株系相似。因此,作者认为,在上海地区番茄上流行并引起坏死病的主要病原可能是ToMV-1株系的1个变异株。     

黄瓜花叶病毒对烟草花叶病毒的干扰作用的研究初报

 于1981年-1985年研究了黄瓜花叶病毒(CMV)对烟草花叶病毒(TMV-T)的干扰作用.用CMV向日蔡环斑花叶分离物(CMV-SRS)为诱导病毒,接种枯斑三生烟十天后,对攻击病毒TMV-T的保护率为85.6%.反之,用TMV-T为诱导病毒,接种枯斑三生烟十天后,对攻击病毒CMV-SRS的保护率为95%(70-100%)证明在CMV-SRS和TMV-T之间,有较强的相互干扰作用.含有卫星核酸的黄瓜花叶病毒CMVS51、及CM-VS52两个分离物以及CMVS51+TMVN-14(烟草花叶病毒弱株系)免疫心叶烟和枯斑三生烟十天后,对强病毒TMV-P有明显的干扰作用.     

北京地区辣椒病毒病毒原种类及黄瓜花叶病毒株系鉴定

近十年来,经鉴定,明确了北京地区辣椒病毒病毒原有7种,即黄瓜花叶病毒(CMV)占病株样品的55％,烟草花叶病毒(TMV)占26％,马铃薯Y病毒(PXY)占13.2％,烟草蚀纹病毒(TEV)占11.8％,马铃薯X病毒(PVX)占10.4％,苜蓿花叶病毒(AMV)占2％,及蚕豆萎蔫病毒(BBWV)占1.4％。其中CMV可划分为4个株系,即重花叶株系,坏死株系,轻花叶株系和带状株系。     

北京地区辣椒病毒病毒原种类及黄瓜花叶病毒株系鉴定

近十年来,经鉴定,明确了北京地区辣椒病毒病毒原有7种,即黄瓜花叶病毒(CMV)占病株样品的55％,烟草花叶病毒(TMV)占26％,马铃薯Y病毒(PXY)占13.2％,烟草蚀纹病毒(TEV)占11.8％,马铃薯X病毒(PVX)占10.4％,苜蓿花叶病毒(AMV)占2％,及蚕豆萎蔫病毒(BBWV)占1.4％。其中CMV可划分为4个株系,即重花叶株系,坏死株系,轻花叶株系和带状株系。     

美国加利福尼亚州的葫芦科植物病毒病——一个不断变化的问题

早期发现的病毒:早在1959年发表的有关加州瓜类病毒病害的综合性报告说明,西瓜花叶病毒(WMV)、黄瓜花叶病毒(CMV)和西葫芦花叶病毒(SqMV)是发生最为普遍的病毒病。在南部沙漠盆地,WMV的株系最为流行,CMV居中,SqMV少见。在加州中部和北部,CMV和SqMV占优势,而WMV次之。10年后发表的报告再次说明,WMV、     

系统侵染的番茄植株中黄瓜花叶病毒的时序变化

 采用实时荧光定量PCR (FQ-PCR)和DAS-ELISA方法,研究了22~26℃温室条件下番茄幼苗中黄瓜花叶病毒CNA株系(CMV-CNA)各基因组RNA组分及其外壳蛋白(CP)含量的动态变化,同时结合同期感病植株症状发展和病情指数,分析并探讨CMV各基因组RNA、CP以及病症显示程度之间的时间效应及其相关性。以18S rRNA为内参照,FQ-PCR相对定量分析结果显示:接种后5~30 d,CMV三分体基因组RNA在系统侵染的番茄组织中负荷量变化趋势大体一致,但是不同时期含量差异显著,均经历对数增长期、稳定期和回落期。其中,以RNA2负荷量变化情况最为平缓。DAS-ELISA检测结果显示:CP含量随接种时间延长而持续升高,但其对数增长趋势相对滞后于基因组RNA。番茄幼苗发病症状与CMV基因组RNA及CP负荷量的变化趋势大体一致,但症状表现时间相对滞后。CMV-CNA株系在番茄幼苗中以基因组RNA、CP以及病症显示先后次序出现高峰期,显示病毒基因组RNA及其CP在植物组织内负荷量的变化与植株症状表现并不同步。其动态变化规律将为研究CMV侵染机制,病毒与寄主互作及防病控病提供量化依据。     

