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Journal of Clinical Microbiology, August 1999, p. 2703-2705, Vol. 37, No. 8
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
TT Virus Infection Is Widespread in the General
Populations from Different Geographic Regions
Kenji
Abe,1,*
Tomoko
Inami,1
Kazue
Asano,2
Chiaki
Miyoshi,3
Naohiko
Masaki,4
Shigeki
Hayashi,4
Ko-ichi
Ishikawa,5
Yutaka
Takebe,5
Khin Maung
Win,6
Abdel Rahman
El-Zayadi,7
Kwang-Hyub
Han,8 and
David Y.
Zhang9
Department of
Pathology1 and AIDS Research
Center,5 National Institute of Infectious
Diseases, and Bureau of International
Cooperation3 and Division of
Gastroenterology,4 International Medical Center
of Japan, Tokyo, and Department of Pediatrics, Seirei
Hamamatsu General Hospital, Shizuoka,2 Japan;
Department of Hepatology, Yangon General Hospital, Yangon,
Myanmar6; Department of Internal
Medicine, Faculty of Medicine, Ain Shams University, Cairo,
Egypt7; Department of Internal Medicine,
Yonsei University College of Medicine, Seoul,
Korea8; and Department of Pathology, The
Mount Sinai Medical Center of the City University of New York, New
York, New York9
Received 22 February 1999/Returned for modification 28 April
1999/Accepted 18 May 1999
 |
ABSTRACT |
By PCR screening, we found an extremely high prevalence of TT virus
(TTV) in the general populations from different geographic regions.
This suggests that TTV may be a common DNA virus with no clear disease
association in humans. TTV genotyping by phylogenetic analysis was also performed.
| |
TEXT |
In 1997, the genome of a novel DNA
virus, termed the TT virus (TTV), isolated from the sera of patients
with posttransfusion non-A-G hepatitis, was sequenced by
representational difference analysis (6, 7). Very recently,
the complete nucleotide sequence of TTV was reported by two different
groups (2, 3). TTV is an unenveloped circular
single-stranded DNA virus and comprises 3,852 nucleotides, with an
isopycnic density of 1.31 to 1.34 g/ml in CsCl (2, 3). The
TTV genome has three possible open reading frames, capable of encoding
770, 202, and 105 amino acids, respectively (2). Due to the
genome structure and its banding in buoyant density gradient
centrifugation, TTV is related among the known animal virus families to
the Circoviridae family (2, 3, 7, 9). Despite TTV
being a DNA virus, its sequence has a wide range of sequence
divergence, allowing classification into several genotypes (7,
11). TTV sequences were detected in sera and liver tissues from
liver disease patients, suggesting that TTV could be responsible for
some acute and chronic liver disease of unknown etiology (1,
7). On the other hand, it has been reported elsewhere that TTV
infection does not induce significant liver damage (5).
However, the epidemiology, clinical significance, and transmission
patterns of TTV remain unclear. To clarify the characterization of
seroepidemiology of TTV, we carried out PCR screening for TTV in
individuals, including healthy populations from different geographic regions.
We collected serum samples from individuals in Japan (233 individuals
without liver disease), Myanmar (51 healthy individuals and 92 liver
disease patients), Nepal (177 blood donors), Egypt (95 blood donors),
Bolivia (95 blood donors), Vietnam (62 high-risk individuals consisting
of medical staff), Korea (73 hemodialysis patients), Cambodia (8 human
immunodeficiency virus [HIV]-infected patients), Ghana (95 HIV-infected patients), and the United States (68 HIV-infected
patients). Informed consent was obtained from participants in this
study. The serum samples were stored at 
20°C or below until assayed.
DNA was extracted from 100 µl of serum samples with a nucleic acid
extraction kit (SepaGene RV-R; Sanko Junyaku Co., Ltd., Tokyo, Japan).
The resulting pellet was resuspended in RNase- and DNase-free water and
then subjected to PCR as described by Takahashi et al. (10).
In brief, the thermocycler was programmed first to preheat at 95°C
for 10 min to activate AmpliTaq Gold DNA polymerase (Perkin-Elmer,
Norwalk, Conn.), and then samples were subjected to 55 cycles
consisting of 94°C for 20 s, 60°C for 20 s, and 72°C
for 30 s with a Perkin-Elmer 9600 or 9700 thermal cycler. The
sequences of the TTV-specific primers were
5'-GCTACGTCACTAACCACGTG-3' (T801, sense primer, nucleotides
6 to 25) and 5'-CTBCGGTGTGTAAACTCACC-3' (T935, antisense
primer, nucleotides 185 to 204; B = G, C, or T) as designed by
Takahashi et al. (10) in the 5'-end region of the TA278
isolate. The PCR products were detected by electrophoresis on 2%
agarose gels, stained with ethidium bromide, and photographed under UV light.
