Chironomidae, or non-biting midges, are found worldwide in a wide variety of aquatic habitats. During periods of mass adult eclosion they can become a nuisance and health hazard. Current control methods target the aquatic larval stage and include the use of insect growth regulators or insecticides, which may be prohibited in certain environments or affect non-target organisms. The aim of this study was to investigate whether entomopathogenic nematodes (EPNs) of the families Steinernematidae and Heterorhabditidae, currently employed for control of terrestrial crop pests, could be used as a viable biocontrol for the aquatic larval stages of the Chrionomidae, offering an alternative to current chemical methods. We demonstrate that Steinernema feltiae (Filipjev, 1934), Steinernema carpocapsae (Weiser, 1955), Steinernema kraussei (Steiner, 1923) and Heterorhabditis bacteriophora (Poinar, 1975) are able to survive in water up to 96 h and are able to parasitize and kill Chironomus plumosus (Linnaeus, 1758) larvae, with mortality observed after just 24 h exposure and with < 20% survival after 4 days. We also show that following application to the water column, EPNs sink to the bottom of the lentic water body and can remain alive for more than 96 h. Taken together, we believe that several EPN species could be developed as a valid form of biocontrol for Chironomidae.