Hepatitis Delta Virus Clade 8 Is the Predominant Clade Circulating in Botswana amongst People Living with HIV.

Hepatitis delta virus (HDV) co-infections more often result in severe hepatitis compared to hepatitis B virus (HBV) infections alone. Despite a high HDV prevalence (7.1%), information regarding circulating HDV clades is very limited in Botswana. We extracted total nucleic acid from confirmed HDV-positive samples and quantified their viral load. We then sequenced the large hepatitis delta antigen (L-HDAg) using Oxford Nanopore Technology (ONT). Genotyping was performed using the HDV Database, and HDV mutation profiling was performed on AliView. All participants with HBV genotypic information belonged to sub-genotype A1, and 80% (4/5) of them had a higher HDV viral load and a lower HBV viral load. We sequenced 75% (9/12) of the HDV-positive samples, which belonged to HDV clade 8. A total of 54 mutations were discovered, with the most prevalent being Q148R (16%), D149P (16%) and G151D (16%). Known mutations such as S117A, K131R, R139K and G151D were detected, while the other mutations were novel. Our results reveal that HDV clade 8 is the predominant clade in Botswana. The significance of all mutations remains unclear. Future studies with a larger sample size to detect other HDV clades that might be circulating in Botswana and functionally characterize the detected mutations are warranted.

An hepatitis B and D virus infection model using human pluripotent stem cell-derived hepatocytes.

Current culture systems available for studying hepatitis D virus (HDV) are suboptimal. In this study, we demonstrate that hepatocyte-like cells (HLCs) derived from human pluripotent stem cells (hPSCs) are fully permissive to HDV infection across various tested genotypes. When co-infected with the helper hepatitis B virus (HBV) or transduced to express the HBV envelope protein HBsAg, HLCs effectively release infectious progeny virions. We also show that HBsAg-expressing HLCs support the extracellular spread of HDV, thus providing a valuable platform for testing available anti-HDV regimens. By challenging the cells along the differentiation with HDV infection, we have identified CD63 as a potential HDV co-entry factor that was rate-limiting for HDV infection in immature hepatocytes. Given their renewable source and the potential to derive hPSCs from individual patients, we propose HLCs as a promising model for investigating HDV biology. Our findings offer new insights into HDV infection and expand the repertoire of research tools available for the development of therapeutic interventions.

[Chronic HBV and HDV infection]. / Chronische Hepatitis-B- und Hepatitis-D-Virusinfektion.

About 0,5% of the population in Germany has a chronic hepatitis B virus (HBV) infection. Untreated, chronic HBV infection can progress to liver cirrhosis and hepatocellular carcinoma (HCC). If diagnosed early, antiviral therapy can effectively prevent liver disease progression, but a cure is currently hardly achievable. About 5% of those chronically infected with HBV are also co-infected with the hepatitis D virus (HDV). HBV/HDV co-infection leads to liver cirrhosis in approximately 50% of patients within 5-10 years. Since 2020, the cell entry inhibitor bulevirtide is available as a specific therapy for HBV/HDV co-infection.

Enhancing linkage to care for hepatitis B, D, and C patients: A retrospective-prospective study.

BACKGROUND: The WHO has set a goal to decrease viral hepatitis-related fatalities by 65% by 2030. AIMS: To locate and retrieve to care all individuals diagnosed with hepatitis B, D or C, and investigate why they were not linked to appropriate medical management. METHODS: We conducted a retrospective-prospective search for patients with hepatitis B, D or C virus (HBV, HDV and HCV) infection in the central laboratory database of the Barcelona northern health area (catchment population, 450,000). We reviewed records and contacted candidates for retrieval who were offered a specialist medical visit. RESULTS: We reviewed records of 3731 patients with viral hepatitis (January 2019-December 2022): 1763 HBsAg+, 69 anti-HDV+ and 1899 HCV-RNA+. Among these, 644 (37%) HBV, 20 (29%) HDV and 1116 (56%) HCV patients were not currently linked to care. The proportion of patients receiving appropriate care was higher in HBV and HDV (p < 0.0001), and a higher percentage of unlinked hepatitis C patients had low life expectancy/comorbidities (39%; p < 0.0001). After implementing the linkage strategy, 254 HBV, 16 HDV and 54 HCV patients were successfully reintegrated into care. Among 1780 patients requiring linkage, 638 (35.8%) had moved to another health area or were missing essential contact data. CONCLUSIONS: Among patients with viral hepatitis who required appropriate specialist care and were eligible for contact, 64% with HBV, 100% with HDV and 27% with HCV were successfully reintegrated into care. Overall, 324 (47.2%) eligible patients were linked to care, indicating the success of this strategy.

