TT virus (TTV) DNA was quantitated in the serum and nine autopsy tissues (bone marrow, lymph node, muscle, thyroid gland, lung, liver, spleen, pancreas, and kidney) obtained from each of three TTV-infected subjects by real-time polymerase chain reaction (PCR), which can detect all TTV genotypes. TTV DNA was detected in all examined tissues, with the viral load being equal to or up to 300 times higher than that in the corresponding serum (2.1 x 10(5) to 5.3 x 10(7) copies/g vs 1.2-3.9 x 10(5) copies/ml). Generally, the TTV viral load was higher in the bone marrow, lung, spleen, and liver than in the other tissues, although it varied by individual. Restriction fragment length polymorphism (RFLP) analysis of the PCR-amplified TTV DNA of 3.3 kilobases (kb) revealed considerable differences among the TTVs in the serum and tissue specimens from each subject. Further, the 3.3-kb amplicons from the serum and tissue specimens from one subject were molecularly cloned, and 30 clones each from the serum and each tissue specimen were subjected to RFLP and sequence analysis (total, 300 clones): the TTV clones were classified into six genotypes including four novel genotypes. The genotypic variability was remarkable: each specimen had one to five TTV genotypes at different frequencies. TTV DNA in replicative intermediate forms and TTV mRNA were detectable in all tissues tested. These results indicate the broad, uneven distribution of TTV genotypes in tissues and suggest that viral replication takes place in multiple tissues at distinct levels in infected individuals.
Copyright 2001 Academic Press.