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high sensitivity have a low occurrence of false negatives. Sensitivity should not be confused with 1010 the limit of detection. The equation for sensitivity is given by: 1011 where TP = true positives and FN = false negatives. 1012 ( gg ) Signature .—One or more oligonucleotide sequences (i.e. forward primer and/or reverse 1013 primer and/or probe) that experimentally detects (e.g., by PCR or probe-based methods) most or 1014 all of the desired target organism sequences (i.e. inclusivity panel) and does not detect most or all 1015 non-target sequences (exclusivity panel, near neighbors, and environmental panel). This 1016 definition deviates from the traditional definition of a signature as a single oligonucleotide 1017 sequence that is “present” in all targets and “absent” in all non-targets. This traditional definition 1018 is problematic due to the various definitions of “presence” and “absence” depending on the 1019 threshold parameters set for example, 1) Edit distance; 2) BLAST alignment score; 3) Melting 1020 temperature or delta G; 4) Impact of 3’ terminal primer mismatches. 1021 ( hh ) Signature erosion .—Emergence of mutations in the sequences of the signature (primers 1022 and probes)that may lead to assay failure. This may happen via the natural course of evolution, 1023 especially in viruses, due to genetic drift or shift or deliberate acts. 1024 ( ii ) Specificity .—The true negative rate is the fraction of tests that are true negative divided 1025 by the actual number of negative samples (i.e., true negative plus false positive). Assays with 1026 high specificity have a low occurrence of false positives. The equation for specificity is given by: 1027

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where TN = true negatives and FP = false positives.

( jj ) Target Selection .—Entails identifying suitable “unique regions” in the genome of interest

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for assay design. These regions may be longer than the actual PCR amplicon.

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