Mutations in the envelope protein of japanese encephalitis virus affect entry into cultured cells and virulence in mice

The premembrane and envelope (E) genes of a full‐length cDNA clone of the dengue type‐4 (DEN4) virus 814669 strain were replaced with those of the Japanese encephalitis (JE) virus JaOH0566 strain. The in vitro‐synthesized RNA transcripts prepared from chimeric cDNA were used to transfect mosquito C6/36 cells. A viable chimeric virus (designated DEN4/JE) was recovered. Unexpectedly, DEN4/JE exhibited restricted growth in Vero cells. After a serial passage in Vero cells, the Vero‐adapted chimeras were obtained (two clones, designated Strain I and Strain II, respectively). The entire genomes of DEN4/JE, Strain I, and Strain II were sequenced and compared. There were multiple mutations, but amino acid substitutions occurred only in E and nonstructural (NS) protein NS4B. Our findings in this study indicate that the 5′ nontranslated region, E, and NS4B may be involved in Vero cell adaptation in this chimeric system.

Section: Discussionmentioning

confidence: 99%

Molecular Basis for Adaptation of a Chimeric Dengue Type‐4/Japanese Encephalitis Virus to Vero Cells

Tang

Eshita

Tadano

et al. 2005

“…Amino-acid substitutions at position E-366 were also found in peptides with T-helper cell epitopes for Murray Valley encephalitis (MVE) virus (36). Phenotype changes among flaviviruses have also been found to be associated with the amino-acid alteration of E proteins (7,19,24,34).…”

Section: Discussionmentioning

confidence: 99%

Antigenic and Genetic Variations among Japanese Encephalitis Virus Strains Belonging to Genotype 1

Ali

Igarashi

1997

Hyperimmune antisera against four Japanese encephalitis (JE) virus strains, ThCMAr4492 and ThCMAr6793 from Thailand and Nakayama and JaGAr01 from Japan, were used to analyze the antigenic relationships among 12 Thai strains belonging to genotype 1, and two Japanese strains and one Chinese strain belonging to genotype 3. The antiserum for ThCMAr6793 significantly neutralized nine of the 12 Thai strains, none of which was significantly neutralized by antisera for the Nakayama and JaGAr01 strains. The antiserum for ThCMAr4492 neutralized only its homologous strain; therefore, ThCMAr4492 was antigenically different from all other strains. Two Thai strains (Subin and KE-093/83) were significantly less neutralized by all four of the antisera tested. In the deduced amino-acid sequence of the E protein, the 12 Thai strains revealed 100 to 98.2 % identity among them and 90.0 to 98.8 % identity with the published strains, respectively. Among significant amino-acid substitutions, three residues at positions E-222, E-327 and E-366 were shared by all of the Thai strains, whereas residues at E-89, E-123, E-131, E-178, E-293, E-351 and E-373 seemed to be strain-specific. The amino acids at positions E-178, E-327, E-351, E-373 and E-366 are found either in the peptides with functional T-helper cell epitopes or in the ectodomain of the E protein of other flaviviruses. These amino acids may therefore be responsible for determining the antigenic heterogeneity of these strains.

“…Molecular markers of virulence and attenuation have most frequently been shown to occur among flaviviruses (5,11,14,16,19,31,41). Recent studies (24) on 11 strains of DEN-2 virus showed that several structural differences in viral genome could be responsible for the pathogenesis of severe disease in Southeast Asia.…”

mentioning

confidence: 99%

Severity‐Related Molecular Differences among Nineteen Strains of Dengue Type 2 Viruses

Pandey

Igarashi

2000

Comparative nucleotide sequencing was carried out on dengue type 2 virus (DEN-2) strains isolated from patients in Northeast Thailand during the epidemic season in 1993. The patients exhibited different clinical manifestations ranging from dengue fever (DF) to dengue haemorrhagic fever (DHF)/dengue shock syndrome (DSS). The results classified 19 DEN-2 strains into 3 subtypes according to nonsynonymous amino acid replacements. The strain isolated from a DSS patient eliciting secondary serological response belonged to subtype I, whereas 13 strains isolated from DHF patients with secondary response and 2 strains from DF patients with primary response belonged to subtype II. On the other hand, 3 strains isolated from DF cases evoking either primary or secondary response belonged to subtype III. These results suggest that subtype III virus infection could result in clinically milder manifestation irrespective of the serological response compared with subtype I or II viruses. The RNA secondary structure predicted for the 3' noncoding region showed 4 different structures (A, B, C, and D). The result also indicates that different subtypes of DEN-2 serotypes are circulating in a single epidemic in Thailand.