scholarly journals High-Throughput Sequencing Facilitates Discovery of New Plant Viruses in Poland

Plants ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 820
Author(s):  
Julia Minicka ◽  
Aleksandra Zarzyńska-Nowak ◽  
Daria Budzyńska ◽  
Natasza Borodynko-Filas ◽  
Beata Hasiów-Jaroszewska
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Plant Viruses ◽  
Yellow Mosaic Virus ◽  
Virus Species ◽  
Phylogenetic Groups ◽  
Accurate Identification ◽  
Detection And Identification ◽  
New Finding ◽  
Recent Occurrence

Viruses cause epidemics on all major crops of agronomic importance, and a timely and accurate identification is essential for control. High throughput sequencing (HTS) is a technology that allows the identification of all viruses without prior knowledge on the targeted pathogens. In this paper, we used HTS technique for the detection and identification of different viral species occurring in single and mixed infections in plants in Poland. We analysed various host plants representing different families. Within the 20 tested samples, we identified a total of 13 different virus species, including those whose presence has not been reported in Poland before: clover yellow mosaic virus (ClYMV) and melandrium yellow fleck virus (MYFV). Due to this new finding, the obtained sequences were compared with others retrieved from GenBank. In addition, cucurbit aphid-borne yellows virus (CABYV) was also detected, and due to the recent occurrence of this virus in Poland, a phylogenetic analysis of these new isolates was performed. The analysis revealed that CABYV population is highly diverse and the Polish isolates of CABYV belong to two different phylogenetic groups. Our results showed that HTS-based technology is a valuable diagnostic tool for the identification of different virus species originating from variable hosts, and can provide rapid information about the spectrum of plant viruses previously not detected in a region.

scholarly journals Assessment of Common Soybean-Infecting Viruses in Ohio, USA, Through Multi-site Sampling and High-Throughput Sequencing

2016 ◽  
Vol 17 (2) ◽  
pp. 133-140 ◽  
Author(s):  
Junping Han ◽  
Leslie L. Domier ◽  
Bryan J. Cassone ◽  
Anne Dorrance ◽  
Feng Qu
Keyword(s):  
Mosaic Virus ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Virus Disease ◽  
Soybean Mosaic Virus ◽  
Alfalfa Mosaic Virus ◽  
Plant Viruses ◽  
Yellow Mosaic Virus ◽  
Tobacco Streak Virus ◽  
Streak Virus

Multi-site sampling was conducted during 2011 and 2012 to assess the scope of virus disease problems of soybean in Ohio, USA. A total of 259 samples were collected from 80 soybean fields distributed in 42 Ohio counties, accounting for more than 90% of major soybean-growing counties in Ohio. A high-throughput RNA-Seq approach was adopted to identify all viruses in the samples that share sufficient sequence similarities with known plant viruses. To minimize sequencing costs, total RNA extracted from up to 20 samples were first pooled to make up regional pools, resulting in eight regional pools per year in both 2011 and 2012. These regional pools were further pooled into two yearly master pools of RNA, and sequenced using the Illumina's HiSeq2000 platform. Bioinformatic analyses of sequence reads led to the identification of signature sequences of nine different viruses. The originating locations of these viruses were then mapped with PCR or RT-PCR. This study confirmed the widespread distribution of Bean pod mottle virus, Soybean vein necrosis virus, Tobacco ringspot virus, and Tobacco streak virus in Ohio. It additionally revealed occasional association of Alfalfa mosaic virus, Bean yellow mosaic virus, Clover yellow vein virus, Soybean mosaic virus, and Soybean Putnam virus with Ohio soybean. This is the first statewide survey of soybean viruses in Ohio, and provides the much-needed baseline information for management of virus diseases of soybean. Accepted for publication 20 May 2016. Published 10 June 2016.

scholarly journals Quality Assessment and Validation of High-Throughput Sequencing for Grapevine Virus Diagnostics

Viruses ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 1130
Author(s):  
Nourolah Soltani ◽  
Kristian A. Stevens ◽  
Vicki Klaassen ◽  
Min-Sook Hwang ◽  
Deborah A. Golino ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Plant Viruses ◽  
Molecular Diagnostic ◽  
Virus Species ◽  
Certification Programs ◽  
Performance Specifications ◽  
Virus Diagnostics ◽  
Diagnostic Research ◽  
And Performance

