Mechanisms And Inheritance Of Propoxur Resistance In Housefly,Musca Domestica(Diptera:Muscidae)

The housefly is a worldwide human and animal health pests,it can transmit many human and animal intestinal diseases.For many decades,insecticides have been extensively used in the control of houseflies.The application of insecticides has resulted in the development chemical of insecticides resistance,such as organophosphorus,carbamates and pyrethroid insecticides.Life tables of PSS,CFD and N-PRR strains have been established.The CFD strain had longer larva duration,pupa duration and less fecundity egg in comparison with the PSS strain.The relative fitness in the CFD strain was 0.85 relative to the PSS strain.Results showed no significant differences in the net reproductive rate(Ro)and the intrinsic rate of increase(rm)between the PSS and N-PRR strain,indicating that fitness cost in the CFD strain was derived from genetic background in the field,and irrelevant to propoxur resistance in houseflies.Propoxur resistance was controlled by a major,incompletely recessive and autosomal factor in housefly,as determined by comparisons of the log dose-probit lines for PSS,N-PRR strain and progenies of reciprocal crosses,backcrossed and self-crosses.The N-PRR strain also showed different levels of resistance to various insecticides.High level of cross resistance was observed to phoxim(57.76-fold)and azamethiphos(57.54-fold).The cross resistance to bifenthrin and deltmathrin was only 12.06-and 4.28-fold,respectively.There was no cross resistance to beta-cypermethrin in N-PRR strain.S,S,S-tributyl phosphorotrithioate(DEF)had no significant synergism to propoxur in the N-PRR strain.But Diethyl maleate(DEM),the inhibitor of glutathione S-transferase,showed extremely synergism(>213.27-fold)to propoxur in the N-PRR strain,compared to 0.38-fold in the PSS strain.Synergist Piperonyl butoxide(PBO)increased propoxur toxicity>369.48-fold in the N-PRR strain as compared to 14.70-fold in the PSS strain.Results of biochemical assays revealed that glutathione S-transferase activity in the N-PRR strain was significantly higher than that in the PSS strain.But there was no significant difference in carboxylesterase and cytochrome P450 between the N-PRR and PSS strain.The Michaelis constant Km and maximum velocity Vma,in the N-PRR strain were 7.52 and 2.16-fold higher than that in the PSS strain,which indicated that the affinity of acetylcholinesterase in the N-PRR strain to the substrate reduced compared with that in the PSS strain,but hydrolysis ability of acetylcholinesterase to the substrate increased.Bimolecular rate constants Ki to five insecticides of the N-PRR strain were lower than that of the PSS strain,indicating that the sensitivity of the acetylcholinesterase was negatively correlated with the resistance level to propoxur in the N-PRR strain.CEA affinity chromatography and procainamide affinity chromatography were applied to purify acetylcholinesterase from the PSS and N-PRR strain in house flies.The purification ratios of procainamide affinity chromatography were 140.3 and 95.5 for the PSS and N-PRR strain,respectively.The sensitivity of purified acetylcholinesterase to five insecticides in the N-PRR strain decreased significantly compared with the PSS strain.The biochemical results suggested that qualitative change of acetylcholinesterase is the main mechanism to propoxur resistance in houseflies.Acetylcholinesterase gene expression levels using real-time quantitative PCR showed that relative expression of mRNA is 2.83-,0.85-and 1.56-fold for males in PSS strain,females in N-PRR strain and males in N-PRR strain compared to the females in the PSS strain,respectively.It indicated that quantitative change of acetylcholinesterase was not the mechanism of propoxur resistance in houseflies.Transcriptome sequencing analysis showed that 130 genes were significantly differentiate expression in the female housefly between the PSS and N-PRR strain,which contained 23 enriched GO terms and 3 pathways.They had 43 significantly different genes in expression related to the cytochrome P450 gene.There were 409 genes significantly different to male housefly between the PSS and N-PRR strain,which had 60 significantly enriched GO terms and 10 pathways.Among these genes,55 genes were related to the cytochrome P450 gene and 3 genes were related to the glutathione S-transferase.Therefore,the sensitivity reduction of acetylcholinesterase might confer the high levels of resistance to propoxur in the N-PRR strain of houseflies.The increase in glutathione S-transferase activity might play a role of propoxur resistance in houseflies.