Synergistic toxicity and physiological impact of imidacloprid alone and binary mixtures with seven representative pesticides on honey bee (Apis mellifera)

Imidacloprid is the most widely used insecticide in the world. In this study, we used spraying methods to simulate field exposures of bees to formulated imidacloprid (Advise® 2FL) alone and binary mixtures with seven pesticides from different classes. Synergistic toxicity was detected from mixtures of Advise (58.6 mg a.i./L imidacloprid)+Domark (512.5 mg a.i. /L tetraconazole), Advise+Transform (58.5 mg a.i./L sulfoxaflor), and Advise+Vydate (68 mg a.i./L oxamyl), and mortality was significantly increased by 20%, 15%, and 26% respectively. The mixtures of Advise+Bracket (88.3 mg a.i./L acephate) and Advise+Karate (62.2 mg a.i./L L-cyhalothrin) showed additive interaction, while Advise+Belay (9.4 mg a.i./L clothianidin) and Advise+Roundup (1217.5 mg a.i./L glyphosate) had no additive/synergistic interaction. Spraying bees with the mixture of all eight pesticides increased mortality to 100%, significantly higher than all other treatments. Except Bracket which significantly suppressed esterase and acetylcholinesterase (AChE) activities, other treatments of Advise-only and mixtures with other pesticides did not suppress enzyme activities significantly, including invertase, glutathione S-transferase (GST), and esterase and AChE. Immunity-related phenoloxidase (PO) activities in survivors tended to be more variable among treatments, but mostly still statistically similar to the control. By using specific enzyme inhibitors, we demonstrated that honey bees mainly rely on cytochrome P450 monooxygenases (P450s) for detoxifying Advise, while esterases and GSTs play substantially less roles in the detoxification. This study provided valuable information for guiding pesticide selection in premixing and tank mixing in order to alleviate toxicity risk to honey bees. Our findings indicated mixtures of Advise with detoxification-enzyme-inducing pesticides may help bees to detoxify Advise, while toxicity synergists may pose further risk to bees, such as the Bracket which not only suppressed esterase and AChE activities, but also increased toxicity to bees.