Bipartite Begomovirus Research Articles

Emergence of tomato leaf curl New Delhi virus in Italy: estimation of incidence and genetic diversity

Tomato leaf curl New Delhi virus(ToLCNDV) is a bipartite begomovirus (familyGeminiviridae) identified for the first time in 1995 in Asia, from where it spread into several countries of the Mediterranean basin. ToLCNDVwas found in Spain in 2012, and subsequently in Tunisia and Italy. The first outbreak in Italy occurred at the end of 2015 in Trapani province (Sicily) on zucchini squashes. Then in 2016, ToLCNDVwas found in infected zucchini plants in Campania, Lazio and Sardinia regions, and in 2017 in Calabria. This study addressed the dispersion and genetic diversity of ToLCNDVisolates in Italy. A total of 1400 plants were analysed. Phylogenetic analysis showed low variability among the Italian isolates, probably as a consequence of the recent introduction and rapid spread of this virus in Italy. Two statistically significant clusters were reported: one grouping only Italian isolates and the other grouping Italian, Spanish, Tunisian and Moroccan isolates. Furthermore, the highest incidence of ToLCNDVwas observed in Sicily, although the disease also appears to be critical in other Italian regions. In this work, a high efficiency of ToLCNDVmechanical transmission intoCucurbita pepo,Cucumis melo inodorusandCucumis melo cantalupensishas been demonstrated. The rapid spread of ToLCNDVin the Mediterranean basin represents a threat for horticultural production, thus it is very important to develop suitable crop management practices, applying genetic resistance strategies and more restrictive phytosanitary measures.

Molecular characterization of Mungbean yellow mosaic India virus infecting tomato (Solanum lycopersicum L.)

The severe mosaic and mild leaf curl symptoms were observed on tomato crops in the agricultural farm at Chitrakoot, Madhya Pradesh, India. Based on typical viral symptoms and high populations of whiteflies in the field, the association of begomovirus was suspected. The identification of a bipartite begomovirus associated with the disease was performed by sequence analyses of their cloned full-length genome. The complete genome sequences were submitted in GenBank under accession numbers MF683072 (DNA-A) and MF683073 (DNA-B). The sequence analyses of both genome of bipartite begomovirus revealed highest nucleotide sequence identity and close phylogenetic relationships with Mungbean yellow mosaic India virus (MYMIV) reported from India, therefore, following the ICTV guidelines, the begomovirus under study was identified as an isolate of MYMIV. Best of our knowledge, this is the first report of MYMIV infecting tomato.

A Novel Strain of Pepper Leafroll Virus Infecting Common Bean and Soybean in Ecuador

