scholarly journals Characterization of Spanish Olive Virome by High Throughput Sequencing Opens New Insights and Uncertainties

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2233
Author(s):  
Ana Belén Ruiz-García ◽  
Celia Canales ◽  
Félix Morán ◽  
Manuel Ruiz-Torres ◽  
Magdalena Herrera-Mármol ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Biological Significance ◽  
Euphyllura Olivina ◽  
Leaf Yellowing ◽  
Full Length Sequence ◽  
Pros And Cons ◽  
Olive Trees ◽  
First Time ◽  
Viral Sequences

The use of high throughput sequencing (HTS) for the analysis of Spanish olive trees showing leaf yellowing discoloration, defoliation, and/or decline has provided new insights into the olive viruses present in Spain and has opened discussions about the pros and cons of these technologies for diagnostic purposes. In this study, we report for the first time in Spanish orchards the presence of olive leaf yellowing-associated virus (OLYaV), for which the second full coding sequence has been determined. This virus has also been detected in a putative vector, the psyllid Euphyllura olivina. In addition, the presence in Spain of Olea europaea geminivirus (OEGV), recently reported in Italy, has been confirmed, and the full-length sequence of two isolates was obtained by HTS and Sanger sequencing. These results, as well as the detection of other viral sequences related to olive latent virus 3 (OLV-3) and olive viral satellite RNA, raises questions on the biological significance of the findings, about the requirement of standardization on the interpretation of HTS results, and the necessity of additional tests to confirm the relevance of the HTS detection of viral sequences.

scholarly journals Virus Surveys in Olive Orchards in Greece Identify Olive Virus T, a Novel Member of the Genus Tepovirus

Pathogens ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 574
Author(s):  
Evanthia Xylogianni ◽  
Paolo Margaria ◽  
Dennis Knierim ◽  
Kyriaki Sareli ◽  
Stephan Winter ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
Leaf Roll ◽  
Virus Infections ◽  
Northern Greece ◽  
Coat Protein Region ◽  
Cherry Leaf Roll Virus ◽  
Leaf Yellowing ◽  
Olive Trees ◽  
Olive Varieties ◽  
Viral Sequences

Field surveys were conducted in Greek olive orchards from 2017 to 2020 to collect information on the sanitary status of the trees. Using a high-throughput sequencing approach, viral sequences were identified in total RNA extracts from several trees and assembled to reconstruct the complete genomes of two isolates of a new viral species of the genus Tepovirus (Betaflexiviridae), for which the name olive virus T (OlVT) is proposed. A reverse transcription–polymerase chain reaction assay was developed which detected OlVT in samples collected in olive growing regions in Central and Northern Greece, showing a virus prevalence of 4.4% in the olive trees screened. Sequences of amplified fragments from the movement–coat protein region of OlVT isolates varied from 75.64% to 99.35%. Three olive varieties (Koroneiki, Arbequina and Frantoio) were infected with OlVT via grafting to confirm a graft-transmissible agent, but virus infections remained latent. In addition, cucumber mosaic virus, olive leaf yellowing-associated virus and cherry leaf roll virus were identified.

scholarly journals High-Throughput Sequencing Reveals Further Diversity of Little Cherry Virus 1 with Implications for Diagnostics

Viruses ◽  
2018 ◽  
Vol 10 (7) ◽  
pp. 385 ◽  
Author(s):  
Asimina Katsiani ◽  
Varvara Maliogka ◽  
Nikolaos Katis ◽  
Laurence Svanella-Dumas ◽  
Antonio Olmos ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Sour Cherry ◽  
Prunus Species ◽  
Cherry Tree ◽  
High Genetic Diversity ◽  
Viral Genomes ◽  
Virus Diversity ◽  
First Time

Little cherry virus 1 (LChV1, Velarivirus, Closteroviridae) is a widespread pathogen of sweet or sour cherry and other Prunus 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 Prunus 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.

scholarly journals Rice Stripe Tenuivirus Has a Greater Tendency To Use the Prime-and-Realign Mechanism in Transcription of Genomic than in Transcription of Antigenomic Template RNAs

2017 ◽  
Vol 92 (1) ◽  
Author(s):  
Xiaojuan Liu ◽  
Jing Jin ◽  
Ping Qiu ◽  
Fangluan Gao ◽  
Wenzhong Lin ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Rna Viruses ◽  
Viral Mrnas ◽  
Viral Pathogens ◽  
Promising Target ◽  
Cellular Mrnas ◽  
Sequencing Strategy ◽  
First Time ◽  
Negative Sense

