Croton Yellow Vein Mosaic Virus Research Articles

Distinct tomato yellow leaf curl Chuxiong virus isolated from whiteflies and plants in China and its symptom determinant and suppressor of post-transcriptional gene silencing

A begomovirus isolated from whiteflies (Bemisia tabaci) and tomato, sweet potato in China was found to be representative of a distinct begomovirus species, for which the name tomato yellow leaf curl Chuxiong virus (TYLCCxV) is proposed. The results of genomic identification and sequence comparison showed that TYLCCxV shares the highest complete nucleotide sequence identity (88.3%) with croton yellow vein mosaic virus (CroYVMV), and may have originated from the recombination between synedrella leaf curl virus (SyLCV) and squash leaf curl Yunnan virus (SLCuYV). Agrobacterium-mediated inoculation showed that TYLCCxV is highly infectious for a range of plant species, producing upward leaf curling, leaf crumpling, chlorosis, distortion, and stunt symptoms in Solanum lycopersicum plants. The results of Southern blot indicated that TYLCCxV is capable of efficiently replicating two heterologous betasatellites. The inoculation of PVX::C4 on Nicotiana benthamiana induced upward leaf curling and stem elongation symptoms, suggesting that TYLCCxV C4 functions as a symptom determinant. TYLCCxV V2 is an important virulence factor that induces downward leaf curling symptoms, elicits systemic necrosis, and suppresses local and systemic GFP silencing in co-agroinfiltrated N. benthamiana and transgenic 16c plants. Considering the multifunctional virulence proteins V2 and C4, the possibility of TYLCCxV causing devastating epidemics on tomato in China is discussed.

Mixed infection of a new begomovirus, Jatropha leaf curl Guntur virus and recombinant/chimeric jatropha leaf curl Gujarat virus in Jatropha gossypiifolia.

The Jatropha gossypiifolia plant showing the severe leaf curl symptoms grown in the borders of chilli fields in Guntur, Andhra Pradesh, India was collected. The infection of begomovirus was detected using the degenerate primers followed by rolling circle amplification (RCA). The RCA products digested with KpnI and EcoRI showing the unit length of the begomovirus genome were cloned in pUC19 and sequenced to obtain the complete begomoviral genome. The sequence information of DNA-A of the two clones GuWC10 contained 2794 nt (MZ217773) and an incomplete genome GuWC3 with 2337 nt (MZ217772). The BLAST analysis of GuWC3 and GuWC10 sequence showed 85·57% identity with jatropha leaf curl Gujarat virus (JLCGV) and 82·68% identity with croton yellow vein mosaic virus (CroYVMV), respectively. The sequence analysis also showed that the GuWC10 clone had a 177 bp recombinant/chimeric sequence of JLCGV while the other region containing 2611 bp showed 92·63% identity with papaya leaf curl virus (PaLCuV/PK). However, the global alignment of the GuWC10 sequence showed a maximum of 80·60% identity with croton yellow vein virus (CroYVV) (FN645902), CroYVMV (JN817516) and PaLCuV/PK (KY978407). The second clone GuWC3 although shorter in length had recombinant sequences of JLCGV, jatropha leaf curl virus (JLCuV/ND) and okra enation leaf curl virus (OELCuV). The nucleotide sequence identity among the GuWC10 and GuWC3 was 71·9%. The phylogenetic analysis placed both the viral strains in the same clade located between PaLCuV/PK and JLCuV clades. According to the ICTV species demarcation criteria of 91% DNA-A sequence identity, the present isolate was considered as a new species of begomovirus and the name Jatropha leaf curl Guntur virus was proposed. This is the first report of a new begomovirus species infecting J. gossypiifolia and the study also reports a mixed infection of Jatropha leaf curl Guntur virus with a recombinant/chimeric JLCGV in the host J. gossypiifolia. Present study suggests the role of weed Jatropha in harbouring begomoviruses and probable source for viral recombination.

Identification and Functional Analysis of Four RNA Silencing Suppressors in Begomovirus Croton Yellow Vein Mosaic Virus.

