Tentative description of Hippeastrum latent virus in Hippeastrum hybridum plants and differentiation from Hippeastrum mosaic virus

In mosaic-diseased plants ofHippeastrum hybridum two viruses were found. One virus with a normal length of 706 nm caused local lesions onHyoscyamus niger test plants and mosaic symptoms in the leaves ofH. hybridum. This virus was identified with theHippeastrum mosaic virus (HMV) (*/*∶*/*∶E/E∶S/*) and had a dilution end point between 10^?3 and 10^?4, a thermal inactivation point between 55–60°C and a longevity at room temperature of 28–32 hours. The second virus had a normal length between 584 and 611 nm depending on the method used. It caused local lesions onGomphrena globosa andChenopodium quinoa leaves, and after inoculation ofH. hybridum was found to be present without showing symptoms. It was readily purified from inoculated leaf tissue ofC. quinoa andNicotiana clevelandii by differential centrifugation and ofH. hybridum by density-gradient centrifugation. Purified virus had an absorption minimum at 242 nm, a maximum at 262 nm and a 260/280 absorption ratio of 1.19. The dilution end point was between 10^?3 and 10^?4, the thermal inactivation point between 70 and 80°C and the longevity in vitro at room temperature 28–32 hours. Although no direct comparisons have been made with other members of the potexvirus group, the virus seems to be a new one now namedHippeastrum latent virus. Both viruses were not seed-borne.     

Investigation ofHippeastrum mosaic virus inHippeastrum hybridum

An attempt was made to identify a mosaic disease inHippeastrum hybridum. Infectious virus material could be demonstrated in roots, leaves, stem, perianth, stamen and pistil. Inclusion bodies were found in the epidermis of leaves, stem, spatha leaves and in the perianth. Virus concentration in a young stage was high but decreased by aging of the perianth. Efforts to transmit the virus by aphids failed. However, the virus was transmitted by seed in a few cases. Plants of 30 species reacted negatively upon inoculation with the virus.Hippeastrum hybridum, Gomphrena globosa, Chlorophytum spec. andLycopersicum esculentum could be infected experimentally. On account of the host range and presence of inclusion bodies the mosaic symptoms inHippeastrum are not caused by tomato spotted wilt virus or Cucumber mosaic virus. Results suggest that the virus under investigation is theHippeastrum mosaic virus. Dr.M. K. Corbett, Wageningen, succeeded in purifying the virus by density gradient centrifugation. Whe preparation contained flexuous rod particles. Plants ofDatura stramonium, Nicotiana glutinosa andN. tabacum Samsun could be infected. Within two weeks after inoculation with purified virus solution these plants showed systemic symptoms.

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Samenvatting In een kwekerij te Hoorn werden planten vanHippeastrum hybridum aangetroffen, die mozaïekverschijnselen vertoonden. De oorzaak hiervan werd nagegaan. Infectieus virusmateriaal kon worden aangetoond in wortels, bladeren, bloemstengel, bloemdekbladen, meeldraden en stijl. Celinsluitsels kwamen voor in de epidermis van bladeren en stengel en in de bloemschede. De aanwezigheid van insluitsels in het bloemdek was afhankelijk van de ouderdom van de bloem. Er schijnt een omgekeerd evenredige relatie te bestaan tussen de virusconcentratie en het aantal insluitsels in bloemen.Pogingen om het virus over te brengen door bladluizen mislukten. In enkele gevallen had zaadoverdracht plaats.Dertig plantesoorten reageerden negatief op een inoculatie met virushoudend sap.Hippeastrum hybridum, Gomphrena globosa, Chlorophytum spec. enLycopersicum esculentum konden wel worden geïnfecteerd. Gezien de waardplantenreeks en het voorkomen van celinsluitsels kunnen de mozaïeksymptomen inHippeastrum niet toegeschreven worden aan Tomato spotted wilt-virus of het komkommer-mozaïek-virus, maar is het waarschijnlijk dat zij worden veroorzaakt door hetHippeastrum-mozaïek-virus, reeds beschreven doorBrierley (1948),Johnson (1951) enProcenko & Procenko (1964).Dr.M. K. Corbett, Wageningen, slaagde erin het virus te zuiveren door middel van density gradient-centrifugering.Datura stramonium, N. glutinosa enN. tabacum Samsun werden systemisch ziek binnen twee weken na inoculatie met de gezuiverde virus-oplossing.

