A new strain of tomato severe leaf curl virus and a unique variant of tomato yellow leaf curl virus from Mexico

The complete genome sequence of a distinct variant of tomato yellow leaf curl virus-Israel (TYLCV-IL) and the DNA-A sequence of a new strain of tomato severe leaf curl virus (ToSLCV) isolated in San Luis Potosi, Mexico, are described and analyzed. The TYLCV-IL[MX:SLP:11] variant differs from all TYLCV-IL isolates described so far by a unique 42-nt duplicated sequence comprising a part of the conserved stem-loop element of the virion-strand replication origin and adjacent regulatory sequences. TYLCV-IL[MX:SLP:11] was associated with tomato chino La Paz virus (ToChLPV-B[MX:SLP:11]) in a Solanum pimpinellifolium plant, and with pepper huasteco yellow vein virus (PHYVV-[MX:SLP:11]) and ToSLCV-GT[MX:SLP:11] in a Solanum lycopersicum plant. In addition, a distinct ToSLCV exhibiting low sequence identity (<89?%) to other ToSLCV isolates from Mexico was found in a tomato plant collected in the same field. Sequence analysis of this new ToSLCV strain indicates that it is a recombinant of close relatives of ToSLCV-GT[MX:SLP:11] and ToChLPV-B[MX:SLP:11] found in mixed infections with TYLCV-IL[MX:SLP:11].     

Only minimal regions of tomato yellow leaf curl virus (TYLCV) are required for replication,expression and movement

The IL-60 platform, consisting of a disarmed form of tomato yellow leaf curl virus (TYLCV) and auxiliary components, was previously developed as a nontransgenic universal vector system for gene expression and silencing that can express an entire operon in plants. IL-60 does not allow rolling-circle replication; hence, production of viral single-stranded (ss) DNA progeny is prevented. We used this double-stranded (ds) DNA-restricted platform (uncoupled from the dsDNA→ssDNA replication phase of progeny viral DNA) for functional genomics studies of TYLCV. We report that the noncoding 314-bp intergenic region (IR) is the only viral element required for viral dsDNA replication. None of the viral genes are required, suggesting recruitment of host factors that recognize the IR. We further show that IR-carrying reporter genes are also capable of replication but remain confined to the cells into which they were introduced. Only two sense-oriented viral genes (V1 and V2) need to be added to the IR-carrying construct for expression and movement. Hence, any IR-dsDNA construct supplemented with V1 and V2 becomes a replication-competent, mobile and expressing plant plasmid. All viral functions (replication, expression and movement) are determined by the IR and the sense-oriented genes. The complementary-oriented viral genes have auxiliary roles in the late phase of the virus “life cycle”. The previously reported involvement of some viral genes in expression and movement is therefore revised.     

Biolistic inoculation of plants with tomato yellow leaf curl virus DNA

Tomato yellow leaf curl virus (TYLCV) full-length DNA was amplified by PCR and cloned into a bacterial plasmid. The cloned TYLCV DNA was excised from the plasmid, ligated and the resulting monomeric circular double-stranded TYLCV DNA was used to inoculate tomato (Solanum lycopersicom) and datura (Datura stramonium) plants by particle bombardment. The bombarded plants produced typical disease symptoms, similar to those produced following whitefly-mediated inoculation, albeit 5-7 days later than whitefly-inoculated plants. The success rate of inoculating tomato plants by particle bombardment averaged 37%, whereas with datura plants, it averaged 85%. With whitefly-mediated inoculation of TYLCV, the success rate of inoculation was also higher in datura plants than in tomato plants. Bombardment of datura plants with a linear form of TYLCV DNA also resulted in viral infection, with an inoculation success rate similar to that with the closed-circular TYLCV DNA. Bombarding datura plants with the bacterial plasmid containing the cloned TYLCV DNA did not result in viral infection, but bombardment with a bacterial plasmid containing a cloned dimer of TYLCV DNA yielded an infection rate of 50-100%. This is the first report of TYLCV inoculation of plants using particle bombardment of a cloned monomeric linear or closed-circular form of TYLCV double-stranded DNA.     

