Effects Of Carbendazim Stress On The Health Of Honeybee(Apis Mellifera Ligustica)

In recent years,carbendazim in agricultural production in our country is becoming more and more frequent application of a kind of fungicides.At the same time,the effect of the sublethal dose of pesticides on honeybee colonies has been constantly expanding.Rapeseed and other crops are widely used,it has a great influence on the pattern of turning the ground to release honeybees for chasing flowers and honey in China.Some studies have shown that carbendazim has the highest detection rate in pollen.As one of the most abundant bee species in China Apis m ellifera ligustica is very important to China’s agricultural production and ecological environment.When a large number of honeybees disappear in foreign countries,the mass death of honeybees during the overwintering period in China gradually attracts people’s attention to the colony health.There are few studies on the effect of carbendazim on honeybees at home and abroad;In this study,the effects of carbendazim sublethal dose on the growth and development of honeybee(Apis mellifera ligustica)larvae and the activity of detoxification enzymes were studied;Effects of carbendazim in five groups(CK:0 ng/mg,A:5 ng/mg,B:50 ng/mg,C:500 ng/mg,D:5000 ng/mg)on intestinal microorganisms of honeybees(Apis mellifera ligustica)under different concentrations;analysis of brain transcriptome of newly emerged bees exposed to sublethal doses of carbendazim at developmental stage.These three main aspects explain the impact of carbendazim stress on the health of honeybee(Apis mellifera ligustica),and are intended to provide guidance for the agricultural application of carbendazim.The results of the study are as following:1.Carbendazim at absolute sublethal doses(referred to the effective concentrations of carbendazim:250 ng/mg(A)and 750 ng/mg(B),respectively),the results on the growth and development of Apis mellifera ligustica larvae and the activity of detoxifying enzymes showed,under this sublethal dose of carbendazepam,the effect on the pupal weight,pupation rate and eclosion rate of 1-day-old pupae of honeybee larvae was not significant(P>0.05).However,the concentration of total protein and the activity of total superoxide dismutase(T-SOD)in honeybee larvae were significantly affected,the activity of superoxide dismutase in bee larvae was significantly increased after carbendazim treatment(P<0.05),at 750 ng/mg(B),the total protein concentration was significantly reduced compared with the control group(P<0.05).In this experiment,the concentration of carbendazim could significantly inhibit(P<0.01)Ecdysone(Ecd)and significantly promote(P<0.05)the secretion of Juvenile hormone(JH).Meanwhile,the detoxifying enzyme system of bees was also greatly affected,when the concentration of carbendazim was 250 ng/mg(A),cytochrome P450(CYP450)and carboxylesterase(CarE)activities were significantly increased compared with the control group(P<0.05)2.Results of the analysis of the effects of carbendazim on the intestinal microorganisms of honeybee(Apis mellifera ligustica)under five groups of different concentrations showed that compared with CK,the abundance of intestinal flora Firmicutes in group A increased,while that in group B,C and D decreased;Proteobacteria in the intestinal flora of group A decreased,while group B,C and D increased.Through the analysis of Alpha Diversity analysis index,the diversity and richness of intestinal flora in the treatment groups of each experimental treatment group changed but did not reach a significant level,comparative analysis of the differences between groups,such as NMDS T_test,showed that only group C significantly increased the abundance of proteobacteria in the composition of intestinal microorganisms(P<0.05),and significantly decreased the abundance of firmicutes(P<0.05).It can affect the honeybees’ intake of food nutrients and damage the immune system,leading to colony collapse.3.Analysis of brain transcriptome of newly emerged bees exposed to sublethal doses of carbendazim at developmental stage found that of the 247 DEGs identified,187 were up-regulated and 60 were down-regulated.The classification of biological functions mainly consists of three aspects:molecular functional biological processes and cellular components.Exposure to carbendazim during larval stages alters expression of major royal jelly protein(MRJPs)encoding genes in adult honeybee brains.DEGs successfully mapped 692 significantly rich GO terms(Q<0.05),and selected the top 10 rich terms in each category,and then used KEGG analysis,which mainly affects the degradation of tryptophan such as leucine.Metabolism and the Generation of Ribosomes in Eukaryotes.Under the stress of carbendazim,the expression of cytochrome P450,such as P450 6AS5(GB49890)and 4aa1(GB49626),was up-regulated,and apolipoprotein D(GB54020)encoding one of the 16 transporter and receptor genes was down-regulated.Changes in these genes may adversely affect the detoxification system and lipid metabolism of bees.Certain genes(dopamine transporter(DAT,GB40867),SIF amide(GB40093),etc.)related to brain development of newly emerged bees were significantly down-regulated under carbendazim exposure(P<0.05),genes associated with brain-related and neuropeptide signaling pathways(GO:0007420 and GO:0007218)are also significantly downregulated(P<0.05),which may adversely affect the neural signaling pathways and sensory systems of honeybees’ brain development.