T1764G1766 core promoter double mutants are restricted to Hepatitis B virus strains with an A1757 and are common in genotype D

Hossein Sendi1,2,3, Marjan Mehrab-Mohseni1,3, Mohammad R. Zali3, Helene Norder1,2 and Lars O. Magnius1,2

1 Department of Virology, Swedish Institute for Infectious Disease Control, SE-171 82 Solna, Sweden
2 Microbiology and Tumor Biology Center, Karolinska Institutet, SE-171 77 Stockholm, Sweden
3 Research Center for Gastroenterology and Liver Diseases, Shaheed Beheshti University of Medical Sciences, Taleghani Hospital, Tabnak Avenue, 19857 Tehran, Iran

Correspondence
Lars O. Magnius
Lars.Magnius{at}smi.ki.se


   ABSTRACT
Top
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
 
To investigate the role of pre-core and basal core promoter (BCP) mutants in hepatitis B e antigen (HBeAg)-negative chronic hepatitis B (e-CHB) in Iran, Hepatitis B virus strains from 30 patients and 42 anti-HBe-positive asymptomatic carriers (ASCs) were characterized. G1896A pre-core stop mutants, detected in 77 % of e-CHB patients and 85 % of ASCs, showed no association with virus load or aminotransferase levels. Twenty per cent of e-CHB patients and 31 % of ASCs harboured T1762A1764 mutants. When this double mutation was associated with G1757, it was linked to a higher virus load in patients than when it was associated with A1757 (105·2±1·8 vs 103·2±0·8 copies ml–1; P=0·004). Interestingly, the most common BCP mutations were T1764 and G1766, which were present in 33 % of e-CHB patients and 29 % of ASCs. These were associated with higher virus load and aminotransferase levels compared with patients lacking core promoter mutations, although this was not significant. The T1764G1766 double mutation was only present in strains with A1757 (P<0·001), which is more frequent in strains of genotype D than in those belonging to other genotypes. On the other hand, the T1762A1764 double mutation was found more frequently in association with G1757 than with A1757. The T1762A1764 double mutation forms a binding site for hepatocyte nuclear factor 1 (HNF1), which is constrained by A1757. However, the T1764G1766 double mutant may form a binding site for HNF3. Thus, position 1757 affects the emergence of promoter double mutants and would predict a relative genotypic restriction of both the T1762A1764 and the T1764G1766 double mutants.

The GenBank/EMBL/DDBJ accession numbers for the sequences and primer sequences reported in this study are DQ102486–DQ102557.


   INTRODUCTION
Top
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
 
Hepatitis B virus (HBV) infection is a global health problem and there are more than 350 million chronic carriers of this virus. During the course of chronic hepatitis B (CHB), a large proportion of the carriers achieve significant reduction in virus replication with loss of HBe antigen (HBeAg) and seroconversion to its antibody, anti-HBe (Realdi et al., 1980; Hoofnagle et al., 1981; Fattovich et al., 1986). Several HBeAg-negative subjects have persistent or intermittently high HBV replication associated with liver inflammation and ongoing fibrosis (Hadziyannis et al., 1983; Bonino et al., 1986). This form of CHB, defined as HBeAg-negative CHB (e-CHB) (Hadziyannis, 1995), is mostly associated with mutations in the basal core promoter (BCP) and pre-core regions that result in reduction or prevention of HBeAg synthesis without affecting the replicative ability of the virus. The most well known is a G to A mutation at nt 1896 in the pre-core region (Carman et al., 1989), which prevents HBeAg production by converting codon 28 of this region into a stop codon with premature termination of HBeAg translation. Pre-core mutants appear early in the HBeAg-positive phase, being detectable in minute amounts together with the predominant wild-type of HBV. However, they will be selected for and predominate over the wild-type virus during the phase of seroconversion of HBeAg to anti-HBe, when immune tolerance to HBV is lost (Okamoto et al., 1990; Hadziyannis, 1995; Chang et al., 1998).

Mutations in the BCP have also been reported in patients with e-CHB or fulminant hepatitis (Okamoto et al., 1994; Kurosaki et al., 1996; Kidd-Ljunggren et al., 1997; Chan et al., 1999). The most commonly detected mutations within the BCP region are an A1762T transversion and a G1764A transition. In vitro and in vivo studies have shown that these mutations decrease the transcription of pre-core mRNA and hence the secretion of HBeAg (Buckwold et al., 1996; Günther et al., 1998; Laras et al., 2002).