A worldwide survey of tomato yellow leaf curl viruses

Summary.  The name tomato yellow leaf curl virus (TYLCV) has been given to several whitefly-transmitted geminiviruses affecting tomato cultures in many tropical and subtropical regions. Hybridization tests with two DNA probes derived from a cloned isolate of TYLCV from Israel (TYLCV-ISR) were used to assess the affinities of viruses in naturally infected tomato plants with yellow leaf curl or leaf curl symptoms from 25 countries. Probe A which included most of the intergenic region was expected to detect only isolates closely related to TYLCV-ISR, especially after high stringency washes. In contrast probe B, which included the full-length genome, was expected to detect a wide range of whitefly-transmitted geminiviruses. Tomato samples from six countries in the Middle East, from Cuba or the Dominican Republic proved to be closely related to TYLCV-ISR and probably were infected by strains of the same virus. Samples from Senegal and Cape Verde Islands were also related to the Middle Eastern virus. Samples from nine other countries in the western Mediterranean area, Africa, or South-East Asia were more distantly related and probably represent one or more additional geminivirus species. Samples from five countries in Africa, Central or South America gave hybridization signals with the full-length viral genome, only after low stringency wash, indicating that these samples were infected by remote viruses. These results were supported by DNA and protein sequence comparison, which indicate that tomato geminiviruses fall into three main clusters representing viruses from 1) the Mediterranean/Middle East/African region, 2) India, the Far East and Australia, and 3) the Americas. Within the first cluster, two sub-clusters of viruses from the western Mediterranean or from the Middle East/Caribbean Islands were distinguished. The incidence of tomato yellow leaf curl diseases has increased considerably between 1990 and 1996. Accepted January 28, 1997; Received April 19, 1996     

Predominance of tomato leaf curl Gujarat virus as a monopartite begomovirus: association with tomato yellow leaf curl Thailand betasatellite

Tomato leaf curl is a serious malady in the state of Maharashtra, India, causing nearly 100 % yield loss. An extensive survey was done in the affected fields of tomato in the year 2008, and members of three species of begomoviruses were identified as causing the disease. More than 60 % of the samples from diseased plants were infected with tomato leaf curl Gujarat virus (ToLCGuV). Isolates collected from these fields differed from the Varanasi isolate of ToLCGuV in not having a DNA B component. Instead, they were like typical Old World monopartite begomoviruses in that they were associated with only one betasatellite, tomato yellow leaf curl Thailand betasatellite (TYLCTHB). ToLCGuV alone is readily infectious, expressing systemic symptoms in Nicotiana benthamiana and tomato. Co-inoculation of ToLCGuV with TYLCTHB, increased symptom severity and reduced the incubation time required for symptom expression. ToLCGuV successfully interacted with heterologous DNA B component of ToLCNDV [IN:Pun:JID:08], and co-inoculation of these two resulted in yellow mottling symptoms that were typical of DNA B.     

Lamium amplexicaule (Lamiaceae): a weed reservoir for tomato yellow leaf curl virus (TYLCV) in Korea

After the first identification of tomato yellow leaf curl virus (TYLCV) in the southern part of Korea in 2008, TYLCV has rapidly spread to tomato farms in most regions of Korea. From 2008 to 2010, a survey of natural weed hosts that could be reservoirs of TYLCV was performed in major tomato production areas of Korea. About 530 samples were collected and identified as belonging to 25 species from 11 families. PCR and Southern hybridization were used to detect TYLCV in samples, and replicating forms of TYLCV DNA were detected in three species (Achyranthes bidentata, Lamium amplexicaule, and Veronica persica) by Southern hybridization. TYLCV transmission mediated by Bemisia tabaci from TYLCV-infected tomato plants to L. amplexicaule was confirmed, and TYLCV-infected L. amplexicaule showed symptoms such as yellowing, stunting, and leaf curling. TYLCV from infected L. amplexicaule was also transmitted to healthy tomato and L. amplexicaule plants by B. tabaci. The rate of infection of L. amplexicaule by TYLCV was similar to that of tomato. This report is the first to show that L. amplexicaule is a reservoir weed host for TYLCV.     

