In latest years, the importance of the Melaleuca alternifolia essential oil (EO) has been greatly empathised due to its anti-microbial and anti-inflammatory effects, as well as to its toxic properties towards many arthropods of great medical and veterinary importance. In this research, the EO extracted from aerial parts of M. alternifolia was evaluated for its toxicity against larvae of the most invasive mosquito worldwide, Aedes albopictus (Diptera: Culicidae), and towards adults of the water flea, Daphnia magna (Cladocera: Crustacea), a non-target aquatic organism that share the same ecological niche of A. albopictus. The chemical composition of M. alternifolia EO was investigated by GC-MS analysis. Tea tree EO was mainly composed by oxygenated monoterpenes, with 1,8-cineole as the major constituent. M. alternifolia EO exerted toxic activity against A. albopictus larvae, with a LC50 = 267.130 ppm. However, this EO had a remarkable acute toxicity also towards adults of the non-target arthropod D. magna, with a LC50 = 80.636 ppm. This research provide useful information for the development of newer and safer mosquito control tools, highlighting that the non-target effects against aquatic organisms that share the same ecological niche of A. albopictus larvae are crucial in the development of ecofriendly mosquito control strategies. Further research is needed to investigate the chronic and/or reproductive toxicity of M. alternifolia EO both towards target and non-target aquatic arthropods.