Mercurialis ambigua and Solanum luteum: Two Newly Discovered Natural Hosts of Tomato Yellow Leaf Curl Geminiviruses

The yellow leaf curl disease of tomato is caused by a complex of virus species, two of which, tomato yellow leaf curl virus (TYLCV)-Sar and TYLCV-Is, are involved in epidemics of southern Spain. Plants of Mercurialis ambigua and Solanum luteum showing abnormal upward leaf curling and leaf distortion collected in the vicinity of tomato crops were found to be naturally infected with TYLCV-Is and TYLCV-Sar, respectively. These weed species, as well as Datura stramonium and S. nigrum, which had also been found to be naturally infected by TYLCVs in the same region in previous studies, were tested for susceptibility to TYLCV-Sar or TYLCV-Is by Agrobacterium tumefaciens-mediated and by Bemisia tabaci inoculation. Results indicated that both TYLCV-Sar and TYLCV-Is were able to infect D. stramonium and M. ambigua, whereas only TYLCV-Sar infected S. nigrum and S. luteum. Implications for the epidemiology of TYLCV are discussed. This is the first report of M.ambigua and S. luteum as hosts of TYLCV.     

A selective barrier in the midgut epithelial cell membrane of the nonvector whitefly <Emphasis Type="Italic">Trialeurodes vaporariorum</Emphasis> to <Emphasis Type="Italic">Tomato yellow leaf curl virus</Emphasis> uptake

We studied the presence of a potential transmission barrier that blocks Tomato yellow leaf curl virus in the nonvector greenhouse whitefly, Trialeurodes vaporariorum. Because T. vaporariorum can ingest and retain the virus after acquisition feeding on an infected plant, comparable to the vector whitefly Bemisia tabaci, circumstance evidence suggested that a transmission barrier presents at location(s) where the virus moves from the digestive tract lumen to the hemolymph. To provide direct evidence for the site of a transmission barrier in the nonvector insect, we compared the accumulation levels and localization of the virus between the two species of whiteflies. Quantitative real-time and conventional PCR analysis showed that accumulation of the virus during acquisition feeding and retention after a short acquisition period were indistinguishable between the two species, but the circulation of the virus within the whiteflies differed significantly between the species. In an immunofluorescence analysis using an antibody specific to the coat protein of the virus, the virus was restricted to the luminal surface of the midgut epithelial cells and did not enter their cytoplasm or that of the salivary glands in T. vaporariorum. In contrast, the virus was localized within the cytoplasm of the midgut epithelial cells and in the paired salivary glands of B. tabaci adults. This direct evidence shows that a selective transmission barrier at the luminal membrane surface of midgut epithelial cells in the nonvector whitefly blocks entrance of the virus into the midgut epithelial cells, resulting in incompetence as a vector of the virus.     

Differentiation of three whitefly-transmitted geminiviruses from the Republic of Yemen

Three viruses collected in southern Yemen in 1990, infecting watermelon, tobacco and tomato were shown to be transmitted by the whiteflyBemisia tabaci and to have particle morphologies typical of geminiviruses. Colonies ofB. tabaci collected from different locations and from different hosts were used in virus transmission tests with the same host range of plants. Colonies established from both watermelon and cotton in the Yemen were identified as the squash silverleaf-inducing B biotype. The culture host of the colony did not influence virus acquisition and transmission efficiencies to and from other hosts. The tobacco and tomato geminiviruses had a similar host range, but differed in their severity in some hosts. Both these viruses differed from the watermelon geminivirus in host range and symptoms.Datura stramonium, an alternative host for all three viruses, could be co-infected by the watermelon and tobacco viruses.B. tabaci was able to acquire both viruses from the co-infectedD. stramonium and infect seedlings of either original host plant species with their respective viruses orD. stramonium with both. The viruses were identified as watermelon chlorotic stunt virus, tobacco leaf curl virus and tomato yellow leaf curl virus and were distinguished by cross hybridisation.     