The prevalence of pre-core and BCP mutations in HBV strains from Iranian e-CHB patients and asymptomatic carriers (ASCs) and their association with HBV DNA and aminotransferase levels are investigated in this study.


   METHODS
Top
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
 
Patients.
The study included 97 patients who were negative for HBeAg and positive for anti-HBe referred to the Hepatology Clinic of Taleghani General Hospital, Tehran, Iran, between March 2000 and April 2003. All patients had been HBsAg-positive for more than 6 months, were tested for HBeAg and anti-HBe, and underwent biochemical liver tests including those for serum albumin, bilirubin, aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase. None of the patients were co-infected with Hepatitis C virus or Hepatitis delta virus, had other concomitant causes of liver diseases, had been treated with interferon or lamivudine, or had received chemotherapy or immunosuppressive drugs.

Based on clinical criteria, the patients were grouped as follows: (i) e-CHB patients with elevated aminotransferase levels above twice the normal range for more than 6 months or clinical or histological signs of chronic liver disease; and (ii) ASCs without signs of disease activity, as determined by elevated ALT or AST levels. Thirty-five patients were categorized as e-CHB, whereas 62 were classified as ASCs. All patients had given their consent for enrolling in the study.

Identification of HBV basic core promoter and pre-core mutations and HBV genotypes.
HBV DNA was extracted from 200 µl serum with a QIAamp DNA mini kit (Qiagen). Pre-core amplification was carried out with primers hepA and hep66 and was nested with primers hepA and hep68 (Arauz-Ruiz et al., 1997). For core promoter amplification, hep64 and hep70 were used in the first round of PCR and hep64 and hep54 were used in the nested PCR. Amplification of the S region was done by using primers hep75 and 73b for the first PCR and hep3/hep33 and hep4/hep34 for nesting, respectively (Norder et al., 1992). PCR amplificates were purified with a GFX PCR DNA and gel-band purification kit (Amersham Biosciences). Purified products were used as templates in the sequencing reaction using the dideoxynucleotide chain-termination method with an ABI PRISM BigDye terminator cycle-sequencing reaction kit (version 3; Applied Biosystems) and the primers used in the PCR were used as sequencing primers. All amplification products were sequenced bi-directionally. An ABI PRISM 3100 genetic analyser (Applied Biosystems) was used for electrophoresis and data collection. Sequences obtained were edited by using the SeqMan program in the LASERGENE package (DNASTAR).

HBV DNA quantification.
HBV DNA quantification was carried out by using Affigene HBV VL kits (Sangtec Molecular Diagnostics) according to the manufacturer's instructions. In each set of PCR-ELISA, five different standards from the kit with 0, 12, 60, 600, 6x103 and 6x105 copies ml–1 were run in parallel with the samples. The limit of detection was 500 copies ml–1 and the quantification range was 2x103–4x107 copies ml–1.

Statistical analysis.
Statistical analysis was performed with {chi}2 or Fisher's exact tests and with the independent t-test for continuous variables using the SPSS version 11.0 software package. P values (two-tailed) less than 0·05 were considered statistically significant. Logarithms of HBV DNA levels were used for the statistical comparisons.


   RESULTS
Top
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
 
Demographic, biochemical and virological data of the patients are shown in Table 1. There was no significant difference in mean age or in male to female ratio between ASCs and e-CHB patients. HBV DNA levels were significantly higher in e-CHB patients than in ASCs (P<0·005).


View this table:
[in this window]
[in a new window]
 
Table 1. Demographic and biochemical data and HBV DNA levels in e-CHB patients and ASCs

 
HBV genotypes
The whole S region was sequenced for the 72 strains for which the BCP region could be sequenced. All were found to belong to HBV genotype D based on phylogenetic analysis (data not shown).

