Evaluation of Viral RNA Recovery Methods in Vectors by Metagenomic Sequencing.

Akello, Joyce Odeke; Leib, Stephen L; Engler, Olivier; Beuret, Christian (2020). Evaluation of Viral RNA Recovery Methods in Vectors by Metagenomic Sequencing. Viruses, 12(5) Molecular Diversity Preservation International MDPI 10.3390/v12050562

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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.

Item Type:

Journal Article (Original Article)


04 Faculty of Medicine > Service Sector > Institute for Infectious Diseases

Graduate School:

Graduate School for Cellular and Biomedical Sciences (GCB)

UniBE Contributor:

Akello, Joyce Odeke and Leib, Stephen L


500 Science > 570 Life sciences; biology
600 Technology > 610 Medicine & health




Molecular Diversity Preservation International MDPI




Stephen Leib

Date Deposited:

08 Jun 2020 10:45

Last Modified:

08 Jun 2020 10:45

Publisher DOI:


PubMed ID:


Uncontrolled Keywords:

NGS metagenomics sample processing vector-borne viruses vectors





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