scholarly journals Probing the Viromic Frontiers

mBio ◽  
2015 ◽  
Vol 6 (6) ◽  
Author(s):  
Angela L. Rasmussen
Keyword(s):  
New Genus ◽  
High Throughput Sequencing ◽  
Screening Method ◽  
Virus Detection ◽  
Clinical Samples ◽  
New Paradigm ◽  
High Background ◽  
Sequencing Platform ◽  
The Family ◽  
Viral Sequences

ABSTRACT Modern molecular technology, and particularly high-throughput sequencing (HTS), has revolutionized virus discovery and expanded the depth and breadth of the virome. Recent HTS was used to identify and discover a previously undescribed member of the family Flaviviridae that has genomic features characteristic of both hepaciviruses and pegiviruses. This virus, designated human hepegivirus-1 (HHpgV-1), may represent a previously undescribed new genus in the Flaviviridae family with implications for public health and blood supply safety. Detecting uncharacterized viruses such as HHpgV-1 in clinical samples requires an unbiased screening method that is as sensitive as PCR, while simultaneously detecting multiple rare viral sequences. The virome-capture-sequencing platform for vertebrate viruses (VirCapSeq-VERT) uses positive-selection oligonucleotide capture to sensitively detect sequences from every known vertebrate virus, even in high-background specimens with low-abundance viruses. VirCapSeq-VERT can also detect uncharacterized viruses with sequence homology to known viruses, enabling a new paradigm for virus detection.

scholarly journals Genomic Analyses of Cherry Rusty Mottle Group and Cherry Twisted Leaf-Associated Viruses Reveal a Possible New Genus Within the FamilyBetaflexiviridae

2015 ◽  
Vol 105 (3) ◽  
pp. 399-408 ◽  
Author(s):  
D. E. V. Villamor ◽  
J. Susaimuthu ◽  
K. C. Eastwell
Keyword(s):  
New Genus ◽  
High Throughput Sequencing ◽  
Prunus Avium ◽  
Triple Gene Block ◽  
Mottle Virus ◽  
Virus Species ◽  
Gene Block ◽  
Green Ring ◽  
The Family ◽  
Prunus Spp

It is demonstrated that closely related viruses within the family Betaflexiviridae are associated with a number of diseases that affect sweet cherry (Prunus avium) and other Prunus spp. Cherry rusty mottle-associated virus (CRMaV) is correlated with the appearance of cherry rusty mottle disease (CRMD), and Cherry twisted leaf-associated virus (CTLaV) is linked to cherry twisted leaf disease (CTLD) and apricot ringpox disease (ARPD). Comprehensive analysis of previously reported full genomic sequences plus those determined in this study representing isolates of CTLaV, CRMaV, Cherry green ring mottle virus, and Cherry necrotic rusty mottle virus revealed segregation of sequences into four clades corresponding to distinct virus species. High-throughput sequencing of RNA from representative source trees for CRMD, CTLD, and ARPD did not reveal additional unique virus sequences that might be associated with these diseases, thereby further substantiating the association of CRMaV and CTLaV with CRMD and CTLD or ARPD, respectively. Based on comparison of the nucleotide and amino acid sequence identity values, phylogenetic relationships with other triple-gene block-coding viruses within the family Betaflexiviridae, genome organization, and natural host range, a new genus (Robigovirus) is suggested.

scholarly journals A Novel Subtype of Bovine Hepacivirus Identified in Ticks Reveals the Genetic Diversity and Evolution of Bovine Hepacivirus

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2206
Author(s):  
Jian-Wei Shao ◽  
Luan-Ying Guo ◽  
Yao-Xian Yuan ◽  
Jun Ma ◽  
Ji-Ming Chen ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
High Throughput Sequencing ◽  
Genetic Recombination ◽  
Purifying Selection ◽  
Genotype 1 ◽  
New Members ◽  
Persistent Infections ◽  
The Family ◽  
Viral Sequences ◽  
Demarcation Criteria

