Direct quantification of waterborne viruses via high-temperature and high-pressure treatment: a simplified nucleic...

High-temperature and high-pressure treatment (HTP) is a physical lysis method that enables direct quantification of viral genomes from environmental samples without nucleic acid extraction. This study assessed the applicability of HTP for rapid virus detection using pepper mild mottle virus (PMMoV) as a model. Optimal conditions for PMMoV quantification were identified as 120 °C for 15 s, yielding results comparable to conventional RNA extraction methods. RNA integrity analysis revealed that temperatures above 140 °C may cause genome degradation, supporting the selection of 120 °C as the optimal setting. The method demonstrated high reproducibility across qPCR and dPCR assays and enabled stable quantification with minimal interference from wastewater matrix components. In addition, additional virus species were evaluated to assess the performance of HTP beyond PMMoV under the tested conditions. These findings highlight the potential of HTP as a simplified workflow without nucleic acid extraction for environmental virus monitoring. Future research should focus on expanding its use to diverse virus types and integrating the method into automated platforms for real-time monitoring.