A New Begomovirus Species Causing Tomato Leaf Curl Disease in Varanasi, India

In November 2001, a leaf curl disease of tomato, manifested by yellowing of leaf lamina, upward leaf curling, leaf distortion, shrinking of leaf surface, and stunted plant growth was observed in tomato-growing areas in the Varanasi and Mirzapur districts of eastern Uttar Pradesh, India, which caused yield losses up to 100%. The causal agent was infective to tomato cv. Punjab Chuhara by whiteflies and grafting. Inoculated plants developed symptoms observed in naturally infected tomatoes. Viral DNA was isolated from artificially inoculated tomato plants using 1% CTAB (2) followed by a concentration of supercoiled DNA by alkaline denaturation (1). A geminivirus was confirmed by polymerase chain reaction using DNA-A degenerate primers (3), and a 550-bp amplified product was obtained from artificially and naturally infected plants. Full-length viral genomes of DNA-A and DNA-B were cloned in plasmid pUC18 at HindIII and XbaI sites, respectively. Partial tandem dimers of the viral clones were infective to Nicotiana benthamiana and tomato cv. Organ Spring through particle bombardment. Infected N. benthamiana plants exhibited downward and upward leaf curling, big veins, leaf puckering with interveinal chlorosis, and stunting. On tomato, symptoms were the same as those seen on naturally infected plants. Cloned DNA also infected Capsicum annuum cv. California Wonder (upward leaf curling and stunting) and tobacco cv. Xanthi (leaf curling and crinkling) but failed to infect Phaseolus vulgaris, okra, cotton, and N. glutinosa. The Varanasi isolate was sap transmissible (0.1 M potassium phosphate buffer, pH 7.0) from the bombarded plants to N. benthamiana and tomato cv. Organ Spring. DNA-A alone infected N. benthamiana (upward leaf curling and big veins) and tomato cv. Organ Spring (mild leaf curl), but symptoms were delayed and milder. Full-length genome sequencing revealed DNA-A (AY190290) contained 2,757 nt and DNA-B (AY190291) contained 2,688 nt. DNA-A of the Varanasi isolate shares 98.4% identity with a DNA-A sequence (AF449999) obtained from a tomato showing leaf curl symptoms from the same region and 97.1% identity with an isolate from Gujarat (900 km from Varanasi). All three sequences represent isolates of the same species, herein called Tomato leaf curl Gujarat virus, based on the priority of submission of the DNA sequence for the Gujarat region (ToLCGV; AF 413671). All isolates noted were obtained from GenBank. However, except for the DNA-A sequence, no other information is available for these ToLCGV isolates. DNA-A of the ToLCGV-Varanasi isolate shares 66.8% identity with Tomato leaf curl New Delhi virus, severe strain (ToLCNdV-Svr) (U15015), and 84.1% with Tomato leaf curl Karnataka virus (U38239). No DNA-B has been reported for these two ToLCGV isolates, and no infectious clone proving the etiology of the disease has been constructed, except for ToLCGV-Varanasi. DNA-B of ToLCGV-Varanasi shares 79.2% homology with ToLCNdV-Svr and 84.1% with ToLCNdV-Luc (X89653). These results suggest that the isolate from Varanasi belongs to ToLCGV, a previously undescribed geminivirus species causing a devastating tomato leaf curl disease in Gujarat and Uttar Pradesh. References: (1) H. C. Birnboim and J. Doly. Nucleic Acids Res. 7:1513, 1979. (2) K. M. Srivastava et al. J. Virol. Methods 51:297, 1995. (3) S. D. Wyatt and J. K. Brown. Phytopathology 86:1288, 1996.

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Evidence for the Association of a Bipartite Geminivirus with Tomato Leaf Curl Disease in Pakistan

Plant Disease ◽

10.1094/pdis.1997.81.8.958c ◽

1997 ◽

Vol 81 (8) ◽

pp. 958-958 ◽

Cited By ~ 11

Author(s):

S. Mansoor ◽

S. H. Khan ◽

M. Saeed ◽

A. Bashir ◽

Y. Zafar ◽

...

