Evaluation of viral RNA recovery methods in vectors by metagenomic sequencing.
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Author(s)
Akello, JO
Leib, SL
Engler, O
Beuret, C
Type
Journal Article
Abstract
Identification and characterization of viral genomes in vectors including ticks and mosquitoes positive for pathogens of great public health concern using metagenomic next generation sequencing (mNGS) has challenges. One such challenge is the ability to efficiently recover viral RNA which is typically dependent on sample processing. We evaluated the quantitative effect of six different extraction methods in recovering viral RNA in vectors using negative tick homogenates spiked with serial dilutions of tick-borne encephalitis virus (TBEV) and surrogate Langat virus (LGTV). Evaluation was performed using qPCR and mNGS. Sensitivity and proof of concept of optimal method was tested using naturally positive TBEV tick homogenates and positive dengue, chikungunya, and Zika virus mosquito homogenates. The amount of observed viral genome copies, percentage of mapped reads, and genome coverage varied among different extractions methods. The developed Method 5 gave a 120.8-, 46-, 2.5-, 22.4-, and 9.9-fold increase in the number of viral reads mapping to the expected pathogen in comparison to Method 1, 2, 3, 4, and 6, respectively. Our developed Method 5 termed ROVIV (Recovery of Viruses in Vectors) greatly improved viral RNA recovery and identification in vectors using mNGS. Therefore, it may be a more sensitive method for use in arbovirus surveillance.
Date Issued
2020-05-19
Online Publication Date
2021-06-24T15:22:02Z
Date Acceptance
2020-05-18
ISSN
1999-4915
Publisher
MDPI AG
Start Page
1
End Page
17
Journal / Book Title
Viruses
Volume
12
Issue
5
Copyright Statement
© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/)
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/)
License URI
Identifier
https://www.mdpi.com/1999-4915/12/5/562
https://www.ncbi.nlm.nih.gov/pubmed/32438629
v12050562
Subjects
NGS
metagenomics
sample processing
vector-borne viruses
vectors
Animals
Chikungunya Fever
Chlorocebus aethiops
Encephalitis Viruses, Tick-Borne
Encephalitis, Tick-Borne
Genome, Viral
High-Throughput Nucleotide Sequencing
Metagenomics
Mosquito Vectors
RNA, Viral
Real-Time Polymerase Chain Reaction
Ticks
Vero Cells
Zika Virus
Zika Virus Infection
Vero Cells
Animals
Ticks
Encephalitis Viruses, Tick-Borne
Encephalitis, Tick-Borne
RNA, Viral
Genome, Viral
Metagenomics
High-Throughput Nucleotide Sequencing
Real-Time Polymerase Chain Reaction
Chikungunya Fever
Zika Virus
Zika Virus Infection
Mosquito Vectors
Chlorocebus aethiops
NGS
metagenomics
sample processing
vector-borne viruses
vectors
Animals
Chikungunya Fever
Chlorocebus aethiops
Encephalitis Viruses, Tick-Borne
Encephalitis, Tick-Borne
Genome, Viral
High-Throughput Nucleotide Sequencing
Metagenomics
Mosquito Vectors
RNA, Viral
Real-Time Polymerase Chain Reaction
Ticks
Vero Cells
Zika Virus
Zika Virus Infection
0605 Microbiology
Publication Status
Published
Country
Switzerland
Date Publish Online
2020-05-19