A novel subgenotype I3 of hepatitis B virus in Guangxi, China: a 15-year follow-up study.

Genotype I of hepatitis B virus (HBV) was proposed recently following sequencing of complete HBV genomes from Vietnam and Laos. However, its long-term molecular evolution is unknown. The objectives of this study were to study the molecular evolution of this genotype from an asymptomatic HBsAg carrier from the Long An cohort over a 15-year period was studied using both NGS and clone-based sequencing. The number of complete genome sequences obtained in 2004, 2007, 2013, and 2019 are 17, 20, 19, and 10, respectively. All strains belong to subgenotype I1, except for six (five from 2007 and one from 2019) and 8 further strains from 2007 which form a cluster branching out from other subgenotype I sequences, supported by a 100% bootstrap value. Based on complete genome sequences, all of the estimated intragroup nucleotide divergence values between these strains and HBV subgenotypes I1-I2 exceed 4%. These strains are recombinants between genotype I1 and subgenotype C but the breakpoints vary. The median intrahost viral evolutionary rate in this carrier was 3.88E-4 substitutions per site per year. The Shannon entropy (Sn) ranged from 0.55 to 0.88 and the genetic diversity, D, ranged from 0.0022 to 0.0041. In conclusion, our data provide evidence of novel subgenotypes. Considering that the 8 strains disappeared after 2007, while one of the 6 strains appears again in 2019, we propose these 6 strains as a new subgenotype, provisionally designated HBV subgenotype I3 and the 8 strains as aberrant genotype.

Infection with Hepatitis B Virus May Increase the Serum Concentrations of Osteopontin.

BACKGROUND: Serum osteopontin (OPN) concentrations were found to be significantly increased in patients infected with hepatitis B virus (HBV) and patients with hepatocellular carcinoma (HCC). OBJECTIVE: The aim of this study was to determine the association among HCC, OPN, and HBV. METHODS: Two hundred and forty-one subjects were recruited and divided into 6 groups: healthy controls, asymptomatic HBsAg carriers, HBsAg (-) patients with other tumors, HBsAg (+) chronic liver disease patients, HBsAg (+) patients with HCC, and HBsAg (-) patients with HCC or liver cirrhosis (LC). Serum concentrations of OPN and HBsAg were measured and analyzed. RESULTS: OPN concentrations in the HBsAg (+) HCC group were significantly higher than the healthy control group and the HBsAg (-) patients with other cancers (both p = 0.0001). The OPN concentrations of the HBsAg (-) patients with HCC or LC also did not differ significantly from those of the healthy control group (p = 0.075). There is a correlation between the titer of HBsAg and concentrations of OPN in all 3 HBsAg (+) groups (all p values <0.05). CONCLUSIONS: Infection with HBV may increase the serum concentrations of OPN. The association of OPN and HCC may be not attributable to tumor development per se but, rather, to HBV infection.

Novel subgenotype D11 of hepatitis B virus in NaPo County, Guangxi, bordering Vietnam.

Hepatitis B virus has been classified into 10 genotypes and 48 subgenotypes worldwide. We found previously, through polymerase chain reaction (PCR) amplification of a sample collected in 2011, that an HBsAg carrier was infected with two genotypes (B and D) of HBV. We carried out cloning, sequencing and phylogenetic analysis of the complete genomes and, for confirmation, analysed a sample collected from the same individual in 2018. Fifteen complete sequences were obtained from each sample. The carrier was infected in 2011 by genotypes B and D and by various recombinants, but only genotype D was present in 2018. The major and minor parents of the recombinants are genotypes B and D, respectively, although the recombination breakpoints vary among them. All 23 genotype D isolates form a cluster, branching out from other subgenotype D sequences and supported by a 100 % bootstrap value. Based on complete genome sequences, almost all of the estimated intragroup nucleotide divergence values between our isolates and HBV subgenotypes D1-D10 exceed 4 %. Compared to the other subgenotypes (D1-D10), 35 unique amino acids were present in our isolates. Our data provide evidence for a novel subgenotype, provisionally designated HBV subgenotype D11.

