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1.
Arch Virol ; 165(7): 1659-1665, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32405827

RESUMO

In this work, a begomovirus isolated from a bean plant coinfected with the potyviruses bean common mosaic virus and bean common mosaic necrosis virus was characterized. The three viruses were detected by high-throughput sequencing and assembly of total small RNAs, but the begomovirus-related contigs did not allow precise identification. Molecular analysis based on standard DNA amplification techniques revealed the presence of a single bipartite virus, which is a novel begomovirus according to the current taxonomic criteria. Infectious clones were generated and agroinoculated into Phaseolus vulgaris and Nicotiana benthamiana plants. In all cases, viral DNA-A and DNA-B were detected in new growths, but no symptoms were observed, thus indicating that this virus produces asymptomatic infections in both host species.


Assuntos
Begomovirus/isolamento & purificação , Phaseolus/virologia , Doenças das Plantas/virologia , Potyvirus/fisiologia , Tabaco/virologia , Begomovirus/classificação , Begomovirus/genética , Begomovirus/fisiologia , Coinfecção/virologia
2.
Phytopathology ; 110(1): 121-129, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31584339

RESUMO

Isolates of the Tomato yellow leaf curl virus (TYLCV) species (genus Begomovirus, family Geminiviridae) infect tomato crops worldwide, causing severe economic damage. Members of the whitefly Bemisia tabaci sibling species group are the vector of begomoviruses, including TYLCV. However, transmission of isolates of the type strain (Israel [IL]) of TYLCV (TYLCV-IL) by tomato seed has recently been reported based on infections occurring in Korea. Because of the consequences of this finding on the epidemiology and control of the disease caused by TYLCV and on the seed market, it was considered essential to revisit and expand those results to other tomato-growing areas. TYLCV DNA content was detected in tomato and Nicotiana benthamiana seed collected from plants naturally or experimentally infected with TYLCV-IL, supporting its seedborne nature. The TYLCV-IL replication detected in tomato and N. benthamiana flower reproductive organs demonstrated close association of this virus with the seed during maturation. However, the significant reduction of TYLCV DNA load after surface disinfections of tomato seed suggests that most of the virus is located externally, as contaminant of the seed coat. Transmission assays, carried out with seven tomato genotypes and more than 3,000 tomato plants, revealed no evidence of seed transmission from "surface-disinfected" or untreated seed for two Mediterranean isolates of TYLCV-IL. Similar results were also obtained for seed collected from TYLCV-IL-infected N. benthamiana plants. The results support the conclusion that TYLCV-IL is seedborne but is not seed transmitted in tomato or N. benthamiana, suggesting that transmission through seed is not a general property of TYLCV.


Assuntos
Begomovirus , Lycopersicon esculentum , Sementes , Begomovirus/fisiologia , Genótipo , Israel , Lycopersicon esculentum/genética , Lycopersicon esculentum/virologia , Doenças das Plantas/virologia , República da Coreia , Sementes/virologia
3.
Mol Plant Microbe Interact ; 33(1): 87-97, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31638467

RESUMO

Transgenic approaches employing RNA interference (RNAi) strategies have been successfully applied to generate desired traits in plants; however, variations between RNAi transgenic siblings and the ability to quickly apply RNAi resistance to diverse cultivars remain challenging. In this study, we assessed the promoter activity of a cauliflower mosaic virus 35S promoter (35S) and a phloem-specific promoter derived from rice tungro bacilliform virus (RTBV) and their efficacy to drive RNAi against the endogenous glutamate-1-semialdehyde aminotransferase gene (GSA) that acts as a RNAi marker, through chlorophyll synthesis inhibition, and against tomato yellow leaf curl Thailand virus (TYLCTHV), a begomovirus (family Geminiviridae) reported to be the prevalent cause of tomato yellow leaf curl disease (TYLCD) in Taiwan. Transgenic Nicotiana benthamiana expressing hairpin RNA of GSA driven by either the 35S or RTBV promoter revealed that RTBV::hpGSA induced stronger silencing along the vein and more uniformed silencing phenotype among its siblings than 35S::hpGSA. Analysis of transgenic N. benthamiana, 35S::hpTYLCTHV, and RTBV::hpTYLCTHV revealed that, although 35S::hpTYLCTHV generated a higher abundance of small RNA than RTBV::hpTYLCTHV, RTBV::hpTYLCTHV transgenic plants conferred better TYLCTHV resistance than 35S::hpTYLCTHV. Grafting of wild-type (WT) scions to TYLCTHV RNAi rootstocks allowed transferable TYLCTHV resistance to the scion. A TYLCTHV-inoculation assay showed that noninfected WT scions were only observed when grafted to RTBV::hpTYLCTHV rootstocks but not 35S::hpTYLCTHV nor WT rootstocks. Together, our findings demonstrate an approach that may be widely applied to efficiently confer TYLCD resistance.