Impact of Hepatitis Delta Virus Infection on the Selection of Hepatitis B Surface Antigen Mutations.

The interaction of multiple viruses in one host is thought to enhance the development of mutations. However, the impact of hepatitis D virus (HDV) positivity on the development of unique hepatitis B virus (HBV) mutations among people living with human immunodeficiency virus (HIV) (PLWH) remains poorly understood in African countries, including Botswana. We used HBV sequences generated from the Botswana Combination Prevention Project (BCPP), which is the largest pair-matched cluster-randomized HIV trial in Botswana. Only participants with available HBV sequences (n = 55) were included in our study ([HIV/HBV-positive (n = 50) and HIV/HBV/HDV-positive (n = 5)]. Geno2pheno was used to determine HBV genotypes, and HBV surface region sequences (all subgenotype A1) were aligned in AliView for mutational analysis, while the impact of mutations was assessed using Phyre2. Our results identified 182 common mutations between the two groups. In the HIV/HBV/HDV cohort, only three mutations (L95W, W156Q, C221Y) were classified as deleterious, with only L95W being the most frequent. In the HIV/HBV cohort, four mutations (W199R, C221A, C221S, W223G) were also classified as deleterious. Our results demonstrate the presence of unique HBV mutations among the HIV/HBV/HDV-positive cohort. Functional characterization of these mutations is recommended to determine their effect on HDV.

The diagnostic cascade for patients with hepatitis delta infection in France, 2018-2022: A cross-sectional study.

BACKGROUND AND AIMS: Chronic hepatitis D infection is the most severe form of viral hepatitis and can rapidly progress to cirrhosis or hepatocellular carcinoma. Despite recommendations for systematic screening of hepatitis B surface antigen (HBsAg)-positive individuals, data from real-world studies have reported a low frequency of hepatitis D (or delta) virus (HDV) screening. Our cross-sectional analysis evaluated the diagnostic cascade for hepatitis D infection in tertiary centres and described the characteristics of HDV-positive patients. METHODS: A total of 6772 individuals who tested HBsAg positive for the first time between 2018 and 2022 were retrospectively included. Demographic, clinical and laboratory data were analysed. RESULTS: A total of 5748 HBsAg-positive individuals (84.9%) were screened for HDV infection. The screening rate varied from 63% to 97% according to the screening strategy used in the centres including or not HDV reflex testing. The prevalence of HDV infection was 6.3%. HDV RNA levels were determined in 285 of the 364 (78.3%) HDV antibody screening-positive patients, and 167 (58.6%) had active HDV infection. 66.8% were males, with a mean age of 44.9 years. A total of 97.5% were born abroad, and 92.9% were HBeAg negative. At the time of diagnosis, HDV RNA levels were 6.0 Log UI/mL; 60.1% had alanine aminotransferase >40 U/L, and 56.3% had significant fibrosis (≥F2), including 41.6% with cirrhosis. The most common genotype was HDV-1 (75.4%). Coinfections were not uncommon: 7.4% were HIV positive, and 15.0% were HCV antibody positive. CONCLUSIONS: The present study highlights the need for increased screening and monitoring of HDV infection. Reflex testing helps to identify HDV-infected individuals.

Hepatitis Delta Virus Surveillance Practice among Clinicians in Nigeria: A Cross-Sectional Survey.

