Anastrepha obliqua (Macquart) (Diptera: Tephritidae), the West Indian fruit fly, is a frugivorous pest that occasionally finds its way to commercial growing areas outside its native distribution. It inhabits areas in Mexico, Central and South America, and the Caribbean with occasional infestations having occurred in the southern tier states (California, Florida, and Texas) of the United States. This fly is associated with many plant species and is a major pest of mango and plum. We examine the genetic diversity of the West Indian fruit fly based on mitochondrial COI and ND6 DNA sequences. Our analysis of 349 individuals from 54 geographic collections from Mexico, Central America, the Caribbean, and South America detected 61 haplotypes that are structured into three phylogenetic clades. The distribution of these clades among populations is associated with geography. Six populations are identified in this analysis: Mesoamerica, Central America, Caribbean, western Mexico, Andean South America, and eastern Brazil. In addition, substantial differences exist among these genetic types that warrants further taxonomic review.