The aim of the present work was to assess the effects of landscape and pesticides on honey bee survival and physiological stress. Integrated use of acetylcholinesterase and detoxification enzymes was tested on honey bee brains for detecting possible exposure to pesticides. Foragers were tracked in agricultural and non-agricultural landscapes in West Tennessee (USA) and then recovered for molecular and chemical analyses. In addition, four honey bee cohorts were fed imidacloprid in the laboratory ad libitum for several weeks and were analyzed by RT-qPCR for gene expression. Pesticides were identified at different concentrations in both crop flowers and recovered foragers. No significant differences in foragers’ mortality were found among locations. Acetylcholinesterase and detoxification genes showed no response to exposure to pesticides except for GstS3 and GstS4. Our results suggest that none of the studied genes make suitable biomarkers for honey bee exposed to pesticides.