ABSTRACTMost segmented negative-sense RNA viruses employ a process termed cap snatching, during which they snatch capped RNA leaders from host cellular mRNAs and use the snatched leaders as primers for transcription, leading to the synthesis of viral mRNAs with 5′ heterogeneous sequences (HSs). With traditional methods, only a few HSs can be determined, and identification of their donors is difficult. Here, the mRNA 5′ ends ofRice stripe tenuivirus(RSV) andRice grassy stunt tenuivirus(RGSV) and those of their host rice were determined by high-throughput sequencing. Millions of tenuiviral HSs were obtained, and a large number of them mapped to the 5′ ends of corresponding host cellular mRNAs. Repeats of the dinucleotide AC, which are complementary to the U1G2of the tenuiviral template 3′-U1G2U3G4UUUCG, were found to be prevalent at the 3′ termini of tenuiviral HSs. Most of these ACs did not match host cellular mRNAs, supporting the idea that tenuiviruses use the prime-and-realign mechanism during cap snatching. We previously reported a greater tendency of RSV than RGSV to use the prime-and-realign mechanism in transcription with leaders cap snatched from a coinfecting reovirus. Besides confirming this observation in natural tenuiviral infections, the data here additionally reveal that RSV has a greater tendency to use this mechanism in transcribing genomic than in transcribing antigenomic templates. The data also suggest that tenuiviruses cap snatch host cellular mRNAs from translation- and photosynthesis-related genes, and capped RNA leaders snatched by tenuiviruses base pair with U1/U3or G2/G4of viral templates. These results provide unprecedented insights into the cap-snatching process of tenuiviruses.IMPORTANCEMany segmented negative-sense RNA viruses (segmented NSVs) are medically or agriculturally important pathogens. The cap-snatching process is a promising target for the development of antiviral strategies against this group of viruses. However, many details of this process remain poorly characterized. Tenuiviruses constitute a genus of agriculturally important segmented NSVs, several members of which are major viral pathogens of rice. Here, we for the first time adopted a high-throughput sequencing strategy to determine the 5′ heterogeneous sequences (HSs) of tenuiviruses and mapped them to host cellular mRNAs. Besides providing deep insights into the cap snatching of tenuiviruses, the data obtained provide clear evidence to support several previously proposed models regarding cap snatching. Curiously and importantly, the data here reveal that not only different tenuiviruses but also the same tenuivirus synthesizing different mRNAs use the prime-and-realign mechanism with different tendencies during their cap snatching.

scholarly journals Detection Of Genomic Variants Of SARS-CoV-2 Circulating In Wastewater By High-Throughput Sequencing

2021 ◽  
Author(s):  
Alba Pérez-Cataluña ◽  
Álvaro Chiner-Oms ◽  
Enric Cuevas-Ferrando ◽  
Azahara Díaz-Reolid ◽  
Irene Falcó ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Community Level ◽  
Low Frequencies ◽  
Spike Gene ◽  
Genomic Variants ◽  
Clinical Databases ◽  
Novel Variants ◽  
First Time

The use of SARS-CoV-2 metagenomics in wastewater can allow the detection of variants circulating at community level. After comparing with clinical databases, we identified three novel variants in the spike gene, and six new variants in the spike detected for the first time in Spain. We finally support the hypothesis that this approach allows the identification of unknown SARS-CoV-2 variants or detected at only low frequencies in clinical genomes.Abstract Figure

scholarly journals Viral Sequences Detection by High-Throughput Sequencing in Cerebrospinal Fluid of Individuals with and without Central Nervous System Disease

Genes ◽  
2019 ◽  
Vol 10 (8) ◽  
pp. 625 ◽  
Author(s):  
Schibler ◽  
Brito ◽  
Zanella ◽  
Zdobnov ◽  
Laubscher ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Cerebrospinal Fluid ◽  
Nervous System ◽  
High Throughput ◽  
Clinical Microbiology ◽  
High Throughput Sequencing ◽  
Sequence Data ◽  
Unknown Origin ◽  
Cns Inflammation ◽  
Viral Sequences

: Meningitis, encephalitis, and myelitis are various forms of acute central nervous system (CNS) inflammation, which can coexist and lead to serious sequelae. Known aetiologies include infections and immune-mediated processes. Despite advances in clinical microbiology over the past decades, the cause of acute CNS inflammation remains unknown in approximately 50% of cases. High-throughput sequencing was performed to search for viral sequences in cerebrospinal fluid (CSF) samples collected from 26 patients considered to have acute CNS inflammation of unknown origin, and 10 patients with defined causes of CNS diseases. In order to better grasp the clinical significance of viral sequence data obtained in CSF, 30 patients without CNS disease who had a lumbar puncture performed during elective spinal anaesthesia were also analysed. One case of human astrovirus (HAstV)-MLB2-related meningitis and disseminated infection was identified. No other viral sequences that can easily be linked to CNS inflammation were detected. Viral sequences obtained in all patient groups are discussed. While some of them reflect harmless viral infections, others result from reagent or sample contamination, as well as index hopping. Altogether, this study highlights the potential of high-throughput sequencing in identifying previously unknown viral neuropathogens, as well as the interpretation issues related to its application in clinical microbiology.