Croton yellow vein mosaic virus (CYVMV), a species in the genus Begomovirus, is a prolific monopartite begomovirus in the Indian sub-continent. CYVMV infects multiple crop plants to cause leaf curl disease. Plants have developed host RNA silencing mechanisms to defend the threat of viruses, including CYVMV. We characterized four RNA silencing suppressors, namely, V2, C2, and C4 encoded by CYVMV and betasatellite-encoded C1 protein (βC1) encoded by the cognate betasatellite, croton yellow vein betasatellite (CroYVMB). Their silencing suppressor functions were verified by the ability of restoring the β-glucuronidase (GUS) activity suppressed by RNA silencing. We showed here for the first time that V2 was capable of self-interacting, as well as interacting with the V1 protein, and could be translocalized to the plasmodesmata in the presence of CYVMV. The knockout of either V2 or V1 impaired the intercellular mobility of CYVMV, indicating their novel coordinated roles in the cell-to-cell movement of the virus. As pathogenicity determinants, each of V2, C2, and C4 could induce typical leaf curl symptoms in Nicotiana benthamiana plants even under transient expression. Interestingly, the transcripts and proteins of all four suppressors could be detected in the systemically infected leaves with no correlation to symptom induction. Overall, our work identifies four silencing suppressors encoded by CYVMV and its cognate betasatellite and reveals their subcellular localizations, interaction behavior, and roles in symptom induction and intercellular virus movement.

A new variant of Croton yellow vein mosaic virus naturally infecting wild sunflower in India.

During 2015-2016, wild sunflower (Verbesina encelioides) was observed with bright yellow vein mosaic symptims in Noida, Uttar Pradesh, India. The initial analysis by PCR with a pair of coat protein based primers revealed the association of a begomovirus. Further, the virus was identified by rolling circle amplification and cloning of the complete genome. The DNA-A (2671 nucleotides, MH359168) with the genome organisation typical of the Old World begomovirus shared 98% sequence identity with that of Croton yellow vein mosaic virus (CYVMV) reported from India. The betasatellite (MH359169) associated with the disease shared 92% sequence identity with papaya leaf curl betasatellite (KY825245). The results showed that the wild sunflower is a new alternate host of CYVMV. The study also revealed a natural association of heterologous betasatellite with CYVMV in wild sunflower exhibiting yellow vein mosaic symptoms.

Natural infection of croton yellow vein mosaic virus and its cognate betasatellite in germplasm of different Crambe spp in India

Crambe is an important crop grown worldwide for industrial oil and seed meal. Besides the fungal and bacterial diseases, the crop is reported to be infected by tobacco mosaic virus, beet western yellows virus and turnip mosaic virus under experimental condition. Till now, there was no report of natural infection of any begomovirus in this crop. In the present study, a leaf curl disease was observed in germplasm accessions of three species of Crambe (C. abyssinica, C. glabrata and C. hispanica). Based on the symptoms and presence of whitefly population in the field, begomovirus infection was suspected. Molecular characterization through RCA approach, indicated presence of croton yellow vein mosaic virus (CYVMV, KJ747958) and croton yellow vein mosaic betasatellite (CroYVMB, KM229762). Co-agroinoculation of partial dimeric construct of CYVMV with complete dimeric construct of CroYVMB, produced typical leaf curl symptoms in C. abyssinica, whereas, agroinoculation of partial dimeric construct of CYVMV alone could not produce symptoms in the same plant. In contrast, the CYVMV construct alone could produce symptom in Nicotiana benthamiana, a model host for plant virus studies. In N. benthamiana co-inoculation of CroYVMV with CYVMV construct develop more severe symptoms. However, neither the CYVMV construct alone nor the co-inoculation with CroYVMB produce any symptom in Arabidopsis thaliana even with different methods of inoculation. Inoculated Arabidopsis thaliana also did not yield any amplification of the virus as assessed through PCR and rolling circle amplification (RCA). Thus it confirmed that for successful infection in crambe, CYVMV requires betasatellite, while in N. benthamiana, it does not require betasatellite for symptom induction and in Arabidopsis thaliana CYVMV alone or in presence of betasatellite did not replicate and produce any symptom. This study constitutes the first confirmed record of natural infection of a begomovirus in crambe and further confirmed that cognate betasatellite of CYVMV has differential role in infectivity in different hosts.

