title: Partial sequencing of the VP2 capsid gene for direct enterovirus genotyping in clinical specimens.
abstract: Typing of human enterovirus (EV) remains a major goal for diagnostic and epidemiological purposes. Whereas sequencing of the VP1 coding region is the reference standard for EV typing, a method relying on sequencing of the VP2 coding region has been proposed as an alternative; however, this has been validated only on cell culture supernatants. To avoid the selection of cultivable strains and to quicken the identification step, a new semi-nested PCR method targeting the VP2 region was developed by use of the CODEHOP strategy. After validation of the method on reference and clinical strains, a total of 352 clinical specimens found to be positive for EV RNA (138 with the GeneXpert EV kit and 214 with the Enterovirus R-gene kit) during a 3-year period (2010-2012) were analysed prospectively for VP2 genotyping. Overall, 204 (58%) specimens were typeable. A higher proportion of throat swab/stool specimens than of cerebrospinal fluid (CSF) specimens was found to be typeable (94 of 142 (66.2%) vs. 83 of 169 (49.1%), respectively, p <0.01 by the chi-square test). Moreover, the median Ct value obtained was lower for typeable specimens than for untypeable specimens (32.20 vs. 33.01, p <0.05, and 25.96 vs. 31.74, p <0.001, for the GeneXpert and R-gene tests, respectively, by the Mann-Whitney-Wilcoxon test). These results suggest that, in cases of EV meningitis, a peripheral specimen (i.e. throat swab or stool) that is susceptible to exhibiting a higher viral load should be used in preference to CSF for identifying the causative EV genotype by use of the VP2 typing method without cell culture isolation.
authors.affiliation: EA-3064, Groupe Immunité des Muqueuses et Agents Pathogènes (GIMAP), Faculty of Medicine of Saint-Etienne, University of Lyon, Saint-Etienne, France; Laboratory of Transmissible Diseases and Biologically Active Substances, Faculty of Pharmacy, Monastir, Tunisia.
authors.forename: W
authors.initials: W
authors.lastname: Ibrahim
authors.forename: N
authors.initials: N
authors.lastname: Boukhadra
authors.forename: D
authors.initials: D
authors.lastname: Nasri-Zoghlami
authors.forename: P
authors.initials: P
authors.lastname: Berthelot
authors.forename: S
authors.initials: S
authors.lastname: Omar
authors.forename: T
authors.initials: T
authors.lastname: Bourlet
authors.forename: B
authors.initials: B
authors.lastname: Pozzetto
authors.forename: S
authors.initials: S
authors.lastname: Pillet
mesh_terms: D036022:Capsid Proteins / Q000235:genetics*; D002555:Cerebrospinal Fluid / Q000821:virology; D004770:Enterovirus / Q000145:classification* / Q000235:genetics*; D004769:Enterovirus Infections / Q000821:virology*; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D008969:Molecular Sequence Data; D010614:Pharynx / Q000821:virology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011446:Prospective Studies; D012367:RNA, Viral / Q000235:genetics; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D036022:Capsid Proteins; D012367:RNA, Viral
keywords: Clinical specimen; VP2 capsid protein; human enterovirus; molecular epidemiology; typing method; viral load