Molecular epidemiology of hepatitis B among Indigenous Australians in Queensland and the Torres Strait Islands.

BACKGROUND: Chronic hepatitis B virus (HBV) infection is a major health problem for all Indigenous Australians. Post-2000, Hepatitis B surface antigen prevalence has decreased, although remaining four times higher among Indigenous compared with non-Indigenous people. AIMS: This study aimed to characterise the HBV from Indigenous populations in Queensland and the Torres Strait Islands. METHODS: Serum samples were collected, with consent, from people within Queensland Indigenous communities prior to 1990 as part of the Queensland Health vaccination programme. Ethics approval was subsequently obtained to further characterise the HBV from 93 of these stored samples. HBV DNA was extracted and genotype was obtained from 82 samples. HBV full genome sequencing was carried out for a subset of 14 samples. RESULTS: Seventy-eight samples were identified as genotype C (2 × C12, 3 × C13 and 73 × C14), one sample as genotype A (A2) and three samples as genotype D (1 × D2, 1 × D3 and 1 × D4). The HBV/C sequences identified were most closely related to sequences isolated from Papua New Guinea and Indonesia (Papua Province). CONCLUSIONS: The HBV isolated from the Torres Strait Islanders was notably different to the HBV/C4 strain isolated from Indigenous people of mainland northern Australia, with no evidence of recombination. This reflects the differences in culture and origin between Torres Strait Islanders and mainland Indigenous people.

Immunogenicity of Wild Type and Mutant Hepatitis B Surface Antigen Virus-like Particles (VLPs) in Mice with Pre-Existing Immunity against the Wild Type Vector.

Virus-like particles (VLPs), composed of the small hepatitis B virus surface antigen (HBsAgS), are the antigenic components of the hepatitis B virus (HBV) vaccine and represent the backbones for a chimeric anti-malaria vaccine and various vaccine candidates. Biological vectors have to face pre-existing anti-vector immune responses due to previous immune exposure. Vector recognition after natural infections or vaccinations can result in unwarranted outcomes, with compromising effects on clinical outcomes. In order to evaluate the impact of a pre-existing anti-HBsAgS immune response, we developed mutant VLPs composed of subunits with reduced HBsAgS-specific antigenicity. The insertion of a Plasmodium falciparum circumsporozoite protein (CSP)-derived epitope as a read-out allowed the assessment of wild type (wt) and mutant VLPs in the context of a pre-existing immune response. Mutant and wt VLP platforms with a CSP-epitope insert are immunogenic and have the ability to generate anti-CSP antibody responses in both naïve BALB/c mice and mice with a pre-existing anti-HBsAgS immune response, but with superior anti-CSP responses in mice with a pre-existing immunity. The data indicate that previous HBsAgS exposure facilitates enhanced antibody responses against foreign epitopes delivered by the HBsAgS platform, and, in this context, the state of immune sensitization alters the outcome of subsequent vaccinations.

A phase 2, open-label, randomized, multiple-dose study evaluating Inarigivir in treatment-naïve patients with chronic hepatitis B.

BACKGROUND/AIMS: Novel agents acting against hepatitis B virus (HBV) are needed to improve HBsAg seroclearance or termed as 'functional cure'. Inarigivir (retinoic acid-inducible gene I agonist) has immunomodulatory and direct antiviral actions against HBV. We aimed to determine the safety and efficacy of Inarigivir for the treatment of HBV infection. PATIENTS/METHODS: 80 treatment-naïve patients were randomized in 4 ascending dose cohorts to receive 12 weeks of Inarigivir 25, 50, 100, 200 mg or placebo in a ratio of 4:1. All patients were then given tenofovir for another 12 weeks. RESULTS: Least squares (LS) mean reductions in HBV DNA from baseline increased with higher doses of Inarigivir (0.6116 in 25 mg and 1.5774 in 200 mg groups vs. 0.0352 in placebo group) (95% CI 0.9518-0.2011 and 1.921-1.1634 respectively). LS mean changes in HBV RNA and HBsAg from baseline ranged from -0.3856 to -0.5794 versus -0.1474 and -0.0956 to -0.1818 versus +0.0026 in Inarigivir-treated versus placebo groups respectively. During the tenofovir-treated period, LS mean reductions in HBsAg in the Inarigivir-treated groups ranged from 0.1709 to 0.3529 versus 0.1984 in the placebo group. Inarigivir-treated groups showed mean reductions in ALT from baseline between 23.3 and 33.8 versus 0.7 U/L in the placebo group. Treatment-emergent adverse events related to Inarigivir and placebo occurred in 4.7% and 6.3% patients respectively. CONCLUSIONS: Twelve-week Inarigivir up to 200 mg dose was associated with a reduction of HBV DNA, HBV RNA and antigen levels. A trend for greater HBsAg reduction was observed in Inarigivir pre-treated patients after switching to tenofovir.