Assuntos
Begomovirus , Resistência à Doença , Lycopersicon esculentum , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas , Begomovirus/fisiologia , Resistência à Doença/genética , Lycopersicon esculentum/genética , Lycopersicon esculentum/virologia , Floema/genética , Regiões Promotoras Genéticas/genética , RNA/genética
4.
Phytopathology ; 110(1): 130-145, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31573394

RESUMO

Epidemics of tomato yellow leaf curl virus (TYLCV; species Tomato yellow leaf curl begomovirus) have been problematic to tomato production in the southeastern United States since the first detection of the virus in Florida in the late 1990s. Current strategies for management focus on farm-centric tactics that have had limited success for controlling either TYLCV or its whitefly vector. Areawide pest management (AWPM)-loosely defined as a coordinated effort to implement management strategies on a regional scale-may be a viable management alternative. A prerequisite for development of an AWPM program is an understanding of the spatial and temporal dynamics of the target pathogen and pest populations. The objective of this study was to characterize populations of whitefly and TYLCV in commercial tomato production fields in southwestern Florida and utilize this information to develop predictors of whitefly density and TYLCV disease incidence as a function of environmental and geographical factors. Scouting reports were submitted by cooperating growers located across approximately 20,000 acres in southwestern Florida from 2006 to 2012. Daily weather data were obtained from several local weather stations. Moran's I was used to assess spatial relationships and polynomial distributed lag regression was used to determine the relationship between weather variables, whitefly, and TYLCV. Analyses showed that the incidence of TYLCV increased proportionally with mean whitefly density as the season progressed. Nearest-neighbor analyses showed a strong linear relationship between the logarithms of whitefly densities in neighboring fields. A similar relationship was found with TYLCV incidences. Correlograms based on Moran's I showed that these relationships extended beyond neighboring fields and out to approximately 2.5 km for TYLCV and up to 5 km for whitefly, and that values of I were generally higher during the latter half of the production season for TYLCV. Weather was better at predicting whitefly density than at predicting TYLCV incidence. Whitefly density was best predicted by the number of days with an average temperature between 16 and 24°C (T16to24), relative humidity (RH) over the previous 31 days, and vapor pressure deficit over the last 8 days. TYLCV incidence was best predicted by T16to24, RH, and maximum wind speed over the previous 31 days. Results of this study helped to identify the extent to which populations of whitefly and TYLCV exist over the agricultural landscape of southwestern Florida, and the environmental conditions that favor epidemic growth. This information was used to propose an approach to AWPM for timing control measures for managing TYLCV epidemics.


Assuntos
Begomovirus , Hemípteros , Lycopersicon esculentum , Doenças das Plantas , Animais , Begomovirus/fisiologia , Florida , Hemípteros/virologia , Lycopersicon esculentum/virologia , Doenças das Plantas/virologia , Densidade Demográfica , Sudeste dos Estados Unidos , Fatores de Tempo
5.
BMC Plant Biol ; 19(1): 556, 2019 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-31842757

RESUMO

BACKGROUND: While virus-vector-host interactions have been a major focus of both basic and applied ecological research, little is known about how different levels of plant defense interact with prior herbivory to affect these relationships. We used genetically-modified strains of tomato (Solanum lycopersicum) varying in the jasmonic acid (JA) plant defense pathways to explore how plant defense and prior herbivory affects a plant virus (tomato yellow leaf curl virus, 'TYLCV'), its vector (the whitefly Bemisia tabaci MED), and the host. RESULTS: Virus-free MED preferred low-JA over high-JA plants and had lower fitness on high-JA plants. Viruliferous MED preferred low-JA plants but their survival was unaffected by JA levels. While virus-free MED did not lower plant JA levels, viruliferous MED decreased both JA levels and the expression of JA-related genes. Infestation by viruliferous MED reduced plant JA levels. In preference tests, neither virus-free nor viruliferous MED discriminated among JA-varying plants previously exposed to virus-free MED. However, both virus-free and viruliferous MED preferred low-JA plant genotypes when choosing between plants that had both been previously exposed to viruliferous MED. The enhanced preference for low-JA genotypes appears linked to the volatile compound neophytadiene, which was found only in whitefly-infested plants and at concentrations inversely related to plant JA levels. CONCLUSIONS: Our findings illustrate how plant defense can interact with prior herbivory to affect both a plant virus and its whitefly vector, and confirm the induction of neophytadiene by MED. The apparent attraction of MED to neophytadiene may prove useful in pest detection and management.


