SARS-CoV-2 whole-genome sequencing using reverse complement PCR: For easy, fast and accurate outbreak and variant analysis.

Coolen, Jordy P. M.; Wolters, Femke; Tostmann, Alma; Groningen, Lenneke F.J. van; Bleeker‐Rovers, Chantal P.; Tan, Edward; Geest-Blankert, Nannet van der; Hautvast, Jeannine L A; Hopman, Joost; Wertheim, Heiman; Rahamat‐Langendoen, Janette; Storch, Marko; Melchers, Willem J. G.

doi:10.1016/j.jcv.2021.104993

Cited by 23 publications

(36 citation statements)

References 45 publications

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“…Sequence reads were cleaned, trimmed and mapped to the SARS-CoV-2 Wuhan-Hu-1 reference genome (NC_045512.2). 7 For each sample, genomic variants were called and filtered by quality, with high quality variant calls being used to generate the consensus genome sequence for each sample. The consensus genome sequence was then processed using Pangolin, 8 a widely used computational tool that assigns the most likely lineage to a given SARS-CoV-2 genome sequence according to the Pango dynamic lineage nomenclature scheme.…”

Section: Methodsmentioning

confidence: 99%

A Randomised -Controlled Phase 2 trial of Molnupiravir in Unvaccinated and Vaccinated Individuals with Early SARS-CoV-2

Khoo

Fitzgerald

Saunders

et al. 2022

Preprint

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Background Molnupiravir was licensed for treating high-risk patients with COVID-19 based on data from unvaccinated adults. AGILE CST-2 (NCT04746183) Phase II reports safety and virological efficacy of molnupiravir in vaccinated and unvaccinated individuals. Methods Adult out-patients with PCR-confirmed SARS-CoV-2 infection within five days of symptom onset were randomly assigned 1:1 to receive molnupiravir (800mg twice daily for five days) or placebo. The primary outcome was time to swab PCR-negativity, compared using a Bayesian model for estimating the probability of a superior virological response (Hazard Ratio>1) for molnupiravir over placebo. Secondary outcomes included change in viral titre at day 5, safety and tolerability, clinical progression and patient reported outcome measures. We analysed outcomes after the last participant reached day 29. Findings Of 180 participants randomised (90 molnupiravir, 90 placebo), 50% were vaccinated. Infections with SARS-CoV-2 variants Delta (40%), Alpha (21%), Omicron (21%) and EU1 (16%) were represented. The median time to negative-PCR was 8 versus 11 days for molnupiravir and placebo (HR=1.30, 95% CrI 0.92-1.71, p=0.7 by Logrank and p=0.03 by Breslow-Gehan tests). Although small numbers precluded subgroup analysis, no obvious differences were observed between vaccinated and unvaccinated participants. Using a two-point prior the probability of molnupiravir being superior to placebo (HR>1) was 75.4%, which was just below our defined threshold of 80% for establishing superiority. Using an uninformative continuous prior, the probability of HR>1 was 94.7%. As an exploratory analysis, the change in viral titre on day 5 (end of treatment) was significantly greater with molnupiravir compared with placebo. A total of 4 participants reported severe adverse events (grade 3+), 3 of whom were in the placebo arm. Interpretation We found molnupiravir to be well-tolerated, with evidence for high probability of antiviral efficacy in a population of vaccinated and unvaccinated individuals infected with a broad range of viral variants.

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Section: Methodsmentioning

confidence: 99%

A Randomised -Controlled Phase 2 trial of Molnupiravir in Unvaccinated and Vaccinated Individuals with Early SARS-CoV-2

Khoo

Fitzgerald

Saunders

et al. 2022

Preprint

View full text Add to dashboard Cite

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“…Samples with low RNA concentration (Ct ≥ 30) often produce low-quality or incomplete genomes without targeted intervention to improve the results of these samples 12–14 .We observed that such high-Ct samples generally produce fewer on-target sequencing reads than low-Ct samples. Consequently, we wanted to test whether greatly increasing sequencing depth of these samples would allow high Ct samples to be reconstructed.…”