Pre-core mutations
The pre-core region of the infecting strain could be amplified from serum of 80 % of the patients (Table 2). A high proportion (82 %) of the patients harboured A1896 pre-core stop mutants, whereas 37 % harboured G1899A mutants with no significant differences in frequencies between e-CHB patients and ASCs. The A1899 mutation was mostly accompanied by the A1896 mutation (Fig. 1). There was no difference in mean age between patients with A1896 mutants and those without, but the patients with A1899 mutants were older than the other patients (43 vs 35 years; P=0·014). There was a slightly, although not significantly, higher mean level of ALT in e-CHB patients with pre-core stop mutants than in patients without these mutants (190 vs 135 IU). All strains had T at nucleotide positions 1850 and 1858, which is as expected for genotype D.


View this table:
[in this window]
[in a new window]
 
Table 2. Frequency of pre-core/BCP mutants according to clinical status

 


View larger version (46K):
[in this window]
[in a new window]
 
Fig. 1. Sequences of the basal core promoter (BCP) positions 1746–1790 and the corresponding part of the X protein plus the substitutions at positions 1896 and 1899 in the pre-core region of 72 HBV strains, for which sequences were available from both regions. The binding sites for hepatocyte nuclear factors 1 (HNF1) and 3 (HNF3) are aligned with the corresponding region of the HBV genome for strains with T1762A1764 and T1764G1766 double mutants, respectively. The symbols for nucleotide ambiguities are as follows: V, A/C/G; W, A/T; R, A/G; K, G/T; Y, C/T.

 
One ASC had a G1816T mutation, thus changing the Met start codon into Ile in the pre-core region, and one of the e-CHB patients had a C1817T mutation, thus changing the Gly at codon 2 into a stop codon. Neither of these patients harboured strains with A1896 or A1899 mutations.

Core promoter mutations
The core promoter regions of the infecting strains could be sequenced from serum for 74 % of the patients (Table 2). The sequences of the region bound by nt 1742 and 1791 for these strains are shown in Fig. 1. A high proportion (81 %) of the patients had strains with at least one nucleotide substitution in the BCP region. The most common mutation was C1766G (38 % patients), followed by G1764A (32 %), G1764T (31 %), A1762T (29 %) and T1753C (20 %), with no significant difference in frequencies between e-CHB patients and ASCs; a G1757A substitution was found in 71 % of the strains and was considered to be a genotype D wild-type substitution.

Nineteen patients (26 %) had strains with T1762A1764 mutations in the core promoter regions. The T1762A1764 double mutation was found more frequently in conjunction with G1757 than with A1757 (Table 4; P<0·001, Fisher's exact test). The 12 patients with this double mutation plus G1757 had the highest virus loads and also significantly higher virus loads than six of seven patients with this double mutation plus A1757 [105·2±1·8 vs 103·2±0·8 copies ml–1; P=0·004 (Table 3)]. One patient in the latter group had deletions in the internal part of the core region and had a high virus titre (107·6 copies ml–1) and was not included in the comparison. Patients with G1757 were older than patients with A1757 (50 vs 29 years; P=0·002). A mutation to C1753 was significantly more often present in strains with T1762A1764 than in strains lacking this double mutation, irrespective of A/G1757 (P<0·001). Nine patients (three e-CHB and six ASCs) harbouring strains with the mutational pattern C1753T1762A1764 accompanied by G1757 had higher HBV DNA levels (105·2 vs 103·3 copies ml–1) and were older (52 vs 28 years) than the three patients, all ASCs, with the pattern C1753T1762A1764 accompanied by A1757. This difference in age was significant (P=0·003).


View this table:
[in this window]
[in a new window]
 
Table 4. Prevalence of different promoter mutations according to the substitutions at nt 1757

 

View this table:
[in this window]
[in a new window]
 
Table 3. Relationship between pre-core/BCP mutants and age, sex ratio, and transferase and HBV DNA levels in patients of different clinical states

Pre-core and BCP sequence information is based on 78 and 72 patients, respectively. M/F, Male to female ratio; VL, virus load given as log mean±SD copies ml–1. Items in bold are significantly statistically different from non-bold items with the same italic superscripts at the following P values: a1, P=0·014; a2, P=0·005; a3, P=0·007; b1, P=0·027; b2, P=0·001; g, P=0·020; d, P=0·023; e1, P=0·002; e2, P=0·039; e3, P=0·007; m1, P=0·004; m2, P=0·049; m3, P=0·019.