Assuntos
Antibiose , Begomovirus/fisiologia , Ciclopentanos/metabolismo , Hemípteros/fisiologia , Herbivoria , Lycopersicon esculentum/fisiologia , Oxilipinas/metabolismo , Doenças das Plantas/virologia , Animais , Lycopersicon esculentum/imunologia , Lycopersicon esculentum/virologia , Imunidade Vegetal , Plantas Geneticamente Modificadas/imunologia , Plantas Geneticamente Modificadas/fisiologia , Plantas Geneticamente Modificadas/virologia , Transdução de Sinais
6.
PLoS One ; 14(12): e0226783, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31869401

RESUMO

Cassava production in Africa is constrained by cassava mosaic disease (CMD) that is caused by the Cassava mosaic virus (CMV). The aim of this study was to evaluate the responses of a range of commonly cultivated West African cassava cultivars to varying inoculum doses of African cassava mosaic virus (ACMV). We grafted 10 cultivars of cassava plants with different inoculum doses of CMV (namely two, four, or six CMD-infected buds) when the experimental plants were 8, 10, or 12 weeks old, using non-inoculated plants as controls. Three cultivars showed disease symptoms when grafted with two buds, and four cultivars showed disease symptoms when grafted with four or six buds. Most cultivars became symptomatic six weeks after inoculation, but one ('TMS92/0326') was symptomatic two weeks after inoculation, and two ('Ntollo' and 'Excel') were symptomatic after four weeks. Root weight tended to be lower in the six-bud than in the two-bud dose, and disease severity varied with plant age at inoculation. These results indicate that the level of CMD resistance in cassava cultivars varies with inoculum dose and timing of infection. This will allow appropriate cultivars to be deployed in each production zone of Africa in accordance with the prevalence of CMD.


Assuntos
Begomovirus/fisiologia , Manihot/fisiologia , Manihot/virologia , Doenças das Plantas/virologia , África , Resistência à Doença , Manihot/anatomia & histologia , Raízes de Plantas/anatomia & histologia , Raízes de Plantas/fisiologia , Raízes de Plantas/virologia
7.
Virol J ; 16(1): 130, 2019 11 07.
Artigo em Inglês | MEDLINE | ID: mdl-31699111

RESUMO

BACKGROUND: MicroRNAs (miRNAs) are a class of 21-24 nucleotide endogenous non-coding small RNAs that play important roles in plant development and defense responses to biotic and abiotic stresses. Tobacco curly shoot virus (TbCSV) is a monopartite begomovirus, cause leaf curling and plant stunting symptoms in many Solanaceae plants. The betasatellite of TbCSV (TbCSB) induces more severe symptoms and enhances virus accumulation when co-infect the plants with TbCSV. METHODS: In this study, miRNAs regulated by TbCSV and TbCSB co-infection in Nicotiana benthamiana were characterized using high-throughput sequencing technology. RESULTS: Small RNA sequencing analysis revealed that a total of 13 known miRNAs and 42 novel miRNAs were differentially expressed in TbCSV and TbCSB co-infected N. benthamiana plants. Several potential miRNA-targeted genes were identified through data mining and were involved in both catalytic and metabolic processes, in addition to plant defense mechanisms against virus infections according to Gene Ontology (GO) analyses. In addition, the expressions of several differentially expressed miRNAs and their miRNA-targeted gene were validated through quantitative real time polymerase chain reaction (qRT-PCR) approach. CONCLUSIONS: A large number of miRNAs are identified, and their target genes, functional annotations also have been explored. Our results provide the information on N. benthamiana miRNAs and would be useful to further understand miRNA regulatory mechanisms after TbCSV and TbCSB co-infection.


