title: Typing of human enterovirus by partial sequencing of VP2.
abstract: The sequencing of the VP1 hypervariable region of the human enterovirus (HEV) genome has become the reference test for typing field isolates. This study describes a new strategy for typing HEV at the serotype level that uses a reverse transcription-PCR assay targeting the central part of the VP2 capsid protein. Two pairs of primers were used to amplify a fragment of 584 bp (with reference to the PV-1 sequence) or a part of it (368 bp) for typing. For a few strains not amplified by the first PCR, seminested primers enhanced the sensitivity (which was found to be approximately 10(-1) and 10(-4) 50% tissue culture infective dose per reaction tube for the first and seminested assay, respectively). The typing method was then applied to 116 clinical and environmental strains of HEV. Sixty-one typeable isolates were correctly identified at the serotype level by comparison to seroneutralization. Forty-eight of 55 "untypeable" strains (87.3%) exhibited the same serotype using VP1 and VP2 sequencing methods. For six strains (four identified as EV-71, one as E-9, and one as E-30 by the VP2 method), no amplification was obtained by the VP1 method. The last strain, typed as CV-B4 by VP1 and CV-B3 by VP2 and monovalent antiserum, could exhibit recombination within the capsid region. Although the VP2 method was tested on only 36 of the 68 HEV serotypes, it appears to be a promising strategy for typing HEV strains isolated on a routine basis. The good sensitivity of the seminested technique could avoid cell culture and allow HEV typing directly from PCR products.
authors.affiliation: Laboratory of Bacteriology-Virology, GIMAP EA3064, Faculty of Medicine Jacques Lisfranc, 15, rue Ambroise Paré, 42023 Saint-Etienne, France.
authors.forename: Dorsaf
authors.initials: D
authors.lastname: Nasri
authors.forename: Lamjed
authors.initials: L
authors.lastname: Bouslama
authors.forename: Shabir
authors.initials: S
authors.lastname: Omar
authors.forename: Henia
authors.initials: H
authors.lastname: Saoudin
authors.forename: Thomas
authors.initials: T
authors.lastname: Bourlet
authors.forename: Mahjoub
authors.initials: M
authors.lastname: Aouni
authors.forename: Bruno
authors.initials: B
authors.lastname: Pozzetto
authors.forename: Sylvie
authors.initials: S
authors.lastname: Pillet
mesh_terms: D036022:Capsid Proteins / Q000235:genetics*; D016000:Cluster Analysis; D004770:Enterovirus / Q000145:classification* / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D010802:Phylogeny; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D012703:Serotyping
chemical_list: D036022:Capsid Proteins