Perennial crops, such as fruit trees, are infected by many viruses, which are transmitted through vegetative propagation and grafting of infected plant material. Some of these pathogens cause severe ...crop losses and often reduce the productive life of the orchards. Detection and characterization of these agents in fruit trees is challenging, however, during the last years, the wide application of high-throughput sequencing (HTS) technologies has significantly facilitated this task. In this review, we present recent advances in the discovery, detection, and characterization of fruit tree viruses and virus-like agents accomplished by HTS approaches. A high number of new viruses have been described in the last 5 years, some of them exhibiting novel genomic features that have led to the proposal of the creation of new genera, and the revision of the current virus taxonomy status. Interestingly, several of the newly identified viruses belong to virus genera previously unknown to infect fruit tree species (e.g.,
,
) a fact that challenges our perspective of plant viruses in general. Finally, applied methodologies, including the use of different molecules as templates, as well as advantages and disadvantages and future directions of HTS in fruit tree virology are discussed.
Several new full genome sequences of olive viruses came to light recently via high-throughput sequencing (HTS) analysis. In this study, total RNA HTS analysis of two Greek olive trees revealed the ...presence of an olive virus T (OlVT) isolate and an olive leaf yellowing-associated virus (OLYaV) isolate. The full viral genome of OlVT isolate (50Ch) is composed of 6862 nucleotides encoding for three proteins (replicase, movement protein, and capsid protein) with typical betaflexiviruses’ genomic features. However, both sequence and phylogenetic data analysis exhibited high levels of variability between 50Ch and the previously characterized OlVT isolates. In addition, the almost full genome of the Greek OLYaV isolate (OL2) was obtained, which is composed of 16,693 nucleotides encoding for 11 open reading frames (ORFs) and shares common genomic features with the recently characterized OLYaV isolates from Spain and Brazil. Sequence and phylogenetic analysis revealed high similarity between these three isolates. Due to problems encountered with the detection of both viruses, new nested RT-PCR assays were developed and applied. In addition, recombination events were observed in OlVT isolates (50Ch GR-168), thus highlighting the potential role of this mechanism in the evolution of the virus. This study is adding further knowledge to the limited information available about these recently characterized olive infecting viral pathogens and highlights their widespread distribution in Greece, one of the most important olive producing countries of the world.
Cucurbit chlorotic yellows virus (CCYV) and cucurbit yellow stunting disorder virus (CYSDV) are two closely related criniviruses that often coinfect cucurbits and are associated with cucurbit yellows ...disease. Both viruses are distributed worldwide and are transmitted in a semipersistent manner by the whitefly vectors
MED or MEAM1. The major goal of this study was to provide insight into the interaction of CCYV and CYSDV in cucumber and to study the effect on transmission by
MED. The titers of both viruses were estimated in single- and dually infected cucumber plants via reverse transcription PCR assays. In mixed infections, the accumulation of both viruses was significantly decreased. When
MED adults were placed on cucumber infected with both viruses, their simultaneous transmission efficiency was significantly higher, whereas transmission efficiency of each individual virus was low. Moreover, nonviruliferous whiteflies preferentially settled on crinivirus-infected cucumber plants, whereas viruliferous whiteflies were attracted by healthy cucumber plants. Finally, the titer of both viruses was calculated in five commercial cucumber hybrids, followed by subsequent transmission experiments. Our results show that although the titers of CYSDV and CCYV were significantly lower in mixed infections in cucumbers, their simultaneous transmission increased.