Comparison of Resistance to Tomato Leaf Curl Virus (India) and Tomato Yellow Leaf Curl Virus (Israel) among Lycopersicon Wild Species, Breeding Lines and Hybrids

The objective of this study was to screen wild and domesticated tomatoes for resistance to Tomato yellow leaf curl virus, Israel (TYLCV-Is) and Tomato leaf curl virus from Bangalore isolate 4, India (ToLCV-[Ban4]) to find sources of resistance to both viruses. A total of 34 tomato genotypes resistant/tolerant to TYLCV-Is were screened for resistance to ToLCV-[Ban4] under glasshouse and field conditions at the University of Agricultural Sciences, Bangalore, India. Resistance was assessed by criteria like disease incidence, symptom severity and squash-blot hybridization. All the tomato genotypes inoculated with ToLCV-[Ban4] by the whitefly vector Bemisia tabaci (Gennadius) produced disease symptoms. In some plants of the lines 902 and 910, however, the virus was not detected by hybridization. The tomato genotypes susceptible to ToLCV-[Ban4] by whitefly-mediated inoculation were also found susceptible to the virus under field conditions. However, there were substantial differences between genotypes in disease incidence, spread, symptom severity and crop yield. Despite early disease incidence, many genotypes produced substantially higher yields than the local hybrid, Avinash-2. Sixteen tomato genotypes from India resistant/tolerant to ToLCV-[Ban4] were also tested for TYLCV-Is resistance at the Hebrew University of Jerusalem, Rehovot, Israel. Accessions of wild species, Lycopersicon hirsutum LA 1777 and PI 390659 were the best sources of resistance to both viruses. Lines 902 and 910, which were, resistant to TYLCV-Is were only tolerant to ToLCV-[Ban4] and accession Lycopersicon peruvianum CMV Sel. INRA, resistant to ToLCV-[Ban4], was only tolerant to TYLCV-Is. Implications of using the resistant lines in breeding programme is discussed.     

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.     

Evidence from nucleic acid hybridization tests for geminivirus infection of ornamental crotons in India

Reduced growth of plants and excessive generalised and spotty yellowing of leaves occurred in ornamental crotons that were infected with a geminivirus, which was detected by spot hybridization tests using a cloned probe derived from DNA-A of acalypha yellow mosaic geminivirus. The croton virus was transmitted by the whitefly,Bemisia tabaci, toNicotiana tabacum andAcalypha indica, in which it caused leaf curl and yellow mosaic, respectively, but it was not transmitted to five other species. Infected crotons may therefore serve as reservoirs of a geminivirus that is able to infect a crop species.     

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.     

Relationships of the Sardinian isolate of tomato yellow leaf curl geminivirus with its whitefly vectorBemisia tabaci Gen

Relationships of an Italian isolate of tomato yellow leaf curl geminivirus from Sardinia (TYLCV-S) with its whitefly vectorBemisia tabaci were studied by means of experimental transmissions from tomato to tomato plants. TYLCV-S was confirmed to be transmitted in a persistent, circulative manner. The minimum latent period in the vector was between 17 and 20 h from the beginning of the acquisition access period (AAP). The maximum retention of infectivity was 8 days from the end of the AAP. Both acquisition and inoculation feeding times influenced the detected proportion of infective insects, with patterns well described by an exponential model. Acquisition was more efficient than inoculation. Males were significantly less efficient vectors than females. Nymphs were as efficient as adults in acquiring the virus. The length of AAP influenced both the retention of infectivity, and the pattern of transmission in serial transfer transmission tests with individual females. No significant difference in transmission efficiency was detected between two colonies ofB. tabaci, one inducing typical silverleaf symptoms on squash, the other inducing only mild symptoms with more than 50 whiteflies per plant. The phenomenon of periodic acquisition was not unequivocally proved for TYLCV-S.Research supported by the National Research Council of Italy, Special Project RAISA, Sub-project N. 2. Paper N. 1961.Supported by a grant from the Istituto Agronomico Mediterraneo, Valenzano (Bari), Italy.     

南疆温室番茄黄化曲叶病病毒种类的分子鉴定

为明确南疆温室番茄黄化曲叶病的病毒种类,利用双生病毒的兼并引物通过PCR扩增,对采集的20个番茄病株进行了分子检测.从20个病株中均扩增到约500 bp的目标片段,对其中4株进行克隆和测序,其相互间序列同源性为97.1％ ～99.3％,与番茄黄化曲叶病毒(Tomato yellow leaf curl virus,TYLCV)的同源性较高,为98.6％ ～ 99.5％.随机选取莎车分离物KS2-5进行全基因组的克隆和测序,KS2-5 DNA全长为2781 nt(序列号:JQ807735),具有典型的双生病毒基因组特征,与TYLCV其它分离物同源性达到98.9％～99.5％,而与其它粉虱传双生病毒的序列同源性较低,为68.3％ ～75.5％,表明危害南疆温室番茄的病毒种类为番茄黄化曲叶病毒TYLCV.     

