Recovery comparison of two virus concentration methods from wastewater using cell culture and real-time PCR

Abstract

Enteric viruses are shed in the feces and may be present in environmental waters. Their detection in wastewater, even at low concentration, is a major challenge. In this study, recoveries of Echovirus 7 (EV7), virions and RNA in wastewater, using virus concentration methods were determined to evaluate the detection of infectious viruses and the possibility of recovering viral genomes. Two virus concentration methods, PEG precipitation method and two-phase separation method, were applied to recovery experiments of EV7-virions from wastewater, in parallel with recovery experiments of EV7 RNA. The titration of EV7 virions was carried out by cell culture using human rhabdomyosarcoma tumor tissue and the EV7 RNA quantification was performed by real-time PCR. The mean recovery yields of EV7 virions using the PEG precipitation method and the two-phase separation method were 78.5 ± 10.99 and 83.1 ± 0.28 %, respectively. Besides, EV7 RNA recoveries obtained using the PEG precipitation method were four times higher than those using the two-phase separation method. According to our results, the two methods enable to concentrate both infectious viruses and viral genomes. Moreover, considering the protocol time and cost together with the ratio of the EV7 virion recovery to the EV7 RNA recovery, the two-phase separation method (83.1/2.71 %, or 30.6) seems to be more appropriate for selective concentration of viral virions than the PEG precipitation method (78.5/10.33 %, or 7.6).