Comparison of ordinary reverse transcription real-time polymerase chain reaction (qRT-PCR) with a newly developed one-step single-tube nested real-time RT-PCR (OSN-qRT-PCR) for sensitive detection of SARS-CoV-2 in wastewater.
Environ Sci Pollut Res Int
; 30(42): 95579-95589, 2023 Sep.
Article
em En
| MEDLINE
| ID: mdl-37553492
ABSTRACT
Wastewater monitoring has proven to be an important approach to detecting and controlling the development of the SARS-CoV-2 pandemic. Various tests based on reverse transcription real-time PCR (qRT-PCR) have been developed and used for the detection of SARS-CoV-2 in wastewater samples. In this study, we attempted to increase the sensitivity of qRT-PCR by developing a one-step single-tube nested qRT-PCR assay (OSN-qRT-PCR). Two variants were developed, oriented to nucleocapsid phosphoprotein gene (N) and to spike protein gene (S), respectively. The performance of conventional qRT-PCR assays oriented to these genes with two novel OSN-qRT-PCR assays were firstly optimized using wastewater artificially contaminated with two encapsidated RNA mimic systems harboring a portion either N or S gene (ENRM and ESRM, respectively). The assays were coupled to a polyethylene glycol-based RNA precipitation/extraction method and applied to detect SARS-CoV-2 in wastewater samples from four cities in Slovakia. Both novel OSN-qRT-PCR assays demonstrated higher detection rates than the ordinary qRT-PCR counterparts. The virus levels in the analyzed wastewater samples had a high or very high relation with the numbers of clinical cases in the monitored regions. In fact, correlation with a 3-, 4-, or 5-day temporal offset was revealed. The OSN-qRT-PCR assays demonstrated robustness, mainly in samples with low viral loads.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
SARS-CoV-2
/
COVID-19
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Revista:
Environ Sci Pollut Res Int
Assunto da revista:
SAUDE AMBIENTAL
/
TOXICOLOGIA
Ano de publicação:
2023
Tipo de documento:
Article
País de afiliação:
Eslováquia