     

Hippeastrum mosaic virus and another filamentous virus in Eucharis grandiflora

Two viruses were found in mosaic-diseased plants ofEucharis grandiflora in a glasshouse of the laboratory. One virus with a normal particle length of 733 nm caused local lesions onHyoscyamus niger and mosaic symptoms in leaves of healthy-lookingEucharis andHippeastrum plants. On the basis of its host range, physical properties and serology it was identified asHippeastrum mosaic virus, a member of the potyvirus group. This was confirmed by the presence of spherical nuclear inclusions and pinwheels in different tissues of diseasedEucharis plants. The second virus with a normal particle length of 598 nm was present in both healthy-looking and mosaic-diseasedEucharis plants, and it inconsistently induced local lesions onGomphrena globosa. According to its morphology and its reaction onGomphrena, it might be identical or related toHippeastrum latent virus. Crystal-like inclusions were observed in the cytoplasm of cells of both healthy-looking and mosaic-showingEucharis leaves. As no virus-free seedlings ofEucharis were available, the virus nature of these inclusions could not be established.     

Tobacco mosaic virus in Pythium spec

Cultures of the soil inhabiting fungusPythium spec. were inoculated in vitro with tobacco mosaic virus. Virus could be demonstrated in the mycelia from 4 days on after inoculation. In 15 days old cultures the virus concentration in the mycelium was higher than in the liquid culture medium. It is not yet clear whether the virus only accumulates, or also multiplies in the mycelium. After growth on solid medium infected mycelia still contained virus indicating that the virus is able to persist and possibly also to multiply in the hyphae.Samenvatting Cultures van de bodemschimmelPythium spec. werden in vitro geïnoculeerd met tabaksmozaïekvirus. In enkele gevallen kon al 4 dagen na inoculatie virus in mycelium worden aangetoond. In 15 dagen oude cultures bevatte het mycelium meer virus dan de cultuurvloeistof. Het is nog niet bekend of het virus zich slechts ophoopt of zich ook vermeerdert in het mycelium. Na groei op een agarmedium gedurende 7 dagen bevatte het mycelium nog virus, wat er op zou wijzen dat het virus in staat is zich te handhaven in de hyfen en zich daarin wellicht ook te vermeerderen.     

Hyoscyamus niger,a useful local lesion host for a mosaic virus in Hippeastrum

Samenvatting Bladeren vanHyoscyamus niger planten vertoonden chlorotische lokale lesies 7 dagen na inoculatie met sap van mozaïekziekeHippeastrum-planten. Met sap vanHippeastrum-planten zonder mozaïeksymptomen werd geen reactie verkregen. In dooppreparaten gemaakt van chlorotische lesies inHyoscyamus-bladeren en van bladeren vanHippeastrum planten met mozaïeksymptomen werden draadvormige virusdeeltjes van 648–772 nm aangetroffen. Het virus kon met sap geperst uit geïnoculeerdeHyoscyamus-planten weer op gezondeHippeastrum-planten worden overgebracht.     

烟草品种对烟草花叶病毒病和黄瓜花叶病毒病的抗性鉴定

由烟草普通花叶病毒(TMV)和烟草黄瓜花叶病毒(CMV)引致的烟草病毒病是世界烟草主产区普遍发生且危害严重的侵染性病害,每年给烟叶生产造成了重大的经济损失。本文采用温室苗期接种鉴定的方法,对16份烟草种质进行了TMV和CMV的抗病性鉴定。结果表明:不同的烟草品种对TMV和CMV的抗病性存在较大差异。在供试种质中,对TMV表现免疫的有‘牛耳烟’、‘8301’、‘台烟7号’、‘三生-NN’共4份材料;表现抗病的有‘吉烟5号’、‘双抗70’、‘大护脖香’、‘秦烟95’共4份材料;表现中抗的有‘铁把子’、‘中烟15’、‘秦烟98’、‘中烟98’共4份材料;表现中感的有‘NC89’、‘翠碧1号’、‘云烟97’共3份材料;表现感病的只有‘秦烟97’。对CMV表现中抗的材料有1份,是‘铁把子’;表现中感的有‘秦烟95’、‘三生-NN’、‘8301’、‘牛耳烟’、‘翠碧1号’共5份材料;表现感病的有‘秦烟98’、‘云烟97’、‘中烟98’、‘NC89’、‘大护脖香’、‘双抗70’、‘秦烟97’、‘中烟15’、‘台烟7号’、‘吉烟5号’共10份材料。研究发现,‘铁把子’是兼抗这两种病毒病的材料。本研究明确了我国16个烟草品种资源的抗病性水平,为抗耐病品种的利用与品种合理布局提供科学依据,同时为烟草抗病毒病育种的亲本选择提供抗源信息。     