Symptoms resembling those caused by begomoviruses (Geminiviridae: Begomovirus) have been observed in common bean and other leguminous crops in Ecuador in the last few years. The only begomovirus species found infecting legumes (common bean and cowpea) in the country to date is cabbage leaf curl virus (CabLCV) (Fiallo-Olive et al. 2018). During May to June 2017, a survey of leguminous plants was conducted in the Ecuadorian localities of Santa Elena (Santa Elena province), Milagro (Guayas province), and Pallatanga (Chimborazo province). Twenty-four plants showing leaf deformation and/or mosaic symptoms were sampled, including common bean (Phaseolus vulgaris, n = 21), soybean (Glycine max, n = 2), and pigeon pea (Cajanus cajan, n = 1). Most common bean plants were infested with whiteflies (Hemiptera: Aleyrodidae) that were also collected. Total DNA was extracted from leaf samples and used as template in polymerase chain reaction (PCR) using specific primers for CabLCV (MA2540 [5′-GATGTTGAAGCATCTGCAAACATTC-3′] and MA2541 [5′-CTAGGAACATCAGGGCTTCTCAAC-3′]) (Fiallo-Olive et al. 2018). All samples were negative for the presence of CabLCV. DNA extracts were then used as templates in rolling-circle amplification (RCA) using φ29 DNA polymerase (TempliPhi kit, GE Healthcare). Digestion of RCA products with a set of restriction enzymes yielded identical restriction patterns for six samples (five common bean and one soybean), suggesting the presence of a bipartite begomovirus. BamHI and NcoI fragments of approximately 2.6 kbp from a common bean plant were cloned in pBluescript II SK(+), and two selected clones were sequenced (Macrogen, Seoul, South Korea). BLAST analysis showed that the clones corresponded to begomoviral DNA-A (BamHI, 2,580 nt, GenBank accession no. MH481901) and DNA-B (NcoI, 2,561 nt, MH481902). Pairwise identity scores with isolates selected after BLAST analysis were calculated with SDT (Muhire et al. 2014). Cloned DNA-A and DNA-B showed the highest identity (93 and 92%, respectively) with pepper leafroll virus (PepLRV) (KC769819 and KC769820), a bipartite begomovirus identified infecting pepper, common bean, and tomato in Peru (Martinez-Ayala et al. 2014). To confirm the identity of the begomovirus present in the rest of the samples, PCR was performed with specific primers whose design was based on the cloned PepLRV DNA-A (MA2602 [5′-GTTAATGGACAATGAGGGTATGTG-3′] and MA2603 [5′-CACGAGGAGAATAATTGGCGTTAC-3′]) using Taq DNA polymerase (BIOTAQ, Bioline) and thermocycler conditions consisting of 35 cycles with 45 s annealing steps at 57°C. DNA fragments of the expected size (452 bp) were amplified from the six samples, and direct sequencing showed them to be identical to the cloned PepLRV DNA-A, supporting the presence of this virus. Based on the species/strain demarcation threshold for the viruses in the genus Begomovirus (Brown et al. 2015), the isolate characterized here belongs to a new strain of PepLRV. DNA was extracted from individual whiteflies (n = 42) using a Chelex-based method, and PCR was carried out using mitochondrial cytochrome oxidase I gene primers, C1-J-2195 (5′-TTGATTTTTTGGTCATCCAGAAGT-3′) and Btab-uni-PrimerR (5′-CTTAAATTTACTGCACTTTCTGCCAYATTAG-3′). Direct sequencing of PCR products showed that seven sequences (one deposited in GenBank with accession no. MH481903) were identical to numerous sequences of Bemisia tabaci Middle East-Asia Minor 1 (MEAM1, formerly B biotype), known to be an efficient vector of begomoviruses. To our knowledge, this is the first report of PepLRV in Ecuador and of soybean as a host for the virus, which had been identified only in Peru to date. Also, this is the first molecular evidence of the presence of B. tabaci MEAM1 in the country.

Occurrence and molecular detection of bipartite begomovirus on tomato in Southern India

Begomoviruses are major constraint in tomato production across the world. In India, tomato production is affected by both monopartite and bipartite begomoviruses. Monopartite were reported to infect tomato crop across India. However, bipartite were reported to infect tomato in Nothern India. In the current study, seventy begomovirus symptomatic tomato samples were collected from southernstates of India and analysed using PCR-based specific primer. The analysis revealed that 45 of them were monopartite, 23 of them were bipartite and 8 bipartite also showed the presence of betasatellite. This is the first report of bipartite begomovirus infecting tomato in Southern India.

New approach for the construction of infectious clones of a circular DNA plant virus using Gibson Assembly

Viruses belonging to the genus Begomovirus (family Geminiviridae) have circular single-strand DNA genomes encapsidated into quasi-icosahedral particles, and are transmitted by whiteflies of the Bemisia tabaci complex. Biological and molecular properties of begomoviruses have been studied efficiently with infectious clones containing dimeric genomic components. However, current approaches employing enzymatic digestion and ligation to binary vectors are laborious, mostly due to many cloning steps or partial digestion by restriction enzyme. Here, an infectious clone of the bipartite begomovirus Bean golden mosaic virus (BGMV) was obtained using PCR and Gibson Assembly (GA). Common bean (Phaseolus vulgaris) seedlings displayed severe yellow mosaic and stunt symptoms 15 days after agroinoculation with DNA-A and DNA-B of BGMV. The approach based on PCR-GA protocol is a fast and useful tool to obtain infectious clones of a circular DNA plant virus.

