Whole genome analysis of hepatitis B virus before and during long-term therapy in chronic infected patients: Molecular characterization, impact on treatment and liver disease progression

Abstract

Hepatitis B virus (HBV) infection remains a serious public health concern worldwide despite the availability of an efficient vaccine and the major improvements in antiviral treatments. The aim of the present study is to analyze the mutational profile of the HBV whole genome in ETV non-responder chronic HBV patients, in order to investigate antiviral drug resistance, immune escape, and liver disease progression to Liver Cirrhosis (LC) or Hepatocellular Carcinoma (HCC). Blood samples were collected from five chronic hepatitis B patients. For each patient, two plasma samples were collected, before and during the treatment. Whole genome sequencing was performed using Sanger technology. Phylogenetic analysis comparing the studied sequences with reference ones was used for genotyping. The mutational profile was analyzed by comparison with the reference sequence M32138. Genotyping showed that the studied strains belong to subgenotypes D1, D7, and D8. The mutational analysis showed high genetic variability. In the RT region of the polymerase gene, 28 amino acid (aa) mutations were detected. The most significant mutations were the pattern rtL180M + rtS202G + rtM204V, which confer treatment resistance. In the S gene, 35 mutations were detected namely sP120T, sT126S, sG130R, sY134F, sS193L, sI195M, and sL216stop were previously described to lead to vaccine, immunotherapy, and/or diagnosis escape. In the C gene, 34 mutations were found. In particular, cG1764A, cC1766G/T, cT1768A, and cC1773T in the BCP; cG1896A and cG1899A in the precore region and cT12S, cE64D, cA80T, and cP130Q in the core region were associated with disease progression to LC and/or HCC. Other mutations were associated with viral replication increase including cT1753V, cG1764A/T, cC1766G/T, cT1768A, and cC1788G in the BCP as well as cG1896A and cG1899A in the precore region. In the X gene, 30 aa substitutions were detected, of which substitutions xT36D, xP46S, xA47T, xI88F, xA102V, xI127T, xK130M, xV131I, and xF132Y were previously described to lead to LC and/or HCC disease progression. In conclusion, our results show high genetic variability in the long-term treatment of chronic HBV patients causing several effects. This could contribute to guiding national efforts to optimize relevant HBV treatment management in order to achieve the global hepatitis elimination goal by 2030.

Introduction

Hepatitis B virus (HBV) infection remains a serious public health concern worldwide despite the availability of an efficient vaccine and the major improvements in antiviral treatments. The World Health Organization (WHO) estimates that, in 2021, approximately 296 million persons are chronic HBV carriers. Among them, 820,000 represent a high risk of mortality caused by developing progressive liver diseases including hepatocellular carcinoma (HCC) and liver cirrhosis (LC) (WHO, 2021).

The genome of HBV is a circular DNA partially double-stranded of 3.2 kb and classified into 10 genotypes from A to J (Sunbul, 2014). It is organized into four main open overlapped reading frames (ORFs; pre-S1/pre-S2/S, pre-C/C, P, and X), encoding several proteins including the surface proteins S, M, and L holding the HBs antigen (HBsAg), the precore/core proteins holding HBeAg and HBcAg antigens, the polymerase (P), and the X protein holding the antigen HBxAg. Thus, mutations that occur in one gene can result in significant changes in the other overlapping genes.

Long-term treatment of HBV chronic patients with the available antiviral molecules can lead to the emergence of mutations throughout the whole genome. Mutations that occur within the reverse transcriptase (RT) domain of the P gene, target of antiviral treatment, may lead to treatment failure (Locarnini and Mason, 2006). Potential resistance-related mutations are grouped into 4 categories, primary mutations (category 1) could reduce antiviral susceptibility and HBV replication fitness. Secondary/compensatory mutations (category 2) developed subsequently and could restore functional defects in the RT activity of HBV caused by primary mutations. Putative antiviral resistance mutations (category 3) were reported as possible drug-resistant mutations but not verified experimentally and may be related to prolonged treatment or replication compensation. Pre-treatment mutations (category 4) could be found among treatment-naive patients but their role in antiviral treatment resistance has not been elucidated (Liu et al., 2010; Ciftci et al., 2014).

Moreover, mutations that emerge throughout a prolonged therapy could affect not only the RT region (Locarnini and Mason, 2006) but also the different overlapping genes. Therefore, such variations might result in hepatitis B immunoglobulin (HBIG) therapy escape, vaccine escape, misdiagnosis, and immune escape. They also could enhance viral replication capacity and viral persistence leading to the progression of severe liver diseases such as HCC or LC (Sheldon et al., 2006; Sheldon, 2008; Rajoriya et al., 2017).

On the other hand, it has been found that the presence of pre-existing naturally occurring mutations in treatment-naive patients may influence the efficacy of antiviral treatments. Therefore, knowledge of the mutational profile by whole genome sequencing of the HBV genome, for chronically infected patients, is of great interest for a complete diagnosis toward an efficient therapy scheme.

For HBV chronic patients in Tunisia, the national therapeutic schema is based on Entecavir (ETV) as a first-line of HBV treatment, and it is fully covered by the National Health Insurance Fund (NHIF), in case of resistance, Tenofovir disoproxil fumarate (TDF), is recommended alone or combined to ETV. However, the TDF is not covered by the NHIF.

The aim of the present study is to analyze the mutational profile through the HBV whole genome in ETV non-responder chronic HBV patients, in order to investigate antiviral drug resistance, immune escape, and liver disease progression to LC or HCC.

Materials and methods

Patients and samples

HBV chronic patients with quantifiable viral load and suspected to be ETV non-responders after viral breakthrough were included in the study. Blood samples were collected from five chronic hepatitis B patients investigated during the routine diagnostic activity of the Laboratory of Clinical Virology in Pasteur Institute of Tunis. For each included patient two plasma samples were collected: one before treatment as part of the pre-treatment diagnostic, and one during the treatment upon request of the treating physician. The period separating the second sample from the date of treatment beginning ranging between 14 and 72 months depending on the time of the viral breakthrough for each patient. Virological and clinical data are shown in Table 1.

Methods

DNA extraction, amplification, and sequencing

DNA was extracted from 200 μl of plasma using the Qiagen QIAamp® DNA extraction kit (QIAGEN® Inc., Hilden, Germany) according to the manufacturer’s instructions. Three pairs of primers previously described by Chekaraou et al. (2010) were used to amplify 3 overlapping amplicons of 1,228-bp (nt 2,817–863), 1,253 bp (nt 448–1,701), and 1,653 bp (nt 1,609–80) covering the whole HBV genome as shown in Figure 1.

Figure 1

PCR reactions were performed in 50 μl of reaction mixture containing 1X polymerase buffer, 1.5 mM MgCl2, 0.2 mM dNTPs, 0.2 μM of each primer, 1.25 U of Taq Core MP® (Applied Biosystems) and nuclease-free water. The amount of DNA extract added varied between 10 to 35 μl depending on the viral load. PCR cycling was as follows: 94°C for 5 min, 40 cycles (94°C for 1 min, 56°C/57°C/62.5°C for regions 1, 2, and 3, respectively, 72°C for 1 min) with a final extension step at 72°C for 10 min. PCR products were analyzed by electrophoresis on 1% agarose gels stained with 1,25X of Red gel™ dye Nucleic Acid (Biotium®) and visualized by UV transilluminator.

The purified template DNA was sequenced using a BigDye Terminator Ready Reaction Cycle Sequencing Kit (Applied Biosystems) using the same primers pairs on an ABI Prism 3130 Genetic Analyzer (Applied Biosystems).

HBV genome assembly, genotyping, and subtyping

The obtained overlapping sequences were then assembled using BLAST multiple sequences software by comparison with a reference sequence (M32138).¹ The generated final sequences were submitted to Genbank under accession numbers: MT591274-MT591281 and OP121186.

Sequence alignment was performed with MAFFT online server using default parameters² by comparing the obtained genomic sequences with 58 reference sequences representing the 10 HBV genotypes (A–J) and their corresponding subgenotypes. The resulting alignment was used to build a maximum likelihood phylogenetic tree using the IQ-TREE web server, supported by 1,000 bootstrap replicates.³ The phylogenetic tree was then visualized using Figtree software.⁴ The tree was rooted using the midpoint rooting method. Genotypes were also confirmed by the National Center for Biotechnology Information’s (NCBI) E-genotype online software.⁵ HBV subtypes were inferred from sequences of the S gene by identifying amino acids (aa) at positions 122, 160, 127, 140, and 159 according to an algorithm previously described (Purdy et al., 2006).

Mutation analysis

Mutational profiles of the nucleotide or amino acid sequences were determined by comparing each gene (P, S, C, and X) before and during treatment with the corresponding reference sequence using Mega 7.026 (Kumar et al., 2016). Mutations’ impacts on treatment, immune response, and liver disease progression were analyzed based on the literature.

Results

HBV whole genome assembly

Whole genome sequences were obtained before and during treatment for 3 patients (1, 2, and 3). For the two remaining patients (4 and 5) we succeeded to obtain the whole genome before treatment. During treatment, the obtained sequence of patient 4 was lacking 408 bp (from nucleotide 45 to nt 453) and for patient 5 we could not be able to amplify the HBV genome which could be due to the low viral load.