Asymptomatic hepatitis B carriers who were vaccinated at birth.

The long-term persistence of immunity following universal infant immunization against hepatitis B virus (HBV) and the need for a subsequent booster dose in adolescence remain under debate. With data derived from Long'an County, Guangxi, China, we reported previously that the prevalence of hepatitis B surface antigen (HBsAg) among adults born from 1987 to 1993 increases with age, although these individuals had received a first dose of the vaccine within 24 hours of birth. Here, we sought the source of transmission by comparison of genotypes among their family members using phylogenetic analysis of complete HBV S gene sequences. For comparison, we screened 2199 vaccinated individuals aged 5 to 17 in Cang Wu County and 1592 vaccinated individuals aged 3 to 7 in Ling Shan County in Guangxi for HBsAg carriers and investigate their family members. In total, 50 asymptomatic HBsAg carriers who were vaccinated at birth and 152 family members were analyzed. The results showed that 25% (95% CI: 6.0-44.0) of the HBsAg-positive children had not acquired their HBV infection from their mothers. This phenomenon showed a trend that increases with age. Antibody escape mutations were detected in 22.9% (95% CI: 11.0-34.8) of the isolates. In conclusion, a booster dose may be necessary for adolescence who were vaccinated at birth in highly endemic countries.

High Prevalence of HBV Lamivudine-Resistant Mutations in HBV/HIV Co-Infected Patients on Antiretroviral Therapy in the Area with the Highest Prevalence of HIV/HBV Co-Infection in China.

OBJECTIVES: We aimed to determine the prevalence of hepatitis B virus (HBV) drug-resistant mutations in patients co- infected with HBV/human immunodeficiency virus (HIV), including both drug-naïve subjects and those who received antiretroviral therapy (ART) in Guangxi, where the prevalence of HIV/HBV co-infection is highest in China. METHODS: Two hundred and three subjects co-infected with HBV/HIV were recruited, including 123 drug-naïve patients (group 1) and 80 who received ART (group 2). The polymerase gene of HBV in the serum of all study subjects was analysed. RESULTS: The results showed that the prevalence of HBV drug-resistant mutations in group 2 (76.5%, 95% CI 56.3-96.7) was significantly higher than that in group 1 (1.4%, 95% CI -1.4 to 4.2; χ2 = 50.955, p < 0.05). The major pattern of lamivudine (3TC)-resistant mutations is L180M+M204I+L80I (35.7%). In total, 95% of subjects with resistant mutations had cross-resistance to telbivudine and entecavir. No putative tenofovir disoproxil fumarate (TDF) resistance change was found. Five subjects (6.5%) in group 2 had HBV viral loads over 10 × 106 copies/mL. Four of them had 3TC-resistant mutations. Multivariate analysis showed that ART was the only factor associated with the development of drug-resistant mutations. CONCLUSION: Treating HIV in HIV/HBV co-infection with antiretroviral agents may result in a very high prevalence of HBV 3TC-resistant mutations. TDF could not completely suppress HBV replication.

Hepatitis B Virus Core Promoter Double Mutations (A1762T, G1764A) Are Associated with Lower Levels of Serum Dihydrolipoyl Dehydrogenase.