scholarly journals The Diversity and Distribution of Viruses Associated with Culex annulirostris Mosquitoes from the Kimberley Region of Western Australia

Viruses ◽  
2020 ◽  
Vol 12 (7) ◽  
pp. 717 ◽  
Author(s):  
Simon H. Williams ◽  
Avram Levy ◽  
Rachel A. Yates ◽  
Nilusha Somaweera ◽  
Peter J. Neville ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
High Throughput ◽  
Western Australia ◽  
High Throughput Sequencing ◽  
Marked Variation ◽  
Chain Reaction ◽  
Virus Prevalence ◽  
Culex Annulirostris ◽  
Polymerase Chain ◽  
Viral Sequences

Metagenomics revealed an impressive breadth of previously unrecognized viruses. Here, we report the virome of the Culex annulirostris Skuse mosquito, an important vector of pathogenic arboviruses in Australia. Mosquitoes were collected from three sites in the Kimberley region of Western Australia. Unbiased high-throughput sequencing (HTS) revealed the presence of 16 novel viral sequences that share less than 90% identity with known viruses. None were closely related to pathogenic arboviruses. Viruses were distributed unevenly across sites, indicating a heterogeneous Cx. annulirostris virome. Polymerase chain reaction assays confirmed HTS data and identified marked variation between the virus prevalence identified at each site.

scholarly journals High-Throughput Sequencing for Detection of Subpopulations of Bacteria Not Previously Associated with Artisanal Cheeses

2012 ◽  
Vol 78 (16) ◽  
pp. 5717-5723 ◽  
Author(s):  
Lisa Quigley ◽  
Orla O'Sullivan ◽  
Tom P. Beresford ◽  
R. Paul Ross ◽  
Gerald F. Fitzgerald ◽  
...  
Keyword(s):  
High Throughput ◽  
Significant Influence ◽  
High Throughput Sequencing ◽  
Microbial Populations ◽  
Bacterial Populations ◽  
Content Type ◽  
Pasteurized Milk ◽  
First Time

ABSTRACTHere, high-throughput sequencing was employed to reveal the highly diverse bacterial populations present in 62 Irish artisanal cheeses and, in some cases, associated cheese rinds. Using this approach, we revealed the presence of several genera not previously associated with cheese, includingFaecalibacterium,Prevotella, andHelcococcusand, for the first time, detected the presence ofArthrobacterandBrachybacteriumin goats' milk cheese. Our analysis confirmed many previously observed patterns, such as the dominance of typical cheese bacteria, the fact that the microbiota of raw and pasteurized milk cheeses differ, and that the level of cheese maturation has a significant influence onLactobacilluspopulations. It was also noted that cheeses containing adjunct ingredients had lower proportions ofLactococcusspecies. It is thus apparent that high-throughput sequencing-based investigations can provide valuable insights into the microbial populations of artisanal foods.

scholarly journals Use of High-Throughput Sequencing and Two RNA Input Methods to Identify Viruses Infecting Tomato Crops

2021 ◽  
Vol 9 (5) ◽  
pp. 1043
Author(s):  
Ayoub Maachi ◽  
Covadonga Torre ◽  
Raquel N. Sempere ◽  
Yolanda Hernando ◽  
Miguel A. Aranda ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Tomato Fruit ◽  
Virus Detection ◽  
Read Depth ◽  
Cdna Libraries ◽  
Virus Species ◽  
Total Rna ◽  
Viral Genomes ◽  
First Time

We used high-throughput sequencing to identify viruses on tomato samples showing virus-like symptoms. Samples were collected from crops in the Iberian Peninsula. Either total RNA or double-stranded RNA (dsRNA) were used as starting material to build the cDNA libraries. In total, seven virus species were identified, with pepino mosaic virus being the most abundant one. The dsRNA input provided better coverage and read depth but missed one virus species compared with the total RNA input. By performing in silico analyses, we determined a minimum sequencing depth per sample of 0.2 and 1.5 million reads for dsRNA and rRNA-depleted total RNA inputs, respectively, to detect even the less abundant viruses. Primers and TaqMan probes targeting conserved regions in the viral genomes were designed and/or used for virus detection; all viruses were detected by qRT-PCR/RT-PCR in individual samples, with all except one sample showing mixed infections. Three virus species (Olive latent virus 1, Lettuce ring necrosis virus and Tomato fruit blotch virus) are herein reported for the first time in tomato crops in Spain.

scholarly journals Identification and Characterization of Citrus Concave Gum-Associated Virus Infecting Citrus and Apple Trees by Serological, Molecular and High-Throughput Sequencing Approaches

