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Background: Although nucleos(t)ide analogues (NAs) are thought to reduce the risk of hepatitis B virus (HBV)‐related hepatocellular carcinoma (HCC), the effect of NA discontinuation on the prognosis of HBV‐related HCC after hepatectomy is rarely reported. We aimed to investigate the potential for hepatitis B virus e antigen (HBeAg)‐negative HBV‐related HCC patients to discontinue NAs based on preoperative hepatitis B virus surface antigen (HBsAg) status. Methods: This historical cohort study involved 1232 NA‐treated HBeAg‐negative patients who underwent curative hepatectomy for HBV‐related HCC from 2014 to 2019. The recurrence‐free survival (RFS) and overall survival (OS) of patients discontinuing NAs before surgery were compared with those continuing NAs. Propensity score matching (PSM) was used to balance baseline characteristics. Results: Of all enrolled patients, 839 (68.1%) patients continued NAs, and 393 (31.9%) patients discontinued NAs. Continuation of NAs was identified as an independent risk factor for RFS (HR 2.047, 95% CI 1.348–3.109, p < 0.001 before PSM and HR 2.756, 95% CI 1.537–4.942, p < 0.001 after PSM) in HBsAg‐negative patients. Similarly, subgroup survival analyses showed that NA discontinuation was associated with better RFS (p = 0.029 before PSM and p < 0.001 after PSM) and comparable OS (p = 0.935 before PSM and p = 0.115 after PSM) than NA continuation in HBsAg‐negative patients. The interaction between HBsAg status and continuation or discontinuation of NAs was significant (p for interaction <0.001). Conclusions: These findings demonstrate the potential for HBeAg‐negative HBV‐related HCC patients who have achieved HBsAg seroclearance to discontinue NAs under strict monitoring. [ABSTRACT FROM AUTHOR]

A highly sensitive and reliable Hepatitis B virus surface antigen (HBsAg) measurement is essential to universal screening, timely diagnosis, and management of Hepatitis B virus (HBV) infection. This study aimed to evaluate the performance of MAGLUMI HBsAg chemiluminescence immunoassay (CLIA). MAGLUMI HBsAg (CLIA) was compared against ARCHITECT HBsAg. 411 HBsAg positive samples, including different stages of infection, genotypes, subtypes, mutants, and 30 seroconversion panels were tested to evaluate diagnostic sensitivity. Diagnostic specificity was evaluated by testing 205 hospitalized samples and 5101 blood donor samples. Precision, limit of blank (LoB), limit of detection (LoD), and linearity were also verified. The diagnostic sensitivity of the MAGLUMI HBsAg (CLIA) was 100% with better seroconversion sensitivity than ARCHITECT HBsAg. The MAGLUMI HBsAg (CLIA) has optimal detection efficacy for HBV subgenotypes samples. The analytical sensitivity is 0.039 IU/mL. The initial diagnostic specificity is 99.63% on blood donors and 96.59% on hospitalized samples. The verification data demonstrated high repeatability, a LoB of 0.02 IU/mL, LoD of 0.05 IU/mL and an excellent linearity of 0.050–250 IU/mL (R2 = 0.9946). The MAGLUMI HBsAg (CLIA) is proved a highly sensitive and reliable assay with optimal subgenotype detection efficacy. [ABSTRACT FROM AUTHOR]

Abstract Background Although nucleos(t)ide analogues (NAs) are thought to reduce the risk of hepatitis B virus (HBV)‐related hepatocellular carcinoma (HCC), the effect of NA discontinuation on the prognosis of HBV‐related HCC after hepatectomy is rarely reported. We aimed to investigate the potential for hepatitis B virus e antigen (HBeAg)‐negative HBV‐related HCC patients to discontinue NAs based on preoperative hepatitis B virus surface antigen (HBsAg) status. Methods This historical cohort study involved 1232 NA‐treated HBeAg‐negative patients who underwent curative hepatectomy for HBV‐related HCC from 2014 to 2019. The recurrence‐free survival (RFS) and overall survival (OS) of patients discontinuing NAs before surgery were compared with those continuing NAs. Propensity score matching (PSM) was used to balance baseline characteristics. Results Of all enrolled patients, 839 (68.1%) patients continued NAs, and 393 (31.9%) patients discontinued NAs. Continuation of NAs was identified as an independent risk factor for RFS (HR 2.047, 95% CI 1.348–3.109, p

Though the world-wide hepatitis B virus (HBV) vaccination program has been well completed for almost thirty years in many nations, almost HBV-related hepatocellular carcinoma (HCC) occurs in unvaccinated middle-aged and elderly adults. Apparently, treating 80% of qualified subjects could decrease HBV-related mortality by 65% in a short period. Nevertheless, globally, only 2.2% of CHB patients undergo antiviral therapy. The HBV markers related to HCC occurrence and prevention are as follows: the HCC risk is the highest at a baseline of HBV DNA of 6–7 log copies/mL, and it is the lowest at a baseline of an HBV DNA level of >8 log copies/mL and ≤4 log copies/mL (parabolic, and not linear pattern). The titer of an HBV core-related antigen (HBcrAg) reflecting the amount of HBV covalently closed circular DNA (ccc DNA) in the liver is related to HCC occurrence. The seroclearance of HBs antigen (HBsAg) is more crucial than HBV DNA negativity for the prevention of HCC. In terms of the secondary prevention of hepatitis B-related HCC involving antiviral therapies with nucleos(t)ide analogues (NAs), unsolved issues include the definition of the immune-tolerant phase; the optimal time for starting antiviral therapies with NAs; the limits of increased aminotransferase (ALT) levels as criteria for therapy in CHB patients; the normalization of ALT levels with NAs and the relation to the risk of HCC; and the relation between serum HBV levels and the risk of HCC. Moreover, the first-line therapy with NAs including entecavir (ETV), tenofovir disoproxil fumarate (TDF), and tenofovir alafenamide (TAF) remains to be clarified. Discussed here, therefore, are the recent findings of HBV markers related to HCC occurrence and prevention, unsolved issues, and the current secondary antiviral therapy for the prevention of HBV-related HCC. [ABSTRACT FROM AUTHOR]

Hepatitis delta virus (HDV) coinfected with HBV causes severe viral hepatitis, however, the number of HDV infection may be underestimated. In the present study, we enrolled 1,141,331 blood donations, routinely tested for HBsAg and/ or HBV DNA, from 21 blood establishments in China. 2,690 donors were HBsAg and/or HBV DNA positive after screening tests. After verification of HBsAg and HBV DNA, 1,490 samples were HBsAg confirmed-positive, including 1,459 HBV DNA-positive samples, and 825 samples were seronegative but HBV DNA positive. We first analyzed demographic characteristics of involved 2,690 donors with different HBV infection status and found the proportions of males, the older donors, workers and farmers were higher in HBsAg-/HBV DNA+ group. Then we evaluated specificity of HDV IgG and IgM antibody assays with 375 HBsAg and HBV DNA confirmed-negative samples, and 374 were tested negative using the two assays, respectively, suggesting a specificity of 99.73% for both assays (374/375, 95% Cl: 98.51–99.95%). Subsequently, we tested for HDV IgG and IgM of 2,315 HBsAg and/or HBV DNA confirmed-positive samples, and nine showed reactivity for IgG, while two were reactive for IgM. All these 11 reactive samples were tested again with another HDV pan-Ig and IgM testing assays and HDV RNA, and only one donor was identified as HDV IgG positive and HDV RNA negative, showing an HDV seroprevalence of 0.067% (95%CI: 0.012–0.38%) among HBsAg-positive blood donors in China. The positive donor was followed up for 2 years after the donation date, and decreased antibody titer of HDV IgG and HBsAg conversion were observed, and the infection status of the donor was HDV infection with recovery and occult hepatitis B virus infection with genotype C2. These results indicated a low seroprevalence of HDV infection among blood donors and a low risk of HDV transmission through blood transfusion in China. [ABSTRACT FROM AUTHOR]

