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Based on nucleotide sequences, we re-identified representative samples of Phlebotomus perfiliewi originating from two different biogeographical areas of Tunisia, whose populations had previously been identified based on morphological criteria. A partial region of the mitochondrial DNA cytochrome b gene was targeted, and sandfly species was determined by analogy with DNA sequences available in the GenBank database via a BLAST analysis, taking into account the query coverage and percentage identity. The recognized species presents the most substantial homology with the analyzed sequence. The results of the molecular identification showed complete agreement with the morphological identifications, and Phlebotomus perfiliewi is genetically variable.