BACKGROUND: The near total absence of routine Hepatitis Delta Virus (HDV) screening in many countries in sub-Saharan Africa is a major challenge to understanding the burden of HDV in the region. AIM: To evaluate Hepatitis Delta Virus screening practices and associated factors among clinicians in Nigeria. METHODS: A cross-sectional study was conducted in June-July 2022, in which a self-administered questionnaire that inquired about HDV awareness, screening practices, and treatment options was shared electronically with consenting clinicians practicing in Nigeria. At the end of the survey, data was analyzed using descriptive and inferential statistics. The level of significance was set at 0.05. RESULTS: At the end of the survey, 210 of the 213 responses retrieved from respondents were analyzed. The respondent's mean age was 38.60 ± 7.27 years with a male-to-female ratio of 1:2.5. They comprised 13.8% gastroenterologists and 86.2% respondents in other areas of clinical medicine. The study showed that 89.5% of the respondents knew that HDV infection occurs only in hepatitis B virus (HBV)-infected individuals. Most (91.4%) respondents do not screen for HDV in chronic HBV patients, mainly due to the non-availability of screening tools and lack of awareness of any screening test for HDV. Research interest was reported as the reason for screening among clinicians who had ever screened for HDV. Pegylated interferon was the main regimen used for treatment by 87.5% of respondents. About 2% did not know treatment options for HDV. A significant association between knowledge of HDV infection and area of specialty, as well as the nature of medical practice was noted (P = 0.008 and 0.013, respectively). CONCLUSION: The study showed a high level of awareness of HDV dependency on HBV, for natural infection to occur. However, it documented very minimal HDV screening in clinical settings and factors affecting screening among clinicians.

The Prevalence and Molecular Epidemiology of Hepatitis Delta Virus in Nigeria: The Results of a Nationwide Study.

Hepatitis delta virus (HDV) is a satellite of hepatitis B virus (HBV), which requires the HBV surface antigen (HBsAg) for its assembly and propagation. Although countries affected by HBV infection in Africa are well identified, data on HDV infection are still scarce, like in Nigeria, where HBV infection is endemic. In this study, we aimed to determine the prevalence of HDV infection and identify the circulating genotypes/strains in the country. A nationwide study was performed on 1281 HBsAg-positive samples collected from patients across eleven sites drawn from the six geopolitical zones in Nigeria. Anti-HDV antibody (HDV-Ab) screening and HDV-RNA viral load quantification were performed using a commercial ELISA assay and real-time RT-PCR kit, respectively. HDV genotyping was performed by the Sanger sequencing of amplicons from the so-called R0 region of the viral genome, followed by phylogenetic analyses. Of the 1281 HBsAg-positive samples, 61 (4.8%) were HDV-Ab positive, among which, 12 (19.7%) were HDV-RNA positive. Genotypes were obtained for nine of them: seven "African" HDV-1, one "Asian/European" HDV-1 and one HDV-6. This study shows that Nigeria is a country of low HDV prevalence where mainly "African" genotype-1 strains are circulating.

vHDvDB 2.0: Database and Group Comparison Server for Hepatitis Delta Virus.

The hepatitis delta virus (HDV) is a unique pathogen with significant global health implications, affecting individuals who are coinfected with the hepatitis B virus (HBV). HDV infection has profound clinical consequences, manifesting either as coinfection with HBV, resulting in acute hepatitis and potential liver failure, or as superinfection in chronic HBV cases, substantially increasing the risk of cirrhosis and hepatocellular carcinoma. Given the complex dynamics of HDV infection and the urgent need for advanced research tools, this article introduces vHDvDB 2.0, a comprehensive HDV full-length sequence database. This innovative platform integrates data preprocessing, secondary structure prediction, and epidemiological research tools. The primary goal of vHDvDB 2.0 is to consolidate HDV sequence data into a user-friendly repository, thereby facilitating access for researchers and enhancing the broader scientific understanding of HDV. The significance of this database lies in its potential to streamline HDV research by providing a centralized resource for analyzing viral sequences and exploring genotype-specific characteristics. It will also enable more in-depth research within the HDV sequence domains.

Assessing the diagnostic accuracy of serological tests for hepatitis delta virus diagnosis: a systematic review and meta-analysis.

Hepatitis Delta Virus (HDV), a satellite virus of Hepatitis B virus, exacerbates liver damage in affected individuals. Screening for HDV antibodies in HBsAg positive patients is recommended, but the diagnostic accuracy of serological tests remains uncertain. This review aimed to assess the diagnostic accuracy of serological tests for HDV. We searched PubMed, Web of Science, Cochrane Central Register of Controlled Trials, Scopus etc. for relevant studies. Studies measuring the sensitivity and specificity of serological HDV tests against PCR as a reference standard were included. Pooled sensitivity and specificity for each test method and sero-marker were calculated. The review included six studies with 11 study arms, evaluating ARCHITECT immunoassay, EIA, ELISA, QMAC, RIA, and Western Blot test methods targeting Anti-HDV IgG, Total anti-HDV and Anti-HDV IgM. Sensitivities for Anti-HDV IgG, Total Anti-HDV and Anti-HDV IgM, tests were 97.4%, 51.9%, and 62.0%, respectively, with specificities of 95.3%, 80.0%, and 85.0%. Our findings, with its limited number of studies, suggest that HDV serological tests, particularly those identifying Anti IgG exhibit high accuracy and can serve as effective screening tools for HDV.