Preventing early childhood transmission of hepatitis B in remote aboriginal communities in Northern Australia.

BACKGROUND: Chronic hepatitis B is a public health concern in Aboriginal communities in the Northern Territory of Australia with prevalence almost four times the non-Aboriginal population. Infection is suspected to mainly occur in early life, however, the mode of transmission and vaccine effectiveness is not known in this population. WHO has set a target for hepatitis B elimination by 2030; elimination in this disproportionately affected population in Australia will require understanding of the modes of transmission and vaccine effectiveness. METHODS: We conducted the study at four very remote Aboriginal communities. We approached mothers who had chronic hepatitis B and had given birth between 1988 and 2013 for consent. We obtained hepatitis B serology, immunisation and birth details from the medical record. If both mother and child had hepatitis B viral DNA detected, we performed viral whole genome sequencing. RESULTS: We approached 45 women for consent, of whom 23 agreed to participate. We included 20 mothers and 38 of their children. Of the 20 included mothers, 5 (25%) had children who were hepatitis B immune by exposure and 3 (15%) had children with evidence of chronic hepatitis B infection at the time of assessment. Hepatitis B immunoglobulin (HBIg) had been given at birth in 29/38 (76.3, 95% CI 59.8-88.6) children, and 26 children (68.4, 95% CI 51.3-82.5) were fully vaccinated. Of the 3 children who had chronic hepatitis B, all had received HBIg at birth and two were fully vaccinated. Of the 5 who were immune by exposure, 4 had received HBIg at birth and one was fully vaccinated. Whole genome sequencing revealed one episode of definite mother to child transmission. There was also one definite case of horizontal transmission. CONCLUSIONS: Chronic hepatitis B in this context is a sensitive issue, with a high proportion of women refusing consent. Although uncommon, there is ongoing transmission of hepatitis B to Aboriginal children in remote northern Australia despite vaccination, and this is likely occurring by both vertical and horizontal routes. Prevention will require ongoing investment to overcome the many barriers experienced by this population in accessing care.

Hepatitis B Virus Flares After Nucleot(s)ide Analogue Cessation Are Associated With Activation of Toll-Like Receptor Signaling Pathways.

BACKGROUND: We evaluated the patterns of peripheral Toll-like receptor (TLR) signaling activity and the expression of TLRs and natural killer (NK) cell activation in a cohort of patients experiencing severe hepatitis flares after stopping nucleot(s)ide analogues (NAs) therapy. METHODS: Samples were collected longitudinally from patients with chronic hepatitis B who were enrolled in a prospective study of NA discontinuation. Patients experiencing hepatitis flares were compared with patients with normal alanine aminotransferase. Peripheral blood mononuclear cells (PBMCs) were stimulated with TLR ligands and cytokine secretion in the cell culture supernatant measured. Expression of TLR2/4, NKG2D, NKp46, and triggering receptor expressed on myeloid cells 1 (TREM-1) on monocytes, NK, and NK-T cells was measured. RESULTS: Seventeen patients with severe reactivation hepatitis flares were compared to 12 nonflare patients. Hepatitis flares were associated with increased activity of TLR2-8 and TLR9 signaling in PBMCs at the time of peak flare compared to baseline. Hepatitis flares were also associated with (1) upregulation of TLR2 and (2) TREM-1 receptor expression on NK. There were no differences at baseline between flare patients and nonflare patients. CONCLUSIONS: Hepatitis flares off NA therapy have a significant innate inflammatory response with upregulation of TLR signaling on peripheral monocytes and TLR2 and TREM-1 expression on NK cells. This implicates the innate immune system in the immunopathogenesis of hepatitis B flares.

