Ryo Yanagawa

Inhibition of virus-induced hemolysis with monoclonal antibodies to different antigenic areas on the hemagglutinin molecule of A/seal/Massachusetts/1/80 (H7N7) influenza virus

SummaryA/seal/Massachusetts/1/80 (H7N7) influenza virus caused maximal hemolysis at pH 5.9. Monoclonal antibodies to each of the four nonoverlapping antigenic areas on the hemagglutinin molecule of the virus inhibited the hemolysis whereas those belonging to two of the groups did not inhibit hemagglutination of the virus. Hemolysis also occurred when the virus was incubated at pH 5.9 prior to addition of erythrocytes. Such hemolysis caused by acid-treated virus was inhibited with the antibodies as well. At pH 5.9, hemagglutination of neither intact virus nor hemagglutinin rosettes was inhibited with any of the monoclonal antibodies, indicating conformational change in the hemagglutinin molecule, at this pH. On the other hand, hemagglutination-inhibition was observed when the antigens were incubated with the monoclonal antibodies at pH 7.0 and then the pH was later shifted to 5.9, suggesting that antibody-binding interferes with the conformational change in the hemagglutinin molecule at pH 5.9. The present findings indicate that antibodies to the hemagglutinin of influenza virus can inhibit virus-induced hemolysis by blocking conformational change in the hemagglutinin molecule and blocking later step of fusion than the conformational change, in addition to blocking attachment of virus to the receptor of erythrocytes.

Antigenic mapping and functional analysis of the F protein of Newcastle disease virus using monoclonal antibodies

SummaryTwelve monoclonal antibodies to the F protein of a velogenic strain of Newcastle disease virus (NDV) were established. Each of these antibodies inhibited virus-induced plaque formation in BHK-21 cells. Seven antibodies neutralized viral infectivity in eggs; thus, antigenic variants could be selected with these antibodies and used for antigenic mapping. Based on the reactivity of the antigenic variants with the antibodies used in selection, 4 distinct antigenic sites (I–IV) were defined on the F protein molecule. In competitive binding assay, sites II and III were found to be spatially close to each other. Each antibody to sites I, II and III inhibited both virus-induced hemolysis of chicken erythrocytes and syncytium formation of BHK-21 cells. On the other hand, some of the antibodies to site IV selectively inhibited either hemolysis or cell fusion. This finding may indicate that the fusion of the viral envelope with erythrocytes and host cell membrane is modulated through different ways. Comparative analysis of different NDV strains using monoclonal antibodies to each of the different antigenic sites showed that the antigenicity of the F protein is highly conserved.

Leptospiral attachment to extracellular matrix of mouse fibroblast (L929) cells

The attachment of leptospires to the extracellular matrix (ECM) remaining after mouse fibroblast (L929) cells on coverslips had been solubilized with Triton X-100 was examined. Each highly virulent line of Leptospira interrogans serovar copenhageni, canicola and pomona attached to ECM more effectively than intermediately virulent and avirulent lines of the same strains, suggesting a correlation between virulence and attachment to ECM. Inhibition of the attachment of highly virulent copenhageni to ECM was found in the presence of the homologous immunoglobulin G Fab fragment.

Contact Infection of Mink with 5 Subtypes of Avian Influenza Virus

SummaryAvian influenza viruses of H3N8, H11N4, H7N7, H8N4, and H5N3 infected mink by contact.

Corynebacterium pilosum and Corynebacterium cystitidis, Two New Species from Cows

Two strains of corynebacteria were isolated from cows that showed signs of cystitis and pyelonephritis. According to the results of numerical-analysis and deoxyribonucleic acid homology studies, these strains differed from the known species of Corynebacterium parasitic on or pathogenic to humans and/or other animals. These strains are regarded as belonging to two new species, for which the names C. pilosum and C. cystitidis are proposed. The type strains of these species are 46 Hara (= ATCC 29592) and 42 Fukuya (= ATCC 29593), respectively.