Interferon alpha induces a stronger antiviral effect than interferon lambda in HBV/HDV infected humanized mice.

Recent studies indicate that treatment of chronic hepatitis D virus (HDV) with either pegylated interferon (IFN)λ or pegylated IFNα monotherapy leads to a dramatic decline in HDV RNA. Herein, we investigated the innate antiviral efficacy of IFNλ and IFNα in humanized mice that lack an adaptive immune response. Humanized mice were either co-infected with hepatitis B virus (HBV) and HDV simultaneously, or HDV infection was performed subsequent to HBV infection (i.e., superinfected). After steady viral replication was achieved, mice received either IFNλ (n = 6) or IFNα (n = 7) for 12 (or 13) weeks. Pretreatment median levels of serum HBV DNA (8.8 [IQR:0.2] log IU/ml), HDV RNA (9.8 [0.5] log IU/ml), HBsAg (4.0 [0.4] log IU/ml) and human albumin, hAlb (6.9 [0.1] log ng/mL) were similar between mice treated with IFNα or IFNλ and between those coinfected versus superinfected. Compared to mice treated with IFNλ, mice treated with IFNα had a significantly greater decline in HBV, HDV, and HBsAg levels. In conclusion, IFNα induces stronger inhibition of HBV and HDV than IFNλ in humanized mice that lack an adaptive immune response. Further studies are needed to assess the respective role of the combined innate-and adaptive-immune systems in the treatment of HBV and HDV with IFNα and IFNλ.

Outcomes of a 1-Year Pilot Study Implementing Universal Hepatitis Delta Virus Testing Among Veterans With Chronic Hepatitis B.

Current US guidelines recommend risk-based testing for hepatitis delta virus (HDV) in persons with chronic hepatitis B (CHB). While there is debate as to whether a risk-based or universal testing approach is most effective, limited data exist on universal HDV testing programs in the United States. We performed a 1-year pilot study evaluating the outcomes of a universal HDV testing approach among US veterans with CHB. All consecutive adults with CHB receiving care at hepatology clinics at a single-centre Veterans Affairs Health System from 1 October 2022 to 30 September 2023 were prospectively tested for anti-HDV antibody (anti-HDV). Patients who were anti-HDV Ab-positive were subsequently tested for HDV RNA. Comparison of HDV testing between groups utilised chi-square testing. A total of 91 consecutive CHB patients (90.0% male, mean age 60.9 ± 14.1 years, 73.9% Asian, 26.1% non-Asia, 16.5% cirrhosis and 17.1% with active or past history of drug use) had anti-HDV ordered. Overall, 76.9% (n = 70) completed anti-HDV testing; 4.3% (n = 3) were positive. HDV RNA testing was ordered in all three patients; two patients completed HDV RNA testing and one had detectable HDV RNA. No significant differences in completion of anti-HDV testing was observed by age, sex, race/ethnicity, cirrhosis status or drug use history. Among a single-centre prospective cohort study piloting a universal HDV testing approach, one patient with viremic HDV was identified. Implementing true reflex testing of all CHB patients with anti-HDV, followed by automated HDV RNA testing for anti-HDV-positive patients would improve the HDV testing cascade and timely diagnosis of HDV.

Clinical Prevalence of Hepatitis D Virus Among Hepatitis B Patients in Sulaymaniyah Governorate, Northern Iraq.

