Metabolomics Studies Of Azadirachtin On Bactrocera Dorsalis Larvae

Bactrocera dorsalis is a worldwide pest caused serious economic losses.As the most excellent biopesticide,azadirachtin has good biological activity against more than 10 orders and 400 species pests.However,the study of azadirachtin against B.dorsalis larvae is rare.Until recently although it is reported that azadirachtin could affect the metabolites of carbohydrates,fatty acids,amino acids,cholesterol,uric acid and urea,these studies mainly focus on its effect on special metabolites and their content.Moreover,the biological meaning of these differentially expressed metabolites have not been deeply and systematically analyzed.Azadirachtin could disrupt insects’ internal secretion,and affect the synthesis and release of hormones,causing block of the metamorphosis,thus interfering with insects’ normal growth and development.But what are the differences among azadirachtin acting as one insect growth regulator with the juvenile hormone and the molting hormone on insects’ metabolites and pathways.It has not been reported until recently.This dissertation based on the biological activity of azadirachtin against B.dorsalis larvae,through introducing the UPLC-MS/MS-based and GC-MS-based metabolomics approach,after metabolites extracting and derivatization,raw data collection,metabolites qualitative and quantitative analysis,univariate and multivariate statistical analysis,and metabolic pathway analysis,the integrative changes of azadirachtin on the metabolites and pathways of B.dorsalis larvae have been studied.So does its comparative effects on the metabolites and pathways of B.dorsalis larvae with pyriproxyfen and tebufenozide.Comparing with pyriproxyfen and tebufenozide,the developmental duration of B.dorsalis larvae in azadirachtin treated group is significantly longer than that of pyriproxyfen and tebufenozide group.But the survival and pupal weight in azadirachtin treated group is significantly lower than that of pyriproxyfen and tebufenozide group.These suggested that endogenous metabolites and pathways of larvae in different groups should been different.A total of 21 and 13 differentially regulated metabolites were screened from UPLCMS/MS and GC-MS based metabolomics,respectively.And the 7 key metabolic pathways of histidine metabolism,D-glutamine and D-glutamate metabolism,biotin metabolism,ascorbate and aldarate metabolism,pentose and glucuronate interconversions,and alanine,aspartate and glutamate metabolism in B.dorsalis larvae were screened as the key metabolic pathway significantly affected by azadirachtin.A total of 16,13 and 10 differentially regulated metabolites were screened from the comparative metabolomics study of azadiracthin,pyriproxyfen and tebufenozide,respectively.The three pathways of cyanoamino acid metabolism,ascorbate and aldarate metabolism,and pentose and glucuronate interconversions in B.dorsalis larvae were screened as the key metabolic pathway significantly affected by azadirachtin;the two pathways of cyanoamino acid metabolism,and pentose and glucuronate interconversions were screened as the key metabolic pathway significantly affected by pyriproxyfen;the two pathways of propanoate metabolism and beta-alanine metabolism were screened as the key metabolic pathway significantly affected by tebufenozide.A total of 10,8 and 3 differentially regulated fatty acids were screened from the comparative study of azadiracthin,pyriproxyfen and tebufenozide,respectively.Although these differentially regulated fatty acids in azadirachtin group could enrich into the three pathways of fatty acid elongation,fatty acid degradation and fatty acid biosynthesis,the enrichment analysis results were not significant.Those differentially regulated fatty acids in pyriproxyfen and tebufenozide group could not enrich into any pathways.The results of laboratory biological activity showed that comparing with pyriproxyfen and tebufenozide,azadirachtin has a good biological activity against B.dorsalis larvae.It could be considered to be included in the integrated control of B.dorsalis on the basis of the application technology having been solved.By introducing metabolomics approach into the research of azadirachtin and botanical pesticides,it has showed that azadirachtin have obvious effects on the amino acid,carbohydrate,nucleotide,fatty acid,and vitamin and cofactor metabolism of B.dorsalis larvae.It has enriched the mechanism knowledge of azadirachtin on insects.Through comparing the changes in the two perspectives of endogenous metabolites and pathway of B.dorsalis larvae,it could conclude that azadirachtin shows similar main metabolic effects in B.dorsalis larvae as pyriproxyfen,but shows a different main metabolic effects with tebufenozide.This not only confirmed the feasibility of using metabolomics approach to study the effect of chemical pesticides on insect metabolism,but also enriched the knowledge of the mechanism of pyriproxyfen and tebufenozide on insects.