In 2020, a new pepper disease was recorded in a greenhouse crop in Drama, Greece, exhibiting mild concentric chlorotic rings and deformation on the fruits. High‐throughput sequencing (HTS) of total ...RNA from a plant with symptoms revealed three viral RNA segments closely related (>90% nucleotide identity) to those of four recently identified Ilarvirus species, Solanum nigrum ilarvirus 1 (SnIV1), surrounding legume associated ilarvirus (sLaIV), Erysiphe necator associated ilar‐like virus (ENaIlV1) and grapevine associated ilarvirus (GaIV). Nucleotide comparisons indicated that SnIV1, sLaIV, ENaIlV1, GaIV and the herein detected ilarvirus are isolates of the same virus species. All these ilarviruses were also discovered with the aid of HTS on different hosts; however, no data existed so far regarding their putative association with a disease. To fulfil Koch's postulates, we conducted mechanical inoculation to indicator plants and cultivated solanaceous species. The virus was transmitted only to Nicotiana benthamiana and Capsicum annuum ‘Arlequin’ plants; mild concentric rings, like the original symptoms, also appeared in the fruits of the pepper plants. We suggest that the new ilarvirus isolate PD be classified as SnIV1. However, the nomenclature of the species, including all the recently determined ilarviruses, should be ultimately addressed by the relevant ICTV group. Finally, a small‐scale survey revealed that SnIV1‐PD is currently not present in other geographical regions; nonetheless, more surveys are needed in Greece and abroad to unravel the geographical distribution, the biological characteristics and the evolutionary history of this virus.
A viral isolate, closely related to a group of newly identified ilarviruses, is inducing mild concentric chlorotic rings and deformation on the fruits of pepper.
In 2018, typical symptoms of sweet potato virus disease (SPVD) such as purple ringspots, distortion and crinkling of the leaves appeared on a sweet potato crop located in southern Greece. ...High‐throughput sequencing (HTS) analysis revealed the presence of seven viruses, sweet potato feathery mottle virus (SPFMV), sweet potato chlorotic stunt virus (SPCSV), sweet potato virus C (SPVC), sweet potato virus G (SPVG), sweet potato virus 2 (SPV2), sweet potato leaf curl virus (SPLCV) and sweet potato pakakuy virus (SPPV). In silico analysis of the HTS data in combination with Sanger sequencing succeeded in obtaining the complete genome sequences of all seven viruses. Nucleotide sequence comparisons of complete viral genomes showed high similarity of all detected potyviruses and SPLCV with homologous sequences retrieved from GenΒank, whereas a new variant of SPCSV RNA1 was characterized and successfully transmitted onto Nicotiana benthamiana plants with the whitefly species Bemisia tabaci MED and MEAM1. Moreover, the Greek strain of SPPV showed higher similarity with SPPV strain B, even though it possesses a variable RT‐RNase H domain. Further phylogenetic analysis clustered Greek sweet potato virus isolates into various phylogenetic groups, whereas few new clades were formed in the cases of SPFMV, SPVG and SPCSV RNA1. Small‐scale surveys on three different sweet potato crops exhibiting typical, mild or no symptoms also indicated the close association of SPCSV with SPVD. Taken together, this study characterizes for the first time the virome of sweet potato crops in Greece and provides a glimpse into the evolutionary relationships of the identified viruses.
Identification and characterization of sweet potato viruses in Greece using high‐throughput sequencing, phylogenetic analysis and vector transmission revealed seven species and a glimpse into their evolutionary relationships.
Control of viruses infecting grapevine Maliogka, Varvara I; Martelli, Giovanni P; Fuchs, Marc ...
Advances in virus research,
01/2015, Letnik:
91
Journal Article
Recenzirano
Grapevine is a high value vegetatively propagated fruit crop that suffers from numerous viruses, including some that seriously affect the profitability of vineyards. Nowadays, 64 viruses belonging to ...different genera and families have been reported in grapevines and new virus species will likely be described in the future. Three viral diseases namely leafroll, rugose wood, and infectious degeneration are of major economic importance worldwide. The viruses associated with these diseases are transmitted by mealybugs, scale and soft scale insects, or dagger nematodes. Here, we review control measures of the major grapevine viral diseases. More specifically, emphasis is laid on (i) approaches for the production of clean stocks and propagative material through effective sanitation, robust diagnosis, as well as local and regional certification efforts, (ii) the management of vectors of viruses using cultural, biological, and chemical methods, and (iii) the production of resistant grapevines mainly through the application of genetic engineering. The benefits and limitations of the different control measures are discussed with regard to accomplishments and future research directions.