OBJECTIVES: The aim of this study was to identify serum proteins with differential concentrations between hepatocellular carcinoma (HCC) patients and HBsAg asymptomatic carriers among individuals infected with hepatitis B virus (HBV) with basal core promoter (BCP) double mutations (A1762T, G1764A). METHODS: iTRAQ and liquid chromatography-tandem mass spectrometry were used to identify differentially expressed protein, and an ELISA test was used for the validation test. RESULTS: The total number of proteins identified was 1,125, of which 239 showed statistically significant differences in their expression. The relative concentrations of serum dihydrolipoyl dehydrogenase (DLD), which showed the most significant correlation with liver diseases and infection, were significantly lower in HCC patients than asymptomatic HBsAg carriers and individuals negative for HBsAg. However, only the difference between HCC patients with BCP double mutations and HBsAg-negative individuals could be confirmed by ELISA. Meanwhile, we found that the concentrations of serum DLD in those infected with HBV with BCP double mutations were significantly lower than in individuals with the wild-type BCP. However, the difference in the concentrations of serum DLD between individuals with wild-type BCP and those negative for HBsAg was not significant. CONCLUSIONS: HBV with BCP double mutations are associated with lower concentrations of serum DLD.

The correlation between serum HBsAg levels and viral loads depends upon wild-type and mutated HBV sequences rather than the HBeAg/anti-HBe status.

Despite several studies regarding the correlation between serum HBsAg titers and viral loads, the association remains uncertain. Eighty-nine individuals were selected randomly from a Chinese cohort of 2,258 subjects infected persistently with hepatitis B virus (HBV) for cross-sectional and longitudinal analysis. Viral loads of mutant HBV are lower than those of wild type HBV. The serum HBsAg titers correlate positively with viral loads in both HBeAg positive and negative subjects (r = 0.449, P = 0.013; r = 0.300, P = 0.018, respectively). No correlation between serum HBsAg titer and viral loads was found in any of the four phases of chronic HBV infection. The serum HBsAg titers correlate positively with viral loads in the group with wild type sequences of the PreS/S, basal core promoter (BCP), and preC regions of HBV(r = 0.502, P = 0.040). However, the correlation was not seen in the group with mutations in these regions (r = 0.165, P = 0.257). The correlation between HBsAg titers and viral loads was seen in individuals with wild type PreS/S sequences but not in the subgroup with BCP double mutations or PreC stop mutation, although their sequences in the preS/S regions were wild type. All these findings were confirmed by the longitudinal analysis. In conclusion, the correlation between serum HBsAg levels and viral loads may not differ between HBeAg positive and negative individuals but may depend on wild-type or mutated genomic sequences. Therefore, HBsAg quantitation may be used as a surrogate for viral loads in only wild-type HBV infections.

Hepatitis B virus candidate subgenotype I1 varies in distribution throughout Guangxi, China and may have originated in Long An county, Guangxi.

Sequencing of the complete hepatitis B virus (HBV) genomes from Vietnam, China and Laos led to the identification of a complex recombinant, referred to initially as an aberrant genotype and later proposed to be a new genotype, I. However, epidemiological data regarding this new genotype are lacking. A cross-sectional study was carried out to investigate the epidemiology of HBV candidate genotype I in Guangxi, China using stratified, random cluster sampling. Four thousand five hundred thirteen subjects were recruited from five counties within Guangxi. Three genotypes, B, C, and I, were identified with a prevalence of 32.6% (114/350), 64% (224/350), and 3.4% (12/350), respectively. All the genotype I isolates belong to candidate subgenotype I1 and were found in Bing Yang (15.3%, 9/59) and Na Po (5.0%, 3/60) counties only. The prevalence of this subgenotype is significantly higher in males (5.1%, 10/195) than in females (1.3%, 2/155; X(2) = 3.959, P < 0.05) but does not differ significantly with age. It was found in the Han (4.5%, 9/201) and Zhuang (3.1%, 3/97) ethnic populations only. There is no significant difference from other genotypes in the prevalence of HBV serological markers. Phylogeographic analysis revealed that genotype I1 likely arose in Long An county, then spread later to Bing Yang, Na Po counties and elsewhere in southeast Asia. In conclusion, the distribution of candidate genotype I within Guangxi is not even and it is highly endemic in some counties. Its prevalence is associated with gender and ethnicity. Subgenotype I1 likely originated in Long An county.

Combined core promoter mutations and pre-S deletion of HBV may not increase the risk of HCC: a geographical epidemiological study in Guangxi, China.