A WW Domain-Containing Protein Forms Immune Nuclear Bodies against Begomoviruses

The bipartite begomoviruses (Geminiviridae family), which are DNA viruses that replicate in the nucleus of infected cells, encode the nuclear shuttle protein (NSP) to facilitate the translocation of viral DNA from the nucleus to the cytoplasm via nuclear pores. This intracellular trafficking of NSP–DNA complexes is accessorized by the NSP-interacting guanosine triphosphatase (NIG) at the cytosolic side. Here, we report the nuclear redistribution of NIG by AtWWP1, a WW domain-containing protein that forms immune nuclear bodies (NBs) against begomoviruses. We demonstrated that AtWWP1 relocates NIG from the cytoplasm to the nucleus where it is confined to AtWWP1-NBs, suggesting that the NIG-AtWWP1 interaction may interfere with the NIG pro-viral function associated with its cytosolic localization. Consistent with this assumption, loss of AtWWP1 function cuased plants more susceptible to begomovirus infection, whereas overexpression of AtWWP1 enhanced plant resistance to begomovirus. Furthermore, we found that a mutant version of AtWWP1 defective for NB formation was no longer capable of interacting with and relocating NIG to the nucleus and lost its immune function against begomovirus. The antiviral function of AtWWP1-NBs, however, could be antagonized by viral infection that induced either the disruption or a decrease in the number of AtWWP1-NBs. Collectively, these results led us to propose that AtWWP1 organizes nuclear structures into nuclear foci, which provide intrinsic immunity against begomovirus infection.

Tomato leaf curl Mahé virus: a novel tomato-infecting monopartite begomovirus from the Seychelles.

This is the first description of the complete genome sequence of a new monopartite begomovirus isolated from tomato with symptoms of tomato (yellow) leaf curl disease collected in the Seychelles. The DNA-A-like nucleotide sequences share the highest nucleotide sequence identity (84%) with tomato leaf curl Anjouan virus (ToLCAnjV) from the Comoros islands. Phylogenetic analysis confirmed its recombinant nature and its relationship to Old World monopartite and bipartite begomoviruses. This discovery of a new member of a species confirms the high genetic diversity of begomoviruses in the south-western Indian Ocean islands.

First natural co-occurrence of tomato leaf curl New Delhi virus DNA-A and chili leaf curl betasatellite on tomato plants (Solanum lycopersicum L.) in India

Tomato leaf curl disease (ToLCD) affected 25% of the tomato crop in Chitrakoot, India and symptomatic leaves were collected for molecular assay. The complete sequences of bipartite begomovirus DNA-A and a betasatellite DNA were amplified. In a sequence analysis, begomovirus DNA-A and betasatellite shared highest sequence identity (91–99%) with Tomato leaf curl New Delhi virus (ToLCNDV) DNA-A and chili leaf curl betasatellite (ChLCB), respectively. The virus was transmitted by whitefly to tomato plants and caused ToLCD symptoms with 70% transmission rate. To our knowledge, this is the first report of the natural occurrence of ToLCNDV and ChLCB in India.

Croton golden mosaic virus: a new bipartite begomovirus isolated from Croton hirtus in Colombia.

A new begomovirus infecting Croton hirtus (Euphorbiaceae) from Colombia has been characterized. The complete DNA-A and DNA-B components were determined to be 2613 and 2551nt in length, respectively, showing the typical genome organization of bipartite New World begomoviruses. DNA-A showed the highest nucleotide sequence identity (81.2%) to sida yellow mottle virus (JN411687), a begomovirus isolated from Sida rhombifolia in Cuba. Based on the current ICTV species demarcation criterion for the genus Begomovirus, we report a new member of this genus infecting C. hirtus. We propose that it be named Croton golden mosaic virus (CroGMV), based on the symptoms observed in the weed. Phylogenetic analysis revealed that CroGMV segregates into a single clade and has some relationship with viruses from Central America and the Caribbean. CroGMV is the first Croton-infecting bipartite begomovirus reported in the world.

