title: Critical studies on binding-based RT-PCR detection of infectious Noroviruses.
abstract: Attempts were made to discriminate between infectious and non-infectious Noroviruses (NoVs) based on their viral binding properties followed by reverse transcription polymerase chain reaction (RT-PCR). Murine norovirus-1 (MNV-1) was employed as a surrogate to test the principle. Detection of both infectious and inactivated MNV-1 was investigated by the plaque assay, RT-PCR and binding-based RT-PCRs. The cell line RAW 264.7 and the ganglioside GD1a were used as binding receptors respectively in combination with RT-PCR. In the second stage of testing, similar approaches were applied to the two main genogroups of human NoVs (GI and GII). Differentiated Caco-2 cells and pig gastric mucin were tested as the binding receptors. Bovine serum albumin (BSA) was used as a non-specific binding control. In this study, the binding-based RT-PCRs decreased the detection of non-infectious NoVs by 1-3-log(10) while all infectious viral particles were detected. No significant difference was observed between the binding-based RT-PCRs within the concentration range investigated, except the binding level of human NoVs GII to pig gastric mucin was higher than to differentiated Caco-2 cells and BSA. This study indicates an improvement in the evaluation of the infectivity of non-cultivable human NoVs. This is also a comprehensive study on both specific and non-specific binding properties of NoVs.
authors.affiliation: Laboratory of Food Microbiology and Food Preservation, Faculty of Bioscience Engineering, Ghent University, Coupure links 653, B-9000 Ghent, Belgium.
authors.forename: Dan
authors.initials: D
authors.lastname: Li
authors.forename: Leen
authors.initials: L
authors.lastname: Baert
authors.forename: Els
authors.initials: E
authors.lastname: Van Coillie
authors.forename: Mieke
authors.initials: M
authors.lastname: Uyttendaele
mesh_terms: D000818:Animals; D018938:Caco-2 Cells; D017250:Caliciviridae Infections / Q000175:diagnosis / Q000821:virology; D005677:G(M1) Ganglioside / Q000031:analogs & derivatives / Q000378:metabolism; D005752:Gastric Mucins / Q000378:metabolism; D006801:Humans; D051379:Mice; D029322:Norovirus / Q000145:classification / Q000302:isolation & purification* / Q000378:metabolism / Q000472:pathogenicity; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D011991:Receptors, Virus / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013552:Swine; D010948:Viral Plaque Assay; D053585:Virus Attachment; D038301:Virus Inactivation
chemical_list: D005752:Gastric Mucins; D012367:RNA, Viral; D011991:Receptors, Virus; C064020:ganglioside GD1alpha; D012710:Serum Albumin, Bovine; D005677:G(M1) Ganglioside