BACKGROUND: Although persistent hepatitis B virus (HBV) infection is a major cause of hepatocellular carcinoma (HCC), the mechanisms of oncogenesis remain obscure. AIMS: To determine whether the findings that HBV basal core promoter (BCP) A1762T, G1764A double mutations, pre-S deletions and a combination of both are risk factors of HCC are supported by geographical epidemiology. METHODS: Study subjects were recruited from Long An county, where the incidence of HCC is the highest, and five other counties in Guangxi, where the HCC incidence is lower and varies among them. The Pre-S region and BCP of HBV from all study subjects were amplified and sequenced and the data were analysed using chi-squared tests. RESULTS: The prevalence of BCP and pre-S mutations differs significantly (χ(2) = 9.850, 5.135, respectively, all P < 0.01) between Long An and the other counties. However, the prevalence of combined BCP and pre-S mutations does not differ significantly (χ(2) = 1.510, P > 0.05). These mutations are less frequent in the young but the prevalence of pre-S deletions does not increase with age. The prevalence of these mutations does not differ significantly between men and women but is significantly higher in Zhuang than the other ethnic populations. Among the other five counties, the prevalence of BCP mutations in counties where the HCC incidence is high is significantly higher than that of counties where the HCC incidence is low. CONCLUSIONS: Combined BCP double mutations and pre-S deletion may not increase the risk of HCC, although these mutations are a risk factor of HCC when they present alone.

Low host immune pressure may be associated with the development of hepatocellular carcinoma: a longitudinal analysis of complete genomes of the HBV 1762T, 1764A mutant.

Background: It has been reported that hepatitis B virus (HBV) double mutations (A1762T, G1764A) are an aetiological factor of hepatocellular carcinoma (HCC). However, it is unclear who is prone to develop HCC, among those infected with the mutant. Exploring HBV quasispecies, which are strongly influenced by host immune pressure, may provide more information about the association of viral factors and HCC. Materials and methods: Nine HCC cases and 10 controls were selected from the Long An cohort. Serum samples were collected in 2004 and 2019 from subjects with HBV double mutations and the complete genome of HBV was amplified and sequenced using next-generation sequencing (NGS). Results: The Shannon entropy values increased from 2004 to 2019 in most cases and controls. There was no significant difference in mean intrahost quasispecies genetic distances between cases and controls. The change in the values of mean intrahost quasispecies genetic distances of the controls between 2004 and 2019 was significantly higher than that of the cases (P<0.05). The viral loads did not differ significantly between cases and controls in 2004(p=0.086) but differed at diagnosed in 2019 (p=0.009). Three mutations occurring with increasing frequency from 2004 to 2019 were identified in the HCC cases, including nt446 C→G, nt514 A→C and nt2857T→C. Their frequency differed significantly between the cases and controls (P<0.05). Conclusions: The change in the values of mean intrahost quasispecies genetic distances in HCC was smaller, suggesting that HBV in HCC cases may be subject to low host immune pressure. Increasing viral loads during long-term infection are associated with the development of HCC. The novel mutations may increase the risk for HCC.

Prevalence of hepatitis B virus infection in a highly endemic area of southern China after catch-up immunization.

The Chinese national goals for control of hepatitis B virus (HBV) infection were to achieve a prevalence of HBsAg below 7% for the entire population, and 1% for children under 5-year old, by 2010. To determine whether Guangxi, a multi-minority province with a low socio-economic status and a very high prevalence of HBV, achieved this goal, a seroepidemiological survey of HBV infection was carried out using stratified, random cluster sampling. The results show that the overall prevalence of HBsAg is 9.16% [95% confidence interval (CI) = 8.32-10%]. The prevalence in males (10.96%, 95% CI = 9.64-12.28%) is significantly higher than in females (7.71%, 95% CI = 6.64-8.78%; χ(2) = 10.5923, P < 0.05). The prevalence in children under 5-year old is 3.62% (95% CI = 0.60-6.64%) and increases with age. The prevalence of HBsAg in non-immunized individuals is significantly higher than in those immunized completely, although not within 24 hr of birth (χ(2) = 31.426, P < 0.05); a significant difference was found in those below the age of 20 years but not in older persons. Gender, age, immunization history, and familial HBsAg carriers are risk factors for infection. In conclusion, this study indicates that Guangxi has not reached the goal for the control of HBV infection. Catch-up HBV immunization may not protect adults effectively against infection in highly endemic regions.