Passion Fruit Chlorotic Mottle Virus: Molecular Characterization of a New Divergent Geminivirus in Brazil.

Brazil is one of the major passion fruit producers worldwide. Viral diseases are among the most important constraints for passion fruit production. Here we identify and characterize a new passion fruit infecting-virus belonging to the family Geminiviridae: passion fruit chlorotic mottle virus (PCMoV). PCMoV is a divergent geminivirus unlike previously characterized passion fruit-infecting geminiviruses that belonged to the genus Begomovirus. Among the presently known geminiviruses, it is most closely related to, and shares ~62% genome-wide identity with citrus chlorotic dwarf associated virus (CCDaV) and camelia chlorotic dwarf associated virus (CaCDaV). The 3743 nt PCMoV genome encodes a capsid protein (CP) and replication-associated protein (Rep) that respectively share 56 and 60% amino acid identity with those encoded by CaCDaV. The CPs of PCMoV, CCDaV, and CaCDaV cluster with those of begomovirus whereas their Reps with those of becurtoviruses. Hence, these viruses likely represent a lineage of recombinant begomo-like and becurto-like ancestral viruses. Furthermore, PCMoV, CCDaV, and CaCDaV genomes are ~12–30% larger than monopartite geminiviruses and this is primarily due to the encoded movement protein (MP; 891–921 nt) and this MP is most closely related to that encoded by the DNA-B component of bipartite begomoviruses. Hence, PCMoV, CCDaV, and CaCDaV lineage of viruses may represent molecules in an intermediary step in the evolution of bipartite begomoviruses (~5.3 kb) from monopartite geminiviruses (~2.7–3 kb). An infectious clone of PCMoV systemically infected Nicotiana benthamina, Arabidopsis thaliana, and Passiflora edulis.

Complete genome sequence of a new bipartite begomovirus infecting Boehmeria leiophylla in China.

A bipartite begomovirus was identified from a Boehmeria leiophylla plant sample exhibiting yellow mosaic symptoms collected in Nabanhe National Nature Reserve, Xishuangbanna, Yunnan, China. Five complete DNA-A and four DNA-B genome sequences were obtained by rolling-circle amplification (RCA), cloned, and sequenced. All DNA-A sequences were determined to be 2759 nucleotides in size, sharing 99.9%-100% nucleotide sequence identity with each other. The DNA-B sequences were comprised of 2673 nucleotides, sharing 98.6-100% nucleotide sequence identity with each other. Genomic organization of the begomovirus was typical of Old World bipartite begomoviruses. Sequence analysis revealed 81.84% nucleotide sequence identity to tomato leaf curl Hsinchu virus (ToLCHsV) from China for the DNA A component and 67.23% identity to the squash leaf curl China virus (SLCCNV) from India for the DNA B component. The sequence comparisons suggest that this bipartite begomovirus represents a novel species for which we propose the name "Ramie yellow mosaic virus".

Alteraciones anatómicas e hiperplasia inducidas por Euphorbia mosaic virus aislamiento Yucatan Peninsula en el mesófilo

<p><em>Euphorbia mosaic virus</em> aislamiento Yucatan Peninsula es un begomovirus bipartita con una amplia gama de hospedantes experimentales, que incluyen especies de importancia económica, como frijol, chile y tomate, entre otras. Este virus provocó síntomas severos en hojas de <em>Nicotiana benthamiana</em>, pérdida de identidad celular y tisular, deformaciones de la lámina foliar y de la nervadura central, cambios en la estructura del mesó- filo y proliferación celular, a diferencia de otros begomovirus bipartitas. Activó la expresión de los genes cycA1, cycA2 y cdkB2.2, implicados en la transición G2/M del ciclo celular mitótico y se localizó en el mesófilo de las hojas. EuMV-YP es el primer ejemplo de un begomovirus bipartita capaz de inducir la proliferación celular.</p>

Minimal genomic variability in Merremia mosaic virus isolates endemic in Merremia spp and cultivated tomato in Puerto Rico.

