Detection and molecular characterization of viruses associated with tomato yellow leaf curl disease in cucurbit crops in Jordan

In this study, Tomato yellow leaf curl Sardinia virus (TYLCSV) and the strains Israel and Mild of Tomato yellow leaf curl virus (TYLCV-IL, TYLCV-Mld) were detected for the first time in four cucurbit crops in Jordan by nested polymerase chain reaction (nPCR). These viruses cause the tomato yellow leaf curl disease (TYLCD) in tomato. Cucumber, squash, melon and watermelon plants inoculated with TYLCV-IL[JO:Cuc], TYLCV-Mld, TYLCSV-IT[IT:Sar:88] and the Jordanian isolate of TYLCV (TYLCV-JV) did not show disease symptoms. However, virus-specific fragments were detected in uppermost leaves of symptomless plants by nPCR. A whitefly transmission test showed that Bemisia tabaci could transmit TYLCV-Mld from cucumber into tomato and jimsonweed plants. However, all infected tomato plants remained symptomless. In addition, results of semi-quantitative PCR (sqPCR) analysis showed that the relative amount of TYLCV-Mld DNA acquired by B. tabaci from cucumber plants was less than that acquired from tomato plants.     

Identification of whitefly transmitted tomato yellow leaf curl geminivirus from Iran and a survey of its distribution with molecular probes

A virus causing a disease of tomato, prevalent in the southern provinces of Iran, with symptoms of leaf-curling, stunting, reduction of leaf size, leaf corrugation, shortening of internodes and severe reduction in fruit yield, was shown to be transmissible to healthy tomato plants by grafting and by whiteflies ( Bemisia tabaci ), but not by sap inoculation. Geminivirus DNA was detected in extracts of diseased tomato plants by dot-blot hybridization assays using as probes full-length cloned DNA of Australian, Italian (Sardinian) or Jordanian strains of tomato yellow leaf curl virus (TYLCV). Geminivirus coat protein was detected in whitefly inoculated plants by dot immunobinding assay using polyclonal antibody raised against Jordanian TYLCV. A limited survey using the dot-blot hybridization assay for virus detection indicated the presence of the virus in tomato-growing provinces of southern but not northern Iran. Whitefly transmission experiments to tomato under controlled greenhouse conditions showed that some isolates of TYLCV-like geminiviruses from different parts of Iran differ in symptomatology.     

Tomato leaf curl geminivirus in Australia: occurrence, detection, sequence diversity and host range

The occurrence of whitefly transmitted geminiviruses in Australia was studied using a mixed DNA probe capable of detecting a range of distinct geminiviruses. The only geminivirus species detected was Tomato leaf curl virus (TLCV), which is spread across a vast geographical region of far-northern coastal Australia, an area inhabited by the Australasian-Oceania biotype of Bemisia tabaci . The newly introduced silverleaf whitefly, B. tabaci biotype B, forms high population densities in the eastern coastal region of Queensland and is currently located approximately 150 km from the nearest known TLCV-infected area. The viral host range appeared to be narrow and of 58 species of crop plants and weeds inoculated using the B biotype, only 11 became infected with the virus, including five that did not show foliar symptoms. A DNA fragment of 694 nt, including the complete C4 open reading frame (ORF), the overlapping N-terminal part of the C1 ORF and the viral iterons involved in replication, was amplified from 11 TLCV field isolates and sequenced. Sequence analysis revealed an overall sequence variation of up to 14% in this region, as well as the presence of distinct viral iterons.     

