Inhibition of hepatitis B virus assembly with synthetic peptides derived from the viral surface and core antigens

The long surface antigen (L-HBsAg) of hepatitis B virus (HBV) plays a central role in the production of infectious virions. During HBV morphogenesis, both the PreS and S domains of L-HBsAg form docking sites for the viral nucleocapsids. Thus, a compound that disrupts the interaction between the L-HBsAg and nucleocapsids could serve as a therapeutic agent against the virus based upon inhibition of morphogenesis. Synthetic peptides correspond to the binding sites in L-HBsAg inhibited the association of L-HBsAg with core antigen (HBcAg). A synthetic peptide carrying the epitope for a monoclonal antibody to the PreS1 domain competed weakly with L-HBsAg for HBcAg, but peptides corresponding to a linear sequence at the tip of the nucleocapsid spike did not, showing that the competing peptide does not resemble the tip of the spike

Nucleotide Sequence Analysis of the HN Gene of the Heat Resistant NDV Strain Af2240 and Substrain V4-UPM

Newcastle disease virus (NOV) is the prototy peavian Paramyxovirus which causes a major disease in poultry. The highlyvi rulentv elogenic NDV strain AF 2240 andavi rulent substrainV4 - UPM possess thermost a blehaemag glutination andneura minidase activities compared to other strains. Therefore,the purpose of this study is to sequence the ha emagglutinin- neura minidase(HN)genes of AF 2240 and V4-UPM. The virus was propagated in the allantoic fluid of 9-10day-old embryonated eggs. The allantoic fluid was harvested after 72 hours incubation at 37°C. Viral purification was carried out on 30% to 60% (N/V ) sucrose density gradient centrifugation at 2 75,000 x g, for 4 hour sat 4°C. The viral RNA was extracted using the hot phenol extraction method. RNA sequencing was performed directly on the genomic RNA using the dideoxy chain termination method. The nucleotide sequence was then confirmed by cycle sequencing and analysed using the computer programs of RIBIODNASIS and MicroGenie which were linked to the Data Bank

Comparison between Phage-ELISA and Phage Dot-Blot Assay Methods for the Detection of Hepatitis B Surface Antigen and its Antibodies in Human Serum

A modified phage-enzyme link immunosorbent assay (phage-ELISA) and a phage dot-blot assay specific for hepatitis B surface antigen (HBsAg) and its antibody were developed by using phage display technology. The phage-ELISA and phage dot-blot assays enabled to detect HBsAg and anti-HBsAg in human sera, and compatible to commercial detection kit. The fusion phages were immobilized onto microtiter plate wells and nitrocellulose membrane sheets, then blocked with 10% milk diluent, and added with human serum at dilution of 1:5000. The absorbance at 405 nm was determined once the colour changes formed. The same human serum also applied on the commercial diagnostic kit for comparison. The statistical analysis was carried out using ANOVA and T Test (LSD) for variable comparison between phage-ELISA and phage dot-blot assays. Based on these studies, the phage-ELISA was found to be more sensitive compared to phage dot-blot assay as the detection of HBsAg in human sera was about 80% as compared to 51.7% by using phage dot-blot assay. Meanwhile, the sensitivity for detection of anti-HBsAg by using phage-ELISA was slightly higher which showed about 83.3%. However, the sensitivity of the assay was dropped almost half when using phage dot-blot assay. Therefore, they are practical to be used as a reliable alternative way for the detection HBsAg and its antibody in human sera

Newcastle disease virus: macromolecules and opportunities

Over the past two decades, enormous advances have occurred in the structural and biological characterization of Newcastle disease virus (NDV). As a result, not only the complete sequence of the viral genome has been fully determined, but also a clearer understanding of the viral proteins and their respective roles in the life cycle has been achieved. This article reviews the progress in the molecular biology of NDV with emphasis on the new technologies. It also identifies the fundamental problems that need to be addressed and attempts to predict some research opportunities in NDV that can be realized in the near future for the diagnosis, prevention and treatment of disease(s)

Recombinant hepatitis B virus core particles: association, dissociation and encapsidation of green fluorescent protein.