Merremia mosaic virus (MerMV), a bipartite begomovirus, was identified for the first time as a pathogen of commercial tomato plantings. Infection of tomato by MerMV caused mild leaf curling and yellow foliar mosaic symptoms. Herein, the MerMV was identified in symptomatic Merremia quinquefolia and M. aegyptia (Convolvulaceae) plants exhibiting bright yellow or yellow-green foliar mosaic symptoms, respectively. The full-length begomoviral components were amplified from total DNA isolated from two wild species of Merremia and commercial tomato plants during 1991-1998. The DNA was subjected to rolling circle amplification, restriction digestion, and DNA sequencing. The resultant 19 and 26 apparently full-length DNA-A and DNA-B components were~2557 and~2492 bases, respectively. The 140-base common region was 97.9% identical between DNA-A and -B components, a predictive evidence for cognate DNA-A and -B components. Although the DNA-A components were highly conserved at 96-100%, the DNA-B components diverged at~89 to 100%, respectively. The overall clonal genomic features strongly suggested that MerMV lineage has been under host-selection for some time, and only recently, has undergone a host-shift, putatively, from wild convolvulaceous species to tomato (Solanaceae). Phylogenetically, MerMV grouped with other bipartite begomoviruses indigenous to the Caribbean region, with MerMV DNA-A components forming three clusters, and the DNA-B components grouped in one clade. Both clades contained only one closet relative, an isolate of MerMV from Venezuela, MerMV-VE. Biolistic inoculation of M. quinquefolia and tomato seedlings with the DNA-A and -B components of PR68 and PR80 resulted in development of symptoms like those observed in naturally-infected species, respectively.

AV2 protein of tomato leaf curl Palampur virus promotes systemic necrosis in Nicotiana benthamiana and interacts with host Catalase2

Tomato leaf curl Palampur virus (ToLCPalV) is a whitefly-transmitted, bipartite begomovirus. Here, we demonstrated that ectopic expression of AV2 from a Potato virus X (PVX)-based vector accelerated systemic necrosis and reactive oxygen species (ROS) accumulation in Nicotiana benthamiana. Furthermore, 10 amino acids from N-terminal region of AV2 were found to be associated with the systemic necrosis symptom/phenotype. Mutational studies of ToLCPalV infectious clones lacking the AV2 revealed that AV2 is essential for the systemic movement of DNA-A, symptom severity and viral DNA accumulation. In a yeast two-hybrid assay, Catalase2 (Cat2) was found to associate with AV2 protein. Further, silencing of Cat2 resulted in appearance of necrotic lesions on N. benthamiana and these plants were highly susceptible to ToLCPalV infection in comparison to control plants. Infection ToLCPalV on Solanum lycopersicum resulted in downregulation of Cat2 transcripts, followed by accumulation of ROS and stress marker transcripts. The AV2 protein also suppressed virus-induced gene silencing (VIGS) of the Phytoene desaturase (PDS) gene. Our results show that AV2 is essential for the pathogenicity, systemic movement and suppression of gene silencing in the host. Altogether, our findings suggest that interactions between AV2 and Cat2 might play a crucial role in the establishment of ToLCPalV infection.

Detection and characterization of tomato leaf curl New Delhi virus association with mosaic disease of ivy gourd (Coccinia grandis (L.) Voigt) in North India

