Honey bees (Apis mellifera) play a pivotal role in agricultural production worldwide, primarily through the provision of pollination services. But despite their importance, honey bee health continues to be threatened by many factors, including parasitization by the mite Varroa destructor, poor queen quality, and pesticide exposure. Accumulation of pesticides in the hive’s comb matrix over time inevitably leads to the exposure of developing brood, including queens, to wax contaminated with multiple compounds. Here, we characterized the brain transcriptome of queens that were reared in wax contaminated with pesticides commonly found in commercial beekeeping operations including either (a) a combination of 204,000 ppb of tau-fluvalinate and 91,900 ppb of coumaphos (“FC” group), (b) a combination of 9,800 ppb of chlorpyrifos and 53,700 ppb of chlorothalonil (“CC” group), or (c) 43,000 ppb of amitraz (“A” group). Control queens were reared in pesticide-free wax. Adult queens were allowed to mate naturally before being dissected. RNA isolated from brain tissue from three individuals per treatment group was sequenced using three technical replicates per queen. Using a cutoff log2 fold-change value of 1.5, we identified 247 differentially expressed genes (DEGs) in the FC group, 244 in the CC treatment group, and 668 in the A group, when comparing each group to the control. This is the first study to examine the sublethal effects of pesticides commonly found in wax (particularly amitraz) on the queen’s brain transcriptome. Future studies should further explore the relationship between our molecular findings and the queen’s behavior and physiology.

Hookworms are infectious parasites that affect more than 500 million people worldwide, causing extensive morbidity and economic burden in developing countries. Mass drug administration is a current control method, but resistance is of concern. Many hookworm species, including those in the genus Ancylostoma, are specialists and require specific host signals to continue development upon infection. Little is known about the signaling mechanisms, although evolutionary conservation between parasitic nematodes and the model organism C. elegans suggests that G‐protein coupled receptors (GPCRs) expressed in amphid neurons are chemosensory detection candidates. The amphid neurons of C. elegans are exposed to the environment, even during the dauer stage, an alternate phase of larval development that is similar to the parasitic infective juvenile stage. Previous experiments involving A. ceylanicum transcriptional GFP fusions in C. elegans identified putative GPCR genes expressed in neurons during dauer stage. To confirm that hookworm proteins are neuronally localized under dauer conditions, a translational GFP fusion of a putative hookworm GPCR was generated. The entire coding sequence, plus introns, was cloned using homologous recombination‐based, yeast assembly methods to create a C‐terminal GFP fusion. The putative A. ceylanicum promoter consisted of the upstream intergenic region, which is approximately 5000 base pairs, and the transcriptional terminator from C. elegans unc‐54 was used. To generate the expression plasmid, amplicons containing homology to the adjacent segment in the assembly were amplified using polymerase chain reaction (PCR) and transformed into yeast. The plasmid was verified by junction‐confirming PCR analysis, restriction enzyme mapping, and sequencing. After microinjection, GFP expression in amphid neurons of stably transformed C. elegans during the dauer stage would support the hypothesis that this putative GPCR plays a role in reception of host signaling molecules and may provide a possible therapeutic target.Support or Funding InformationThis work was supported by the Salisbury University Faculty Mini‐Grant Program.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Honeybees, Apis mellifera, are crucial pollinators in our world today. Approximately one third of the world's agriculture depends upon pollination by insects to increase crop yields. During their flights, which can cover up to five miles, bee flight muscles oxidize carbohydrates to produce ATP, through the processes of glycolysis in the cytoplasm and Krebs cycle/oxidative phosphorylation in the mitochondria. To transport pyruvate, the product of glycolysis, from the cytoplasm through the inner mitochondrial membrane, a transporter protein known as the mitochondrial pyruvate carrier (MPC), is required. If this transporter protein is inhibited, honeybees can't produce adequate energy for flight. One way this transporter could be impaired is by a class of pesticides called neonicotinoids, particularly Imidacloprid. This pesticide is the biggest concern to the honeybee because it is used on about 95% of corn and canola crops, and many fruits and vegetables. The seeds are coated in the pesticide, and it is assimilated and incorporated into the pollen and nectar which is consumed by the bees. Previous studies have shown that neonicotinoids affect the mitochondria by depolarizing the inner mitochondrial membrane potential which disrupts the proton gradient necessary for chemiosmosis. This could potentially have other effects within the mitochondria during the process of cellular respiration. In our experiment, we are testing the potential effects of Imidacloprid on the mitochondrial function of honeybees by examining rates of respiration and the expression levels of the MPC gene. We hypothesize that honeybees will up‐regulate the MPC as a response to the pesticide in order to increase their respiration rates and maintain ATP production. To test our hypothesis, we fed bees sugar solutions containing Imidacloprid (25ppb) and a control group. After 4 days of exposure RNA was extracted and converted to cDNA for quantitative PCR for control and 25ppb treatments. The qPCR results will allow us to determine the level of MPC gene expression within the control and exposed bees which we can then compare to determine if the gene was up‐ or down‐regulated. Mitochondria isolated from each group were tested in vitro using an Oxygraphplus to examine potential effects of Imidacloprid on respiration rates. Mitochondrial respiration rates for state 4 in our control and exposed honeybees were similar, 58.9nmol O2/min*mg protein, control, and 56.93nmol O2/min*mg protein, exposed. In our state 3 respiration our exposed honeybees had a higher rate, 104.94nmol O2/min*mg protein, as compared to our control group, 92.4nmol O2/min*mg protein. No statistical difference was observed between the control and exposed honeybees state 3, p=0.22, or state 4, p=0.26, respiration rates. We are currently running qPCR to test for changes in MPC gene expression in response to Imidacloprid exposure.Support or Funding InformationWe would like to thank Henson School of Science and Technology and Salisbury University's Green Fund for all of their support and funding through out this project.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.