Molecular characterization and pathogenicity of tomato yellow leaf curl virus in China

Several tomato production regions in China were surveyed for tomato yellow leaf curl disease (TYLCD), and 31 tomato leaf samples showing TYLCD-like symptoms were collected. The partial or full-length genomes of these isolates were sequenced and tomato yellow leaf curl virus (TYLCV) was detected in Shanghai, Zhejiang, Jiangsu Shandong and Hebei provinces of China. The TYLCV isolates found in China share high sequence identity (>98%) and have more than 97% sequence identity with TYLCV-IL[IL:Reo] (X15656). Phylogenetic relationship analysis reveals that although with little genetic variability, they can form two groups and all the TYLCV isolates in China belong to the group I. An infectious clone of TYLCV-[CN:SH2] (AM282874) was constructed and agro-inoculated into Nicotiana benthamiana, N. tabacum Samsun, N. glutinosa, Solanum lycopersicum, Petunia hybrida, Cucumis sativus, Gossypium hirsutum, S. melongena, and Capsicum annuum. TYLCV-[CN:SH2] can induce severe leaf curling and stunting symptoms in these plants except C. sativus, G. hirsutum, S. melongena and C. annuum. We verified that TYLCV can trans-replicate tomato yellow leaf curl China virus DNA-β in N. benthamiana and S. lycopersicum and induced more severe symptoms with distortion and yellow vein.     

Genetic diversity of tomato-infecting Tomato yellow leaf curl virus (TYLCV) isolates in Korea

Epidemic outbreaks of Tomato yellow leaf curl virus (TYLCV) diseases occurred in greenhouse grown tomato (Solanum lycopersicum) plants of Busan (TYLCV-Bus), Boseong (TYLCV-Bos), Hwaseong (TYLCV-Hwas), Jeju Island (TYLCV-Jeju), and Nonsan (TYLCV-Nons) in Korea during 2008-2009. Tomato disease by TYLCV has never occurred in Korea before. We synthesized the full-length genomes of each TYLCV isolate from the tomato plants collected at each area and determined their nucleotides (nt) sequences and deduced the amino acids of six open reading frames in the genomes. TYLCV-Bus and -Bos genomes shared higher nt identities with four Japanese isolates -Ng, -Omu, -Mis, and -Miy. On the other hand, TYLCV-Hwas, -Jeju, and -Nons genomes shared higher nt identities with five Chinese isolates TYLCV-AH1, -ZJ3, -ZJHZ12, -SH2, -Sh10, and two Japanese isolates -Han and -Tosa. On the basis of a neighbor-joining tree, five Korean TYLCV isolates were separated into three clades. TYLCV-Bus and -Bos formed the first clade, clustering with four Japanese isolates TYLCV-Mis, -Omu, -Ng, and -Miy. TYLCV-Jeju and -Nons formed the second clade, clustering with two Chinese isolates -ZJHZ212 and -Sh10. TYLCV-Hwas was clustered with two Japanese isolates -Han and -Tosa and three Chinese isolates -AH1, -ZJ3, and -SH2. Two fragments that had a potentially recombinant origin were identified using the RDP, GENECONV, BootScan, MaxChi, Chimaera, SiScan, and 3Seq methods implemented in RDP3.41. On the basis of RDP analysis, all TYLCV isolates could originated from the interspecies recombination between TYLCV-Mld[PT] isolated from Portugal as a major parent and TYLCTHV-MM isolated from Myanmar as a minor parent.     

Real-time PCR for the quantitation of Tomato yellow leaf curl Sardinia virus in tomato plants and in Bemisia tabaci

Tomato yellow leaf curl Sardinia virus (TYLCSV) (Geminiviridae) is an important pathogen severely affecting tomato production in the Mediterranean basin. Although diagnostic protocols are available for its detection in plants and its vector Bemisia tabaci (Gennadius), suitable tools for estimating and comparing viral loads in plant and insect tissues are needed. In this paper, real-time PCR methods are described for quantitation of TYLCSV in both tomato plant and whitefly extracts. The DNA extraction method was optimised on TYLCSV-infected tomato tissue. The amount of virus was determined using specific primers and probe and standardised to the amount of DNA present in each sample, using selected endogenous tomato or Bemisia genes as internal references. The distribution of TYLCSV was relatively quantified within the four uppermost leaves of plants. An absolute estimation of the amount of TYLCSV in the first leaf below the apex was obtained. The kinetics of virus retention within different batches of viruliferous whiteflies was also analysed. The real-time PCR was 2200-fold more sensitive than membrane hybridisation, allowing detection of as few as 10 viral copies in a sample. These methods are potentially suitable for several applications, such as plant breeding for resistance, analysis of virus replication, and virus-vector interaction studies.     

