Attenuation markers of a candidate dengue type 2 vaccine virus, strain 16681 (PDK-53), are defined by mutations in the 5 ' noncoding region and nonstructural proteins 1 and 3
Autor(es): Butrapet, S; Huang, CYH; Pierro, DJ; Bhamarapravati, N; Gubler, DJ; Kinney, RM
Resumo: The genome of a candidate dengue type 2 (DEN-2) vaccine virus, strain PDK-53, differs from its DEN-2 16681 parent by nine nucleotides. Using infectious cDNA clones, we constructed 18 recombinant 16681/PDK-53 viruses to analyze four 16681-to-PDK-53 mutations, including 5' noncoding region (5'NC)-57 C-to-T, premembrane (prM)-29 Asp-to-Val (the only mutation that occurs in the structural proteins), nonstructural protein 1 (NS1)-53 Gly to-Asp, and NS3-250 Glu-to-Val, The viruses were studied for plaque size, growth rate, and temperature sensitivity in LLC-MK(2) cells, growth rate in C6/36 cells, and neurovirulence in newborn mice. All of the viruses replicated to peak titers of 10(7.3) PFU/ml or greater in LLC-MK(2) cells, The crippled replication of PDK-53 virus in C6/36 cells and its attenuation for mice were determined primarily by the 5'NC-57-T and NS1-53-Asp mutations. The temperature sensitivity of PDK-53 virus was attributed to the NS1-53-Asp and NS3-250-Val mutations. The 5'NC-57, NS1-53, and NS3-250 loci all contributed to the small-plaque phenotype of PDK-53 virus. Reversions at two or three of these loci in PDK-53 virus were required to reconstitute the phenotypic characteristics of the parental 16681 virus. The prM-29 locus had little or no effect on viral phenotype, Sequence analyses showed that PDK-53 virus is genetically identical to PDK-45 virus. Restriction of the three major genetic determinants of attenuation markers to nonstructural genomic regions makes the PDK-53 virus genotype attractive for the development of chimeric DEN virus vaccine candidates.
Palavras-Chave: Yellow fever - Virus; Full length Cdna; Antibody - Dependent Enhancement; Japanese Encephalitis - Virus; Tick borne encephalitis; Aedes aegypti; Genetic - Determinants; Mouse neurovirulence; Secondary structure; Poliovirus vaccine
Imprenta: Journal of Virology, v. 74, n. 7, p. 3011-3019, 2000
Identificador do objeto digital: 10.1128/JVI.74.7.3011-3019.2000
Descritores: Aedes aegypti - Cell ; Aedes aegypti - Genome ; Aedes aegypti - Neurovirulence ; Aedes aegypti - Virus
Data de publicação: 2000