百合黄瓜花叶病毒及其检测

应用酶联免疫吸附法(ELISA)检测了浙江丽水和杭州的东方百合、亚洲百合等栽培品种上黄瓜花叶病毒(CMV),64个田间样品中有26个带毒,检出率为40.6%;dsRNA检测结果与已知CMV-FQS株系的电泳条带相同,但百合组织中CMVdsRNA含量较低;电镜观察,受CMV侵染的样品中大多含有线状病毒,复合侵染率为35.9%;寄主反应测定显示从百合植株上获得的CMV株系均不能通过汁液摩擦接种侵染昆诺藜、苋色藜、普通烟、心叶烟等6科10种指示植物。     

马来西亚某些蔬菜作物病毒病的发生情况

马来西亚辣椒、甜椒、番茄、茄子、菠菜和黄瓜病毒病发生情况调查表明:80%以上的辣椒感染花叶病,并导致辣椒减产。鉴定了9个辣椒病株标样,其中3个标样由辣椒脉斑驳病毒(Chilli veinal mottle virus,CVMV)单独侵染,其余为两种病毒复合侵染,CVMV 和黄瓜花叶病毒普通株系(CMV—O)、CVMV 和烟草花叶病毒番茄株系(TMV-t)、CVMV 和 CMV 的辣椒株系(CMV-C)复合侵染分别占标样数的2,2,1,1。甜椒病毒病的发生率低于辣椒(由于绝大部分甜椒在塑料大棚栽培的缘故),鉴定     

瞬时表达靶向TMV外壳蛋白基因的siRNA能干扰病毒侵染

 RNA干扰(RNA interference,RNAi)是与内源性mRNA编码区某段序列同源的双链RNA导入细胞后,该mRNA发生特异性降解,从而导致该基因表达沉默的现象。小干扰RNA(small interfering RNA,siRNA)作为RNAi途径的重要中介,已被广泛应用于动、植物抗病毒治疗研究。本文以烟草花叶病毒(Tobacco mosaic virus,TMV)外壳蛋白基因为靶位,设计合成表达小干扰RNA的寡核苷酸,亚克隆到植物双元表达载体pBI121中,直接转化根癌农杆菌。通过根癌农杆菌介导的瞬时表达法,研究了同源于TMV外壳蛋白的siRNA对TMV侵染的干扰作用。结果表明,瞬时表达的siRNA能够特异性干扰TMV侵染。含有重组表达载体pBI121/siRNA的根癌农杆菌渗入普通烟植株,在TMV接种后14d其上部叶片没有表现典型的花叶症状。对这些叶片进行Northern杂交试验也没有检测到TMV病毒的RNA积累或仅有很少量的积累。在枯斑寄主心叶烟上,siRNA的瞬时表达可使TMV侵染后的枯斑数明显减少,甚至不产生枯斑。此外,同源于TMV外壳蛋白的siRNA瞬时表达对非同源的黄瓜花叶病毒(Cucumber mosaic virus,CMV)没有抑制作用,表明siRNA的干扰作用具有高度的同源依赖性。     

Characteristics of a virus inhibitor from the leaf extract of Celosia cristata

A virus inhibitor in Celosia was shown to have moderate thermal stability and resistance to dilution and drying. It was non-dialysable and non-sedimentable and showed glycoproteinaceous characteristics with a molecular weight between 21 000 and 22 000. The inhibitory activity was not reversible by simultaneous application of actinomycin-D to the leaf surface of Nicotiana glutinosa and the virus was fully recovered from a mixture of tobacco mosaic virus and inhibitor, indicating that the inhibitor did not appear to act on the virus directly but interfered with virus establishment in the host.     

黄瓜花叶病毒甜菜分离物的分离鉴定

 从新疆石河子地区春季采种甜菜和夏播母根用甜菜上分离到3个病毒分离物,代号为石-B-2、石-B-5和石-B-D,自然感染甜菜引起叶片黄色花叶、扭曲、皱缩和植株严重矮化。病毒粒体球状、直径28~30 nm、均含有分子量约为2 9k D外壳蛋白、3种相同的较大片段的双链RNA (即3400 bp的RNA1、3100 bp的RNA₂、2300 bp的RNA₃)和片段大小约400 bp的小RNA,石-B-2和石-B-D具有1100 bp的RNA4,在ELISA和琼脂双扩散试验中,3个分离物均与CMV-83抗血清有特异反应,初步认为这3个球状病毒分离物属于CMV,但它们具有狭窄的寄主范围,不感染心叶烟、蔓陀罗、番茄,在寄主反应、双链RNA4和卫星RNA的大小上明显不同于国内外报道的CMV株系或分离物,同时说明侵染甜菜的CMV可能存在着株系分化。     