HBV genotyping and subtyping

Phylogenetic analysis (Figure 2) showed that all the sequences belong to genotype D. Subgenotyping showed that patients 1 and 4 were infected with subgenotype D7; patients 2 and 3 with D1 and patient 5 with D8, supported by high bootstrap values: 100, 100, and 85, respectively.

Figure 2

Subtyping showed that the studied HBV strains belonged to the ayw2 subtype based on Arg122, Lys160, Pro127, Tyr140, and Gly159 positions.

Genetic variability in the P, S, C, and X genes

Mutational profile of the RT region in the polymerase gene

The mutational analysis of the RT region revealed a total of 28 aa substitutions ranging between 7 and 12 per patient, among them, several potential resistance-related mutations were detected. Primary mutations (category 1), rtS202G and rtM204V, occurred in patients 2 and 3 during treatment. Secondary/compensatory mutations (category 2), were found in 8 aa replacements; 5 were detected before treatment (rtL91I and rtT128N in patient 2; rtQ149K and rtP237T in patients 1, 4, and 5; rtQ267H in patients 4 and 5) and 3 changes emerged during treatment (rtL180M in patients 2 and 3; rtQ215S and rtF221Y in patient 2).

Three putative antiviral resistance mutations (category 3) were detected: rtR153W in treatment-naïve patients 1, 4, and 5 as well as rtD134E and rtC256S during treatment in patients 3 and 1, respectively. Three pre-treatment mutations (category 4) were also found: rtR110G and rtI266R in patient 1 and rtD263E in patient 5.

Mutations that did not fit categories 1 to 4 were classified into “novel amino acid substitutions” and were observed in 12 aa positions. Six variations namely rtE11D, rtH54Y, rtW257Y, rtD263E, rtQ267Y, and rtE271D were found in treatment-naïve patients and six variations namely rtL145M, rtL260F, rtQ267R, rtK270R, rtM309K, and rtN337T occurred during treatment. The aa changes detected in the RT region of the P gene are mentioned in Table 2; Figure 3.

Figure 3

Mutational analysis of the pre-S/S coding regions

A total of 35 aa substitutions were observed in the whole S gene ranging between 3 and 18 mutations per patient, most of them (n = 16) were located in the S region. In the pre-S1 and pre-S2 regions, n = 9 and n = 10 substitutions were observed, respectively. Mutations detected in the S gene are summarized in Table 3; Figure 3. Out of the 16 aa changes in the S region, 10 were clustered in HBsAg epitopes including B-cells epitopes (aa100-160) as follows: 6 (sN3T, sL42R, sL49R, sT57I, sC76Y, and sQ101H) in HBs1 (upstream of aa120); 1 (sP120T) in HBs2 (aa120–123) and 3 (sT126S, sG130R and sY134F) in HBs3 (aa124–137). Furthermore, the major hydrophilic region (MHR, aa99–169) of the S region had accumulated 5 aa variations of which three were within the HBsAg “a” determinant region (aa124–147).

In addition, nine mutations out of the 16 substitutions in the S gene occurred in different CD4 and CD8 recognition epitopes with the following distribution: 6 aa changes (sL42R, sL49R, sT57I, sS193L, sI195M, and sL216*) in T-helper CD4 epitopes (aa21–65/aa186-197/aa215–223) and 3 (sS207R, sP211R, sL213I) within cytotoxic T lymphocyte CD8 epitopes (aa206-215).

Immune escape mutations (sQ101H, sG130R, sY134F, sS207R, and sL213I) were detected in patients 2 and 5, HBsAg vaccine escape mutations (sP120T, sT126S, sS193L, and sI195M) were found in patients 2 and/or 3, HBIG immunotherapy escape mutations (sP120T and sT126S) were detected in patients 2 and 3, respectively, and misdiagnosis mutations (sP120T, sT126S, sI195M, and sL216stop) were observed in patients 2 and/or 3. Other mutations such as sN3T, sL42R, sC76Y, and sP211R are either not reported or with unknown impacts are also detected in our study.

Mutational analysis of the basal core promotor BCP, precore, and core coding regions

The analysis of the 9 HBV genomic sequences bearing the BCP, pre-C, and Core regions is summarized in Table 4. In the BCP region, 14 different aa mutations were identified over 10 sites ranging between 2 and 8 per patient. Nucleotide mutation G1757A was detected in patients 2, 3, 4, and 5; A1762T, G1764A, and T1753V were detected in patients 1, 3, and 5; C1773T in patients 2 and 3, G1764T/C1766G in patient 2 and C1766T/T1768A in patient 3. The double mutation G1764A/A1762T was found in patients 1, 3, and 5.

In the precore region, 2 mutations were identified: G1896A (patient 2) and G1899A (patients 1, 2, and 3). Whereas, 22 mutations were observed in the core region ranging between 2 and 8 per patient. Among them, several amino acid substitutions were found within different antigen immunogenic epitopes. In particular, 4 aa changes were detected in the T-helper CD4 epitopes (aa 35–45 and 48–69; cE40D/Q, cE64D, cT67N, cA69G), 5 in the B-cell epitopes (aa76–89, 105–116, 130–135; cP79Q, cA80T/S, cV85I, cI116V and cP130Q) and 7 in the CTL CD8 epitopes (aa 18–27, 50–69, 74–83, 141–151; cS21T, cE64D, cT67N, cA69G, cV74S/G, cA80T/S, and cR151Q). The detected nucleotide/aa mutations found in the C gene are presented in Table 4; Figure 3.

Mutational analysis in the X coding region

In total, 30 aa substitutions were found in the X gene region ranging between 4 and 14 aa substitutions per patient of which 29 were before treatment beginning and only 1 was during it. Among them 10 variations were detected in the B-cell epitope (aa 29–48) namely: xL34I, xT36D/G, xS38P, xS39P, xP40S, xS41P, xL42P, xS43P, xP46S and xA47T. Four mutations; xK95N, xL98I, xA102, and xT105M; were detected within the T-helper CD4 epitope (aa 91–105) and 2 substitutions (xD119N and xL123W) were detected in CTL CD8 epitope (aa115–123).

As BCP overlaps partially with the HBx coding sequence, mutations at nucleotide positions T1753C/A/G, T1762T, G1764A/T, and C1788G; induce amino acid changes xI127T/D/G, xK130M, xV131I/L and xH139D near the C-terminus of the HBx protein, respectively.

The amino acid substitutions detected in the HBV X gene and their impact are summarized in Table 5; Figure 3.

Discussion

In the present study, we have succeeded to generate the whole HBV genome by amplifying 3 overlapping PCR products covering the entire genome (3.2 kb) using Sanger technology. This technology remains of great importance despite the transition of most laboratories to Next generation sequencing (NGS) technologies. In fact, for small genomes, such as HBV, the Sanger technology is cost effective and more efficient for low viral loads < 10³ IU/ml.

The whole genome was assembled for the 5 patients before the treatment and for 4 patients during the treatment. HBV genome was used for genotyping as well as to study the mutational profile in all the genes (S, P, X, and C) in order to give scientific proof of antiviral treatment resistance.

Genotyping showed that genotype D was detected in the 5 studied patients. This genotype was previously described as a predominant HBV genotype in Tunisia and the Maghreb region as well as in the Middle East with a low co-circulation rate of genotype E (Ayed et al., 2007; Ezzikouri et al., 2008; Ouneissa et al., 2013).

Subgenotypes D1 and D7, found in the present study, were previously described as the most prevalent subgenotypes circulating in Tunisia (Meldal et al., 2009). However, subgenotype D8 is to our knowledge detected for the first time in Tunisia. This subgenotype has been firstly detected in Niger and has been described as a recombinant strain between genotypes D and E (Chekaraou et al., 2010). The recombination analysis of the detected D8 strain, using the NCBI viral genotyping tool, was in line with the previous findings. Further studies are needed on larger population size to estimate the prevalence of this subgenotype in Tunisia.

In the second part of the present study, we have analyzed the mutational profile of all HBV genes P, S, C, and X.

The mutational profile of the RT region in the P gene showed high genetic variability with 28 different mutations. Before the treatment, 14 aa mutations were detected of which patient 2 had already 2 secondary/compensatory substitutions: rtL91I and T128N, described to be a resistance mutation to ETV and/or to LMV, respectively (Torresi et al., 2002a; Mahabadi et al., 2013; Ziaee et al., 2016). For the remaining patients, four mutations were detected and reported to be resistant to at least one of the following antivirals: rtQ267H in patients 4 and 5 to LMV and LdT; rtP237T in patients 1, 4, and 5 to ADV; rtR153W in patients 1, 4, and 5 in addition to rtD263E in patient 5 potentially to TDF (Pollicino et al., 2009; Qin et al., 2013b; Bakhshizadeh et al., 2015; Mokaya et al., 2020). The eight remaining substitutions were not previously described to have an impact on antiviral treatment.

