Rapid, Sensitive, and Specific Severe Acute Respiratory Syndrome Coronavirus 2 Detection: A Multicenter Comparison Between Standard Quantitative Reverse-Transcriptase Polymerase Chain Reaction and CRISPR-Based DETECTR
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Abstract
Background
Recent advances in CRISPR-based diagnostics suggest that DETECTR, a combination of reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) and subsequent Cas12 bystander nuclease activation by amplicon-targeting ribonucleoprotein complexes, could be a faster and cheaper alternative to quantitative reverse-transcription polymerase chain reaction (qRT-PCR) without sacrificing sensitivity and/or specificity.
Methods
In this study, we compare DETECTR with qRT-PCR to diagnose coronavirus disease 2019 on 378 patient samples. Patient sample dilution assays suggest a higher analytical sensitivity of DETECTR compared with qRT-PCR; however, this was not confirmed in this large patient cohort, where we report 95% reproducibility between the 2 tests.
Results
These data showed that both techniques are equally sensitive in detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) providing additional value of DETECTR to the currently used qRT-PCR platforms. For DETECTR, different guide ribonucleic acids can be used simultaneously to obviate negative results due to mutations in N-gene. Lateral flow strips, suitable as a point-of-care test, showed a 100% correlation to the high-throughput DETECTR assay. More importantly, DETECTR was 100% specific for SARS-CoV-2 relative to other human coronaviruses.
Conclusions
Because there is no need for specialized equipment, DETECTR could be rapidly implemented as a complementary technically independent approach to qRT-PCR thereby increasing the testing capacity of medical microbiological laboratories and relieving the existent PCR platforms for routine non-SARS-CoV-2 diagnostic testing.
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SciScore for 10.1101/2020.07.27.20147249: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Software and Algorithms Sentences Resources All statistics were analyzed in Graphpad Prism version 8.0.2. Graphpad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)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: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from Trial…
SciScore for 10.1101/2020.07.27.20147249: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Software and Algorithms Sentences Resources All statistics were analyzed in Graphpad Prism version 8.0.2. Graphpad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)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: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.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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SciScore for 10.1101/2020.07.27.20147249: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding -7 -16 In a pilot experiment we blindly tested a small cohort of patient samples including four positive, four negative and four samples with not interpretable (NI) qRT-PCR results. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Software and Algorithms Sentences Resources All statistics were analyzed in Graphpad Prism version 8.0.2. Graphpad Prismsuggested: (GraphPad Prism, SCR_002798)This emphasizes the importance of a separate amplification step (PCR or LAMP) prior to Cas12 detection (Figure 1C and … SciScore for 10.1101/2020.07.27.20147249: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding -7 -16 In a pilot experiment we blindly tested a small cohort of patient samples including four positive, four negative and four samples with not interpretable (NI) qRT-PCR results. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Software and Algorithms Sentences Resources All statistics were analyzed in Graphpad Prism version 8.0.2. Graphpad Prismsuggested: (GraphPad Prism, SCR_002798)This emphasizes the importance of a separate amplification step (PCR or LAMP) prior to Cas12 detection (Figure 1C and Supplemental figure 1D). LAMPsuggested: (LAMP, SCR_001740)Data from additional tools added to each annotation on a weekly basis.
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