Hepaciviruses represent a group of viruses that pose a significant threat to the health of humans and animals. New members of the genus Hepacivirus in the family Flaviviridae have recently been identified in a wide variety of host species worldwide. Similar to the Hepatitis C virus (HCV), bovine hepacivirus (BovHepV) is hepatotropic and causes acute or persistent infections in cattle. BovHepVs are distributed worldwide and classified into two genotypes with seven subtypes in genotype 1. In this study, three BovHepV strains were identified in the samples of ticks sucking blood on cattle in the Guangdong province of China, through unbiased high-throughput sequencing. Genetic analysis revealed the polyprotein-coding gene of these viral sequences herein shared 67.7–84.8% nt identity and 76.1–95.6% aa identity with other BovHepVs identified worldwide. As per the demarcation criteria adopted for the genotyping and subtyping of HCV, these three BovHepV strains belonged to a novel subtype within the genotype 1. Additionally, purifying selection was the dominant evolutionary pressure acting on the genomes of BovHepV, and genetic recombination was not common among BovHepVs. These results expand the knowledge about the genetic diversity and evolution of BovHepV distributed globally, and also indicate genetically divergent BovHepV strains were co-circulating in cattle populations in China.

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.

scholarly journals A Versatile Processing Workflow to Enable Pathogen Detection in Clinical Samples from Organs Using VIDISCA

Diagnostics ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 791
Author(s):  
Alba Folgueiras-González ◽  
Robin van den Braak ◽  
Martin Deijs ◽  
Lia van der Hoek ◽  
Ad de Groof
Keyword(s):  
High Throughput ◽  
Pathogen Detection ◽  
High Throughput Sequencing ◽  
Genetic Material ◽  
Virus Detection ◽  
Detection Sensitivity ◽  
Clinical Samples ◽  
Tissue Samples ◽  
Sequencing Technologies ◽  
Organic Extraction

In recent years, refined molecular methods coupled with powerful high throughput sequencing technologies have increased the potential of virus discovery in clinical samples. However, host genetic material remains a complicating factor that interferes with discovery of novel viruses in solid tissue samples as the relative abundance of the virus material is low. Physical enrichment processing methods, although usually complicated, labor-intensive, and costly, have proven to be successful for improving sensitivity of virus detection in complex samples. In order to further increase detectability, we studied the application of fast and simple high-throughput virus enrichment methods on tissue homogenates. Probe sonication in high EDTA concentrations, organic extraction with Vertrel™ XF, or a combination of both, were applied prior to chromatography-like enrichment using Capto™ Core 700 resin, after which effects on virus detection sensitivity by the VIDISCA method were determined. Sonication in the presence of high concentrations of EDTA showed the best performance with an increased proportion of viral reads, up to 9.4 times, yet minimal effect on the host background signal. When this sonication procedure in high EDTA concentrations was followed by organic extraction with Vertrel™ XF and two rounds of core bead chromatography enrichment, an increase up to 10.5 times in the proportion of viral reads in the processed samples was achieved, with reduction of host background sequencing. We present a simple and semi-high-throughput method that can be used to enrich homogenized tissue samples for viral reads.

Occurrence and distribution of Actinidia viruses in Shaanxi Province of China

Plant Disease ◽  
2020 ◽  
Author(s):  
Zhao Lei ◽  
Mengji Cao ◽  
Qianru Huang ◽  
Yicheng Wang ◽  
Jiaxiu Sun ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
Virus Detection ◽  
International Committee ◽  
Shaanxi Province ◽  
Transmission Electron ◽  
Viral Particles ◽  
The Family ◽  
Novel Virus ◽  
Actinidia Spp ◽  
Yellowing Virus