During the treatment, 14 additional aa substitutions occurred. The most significant ones were rtM204V, rtL180M, and rtS202G detected in patients 2 and 3. Indeed, it has been described that the rtM204V substitution is usually associated with the compensatory mutation rtL180M, which restores the replication capacity of rtM204V mutants (Tenney et al., 2004). Thus, the pattern rtL180M, rtS202G, and rtM204V act synergistically not only to increase viral load but also to reduce treatment susceptibility and confer cross-resistance to ETV, TDF, LMV, and LdT (Kamiya, 2003; Li et al., 2005; He et al., 2015; Mokaya et al., 2020). Other emerged aa variations have been detected in our patients and previously described as resistance mutations that reduce the affinity and susceptibility to antiviral drugs namely rtQ215S and rtC256S to LMV; rtQ215S and rtF221Y to ADV; rtD134E, rtQ215S, and rtC256S to TDF (Moriconi et al., 2007; Amini-Bavil-Olyaee et al., 2009; Liu et al., 2009; Pollicino et al., 2009; Ciftci et al., 2014; Park et al., 2019; Mokaya et al., 2020).

Thus, our results support the need to introduce HBV genome sequencing as a pre-treatment diagnosis to predict potential resistance to available antiviral molecules, as well as to monitor the evolution of treatment response.

In addition, we have studied the mutational profile in the preS1, preS2, and S genes. As the coding sequence of the HBsAg is completely overlapped with the RT domain of the HBV polymerase, some mutations occurring in the RT region may lead to the emergence of escape mutants in the S region and vice versa. Thus, rtT128N, rtD134E, rtS202G, rtM204V, rtQ215S and rtF221Y substitutions observed in the RT region result in sP120T, sT126S, sS193L sI195M, sS207R and sL213I in the HBsAg gene, respectively. These mutations in addition to sT57I, sQ101H, sG130R, sY134F, and sL216stop could alter the antigenicity of HBsAg and reduce its expression and/or recognition by antibodies. Therefore, they could induce immune, vaccine, HBIG therapy, and/or diagnosis escape as well as influence HBsAg expression and treatment efficacy (Moerman et al., 2004; Sitnik et al., 2004; Bahramali et al., 2008; Amini-Bavil-Olyaee et al., 2010; Coppola, 2015; Ziaee et al., 2016; Rendon et al., 2017; Tokgöz et al., 2018; Aydın et al., 2019; Hosseini et al., 2019; Duda, 2020).

Regarding the mutational profile of BCP (nt 1,742–1,849), precore (nt 1,814–1,900), and core regions that code HBeAg and HBcAg proteins, the double mutants A1762T/G1764A, G1764T/C1766G and C1766T/T1768A, as well as the single mutations A1752G, T1753V (C/A/G), C1766T and C1788G, detected in BCP region, have been reported to enhance viral replication and/or reduce HBeAg synthesis by suppressing the transcription of the pre-C region (Parekh et al., 2003; Sendi et al., 2005; Poustchi et al., 2008; Tong et al., 2013; Caligiuri et al., 2016; Lazarevic et al., 2019). The single nucleotide mutations G1896A and G1899A in the precore region have been suggested to be mutational hotspots occurring most frequently in genotype D and were previously reported in Tunisian studies with an occurrence alone or in association (Triki et al., 2000; Bahri et al., 2006; Ayed et al., 2007; Poustchi et al., 2008; Ouneissa et al., 2012). These mutants result in a stop codon at position W28* and a substitution at position G29D, respectively, leading to the production of a truncated precore protein and then the abolition of HBeAg expression (Kobayashi et al., 2003; Thompson et al., 2010; Ducancelle et al., 2016). These variations are the major immune escape mutants of HBV as HBeAg is the main target for both cellular and humoral immune responses leading to a higher risk of liver HCC and LC progression (Tong et al., 2005; Liao et al., 2012; Suppiah et al., 2015; Pahal et al., 2016). In addition, precore mutants impose serious consequences on the treatment and enhance viral replication (Ouneissa et al., 2012; Kargar Kheirabad et al., 2017; Boyd et al., 2018).

Concerning the core mutations, cT67N within the T-helper CD4 epitope might be able to escape the host immune response (Datta et al., 2014; Saha et al., 2014). Moreover, cV74G, cP79Q, and cA80T mutations are known to reduce both HBe and HBc antigenicity (Pollicino et al., 2007; Huang et al., 2014). In addition, cA80T has resulted in the production of altered and truncated HBcAg protein leading potentially to abnormal immune reaction and negativity of anti-HBc (Bajpai et al., 2017).

In the last part of this study, we studied the mutational profile in the X gene. Substitutions xS43P and xP46S located in the B-cell epitope were detected in our study and have been suggested to be related with immune escape (Putri et al., 2019). Mutations xP46S, xA47T, xI88F, xA102V, xI127T, xK130M, xV131I, and xF132Y, were previously reported as significant HCC-related HBx mutants alone or combined such as (I127T + K130M + V131I) in patients 1, 3 and 5 and (xK130M + xV131I + xF132Y) in patient 3 (Pollicino et al., 2007; Ghosh et al., 2012; Ali et al., 2014; Al-Qahtani et al., 2017). Moreover, the double mutant xK130M + xV131I has been suggested to exacerbate the host’s immune response, increase viral replication, and lead to a truncated HBx protein (Wungu et al., 2019). In addition, it is associated with the activation of proto-oncogenes and inactivation of the tumor suppressor gene leading to a rapid progression of liver cirrhosis and/or HCC cell invasion and metastasis (Wang et al., 2016).

Several mutations previously reported to be significantly associated with an increased risk of severe liver disease progression to HCC and/or LC progression were also detected in other genes namely (rtD134E/rtF221Y/rtM204V/rtM309k) in the RT region; (sF22L) in the preS2 region; (sL49R, sL213I and sL216*) in the S region; (C1766T/T1768A double mutant, C1773T, C1799G, and C1766G) in the BCP region; and (cT12S/cE64D/cT67N/cA80T/cP130Q) in the core region of the C gene. These HCC-related mutations could be used as markers of HCC evolution in particular rtF221Y mutant which has been indicated as an independent risk factor for poor overall survival (Jammeh et al., 2008; Yin et al., 2011; Kitab et al., 2012; Zheng et al., 2012; Gopalakrishnan, 2013; Tong et al., 2013; Datta et al., 2014; Chaouch et al., 2016; Nishizawa et al., 2016; Kim et al., 2017; Li et al., 2017; Al-Qahtani et al., 2018; Choi et al., 2018; Hosseini et al., 2019). In contrast, the early development of G1757A in the BCP reduces the oncogenic potential of HBV suggesting that it might be a protective biomarker in chronic hepatitis B (Poustchi et al., 2008; Mohamadkhani et al., 2011; Ducancelle et al., 2013).

In addition to the commonly mentioned substitutions in all genes (P, S, C, and X), several nucleotide/amino acid substitutions have been detected in our patients (see Tables 2–5) but have never been reported previously or have been reported with unknown impact. Therefore, further studies are necessary to better understand and elucidate the effect of these mutations on HBV treatment, antigenicity, and disease evolution.

Conclusion

In conclusion, we would propose the whole genome sequencing as a pre-treatment diagnosis to predict potential resistance to available antiviral molecules, as well as to monitor the evolution of treatment response and prevent progression to cirrhosis or hepatocellular carcinoma. Thus, this could contribute to guiding national efforts to optimize relevant HBV treatment management in order to achieve the global hepatitis elimination goal by 2030.

Funding

This work was funded by the Tunisian Ministry of Higher Education [Programme d’Encouragement des Jeunes Chercheurs PEJC, 1ère Edition (2018; project code: 18PJEC07-09)], the Research Laboratory LR20IPT02: “Virus, Vectors and Hosts: One Health approach and technological innovation for a better health,” and the Clinical Investigation Center (CIC).

Publisher’s note

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.

References

• 1

  Abdel HamidM.SalamaA. (2018). X Gene variability and genotyping of hepatitis B virus (HBV) in sudanese patients with liver diseases, Khartoum, Sudan. Available at: https://www.researchgate.net/publication/325477946_X_GENE_VARIABILITY_AND_GENOTYPING_OF_HEPATITIS_B_VIRUS_HBV_IN_SUDANESE_PATIENTS_WITH_LIVER_DISEASES_KHARTOUM_SUDAN (Accessed October 22, 2020).

  • Google Scholar

• 2

  Al-QahtaniA. A.Al-AnaziM. R.NazirN.GhaiR.AbdoA. A.SanaiF. M.et al. (2017). Hepatitis B virus (HBV) X gene mutations and their association with liver disease progression in HBV-infected patients. Oncotarget8, 105115–105125.