Tomato leaf curl Guangxi virus is a distinct monopartite begomovirus species

Three begomovirus isolates were obtained from tomato plants showing leaf curl symptoms in Guangxi province of China. Typical begomovirus DNA components representing the three isolates (GX-1, GX-2 and GX-3) were cloned and their full-length sequences were determined to be 2752 nucleotides. Nucleotide identities among the three viral sequences were 98.9–99.7%, but all shared <86.7% nucleotide sequence identity with other reported begomoviruses. The sequence data indicated that GX-1, GX-2 and GX-3 are isolates of a distinct begomovirus species for which the name Tomato leaf curl Guangxi virus (ToLCGXV) is proposed. Further analysis indicated that ToLCGXV probably originated through recombination among viruses related to Ageratum yellow vein virus, Tomato leaf curl China virus and Euphorbia leaf curl virus. PCR and Southern blot analyses demonstrated that isolates GX-1 and GX-2 were associated with DNAβ components, but not isolate GX-3. Sequence comparisons revealed that GX-1 and GX-2 DNAβ components shared the highest sequence identity (86.2%) with that of Tomato yellow leaf curl China virus (TYLCCNV). An infectious construct of ToLCGXV isolate GX-1 (ToLCGXV-GX) was produced and determined to be highly infectious in Nicotiana benthamiana, N. glutinosa, tobacco cvs. Samsun and Xanthi, tomato and Petunia hybrida plants inducing leaf curl and stunting symptoms. Co-inoculation of tomato plants with ToLCGXV-GX and TYLCCNV DNAβ resulted in disease symptoms similar to that caused by ToLCGXV-GX alone or that observed in infected field tomato plants.     

Pymetrozine interferes with transmission of<Emphasis Type="Italic">Tomato yellow leaf curl virus</Emphasis> by the whitefly<Emphasis Type="Italic">Bemisia tabaci</Emphasis>

Pymetrozine, a novel compound belonging to the class pyridine-azomethines, is a feeding inhibitor labeled for use against plant pests in the order Hemiptera. Pymetrozine was evaluated for its ability to interfere with whitefly transmission of the begomovirusTomato yellow leaf curl virus (TYLCV). Pymetrozine was applied as Fulfill ^TM 50 WG at two rates (0.291 and 0.582 g formulationl ⁻¹) to tomato seedlings with four to six true leaves. Viruliferous whiteflies (three to five per plant) were added 1, 4, 7 and 11 d after a single application of pymetrozine, and transmission rates were determined 4 wk after the addition of whiteflies. Pymetrozine provided protection against transmission of TYLCV by viruliferous whiteflies for up to 1 wk after a single apliation. No phytotoxicity was observed on tomato transplants. These results indicate that pymetrozine could be an effective tool for tomato transplant producers to protect susceptible transplants from infection by begomoviruses, such as TYLCV. Pymetrozine might also work well as part of an integrated approach to begomovirus management in greenhouse tomato fruit production. http://www.phytoparasitica.org positing Oct. 20, 2003.     

Characterisation of a Quantitative Resistance to Vector Transmission of Tomato yellow leaf curl virus in Lycopersicon pimpinellifolium

Two wild genotypes from the same species Lycopersicon pimpinellifolium, WVA106 (susceptible) and INRA-Hirsute (so-called ‘resistant’), were compared with respect to their reaction to Tomato yellow leaf curl virus isolate Réunion (TYLCV-Mld[RE]), using both whitefly-mediated inoculation and graft inoculation. Disease incidence and symptom severity were scored. Presence and quantification of viral DNA were assessed by dot blot hybridisation. Upon insect inoculation, accession INRA-Hirsute showed a moderate resistance against TYLCV that was overcome by a high inoculation pressure obtained by increasing the cumulative number of inoculative whiteflies. Temporal analyses of the disease progress in relation to this criterion exhibited that the protection was quantitative, mainly reducing the TYLCV-Mld[RE] incidence by at maximum 50% at low inoculation pressure. When graft inoculated, the final TYLCV-Mld[RE] disease incidence was 100% in both susceptible and resistant genotypes with severe symptoms, suggesting a reduction of virus transmission by a vector resistance as a possible mechanism. Implications of using such type of resistance in breeding programmes are discussed.     

