Interactions of Pesticides with Drug Efflux Transporter P‐glycoprotein in the European Honeybee

Modern agriculture depends on pollinators such as the European Honeybee, Apis mellifera, to produce crops. It is estimated that almost $20 Billion a year in US agricultural production can be contributed to honeybees. However, global bee populations have been in recent decline and the high use of agricultural pesticides have been suggested to directly or indirectly affect bee colony health. Yet, little is known about the types and levels of pesticides accumulation in bees and the underlying mechanism of uptake and distribution. Multidrug resistance proteins from the ABC family, such as P‐glycoprotein (P‐gp), are a first line of defense in biological barriers and responsible for the efflux of drugs and toxins in all organisms. This study aims to determine the molecular interactions of certain prevalent pesticides with the honeybee P‐glycoprotein to predict the bioaccumulation potential of these compounds, including neonicotinoids, glyphosate and fipronil. We are currently cloning honeybee P‐gp from forager bee specimen. Next, we will express and purify honeybee P‐gp and use an optimized ATPase activity assay to define inhibitors, activators and non‐interacting compounds. Compounds identified as P‐gp inhibitors or non‐interacting chemicals will be infused into sugar water and fed to living bees. Total body burden of these compounds will then be determined using LC‐ and GC mass spectrometry. Finally, we will dissect bees and separate the digestive system (foregut, midgut, hindgut) to validate P‐gp expression using Western immunoblotting. Our preliminary data has shown that a commercially available human P‐gp epitope antibody can recognize honeybee P‐gp. Future experiments will aim at studying behavioral changes upon P‐gp inhibitory pesticide exposure, including changes in flight paths and waggle dance in living colonies. In addition, we will determine the interactions of these pesticides with another key efflux transporter in bees, the multidrug resistance protein 1 or MRP1. The results of this study will help to identify the bioaccumulation potential of currently used agricultural pesticides and guide the rational design and use of compounds that are better eliminated by bees. Finally, it will inform farmers, stakeholders and governmental agencies about non‐lethal combinations of pesticides often used as mixtures for proper crop protection. Support or Funding Information This study is supported by the Principle Investigator Start‐up funds from UC Davis.