Nucleotide sequences of the envelope protein genes of 19 geographically and temporally distinct dengue (DEN)-4 viruses were determined. Nucleic acid sequence comparison revealed that the identity among the DEN-4 viruses was greater than 92%. Similarity among deduced amino acids was between 96 and 100%; in most cases identical amino acid substitutions occurred among viruses from similar geographical regions. Alignment of nucleic acid sequences followed by parsimony analysis generated phylogenetic trees, which indicated that geographically independent evolution of DEN-4 viruses had occurred. DEN-4 viruses were separated into two genetically distinct subtypes (genotypes). Genotype-1 contains viruses from the Philippines, Thailand and Sri Lanka; genotype-2 consists of viruses from Indonesia, Tahiti, the Caribbean Islands (Puerto Rico, Dominica) and Central and South America.