Analysis of entire hepatitis B virus genomes reveals reversion of mutations to wild type in natural infection, a 15 year follow-up study.

It has been reported that some mutations in the genome of hepatitis B virus (HBV) may predict the outcome of the virus infection. However, evolutionary data derived from long-term longitudinal analysis of entire HBV genomes using next generation sequencing (NGS) remain rare. In this study, serum samples were collected from asymptomatic hepatitis B surface antigen (HBsAg) carriers from a long-term prospective cohort. The entire HBV genome was amplified by polymerase chain reaction (PCR) and sequenced using NGS. Twenty-eight time series serum samples from nine subjects were successfully analysed. The Shannon entropy (Sn) ranged from 0 to 0.89, with a median value of 0.76, and the genetic diversity (D) ranged from 0 to 0.013, with a median value of 0.004. Intrahost HBV viral evolutionary rates ranged from 2.39E-04 to 3.11E-03. Double mutations at nt1762(A â†’ T) and 1764(G â†’ A) and a stop mutation at nt1896(G â†’ A) were seen in all sequences from subject BO129 in 2007. However, in 2019, most sequences were wild type at these positions. Deletions between nt 2920-3040 were seen in all sequences from subject TS115 in 2007 and 2013 but these were not present in 2004 or 2019. Some sequences from subject CC246 had predicted escape substitutions (T123N, G145R) in the surface protein in 2004, 2013 and 2019 but none of the sequences from 2007 had these changes. In conclusion, HBV mutations may revert to wild type in natural infection. Clinicians should be wary of predicting long-term prognoses on the basis of the presence of mutations.

A complex hepatitis B virus (X/C) recombinant is common in Long An county, Guangxi and may have originated in southern China.

Recently, a complex (X/C) hepatitis B virus (HBV) recombinant, first reported in 2000, was proposed as a new genotype; although this was refuted immediately because the strains differ by less than 8 % in nucleotide distance from genotype C. Over 13.5 % (38/281) of HBV isolates from the Long An cohort in China were not assigned to a specific genotype, using current genotyping tools to analyse surface ORF sequences, and these have about 98 % similarity to the X/C recombinants. To determine whether this close identity extends to the full-length sequences and to investigate the evolutionary history of the Long An X/C recombinants, 17 complete genome sequences were determined. They are highly similar (96-99 %) to the Vietnamese strains and, although some reach or exceed 8 % nucleotide sequence difference from all known genotypes, they cluster together in the same clade, separating in a phylogenetic tree from the genotype C branch. Analysis of recombination reveals that all but one of the Long An isolates resembles the Vietnamese isolates in that they result from apparent recombination between genotype C and a parent of unknown genotype (X), which shows similarity in part to genotype G. The exception, isolate QL523, has a greater proportion of genotype C parent. Phylogeographic analysis reveals that these recombinants probably arose in southern China and spread later to Vietnam and Laos.

Changing risk factors for hepatocellular carcinoma in hyperendemic regions in the era of universal hepatitis B vaccination.