  • Google Scholar

• 3

  AliA.Abdel-HafizH.SuhailM.Al-MarsA.ZakariaM. K.FatimaK.et al. (2014). Hepatitis B virus, HBx mutants and their role in hepatocellular carcinoma. World J. Gastroenterol.20, 10238–10248. doi: 10.3748/wjg.v20.i30.10238

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 4

  Al-QahtaniA. A.Al-AnaziM. R.NazirN.AbdoA. A.SanaiF. M.Al-HamoudiW. K.et al. (2018). The correlation between Hepatitis B virus Precore/Core mutations and the progression of severe liver disease. Front. Cell. Infect. Microbiol.8. doi: 10.3389/fcimb.2018.00355

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 5

  Amini-Bavil-OlyaeeS.HerbersU.MohebbiS. R.SabahiF.ZaliM. R.LueddeT.et al. (2009). Prevalence, viral replication efficiency and antiviral drug susceptibility of rtQ215 polymerase mutations within the hepatitis B virus genome. J. Hepatol.51, 647–654. doi: 10.1016/j.jhep.2009.04.022

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 6

  Amini-Bavil-OlyaeeS.VucurM.LueddeT.TrautweinC.TackeF. (2010). Differential Impact of Immune Escape Mutations G145R and P120T on the Replication of Lamivudine-Resistant Hepatitis B Virus e Antigen-Positive and -Negative Strains. J. Virol.84, 1026–1033. doi: 10.1128/JVI.01796-09

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 7

  AraujoN. M.Branco-VieiraM.SilvaA. C. M.PilottoJ. H.GrinsztejnB.de AlmeidaA. J.et al. (2008). Occult hepatitis B virus infection in HIV-infected patients: evaluation of biochemical, virological and molecular parameters. Hepatol. Res.38, 1194–1203. doi: 10.1111/j.1872-034X.2008.00392.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 8

  ArtariniA.Geby JessicaH.Rini KartikasariR.RianiC.Soefie RetnoningrumD. (2016). Detection of Hepatitis B virus X gene mutation from local clinical samples. Microbiol Indones.10, 9–14. doi: 10.5454/mi.10.1.2

  • CrossRef
  • Google Scholar

• 9

  AydınM.TekinS.SayanM.AkhanS. (2019). Molecular characterization of Hepatitis B virus strains isolated from chronic Hepatitis B patients in southeastern region of Turkey. Viral Hepat. J.25, 40–44. doi: 10.4274/vhd.galenos.2019.2019.0015

  • CrossRef
  • Google Scholar

• 10

  AyedK.GorgiY.Ayed-JendoubiS.AouadiH.SfarI.NajjarT.et al. (2007). Hepatitis B virus genotypes and precore/core-promoter mutations in Tunisian patients with chronic hepatitis B virus infection. J. Infect.54, 291–297. doi: 10.1016/j.jinf.2006.05.013

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 11

  AzarkarZ.ZiaeeM.EbrahimzadehA.SharifzadehG.JavanmardD. (2018). Epidemiology, risk factors, and molecular characterization of occult hepatitis B infection among anti-hepatitis B core antigen alone subjects. J. Med. Virol.91.

  • Google Scholar

• 12

  BahramaliG.SadeghizadehM.Amini-Bavil-OlyaeeS.AlvaianS. M.Behzad-BehbahaniA.AdeliA.et al. (2008). Clinical, virologic and phylogenetic features of hepatitis B infection in Iranian patients. World J. Gastroenterol.14, 5448–5453. doi: 10.3748/wjg.14.5448

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 13

  BahriO.CheikhI.HajjiN.DjebbiA.MaamouriN.SadraouiA.et al. (2006). Hepatitis B genotypes, precore and core promoter mutants circulating in Tunisia. J. Med. Virol.78, 353–357. doi: 10.1002/jmv.20554

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 14

  BajpaiV.GuptaE.KunduN.SharmaS.ShashtryS. M. (2017). Hepatitis B core antibody negativity in a chronic hepatitis b infected patient: report of an unusual serological pattern. J. Clin. Diagn. Res.11, DD04–DD06. doi: 10.7860/JCDR/2017/26821.10498

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 15

  BakhshizadehF.HekmatS.KeshvariM.AlavianS. M.MostafaviE.KeivaniH.et al. (2015). Efficacy of tenofovir disoproxil fumarate therapy in nucleoside-analogue naive Iranian patients treated for chronic hepatitis B. Hepat. Mon.15. doi: 10.5812/hepatmon.15(5)2015.25749

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 16

  BiswasA.PanigrahiR.ChandraP. K.BanerjeeA.DattaS.PalM.et al. (2013). Characterization of the occult hepatitis B virus variants circulating among the blood donors from eastern India. Sci. World J. doi: 10.1155/2013/212704

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 17

  BoydA.KouaméM. G.HoughtalingL.MohR.GabillardD.MaylinS.et al. (2019). Hepatitis B virus activity in untreated hepatitis B e antigen-negative human immunodeficiency virus-hepatitis B virus co-infected patients from sub-Saharan Africa. Trans. R. Soc. Trop. Med. Hyg.113, 437–445.

  • Google Scholar

• 18

  BoydA.MohR.MaylinS.Abdou ChekaraouM.MahjoubN.GabillardD.et al. (2018). Precore G1896A mutation is associated with reduced rates of HBsAg seroclearance in treated HIV hepatitis B virus co-infected patients from Western Africa. J. Viral Hepat.25, 1121–1131. doi: 10.1111/jvh.12914

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 19

  CaligiuriP.CerrutiR.IcardiG.BruzzoneB. (2016). Overview of hepatitis B virus mutations and their implications in the management of infection. World J. Gastroenterol.22, 145–154. doi: 10.3748/wjg.v22.i1.145

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 20

  ChaouchH.TaffonS.VillanoU.EquestreM.BruniR.BelhadjM.et al. (2016). Naturally occurring surface antigen variants of Hepatitis B virus in Tunisian patients. Intervirology59, 36–47. doi: 10.1159/000445894

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 21

  ChaudhuriV.TayalR.NayakB.AcharyaS. K.PandaS. K. (2004). Occult hepatitis B virus infection in chronic liver disease: full-length genome and analysis of mutant surface promoter. Gastroenterology127, 1356–1371. doi: 10.1053/j.gastro.2004.08.003

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 22

  ChekaraouM. A.BrichlerS.MansourW.GalF.GarbaA.DényP.et al. (2010). A novel hepatitis B virus (HBV) subgenotype D (D8) strain, resulting from recombination between genotypes D and E, is circulating in Niger along with HBV/E strains. J. Gen. Virol.91, 1609–1620. doi: 10.1099/vir.0.018127-0

  • CrossRef
  • Google Scholar

• 23

  ChenC. H.LeeC. M.LuS. N.ChangchienC. S.EngH. L.HuangC. M.et al. (2005). Clinical significance of hepatitis B virus (HBV) genotypes and precore and core promoter mutations affecting HBV e antigen expression in Taiwan. J. Clin. Microbiol.43, 6000–6006. doi: 10.1128/JCM.43.12.6000-6006.2005

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 24

  ChogaW. T.AndersonM.ZumbikaE.PhiniusB. B.MbangiwaT.BhebheL. N.et al. (2020). In Silico prediction of human leukocytes antigen (HLA) class II binding Hepatitis B virus (HBV) peptides in Botswana. Viruses12:731. doi: 10.3390/v12070731

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 25

  ChoiY. M.LeeS. Y.KimB. J. (2018). Naturally occurring hepatitis B virus reverse transcriptase mutations related to potential antiviral drug resistance and liver disease progression. World J. Gastroenterol.24, 1708–1724. doi: 10.3748/wjg.v24.i16.1708

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 26

  CiancioA.SmedileA.RizzettoM.LaggetM.GerinJ.KorbaB. (2004). Identification of HBV DNA Sequences That Are Predictive of Response to Lamivudine Therapy. Hepatology39, 64–73. doi: 10.1002/hep.20019

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 27

  CiftciS.KeskinF.CakirisA.AkyuzF.PinarbasiB.AbaciN.et al. (2014). Analysis of potential antiviral resistance mutation profiles within the HBV reverse transcriptase in untreated chronic hepatitis B patients using an ultra-deep pyrosequencing method. Diagn. Microbiol. Infect. Dis.79, 25–30. doi: 10.1016/j.diagmicrobio.2014.01.005

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 28

  ColagrossiL.HermansL. E.SalpiniR.Di CarloD.PasS. D.AlvarezM.et al. (2018). Immune-escape mutations and stop-codons in HBsAg develop in a large proportion of patients with chronic HBV infection exposed to anti-HBV drugs in Europe. BMC Infect. Dis.18:251. doi: 10.1186/s12879-018-3161-2

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 29

  CoppolaN. (2015). Clinical significance of hepatitis B surface antigen mutants. World J. Hepatol.7:2729. doi: 10.4254/wjh.v7.i27.2729

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 30

  DattaS.GhoshA.DasguptaD.GhoshA.RoychoudhuryS.RoyG.et al. (2014). Novel point and combo-mutations in the genome of hepatitis B virus-genotype D: characterization and impact on liver disease progression to hepatocellular carcinoma. PLoS One9. doi: 10.1371/journal.pone.0110012

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 31

  DelaneyW. E.LocarniniS.ShawT. (2001). Resistance of hepatitis B virus to antiviral drugs: Current aspects and directions for future investigation. Antivir. Chem. Chemother.12, 1–35. doi: 10.1177/095632020101200101

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 32

  DucancelleA.AbgueguenP.BirguelJ.MansourW.PivertA.le Guillou-GuillemetteH.et al. (2013). High Endemicity and Low Molecular Diversity of Hepatitis B Virus Infections in Pregnant Women in a Rural District of North Cameroon. PLoS One8:80346

  • Google Scholar

• 33

  DucancelleA.PivertA.BertraisS.BoursierJ.BalanV.VeillonP.et al. (2016). Different precore/core mutations of hepatitis B interact with, limit, or favor liver fibrosis severity. J. Gastroenterol. Hepatol.31, 1750–1756. doi: 10.1111/jgh.13338

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 34

  DudaA. (2020). Influence de la variabilité des glycoprotéines d’enveloppe du virus de l’hépatite B sur la clairance de l’Ag HBs chez des patients co-infectés par le VIH suivis au CHU de Nancy. Available at: http://www.culture.gouv.fr/culture/infos-pratiques/droits/protection.htm (Accessed September 29, 2020).