The recombinant hepatitis B virus (HBV) core antigen (HBcAg) expressed in Escherichia coli self-assembles into icosahedral capsids of about 35 nm which can be exploited as gene or drug delivery vehicles. The association and dissociation properties of the C-terminally truncated HBcAg with urea and guanidine hydrochloride (GdnHCl) were studied. Transmission electron microscopy (TEM) revealed that the dissociated HBcAg was able to re-associate into particles when the applied denaturing agents were physically removed. In order to evaluate the potential of the particles in capturing molecules, purified green fluorescent protein (GFP) was applied to the dissociated HBcAg for encapsidation. The HBcAg particles harbouring the GFP molecules were purified using sucrose density gradient ultracentrifugation and analysed using native agarose gel electrophoresis and TEM. A method for the encapsidation of GFP in HBcAg particles which has the potential to capture drugs or nucleic acids was established

Fighting the hepatitis B virus: past, present & future

Hepatitis B virus (HBV) is one of the greatest killers of human beings. The production of recombinant HBV vaccine via genetic engineering and mass immunization of the world population with the vaccine have had huge success in bringing down the number of infected people. However, until now there is no effective treatment for HBV infections. Nucleoside analogues are commonly used to treat chronic hepatitis B patients, but prolonged treatments have resulted in the selection of drug resistant strains and vaccine escape mutants. More importantly, HBV is linking arms with the human immunodeficiency virus (HIV) to kill more humans. Many dedicated scientists and highly motivated doctors have risked their lives to control the spread of HBV and search for a cure for this deadly virus. Their ultimate goal is to eradicate the virus from this planet and make the world a better place for everyone. Therefore, the main aim of this talk is to provide a comprehensive review of their contributions in the development of vaccines, diagnostic assays, therapeutics and drug delivery systems to fight HBV

Automatic Detection and Analysis of Outdated Documentation in GitHub Repositories

Outdated documentation is a pervasive problem in software development, preventing effective use of software, and misleading users and developers alike. We posit that one possible reason why documentation becomes out of sync so easily is that developers are unaware of when their source code modifications render the documentation obsolete. Ensuring that the documentation is always in sync with the source code takes considerable effort, especially for large codebases. To address this situation, we propose an approach that can automatically detect code element references that survive in the documentation after all source code instances have been deleted. In this work, we analysed more than 3,000 GitHub projects and found that most projects contain at least one outdated code element reference at some point in their history. We submitted GitHub issues to real-world projects containing outdated references detected by our approach, some of which have already led to documentation fixes. As an initiative toward keeping documentation in software repositories up-to-date, we have made our implementation available and created a tool for developers to scan their GitHub projects for outdated code element references. Lastly, we extended our approach to detect outdated references to code elements in over 2,000 images present in software documentation.Thesis (MPhil) -- University of Adelaide, School of Computer and Mathematical Sciences, 202

Detection and precipitation of hepatitis B core antigen using a fusion bacteriophage

The nucleocapsids of hepatitis B virus (HBV) are made of 180 or 240 subunits of core proteins or known as core antigens (HBcAg). A fusion bacteriophage bearing the WSFFSNI sequence that interacts tightly to HBcAg was employed as a diagnostic reagent for the detection of the antigen using the phage-enzyme-linked immunosorbent (phage-ELISA), dot blot and immunoprecipitation assays. The results from phage-ELISA and dot blot assay showed that as low as 10 ng of HBcAg can be detected optimally by 1.0×1012 pfu/ml fusion M13 bacteriophage. The sensitivity of the dot blot assay corresponds with that of the phage-ELISA. HBcAg in HBV positive serum samples can also be detected using the fusion phage via the phage-ELISA and phage-dot blot assay. The phage cross-linked to cyanogen bromide (CNBr) activated agarose can also be used to precipitate HBcAg in bacterial lysate. The optimum amount of phage needed for cross-linking to 1 g of agarose is about 7.0×106 pfu/ml which could also precipitate purified and unpurified HBcAg in bacterial lysate. This study demonstrates the potential of fusion bacteriophage bearing the sequence WSFFSNI as a diagnostic reagent and a ligand for the detection and purification of HBcAg respectively