Combination treatments including the small-interfering RNA JNJ-3989 induce rapid and sometimes prolonged viral responses in patients with CHB.

BACKGROUND & AIMS: RNA interference therapy has been shown to reduce hepatitis B surface antigen (HBsAg) levels in preclinical models, which could confer functional cure in patients with chronic hepatitis B. This phase IIa trial (ClinicalTrials.gov Identifier: NCT03365947) assessed the safety and efficacy of the small-interfering RNA JNJ-73763989 (JNJ-3989) plus a nucleos(t)ide analogue (NA), with/without the capsid assembly modulator JNJ-56136379 (JNJ-6379) in patients with chronic hepatitis B. METHODS: Treatment-naïve and NA-suppressed patients received 3 subcutaneous JNJ-3989 doses every week (QW; 100, 200, or 300 mg), 2 weeks (Q2W; 100 mg) or 4 weeks (Q4W; 25, 50, 100, 200, 300, or 400 mg), or JNJ-3989 Q4W (200 mg) plus oral JNJ-6379 250 mg daily for 12 weeks. Patients received NAs throughout. RESULTS: Eighty-four patients were recruited. All treatments were well tolerated, with all 5 serious adverse events considered unrelated to study drugs. JNJ-3989 100 to 400 mg Q4W resulted in HBsAg reductions ≥1 log10 IU/ml from baseline in 39/40 (97.5%) patients at the nadir. All patients receiving the triple combination (n = 12) had HBsAg reductions ≥1 log10 IU/ml from baseline at the nadir. HBsAg reductions were similar for HBeAg-positive (n = 21) and HBeAg-negative (n = 47) patients in all JNJ-3989 Q4W treatment arms, including the triple combination (n = 68). Smaller HBsAg reductions were seen with 25 mg (n = 8) and 50 mg (n = 8) than with 100 to 400 mg (n = 40). Shorter dosing intervals (QW [n = 12] and Q2W [n = 4]) did not improve response vs. Q4W dosing. HBsAg reductions ≥1 log10 IU/ml from baseline persisted in 38% of patients 336 days after the last JNJ-3989 dose. CONCLUSIONS: JNJ-3989 plus an NA, with/without JNJ-6379, was well tolerated and resulted in HBsAg reductions up to 336 days after the last JNJ-3989 Q4W dose. CLINICAL TRIAL NUMBER: NCT03365947. LAY SUMMARY: Hepatitis B virus affects people's livers and produces particles called hepatitis B surface antigen (HBsAg) that damage a person's liver and can help the virus infect a person for a long time, known as chronic hepatitis B (CHB). In this study, a new treatment called JNJ-3989 was assessed (in combination with normal treatment known as nucleos(t)ide analogues), for its safety and effectiveness in reducing the number of HBsAg particles in people with CHB. The results of this study showed that treatment with JNJ-3989 could be safe for people with CHB, lowered their HBsAg levels, and kept HBsAg levels lowered for 336 days in 38% of patients after receiving their last dose of JNJ-3989.

Secreted hepatitis B virus splice variants differ by HBV genotype and across phases of chronic hepatitis B infection.

Chronic hepatitis B (CHB) is characterized by progression through different phases of hepatitis B virus (HBV) infection and disease. Although not necessary for HBV replication, there is increasing evidence that HBV splice variants are associated with liver disease progression and pathogenesis. However, there have been no studies till date on the frequency or diversity of splice variants for different HBV genotypes across the phases of CHB. Next generation sequencing data from 404 patient samples of HBV genotype A, B, C or D in Phase I, Phase II or Phase IV of CHB was analysed for HBV splice variants using an in house bioinformatics pipeline. HBV splice variants differed in frequency and type by genotype and phase of natural history. Splice variant Sp1 was the most frequently detected (206/404, 51% of patients), followed by Sp13 (151/404 37% of patients). The frequency of variants was generally highest in Phase II (123/165, 75% of patients), a phase typically associated with enhanced immune activation, followed by Phase I (69/99, 70% of patients). Splice variants were associated with reduced hepatitis B e antigen (HBeAg) levels and statistically reduced likelihood of achieving HBsAg loss (functional cure) in Phase II patients for Sp1 and Sp13 (p = .0014 and .0156, respectively). The frequency of HBV splice variants in patient serum differed markedly by HBV genotype and phase of CHB natural history. The increased levels of HBV splice variants detected in CHB phase II patients compared with the higher replicative Phase I in particular warrants further investigation.