The efficacy of antisense-based construct for inducing resistance against Croton yellow vein mosaic virus in Nicotiana tabacum.

Begomoviruses have increased pathogenicity because of their adaptation to a wide host range; consequently, these viruses cause a major loss to agroeconomic crops worldwide. In this study, we designed a gene construct representing an antisense coat protein gene. We also analyzed the efficacy of the induced resistance against Croton yellow vein mosaic virus (CrYVMV) affecting papaya in Nicotiana tabacum plants. Positive control plants developed typical leaf curl symptoms, whereas transgenic plants were symptomless. Moreover, the key component (i.e., short interfering RNA) of the antisense pathway was upregulated in transgenic plants. This finding demonstrates the activation of the gene silencing mechanism in transgenic plants. Thus, these results confirm that our construct is functional and effectively induces transient resistance against CrYVMV infections.

First Report of Tomato leaf curl New Delhi virus on Calotropis procera, a Weed as Potential Reservoir Begomovirus Host in Pakistan

HomePlant DiseaseVol. 101, No. 6First Report of Tomato leaf curl New Delhi virus on Calotropis procera, a Weed as Potential Reservoir Begomovirus Host in Pakistan Previous DISEASE NOTES OPENOpen Access licenseFirst Report of Tomato leaf curl New Delhi virus on Calotropis procera, a Weed as Potential Reservoir Begomovirus Host in PakistanS. S. Zaidi, S. Shakir, H. J. Malik, M. Farooq, I. Amin, and S. MansoorS. S. Zaidihttp://orcid.org/0000-0002-2976-4624Search for more papers by this author, S. ShakirSearch for more papers by this author, H. J. MalikSearch for more papers by this author, M. FarooqSearch for more papers by this author, I. AminSearch for more papers by this author, and S. MansoorSearch for more papers by this authorAffiliationsAuthors and Affiliations S. S. Zaidi , Agricultural Biotechnology Division, National Institute for Biotechnology and Genetic Engineering (NIBGE), Faisalabad, Pakistan S. Shakir , Virology Laboratory, Center for Agricultural Biochemistry and Biotechnology (CABB), University of Agriculture, Faisalabad, Pakistan, and Agricultural Biotechnology Division, National Institute for Biotechnology and Genetic Engineering (NIBGE), Faisalabad, Pakistan H. J. Malik M. Farooq I. Amin S. Mansoor , Agricultural Biotechnology Division, National Institute for Biotechnology and Genetic Engineering (NIBGE), Faisalabad, Pakistan. Published Online:19 Apr 2017https://doi.org/10.1094/PDIS-10-16-1539-PDNAboutSections ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InRedditEmailWechat In 2015–16, during field surveys of Sindh and Punjab provinces of Pakistan to observe virus symptoms on cultivated crops and nearby weeds, symptoms of leaf yellowing and yellow mosaic were observed on leaves of Calotropis procera growing around the cultivated fields. C. procera, also known as rubber bush or aak, is a common weed in Pakistan. To detect the possible presence of begomoviruses, five symptomatic plants and one asymptomatic plant were collected from Punjab and Sindh, and total DNA was extracted. Southern blot hybridization was performed using a DIG labeled probe to the Tomato leaf curl New Delhi virus (ToLCNDV) C1 gene. A strong hybridization signal was observed in symptomatic plants, while there was no signal in asymptomatic plants collected from the same fields. To further confirm the association of ToLCNDV, rolling circle amplification (RCA) was performed to enrich for the circular DNA molecules of the begomovirus genome, using phi29 DNA polymerase (Thermo Fisher Scientific, Waltham, MA). The RCA product was purified and sequenced by the Illumina MiSeq sequencing system. The Illumina NeoPrep automation system (Illumina, San Diego, CA) was used with the Illumina TruSeq Nano DNA Library Prep Kit (NP-101-1001). After sequencing, adapters were trimmed by the MiSeq Reporter Software and sequencing data were analyzed by the CLC Genomics Workbench 7.5 (https://www.qiagenbioinformatics.com). De novo assemblies were prepared following the standard quality parameters (quality score: 0.001 and Phred score: 30). The NCBI BLASTn results of assembled de novo reads confirmed the presence of ToLCNDV DNA-A and ToLCNDV DNA-B (sequence available in the database with the accession number KX710158 and KX710159, respectively). The species demarcation of ToLCNDV was also confirmed using the Sequence Demarcation Tool (SDT) program (http://web.cbio.uct.ac.za/∼brejnev/), following the revised taxonomic criteria of begomoviruses (Brown et al. 2015). The nBLAST analysis of ToLCNDV DNA-A reported in this study indicated that it was 98% identical to the ToLCNDV DNA-A reported previously in chili from Khanewal, Pakistan (DQ116880); while the nBLAST analysis of ToLCNDV DNA-B reported in this study indicated that it was 95% identical to the ToLCNDV DNA-B reported previously in Cucurbita pepo from Rawalpindi, Pakistan (KT948073). No evidence for the presence of alphasatellites or betasatellite either in the Illumina data or by PCR using universal primers for amplification of alphasatellites or betasatellites was found. Whitefly transmitted ToLCNDV (Gilbertson et al. 2015) infects several important crops like tomato, potato, cucurbits (Zaidi et al. 2016a), and cotton (Zaidi et al. 2016b). The occurrence of ToLCNDV on C. procera indicates the increasing host range of ToLCNDV, which is serious threat for other economically important crops. C. procera has been reported as a potential host for Papaya leaf curl virus (JQ407224; formerly known as Croton yellow vein mosaic virus) and croton yellow vein mosaic betasatellite (HQ631430) in India (Prajapat et al. 2012). C. procera is a common weed and may therefore serve as an alternative virus reservoir, with an important role in the epidemiology of the disease. To our knowledge, this is the first report of ToLCNDV infecting C. procera in Pakistan.

