Capsicum Species: Symptomless Hosts and Reservoirs of Tomato yellow leaf curl virus

ABSTRACT Five Capsicum species were tested for susceptibility to Tomato yellow leaf curl virus (TYLCV) and the mild strain of TYLCV (TYLCV-Mld). TYLCV was able to infect 30 of 55 genotypes of C. annuum, one of six genotypes of C. chinense, one of two genotypes of C. baccatum, and the only genotype of C. frutescens tested but was unable to infect the one genotype of C. pubescens tested. This is the first evidence for the susceptibility of C. baccatum, C. chinense, and C. frutescens to TYLCV. Unlike TYLCV isolates, TYLCV-Mld was unable to infect C. chinense. No host differences were observed between the Israeli and Florida isolates of TYLCV. None of the Capsicum species showed symptoms after infection with TYLCV or TYLCV-Mld. TYLCV was detected in fruits of C. annuum, but whiteflies were unable to transmit virus from fruits to plants. White-flies were able to transmit both TYLCV and TYLCV-Mld from infected pepper plants to tomato plants. Pepper plants in research plots were found infected with TYLCV at rates as much as 100%. These data demonstrate the ability of some genotypes of pepper to serve as reservoirs for the acquisition and transmission of TYLCV and TYLCV-Mld.     

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.     

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.     

Pepper (Capsicum annuum) Is a Dead-End Host for Tomato yellow leaf curl virus

ABSTRACT Tomato yellow leaf curl (TYLC) is one of the most devastating pathogens affecting tomato (Lycopersicon esculentum) worldwide. The disease is caused by a complex of begomovirus species, two of which, Tomato yellow leaf curl Sardinia virus (TYLCSV) and Tomato yellow leaf curl virus (TYLCV), are responsible for epidemics in Southern Spain. TYLCV also has been reported to cause severe damage to common bean (Phaseolus vulgaris) crops. Pepper (Capsicum annuum) plants collected from commercial crops were found to be infected by isolates of two TYLCV strains: TYLCV-Mld[ES01/99], an isolate of the mild strain similar to other TYLCVs isolated from tomato crops in Spain, and TYLCV-[Alm], an isolate of the more virulent TYLCV type strain, not previously reported in the Iberian Peninsula. In this work, pepper, Nicotiana benthamiana, common bean, and tomato were tested for susceptibility to TYLCV-Mld[ES01/99]and TYLCV-[Alm] by Agrobacterium tumefaciens infiltration, biolistic bombardment, or Bemisia tabaci inoculation. Results indicate that both strains are able to infect plants of these species, including pepper. This is the first time that infection of pepper plants with TYLCV clones has been shown. Implications of pepper infection for the epidemiology of TYLCV are discussed.     

Effect of Host Plant Resistance to Tomato yellow leaf curl virus (TYLCV) on Virus Acquisition and Transmission by Its Whitefly Vector

ABSTRACT The effect that Tomato yellow leaf curl virus (TYLCV)-infected resistant tomato plants may have on virus epidemiology was studied. Four tomato genotypes that exhibit different levels of viral resistance, ranging from fully susceptible to highly resistant, served as TYLCV-infected source plants. Viral acquisition and transmission rates by white-flies following feeding on the different source plants were evaluated. TYLCV transmission rate by whiteflies that had fed on infected source plants 21 days postinoculation (DPI), shortly after the appearance of TYLCV symptoms, was negatively correlated with the level of resistance displayed by the source plant. Therefore, the higher the resistance, the lower the transmission rate. In addition, TYLCV DNA accumulation was shown to be lower in the resistant source plants compared with the susceptible plants. Whitefly survival rate, following feeding on source plants 21 DPI, was similar for all the cultivars tested. Significant differences in whitefly survival were found, however, following feeding on the infected source plants at 35 DPI; here, whitefly survival rate increased with higher levels of resistance displayed by the source plant. At 35 DPI, the susceptible plants had developed severe TYLCV disease symptoms, and transmission rates from these plants were the lowest, presumably due to the poor condition of these plants. Transmission rates from source plants displaying a medium level of resistance level were highest, with rates declining following feeding on source plants displaying higher levels of TYLCV resistance. TYLCV DNA accumulation in whiteflies following feeding on infected source plants at both 21 and 35 DPI was directly correlated with viral DNA accumulation in source plants. Results show that, in essence, the higher the resistance expressed, the less suitable the plant was as a viral source. Consequently, following acquisition from a highly resistant plant, TYLCV transmission by whiteflies will be less efficient.     