 
Thirty-one per cent of patients had strains with another double mutation, i.e. T1764G1766 (Table 2). Only strains with A1757 had T1764G1766 mutations (P<0·001, Fisher's exact test). None of the strains with T1762A1764 mutations had G1766 and none of the strains with T1764G1766 mutations had T1762 (Table 4; P<0·001, Fisher's exact test). e-CHB patients harbouring T1764G1766 mutants tended to have higher HBV DNA and aminotransferase levels than patients with strains that had wild-type BCP sequences, although these differences were not significant (Table 3).

Two e-CHB patients had the triple mutational pattern T1762T1766A1768 (Fig. 1). One of them had a high virus load, 105·7 copies ml–1. None of the ASCs had this mutational pattern.


   DISCUSSION
Top
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
 
The pre-core A1896 mutants may be detected in 20–95 % of HBeAg-negative patients worldwide, with geographical variation in its prevalence due to its emergence being restricted mainly to HBV genotypes B–E, which have T1858 (Li et al., 1993). It is highly prevalent in the Mediterranean countries (Funk et al., 2002), where genotype D predominates (Norder et al., 1993).

Despite a high prevalence of hepatitis B in the Middle East, few studies have been conducted on the relative role of pre-core and core promoter mutants in e-CHB patients in this region. In a recent study of 23 HBV strains all belonging to genotype D from Iranian anti-HBe-positive patients, 56 % were reported to have A1896 pre-core mutations (Amini-Bavil-Olyaee et al., 2005).

In our study, the overall prevalence of pre-core mutants was 87 %, including the two other mutants in the pre-core region, one of which abolished the pre-core start codon. A similar prevalence of pre-core mutants has been reported in Turkey (Bozdayi et al., 1999). The e-CHB patients with A1896 mutants did not have higher levels of HBV DNA or aminotransferases compared with the e-CHB patients who lacked these mutants, which is in agreement with other studies (Okamoto et al., 1990; Takeda et al., 1990; Gandhe et al., 2003). There are, however, studies showing that e-CHB with pre-core mutants frequently take a more aggressive course, with the development of cirrhosis and/or hepatocellular carcinoma (HCC) (Bonino et al., 1986; Brunetto et al., 1991; Naoumov et al., 1992).

The A1899 mutant, detected in 37 % of patients, has been associated with increased severity of liver disease (Carman et al., 1989; Tillmann et al., 1995; Chan et al., 2000). In our study, patients with A1899 mutants were older than the other patients, suggesting that these mutants evolve in patients with a longer history of infection and who therefore will have more advanced liver disease more frequently. In a study from Spain, all e-CHB patients with T1762 and A1899 mutations associated with A1764 and A1896 mutations had liver cirrhosis and it was concluded that the increased risk of severe liver disease was due to enhanced HBV replication (Jardi et al., 2004). In our study, both T1762 and A1899 mutants were found in older patients, which may explain why they have been found more frequently in cirrhotic patients in the Spanish study.

Eighty-one per cent of our patients harboured strains with at least one substitution in the region bound by nt 1742 and 1791, which is important in the control of pre-C RNA transcription (Yuh et al., 1992). Clinical studies have correlated core promoter mutants with virus load, liver disease and response to antiviral therapy (Chan et al., 1999; Baptista et al., 1999; Erhardt et al., 2000). Most of these have focused on T1762A1764 mutants and have shown an association between these mutants and the severity of liver disease (Sato et al., 1995; Baumert et al., 1996; Scaglioni et al., 1997; Kao et al., 2003). In our study, the T1762A1764 double mutant was more frequent in older patients, which is in accordance with a recent study from Albania suggesting that longer duration of infection could be important in the emergence of these mutants (Kondili et al., 2005). e-CHB patients harbouring strains with T1762A1764 mutations associated with G1757 were found to have higher HBV DNA levels than patients with T1762A1764 mutations associated with A1757. A mutation to C1753 was present significantly more often in strains with T1762A1764 than in strains lacking this double mutation. The triple C1753T1762A1764 mutant has previously been isolated from HBeAg-positive patients and yielded highly replicating clones, but showed only a minor reduction in HBeAg expression, which is similar to isolates with only the T1762A1764 double mutation (Parekh et al., 2003). When patients harbouring strains with C1753T1762A1764 associated with G1757, which is less prevalent in genotype D, were compared with patients harbouring strains with C1753T1762A1764 associated with A1757, the latter had lower virus load, indicating that the T1762A1764 mutation was less efficient when associated with A1757 and in conjunction with C1753.