ABSTRACT Monocytes play an important role in the control of microbial infection, but monocyte biology during chronic hepatitis B virus (HBV) infection (CHI) remains inadequately studied. We investigated the frequency, phenotype, and functions of monocyte subsets in different phases of CHI, namely, immune tolerance (IT), hepatitis B early antigen (HBeAg)-positive/HBeAg-negative chronic hepatitis B (EP-/EN-CHB, respectively), and inactive carrier (IC), identified factors responsible for their functional alterations, and determined the impact of antiviral therapy on these cells. Flow cytometric analysis indicated that HLA-DR+ CD14++ CD16− classical monocytes were significantly reduced while HLA-DR+ CD14++ CD16+ intermediate and HLA-DR+ CD14+ CD16++ nonclassical monocytes were expanded in IT and EP-/EN-CHB compared with those in IC and healthy controls (HC). In comparison to IC/HC, monocytes in IT and CHB exhibited diminished expression of Toll-like receptor 2 (TLR-2)/TLR-4/TLR-9 and cytokines interleukin-12 (IL-12)/tumor necrosis factor alpha (TNF-α)/IL-6 but produced higher levels of IL-10/transforming growth factor β (TGF-β). Further, monocytes in CHB/IT showed impaired phagocytosis and oxidative response relative to those in IC/HC. In vitro assays indicated that high titers of hepatitis B surface antigen (HBsAg) present in IT/CHB and of IL-4 in CHB triggered the functional defects in monocytes via induction of β-catenin. Additionally, monocyte-derived M1 macrophages of CHB/IT produced fewer proinflammatory and more anti-inflammatory cytokines than those of IC/HC, while in CHB/IT, the monocytes skewed the differentiation of CD4+ T cells more toward regulatory T cells and a Th2 phenotype. Moreover, monocytes in CHB and IT overexpressed chemokine receptor CCR2, which coincided with increased intrahepatic accumulation of β-catenin+ CD14+ cells. One year of tenofovir therapy failed to normalize monocyte functions or reduce serum HBsAg/IL-4 levels. Taken together, monocytes are functionally perturbed mostly in IT and EP-/EN-CHB phases. Targeting intramonocytic β-catenin or reducing HBsAg/IL-4 levels might restore monocyte function and facilitate viral clearance. IMPORTANCE Chronic HBV infection (CHI) is a major cause of end-stage liver disease for which pharmacological treatments currently available are inadequate. Chronically HBV-infected patients fail to mount an efficient immune response to the virus, impeding viral clearance and recovery from hepatitis. Monocytes represent a central part of innate immunity, but a comprehensive understanding on monocyte involvement in CHI is still lacking. We here report a multitude of defects in monocytes in chronically HBV-infected patients that include alteration in subset distribution, Toll-like receptor expression, cytokine production, phagocytic activity, oxidative response, migratory ability, polarization of monocyte-derived macrophages, and monocyte–T-cell interaction. We demonstrated that high levels of hepatitis B virus surface antigen and IL-4 potentiate these defects in monocytes via β-catenin induction while therapy with the nucleotide analog tenofovir fails to restore monocyte function. Our findings add to the continuing effort to devise new immunotherapeutic strategies that could reverse the immune defects in CHI.

Background: Hepatitis B virus surface antigen (HBsAg) is an important protein in both diagnosis and prevention of hepatitis B infection. In the current study, a piezoelectric immunosensor based on antibody-antigen interaction was designed to detect HBsAg. A quartz crystal microbalance system was employed to detect antibody-antigen interaction. Methods: At first, an oscillator was designed to measure the resonant frequency affected by the reactants using IC 74LVC1GX04. Antibody against HBsAg was immobilized on 10 MHz AT-cut quartz crystal. The surface modifications were monitored by atomic force microscopy (AFM) and contact angle measurements. Different concentrations of antibody were used for surface immobilization and the frequency shifts were assessed. The system stability was studied by evaluating the stability of the crystal and the immobilized antibody. The adsorption of antibody onto the crystal was analyzed using AFM and changes in the resonance frequency. Further, a direct immunoassay was performed with this immobilized antibody to identify HBsAg solutions at different concentrations. Finally, specific and non-specific responses were investigated using hepatitis B (HBsAg) and hepatitis C (HCV Ag) antigens, respectively. Results: Antibodies against HBsAg were successfully immobilized on 10 MHz AT-cut quartz crystal. The stability tests of crystal immobilized with antibody and unimmobilized crystal revealed that both forms of crystals were stable. Theoretical and experimental frequency assays were compared. A decrease in the contact angle indicated the hydrophilicity of surface after modifications. AFM images illustrated a more uniform surface after antibody adsorption and the surface roughness (RMS) reduced from 1.13 to 0.99 nm. Changes in the frequency were detected after the physical adsorption of HBsAb on the designed chip. The standard curve of antigen revealed the frequency changes depend on concentration of antigen. Finally, the specificity test confirmed the specificity of the designed biosensor for the detection of HBsAg from HCV Ag. The quantization of immobilized antibody was characterized by the frequency shift of the QCM. The obtained results were compared with ELISA assay. The correlation coefficients of HBsAg dilution between QCM and ELISA was 0.9821. Conclusions: This study is a new step to meet the challenges regarding HBsAg detection. Physical adsorption used in this study was effective as the simplest immobilization method to design a QCM-based immunosensor for HBsAg detection. Facilitated, fast, and simple detection of HBsAg by an antibody-based QCM biosensor is our main objective. [ABSTRACT FROM AUTHOR]

Background: Many efforts are currently focused on functional treatment of the hepatitis B virus (HBV). This can be done by suppressing the secretion of HBV surface antigen (HBsAg). Scientific communities are very interested in natural products in that respect. Objective: Use of root extract of Havachoobe (Onosma dichroanthum BoissI), a Northern Iranian native medical herb, for assessment of its anti-HBsAg secretion activity. Methods: Havachoobe had been bought at a nearby apothecary store. Plant root extract was obtained using a hydroalcoholic process. Cytotoxic activity of the extract was examined on PLC/PRF/5 cells using MTT assay. ELISA has been used to measure HBsAg in the treated cell line supernatants. In addition, real-time PCR analysis was performed to evaluate the expression of HBsAg before and after treatment of Onosma in vitro. Results: The results showed very low root extract cytotoxicity at concentrations under 8 μg/mL. Tissue culture infectious dose 50 was obtained at 63.78 μg/mL. In a dose-dependent and time-dependent manner, a significantly reduced HBsAg secretion was observed at a concentration of 8 ppm at 12 h post-treatment. The real-time PCR result showed relative decreased HBsAg expression at all doses at 12 h post-treatment time. Discussion: In this study, we first reported anti-HBsAg activity on an Iranian herbal medicine. Havachoobe root extract was shown to be able to inhibit HBsAg in a dose-dependent and time-dependent manner. We find the extract exerts its inhibitory effect of HBsAg by targeting transcription of HBsAg. [ABSTRACT FROM AUTHOR]

Background: Cytokines play an important role in occurrence and recovery of hepatitis B virus (HBV) infection. The aim of this study was to investigate the changes of cytokines concentration and its correlation to alanine aminotransferase (ALT), HBV deoxyribonucleic acid (HBV-DNA), hepatitis B envelope antigen (HBeAg), and HBV surface antigen (HBsAg) in the development of chronic hepatitis B (CHB). Methods: Thirteen healthy individuals (HI), 30 chronic HBV-infected patients in immune tolerant (IT) phase, and 55 CHB patients were enrolled between August 2015 and May 2017. The peripheral blood samples were collected from all individuals. The levels of interferon (IFN)-α2, interleukin (IL)-10, transforming growth factor (TGF)-β1, HBV-DNA, HBsAg, and HBeAg and liver function were measured. The quantitative determinations of cytokines levels, including IFN-α2, IL-10, and TGF-β1 were performed using Luminex multiplex technology. The correlation of cytokines to ALT, HBV-DNA, HBsAg, and HBeAg was analyzed by linear regression analysis. Results: IFN-α2 levels were similar between HI and IT groups (15.35 [5.70, 67.65] pg/ml vs. 15.24 [4.07, 30.73] pg/ml, Z = −0.610, P = 0.542), while it elevated significantly in CHB group (35.29 [15.94, 70.15] pg/ml vs. 15.24 [4.07, 30.73] pg/ml; Z = −2.522, P = 0.012). Compared with HI group (3.73 [2.98, 11.92] pg/ml), IL-10 concentrations in IT group (5.02 [2.98, 10.11] pg/ml), and CHB group (7.48 [3.10, 18.00] pg/ml) slightly increased (χ2 = 2.015, P = 0.365), and there was no significant difference between IT and CHB group (Z = −1.419, P = 0.156). The TGF-β1 levels among HI (3.59 ± 0.20 pg/ml), IT (3.62 ± 0.55 pg/ml), and CHB groups (3.64 ± 0.30 pg/ml) were similar (χ2 = 2.739, P = 0.254). In all chronic HBV-infected patients (including patients in IT and CHB groups), the elevation of IFN-α2 level was significantly associated with ALT level (β= 0.389, t = 2.423, P = 0.018), and was also negatively correlated to HBV-DNA load (β = −0.358, t = −2.308, P = 0.024), HBsAg (β = −0.359, t = −2.288, P = 0.025), and HBeAg contents (β = −0.355, t = −2.258, P = 0.027). However, when both ALT level and cytokines were included as independent variable, HBV-DNA load, HBsAg, and HBeAg contents were only correlated to ALT level (β = −0.459, t = −4.225, P = 0.000; β = −0.616, t = −6.334, P = 0.000; and β = −0.290, t = −2.433, P = 0.018; respectively). Conclusions: IFN-α2 elevation was associated with ALT level in patients with chronic HBV infection. However, in CHB patients, only ALT level was correlated to HBV-DNA, HBsAg and HBeAg contents.

