A simplified, amplicon-based method for whole genome sequencing of human respiratory syncytial viruses.
J Clin Virol
; 161: 105423, 2023 04.
Article
en En
| MEDLINE
| ID: mdl-36934591
ABSTRACT
BACKGROUND:
Human Respiratory Syncytial Virus (RSV) infections pose a significant risk to human health worldwide, especially for young children. Whole genome sequencing (WGS) provides a useful tool for global surveillance to better understand the evolution and epidemiology of RSV and provide essential information that may impact on antibody treatments, antiviral drug sensitivity and vaccine effectiveness.OBJECTIVES:
Here we report the development of a rapid and simplified amplicon-based one-step multiplex reverse-transcription polymerase chain reaction (mRT-PCR) for WGS of both human RSV-A and RSV-B viruses. STUDYDESIGN:
Two mRT-PCR reactions for each sample were designed to generate amplicons for RSV WGS. This new method was tested and evaluated by sequencing 206 RSV positive clinical samples collected in Australia in 2020 and 2021 with RSV Ct values between 10 and 32.RESULTS:
In silico analysis and laboratory testing revealed that the primers used in the new method covered most of the currently circulating RSV-A and RSV-B. Amplicons generated were suitable for both Illumina and Oxford Nanopore Technologies (ONT) NGS platforms. A success rate of 83.5% with a full coverage for the genome of 98 RSV-A and 74 RSV-B was achieved from all clinical samples tested.CONCLUSIONS:
This assay is simple to set up, robust, easily scalable in sample preparation and relatively inexpensive, and as such, provides a valuable addition to existing NGS RSV WGS methods.Palabras clave
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Virus Sincitial Respiratorio Humano
/
Infecciones por Virus Sincitial Respiratorio
Tipo de estudio:
Diagnostic_studies
Límite:
Child
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Child, preschool
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Humans
Idioma:
En
Revista:
J Clin Virol
Asunto de la revista:
VIROLOGIA
Año:
2023
Tipo del documento:
Article