TT Virus Infection in Patients with Hepatitis C: Frequency, Persistence, and Sequence Heterogeneity
William L. Irving
1
3
Jonathan K. Ball
1
3
Selina Berridge
1
3
Rebecca Curran
1
3
Anna M. Grabowska
1
3
Claire L. Jameson
1
3
Keith R. Neal
1
2
Steven D. Ryder
0
1
Brian J. Thomson
1
3
for the Trent HCV Study Group
1
4
0
Medicine, University Hospital, Queen's Medical Centre
,
Nottingham
,
United Kingdom
1
Received 9 September 1998; revised 18 February 1999; electronically pub- lished 14 May 1999. Presented in part: American Association for the Study of Liver Diseases
,
Chicago
,
November 1998. All serum samples from patients described were obtained with informed consent and with the approval of the relevant local hospital ethics committee. Grant support: National Health Service Executive Trent and Sir Jules Thorn Charitable Trust
2
Public Health Medicine and Epidemiology
3
Microbiology and Infectious Diseases
4
Study group members are listed after the text. ogy, Queen's Medical Centre
,
Nottingham NG7 2UH
,
UK
TT virus (TTV) was recently identified in the serum of a patient with hepatitis. The role of TTV in liver disease has not been established. Three polymerase chain reaction (PCR) protocols were used to detect TTV DNA in sera of persons infected with hepatitis C virus (HCV) and in blood donors. Sera from 11.5% of HCV-infected patients and 7.7% of blood donors were positive by protocols 1 or 2. In contrast, 48.7% and 57.7% of sera, respectively, were positive when tested by protocol 3. There was no difference in the severity of hepatitis in persons coinfected with TTV and HCV when compared with those infected with HCV alone, regardless of which TTV PCR protocol was used. TTV DNA persisted in serum samples taken up to 6 years apart in individual patients. Sequence analysis indicated that most viral sequences were distinct between patients, and there was evidence of genetic heterogeneity and viral evolution within individuals.
-
TT virus (TTV), named with the initials of the person in
whose serum it was first detected, was recently identified by
representational difference analysis in the posttransfusion
serum of a patient with transfusion-acquired hepatitis and
subsequently demonstrated by polymerase chain reaction (PCR)
in sera from 3 of 5 patients with posttransfusion hepatitis [1].
In 1 of these patients, TTV DNA was detectable in serum
samples taken up to 21 weeks after transfusion, suggesting that
in at least some persons TTV infection can become chronic.
Genoprevalence rates of TTV DNA in the sera of normal blood
donors vary geographically, for example, from as low as 1% in
the United Kingdom and United States [2, 3] to 12% in Japan
[4] and 36% in Thailand [5]. In addition, TTV DNA detection
rates are highly dependent on the primer sequences used for
the PCR assays: Takahashi et al. [6] reported an astonishing
92% positivity rate among healthy Japanese subjects using a
novel set of primers.
TTV has the properties of an unenveloped, single-stranded
DNA virus, possibly belonging to the Parvoviridae or
Circoviridae families [4]. Sequence analysis of viral DNA from a
number of different sera demonstrates the existence of 26
distinct TTV genotypes, each of which may exist as a variety of
subtypes [2, 4, 7, 8]. The role of TTV as a cause of liver disease,
either as a sole agent or as a cofactor in other forms of chronic
hepatitis, is unknown.
Hepatitis C virus (HCV) is the most common cause of
chronic viral hepatitis in the developed world. At least 20% of
persons infected with HCV develop chronic liver disease within
20 years (e.g., cirrhosis and hepatocellular carcinoma) [9, 10].
The factors that determine the progression of HCV disease in
the liver, however, remain largely unknown. We tested a large
number of sera for the presence of TTV DNA, to determine
the frequency of coinfection in persons known to be chronically
infected with HCV and to assess the effect of coinfection on
the severity of liver disease in such persons. The availability of
stored sera obtained over several years from these patients also
allowed us to examine the chronicity of TTV infection in this
setting. In addition, we conducted preliminary comparative
sequence analysis of the virus populations present within the
infected subjects.
Methods
Sera. Serum samples stored in untouched aliquots at 2807C
were derived from patients with HCV infection (n 5 122).
Eightytwo of these sera were previously shown to contain antibodies
(second generation ELISA; Ortho, Amersham, UK) to both HCV
and HCV RNA, as assessed by reverse transcriptase (RT)PCR
(Roche Amplicor, Welwyn Garden City, UK). The remaining 40
sera were HCV antibody-positive but negative by RT-PCR. None
of the HCV-infected patients were hemophiliacs. Diagnostic liver
biopsy had been performed on 77 of the RT-PCR positive patients,
and liver disease severity was assessed by the Knodell score [11].
In addition, stored sera from 26 unselected blood donors were
tested.
TTV DNA detection. DNA was extracted from 150 mL of
serum by means of the Nucleospin Virus kit (Biogene, Kimb (...truncated)