Keyword(s):

Tomato Leaf ◽

Cotton Leaf ◽

Degenerate Primers ◽

Tomato Plants ◽

Tomato Leaf Curl Disease ◽

Eastern Hemisphere ◽

Tomato Leaf Curl ◽

Leaf Curl Virus ◽

Leaf Curl Disease ◽

Leaf Curl

Tomato leaf curl disease is the most important constraint on tomato production in Pakistan, where it is found throughout the country. The disease, which occurs in high incidence in Punjab and Sindh provinces, causes 30 to 40% yield losses in the spring crop and uneconomically high losses when grown as an autumn crop. The symptoms of the disease include upward or downward leaf curling, vein thickening, and stunting of the plant. The disease is transmitted by Bemisia tabaci whiteflies (non-B, biotype K) and is suspected to be caused by a geminivirus. For the detection of geminivirus, total DNA was extracted from infected plants, fractionated in an agarose gel, transferred to a nylon membrane, and Southern blotted. A full-length clone of DNA-A of cotton leaf curl virus from Pakistan (S. Mansoor, I. Bedford, M. S. Pinner, A. Bashir, R. Briddon, J. Stanley, Y. Zafar, K. A. Malik, and P. G. Markham, unpublished) was labeled with [32P]dCTP by the oligo-labeling method and hybridized at medium stringency. Geminivirus DNA forms that are normally found in infected plants were detected in plants with tomato leaf curl disease but not in healthy plants. To further confirm the presence of a whiteflytransmitted geminivirus, universal primers for dicot-infecting geminiviruses (1) were used in polymerase chain reaction (PCR) and a product of expected size (approximately 2.7 kb) was detected. The 2.7-kb PCR-amplified DNA from diseased tomato plants was labeled with [32P]dCTP and used as probe in Southern hybridization. This probe also detected geminivirus DNA forms at medium stringency. Both monopartite and bipartite geminiviruses transmitted by whiteflies have been reported to cause leaf curl symptoms on tomato from the Eastern hemisphere. Degenerate primers (PBLv2040 and PCRc1), which amplify B component DNA, were used to determine if tomato leaf curl was monopartite or bipartite (2). A product of expected size (0.65 kb) was amplified, suggesting this virus to be bipartite. DNA-B PCR product obtained from diseased tomato plants was hybridized as described above and detected geminivirus DNA forms at medium stringency. Samples of diseased tomato plants were collected from tomato fields throughout Punjab. DNA-A was detected in all 20 samples whereas DNA B was detected in 17 samples when hybridized by dot blot method at medium stringency. Our data show that tomato leaf curl virus from Pakistan is a bipartite geminivirus. This is the first evidence for a bipartite geminivirus in tomato plants from Pakistan. References: (1) R. W. Briddon and P. G. Markham. Mol. Biotechnol. 1:202, 1993. (2) M. R. Rojas et al. Plant Dis. 77:340, 1993.

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A New Begomovirus Species Causing Tomato Leaf Curl Disease in Patna, India

Plant Disease ◽

10.1094/pdis-93-5-0545b ◽

2009 ◽

Vol 93 (5) ◽

pp. 545-545 ◽

Cited By ~ 9

Author(s):

P. Kumari ◽

B. Chattopadhyay ◽

A. K. Singh ◽

S. Chakraborty

Keyword(s):

