Ghir-Karzin district is one of the most important endemic foci of visceral leishmaniaisis (VL) in Fars province of southern Iran. To identify the vector(s) of Leishmania in this focus, a total of 2,539 sand flies were collected during June to September 2005. Eleven species (six Phlebotomus and five Sergentomyia) were identified, of which Phlebotomus papatasi Scopoli and Phlebotomus major Annandale were the first (37.4%) and third (11.2%) most common species, respectively. Natural leptomonad infections were observed in two P. papatasi (4.25%) and three P. major (6.65%) specimens on dissection and microscopic examination. Using a Leishmania genus-specific standard polymerase chain reaction (PCR; primers RV1-RV2) and a species-specific nested-PCR (primers LINR4, LIN17, and LIN 19), Leishmania infantum and Leishmania major kinetoplast minicircle DNA was detected in 6 of 72 P. major (8.3%) and 4 of 65 P. papatasi (6.1%), respectively. This is the first detection of L. infantum in P. major, implicating this sand fly as a probable vector of VL in Iran.