Kiwifruit (Actinidia spp.) is an economically important fruit crop globally. China is the largest kiwifruit-growing country in the world, and Shaanxi Province is the major kiwifruit-growing region in China. A systematic survey detected various symptoms in kiwifruit plants grown in a commercial kiwifruit field in Shaanxi Province. Samples were collected from kiwifruit plants showing symptoms and used for virus detection by high-throughput sequencing. In addition to ten known kiwifruit viruses, three new viruses were detected and tentatively named Actinidia yellowing ringspot virus (AYRSpV), Actinidia yellowing virus 1 (AcYV1) and Actinidia yellowing virus 2 (AcYV2). The genome sequences of the three new viruses and four known viruses were determined. Based on the demarcation criteria of the International Committee on Taxonomy of Viruses (ICTV), AYRSpV might be a new member of the genus Ilarvirus in the family Bromoviridae, AcYV1 might be a new virus of the genus Waikavirus in the family Secoviridae, and AcYV2 might be a novel virus in the family Tombusviridae. Spherical viral particles were found in the samples infected with AYRSpV, AcYV1 and AcYV2 by transmission electron microscopy. Further analysis showed that all thirteen viruses can infect both A. deliciosa and A. chinensis, but the incidences of these infections vary among different kiwifruit cultivars in different regions. These results provide valuable information for understanding the viriome of kiwifruit in China.

scholarly journals Adventitious Virus Detection in Cells by High-Throughput Sequencing of Newly Synthesized RNAs: Unambiguous Differentiation of Cell Infection from Carryover of Viral Nucleic Acids

mSphere ◽  
2019 ◽  
Vol 4 (3) ◽  
Author(s):  
Justine Cheval ◽  
Erika Muth ◽  
Gaëlle Gonzalez ◽  
Muriel Coulpier ◽  
Pascale Beurdeley ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Rna Virus ◽  
Virus Detection ◽  
Analytical Sensitivity ◽  
Cell Infection ◽  
Biological Products ◽  
Viral Sequences ◽  
In Cells

ABSTRACTThe use of high-throughput sequencing (HTS) to identify viruses in biologicals differs from current molecular approaches, since its use enables an unbiased approach to detection without the need to design specific primers to preamplify target sequences. Its broad range of detection and analytical sensitivity make it an important tool to ensure that biologicals are free from adventitious viruses. Similar to other molecular methods, however, identification of viral sequences in cells by HTS does not prove viral infection, since this could reflect carryover of inert viral sequences from reagents or other sources or the presence of transcriptionally inactive cellular sequences. Due to the broad range of detection associated with HTS, the above can potentially be perceived as a drawback for the testing of pharmaceutical biological products using this method. In order to avoid the identification of inert viral sequences, we present a methodology based on metabolic RNA labeling and sequencing, which enables the specific identification of newly synthesized viral RNAs in infected cells, resulting in the ability to unambiguously distinguish active infection by DNA or RNA viruses from inert nucleic acids. In the present study, we report the ability to differentiate Vero cells acutely infected by a single-stranded positive-sense RNA virus (tick-borne encephalitis virus) from cells which have been in contact with nonreplicating virus particles. Additionally, we also found a laboratory contamination by the squirrel monkey retrovirus of our Vero cell line, which was derived from an Old World (African green) monkey, a type of contamination which until now has been identified only in cells derived from primates from the New World.IMPORTANCEThe use of high-throughput sequencing (HTS) to identify viral contamination of biological products is extremely sensitive and provides a broad range of detection. Nevertheless, viral sequences identified can also be inert. Examples include contamination resulting from reagents or the presence of inactivated viruses in animal-derived components of the cell culture medium. We therefore developed a method that relies on the sequencing of newly synthesized RNAs, an unequivocal sign of the presence of a transcriptionally active virus. This improvement in the specificity of viral testing increases the acceptability of HTS as a standard test for cells used in manufacturing biologicals and in biotherapies.

scholarly journals Virome Capture Sequencing Enables Sensitive Viral Diagnosis and Comprehensive Virome Analysis

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Thomas Briese ◽  
Amit Kapoor ◽  
Nischay Mishra ◽  
Komal Jain ◽  
Arvind Kumar ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Viral Infections ◽  
Limit Of Detection ◽  
Virus Detection ◽  
Fold Increase ◽  
Sequencing Platform ◽  
Oligonucleotide Library ◽  
Capture Sequencing