  • Google Scholar

• 35

  ElkadyA.TanakaY.KurbanovF.OynsurenT.MizokamiM. (2008). Virological and clinical implication of core promoter C1752/V1753 and T1764/G1766 mutations in hepatitis B virus genotype D infection in Mongolia. J. Gastroenterol. Hepatol.23, 474–481. doi: 10.1111/j.1440-1746.2008.05321.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 36

  EzzikouriS.CheminI.ChafikA.WakrimL.NourlilJ.MalkiA.et al. (2008). Genotype determination in Moroccan hepatitis B chronic carriers. Infect. Genet. Evol.8, 306–312. doi: 10.1016/j.meegid.2008.01.010

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 37

  FangZ. L.SabinC. A.DongB. Q.GeL. Y.WeiS. C.ChenQ. Y.et al. (2008). HBV A1762T, G1764A mutations are a valuable biomarker for identifying a subset of male HBsAG carriers at extremely high risk of hepatocellular carcinoma: a prospective study. Am. J. Gastroenterol.103, 2254–2262. doi: 10.1111/j.1572-0241.2008.01974.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 38

  FeeneyS. A.MccaugheyC.WattA. P.AgnafM. R. E.McdougallN.WendU. C.et al. (2013). Reactivation of occult hepatitis B virus infection following cytotoxic lymphoma therapy in an anti-HBc negative patient. J. Med. Virol.85, 597–601. doi: 10.1002/jmv.23513

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 39

  GermerJ. J.CharltonM. R.IshitaniM.ForehandC. D.PatelR. (2003). Characterization of Hepatitis B Virus Surface Antigen and Polymerase Mutations in Liver Transplant Recipients Pre- and Post-Transplant. Am. J. Transplant.3, 743–753. doi: 10.1034/j.1600-6143.2003.00149.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 40

  GhoshS.MondalR. K.BanerjeeP.NandiM.SarkarS.DasK.et al. (2012). Tracking the naturally occurring mutations across the full-length genome of hepatitis B virus of genotype D in different phases of chronic e-antigen-negative infection. Clin. Microbiol. Infect.18, E412–E418. doi: 10.1111/j.1469-0691.2012.03975.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 41

  Gil-GarcíaA. I.MadejónA.Francisco-RecueroI.López-LópezA.VillafrancaE.RomeroM.et al. (2019). Prevalence of hepatocarcinoma-related hepatitis B virus mutants in patients in grey zone of treatment. World J. Gastroenterol.25, 5883–5896. doi: 10.3748/wjg.v25.i38.5883

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 42

  GopalakrishnanD. (2013). Hepatitis B virus subgenotype A1 predominates in liver disease patients from Kerala, India. World J. Gastroenterol.19:9294. doi: 10.3748/wjg.v19.i48.9294

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 43

  GüntherS.von BreunigF.SantantonioT.JungM. C.GaetaG. B.FischerL.et al. (1999). Absence of mutations in the YMDD motif/B region of the hepatitis B virus polymerase in famciclovir therapy failure. J. Hepatol.30, 749–754. doi: 10.1016/S0168-8278(99)80124-X

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 44

  HeX.WangF.HuangB.ChenP.ZhongL. (2015). Detection and analysis of resistance mutations of hepatitis B virus. Int. J. Clin. Exp. Med.8, 9630–9639. PMID:

  • Pubmed Abstract
  • Google Scholar

• 45

  HomsM.ButiM.TaberneroD.QuerJ.SanchezA.CorralN.et al. (2012). Quasispecies dynamics in main core epitopes of hepatitis B virus by ultra-deep-pyrosequencing. World J. Gastroenterol.18, 6096–6105. doi: 10.3748/wjg.v18.i42.6096

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 46

  HomsM.JardiR.ButiM.SchaperM.TaberneroD.Fernandez-FernandezP.et al. (2011). HBV core region variability: effect of antiviral treatments on main epitopic regions. Antivir. Ther.16, 37–49. doi: 10.3851/IMP1701

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 47

  HorikitaM.ItohS.YamamotoK.ShibayamaT.TsudaF.OkamotoH. (1994). Differences in the entire nucleotide sequence between hepatitis B virus genomes from carriers positive for antibody to hepatitis B e antigen with and without active disease. J. Med. Virol.44, 96–103. doi: 10.1002/jmv.1890440118

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 48

  HosseiniS. Y.SanaeiN.FattahiM. R.Malek-HosseiniS. A.SarvariJ. (2019). Association of HBsAg mutation patterns with hepatitis B infection outcome: Asymptomatic carriers versus HCC/cirrhotic patients. Ann. Hepatol.18, 640–645. doi: 10.1016/j.aohep.2018.12.006

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 49

  HuangY.TongS.TaiA. W.HussainM.LokA. S. F. (2011). Hepatitis B virus core promoter mutations contribute to hepatocarcinogenesis by deregulating SKP2 and its target, p21. Gastroenterology141, 1412–1421.e5. doi: 10.1053/j.gastro.2011.06.048

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 50

  HuangF. Y.WongD. K. H.SetoW. K.ZhangA. Y.LeeC. K.LinC. K.et al. (2014). Sequence variations of full-length hepatitis B virus genomes in Chinese patients with HBsAg-negative hepatitis B infection. PLoS One9. doi: 10.1371/journal.pone.0115743

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 51

  IsmailA. M.SamuelP.EapenC. E.KannangaiR.AbrahamP. (2011). Antiviral resistance mutations and genotype-associated amino acid substitutions in treatment-Naïve Hepatitis B virus-infected individuals from the Indian subcontinent. Intervirology55, 36–44.

  • Google Scholar

• 52

  JammehS.TavnerF.WatsonR.ThomasH. C.KarayiannisP. (2008). Effect of basal core promoter and pre-core mutations on hepatitis B virus replication. J. Gen. Virol.89, 901–909. doi: 10.1099/vir.0.83468-0

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 53

  JavanmardD.NiyaM. H. K.KalafkhanyD.NajafiM.ZiaeeM.BabaeiM. R.et al. (2020). Downregulation of gsk3β and upregulation of urg7 in hepatitis b-related hepatocellular carcinoma. Hepat. Mon.20, 1–11.

  • Google Scholar

• 54

  KamiyaN. (2003). The mechanisms of action of antivirals against hepatitis B virus infection. J. Antimicrob. Chemother.51, 1085–1089. doi: 10.1093/jac/dkg236

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 55

  KaratayliE.KaratayliS. C.CinarK.GokahmetogluS.GüvenK.IdilmanR.et al. (2012). Molecular characterization of a novel entecavir mutation pattern isolated from a multi-drug refractory patient with chronic hepatitis B infection. J. Clin. Virol.53, 130–134. doi: 10.1016/j.jcv.2011.10.011

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 56

  Kargar KheirabadA.FarshidfarG.NasrollaheianS.GouklaniH. (2017). Prevalence and Characteristics of Precore Mutation in Iran and Its Correlation with Genotypes of Hepatitis B. Electron. Physician9, 4114–4123. doi: 10.19082/4114

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 57

  KatsoulidouA.ParaskevisD.MagiorkinisE.MoschidisZ.HaidaC.HatzitheodorouE.et al. (2009). Molecular characterization of occult hepatitis B cases in Greek blood donors. J. Med. Virol.81, 815–825. doi: 10.1002/jmv.21499

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 58

  KimJ. H.JungY. K.JooM. K.KimJ. H.YimH. J.ParkJ. J.et al. (2010). Hepatitis B viral surface mutations in patients with adefovir resistant chronic hepatitis B with A181T/V polymerase mutations. J. Korean Med. Sci.25, 257–264. doi: 10.3346/jkms.2010.25.2.257

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 59

  KimH.LeeS. A.KimB. J. (2016). X region mutations of hepatitis B virus related to clinical severity. World J. Gastroenterol.22, 5467–5478. doi: 10.3748/wjg.v22.i24.5467

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 60

  KimJ. E.LeeS. Y.KimH.KimK. J.ChoeW. H.KimB. J. (2017). Naturally occurring mutations in the reverse transcriptase region of hepatitis B virus polymerase from treatment-naïve Korean patients infected with genotype C2. World J. Gastroenterol.23, 4222–4232. doi: 10.3748/wjg.v23.i23.4222

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 61

  KimH.LeeS. A.KimD. W.LeeS. H.KimB. J. (2013). Naturally occurring mutations in large surface genes related to occult Infection of Hepatitis B virus Genotype C. Blackard J, editor. PLoS One8:e54486. doi: 10.1371/journal.pone.0084194

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 62

  KitabB.Essaid El FeydiA.AfifiR.TrepoC.BenazzouzM.EssamriW.et al. Variability in the Precore and Core Promoter Regions of HBV Strains in Morocco: Characterization and Impact on Liver Disease Progression. RayR. editor. PLoS One. (2012). 7:e42891, doi: 10.1371/journal.pone.0042891.

