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Neonicotinoid seed treatments are frequently used in cotton (Gossypium hirsutum L. [Malvales: Malvaceae]) production to provide protection against early-season herbivory. However, there is little known about how these applications affect extrafloral nectar (EFN), an important food resource for arthropod natural enemies. Using enzyme-linked immunosorbent assays, we found that neonicotinoids were translocated to the EFN of clothianidin- and imidacloprid-treated, greenhouse-grown cotton plants at concentrations of 77.3 ± 17.3 and 122.6 ± 11.5 ppb, respectively. We did not find differences in the quantity of EFN produced by neonicotinoid-treated cotton plants compared to untreated controls, either constitutively or after mechanical damage. Metabolomic analysis of sugars and amino acids from treated and untreated plants did not detect differences in overall composition of EFN. In bioassays, female Cotesia marginiventris (Cresson) (Hymenoptera: Braconidae) parasitoid wasps that fed on EFN from untreated, clothianidin-treated, or imidacloprid-treated plants demonstrated no difference in mortality or parasitization success. We also conducted acute toxicity assays for C. marginiventris fed on honey spiked with clothianidin and imidacloprid and established LC50 values for male and female wasps. Although LC50 values were substantially higher than neonicotinoid concentrations detected in EFN, caution should be used when translating these results to the field where other stressors could alter the effects of neonicotinoids. Moreover, there are a wide range of possible sublethal impacts of neonicotinoids, none of which were explored here. Our results suggest that EFN is a potential route of exposure of neonicotinoids to beneficial insects and that further field-based studies are warranted. [ABSTRACT FROM AUTHOR]