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Vicente Pallas - One of the best experts on this subject based on the ideXlab platform.
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Polyvalent detection of twelve Viruses and four Viroids affecting tomato by using a unique polyprobe
European Journal of Plant Pathology, 2019Co-Authors: Jesús Ángel Sánchez-navarro, Lorena Corachán, Isabel Font, Ana Alfaro-fernández, Vicente PallasAbstract:Non-radioactive molecular hybridization represents an attractive approach for the detection of multiple plant virus and/or Viroids and a good alternative to the more extended serological and PCR-based detection methods. The use of polyprobes or riboprobes carrying partial sequences of different plant Viruses or Viroids fused in tandem, has permitted the detection of up to 10 different pathogens or the development of genus-specific probes. In the present article, the polyprobe technology has been adapted for the detection of the main Viruses and Viroids affecting tomato crops. To do this, three polyprobes have been developed covering four Viroids (Poly4), twelve Viruses (Poly12) or the four Viroids plus the twelve Viruses (poly16). The detection limit of the three polyprobes was comparable to the individual probes allowing the detection of up to 0,2 pg/μl of viral or viroidal RNA. A survey of 50 field samples revealed that all positive samples detected with the individual probes were also detected with the corresponding poly12 (98%) or poly16 (100%) probes. The analysis of tomato seeds revealed that both, single and polyprobes, were able to detect an infected seed in a pool of 250 healthy seeds. Finally, a ring-test analysis among six laboratories revealed a high reproducibility of the non-radioactive molecular hybridization procedure using the three polyprobes. The use of this technology in the routine analysis of tomato samples is discussed.
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recent advances on the multiplex molecular detection of plant Viruses and Viroids
Frontiers in Microbiology, 2018Co-Authors: Vicente Pallas, J A Sancheznavarro, D JamesAbstract:Plant Viruses are still one of the main contributors to economic losses in agriculture. It has been estimated that plant Viruses can cause as much as 50 billion euros loss worldwide, per year. This situation may be worsened by recent climate change events and the associated changes in disease epidemiology. Reliable and early detection methods are still one of the main and most effective actions to develop control strategies for plant viral diseases. During the last years, considerable progress has been made to develop tools with high specificity and low detection limits for use in the detection of these plant pathogens. Time and cost reductions have been some of the main objectives pursued during the last few years as these increase their feasibility for routine use. Among other strategies, these objectives can be achieved by the simultaneous detection and (or) identification of several Viruses in a single assay. Nucleic acid-based detection techniques are especially suitable for this purpose. Polyvalent detection has allowed the detection of multiple plant Viruses at the genus level. Multiplexing RT polymerase chain reaction (PCR) has been optimized for the simultaneous detection of more than 10 plant Viruses/Viroids. In this short review, we provide an update on the progress made during the last decade on techniques such as multiplex PCR, polyvalent PCR, non-isotopic molecular hybridization techniques, real-time PCR, and array technologies to allow simultaneous detection of multiple plant Viruses. Also, the potential and benefits of the powerful new technique of deep sequencing/next-generation sequencing are described.
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survey of stone fruit Viruses and Viroids in chile
Journal of Plant Pathology, 2016Co-Authors: Nicola Fiore, J A Sancheznavarro, Alan Zamorano, Ana Maria Pino, F Gonzalez, I M Rosales, Vicente PallasAbstract:Stone fruits orchards from three Chilean regions were visited to collect leaf samples which were tested for the presence of the most important Viruses and Viroids. Molecular nonradioactive hybridization (MH) testing of 2,456 samples and confirmatory RT-PCR of some MH-negative samples gave the following infection rates: 31.5% for Prunus necrotic ringspot virus (PNRSV); 31.3% for Prune dwarf virus (PDV); 25.7% for Peach latent mosaic viroid (PLMVd); 3.1% for Tomato ringspot virus (ToRSV) only in rootstocks; 1.8% for Plum pox virus (PPV); 1.2% for Hop stunt viroid (HSVd); 1.1% for Apple mosaic virus (ApMV); and 0.9% for Apple chlorotic leaf spot virus (ACLSV). American plum line pattern virus (APLPV) was not detected. The overall infection rate in the surveyed orchards was 29.6%, specifically 28.2% (Region Metropolitana), 23.4% (Valparaiso) and 33% (Bernardo O’Higgins) in the three regions surveyed. ApMV and HSVd, detected only in peach and nectarine, are new records in stone fruits in Chile.