For personalized treatment of chronic HBV-infected patients, it is necessary to identify hepatitis B virus (HBV) surface antigen (HBsAg) subtypes and HBV genotypes circulating among infected individuals. To this end, a laboratory version of the test kit for HBsAg subtype and HBV genotype identification in human blood serum/plasma using enzyme-linked immunosorbent assay (ELISA) and a custom monoclonal antibody (MAb) panel has been developed. With this kit version, it appears possible to identify the main HBsAg subtypes (ayw2, ayw3, adw2, and adrq+) and HBV genotypes (A, C, and D) circulation in Russian Federation. The aim of the work was to test the laboratory-developed ELISA kits for HBsAg subtyping and HBV genotyping in human HBsAg-positive blood plasma samples. A total of 146 HBsAg-positive plasma samples were tested. For 108 samples with detectable viral DNA, the results of HBsAg subtyping and HBV genotyping using the suggested MAb panel were compared with the results of genomic DNA sequencing for HBV isolates. For 38 HBsAg-positive samples in which HBV DNA could not be detected, ELISA, using the highly specific MAb panel kindly provided by P. Swenson, PhD, was employed as a reference technique. The study demonstrated that for the group of HBsAg-positive samples with detectable HBV DNA, the coincidence between the results obtained using the suggested MAb panel and the results of HBV DNA genome sequence analysis was 85% for HBsAg subtyping and 98% for HBV genotyping. For the group of HBsAg-positive samples with undetectable HBV DNA, the coincidence between the two techniques (the one using the suggested MAb panel and one using Swenson's MAb panel) was 95% in the case of HBsAg subtyping and 100% for HBV genotyping. The reported results allow recommending the suggested laboratory ELISA test kit for HBsAg subtype and HBV genotype identification in human blood plasma as an alternative or a complementary technique to genomic analysis in order to study HBV characteristics especially in those cases when HBV DNA isolation appears impossible. [ABSTRACT FROM AUTHOR]

• Attaching aptamers to agarose through DNA tetrahedral structure to build a novel type of immunosorbent. • This immunosorbent could remove approximately 90% of the HBsAg from human plasma samples. • Better adsorption for samples with high concentration of HBsAg. • The immunosorbents are noncytotoxic. DNA nanostructures are effective intermediates for immobilizing biomolecules because of their high level of controllability, the flexibility to create desired nanostructures through precision "bottom-up" assembly and the convenience to building fine nanostructures. The purpose of this study was to demonstrate the potential use of aptamers bound to agarose through a DNA tetrahedral structure to make a novel type of immunosorbent for the removal of hepatitis B virus surface antigens. Our results show that the proposed immunosorbent exhibits favorable biocompatibility and specificity. Electrophoresis and confocal microscopy were used to confirm the formation of immunosorbents. The ARCHITECT HBsAg assay was performed to quantitatively measure hepatitis B surface antigen. The cytotoxic potential of the immunosorbent was determined by an in vitro viability assay using V79 lung fibroblasts, demonstrating that the immunosorbents are noncytotoxic. The high adsorption capacity of the novel DNA nanostructure-based adsorbents towards hepatitis B surface antigen indicated the potential application of these materials for the treatment of Hepatitis B infection. [ABSTRACT FROM AUTHOR]

Background: Transfusion-transmissible hepatitis B virus (HBV) infection is a major problem worldwide. Recently, confirmatory nucleic acid tests (NATs) for HBV DNA have been employed in several countries. We assessed the prevalence and yearly trends of HBV infection in blood donors in the Eastern Province of Saudi Arabia, screening for HBV surface antigen (HBsAg), antibody against HBV core antigen (anti-HBc), and HBV DNA. Methods: Between 2011 and 2015, a total of 22,842 donors were screened for HBsAg, anti-HBc, and HBV DNA using the HBsAg Qualitative II kit (Abbott, Ireland Diagnostics Division, Sligo, Ireland), ARCHITECT Anti-hepatitis B core antigen antibody (HBc) II Assay kit (Abbott GmbH & Co. KG, Wiesbaden, Germany), and NAT Procleix Ultrio Elite Assay kit (Grifols Diagnostic Solutions Inc., Los Angeles, CA, USA), respectively. Results: A total of 739 (3.24%) donors were HbsAg(+), anti-HBc(+), or HBV DNA(+); 63 (0.28%) were HbsAg(+), anti-HBc(+), and HBV DNA(+). Twelve (0.05%) were anti-HBc(+) and HBV DNA(+) but HBsAg(-); they were considered to have occult infection. Further, 664 (2.91%) were HBsAg(-) but anti-HBc(+), indicating chronic or resolving infection. HBV prevalence increased significantly from 2011 to 2012, increased marginally till 2013, and showed a decreasing trend from 2013 (P >0.05). Conclusions: The five-year prevalence of HBV infection among blood donors in the Eastern Province of Saudi Arabia (3.24%) is lower than that reported for other regions in the country. The occult HBV infection rate of 0.05% emphasizes the importance of NATs in isolating potential infectious blood units. [ABSTRACT FROM AUTHOR]

This review article deals with the possibility of using the quantitative determination of HBsAg (qHBsAg) to monitor the natural history of the disease. The necessity of unification of qHBsAg use to monitor the natural history of the disease and antiviral therapy, with insuring recommendations in the continental guidelines for HBV-infection, is indicated.

This study was to elucidate longitudinally quantitative changes of hepatitis B virus (HBV) surface antigen (HBsAg) and HBV DNA in elder HBsAg carriers in a community. Among 1002 residents screened for HBsAg in 2005, 405 responded to this follow-up study in 2010. Fifty-nine (14.6%) were HBsAg carriers in 2005; HBsAg quantification and HBV DNA were measured. HBsAg quantification (cutoff 1600 IU/mL) and HBV DNA (cutoff 2000 IU/mL) were combined to stratify the participants between two screens. A total of 30 men and 29 women with a mean age of 63.9 ± 7.9 years were enrolled. Quantitative levels of HBsAg and HBV DNA were significantly correlated in 2005 (r = 0.509, p

Abstract: A critical challenge in producing an antibody‐based assay with the highest reproducibility and sensitivity is the strategy to immobilize antibodies to solid phase. To date, numerous methods of antibody immobilization were reported but each was subjected to its advantages and limitations. The current study proposes a new potential antibody binding protein, the human neonatal fragment crystallizable (Fc) receptor. This protein has shown its high affinity to the Fc of antibody either in vivo or in vitro. Human neonatal Fc receptor is a heterodimer constructed by p51 α‐heavy chain and β2‐microglobulin light chain; however, the binding sites toward the antibody are located in the p51 α‐heavy chain. Hence, vector cloning and recombinant protein expression were carried out to express the p51 α‐heavy chain of the human neonatal Fc receptor (hFcRn‐α). The recombinant protein expressed, hFcRn‐α, was adopted to pin rabbit IgG against hepatitis B virus surface antigen to a solid phase. A sandwich enzyme‐linked immunosorbent assay was further developed to evaluate the efficiency of hFcRn‐α‐directed immobilization in antigen detection. The result was compared with the conventional physical adsorption method. The findings demonstrated that human neonatal Fc receptor was efficient in pinning antibodies and generating higher signals compared with the physical adsorption of antibody. [ABSTRACT FROM AUTHOR]

Herein, we present a novel sandwich fluorescence enzyme linked immunosorbent assay (ELISA) for highly sensitive detection of Hepatitis B virus surface antigen (HBsAg) based on glucose oxidase (GOx)-induced fluorescence quenching of mercaptopropionic acid-modified CdTe quantum dots (MPA-QDs). In this system, hydrogen peroxide (H 2 O 2 ) sensitive MPA-QDs was used as a signal output, and glucose oxidase (GOx) was used as label which can generate H 2 O 2 via catalytic oxidation of glucose. The proposed method showed dynamic linear detection of HBsAg both in the range of 47 pg mL −1 ~ 380 pg mL −1 and 0.75 ng mL −1 ~ 12.12 ng mL −1 . The detection limit of the proposed fluorescence ELISA was 1.16 pg mL −1 , which was approximately 430-fold lower than that of horseradish peroxidase (HRP)-based conventional ELISA. The average recoveries for HBsAg-spiked serum samples ranged from 98.0% to 126.8% with the relative standard derivation below 10%, thus indicating acceptable precision and high reproducibility of the proposed fluorescence ELISA for HBsAg detection. Additionally, the developed method showed no false positive results analyzing 35 real HBsAg-negative serum samples, and exhibited excellent agreement (R 2 =0.9907) with a commercial time-resolved fluorescence immunoassay (TRFIA) kit for detecting 31 HBsAg-positive serum samples. In summary, the proposed method based on fluorescence quenching of H 2 O 2 sensitive QDs is considerably to be an excellent biodetection platform with ultrahigh sensitivity, good accuracy and excellent reliability. [ABSTRACT FROM AUTHOR]