Sixteen ivy gourd ( Coccinia grandis (L.) Voigt) plant samples showing severe mosaic symptoms were collected from New Delhi and Varanasi (Uttar Pradesh) in India. Begomovirus infection was confirmed by PCR using begomovirus-specific primers. Amplified PCR products (1.2 kb fragments) were cloned and the sequence was characterized. Based on sequence analysis, begomovirus associated with the majority of ivy gourd samples (16) was found to be a member of a bipartite begomovirus species, which is closely related to tomato leaf curl New Delhi virus (ToLCNDV). Therefore, two samples of ivy gourd, IVG1-ND and IVG2-Var, were selected for full-length genome (DNA-A and DNA-B-like sequence) amplification by the rolling circle DNA amplification (RCA) method. Sequence analysis performed using the Species Demarcation Tool (SDT) program revealed that they share 89.5-91.3% (IVG1-ND) and 93.4-96.8% (IVG2-Var) nucleotide (nt) identity with the DNA-A-like sequence of ToLCNDV isolated from cucurbits and chilli, respectively. The IVG1-ND and IVG2-Var isolates shared 90% nt identity among themselves, indicating that they are two different strains of ToLCNDV. Similarly, SDT analysis of the DNA-B-like sequence of IVG1-ND and IVG2-Var exhibited showed 82.7-93.3% nt identity with the DNA-B-like sequences of ToLCNDV infecting cucurbits. The recombination analysis of DNA-A and DNB-B-like sequences showed that the greater part of their genome most likely originated from previously reported begomoviruses that are known to infect chilli and cucurbits through recombination. https://doi.org/10.2298/ABS170616051V Received: June 16, 2017; Revised: November 16, 2017; Accepted: November 29, 2017; Published online: December 21, 2017 How to cite this article: Venkataravanappa V, Narasimha Reddy LRC, Saha S, Subbanna SK, Manem KR. Detection and characterization of tomato leaf curl New Delhi virus association with mosaic disease of ivy gourd ( Coccinia grandis (L.) Voigt) in North India. Arch Biol Sci. 2018;70(2):339-47.

Molecular detection and characterization of phytoplasma in association with begomovirus in eggplant.

The samples from eggplants showing mixed symptoms of little leaf and mosaic were collected from two districts (Mirzapur and Varanasi) of Uttar Pradesh, India. The total nucleic acid extracted from these samples was amplified by PCR using universal 16S rRNA primers specific to phytoplasma and primers specific to DNA-A-like sequence of begomovirus. A total of eighteen eggplant samples showing the symptoms of little leaf and mosaic tested positive for the presence of both begomovirus and phytoplasma. The phytoplasma associated with the mixed symptoms of mosaic and little leaf in the eggplant samples was identified as a member belonging to Clover proliferation group (16SrVI) (nucleotide sequence identity of 97.5-97.8%). The characterized begomovirus from the eggplant samples was identified as a strain of previously described bipartite begomovirus tomato leaf curl Palampur virus (ToLCPalV) (92.5-94.1% nucleotide sequence identity), which is known to infect cucurbits and solanaceous crops in India and Ireland. Further, putative recombination events were detected within the 16S rRNA gene F2n/R2 fragment of phytoplasma and DNA-A of strain of ToLCPalV. Most of the sequence variations observed within the phytoplasma were due to intra and interspecific recombination events between eggplant little leaf-16SrVI-D, Ca. P. asteris-16SrI and Ca. P. pruni-16SrIII. Similarly, most of the DNA fragments of newly characterized strain of ToLCPalV appear to have been derived from tomato leaf curl New Delhi virus (ToLCNDV), squash leaf curl China virus (SLCCNV) and ToLCPalV like ancestors. This perhaps is the first evidence of mixed infection of both phytoplasma-begomovirus in eggplant in India.

Breeding tomato (Solanum lycopersicum L.) for resistance to biotic and abiotic stresses

Tomato (Solanum lycopersicum L.) is an important vegetable crop cultivated in the tropical and sub-tropical regions of the world. Low productivity in India is due to occurrence of both biotic and abiotic stresses. Among the biotic stresses, tomato leaf curl disease, bacterial wilt, early blight and Groundnut Bud Necrosis Virus disease have become serious production constraints causing considerable yield loss in the major tomato growing areas of the country. Adoption of multiple disease resistant varieties or F1 hybrids would be the most appropriate way to address these diseases. At ICAR-IIHR, Bengaluru systematic breeding strategies were employed to pyramid genes for resistance to early blight, bacterial wilt and tomato leaf curl diseases and to develop advanced breeding lines& F1 hybrids with triple disease resistance. Stable source of resistance to early blight and bi-partite begomo-virus (Tomato Leaf Curl New Delhi Virus) has been identified in Solanum habrochaites LA-1777. Validation with molecular markers linked to tomato leaf curl virus resistance revealed that LA-1777 carryTy2 and other putative resistant genes. Several high yielding dual purpose hybrids were also developed for fresh market and processing with high level of resistance to multiple diseases. Cherry tomato lines have also been bred for high TSS, total carotenoids, total phenols, flavonoids, vitamin C, acidity and lycopene content. IIHR-249-1, IIHR-2101 (Solanum habrochaites LA-1777), IIHR- 2866 and IIHR-2864 recorded high values for quality parameters like total carotenoids, lycopene, vitamin C, total phenols, flavonoids and TSS. Drought tolerant root stock has been developed by an interspecific cross between S. habrochaites LA-1777 and S. lycopersicum (15 SB SB). Resistant sources have also been identified against Tuta absoluta, a serious insect pest reported from major tomato growing areas in the country in recent time. High temperature tolerant breeding lines are in pipe line.