Stopping nucleot(s)ide analogues in non-cirrhotic HBeAg-negative chronic hepatitis B patients: HBsAg loss at 96 weeks is associated with low baseline HBsAg levels.

BACKGROUND AND AIMS: Current guidelines recommend long-term nucleot(s)ide analogue (NA) therapy for patients with HBeAg-negative chronic hepatitis B (CHB). However, disease remission has been described after stopping NA therapy, as well as HBsAg loss. METHODS: We performed a prospective multi-centre cohort study of stopping NA therapy. Inclusion criteria were HBeAg-negative CHB, the absence of cirrhosis and HBVDNA5× ULN occurred in 35 (32%); ALT flares were not associated with HBsAg loss. There were no unexpected safety issues. CONCLUSION: Virological reactivation was very common after stopping NA therapy and occurred earlier after stopping TDF versus ETV. The majority of patients had ALT <2× ULN at week 96, but only one-third achieved disease remission and HBsAg loss was rare. Very low HBsAg levels at baseline were uncommon but predicted for HBsAg loss and disease remission.

Role of anti-HBs in functional cure of HBeAg+ chronic hepatitis B patients infected with HBV genotype A.

BACKGROUND & AIMS: HBsAg-specific antibody responses are difficult to detect during chronic hepatitis B infection (CHB) and are often overlooked. The aim of this study was to examine whether anti-HBs may be involved in functional cure (FC) by profiling anti-HBs responses in patients with CHB using a panel of specific assays. METHODS: Longitudinal serum samples were obtained from 25 patients with CHB who were infected with HBV genotype A and were undergoing nucleos(t)ide analogue (NA) treatment: 14 achieved FC while 11 remained infected (non-FC). Anti-HBs immune complexes (HBsAg-IC), FcγRIIIa dimer binding, epitope specificity and neutralisation efficacy were measured. RESULTS: HBsAg-IC peaks were detected prior to HBsAg loss in 10/14 FC patients. These HBsAg-IC peaks overlapped with either an alanine aminotransferase (ALT) flare (8/10 patients), or a rise in ALT (2/10 patients). HBsAg-IC peaks were detected in 7/11 non-FC patients, but were not associated with an ALT flare. FCγRIIIa binding was detected in 9/14 FC patients, independent from detection of overlapping HBsAg-IC/ALT peaks. FC patients had stable HBsAg epitope occupancy across the study, whereas non-FC patients had a reduction in HBsAg epitope occupancy within the first 12-24 weeks of NA treatment. Convalescent sera from FC patients recognised more HBsAg epitopes and neutralised HBV infection more potently than anti-HBs derived from vaccinees. Neutralisation potency appeared to increase post-HBsAg loss in 4/5 FC patients examined. CONCLUSIONS: Using these assays, we confirm that anti-HBs responses are present and fluctuate over time in this cohort of patients with HBeAg+ CHB, who were infected with HBV genotype A and treated with NAs. Key anti-HBs profiles associated with either FC or failure to achieve FC were also identified, suggesting a role for anti-HBs responses in FC. LAY SUMMARY: Using a panel of assays to characterise hepatitis B surface antibody (anti-HBs) responses in a group of patients with chronic hepatitis B, we identified anti-HBs profiles associated with either functional cure, or failure to achieve functional cure. Functional cure was associated with immune complex peaks which overlapped with alanine aminotransferase flares. Conversely, in those who did not achieve functional cure, immune complex peaks were present, but were not associated with alanine aminotransferase flares, and a decline in anti-HBs diversity was observed early during treatment.

Evaluation of dried blood spots for hepatitis B and D serology and nucleic acid testing.