Abstract: A sensitive electrochemical immunosensor for Hepatitis B virus surface antigen (HBsAg) detection was fabricated based on hemin/G‐quadruplex interlaced onto Fe3O4‐AuNPs or hemin ‐amino‐reduced graphene oxide nanocomposite (H‐amino‐rGO‐Au). G‐quadruplex DNAzyme, which is composed of hemin and guanine‐rich nucleic acid, is an effective signal amplified tool for its outstanding peroxidase activity and Fe3O4‐AuNPs or (H‐amino‐rGO‐Au) nanocomposites with quasi‐enzyme activity provide appropriate support for the immobilization of hemin/G‐quadruplex. The target protein was sandwiched between the primary antibody immobilized on the GO and secondary antibody immobilized on the Fe3O4‐AuNPs or (H‐amino‐rGO‐Au) nanocomposites and glutaraldehyde was used as linking agent for the immobilization of primary antibody on the surface of GO. Both Fe3O4‐AuNPs and H‐amino‐rGO‐Au nanocomposite and also hemin/G‐quadruplex can cooperate the electrocatalytic reduction of H2O2 in the presence of methylene blue as mediator. The proposed immunosensor has a wide linear dynamic range of 0.1 pg/ml to 300 pg/ml with a detection limit of 60 fg/ml when Fe3O4‐AuNPs was used for immobilization of hemin/G‐quadruplex, while the dynamic range and DL were 0. 1–1000 pg/mL and 10 fg/mL, respectively in the presence of H‐amino‐rGO‐ Au nanocomposite as platform for immobilizing of hemin/G‐quadruplex. The proposed immunosensor was also used for analysis of HBsAg in spiked human serum samples with satisfactory results. [ABSTRACT FROM AUTHOR]

The replication of hepatitis B virus (HBV) may be modulated by a variety of cell signaling pathways, including the phosphatidylinositol 3‑kinase (PI3K)‑RAC‑α serine/threonine‑protein kinase (AKT)‑serine/threonine‑protein kinase mTOR (mTOR) pathway. The aim of the present study was to determine the regulatory effects of this pathway on the infection and replication of HBV. The results indicated that the HBV entry process may activate the AKT pathway, as demonstrated by AKT phosphorylation in HBV natural infection. However, inhibition of AKT phosphorylation by short‑term treatment with AKT inhibitors was unable to block HBV entry, which suggested that AKT activation induced by HBV infection is not essential for viral entry process. Prolonged treatment with PI3K‑AKT‑mTOR pathway inhibitors markedly promoted HBV replication in HBV replicating and natural infection models. The PI3K‑AKT‑mTOR pathway was therefore identified to be a negative regulator of HBV replication. These inhibitors enhanced the replication and transcription of HBV in an HBx‑dependent way. The results additionally indicated that a PI3K inhibitor, Ly294002, inhibited the secretion of the small surface antigen of HBV in a PI3K‑AKT‑independent manner. The inhibitor Ly294002 may be used as a tool for the drug development of surface antigen secretion inhibitors. [ABSTRACT FROM AUTHOR]

Bio-nanocapsules (BNCs) consisting of hepatitis B virus surface antigen (HBsAg) L proteins and phospholipids are used as efficient non-viral carriers for liver-specific delivery of genes and drugs. Considering the administration to HB vaccinees and HB patients, endogenous anti-HBsAg immunoglobulins (HBIGs) may reduce the delivery efficacy and prevent repetitive administration. Therefore, low immunogenic BNCs were generated by inserting two point mutations in the HBsAg L protein, which were found in HBV escape mutants. Escape mutant-type BNC (emBNC) showed 50% lower HBIG binding capacity than that of parental BNC (wtBNC). It induced HBIG production to a lesser extent than that associated with wtBNC in BALB/c mice. The emBNC could accumulate into human hepatocyte-derived tumor in mice pre-treated with HBIGs. The complex of emBNC and cationic liposomes could deliver plasmid DNA to HepG2 cells efficiently in the presence of HBIGs. Thus, emBNC could evade HBIG-neutralizing antibodies, expanding the clinical utility of BNC-based nanomedicine. [ABSTRACT FROM AUTHOR]

Background: Hepatitis B is the world's most common serious liver infection. Infection by hepatitis B virus during pregnancy has high rate of vertical transmission and adverse effect on both the mother and child. Data on seroprevalence and associated factors among pregnant women in Dawuro or surrounding was scarce. Therefore, the aim of this study was to assess prevalence of hepatitis B virus and factors associated among pregnant women in Dawuro Zone. Methods: Institution based cross-sectional study was conducted from 1st April to May 31, 2015 in Dawuro zone. Venous blood was collected, plasma was separated and hepatitis B virus surface antigen was screened using rapid test. Logistic regression was employed with 95% CI and p value of <0.05 was used as statistically significant. Results: Data were collected on 289 women. The overall prevalence from this study was (p=3.5%, 95% CI 1.4-5.6%) multivariate analysis using logistic regression showed multiple sexual partner (AOR=6.923; 95% CI 1.685-28.441), and abortion history (AOR = 4.975; 95% CI 1.21-20.456), were significantly associated with hepatitis B virus surface antigen (HBsAg) infection. Conclusion: This seroprevalence was categorized as intermediate endemicity according to WHO classification criteria. Health education on sexual transmission of hepatitis B virus and inclusion of screening hepatitis b virus as routine antenatal care service is recommended. [ABSTRACT FROM AUTHOR]

Here we prepared an electrochemical immunosensor employing Au sheet as working electrode, Fe 3 O 4 magnetic nanoparticles (MNPs) as supporting matrix and hemin/G-quadruplex DNAzyme as signal amplifier for determination of hepatitis B virus surface antigen (HBsAg). First, the primary antibody of HBs (Ab 1 ) was immobilized on the surface of the carboxyl-modified MNPs. Then, the assembly of antibody and alkylthiol/G-quadruplex DNA/hemin on gold nanoparticles was used as bio-bar-coded nanoparticle probe. Protein target was sandwiched between the primary antibody of HBs (Ab 1 ) immobilized on the MNPs and hemin bio-bar-coded AuNPs probe labeled antibody (Ab 2 ). Hemin/G-quadruplex structure as HRP mimicking-DNAzyme significantly improved the catalytic reduction of H 2 O 2 by oxidation of methylene blue (MB). Square wave voltammetry signals of MB provided quantitative measurements of HBsAg with a linear concentration range of 0.3–1000 pgmL −1 and detection limit of 0.19 pgmL −1 . Due to efficient catalytic activity of HRP mimicking-DNAzyme, the proposed immunosensor exhibited high sensitivity and it holds great promise for clinical application and provides a new platform for immunosensor development and fast disease diagnosis. [ABSTRACT FROM AUTHOR]

Background: Although Egypt had adopted implementation of routine infant hepatitis B virus (HBV) vaccination in 1992, its effectiveness is not evaluated on a national scale. Assessment of early and long-term seroprotection after compulsory vaccination is an important measure for monitoring the success of the vaccination program. Aim: The aim of this study was to assess HBV seroprotection and immune memory in children and adolescents who were vaccinated during infancy in Cairo Governorate. Materials and Methods: The study was carried out in two phases. The first phase was a cross-sectional study carried out in five districts in Cairo Governorate, recruiting 819 children in the age range of 9 months to 16 years. All children had received full doses of the compulsory HBV vaccination. Serum samples were taken from each child and assessed for antibody against hepatitis B virus surface antigen (anti-HBs) titer; total antibodies against HBV core antigen, and HBV surface antigen. HBV DNA was investigated by real-time PCR for those who were HBV core antigen or HBV surface antigen positive. In the second phase, nonseroprotected children (anti-HBs <10 IU/l) received HBV booster dose. Anti-HBs titer was reassessed after 4 weeks to identify anamnestic response. Individuals showing antibody concentrations of less than 10 IU/l were then given an additional complete course of vaccination. Results: Four out of 819 children had HBV breakthrough infection. The seroprotection rate was 60.7%, and was significantly higher among children aged less than 5 years compared to the older age groups and among boys compared to girls. Multivariate logistic analysis showed age as the only independent predictor of low anti-HBs titer. About 95% of nonseroprotected children developed anamnestic response postbooster. Anti-HBs geometric mean titer (GMT) increased significantly from pre-booster (13.8±16.9IU/L) compared to post-booster (307±6.0IU/L, P<0.001). Anti-HBs GMT was significantly higher among children with prebooster anti-HBs level =1 IU/l (424.9±4.4 IU/l) compared to children with undetectable level (178.3±8.3). Conclusion: Despite waning of anti-HBs over time, long-term protection still exists. The high anamnestic response rate signifies the existence of immune memory and giving a booster dose is not recommended. However, we suggest that prolonged follow up and surveillance of vaccinees immunized at an early age should be continued. [ABSTRACT FROM AUTHOR]

