Multicenter international assessment of a SARS-CoV-2 RT-LAMP test for point of care clinical application

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1. Version published to 10.1371/journal.pone.0268340

   May 11, 2022
2. 

   ScreenIT

   Mar 3, 2022

   SciScore for 10.1101/2022.02.27.22271548: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Ethics	IRB: These samples were surplus diagnostic materials that were analyzed anonymously, and no specific approval from Research Ethics Board (REB) was required.
   Sex as a biological variable	not detected.
   Randomization	Batch 2: 120 positive and 120 negative samples were randomly chosen, and re-evaluated to confirm SARS-CoV-2 condition and sample quality.
   Blinding	not detected.
   Power Analysis	not detected.

   Table 2: Resources

   Software and Algorithms
   Sentences	Resources
   The RT-PCR reaction was carried out with CFX 384 Real-Time System (Bio-Rad Laboratories) operated with Bio-Rad CFX Manager 3.1, and the plate reading was set as defined by the kit instructions for all channel reading.	Bio-Rad Laboratories suggeste…

   SciScore for 10.1101/2022.02.27.22271548: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Ethics	IRB: These samples were surplus diagnostic materials that were analyzed anonymously, and no specific approval from Research Ethics Board (REB) was required.
   Sex as a biological variable	not detected.
   Randomization	Batch 2: 120 positive and 120 negative samples were randomly chosen, and re-evaluated to confirm SARS-CoV-2 condition and sample quality.
   Blinding	not detected.
   Power Analysis	not detected.

   Table 2: Resources

   Software and Algorithms
   Sentences	Resources
   The RT-PCR reaction was carried out with CFX 384 Real-Time System (Bio-Rad Laboratories) operated with Bio-Rad CFX Manager 3.1, and the plate reading was set as defined by the kit instructions for all channel reading.	Bio-Rad Laboratories suggested: (Bio-Rad Laboratories, RRID:SCR_008426) Bio-Rad CFX Manager suggested: None CFX suggested: None
   Evaluation of RT-LAMP performance with receiver operating characteristic (ROC) curve analysis: For all the positive and negative clinical NP samples confirmed by BGI RT-PCR kit, RT- LAMP TTR or slope20-40 was plotted in functions of the true positive rate (Sensitivity) and the false positive rate (1-Specificity) for ROC curve analysis using MedCalc software [31].	MedCalc suggested: (MedCalc, RRID:SCR_015044)

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   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).

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   Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:

   Although RT-PCR with purified RNA is the gold standard to confirm SARS-CoV-2 infection, a major limitation is its long turnaround time, especially outside of larger urban centers, compromising test efficacy in terms of timely self-isolation and contact tracing. Rapid and economical PoC tests for SARS-CoV-2, together with masking and social distancing, are necessary to stop community transmission of the disease [51, 52]. With this in mind and recognizing that minimum sample manipulation is essential for PoC tests [15], we next optimized RT-LAMP for raw Canadian NP swab samples without RNA extraction. The best condition used multiplexed Gene E1 and ORF1a primers and supplementation with 0.5M betaine and 0.25% Igepal CA-630. In < 32 mins, the optimized RT-LAMP detected samples with Ct ≤ 25 (viral load ≥ 7.9 × 106 copies/mL) with 100% sensitivity, samples with 25 < Ct ≤ 27.2 (viral load: 2.6 × 106 - 7.9 × 106 copies/mL) with 80% sensitivity and samples with 27.2 < Ct ≤ 29.2 (viral load: 5.0 × 105 - 2.0 × 106 copies/mL) with 31.8%. However, this direct test failed to detect SARS-CoV-2 in samples with Ct ≥ 30 (viral load ≤ 3.2 × 105 copies/mL). For all the samples, the RT-LAMP detected human ACTB in less than 30 minutes. Detection of high titer samples was also demonstrated in labs in Colombia and Ecuador. In the former, an initial survey of 41 negative and 118 positive selected samples defined a cutoff TTR of 25 minutes, then a simulation detection test with 88 positive and 120 ne...

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   Results from TrialIdentifier: No clinical trial numbers were referenced.

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   Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

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   Results from JetFighter: We did not find any issues relating to colormaps.

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   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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   Results from scite Reference Check: We found no unreliable references.

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3. 

   Version published to 10.1101/2022.02.27.22271548 on medRxiv

   Feb 27, 2022