Genetic diversity of Croton Yellow Vein Mosaic Virus and betasatellites associated with yellow vein mosaic disease of Sida acuta in India

Genetic diversity of Croton Yellow Vein Mosaic Virus and betasatellites associated with yellow vein mosaic disease of Sida acuta in India

Agroinfection of tobacco by croton yellow vein mosaic virus and designing of a replicon vector for expression of foreign gene in plant.

Croton yellow vein mosaic virus (CYVMV, genus Begomovirus family Geminiviridae) is a proliferating begomovirus in the Indian sub-continent. The infectious constructs in binary vector was developed against the CYVMV genome and its associated betasatellite. Agroinoculation of the genomic construct of CYVMV produced leaf curl symptoms alone in three species of tobacco, Nicotiana tabacum, N. benthamiana and N. glutinosa. Co-inoculation of betasatellite enhanced the severity of the disease and reduced the incubation time. Based on the infectious clone, a replicon vector pCro, with only the ability to replicate inside the plant was developed. In pCro vector, CP and V2 ORFs from genome of CYVMV was deleted, which resulted localised replication of the molecule with no visible symptoms. Besides the partial CYVMV genome, pCro alsohas a cassette containing a double 35S promoter, multiple cloning sites and a NOS terminator to overexpress any foreign protein in plant. Episomal release of the replicon from the binary vector backbone after agroinoculation was detected by PCR. A GFP gene was cloned in pCro vector (pCro-GFP) and agroinoculated to N. tabacum resulted in localized expression of GFP at 5dpi. The CYVMV replicon vector will be a useful tool for studying functional genomics, vaccine expression and gene silencingin plant.

First Report of a Croton yellow vein mosaic virus (CYVMV) Associated with Tomato Leaf Curl Disease in India

AbstractLeaf curl disease symptoms were observed in tomato crop grown in a tomato field at Matera district of Bahraich, India, in March 2013 with an 85% disease incidence. The infected plants exhibited leaf curl symptoms accompanied with puckering, vein swelling and stunting of the whole plant. PCR carried out with begomovirus coat protein gene and DNA beta‐specific primer sets resulted in positive amplification of ~775 bp and 1.35 kbp, respectively, with all symptom‐bearing plant samples. BLASTn and phylogenetic analyses of CP gene sequences showed highest and close relationship with Croton yellow vein mosaic virus (CYVMV) isolates, while the phylogenetic study of betasatellite sequence showed distinct relationships with other begomovirus associated betasatellites reported from India and abroad. This is a first report of a CYVMV associated with tomato leaf curl disease in India.