Hepatitis D virus (HDV), which occurs as a co-infection with the hepatitis B virus (HBV), is a significant public health burden. Currently, there is a scarcity of data regarding this co-infection in the developing countries. This study aims to address the clinical prevalence of HDV among HBV-infected patients in Sulaymaniyah Governorate, Iraq. This prospective cross-sectional study, conducted from May to November 2022, screened HBV DNA-positive patients visiting Sulaimani Teaching Hospital in Sulaymaniyah governorate, Iraq, for anti-HDV antibodies and HDV RNA. The study included 150 confirmed HBV DNA-positive patients. Of these, 54.7% were male. The mean age of the patients was 49.1 ± 10.1 (18-68). Serological assessment found that 23 (15.3%) of the patients had anti-HDV IgG antibodies, suggesting past or chronic HDV infection, while 16 (10.7%) tested positive for anti-HDV IgM, indicating recent/acute infection. Further molecular analysis confirmed HDV RNA in 15 (10%) of HBV patients, indicating real HDV prevalence. The prevalence of anti-HDV and HDV RNA did not significantly differ by age, gender, marital status, residency, medical, family or medical history (p > 0.05). In conclusion, this study demonstrated a relatively high HDV prevalence among HBV patients in Sulaymaniyah Governorate, Iraq, at 10%, which stresses the need for better screening, health strategies and focused research to combat its impact.

Prevalence of Hepatitis D in People Living with HIV: A National Cross-Sectional Pilot Study.

This study assesses the prevalence of hepatitis D virus (HDV) in people living with HIV (PLWHIV) in Greece. Given the compounding effects of HDV and hepatitis B (HBV) on liver disease progression, as well as the emergence of new therapeutic options such as bulevirtide, understanding regional disparities and the epidemiological impact of such co-infections is vital. A cross-sectional analysis was conducted utilizing 696 serum samples from PLWHIV attending five major university hospitals. The methodology included HDV antibody detection by ELISA and HDV RNA confirmation. Of the 30 HBsAg-positive samples analyzed, the study population was primarily male (93%), with a median age of 54 years. Participants had been on antiretroviral therapy for a median of 10 years, and the median CD4 count was 738 (539-1006) copies/mL. Additional serological findings revealed a 7% prevalence of hepatitis C virus (HCV) IgG antibodies and a 55% prevalence of hepatitis A virus (HAV) IgG antibodies. Seroreactivity for syphilis (RPR/VDRL/TPHA positive) was identified in 33% of the participants. The results indicated a low HDV prevalence, with only one individual (3%) testing positive for anti-HDV IgG antibodies and none for HDV RNA. This indicates a lower prevalence of HDV among PLWHIV with chronic HBV in Greece compared to global data.

Assessment of performance and comparison of three commercial HDV RNA detection assays: implications for diagnosis and treatment monitoring.

Objectives: Precise HDV-RNA detection and quantification are pivotal for diagnosis and monitoring of response to newly approved treatment. We evaluate the performance of three HDV RNA detection and quantification assays. Methods: Hepatitis Delta RT-PCR system kit, EurobioPlex HDV assay, and RoboGene HDV RNA Quantification kit 2.0 were used for testing 151 HBsAg-positive samples, 90 HDV-RNA negative and 61 HDV-RNA positive. We also evaluated serial dilutions of the WHO international standard for HDV, PEI 7657/12. All HDV-RNA positive samples were genotyped using a next-generation sequencing strategy. Results: Qualitative results indicated a 100% concordance between tests. Quantitative results correlated well, r2 = 0.703 (Vircell-vs-Eurobio), r2 = 0.833 (Vircell-vs-RoboGene), r2 = 0.835 (Robogene-vs-Eurobio). Bias index was 2.083 (Vircell-vs-Eurobio), -1.283 (Vircell-vs-RoboGene), and -3.36 (Robogene-vs-Eurobio). Using the WHO IS, Vircell overestimated the viral load by 0.98 log IU/mL, Eurobio by 1.46 log IU/mL, and RoboGene underestimated it by 0.98 log IU/mL. Fifty-nine samples were successfully genotyped (Genotype 1, n=52; Genotype 5, n=7; Genotype 6, n=1), with similar results for correlation and bias. Conclusion: This study underscores the necessity of using reliable HDV-RNA detection and quantification assays, as evidenced by the high concordance rates in qualitative detection and the observed variability in quantitative results. These findings highlight the importance of consistent assay use in clinical practice to ensure accurate diagnosis and effective treatment monitoring of HDV infection.

Method for Quantitative HDV RNA Detection: I, Manual Workflow (Serum and Liver Tissue) and II, Fully Automated High Throughput Workflow for Diagnostic Use.