Plants ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 2390
Author(s):  
Maria Minutolo ◽  
Maria Cinque ◽  
Michela Chiumenti ◽  
Francesco Di Serio ◽  
Daniela Alioto ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Rna Virus ◽  
Polyclonal Antiserum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Apple Trees ◽  
The Usa ◽  
Tissue Blot Immunoassay ◽  
Apple Cultivars ◽  
First Time

Citrus concave gum-associated virus (CCGaV) is a negative-stranded RNA virus, first reported a few years ago in citrus trees from Italy. It has been reported in apple trees in the USA and in Brazil, suggesting a wider host range and geographic distribution. Here, an anti-CCGaV polyclonal antiserum to specifically detect the virus has been developed and used in a standard double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) that has been validated as a sensitive and reliable method to detect this virus both in citrus and apple trees. In contrast, when the same antiserum was used in direct tissue-blot immunoassay, CCGaV was efficiently detected in citrus but not in apple. Using this antiserum, the first apple trees infected by CCGaV were identified in Italy and the presence of CCGaV in several apple cultivars in southern Italy was confirmed by field surveys. High-throughput sequencing (HTS) allowed for the assembling of the complete genome of one CCGaV Italian apple isolate (CE-c3). Phylogenetic analysis of Italian CCGaV isolates from apple and citrus and those available in the database showed close relationships between the isolates from the same genus (Citrus or Malus), regardless their geographical origin. This finding was further confirmed by the identification of amino acid signatures specific of isolates infecting citrus or apple hosts. Analysis of HTS reads also revealed that the CE-c3 Italian apple tree, besides CCGaV, was simultaneously infected by several viruses and one viroid, including apple rubbery wood virus 2 which is reported for the first time in Italy. The complete or almost complete genomic sequences of the coinfecting agents were determined.

scholarly journals A Reference Viral Database (RVDB) To Enhance Bioinformatics Analysis of High-Throughput Sequencing for Novel Virus Detection

mSphere ◽  
2018 ◽  
Vol 3 (2) ◽  
Author(s):  
Norman Goodacre ◽  
Aisha Aljanahi ◽  
Subhiksha Nandakumar ◽  
Mike Mikailov ◽  
Arifa S. Khan
Keyword(s):  
High Throughput ◽  
Selection Criteria ◽  
High Throughput Sequencing ◽  
Bioinformatics Analysis ◽  
Virus Detection ◽  
Sequence Diversity ◽  
Endogenous Retroviruses ◽  
Viral Sequence ◽  
Novel Virus ◽  
Viral Sequences

ABSTRACTDetection of distantly related viruses by high-throughput sequencing (HTS) is bioinformatically challenging because of the lack of a public database containing all viral sequences, without abundant nonviral sequences, which can extend runtime and obscure viral hits. Our reference viral database (RVDB) includes all viral, virus-related, and virus-like nucleotide sequences (excluding bacterial viruses), regardless of length, and with overall reduced cellular sequences. Semantic selection criteria (SEM-I) were used to select viral sequences from GenBank, resulting in a first-generation viral database (VDB). This database was manually and computationally reviewed, resulting in refined, semantic selection criteria (SEM-R), which were applied to a new download of updated GenBank sequences to create a second-generation VDB. Viral entries in the latter were clustered at 98% by CD-HIT-EST to reduce redundancy while retaining high viral sequence diversity. The viral identity of the clustered representative sequences (creps) was confirmed by BLAST searches in NCBI databases and HMMER searches in PFAM and DFAM databases. The resulting RVDB contained a broad representation of viral families, sequence diversity, and a reduced cellular content; it includes full-length and partial sequences and endogenous nonretroviral elements, endogenous retroviruses, and retrotransposons. Testing of RVDBv10.2, with an in-house HTS transcriptomic data set indicated a significantly faster run for virus detection than interrogating the entirety of the NCBI nonredundant nucleotide database, which contains all viral sequences but also nonviral sequences. RVDB is publically available for facilitating HTS analysis, particularly for novel virus detection. It is meant to be updated on a regular basis to include new viral sequences added to GenBank.IMPORTANCETo facilitate bioinformatics analysis of high-throughput sequencing (HTS) data for the detection of both known and novel viruses, we have developed a new reference viral database (RVDB) that provides a broad representation of different virus species from eukaryotes by including all viral, virus-like, and virus-related sequences (excluding bacteriophages), regardless of their size. In particular, RVDB contains endogenous nonretroviral elements, endogenous retroviruses, and retrotransposons. Sequences were clustered to reduce redundancy while retaining high viral sequence diversity. A particularly useful feature of RVDB is the reduction of cellular sequences, which can enhance the run efficiency of large transcriptomic and genomic data analysis and increase the specificity of virus detection.

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