Table 5.

Clinical performance of the developed multiplex qRT-PCR assays for Omicron subvariants (BA.1 and BA.2) and recombinant variants (XD and XE) of SARS-CoV-2.

Omicron subvariant BA.1	Sanger sequencing		Performance characteristics
	Positive	Negative	Sensitivity (%)	Specificity (%)	Youden’s Index	PPV (%)	NPV (%)	Agreement (%)	Kappa
The developed assay
Positive	12	0	92.3	100	0.923	100	92.3	99.5	0.955 (P>0.05)
Negative	1	92
Total	13	92
Omicron subvariant BA.2	Sanger sequencing		Performance characteristics
	Positive	Negative	Sensitivity (%)	Specificity (%)	Youden’s Index	PPV (%)	NPV (%)	Agreement	Kappa
The developed assay
Positive	2	0	100	100	1.000	100	100	100	1.000 (P>0.05)
Negative	0	103
Total	2	103
Recombinant variant XD	Sanger sequencing		Performance characteristics
	Positive	Negative	Sensitivity (%)	Specificity (%)	Youden’s Index	PPV (%)	NPV (%)	Agreement	Kappa
The developed assay
Positive	1	0	100	100	1.00	100	100	100	1.000 (P>0.05)
Negative	0	104
Total	1	104
Recombinant variant XE	Sanger sequencing		Performance characteristics
	Positive	Negative	Sensitivity (%)	Specificity (%)	Youden’s Index	PPV (%)	NPV (%)	Agreement	Kappa
The developed assay
Positive	1	0	100	100	1.00	100	100	100	1.000 (P>0.05)
Negative	0	104
Total	1	104

Sensitivity =TP/(TP+FN) × 100%, specificity = TN/(TN+FP) × 100%, PPV (positive predictive value) = TP/(TP+FP) × 100%, NPV (negative predictive value) = TN/(TN+FN) × 100%, Agreement = (TP+TN)/(TP+FP+TN+FN) × 100%, whereas TP, TN, FP and FN represent the number of true positive, true negative, false positive, and false negative, respectively. Youden’s Index = (sensitivity + positivity)-1. Kappa index, assessing the agreement between Sanger sequencing and the developed multiplex qRT-PCR assays, was calculated by the Kappa and the McNemar crosstab statistics implemented in SPSS 22.0.