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Simultaneous detection of eight Viruses and two Viroids affecting stone fruit trees by using a unique polyprobe
European Journal of Plant Pathology, 2012Co-Authors: Ana Peiró, Vicente Pallas, Jesús Ángel Sánchez-navarroAbstract:The use of riboprobes carrying partial sequences of different plant Viruses or Viroids fused in tandem, has permitted the simultaneous detection of up to six different pathogens using a non-radioactive molecular hybridization procedure. In the present report, we describe the development of a unique polyprobe (poly10) with the capacity to detect Viruses and Viroids commonly found infecting fruit trees. The poly10 covers eight Viruses: Apple mosaic virus (ApMV), Prunus necrotic ringspot virus (PNRSV), Prune dwarf virus (PDV), American plum line pattern (APLPV), Plum pox virus (PPV), Apple chlorotic leaf spot virus (ACLSV), Apricot latent virus (ApLV), Plum bark necrosis and stem pitting-associated virus (PBNSPaV) and two Viroids: Hop stunt viroid (HSVd) and Peach latent mosaic viroid (PLMVd). Poly10 is comparable to the individual riboprobes in terms of end-point dilution limit and specificity, allowing the detection of up to 2.5 picograms of viral or viroidal RNA. However, the polyprobe requires a hybridization temperature of 60°C instead of the standard 68°C. The validation of the new simultaneous detection strategy was confirmed by the analysis of 60 field samples, which came from seven different hosts. The use of the polyprobe as an alternative to other routinely used detection methods is discussed.
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systematic search for recombination events in plant Viruses and Viroids
1997Co-Authors: Thierry Candresse, Frederic Revers, Olivier Le Gall, Sandra A Kofalvi, Jose F Marcos, Vicente PallasAbstract:One of the key issues regarding the biosafety of transgenic plants expressing plant virus genetic information concerns the potential for recombination between transgene-derived sequences and an incoming, more or less related virus. With no precise information currently available on the rate of appearance and potential fitness of such recombinants, the hypothesis that all viable possible recombinants have already been produced and selected in nature in the course of natural multiple infections has been the subject of a debate. One way to further our understanding of these mechanisms is to perform systematic searches, in order to identify natural viral isolates deriving from a recombination event and, therefore, make a first evaluation of the frequency of recombinant isolates in natural viral populations.
Hoseong Choi - One of the best experts on this subject based on the ideXlab platform.
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identification of Viruses and Viroids infecting tomato and pepper plants in vietnam by metatranscriptomics
International Journal of Molecular Sciences, 2020Co-Authors: Hoseong Choi, Won Kyong Cho, Phutri Tran, Lakha Salaipeth, Haeryun Kwak, Hongsoo Choi, Kookhyung KimAbstract:Tomato (Lycopersicum esculentum L.) and pepper (Capsicum annuum L.) plants belonging to the family Solanaceae are cultivated worldwide. The rapid development of next-generation sequencing (NGS) technology facilitates the identification of Viruses and Viroids infecting plants. In this study, we carried out metatranscriptomics using RNA sequencing followed by bioinformatics analyses to identify Viruses and Viroids infecting tomato and pepper plants in Vietnam. We prepared a total of 16 libraries, including eight tomato and eight pepper libraries derived from different geographical regions in Vietnam. We identified a total of 602 virus-associated contigs, which were assigned to 18 different virus species belonging to nine different viral genera. We identified 13 different Viruses and two Viroids infecting tomato plants and 12 Viruses and two Viroids infecting pepper plants with Viruses as dominantly observed pathogens. Our results showed that multiple infection of different viral pathogens was common in both plants. Moreover, geographical region and host plant were two major factors to determine viral populations. Taken together, our results provide the comprehensive overview of viral pathogens infecting two important plants in the family Solanaceae grown in Vietnam.