The T1764G1766 mutant has previously been found in a child with HCC and was shown to give a 2·8-fold increase in core promoter activity. This mutant was suggested to form a putative new binding site for the transcription factor hepatocyte nuclear factor 3 (HNF3) (Gerner et al., 1999). Although this previously seemed to be a rare mutation, it was found in almost one-third of our patients. These patients had higher virus load and aminotransferase levels than patients lacking core promoter mutations, although these differences were not significant.

Two e-CHB patients were found to harbour a unique triple mutant, T1762T1766A1768, in one case associated with a high virus load. The T1766A1768 double mutant has previously been reported in an outbreak of fulminant hepatitis and was shown to display a more than 10-fold enhancement in promoter activity, whereas the T1762A1764 mutant showed just about threefold enhancement compared with wild-type clones (Baumert et al., 1996; Gerner et al., 1999).

The T1762A1764 mutant changes two codons in the X protein with the substitutions Met130Ile131, which would decrease the transactivating activity of the X gene product on both pre-core and core transcription (Li et al., 1999). It has been suggested that the T1762A1764 mutant will restore core transcription, but not pre-core transcription, by creating an HNF1 site (Li et al., 1999). The T1764G1766 mutant will create a substitution to Leu131 in the X gene product, which might also suppress both pre-core and core transcription. The latter could then be restored by the T1764G1766 mutation by creating an HNF3 site (Gerner et al., 1999). The combined putative mutational patterns T1762A1764G1766 or T1762T1764G1766 would not generate compensatory HNF1- or HNF3-binding sites and would therefore not be selected for, which explains their absence in this study.

Most interestingly, only patients harbouring strains with A1757 had T1764G1766 mutations. As the vast majority of HBV strains belonging to genotypes A to C have G1757 and it is only in genotypes D and E that an A is more common than G at this position (Table 5), the T1764G1766 mutant is likely to occur mainly in genotype D and possibly genotype E. As G1757 forms the 5' end of the HNF1-binding site, T1762A1764 mutations are more efficient when there is a G1757 instead of an A1757. However, when there is an A1757, the emergence of the T1764G1766 mutation will create an HNF3 site and will be more efficient than the T1762A1764 mutation in conjunction with A1757 in compensatory upregulation of HBcAg production. The most likely reason for the complete absence of the T1764G1766 mutation in association with G1757 is the high efficiency of the T1762A1764 mutation in this molecular context. This view is supported by HBV DNA levels being highest for T1762A1764 associated with G1757, intermediate for T1764G1766 and lowest for T1762A1764 associated with an A1757.


View this table:
[in this window]
[in a new window]
 
Table 5. Prevalence of A1757 in different genotypes based on the 234 complete human HBV genomes investigated by Norder et al. (2004)

 
In conclusion, A1757 makes the T1764G1766 double mutation more efficient than the T1762A1764 double mutation at increasing core transcription and virus replication. In agreement with this view, the T1762A1764 double mutation was found more frequently in conjunction with G1757 than in conjunction with A1757. Apart from the described emergence of the pre-core 1896 stop mutation being restricted to HBV genotypes with a T1858, other mutational constraints depending on a single silent nucleotide polymorphism within the virus genome have not been described previously.


   ACKNOWLEDGEMENTS
 
We want to acknowledge Drs Hamid Mohaghegh and Hooman Nafisi for serum sampling for this study and also Drs Saeed Shahraz and Babak Noorinayer for their coordination of the research. This study was funded by grants from the Swedish Institute 00475/2004 (H. S.) and the Ministry of Health, the Islamic Republic of Iran (M. R. Z.).