Tracing the geminivirus-whitefly transmission pathway by polymerase chain reaction in whitefly extracts, saliva, hemolymph, and honeydew

ABSTRACT A membrane feeding system and polymerase chain reaction (PCR) were used to track squash leaf curl virus (SLCV) DNA in whole whitefly body extracts and in saliva, honeydew, and hemolymph of its whitefly vector, Bemisia tabaci, and a whitefly nonvector, Trialeurodes vaporariorum. SLCV ingestion was monitored by PCR in whiteflies that were given acquisition access periods (AAPs) ranging from 0.5 to 96 h on virus-infected plants. SLCV detection by PCR in whole body extracts was considered reflective of virus ingestion. As whiteflies were given longer AAPs, the number of whiteflies that ingested SLCV increased. SLCV DNA was detected in honeydew of vector and nonvector whiteflies, indicating that virions, viral DNA, or both passed unimpeded through the digestive system. SLCV DNA was detected in saliva and hemolymph of B. tabaci, but not in these fractions from nonvector whiteflies, despite virus ingestion by both. Although vector and nonvector whiteflies both ingested SLCV, only in the vector, B. tabaci, did virus cross the gut barrier, enter the hemolymph, or pass into the salivary system. These results suggest that digestive epithelia of nonvector whiteflies did not permit SLCV passage from the gut to hemocoel, whereas virus effectively crossed the analogous gut barrier in vector whiteflies.     

Solanum nigrum: an indigenous weed reservoir for a tomato yellow leaf curl geminivirus in southern Spain

A tomato yellow leaf curl geminivirus (TYLCV-AL), was first identified in tomato plants in Almeria, southern Spain in 1992. This virus is transmitted by the tobacco whitefly, Bemisia tabaci (Gennadius), and is presently infecting tomato crops throughout the south eastern region of Spain. Solanum nigrum, collected from a field in south east Spain and exhibiting leaf curl symptoms, was squash blotted onto nylon membrane and gave a positive signal when hybridised to a TYLCV-Is DNA probe. Laboratory tests showed B. tabaci to transmit TYLCV-AL from infected tomato plants to S. nigrum seedlings. The virus could then be acquired by B. tabaci and transmitted back from infected S. nigrum plants to tomato, inducing typical TYLCV symptoms. These results indicate the importance of S. nigrum as a weed host/reservoir for a TYLCV and its possible role in the spread of this virus within Europe.     

B型烟粉虱携带传播烟草曲茎病毒的能力

B型烟粉虱近20年来入侵许多国家和地区,已成为重要的世界性作物害虫,其主要危害方式之一是传播双生病毒,造成作物病毒病大发生.应用PCR技术,研究了入侵我国的B型烟粉虱个体获取、存留及传播烟草曲茎病毒（Tobacco curly shoot virus,TbCSV）的能力.B型烟粉虱在感染TbCSV的普通烟毒株上饲毒30 min时,就可在40%的个体内检测到TbCSV DNA,个体带毒率随饲毒时间延长而增加,饲毒12 h后,带毒率达100%;TbCSV DNA在B型烟粉虱体内可存留10天左右,但不能终身存留;传毒处理植株的发病症状及PCR检测结果表明,B型烟粉虱是TbCSV的传播媒介.     

Yellow leaf curl and other virus diseases of tomato in Cyprus

Outdoor and greenhouse tomato plantings in the main production centres of Cyprus were surveyed for virus diseases during 1978–1982. A disease characterized by prominent leaf yellowing and curling was the most prevalent and damaging. The causal agent was identified as tomato yellow leaf curl virus (TYLCV) because it infected Datura stramonium but not the tobacco cultivars Havana 423 or Virginia, was graft-transmissible but not mechanically or seed-transmissible, and was transmitted persistently by the tobacco whitefly, Bemisia tabaci. Minimum acquisition and inoculation feeding periods were 20–30 and 10–20 min. respectively; the latent period was 21–24 h. The vector was infective for about 12 days, but there was no transovarial transmission to progeny.
Four other, mechanically transmissible, viruses were also isolated from tomato and identified on the basis of symptomatology, indicator host reactions and serology; tobacco mosaic virus (TMV), potato virus Y (PVY). potato virus X (PVX), and cucumber mosaic virus (CMV). These viruses, especially TMV, were commonly associated with mosaic symptoms and occasionally with other leaf or fruit disorders; coinfection with TYLCV occurred frequently.     

