TT virus is shown in the liver by in situ hybridization with a PCR-generated probe from the serum TTV-DNA.
OBJECTIVE
It has been a conflicting issue whether TT virus (TTV), a newly isolated DNA virus from a patient with liver injury of unknown cause, is a causative agent of acute and/or chronic hepatitis. TT Virus DNA titers were shown to be 10-100-fold greater in liver tissue than in serum, whereas the majority of TTV-positive cases had no biochemical or histological evidence of significant liver damage. We therefore attempted in situ hybridization to investigate whether TTV is hepatotropic.


METHODS
Because of the marked divergence in TTV genome types, a template for TTV-DNA (coding region for N22 clone) was amplified and labeled with digoxigenin-dUTP by using hemi-nested PCR from the serum, then DNA probes were applied to the liver sections of the same case. After hybridization, the probes were visualized immunohistochemically. Besides TTV-DNA-negative cases, competitive inhibition experiments with unlabeled probes were performed to confirm the specificity.


RESULTS
There were no positive signals in the negative controls, and the intensity of positive signals was markedly diminished in the competitive inhibition experiments. No cross-hybridization with different genotype probes also confirms the specificity. Under the optimal conditions, the positive signals were located in the cytoplasm of the hepatocytes in eight of nine TTV-DNA-positive cases. The signals were not seen in non-parenchymal cells of the liver.


CONCLUSIONS
TT Virus is proved to be hepatotropic by in situ hybridization.
Ohbayashi
H
H
Second Department of Internal Medicine, Faculty of Medicine, Tokyo Medical and Dental University, Japan.
Tanaka
Y
Y
Ohoka
S
S
Chinzei
R
R
Kakinuma
S
S
Goto
M
M
Watanabe
M
M
Marumo
F
F
Sato
C
C
D004279:DNA, Viral / Q000097:blood; D004587:Electrophoresis, Agar Gel; D006801:Humans; D017403:In Situ Hybridization; D008099:Liver / Q000821:virology*; D015335:Molecular Probes; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D022783:Torque teno virus / Q000235:genetics / Q000302:isolation & purification*
D004279:DNA, Viral; D015335:Molecular Probes