   REFERENCES
Top
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
 
Amini-Bavil-Olyaee, S., Sarrami-Forooshani, R., Mahboudi, F., Sabahi, F., Adeli, A., Noorinayer, B., Azizi, M. & Reza Zali, M. (2005). Genotype characterization and phylogenetic analysis of hepatitis B virus isolates from Iranian patients. J Med Virol 75, 227–234.[CrossRef][Medline]

Arauz-Ruiz, P., Norder, H., Visoná, K. A. & Magnius, L. O. (1997). Genotype F prevails in HBV infected patients of hispanic origin in Central America and may carry the precore stop mutant. J Med Virol 51, 305–312.[CrossRef][Medline]

Baptista, M., Kramvis, A. & Kew, M. C. (1999). High prevalence of 1762T 1764A mutations in the basic core promoter of hepatitis B virus isolated from black Africans with hepatocellular carcinoma compared with asymptomatic carriers. Hepatology 29, 946–953.[CrossRef][Medline]

Baumert, T. F., Rogers, S. A., Hasegawa, K. & Liang, T. J. (1996). Two core promoter mutations in a hepatitis B virus strain associated with fulminant hepatitis result in enhanced viral replication. J Clin Invest 98, 2268–2276.[Abstract/Free Full Text]

Bonino, F., Rosina, F., Rizzetto, M., Rizzi, R., Chiaberge, E., Tardanico, R., Callea, F. & Verme, G. (1986). Chronic hepatitis in HBsAg carriers with serum HBV-DNA and anti-HBe. Gastroenterology 90, 1268–1273.[Medline]

Bozdayi, A. M., Bozkaya, H., Türkyilmaz, A., Aslan, N., Verdi, H., Kence, A. & Uzunalimoglu, Ö. (1999). Polymorphism of precore region of hepatitis B virus DNA among patients with chronic HBV infection in Turkey. Infection 27, 357–360.[CrossRef][Medline]

Brunetto, M. R., Giarin, M. M., Oliveri, F. & 8 other authors (1991). Wild-type and e antigen-minus hepatitis B viruses and course of chronic hepatitis. Proc Natl Acad Sci U S A 88, 4186–4190.[Abstract/Free Full Text]

Buckwold, V. E., Xu, Z., Chen, M., Yen, T. S. B. & Ou, J.-H. (1996). Effects of a naturally occurring mutation in the hepatitis B virus basal core promoter on precore gene expression and viral replication. J Virol 70, 5845–5851.[Abstract]

Carman, W. F., Jacyna, M. R., Hadziyannis, S., Karayiannis, P., McGarvey, M. J., Makris, A. & Thomas, H. C. (1989). Mutation preventing formation of hepatitis B e antigen in patients with chronic hepatitis B infection. Lancet ii, 588–591.

Chan, H. L. Y., Hussain, M. & Lok, A. S. F. (1999). Different hepatitis B virus genotypes are associated with different mutations in the core promoter and precore regions during hepatitis B e antigen seroconversion. Hepatology 29, 976–984.[CrossRef][Medline]

Chan, H. L. Y., Leung, N. W. Y., Hussain, M., Wong, M. L. & Lok, A. S. F. (2000). Hepatitis B e antigen-negative chronic hepatitis B in Hong Kong. Hepatology 31, 763–768.[CrossRef][Medline]

Chang, M. H., Hsu, H. Y., Ni, Y. H., Tsai, K. S., Lee, P. I., Chen, P. J., Hsu, Y. L. & Chen, D. S. (1998). Precore stop codon mutant in chronic hepatitis B virus infection in children: its relation to hepatitis B e seroconversion and maternal hepatitis B surface antigen. J Hepatol 28, 915–922.[CrossRef][Medline]

Erhardt, A., Reineke, U., Blondin, D., Gerlich, W. H., Adams, O., Heintges, T., Niederau, C. & Häussinger, D. (2000). Mutations of the core promoter and response to interferon treatment in chronic replicative hepatitis B. Hepatology 31, 716–725.[CrossRef][Medline]

Fattovich, G., Rugge, M., Brollo, L., Pontisso, P., Noventa, F., Guido, M., Alberti, A. & Realdi, G. (1986). Clinical, virologic and histologic outcome following seroconversion from HBeAg to anti-HBe in chronic hepatitis type B. Hepatology 6, 167–172.[Medline]

Funk, M. L., Rosenberg, D. M. & Lok, A. S. F. (2002). World-wide epidemiology of HBeAg-negative chronic hepatitis B and associated precore and core promoter variants. J Viral Hepat 9, 52–61.[CrossRef][Medline]

Gandhe, S. S., Chadha, M. S., Walimbe, A. M. & Arankalle, V. A. (2003). Hepatitis B virus: prevalence of precore/core promoter mutants in different clinical categories of Indian patients. J Viral Hepat 10, 367–382.[CrossRef][Medline]

