A mitotoxic fungicide alters post-ingestive glucose signals necessary for associative learning in honey bees

The Proboscis Extension Reflex (PER) paradigm trains honey bees to associate an odor with a sugar reward and is commonly used to assess impacts on associative learning after exposure to pesticides. While the effects of some types of pesticides have been well-investigated, relatively little attention has been focused on fungicides that are applied to flowering crops. We have previously shown that consumption of field-relevant concentrations of the fungicide Pristine® (active ingredients: 25.2% boscalid, 12.8% pyraclostrobin) impairs honey bee performance in an associative learning assay, but the mechanism of its action has not been investigated. We hypothesized that Pristine® interferes with carbohydrate absorption and/or regulation, thereby disrupting the post-ingestive feedback mechanisms necessary for robust learning. To test this hypothesis, we measured hemolymph glucose and trehalose levels at five time points during the ten minutes after bees consumed a sucrose solution. Pristine®-exposed bees had elevated baseline glucose concentrations in the hemolymph relative to control bees. Hemolymph glucose levels rose significantly within five minutes of feeding in control bees, but not in Pristine®-fed bees. These data suggest that the post-ingestive feedback mechanisms necessary for robust learning are disrupted in bees that have consumed this fungicide, providing a plausible mechanistic explanation for its effects on learning performance in the PER assay. Pristine®-exposed bees may have elevated hemolymph glucose levels because the fungicide elicits an inflammatory response. These results provide additional mechanistic understanding of the negative physiological effects of mitotoxic fungicides on this important pollinator.