Metavirome sequencing to evaluate norovirus diversity in sewage and related bioaccumulated oysters

Metagenomic sequencing is a promising method to determine the virus diversity in environmental samples such as sewage or shellfish. However, to identify the short RNA genomes of human enteric viruses among the large diversity of nucleic acids present in such complex matrices, method optimization is still needed. This work presents methodological developments focused on norovirus, a small ssRNA non-enveloped virus known as the major cause of human gastroenteritis worldwide and frequently present in human excreta and sewage. Different elution protocols were applied and Illumina MiSeq technology were use

Characterization of Norovirus and Other Human Enteric Viruses in Sewage and Stool Samples Through Next-Generation Sequencing

This study aimed to optimize a method to identify human enteric viruses in sewage and stool samples using random primed next-generation sequencing. We tested three methods, two employed virus enrichment based on the binding properties of the viral capsid using pig-mucin capture or by selecting viral RNA prior to library preparation through a capture using the SureSelect target enrichment. The third method was based on a non-specific biophysical precipitation with polyethylene glycol. Full genomes of a number of common human enteric viruses including norovirus, rotavirus, husavirus, enterovirus and astrovirus were obtained. In stool samples full norovirus genome were detected as well as partial enterovirus genome. A variety of norovirus sequences was detected in sewage samples, with genogroup II being more prevalent. Interestingly, the pig-mucin capture enhanced not only the recovery of norovirus and rotavirus but also recovery of astrovirus, sapovirus and husavirus. Documenting sewage virome using these methods provides information for molecular epidemiology and may be useful in developing strategies to prevent further spread of viruses