To determine the genotype, TTV DNA was amplified by nested PCR with
primers NG059 and RD038 for the outer primer pairs (377 bases) and
NG061 and NG063 for the inner primer pairs (271 bases) as designed by
Okamoto et al. (7) in the ORF1 region of the TA278 isolate.
Amplified PCR products were subjected to direct sequencing, and
then phylogenetic analysis was performed as reported previously
(4). Twenty previously reported TTV sequences were obtained
from the GenBank database and used for comparison with the sequence of
the isolate in this study.
Statistical analyses were performed by the chi-square test or Fisher's
exact test. A difference with a P value of <0.05 was considered significant.
As shown in Table 1, a very high
prevalence of TTV infection was found in tested individuals, including
healthy populations, from 10 different countries. In the Japanese
study, TTV DNA was detected significantly more often in the groups of
people over 10 years of age (P < 0.05) (Table
2). On the other hand, the prevalence of
TTV in individuals from other countries had already reached nearly 80%
or more in the age groups over 10 years old (differences between the
age groups were not statistically significant). Furthermore, the TTV
genome could be classified into at least six different genotypes by
phylogenetic analysis, and the major genotypes are type 1 and type 2 (Fig. 1). However, there is no correlation between major genotypes and geographic origin.
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FIG. 1.
Phylogram generated by neighbor-joining analysis of
genetic distances in the ORF1 region of TTV isolates. Isolates
determined in this study are presented in boldface. The
database-derived isolates and their accession numbers were as follows:
G97801 (AB011486), TA278 (AB008394), G103301 (AB011487), N22
(AB017767), G10491 (AB011489), TY96117 (AB011494), G88801 (AB011491),
G102001 (AB011488), G105001 (AB011490), TS003 (AB017770), NA004
(AB017771), PT3 (AB017768), AF079541 (AF079541), AF079542 (AF079542),
AF079543 (AF079543), COL139 (AB01635), JaM21 (AB017887), JaNBNC10
(AB018961), JaM28 (AB017888), and JaM18 (AB017886). UK, United Kingdom;
USA, United States of America.
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|
Although the TTV PCR system has been established already by Nishizawa
et al. (6) and Okamoto et al. (7), there have been problems associated with low sensitivity and unstable reaction. To
resolve this problem, Takahashi et al. recently reported that TTV PCR
with a new primer combination is more sensitive and stable than the
previous system (10). Using the new PCR system, they reported that TTV DNA was identified in 92% of 100 healthy individuals in Japan (10). In the present study, we found that TTV
viremia is widespread in the general populations in many countries.
Such an extremely high prevalence of TTV infection in the general
population suggests that TTV may be transmissible not only in the blood
but also by a nonparenteral route. Indeed, Okamoto et al. reported that
TTV was excreted into the feces, thereby suggesting that TTV may be
transmitted not only parenterally but also nonparenterally by a
fecal-oral route (8). Molecular analysis of TTV genomes showed that genotypes 1 and 2 of TTV were widespread worldwide.
In conclusion, TTV viremia is widespread, with a very high incidence in
general populations worldwide. This suggests that TTV is a common virus
and may be a nonpathogenic DNA virus in humans, although the pathogenic
role of TTV still remains to be investigated.
Nucleotide sequence accession numbers.
The nucleotide sequence
data reported in this paper have been submitted to the DDBJ, EMBL, and
GenBank databases under accession no. AB023311 through AB023352.
| |
ACKNOWLEDGMENTS |
We thank Takeshi Kurata for his continuous encouragement during
this study and Nami Konomi, Hideo Naito, Mari Yamaguchi, and Mitsugu
Usui for their kind cooperation.
This study was supported in part by grants-in-aid for science research
from the Ministry of Education, Science and Culture of Japan and the
Ministry of Health and Welfare of Japan.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Department of
Pathology, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku-ku, Tokyo 162-8640, Japan. Phone: (81) 3-5285-1111, ext. 2624. Fax: (81) 3-5285-1189 or (81) 3-5285-1150. E-mail:
kenjiabe{at}nih.go.jp.
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Journal of Clinical Microbiology, August 1999, p. 2703-2705, Vol. 37, No. 8
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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