The hepatitis delta virus (HDV) is a small RNA virus (1700 base pairs), which uses the surface proteins of the hepatitis B virus (HBV) as an envelope. Accurate and reliable quantitative detection of HDV RNA is central for scientific and translational clinical research or diagnostic purposes. However, HDV poses challenges for nucleic acid amplification techniques: (1) the circular genome displays high intramolecular base pairing; (2) high content of cytosine and guanine; and (3) enormous genomic diversity among the eight known HDV genotypes (GTs). Here, we provide step-by-step instructions for (A) a manual workflow to perform a quantitative HDV reverse transcription (RT)-PCR from serum and liver tissue and (B) a quantitative HDV RT-PCR assay with whole process control to be used for serum or plasma samples run on a fully automated system. Both assays target the conserved ribozyme region and demonstrate inclusivity for all eight HDV GTs. The choice of assay depends on the experimental needs and equipment availability. While the former is ideal for scientific research laboratories, the latter provides a useful tool in the field of translational research or diagnostics.

[Seroprevalence and Risk Factors of Hepatitis B, C and D in Adults in Trabzon, Türkiye]. / Trabzon'da 20 Yas Üstü Eriskinlerde Hepatit B, Hepatit C ve Hepatit D Seroprevalansi ve Risk Faktörleri.

Viral hepatitis are infections that can cause liver damage, become chronic, lead to cirrhosis, hepatocellular carcinoma and ultimately result in death due to their ability to spread in the community through blood and infected body fluids. The aim of this study was to determine the seroprevalence of hepatitis B (HBV), hepatitis C (HCV), and hepatitis D (HDV) transmitted through blood among individuals living in Trabzon province and to examine the factors potentially associated with seroprevalence. This cross-sectional study was conducted in Trabzon province, located in the northeast of Türkiye, including a total of 10 districts, including the central district. Since seroprevalence was calculated for HBV, HCV, and HDV in the study, the sample size was separately calculated for each, and the calculated maximum sample size of 1116 was accepted as the minimum sample size for the study. The study was completed with 1502 participants. Serological tests for HBV included HBsAg, anti-HBs, and anti-HBc IgG; for HCV, anti-HCV; and for HDV, anti-HDV were analysed. Data were evaluated for HBV risk factors using univariate analyses with Chi-square test and for multiple analyses using enter model logistic regression analysis. The mean age of the participants was 45.7 ± 16.6 years, with 767 (51.1%) being female. The prevalence of HBV seropositivity, indicating vaccination, was 23.0%, while the seroprevalence of HBV among unvaccinated adults was 27.4%. HBsAg positivity was 5.1%, and isolated anti-HBc IgG positivity was 4.2%. The proportion of individuals with HBsAg in the gray zone was 0.5%, while the positivity rates for anti-HBs and anti-HBc IgG (indicating past infection) were 17.6%. The prevalence of anti-HCV was six per thousand, while anti-HDV was not detected in the analyses. HBsAg positivity and co-infection with HCV were found in one person, and among the nine individuals positive for anti-HCV, isolated anti-HBc IgG positivity was detected in three. Increasing age, presence of a person with jaundice in the family, presence of diabetes mellitus, alcohol use and cupping therapy were identified as risk factors for HBV in the logistic regression analysis. Risk factors for HCV in univariate analyses were being over 40 years old, presence of hepatic steatosis and receiving dialysis treatment. The results of the study indicate that despite being included in our vaccination schedule and the administration of vaccines to high-risk adults, HBV still requires intensive attention as a public health problem. HCV, lacking a vaccine has been evaluated as an infectious agent that needs to be taken into consideration due to its potential risks and requires the complete implementation of individual and social precautions.

Seroprevalence of hepatitis D virus in Bulgarian outpatients with history of liver dysfunction.

The aim of this research was to define the prevalence of antibodies against hepatitis D virus (anti-HDV Ab) in a group of 26 outpatients with liver dysfunction in northeastern Bulgaria. Serum samples were obtained from April 2022 to December 2023 in the "Status" Medical Diagnostic Laboratory, Varna, Bulgaria. We found seroprevalence of anti-HDV Ab in 15.4% (CI: 4.3-34.8%) of the target population. Age and gender had no significant role in HDV seropositivity.