Molecular characterization of a new species of begomovirus associated with yellow vein mosaic of bhendi (Okra) in Bhubhaneswar, India

Yellow vein mosaic disease of okra is major constraint for the production of okra in India. It is caused by whitefly transmitted begomoviruses and results in severe economic losses of the crop. In the present study a new begomovirus (OYBHU isolate) infecting okra showing yellow vein symptoms from Bhubhaneswar, India was characterized. The complete genome sequence (homologous to DNA-A) was determined and it comprised 2757 nucleotides. Attempts to amplify DNA- B were not successful. However, the association of a betasatellite suggests that the isolate is monopartite and typical of the begomoviruses infecting malvaceous hosts in the Old World. The genome organization is characteristic of begomoviruses, encoding seven ORFs with two ORFs [AV1 (CP) and AV2] in virion sense strand and five ORFs (AC1-AC5) in complementary strand. Comparisons of the virus genome sequence with other known begomoviruses suggests that OYBHU virus isolate is distinct, with Croton yellow vein mosaic virus being its nearest relative, sharing the 81.4 to 86.1 % sequence identity. This is further supported by phylogenetic analyses with clustering of BYVBV on a well supported branch separated from all other begomoviruses. The name Bhendi yellow vein Bhubhaneswar virus (BYVBV) is proposed for virus isolate OYBHU with the additional descriptor [India: Bhubhaneswar:OYBHU:06]. Recombination analyses with BYVBV suggest that it is a recombinant and may have originated by the exchange of genomic segments between the begomoviruses Croton yellow vein mosaic virus, Bhendi yellow vein mosaic virus, Cotton leaf curl multan virus and Mesta yellow vein mosaic virus.

A Leaf Curl Disease in Germplasm of Rapeseed‐Mustard in India: Molecular Evidence of a Weed‐Infecting Begomovirus–Betasatellite Complex Emerging in a New Crop

AbstractEvaluation of 130 accessions of rapeseed‐mustard germplasm grown at the National Bureau of Plant Genetic Resources, New Delhi, India during the winter season (2011–2012) revealed the occurrence of a leaf curl disease in seven accessions. The occurrence of the disease was observed in another 62 of 525 accessions evaluated during 2012–2013. The association of a monopartite begomovirus and betasatellite was established with the symptomatic plants by whitefly transmission and PCR amplification. The complete nucleotide sequences of the begomovirus (JX270684, 2745 nucleotides), obtained by rolling circle amplification, showed the highest sequence identity (98.1%) with the weed‐infecting begomovirus, Croton yellow vein mosaic virus. Analysis of recombination indicated the probable occurrence of many overlapping inter‐ and intraspecific recombination events. The sequence of betasatellite (JX270685, 1355 nucleotides) showed the highest sequence identity (95.7%) with Croton yellow vein mosaic betasatellite. Begomoviruses were not previously known to naturally infect rapeseed‐mustard. This is the first report of the emergence of a weed‐infecting begomovirus–betasatellite complex in rapeseed‐mustard germplasm in India and raises the concern on utilization of such susceptible germplasm in crop improvement programmes.

Host range and genetic diversity of croton yellow vein mosaic virus, a weed-infecting monopartite begomovirus causing leaf curl disease in tomato