Transmission of Tomato Yellow Leaf Curl Virus by two invasive biotypes and a Chinese indigenous biotype of the whitefly Bemisia tabaci

The global invasion of certain Bemisia tabaci biotypes provides opportunities to compare the competency of virus transmission between invasive and indigenous biotypes. Here we report on the acquisition, retention and transmission of Tomato Yellow Leaf Curl Virus (TYLCV) by the invasive B, Q and indigenous ZHJ2 biotypes of B. tabaci from Zhejiang, China. For all whitefly biotypes, TYLCV DNA was detected within a 30-min acquisition access period (AAP) to infected leaves. The percentage of adults with viral DNA increased with the length of AAP and reached 100% after 10–12 h. Following acquisition, viruliferous B, Q and ZHJ2 adults retained TYLCV DNA for the rest of their lives. Transmission was achieved with one B/Q adult per plant at the frequency of 50–55%, which rose to 100% with 10 insects per plant. In contrast, transmission of the virus was not observed with one ZHJ2 adult per plant in the experiments, and the transmission frequency rose to only 30–45% when whitefly adults increased to 5–10 per plant. These new data will help in the determination of the pest status of the whitefly biotypes as virus vectors in the regions of invasion, and so help in the development of management strategies.     

Endophytes of <Emphasis Type="Italic">Lippia citriodora</Emphasis> (Syn. <Emphasis Type="Italic">Aloysia triphylla</Emphasis>) enhance its growth and antioxidant activity

Sweet pepper (Capsicum annuum) is a popular crop worldwide and an asymptomatic host of the begomovirus (Geminiviridae) Tomato yellow leaf curl virus (TYLCV). A previous study showed that TYLCV could be transmitted by the seeds of tomato plants, but this phenomenon has not been confirmed in other plants. In 2015, four different cultivars of sweet pepper (‘Super Yellow,’ ‘Super Red,’ ‘Sunnyez’ and ‘Cupra’) known to be susceptible to TYLCV were agro-inoculated with a TYLCV infectious clone. Three months after inoculation, the leaves of the ‘Super Yellow’ cultivar showed 80% (8/10) susceptibility and the other three sweet pepper cultivars showed 30 to 50% susceptibilities. All of the ‘Super Yellow’ seed bunches (five seeds per bunch) from plants whose leaves were confirmed to be TYLCV-infected were also TYLCV-infected (8/8). The seeds of other cultivars showed 20 to 40% susceptibilities. Virus transmission rates were also verified with 10 bunches of seedlings for each cultivar (five seedlings per pool). Eight bunches of ‘Super Yellow’ seedlings (8/10) were confirmed to be TYLCV-infected and one to three bunches of each of the other cultivar seedlings were also infected. Viral replication in TYLCV-infected seeds and seedlings was confirmed via strand-specific amplification using virion-sense- and complementary-sense-specific primer sets. This is the first report of TYLCV seed transmission in sweet pepper plants and among non-tomato plants. Because sweet pepper is an asymptomatic host of TYLCV, seeds infected with TYLCV could act as a silent invader of tomatoes and other crops.     

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.     

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.     

Epidemiological studies of the tomato yellow leaf curl virus (TYLCV) in the Jordan Valley,Israel

The spread of tomato yellow leaf curl virus (TYLCV) is significantly correlated with the population size of its vector,Bemisia tabaci Genn. The perennial weedCynanchum acutum L. and the annual weedMalva parviflora L. were found to be natural hosts of TYLCV in the Jordan Valley.C. acutum is not a preferred host forB. tabaci, but the whitefly feeds on it sufficiently long to acquire the virus. Whiteflies marked with fluorescent dust while feeding naturally onC. acutum along the banks of the Jordan River, were subsequently trapped within the main tomato-production area 7 km away. An increase in theB. tabaci population and in TYLCV infectivity was found in plots surrounded by windbreaks. The epidemiological cycle of TYLCV is described and cultural control measures are suggested.     

番茄黄化曲叶病毒与番茄褪绿病毒复合侵染对番茄黄化曲叶病毒传播的影响

番茄黄化曲叶病毒(tomato yellow leaf curl virus, TYLCV)是一种由烟粉虱传播的单链环状DNA病毒, 在田间可与多种病毒发生复合侵染, 如番茄褪绿病毒(tomato chlorosis virus, ToCV)等?本文对比了TYLCV单独侵染和TYLCV与ToCV复合侵染对烟粉虱获取和传播TYLCV的影响?结果表明, 与取食TYLCV单独侵染的番茄相比, 取食复合侵染番茄的烟粉虱对TYLCV的传毒率显著提高, 且番茄植株和烟粉虱体内TYLCV的病毒积累量也显著提高?试验结果说明复合侵染会提高烟粉虱的传毒率, 促进TYLCV的发生与流行?     