Areas with the highest burden of hepatitis B virus (HBV) infection are often low-middle-income countries with limited access to diagnosis due to isolation, affordability, and/or feasibility. Dried blood spots (DBSs) provide an alternative for remote areas where collection and transportation of serum is impractical. In this study, the application of DBS for serological and molecular detection of HBV and hepatitis D virus (HDV) was evaluated. Hepatitis B surface antigen was detected in 87 of 91 (95.6%) DBS. Seventeen of 21 (81%) had detectable HBeAg and 52 of 71 (73.2%) were anti-HBe positive. Anti-HD was detectable in 11 of 12 (91.6%) spiked control DBS after an initial failure to detect in patient DBS. HBV DNA was detected from 50 of 70 (71.4%) DBS with serum loads greater than 200 IU/mL in an in-house assay and 18 of 24 (75%) DBS with loads exceeding 389 IU/mL in a commercial assay. Using linear regression, HBV DNA loads from DBS were able to predict serum loads in 46 of 50 (92%) samples to within 1 log of actual serum load. HDV RNA was detected in 42 of 47 (89%) DBS with serum levels greater than 7200 IU/mL. DBSs are recommended for diagnosis of HBV, monitoring, and detection of high loads in pregnant women where peripheral blood testing remains unfeasible. Detection of HDV RNA from DBS may prove useful in endemic areas.

Long-term serological, virological and histological responses to RNA inhibition by ARC-520 in Chinese chronic hepatitis B patients on entecavir treatment.

OBJECTIVE: We examined the serological, virological (in serum and liver) and histological profiles in chronic hepatitis B virus (HBV) patients during and after completion of multiple dose (MD) ARC-520. DESIGN: The present phase 1b study was a multidose, open-label extension cohort of patients that had received single dose ARC-520 in our previous study. Eight patients received 4-9 4 weekly doses of MD ARC-520 and entecavir. Liver biopsies were performed in six patients. Intrahepatic and serum HBV DNA, HBV RNA and viral antigens were measured. RESULTS: All patients had 28.9-30.4 months of follow-up after the last MD. All three hepatitis B e antigen (HBeAg)-positive patients had profound reductions in hepatitis B surface antigen (HBsAg), HBeAg, hepatitis B core-related antigen and HBV RNA with two undergoing HBeAg seroconversion. One further achieved HBsAg seroconversion (anti-HBs level of 25.1 IU/L) and the remaining two had HBsAg reductions of -1.7 and -3.5 log IU/mL >30 months after MD. Among the five HBeAg-negative patients, four had modest HBsAg reduction >29 months after completion of MD and one achieved HBsAg seroconversion (anti-HBs level of 152.5 IU/L) and was negative for liver HBsAg staining. Entecavir was successfully stopped in this patient 12 months after HBsAg seroconversion. Temporally related alanine aminotransferase elevations preceded by HBsAg reductions were observed in three patients suggesting immune activation. HBcAg staining was negative in all six biopsied patients. Two patients with <10% HBsAg positive staining of hepatocytes had correspondingly low serum HBsAg levels of 1.5 and 11.5 IU/mL. CONCLUSIONS: MD ARC-520 therapy achieved sustained and profound reductions of viral antigens and HBV RNA. HBsAg seroclearance was achievable. TRIAL REGISTRATION NUMBER: NCT02065336.

Origins and Evolution of the Primate Hepatitis B Virus.

Recent interest in the origins and subsequent evolution of the hepatitis B virus (HBV) has strengthened with the discovery of ancient HBV sequences in fossilized remains of humans dating back to the Neolithic period around 7,000 years ago. Metagenomic analysis identified a number of African non-human primate HBV sequences in the oldest samples collected, indicating that human HBV may have at some stage, evolved in Africa following zoonotic transmissions from higher primates. Ancestral genotype A and D isolates were also discovered from the Bronze Age, not in Africa but rather Eurasia, implying a more complex evolutionary and migratory history for HBV than previously recognized. Most full-length ancient HBV sequences exhibited features of inter genotypic recombination, confirming the importance of recombination and the mutation rate of the error-prone viral replicase as drivers for successful HBV evolution. A model for the origin and evolution of HBV is proposed, which includes multiple cross-species transmissions and favors subsequent recombination events that result in a pathogen and can successfully transmit and cause persistent infection in the primate host.

