Assessment of lethal and sublethal effects of imidacloprid, ethion, and glyphosate on aversive conditioning, motility, and lifespan in honey bees (Apis mellifera L.).

Honeybees (Apis mellifera) play an important role in agriculture worldwide. Several factors including agrochemicals can affect honey bee health including habitat fragmentation, pesticide application, and pests. The growing human population and subsequent increasing crop production have led to widespread use of agrochemicals and there is growing concern that pollinators are being negatively impacted by these pesticides. The present study compares acute exposure to imidacloprid (0.2 and 0.4 mgL-1), ethion (80 and 106.7 mgL-1) or glyphosate (0.12 and 0.24 mgL-1) on aversive learning and movement, to chronic exposure at these and higher concentrations on movement, circadian rhythms, and survival in honey bee foragers. For acute learning studies, a blue/yellow shuttle box experiment was conducted; we observed honey bee choice following aversive and neutral stimuli. In learning studies, control bees spent >50% of the time on yellow which is not consistent with previous color bias literature in the subspecies or region of the study. The learning apparatus was also used to estimate mobility effects within 20 min of exposure. Chronic exposure (up to 2 weeks) with the above metrics was recorded by an automated monitoring system. In chronic exposure experiments, RoundUp®, was also tested to compare to its active ingredient, glyphosate. We found that imidacloprid and ethion have negative impacts on aversive learning and movement following a single-dose and that chronic exposure effects were dose-dependent for these two insecticides. In contrast, glyphosate had no effect on learning and less of an effect on movement; RoundUp® showed dose-dependent results on circadian rhythmicity. Overall, the results suggest that short-term exposure to imidacloprid and ethion adversely affect honey bee foragers and chronic exposure to glyphosate may affect pollination success.

Purification and characterization of a phenoloxidase in the hemocytes of Ephestia kuehniella Zeller (Lepidoptera: Pyralidae): effects of insect growth regulators and endogenous inhibitors.

A phenoloxidase was extracted and purified from hemocytes of Ephestia kuehniella by using ammonium sulfate, Sepharyl G-100 and DEAE-Cellulose fast flow chromatographies. At the final stage of purification, a protein was purified by molecular mass of 78.5 kDa, specific activity of 1.17 U/mg protein, recovery of 20.48% and purification fold of 16.71. The purified PO showed the highest activity at pH 4-5 and temperatures of 35-40 °C. Na(+), K(+), Mn(+), Zn(2+) and Mg(2+) decreased activity of the purified PO but Ca(2+) and Cu(2+) increased the enzymatic activity. EDTA (General chelating agent), DTC (Copper chelating agent) and EGTA (Calcium chelating agent) significantly decreased PO activity but TTHA (Magnesium chelating agent) showed no statistically significant effects. Kinetic parameters of the purified enzyme showed the highest Vmax when L-DOPA was used as substrate but no significant differences were observed in case of Km for used L-DOPA, pyrocatechol and hydroquinone. In vitro inhibition of the purified PO by using two insect growth regulators, Hexaflumuron and Pyriproxyfen, revealed IC50 of 96.41 and 38.59 µg/ml for these compounds, respectively. Kinetic studies using different concentrations of L-DOPA and IC50 concentrations of the two IGRs revealed the increase of Km value versus control and competitive inhibition. Finally, column chromatography of hemolymph revealed peak III showing endogenous inhibitors of phenoloxidase by molecular weight of 27.3 that showed competitive inhibition on the PO.