Table 2.
Inclusivity evaluation of primers and probe sets using SCREENED with the first SARS-CoV-2 genome batch 07042020 for all the RT-qPCR tests.
| Assay | Target | Genomes with Mismatches in the First Five Nucleotides of the Primer’s 3′ End | Genomes with >10% Mismatches in the Annealing Sites of Primers and Probes | False Negative Results *** | Inclusivity | ||
|---|---|---|---|---|---|---|---|
| Number * | Modifications ** | Number * | Modifications ** | ||||
| 1 | ORF1b | 1 | Fw GTGGGTTTTACATTTAA | 0 | - | 1 | 99.96% |
| 1 | N | 1 | Rv CAGACATTTTGCTCTCAAACTG | 358 | Fw AACGAACTTCTCCTGCTAGAAT | 359 | 86.03% |
| 2 | RdRp-P1 | 0 | - | 1 | Pb CCAGGTGGGACCTCATCAGGAGATGC | 1 | 99.96% |
| 2 | RdRp-P2 | 0 | - | 0 | - | 0 | 100% |
| 2 | E | 0 | - | 0 | - | 0 | 100% |
| 2 | N | 4 | Fw CACATTGGCACCCGTAATC | 0 | - | 5 | 99.81% |
| 1 | Rv GAGGAACGAGAAGAGACTTG | ||||||
| 3 | RdRp-IP2 | 3 | Rv CTCCCTTTGTTGTGTTAT | 0 | - | 3 | 99.88% |
| 3 | RdRp-IP4 | 0 | - | 0 | - | 0 | 100% |
| 3 | E | 0 | - | 0 | - | 0 | 100% |
| 4 | N-1 | 7 | Rv TCTGGTTACTGCCAGTTGAACCTG | 0 | - | 7 | 99.73% |
| 4 | N-2 | 1 | Fw TTACAAACATTGGCCTCAAA | 0 | - | 1 | 99.96% |
| 4 | N-3 | 0 | - | 0 | - | 0 | 100% |
| 5 | N | 3 | Fw AAATTTTGGGGACCATGAAC | 0 | - | 8 | 99.69% |
| 4 | Fw AAATTTTGGGGACCAGGAAT | ||||||
| 1 | Rv TGGCACCTGTGTAGGTAAAC | ||||||
| 6 | ORF1b/nsp14 | 0 | - | 0 | - | 0 | 100% |
| 6 | N | 2 | Rv CGAAGGTGTGACTTCAATG | 0 | - | 2 | 99.92% |
| 7 | N | 7 | Fw CGTTTGGTGGACCCTCAGGT | 0 | - | 7 | 99.73% |
| 8 | RdRp/Hel | 0 | - | 0 | - | 0 | 100% |
| 8 | S | 0 | - | 0 | - | 0 | 100% |
| 8 | N | 1 | Fw GCGTTCTTCGGAATGTCT | 0 | - | 1 | 99.96% |
| 9 | ORF1a | 0 | - | 0 | - | 0 | 100% |
| 10 | RdRp | 8 | Rv TGCATTAACATTGGCCGTAA | 0 | - | 8 | 99.69% |
| 10 | S | 0 | - | 0 | - | 0 | 100% |
| 10 | E | 0 | - | 0 | - | 0 | 100% |
| 10 | N | 1 | Rv GTTGCGACTACGTGATGAGT | 0 | - | 1 | 99.96% |
| 11 | N-1 | 3 | Fw GCCTCTTCTCGTTCCTCACCAC | 0 | - | 8 | 99.69% |
| 5 | FW GCCTCTTCTCGTTCCTCATTAC | ||||||
| 11 | N-2 | 3 | Fw AGCCTCTTCTCGTTCCTCACCAC | 0 | - | 8 | 99.69% |
| 5 | Fw AGCCTCTTCTCGTTCCTCATTAC | ||||||
| 11 | ORF1a-3 | 0 | - | 0 | - | 0 | 100% |
| 11 | ORF1a-4 | 0 | - | 0 | - | 0 | 100% |
| 11 | S-5 | 9 | Fw CAGGTATATGTGCTAGTTATCACAC | 0 | - | 11 | 99.57% |
| 1 | Fw CAGGTATATGCGCTAGTTATCATAC | ||||||
| 1 | Fw CAGGTATATGCGCTAGTTATCGGAC | ||||||
| 11 | S-6 | 9 | Fw GCAGGTATATGTGCTAGTTATCAC | 0 | - | 11 | 99.57% |
| 1 | Fw GCAGGTATATGCGCTAGTTATCGT | ||||||
| 1 | Fw GCAGGTATATGCGCTAGTTATCGG | ||||||
| 12 | E | 0 | - | 0 | - | 0 | 100% |
-: No modifications retrieved by SCREENED. *: Number of genomes having the type of mismatches investigated from a total of 2569 unique representative genomes. **: Detected mismatches are indicated in bold in the 5′-3′ sequence of forward primers (Fw), reverse primers (Rv), and probes (Pb). The mismatch in bold, and underlined is an additional SNP not specific to the parameters investigated by SCREENED. ***: The total number of genomes that did not produce a positive signal for the evaluated primers and probe sets considered as false negative results. No false negative results were obtained due to minimum alignment length lower than 90%.