Mutation-Specific SARS-CoV-2 PCR Screen: Rapid and Accurate Detection of Variants of Concern and the Identification of a Newly Emerging Variant with Spike L452R Mutation
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Abstract
The emergence of more transmissible and/or more virulent severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) variants of concern (VOC) has triggered intensive genomic surveillance, which is costly and difficult to sustain operationally over the long term. To address this problem, we developed a set of four multiplex mutation-specific PCR-based assays with same-day reporting that can detect five VOC and three variants of interest (VOI), as defined in the March 2021 guidelines from the U.S.
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SciScore for 10.1101/2021.04.22.21255574: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable not detected. Randomization The set comprised 247 samples, including 156 SARS-CoV-2 positive swabs from February 2nd to April 1st 2021 that included all adequate samples with Ct values less than 35, and 91 samples collected from Jan 1st 2021 to Feb 1st 2021 with Ct values less than 35 that were randomly selected from adequate residual samples. Blinding not detected. Power Analysis not detected. Table 2: Resources
Software and Algorithms Sentences Resources Mutation Screening Assays: SARS-CoV-2 positive sample were screened by four multiplex rt-PCR assays, with results available same day. Mutation Screening Assayssuggested: (NIDDK Information Network, RRID:SCR_00…SciScore for 10.1101/2021.04.22.21255574: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable not detected. Randomization The set comprised 247 samples, including 156 SARS-CoV-2 positive swabs from February 2nd to April 1st 2021 that included all adequate samples with Ct values less than 35, and 91 samples collected from Jan 1st 2021 to Feb 1st 2021 with Ct values less than 35 that were randomly selected from adequate residual samples. Blinding not detected. Power Analysis not detected. Table 2: Resources
Software and Algorithms Sentences Resources Mutation Screening Assays: SARS-CoV-2 positive sample were screened by four multiplex rt-PCR assays, with results available same day. Mutation Screening Assayssuggested: (NIDDK Information Network, RRID:SCR_001606)All screening positive samples and a similar number of screening negative samples collected in the same period were sent for WGS. WGSsuggested: NoneGenomic Analyses and Strain Typing: Sequence analysis tools include custom pipelines utilizing GATK and Mutect2 (Broad Institute) and Dragen SARS-COVID variant detection (for CovidSeq assay). GATKsuggested: (GATK, RRID:SCR_001876)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:There are some limitations to a reflex mutant-PCR panel strategy. First, many diagnostic PCR platforms can deplete swab material, leaving an inadequate volume of residual sample for a multi-tube mutation screen. These assays also did not perform well when the diagnostic PCR Ct value is greater than 35 cycles. However, this problem is shared with amplicon-based WGS [25]. As demonstrated by our finding with L452R-positives, new variants of SARS-CoV-2 are still emerging so confirmation of some results by WGS will still be required. Finally, to evaluate for novel mutations, a sampling of screen-negative specimens for WGS will be an important component of a full surveillance strategy. In summary, the use of a panel of multiplex, mutant-specific rt-PCR assays represents an ideal balance of cost, turn-around-time and accuracy for detecting SARS-CoV-2 VOC and VOI. The technology is amenable for use in a larger number of clinical laboratories than next-generation sequencing. In our current environment requiring rapid strain typing to guide both treatment decisions and public health measures, such a rapid and accessible approach will be essential.
Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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