Assuntos
Begomovirus/fisiologia , MicroRNAs/genética , RNA de Plantas/genética , Vírus Satélites/fisiologia , Tabaco/genética , Tabaco/virologia , Coinfecção , Regulação da Expressão Gênica de Plantas , Genes de Plantas/genética , MicroRNAs/metabolismo , Doenças das Plantas/genética , Doenças das Plantas/virologia , RNA de Plantas/metabolismo
8.
Plant Dis ; 103(11): 2913-2919, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31436474

RESUMO

Thirty-one melon accessions were screened for resistance to the begomoviruses Melon chlorotic mosaic virus (MeCMV) and Tomato leaf curl New Delhi virus (ToLCNDV). Five accessions presented nearly complete resistance to both viruses. Accession IC-274014, showing the highest level of resistance to both viruses, was crossed with the susceptible cultivar Védrantais. The F1, F2, F3/F4, and both backcross progenies were mechanically inoculated with MeCMV. Plants without symptoms or virus detection by enzyme-linked immunosorbent assay and/or PCR were considered as resistant. The segregations were compatible with two recessive and one dominant independent genes simultaneously required for resistance. Inheritance of resistance to ToLCNDV in the F2 was best explained by one recessive gene and two independent dominant genes simultaneously required. Some F3 and F4 families selected for resistance to MeCMV also were resistant to ToLCNDV, suggesting that common or tightly linked genes were involved in resistance to both viruses. We propose the names begomovirus resistance-1 and Begomovirus resistance-2 for these genes (symbols bgm-1 and Bgm-2). Resistance to MeCMV in IC-274014 was controlled by bgm-1, Bgm-2, and the recessive gene melon chlorotic mosaic virus resistance (mecmv); resistance to ToLCNDV was controlled by bgm-1, Bgm-2, and the dominant gene Tomato leaf curl New Delhi virus resistance (Tolcndv).


Assuntos
Begomovirus , Cucurbitaceae , Resistência à Doença , Begomovirus/fisiologia , Cucurbitaceae/virologia , Resistência à Doença/genética
9.
BMC Genomics ; 20(1): 654, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31416422

RESUMO

BACKGROUND: Cucurbit yellow stunting disorder virus (CYSDV; genus Crinivirus, Closteroviridae) is transmitted in a semipersistent manner by the whitefly, Bemisia tabaci, and is efficiently transmitted by the widely prevalent B. tabaci cryptic species, MEAM1. In this study, we compared transcriptome profiles of B. tabaci MEAM1, after 24 h, 72 h and 7 days of acquisition feeding on melon plants infected with CYSDV (CYSDV-whiteflies) with those fed on virus-free melon, using RNA-Seq technology. We also compared transcriptome profiles with whiteflies fed on tomato plants separately infected with Tomato chlorosis virus (ToCV), a crinivirus closely related to CYSDV, and Tomato yellow leaf curl virus (TYLCV), a member of the genus Begomovirus, which has a distinctly different mode of transmission and their respective virus-free controls, to find common gene expression changes among viruliferous whiteflies feeding on different host plants infected with distinct (TYLCV) and related (CYSDV and ToCV) viruses. RESULTS: A total of 275 differentially expressed genes (DEGs) were identified in CYSDV-whiteflies, with 3 DEGs at 24 h, 221 DEGs at 72 h, and 51 DEGs at 7 days of virus acquisition. Changes in genes encoding orphan genes (54 genes), phosphatidylethanolamine-binding proteins (PEBP) (20 genes), and AAA-ATPase domain containing proteins (10 genes) were associated with the 72 h time point. Several more orphan genes (20 genes) were differentially expressed at 7 days. A total of 59 common DEGs were found between CYSDV-whiteflies and ToCV-whiteflies, which included 20 orphan genes and 6 lysosomal genes. A comparison of DEGs across the three different virus-host systems revealed 14 common DEGs, among which, eight showed similar and significant up-regulation in CYSDV-whiteflies at 72 h and TYLCV-whiteflies at 24 h, while down-regulation of the same genes was observed in ToCV-whiteflies at 72 h. CONCLUSIONS: Dynamic gene expression changes occurred in CYSDV-whiteflies after 72 h feeding, with decreased gene expression changes associated with 7 days of CYSDV acquisition. Similarities in gene expression changes among CYSDV-whiteflies, ToCV-whiteflies and TYLCV-whiteflies suggest the possible involvement of common genes or pathways for virus acquisition and transmission by whiteflies, even for viruses with distinctly different modes of transmission.