Background and aim To prospectively evaluate the efficacy of vaccine alone compared with vaccine plus HBIG for preventing HBV transmission in neonates of HBsAg (+)/HBeAg (−) mothers. Methods Combined immunization is currently recommended for neonates of HBsAg (+) mothers in China. As a result, a randomized design is infeasible due to ethical reasons. In practice, Guangxi Zhuang Autonomous Region and Jiangsu Province implement vaccine alone and vaccine plus HBIG strategies for neonates born to HBsAg (+)/HBeAg (−) mothers, respectively. We alternatively enrolled neonates of HBsAg (+)/HBeAg (−) mothers from these two regions. Three doses of a recombinant yeast-derived hepatitis B vaccine were given at 0, 1 and 6 months with or without HBIG at birth. Results At 7 months, sera were collected from 132 neonates in Guangxi Zhuang Autonomous Region and 752 neonates in Jiangsu Province. Baseline characteristics of both mothers and neonates were comparable in the two regions. No differences were revealed regarding the occurrence of perinatal HBV transmission with or without HBIG at birth [0.1% (1/752) vs. 0.0% (0/132), p = 1.000]. The anti-HBs response rates were 97.7% (129/132) and 98.5% (740/751) for the neonates with vaccine alone and with HBIG ( p = 0.758), respectively. Vaccine alone induced a significantly higher anti-HBs GMC as compared to vaccine plus HBIG at 7 months of age (1555.3 mIU/mL vs. 654.9 mIU/mL, p < 0.0001). At 12 months of age, protective levels of anti-HBs remained in 97.4% (596/612) and 98.3% (118/120) of the neonates receiving and not receiving HBIG, respectively ( p = 0.771). The neonates receiving combined prophylaxis had a markedly lower anti-HBs GMC (210.7 mIU/mL vs. 297.0 mIU/mL, p = 0.011). Horizontal HBV transmission occurred in none of the successfully immunized neonates for both compared groups at 12 months of age. Conclusions Vaccine alone may be enough for preventing HBV transmission in neonates of HBsAg (+)/HBeAg (−) mothers. [ABSTRACT FROM AUTHOR]

In this paper, a novel time-resolved fluorescence immunoassay (TRFIA) is described that allows the simultaneous quantitative detection of hepatitis B virus surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in human serum to aid the diagnosis and monitoring of hepatitis B virus infection. The proposed method was developed based on a two-step sandwich immunoassay protocol in which monoclonal antibodies against HBsAg and HBeAg were co-coated in 96 microtitration wells, then tracer polyclonal antibodies against HBsAg labeled with samarium and tracer monoclonal antibodies against HBeAg labeled with europium chelates were used for detection. The detection range was 0.1-150 IU/mL for HBsAg and 0.5-160 PEIU/mL for HBeAg, and the detection limits were 0.03 IU/L and 0.09 PEIU/ml, respectively. The intra- and inter-assay coefficients of variation were below 8 % for both virus antigens. The dilution linearity and accuracy of the assay were satisfactory. No statistically significant differences were observed in sensitivity or specificity for the serum samples between the dual-label TRFIA and a commercial single-label TRFIA. These results demonstrate that an effective, reliable and convenient HBsAg/HBeAg dual-label TRFIA was successfully developed that may be clinically applicable for blood screening to monitor the course of hepatitis B virus infection and predict treatment responses. [ABSTRACT FROM AUTHOR]

Objective: To determine the frequency of hepatitis B virus surface antigen (HBsAg) and hepatitis C virus (HCV) antibodies in children with hematological disorders having multiple transfusions. Methodology: This descriptive cross-sectional study was conducted in the Department of Pediatrics, Lady Reading Hospital, Peshawar from December 10, 2016 to June 10, 2017. A total of 230 children with different hematological disorders who received two or more blood transfusions during the past two years were included in the study. Each child was screened for HBsAg and anti-HCV antibodies using 3rd generation commercial ELISA micro-plate kits (DIA.PRO, ITALY) according to the manufacturer instructions. Results: Out of 230, 122 (53%) patients were males and 108 (47%) were females. Their mean age was 10 ±10.86 years. In this study, 23% of patients were positive for anti-HCV antibodies, while 7% of patients were positive for HBsAg. Out of 53 patients positive for hepatitis C antibodies, 55% were males. Similarly, out of 16 patients positive for HBsAg, 56% were males. Conclusion: The frequency of anti-HCV antibodies was 23% and HBsAg was positive in 7% of all children who received multiple transfusions in the past two years. [ABSTRACT FROM AUTHOR]

• Tenofovir alafenamide (TAF)-containing antiretroviral therapy effectively suppressed lamivudine-resistant hepatitis B virus (HBV) in people living with human immunodeficiency virus (HIV). • The decrease in hepatitis B virus surface antigen (HBsAg) level was minimal, and HBsAg loss occurred rarely (2.2%) after switching to a TAF-containing regimen. • Hepatitis B virus envelope antigen seroconversion occurred more frequently in patients with HIV co-infected with lamivudine-susceptible HBV than in those co-infected with lamivudine-resistant HBV. • Switching from a tenofovir-disoproxil-fumarate- to a TAF-containing regimen led to improved proteinuria and bone density in patients co-infected with HIV and HBV. Data on the effectiveness of tenofovir alafenamide (TAF) against lamivudine-resistant (LAM-R) hepatitis B virus (HBV) among patients co-infected with human immunodeficiency virus (HIV) and HBV are limited. Between April and December 2018, HIV-positive patients co-infected with LAM-R or lamivudine-susceptible (LAM-S) HBV who switched from tenofovir-disoproxil-fumarate-containing antiretroviral therapy (ART) to TAF-containing ART were followed for 96 weeks. Plasma HBV and HIV loads, HBV serological markers, and liver function before and after the switch were analysed. In total, 182 patients co-infected with HIV and HBV were included in this study: 45 with LAM-R HBV and 137 with LAM-S HBV. At baseline, 28.9% and 7.4% of patients in the LAM-R and LAM-S groups, respectively, tested positive for hepatitis B virus envelope antigen (HBeAg) (P <0.001), and the respective percentages of patients who had achieved plasma HBV DNA <20 IU/mL were 95.5% and 97.1%. At weeks 48 and 96, 100% and 94.9% of patients in the LAM-R group, respectively, and 97.1% and 95.6% of patients in the LAM-S group, respectively, maintained plasma HBV DNA <20 IU/mL. Lamivudine resistance of HBV and baseline hepatitis B virus surface antigen (HBsAg) level were associated with HBsAg decrement at week 96 at a degree of 0.25 log 10 IU/mL [95% confidence interval (CI) 0.059–0.246] and 0.22 log 10 IU/mL (per 1-log 10 IU/mL increase, 95% CI 0.018–0.101), respectively. At week 96, 2.2% (4/182) of patients had HBsAg loss; no patients in the LAM-R group and 25.0% (2/8) of patients in the LAM-S group had HBeAg seroconversion. Switching to TAF-containing regimens maintained high rates of HBV viral suppression in patients co-infected with either LAM-R or LAM-S HBV. The decrease in HBsAg was minimal, and HBsAg seroconversion occurred infrequently. [ABSTRACT FROM AUTHOR]

Immunogenicity and safety of different adjuvants combined with a model antigen (HBsAg) were compared. Healthy HBV-naïve adults were randomized to receive HBs adjuvanted with alum or Adjuvant Systems AS01B, AS01E, AS03A or AS04 at Days 0 and 30. Different frequencies of HBs-specific CD4 + T cells 14 days post dose 2 but similar polyfunctionality profiles were induced by the different adjuvants with frequencies significantly higher in the AS01B and AS01E groups than in the other groups. Antibody concentrations 30 days post-dose 2 were significantly higher in AS01B, AS01E and AS03A than in other groups. Limited correlations were observed between HBs-specific CD4 + T cell and antibody responses. Injection site pain was the most common solicited local symptom and was more frequent in AS groups than in alum group. Different adjuvants formulated with the same antigen induced different adaptive immune responses and reactogenicity patterns in healthy naïve adults. The results summary for this study (GSK study number 112115 – NCT# NCT00805389 ) is available on the GSK Clinical Study Register and can be accessed at www.gsk-clinicalstudyregister.com . [ABSTRACT FROM AUTHOR]