(LChV1,
,
) is a widespread pathogen of sweet or sour cherry and other
species, which exhibits high genetic diversity and lacks a putative efficient transmission vector. Thus far, four distinct ...phylogenetic clusters of LChV1 have been described, including isolates from different
species. The recent application of high throughput sequencing (HTS) technologies in fruit tree virology has facilitated the acquisition of new viral genomes and the study of virus diversity. In the present work, several new LChV1 isolates from different countries were fully sequenced using different HTS approaches. Our results reveal the presence of further genetic diversity within the LChV1 species. Interestingly, mixed infections of the same sweet cherry tree with different LChV1 variants were identified for the first time. Taken together, the high intra-host and intra-species diversities of LChV1 might affect its pathogenicity and have clear implications for its accurate diagnostics.
High-throughput sequencing (HTS) was used to construct the virome profile of an old grapevine-leafroll-diseased grapevine (
Vitis vinifera
).
De novo
assembly of HTS data showed a complex infection, ...including a virus sequence with similarity to viruses of the genus
Badnavirus,
family
Caulimoviridae
. The complete genome sequence of this virus consists of 7090 nucleotides and has four open reading frames (ORFs). Genome organisation and phylogenetic analysis identify this virus as a divergent variant of grapevine Roditis leaf discoloration-associated virus (GRLDaV) with 90% nucleotide sequence identity to isolate w4 (NC_027131). This is the first genome sequence of a South African variant of GRLDaV.
In this study, samples collected from eight sweet cherry trees in northern Greece were analyzed by high-throughput sequencing for the presence of viruses. Bioinformatic analysis revealed the presence ...of divergent isolates of cherry latent virus 1 (CLV-1), a recently identified trichovirus in a sweet cherry accession imported into the USA from the Republic of Georgia. The complete genome sequences of seven CLV-1 isolates were determined, and phylogenetic analysis indicated that they belonged to a separate clade from the previously characterized Georgian isolate. A small-scale survey confirmed the presence of CLV-1 in 47 out of 151 sweet cherry samples tested, and partial sequencing of 15 isolates showed a high degree of nucleotide sequence similarity among them.
Three duplex real-time reverse-transcription polymerase chain reaction (real-time RT-PCR) assays based on TaqMan chemistry, were developed for the simultaneous detection and specific quantification ...of apple chlorotic leafspot virus (ACLSV), plum pox virus (PPV), prunus necrotic ringspot virus (PNRSV), prune dwarf virus (PDV), peach latent mosaic viroid (PLMVd) and the European stone fruit yellows (ESFY) phytoplasma, which are considered among the most important pathogens affecting stone fruit trees. The quantitative RT-PCR (RT-qPCR) assays were optimized using RNA transcripts (linearized plasmid was used for the assay optimization of the ESFY phytoplasma) of known concentrations. No differences in sensitivity were recorded between the duplex and singleplex RT-qPCR assays. The amplification efficiency of the duplex assays reached 91.1–95.8%, while the linear range of quantification was from 20 to 2 × 107 RNA/linearized plasmid transcripts for PLMVd and ESFY phytoplasma, 40 to 4 × 107 RNA transcripts for ACLSV, PPV and PDV, and 102 to 108 RNA transcripts for PNRSV, respectively. The duplex RT-qPCR assays, which were validated using both characterized isolates from all pathogens and field samples from Prunus species in Northern Greece, exhibited a broad detection range. Overall, the developed methods comprise useful tools that could be applied for the simultaneous and reliable detection of graft-transmissible pathogens in certification programs of Prunus spp.
•Three new duplex RT-qPCR assays were developed for the detection of important graft-transmissible pathogens of stone fruits.•The methods exhibit high diagnostic sensitivity and specificity.•The developed assays comprise useful tools that could be applied for virus indexing in certification programs of Prunus spp.
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