Molecular detection of rotavirus
Details of nucleic acid extractions, reverse transcription, and rotavirus molecular detection were described previously (19). Briefly, total nucleic acid was extracted from 30% rectal swab suspension or 10% stool suspension, prepared using NucliSENS® lysis buffer (BioMérieux, Laval, QC, Canada) by NucliSENS® easyMAG® (BioMérieux, Laval, QC, Canada) according to manufacturer’s instructions. All resultant nucleic acid extracts were immediately stored at -70°C until testing. cDNA was synthesized from viral RNA using SuperScriptTM II reverse transcriptase (Invitrogen, MA, USA). Group A rotaviruses in stool and swabs were identified by an in-house reverse transcription real-time PCR (RT-qPCR) assay, the Gastroenteritis Viral Panel (GVP), and a multi-target bead-based molecular assay, the Luminex Gastrointestinal Pathogen Panel (GPP) (Luminex Molecular Diagnostics, ON, Canada), as described previously (28).