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 63

  KobayashiM.AraseY.IkedaK.TsubotaA.SuzukiY.SaitohS.et al. (2003). Precore wild-type hepatitis B virus with G1896 in the resolution of persistent hepatitis B virus infection. Intervirology46, 157–163. doi: 10.1159/000071456

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 64

  KumarS.StecherG.TamuraK.DudleyJ. (2016). MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets Downloaded from. Mol. Biol. Evol.33, 1870–1874. doi: 10.1093/molbev/msw054

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 65

  KweiK.TangX.LokA. S.SureauC.GarciaT.LiJ.et al. (2013). Impaired Virion secretion by Hepatitis B virus immune escape mutants and its rescue by wild-type envelope proteins or a second-site mutation. J. Virol.87, 2352–2357. doi: 10.1128/JVI.02701-12

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 66

  LagoB. V.MelloF. C.RibasF. S.ValenteF.SoaresC. C.NielC.et al. Analysis of Complete Nucleotide Sequences of Angolan Hepatitis B Virus Isolates Reveals the Existence of a Separate Lineage within Genotype E. CheminI. A., editor. PLoS One. (2014). 9:e92223, doi: 10.1371/journal.pone.0092223.

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 67

  LazarevicI.BankoA.MiljanovicD.CupicM. (2019). Immune-escape hepatitis B virus mutations associated with viral reactivation upon immunosuppression. Viruses11:778. doi: 10.3390/v11090778

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 68

  LengX. H.ChenE. Q.DuL. Y.BaiL.GongD. Y.ChengX.et al. (2015). Biological characteristics of the A1762T/G1764A mutant strain of hepatitis B virus in vivo. Mol. Med. Rep.12, 5141–5148. doi: 10.3892/mmr.2015.4072

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 69

  LeónB.TaylorL.VargasM.LuftigR. B.AlbertazziF.HerreroL.et al. (2005). HBx M130K and V131I (T-A) mutations in HBV genotype F during a follow-up study in chronic carriers. Virol. J.4:60

  • Google Scholar

• 70

  LiM. W.HouW.WoJ. E.LiuK. Z. (2005). Character of HBV (hepatitis B virus) polymerase gene rtM204V/I and rtL180M mutation in patients with lamivudine resistance. J. Zhejiang Univ. Sci.6, 664–667. doi: 10.1631/jzus.2005.B0664

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 71

  LiH.JiaJ.WangM.WangH.GuX.FangM.et al. (2017). F221Y mutation in hepatitis B virus reverse transcriptase is associated with hepatocellular carcinoma prognosis following liver resection. Mol. Med. Rep.15, 3292–3300. doi: 10.3892/mmr.2017.6362

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 72

  LiX. Y.LiangC. H.ParkmanV.LvZ. T. (2018). The association between TNF-a 238A/G and 308A/G polymorphisms and juvenile idiopathic arthritis: An updated PRISMA-compliant meta-analysis. Medicine97. doi: 10.1097/MD.0000000000013964

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 73

  LiX. G.LiuB. M.XuJ.LiuX. E.DingH.LiT. (2012). Discrepancy of potential antiviral resistance mutation profiles within the HBV reverse transcriptase between nucleos(t)ide analogue-untreated and -treated patients with chronic hepatitis B in a hospital in China. J. Med. Virol.84, 207–216. doi: 10.1002/jmv.23182

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 74

  LiaoY.HuX.ChenJ.CaiB.TangJ.YingB.et al. (2012). Precore mutation of hepatitis B virus may contribute to hepatocellular carcinoma risk: evidence from an updated meta-analysis. PLoS One7. doi: 10.1371/journal.pone.0038394

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 75

  LinX.XuX.HuangQ. L.LiuY. Q.ZhengD. L.ChenW. N.et al. (2005). Biological impacts of “hot-spot” mutations of hepatitis B virus X proteins are genotype B and C differentiated. World J. Gastroenterol.11, 4703–4708. doi: 10.3748/wjg.v11.i30.4703

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 76

  LiuB. M.LiT.XuJ.LiX. G.DongJ. P.YanP.et al. (2010). Characterization of potential antiviral resistance mutations in hepatitis B virus reverse transcriptase sequences in treatment-naïve Chinese patients. Antiviral Res.85, 512–519. doi: 10.1016/j.antiviral.2009.12.006

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 77

  LiuS.ZhangH.GuC.YinJ.HeY.XieJ.et al. (2009). Associations between hepatitis B virus mutations and the risk of hepatocellular carcinoma: a meta-analysis. J. Natl. Cancer Inst.101, 1066–1082.

  • Google Scholar

• 78

  LocarniniS.MasonW. S. (2006). Cellular and virological mechanisms of HBV drug resistance. J. Hepatol.44, 422–431. doi: 10.1016/j.jhep.2005.11.036

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 79

  LocarniniS.McMillanJ.BartholomeuszA. (2003). The hepatitis B virus and common mutants. Semin. Liver Dis.23, 5–20.

  • Google Scholar

• 80

  MahabadiM.NorouziM.AlavianS. M.SamimiradK.AzadT. M.SaberfarE.et al. (2013). Drug-related mutational patterns in hepatitis B virus (HBV) reverse transcriptase proteins from Iranian treatment-naïve chronic HBV patients. Hepat. Mon.13. doi: 10.5812/hepatmon.6712

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 81

  ManiM.VijayaraghavanS.SaranganG.BaraniR.AbrahamP.SrikanthP. (2019). Hepatitis B virus X protein: the X factor in chronic hepatitis B virus disease progression. Indian J. Med. Microbiol. doi: 10.4103/ijmm.IJMM_19_421

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 82

  MeldalB. H. M.MoulaN. M.BarnesI. H. A.BoukefK.AllainJ. P. (2009). A novel hepatitis B virus subgenotype, D7, in Tunisian blood donors. J. Gen. Virol.90, 1622–1628. doi: 10.1099/vir.0.009738-0

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 83

  MoermanB.MoonsV.SommerH.SchmittY.StetterM. (2004). Evaluation of sensitivity for wild type and mutant forms of hepatitis B surface antigen by four commercial HBsAg assays. Clin. Lab.50, 159–162. PMID:

  • Pubmed Abstract
  • Google Scholar

• 84

  MohamadkhaniA.MontazeriG.PoustchiH. (2011). The importance of hepatitis B virus genome diversity in basal core promoter region. Middle East J. Dig. Dis.3, 13–19. PMID:

  • Pubmed Abstract
  • Google Scholar

• 85

  MokayaJ.MapongaT. G.McNaughtonA. L.van SchalkwykM.HugoS.SingerJ. B.et al. (2020). Evidence of tenofovir resistance in chronic hepatitis B virus (HBV) infection: An observational case series of South African adults. J. Clin. Virol.129. doi: 10.1016/j.jcv.2020.104548

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 86

  MokayaJ.McNaughtonA.BesterP.GoedhalsD.BarnesE.MarsdenB.et al. (2019). Hepatitis B virus resistance to tenofovir: fact or fiction? A synthesis of the evidence to date. medRxiv [Preprint].

  • Google Scholar

• 87

  MokayaJ.McnaughtonA. L.BesterP. A.GoedhalsD.BarnesE.MarsdenB. D.et al. (2020). Hepatitis B virus resistance to tenofovir: fact or fiction? A systematic literature review and structural analysis of drug resistance mechanisms [version 1; peer review: awaiting peer review].

  • Google Scholar

• 88

  MondalR. K.KhatunM.GhoshS.BanerjeeP.DattaS.SarkarS.et al. (2015). Immune-driven adaptation of hepatitis B virus genotype D involves preferential alteration in B-cellepitopes and replicative attenuation-an insight from human immunodeficiency virus/hepatitis B virus coinfection. Clin. Microbiol. Infect.21, 710.e11–710.e20. doi: 10.1016/j.cmi.2015.03.004

  • CrossRef
  • Google Scholar

• 89

  MoriconiF.ColombattoP.CocoB.CiccorossiP.OliveriF.FlichmanD.et al. (2007). Emergence of hepatitis B virus quasispecies with lower susceptibility to nucleos(t)ide analogues during lamivudine treatment. J. Antimicrob. Chemother.60, 341–349. doi: 10.1093/jac/dkm187

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 90

  MukaideM.TanakaY.Shin-IT.YuenM. F.KurbanovF.YokosukaO.et al. (2010). Mechanism of entecavir resistance of hepatitis B virus with viral breakthrough as determined by long-term clinical assessment and molecular docking simulation. Antimicrob. Agents Chemother.54, 882–889. doi: 10.1128/AAC.01061-09

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 91

  NgL. F. P.ChanM.ChanS. H.ChengP. C. P.LeungE. H. C.ChenW. N.et al. (2005). Host heterogeneous ribonucleoprotein K (hnRNP K) as a potential target to suppress hepatitis B virus replication. PLoS Med.2, 0673–0683.