Pathogenicity and insect transmission of a begomovirus complex between tomato yellow leaf curl virus and Ageratum yellow vein betasatellite

Tomato yellow leaf curl virus (TYLCV) and Ageratum yellow vein betasatellite (AYVB) are members of the genus Begomovirus (family Geminiviridae). TYLCV and AYVB have been found in Japan over the last 15 years, and are associated with tomato leaf curl and the tomato yellow leaf curl diseases (TYLCD). AYVB is also associated with some monopartite begomoviruses. We have cloned both TYLCV and AYVB and demonstrated that TYLCV can trans-replicate with AYVB in Nicotiana benthamiana and tomato plants. A mixed infection of TYLCV and AYVB induced more severe symptoms of upward leaf curl, stunting, vein thickening, and swelling compared with TYLCV infection alone. The symptoms induced by infection of AYVB included a rise in abnormal cell proliferation, and pigmentation around leaf vein tissues. This is the first study to show that a complex of TYLCV and AYVB can be transmitted by vector insects among tomato plants. These results indicate that TYLCV possesses the potential to induce severe TYLCD by associating with AYVB.     

Detection of tomato yellow leaf curl virus by loop-mediated isothermal amplification reaction

The genomic DNA molecule of tomato yellow leaf curl virus (TYLCV), a whitefly-transmitted geminivirus, was amplified from total DNA extracts of TYLCV-infected tomato (Lycopersicon esculentum) by the use of loop-mediated isothermal amplification (LAMP). The procedure was also used to amplify TYLCV DNA from total DNA extracts of individual whiteflies (Bemisia tabaci) that had fed on TYLCV-infected plants. One of the characteristics of the LAMP method is its ability to synthesize an extremely large amount of DNA. Accordingly, a large amount of by-product, pyrophosphate ion, is produced yielding a white precipitate of magnesium pyrophosphate in the reaction mixture. The presence or absence of this white precipitate allows easy detection of amplification of TYLCV genomic DNA without gel electrophoresis.     

Development of a real-time PCR for Tomato yellow leaf curl Sardinia virus

Recently, tomato yellow leaf curl disease has become important for the tomato grown both in greenhouse and field conditions in Tunisia. Here, we describe a rapid, specific, reliable, and sensitive real-time PCR, based on TaqMan chemistry, for Tomato yellow leaf curl Sardinia virus (TYLCSV). This method proved suitable for the detection and quantification of this virus in tomato, pepper and bean plants. It detected the virus even in the samples that were negative by conventional assays.     

Interaction of tomato yellow leaf curl virus with diverse betasatellites enhances symptom severity

The complete nucleotide sequence was determined for a begomovirus isolated from tomato exhibiting leaf curling and yellowing symptoms in Tochigi Prefecture in Japan. The genome organization of this virus was similar to those of other Old World monopartite begomoviruses. Neither a DNA betasatellite nor a DNA-B component was detected. It had the highest total nucleotide sequence identity (99%) with tomato yellow leaf curl virus-Israel[Japan:Tosa:2005] (TYLCV-IL[JR:Tos:05]) and TYLCV-Israel[Japan:Haruno:2005] (TYLCV-IL[JR:Han:05]). Its coat protein V1 also showed an identical amino acid sequence with those of TYLCV-IL[JR:Tos:05] and TYLCV-IL[JR:Han:05]. Thus, the begomovirus was determined to be an isolate of TYLCV-IL designated as TYLCV-Israel[Japan:Tochigi:2007] (TYLCV-IL[JR:Toc:07]). We investigated the interaction of TYLCV-IL[JR:Toc:07] with two known satellites associated with tomato yellow dwarf disease in Japan, tobacco leaf curl Japan betasatellite [Japan:Ibaraki:2006] and honeysuckle yellow vein mosaic betasatellite [Japan:Nara:2006], as well as with tomato leaf curl Philippines betasatellite [Philippines:Laguna1:2008], in tomato and Nicotiana benthamiana plants. TYLCV-IL[JR:Toc:07] trans-replicated these betasatellites, inducing more severe tomato yellow leaf curl disease-related symptoms than TYLCV-IL[JR:Toc:07] alone.     