Molecular Phylogenetics of Hepatitis D Virus in New Zealand and the Implications for Pacific Island Countries.

Hepatitis delta virus (HDV) is considered a satellite virus that requires hepatitis B virus surface antigen for infectivity. HDV is endemic in some Pacific Island (PI) countries, including Kiribati and Nauru, with a unique genotype 1, "Pacific clade." The aims of this study were to determine the HDV genotypes in New Zealand and investigate the link of strains to other PI countries and the rest of the world through phylogenetics. Sequencing and phylogenetic analyses were performed on 16 HDV-positive serum samples from 14 individuals collected between 2009 and 2014 at Auckland Hospital. Thirteen of 14 strains were confirmed as genotype 1 and 1 was genotype 5. Eleven of the 13 genotype 1 strains clustered with the Pacific clade. These were isolated from subjects born in Samoa, Kiribati, Tuvalu, and Niue. Another genotype 1 strain isolated from a Maori health-care worker clustered most closely with a European strain. There was an African genotype 1 and genotype 5 from African-born subjects with HIV coinfection. This study supports the probable transmission of HDV Pacific clade around the PI from Micronesia to Polynesia. The data also confirm the need to screen hepatitis B surface antigen-positive individuals for HDV.

Analysis of Hepatitis B Virus Haplotype Diversity Detects Striking Sequence Conservation Across Genotypes and Chronic Disease Phase.

BACKGROUND AND AIMS: We conducted haplotype analysis of complete hepatitis B virus (HBV) genomes following deep sequencing from 368 patients across multiple phases of chronic hepatitis B (CHB) infection from four major genotypes (A-D), analyzing 4,110 haplotypes to identify viral variants associated with treatment outcome and disease progression. APPROACH AND RESULTS: Between 18.2% and 41.8% of nucleotides and between 5.9% and 34.3% of amino acids were 100% conserved in all genotypes and phases examined, depending on the region analyzed. Hepatitis B e antigen (HBeAg) loss by week 192 was associated with different haplotype populations at baseline. Haplotype populations differed across the HBV genome and CHB history, this being most pronounced in the precore/core gene. Mean number of haplotypes (frequency) per patient was higher in immune-active, HBeAg-positive chronic hepatitis phase 2 (11.8) and HBeAg-negative chronic hepatitis phase 4 (16.2) compared to subjects in the "immune-tolerant," HBeAg-positive chronic infection phase 1 (4.3, P< 0.0001). Haplotype frequency was lowest in genotype B (6.2, P< 0.0001) compared to the other genotypes (A = 11.8, C = 11.8, D = 13.6). Haplotype genetic diversity increased over the course of CHB history, being lowest in phase 1, increasing in phase 2, and highest in phase 4 in all genotypes except genotype C. HBeAg loss by week 192 of tenofovir therapy was associated with different haplotype populations at baseline. CONCLUSIONS: Despite a degree of HBV haplotype diversity and heterogeneity across the phases of CHB natural history, highly conserved sequences in key genes and regulatory regions were identified in multiple HBV genotypes that should be further investigated as targets for antiviral therapies and predictors of treatment response.

Hepatitis B related dilemmas in the renal unit.

Testing for hepatitis B in dialysis patients is routine, but newer and more sensitive detection methods mean that there is sometimes confusion around viral loads and occult infection. There are frequently difficult choices surrounding isolation and treatment. Here we describe the use of HBV serology and DNA testing in decisions around patients with end-stage renal disease. We also suggest isolation decisions based on our current understanding of the virus and its infectivity.

Real-world application of the Xpert® HBV viral load assay on serum and dried blood spots.