Evaluation of Seed Transmission of Turnip yellow mosaic virus and Tobacco mosaic virus in Arabidopsis thaliana

ABSTRACT The mechanism of virus transmission through seed was studied in Arabidopsis thaliana infected with Turnip yellow mosaic virus (TYMV) and Tobacco mosaic virus (TMV). Serological and biological tests were conducted to identify the route by which the viruses reach the seed and subsequently are located in the seed. Both TYMV and TMV were detected in seed from infected plants, however only TYMV was seed-transmitted. This is the first report of transmission of TYMV in seed of A. thaliana. Estimating virus seed transmission by grow-out tests was more accurate than enzyme-linked immunosorbent assay due to the higher frequency of antigen in the seed coat than in the embryo. Virus in the seed coat did not lead to seedling infection. Thus, embryo invasion is necessary for seed transmission of TYMV in A. thaliana. Crosses between healthy and virus-infected plants indicated that TYMV from either the female or the male parent could invade the seed. Conversely, invasion from maternal tissue was the only route for TMV to invade the seed. Pollination of flowers on healthy A. thaliana with pollen from TYMV-infected plants did not result in systemic infection of healthy plants, despite TYMV being carried by pollen to the seed.     

用甘蔗花叶病毒生物防治烟草蚀纹病毒病

 烟草是我国重要的经济作物,2004年种植面积达100万hm²,占全国耕地面积1%左右。     

嘧肽霉素抗烟草花叶病毒作用机理初步研究

 以烟草花叶病毒(TMV)/烟草为测试体系,用³H-尿苷和³H-亮氨酸2种同位素示踪的方法研究了4%嘧肽霉素水剂抗TMV的作用机理。通过³H-尿苷示踪实验发现:无论是叶盘法还是整株法测定,嘧肽霉素均能抑制病毒对³H-尿苷的利用,抑制率分别为72.43%和68.95%;而对寄主植物RNA合成,2种方法的抑制率分别为8.09%和11.02%。通过³H-亮氨酸示踪结果可以看出:嘧肽霉素较强地抑制了病毒对³H-亮氨酸的吸收,但能促进寄主对³H-亮氨酸的吸收。³H-尿苷和³H-亮氨酸分别是RNA和蛋白质合成的前体物,因此,嘧肽霉素通过抑制病毒RNA复制和蛋白质的合成来扰乱病毒的增殖,从而减弱病害的发生。     

地高辛标记的cDNA探针检测烟草花叶病毒、黄瓜花叶病毒及马铃薯Y病毒

 根据已发表的烟草花叶病毒(Tobacco mosaic virus,TMV)、黄瓜花叶病毒(Cucumber mosaic virus,CMV)和马铃薯Y病毒(Potato virus Y,PVY)的外壳蛋白基因序列,设计特异引物,分别以提取的TMV、CMV和PVY侵染的病叶总RNA为模板,反转录PCR进行体外扩增,分别得到长度为0.44、0.77、0.80 kb的目的片段,并克隆到pGEM-T easy质粒载体上,以构建的重组质粒为模板,用PCR方法合成了相应的地高辛标记的双链DNA探针。以合成的探针通过斑点杂交技术检测烟草病叶总RNA和烟草病叶汁液。TMV、CMV和PVY的3种地高辛探针检测各自感染的烟草病叶总RNA的稀释低限分别为1:1000、1:10000、1:320,检测各自侵染烟草病汁液的最大稀释倍数分别为1:100、1:100、1:10,而每种探针与健康烟草和其它2种病毒的反应均为阴性。     