  • Google Scholar

• 92

  NishizawaT.HoshinoT.NaganumaA.KobayashiT.NagashimaS.TakahashiM.et al. (2016). Enhanced pregenomic RNA levels and lowered precore mRNA transcription efficiency in a genotype A hepatitis B virus genome with C1766T and T1768A mutations obtained from a fulminant hepatitis patient. J. Gen. Virol.97, 2643–2656. doi: 10.1099/jgv.0.000566

  • CrossRef
  • Google Scholar

• 93

  OlingerC. M.WeberB.OtegbayoJ. A.AmmerlaanW.Van Der Taelem-BruléN.MullerC. P. (2007). Hepatitis B virus genotype E surface antigen detection with different immunoassays and diagnostic impact of mutations in the preS/S gene. Med. Microbiol. Immunol.196, 247–252. doi: 10.1007/s00430-007-0050-5

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 94

  OlusolaB. A.FaneyeA. O.OluwasemowoO. O.MotayoB. O.AdebayoS.Oludiran-AyoadeA. E.et al. (2021). Profiles of mutations in hepatitis B virus surface and polymerase genes isolated from treatment-naïve Nigerians infected with genotype E. J. Med. Microbiol.70. doi: 10.1099/jmm.0.001338

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 95

  OuneissaR.BahriO.Alaya-BouafifN.Ben ChouaiebS.YahiaA.Ben SadraouiA.et al. (2012). Frequency and clinical significance of core promoter and precore region mutations in Tunisian patients infected chronically with hepatitis B. J. Med. Virol.84, 1719–1726. doi: 10.1002/jmv.23394

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 96

  OuneissaR.BahriO.Ben YahiaA.TouziH.Msaddak AzouzM.Ben MamiN.et al. (2013). Evaluation of PCR-RFLP in the Pre-S Region as Molecular Method for Hepatitis B Virus Genotyping. Hepat. Mon.13. doi: 10.5812/hepatmon.11781

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 97

  PahalV.SinghJ.DadhichK. S. (2016). Hepatitis B virus core promoter and precore mutations and their relatedness to genotypes and disease pathogenesis. Pelagia Res. Lib. Adv. Appl. Sci. Res.7, 70–80.

  • Google Scholar

• 98

  ParekhS.ZoulimF.AhnS. H.TsaiA.LiJ.KawaiS.et al. (2003). Genome replication, Virion secretion, and e antigen expression of naturally occurring Hepatitis B virus Core promoter mutants. J. Virol.77, 6601–6612. doi: 10.1128/JVI.77.12.6601-6612.2003

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 99

  ParkE. S.LeeA. R.KimD. H.LeeJ. H.YooJ. J.AhnS. H.et al. (2019). Identification of a quadruple mutation that confers tenofovir resistance in chronic hepatitis B patients. J. Hepatol.70, 1093–1102. doi: 10.1016/j.jhep.2019.02.006

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 100

  PollicinoT.IsgròG.di StefanoR.FerraroD.MaimoneS.BrancatelliS.et al. (2009). Variability of reverse transcriptase and overlapping S gene in hepatitis B virus isolates from untreated and lamivudine-resistant chronic hepatitis B patients. Antivir. Ther.14, 649–654. doi: 10.1177/135965350901400504

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 101

  PollicinoT.RaffaG.CostantinoL.LisaA.CampelloC.SquadritoG.et al. (2007). Molecular and functional analysis of occult hepatitis B virus isolates from patients with hepatocellular carcinoma. Hepatology45, 277–285. doi: 10.1002/hep.21529

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 102

  PourkarimM. R.VergoteV.Amini-Bavil-OlyaeeS.SharifiZ.SijmonsS.LemeyP.et al. (2014). Molecular characterization of hepatitis B virus (HBV) strains circulating in the northern coast of the Persian Gulf and its comparison with worldwide distribution of HBV subgenotype D1. J. Med. Virol.86, 745–757. doi: 10.1002/jmv.23864

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 103

  PoustchiH.MohamadkhaniA.BowdenS.MontazeriG.AyresA.RevillP.et al. (2008). Clinical significance of precore and core promoter mutations in genotype D hepatitis B-related chronic liver disease. J. Viral Hepat.15, 753–760. doi: 10.1111/j.1365-2893.2008.00998.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 104

  PurdyM. A.TalekarG.SwensonP.AraujoA.FieldsH. (2006). A new algorithm for deduction of hepatitis B surface antigen subtype determinants from the amino acid sequence. Intervirology50, 45–51.

  • Google Scholar

• 105

  PutriW. A.YanoY.YamaniL. N.LusidaM. I.SoetjiptoL. Y.et al. (2019). Association between HBx variations and development of severe liver disease among indonesian patients. Kobe J. Med. Sci.65, 28–35.

  • Google Scholar

• 106

  QinB.PeiR. J.HeT. T.HuangZ. H.PanG. S.TuC. Y.et al. (2013a). Polymerase mutations rtN238R, rtT240Y and rtN248H of hepatitis B virus decrease susceptibility to adefovir. Chin. Sci. Bull.58, 1760–1766. doi: 10.1007/s11434-013-5770-x

  • CrossRef
  • Google Scholar

• 107

  QinB.ZhangB.ZhangX.HeT.XuW.FuL.et al. (2013b). Substitution Rtq267h of hepatitis B virus increases the weight of replication and lamivudine resistance. Hepat. Mon.13:12160. doi: 10.5812/hepatmon.12160

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 108

  QuarleriJ. (2014). Core promoter: a critical region where the hepatitis B virus makes decisions. World J. Gastroenterol.20, 425–435. doi: 10.3748/wjg.v20.i2.425

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 109

  Quiros-RoldanE.CalabresiA.LapadulaG.TirelliV.CostarelliS.CologniG.et al. (2008). Evidence of long-term suppression of hepatitis B virus DNA by tenofovir as rescue treatment in patients coinfected by HIV. Antivir. Ther.13, 341–348. doi: 10.1177/135965350801300315

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 110

  RajoriyaN.CombetC.ZoulimF.JanssenH. L. A. (2017). How viral genetic variants and genotypes influence disease and treatment outcome of chronic hepatitis B. Time for an individualised approach?J. Hepatol.67, 1281–1297. doi: 10.1016/j.jhep.2017.07.011

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 111

  RendonJ. C.Cortes-ManceraF.Restrepo-GutierrezJ. C.HoyosS. (2017). Navas MC. Molecular characterization of occult hepatitis B virus infection in patients with end-stage liver disease in Colombia. PLoS One12:e0180447. doi: 10.1371/journal.pone.0180447

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 112

  SahaD.PalA.BiswasA.PanigrahiR.SarkarN.DasD.et al. (2014). Molecular characterization of HBV strains circulating among the treatment-naive HIV/HBV co-infected patients of eastern India. PLoS One9. doi: 10.1371/journal.pone.0090432

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 113

  SalarneiaF.ZhandS.KhodabakhshiB.TabarraeiA.VakiliM. A.JavidN.et al. (2016). Mutations at nucleotide 1762, 1764 and 1766 of Hepatitis B virus X gene in patients with chronic Hepatitis B and Hepatitis B-related cirrhosis. Med. Lab. J10, 31–35.

  • Google Scholar

• 114

  SchildgenO. (2007). Novel lamivudine resistance [3]. Antimicrob. Agents Chemother.51:4533. doi: 10.1128/AAC.00840-07

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 115

  SendiH.Mehrab-MohseniM.ZaliM. R.NorderH.MagniusL. O. (2005). T1764G1766 core promoter double mutants are restricted to Hepatitis B virus strains with an A1757 and are common in genotype D. J. Gen. Virol.86, 2451–2458. doi: 10.1099/vir.0.81023-0

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 116

  ShawT.BartholomeuszA.LocarniniS. (2006). HBV drug resistance: mechanisms, detection and interpretation. J. Hepatol.44, 593–606. doi: 10.1016/j.jhep.2006.01.001

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 117

  SheldonJ. (2008). Soriano V. Hepatitis B virus escape mutants induced by antiviral therapy. J. Antimicrob. Chemother.61, 766–768. doi: 10.1093/jac/dkn014

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 118

  SheldonJ.RodèsB.ZoulimF.BartholomeuszA.SorianoV. (2006). Mutations affecting the replication capacity of the hepatitis B virus [Internet]. J. Viral Hepat.13, 427–434. doi: 10.1111/j.1365-2893.2005.00713.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 119

  SitnikR.PinhoJ. R. R.BertoliniD. A.BernardiniA. P.da SilvaL. C.CarrilhoF. J. (2004). Hepatitis B virus genotypes and precore and core mutants in Brazilian patients. J. Clin. Microbiol.42, 2455–2460. doi: 10.1128/JCM.42.6.2455-2460.2004

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 120

  SominskayaI.MihailovaM.JansonsJ.LegzdinaD.SudmaleG.PumpensP.et al. (2011). Hepatitis B virus genotypes in Latvia. Open Hepatol. J.3. doi: 10.2174/1876517301103010007

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 121

  SunbulM. (2014). Hepatitis B virus genotypes: global distribution and clinical importance. World J. Gastroenterol.20:5427. doi: 10.3748/wjg.v20.i18.5427

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 122

  SunturB. M.SayanM.KayaH.ÜnalN. (2019). Overlapping Pol/S gene analysis in chronic hepatitis B patients with coexisting Hbsag and Anti-HBs. Acta Medica Mediterranea.35:3113

  • Google Scholar

• 123

  SuppiahJ.ZainR. M.BahariN.NawiS. H.SaatZ. (2015). G1896A precore mutation and association with HBeAG status, genotype and clinical status in patients with chronic hepatitis B. Hepat. Mon.15. doi: 10.5812/hepatmon.31490

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 124

  TaghiabadiM.HosseiniS. Y.GorzinA. A.TaghaviS. A.MonavariS. H. R.SarvariJ. (2019). Comparison of pre-S1/S2 variations of hepatitis B virus between asymptomatic carriers and cirrhotic/hepatocellular carcinoma-affected individuals. Clin Exp Hepatol.5, 161–168. doi: 10.5114/ceh.2019.84781

