Fruit flies (Diptera: Tephritidae) are a species-rich and economically important group. The phylogenetic relationships among the many taxa are still to be fully resolved and the monophyly of several groups is still to be confirmed. This paper reports a study of the phylogenetic relationships among 23 economically important tephritid species (representing several major lineages of the family) which examines the sequence of a region of mitochondrial DNA encompassing the cytb, tRNASer and ND1 genes. Substitutions characteristic of particular taxa were found that could help classify members of the family at any developmental stage. The trees obtained by the maximum parsimony, neighbour joining and maximum likelihood methods were generally compatible with present morphological classification patterns. However, the data reveal some characteristics of the phylogenetic relationships of this family that do not agree with present classifications. The results support the probable non-monophyletic nature of the subfamily Trypetinae and suggest that Bactrocera cucurbitae (Coquillet) is more closely related to the genus Dacus than to other species of Bactrocera.