Viral Genome ◽

Tandem Repeats ◽

Tomato Leaf ◽

Begomovirus Species ◽

Full Length ◽

Degenerate Primers ◽

Viral Dna ◽

Tomato Leaf Curl ◽

Leaf Curl Disease ◽

Leaf Curl

During December of 2007, a severe leaf curl disease of tomato (ToLCD) occurred in tomato-growing areas in the Patna District of Bihar, India. Viral DNA was isolated from symptomatic tomato plants (2) and begomovirus association was confirmed by PCR using DNA-A degenerate primers (3). Isolated viral DNA was restricted with KpnI and full-length genome was cloned in pUC18. DNA-β was amplified using degenerate primers (1) and cloned in pTZ57RT. Partial tandem repeats of viral genome and DNA-β could infect Nicotiana benthamiana and tomato through Agrobacterium-mediated inoculation. Infected test plants exhibited typical symptoms characteristic of ToLCD. Full-length viral genome (GenBank Accession No. EU862323) consists of 2,752 nt and showed the highest identity (85.8%) with Tomato leaf curl Laos virus-[Laos] (GenBank Accession No. AF195782). The satellite DNA-β component (GenBank Accession No. EU862324) consists of 1,349 nt and showed the highest identity (75.8%) with Tomato leaf curl Joydebpur betasatellite (GenBank Accession No. AJ966244). On the basis of the ICTV species demarcation criteria of 89% of DNA-A sequence identity, the present isolate was considered as a new begomovirus species and named Tomato leaf curl Patna virus (ToLCPaV). Since the isolated DNA-β satellite shares less than 78% identity, it is considered a new species of betasatellite and the name, Tomato leaf curl Patna betasatellite (ToLCPaB) is proposed. These results show that severe ToLCD in Patna is caused by a newly identified species of begomovirus and betasatellite. References: (1) R. W. Briddon et al. Mol. Biotechnol 20:315, 2002. (2) S. Chakraborty et al. Phytopathology 93:1485, 2003. (3) S. D. Wyatt and J. K. Brown. Phytopathology 86:1288, 1996.

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Molecular Characterization of a Begomovirus, α-Satellite, and β-Satellite Associated with Leaf Curl Disease of Parthenium hysterophorus in India

Plant Disease ◽

10.1094/pdis-09-15-0982-re ◽

2016 ◽

Vol 100 (11) ◽

pp. 2299-2305 ◽

Cited By ~ 9

Author(s):

Susheel Kumar ◽

Ashish Srivastava ◽

Meraj Jaidi ◽

Puneet Singh Chauhan ◽

S. K. Raj

Keyword(s):

Satellite Dna ◽

Rolling Circle Amplification ◽

Tomato Leaf ◽

Parthenium Hysterophorus ◽

Alternate Host ◽

Invasive Weed ◽

Tomato Leaf Curl Disease ◽

Tomato Leaf Curl ◽

Leaf Curl Disease ◽

Leaf Curl

Parthenium hysterophorus plants exhibiting severe leaf curl and stunting symptoms were observed near agriculture fields in Lucknow, India. The association of a begomovirus, β-satellite, and α-satellite with these symptoms of a Parthenium disease was investigated by sequence analyses of virus and satellite DNA amplified by rolling circle amplification and polymerase chain reaction. The highest sequence identities and closest phylogenetic relationships for the begomovirus, β-satellite, and α-satellite detected in P. hysterophorus plants were to Tomato leaf curl virus (ToLCV), papaya leaf curl β-satellite (PaLCuB), and Ageratum yellow vein India α-satellite (AYVIA), respectively. These findings identified the virus and satellites infecting the Parthenium sp. as ToLCV, PaLCuB, and AYVIA, respectively. P. hysterophorus and tomato seedlings infected with cloned ToLCV, PaLCuB, and AYVIA by agroinoculation developed leaf curl symptoms, whereas plants infected with ToLCV alone or with ToLCV and AYVIA developed mild yellowing. The results show that this complex infects and causes disease in P. hysterophorus and tomato. P. hysterophorus is an invasive weed commonly found around agricultural fields and along roadsides in India. These results indicate that P. hysterophorus plants infected with ToLCV and associated satellite DNA act as an alternate host (reservoir), and that could lead to increased incidence of tomato leaf curl disease.

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