American tegumentary leishmaniasis, a zoonotic disease caused by theLeishmaniagenus, poses significant challenges in treatment, including administration difficulty, low efficacy, and parasite resistance. Novel compounds or associations offer alternative therapies, and natural products such as oregano essential oil (OEO), extracted fromOriganum vulgare, have been extensively researched due to biological effects, including antibacterial, antifungal, and antiparasitic properties. Silver nanoparticles (AgNp), a nanomaterial with compelling antimicrobial and antiparasitic activity, have been shown to exhibit potent leishmanicidal properties. We evaluated the in vitro effect of OEO and AgNp-Bio association onL. amazonensisand the death mechanisms of the parasite involved. Our results demonstrated a synergistic antileishmanial effect of OEO + AgNp on promastigote forms andL. amazonensis-infected macrophages, which induced morphological and ultrastructural changes in promastigotes. Subsequently, we investigated the mechanisms underlying parasite death and showed an increase in NO, ROS, mitochondrial depolarization, accumulation of lipid-storage bodies, autophagic vacuoles, phosphatidylserine exposure, and damage to the plasma membrane. Moreover, the association resulted in a reduction in the percentage of infected cells and the number of amastigotes per macrophage. In conclusion, our findings establish that OEO + AgNp elicits a late apoptosis-like mechanism to combat promastigote forms and promotes ROS and NO production in infected macrophages to target intracellular amastigote forms.