Environmental surveillance of circulation of enteroviruses-Public health effects

Abstract

Objective: This study presents a new approach for the molecular "serotyping" of enteroviruses concentrated and isolated from urban sewage. Method: Samples were collected from the Nicosia Sewage Treatment Plant in Cyprus. Viruses were adsorbed on cellulose nitrate membrane filters, isolated from the membrane filter using the VIRADEN method and identified by RT-PCR, followed by 5-UTR RFLP analysis and partial sequencing of the VP1 protein coding region. Results: In total, 42 enteroviruses were isolated. Initial sub-grouping based on HpaII restriction profile showed that all isolates belonged to the same genetic subcluster of non-polio enteroviruses. Partial VP1 sequencing revealed that isolates belonged to serotypes CBV4 (33 isolates, 79%), CBV1 (8 isolates, 19%) and Echovirus 7 (1 isolate, 2%). All isolates of the same serotype had highly similar VP1 sequences. The HpaII digests predicted the genetic sub-cluster in all cases. Finally, phylogenetic analysis revealed that the isolates correlate with strains of the same serotype isolated during the last four decades in Europe, Asia, Northern Africa and the Middle East, implying a possible epidemiological relationship. Conclusions: In conclusion, the results confirm that this methodology is highly promising for environmental surveillance of enterovirus circulation and epidemiology. 5-UTR RFLP analysis with HpaII is useful for initial sub-classification, while partial VP1sequencing is efficient for molecular serotyping.