Direct PCR for Rapid and Safe Pathogen Detection: Laboratory Evaluation Supporting Field Use in Infectious Disease Outbreak

Rapid, safe, and field-deployable molecular diagnostics are crucial for the effective management of infectious disease outbreaks, particularly those involving highly infectious pathogens, which can produce clinical symptoms similar to less infectious pathogens, thus raising potential biosafety concerns. In this study, we evaluated DNA/RNA Defend Pro (DRDP) buffer, a novel viral-inactivating transport medium designed to stabilize nucleic acids and allow direct PCR without nucleic acid extraction. To ensure critical qPCR parameters were not compromised by using DRDP, we conducted serial dilution tests using herpes simplex viruses 1 and 2 (HSV-1, HSV-2) and varicella-zoster virus (VZV), comparing DRDP to standard universal transport medium (UTM). Detection sensitivity, determined by cycle quantification (Cq) values, favored DRDP, as UTM samples required a 2–3-fold dilution to mitigate PCR inhibition. DRDP maintained reliable PCR compatibility at reaction volumes containing up to 25% buffer. At higher DRDP concentrations (30–35%), PCR inhibition occurred due to EDTA content but was fully reversible by adding supplemental magnesium. Furthermore, DRDP samples did not require an initial 95 °C thermal lysis step, thus simplifying the procedure without reducing PCR sensitivity or efficiency.