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In silico identification of Viruses and Viroids infecting grapevine cultivar cabernet sauvignon using a grapevine transcriptome
Journal of Plant Pathology, 2018Co-Authors: Yeonhwa Jo, Hoseong Choi, Myung-kyu Song, Jae-seong ParkAbstract:Next-generation sequencing (NGS)-based studies have reported a large amount of transcriptome data. In fact, plant transcriptome data sometimes contain sequence reads associated with Viruses and Viroids. Recently, we screened several grapevine transcriptomes to identify Viruses and Viroids that infect grapevine and we selected a transcriptome of the grapevine cultivar Cabernet Sauvignon for an in silico data analysis. After de novo assembly of raw data, the obtained contigs were blasted against a virus reference database. We identified five Viruses and two Viroids that infected the grapevine. We discovered that 2.16% of the transcriptome was comprised of viral RNAs, with more than half of the viral reads deriving from grapevine rupestris stem pitting-associated virus. However, the RNA populations of the two Viroids were more than half of the total viral population based on the viral RNA copy number. Moreover, we identified single nucleotide polymorphisms for each virus and viroid and performed phylogenetic analyses. Taken together, we report the successful application of plant transcriptome data using bioinformatics analyses in the identification of five Viruses and two Viroids in the grapevine cultivar Cabernet Sauvignon, which is one of the most important grapevine cultivars in the world.
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Peach RNA viromes in six different peach cultivars
Scientific Reports, 2018Co-Authors: Yeonhwa Jo, Ju-yeon Yoon, Sen Lian, Hoseong Choi, Seung-kook ChoiAbstract:Many recent studies have demonstrated that several known and unknown Viruses infect many horticultural plants. However, the elucidation of a viral population and the understanding of the genetic complexity of viral genomes in a single plant are rarely reported. Here, we conducted metatranscriptome analyses using six different peach trees representing six individual peach cultivars. We identified six Viruses including five Viruses in the family Betaflexiviridae and a novel virus belonging to the family Tymoviridae as well as two Viroids. The number of identified Viruses and Viroids in each transcriptome ranged from one to six. We obtained 18 complete or nearly complete genomes for six Viruses and two Viroids using transcriptome data. Furthermore, we analyzed single nucleotide variations for individual viral genomes. In addition, we analyzed the amount of viral RNA and copy number for identified Viruses and Viroids. Some Viruses or Viroids were commonly present in different cultivars; however, the list of infected Viruses and Viroids in each cultivar was different. Taken together, our study reveals the viral population in a single peach tree and a comprehensive overview for the diversities of viral communities in different peach cultivars.
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in silico approach to reveal viral populations in grapevine cultivar tannat using transcriptome data
Scientific Reports, 2015Co-Authors: Hoseong Choi, Ju-yeon Yoon, Seung-kook Choi, Jin Kyong Cho, Won Kyong ChoAbstract:Viruses are ubiquitous and present in a wide range of settings, from living organisms to various environments. Although Viruses are regarded as important pathogens in higher plants, viral populations in specific host plants have not yet been fully examined. This study revealed viral populations in grape berries obtained from a cultivar from a single vineyard using currently available grapevine transcriptomes. Eight Viruses and two Viroids were identified using 11 grapevine libraries. Virus-associated sequences in each transcriptome ranged from 0.2% (seed) to 8.8% (skin). The amount of viral RNAs and virus copy numbers was quantified, thus revealing the dominant virus or viroid in each individual library. In addition, five viral genomes were successfully assembled de novo using transcriptome data. Phylogenetic analyses revealed that the Viruses and Viroids might have originated from Europe, along with the host. Single nucleotide variation studies revealed the quasispecies of RNA Viruses. Taken together, this study defines complex viral populations in three different grape tissues from a single vineyard.
Pallás Benet Vicente - One of the best experts on this subject based on the ideXlab platform.
