Detection of hepatitis delta virus RNA by a nonradioactive in situ hybridization procedure.
A digoxigenin-tailed, synthetic oligodeoxynucleotide was used to detect genomic hepatitis delta virus (HDV) RNA in formalin-fixed, paraffin-embedded liver sections by a nonisotopic in situ hybridization (NISH) procedure. Twenty-three liver samples from chronically HDV-infected patients were studied. Eight liver specimens from humans and chimpanzees without markers of active HDV infection served as negative controls. In three samples, the NISH findings were compared with characteristic nuclear features and with the distribution of the HDV encoded antigen, HDAg, as detected by direct immunofluorescence. All samples from HDV-infected patients were positive for HDV RNA by NISH. The viral genome was exclusively observed within the hepatocytic nuclei. No enzymatic reaction was detected after hybridization with the negative controls. "Sanded" nuclei, a cytopathologic change associated with HDV infection, were HDV RNA-positive, but only a small percentage of infected cells showed that feature. Hepatocytes containing the HDV RNA were sometimes binucleated or exhibited giant nuclei. When HDAg and HDV RNA were sequentially detected within the same sections, the localization of the viral antigen almost completely overlapped with the expression of the HDV transcripts, and vice versa. In conclusion, detection of intrahepatic HDV RNA by NISH is a rapid, sensitive, and specific technique that is easily applicable to routine histopathology and allows correlation of HDV with the morphology of hepatocyte nuclei.
Pacchioni
D
D
Department of Biomedical Sciences and Human Oncology, University of Turin School of Medicine, Italy.
Negro
F
F
Chiaberge
E
E
Rizzetto
M
M
Bonino
F
F
Bussolati
G
G
D000956:Antigens, Viral / Q000032:analysis; D001483:Base Sequence; D005455:Fluorescent Antibody Technique; D003699:Hepatitis D / Q000175:diagnosis* / Q000473:pathology; D003698:Hepatitis Delta Virus / Q000235:genetics* / Q000276:immunology; D006801:Humans; D007124:Immunoenzyme Techniques; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D012367:RNA, Viral / Q000032:analysis*
D000956:Antigens, Viral; D012367:RNA, Viral