Assuntos
Crinivirus/fisiologia , Cucurbitaceae/virologia , Hemípteros/metabolismo , Doenças das Plantas/virologia , Animais , Begomovirus/fisiologia , Regulação da Expressão Gênica , Hemípteros/genética , Hemípteros/virologia , Lycopersicon esculentum/virologia , RNA-Seq , Fatores de Tempo , Transcriptoma
10.
J Agric Food Chem ; 67(33): 9241-9253, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31369258

RESUMO

Antiviral compounds targeting viral replicative processes have been studied as an alternative for the control of begomoviruses. Previously, we have reported that the peptide AmPep1 has strong affinity binding to the replication origin sequence of tomato yellow leaf curl virus (TYLCV). In this study, we describe the mechanism of action of this peptide as a novel alternative for control of plant-infecting DNA viruses. When AmPep1 was applied exogenously to tomato and Nicotiana benthamiana plants infected with TYLCV, a decrease in the synthesis of the two viral DNA strands (CS and VS) was observed, with a consequent delay in the development of disease progress in treated plants. The chemical mechanism of action of AmPep1 was deduced using Raman spectroscopy and molecular modeling showing the formation of chemical interactions such as H bonds and electrostatic interactions and the formation of π-π interactions between both biomolecules contributing to tampering with the viral replication.


Assuntos
Amaranthus/química , Antivirais/química , Antivirais/farmacologia , Begomovirus/efeitos dos fármacos , Peptídeos/química , Peptídeos/farmacologia , RNA Viral/química , Replicação Viral/efeitos dos fármacos , Begomovirus/química , Begomovirus/genética , Begomovirus/fisiologia , Sequências Repetidas Invertidas/efeitos dos fármacos , Lycopersicon esculentum/virologia , Doenças das Plantas/virologia , Proteínas de Plantas/química , RNA Viral/genética , Tabaco/virologia
11.
J Biol Dyn ; 13(sup1): 325-353, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31149889

RESUMO

Vector-transmitted diseases of plants have had devastating effects on agricultural production worldwide, resulting in drastic reductions in yield for crops such as cotton, soybean, tomato, and cassava. Plant-vector-virus models with continuous replanting are investigated in terms of the effects of selection of cuttings, roguing, and insecticide use on disease prevalence in plants. Previous models are extended to include two replanting strategies: frequencyreplanting and abundance-replanting. In frequency-replanting, replanting of infected cuttings depends on the selection frequency parameter ε, whereas in abundance-replanting, replanting depends on plant abundance via a selection rate parameter also denoted as ε. The two models are analysed and new thresholds for disease elimination are defined for each model. Parameter values for cassava, whiteflies, and African cassava mosaic virus serve as a case study. A numerical sensitivity analysis illustrates how the equilibrium densities of healthy and infected plants vary with parameter values. Optimal control theory is used to investigate the effects of roguing and insecticide use with a goal of maximizing the healthy plants that are harvested. Differences in the control strategies in the two models are seen for large values of ε. Also, the combined strategy of roguing and insecticide use performs better than a single control.


Assuntos
Agricultura/métodos , Produtos Agrícolas/virologia , Vetores de Doenças , Modelos Biológicos , Doenças das Plantas/prevenção & controle , Doenças das Plantas/virologia , Animais , Begomovirus/fisiologia , Hemípteros/fisiologia , Inseticidas/toxicidade , Manihot/parasitologia , Manihot/virologia , Análise Numérica Assistida por Computador
12.
Virology ; 531: 240-247, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30933715

RESUMO

Begomoviruses contain some of the most damaging viral disease agents of crops worldwide, and are transmitted by whiteflies of the Bemisia tabaci species complex. During the last 20 years, transovarial transmission of tomato yellow leaf curl virus (TYLCV) has been reported in two invasive species of the B. tabaci complex. To further decipher the importance of this mode of transmission, we analyzed transovarial transmission of TYLCV by seven whitefly species indigenous to China. TYLCV virions were detected in eggs of all species except one, and in nymphs of two species, but in none of the ensuing adults of all seven species. Our results suggest that these indigenous whiteflies are unable to transmit TYLCV, a begomovirus alien to China, via ova to produce future generations of viruliferous adults, although most of the species exhibit varying ability to carry over the virus to the eggs/nymphs of their offspring via transovarial transmission.


Assuntos
Begomovirus/fisiologia , Hemípteros/fisiologia , Hemípteros/virologia , Insetos Vetores/fisiologia , Insetos Vetores/virologia , Lycopersicon esculentum/virologia , Doenças das Plantas/virologia , Animais , China , Hemípteros/classificação , Hemípteros/genética , Insetos Vetores/classificação , Insetos Vetores/genética
13.
Plant Dis ; 103(6): 1181-1188, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30908127