辣椒上烟草轻型绿花叶病毒的鉴定

 辣椒源自南美洲,属于茄科辣椒属（Capsicum L.）,为重要经济作物。病毒病是影响辣椒生产的主要病害,侵染辣椒的病毒有40余种［1］,烟草轻型绿花叶病毒（Tobacco mild green mosaic virus, TMGMV）是近年来在辣椒上发生和危害报道较多的病毒之一。TMGMV是McKinney［2］于1935年在烟草属植物（Nicotiana gluanca）上首次发现的,为烟草花叶病毒属（Tobamovirus）成员。除辣椒外,番茄［3］、凤仙花、蓝眼菊和矮牵牛也是TMGMV的自然寄主。     

烟草花叶病毒丁香分离物的分离与鉴定

 从表现花叶症状的丁香病株上获得一病毒分离物,其在电镜下为约300 nm×18nm的杆状粒子;电泳分析表明感病组织中ds RNA大约为6.4kbp,而其外壳蛋白分子量约为17.6k Da。以上实验结果初步将该病毒分离物鉴定为烟草花叶病毒属(Tobamovirus)。根据该属病毒复制酶基因序列设计通用引物,进行RT-PCR检测,扩增出约1000 bp的预期特异片段(Gen Bank AY566703)。将PCR产物克隆后测序,序列分析表明,与从蚕豆中分离的TMV-B株系序列(Gen Bank AJ011933.1)同源性为99.90%。根据烟草花叶病毒(Tobacco mosaic virus,TMV)的RNA CP基因序列设计引物,进行RT-PCR,扩增出约800 bp的预期特异片段(Gen Bank AY56672),序列分析表明,与TMV-B株系序列(Gen Bank AJ011933.1)同源性达99%,上述实验结果表明,该病毒分离物为TMV。由于该分离物与TMV-B在指示植物上的症状存在明显差异,所以,作者把该分离物暂命名为TMV-S。     

基于全基因组编码区序列的烟草花叶病毒分子进化分析

为揭示烟草花叶病毒分子进化特征,从GenBank中下载已报道的烟草花叶病毒全基因组编码区序列,进行重组、系统发育、遗传变异、种群结构和基因漂流等分析。结果表明:突变和负选择作用是驱动烟草花叶病毒进化的主要作用力,种群结构稳定,处于扩张趋势中,基因变异程度低,重组在烟草花叶病毒分子进化中作用不明显。烟草花叶病毒不同分离物形成3个有一定地理相关性的种群ChinaⅠ、ChinaⅡ和Europe。欧洲分离物核苷酸序列多样性比中国的差异大,Europe和ChinaⅠ种群可能受到遗传漂变影响,Europe与ChinaⅡ、ChinaⅠ与ChinaⅡ之间存在发生基因交流的渠道。     

寡聚半乳糖醛酸诱导烟草抗烟草花叶病毒研究初探

用寡聚半乳糖醛酸诱导烟草抗烟草花叶病毒,田间和温室试验结果表明,寡聚半乳糖醛酸50μg/mL的诱抗效果最好,对烟草花叶病毒引起的枯斑的抑制率为64.4%。寡聚半乳糖醛酸可以诱导烟草植株抗性酶超氧化物歧化酶和过氧化氢酶活性的升高。     

