To demonstrate how the algorithm handled closely related genetic species, a sample of a non-targeted pathogen was considered using multiplex PCR (via method in Lin et al., submitted for publication). For Variola major virus, one of the biothreat pathogens on the RPMv.1, the validation runs demonstrated that Variola major virus purified DNA templates of plasmids were always positively identified when detected (Table 3) . Table 4 shows the results when purified Vaccinia genomic DNA was spiked into nasal washes and processed at various concentrations using multiplex PCR. The array has two ProSeqs from hemagglutinin (VMVHA, $500 bp) and cytokine response modifier B (VMVcrmB, $300 bp) genes for Variola major virus detection. The percentage of the ProSeq that hybridizes is sufficient that if hybridization patterns were only considered one might assume that this tile is identifying the presence of its target. This would indicate that reference sequence selected was not the best choice. However, when our algorithm was applied none of the samples is in fact identified as Variola major or minor virus. Vaccinia was always one of the Orthopoxvirus species listed with the highest scores for VMVcrmB ProSeq, but in only seven cases was it uniquely identified as the probable species detected. In only one sample at the lowest concentration and fraction of VMVcrmB hybridizing, did this ProSeq even identify Variola major and minor virus as one among the Orthopoxvirus species that could be the cause of the hybridization. The lower limit of detection for the amplification method used was between this concentration and the one above it for Variola major itself. Table 2 showed that HybSeqs split into multiple SubSeqs were capable of very specific identification.