An amplicon-based protocol for whole-genome sequencing of human respiratory syncytial virus subgroup A.
Biol Methods Protoc
; 9(1): bpae007, 2024.
Article
em En
| MEDLINE
| ID: mdl-38371356
ABSTRACT
It is convenient to study complete genome sequences of human respiratory syncytial virus (hRSV) for ongoing genomic characterization and identification of highly transmissible or pathogenic variants. Whole genome sequencing of hRSV has been challenging from respiratory tract specimens with low viral loads. Herein, we describe an amplicon-based protocol for whole genome sequencing of hRSV subgroup A validated with 24 isolates from nasopharyngeal swabs and infected cell cultures, which showed cycle threshold (Ct) values ranging from 10 to 31, as determined by quantitative reverse-transcription polymerase chain reaction. MinION nanopore generated 3200 to 5400 reads per sample to sequence over 93% of the hRSV-A genome. Coverage of each contig ranged from 130× to 200×. Samples with Ct values of 20.9, 25.2, 27.1, 27.7, 28.2, 28.8, and 29.6 led to the sequencing of over 99.0% of the virus genome, indicating high genome coverage even at high Ct values. This protocol enables the identification of hRSV subgroup A genotypes, as primers were designed to target highly conserved regions. Consequently, it holds potential for application in molecular epidemiology and surveillance of this hRSV subgroup.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
Biol Methods Protoc
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
México
País de publicação:
Reino Unido