Maintenance of Cotton Leaf Curl Multan Betasatellite by Tomato Leaf Curl New Delhi Virus-Analysis by Mutation.

Viruses of the genus Begomovirus (family Geminiviridae) are economically important phytopathogens that are transmitted plant-to-plant by the whitefly Bemisia tabaci. Most Old World (OW) begomoviruses are monopartite and many of these interact with symptoms and host range determining betasatellites. Tomato leaf curl New Delhi virus (ToLCNDV) is one of only a few OW begomoviruses with a bipartite genome (components known as DNA A and DNA B). Four genes [AV2, coat protein (CP), transcriptional-activator protein (TrAP), and AC4] of ToLCNDV were mutated and the effects of the mutations on infectivity, symptoms and the ability to maintain Cotton leaf curl Multan betasatellite (CLCuMuB) were investigated. Infectivity and virus/betasatellite DNA titer were assessed by Southern blot hybridization, PCR, and quantitative PCR. The results showed TrAP of ToLCNDV to be essential for maintenance of CLCuMuB and AV2 to be important only in the presence of the DNA B. AC4 was found to be important for the maintenance of CLCuMuB in the presence of, but indispensable in the absence of, the DNA B. Rather than being required for maintenance, the CP was shown to possibly interfere with maintenance of the betasatellite. The findings show that the interaction between a bipartite begomovirus and a betasatellite is more complex than just trans-replication. Clearly, multiple levels of interactions are present and such associations can cause additional significant losses to crops although the interaction may not be stable.

Natural Infection and Recombination Analysis of Bipartite Begomovirus and its Cognate Beta-satellite in Benincasa hispida

Natural Infection and Recombination Analysis of Bipartite Begomovirus and its Cognate Beta-satellite in Benincasa hispida

Characterization of tomato leaf curl purple vein virus, a new monopartite New World begomovirus infecting tomato in Northeast Brazil.

A new begomovirus species was identified from tomato plants with upward leaf curling and purple vein symptoms, which was first identified in the Piaui state of Northeast (NE) Brazil in 2014. Tomato leaf samples were collected in 2014 and 2016, and PCR with degenerate primers revealed begomovirus infection. Rolling circle amplification and restriction enzyme digestion indicated a single genomic DNA of ~ 2.6kb. Cloning and sequencing revealed a genome organization similar to DNA-A components of New World (NW) bipartite begomoviruses, with no DNA-B. The complete nucleotide sequence had the highest identity (80%) with the DNA-A of Macroptilium yellow spot virus (MacYSV), and phylogenetic analyses showed it is a NW begomovirus that clusters with MacYSV and Blainvillea yellow spot virus, also from NE Brazil. Tomato plants agroinoculated with a dimeric clone of this genomic DNA developed upward leaf curling and purple vein symptoms, indistinguishable from those observed in the field. Based on agroinoculation, this virus has a narrow host range, mainly within the family Solanaceae. Co-inoculation experiments with tomato severe rugose virus and tomato mottle leaf curl virus, the two predominant begomoviruses infecting tomato in Brazil, revealed a synergistic interaction among these begomoviruses. The name Tomato leaf curl purple vein virus (ToLCPVV) is proposed for this new begomovirus.