Pcr-amplification of tomato yellow leaf curl virus (TYLCV) DNA from squashes of plants and whitefly vectors: Application to the study of TYLCV acquisition and transmission

DNA of tomato yellow leaf curl virus (TYLCV), a geminivirus transmitted by the whitefly Bemisia tabaci, was amplified from squashes of infected tomato plants and of viruliferous vectors using the polymerase chain reaction (PCR). Samples of infected tissues as small as 1 mm² were squashed onto a nylon membrane. A 1 × 2 mm strip containing the squash was introduced into a 25 µl PCR reaction mix. The reaction products were subjected to gel electrophoresis, blotted and hybridized with a radiolabeled virus-specific DNA probe. TYLCV DNA was amplified from squashes of leaves, roots, and stem of infected tomato and from individual viruliferous whiteflies. The same squash could be used several times to amplify different virus DNA fragments with various sets of primers. Thus plant and insect squashes can be used as templates for the amplification of geminiviral DNA with no need to prepare tissue extracts or purify nucleic acids. The squash-PCR procedure was applied to study whitefly transmission of TYLCV. Tomato plants were inoculated by placing a single viruliferous insect in the center of a young leaflet. In some plants TYLCV DNA was detected at the site of inoculation as early as 5 min after the beginning of the access feeding and in all plants after 30 min. The squash-PCR procedure also was applied to the study of TYLCV acquisition by the insect vector. TYLCV DNA was detected in the head of whiteflies as early as 5 min after the beginning of the access feeding on infected tomato plants. Viral DNA was detected in the thorax after 10 min and in the abdomen after 25 min.     

New natural hosts of Tomato yellow leaf curl virus identified in and near tomato-growing greenhouses in eastern China

Tomato yellow leaf curl virus (TYLCV) causes huge losses to tomato production worldwide. In July 2011 and July–August 2012, we screened for potential TYLCV hosts in a tomato-growing area in Shandong Province, the core vegetable-producing region in China. PCR detection showed that 5 species of plants, Zinnia elegans, Acalypha australis, Gossypium hirsutum, Abutilon theophrasti, and Nicotiana tabacum, were infected. Full genomic sequences of the new TYLCV isolates were obtained and submitted for sequence analysis. Sequence alignment and similarity analysis showed that they all belonged to the TYLCV-IL strain.     

Preparation and epitope characterization of monoclonal antibodies suitable for detection of <Emphasis Type="Italic">Tomato yellow leaf curl virus</Emphasis>

Tomato yellow leaf curl virus (TYLCV) is a begomovirus that seriously threatens tomato crops worldwide. Current immunodiagnostic methods for this pathogen employ commercially produced mAbs raised against TYLCV. However, despite the existence of these mAbs, little information regarding their characterization or strategy of production has been published. In addition, research on TYLCV would certainly benefit were more mAbs available, thus allowing efficient examination of the virus life cycle, modes of pathogenesis and possibly the development of resistant cultivars. The coat protein (CP) of TYLCV is the only known building block of the viral capsid. Therefore, in this study we used CP as an immunogen for the production of novel mAbs. We employed a strategy in which the CP was truncated at its C-terminus to avoid intra- and inter-protein interactions that could impair epitope exposure. For the same reason, we used a denaturated antigen to expose linear epitopes during the immunization. This effort yielded three mAbs: they were characterized biochemically and immunologically, and their epitopes were mapped. Possible applications of these mAbs are discussed.     

Tomato yellow leaf curl virus accumulates in vesicle-like structures in descending and ascending midgut epithelial cells of the vector whitefly, Bemisia tabaci, but not in those of nonvector whitefly Trialeurodes vaporariorum

Begomoviruses are transmitted by a single species of vector insect, the whitefly Bemisia tabaci, in a circulative manner. However, the mechanisms of this strict vector specificity have not been clarified. By immunoelectron microscopy, we showed that a begomovirus, Tomato yellow leaf curl virus (TYLCV), can enter midgut epithelial cells of the vector whitefly B. tabaci but not those of a nonvector whitefly, Trialeurodes vaporariorum, belonging to the same family. In midgut epithelial cells of viruliferous B. tabaci, the virus was localized in vesicle-like structures, suggesting endocytosis as an entry mechanism. These structures were also observed in midgut cells of nonviruliferous B. tabaci that had fed on healthy plants and in those of the nonvector T. vaporariorum that had fed on virus-infected plants. Vesicles containing TYLCV particles were observed most frequently in cells in the anterior part of the descending midgut, suggesting that this is the major entry site. These results clearly demonstrated that the virus-containing vector and nonvector whiteflies differ in the cellular localization of the virus and strongly suggest that a critical step in determining the vector insect specificity of begomoviruses is the entry of the viruses into midgut epithelial cells.     