<Emphasis Type="Italic">Physalis ixocarpa</Emphasis> and <Emphasis Type="Italic">P. peruviana</Emphasis>, new natural hosts of <Emphasis Type="Italic">Tomato chlorosis virus</Emphasis>

Tomato chlorosis virus causes yellow leaf disorder epidemics in many countries worldwide. Plants of Physalis ixocarpa showing abnormal interveinal yellowing and plants of Physalis peruviana showing mild yellowing collected in the vicinity of tomato crops in Portugal were found naturally infected with ToCV. Physalis ixocarpa and P. peruviana were tested for susceptibility to ToCV by inoculation with Bemisia tabaci, Q biotype. Results confirmed that ToCV is readily transmissible to both species. The infection was expressed in P. ixocarpa by conspicuous interveinal yellow areas on leaves that developed into red or brown necrotic flecks, while P. peruviana test plants remained asymptomatic. Infected plants of both P. ixocarpa and P. peruviana served as ToCV sources for tomato infection via B. tabaci transmission. This is the first report of P. ixocarpa and P. peruviana as natural hosts of ToCV.     

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.     

烟粉虱传播的番茄褪绿病毒和番茄黄化曲叶病毒对不同番茄品种的复合侵染

为明确烟粉虱传播的番茄褪绿病毒(Tomato chlorosis virus,ToCV)与番茄黄化曲叶病毒(Tomato yellow leaf curl virus,TYLCV)对不同番茄品种的复合侵染情况,于2015年11月在山东省寿光市温室内采集13个番茄品种共390份疑似发病植株叶片,对不同番茄品种的TYLCV抗性和2种病毒的复合侵染以及温室内发病番茄植株上烟粉虱成虫的带毒率进行检测。结果表明,采集的13个番茄品种经分子标记检测鉴定均为TYLCV杂合抗性;不同番茄品种ToCV与TYLCV的复合侵染率存在明显差异,大果番茄粉宴和贝瑞上复合侵染率最高可达73.3%,而樱桃番茄八喜上未检测到这2种病毒的复合侵染。此外,在发病番茄植株上采集的烟粉虱成虫体内可检测到2种病毒,其中烟粉虱ToCV带毒率为90.7%,TYLCV带毒率为80.0%,同时检测到ToCV与TYLCV的概率为71.3%。表明ToCV和TYLCV的复合侵染在山东省番茄生产中普遍发生,烟粉虱可同时携带这2种病毒并广泛传播。     

Visualisation of <Emphasis Type="Italic">hrp</Emphasis> gene expression in <Emphasis Type="Italic">Xanthomonas euvesicatoria</Emphasis> in the tomato phyllosphere

The plasmid pUFZ75 conferred constitutive GFP expression on the bacterial pathogen Xanthomonas euvesicatoria (syn. X. campestris pv. vesicatoria). Colonisation of the tomato phyllosphere and invasion of tomato leaves by X. euvesicatoria was examined using both fluorescence and confocal laser scanning microscopy. Xanthomonas euvesicatoria established a limited population on the tomato leaf surface, primarily occupying the depressions between epidermal cells and around the stomata, prior to invasion of the leaf via the stomata and subsequent growth within the substomatal chamber and the leaf apoplast. Additionally, hrp-gfp fusions were used to report on the temporal and spatial expression of hrp genes during epiphytic colonisation and invasion. Xanthomonas euvesicatoria cells carrying hrpG- and hrpX-gfp reporter constructs were not fluorescent in vitro on non-hrp-inducing LB agar but did exhibit a low level of fluorescence on the leaf surface within 24 h of inoculation, particularly in the vicinity of stomata. Cells carrying hrpG- and hrpX-gfp fusions exhibited high levels of fluorescence 72 h after inoculation in the substomatal chamber and the leaf apoplast. Cells carrying the hrpF-gfp fusion were slightly fluorescent on LB agar and showed no further increase in fluorescence on the leaf surface by 24 h after inoculation, but did show a significant increase in fluorescence 72 h after inoculation in the substomatal chamber and apoplast. The apparent low-level induction of the regulators hrpG and hrpX on the tomato leaf surface may suggest that some of the genes of the X. euvesicatoria HrpG/HrpX regulon are up- or down-regulated prior to invasion of the stomata while still on the leaf surface.     

Arabidopsis thaliana,an experimental host for tomato yellow leaf curl disease‐associated begomoviruses by agroinoculation and whitefly transmission