Development of High-Throughput Sequencing (HTS), also known as next generation sequencing, revolutionized diagnostic research of plant viruses. HTS outperforms bioassays and molecular diagnostic assays that are used to screen domestic and quarantine grapevine materials in data throughput, cost, scalability, and detection of novel and highly variant virus species. However, before HTS-based assays can be routinely used for plant virus diagnostics, performance specifications need to be developed and assessed. In this study, we selected 18 virus-infected grapevines as a test panel for measuring performance characteristics of an HTS-based diagnostic assay. Total nucleic acid (TNA) was extracted from petioles and dormant canes of individual samples and constructed libraries were run on Illumina NextSeq 500 instrument using a 75-bp single-end read platform. Sensitivity was 98% measured over 264 distinct virus and viroid infections with a false discovery rate (FDR) of approximately 1 in 5 positives. The results also showed that combining a spring petiole test with a fall cane test increased sensitivity to 100% for this TNA HTS assay. To evaluate extraction methodology, these results were compared to parallel dsRNA extractions. In addition, in a more detailed dilution study, the TNA HTS assay described here consistently performed well down to a dilution of 5%. In that range, sensitivity was 98% with a corresponding FDR of approximately 1 in 5. Repeatability and reproducibility were assessed at 99% and 93%, respectively. The protocol, criteria, and performance levels described here may help to standardize HTS for quality assurance and accreditation purposes in plant quarantine or certification programs.

scholarly journals Direct Metatranscriptomic Survey of the Sunflower Microbiome and Virome

Viruses ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 1867
Author(s):  
Ziyi Wang ◽  
Achal Neupane ◽  
Jiuhuan Feng ◽  
Connor Pedersen ◽  
Shin-Yi Lee Marzano
Keyword(s):  
South Dakota ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Genetic Resource ◽  
De Novo ◽  
Fungal Infections ◽  
Yellow Mosaic Virus ◽  
Protein Database ◽  
Dna Virus ◽  
Field Production

Sunflowers (Helianthus annuus L.) are susceptible to multiple diseases in field production. In this study, we collected diseased sunflower leaves in fields located in South Dakota, USA, for virome investigation. The leaves showed visible symptoms on the foliage, indicating phomopsis and rust infections. To identify the viruses potentially associated with the disease diagnosed, symptomatic leaves were obtained from diseased plants. Total RNA was extracted corresponding to each disease diagnosed to generate libraries for paired-end high throughput sequencing. Short sequencing reads were assembled de novo and the contigs with similarities to viruses were identified by aligning against a custom protein database. We report the discovery of two novel mitoviruses, four novel partitiviruses, one novel victorivirus, and nine novel totiviruses based on similarities to RNA-dependent RNA polymerases and capsid proteins. Contigs similar to bean yellow mosaic virus and Sclerotinia sclerotiorum hypovirulence-associated DNA virus were also detected. To the best of our knowledge, this is the first report of direct metatranscriptomics discovery of viruses associated with fungal infections of sunflowers bypassing culturing. These newly discovered viruses represent a natural genetic resource from which we can further develop potential biopesticide to control sunflower diseases.

scholarly journals Double-Stranded RNA High-Throughput Sequencing Reveals a New Cytorhabdovirus in a Bean Golden Mosaic Virus-Resistant Common Bean Transgenic Line

Viruses ◽  
2019 ◽  
Vol 11 (1) ◽  
pp. 90 ◽  
Author(s):  
Dione M. T. Alves-Freitas ◽  
Bruna Pinheiro-Lima ◽  
Josias C. Faria ◽  
Cristiano Lacorte ◽  
Simone G. Ribeiro ◽  
...  
Keyword(s):  
Phaseolus Vulgaris ◽  
Common Bean ◽  
Mosaic Virus ◽  
High Throughput ◽  
Transgenic Line ◽  
High Throughput Sequencing ◽  
Open Reading Frames ◽  
Virus Species ◽  
Homologous Proteins ◽  
Bean Golden Mosaic Virus

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.

scholarly journals Rapid Detection and Identification of Infectious Pathogens Based on High-throughput Sequencing

2015 ◽  
Vol 128 (7) ◽  
pp. 877-883 ◽  
Author(s):  
Pei-Xiang Ni ◽  
Xin Ding ◽  
Yin-Xin Zhang ◽  
Xue Yao ◽  
Rui-Xue Sun ◽  
...  
Keyword(s):  
High Throughput ◽  
Rapid Detection ◽  
High Throughput Sequencing ◽  
Detection And Identification

scholarly journals High-Throughput Sequencing (HTS) of newly synthetized RNAs enables one shot detection and identification of live mycoplasmas and differentiation from inert nucleic acids

Biologicals ◽  
2020 ◽  
Vol 65 ◽  
pp. 18-24
Author(s):  
Céline Desbrousses ◽  
Fabienne Archer ◽  
Adélie Colin ◽  
Alexandra Bobet-Erny ◽  
Angélique Champavère ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Detection And Identification ◽  
Shot Detection

scholarly journals Virus Detection by High-Throughput Sequencing of Small RNAs: Large-Scale Performance Testing of Sequence Analysis Strategies

