Preliminary Study On The Fumigation Activity And Mechanism Of The Mixed Use Of Ethyl Formate And Methyl Isothiocyanate On Phyllostachys Vulgaris

Acanthoscelies obtectus(Say)belongs to Acanthoscelides of Bruchidae,Coleoptera.It is an important pest endangering a variety of beans.It has fast spreading speed,complex transmission routes and strong ecological adaptability.It has been listed as an important quarantine pest in China.The A.obtectus(Say)feeds on bean seeds by larvae,the Phaseolus vulgaris loses the edible value and the commodity value after being eaten by the A.obtectus(Say)larvae.By studying the mixed fumigation effect of ethyl formate plus methyl isothiocyanate on A.obtectus(Say),an optimal formulation with high toxicity,strong quick effect and low cost was selected.The fumigation effect of the optimal formula on A.obtectus(Say)at different development stages(egg,larvae,pupae and adult)was systematically studied.The molecular mechanism of fumigation and insecticidal effect of the optimal formula on A.obtectus(Say)was studied by transcriptome analysis,which provided theoretical basis for comprehensive control of A.obtectus(Say).1?Study on the optimal fumigation formula of ethyl formate plus methyl isothiocyanate for the adults of Acanthoscelides obtectus(Say)In order to explore the fumigation effect of ethyl formate plus methyl isothiocyanate on the adults of Acanthoscelides obtectus(Say)and the optimal formula for prevention and control.Based on the measurement of the median lethal concentrations of ethyl formate plus methyl isothiocyanate by wide-port bottle closed fumigation method,the two-factor quadratic regression rotation combination design was used to obtain the optimal formula for the control effect and economic benefit according to the La Grange extremum principle.There was a great difference in the toxicity of the two fumigants to the adults of A.obtectus.The insecticidal effect of methyl isothiocyanate was the best(LC₅₀value was 0.667 mg/L),followed by ethyl formate(LC₅₀value was 20.59?L/L).A quadratic polynomial can be obtained by the two-factor quadratic regression rotation combination.According to the optimization objective function and constraint function,the dosage of ethyl formate in the mixed solution is 102.702?L,and that of methyl isothiocyanate is 3.0263 mg.When the ratio of ethyl formate and methyl isothiocyanate was 33.94?1,the best economic benefit and insecticidal effect were obtained.2?Fumigation activity of the optimum formula of ethyl formate plus methyl isothiocyanate on Acanthoscelides obtectus(Say)In this paper,we sealed jar to fumigate Acanthoscelides obtectus(Say).The killing effects of effects mixed fumigation of ethyl formate plus methyl isothiocyanate(mass ratio 97:3)on A.obtectus were measured under different concentrations,time and temperature conditions.The results showed that the treatment time and temperature significantly affected the mixed fumigation on the adults.The corrected mortality of adults could reach 100%when treated with concentration of 20?L/L of mixed fumigation for more than 60 hours at 25?.The better fumigation effect was obtained at relatively low temperature(15-20?)than that at relatively high temperature(25-35?).When the fumigation temperature was 20?,the fumigation effect of the mixture fumigation was egg>adult>larva>pupae in descending order.3?The control effect of ethyl formate plus methyl isothiocyanate mixed with optimal formula on Acanthoscelides obtectus(Say)in simulated warehouse.The results showed that the mixture had the best fumigation effect on eggs,followed by adults,larvae and pupae in simulated warehouse.When the concentration was 4?L/L,the corrected mortality of eggs in the middle and lower layers of the simulated warehouse reached 100%.When the concentration was 20?L/L,the corrected mortality of adults in the lower layer of the simulated warehouse reached100%,and there were a few surviving adults in the middle layer.When the concentration was 70?L/L,the best fumigation effect of larvae and pupae in the simulated warehouse was in the lower layer,and the corrected mortality was 57.97%and 23.54%.4?Transcriptome analysis of Acanthoscelides obtectus(Say)under fumigant stress.We used transcriptome analysis to compare the expression patterns of resistance and stress-response genes in A.obtectus subjected to ethyl formate,methyl isothiocyanate,and ethyl formate plus methyl isothiocyanate(mass ratio 97:3),to determine the molecular mechanisms of resistance to these fumigant.After removing the low-quality sequence and further assembling,33537 unigenes were obtained,the average length is 1,395 bp.According to the homology analysis of unigenes annotated in Nr database,58.89%of unigenes showed the highest homology with Callosobruchus maculatus,while 11.21%of unigenes have higher homology with related genes of Anoplophora glabripennis.A comparative analysis of gene expression under ethyl formate,methyl isothiocyanate,and ethyl formate plus methyl isothiocyanate(mass ratio 97:3)stress revealed 106,193,and 637 upregulated unigenes;65,100,and 1,108 downregulated unigenes.Lysine degradation and RNA transport signaling pathways were significantly enriched after treatment with the three chemicals.After ethyl formate treatment,three detoxification enzyme genes were significantly downregulated,including one P450family gene,one Car E family gene and one EST family gene.After methyl isothiocyanate treatment,three detoxification enzyme genes were significantly downregulated,including two P450 family genes and one GST family gene.Four P450 family genes were significantly downregulated after treatment with ethyl formate plus methyl isothiocyanate(mass ratio 97:3).These signaling pathways and related genes play important roles in response to fumigation stress.In order to verify the reliability of RNA-seq results,fluorescence quantitative PCR was used to verify the results of differential gene expression analysis.RT-PCR and RNA-seq results were consistent,which indicated that RNA-seq results were reliable.The molecular mechanism of fumigation and insecticidal effect of ethyl formate plus methyl isothiocyanate on A.obtectus was preliminarily studied.