Croton yellow vein mosaic virus (CYVMV) is a widely occurring begomovirus in Croton bonplandianum, a common weed in the Indian subcontinent. In this study, CYVMV (genus Begomovirus, family Geminiviridae) was transmitted by whiteflies (Bemisia tabaci) to as many as 35 plant species belonging to 11 families, including many vegetables, tobacco varieties, ornamentals and weeds. CYVMV produced bright yellow vein symptoms in croton, whereas in all the other host species, the virus produced leaf curl symptoms. CYVMV produced leaf curl in 13 tobacco species and 22 cultivars of Nicotiana tabacum and resembled tobacco leaf curl virus (TobLCV) in host reactions. However, CYVMV was distinguished from TobLCV in four differential hosts, Ageratum conyzoides, C. bonplandianum, Euphorbia geniculata and Sonchus bracyotis. The complete genome sequences of four isolates originating from northern, eastern and southern India revealed that a single species of DNA-A and a betasatellite, croton yellow vein mosaic betasatellite (CroYVMB) were associated with the yellow vein mosaic disease of croton. The sequence identity among the isolates of CYVMV DNA-A and CroYVMB occurring in diverse plant species was 91.8-97.9 % and 83.3-100 %, respectively. The CYVMV DNA-A and CroYVMB generated through rolling-circle amplification of the cloned DNAs produced typical symptoms of yellow vein mosaic and leaf curling in croton and tomato, respectively. The progeny virus from both the croton and tomato plants was transmitted successfully by B. tabaci. The present study establishes the etiology of yellow vein mosaic disease of C. bonplandianum and provides molecular evidence that a weed-infecting monopartite begomovirus causes leaf curl in tomato.

Complete nucleotide sequences of two begomoviruses infecting Madagascar periwinkle (Catharanthus roseus) from Pakistan

Though Catharanthus roseus (Madagascar periwinkle) is an ornamental plant, it is famous for its medicinal value. Its alkaloids are known for anti-cancerous properties, and this plant is studied mainly for its alkaloids. Here, this plant has been studied for its viral diseases. Complete DNA sequences of two begomoviruses infecting C. roseus originating from Pakistan were determined. The sequence of one begomovirus (clone KN4) shows the highest level of nucleotide sequence identity (86.5%) to an unpublished virus, chili leaf curl India virus (ChiLCIV), and then (84.4% identity) to papaya leaf curl virus (PaLCV), and thus represents a new species, for which the name "Catharanthus yellow mosaic virus" (CYMV) is proposed. The sequence of another begomovirus (clone KN6) shows the highest level of sequence identity (95.9% to 99%) to a newly reported virus from India, papaya leaf crumple virus (PaLCrV). Sequence analysis shows that KN4 and KN6 are recombinants of Pedilanthus leaf curl virus (PedLCV) and croton yellow vein mosaic virus (CrYVMV).

Biology and interactions of two distinct monopartite begomoviruses and betasatellites associated with radish leaf curl disease in India.

BackgroundEmerging whitefly transmitted begomoviruses are major pathogens of vegetable and fibre crops throughout the world, particularly in tropical and sub-tropical regions. Mutation, pseudorecombination and recombination are driving forces for the emergence and evolution of new crop-infecting begomoviruses. Leaf curl disease of field grown radish plants was noticed in Varanasi and Pataudi region of northern India. We have identified and characterized two distinct monopartite begomoviruses and associated beta satellite DNA causing leaf curl disease of radish (Raphanus sativus) in India.ResultsWe demonstrate that RaLCD is caused by a complex of two Old World begomoviruses and their associated betasatellites. Radish leaf curl virus-Varanasi is identified as a new recombinant species, Radish leaf curl virus (RaLCV) sharing maximum nucleotide identity of 87.7% with Tomato leaf curl Bangladesh virus-[Bangladesh:2] (Accession number AF188481) while the virus causing radish leaf curl disease-Pataudi is an isolate of Croton yellow vein mosaic virus-[India] (CYVMV-IN) (Accession number AJ507777) sharing 95.8% nucleotide identity. Further, RDP analysis revealed that the RaLCV has a hybrid genome, a putative recombinant between Euphorbia leaf curl virus and Papaya leaf curl virus. Cloned DNA of either RaLCV or CYVMV induced mild leaf curl symptoms in radish plants. However, when these clones (RaLCV or CYVMV) were individually co-inoculated with their associated cloned DNA betasatellite, symptom severity and viral DNA levels were increased in radish plants and induced typical RaLCD symptoms. To further extend these studies, we carried out an investigation of the interaction of these radish-infecting begomoviruses and their associated satellite, with two tomato infecting begomoviruses (Tomato leaf curl Gujarat virus and Tomato leaf curl New Delhi virus). Both of the tomato-infecting begomoviruses showed a contrasting and differential interaction with DNA satellites, not only in the capacity to interact with these molecules but also in the modulation of symptom phenotypes by the satellites.ConclusionThis is the first report and experimental demonstration of Koch's postulate for begomoviruses associated with radish leaf curl disease. Further observations also provide direct evidence of lateral movement of weed infecting begomovirus in the cultivated crops and the present study also suggests that the exchange of betasatellites with other begomoviruses would create a new disease complex posing a serious threat to crop production.

