Each candidate primer pair was first tested for its specificity and sensitivity of PCR amplification. This was assessed in a real-time PCR cross-testing against human PBMC DNA (50 ng/PCR reaction) from a healthy donor, DNA from human cell cultures infected by various target viruses, or human DNA spiked with a known amount of genome copies of various viruses. It was important to ensure that the selected primers targeting a specific virus would not non-specifically amplify any DNA in a sample. The melting temperature (Tm) peak of the product amplified from the target virus should be clearly differentiable from the Tm peak of primer dimers or any non-specific products produced in PCR. DNA or RNA panels were created from the cloned synthetic templates (listed in Table S1 ) by spiking of 2-10 4 genome copies of each virus into 50 ng of human background DNA. The sensitivity of each candidate primer set for each target virus was assessed using these panels.