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Polyvalent detection of twelve Viruses and four Viroids affecting tomato by using a unique polyprobe
Springer, 2020Co-Authors: Sánchez-navarro J. A., Corachán Lorena, Font Isabel, Alfaro-fernández Ana, Pallás Benet VicenteAbstract:Non-radioactive molecular hybridization represents an attractive approach for the detection of multiple plant virus and/or Viroids and a good alternative to the more extended serological and PCR-based detection methods. The use of polyprobes or riboprobes carrying partial sequences of different plant Viruses or Viroids fused in tandem, has permitted the detection of up to 10 different pathogens or the development of genus-specific probes. In the present article, the polyprobe technology has been adapted for the detection of the main Viruses and Viroids affecting tomato crops. To do this, three polyprobes have been developed covering four Viroids (Poly4), twelve Viruses (Poly12) or the four Viroids plus the twelve Viruses (poly16). The detection limit of the three polyprobes was comparable to the individual probes allowing the detection of up to 0,2 pg/μl of viral or viroidal RNA. A survey of 50 field samples revealed that all positive samples detected with the individual probes were also detected with the corresponding poly12 (98%) or poly16 (100%) probes. The analysis of tomato seeds revealed that both, single and polyprobes, were able to detect an infected seed in a pool of 250 healthy seeds. Finally, a ring-test analysis among six laboratories revealed a high reproducibility of the non-radioactive molecular hybridization procedure using the three polyprobes. The use of this technology in the routine analysis of tomato samples is discussed.This work was supported by grant BIO2017–88321-R from the Spanish Dirección General de Investigación Científica y Técnica (DGICYT) and the Prometeo Program GV2015/010 from the Generalitat Valenciana
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Survey of Stone fruit Viruses and Viroids in Chile
EDIZIONE ETS PISA ITALY, 2016Co-Authors: Fiore Nicola, Zamorano A., Pino A.m., González F., Rosales I.m., Sanchez Navarro, Jesus Angel, Pallás Benet VicenteAbstract:[EN] Stone fruits orchards from three Chilean regions were visited to collect leaf samples which were tested for the presence of the most important Viruses and Viroids. Molecular nonradioactive hybridization (MH) testing of 2,456 samples and confirmatory RT-PCR of some MH-negative samples gave the following infection rates: 31.5% for Prunus necrotic ringspot virus (PNRSV); 31.3% for Prune dwarf virus (PDV); 25.7% for Peach latent mosaic viroid (PLMVd); 3.1% for Tomato ringspot virus (ToRSV) only in rootstocks; 1.8% for Plum pox virus (PPV); 1.2% for Hop stunt viroid (HSVd); 1.1% for Apple mosaic virus (ApMV); and 0.9% for Apple chlorotic leaf spot virus (ACLSV). American plum line pattern virus (APLPV) was not detected. The overall infection rate in the surveyed orchards was 29.6%, specifically 28.2% (Region Metropolitana), 23.4% (Valparaiso) and 33% (Bernardo O'Higgins) in the three regions surveyed. ApMV and HSVd, detected only in peach and nectarine, are new records in stone fruits in Chile.This work was funded in equal parts by Chilean scholarship program for graduate students, 21060757, CONICYT, Chile; Project CSIC 2004CL0030; Project SAG C4-89-14-15.Fiore, N.; Zamorano, A.; Pino, A.; González, F.; Rosales, I.; Sanchez Navarro, JA.; Pallás Benet, V. (2016). Survey of Stone fruit Viruses and Viroids in Chile. JOURNAL OF PLANT PATHOLOGY. 98(3):631-635. doi:10.4454/JPP.V98I3.032S63163598
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Simultaneous detection of eight Viruses and two Viroids affecting stone fruit trees by using a unique polyprobe
