A comparison of two extraction methods for the detection of Enteroviruses in raw sludge

Abstract

The aim of this study was to compare two viral extraction methods for the detection of naturally occurring Enteroviruses in raw sludge. The first method (M1) is based on an ultracentrifugation step. In the second one (M2), viral RNA was extracted directly after viral elution from suspended solids. Genomes of enteroviruses were quantified by a quantitative real time PCR (qRT-PCR) in sludge samples. Somatic coliphages and F-specific RNA phages, considered as viral indicators of enteric viruses in sludge, were enumerated by the double layer agar technique. Results showed that direct assay of RNA extraction yielded higher genomic copies of enteric viruses (with an average of 5.07Log10genomiccopies/100mL). After the ultracentrifugation assay in the second method, genomic copies number decreases (with an average of 4.39Log10genomiccopies/100mL). This can be explained by an eventual concentration of inhibitors existing in sludge samples. Phages enumeration results showed their presence in all sludge samples with an average of (5.69Log10PFU/100mL) for somatic coliphages and (4Log10PFU/100mL) for F-specific RNA phages. This emphasizes the use of somatic coliphages as viral indicators for enteroviruses in environmental samples and especially in raw sludge samples in wastewater treatment plants prior to agricultural use.