RESUMO

Epidemics of tomato yellow leaf curl disease (TYLCD) caused by tomato yellow leaf curl-like begomoviruses (genus Begomovirus, family Geminiviridae) severely damage open field and protected tomato crops worldwide. Intensive application of insecticides against the whitefly vector Bemisia tabaci is generally used as control strategy to reduce TYLCD impact. This practice, however, is frequently ineffective and has a negative impact on the environment and human health. TYLCD-resistant varieties are commercially available, but cultivation of susceptible traditional tasting ones is also requested if possible. For susceptible tomatoes, here we show that using whitefly optical barriers by means of UV-blocking plastics in protected crops can contribute to reducing TYLCD damage and increasing commercial fruit yield. Moreover, induction of systemic acquired resistance by application of the elicitor of plant defense acibenzolar-S-methyl was effective to reduce yield losses when viral pressure was moderate. Interestingly, combining both practices in protected tomato crops can result in a significant TYLCD control. Therefore, these control practices are proposed to be used commercially as management alternatives to include in integrated management of TYLCD.


Assuntos
Begomovirus , Resistência à Doença , Hemípteros , Lycopersicon esculentum , Animais , Begomovirus/fisiologia , Produtos Agrícolas/virologia , Hemípteros/virologia , Lycopersicon esculentum/virologia
14.
Acta Virol ; 63(1): 26-35, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30879310

RESUMO

Whitefly-transmitted viruses of the genus Begomovirus (the family Geminiviridae) have become a limiting factor for agricultural productivity in many warmer parts of the world. The economies of Pakistan and India have, since the early 1990s, suffered losses due to cotton leaf curl disease (CLCuD). The disease is caused by begomoviruses, the most important of which at this time is cotton leaf curl Kokhran virus strain Burewala (CLCuKoV-Bu), and a disease-specific betasatellite, cotton leaf curl Multan betasatellite (CLCuMuB). Efforts to minimize losses due to CLCuD rely mainly on the use of insecticides to kill the whitefly vector; no resistant cotton varieties are currently commercially available. The study described here has investigated RNA interference technology for its potential to yield resistance against CLCuKoV-Bu and three other begomoviruses; CLCuKoV, tomato leaf curl New Delhi virus (ToLCNDV) and Pedilanthus leaf curl virus (PeLCV). Three fragments of the virion-sense V2 gene of CLCuKoV-Bu were transformed into Nicotiana benthamiana in antisense orientation and transgenic lines expressing virus-specific short RNAs were assessed for their ability to yield resistance. Only CLCuKoV-Bu with the V2 sequence closest to the promoter was resistant. Inoculation of CLCuKoV-Bu with CLCuMuB into transgenic plants did not significantly affect the outcome, although viral DNA was detected in number of plants, suggesting that the betasatellite may impair RNAi resistance. Overall the results indicate that targeting the 5' end of V2 gene using antisense-RNA has the potential to deliver resistance against begomoviruses and that RNAi-based resistance imparts some degree of resistance to heterologous viruses. Keywords: geminivirus; begomovirus; RNAi; resistance; CLCuKoV-Burewala; CLCuMuB.


Assuntos
Begomovirus , Resistência à Doença , Tabaco , Begomovirus/genética , Begomovirus/fisiologia , DNA Viral/genética , Resistência à Doença/genética , Doenças das Plantas/genética , Doenças das Plantas/virologia , Plantas Geneticamente Modificadas , Interferência de RNA , Tabaco/genética , Tabaco/virologia
15.
Acta Virol ; 63(1): 36-44, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30879311

RESUMO

Alphasatellites, formerly known as DNA 1, are a satellite-like components associated with begomoviruses (the family Geminiviridae) that require betasatellite for symptom induction but depend on DNA-A for systemic movement. We have converted alphasatellite into gene-silencing vector (modified alphasatellite (∆DNA 1)) by deleting its A-rich region that does not affect the replication nor the movement of the helper virus. Insertion of a transgene green florescence protein (GFP) into ∆DNA 1 resulted in the silencing g of the cognate gene in Nicotiana benthamiana. The silencing persisted for more than one and half month and was associated with the decreased level of mRNA of the target gene. This satellite-like DNA vector induced gene silencing (VIGS) promises to be applicable to other begomovirus/alphasatellite systems, thereby providing the powerful approach to gene discovery and the analysis of gene functions in malvaceous crops. Keywords: cotton; begomovirus; alphasatellite; RNAi.