Transreplication of a Tomato yellow leaf curl Thailand virus DNA-B and replication of a DNAbeta component by Tomato leaf curl Vietnam virus and Tomato yellow leaf curl Vietnam virus

The genomes of two tomato-infecting begomoviruses from Vietnam were cloned and sequenced. A new variant of Tomato leaf curl Vietnam virus (ToLCVV) consisting of a DNA-A component and associated with a DNAbeta molecule as well as an additional begomovirus tentatively named Tomato yellow leaf curl Vietnam virus (TYLCVV) consisting also of a DNA-A component were identified. To verify if monopartite viruses occurring in Vietnam and Thailand are able to transreplicate the DNA-B component of Tomato yellow leaf curl Thailand virus-[Asian Institute of Technology] (TYLCTHV-[AIT]) infectivity assays were performed via agroinoculation and mechanically. As result, the DNA-B component of TYLCTHV-[AIT] was transreplicated by different DNA-A components of viruses from Vietnam and Thailand in Nicotiana benthamiana and Solanum lycopersicum. Moreover, the TYLCTHV-[AIT] DNA-B component facilitated the mechanical transmission of monopartite viruses by rub-inoculation as well as by particle bombardment in N. benthamiana and tomato plants. Finally, defective DNAs ranging from 735 to 1457 nucleotides were generated in N. benthamiana from those combinations containing TYLCTHV-[AIT] DNA-B component.     

Application of Phi29 DNA polymerase in identification and full-length clone inoculation of tomato yellow leaf curl Thailand virus and tobacco leaf curl Thailand virus

Summary. Tomato plants grown in greenhouses in Thailand developed typical symptoms of a tomato yellow leaf curl Thailand virus (TYLCTHV) infection. After confirmation by ELISA, a Phi29 DNA polymerase approach was chosen for further molecular analysis of TYLCTHV. Total DNA purified from infected tomato leaves was subjected to rolling-circle amplification (RCA) of DNA-A and DNA-B of TYLCVTHV. In addition, a new monopartite geminivirus with a putative recombinant background was identified by RCA and tentatively named tobacco leaf curl Thailand virus (TbLCTHV). To confirm the composition of both geminiviruses, full-length clones were established and used for inoculation of Nicotiana benthamiana by particle bombardment or agroinfection. When TYLCTHV DNA-A and DNA-B were applied together by particle bombardment or agroinfection, severe stunting, yellowing, and leaf curling were observed. Whereas TYLCTHV DNA-A and TbLCTHV revealed no infection after'particle bombardment, similar symptoms in N. benthamiana, like leaf upward curling and yellowing were observed following agroinfection. DNA components of TYLCTHV DNA-A and DNA-B were excised from their respective plasmids, ligated, and amplified by Phi29 DNA polymerase. The ability of viral concatamere inoculation was evaluated in particle co-bombardment experiments on N. benthamiana. Thus, particle bombardment of RCA-derived multimeric products proved to be at least as effective as inoculation with a partial repeat construct and tenfold as effective as inoculation with excised unit-lengths of DNA-A and DNA-B of TYLCVTHV when using each DNA component in an amount of 5 ng.     

Characterization and distribution of tomato yellow margin leaf curl virus, a begomovirus from Venezuela

A begomovirus causing mottling and leaf deformation in tomato from the State of Mérida was cloned and sequenced. The virus has a bipartite genome comprised of a DNA-A (2,572 nucleotides) and a DNA-B (2,543 nucleotides) with a genome organization typical of New World begomoviruses. Both components share a common region of 115 nucleotides with 98 % sequence identity. Phylogenetic analysis indicated that while no virus sequences were closely related, the A component was distantly related to those of two other tomato-infecting viruses, tomato leaf deformation virus and Merremia mosaic virus; and the DNA-B, to those of pepper huasteco yellow vein virus and Rhynchosia golden mosaic Yucatan virus. The DNA-A and DNA-B sequences were submitted to GenBank (accession no. AY508993 and AY508994, respectively) and later accepted by the International Committee on Taxonomy of Viruses as the genome of a member of a unique virus species with the name Tomato yellow margin leaf curl virus (TYMLCV). Tomato (Solanum lycopersicum L. ‘Fl. Lanai’) plants inoculated with cloned TYMLCV DNA-A and DNA-B became systemically infected and showed chlorotic margins and leaf curling. The distribution of TYMLCV in tomato-producing states in Venezuela was determined by nucleic acid spot hybridization analysis of 334 tomato leaf samples collected from ten states using a TYMLCV-specific probe and confirmed by PCR and sequencing of the PCR fragment. TYMLCV was detected in samples from the states of Aragua, Guárico, and Mérida, suggesting that TYMLCV is widely distributed in Venezuela.     