BACKGROUND: LongAn, Guangxi, was the first county in China to implement universal childhood hepatitis B virus (HBV) immunization. We aimed to determine its long-term effects in preventing hepatocellular carcinoma (HCC) 32 years after the immunization programme was launched. METHODS: Information on HCC deaths for LongAn and its neighbouring county, BinYang (where universal hepatitis B vaccination was not started till 2002), were obtained from the national mortality surveillance system. The data were analysed using Poisson regression. RESULTS: The overall age-adjusted mortalities of HCC in LongAn and BinYang during 2017-2018 were 53.3/100,000 and 45.4/100,000, respectively. The mortality of males aged 20-29 years in LongAn, who were vaccinated at birth, was lower (2.7/100,000, 95%CI 0.8-4.5) than that of males in BinYang, who were not vaccinated (4.7/100,000, 95%CI 3.2-6.3). In LongAn, the HCC mortality in adults aged 20-29 years declined significantly from 7.9/100,000 (95%CI 4.4-11.4) in 2004 to 1.4/100,000 (95%CI 0.4-2.4) in 2017-2018 (χ2 = 5.554, p = 0.018). Among those vaccinated at birth, the HCC mortality in mountainous areas, where dietary exposure to aflatoxins is more common, is higher (9.0/100,000, 95%CI 4.5-13.5) than in low-lying areas (6.5/100,000, 95%CI 3.6-9.4) (χ2 = 0.2393, p = 0.618). CONCLUSION: Immunization of infants against HBV has reduced their risk of developing HCC as children and young adults but could not prevent all cases of HCC, suggesting that the major risk factor for HCC in hyperendemic regions is shifting from HBV to other factors. Additional prevention strategies for HCC will be needed in the future.

[A prospective study of the relationship between serum hepatitis B virus DNA and the risk of primary liver cancer].

OBJECTIVE: To determine the relationship between the serum hepatitis B virus (HBV) DNA and the risk of primary liver cancer (PLC). METHODS: Farmers aged 30 to 55 years in Long An county were recruited in this study Blood samples were collected and the sera were tested for HBsAg using Enzyme-Linked ImmunoSorbent Assay (ELISA), and the HBsAg-positive sera were further tested for viral DNA using nested polymerase chain reaction (nPCR). The study subjects were divided into three groups. The first group was positive for both HBsAg and HBV DNA. The second group was positive for HBsAg but negative for HBV DNA. Age-, sex-, residence-matched HBsAg negative controls for group 1 and group 2 were enrolled in the third group. The cohort was followed up for four years. RESULTS: The positive rate of HBsAg in these farmers was 14.52% (3975/27,379), and the HBV DNA positive rate in HBsAg positive subjects was 40.35% (1604/3975). The total PLC incidence rate in Group 1 and 2 was 672.45 /100,000 person-years (PY), significantly higher than that in Group3 (17.19 /100,000 PY). The relative risk (RR) was 39.123, and the 95% confidence interval (CI) was 9.018-159.146. The PLC incidence rate of Group 1 (984.03/100,000 PY) was significantly higher than that of Group2 (324.38 /100,000 PY). The RR was 3.034, and the 95% CI was 1.795-5.125. Multivariate analyses of Group1 and 2 with Cox model showed that sex, age, serum HBV DNA, and family history of PLC were independent risk factors of PLC. CONCLUSION: HBV DNA and HBsAg positive subjects have a higher chance to develop PLC than HBV DNA negative-, HBsAg positive subjects.

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.