Gerner, P., Lausch, E., Friedt, M., Tratzmüller, R., Spangenberg, C. & Wirth, S. (1999). Hepatitis B virus core promoter mutations in children with multiple anti-HBe/HBeAg reactivations result in enhanced promoter activity. J Med Virol 59, 415–423.[CrossRef][Medline]

Günther, S., Piwon, N. & Will, H. (1998). Wild-type levels of pregenomic RNA and replication but reduced pre-C RNA and e-antigen synthesis of hepatitis B virus with C(1653)->T, A(1762)->T and G(1764)->A mutations in the core promoter. J Gen Virol 79, 375–380.[Abstract]

Hadziyannis, S. J. (1995). Hepatitis B e antigen negative chronic hepatitis B: from clinical recognition to pathogenesis and treatment. Viral Hepatitis Rev 1, 7–36.

Hadziyannis, S. J., Lieberman, H. M., Karvountzis, G. G. & Shafritz, D. A. (1983). Analysis of liver disease, nuclear HBcAg, viral replication, and hepatitis B virus DNA in liver and serum of HBeAg vs. anti-HBe positive carriers of hepatitis B virus. Hepatology 3, 656–662.[Medline]

Hoofnagle, J. H., Dusheiko, G. M., Seeff, L. B., Jones, E. A., Waggoner, J. G. & Bales, Z. B. (1981). Seroconversion from hepatitis B e antigen to antibody in chronic type B hepatitis. Ann Intern Med 94, 744–748.[Medline]

Jardi, R., Rodriguez, F., Buti, M. & 7 other authors (2004). Mutations in the basic core promoter region of hepatitis B virus. Relationship with precore variants and HBV genotypes in a Spanish population of HBV carriers. J Hepatol 40, 507–514.[CrossRef][Medline]

Kao, J.-H., Chen, P.-J., Lai, M.-Y. & Chen, D.-S. (2003). Basal core promoter mutations of hepatitis B virus increase the risk of hepatocellular carcinoma in hepatitis B carriers. Gastroenterology 124, 327–334.[CrossRef][Medline]

Kidd-Ljunggren, K., Öberg, M. & Kidd, A. H. (1997). Hepatitis B virus X gene 1751 to 1764 mutations: implications for HBeAg status and disease. J Gen Virol 78, 1469–1478.[Abstract]

Kondili, L. A., Brunetto, M. R., Maina, A. M., Argentini, C., Chionne, P., La Sorsa, V., Resuli, B., Mele, A. & Rapicetta, M. (2005). Clinical and molecular characterization of chronic hepatitis B in Albania: a country that is still highly endemic for HBV infection. J Med Virol 75, 20–26.[CrossRef][Medline]

Kurosaki, M., Enomoto, N., Asahina, Y., Sakuma, I., Ikeda, T., Tozuka, S., Izumi, N., Marumo, F. & Sato, C. (1996). Mutations in the core promoter region of hepatitis B virus in patients with chronic hepatitis B. J Med Virol 49, 115–123.[CrossRef][Medline]

Laras, A., Koskinas, J. & Hadziyannis, S. J. (2002). In vivo suppression of precore mRNA synthesis is associated with mutations in the hepatitis B virus core promoter. Virology 295, 86–96.[CrossRef][Medline]

Li, J.-S., Tong, S.-P., Wen, Y.-M., Vitvitski, L., Zhang, Q. & Trépo, C. (1993). Hepatitis B virus genotype A rarely circulates as an HBe-minus mutant: possible contribution of a single nucleotide in the precore region. J Virol 67, 5402–5410.[Abstract]

Li, J., Buckwold, V. E., Hon, M. & Ou, J. (1999). Mechanism of suppression of hepatitis B virus precore RNA transcription by a frequent double mutation. J Virol 73, 1239–1244.[Abstract/Free Full Text]

Naoumov, N. V., Schneider, R., Grotzinger, T., Jung, M. C., Miska, S., Pape, G. R. & Will, H. (1992). Precore mutant hepatitis B virus infection and liver disease. Gastroenterology 102, 538–543.[Medline]

