Currently, the potential impact of insecticide mixtures to nontarget organisms is largely unknown, and additional study is needed. The present study investigated the mixture toxicity of the organophosphate insecticide phostebupirim and the pyrethroid insecticide cyfluthrin using 4 nontarget species including Daphnia magna, Hyalella azteca, Pimephales promelas (fathead minnow), and Danio rerio (zebrafish). For each species, the toxicity of equipotent mixtures was compared with the expected toxicity estimated using the independent action (IA) and concentration addition (CA) models. Lethal and sublethal responses to D. magna and H. azteca were best described with the IA model. For both fish species, mixture toxicity was significantly higher than that estimated using either mixture model. The synergism noted in fish exposed to the combination of phostebupirim and cyfluthrin was confirmed by exposing P. promelas larvae to a nontoxic dose of phostebupirim and a range of toxic cyfluthrin concentrations, and vice versa. Sublethal and lethal concentrations to fish were up to 7 times lower for the mixture than in concurrently run individual compound exposures. Potential mechanisms for the synergistic responses found in fish are presented. Environ Toxicol Chem 2017;36:1947-1954. © 2016 SETAC.