Clinical use of targeted high-throughput whole-genome sequencing for a dengue virus variant

Lee, Chun Kiat; Chua, Cui Wen; Chiu, Lily; Koay, Evelyn Siew-Chuan

doi:10.1515/cclm-2016-0660

Cited by 4 publications

(3 citation statements)

References 7 publications

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“…Historically, segmental reverse transcription-PCR (RT-PCR) combined with Sanger DNA sequencing of the DENV E gene (about 1,600 bp to 1,800 bp) was successfully applied ( 10 , 30 ). With the requirement of accuracy in molecular epidemiological studies, WGS of DENV has been gradually applied ( 22 , 25 , 31 – 34 ). NGS technologies have recently enabled large-scale surveillance of infectious diseases, and the power and utility of NGS are based on its massively parallel interrogation of nucleic acids.…”

Section: Discussionmentioning

confidence: 99%

A Serotype-Specific and Multiplex PCR Method for Whole-Genome Sequencing of Dengue Virus Directly from Clinical Samples

Jiang

et al. 2022

Microbiol Spectr

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Section: Discussionmentioning

confidence: 99%

A Serotype-Specific and Multiplex PCR Method for Whole-Genome Sequencing of Dengue Virus Directly from Clinical Samples

Jiang

et al. 2022

Microbiol Spectr

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“…Analysis of the DNA sequence of an organism's genome, in its entirety and determined at a single time, is known as whole genome sequencing (WGS). Its application to DENV reveals the complete sequence of the viral genome and facilitates molecular characterization of the virus and identification of significant genetic markers that may be associated with virulence properties [46,47]. An initial structural comparison between genomes of DENV isolates recovered from cases of DF and DHF revealed that charge differences among six encoded amino acids existed in structural (prM and E) and NS (NS4b and NS5) genes [48].…”

Section: Role Of the Viral Genome In Denv Virulencementioning

confidence: 99%

Dengue Serotypes in Bangladesh: Whole Genome Sequencing and Comparative Genomics Facilitates Pathogenesis and Epidemiology Studies and Informs Improved Disease Control

Tabassum¹,

Taylor‐Robinson²

2019

Microbiol Infect Dis

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Dengue virus (DENV) is one of the most extensive arthropod-borne (arbo)viruses worldwide, especially prevalent in tropical and subtropical countries, and which is responsible for causing an array of clinical disease manifestations in humans. The increasing frequency of dengue outbreaks and the expanding range over which both the virus and its vector of transmission, female Aedes mosquitoes, are endemic, are pressing global public health concerns. The ongoing lack of either an efficacious vaccine or antiviral drug has contributed to the escalating problem. Inadequate knowledge of DENV genomic architecture and pathogenesis has proven to be a major drawback when designing effective prevention and treatment options. Mounting case numbers of dengue infection are reported each year in Bangladesh, yet no full-length genome sequence data are available for DENV isolates from this densely populated, developing South Asian nation. Sequencing and characterization of the whole genome of Bangladeshi DENV isolates of different virus serotypes is therefore an important priority in order to identify therapeutic target determinants against which to develop effective measures to combat the disease.

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“…Alternatively, HTS can be used in a targeted metagenomics approach in which a PCR step is used to pre-amplify viral sequences before sequencing. In recent years, HTS has been successfully used as a tool for identification of DENV directly from clinical samples [6,7]. This also allows the rapid identification of the serotype and genotype important for disease management as the genotype may be associated with disease outcome [8].…”

Section: Introductionmentioning

confidence: 99%

DEN-IM: Dengue Virus identification from shotgun and targeted metagenomics

Mendes

Lizarazo

Machado

et al. 2019

Preprint

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Dengue virus (DENV) represents a public health and economic burden in affected countries. The availability of genomic data is key to understand viral evolution and dynamics, supporting improved control strategies. Currently, the use of High Throughput Sequencing (HTS) technologies, which can be applied both directly to patient samples (shotgun metagenomics) and to PCR amplified viral sequences (targeted metagenomics), is the most informative approach to monitor the viral dissemination and genetic diversity.Despite many advantages, these technologies require bioinformatics expertise and appropriate infrastructure for the analysis and interpretation of the resulting data. In addition, the many software solutions available can hamper reproducibility and comparison of results.Here we present DEN-IM, a one-stop, user-friendly, containerised and reproducible workflow for the analysis of DENV sequencing data, both from shotgun and targeted metagenomics approaches. It is able to infer DENV coding sequence (CDS), identify serotype and genotype, and generate a phylogenetic tree. It can easily be run on any UNIX-like system, from local machines to high-performance computing clusters, performing a comprehensive analysis without the requirement of extensive bioinformatics expertise.Using DEN-IM, we successfully analysed two DENV datasets. The first comprised 25 shotgun metagenomic sequencing samples of varying serotype and genotype, including a spiked sample containing the existing four serotypes. The second dataset consisted of 106 targeted metagenomics samples of DENV 3 genotype III where DEN-IM allowed detection of the intra-genotype diversity.The DEN-IM workflow, parameters and execution configuration files, and documentation are freely available at https://github.com/B-UMMI/DEN-IM. The Dengue virus (DENV), a single-stranded positive-sense RNA virus belonging to the Flavivirus 2 genus, is one of the most prevalent arboviruses and is mainly concentrated in tropical and subtropical 3 regions. Infection with DENV results in symptoms ranging from mild fever to haemorrhagic fever and 4 shock syndrome [1]. Transmission to humans occurs through the bite of Aedes mosquitoes namely 5 Aedes aegypti and Aedes albopictus [2] In 2010, it was predicted that the burden of dengue disease 6 reached 390 million cases per year worldwide [3]. The high morbidity and mortality of dengue makes 7 it the arbovirus with the highest clinical significance [4]. 8 The viral genome of ∼11,000 nucleotides, consists of a Coding Sequence (CDS) of approximately 910.2 Kb that is translated into a single polyprotein encoding three structural proteins (capsid -C, 10 premembrane -prM, envelope -E) and seven non-structural proteins (NS1, NS2A, NS2B, NS3, NS4A, 11 NS4B and NS5. Additionally, the genome contains two Non-Coding Regions (NCRs) at their 5' and 12 3' ends [5]. 13DENV can be classified into four serotypes (1, 2, 3 and 4), differing from each other by 25% to 14 40% at the amino acid level. They are further classified into genotypes that vary by up to ∼...

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Clinical use of targeted high-throughput whole-genome sequencing for a dengue virus variant

Cited by 4 publications

References 7 publications

A Serotype-Specific and Multiplex PCR Method for Whole-Genome Sequencing of Dengue Virus Directly from Clinical Samples

A Serotype-Specific and Multiplex PCR Method for Whole-Genome Sequencing of Dengue Virus Directly from Clinical Samples

Dengue Serotypes in Bangladesh: Whole Genome Sequencing and Comparative Genomics Facilitates Pathogenesis and Epidemiology Studies and Informs Improved Disease Control

DEN-IM: Dengue Virus identification from shotgun and targeted metagenomics

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