侵染苘麻的菜豆金色黄花叶病毒的鉴定及基因组结构分析

为明确引起我国山西晋中地区苘麻叶片表现皱缩和花叶症状的病原物及其基因组分子特征, 本研究利用双生病毒简并引物扩增获得病毒基因组部分序列,经测序、比对后设计特异性引物扩增病毒基因组序列, 进而通过生物信息学方法构建系统发育树并进行序列分析。结果表明:引起苘麻叶片皱缩、花叶的病原物为番茄黄化曲叶病毒(tomato yellow leaf curl virus, TYLCV), 将该分离物命名为TYLCV-Abu, GenBank登录号为OP293347, 但未扩增到β卫星。该病毒DNA-A基因组全长为2 782 bp, 含有6个开放阅读框。TYLCV-Abu分离物与TYLCV茄子分离物KSQ1-3(GenBank登录号KC428753)的核苷酸序列一致性最高, 为98.99%, 其中C4和V2编码的蛋白变异较大。重组结果分析显示,分离物TYLCV-Abu是由TYLCV-F(GenBank登录号KY971326)和TYLCV-KSQ1-3重组得到, 重组区域为其基因组2 617－2 782 nt区域。这是首次从苘麻样品中扩增到TYLCV全基因组序列并进行分析。     

A simple, efficient agroinoculation soaking procedure for Tomato yellow leaf curl virus

Tomato yellow leaf curl virus (TYLCV, genus Begomovirus, family Geminiviridae), poses a serious threat to tomato crops in tropical and subtropical regions. We developed a simple agroinoculation method with an infectious clone of TYLCV. Dipping of excised sections of susceptible tomato shoots in an Agrobacterium suspension successfully introduced the replicating virus with high efficiency. An additional vacuum treatment for 5 min ensured uniform infection without escapes, allowing evaluation of differences in TYLCV resistance among tomato cultivars. The method can be used in laboratory experiments for virological studies and in breeding programs for resistant cultivars.     

First record of the Q Biotype of the sweetpotato whitefly, Bemisia tabaci, intercepted in the UK

For the UK, Bemisia tabaci poses a threat primarily to protected vegetable crops due to the transmission of several plant-pathogenic viruses. There are at least 24 different biotypes of B. tabaci that cannot be differentiated through morphological traits. The B (Middle East-Asia Minor 1 species) and Q (Mediterranean species) biotypes are widely considered to be the most important and, as such, the ability to rapidly and precisely biotype B. tabaci interceptions is vital when developing effective control strategies. Intercepted adult/pupal B. tabaci received from the UK Plant Health and Seeds Inspectorate (PHSI) during 2002–2003 (n?=?60) and 2010–2011 (n?=?42) were both biotyped and tested for the presence of Tomato yellow leaf curl virus (TYLCV) and Tomato yellow leaf curl Sardinia virus (TYLCSV) using a real-time PCR assay based on TaqMan? chemistry. The positive results indicated that during 2002–2003 the Q biotype comprised 68.3?% of the interceptions whilst in 2010–2011 it comprised 66.7?% of the B. tabaci samples intercepted. Only three of the B biotypes collected during 2002–2003 were positive for TYLCSV, two originating from Israel and the other of unknown origin. The implications in regards to pest management of the insect are discussed.     

Suppressive effect of acetylated glyceride (Bemidetach™EC) on <Emphasis Type="Italic">Tomato yellow leaf curl virus</Emphasis> transmitted by the whitefly <Emphasis Type="Italic">Bemisia tabaci</Emphasis> on greenhouse tomato plants

Acetylated glyceride (Bemidetach?EC)—a food additive—repels adult sweet potato whiteflies (Bemisia tabaci) and inhibits their mating behavior. We evaluated the effects of acetylated glyceride spraying of greenhouse-grown tomato plants on infestation with B. tabaci and the occurrence of Tomato yellow leaf curl virus (TYLCV) disease under commercial-like conditions. The abundance of adult B. tabaci was significantly reduced by three sprayings of acetylated glyceride, and the TYLCV incidence was significantly suppressed to less than 30% of that in the untreated control. These results suggest that acetylated glyceride sprays suppress the secondary spread of TYLCV in greenhouse-grown tomatoes by lessening B. tabaci adult density.