应用PCR方法检测烟粉虱传双生病毒

番茄黄化曲叶病毒属于双生病毒,根据其外壳蛋白基因保守序列设计简并引物,对杭州和海宁地区番茄上疑似番茄黄化曲叶病毒进行PCR检测,获得产物克隆到pMD18-T载体中,经测序和序列分析,表明杭州和海宁地区发生的病毒是番茄黄化曲叶病毒,并且两地区病毒核苷酸序列同源性达97.5%。     

Dual begomovirus infections and high Bemisia tabaci populations: two factors driving the spread of a cassava mosaic disease pandemic

A cassava mosaic disease (CMD) pandemic currently affects much of East and Central Africa. To understand the factors driving the pandemic's continued spread, complementary data sets were collected from cassava plots, planted with healthy cuttings, at eight sites along a north–south transect in southern Uganda, through the pandemic's leading edge. Data were collected on virus incidence, symptom severity, populations of the whitefly vector, Bemisia tabaci , their infectivity and ability to transmit different viruses. In 1996, 6 months after planting, CMD incidences were highest at sites 1 and 2, then decreased progressively until site 6, and remained low at sites 7 and 8. The largest B. tabaci populations also occurred at northernmost sites, 1–3. In 1997, CMD incidence increased significantly at sites 5–8 and this was associated with significant increases in the B. tabaci populations. The pandemic's spread was also associated with significant increases in the percentage of dual infections of East African cassava mosaic virus -Uganda and African cassava mosaic virus , which caused the severest symptoms and the greatest reduction in leaf area. Whitefly adults collected from within the pandemic area were infective, whereas those collected ahead of the pandemic were not. The transmission rate of African cassava mosaic virus from plants with dual infections was significantly less than that of East African cassava mosaic virus -Uganda, which may explain the latter's predominance within the pandemic. These results show that the arrival of East African cassava mosaic virus -Uganda into areas affected previously only by African cassava mosaic virus , has resulted in novel virus/vector/host–plant interactions that drive the pandemic's continued spread.     

Comparative whitefly transmission of Tomato chlorosis virus and Tomato infectious chlorosis virus from single or mixed infections

Tomato chlorosis virus (ToCV) and Tomato infectious chlorosis virus (TICV) are two criniviruses that are emerging worldwide, and induce similar yellowing diseases in tomato crops. While TICV is transmitted only by Trialeurodes vaporariorum , ToCV is transmitted by three whitefly species in two genera Trialeurodes vaporariorum , T. abutilonea and Bemisia tabaci . The efficiency of transmission by T. vaporariorum from plants infected by one virus or by both was compared, and the probability of virus transmission by a single whitefly was derived from group testing experiments. The estimated transmission probabilities ranged from 0·01 to 0·13, and were not significantly different between ToCV and TICV, or between single and mixed infections. Experiments using B. tabaci as a vector and source plants infected by TICV and ToCV did not reveal any functional trans-complementation for transmission of TICV by ToCV, suggesting that if this phenomenon occurs in nature, it is at a very low frequency. Possible reasons why TICV did not establish in southern France while ToCV is now endemic are discussed.     

Emergence of a New Cucurbit-Infecting Begomovirus Species Capable of Forming Viable Reassortants with Related Viruses in the Squash leaf curl virus Cluster