Assuntos
Begomovirus , Resistência à Doença , Inativação Gênica , Tabaco , Begomovirus/genética , Begomovirus/fisiologia , DNA Satélite/genética , Resistência à Doença/genética , Vetores Genéticos/genética , Tabaco/genética , Tabaco/virologia
16.
PLoS One ; 14(2): e0210011, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30730891

RESUMO

Cotton is a commercial and economically important crop that generates billions of dollars in annual revenue worldwide. However, cotton yield is affected by a sap-sucking insect Bemisia tabaci (whitefly), and whitefly-borne cotton leaf curl disease (CLCuD). The causative agent of devastating CLCuD is led by the viruses belonging to the genus Begomovirus (family Geminiviridae), collectively called cotton leaf curl viruses. Unfortunately, the extensively cultivated cotton (Gossypium hirsutum) species are highly susceptible and vulnerable to CLCuD. Yet, the concomitant influence of whitefly and CLCuD on the susceptible G. hirsutum transcriptome has not been interpreted. In the present study we have employed an RNA Sequencing (RNA-Seq) transcriptomics approach to explore the differential gene expression in susceptible G. hirsutum variety upon infection with viruliferous whiteflies. Comparative RNA-Seq of control and CLCuD infected plants was done using Illumina HiSeq 2500. This study yielded 468 differentially expressed genes (DEGs). Among them, we identified 220 up and 248 downregulated DEGs involved in disease responses and pathogen defense. We selected ten genes for downstream RT-qPCR analyses on two cultivars, Karishma and MNH 786 that are susceptible to CLCuD. We observed a similar expression pattern of these genes in both susceptible cultivars that was also consistent with our transcriptome data further implying a wider application of our global transcription study on host susceptibility to CLCuD. We next performed weighted gene co-expression network analysis that revealed six modules. This analysis also identified highly co-expressed genes as well as 55 hub genes that co-express with ≥ 50 genes. Intriguingly, most of these hub genes are shown to be downregulated and enriched in cellular processes. Under-expression of such highly co-expressed genes suggests their roles in favoring the virus and enhancing plant susceptibility to CLCuD. We also discuss the potential mechanisms governing the establishment of disease susceptibility. Overall, our study provides a comprehensive differential gene expression analysis of G. hirsutum under whitefly-mediated CLCuD infection. This vital study will advance the understanding of simultaneous effect of whitefly and virus on their host and aid in identifying important G. hirsutum genes which intricate in its susceptibility to CLCuD.


Assuntos
Begomovirus/fisiologia , Gossypium/genética , Gossypium/virologia , Hemípteros/fisiologia , Doenças das Plantas/genética , Doenças das Plantas/virologia , Transcriptoma , Animais , Resistência à Doença , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Redes Reguladoras de Genes , Genes de Plantas , Insetos Vetores/fisiologia
17.
Viruses ; 11(1)2019 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-30669683

RESUMO

Using double-strand RNA (dsRNA) high-throughput sequencing, we identified five RNA viruses in a bean golden mosaic virus (BGMV)-resistant common bean transgenic line with symptoms of viral infection. Four of the identified viruses had already been described as infecting common bean (cowpea mild mottle virus, bean rugose mosaic virus, Phaseolus vulgaris alphaendornavirus 1, and Phaseolus vulgaris alphaendornavirus 2) and one is a putative new plant rhabdovirus (genus Cytorhabdovirus), tentatively named bean-associated cytorhabdovirus (BaCV). The BaCV genome presented all five open reading frames (ORFs) found in most rhabdoviruses: nucleoprotein (N) (ORF1) (451 amino acids, aa), phosphoprotein (P) (ORF2) (445 aa), matrix (M) (ORF4) (287 aa), glycoprotein (G) (ORF5) (520 aa), and an RNA-dependent RNA polymerase (L) (ORF6) (114 aa), as well as a putative movement protein (P3) (ORF3) (189 aa) and the hypothetical small protein P4. The predicted BaCV proteins were compared to homologous proteins from the closest cytorhabdoviruses, and a low level of sequence identity (15⁻39%) was observed. The phylogenetic analysis shows that BaCV clustered with yerba mate chlorosis-associated virus (YmCaV) and rice stripe mosaic virus (RSMV). Overall, our results provide strong evidence that BaCV is indeed a new virus species in the genus Cytorhabdovirus (family Rhabdoviridae), the first rhabdovirus to be identified infecting common bean.


