title: Detection of group B and C rotaviruses by polymerase chain reaction.
abstract: We adapted the polymerase chain reaction (PCR) to detect the noncultivatable group B and C rotaviruses and introduced a simple and convenient technique to purify viral RNA from stool specimens. Double-stranded RNA present in stool extracts was purified by adsorption to hydroxyapatite and was used as the template for reverse transcription and polymerase amplification. Primer pairs specific for group B (gene 8) and group C (gene 6) rotaviruses were selected to amplify group-characteristic sizes of cDNA copies readily identifiable in ethidium bromide-stained agarose gels. These primer pairs were used separately in individual PCR assays or were pooled with a primer pair specific for group A rotavirus (gene 9) in a combined PCR assay for the simultaneous detection of all three rotavirus groups. The method was very sensitive and was used to identify both human and porcine strains of group B and C rotaviruses in stool specimens. A second PCR amplification with internal group-specific primers served to increase further the sensitivity of the test and to confirm the diagnostic results obtained in the first amplification.
authors.affiliation: Division of Viral and Rickettsial Diseases, Centers for Disease Control, Atlanta, Georgia 30333.
authors.forename: V
authors.initials: V
authors.lastname: Gouvea
authors.forename: J R
authors.initials: JR
authors.lastname: Allen
authors.forename: R I
authors.initials: RI
authors.lastname: Glass
authors.forename: Z Y
authors.initials: ZY
authors.lastname: Fang
authors.forename: M
authors.initials: M
authors.lastname: Bremont
authors.forename: J
authors.initials: J
authors.lastname: Cohen
authors.forename: M A
authors.initials: MA
authors.lastname: McCrae
authors.forename: L J
authors.initials: LJ
authors.lastname: Saif
authors.forename: P
authors.initials: P
authors.lastname: Sinarachatanant
authors.forename: E O
authors.initials: EO
authors.lastname: Caul
mesh_terms: D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics; D005814:Genes, Viral; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D012401:Rotavirus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D012400:Rotavirus Infections / Q000382:microbiology; D012680:Sensitivity and Specificity
chemical_list: D004279:DNA, Viral; D012367:RNA, Viral