Norder, H., Hammas, B., Löfdahl, S., Couroucé, A.-M. & Magnius, L. O. (1992). Comparison of the amino acid sequences of nine different serotypes of hepatitis B surface antigen and genomic classification of the corresponding hepatitis B virus strains. J Gen Virol 73, 1201–1208.[Abstract]

Norder, H., Hammas, B., Lee, S.-D., Bile, K., Couroucé, A.-M., Mushahwar, I. K. & Magnius, L. O. (1993). Genetic relatedness of hepatitis B viral strains of diverse geographical origin and natural variations in the primary structure of the surface antigen. J Gen Virol 74, 1341–1348.[Abstract]

Norder, H., Couroucé, A.-M., Coursaget, P., Echevarria, J. M., Lee, S.-D., Mushahwar, I. K., Robertson, B. H., Locarnini, S. & Magnius, L. O. (2004). Genetic diversity of hepatitis B virus strains derived worldwide: genotypes, subgenotypes, and HBsAg subtypes. Intervirology 47, 289–309.[CrossRef][Medline]

Okamoto, H., Yotsumoto, S., Akahane, Y. & 7 other authors (1990). Hepatitis B viruses with precore region defects prevail in persistently infected hosts along with seroconversion to the antibody against e antigen. J Virol 64, 1298–1303.[Medline]

Okamoto, H., Tsuda, F., Akahane, Y., Sugai, Y., Yoshiba, M., Moriyama, K., Tanaka, T., Miyakawa, Y. & Mayumi, M. (1994). Hepatitis B virus with mutations in the core promoter for an e antigen-negative phenotype in carriers with antibody to e antigen. J Virol 68, 8102–8110.[Abstract]

Parekh, S., Zoulim, F., Ahn, S. H. & 7 other authors (2003). Genome replication, virion secretion, and e antigen expression of naturally occurring hepatitis B virus core promoter mutants. J Virol 77, 6601–6612.[Abstract/Free Full Text]

Realdi, G., Alberti, A., Rugge, M., Bortolotti, F., Rigoli, A. M., Tremolada, F. & Ruol, A. (1980). Seroconversion from hepatitis B e antigen to anti-HBe in chronic hepatitis B virus infection. Gastroenterology 79, 195–199.[Medline]

Sato, S., Suzuki, K., Akahane, Y. & 8 other authors (1995). Hepatitis B virus strains with mutations in the core promoter in patients with fulminant hepatitis. Ann Intern Med 122, 241–248.[Abstract/Free Full Text]

Scaglioni, P. P., Melegari, M. & Wands, J. R. (1997). Biologic properties of hepatitis B viral genomes with mutations in the precore promoter and precore open reading frame. Virology 233, 374–381.[CrossRef][Medline]

Takeda, K., Akahane, Y., Suzuki, H., Okamoto, H., Tsuda, F., Miyakawa, Y. & Mayumi, M. (1990). Defects in the precore region of the HBV genome in patients with chronic hepatitis B after sustained seroconversion from HBeAg to anti-HBe induced spontaneously or with interferon therapy. Hepatology 12, 1284–1289.[Medline]

Tillmann, H., Trautwein, C., Walker, D., Michitaka, K., Kubicka, S., Boker, K. & Manns, M. (1995). Clinical relevance of mutations in the precore genome of the hepatitis B virus. Gut 37, 568–573.[Abstract]

Yuh, C.-H., Chang, Y.-L. & Ting, L.-P. (1992). Transcriptional regulation of precore and pregenomic RNAs of hepatitis B virus. J Virol 66, 4073–4084.[Abstract]

Received 10 March 2005; accepted 23 May 2005.



This Article
Abstract
Full Text (PDF)
Alert me when this article is cited
Alert me if a correction is posted
Citation Map
Services
Email this article to a friend
Similar articles in this journal
Similar articles in PubMed
Alert me to new issues of the journal
Download to citation manager
Google Scholar
Articles by Sendi, H.
Articles by Magnius, L. O.
Articles citing this Article
PubMed
PubMed Citation
Articles by Sendi, H.
Articles by Magnius, L. O.
Agricola
Articles by Sendi, H.
Articles by Magnius, L. O.


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
INT J SYST EVOL MICROBIOL MICROBIOLOGY J GEN VIROL
J MED MICROBIOL ALL SGM JOURNALS