Effects Of Sublethal Concentration Of Thiacloprid On Learning And Memory Of Apis Mellifera Worker Bees

As an important pollinator,bees make a huge contribution to global agricultural production every year.Neonicotinoid insecticides are used in agriculture widely because its highly toxic to insects than to mammals.But neonicotinoid will harm non-target insects（honeybees）,while killing pests.Neonicotinoid insecticides mainly act on the neuronal system of bees by blocking the normal acetylcholine transmission,which could induce apoptosis of neurons,affect learning and memory behaviors,and then impair foraging abilities of bees.Previous studies about the effects of neonicotinoid insecticides on learning and memory of honeybees is mainly focused on imidacloprid and thiamethoxam,there are few studies on relatively lower toxic thiacloprid.In this paper,we treated the Apis mellifera with subleathal dose of thiacloprid to investigate the effect of thiacloprid on development,learning and memory proformances.Furthermore,we used the immunohistochemical staining of neurons to discuss the mechanism of the effect of thiacloprid on the learning and memory of honeybees.Under laboratory conditions,the effects of sublethal concentrations of thiacloprid on learning and memory of emergent bees and collected bees were studied.First,the 48-hour thiacloprid median lethal concentration was determined for the bees in the experimental bee farm.The results showed that the48-hour median lethal concentration of thiacloprid to the bees in the experimental bee farm was 12.87mg/L.Secondly,determined the concentration of thiacloprid used in exposure experiments of emergent bees and collected bees within the range of LC₅₀/10～LC₅₀/100.In the experiment of emergent bees,healthy bee larvae were transferred to the laboratory for in vitro rearing.The treated group larvae were fed with food mixed with thiacloprid,the concentration was0.5,and 1 mg/L,the control group larvae were fed with normal food.After 18 days of culturing in the laboratory,the newly emerging adult were collected and tested for associate olfactory learning and memory behaviors.The results showed that exposure to sublethal doses of thiacloprid at the larval stage reduced the learning efficiency and memory retention time of adults.The bee larvae exposured to 1mg/L of thiacloprid would reduce capped rate and eclosion rate.During this period,the body weight of the larvae did not affect.In the experiment of collected,newly emerged bees were collected and raised in the laboratory.They were exposed to different concentrations of thiacloprid for 12 days.After the end of exposure period,the bees were collected,and they were tested the associated olfactory learning and memory behaviors.The results showed that the adult bees exposured to sublethal concentrations of thiacloprid would reduce the learning efficiency and memory retention time.At the same time,immunohistochemical staining techniques were used to determine the brain structure of the olfactory lobe（AL）,internal chiasma（IC）,external chiasma（OC）,and optic ganglion layer（lamina,The apoptotic rate of nerve cells in 5 regions of LA）and mushroom bodies（MB）.The results of TUNEL staining and Caspase-3 staining showed that honeybees exposured to sublethal concentrations of thiacloprid could increase the apoptosis rate of brain nerve cells.These studies evaluated the risk of thiacloprid to Apis mellifera,and provide insights into neural mechanism of thiacloprid impair honey bees,and thus establish new honey bee safety evaluation guidelines.