ABSTRACT Cucurbit leaf curl virus (CuLCV), a whitefly-transmitted geminivirus previously partially characterized from the southwestern United States and northern Mexico, was identified as a distinct bipartite begomovirus species. This virus has near sequence identity with the previously partially characterized Cucurbit leaf crumple virus from California. Experimental and natural host range studies indicated that CuLCV has a relatively broad host range within the family Cucurbitaceae and also infects bean and tobacco. The genome of an Arizona isolate, designated CuLCV-AZ, was cloned and completely sequenced. Cloned CuLCV-AZ DNA A and B components were infectious by biolistic inoculation to pumpkin and progeny virus was transmissible by the whitefly vector, Bemisia tabaci, thereby completing Koch's postulates. CuLCV-AZ DNA A shared highest nucleotide sequence identity with Squash leaf curl virus-R (SLCV-R), SLCV-E, and Bean calico mosaic virus (BCaMV) at 84, 83, and 80%, respectively. The CuLCV DNA B component shared highest nucleotide sequence identity with BCaMV, SLCV-R, and SLCV-E at 71, 70, and 68%, respectively. The cis-acting begomovirus replication specificity element, GGTGTCCTGGTG, in the CuLCV-AZ origin of replication is identical to that of SLCV-R, SLCV-E, and BCaMV, suggesting that reassortants among components of CuLCV-AZ and these begomoviruses may be possible. Reassortment experiments in pumpkin demonstrated that both reassortants of CuLCV-AZ and SLCV-E A and B components were viable. However, for CuLCV-AZ and SLCV-R, only one reassortant (SLCV-R DNA A/CuLCV-AZ DNA B) was viable on pumpkin, even though the cognate component pairs of both viruses infect pumpkin. These results demonstrate that reassortment among sympatric begomovirus species infecting cucurbits are possible, and that, if generated in nature, could result in begomoviruses bearing distinct biological properties.     

The transmission of geminiviruses by Bemisia tabaci

The cosmopolitan whitefly species, Bemisia tabaci (Gennadius) and Trialeurodes taporariorum (Westwood) have always been regarded as pests to a large range of worldwide crops. Both species are capable of transmitting plant viruses, with T. vaporariorum being the vector of only a few ‘clostero’-like viruses and B. tabaci the vector of viruses in several groups. The largest group of viruses transmitted by B. tabaci are the geminiviruses and B. tabaci is known to transmit around 60 members. Until recently, B. tabaci had been associated with only a limited range of host plants within any one region, although its total potential host range was large. Virus transmission was confined within the plant host range of each regional population of B. tabaci. The emergence of the polyphagous ‘B’ biotype of B. tabaci with its increased host range of more than 600 plant species, has resulted in geminiviruses infecting previously unaffected crops. As the ‘B’ biotype spreads further into Europe, European field and glasshouse crops have been shown to be susceptible to whitefly-transmitted viruses already endemic to other parts of the world. More than 20 colonies of B. tabaci, including both ‘B’ and non-‘B’ biotypes from disparate global locations have been compared for their ability to transmit more than 20 geminiviruses. All but two highly host-specific colonies were capable of transmitting most geminiviruses tested. However, some viruses were transmitted more efficiently than others. The virus coat protein or capsid is essential for vector recognition and transmission. By comparing transmissible viruses at the molecular level to viruses that are no longer whitefly-transmissible, the active epitope on the virus coat protein could be identified for designing future virus control strategies.     

番茄褪绿病毒与番茄黄化曲叶病毒复合侵染对番茄褪绿病毒传播的影响

番茄褪绿病毒Tomato chlorosis virus(ToCV)是一种由烟粉虱Bemisia tabaci传播的正义单链RNA病毒,在田间常与番茄黄化曲叶病毒Tomato yellow leaf curl virus(TYLCV)复合侵染而造成番茄生产上重大的经济损失。为了明确ToCV与TYLCV的复合侵染对烟粉虱传播ToCV所造成的影响,本文采用RT-PCR以及qRT-PCR检测了复合侵染的番茄对烟粉虱获取和传播ToCV的影响。研究表明,烟粉虱取食复合侵染的番茄后对ToCV的传播效率显著提高,仅25头烟粉虱的传毒率即可达到100%,ToCV在烟粉虱以及番茄体内的累积量均显著提高。说明这种复合侵染促进了烟粉虱对ToCV的传播,在田间应当及时防控烟粉虱,警惕病毒与烟粉虱的蔓延。     