Assuntos
Begomovirus/fisiologia , Phaseolus/virologia , Doenças das Plantas/virologia , Vírus de RNA/isolamento & purificação , RNA de Cadeia Dupla/genética , Rhabdoviridae/isolamento & purificação , Resistência à Doença , Genoma Viral , Sequenciamento de Nucleotídeos em Larga Escala , Fases de Leitura Aberta , Filogenia , Folhas de Planta/virologia , Plantas Geneticamente Modificadas/virologia , Vírus de RNA/classificação , RNA Viral/genética , Rhabdoviridae/classificação , Análise de Sequência de DNA , Proteínas Virais/genética
18.
Virology ; 529: 81-90, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30684693

RESUMO

One geminiviral gene encodes the capsid protein (CP), which can appear as several bands after electrophoresis depending on virus and plant. African cassava mosaic virus-Nigeria CP in Nicotiana benthamiana, however, yielded one band (~ 30 kDa) in total protein extracts and purified virions, although its expression in yeast yielded two bands (~ 30, 32 kDa). Mass spectrometry of the complete protein and its tryptic fragments from virions is consistent with a cleaved start M1, acetylated S2, and partial phosphorylation at T12, S25 and S62. Mutants for additional potentially modified sites (N223A; C235A) were fully infectious and formed geminiparticles. Separation in triton acetic acid urea gels confirmed charge changes of the CP between plants and yeast indicating differential phosphorylation. If the CP gene alone was expressed in plants, multiple bands were observed like in yeast. A high turnover rate indicates that post-translational modifications promote CP decay probably via the ubiquitin-triggered proteasomal pathway.


Assuntos
Begomovirus/fisiologia , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Replicação Viral , Sequência de Aminoácidos , DNA Viral , Modelos Moleculares , Fosforilação , Doenças das Plantas/virologia , Conformação Proteica , Isoformas de Proteínas
19.
Genomics ; 111(6): 1333-1342, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-30237075

RESUMO

Phaseolus vulgaris is an economically important legume in tropical and subtropical regions of Asia, Africa, Latin-America and parts of USA and Europe. However, its production gets severely affected by mungbean yellow mosaic India virus (MYMIV). We aim to identify and characterize differentially expressed miRNAs during MYMIV-infection in P. vulgaris. A total of 422 miRNAs are identified of which 292 are expressed in both MYMIV-treated and mock-treated samples, 109 are expressed only in MYMIV-treated and 21 are expressed only in mock-treated samples. Selected up- and down-regulated miRNAs are validated by RT-qPCR. 3367 target ORFs are identified for 270 miRNAs. Selected targets are validated by 5' RLM-RACE. Differentially expressed miRNAs regulate transcription factors and are involved in improving stress tolerance to MYMIV. These findings will provide an insight into the role of miRNAs during MYMIV infection in P. vulgaris in particular and during any biotic stress conditions in Leguminosae family in general.


Assuntos
Begomovirus/fisiologia , Interações Hospedeiro-Patógeno/genética , MicroRNAs/metabolismo , Phaseolus/genética , Phaseolus/virologia , Doenças das Plantas/virologia , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , MicroRNAs/fisiologia , Doenças das Plantas/genética , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de RNA
20.
Arch Insect Biochem Physiol ; 100(2): e21503, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30570176

RESUMO

In transmitting plant viruses, insect vectors undergo physiological and behavioral alterations. The whitefly Bemisia tabaci is a vector of tomato yellow leaf curl virus (TYLCV), causing severe damages to various horticultural crop plants. To determine whether whitefly alteration is specific to vector species, the responses to TYLCV ingestion were compared between B. tabaci and Trialeurodes vaporariorum, a nonvector for TYLCV. The two species were reared on TYLCV-infected and noninfected tomato, a host of TYLCV, and their longevity and fecundity were determined while rearing in either tomato or eggplant, a nonhost of TYLCV. TYLCV-ingested B. tabaci increased their developmental rates but reduced fecundity when they were reared in either tomato or eggplant compared with those of TYLCV-free ones. In contrast, TYLCV-ingested T. vaporariorum did not show any of the aforementioned changes when reared on both plant species. In addition, TYLCV-ingested B. tabaci increased their levels of three heat shock protein genes ( hsp20, hsp70, and hsp90) against thermal stress, whereas TYLCV-ingested T. vaporariorum did not. The presence of TYLCV virions was identified in two colonies of both species via polymerase chain reaction analysis. TYLCV was detected in the whole body, saliva, and eggs of B. tabaci, while TYLCV was detected only in the whole body but not in the saliva and eggs of T. vaporariorum. The present results strongly indicated that TYLCV specifically manipulate physiological processes of the vector species, B. tabaci.


Assuntos
Begomovirus/fisiologia , Hemípteros/virologia , Animais , Interações Hospedeiro-Patógeno , Insetos Vetores , Lycopersicon esculentum
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