Dengue is an acute viral disease transmitted by the Aedesaegypti mosquito which is present in most tropical urban areas of the world. There are four antigenically distinct serotypes, designated dengue-1 (DEN-1), dengue-2 (DEN-2), dengue-3 (DEN-3) and dengue-4 virus (DEN-4). In this study, we determined the serotypes of dengue viruses isolated in Guatemala in 1995–1998, and found that DEN-3 viruses appeared in 1995 and became predominant in the following three years. We then sequenced cDNAs from fifteen DEN-3 isolates recovered during 1996–1998. From the nucleic acid sequences and previously determined DEN-3 sequences, a phylogenetic tree was constructed using the neighbor joining method. The tree indicated that all fifteen isolates and other DEN-3 viruses isolated in Sri Lanka, India, Samoa and Mozambique formed subtype III. More than two decades ago, DEN-3 virus was prevalent in the Caribbean, but the isolates obtained at that time belonged to subtype IV. Therefore, we concluded that the 1996–1998 dengue epidemic in Guatemala was caused by DEN-3 strains, imported from a tropical area of Asia or Africa or from a Pacific island.