Section: Resultsmentioning

confidence: 85%

“…Samples with low RNA concentration (Ct ≥ 30) often produce low-quality or incomplete genomes without targeted intervention to improve the results of these samples [12][13][14] . We observed that such high-Ct samples generally produce fewer on-target sequencing reads than low-Ct samples.…”

Section: Assessment Of Over-sequencing Of High Ct Samples As a Strate...mentioning

confidence: 99%

Development and scaling of a sequencing pipeline for genomic surveillance of SARS-CoV-2 in New York City

Hammerling¹,

Kang²,

Ward³

et al. 2022

Preprint

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In the ongoing COVID-19 pandemic, detecting the appearance and spread of variants of concern (VOC) is a critical capability in the fight to quell the virus and return to normalcy. Genomic surveillance of the emergence, propagation, and geographical spread of VOCs is thus an important tool for public health officials and government leaders to make policy decisions and advise the public. As part of our role as a major SARS-CoV-2 diagnostic testing facility in New York City, the Pandemic Response Lab (PRL) has been performing genomic surveillance on the large number of positive samples processed by the facility on a daily basis from throughout the New York metropolitan area. Here we describe the development and optimization of a high-throughput SARS-CoV-2 genome sequencing facility at PRL serving New York City.

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“… 44 Novel whole‐genome sequencing technologies based on the EasySeqTM RC‐PCR SARS‐CoV‐2 WGS kit and RT‐PCR have proven to be useful for high‐throughput detection of mutant strains of SARS‐CoV‐2. 46 Vega‐Magaña et al designed three specific primers and probes for qRT‐PCR detection based on the N501Y, 69‐70del, K417N, and E484K S mutations, which played an important role in detecting the E484K mutation and P.2 mutant strains. 47 Exploiting the good selectivity and self‐quenching properties ascribed to molecular beacons, researchers developed a two‐tube multiplex qRT‐PCR detection method that can identify present viruses of concern (VOCs) via the detection of eight different mutation sites in the S protein.…”

Section: Nucleic Acid‐based Sars‐cov ‐2 Detectionmentioning

confidence: 99%

An updated review of SARS‐CoV‐2 detection methods in the context of a novel coronavirus pandemic

Zhang

Huang

Zhu

et al. 2022

Bioengineering & Transla Med

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The World Health Organization has reported approximately 430 million confirmed cases of coronavirus disease 2019 (COVID‐19), caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), worldwide, including nearly 6 million deaths, since its initial appearance in China in 2019. While the number of diagnosed cases continues to increase, the need for technologies that can accurately and rapidly detect SARS‐CoV‐2 virus infection at early phases continues to grow, and the Federal Drug Administration (FDA) has licensed emergency use authorizations (EUAs) for virtually hundreds of diagnostic tests based on nucleic acid molecules and antigen–antibody serology assays. Among them, the quantitative real‐time reverse transcription PCR (qRT‐PCR) assay is considered the gold standard for early phase virus detection. Unfortunately, qRT‐PCR still suffers from disadvantages such as the complex test process and the occurrence of false negatives; therefore, new nucleic acid detection devices and serological testing technologies are being developed. However, because of the emergence of strongly infectious mutants of the new coronavirus, such as Alpha (B.1.1.7), Delta (B.1.617.2), and Omicron (B.1.1.529), the need for the specific detection of mutant strains is also increasing. Therefore, this article reviews nucleic acid‐ and antigen–antibody‐based serological assays, and compares the performance of some of the most recent FDA‐approved and literature‐reported assays and associated kits for the specific testing of new coronavirus variants.

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SARS-CoV-2 whole-genome sequencing using reverse complement PCR: For easy, fast and accurate outbreak and variant analysis.

Cited by 23 publications

References 45 publications

A Randomised -Controlled Phase 2 trial of Molnupiravir in Unvaccinated and Vaccinated Individuals with Early SARS-CoV-2

A Randomised -Controlled Phase 2 trial of Molnupiravir in Unvaccinated and Vaccinated Individuals with Early SARS-CoV-2

Development and scaling of a sequencing pipeline for genomic surveillance of SARS-CoV-2 in New York City

An updated review of SARS‐CoV‐2 detection methods in the context of a novel coronavirus pandemic

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