Tomato yellow leaf curl disease is one of the most devastating viral diseases affecting tomato crops worldwide. This disease is caused by several begomoviruses (genus Begomovirus, family Geminiviridae), such as Tomato yellow leaf curl virus (TYLCV), that are transmitted in nature by the whitefly vector Bemisia tabaci. An efficient control of this vector‐transmitted disease requires a thorough knowledge of the plant–virus–vector triple interaction. The possibility of using Arabidopsis thaliana as an experimental host would provide the opportunity to use a wide variety of genetic resources and tools to understand interactions that are not feasible in agronomically important hosts. In this study, it is demonstrated that isolates of two strains (Israel, IL and Mild, Mld) of TYLCV can replicate and systemically infect A. thaliana ecotype Columbia plants either by Agrobacterium tumefaciens‐mediated inoculation or through the natural vector Bemisia tabaci. The virus can also be acquired from A. thaliana‐infected plants by B. tabaci and transmitted to either A. thaliana or tomato plants. Therefore, A. thaliana is a suitable host for TYLCV–insect vector–plant host interaction studies. Interestingly, an isolate of the Spain (ES) strain of a related begomovirus, Tomato yellow leaf curl Sardinia virus (TYLCSV‐ES), is unable to infect this ecotype of A. thaliana efficiently. Using infectious chimeric viral clones between TYLCV‐Mld and TYLCSV‐ES, candidate viral factors involved in an efficient infection of A. thaliana were identified.     

陕西省早春茬番茄定植时间与番茄黄化曲叶病毒病发生的关系

为了明确早春茬番茄定植时间与番茄黄化曲叶病毒病发生的关系,采取小区对比试验对不同定植时间下番茄上烟粉虱种群消长动态、番茄黄化曲叶病毒病发病株率、发病程度及番茄产量和经济效益等进行了系统调查。结果表明:番茄田烟粉虱种群消长动态在年度间基本相同,4月上旬始见,5月中旬达到高峰,烟粉虱发生后15 d左右出现番茄黄化曲叶病毒病症状。4月上旬以前定植番茄黄化曲叶病毒病发生较轻,单位面积产量和经济效益较高,4月上旬以后定植发病较重,产量和经济效益较低。定植时间(X_1)与番茄黄化曲叶病毒发病株率(Y_1)及病情指数(Y_2)均呈显著正相关,其相关关系为Y_1=1.470 4X_1+7.947 2,Y_2=0.876 7X_1-6.441 7。生产实践中采取双膜或三膜覆盖栽培等措施适当提早定植,避免番茄感病期与烟粉虱发生高峰期相遇,在不使用任何防治措施下,可有效预防番茄黄化曲叶病毒病暴发成灾。     

Identification of a CAPS marker tightly linked to the Tomato yellow leaf curl disease resistance gene <Emphasis Type="Italic">Ty-1</Emphasis> in tomato

During the process of breeding programmes, several resistance genes have been introgressed into tomato (Solanum lycopersicum) cultivars from different wild tomato relatives. A number of these resistance genes have been mapped to chromosome 6. Among them, Ty-1 and Mi, which confer resistance to Tomato yellow leaf curl disease and to Meloidogyne spp., respectively, are in most cases incorporated in commercial hybrids. Several molecular markers tightly linked to Mi have been identified. This study was conducted in order to find an informative molecular marker linked to Ty-1. Six markers mapped in the same region as Ty-1 were analysed in plant material carrying different combinations of Ty-1 and Mi alleles. Three of the six markers revealed polymorphism among the assayed accessions. One allele of JB-1 marker showed association with Ty-1. Furthermore, the presence of Mi did not interfere with the results. The analysis of several accessions of wild tomato relatives with the three polymorphic markers allowed the establishment of the origin of the alleles found in cultivated plant material, showing that introgressions from S. lycopersicum, S. pimpinellifolium and S. habrochaites will not interfere with the results of this marker which tags Ty-1. Furthermore this analysis enabled the location of CT21, the RFLP marker from which JB-1 was designed.     

番茄黄化曲叶病毒北京分离物侵染性克隆的构建与其致病性

为明确番茄黄化曲叶病毒北京分离物（Beijing isolate of tomato yellow leaf curl virus,TYLCV-BJ）致病性的强弱,以感染TYLCV-BJ的番茄叶片DNA为模板PCR扩增获得该分离物基因组全长序列,并构建该分离物的侵染性克隆,将其分别接种到番茄、烟草和拟南芥植株上,比较该分离物和TYLCV上海分离物2(TYLCV-Shanghai 2,TYLCV-SH2)致病性的差异。结果显示,该分离物基因组全长序列同TYLCV-SH2的相似度为99.03%,在番茄和烟草植株上TYLCV-BJ比TYLCV-SH2发病更早,症状更重,TYLCV DNA和外壳蛋白积累量更高。TYLCV-BJ可以通过农杆菌Agrobacterium tumefaciens注射法在拟南芥中复制和系统侵染,而TYLCV-SH2不能有效侵染拟南芥。表明TYLCV-BJ的致病性强于TYLCV-SH2,所建立的侵染性克隆有广泛的研究和应用价值。