烟草品种对烟草花叶病毒和黄瓜花叶病毒的抗性鉴定

 2010-2011年采用大田人工接种鉴定的方法,对生产中大面积推广使用的24份烟草品种进行了烟草花叶病毒（TMV）和黄瓜花叶病毒（CMV）的抗性鉴定。结果表明,供试品种对TMV和CMV的抗病性存在较大差异,对TMV表现抗病的有Coker86、吉烟5号、Coker176、CV87、辽烟8号、CV91、中烟90等7份材料;表现中抗的有秦烟98、双抗70、C151等3份材料;表现中感的有秦烟96、G80、金星6007、龙江981、K326、秦烟201、NC89等7份材料;表现感病的有G28、云烟97、净叶黄、红花大金元、RG11、云烟85、云烟87等7份材料。对CMV表现抗病的有Coker86、龙江981、C151、秦烟201、云烟87等5份材料;表现中抗的材料是金星6007;表现中感的有CV91、RG11、Coker176、中烟90、K326、红花大金元、净叶黄、G80、G28等9份材料;表现感病的有秦烟98、云烟85、秦烟96、NC89、双抗70、云烟97、CV87、辽烟8号、吉烟5号等9份材料。其中兼抗TMV和CMV两种病毒病的材料有2份,分别是Coker86和C151。同时研究还发现,抗病性不同的烟草品种在受到病毒危害以后,对烟叶的产量和品质的影响也不同。明确了中国24个烟草品种的抗病性水平,为抗病品种的利用与品种合理布局提供科学依据,同时为烟草抗病毒病育种的亲本选择提供抗源信息。     

壳寡糖诱导烟草抗烟草花叶病毒的超微结构研究

 电镜检查表明,接种烟草花叶病毒(TMV)表现系统侵染的烟草植株叶肉细胞和筛管内有大量病毒粒子和病毒结晶体,细胞质中出现髓鞘样结构、多泡体和次级囊泡。叶片薄壁细胞内叶绿体、线粒体等细胞器变形解体,膜结构增生。经壳寡糖(50 μg/mL)诱导处理的烟草,系统症状减轻,叶肉细胞和筛管中病毒粒子显著减少,偶见病毒结晶体;叶肉细胞中细胞器基本完好,但叶绿体内出现较大的淀粉粒;液泡内和细胞间隙出现大量电子致密物质。在未接毒和未经壳寡糖处理的植株中未见此种电子致密物,其产生可能与诱导抗病性表达有关。     

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。     

大豆凝集素基因lec-s转化烟草>增强对烟草花叶病毒的抗性

 将编码大豆凝集素的lec-s基因插入植物表达载体pBI121中,构建植物重组表达质粒pBI121:: lec-s。由根癌土壤杆菌EHA105介导的叶盘法转化烟草,获得了转基因烟草株系。PCR和RT-PCR检测证明lec-s基因已转入烟草植株中。接种烟草花叶病毒（Tobacco mosaic virus,TMV）进行抗病性试验结果表明,转基因烟草叶片上的病斑数显著减少,说明转基因烟草表现出对TMV的抗性。定量RT-PCR检测发现,接种TMV后,抗病防卫基因（PR-1a、GST1、Pal和hsr515）在转基因烟草叶片中显著上调表达。这些结果表明,大豆凝集素基因lec-s转化烟草可对TMV产生抗性,其作用机制可能在于lec-s基因参与了植物的防卫信号通路,诱导了抗病防卫基因在转基因植株体内的表达,增强了植株对TMV的系统抗性。     

Economic impact of Turnip mosaic virus, Cauliflower mosaic virus and Beet mosaic virus in three Kenyan vegetables

Screenhouse experiments conducted in Kenya showed that inoculation of cabbage seedlings with Turnip mosaic virus (TuMV), either alone, or in combination with Cauliflower mosaic virus (CaMV), reduced the number and weight of marketable harvested heads. When viruses were inoculated simultaneously, 25% of cabbage heads were non-marketable, representing 20-fold loss compared with control. By contrast, inoculation with CaMV alone had insignificant effects on cabbage yield. This suggests that TuMV is the more detrimental of these pathogens, and its management should be a priority. Early exposure to TuMV produced cabbages that were 50% lighter than non-infected plants, but later infection was less damaging suggesting that controlling virus infection at the seedling stage is more important. TuMV was far less damaging to kale than it was to cabbage; although high proportions of TuMV-inoculated kale plants showed symptoms (>90%), the marketability and quality of leaves were not significantly reduced, and no clear relationship existed between timing of infection and subsequent crop losses. Early inoculation of Swiss chard with Beet mosaic virus (BtMV) significantly impaired leaf quality (∼50% reduction in marketable leaf production), but the impact of disease was greatest in plants that had been inoculated at maturity, where average leaf losses were two and a half times those recorded in virus-free plants. Disease-management of BtMV in Swiss chard is important, therefore, not only at the seedling stage, but particularly when plants are transplanted from nursery to field.