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J Virol Methods ; 294: 114174, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-33984396

RESUMEN

There is growing evidence that measurement of SARS-CoV-2 viral copy number can inform clinical and public health management of SARS-CoV-2 carriers and COVID-19 patients. Here we show that quantification of SARS-CoV-2 is feasible in a clinical setting, using a duplex RT-qPCR assay which targets both the E gene (Charité assay) and a human RNA transcript, RNase P (CDC assay) as an internal sample sufficiency control. Samples in which RNase P is not amplified indicate that sample degradation has occurred, PCR inhibitors are present, RNA extraction has failed or swabbing technique was insufficient. This important internal control reveals that 2.4 % of nasopharyngeal swabs (15/618 samples) are inadequate for SARS-CoV-2 testing which, if not identified, could result in false negative results. We show that our assay is linear across at least 7 logs and is highly reproducible, enabling the conversion of Cq values to viral copy numbers using a standard curve. Furthermore, the SARS-CoV-2 copy number was independent of the RNase P copy number indicating that the per-swab viral copy number is not dependent on sampling- further allowing comparisons between samples. The ability to quantify SARS-CoV-2 viral copy number will provide an important opportunity for viral burden-guided public health and clinical decision making.


Asunto(s)
Prueba de Ácido Nucleico para COVID-19/métodos , Prueba de Ácido Nucleico para COVID-19/normas , ARN Viral/genética , SARS-CoV-2/genética , Manejo de Especímenes/normas , COVID-19/diagnóstico , COVID-19/virología , Dosificación de Gen , Genes Esenciales , Humanos , Límite de Detección , ARN Viral/aislamiento & purificación , Estándares de Referencia , Ribonucleasa P/genética , Manejo de Especímenes/métodos , Carga Viral
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