As we strive towards the WHO goal of elimination of viral hepatitis as a public health threat by 2030, implementation of reliable, accurate diagnostic assays is crucial to identify those at risk of disease progression and those at risk of transmission. Ironically those at greatest risk of chronic hepatitis B are often in resource-poor regions with limited access to testing, collection, storage, and/or transportation of peripheral blood. The Xpert® HBV Viral Load assay provides an easy to use, convenient means of measuring load on GeneXpert platforms. In this study, the Xpert assay is evaluated against four commercially available high-throughput assays for Hepatitis B virus (HBV) loads. In addition application of dried blood spots (DBS) for estimation of viral load is assessed on real-world samples collected from a remote Pacific Island, Kiribati. A total of 107 serum/plasma samples were tested in the Xpert HBV load assay and compared with the Abbott m2000, Alinity m, and Roche Cobas CAP/CTM and 6800. Fifty-three DBS were tested in the Xpert assay and compared with matching serum samples. Overall 82% serum/plasma samples demonstrated good correlation between the Xpert and Roche and Abbott assays, to within 0.5 log10 IU/ml. The greatest discrepancies were seen at the limits of quantification of all assays. About 85.4% DBS gave estimable viral loads to within 1 log10 IU/ml of the serum load. The Xpert HBV viral load assay is recommended for all settings but particularly useful for resource-poor settings. Utility of DBS with the Xpert assay provides a simple means for testing in remote settings.

Hepatitis B and D in the Pacific Islands of Kiribati.

BACKGROUND: Historical reports indicate that hepatitis B and hepatitis D are highly endemic in the Pacific Island of Kiribati but current levels are unknown. OBJECTIVES: To determine current prevalence of HBV and HDV in Kiribati, characterize the strains in both mono-infection and co-infection and assess individuals for antiviral therapy. STUDY DESIGN: Sera obtained from 219 patients were screened for HBsAg, HBeAg, HBV DNA, anti-HD, and HDV RNA. 61 HBV isolates were sequenced for genotype, phylogenetic analysis and detection of pre-core and basal core promoter mutations. 82 HDV isolates were also sequenced. RESULTS: 55.7 % HBsAg positive samples had antibodies to HDV and 73.2 % had detectable HDV RNA, indicating that 40.8 % HBsAg-positive individuals had current HBV/HDV co-infection. There were 42 co-infected males and 40 females; the youngest individual was a 4 year-old boy. HBV isolates were genotype D4, and HDV strains formed a distinct Pacific clade of genotype 1. Undetectable HBV DNA loads were statistically more frequent in the co-infected sub-population (p < 0.0001). Basal core promoter and pre-core mutations were present in both mono and co-infection. CONCLUSION: Kiribati has one of the highest HBV/HDV co-infection rates in the world. The epidemiology of co-infection in this population was unusual with males and females equally represented and the presence of co-infection in a 4 year old child suggesting neonatal or early horizontal transmission, which is extremely rare. Coinfection with HDV resulted in statistically significant suppression of HBV DNA levels. The HDV strain identified in Kiribati was unique to the Pacific Islands.

HBV variants are common in the 'immune-tolerant' phase of chronic hepatitis B.

Nucleos(t)ide analogues (NUC) treatment prevents progression of liver fibrosis in subjects with chronic hepatitis B (CHB). However, risk of hepatocellular carcinoma (HCC) persists despite viral suppression. Specific HBV variants have been associated with adverse outcomes, including HCC; however, the frequency of these variants during the seemingly benign immunotolerant (IT) phase is unknown. Next-generation sequencing and detailed virological characterization on a cohort of treatment-naïve IT subjects were performed to determine the frequency of clinically relevant viral variants. Samples from 97 subjects (genotype B/C 55%/45%, median HBV-DNA 8.5 log10 IU/mL, median HBsAg 4.8 log10  IU/mL, median HBeAg 3.6 log10  PEIU/mL) were analysed. Despite subjects being in the IT phase, clinically relevant HBV variants were common at baseline, particularly in the basal core promoter (BCP, overlaps the hepatitis B X (HBx) gene), precore and PreS regions. BCP/HBx variants were independently associated with lower baseline HBeAg, HBsAg and HBV-DNA titres. Precore variants were independently associated with higher baseline ALT. Increased viral diversity was associated with increased age and lower HBV-DNA, HBsAg and HBeAg levels. Low-level (<5%) drug resistance-associated amino acid substitutions in the HBV reverse transcriptase were detected in 9 (9%) subjects at pre-treatment but were not associated with reduced antiviral activity. Future studies should evaluate whether the detection of HBV variant during IT CHB is predictive of progression to immune clearance and poor prognosis, and whether early initiation of antiviral therapy during IT CHB to prevent the selection of HBV variants is clinically beneficial.