Currently, there is a trend toward an increasing use of biopesticides assumed to be environmentally friendly, such as Bacillus thuringiensis (Bt). Studies of the Bt toxicity to nontarget organisms have reported low effects at high exposure levels, which is interpreted as indicating negligible risk to nontarget organisms. We investigated the response of the nontarget organism Daphnia magna to waterborne DiPel ES, a globally used Bt formulation. Neonates and adults were exposed for 48 h to a wide range of concentrations, and immobilization and mortality were monitored. Whole body biomarkers (body weight, protein, chitobiase, catalase, xenobiotic metabolism, and acetylcholinesterase) were measured in the adults. The immobilization and mortality of the neonates were affected in a nonmonotonic and inverted U-shaped pattern with EC50s that were ∼105-fold lower than those reported by the manufacturer. The immobilization of adults demonstrated a similar pattern, but significant mortality was not observed. The biomarker results revealed multiphasic dose-response curves, which suggested toxicity mechanisms that affected various physiological pathways. The main particle size in exposure media was in the size range of bacterial spores and crystal toxins. However, the chemical heterogeneity was nonmonotonic, with a change in the phase at the maximum of toxicity (∼5 μL L-1), which might explain the observed nonmonotonic effects. These results demonstrate the vulnerability of a nontarget organism to a biopesticide that is considered to be safe, while challenging the universal applicability of the central ecotoxicological assumption of monotonicity.