The feeding behavior of sand flies provides valuable information about the vector/host interactions and elucidates the epidemiological patterns of American cutaneous leishmaniasis (ACL) transmission. The aim of this study was to identify the blood meal sources of sand flies in endemic areas of leishmaniasis in Parana State through polymerase chain reaction (PCR) amplification of a prepronociceptin (PNOC) gene fragment and its subsequent DNA sequencing. Moreover, molecular assays were conducted to evaluate the sensitivity and reproducibility of the PNOC gene amplification. Besides that, a time-course digestion test of the blood using sand flies that fed artificially on BALB/c mice was performed. Of 1263 female sand flies collected in the field, 93 (3.6%) specimens were engorged and 27 allowed efficient amplification of the PNOC gene. These flies had fed on equine (Equus caballus), porcine (Sus scrofa) and canine (Canis lupus familiaris) species. The results also showed that the identification of the blood meal sources of the sand flies using the molecular method was directly linked to the level of digestion of the blood (time-course) and not to the amount of blood that had been ingested or to the presence of inhibitors in the blood Ver menos