OBJECTIVES: Surveillance of hepatocellular carcinoma (HCC) can detect small tumors for resection but at a huge cost of health resources. The challenge is to reduce the surveillance population. We reported that 96% of HCC patients but only 24% of controls were infected with the hepatitis B virus (HBV) with A(1762)T, G(1764)A mutations in Guangxi, China. It is likely to be extremely beneficial in terms of cost and resources if a significant number of tumors can be detected early by screening this selected population. Our aim is to test this hypothesis. METHODS: A cohort of 2,258 hepatitis B surface antigen-positive subjects aged 30-55 yr was recruited in Guangxi. Following evaluation of virological parameters at baseline, HCC is diagnosed by 6-monthly measurements of serum alpha-fetoprotein levels and ultrasonographic examinations. RESULTS: Sixty-one cases of HCC were diagnosed after 36 months of follow-up. The HCC rate was higher in the mutant than wild-type group (P < 0.001, rate ratio [RR] 6.23, 95% confidence interval [CI] 2.83-13.68). The HCC rate in the male mutant group was higher than that in the male wild-type group (P < 0.001, RR 11.54, 95% CI 3.58-37.24). Specifically, 93.3% of male cases are infected with the mutant. Multivariate analyses showed that in men, increasing age and A(1762)T, G(1764)A double mutations are independently associated with developing HCC. CONCLUSIONS: HBV A(1762)T, G(1764)A mutations constitute a valuable biomarker to identify a subset of male HBsAg carriers aged >30 yr at extremely high risk of HCC in Guangxi, and likely elsewhere.

Comparison of immunogenicity between hepatitis B vaccines with different dosages and schedules among healthy young adults in China: A 2-year follow-up study.

Immunogenicity of hepatitis B vaccine between 20 µg with 3-dose schedule and 60 µg with 2-dose regimens was compared 2 years after primary immunization. A total of 353 healthy adults aged 18-25 years were enrolled in the study and randomly assigned (1: 1: 1) into 3 vaccine groups: A (20 µg, 0-1-6 month), B (60 µg, 0-1 month) and C (60 µg, 0-2 month). Serum samples were collected at 1 month after a series vaccination and 12 months, 24 months after the first-dose. The GMC level of anti-HBs antibody was measured using Chemiluminescent Microparticle ImmunoAssay (CMIA). There were 59, 45 and 55 vaccinees available to follow-up with 2 year later in vaccine groups A, B and C, respectively. No significant differences existed in sex ratio, age and body mass index (BMI) among vaccinees at month 24 and the corresponding participants at baseline in each group (P > 0.05). The seroprotection rates in group A, B and C were 98.31%, 88.37% and 85.19%, respectively (P = 0.014), reflecting the fact that the rate of group A was significantly higher than that in group C (P = 0.026). Also, the GMC level of anti-HBs antibody in group A was significantly higher than those of other two groups (427.46 mIU/ml vs. 89.74 mIU/ml, 89.80 mIU/ml, respectively; all P < 0.01). This data suggested that the standard 20 µg (0-1-6 month) regimen of hepatitis B vaccine should be recommended as a priority on the premise of complete compliance in adults.

Higher prevalence of cancer related mutations 1762T/1764A and PreS deletions in hepatitis B virus (HBV) isolated from HBV/HIV co-infected compared to HBV-mono-infected Chinese adults.

In the era of combination therapy for human immunodeficiency virus (HIV), liver disease including hepatocellular carcinoma (HCC), are the major causes of death for patients co-infected with hepatitis B virus (HBV) and HIV. However, the mechanisms remain obscure. We aimed to determine whether HCC-related HBV mutations including 1762T/1764A double mutation and pre-S deletions occur more frequently in HBV/HIV co-infected individuals compared to HBV mono-infected individuals. In this study, the basic core promoter (BCP) and the preS/S regions of HBV isolated from 61 pairs of HBV/HIV co-infected and HBV mono-infected participants were analyzed. We found that the prevalence of HBV isolates with 1762T/1764A and/or preS deletion mutations was 37.7% (95% CI: 29.1-46.3). The prevalence of these mutations in HBV/HIV co-infected group (52.5%, 95% CI: 40.0-65.0) was significantly higher than in the HBV mono-infected group (23.0%, 95% CI: 12.4-33.6) (X2=11.307, P<0.05). HBV/HIV co-infection was associated with higher viral loads but these higher viral loads were not associated with the higher prevalence of HCC-related HBV mutations. Individually 1762T1764A (44.3%) or preS deletions (23%) occurred more frequently in isolates from co-infected compared to mono-infected individuals (21.3%, 4.9%, respectively) (X2=7.290, P<0.05; X2=8.270, P<0.05). Moreover, 1762T/1764A and preS deletions occurred more frequently in genotypes C and I compared to genotype B (p<0.05). Multivariate analysis revealed that co-infection with HIV was associated with the development of both 1762T/1764A ((RR: 2.932(1.325-6.488)) and preS deletions ((RR: 5.759(1.562-21.235)). These results demonstrate that co-infection with HIV was associated with increased prevalence of HCC-related mutations in HBV isolates from Chinese patients.