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 125

  TenneyD. J.LevineS. M.RoseR. E.WalshA. W.WeinheimerS. P.DiscottoL.et al. (2004). Clinical emergence of entecavir-resistant hepatitis B virus requires additional substitutions in virus already resistant to lamivudine. Antimicrob. Agents Chemother.48, 3498–3507. doi: 10.1128/AAC.48.9.3498-3507.2004

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 126

  ThompsonA. J. V.NguyenT.IserD.AyresA.JacksonK.LittlejohnM.et al. (2010). Serum hepatitis B surface antigen and hepatitis B e antigen titers: Disease phase influences correlation with viral load and intrahepatic hepatitis B virus markers. Hepatology51, 1933–1944. doi: 10.1002/hep.23571

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 127

  TokgözY.TerlemezS.SayanM.KırdarS. (2018). Investigation of antiviral resistance and escape mutations in children with naive chronic hepatitis B patients and their parents. Turk. J. Pediatr.60, 514–519. doi: 10.24953/turkjped.2018.05.007

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 128

  TongM. J.BlattL. M.KaoJ. H.ChengJ. T.CoreyW. G. (2007). Basal core promoter T1762/A1764 and precore A1896 gene mutations in hepatitis B surface antigen-positive hepatocellular carcinoma: a comparison with chronic carriers. Liver Int.27, 1356–1363. doi: 10.1111/j.1478-3231.2007.01585.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 129

  TongS.KimK. H.ChanteC.WandsJ.LiJ. (2005). Hepatitis B virus e antigen variants. Int. J. Med. Sci.2, 2–7. doi: 10.7150/ijms.2.2

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 130

  TongS.LiJ.WandsJ. R.WenY. (2013). Hepatitis B virus genetic variants: biological properties and clinical implications. Emerg. Microbes Infect.2, 1–11.

  • Google Scholar

• 131

  TorresiJ.Earnest-SilveiraL.CiviticoG.WaltersT. E.LewinS. R.FyfeJ.et al. (2002a). Restoration of replication phenotype of lamivudine-resistant hepatitis B virus mutants by compensatory changes in the “fingers” subdomain of the viral polymerase selected as a consequence of mutations in the overlapping S gene. Virology299, 88–99. doi: 10.1006/viro.2002.1448

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 132

  TorresiJ.Earnest-SilveiraL.DeliyannisG.EdgttonK.ZhuangH.LocarniniS. A.et al. (2002b). Reduced antigenicity of the Hepatitis B virus HBsAg protein arising as a consequence of sequence changes in the overlapping polymerase gene that are selected by lamivudine therapy. Virology293, 305–313. doi: 10.1006/viro.2001.1246

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 133

  TrikiH.Ben SlimaneS.Ben MamiN.SakkaT.Ben AmmarA.DellagiK. (2000). High circulation of hepatitis B virus (HBV) precore mutants in Tunisia, North Africa. Epidemiol. Infect.125, 169–174. doi: 10.1017/S0950268899003921

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 134

  VilletS.OllivetA.PichoudC.BarraudL.VilleneuveJ. P.TrépoC.et al. (2007). Stepwise process for the development of entecavir resistance in a chronic hepatitis B virus infected patient. J. Hepatol.46, 531–538. doi: 10.1016/j.jhep.2006.11.016

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 135

  WangL.HanF.DuanH.JiF.YanX.FanY.et al. (2017). Hepatitis B virus pre-existing drug resistant mutation is related to the genotype and disease progression. J. Infect. Dev. Ctries.11, 727–732. doi: 10.3855/jidc.9021

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 136

  WangY.ZengL.ChenW. (2016). HBV X gene point mutations are associated with the risk of hepatocellular carcinoma: A systematic review and meta-analysis. Mol. Clin. Oncol.4, 1045–1051. doi: 10.3892/mco.2016.847

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 137

  WangQ.ZhangT.YeL.WangW.ZhangX. (2012). Analysis of hepatitis B virus X gene (HBx) mutants in tissues of patients suffered from hepatocellular carcinoma in China. Cancer Epidemiol.36, 369–374. doi: 10.1016/j.canep.2011.11.006

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 138

  WeiC.Yu-tianC.Ji-zhiW.Yong-weiL.GangL. (2011). Characterization of hepatitis virus B isolated from a multi-drug refractory patient. Virus Res.155, 254–258. doi: 10.1016/j.virusres.2010.10.018

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 139

  WestlandC. (2003). Week 48 resistance surveillance in two phase 3 clinical studies of adefovir dipivoxil for chronic hepatitis B. Hepatology38, 96–103. doi: 10.1053/jhep.2003.50288

  • CrossRef
  • Google Scholar

• 140

  WHO (2021). Hepatitis B WHO guidlines. Available at: https://www.who.int/news-room/fact-sheets/detail/hepatitis-b (Accessed April 25, 2022).

  • Google Scholar

• 141

  WuY.GanY.GaoF.ZhaoZ.JinY.ZhuY.et al. (2014). Novel natural mutations in the Hepatitis B virus reverse transcriptase domain associated with hepatocellular carcinoma. PLoS One9. doi: 10.1371/journal.pone.0115141

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 142

  WuJ. F.NiY. H.ChenH. L.HsuH. Y.ChangM. H. (2014). The impact of hepatitis B virus precore/core gene carboxyl terminal mutations on viral biosynthesis and the host immune response. J Infect Dis209, 1374–1381. doi: 10.1093/infdis/jit638

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 143

  WunguC. D. K.AminM.RuslanS. E. N.PurwonoP. B.KholiliU.MaimunahU.et al. (2019). Association between host TNF-α, TGF-β1, p53 polymorphisms, hbv x gene mutation, hbv viral load and the progression of hbv-associated chronic liver disease in indonesian patients. Biomed. Rep.11, 145–153. doi: 10.3892/br.2019.1239

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 144

  XuR.ZhangX.ZhangW.FangY.ZhengS.YuX. F. (2007). Association of human APOBEC3 cytidine deaminases with the generation of hepatitis virus B x antigen mutants and hepatocellular carcinoma. Hepatology46, 1810–1820. doi: 10.1002/hep.21893

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 145

  YamaniL. N.YanoY.UtsumiT.WasityastutiW.RinonceH. T.WidasariD. I.et al. (2017). Profile of mutations in the reverse transcriptase and overlapping surface genes of hepatitis B virus (HBV) in treatment-naïve indonesian HBV carriers. Jpn. J. Infect. Dis.70, 647–655. doi: 10.7883/yoken.JJID.2017.078

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 146

  YanL.ZhangH.MaH.LiuD.LiW.KangY.et al. (2015). Deep sequencing of hepatitis B virus basal core promoter and precore mutants in HBeAg-positive chronic hepatitis B patients. Sci. Rep.9:5

  • Google Scholar

• 147

  YangH.QiX.SabogalA.MillerM.XiongS.DelaneyW. E. (2005). Cross-resistance testing of next-generation nucleoside and nucleotide analogues against lamivudine-resistant HBV. Antivir. Ther.1359

  • Google Scholar

• 148

  YangH.WestlandC. E.DelaneyW. E.HeathcoteE. J.HoV.FryJ.et al. (2002). Resistance surveillance in chronic hepatitis B patients treated with adefovir dipivoxil for up to 60 weeks. Hepatology36, 464–473. doi: 10.1053/jhep.2002.34740

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 149

  YinJ.XieJ.LiuS.ZhangH.HanL.LuW.et al. (2011). Association between the various mutations in viral core promoter region to different stages of hepatitis B, ranging of asymptomatic carrier state to hepatocellular carcinoma. Am. J. Gastroenterol.106, 81–92. doi: 10.1038/ajg.2010.399

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 150

  YuanJ. M.AmbinderA.FanY.GaoY. T.YuM. C.GroopmanJ. D. (2009). Prospective evaluation of hepatitis B 1762 T/1764 a mutations on hepatocellular carcinoma development in Shanghai, China. Cancer Epidemiol. Biomarkers Prev.18, 590–594. doi: 10.1158/1055-9965.EPI-08-0966

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 151

  ZhengJ.ZengZ.ZhangD.YuY.WangF.PanC. Q. (2012). Prevalence and significance of Hepatitis B reverse transcriptase mutants in different disease stages of untreated patients. Liver Int.32, 1535–1542. doi: 10.1111/j.1478-3231.2012.02859.x

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 152

  ZhuR.ZhangH. P.YuH.LiH.LingY. Q.HuX. Q.et al. (2008). Hepatitis B virus mutations associated with in situ expression of hepatitis B core antigen, viral load and prognosis in chronic hepatitis B patients. Pathol. Res. Pract.204, 731–742. doi: 10.1016/j.prp.2008.05.001

  • Pubmed Abstract
  • CrossRef
  • Google Scholar

• 153

  ZiaeeM.JavanmardD.SharifzadehG.NamaeiM. H.AzarkarG. (2016). Genotyping and mutation pattern in the overlapping MHR region of HBV isolates in southern khorasan, eastern Iran. Hepat. Mon.16:37806. doi: 10.5812/hepatmon.37806

  • Pubmed Abstract
  • CrossRef
  • Google Scholar