Aedes aegypti, also known as the yellow fever mosquito, is the main vector of several arboviruses. In Ecuador, dengue and chikungunya are the most prevalent mosquito-borne diseases. Hence, there is a need to understand the population dynamics and genetic structure of the vector in tropical areas for a better approach towards effective vector control programs. This study aimed to assess the genetic diversity of Ae. aegypti, through the analyses of the mitochondrial gene ND4, using a combination of phylogenetic and population genetic structure from 17 sites in Ecuador. Results showed two haplotypes in the Ecuadorian populations of Ae. aegypti. Haplotype 1 was closely related to Ae. aegypti reported from America, Asia, and West Africa. Haplotype 2 was only related to samples from America. The sampled vectors from the diverse localities showed low nucleotide diversity (π = 0-0.01685) and genetic differentiation (FST = 0.152). AMOVA analyses indicated that most of the variation (85-91%) occurred within populations, suggesting that geographical barriers have little effect on the genetic structure of Ecuadorian populations of Ae. aegypti. These results agree with the one main population (K = 1) detected by Structure. Vector genetic identity may be a key factor in the planning of vector control strategies.