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The emergence of the SARS-CoV-2 virus and the exponential growth of COVID-19 cases have created a major crisis for public health systems. The critical identification of contagious asymptomatic carriers requires the isolation of viral nucleic acids, reverse transcription, and amplification by PCR. However, the shortage of specific proprietary reagents or the lack of automated platforms have seriously hampered diagnostic throughput in many countries. Here, we provide a procedure for SARS-CoV-2 detection for diagnostic purposes from clinical samples in the setting of a basic research molecular biology lab. The procedure details the necessary steps for daily analysis of up to 500 clinical samples with a team composed of 12 experienced researchers. The protocol has been designed to rely on widely available reagents and devices, to cope with heterogeneous clinical specimens, to guarantee nucleic acid extraction from very scarce biological material, and to minimize the rate of false-negative results.
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http://dx.doi.org/10.3390/mps3030059 | DOI Listing |
PLOS Glob Public Health
September 2025
Department of Biology, University of Ottawa, Ottawa, Ontario, Canada.
Built environment surveillance has shown promise for monitoring COVID-19 burden at granular geographic scales, but its utility for surveillance across larger areas and populations is unknown. Our study aims to evaluate the role of built environment detection of SARS-CoV-2 for the surveillance of COVID-19 across broad geographies and populations. We conducted a prospective city-wide sampling study to examine the relationship between SARS-CoV-2 on floors and COVID-19 burden.
View Article and Find Full Text PDFJ Infect Dev Ctries
August 2025
Department of Medical Microbiology, Faculty of Medicine, Ege University, Izmir 35100, Turkey.
Introduction: The aim of this study was to compare the performance of different clinical specimens-nasopharyngeal (NP) swabs collected by healthcare professionals (HCP-NP), self-collected nasal swabs (Sc-N), and saliva samples (S)-in diagnostic tests for investigating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA and influenza A/B RNA.
Methodology: These clinical samples were collected from 404 symptomatic cases and tested with the SARS-CoV-2 and influenza A/B RNA tests on the cobas 6800 System of Roche Molecular Systems (Roche Molecular Systems, Pleasanton, USA). The SARS-CoV-2 or influenza virus infection status was determined for all patients based on the predefined criteria and corresponding algorithms.
Microbiol Spectr
September 2025
Laboratoy of Virology, Microbiology Department, Hospital Universitario 12 de Octubre, Madrid, Spain.
Millions of reverse transcription-polymerase chain reaction (RT-PCR) tests have been performed worldwide during the SARS-CoV-2 pandemic, using various protocols. This study evaluates the duration of SARS-CoV-2 RNA detectability by RT-PCR at body temperature and analyzes changes in cycle threshold (Ct) values over time. Positive nasopharyngeal swabs for SARS-CoV-2 RT-PCR ( = 120) with different Ct values were collected from Hospital Universitario 12 de Octubre (Madrid, Spain, 2020).
View Article and Find Full Text PDFMicrobiol Spectr
September 2025
Department of Clinical Microbiology, Hospital Clínic of Barcelona-ISGlobal, University of Barcelona, Barcelona, Spain.
Unlabelled: Accurate methods to assess viral viability are crucial for determining isolation duration and antiviral therapy in immunocompromised patients. Although cell culture (CC) is the gold standard, it has limitations. Cycle threshold (Ct) values from genomic RNA (gRNA) RT-PCR and subgenomic RNA (sgRNA) RT-PCR have been proposed as markers of active viral replication.
View Article and Find Full Text PDF