ABSTRACT Insensitivity and technical complexity have impeded the implementation of high-throughput nucleic acid sequencing in differential diagnosis of viral infections in clinical laboratories. Here, we describe the development of a virome capture sequencing platform for vertebrate viruses (VirCapSeq-VERT) that increases the sensitivity of sequence-based virus detection and characterization. The system uses ~2 million probes that cover the genomes of members of the 207 viral taxa known to infect vertebrates, including humans. A biotinylated oligonucleotide library was synthesized on the NimbleGen cleavable array platform and used for solution-based capture of viral nucleic acids present in complex samples containing variable proportions of viral and host nucleic acids. The use of VirCapSeq-VERT resulted in a 100- to 10,000-fold increase in viral reads from blood and tissue homogenates compared to conventional Illumina sequencing using established virus enrichment procedures, including filtration, nuclease treatments, and RiboZero rRNA subtraction. VirCapSeq-VERT had a limit of detection comparable to that of agent-specific real-time PCR in serum, blood, and tissue extracts. Furthermore, the method identified novel viruses whose genomes were approximately 40% different from the known virus genomes used for designing the probe library. The VirCapSeq-VERT platform is ideally suited for analyses of virome composition and dynamics.IMPORTANCEVirCapSeq-VERT enables detection of viral sequences in complex sample backgrounds, including those found in clinical specimens, such as serum, blood, and tissue. The highly multiplexed nature of the system allows both the simultaneous identification and the comprehensive genetic characterization of all known vertebrate viruses, their genetic variants, and novel viruses. The operational simplicity and efficiency of the VirCapSeq-VERT platform may facilitate transition of high-throughput sequencing to clinical diagnostic as well as research applications.

scholarly journals Assessment of microbial community diversity in lakes of İğneada floodplain forest by metabarcoding approach

2021 ◽  
Vol 4 (4) ◽  
pp. 304-312
Author(s):  
E. Gözde Özbayram ◽  
brahim Halil Miraloğlu ◽  
Bahar İnce
Keyword(s):  
Bacterial Community ◽  
Bacterial Diversity ◽  
High Throughput Sequencing ◽  
Floodplain Forest ◽  
Community Diversity ◽  
Rrna Gene ◽  
Community Patterns ◽  
Biotechnological Potential ◽  
Sequencing Platform ◽  
The Family

This paper aims to contribute to the understanding of bacterial community patterns of the lakes of İğneada Floodplain Forest by metabarcoding approach. Within this scope, surface water samples were collected from three lakes located in the area namely Mert Lake, Hamam Lake, and Saka Lake, and the bacterial diversity was assessed by a high throughput sequencing of the 16S rRNA gene. Chao1 richness and Shannon diversity were higher in Saka Lake indicated a more diverse bacterial community. Proteobacteria was by far the most abundant phyla in all lakes. Although Bacteroidetes and Actinobacteria also dominated the community, their abundances differed in each lake. While the family Burkholderiaceae represented 25% of the bacterial community in Saka Lake, the abundances were 9% and 4% in Hamam Lake and Mert Lake, respectively. This study is one of the first investigations specifically focused on the bacterial communities in three lakes of İğneada Floodplain by next-generation sequencing platform and gave a prescreening of the bacterial diversity. Further studies are required to determine the biotechnological potential of this unique habitat.

scholarly journals Divergent RNA Viruses In Macrophomina phaseolina exhibit potential as virocontrol agents

2020 ◽  
Author(s):  
Jing Wang ◽  
Yunxia Ni ◽  
Xintao Liu ◽  
Hui Zhao ◽  
Yannong Xiao ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
Viral Evolution ◽  
Macrophomina Phaseolina ◽  
Phytopathogenic Fungus ◽  
Fungal Host ◽  
Viral Genomes ◽  
Geographical Regions ◽  
The Family ◽  
Viral Sequences ◽  
Complete Viral Genome