海氏桨角蚜小蜂对不同龄期Q隐种烟粉虱和温室白粉虱的取食和寄生选择

许多粉虱寄生蜂雌蜂对粉虱若虫的攻击方式不仅是寄生,而且还能取食。本文以海氏桨角蚜小蜂Eretmocerus hayati作为测试寄生蜂,Q隐种烟粉虱和温室白粉虱作为寄主,在非选择和双向选择条件下调查了寄生蜂对寄主的取食和寄生选择性。研究结果表明,非选择试验条件下,海氏桨角蚜小蜂48 h内取食1～4龄烟粉虱和温室白粉虱若虫的数量分别为10.7、6.4、6.7、5.0头和4.4、3.7、2.8、2.3头,寄生1～4龄烟粉虱若虫的数量分别为22.1、27.5、25.4、16.5头,但不能寄生各龄期温室白粉虱若虫;而在选择试验条件下,寄生蜂取食1～4龄烟粉虱若虫的数量分别为同龄期温室白粉虱若虫数量的1.8、2.6、2.5、1.5倍。说明寄生蜂的取食寄主能力明显受寄主种类和龄期的影响。海氏桨角蚜小蜂更适合用来防治烟粉虱,但不适于防治温室白粉虱。     

内蒙古和贵州地区粉虱发生对番茄褪绿病毒流行的影响

番茄褪绿病毒Tomato chlorosis virus(ToCV)对我国番茄种植业破坏极大,而粉虱类昆虫是其主要传播介体。本研究调查了我国内蒙古自治区赤峰市和贵州省贵阳市的番茄褪绿病毒病发生情况,发现赤峰市的发病率为73.3%,贵阳市的发病率为53.3%。两个地区均存在Q型烟粉虱和温室白粉虱,赤峰市的Q型烟粉虱和温室白粉虱分别占75.9%和24.1%;贵阳市的Q型烟粉虱和温室白粉虱分别占39.8%和60.2%。对两个地区的两种粉虱种群在室内进行传毒特性研究,结果发现,经过48 h获毒后Q型烟粉虱和温室白粉虱的获毒量没有明显差异,但无论1头或10头粉虱,Q型烟粉虱的传毒量都高于温室白粉虱。结果说明,近年来随着Q型烟粉虱发生的日益严重,番茄褪绿病毒通过Q型烟粉虱的传播值得人们重视和进一步研究。     

Molecular identification of a begomovirus causing yellow vein disease on Calendula officinalis in India

A whitefly transmitted begomovirus was detected by PCR using begomovirus-specific primers from naturally infected Calendula officinalis plants showing yellow vein disease symptoms. An approximately 800 bp PCR amplicon was cloned and sequenced to identify the species of the virus isolate. Analysis of nucleotide sequence data resulted in its identification as the complete coat protein gene open reading frame (CP ORF) of 771 bp, which encoded 256 amino acid residues. The coat protein of the virus isolate shared maximum identities of 96–97% with four strains of Tobacco curly shoot virus (ToCSV) and an Ageratum enation virus (AgEV) during BLAST analysis of sequence data. Nucleotide- and amino acid-based phylogenetic analysis revealed the close relationship of the isolate with ToCSV strains, therefore it has been identified as an isolate of ToCSV and C. officinalis is considered to be a new host of ToCSV begomovirus. Association of a DNA-β molecule with the virus isolate was also detected by PCR and Southern hybridization tests using DNA-β specific primers and probe.     

寄主植物对B型烟粉虱个体发育和种群繁殖的影响

同一原始种群B型烟粉虱[Bemisia tabaci(Gennadius)],分别在甘蓝、一品红、棉花、番茄和黄瓜连续隔离饲养大约20代后形成5种对应的烟粉虱种群,研究其在甘蓝上各虫态大小、存活率、发育历期、成虫寿命和平均产卵量等生物学参数。在(25±1)℃条件下,一品红种群烟粉虱的1、2、3龄若虫体长和宽均显著低于其他4个种群烟粉虱;发育时间以甘蓝和一品红种群烟粉虱(20.44、19.69d)显著高于番茄种群烟粉虱(19.13d);雌虫寿命以甘蓝种群烟粉虱显著低于一品红、棉花、番茄和黄瓜种群烟粉虱;平均单雌产卵量以一品红种群烟粉虱(169.30粒)显著高于其他4个种群烟粉虱;5个种群烟粉虱在甘蓝上的总存活率均无显著差异。     