Leptospiral attachment to cultured cells

Each virulent strain of copenhageni, canicola and pomona of Leptospira interrogans attached effectively to MDCK cells and primary dog kidney cells, while the avirulent or less virulent line of the same strain and avirulent strains belonging to the same serovars and the avirulent reference strains of other serovars did not. Inhibition of the attachment of the virulent copenhageni to MDCK cells was found in the presence of the homologous immunoglobulin G Fab fragment. Strains of L. biflexa attached to the animal cells, but they differed from those of virulent L. interrogans in their capability to attach to glass.

Purification, characterization and serological properties of a glycolipid antigen reactive with a serovar-specific monoclonal antibody against Leptospira interrogans serovar canicola

A glycolipid antigen possessing a serovar-specific antigenic determinant of Leptospira interrogans serovar canicola was purified from a chloroform/methanol extract of the organism. The purification procedures included silicic acid column chromatography and preparative thin-layer chromatography (TLC). Antigenic activity was detected by a TLC-enzyme immunostaining technique using monoclonal antibody CT3, which specifically agglutinates serovar canicola and only weakly serovar sumneri but no other serovars of Leptospira. The purified glycolipid reacted with CT3 antibody, indicating that the glycolipid possessed a serovar-specific antigenic determinant. Infrared spectrum and proton nuclear magnetic resonance analyses showed that the glycolipid contained sugar and lipid moieties, which possessed amide linkages and an acetyl group. Gas-liquid chromatography-mass spectrometry analysis showed that the glycolipid contained two unknown sugars, one of which (unknown sugar II) appeared to be associated with the antigenic determinant specific for canicola. The serovar-specific antigenic determinant was destroyed by mild alkali treatment of the glycolipid. These findings suggested that the antigenic determinant was an alkali-labile moiety which may be related to the unknown sugar II.

Isolation of an Antigenic Oligosaccharide Fraction from Leptospira interrogans Serovar canicola with a Monoclonal Antibody

An oligosaccharide fraction containing the antigenic determinant of lipopolysaccharide antigen (TM antigen) from Leptospira interrogans serovar canicola, recognized by a monoclonal antibody (CT3) which agglutinates serovars canicola and broomi, was isolated by formic acid and successive sulphuric acid hydrolyses. Separation of the antigenic compounds was done by Bio-Gel P-2 and Sephadex G-25 gel filtration, and high-performance liquid chromatography with two different columns. The fraction finally obtained was a mixture of two oligosaccharides, both of which migrated as a single spot having a slightly higher mobility than an authentic tetrasaccharide (stachyose) on thin layer chromatography. The fraction contained rhamnose, arabinose and two major and two minor unknown sugars which were shown to be N- or O-acetylated and/or O-methylated sugars by nuclear magnetic resonance. The fraction inhibited the binding of CT3 antibody with TM antigen in enzyme-linked immunosorbent assay and microscopic agglutination of serovar canicola with the antibody. The inhibitory activity was destroyed by periodate oxidation or mild alkaline treatment, but was resistant to sodium borohydride reduction.

Experimental infection of mink with influenza A viruses

SummaryMink were found to be susceptible to the intranasal inoculation of human, swine, equine and avian influenza A viruses. The viruses were recovered until the 7th post inoculation (p.i.) day from the respiratory tract. The inoculated mink showed antibody response against these viruses. Contact infection in mink with A/Kumamoto/22/77 (H3N2) was possible.

Interference with a conformational change in the haemagglutinin molecule of influenza virus by antibodies as a possible neutralization mechanism

A possible mechanism of neutralization of influenza virus by antibodies to the haemagglutinin molecule is proposed in addition to the generally accepted mechanism of blocking attachment to host cell receptors. This proposed mechanism involves interference with a low-pH-induced conformational change in the haemagglutinin molecule by bivalent binding of antibodies, which results in inhibition of the fusion step in the viral replication process.