The aim of the present study was to investigate the in vitro effects of hepatitis B virus surface antigen (HBsAg) on the immune function of human monocyte-derived dendritic cells (MD-DCs), and the moderating role of T cell immunoglobulin and mucin domain-containing molecule-3 (Tim-3) signaling molecule. The monocytes, obtained from healthy adult peripheral blood, were incubated with recombinant human granulocyte-macrophage colony-stimulating factor and interleukin (IL)-4 to induce DCs. DC-associated cell markers were detected using flow cytometry. MD-DCs were treated with HBsAg (5 μg/ml) in vitro for 48 h and subsequently, cell markers, lymphocyte stimulatory capacity, signaling protein and downstream cytokines were assessed. In addition, a Tim-3 monoclonal antibody was used to inhibit the Tim-3 signaling pathway, and subsequently the immune responses of MD-DCs to HBsAg stimulation were determined using the aforementioned method. The cell phenotype expressions of MD-DCs were all significantly increased with cluster of differentiation (CD)11c at 70.09±0.57%, human leukocyte antigen-DR at 79.83±2.12%, CD80 at 48.33±7.34% and CD86 at 44.21±5.35%. The treatment of MD-DCs with HBsAg resulted in a CD80 and CD86 enhanced expression, enhanced lymphocyte stimulatory capacity, upregulated expression of Tim-3 and nuclear factor-κB (NF-κB), as well as enhanced cytokine secretion of IL-6, IL-10 and interferon (IFN)-γ. However, a reduced immune response of MD-DCs in response to HBsAg stimulation was observed when the Tim-3 signaling pathway was inhibited prior to stimulation. The expression of NF-κB was decreased and the cytokine secretion level of IL-6, IL-10 and IFN-γ were downregulated. The treatment with HBsAg in vitro resulted in an enhanced immune response of MD-DCs, which may be positively regulated by the Tim-3 signaling molecule. [ABSTRACT FROM AUTHOR]

The aim of this study was to prepare poly(lactic-co-glycolic acid) (PLGA) microspheres containing hepatitis B virus surface antigen (HBsAg) using human serum albumin (HSA) as a stabilizer. Lyophilization and emulsification of HBsAg solution with dichloromethane caused a considerable loss of HBsAg antigenicity. Thus, the effects of HSA and trehalose on HBsAg recovery during lyophilization and emulsification were investigated. Adding HSA to HBsAg solutions significantly improved antigen recovery to >90% during lyophilization and emulsification. The effects of co-encapsulated HSA on the characteristics of the PLGA microspheres and stability of HBsAg released from the microspheres were also investigated. The in vitro release test showed that HBsAg was released from the PLGA microspheres continuously over seventy days. A large amount of released HBsAg was inactive without co-encapsulation of HSA. On the contrary, with HSA co-encapsulation, the released HBsAg retained approximately 90% of its antigenicity. The single injection of the HBsAg-HSA-loaded PLGA microspheres in rats resulted in higher anti-HBsAg IgG and Th1 cytokine levels than the single injection of the HBsAg-loaded microspheres or two injections of the conventional aluminum-adjuvanted HBsAg vaccine. Based on these findings, the HBsAg-HSA-loaded PLGA microspheres could be an effective carrier for HBsAg and form a promising depot system. [ABSTRACT FROM AUTHOR]

The Malaria Vaccine Technology Roadmap calls for a 2015 landmark goal of a first-generation malaria vaccine that has protective efficacy against severe disease and death, lasting longer than one year. This review focuses on product development efforts over the last five years of RTS,S, a pre-erythrocytic, recombinant subunit, adjuvanted, candidate malaria vaccine designed with this goal of a first-generation malaria vaccine in mind. RTS,S recently completed a successful pivotal Phase III safety, efficacy and immunogenicity study. Although vaccine efficacy was found to be modest, a substantial number of cases of clinical malaria were averted over a 3–4 years period, particularly in settings of significant disease burden. European regulators have subsequently adopted a positive opinion under the Article 58 procedure for an indication of active immunization of children aged 6 weeks up to 17 months against malaria caused by Plasmodium falciparum and against hepatitis B. Further evaluations of the benefit, risk, feasibility and cost-effectiveness of RTS,S are now anticipated through policy and financing reviews at the global and national levels. [ABSTRACT FROM AUTHOR]

Bio-nanocapsules (BNCs) are a hollow nanoparticle consisting of about 100-nm liposome (LP) embedding about 110 molecules of hepatitis B virus (HBV) surface antigen (HBsAg) L protein as a transmembrane protein. Owing to the human hepatocyte-recognizing domains on the N-terminal region (pre-S1 region), BNCs have recently been shown to attach and enter into human hepatic cells using the early infection mechanism of HBV. Since BNCs could form a complex with an LP containing various drugs and genes, BNC–LP complexes have been used as a human hepatic cell-specific drug and gene-delivery system in vitro and in vivo . However, the role of BNCs in cell entry and intracellular trafficking of payloads in BNC–LP complexes has not been fully elucidated. In this study, we demonstrate that low pH-dependent fusogenic activity resides in the N-terminal part of pre-S1 region (NPLGFFPDHQLDPAFG), of which the first FF residues are essential for the activity, and which facilitates membrane fusion between LPs in vitro . Moreover, BNC–LP complexes can bind human hepatic cells specifically, enter into the cells via clathrin-mediated endocytosis, and release their payloads mostly into the cytoplasm. Taken together, the BNC portion of BNC–LP complexes can induce membrane fusion between LPs and endosomal membranes under low pH conditions, and thereby facilitate the endosomal escape of payloads. Furthermore, the fusogenic domain of the pre-S1 region of HBsAg L protein may play a pivotal role in the intracellular trafficking of not only BNC–LP complexes but also of HBV. [ABSTRACT FROM AUTHOR]

Monocytes play an important role in the control of microbial infection, but monocyte biology during chronic hepatitis B virus (HBV) infection (CHI) remains inadequately studied. We investigated the frequency, phenotype, and functions of monocyte subsets in different phases of CHI, namely, immune tolerance (IT), hepatitis B early antigen (HBeAg)-positive/HBeAg-negative chronic hepatitis B (EP-/EN-CHB, respectively), and inactive carrier (IC), identified factors responsible for their functional alterations, and determined the impact of antiviral therapy on these cells. Flow cytometric analysis indicated that HLA-DR + CD14 ++ CD16 - classical monocytes were significantly reduced while HLA-DR + CD14 ++ CD16 + intermediate and HLA-DR + CD14 + CD16 ++ nonclassical monocytes were expanded in IT and EP-/EN-CHB compared with those in IC and healthy controls (HC). In comparison to IC/HC, monocytes in IT and CHB exhibited diminished expression of Toll-like receptor 2 (TLR-2)/TLR-4/TLR-9 and cytokines interleukin-12 (IL-12)/tumor necrosis factor alpha (TNF-α)/IL-6 but produced higher levels of IL-10/transforming growth factor β (TGF-β). Further, monocytes in CHB/IT showed impaired phagocytosis and oxidative response relative to those in IC/HC. In vitro assays indicated that high titers of hepatitis B surface antigen (HBsAg) present in IT/CHB and of IL-4 in CHB triggered the functional defects in monocytes via induction of β-catenin. Additionally, monocyte-derived M1 macrophages of CHB/IT produced fewer proinflammatory and more anti-inflammatory cytokines than those of IC/HC, while in CHB/IT, the monocytes skewed the differentiation of CD4 + T cells more toward regulatory T cells and a Th2 phenotype. Moreover, monocytes in CHB and IT overexpressed chemokine receptor CCR2, which coincided with increased intrahepatic accumulation of β-catenin + CD14 + cells. One year of tenofovir therapy failed to normalize monocyte functions or reduce serum HBsAg/IL-4 levels. Taken together, monocytes are functionally perturbed mostly in IT and EP-/EN-CHB phases. Targeting intramonocytic β-catenin or reducing HBsAg/IL-4 levels might restore monocyte function and facilitate viral clearance. IMPORTANCE Chronic HBV infection (CHI) is a major cause of end-stage liver disease for which pharmacological treatments currently available are inadequate. Chronically HBV-infected patients fail to mount an efficient immune response to the virus, impeding viral clearance and recovery from hepatitis. Monocytes represent a central part of innate immunity, but a comprehensive understanding on monocyte involvement in CHI is still lacking. We here report a multitude of defects in monocytes in chronically HBV-infected patients that include alteration in subset distribution, Toll-like receptor expression, cytokine production, phagocytic activity, oxidative response, migratory ability, polarization of monocyte-derived macrophages, and monocyte-T-cell interaction. We demonstrated that high levels of hepatitis B virus surface antigen and IL-4 potentiate these defects in monocytes via β-catenin induction while therapy with the nucleotide analog tenofovir fails to restore monocyte function. Our findings add to the continuing effort to devise new immunotherapeutic strategies that could reverse the immune defects in CHI.