RNA Silencing Suppressor Encoded by Betasatellite DNA Associated With Croton Yellow Vein Mosaic Virus

Summary A novel single stranded DNA of approximately 1350 nucleotides in length has been identified associated with infected yellow vein mosaic croton termed as DNA- β, requires the begmovirus for replication, encapsidation, insect transmission and movement in plants. It exhibit the properties of a satellite molecule having approximately 80 nucleotides conserved region which has been suggested to be important in trans-replication of DNA- β by the begmoviruses rep, possibly containing cryptic rep binding sites. In the present study βC1genes encoded by DNA betasatellite were targeted in transient assay for stable transformation to develop resistance against geminivirus.

Chilli leaf curl Palampur virus is a distinct begomovirus species associated with a betasatellite

Capsicum frutescens (chilli) plants exhibiting leaf curl symptoms, collected from Palampur, Himachal Pradesh region of India, were found associated with a begomovirus and a betasatellite‐like molecule. Viral DNA was amplified, cloned and sequenced. The begomoviral genome and the betasatellite consisted of 2775 and 1376 nucleotides respectively. The virus appeared to be monopartite. The genome sequence had <87·9% identity with all other begomovirus sequences, below the threshold for species demarcation, suggesting that the isolate represents a distinct species for which the name Chilli leaf curl Palampur virus (ChiLCPaV) is proposed. Infectious clones consisting of partial tandem repeats of the viral genome (1·9‐mer) and the betasatellite (1·7‐mer) were constructed in the binary vector pCAMBIA‐1300 and agroinoculated to chilli and Nicotiana benthamiana. The viral clone did not produce leaf curl symptoms when inoculated alone but the plants were stunted compared to controls. Typical leaf curling and stunting symptoms were observed when the viral clone was inoculated with the betasatellite. In a phylogenetic analysis with other closely related begomoviruses, the new sequence grouped with an isolate of Papaya leaf curl virus‐[Pakistan:2008] from Rhynchosia capitata and Tomato leaf curl New Delhi virus‐[India:2006] from tomato. The betasatellite showed most identity (94%) with an isolate of Chilli leaf curl betasatellite from Pakistan. Using the rdp3 recombination detection program, a recombination event involving Croton yellow vein mosaic virus‐[India:2007] as a major parent and Tomato leaf curl Karnataka virus‐[India:2007] as a minor parent was detected in the ChiLCPaV sequence. No significant recombination event was detected in the betasatellite sequence.

Complete nucleotide sequence of Croton yellow vein mosaic virus and DNA-β associated with yellow vein mosaic disease of Jatropha gossypifolia in India

A severe yellow vein mosaic disease was noticed on several Jatropha gossypifolia plants growing nearby agriculture fields at Lucknow, India. Diseased plants exhibited yellow vein mosaic, leaf deformation, vein swelling and stunting. A population of whiteflies (Bemisia tabaci) was also noticed in the vicinities; therefore, begomovirus infection was suspected. To confirm begomovirus association, total DNA was isolated from symptomatic leaf samples and subjected to PCR using DNA-A, DNA-B and DNA-β-specific primers. DNA-A and DNA-β was successfully amplified but several attempts failed to amplify DNA-B indicating monopartite nature of the begomovirus. The sequence analysis of amplicons revealed the presence of 2757 nucleotides of DNA-A genome (EU727086) and 1315 nt of DNA-β molecule (EU604296). The sequence analysis of DNA-A (EU727086) revealed the highest 96% identities and closest relationship with Croton yellow vein mosaic virus (CYVMV, AJ507777) infecting Croton bonplandianum in India. The DNA-β (EU604296) showed the highest 96% sequence identity and closest phylogenetic relationship with CYVMV-associated DNA-β (AM410551) isolated from Croton sp. in Pakistan. Based on the highest sequence identities and closest phylogenetic relationships of the DNA-A genome and DNA-β molecule with respective sequences of various isolates of Croton yellow vein mosaic virus, the begomovirus associated with yellow vein mosaic disease of J. gossypifolia was identified as an isolate of Croton yellow vein mosaic virus.

