About 50 Chrysomyinae specimens belonging to three forensic relevant species (Chrysomia albiceps, Phormia regina and Protophormia terraenovae) were collected from different geographical locations in Belgium over the last 5 yr. A 304-bp fragment of their mitochondrial COI gene is sequenced. The monophyletic branches of the phylogenetic tree reveal that this marker is suitable for discrimination between these species. The intra versus interspecific variability marks clear threshold levels for DNA barcoding. Nineteen Chrysomyinae specimens, collected from four locations in France, show mitotypes that are identical or at least very similar to the Belgian mitotypes. Considering additional specimens from outside of Europe reveals no intraspecific geographical variation within C. albiceps and P. terraenovae, whereas P. regina is subbranched in a Belgian-French and a USA-Chinese population.