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Digital PCR is a sensitive new technique for SARS-CoV-2 detection in clinical applications.

ABSTRACT: The global coronavirus disease 2019 (COVID-19) pandemic has posed great challenges in people's daily lives. Highly sensitive laboratory techniques played a critical role in clinical COVID-19 diagnosis and management. In this study the feasibility of using a new digital PCR-based detection assay for clinical COVID-19 diagnosis was investigated by comparing its performance with that of RT-PCR. Clinical patient samples and samples obtained from potentially contaminated environments were analyzed. The study included 10 patients with confirmed COVID-19 diagnoses, 32 validated samples of various types derived from different clinical timepoints and sites, and 148 environmentally derived samples. SARS-CoV-2 nucleic acids were more readily detected in respiratory tract samples (35.0%). In analyses of environmentally derived samples, the positivity rate of air samples was higher than that of surface samples, probably due to differences in virus concentrations. Digital PCR detected SARS-CoV-2 in several samples that had previously been deemed negative, including 3 patient-derived samples and 5 environmentally derived samples. In this study digital PCR exhibited higher sensitivity than conventional RT-PCR, suggesting that it may be a useful new method for clinical SARS-CoV-2 detection. Improvement of SARS-CoV-2 detection would substantially reduce the rates of false-negative COVID-19 test results, in particular those pertaining to asymptomatic carriers.

SUBMITTER: Jiang Y

PROVIDER: S-EPMC7641518 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Digital PCR is a sensitive new technique for SARS-CoV-2 detection in clinical applications.

Jiang Yanfang Y Wang Haifeng H Hao Sijia S Chen Yukun Y He Jiaxue J Liu Yong Y Chen Liguo L Yu Yuanhua Y Hua Shucheng S

Clinica chimica acta; international journal of clinical chemistry 20201104

The global coronavirus disease 2019 (COVID-19) pandemic has posed great challenges in people's daily lives. Highly sensitive laboratory techniques played a critical role in clinical COVID-19 diagnosis and management. In this study the feasibility of using a new digital PCR-based detection assay for clinical COVID-19 diagnosis was investigated by comparing its performance with that of RT-PCR. Clinical patient samples and samples obtained from potentially contaminated environments were analyzed. T ...[more]

PMID: 33159953

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Project description:Wastewater based epidemiology (WBE) has emerged as a strategy to identify, locate, and manage outbreaks of COVID-19, and thereby possibly prevent surges in cases, which overwhelm local to global health care networks. The WBE process is based on assaying municipal wastewater for molecular markers of the SARS-CoV-2 virus. Standard processes for purifying viral RNA from municipal wastewater are often time-consuming and require the handling of large quantities of wastewater, negatively affecting throughput, timely reporting, and safety. We demonstrate here an automated, faster system to purify viral RNA from smaller volumes of wastewater but with increased sensitivity for detection of SARS-CoV-2 markers. We document the effectiveness of this new approach by way of comparison to the PEG/NaCl/Qiagen method prescribed by the State of Michigan for SARS-CoV-2 wastewater monitoring and show its application to several Detroit sewersheds. Specifically, compared to the PEG/NaCl/Qiagen method, viral RNA purification using the PerkinElmer Chemagic™ 360 lowered handling time, decreased the amount of wastewater required by ten-fold, increased the amount of RNA isolated per μl of final elution product by approximately five-fold, and effectively removed ddPCR inhibitors from most sewershed samples. For detection of markers on the borderline of viral detectability, we found that use of the Chemagic™ 360 enabled the measurement of viral markers in a significant number of samples for which the result with the PEG/NaCl/Qiagen method was below the level of detectability. The improvement in detectability of the viral markers might be particularly important for early warning to public health authorities at the beginning of an outbreak. Applied to sewersheds in Detroit, the technique enabled more sensitive detection of SARS-CoV-2 markers with good correlation between wastewater signals and COVID-19 cases in the sewersheds. We also discuss advantages and disadvantages of several automated RNA purification systems, made by Promega, PerkinElmer, and ThermoFisher.

Dataset Information

Digital PCR is a sensitive new technique for SARS-CoV-2 detection in clinical applications.

Publications

Digital PCR is a sensitive new technique for SARS-CoV-2 detection in clinical applications.

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