Des-γ carboxyprothrombin may not be a good biomarker for hepatocellular carcinoma in those chronically infected with hepatitis B virus with basal core promoter double mutations (T

BACKGROUND: The accuracy of des-γ -carboxyprothrombin (DCP) in the detection of hepatocellular carcinoma (HCC) in those infected hepatitis B virus (HBV) from cross-sectional or case-control studies is contradictory. OBJECTIVE: To resolve this contradiction using a prospective study. METHODS: Three hundred male individuals persistently infected with HBV were recruited from the Chinese cohort and followed up once per year from 2012 to 2015. Each subject was screened for HCC by measurements of serum alpha-fetoprotein (AFP), lectin-bound α -fetoprotein (AFP-L3), DCP concentrations and ultrasonographic examinations. RESULTS: Nineteen HCC cases were identified. The area under receiver operating characteristic (AUROC) at first, second and third visit for AFP, AFP-L3 and DCP ranges from 0.710-0.897, 0.566-0.637 and 0.520-0.595, respectively. The rate of elevated DCP is not significantly different between the HCC cases and controls (52.6% vs. 47.4%) (P > 0.05). The incidence of HCC in subjects with elevated DCP is not significantly higher than that of those with normal DCP (9.5% vs. 4.6%) (P > 0.05). The AUROC of combinations of these biomarkers was higher than that of AFP alone at the first visit. However, it was reduced at the second visit. At the third visit, the AUROCs of AFP + DCP and AFP + AFP-L3 + DCP, but not that of AFP + AFP-L3, were higher than that of AFP alone. CONCLUSIONS: AFP but DCP or AFP-L3 remains a valuable biomarker for HCC in those chronically infected with HBV. The combination with AFP-L3 and DCP may not increase the accuracy of AFP in differentiating HCC cases from controls, among those infected with HBV.

The incidence rate over 10 years of naturally occurring, cancer related mutations in the basal core promoter of hepatitis B virus.

Cross-sectional analyses showed that the prevalence of basal core promoter (BCP) double mutations (nt 1762T, 1764A) of hepatitis B virus (HBV) gradually increases with age. We aimed to determine the incidence rate of the mutations over 10 years. Study subjects were selected from the Long An cohort established in 2004, including 59 with HBV with single mutations at nt 1762 or 1764 in the BCP and 342 with wild type BCP sequences at baseline. Their serum samples for analysis were obtained at the 3rd and 10th annual visits, respectively. The results showed that the annual incidence rate of BCP double mutations is 3.8% (95% confidence interval [CI]: 1.4-6.2) and tends to decrease with age. The peak incidence is in the 30-34 years age-group. The incidence rate in HBeAg positive individuals (5.5%) is significantly higher than in those without HBeAg (3.4%) (P<0.05). The incidence rate is significantly higher in genotype C (4.8%) than in genotype B (2.8%) or I (3.1%). The incidence rate of the mutations (6.8%) developing from a single mutation at nt 1762 or 1764 is significantly higher than that (3.8%) from the wild type sequence (P<0.005). The difference in incidence of single mutations between nt 1762 (0.7%) and 1764 (0.03%) is significant (P<0.05). In conclusion, the incidence rate of BCP double mutations tends to decrease with age after the age of 35 years. Viruses with a single mutation at nt 1762 or 1764 are more prone to develop double mutations. Nt 1762 is the more common site of the first mutation.