'Springer Science and Business Media LLC', 2012Co-Authors: Peiró Morell Ana, Pallás Benet Vicente, Sanchez Navarro, Jesus AngelAbstract:The use of riboprobes carrying partial sequences of different plant Viruses or Viroids fused in tandem, has permitted the simultaneous detection of up to six different pathogens using a non-radioactive molecular hybridization procedure. In the present report, we describe the development of a unique polyprobe (poly10) with the capacity to detect Viruses and Viroids commonly found infecting fruit trees. The poly10 covers eight Viruses: Apple mosaic virus (ApMV), Prunus necrotic ringspot virus (PNRSV), Prune dwarf virus (PDV), American plum line pattern (APLPV), Plum pox virus (PPV), Apple chlorotic leaf spot virus (ACLSV), Apricot latent virus (ApLV), Plum bark necrosis and stem pitting-associated virus (PBNSPaV) and two Viroids: Hop stunt viroid (HSVd) and Peach latent mosaic viroid (PLMVd). Poly10 is comparable to the individual riboprobes in terms of end-point dilution limit and specificity, allowing the detection of up to 2.5 picograms of viral or viroidal RNA. However, the polyprobe requires a hybridization temperature of 60 degrees C instead of the standard 68 degrees C. The validation of the new simultaneous detection strategy was confirmed by the analysis of 60 field samples, which came from seven different hosts. The use of the polyprobe as an alternative to other routinely used detection methods is discussed.A. P. M. is the recipient of a JAE-Pre contract from the Consejo Superior de Investigaciones Cientificas (CSIC). We thank L. Corachan for her excellent technical assistance. This work was supported by grant BIO2008-03528 from the Spanish granting agency DGICYT and by grant ACOMP/2010/214 from the Generalitat Valenciana.Peiró Morell, A.; Pallás Benet, V.; Sanchez Navarro, JA. (2012). Simultaneous detection of eight Viruses and two Viroids affecting stone fruit trees by using a unique polyprobe. European Journal of Plant Pathology. 132(4):469-475. doi:10.1007/s10658-011-9893-0S469475132
Maja Ravnikar - One of the best experts on this subject based on the ideXlab platform.
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a framework for the evaluation of biosecurity commercial regulatory and scientific impacts of plant Viruses and Viroids identified by ngs technologies
Frontiers in Microbiology, 2017Co-Authors: Sebastien Massart, Maja Ravnikar, Thierry Candresse, Christophe Lacomme, Lukas Predajna, Jeansebastien Reynard, Artemis Rumbou, Pasquale Saldarelli, Dijana Skoric, Eeva J VainioAbstract:Recent advances in high-throughput sequencing technologies and bioinformatics have generated huge new opportunities for discovering and diagnosing plant Viruses and Viroids. Plant virology has undoubtedly benefited from these new methodologies, but at the same time is also faced with substantial bottlenecks, namely the biological characterization of the newly discovered Viruses and the analysis of their impact at the biosecurity, commercial, regulatory and scientific levels. This paper proposes a scaled and progressive scientific framework for efficient biological characterization and risk assessment when a previously known or a new plant virus is detected by next generation sequencing (NGS) technologies. Four case studies are also presented to illustrate the need for such a framework, and to discuss the scenarios.
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Next generation sequencing for detection and discovery of plant Viruses and Viroids: Comparison of two approaches
Frontiers in Microbiology, 2017Co-Authors: Anja Pecman, Denis Kutnjak, Adrian Fox, Ian Adams, Ion Gutiérrez-aguirre, Neil Boonham, Maja RavnikarAbstract:Next generation sequencing (NGS) technologies are becoming routinely employed in different fields of virus research. Different sequencing platforms and sample preparation approaches, in the laboratories worldwide, contributed to a revolution in detection and discovery of plant Viruses and Viroids. In this work, we are presenting the comparison of two RNA sequence inputs (small RNAs versus ribosomal RNA depleted total RNA) for the detection of plant Viruses by Illumina sequencing. This comparison includes several Viruses, which differ in genome organization and Viroids from both known families. The results demonstrate the ability for detection and identification of a wide array of known plant Viruses/Viroids in the tested samples by both approaches. In general, yield of viral sequences was dependent on viral genome organization and the amount of viral reads in the data. A putative novel Cytorhabdovirus, discovered in this study, was only detected by analysing the data generated from ribosomal RNA depleted total RNA and not from the small RNA dataset, due to the low number of short reads in the latter. On the other hand, for the Viruses/Viroids under study, the results showed higher yields of viral sequences in small RNA pool for Viroids and Viruses with no RNA replicative intermediates (single stranded DNA Viruses).