Tomato yellow leaf curl virus: host range and virus-vector relationships

The Jordanian isolate of tomato yellow leaf curl virus has a narrow host range restricted to a few solanaceous plants. Severe symptoms developed on tomatoes and Datura stramonium , whereas Nicotiana glutinosa and N. tabacum cvs Samsun and Havana 423 were infected without showing symptoms. The whitefly Bemisa tabaci is an efficient vector; a single whitefly was able to transmit the virus. The minimum acquisition and inoculation feeding periods were 60 and 30 min, respectively, and the latent period was 20–24 h. The virus was retained by Bemisia tabaci for 11 days. The host range and virus vector relationships of the Jordanian isolate are reported for the first time.     

Ageratum yellow vein China virus Is a Distinct Begomovirus Species Associated with a DNAbeta Molecule

ABSTRACT Ageratum conyzoides plants exhibiting yellow vein symptoms, collected near Haikou, Hainan Province, China, contained begomoviral DNA-A-like molecules. The complete sequences of the molecules from two samples, Hn2 and Hn2-19, were shown to consist of 2,768 and 2,748 nucelotides (nt), respectively. These sequences have more than 97% nucleotide sequence identity, but less than 86% identity with other reported begomovirus sequences. In line with the taxonomic convention for begomoviruses, Hn2 and Hn2-19 are therefore considered to represent isolates of a distinct begomovirus species, for which the name Ageratum yellow vein China virus (AYVCNV) is proposed. Sequence alignment shows AYVCNV has arisen by recombination among viruses related to Ageratum yellow vein virus, Papaya leaf curl China virus, and an unidentified begomovirus. Southern blot analyses revealed that all plants sampled contained molecules resembling DNAbeta. DNAbeta molecules from three samples were 1,323 or 1,324 nt long and had >98% sequence identity but <81% identity with previously reported DNAbeta sequences. Infectious clones of Hn2 and its associated DNAbeta were constructed and agroinoculated to plants. Hn2 alone caused sporadic asymptomatic systemic infection of Nicotiana benthamiana, N. glutinosa, Lycopersicon esculentum, Petunia hybrida, and A. conyzoides but its accumulation was much enhanced in plants co-inoculated with DNAbeta. The co-inoculated N. benthamiana, N. glutinosa, P. hybrida, and L. esculentum plants developed leaf curling or leaf crinkling symptom; those in A. conyzoides were typical of ageratum yellow vein disease. When the DNAbeta molecules associated with four other Chinese begomoviruses were coinoculated with Hn2 to N. benthamiana and N. glutinosa, the DNAbeta molecules were replicated, and the plants developed systemic symptoms of types that were specific for each DNAbeta. This illustrates that there is less specific interaction between monopartite begomovirus and DNAbeta than between the DNA-A and DNA-B of begomoviruses with bipartite genomes.     

Synergistic Disease in Pepper Caused by the Mixed Infection of Cucumber mosaic virus and Pepper mottle virus

ABSTRACT The occurrence of more than one virus species in a single plant is not uncommon in cultivated and native plant species. A mixed virus infection may lead to greater disease severity than individual viral components and this is sometimes referred to as a synergistic disease. Although, in some cases, synergism has been demonstrated for various plant growth parameters such as plant height, weight, and yield, proof of synergy typically has not been demonstrated for symptom severity when the mixed virus infection was not lethal. We demonstrated synergy in bell pepper plants co-infected with Cucumber mosaic virus (CMV) and Pepper mottle virus (PepMoV) relative to each virus alone for stem height (two of three trials) and aboveground fresh weight (one of three trials) using factorial analysis and Abbott's equation for synergy. This approach allowed affirmation of the type of response (i.e., synergistic rather than antagonistic) and statistical proof of synergy. A detailed evaluation of symptom severity for each viral treatment revealed three phases associated with host plant developmental stages. A numerical symptom severity rating scale was developed and used in each of two equations to demonstrate statistical proof for synergy based on symptom severity for co-infected plants. Virus accumulation in noninoculated leaves was determined by enzyme-linked immunosorbent assay. In singly infected plants, CMV titers declined in mildly symptomatic leaves representing later stages of plant development, but titers increased in similar leaves of co-infected plants. In contrast, PepMoV titers did not differ in singly or co-infected plants.     