2019 ◽  
Vol 109 (3) ◽  
pp. 488-497 ◽  
Author(s):  
Sebastien Massart ◽  
Michela Chiumenti ◽  
Kris De Jonghe ◽  
Rachel Glover ◽  
Annelies Haegeman ◽  
...  
Keyword(s):  
High Throughput ◽  
Large Scale ◽  
High Throughput Sequencing ◽  
Performance Test ◽  
Virus Detection ◽  
Performance Testing ◽  
Plant Viruses ◽  
Reference Sequence ◽  
Bioinformatics Pipeline ◽  
Double Blind

Recent developments in high-throughput sequencing (HTS), also called next-generation sequencing (NGS), technologies and bioinformatics have drastically changed research on viral pathogens and spurred growing interest in the field of virus diagnostics. However, the reliability of HTS-based virus detection protocols must be evaluated before adopting them for diagnostics. Many different bioinformatics algorithms aimed at detecting viruses in HTS data have been reported but little attention has been paid thus far to their sensitivity and reliability for diagnostic purposes. Therefore, we compared the ability of 21 plant virology laboratories, each employing a different bioinformatics pipeline, to detect 12 plant viruses through a double-blind large-scale performance test using 10 datasets of 21- to 24-nucleotide small RNA (sRNA) sequences from three different infected plants. The sensitivity of virus detection ranged between 35 and 100% among participants, with a marked negative effect when sequence depth decreased. The false-positive detection rate was very low and mainly related to the identification of host genome-integrated viral sequences or misinterpretation of the results. Reproducibility was high (91.6%). This work revealed the key influence of bioinformatics strategies for the sensitive detection of viruses in HTS sRNA datasets and, more specifically (i) the difficulty in detecting viral agents when they are novel or their sRNA abundance is low, (ii) the influence of key parameters at both assembly and annotation steps, (iii) the importance of completeness of reference sequence databases, and (iv) the significant level of scientific expertise needed when interpreting pipeline results. Overall, this work underlines key parameters and proposes recommendations for reliable sRNA-based detection of known and unknown viruses.

scholarly journals Recent Advances on Detection and Characterization of Fruit Tree Viruses Using High-Throughput Sequencing Technologies

Viruses ◽  
2018 ◽  
Vol 10 (8) ◽  
pp. 436 ◽  
Author(s):  
Varvara Maliogka ◽  
Angelantonio Minafra ◽  
Pasquale Saldarelli ◽  
Ana Ruiz-García ◽  
Miroslav Glasa ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Infected Plant ◽  
Fruit Trees ◽  
Plant Viruses ◽  
Fruit Tree ◽  
Advantages And Disadvantages ◽  
Sequencing Technologies ◽  
Recent Advances

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., Fabavirus, Luteovirus) 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.

scholarly journals Real time portable genome sequencing for global food security

2018 ◽  
Author(s):  
Laura M. Boykin ◽  
Ammar Ghalab ◽  
Bruno Rossitto De Marchi ◽  
Anders Savill ◽  
James M. Wainaina ◽  
...  
Keyword(s):  
Real Time ◽  
Crop Production ◽  
High Throughput Sequencing ◽  
Smallholder Farmers ◽  
Plant Viruses ◽  
Sub Saharan Africa ◽  
Global Food Security ◽  
Detection And Identification ◽  
Sub Saharan ◽  
Zero Hunger

AbstractThe United Nations has listed Zero Hunger as one of the 17 global sustainable development goals to end extreme poverty by 2030. Plant viruses are a major constraint to crop production globally causing an estimated $30 billion in damage 1 leaving millions of people food insecure 2. In Africa, agriculture employs up to 50% of the workforce, yet only contributes 15% to the GDP on average 3, suggesting that there is low productivity and limited value addition. This can be addressed through continued innovation in the fields of science and technology as suggested in the Science Agenda for Agriculture in Africa (S3A) 4. Sustainable management of plant viruses and their associated vectors must include efficient diagnostics for surveillance, detection and identification to inform disease management, including the development and strategic deployment of virus resistant varieties. To date, researchers have been utilizing conventional methods such as; PCR, qPCR, high throughput sequencing (RNA-Seq, DNA-Seq) and Sanger sequencing for pathogen identification. However, these methods are both costly and time consuming, delaying timely control actions. The emergence of new tools for real-time diagnostics, such as the Oxford Nanopore MinION, have recently proven useful for early detection of Ebola 6 and Zika 7,8, even in low resourced laboratories. For the first time globally, the MinION portable pocket DNA sequencer was used to sequence whole plant virus genomes. We used this technology to identify the begomoviruses causing the devastating CMD which is ravaging smallholder farmers’ crops in sub-Saharan Africa. Cassava, a carbohydrate crop from which tapioca originates, is a major source of calories for over eight hundred (800) million people worldwide. With this technology, farmers struggling with diseased crops can take immediate, restorative action to improve their livelihoods based on information about the health of their plants, generated using a portable, real-time DNA sequencing device.

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