Infectivity of Euphorbia leaf curl virus and interaction with Tomato yellow leaf curl China betasatellite

To investigate the infectivity of Euphorbia leaf curl virus (EuLCV), an infectious clone was constructed and tested by agroinoculation and whitefly inoculation. EuLCV infected Nicotiana benthamiana, N. glutinosa, Solanum lycopersicum, Petunia hybrida efficiently upon agroinoculation and induced leaf curling, vein swelling and stunting in these plants but no symptoms in N. tabacum. Co-inoculation of EuLCV with a betasatellite DNA from an unrelated begomovirus enhanced symptoms in N. benthamiana, N. glutinosa, N. tabacum, S. lycopersicum and P. hybrida plants but had no effect on the accumulation of EuLCV DNA. Euphorbia pulcherrima plants were only infectable by insect transmission from agro-infected P. hybrida as a source. This is the first report about a monopartite begomovirus that has been reintroduced into a plant of the genus Euphorbia.     

A natural recombinant between the geminiviruses Tomato yellow leaf curl Sardinia virus and Tomato yellow leaf curl virus exhibits a novel pathogenic phenotype and is becoming prevalent in Spanish populations

We have examined the consequences of cleaving the fusion glycoprotein (F) of human respiratory syncytial virus (HRSV) at two distinct furin-recognition sites. Purified anchorless F is a mixture of unaggregated cone-shaped molecules and rosettes of lollipop-shaped spikes. The unaggregated molecules contain a proportion of uncleaved F0 and an intermediate, F(delta1-109), cleaved only at site I, residues 106-109. Inhibition of cleavage at site I, by two amino acid changes (R108N/R109N), reduces the proportion of aggregated molecules with a concomitant increase in the amount of unprocessed F0. Inhibition of cleavage at site II, residues 131-136, by deletion of four amino acids (delta131-134), abrogates aggregation of anchorless F and all molecules are seen as individual cone-shaped rods. In vitro cleavage of anchorless F, or mutant delta131-134, with trypsin at 4, 20, or 37 degrees C, under conditions in which cleavage at site II is complete in all molecules, leads to their aggregation in rosettes of lollipop-shaped spikes. Thus, cleavage at site II is required for the structural changes in anchorless F that lead to changes in shape and to aggregation. The segment between sites I and II, residues 110-136, is not associated with anchorless F in the supernatant of infected cell cultures, indicating that it is released from the processed protein when cleavage at sites I and II is completed.     

The begomoviruses Honeysuckle yellow vein mosaic virus and Tobacco leaf curl Japan virus with DNAbeta satellites cause yellow dwarf disease of tomato

The complete nucleotide sequences of two begomoviruses (Nara virus-1 and Nara virus-2), a satellite DNA (DNAbeta-Nara) and defective DNAs were obtained from honeysuckle (Lonicera japonica) showing characteristic yellow vein mosaic symptoms in Nara Prefecture, Japan. One begomovirus (Ibaraki virus) and a satellite DNA (DNAbeta-Ibaraki) was isolated and cloned from honeysuckle plants exhibited typical yellowing of veins and small elliptical shaped enations along veins on the under side of the leaves in Ibaraki Prefecture, Japan. The genome organization of the three viruses is the same as those of other Old World monopartite begomoviruses. Nara virus-1 had overall nucleotide sequence identity with Nara virus-2 of 94% and Ibaraki virus of 90%. DNAbeta-Nara had overall nucleotide sequence identity with DNAbeta-Ibaraki of 83%. Comparison of the nucleotide sequences with other begomoviruses revealed that Nara virus-1 and Nara virus-2 are strains of Honeysuckle yellow vein mosaic virus (HYVMV), hence named as HYVMV-Nara1 and HYVMV-Nara2, whereas Ibaraki virus was a strain of Tobacco leaf curl Japan virus (TbLCJV), designated as TbLCJV-Hs[Iba]. HYVMV-Nara1 and HYVMV-Nara2 have hybrid genomes, which are likely to have formed recombination between HYVMV and TbLCJV. TbLCJV-Hs[Iba] or HYVMV-Nara2 could infect and cause yellowing, leaf crinkling and stunting symptoms when partial tandem dimeric constructs were agroinoculated on tomato plants. However, in the presence of DNAbeta, both TbLCJV-Hs[Iba] or HYVMV-Nara2 produced more severe stunting symptoms in tomato plants. Therefore, these viruses along with their satellites are causal agents of tomato yellow dwarf disease in Japan, and honeysuckle acts as a potential reservoir host. Previously available evidence indicated that DNAbeta elements do not contain iteron sequences of their helper viruses; hence this is the first evidence that DNAbeta satellites have the iteron of their helper virus.