Tobacco curly shoot virus DNAbeta Is Not Necessary for Infection but Intensifies Symptoms in a Host-Dependent Manner

ABSTRACT We demonstrated that only 11 isolates were associated with DNAbeta among 39 Tobacco curly shoot virus (TbCSV)-infected, field-collected samples. An infectious clone of TbCSV-[Y35], an isolate associated with DNAbeta, induced severe upward leaf curling in Nicotiana benthamiana. In the presence of its cognate DNAbeta (TbCSV-[Y35] DNAbeta), the symptom changed to a downward leaf curl. Furthermore, TbCSV-[Y35] alone was able to induce severe symptoms in tobacco and tomato plants, although co-infection with DNAbeta intensified symptom severity in tobacco plants. In contrast to other begomovirus-DNAbeta complexes, the satellite had no effect on the accumulation of TbCSV-[Y35] DNA in systemically infected host plants. The betaC1 mutant caused symptoms comparable to those induced by TbCSV-[Y35] in the absence of DNAbeta. TbCSV-[Y35] can be transmitted between plants by a whitefly vector, regardless of the presence or absence of DNAbeta. For a TbCSV isolate not associated with DNAbeta (TbCSV-[Y1]), systemic infection of N. benthamiana induced symptoms resembling those of TbCSV-[Y35]. Co-infection of TbCSV-[Y1] with TbCSV-[Y35] DNAbeta induced symptoms similar to those following infection by TbCSV-[Y35] and its DNAbeta. This indicates that TbCSV DNAbeta is not necessary for infection but intensifies symptoms in a host-dependent manner. Thus, TbCSV may represent an evolutionary intermediate between the DNAbeta requiring begomoviruses and the truly monopartite begomoviruses. The relevance of these results to our present understanding of the evolution of begomovirus-satellite disease complexes is discussed.     

Further studies on resistance-breaking strains of potato virus X

A stock culture of isolate CP of potato virus X (PVX) maintained by serial subculture in plants of Nicotiana glutinosa was found to contain PVX strain group four in addition to the original strain group two. The group four strain was separated from the mixture by sap-inoculation to potato cultivars Maris Piper and Pentland Dell, both of which carry PVX hypersensitivity genes Nx and Nb , by graft-inoculation to Maris Piper and by sap-inoculation to cultivar Pentland Ivory which carries Nb but not Nx. Strain group four seemed to be a minor component of the strain mixture in N. glutinosa as few potato plants became infected with it when insusceptible plants were sap-inoculated or when sap inoculum was diluted 500 times. The group four strain passed readily through tubers of infected potato plants and was stable on serial sub-culture in N. glutinosa. When stock cultures of PVX group two isolates B and EX kept in N. glutinosa were tested on cultivar Pentland Dell, they also proved to be mixtures of group two and group four, indicating that spontaneous appearance of group four in cultures of group two strains may occur readily.
When group four strains derived from isolate CP and from PVX common strain isolate DX were graft-inoculated to many plants of cultivar Cara, which carries PVX immunity gene Rx , there was no evidence of selection of a strain like HB which overcomes this gene. In mixed infection with isolate DX, HB was still present after passage through two generations of progeny tubers of cultivar Pentland Crown which lacks any resistance genes, indicating that HB is a fully competitive strain.     

Studies on resistance to beet western yellows virus in lettuce (Lactuca sativa) and the occurrence of field sources of the virus

An effective method based on glasshouse and field procedures was developed for screening commercial cultivars and other lettuce types for resistance to beet western yellows virus (BWYV). Field experiments in 1985, 1986 and 1987 showed that lettuce cultivars varied in their reaction to BWYV, but no high levels of resistance were identified in the main commercial types. Crisp types generally showed milder symptoms than butterhead or cos types, but individual butterhead cultivars were identified with resistance equal to the best crisp types. The highest levels of resistance were identified in Batavian type cultivars and extreme resistance or possible immunity was found in Lactuca perennis and L. muralis . BWYV caused yield reductions in some cultivars as high as 63% and reduced maturation by up to 38%, in others. There was no correlation between chlorotic leaf symptom severity and yield reduction.
BWYV was isolated from a range of weed and non-lettuce host species growing near affected lettuce crops. Isolates of BWYV obtained from infected lettuce and brassicas appeared to be similar. They infected sugar beet with difficulty but caused no symptoms, and could only be detected by ELISA serology.