Abstract Macrophomina phaseolina is an important necrotrophic phytopathogenic fungus and cause extensive damage in many oilseed crops. Twelve M. phaseolina isolates with diverse biological phenotypes were selected for a high-throughput sequencing-based metatranscriptomic and bioinformatics analysis to identify viruses infecting M. phaseolina. The analysis identified 40 partial or nearly complete viral genome segments, 31 of which were novel viruses. Among these viral sequences, 43% of the viral genomes were double-stranded RNA (dsRNA), 47% were positive single-stranded RNA (ssRNA+), and the remaining 10% were negative sense-stranded RNA (ssRNA-). The 40 viruses showed affinity to 13 distinct viral lineages, including Bunyavirales (four viruses), Totiviridae (three viruses), Chrysoviridae (five viruses), Partitiviridae (four viruses), Hypoviridae (one virus), Endornaviridae (two viruses), Tombusviridae (three viruses), Narnaviridae (one virus), Potyviridae (one virus), Bromoviridae (one virus), Virgaviridae (six viruses), “Fusagraviridae” (five viruses), and Ourmiavirus (four viruses). Two viruses are closely related to two families, Potyviridae and Bromoviridae, which previously contained no mycovirus species. Moreover, nine novel viruses associated with M. phaseolina were identified in the family Totiviridae, Endornaviridae, and Partitiviridae. Coinfection with multiple viruses is prevalent in M. phaseolina, with each isolate harboring different numbers of viruses, ranging from three to eighteen. Furthermore, the effects of the viruses on the fungal host were analyzed according to the biological characteristics of each isolate. The results suggested that Macrophomina phaseolina hypovirus 2, Macrophomina phaseolina fusagravirus virus 1-5 (MpFV1-5), Macrophomina phaseolina endornavirus 1-2 (MpEV1-2), Macrophomina phaseolina ourmia-like virus 1-3 (MpOLV1-3), Macrophomina phaseolina mitovirus 4 (MpMV4), and Macrophomina phaseolina mycobunyavirus 1-4 (MpMBV1-4) were only detected in hypovirulent isolates. Those viruses associated with hypovirulence might be used as biological control agents as an environmentally friendly alternative to chemical fungicides. These findings considerably expand our understanding of mycoviruses in M. phaseolina and unvailed the presence of a huge difference among viruses in isolates from different hosts in distant geographical regions. Together, the present study provides new knowledge about viral evolution and fungus-virus coevolution.

scholarly journals LABRADOR—A Computational Workflow for Virus Detection in High-Throughput Sequencing Data

Viruses ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2541
Author(s):  
Izabela Fabiańska ◽  
Stefan Borutzki ◽  
Benjamin Richter ◽  
Hon Q. Tran ◽  
Andreas Neubert ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
De Novo ◽  
Virus Detection ◽  
Third Party ◽  
Reference Sequence ◽  
Clinical Samples ◽  
Sequencing Data

High-throughput sequencing (HTS) allows detection of known and unknown viruses in samples of broad origin. This makes HTS a perfect technology to determine whether or not the biological products, such as vaccines are free from the adventitious agents, which could support or replace extensive testing using various in vitro and in vivo assays. Due to bioinformatics complexities, there is a need for standardized and reliable methods to manage HTS generated data in this field. Thus, we developed LABRADOR—an analysis pipeline for adventitious virus detection. The pipeline consists of several third-party programs and is divided into two major parts: (i) direct reads classification based on the comparison of characteristic profiles between reads and sequences deposited in the database supported with alignment of to the best matching reference sequence and (ii) de novo assembly of contigs and their classification on nucleotide and amino acid levels. To meet the requirements published in guidelines for biologicals’ safety we generated a custom nucleotide database with viral sequences. We tested our pipeline on publicly available HTS datasets and showed that LABRADOR can reliably detect viruses in mixtures of model viruses, vaccines and clinical samples.

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