Lead Tolerance And Transport Mechanism In Musca Domestica

Heavy metal pollution mainly refers to the pollution of atmosphere,water,soil and biosphere caused by heavy metals such as lead,cadmium,chromium,copper,cobalt,nickel,mercury,etc.with significant biological toxicity.It is difficult to reduce the content of heavy metals in the environment.Much attention is focus on the bio-transformation of heavy metal toxic forms.Controlling heavy metal pollution with insects is expected to be a new type of environmental management method,which would be effective,economical and safe.Musca domestica tolerant to heavy metals such as copper,zinc,cadmium,and lead.It reduces the toxicity of heavy metals through external excretion,change the enzyme activities of protective enzymes and detoxification enzymes and transfer heavy metals to tissues that are less harmful to the insects.However,how heavy metals enter the rest of insect body through it's intestine? What are the main ion channels and transport proteins it rely on? What parts components of the regulatory network are involved? The answers to these questions are helpful to the understanding of insects' mechanism on heavy metal stress tolerance.In this study,houseflies were used as experimental materials,and on the basis of clarifying the influence of lead on the growth and development,reproduction and nutrient accumulation of housefly,the genes and gut microbes involved in lead transport in the intestine of housefly were identified.The main results are as follows:1 The tolerance of housefly to leadHousefly can complete their entire life history under 20 and 5000 mg/kg lead treatments.Under lead treatment,the lead from in the feed can be transferred to the larvae,but as larvae grow,they can excrete some of the lead.Through monitoring the growth of larvae,it can be found that low-concentration lead treatment would not affect the growth and development of housefly larvae.High-concentration lead treatment significantly reduced the growth rate of larvae body weight,prolonged the development period,and reduced survival rate.Low-and high-concentration lead treatment had no significant effect on the development of pupae.Regarding adult reproduction,low-concentration lead treatment had no significant effect on adult reproduction,while high-concentration lead treatment caused changes in the dynamics of adult oviposition.Compared with the control group,there would be no oviposition peak,and the daily oviposion quantity production and total oviposion quantity were significantly reduced.2 Effects of lead on nutrient accumulation,digestive enzyme activity and gene expression of housefly larvaeThe growth of housefly larvae was related to the accumulation of nutrients.The results of nutrient accumulation,digestive enzyme activity and gene expression in the larvae showed that high-concentration lead treatment decreased the concentration of total sugar and total protein in the larvae,and low-concentration lead treatment decreased the accumulation of total protein in the larvae,but had no significant effect on the content of total sugar and total lipid.The nutrient accumulation of younger larvae was more susceptible to lead treatment.Under lead treatment,the number of fat droplets in the fat body cells of 4-day-old larvae increased,but the granules were smaller and cell distribution was more compact.The activities of larval amylase and trypsin were reduced under both low and high concentration lead treatments.The lipase activity was not affected under low concentration lead treatment,but the activity increased under high concentration lead treatment.Under lead treatment,the expression levels of amylase and lipase genes were down-regulated.Trypsin involved multiple genes,although the gene expression level changes,the changes of different genes were inconsistent.3 Identification and functional study of genes related to lead transport in housefly larvaeThe acidity and alkalinity of the digestive tract of housefly larvae were distributed in sections.The foregut and hindgut were neutral.The midgut was divided into three segments,with weak acidity in the anterior segment,strong acidity in the middle segment and weak alkalinity in the posterior segment.Lead treatment enhanced midgut acidity.The results of RNA-seq showed that under low-concentration lead treatment,the larvae had a large amount of differentially expressed genes related to material metabolism.Under high-concentration lead treatment,the differentially expressed genes were mainly clustered in cell membrane structure,material transport,energy metabolism and digestive enzyme activity.According to the results of the GO function analysis,13 genes involved in transmembrane transport and ion transport were screened out.Among them,ZCP A1 gene is closely related to the midgut lead transport process of house flies.CYT b-c1,GST 2and FER can slightly affect the midgut lead transport process of house flies.4 The effect of gut microbes on the transport of lead in housefly larvaeThe main gut microbes in housefly larvae were Providencia and Enterococcus.Under the lead treatment concentration of 5000 mg/kg,the proportions of Providencia and Ignatzschineria in the larvae decreased,while the proportion of Enterococcus and Klebsiella in the larvae increased.Through the isolation and purification of larval gut microbes,4 lead-resistant strains were obtained(Providencia heimbachae,Klebsiella Pneumoniae,Staphylococcus sciuri and Bordetella sp).When the lead content in the culture medium was 500 mg/L,Bordetella sp,P.heimbachae and S.sciuri can adsorb10% of lead,but K.pneumoniae can not reduce the lead content in the culture medium.This study analyzed the effects of lead on the growth,development and reproduction of housefly from the larvae,pupae,and adults,and clarified the tolerance of housefly to lead.Through the identification of genes and gut microbes related to the intestinal lead transport of housefly,and verify its function in the process of intestinal lead transport.The main genes and gut microbes involved in the intestinal lead transport process of housefly are identified.It is found that at the molecular level,ZCP A1,CYT b-c1,GST 2 and FER were involved in the intestinal lead transport process of housefly,and ZCP A1 played a key role.At the level of gut microbes,three microbial flora: Bordetella sp,P.heimbachae and S.sciuri adsorbed lead when lead level was high.The results of the study provide important evidence for exploring the main channels and transport carriers of lead transport in insects intestines,and provide important clues for revealing the molecular mechanism of insect tolerance to lead.