Studies on the Antigenic Structure of Japanese Encephalitis Virus Using Monoclonal Antibodies

Abstract

The antigenic relationships among 11 strains of Japanese encephalitis ( JE) virus were analyzed by using monoclonal antibodies (NARMA) against the Nakayama-RFVL strain in hemagglutination-inhibition (HI) and neutralization (Nt) tests. Of the 14 JE virus-specific HI antibodies, all except NARMA 5 showed Nt reactivity with the homologous strain. The HI and Nt titers of these antibodies were not parallel. The 14 antibodies included the following characteristic antibodies : NARMA 3 is a species-specific antibody with HI and Nt reactivities against JE virus, NARMA 13 is a species-specific HI antibody, NARMA 6 is a Nakayama strain-specific antibody with HI and Nt reactivities, and NARMA 5 is a Nakayama strain-specific HI antibody. The 11 strains of JE virus were divided into four major antigenic groups. However, slight antigenic differences were found among some strains of the same group. Furthermore, competitive binding assays were performed to determine the distribution of antigenic determinants by enzyme-linked immunosorbent assay. The results suggest the existence of at least five HI sites on the JE virus virion, and indicate that the JE species-specific HI site and the flavivirus genusspecific HI site are topologically distinct.Comparative studies of the antigenic structure of Japanese encephalitis ( JE) virus have been performed by many investigators (1,5,12,17,21) since Hale and Lee (4) observed antigenic variation among the JE virus strains isolated in Malaya. However, the immunological classification of strains of JE virus is not yet clear. In the previous study (13), we produced 26 monoclonal antibodies (NARMA 1 to 26) that reacted with the Nakayama-RFVL strain of JE virus in hemagglutinationinhibition (HI) tests. These antibodies fell into four groups: JE species-, subgroup-, and genus-specific antibodies and antibodies reactive with JE and Murray Valley encephalitis (MVE) viruses. Moreover, the 27 JE virus strains were divided into four major antigenic groups by using five JE species-specific monoclonal antibodies in the HI test, and it was shown that the Nakayama-Yakken strain is a mutant which lacks the Nakayama strain-specific antigen and that the recently isolated strains are immunologically different from the Nakayama or JaGAr 01 strains.In this report, we describe the characteristics of the 14 JE species-specific monoclonal antibodies in the neutralization (Nt) test and the antigenic relationships 106 9

In vitro growth, pathogenicity and serological characteristics of the Japanese encephalitis virus genotype V Muar strain

“…The amino acid differences between the vaccine strains and the GI strains did not exceed 2.4 %, indicating that the difference may not influence the efficacy of the vaccines. Previous studies have identified individual residues within the E protein that are related to the recognition of JEV by neutralizing antibodies (Crill & Chang, 2004;Goncalvez et al, 2008;Kobayashi et al, 1985;Morita et al, 2001;Vogt et al, 2009;Wu et al, 2003). Although the sequence variation between the vaccine strains and GV strains do not correspond to epitope residues, only a limited number of residues could be responsible for the decreased neutralization ability of the vaccines against GV strains.…”

Effect of Cytokines on Japanese Encephalitis Virus Production by Human Monocytes

“…The JE virus Kamiyama strain (15), which was isolated from human brain in Fukuoka (Japan) in 1966, was used in this study. The virus was propagated in C6/36 cells (9) as described previously (15) Virus titration and determination of number of infected cells. The virus titer was assayed by plaque formation on Vero cells by the method of Kimura-Kuroda and Yasui (12) .…”

“…The number of infected monocytes was determined by immunoperoxidase staining using horseradish peroxidase (HRPO)-labeled F(ab')2 fragment of NARMA 7 (JE species-specific monoclonal antibody) (14) at 48 hr after infection as described previously (15) . The infected monocytes were air-dried on glass slides and fixed with acetone at -20 C for 10 min, and reacted with HRPO-F (ab') 2 fragment of NARMA 7 at 37 C for 1 hr.…”

Nucleotide and deduced amino acid sequence of the structural protein genes of Japanese encephalitis viruses from different geographical locations

“…The amino acid change at E51 would change the conformation of the E protein in this region. The 10-fold haemagglutination inhibition activity difference of Narma3 between strains Nakayama and Kamiyama may result from their amino acid difference at E-51 (Kobayashi et al, 1985).…”

“…Antigenic, biochemical and genetic differences between JE virus strains isolated from different areas have been observed by cross-neutralization (Banerjee, 1975), oligonucleotide fingerprinting (Banerjee & Ranadive, 1984;Hori, 1986), monoclonal and polyclonal antibody reactivities (Kobayashi et al, 1984(Kobayashi et al, , 1985Wills et al, 1992) and by limited (240 bp) genomic sequence comparison (Chen et al, 1990(Chen et al, , 1992Huong et al, 1993). Chen et al (1990Chen et al ( , 1992 suggested that JE viruses could be divided into four genotypes using 12% nucteotide divergence as a cut-off point for virus relationships when comparing prM gene segments.…”