Hepatitis delta virus (HDV) coinfected with HBV causes severe viral hepatitis, however, the number of HDV infection may be underestimated. In the present study, we enrolled 1,141,331 blood donations, routinely tested for HBsAg and/or HBV DNA, from 21 blood establishments in China. 2,690 donors were HBsAg and/or HBV DNA positive after screening tests. After verification of HBsAg and HBV DNA, 1,490 samples were HBsAg confirmed-positive, including 1,459 HBV DNA-positive samples, and 825 samples were seronegative but HBV DNA positive. We first analyzed demographic characteristics of involved 2,690 donors with different HBV infection status and found the proportions of males, the older donors, workers and farmers were higher in HBsAg-/HBV DNA+ group. Then we evaluated specificity of HDV IgG and IgM antibody assays with 375 HBsAg and HBV DNA confirmed-negative samples, and 374 were tested negative using the two assays, respectively, suggesting a specificity of 99.73% for both assays (374/375, 95% Cl: 98.51-99.95%). Subsequently, we tested for HDV IgG and IgM of 2,315 HBsAg and/or HBV DNA confirmed-positive samples, and nine showed reactivity for IgG, while two were reactive for IgM. All these 11 reactive samples were tested again with another HDV pan-Ig and IgM testing assays and HDV RNA, and only one donor was identified as HDV IgG positive and HDV RNA negative, showing an HDV seroprevalence of 0.067% (95%CI: 0.012-0.38%) among HBsAg-positive blood donors in China. The positive donor was followed up for 2 years after the donation date, and decreased antibody titer of HDV IgG and HBsAg conversion were observed, and the infection status of the donor was HDV infection with recovery and occult hepatitis B virus infection with genotype C2. These results indicated a low seroprevalence of HDV infection among blood donors and a low risk of HDV transmission through blood transfusion in China., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Chang, Yan, Ji, Sun, Jiang, Lu, Wang L and HBV-Infected Blood Donors Study Group.)

Objectives In the case of coinfection with HIV and hepatitis B virus ( HBV) and/or hepatitis C virus ( HCV), hepatic disease progression is often accelerated, with higher rates of liver cirrhosis and liver-related mortality. We aimed to evaluate the performance of the rapid tests used routinely to detect HBV surface antigen ( HBsAg) and anti- HCV among HIV-infected patients in Guinea- Bissau. Methods Blood samples from HIV-infected patients in Guinea- Bissau were stored after testing for HBsAg and anti- HCV with rapid tests. Samples were subsequently re-tested for HBsAg and anti- HCV in Denmark. Results Two rapid tests were used in Guinea- Bissau: HBsAg Strip Ref 2034 ( VEDA. LAB, Alençon, France; sensitivity 62.3%; specificity 99.2%) and HEPA-SCAN ( Bhat Bio- Tech, Bangalore, India; sensitivity 57.1%; specificity 99.7%). In the two tests the ability to obtain the correct outcome depended on the antigen and antibody concentrations, respectively. Sex, age, CD4 cell count and antiretroviral therapy status did not differ between false negative and true positive samples in either of the tests. The study is limited by a low number of anti- HCV positive samples. Conclusions New diagnostic rapid tests should always be evaluated in the setting in which they will be used before implementation. [ABSTRACT FROM AUTHOR]

Occult hepatitis B virus infection (OBI) is highly prevalent worldwide. In some cases, it is a consequence of infection with variant viruses mutated in the S gene and producing a surface antigen not recognized by diagnostic kits. In most cases, OBI is due to a strong inhibition of hepatitis B virus (HBV) activities exerted by host defense mechanisms. OBI may reactivate in patients undergoing immunosuppressive therapy and/or chemotherapy with the possibility of a consequent development of acute hepatitis that may lead to hepatic failure. Hematological malignancies and therapeutic schedules including rituximab are the conditions most frequently associated with OBI reactivation. However, this event may occur in a large number of additional clinical and therapeutic settings. Identification of patients prone to undergo reactivation is of great importance for promptly starting a proper antiviral therapy that may stop the HBV reactivation and prevent its clinical sequelae. © 2014 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Highlights: [•] Flow cytometry was used for simultaneous detection of multiple analytes using E. coli cells. [•] E. coli cells with and without fluorescence in cytosol were used for multiple analyte detection. [•] E. coli cell with autodisplayed Z-domains was used for hyper sensitive immunoassay. [•] Flow cytometry based multiple detection was demonstrated with HBsAg and CRP as model analytes. [Copyright &y& Elsevier]

DNA damage response and repair pathways are important barriers to carcinogenesis. Here, we show that promyelocytic leukaemia ( PML, also known as TRIM19), involved in sensing DNA damage and executing homologous recombination repair, is down-regulated in non-tumour liver cells surrounding hepatitis B virus ( HBV)-related hepatocellular carcinoma ( HCC). No PML mutation or deletion was found in HBV-infected liver or HCC cells. Immunohistochemical analysis of liver biopsies from patients with breast or liver cancer and HBV reactivation after chemotherapy revealed PML up-regulation and HBV exacerbation in normal liver tissue in response to DNA damage (functional PML), PML down-regulation in HCC peritumour cells associated with high HBsAg accumulation and low HBV replication activity (suppressive PML), and heterogeneous nuclear PML expression in HCC cells that lost HBV DNA and HBsAg and were non-reactive to DNA damage (dysregulated PML). Loss of PML in HBsAg-transgenic mice promoted chromosome breaks in liver cells and accelerated the accumulation of body and liver fat and the development of a liver steatosis-dysplasia-adenoma-carcinoma sequence in an inflammation-independent and male-predominant manner, compared to PML knock-out or HBsAg-transgenic mice during the same time period. These results indicate that PML deficiency facilitates genomic instability and promotes HBsAg-related hepatocarcinogenesis, which also involves androgen and lipid metabolism. These findings uncover a novel PML link between HBV-related tumourigenesis, DNA repair, and metabolism. Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

We developed a fluorescence-activated cell sorter (FACS)-based immunoassay using Escherichia coli with autodisplayed Z-domains as a solid support for detection antibodies as well as a method to optimize the signal-to-noise ratio of fluorescence signals from E. coli. To improve the sensitivity of FACS measurements by eliminating artifactual signals from unrelated small particles in the sample mixture, E. coli with autodisplayed Z-domains was modified to express enhanced green fluorescence protein (eGFP) in the cytosol. By selectively gating the measurement channel for eGFP in the cytosol, we were able to discriminate between green fluorescent E. coli and artifact particles. We confirmed the efficacy of this optimization method for FACS-based immunoassay using several model reactions. Finally, as proof of concept, we used the FACS-based immunoassay and our optimization method for the medical diagnosis of human hepatitis B virus surface antigen. [ABSTRACT FROM AUTHOR]

ABSTRACT Time-resolved fluorometry of lanthanide chelates is one of the most useful non-isotopic detection techniques and has been used in numerous applications in biomedical science. We developed a time-resolved fluoroimmunoassay (TRFIA) to quantify α-fetoprotein (AFP) and hepatitis B virus surface antigen (HBsAg) in human serum. Based on a two-site sandwich protocol, monoclonal antibodies (McAbs) against AFP and HBsAg were co-coated in 96 microtitration wells and tracer McAbs against HBsAg and AFP were labeled with europium (Eu) and samarium (Sm) chelates, respectively. After application of diluted serum samples, Eu3+- and Sm3+-McAbs were added and fluorescence signals of Sm3+ and Eu3+ tracers were collected. Detection limits of AFP and HBsAg were 0.09 mIU/L and 0.01 µg/L, respectively. Measurement ranges of AFP-TRFIA and HBsAg-TRFIA were 1-1000 mIU/L and 0.2-150 µg/L, respectively. Intra- and inter-assay coefficients of variation of AFP-TRFIA were 3.3-4.1% and 5.7-7.2% and for HBsAg-TRFIA were 2.9-3.9% and 4.9-6.8%, respectively. Linear correlation of TRFIA and chemiluminescence immunoassay measurements resulted in a correlation coefficient of 0.9949 for AFP and 0.9940 for HBsAg. For the endurance test, Eu-labeled McAbs were stable for at least one year at −20°C and the results of the TRFIA with the same reagents were also reproducible after one year. The availability of a highly sensitive, reliable and convenient AFP/HBsAg TRFIA will allow the quantification of both AFP and HBsAg, thereby providing diagnostic value in various clinical conditions and could be applied for clinical use. Copyright © 2012 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Abstract: The S domain of hepatitis B virus surface antigen (sHBsAg) is the primary component for vaccine development against virus infection. For stable expression of sHBsAg in recombinant Saccharomyces cerevisiae, new accessory genes necessary for foreign protein expression were screened by DNA microarray. Among 600 genes of interest, genes coding for an activating protein of ATPase in Hsp90 (Aha1p), S. cerevisiae DnaJ (Scj1p), thioredoxin 2 (Trx2p) and a GTPase-activator specific for Sar1 (Sec23p) as well as Pdi1p were selected in transcriptome analysis, which are known to facilitate disulfide bond formation or induce protein transport in the secretion pathway. Individual and combinatorial expression of SEC23, TRX2 and PDI1 increased total sHBsAg concentration by 1.9–6.5-fold, relative to the control strain expressing sHBsAg only. Additionally, moderate expression of Kex2p protease able to cut off the signal peptide enhanced the portion of the authentic sHBsAg to total sHBsAg. Fed-batch fermentation of the S. cerevisiae 2805 strain coexpressing the sHBsAg, SEC23, PDI1 and KEX2 genes resulted in 70.6mg/L final sHBsAg concentration which was 5.6 times higher than that of the control. Transmission electron microscopic analysis of the yeast cells elucidated the effects of the accessory gene coexpression on the intracellular localization of sHBsAg. Like PDI1, coexpression of both SEC23 and/or TRX2 newly isolated in this study is expected to improve the target protein expression in S. cerevisiae. [Copyright &y& Elsevier]