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plant Viruses and Viroids in an aqueous environment survival water mediated transmission and detection
QBOL-EPPO conference on DNA barcoding and diagnostic methods for plant pests, 2012Co-Authors: Natasa Mehle, Jelena Ruscic, Ion Gutierrezaguirre, N Prezelj, D Delic, Urska Vidic, Matevž Rupar, Mladen Krajacic, Maja RavnikarAbstract:Hydroponic systems and intensive irrigation in horticulture, are widely used, and hold the potential for rapid and efficient spread of water- transmissible plant pathogens throughout the whole crop. Although numerous plant Viruses have been detected in aqueous environment, for many of them, the survival in water and the potential for direct transmission through irrigation water are still unknown. Therefore we decided to explore whether the water can be a source of infection with PepMV, PVY and PSTVd, that are relatively stable and contagious Viruses/viroid, and a serious threat to tomato and/or potato production. In irrigation waters, the Viruses are usually present in concentrations lower than the detection limit of classical methods, but that may be sufficient to infect plants ; therefore the development of highly sensitive diagnostic methods is necessary. Small sample volumes may lead to non-representative testing, therefore, a concentration step allowing the handling of larger water volumes may improve water monitoring diagnostic scheme. A fast and efficient way to concentrate highly diluted Viruses consists on using CIM monolithic chromatographic supports. The application of such strategy to the above mentioned plant Viruses/viroid and the results of survival in water at room temperature and transmition efficiency through nutrient solutions in hydroponic cultivation of tomatoes and potatoes will be presented.
Won Kyong Cho - One of the best experts on this subject based on the ideXlab platform.
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identification of Viruses and Viroids infecting tomato and pepper plants in vietnam by metatranscriptomics
International Journal of Molecular Sciences, 2020Co-Authors: Hoseong Choi, Won Kyong Cho, Phutri Tran, Lakha Salaipeth, Haeryun Kwak, Hongsoo Choi, Kookhyung KimAbstract:Tomato (Lycopersicum esculentum L.) and pepper (Capsicum annuum L.) plants belonging to the family Solanaceae are cultivated worldwide. The rapid development of next-generation sequencing (NGS) technology facilitates the identification of Viruses and Viroids infecting plants. In this study, we carried out metatranscriptomics using RNA sequencing followed by bioinformatics analyses to identify Viruses and Viroids infecting tomato and pepper plants in Vietnam. We prepared a total of 16 libraries, including eight tomato and eight pepper libraries derived from different geographical regions in Vietnam. We identified a total of 602 virus-associated contigs, which were assigned to 18 different virus species belonging to nine different viral genera. We identified 13 different Viruses and two Viroids infecting tomato plants and 12 Viruses and two Viroids infecting pepper plants with Viruses as dominantly observed pathogens. Our results showed that multiple infection of different viral pathogens was common in both plants. Moreover, geographical region and host plant were two major factors to determine viral populations. Taken together, our results provide the comprehensive overview of viral pathogens infecting two important plants in the family Solanaceae grown in Vietnam.
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in silico approach to reveal viral populations in grapevine cultivar tannat using transcriptome data
Scientific Reports, 2015Co-Authors: Hoseong Choi, Ju-yeon Yoon, Seung-kook Choi, Jin Kyong Cho, Won Kyong ChoAbstract:Viruses are ubiquitous and present in a wide range of settings, from living organisms to various environments. Although Viruses are regarded as important pathogens in higher plants, viral populations in specific host plants have not yet been fully examined. This study revealed viral populations in grape berries obtained from a cultivar from a single vineyard using currently available grapevine transcriptomes. Eight Viruses and two Viroids were identified using 11 grapevine libraries. Virus-associated sequences in each transcriptome ranged from 0.2% (seed) to 8.8% (skin). The amount of viral RNAs and virus copy numbers was quantified, thus revealing the dominant virus or viroid in each individual library. In addition, five viral genomes were successfully assembled de novo using transcriptome data. Phylogenetic analyses revealed that the Viruses and Viroids might have originated from Europe, along with the host. Single nucleotide variation studies revealed the quasispecies of RNA Viruses. Taken together, this study defines complex viral populations in three different grape tissues from a single vineyard.
