Phenotypic characterization of cell culture-derived hepatitis E virus subjected to different chemical treatments: Application in virus removal via nanofiltration.

Safety evaluation for the hepatitis E virus (HEV) is required for plasma fractionation products. Plasma-derived HEV (pHEV) is quite unique in that it is associated with a lipid membrane, which, when stripped during manufacturing processes, induces morphological changes in the virus, making it difficult to select proper HEV phenotypes for clearance studies. We developed a convenient system for the preparation of a high titer cell culture-derived HEV (cHEV).

Quantitative PCR evaluation of parvovirus B19 removal via nanofiltration.

When human parvovirus B19 (B19) is removed from plasma-derived products by nanofiltration, non-infectious fragmented B19 DNA in filtrate prevents quantitative real time PCR (qPCR) from accurately evaluating reduction of the virus particles. To determine optimal target sequence length for detection of full-length B19 genome in the viral particles by qPCR, we analyzed 4 different sequences ranging from 372 to 1,980 bp and found that a 989 bp sequence shows a well-balanced performance for the sensitivity and the run time. Nuclease treatment of filtrates prior to qPCR is also expected to decrease the influence of the residual B19 DNA, but extremely high protein concentration of plasma-derived products in filtrates may result in incomplete digestion of the B19 DNA.

Hepatitis E virus derived from different sources exhibits different behaviour in virus inactivation and/or removal studies with plasma derivatives.

Hepatitis E virus (HEV) causes viral hepatitis, and is considered a risk factor for blood products. Although some HEV inactivation/removal studies have been reported, detailed investigations of different manufacturing steps as heat treatment, partitioning during cold ethanol fractionation, low pH treatment, and virus filtration have yet to be reported for plasma-derived medicinal products. In this study, human serum- and swine faeces-derived HEVs, with and without detergent treatment, were used.

Mode of swine hepatitis E virus infection and replication in primary human hepatocytes.

The aim of this study was to investigate the infection and replication of swine-derived hepatitis E virus (HEV) in primary cultured human hepatocytes (PHCs). Hepatocytes were cultured from the resected normal livers of patients with metastatic tumours. These cultured hepatocytes were infected with swine-derived genotype 3 or 4 HEV.

Immunization of rabbits with synthetic peptides derived from a highly conserved β-sheet epitope region underneath the receptor binding site of influenza A virus.

Background: There is increasing concern about the speed with which health care providers can administer prophylaxis and treatment in an influenza pandemic. Generally, it takes several months to manufacture an influenza vaccine by propagation of the virus in chicken eggs or cultured cells. Newer, faster protocols for the production of vaccines that induce broad-spectrum immunity are therefore highly desirable.

Induction of anti-influenza immunity by modified green fluorescent protein (GFP) carrying hemagglutinin-derived epitope structure.

The development of vaccination methods that can overcome the emergence of new types of influenza strains caused by escape mutations is desirable to avoid future pandemics. Here, a novel type of immunogen was designed that targeted the conformation of a highly conserved region of influenza A virus hemagglutinin (HA) composed of two separate sequences that associate to form an anti-parallel β-sheet structure. Our previous study identified this β-sheet region as the structural core in the epitope of a characteristic antibody (B-1) that strongly neutralizes a wide variety of strains within the H3N2 serotype, and therefore this β-sheet region was considered a good target to induce broadly reactive immunity against the influenza A virus.

Highly conserved sequences for human neutralization epitope on hemagglutinin of influenza A viruses H3N2, H1N1 and H5N1: Implication for human monoclonal antibody recognition.

The epitope sequences within the hemagglutinin (HA) of influenza A virus H3N2 at amino acid residues 173-181 and 227-239 that forms anti-parallel beta-sheet structure are similarly recognized by human monoclonal antibodies (HuMAbs), B-1 and D-1 that we recently obtained using the peripheral blood lymphocytes from two influenza-vaccinated volunteers. Both HuMAbs showed strong global neutralization of H3N2 strains. Here we show the significant conservation of the beta-sheet region consisting of the above-mentioned two epitope regions in H3N2.

Broad neutralizing human monoclonal antibodies against influenza virus from vaccinated healthy donors.

Human monoclonal antibodies (HuMAbs) prepared from patients with viral infections could provide information on human epitopes important for the development of vaccines as well as potential therapeutic applications. Through the fusion of peripheral blood mononuclear cells from a total of five influenza-vaccinated volunteers, with newly developed murine-human chimera fusion partner cells, named SPYMEG, we obtained 10 hybridoma clones stably producing anti-influenza virus antibodies: one for influenza A H1N1, four for influenza A H3N2 and five for influenza B. Surprisingly, most of the HuMAbs showed broad reactivity within subtype and four (two for H3N2 and two for B) showed broad neutralizing ability.