Abstract: A bio-nanocapsule (BNC) is a hollow nanoparticle consisting of an approximately 100-nm-diameter liposome with about 110 molecules of hepatitis B virus (HBV) surface antigen L protein embedded as a transmembrane protein. BNC can encapsulate various drugs and genes and deliver them specifically to human hepatic cells based on the ability of HBV to recognize human hepatocyte, which is integrated in the N-terminal region of L protein. However, it is elusive whether the cellular attachment and entry into hepatic cells of BNC utilize the early infection mechanism of HBV. In this study, we have found that while all human hepatic cells show distinct affinities for BNC compared to non-hepatic cells, primary hepatocytes shows the highest efficiency for cellular binding and incorporation of BNC. Amounts of BNCs bound weakly and strongly to cell membranes and those entered into the cells varied significantly depending on the types of human hepatic cells. The weak and strong binding modes of BNC are likely mediated through binding to two distinct HBV receptors (heparin-mediated low-affinity and unidentified high-affinity receptors), which play major roles in the early infection mechanism of HBV. The rates of cellular uptake of BNC are similar to those reported for HBV. The BNCs incorporated into the cells are swiftly sorted to either early endosomes or macropinosomes and then to late endosomes and/or lysosomes. These findings strongly suggest that BNC is bound to and incorporated into human hepatic cells according to the early infection mechanism of HBV. [Copyright &y& Elsevier]

Abstract: Bio-nanocapsules (BNCs) are hollow nanoparticles composed of the L protein of hepatitis B virus (HBV) surface antigen (HBsAg), which can specifically introduce genes and drugs into various kinds of target cells. Although the classic electroporation method has typically been used to introduce highly charged molecules such as DNA, it is rarely adopted for proteins due to its very low efficiency. In this study, a novel approach to the preparation of BNC was established whereby a target protein was pre-encapsulated during the course of nanoparticle formation. Briefly, because of the process of BNC formation in a budding manner on the endoplasmic reticulum (ER) membrane, the association of target proteins to the ER membrane with lipidation sequences (ER membrane localization sequences) could directly generate protein-encapsulating BNC in collaboration with co-expression of the L proteins. Since the membrane-localized proteins are automatically enveloped into BNCs during the budding event, this method can be protect the proteins and BNCs from damage caused by electroporation and obviate the need for laborious consideration to study the optimal conditions for protein encapsulation. This approach would be a useful method for encapsulating therapeutic candidate proteins into BNCs. [Copyright &y& Elsevier]

Abstract: Bio-nanocapsules (BNCs) are hollow particles (approx. 50nm diameter) consisting of hepatitis B virus surface antigen (HBsAg) large (L, pre-S1+pre-S2+S) proteins embedded in a unilamellar liposome, sharing the same transmembrane S region with an immunogen of hepatitis B vaccine (i.e., HBsAg small (S) protein particle). BNCs can incorporate drugs and genes into the hollow space and systemic administration of the BNCs can deliver the products to human liver via the human hepatocyte-specific receptor within the pre-S (pre-S1+pre-S2) region displayed on BNC’s surface. Thus, BNCs are expected to offer efficient and safe non-viral nanocarriers to deliver human liver-specific genes and drugs. To date, BNCs have been purified from the crude extract of BNC-overexpressing yeast cells by fractionation with polyethylene glycol followed by one CsCl equilibrium and two sucrose density gradient ultracentrifugation steps. However, the process was inefficient in terms of yield and time, and was not suitable for mass production because of the ultracentrifugation step. Furthermore, trace contamination with yeast-derived proteinases degraded the pre-S region, which is indispensable for liver-targeting, during long-term storage. In this study, we developed a new purification method involving heat treatment and sulfated cellulofine column chromatography to facilitate rapid purification, completely remove proteinases, and enable mass production. In addition, the BNCs were functional for at least 14months after lyophilization with 5% (w/v) sucrose as an excipient. This new process will significantly contribute to the development of forthcoming BNC-based nanomedicines as well as hepatitis B vaccines. [Copyright &y& Elsevier]

Abstract: Background: Unhygienic blood collection in the early 1990s led to blood-borne infections in Central China. This study aimed to estimate human immunodeficiency virus (HIV) co-infection with hepatitis C and B viruses (HCV and HBV) and their risk factors in a rural area of Shanxi Province with a history of commercial blood donation. Methods: A cross-sectional study was conducted in 2004. All adult residents in the target area were invited to participate in the study. Face-to-face interviews were completed and blood specimens were tested for HIV, HCV, and HBV surface antigen (HBsAg). Results: Prevalence rates of HIV, HCV, and HBsAg were 1.3% (40/3 062), 12.7% (389/3 062), and 3.5% (103/2982), respectively. Of the 40 HIV-positive specimens, 85% were HCV positive and 2.5% were HBsAg positive. The history of commercial blood donation was positively associated with HIV, HCV, and HIV/HCV co-infections, but was negatively associated with HBsAg seropositivity. Migration for employment in the last 5 years was positively related to HIV, HBsAg, and HIV/HCV co-infections. Univariate logistic analysis showed that illegal drug use, number of sex partners, extramarital sex behavior, commercial sex behavior, and condom use rate were not related to anti-HIV, anti-HCV, HBsAg seropositivity or their co-infections. Conclusion: The history of commercial blood donation was the main risk factor for HIV, HCV, and HIV/HCV co-infections in this former commercial blood donation area. HIV and HCV prevention and treatment interventions are important in this area. [Copyright &y& Elsevier]

Purpose: Measurement of serum hepatitis B virus surface antigen (HBsAg) levels is important for the management of chronic hepatitis B patients in terms of monitoring response to antiviral therapy. This study aimed to evaluate the diagnostic performance of a new diagnostic kit, which quantitatively measures serum HBsAg level using an immunoradiometric assay (IRMA)-based method. Measurements were compared with those obtained using a chemiluminescent microparticle immunoassay (CMIA)-based method. Methods: The blood samples of 96 patients with chronic hepatitis B were used in this study. Copy numbers of serum hepatitis B virus (HBV) DNA were determined in 23 of these samples. The correlation between and the concordance of IRMA and CMIA results were determined using Pearson's correlation coefficients. P values of 0.05 were considered to be statistically significant throughout. Results: Laboratory diagnoses based on IRMA were wholly in agreement with those based on CMIA. Furthermore, serum HBsAg levels by IRMA were found to be highly correlated with those determined by CMIA (correlation coefficient R = 0.838, p < 0.001). Serum HBsAg level and serum HBV DNA copies were found to be linearly related by both methods ( R = 0.067, p = 0.316 by IRMA, and R = 0.101, p = 0.215 by CMIA). Conclusion: The diagnostic performance of the investigated IRMA method of determining HBsAg levels was found to be comparable with that of a CMIA-based method in chronic hepatitis B patients. [ABSTRACT FROM AUTHOR]

To check feasibility and effectiveness of the α-amylase reporter system, two vectors were designed and tested using hepatitis B virus surface antigen (HBsAg) and Homo sapiens granulocyte-macrophage colony stimulating factor 2 (hGM-CSF2) as a model. By integrating the vector containing two independent cassettes into the same genome locus, high-producing clones of HBsAg (or hGM-CSF2) were screened using the α-amylase as a reporter. Results show there was a positive correlation (Correlation coefficient, R > 0.95) between the yield of recombinant proteins and the α-amylase activity of corresponding transformants, which was independent of the gene dosage. [ABSTRACT FROM AUTHOR]