Molecular characterization of two distinct monopartite begomoviruses infecting tomato in india

BackgroundTomato leaf curl viruses, which are the members of the genus Begomovirus, have emerged as devastating pathogens worldwide causing huge economic losses and threatening production of crops like cassava, cotton, grain legumes and vegetables. Even though the ToLCV isolates from Northern India have been shown to possess bipartite genome (designated as DNA A and DNA B), those from Australia, Taiwan and Southern India have a single genomic component (DNA A). We describe here the genetic diversity of two isolates of monopartite Tomato leaf curl virus infecting tomato in two extreme regions (North and South) of Indian subcontinent.ResultsThe rolling circle amplification (RCA) products obtained from symptomatic samples were digested, cloned and sequenced. The complete DNA sequence of two Tomato leaf curl virus isolates identified as ToLCV-CTM (India, New Delhi, 2005) and ToLCVK3/K5 (India, Kerala, 2008) are reported here. These isolates had the characteristic features of Begomovirus genome organization with six conserved open reading frames (ORFs). The ToLCV-K3 and ToLCV-K5 isolates may be the strains of the same virus since they show sequence homology of 97% over their entire genome. This, according to the guidelines established by the ICTV Geminiviridae Study-Group is higher than threshold (92%) for delineation of different viral variants and hence single, average value has been assigned for all their analyses presented here. The ToLCV-CTM and ToLCV-K3/K5 viruses were found to be monopartite, as neither DNA-B component nor betasatellite associated with begomovirus species, were detected. The complete nucleotide sequence of DNA-A genome of CTM exhibited highest sequence homology (88%) to Croton yellow vein mosaic virus (AJ507777), and of isolates K3/K5 (88.5%) to Tomato leaf curl Pakistan virus (DQ116884). This is less than the threshold value for demarcation of species in the genus Begomovirus.ConclusionK3/K5 and CTM are considered to be novel isolates of Tomato leaf curl virus. Sequence analyses and phylogenetic study indicate that these two ToLCV isolates might have evolved by recombination between viruses related to two or more viral ancestors. The existence of different ToLCV isolates having high genome diversity in India poses a threat to the tomato production in the Asian continent.

MOLECULAR CHARACTERIZATION OF BEGOMOVIRUSES ASSOCIATED WITH PAPAYA LEAF CURL DISEASE IN INDIA

For molecular characterization of papaya leaf curl causing begomoviruses, papaya samples showing symptoms of leaf curl, vein clearing, enations and reduced leaf size were collected from Kodur of Andhra Pradesh, Bangalore of Karnataka, Coimbatore of Tamil Nadu, Lucknow of Uttar Pradesh and Delhi States of India. Using primers specific to begomovirus DNA-A used the total DNA isolated from these samples for begomovirus infection. For molecular differentiation of papaya leaf curl isolates, cloning and sequencing of PCR amplified fragments was done. The complete DNA-A sequence of four papaya leaf curl isolates was characterized. Comparative sequence analysis with 21 other begomoviruses has indicated that the PaLC. Coimbatore isolate has 77.5% nucleotide identity with Ageratum enation virus (AEV), PaLCV.Kodur has 90.3% nucleotide identity with Croton yellow vein mosaic virus (CrYVMV), PaLC.UP13 isolate has 94.7% nucleotide identity with Papaya leaf curl virus (PaLCV) and PaLC.New Delhi isolate has 95.5% nucleotide identity with Tomato leaf curl New Delhi virus (ToLCNDV). Phylogenetic analysis and nucleotide comparison has indicated that in India, papaya is infected with more than three distinct begomoviruses. PaLC.Coimbatore isolate has nucleotide identity less than 89% with another known begomoviruses and named as Papaya leaf curl Coimbatore virus (PaLCCoV) and considered as a new begomovirus infecting papaya in India.