title: Reovirus 3 not detected by reverse transcriptase-mediated polymerase chain reaction analysis of preserved tissue from infants with cholestatic liver disease.
abstract: Reovirus type 3 has been implicated in the origin and pathogenesis of extrahepatic biliary atresia and idiopathic neonatal hepatitis, but routine detection of this virus in hepatobiliary tissues from affected infants by culture and histological techniques has been unsuccessful. In this study, oligonucleotide primers specific to the M3 genome segment of reovirus 3 (Dearing) were used in a reverse transcriptase-mediated polymerase chain reaction technique to develop a sensitive and specific assay for the detection of reovirus 3 RNA in formalin-fixed, paraffin-embedded patient samples. Optimal reaction conditions were determined by testing infected murine tissues and preserved human liver tissue supplemented with reovirus 3. Archival specimens from 50 infants, including 14 with extrahepatic biliary atresia, 20 with idiopathic neonatal hepatitis, and 16 age-matched controls, were evaluated. Successful amplification of human albumin complementary DNA from the preserved tissues confirmed the presence of intact RNA in every patient specimen tested. Analysis of the amplification reactions by agarose gel electrophoresis and Southern blot hybridization detected the presence of reoviral RNA only once in a single patient sample. These results do not support a strong role for reovirus 3 in the development of neonatal cholestatic liver disease. The recent association of other RNA viruses of the Reoviridae family with murine liver disease and human extrahepatic biliary atresia indicates that continued investigation into a viral cause for idiopathic neonatal hepatobiliary disease is warranted.
authors.affiliation: Department of Pediatrics, University of Oklahoma Health Sciences Center, Oklahoma City.
authors.forename: M I
authors.initials: MI
authors.lastname: Steele
authors.forename: C M
authors.initials: CM
authors.lastname: Marshall
authors.forename: R E
authors.initials: RE
authors.lastname: Lloyd
authors.forename: V E
authors.initials: VE
authors.lastname: Randolph
mesh_terms: D000818:Animals; D001483:Base Sequence; D017734:Bile Ducts, Extrahepatic / Q000821:virology; D001656:Biliary Atresia / Q000821:virology; D002779:Cholestasis / Q000821:virology*; D006525:Hepatitis, Viral, Human / Q000821:virology; D006801:Humans; D007231:Infant, Newborn; D012089:Mammalian orthoreovirus 3 / Q000235:genetics / Q000302:isolation & purification*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000235:genetics; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000032:analysis; D012194:RNA-Directed DNA Polymerase; D012088:Reoviridae Infections / Q000821:virology; D012189:Retrospective Studies; D014021:Tissue Preservation
chemical_list: D015345:Oligonucleotide Probes; D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase