Cell surface expression of a chimeric protein containing mouse immunoglobulin G1 Fc domain and its immunological property

Recently we reported that a chimeric molecule containing mouse transferrin receptor and immunoglobulin G1 (IgG1) Fc, mTR-Fc, induced higher immune responses and can be used as a vaccine adjuvant. In this study, the immunological property of the molecule was investigated. Although, the mTR-Fc did not activate complement classical pathway, it was recognized by activated macrophage as like intact IgG Fc, which is recognized by macrophage via Fcgamma receptor. In addition, we found that splenocyte simultaneously exposed to lipopolysaccaride (LPS) and mTR-Fc produced higher amount of interleukin-10, comparing to that exposed to only LPS. These results suggest that the mTR-Fc molecules conserved the IgG Fc property to biasing immune responses via modulation of cytokine production by antigen presenting cell.     

A fusion protein of IgG fc and mouse-derived antigen on the surface of pseudorabies virus particles does not accelerate production of harmful auto-reactive antibodies

Previously we reported that immunization with pseudorabies virus (PRV), harboring chimeric Fc on the surface of the virus particles (PRV/Fc), induced higher immune responses than normal PRV particles. The chimeric Fc was fused with mouse transferrin receptor of transmembrane domain (mTR) and the Fc region of immunoglobulin G1. Since it has been reported that some chimeric protein of Fc and self-antigen induce auto-reactive antibodies, in this present study, we examined whether PRV/Fc induces auto-reactive antibodies that react with mTR. PRV/Fc immunized mice produced higher levels of anti-PRV antibodies and antibodies that reacted with mouse-derived 3T3/A31 cells (A31 cell), compared to normal PRV immunized mice. However, antibodies that reacted with mTR in A31 cells were not detected in both Western blot analyses and indirect immunofluorescence assay. The antibodies reacted with an antigen of approximately 16 kDa in A31 cells, but this antigen has a different molecular mass from that of mTR. The antibody also reacted with the antigen of approximately 16 kDa in RK13 cells in which the virus had been propagated. In addition, antibodies induced by immunization with normal PRV also reacted with the same antigen in A31 and RK13 cells. Moreover, neither kidney disorders, in which high levels of mTR were expressed, nor clinical symptoms of autoimmune diseases were observed in mice immunized with either PRV or PRV/Fc. These results indicated that the antibodies were not induced by mTR-Fc, but were instead induced by trace amounts of RK13 derived antigens contained in PRV or PRV/Fc preparations, and cross-reacted with equivalent molecules in mouse derived A31 cells. Therefore, this study confirmed that immunization with PRV/Fc did not induce harmful auto-reactive antibodies.     

Screening of Neutrophil Activating Factors from a Metagenome Library of Sponge-Associated Bacteria

Marine sponge-associated bacteria are known as bio-active compound produce. We have constructed metagenome libraries of the bacteria and developed a metagenomic screening approach. Activity-based screening successfully identified novel genes and novel enzymes; however, the efficiency was only in 1 out of 10⁴ clones. Therefore, in this study, we thought that bioinformatics could help to reduce screening efforts, and combined activity-based screening with database search. Neutrophils play an important role for the immune system to recognize excreted bacterial by-products as chemotactic factors and are recruited to infection sites to kill pathogens via phagocytosis. These excreted by-products are considered critical triggers that engage the immune system to mount a defense against infection, and identifying these factors may guide developments in medicine and diagnostics. We focused on genes encoding amino acid ligase and peptide synthetase and selected from an in-house sponge metagenome database. Cell-free culture medium of each was used in a neutrophil chemiluminescence assay in luminol reaction. The clone showing maximum activity had a genomic sequence expected to produce a molecule like a phospho-N-acetylmuramyl pentapeptide by the metagenome fragment analysis.     

Genetic characterization of chicken anemia viruses newly isolated from diseased chicks in Japan in 2020

In this study, a total of nine chicken samples obtained from two broiler flocks in Oita and Tottori prefectures in 2020 were examined for Chicken anemia virus (CAV) infection. The samples were collected from clinically suspected flocks and diseased chickens. The CAV genome was detected in all nine samples tested by real-time PCR. Phylogenetic analyses and sequence comparisons of the full-length VP1 gene sequences indicated that all the Japanese CAV strains obtained in this study formed a similar cluster of genotype III and shared high nucleotide (99.62–100%) identity. The current Japanese CAV strains were closely related to Chinese CAV strains but not related to vaccine strains. One positive selection site of VP1 was detected among the Japanese CAV strains.     

Characteristics of effective marine protected areas in Hawaiʻi

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Isolation of avian bornaviruses from psittacine birds using QT6 quail cells in Japan

Avian bornaviruses (ABVs) were recently discovered as the causative agents of proventricular dilatation disease (PDD). Although molecular epidemiological studies revealed that ABVs exist in Japan, no Japanese isolate has been reported thus far. In this study, we isolated four strains of Psittaciform 1 bornavirus from psittacine birds affected by PDD using QT6 quail cells. To our knowledge, this is the first report to isolate ABVs in Japan and to show that QT6 cells are available for ABV isolation. These isolates and QT6 cells would be powerful tools for elucidating the fundamental biology and pathogenicity of ABVs.