烟粉虱取食感染TYLCV番茄对其解毒酶和保护酶活性的影响

为了明确感染番茄黄化曲叶病毒(Tomato yellow leaf curl virus,TYLCV)的番茄对烟粉虱生理防御的影响,本文采用生化分析法研究了烟粉虱取食感染TYLCV的番茄植株后解毒酶和保护酶活性的变化。结果表明,B型烟粉虱在带毒番茄上取食72h及30d(长期饲养种群)后,其谷胱甘肽S-转移酶(GSTs)和羧酸酯酶(CarE)活性呈现先升高后下降的趋势,而P450酶活性则持续上升,取食30d种群是对照种群的1.99倍;三大保护酶系超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、过氧化物酶(POD)则在取食72h后,酶活性高于对照,而在感染TYLCV番茄上取食30d后,保护酶活性则持续高于72h及对照处理,分别是对照种群的1.45倍、8.42倍和5.03倍,差异显著;Q型烟粉虱表现趋势与B型烟粉虱相似。研究结果为进一步明确入侵烟粉虱与双生病毒之间的互作、揭示烟粉虱的适应性机制等补充了资料。     

番茄褪绿病毒侵染黄瓜的首次报道

番茄褪绿病毒Tomato chlorosis virus(ToCV)是严重危害世界经济作物的一种病毒,寄主范围广泛。田间调查发现黄瓜Cucumis sativus表现出叶片黄化、脉间褪绿的疑似番茄褪绿病毒感病症状,同时叶片背面聚集了大量烟粉虱。采用RT-PCR方法对样品叶片和烟粉虱进行检测,ToCV感染率为65%,且发病叶片上烟粉虱携带ToCV。为进一步确定黄瓜是否为番茄褪绿病毒的新寄主,室内利用农杆菌侵染性克隆接种健康黄瓜,结果显示:接种30 d的黄瓜新生叶片出现褪绿症状。采用ToCV HSP70基因的引物对田间黄瓜叶片、烟粉虱和室内黄瓜新生叶片进行RT-PCR,扩增出约450 bp的条带,在NCBI上BLAST显示与KC887999.1的同源性最高,为99%。这些数据表明黄瓜是番茄褪绿病毒的寄主。这是ToCV感染黄瓜的首次报道。     

A general model of plant-virus disease infection incorporating vector aggregation

In plant-virus disease epidemiology, dynamical models have invariably incorporated a bilinear inoculation rate that is directly proportional to both the abundance of healthy (susceptible) hosts and the abundance of infective vectors. Similarly, the acquisition rate is usually assumed to be directly proportional to the abundance of nonviruliferous vectors and that of infectious hosts. These bilinear assumptions have been questioned for certain human diseases, and infection rates that incorporate power parameters of the variables have been proposed. Here, infection rates for plant-virus diseases that are of a more general form than the familiar bilinear terms are examined. For such diseases, the power parameter can be regarded as a measure of the spatial aggregation of the vectors or as a coefficient of interference between them, depending on the context.
Field data of cassava mosaic virus disease (CMD) incidence were examined. When vector population density and disease incidence were high, disease progress curves over the first 6 months from planting could not be explained using models with bilinear infection rates. Incorporation of the new infection terms allowed the range of observed disease progress curve types to be described. New evidence of a mutually beneficial interaction between the viruses causing CMD and the whitefly vector, Bemisia tabaci , has shown that spatial aggregation of the vectors is an inevitable consequence of infection, particularly with a severe virus strain or a sensitive host. Virus infection increases both vector fecundity and the density of vectors on diseased plants. It is postulated that this enhances disease spread by causing an increased emigration rate of infective vectors to other crops. Paradoxically, within the infected crop, vector aggregation reduces the effective contact rate between vector and host and therefore the predicted disease incidence is less than when a bilinear contact rate is used.